@article {70284, title = {Astrocyte-induced internal state transitions reshape brainwide sensory, integrative, and motor computations}, journal = {bioRxiv}, year = {2026}, month = {2026 Feb 05}, abstract = {

Animals rapidly adapt to changing circumstances by shifting how they perceive, integrate, and act. Such flexibility is often attributed to transitions between internal states that exert widespread influence across the brain. Yet the mechanisms that drive state transitions and how they reconfigure brainwide computation remain unclear. Larval zebrafish, when actions are rendered futile by decoupling visual flow feedback from swimming in virtual reality, enter a temporary passive, energy-preserving state. In this state, astrocyte calcium levels are elevated, and swim reinitiation requires greater accumulated visual motion. Using whole-brain, cellular-resolution activity imaging, we observed widespread circuit alterations underlying this disengaged state: neuronal visual responses weakened, visual motion integration over time became dramatically leakier, motor inhibition increased, and motor preparation slowed, together suppressing conversion of sensory evidence into action. Astrocyte calcium rose during futile swimming, tracked the emergence and resolution of these brainwide changes, and was both necessary and sufficient to drive them. Thus, astrocytes orchestrate internal states that profoundly reshape neural computations, most powerfully at intermediate integrative processing stages, to meet changing demands.

}, doi = {10.64898/2026.02.05.704034}, url = {https://www.biorxiv.org/content/early/2026/02/05/2026.02.05.704034}, author = {Lim, Jing-Xuan and Wei, Ziqiang and Narayan, Sujatha and Zhang, Yan and Hasseman, Jeremy P. and Kolb, Ilya and Zheng, Jihong and Sheikhattar, Alireza and Mi, Xuelong and Zheng, Wei and Yang, Xueying and Beriashvili, Mariam and Fleishman, Greg and Wee, Caroline L. and de Zeeuw, Chris and Yu, Guoqiang and Babadi, Behtash and Rubinov, Mikail and Looger, Loren L. and Bergles, Dwight E. and Fitzgerald, James E. and Ahrens, Misha B.} } @article {70003, title = {BPS2026 {\textendash} De novo-designed rhodamine binders for advanced fluorescence microscopy}, journal = {bioRxiv}, year = {2026}, month = {2026 Feb 19}, abstract = {

Fluorescence microscopy is a powerful tool for studying biomolecules in their native environments, offering high spatio-temporal resolution but requiring fluorescent labels. Current live-cell compatible labeling strategies repurpose natural systems, such as fluorescent proteins or proteins that bind fluorescent ligands. While advances have been made to engineer natural proteins into labels with minimal size, high brightness, as well as enhanced thermo- and photostability, these approaches often require trade-offs among desirable properties due to the inherent limitations of re-engineering natural proteins. In this work, we present rhodamine binder (Rhobin) tags{\textemdash}de novo-designed proteins that bind rhodamine-derived fluorophores. Unlike traditional approaches, Rhobin tags were developed by directly incorporating desirable features during the computational design process, which resulted in compact binders with outstanding thermostability. Their nanomolar substrate affinity, rapid labeling kinetics, and orthogonality to established labeling systems such as HaloTag and SNAP-tag facilitate versatile live and fixed-cell imaging of diverse subcellular targets in mammalian cells. Transient fluorophore binding further enables advanced imaging techniques, including live-cell super-resolution STED microscopy with reduced photobleaching and single-molecule localization microscopy in live and fixed cells. To the stability of Rhobin tags under extreme environmental conditions, we demonstrate showcase protein tagging and timelapse imaging in the extremophile\ Sulfolobus acidocaldarius\ living at 75{\textdegree}C, an application previously inaccessible with existing tags. We anticipate that Rhobin tags will become a central component in the toolbox of fluorescent labels and will pave the way for a new generation of modular protein tags and biosensors with tailor-made properties.

}, doi = {10.1016/j.bpj.2025.11.2270}, url = {https://doi.org/10.1016/j.bpj.2025.11.2270}, author = {Yserentant, Klaus and Chen, Yuda and Hong, Kibeom and Kuang, Yiming and Bhowmick, Arghya and Charles-Orszag, Arthur and Lord, Samuel J. and Lu, Lei and Hou, Kaipeng and Mann, Samuel I and Grimm, Jonathan B and Lavis, Luke D and Mullins, R. Dyche and DeGrado, William F and Huang, Bo} } @article {70309, title = {Brain-wide hierarchical and sexually dimorphic tuning for social vocalizations}, journal = {bioRxiv}, year = {2026}, month = {2026 Mar 04}, abstract = {

Acoustic communication is widespread among vertebrates and central to social behavior. Yet how brain-wide circuits identify conspecific signals and distinguish acoustic elements with different, often sex-specific social valence remains poorly understood. Here we present the first whole-brain analysis of neuronal responses to conspecific vocalisations in vertebrates, using the transparent fish Danionella cerebrum. Combining volumetric calcium imaging with playbacks probing the stimulus space of the natural sound repertoire, we uncover an unexpectedly early and specialized processing hierarchy: hindbrain nuclei already segregate vocalization-like pulse trains from tones, midbrain regions sharpen these representations and extract temporal features that define vocalization type, and the central posterior thalamic nucleus responds selectively to conspecific vocalization rates and thus acts as a gate for social sounds. Male and female brains share this early feature code but diverge in diencephalic and telencephalic regions, where identical acoustic features evoke sex-specific population activity patterns that parallel dimorphic behavior. Together, our results provide the first cellular-resolution, brain-wide account of social sound processing in a vertebrate, from early categorical segregation to thalamic gating and sex-specific population responses in social circuits.

}, doi = {10.64898/2026.03.04.709502}, url = {https://www.biorxiv.org/content/early/2026/03/04/2026.03.04.709502}, author = {Henninger, Jorg and Hoffmann, Maximilian and Kadobianskyi, Mykola and Veith, Johannes and Berlage, Caroline and Groneberg, Antonia and Markov, Daniil and Schulze, Lisanne and Svanidze, Ana and Maler, Leonard and Judkewitz, Benjamin} } @article {70303, title = {A dual-color miniature endoscope for calcium imaging in behaving mice}, journal = {bioRxiv}, year = {2026}, month = {2026 Feb 20}, abstract = {

Calcium imaging with miniature endoscopes has become an essential tool in neuroscience, but conventional miniscopes typically record signals from only a single calcium indicator. Here, we present a dual-color miniature endoscope (miniscope) that enables simultaneous calcium imaging from two neuronal populations using spectrally distinct genetically encoded indicators. In freely moving mice, we used this system to record activity from striatal neurons of the direct (dSPN) and indirect (iSPN) pathways. We showed that dSPNs were activated earlier than iSPNs during contraversive movements, with dSPNs preferentially active during acceleration and iSPNs during deceleration. During ipsiversive turns, however, this temporal relationship was reversed. These findings indicate that dSPNs and iSPNs are not concurrently active, but instead exhibit complementary, direction-dependent dynamics that govern movement velocity. Our dual-color miniscope provides a compact, cost-effective platform for simultaneous two-population imaging, offering new opportunities to dissect coordinated activity across neural circuits in freely behaving animals.

}, doi = {10.64898/2026.02.19.706914}, url = {https://www.biorxiv.org/content/early/2026/02/20/2026.02.19.706914}, author = {Zhang, Jinyong and Hong, Feiyang and Kim, Jiwon and Bakhurin, Konstantin and Kim, Namsoo and Yin, Henry H.} } @article {70330, title = {Evolutionary basis of male same-sex sexual behavior by multiple pheromone switches in Drosophila.}, journal = {Curr Biol}, year = {2026}, month = {2026 Mar 20}, abstract = {

Male same-sex sexual behavior (SSB) is widespread among animal species, but its proximate (mechanistic) and ultimate (evolutionary) explanations remain unclear. A prevailing view is that SSB reflects impaired sex recognition, especially in insects. By unbiased behavioral screening, we identified a Drosophila species, D. santomea, in which males seldom attack and spontaneously court males vigorously, in addition to females. Behavioral, chemical, and optogenetic neuronal manipulations indicate that D. santomea males can distinguish conspecific sex and retain functional aggression circuitry. Instead, male SSB reflects three evolved pheromonal changes affecting two separate signaling systems, resulting in both reduced pheromone production and behavioral valence reversal. One of these occurs unexpectedly in females and may have evolved to prevent hybridization with an interfertile, geographically overlapping sibling species. Remarkably, male SSB and similar pheromonal changes also selectively co-occur in D. persimilis, a geographically and phylogenetically distant species and member of another sympatric sibling pair, implying evolutionary convergence in the two young taxa. The results identify a pheromonal mechanism for rapid social evolution in Drosophila and suggest a plausible evolutionary origin for male SSB as arising in concert with female adaptations that ensure reproductive isolation during speciation.

}, issn = {1879-0445}, doi = {10.1016/j.cub.2026.02.046}, url = {https://doi.org/10.1016/j.cub.2026.02.046}, author = {Ouadah, Youcef and Naragon, Thomas H and Smihula, Hayley and Behrman, Emily L and Khallaf, Mohammed A and Ding, Yun and Stern, David L and Parker, Joseph and Anderson, David J} } @article {70286, title = {Extracting large-scale neural activity with Suite2p}, journal = {bioRxiv}, year = {2026}, month = {2026 Feb 06}, abstract = {

Neural recordings using optical methods have improved dramatically. For example, we demonstrate here recordings of over 100,000 neurons from the mouse cortex obtained with a standard commercial microscope. To process such large datasets, we developed Suite2p, a collection of efficient algorithms for motion correction, cell detection, activity extraction and quality control. We also developed new approaches to benchmark performance on these tasks. Our GPU-accelerated non-rigid motion correction substantially outperforms alternative methods, while running over five times faster. For cell detection, Suite2p outperforms the CNMF algorithm in Caiman and Fiola, finding more cells and producing fewer false positives, while running in a fraction of the time. We also introduce quality control steps for users to evaluate performance on their own data, while offering alternative algorithms for specialized types of recordings such as those from one-photon and voltage imaging.

}, doi = {10.64898/2026.02.04.703741}, url = {https://www.biorxiv.org/content/early/2026/02/06/2026.02.04.703741}, author = {Stringer, Carsen and Ki, Chris and DelGrosso, Nicholas and LaFosse, Paul and Zhang, Qingqing and Pachitariu, Marius} } @article {70093, title = {iGABASnFR2 is an improved genetically encoded protein sensor of GABA}, journal = {eLife}, year = {2026}, month = {2026 Mar 18}, abstract = {

Monitoring GABAergic inhibition in the nervous system has been enabled by development of an intensiometric molecular sensor that directly detects GABA. However, the first generation iGABASnFR exhibits low signal-to-noise and suboptimal kinetics, making in vivo experiments challenging. To improve sensor performance, we targeted several sites in the protein for near-saturation mutagenesis and evaluated the resulting sensor variants in a high throughput screening system using evoked synaptic release in primary cultured neurons. This identified a sensor variant, iGABASnFR2, with 4.2-fold improved sensitivity and 20\% faster kinetics, and binding affinity that remained in a range sensitive to changes in GABA concentration at synapses. We also identified sensors with an inverted response, decreasing fluorescence intensity upon GABA binding. We termed the best such negative-going sensor iGABASnFR2n, which can be used to corroborate observations with the positive-going sensor. These improvements yielded a qualitative enhancement of in vivo performance when compared directly to the original sensor. iGABASnFR2 enabled the first measurements of direction-selective GABA release in the retina. In vivo imaging in somatosensory cortex revealed that iGABASnFR2 can report volume-transmitted GABA release following whisker stimulation. Overall, the improved sensitivity and kinetics of iGABASnFR2 make it a more effective tool for imaging GABAergic transmission in intact neural circuits.

}, doi = {10.7554/eLife.108319.3}, url = {https://doi.org/10.7554/eLife.108319.3}, author = {Kolb, Ilya and Hasseman, Jeremy P and Matsumoto, Akihiro and Jensen, Thomas P and Kopach, Olga and Arthur, Benjamin J and Zhang, Yan and Tsang, Arthur and Reep, Daniel and Tsegaye, Getahun and Zheng, Jihong and Patel, Ronak H and Looger, Loren L and Marvin, Jonathan S and Korff, Wyatt L and Rusakov, Dmitri A and Yonehara, Keisuke and Turner, Glenn C} } @article {70241, title = {Machine learning prediction of analyte-induced fluorescence perturbations in DNA-functionalized carbon nanotubes}, journal = {Nano Letters}, year = {2026}, month = {2026 Jan 07}, abstract = {

Single-walled carbon nanotubes (SWCNTs) functionalized with single-stranded DNAs can function as near-infrared nanosensors for molecular analytes. However, predicting which analytes elicit strong optical responses for specific nanosensors remains challenging. We developed machine learning (ML) models to predict analyte-induced fluorescence changes in a DNA{\textendash}SWCNT dopamine nanosensor. Using a data set of 63 small molecules sampling chemical space around dopamine, we encoded analytes with RDKit fingerprints, with or without HOMO and LUMO energies, and applied principal component analysis to identify structural motifs associated with optical response strength. We trained support vector regression and classification models using two strategies: ensembles of 200 models and cross-validation. Regression models achieved mean\ R2\ values of 0.2{\textendash}0.4, with cross-validation outperforming ensembles, while classifiers reached mean F1 scores of \~{}0.8. Cross-validation performed best for predictions on a blind set of 21 molecules. These findings show that ML can capture structure{\textendash}response patterns in modest data sets and guide in silico DNA{\textendash}SWCNT nanosensor design.

}, doi = {10.1021/acs.nanolett.5c05206}, url = {https://doi.org/10.1021/acs.nanolett.5c05206}, author = {Chakraborty, Sayantani and Krasley, Andrew T. and Smith, Colby H. and Beyene, Abraham G. and Vukovi{\'c}, Lela} } @article {70293, title = {Machine learning-guided spatial omics for tissue-scale discovery of cell-type-specific architectures}, journal = {bioRxiv}, year = {2026}, month = {2026 Feb 13}, abstract = {

Multiplexed protein imaging enables spatially resolved analysis of molecular organization in tissues, but existing spatial proteomics platforms remain constrained in scalability, throughput, and integration with RNA measurements and interpretable computational analysis. Here, we present an integrated spatial omics framework that combines highly multiplexed protein and RNA imaging with explainable machine learning to map cell-type-specific molecular and structural architectures at tissue scale. Using this platform, we simultaneously profiled up to 46 proteins and 79 RNA species across \~370,000 cells in intact mouse brain tissue at diffraction-limited subcellular resolution (\~260 nm). We developed a scalable, open-source computational pipeline for large-scale image processing and analysis, and show that nuclear protein and chromatin features alone are sufficient to accurately classify brain cell types and their spatial organization. Incorporation of explainable deep learning further enabled identification of human-interpretable, cell-type-specific subnuclear structural features directly from imaging data, with independent quantitative validation. Together, this integrated experimental and computational framework enables tissue-scale spatial proteomics-based cell-type classification and structural feature discovery, providing a broadly applicable platform for mechanistic studies, high-content screening, and translational applications.

}, doi = {10.64898/2026.02.12.705598}, url = {https://www.biorxiv.org/content/early/2026/02/13/2026.02.12.705598}, author = {Lian, Yumin and Adjavon, Diane-Yayra and Kawase, Takashi and Kim, Jun and Fleishman, Greg and Preibisch, Stephan and Funke, Jan and Liu, Zhe J.} } @article {70266, title = {Mechanically-induced Septin Networks protect nuclear integrity}, journal = {bioRxiv}, year = {2026}, month = {2026 Jan 21}, abstract = {

The cytoskeleton is a key mediator of mechanical interactions in cells, but specific contributions of septins remains unclear. Septins preferentially localize with a subset of actin stress fibers positioned under the nucleus, where they are situated between the membrane and stress fibers. Removing the nucleus from the cell results in the loss of these subnuclear septin-decorated stress fibers. Surprisingly, however, their formation can be rescued using a large glass bead in place of the nucleus. Similarly, applying a compressive force to the cell via confinement, whether externally or through internally generated actomyosin forces, results in increased septin accumulation in regions where the nucleus engages the cell cortex. Finally, loss of septin filaments via knockdown of SEPT7 increases the likelihood of nuclear membrane rupture during confinement. Together these data suggest that septins act as a dynamic mechanosensitive protective mechanism to buffer mechanical forces on the nucleus.

}, doi = {10.64898/2026.01.20.700414}, url = {https://www.biorxiv.org/content/early/2026/01/21/2026.01.20.700414}, author = {Utgaard, Margaret E. and Caillier, Alexia and Chandrasekar, Shreya and Tidei, Joseph J. and Balasubramanian, Harikrushnan and Lee, Rachel M. and Puls, Owen and Khuon, Satya and Aaron, Jesse and Chew, Teng-Leong and Beach, Jordan R. and Oakes, Patrick W.} } @article {70333, title = {Midgestation metabolic constraint in purine metabolism drives distinct strategies for placenta and fetal growth}, journal = {bioRxiv}, year = {2026}, month = {2026 Mar 18}, abstract = {

Purine nucleotides are essential for mammalian development1,2. Purine monophosphates support cell signaling and proliferation and are synthesized by cells through either de novo synthesis or a salvage pathway3. We previously identified a midgestational metabolic transition in mice (gestational days gd10.5{\textendash}11.5) characterized by changes in purine metabolism4. Midgestation is a period of rapid growth for placenta and embryo, yet it remains unclear how the placental tissues expand without directly competing with the embryo for biosynthetic resources. Here, we show that this midgestational metabolic transition is associated with a marked reduction in embryonic expression of purine salvage enzymes, which constrains embryonic metabolism and leads to different strategies for purine synthesis between the placenta and embryo. Midgestation embryos are unable to engage the purine salvage pathway even when de novo purine synthesis is blocked either in vivo or in ex utero embryo culture, whereas placental tissue and trophoblasts retain the capacity to use either pathway. Disruption of de novo purine synthesis in mice causes reduced embryonic growth, impaired axial elongation, and abnormal brain and placental development, which are only partially rescued by supplementation with purine salvage precursors. In human placenta, trophoblast stem cells readily switch between the de novo and salvage pathways based on nutrient availability, and syncytiotrophoblasts (STB) preferentially rely on the salvage pathway. We identified guanosine monophosphate (GMP) as a metabolic checkpoint regulating STB differentiation, with insufficient GMP levels causing degradation of the small GTPase Rheb and failure of mTOR activation. Supplementation of purine salvage substrates restored GMP synthesis and STB differentiation in humans, but not mice. Further, in vivo measurements in humans revealed that maternal circulating hypoxanthine decreases during pregnancy and is further reduced in women with clinically small placentas, highlighting the role of hypoxanthine in supporting placental growth. These results uncover compartmentalized purine salvage between the embryo and placenta as a mechanism that limits competition for biosynthetic resources and enables coordinated growth during mammalian development.

}, doi = {10.64898/2026.03.18.712680}, url = {https://www.biorxiv.org/content/early/2026/03/18/2026.03.18.712680}, author = {Xu, Weizhi and De La Cruz, Nancy and Woods, Andrea and Lokshtanov, Dmitry and Gao, Shihong and Khan, Nawal and Wright, Sylvia and Florian-Rodriguez, Maria E. and McIntire, Donald D. and Duryea, Elaine L and Nelson, David B. and Spong, Catherine Y. and Herrera, Christina L. and Hanna, Jacob H. and Srivatsan, Sanjay and Aguilera-Castrejon, Alejandro and Solmonson, Ashley} } @article {70242, title = {mRNAbow: A versatile gene expression system for multiplexed fluorescent imaging using optimized in vitro transcribed mRNA}, journal = {bioRxiv}, year = {2026}, month = {2026 Jan 02}, abstract = {

Messenger RNA (mRNA) transfection enables rapid, transient protein expression without nuclear entry, providing a powerful alternative to DNA or viral delivery in post-mitotic and otherwise difficult-to-transfect cells. Although in vitro transcribed (IVT) mRNAs have revolutionized therapeutic applications, their adoption in experimental biology remains limited by challenges in synthesis, variability across cell types, and concerns about cytotoxicity. Here, we define design principles that maximize IVT mRNA performance across diverse cellular and organismal systems. Through systematic comparison of capping strategies and base modifications, including N1-methyl-pseudouridine, 5-methylcytidine, and 5-methoxyuridine, we identify modifications that enhance translation while minimizing activation of cellular stress responses. Optimized transcripts drive robust protein expression within four hours, persist for up to one week, and support multiplexed expression of structurally and functionally distinct proteins in mammalian cells, including cancer cell lines, iPSC-derived systems, primary cells, and organoids, as well as in vivo in zebrafish embryos and in less genetically tractable models such as Danionella cerebrum and sea urchin embryos. To further expand accessibility for community use, we developed mRNAbow, a platform for generating low-toxicity mRNAs encoding organelle-targeted fluorescent proteins and biosensors for multiplex imaging, with corresponding plasmids made publicly available. Together, these advances establish a generalizable framework for IVT mRNA design and expand experimental access to synthetic mRNA technologies for dissecting cellular architecture and dynamics.

}, doi = {10.64898/2026.01.02.697412}, url = {https://www.biorxiv.org/content/early/2026/01/02/2026.01.02.697412}, author = {Choi, Heejun and Halanych, Coral and Kasberg, William and Testa, Michael D. and Rubin-Elgressy, Sarah and Nguyen, Phuong and Walpita, Deepika and Tsang, Arthur and Cortes, Dan and Song, Erin Yue and Wu, Haodi and Weissman, Irving L and Espinosa-Medina, Isabel and Satou, Chie and Song, Jia L. and Matus, David Q. and Lippincott-Schwartz, Jennifer} } @article {70297, title = {Nanoscale imaging of expanded cells and proteins with spontaneously blinking dyes}, journal = {bioRxiv}, year = {2026}, month = {2026 Feb 23}, abstract = {

Expansion microscopy (ExM) enables nanoscale imaging on standard microscopes, but combining ExM with single-molecule localization microscopy (SMLM) remains difficult, owing to the incompatibility of expanded hydrogels with photoswitching buffers. Here, we introduce a single-step expansion microscopy method that allows SMLM with spontaneously blinking dyes in 6-14{\texttimes} expanded samples, without re-embedding. We demonstrate nanometer-resolution imaging by resolving the organization of the nuclear pore complex (NPC) and the molecular structure of recombinant homotrimeric proliferating cell nuclear antigen (PCNA).

}, doi = {10.64898/2026.02.23.707413}, url = {https://www.biorxiv.org/content/early/2026/02/23/2026.02.23.707413}, author = {Taban, Danush and Jungblut, Marvin and Budiarta, Made and Helmerich, Dominic A. and Kiesel, Cerridwen and Plutkis, Sarah E. and Lavis, Luke D. and Krah, Donatus and Shaib, Ali H. and Doose, S{\"o}ren and Kollmannsberger, Philip and Rizzoli, Silvio O. and Beliu, Gerti and Sauer, Markus} } @article {70304, title = {Nanoscale imaging of expanded cells and proteins with spontaneously blinking dyes}, journal = {bioRxiv}, year = {2026}, month = {2026 Feb 23}, abstract = {

Expansion microscopy (ExM) enables nanoscale imaging on standard microscopes, but combining ExM with single-molecule localization microscopy (SMLM) remains difficult, owing to the incompatibility of expanded hydrogels with photoswitching buffers. Here, we introduce a single-step expansion microscopy method that allows SMLM with spontaneously blinking dyes in 6-14{\texttimes} expanded samples, without re-embedding. We demonstrate nanometer-resolution imaging by resolving the organization of the nuclear pore complex (NPC) and the molecular structure of recombinant homotrimeric proliferating cell nuclear antigen (PCNA).

}, doi = {10.64898/2026.02.23.707413}, url = {https://www.biorxiv.org/content/early/2026/02/23/2026.02.23.707413}, author = {Taban, Danush and Jungblut, Marvin and Budiarta, Made and Helmerich, Dominic A. and Kiesel, Cerridwen and Plutkis, Sarah E. and Lavis, Luke D. and Krah, Donatus and Shaib, Ali H. and Doose, S{\"o}ren and Kollmannsberger, Philip and Rizzoli, Silvio O. and Beliu, Gerti and Sauer, Markus} } @article {70250, title = {Oxytocin modulation of spinal circuits drives therapeutic benefits of massage}, journal = {bioRxiv}, year = {2026}, month = {2026 Jan 12}, abstract = {

Across social species, social touch enhances well-being and reduces pain {\textemdash} two seemingly distinct benefits that enhance survival. Yet where and how the nervous system integrates these functions, and whether a single mechanism could serve both, remains unknown. Here we show that massage triggers oxytocin release, which shapes both pain and touch reward at the earliest stage of central processing {\textemdash} the spinal cord {\textemdash} through a single, state-dependent circuit mechanism. We report that in humans, massage enhances well-being, effects that correlate with endogenous oxytocin release. In mice, gentle touch activates hypothalamic oxytocin neurons that project directly to the spinal dorsal horn. Genetic manipulation of spinal oxytocin circuits alters behavioral responses to both gentle touch and noxious stimuli. Spinal calcium imaging and slice electrophysiology reveal that oxytocin acts on both excitatory and inhibitory spinal neurons to sculpt the relative activity of spinal ascending systems that convey both social touch and pain to the brain. Extending these findings to humans, we show that oxytocin receptors are also expressed on spinal excitatory and inhibitory neurons, and that endogenous oxytocin during massage correlates with altered spinal touch processing. Thus, spinal oxytocin signaling provides an evolutionarily conserved mechanism for the therapeutic benefits of massage.

}, doi = {10.64898/2026.01.11.698886}, url = {https://www.biorxiv.org/content/early/2026/01/12/2026.01.11.698886}, author = {Bohic, Manon and Salamone, Paula Celeste and Zuo, Wanhong and Negm, Ahmed and Fulton, Sasha Leilani and Du, Shibin and Jayakumar, Swathi and Keating, Jessica and Soubeyre, Vanessa and Gradwell, Mark Andrew and Upadhyay, Aman and Shorter, Lucero and Kim, Joel and Inoue, Yukiko Ueno and Inoue, Takayoshi and Mensch, Brett and Ye, Jiang-Hong and Peirs, Cedric and Poulen, Gaetan and Lonjon, Nicolas and Vachiery-Lahaye, Florence and Bauchet, Luc and Bourinet, Emmanuel and Olausson, Hakan and Abdus-Saboor, Ishmail and Tao, Yuan-Xiang and Boehme, Rebecca and Abraira, Victoria E.} } @article {70294, title = {Phasor analysis of RGB camera data enables fluorescence microscopy unmixing and brightfield segmentation in a commercial microscope}, journal = {bioRxiv}, year = {2026}, month = {2026 Feb 16}, abstract = {

Spectral information plays a crucial role in biological imaging, yet conventional epifluorescence and histological techniques often rely on RGB image acquisition, limiting the resolution of spectrally overlapping components. Here, we present a phasor-based spectral analysis framework adapted for RGB images, enabling unsupervised segmentation and unmixing without the need for hyperspectral systems or sequential acquisition. By applying a discrete Fourier transform to the red, green, and blue intensities at each pixel, we generate a two-dimensional phasor plot where spectral relationships are encoded in modulation and phase. We demonstrate the utility of this approach across three distinct applications: segmentation of lung histology images stained with hematoxylin and eosin to quantify alveolar collapse, analysis of autofluorescence in skin lesions (nevi and melanoma) to highlight pathological spectral signatures, and spectral unmixing in multicolor-labeled U2OS cells to resolve overlapping fluorophores. Our method improves signal separation, reduces noise, and enhances biological interpretability using standard RGB acquisition. These findings establish RGB phasor analysis as a practical and powerful tool for spectral decomposition and segmentation in microscopy, bridging the gap between conventional imaging and advanced spectral analysis.

}, doi = {10.64898/2026.02.13.705652}, url = {https://www.biorxiv.org/content/early/2026/02/16/2026.02.13.705652}, author = {Schuty, Bruno and Garcia, Maria Jose and Khuon, Satya and Malacrida, Leonel S} } @article {70257, title = {Redefining colocalization analysis with a novel phasor mixing coefficient}, journal = {Journal of Cell Science}, volume = {139}, year = {2026}, month = {2026 Jan 19}, pages = {jcs264388}, abstract = {

The first step to probing any potential interaction between two biomolecules is to determine their spatial association. In other words, if two biomolecules localize similarly within a cell, then it is plausible they could interact. Traditionally, this is quantified through various colocalization metrics. These measures infer this association by estimating the degree to which fluorescent signals from each biomolecule overlap or correlate. However, these metrics are, at best, proxies, and they depend strongly on various experimental choices. Here, we define a new strategy that leverages multispectral imaging and phasor analysis, termed the phasor mixing coefficient (PMC). The PMC measures the precise mixing of fluorescent signals in each pixel. We demonstrate how the PMC captures complex biological subtlety by offering two distinct values, a global measure of overall color mixing and the homogeneity thereof. We additionally show that the PMC exhibits less sensitivity to signal-to-noise ratio, intensity threshold and background signal compared to canonical methods. Moreover, this method provides a means to visualize color mixing at each pixel. We show that the PMC offers users a nuanced and robust metric to quantify biological association.

}, issn = {0021-9533}, doi = {10.1242/jcs.264388}, url = {https://doi.org/10.1242/jcs.264388}, author = {Puls, Owen F. and Aaron, Jesse S. and Quarles, Ellen K. and Khuon, Satya and Eisenman, Leanna R. and Kamaid, Andres and Malacrida, Leonel and Chew, Teng-Leong} } @article {70251, title = {SCREWx: A Screwless, Chronic, Recoverable, and Lightweight Neuropixels fixture for freely-moving rodents}, journal = {bioRxiv}, year = {2026}, month = {2026 Jan 06}, abstract = {

High-density Neuropixels probes enable the study of large neural populations with single-cell and sub-millisecond resolution. While single-probe and acute head-fixed experiments have yielded critical scientific insights, understanding the neural mechanisms underlying many complex behaviors requires simultaneous multi-region recordings in freely moving, chronically implanted animals. Various probe fixtures have been developed to enable high-density recording, but existing designs impose critical limitations: their substantial weight restricts the maximum probe count that smaller animals can support, their bulky dimensions constrain the proximity of targeted brain regions, and their complex assembly risks damaging the probe during insertion and recovery. In this paper, we present a lightweight, fully 3D-printable, compact, and screwless fixture for chronic Neuropixels implants in freely moving rodents that features simple mechanisms for stable implantation and safe extraction. Our fixture design enables stable, high-yield single-unit recordings for months-long experiments, along with an 83\% successful probe extraction rate. This fixture design provides a robust and accessible solution for long-term, multi-probe chronic Neuropixels recordings, increasing experimental throughput and enabling more complex experimental designs to investigate brain-wide neural dynamics.

}, doi = {10.64898/2026.01.06.697790}, url = {https://www.biorxiv.org/content/early/2026/01/06/2026.01.06.697790}, author = {Cheng, Alexandra and DeWeese, Tate and Zhou, Yueqing and Sueoka, Yotaro and Koukuntla, Sai and Green, Merrill and Cullen, Kathleen and Knierim, James and Graves, Austin and Harris, Timothy} } @article {70277, title = {Stem cell control and cancer initiation by an autocrine, injury-activated Igf complex}, journal = {bioRxiv}, year = {2026}, month = {2026 Feb 04}, abstract = {

Stem cells rapidly proliferate after injury to repair damaged tissue, and chronic injury predisposes to cancer. However, injury-activated mitogens, the mechanisms that keep them inactive until injury, and their role in cancer are not understood. Here we identify Igf2 as the injury-activated mitogen for neuroendocrine stem cells, a facultative airway stem cell and origin of small cell lung cancer. Igf2 is constitutively produced by the stem cells but sequestered in inactive form by co-expressed Igf binding proteins. Injury releases Igf2 and induces proliferation by activating its receptors and repressing Rb tumor suppressor, which normally enforces stem cell quiescence. Persistent pathway activation initiates oncogenesis. Thus, in addition to its classical hormonal roles in physiology, growth, and aging, Igf operates locally with Igf binding proteins and Rb to control injury-induced stem cell activation and cancer. This pathway may also control related stem cells and cancers of the body and brain.

}, doi = {10.64898/2026.02.02.703150}, url = {https://www.biorxiv.org/content/early/2026/02/04/2026.02.02.703150}, author = {Zhang, Yue and Ouadah, Youcef and Liu, Yin and Kumar, Maya and Morck, Makenna and Krasnow, Mark A} } @article {70329, title = {Unstructured transcription factor interactions enable emergent specificity.}, journal = {Science}, year = {2026}, month = {2026 Mar 19}, pages = {eaeb6487}, abstract = {

How intrinsically disordered regions (IDRs) influence chromatin binding and nuclear organization of transcription factors (TFs) remains unclear. We employed proximity-assisted photoactivation (PAPA), a single-molecule protein-protein interaction sensor, to investigate how IDRs might influence TF interactions with each other and with chromatin in live cells. We found that the Sp1 DNA binding domain (DBD) interacted poorly with chromatin and did not colocalize with Sp1. Weak interaction of the isolated IDR with full-length Sp1 was enhanced by fusion to various unrelated DBDs. Live imaging of polytene chromosomes confirmed that an IDR could confer sharp locus specificity on an otherwise nonspecific DBD. These findings suggest that TF specificity emerges on chromatin when ensembles of diverse, unstructured interactions are scaffolded by transient DNA contacts.

}, issn = {1095-9203}, doi = {10.1126/science.aeb6487}, url = {https://doi.org/10.1126/science.aeb6487}, author = {Abidi, Abrar A and Cattoglio, Claudia and Tang, Natalie N and Fan, Vinson B and Dailey, Gina M and Hay, Amir D and Kunamaneni, Prasanthi and Milkie, Daniel E and Darzacq, Xavier and Betzig, Eric and Tjian, Robert and Graham, Thomas G W} } @article {70285, title = {Whole-brain co-mapping of gene expression and neuronal activity at cellular resolution in behaving zebrafish}, journal = {bioRxiv}, year = {2026}, month = {2026 Feb 10}, abstract = {

The brain{\textquoteright}s capabilities rely on both the molecular properties of individual cells and their interactions across brain-wide networks. However, relating gene expression to activity in individual neurons across the entire brain remains elusive. Here we developed an experimental-computational platform, WARP, for whole-brain imaging of neuronal activity during behavior, expansion-assisted spatial transcriptomics, and cellular-level registration of these two modalities. Through joint analysis of whole-brain neuronal activity during multiple behaviors, cellular gene expression, and anatomy, we identified functions of molecularly defined populations {\textemdash} including luminance coding in a cckb-pou4f2 midbrain population and task-structured activity in pvalb7-eomesa hippocampal-like neurons {\textemdash} and defined over 2,000 other function-gene-anatomy subpopulations. Analysis of this unprecedented multimodal dataset also revealed that most gene-matched neurons showed stronger activity correlations, highlighting a brain-wide role for gene expression in functional organization. WARP establishes a foundational platform and open-access dataset for cross-experiment discovery, high-throughput function-to-gene mapping, unification of cell biology and systems neuroscience, and scalable circuit modeling at the whole-brain scale.

}, doi = {10.64898/2026.02.07.704095}, url = {https://www.biorxiv.org/content/early/2026/02/10/2026.02.07.704095}, author = {Marquez Legorreta, Emmanuel and Fleishman, Greg M. and Hesselink, Luuk W. and Eddison, Mark and Smeets, Kasper and Stringer, Carsen and Keller, Philipp J. and Narayan, Sujatha and Chen, Alex B. and Mensh, Brett D. and Sternson, Scott M. and Englitz, Bernhard and Tillberg, Paul W. and Ahrens, Misha B.} } @article {69560, title = {Activity-dependent synapse elimination requires caspase-3 activation}, journal = {eLife}, year = {2025}, month = {2025 Jul 10}, abstract = {

During brain development, synapses are initially formed in excess and are later eliminated in an activity-dependent manner, with weak synapses being preferentially removed. Previous studies identified glia as mediators of synapse removal, but it is unclear how glia specifically target weak synapses. Here we show that, in the developing mouse visual pathway, inhibiting synaptic transmission induces postsynaptic activation of caspase-3. Caspase-3 is essential for synapse elimination driven by both spontaneous and experience-dependent neural activity. Synapse weakening-induced caspase-3 activation determines the specificity of synapse elimination mediated by microglia but not astrocytes. Furthermore, in a mouse model of Alzheimer{\textquoteright}s disease, caspase-3 deficiency protects against synapse loss induced by amyloid-β deposition. Our results reveal caspase-3 activation as a key step in activity-dependent synapse elimination during development and synapse loss in neurodegeneration.

bioRxiv preprint:\ https://doi.org/10.1101/2024.08.02.606316

}, doi = {10.7554/eLife.101779.2}, url = {https://doi.org/10.7554/eLife.101779.2}, author = {Yu, Zhou and Gutu, Andrian and Kim, Namsoo and O{\textquoteright}Shea, Erin K.} } @article {70129, title = {Alternating angle milling suppresses streaking artifacts in FIB-SEM imaging}, journal = {bioRxiv}, year = {2025}, month = {2025 Sep 26}, abstract = {

Focused ion beam scanning electron microscopy (FIB-SEM) (1{\textendash}3) has been used in life sciences to produce large volumetric datasets with high resolution information on ultrastructure of biological organisms. 3D image acquisition is accomplished by serial removal of thin layers of material using focused ion beam (FIB) milling followed by scanning electron microscopy (SEM) imaging.One of the challenges in the standard FIB-SEM imaging protocol is that FIB milling results in characteristic artifacts, known as {\textquotedblleft}streaks{\textquotedblright} or {\textquotedblleft}curtains{\textquotedblright}. These streaks are caused by non-uniform material removal forming long straight trenches parallel to the FIB milling direction. These artifacts get worse along the milling direction and ultimately limit size of the SEM field of view.Various methods have been proposed to mitigate the streaks in acquired images. While these techniques often provide noticeable visual improvement, the underlying problem remains. The structural information in the {\textquotedblleft}streaked{\textquotedblright} areas is lost due to non-uniform material removal during milling and cannot be fully recovered.We propose a simple modification allowing for a significant reduction of milling non-uniformities of streaks. We demonstrate the effectiveness of this approach on various samples.

}, doi = {10.1101/2025.09.25.678572}, url = {https://www.biorxiv.org/content/early/2025/09/26/2025.09.25.678572}, author = {Shtengel, Gleb and Qiu, Wei and Aaron, Jesse and Crowe, Arthur S. and Polilov, Alexey A. and Karkali, Katerina and Bleck, Christopher K.E. and Hess, Harald H.} } @article {69744, title = {Bio-inspired 3D-printed phantom: Encoding cellular heterogeneity for characterization of quantitative phase imaging}, journal = {Measurement}, volume = {247}, year = {2025}, month = {2025 Apr 15}, pages = {116765}, abstract = {

Quantitative phase imaging (QPI) has proven to be a valuable tool for advanced biological and pharmacological research, providing phase information for the study of cell features and physiology in label-free conditions. The next step for QPI to become a gold standard is the quantitative assessment of the phase gradients over the different microscopy setups. Given the large variety of QPI systems, a systematic comparison is a challenging task, and requires a calibration target representative of the living samples. In this paper, we introduce a tailor-made 3D-printed phantom derived from phase images of eukaryotic cells. It comprises typical morphologies and optical thicknesses found in biological cultures and is characterized with digital holographic microscopy (reference measurements). The performance of three different full field QPI optical systems, in terms of optical path difference and dry mass accuracy, were evaluated. This phantom opens up other possibilities for the validation of reconstruction algorithms and post-processing routines, and paves the way for calibration targets designed ad hoc for specific biological questions.

}, keywords = {Cell phantom, Inter-laboratory comparison, Metrology, Phase calibration target, Quantitative phase imaging, Two-photon polymerization lithography}, issn = {0263-2241}, doi = {10.1016/j.measurement.2025.116765}, url = {https://www.sciencedirect.com/science/article/pii/S0263224125001241}, author = {Sylvia Desissaire and Micha{\l} Ziemczonok and Tigrane Cantat-Moltrecht and Arkadiusz Ku{\'s} and Guillaume Godefroy and Lionel Herv{\'e} and Chiara Paviolo and Wojciech Krauze and C{\'e}dric Allier and Ondrej Mandula and Ma{\l}gorzata Kujawi{\'n}ska} } @article {70045, title = {Cdc42 regulates apical membrane fusion via the Rab11a{\textendash}VAMP2 pathway in salivary gland acinar cells}, journal = {bioRxiv}, year = {2025}, month = {2025 Jul 31}, abstract = {

Epithelial polarity is essential for proper tissue organization and function, yet the molecular mechanisms governing apical membrane formation during secretory epithelial development remain incompletely understood. Here, we investigate the role of the small GTPase Cdc42 in salivary gland acinar cell development using a mouse model designed to knock out Cdc42 specifically at the onset of acinar cell formation. Loss of Cdc42 resulted in defective apical membrane formation accompanied by accumulation of vesicles around the apical lumen. These vesicles contained the apical water channel AQP5 and the apical recycling endosome (ARE) marker Rab11a, while the basolateral transporter NKCC1 retained normal localization, indicating an apical-selective trafficking defect. Importantly, Cdc42 deficiency caused a selective 40\% reduction in the expression of the SNARE protein VAMP2, while other vesicle trafficking proteins including VAMP8, SNAP23, and EEA1 remained unchanged. Our findings reveal that Cdc42 controls apical membrane formation by maintaining VAMP2 expression, which is essential for the fusion of Rab11a-positive recycling endosomes. The accumulation of fusion-incompetent AREs near the apical surface demonstrates the critical role of the Cdc42-VAMP2 pathway in epithelial development. These results provide new insights into how polarity regulators integrate vesicle trafficking and fusion machinery, and may have implications for understanding glandular diseases involving epithelial polarity defects.

}, doi = {10.1101/2025.07.31.667845}, url = {https://www.biorxiv.org/content/early/2025/08/01/2025.07.31.667845}, author = {Shitara, Akiko and Nagase, Haruna and Bleck, Christopher K. E. and Ohno, Yuta and Kataoka, Hideo and Satoh, Keitaro and Kashimata, Masanori} } @article {70185, title = {The CENP-A chaperone complex spatially organizes centromeres}, journal = {bioRxiv}, year = {2025}, month = {2026 Nov 09}, abstract = {

Centromeres, defined by CENP-A-containing nucleosomes, direct the assembly of kinetochores for spindle attachment. In mitosis, CENP-A and the constitutive centromere-associated network (CCAN) of the inner kinetochore are arranged into bipartite subdomains within clearings of chromatin. However, it remains unclear whether any of these features exist before mitosis. We show that in interphase, CENP-A and the CCAN assemble \~200-300 nm shell-like structures that enclose a chromatin-poor central cavity. Strikingly, this cavity is occupied by the interphase-specific CENP-A chaperone complex, which promotes CENP-A assembly once per cell cycle. However, chaperone presence, but not CENP-A incorporation, is required to generate both the shell architecture and the chromatin clearing. The CCAN scaffold CENP-C, which links CENP-A nucleosomes to the chaperone complex, exhibits radial organization spanning the entire structure and is essential for its formation. These data uncover a previously unrecognized structural role for the CENP-A chaperone machinery in establishing interphase centromere architecture and suggest a mechanism by which this machinery configures centromeres for faithful kinetochore assembly and genome stability.

}, doi = {10.1101/2025.11.07.687291}, url = {https://www.biorxiv.org/content/early/2025/11/09/2025.11.07.687291}, author = {Gupta, Hindol and Haase, Julian and Wu, Yicong and Chen, Jiji and Karadimov, George and Shroff, Hari and Kelly, Alexander E.} } @article {69903, title = {Challenges of microscopy technology dissemination to resource-constrained communities.}, journal = {Nat Methods}, year = {2025}, month = {2025 May 06}, abstract = {

No abstract available.

}, issn = {1548-7105}, doi = {10.1038/s41592-025-02690-7}, url = {https://doi.org/10.1038/s41592-025-02690-7}, author = {Aaron, Jesse S and Jacobs, Caron A and Malacrida, Leonel and Keppler, Antje and French, Paul and Fletcher, Daniel A and Wood, Christopher and Brown, Claire M and Wright, Graham D and Ogawa, Satoshi and Maina, Mahmoud and Chew, Teng-Leong} } @article {70026, title = {CHLOK: a chemigenetic multicolor labeling system to visualize neuronal birthdate and circuit integration}, journal = {Research Square}, year = {2025}, month = {2025 Jul 08}, abstract = {

Understanding how neurons integrate into developing circuits and contribute to functional activity is essential for decoding brain development and plasticity. However, current methods to study neuronal integration often suffer from low throughput, limited spatiotemporal resolution, or invasive procedures that hinder in vivo functional analysis. To overcome these challenges, we present a birthdate-labeling strategy, named CHLOK, based on HaloTag technology and a broad palette of fluorescent synthetic dyes. This approach enables precise multicolor labeling of neurons according to their maturation stage and allows flexible integration into functional assays through compatibility with calcium imaging and optogenetics. We validated CHLOK by mapping birthdate-resolved neuronal activity in the developing visual and motor systems of zebrafish larvae. Our results reveal distinct functional contributions of early- versus late-born neurons, providing new insights into the temporal dynamics of circuit formation. Furthermore, we demonstrate the versatility of this approach, showcasing age-specific multicolor calcium and voltage imaging as well as optogenetic manipulation. By overcoming key limitations of existing techniques, CHLOK offers a powerful, versatile and non-invasive tool for studying neural integration, circuit development and function in vivo.

}, doi = {10.21203/rs.3.rs-7039578/v1}, url = {https://doi.org/10.21203/rs.3.rs-7039578/v1}, author = {Faini, Giulia and Tuffery, Matthieu and Saleem, Amna and Zhang, Lixia and Du, Felix and Le Bourdelles, Guillaume and Duroure, Karine and Schreiter, Eric R and Tanese, Dimitrii and Emiliani, Valentina and Del Bene, Filippo and Koyama, Minoru} } @article {69902, title = {Chromatix: A differentiable, GPU-accelerated wave-optics library}, journal = {bioRxiv}, year = {2025}, month = {2025 May 2}, abstract = {

Modern microscopy methods incorporate computational modeling of optical systems as an integral part of the imaging process, either to solve inverse problems or enable optimization of the optical system design. These methods often depend on differentiable simulations of optical systems, yet no standardized framework exists{\textemdash}forcing computational optics researchers to repeatedly and independently implement simulations that are prone to errors, difficult to reuse in other applications, and often computationally suboptimal. These common problems limit the potential impact of computational optics as a field. We present Chromatix: an open-source, GPU-accelerated differentiable wave optics library. Chromatix builds on JAX to enable fast simulation of diverse optical systems and inverse problem solving, scaling these simulations from single-CPU laptops to multi-GPU servers. The library implements various optical elements (e.g., lenses, polarizers and spatial light modulators) and multiple light propagation models (e.g., Fresnel approximation, angular spectrum and off-axis propagation) that can be flexibly combined to model various computational optics applications such as snapshot microscopy, holography, and phase retrieval of multiple scattering samples. These simulations can be automatically parallelized to scale across multiple GPUs with a single-line change to the modeling code, enabling simulation and optimization of previously impractical optical system designs. We demonstrate Chromatix{\textquoteright}s capacity to substantially accelerate optics simulation and optimization on existing methods in computational optics, speeding up optical simulation and optimization from 2-6{\texttimes} on a single GPU to up to 22{\texttimes} on 8 GPUs (depending on the particular system being modeled) compared to the original implementations. Chromatix establishes a standard for wave optics simulations, democratizing access to and expanding the design space of computational optics.i

}, doi = {10.1101/2025.04.29.651152}, url = {https://www.biorxiv.org/content/early/2025/05/02/2025.04.29.651152}, author = {Deb, Diptodip and Both, Gert-Jan and Bezzam, Eric and Kohli, Amit and Yang, Siqi and Chaware, Amey and Allier, C{\'e}dric and Cai, Changjia and Anderberg, Geneva and Eybposh, M. Hossein and Schneider, Magdalena C. and Heintzmann, Rainer and Rivera-Sanchez, Fabrizio A. and Simmerer, Corey and Meng, Guanghan and Tormes-Vaquerano, Jovan and Han, SeungYun and Shanmugavel, Sibi Chakravarthy and Maruvada, Teja and Yang, Xi and Kim, Yewon and Diederich, Benedict and Joo, Chulmin and Waller, Laura and Durr, Nicholas J. and P{\'e}gard, Nicolas C. and La Rivi{\`e}re, Patrick J. and Horstmeyer, Roarke and Chowdhury, Shwetadwip and Turaga, Srinivas C.} } @article {70165, title = {A Class of Markovian self-reinforcing processes with power-law distributions}, journal = {arXiv}, year = {2025}, month = {2025 Oct 21}, abstract = {

Solar flares, email exchanges, and many natural or social systems exhibit bursty dynamics, with periods of intense activity separated by long inactivity. These patterns often follow power- law distributions in inter-event intervals or event rates. Existing models typically capture only one of these features and rely on non-local memory, which complicates analysis and mechanistic interpretation. We introduce a novel self-reinforcing point process whose event rates are governed by local, Markovian nonlinear dynamics and post-event resets. The model generates power-law tails for both inter-event intervals and event rates over a broad range of exponents observed empirically across natural and human phenomena. Compared to non-local models such as Hawkes processes, our approach is mechanistically simpler, highly analytically tractable, and also easier to simulate. We provide methods for model fitting and validation, establishing this framework as a versatile foundation for the study of bursty phenomena.

}, doi = {10.48550/arXiv.2510.19034}, url = {https://arxiv.org/abs/2510.19034}, author = {Pavlo Bulanchuk and Sue Ann Koay and Sandro Romani} } @article {70130, title = {Combinatorial protein barcodes enable self-correcting neuron tracing with nanoscale molecular context}, journal = {bioRxiv}, year = {2025}, month = {2025 Sep 28}, abstract = {

Mapping nanoscale neuronal morphology with molecular annotations is critical for understanding healthy and dysfunctional brain circuits. Current methods are constrained by image segmentation errors and by sample defects (e.g., signal gaps, section loss). Genetic strategies promise to overcome these challenges by using easily distinguishable cell identity labels. However, multicolor approaches are spectrally limited in diversity, whereas nucleic acid barcoding lacks a cell-filling morphology signal for segmentation. Here, we introduce PRISM (Protein-barcode Reconstruction via Iterative Staining with Molecular annotations), a platform that integrates combinatorial delivery of antigenically distinct, cell-filling proteins with tissue expansion, multi-cycle imaging, barcode-augmented reconstruction, and molecular annotation. Protein barcodes increase label diversity by \>750-fold over multicolor labeling and enable morphology reconstruction with intrinsic error correction. We acquired a \~10 million μm3 volume of mouse hippocampal area CA2/3, multiplexed across 23 barcode antigen and synaptic marker channels. By combining barcodes with shape information we achieve an 8x increase in automatic tracing accuracy of genetically labelled neurons. We demonstrate PRISM supports automatic proofreading across micron-scale spatial gaps and reconnects neurites across discontinuities spanning hundreds of microns. Using PRISM{\textquoteright}s molecular annotation capability, we map the distribution of synapses onto traced neural morphology, characterizing challenging synaptic structures such as thorny excrescences (TEs), and discovering a size correlation among spatially proximal TEs on the same dendrite. PRISM thus supports self-correcting neuron reconstruction with molecular context.

}, doi = {10.1101/2025.09.26.678648}, url = {https://www.biorxiv.org/content/early/2025/09/28/2025.09.26.678648}, author = {Park, Sung Yun and Sheridan, Arlo and An, Bobae and Jarvis, Erin and Lyudchik, Julia and Patton, William and Axup, Jun Y. and Chan, Stephanie W. and Damstra, Hugo G.J. and Leible, Daniel and Leung, Kylie S. and Magno, Clarence A. and Meeran, Aashir and Michalska, Julia M. and Rieger, Franz and Wang, Claire and Wu, Michelle and Church, George M. and Funke, Jan and Huffman, Todd and Leeper, Kathleen G.C. and Truckenbrodt, Sven and Winnubst, Johan and Kornfeld, Joergen M.R. and Boyden, Edward S. and Rodriques, Samuel G. and Payne, Andrew C.} } @article {69794, title = {Combining sampling methods with attractor dynamics in spiking models of head-direction systems}, journal = {bioRxiv}, year = {2025}, month = {2025 Feb 26}, abstract = {

Uncertainty is a fundamental aspect of the natural environment, requiring the brain to infer and integrate noisy signals to guide behavior effectively. Sampling-based inference has been proposed as a mechanism for dealing with uncertainty, particularly in early sensory processing. However, it is unclear how to reconcile sampling-based methods with operational principles of higher-order brain areas, such as attractor dynamics of persistent neural representations. In this study, we present a spiking neural network model for the head-direction (HD) system that combines sampling-based inference with attractor dynamics. To achieve this, we derive the required spiking neural network dynamics and interactions to perform sampling from a large family of probability distributions - including variables encoded with Poisson noise. We then propose a method that allows the network to update its estimate of the current head direction by integrating angular velocity samples - derived from noisy inputs - with a pull towards a circular manifold, thereby maintaining consistent attractor dynamics. This model makes specific, testable predictions about the HD system that can be examined in future neurophysiological experiments: it predicts correlated subthreshold voltage fluctuations; distinctive short- and long-term firing correlations among neurons; and characteristic statistics of the movement of the neural activity "bump" representing the head direction. Overall, our approach extends previous theories on probabilistic sampling with spiking neurons, offers a novel perspective on the computations responsible for orientation and navigation, and supports the hypothesis that sampling-based methods can be combined with attractor dynamics to provide a viable framework for studying neural dynamics across the brain.

}, doi = {10.1101/2025.02.25.640158}, url = {https://www.biorxiv.org/content/early/2025/02/26/2025.02.25.640158}, author = {Pjanovic, Vojko and Zavatone-Veth, Jacob A and Masset, Paul and Keemink, Sander W and Nardin, Michele} } @article {69854, title = {Combining spatial transcriptomics and ECM imaging in 3D for mapping cellular interactions in the tumor microenvironment.}, journal = {Cell Syst}, year = {2025}, month = {2025 Apr 09}, pages = {101261}, abstract = {

Tumors are complex ecosystems composed of malignant and non-malignant cells embedded in a dynamic extracellular matrix (ECM). In the tumor microenvironment, molecular phenotypes are controlled by cell-cell and ECM interactions in 3D cellular neighborhoods (CNs). While their inhibition can impede tumor progression, routine molecular tumor profiling fails to capture cellular interactions. Single-cell spatial transcriptomics (ST) maps receptor-ligand interactions but usually remains limited to 2D tissue sections and lacks ECM readouts. Here, we integrate 3D ST with ECM imaging in serial sections from one clinical lung carcinoma to systematically quantify molecular states, cell-cell interactions, and ECM remodeling in CN. Our integrative analysis pinpointed known immune escape and tumor invasion mechanisms, revealing several druggable drivers of tumor progression in the patient under study. This proof-of-principle study highlights the potential of in-depth CN profiling in routine clinical samples to inform microenvironment-directed therapies. A record of this paper{\textquoteright}s transparent peer review process is included in the supplemental information.

}, issn = {2405-4720}, doi = {10.1016/j.cels.2025.101261}, url = {https://doi.org10.1016/j.cels.2025.101261}, author = {Pentimalli, Tancredi Massimo and Schallenberg, Simon and Le{\'o}n-Peri{\~n}{\'a}n, Daniel and Legnini, Ivano and Theurillat, Ilan and Thomas, Gwendolin and Boltengagen, Anastasiya and Fritzsche, Sonja and Nimo, Jose and Ruff, Lukas and Dernbach, Gabriel and Jurmeister, Philipp and Murphy, Sarah and Gregory, Mark T and Liang, Yan and Cordenonsi, Michelangelo and Piccolo, Stefano and Coscia, Fabian and Woehler, Andrew and Karaiskos, Nikos and Klauschen, Frederick and Rajewsky, Nikolaus} } @article {69881, title = {Comparative connectomics of Drosophila descending and ascending neurons.}, journal = {Nature}, year = {2025}, month = {2025 Apr 30}, abstract = {

In most complex nervous systems there is a clear anatomical separation between the nerve cord, which contains most of the final motor outputs necessary for behaviour, and the brain. In insects, the neck connective is both a physical and an information bottleneck connecting the brain and the ventral nerve cord (an analogue of the spinal cord) and comprises diverse populations of descending neurons (DNs), ascending neurons (ANs) and sensory ascending neurons, which are crucial for sensorimotor signalling and control. Here, by integrating three separate electron microscopy (EM) datasets, we provide a complete connectomic description of the ANs and DNs of the Drosophila female nervous system and compare them with neurons of the male nerve cord. Proofread neuronal reconstructions are matched across hemispheres, datasets and sexes. Crucially, we also match 51\% of DN cell types to light-level data defining specific driver lines, as well as classifying all ascending populations. We use these results to reveal the anatomical and circuit logic of neck connective neurons. We observe connected chains of DNs and ANs spanning the neck, which may subserve motor sequences. We provide a complete description of sexually dimorphic DN and AN populations, with detailed analyses of selected circuits for reproductive behaviours, including male courtship (DNa12; also known as aSP22) and song production (AN neurons from hemilineage 08B) and female ovipositor extrusion (DNp13). Our work provides EM-level circuit analyses that span the entire central nervous system of an adult animal.

}, issn = {1476-4687}, doi = {10.1038/s41586-025-08925-z}, url = {https://doi.org/10.1038/s41586-025-08925-z}, author = {St{\"u}rner, Tomke and Brooks, Paul and Serratosa Capdevila, Laia and Morris, Billy J and Javier, Alexandre and Fang, Siqi and Gkantia, Marina and Cachero, Sebastian and Beckett, Isabella R and Marin, Elizabeth C and Schlegel, Philipp and Champion, Andrew S and Moitra, Ilina and Richards, Alana and Klemm, Finja and Kugel, Leonie and Namiki, Shigehiro and Cheong, Han S J and Kovalyak, Julie and Tenshaw, Emily and Parekh, Ruchi and Phelps, Jasper S and Mark, Brandon and Dorkenwald, Sven and Bates, Alexander S and Matsliah, Arie and Yu, Szi-Chieh and McKellar, Claire E and Sterling, Amy and Seung, H Sebastian and Murthy, Mala and Tuthill, John C and Lee, Wei-Chung Allen and Card, Gwyneth M and Costa, Marta and Jefferis, Gregory S X E and Eichler, Katharina} } @article {69997, title = {A connectomic resource for neural cataloguing and circuit dissection of the larval zebrafish brain}, journal = {bioRxiv}, year = {2025}, month = {2025 Jun 16}, abstract = {

We present a correlated light and electron microscopy (CLEM) dataset from a 7-day-old larval zebrafish, integrating confocal imaging of genetically labeled excitatory (vglut2a) and inhibitory (gad1b) neurons with nanometer-resolution serial section EM. The dataset spans the brain and anterior spinal cord, capturing \>180,000 segmented soma, \>40,000 molecularly annotated neurons, and 30 million synapses, most of which were classified as excitatory, inhibitory, or modulatory. To characterize the directional flow of activity across the brain, we leverage the synaptic and cell body annotations to compute region-wise input and output drive indices at single cell resolution. We illustrate the dataset{\textquoteright}s utility by dissecting and validating circuits in three distinct systems: water flow direction encoding in the lateral line, recurrent excitation and contralateral inhibition in a hindbrain motion integrator, and functionally relevant targeted long-range projections from a tegmental excitatory nucleus, demonstrating that this resource enables rigorous hypothesis testing as well as exploratory-driven circuit analysis. The dataset is integrated into an open-access platform optimized to facilitate community reconstruction and discovery efforts throughout the larval zebrafish brain.

Preprint: https://www.biorxiv.org/content/early/2025/06/15/2025.06.10.658982

}, doi = {10.1101/2025.06.10.658982}, url = {https://www.biorxiv.org/content/early/2025/06/15/2025.06.10.658982}, author = {Petkova, Mariela D. and Januszewski, Micha{\l} and Blakely, Tim and Herrera, Kristian J. and Schuhknecht, Gregor F.P. and Tiller, Robert and Choi, Jinhan and Schalek, Richard L. and Boulanger-Weill, Jonathan and Peleg, Adi and Wu, Yuelong and Wang, Shuohong and Troidl, Jakob and Kumar Vohra, Sumit and Wei, Donglai and Lin, Zudi and Bahl, Armin and Tapia, Juan Carlos and Iyer, Nirmala and Miller, Zachary T. and Hebert, Kathryn B. and Pavarino, Elisa C. and Taylor, Milo and Deng, Zixuan and Stingl, Moritz and Hockling, Dana and Hebling, Alina and Wang, Ruohong C. and Zhang, Lauren L. and Dvorak, Sam and Faik, Zainab and King, Kareem I. and Goel, Pallavi and Wagner-Carena, Julian and Aley, David and Chalyshkan, Selimzhan and Contreas, Dominick and Li, Xiong and Muthukumar, Akila V. and Vernaglia, Marina S. and Carrasco, Teodoro Tapia and Melnychuck, Sofia and Yan, TingTing and Dalal, Ananya and DiMartino, James M. and Brown, Sam and Safo-Mensa, Nana and Greenberg, Ethan and Cook, Michael and Finley-May, Samantha and Flynn, Miriam A. and Hopkins, Gary Patrick and Kovalyak, Julie and Leonard, Meghan and Lohff, Alanna and Ordish, Christopher and Scott, Ashley L. and Takemura, Satoko and Walsh, Claire and Walsh, John J. and Berger, Daniel R. and Pfister, Hanspeter and Berg, Stuart and Knecht, Christopher and Meissner, Geoffrey W. and Korff, Wyatt and Ahrens, Misha B. and Jain, Viren and Lichtman, Jeff W. and Engert, Florian} } @article {69783, title = {Cortical control of innate behavior from subcortical demonstration}, journal = {bioRxiv}, year = {2025}, month = {2025 Feb 13}, abstract = {

Motor control in mammals is traditionally viewed as a hierarchy of descending spinal-targeting pathways, with frontal cortex at the top 1{\textendash}3. Many redundant muscle patterns can solve a given task, and this high dimensionality allows flexibility but poses a problem for efficient learning 4. Although a feasible solution invokes subcortical innate motor patterns, or primitives, to reduce the dimensionality of the control problem, how cortex learns to utilize such primitives remains an open question 5{\textendash}7. To address this, we studied cortical and subcortical interactions as head-fixed mice learned contextual control of innate hindlimb extension behavior. Na{\"\i}ve mice performed reactive extensions to turn off a cold air stimulus within seconds and, using predictive cues, learned to avoid the stimulus altogether in tens of trials. Optogenetic inhibition of large areas of rostral cortex completely prevented avoidance behavior, but did not impair hindlimb extensions in reaction to the cold air stimulus. Remarkably, mice covertly learned to avoid the cold stimulus even without any prior experience of successful, cortically-mediated avoidance. These findings support a dynamic, heterarchical model in which the dominant locus of control can change, on the order of seconds, between cortical and subcortical brain areas. We propose that cortex can leverage periods when subcortex predominates as demonstrations, to learn parameterized control of innate behavioral primitives.

}, doi = {10.1101/2025.02.12.637930}, url = {https://www.biorxiv.org/content/early/2025/02/13/2025.02.12.637930}, author = {Keller, Jason A. and Kwak, Iljung S. and Stark, Alyssa K. and Pachitariu, Marius and Branson, Kristin and Dudman, Joshua T.} } @article {69458, title = {Courtship song differs between African and European populations of Drosophila melanogaster and involves a strong effect locus}, journal = {G3 Genes|Genomes|Genetics}, year = {2025}, month = {2025 Mar 07}, abstract = {

The courtship song of\ Drosophila melanogaster\ has long served as an excellent model system for studies of animal communication and differences in courtship song have been demonstrated among populations and between species. Here, we report that flies of African and European origin, which diverged approximately 13,000 years ago, show significant genetic differentiation in the use of slow versus fast pulse song. Using a combination of quantitative trait mapping and population genetic analysis we detected a single strong QTL underlying this trait and we identified candidate genes that may contribute to the evolution of this trait. Song trait variation between parental strains of our recombinant inbred panel enabled detection of genomic intervals associated with six additional song traits, some of which include known courtship-related genes. These findings improve the prospects for further genetic insights into the evolution of reproductive behavior and the biology underlying courtship song.

bioRxiv Preprint:\ https://www.biorxiv.org/content/early/2024/05/17/2024.05.14.594231

}, doi = {10.1093/g3journal/jkaf050}, url = {https://doi.org/10.1093/g3journal/jkaf050}, author = {Lollar, Matthew J. and Kim, Elizabeth and Stern, David L. and Pool, John E.} } @article {69719, title = {Deep learning-based aberration compensation improves contrast and resolution in fluorescence microscopyAbstract}, journal = {Nature Communications}, volume = {16}, year = {2025}, month = {Jan-12-2025}, abstract = {

Optical aberrations hinder fluorescence microscopy of thick samples, reducing image signal, contrast, and resolution. Here we introduce a deep learning-based strategy for aberration compensation, improving image quality without slowing image acquisition, applying additional dose, or introducing more optics. Our method (i) introduces synthetic aberrations to images acquired on the shallow side of image stacks, making them resemble those acquired deeper into the volume and (ii) trains neural networks to reverse the effect of these aberrations. We use simulations and experiments to show that applying the trained {\textquoteleft}de-aberration{\textquoteright} networks outperforms alternative methods, providing restoration on par with adaptive optics techniques; and subsequently apply the networks to diverse datasets captured with confocal, light-sheet, multi-photon, and super-resolution microscopy. In all cases, the improved quality of the restored data facilitates qualitative image inspection and improves downstream image quantitation, including orientational analysis of blood vessels in mouse tissue and improved membrane and nuclear segmentation in\ C. elegans\ embryos.

}, doi = {10.1038/s41467-024-55267-x}, url = {https://www.nature.com/articles/s41467-024-55267-xhttps://www.nature.com/articles/s41467-024-55267-x.pdfhttps://www.nature.com/articles/s41467-024-55267-x.pdfhttps://www.nature.com/articles/s41467-024-55267-x}, author = {Guo, Min and Wu, Yicong and Hobson, Chad M. and Su, Yijun and Qian, Shuhao and Krueger, Eric and Christensen, Ryan and Kroeschell, Grant and Bui, Johnny and Chaw, Matthew and Zhang, Lixia and Liu, Jiamin and Hou, Xuekai and Han, Xiaofei and Lu, Zhiye and Ma, Xuefei and Zhovmer, Alexander and Combs, Christian and Moyle, Mark and Yemini, Eviatar and Liu, Huafeng and Liu, Zhiyi and Benedetto, Alexandre and La Riviere, Patrick and Col{\'o}n-Ramos, Daniel and Shroff, Hari} } @article {69457, title = {Deep-tissue transcriptomics and subcellular imaging at high spatial resolution}, journal = {Science}, year = {2025}, month = {2025 Feb 20}, abstract = {

Limited color channels in fluorescence microscopy have long constrained spatial analysis in biological specimens. Here, we introduce cycle Hybridization Chain Reaction (HCR), a method that integrates multicycle DNA barcoding with HCR to overcome this limitation. cycleHCR enables highly multiplexed imaging of RNA and proteins using a unified barcode system. Whole-embryo transcriptomics imaging achieved precise three-dimensional gene expression and cell fate mapping across a specimen depth of ~310 μm. When combined with expansion microscopy, cycleHCR revealed an intricate network of 10 subcellular structures in mouse embryonic fibroblasts. In mouse hippocampal slices, multiplex RNA and protein imaging uncovered complex gene expression gradients and cell-type-specific nuclear structural variations. cycleHCR provides a quantitative framework for elucidating spatial regulation in deep tissue contexts for research and potentially diagnostic applications.

Preprint: https://doi.org/10.1101/2024.05.17.594641

}, doi = { 10.1126/science.adq2084}, url = {https://www.science.org/doi/10.1126/science.adq2084}, author = {Gandin, Valentina and Kim, Jun and Yang, Liang-Zhong and Lian, Yumin and Kawase, Takashi and Hu, Amy and Rokicki, Konrad and Fleishman, Greg and Tillberg, Paul and Aguilera Castrejon, Alejandro and Stringer, Carsen and Preibisch, Stephan and Liu, Zhe J.} } @article {70054, title = {Design of Orthogonal Far-Red, Orange and Green Fluorophore-binding Proteins for Multiplex Imaging}, journal = {bioRxiv}, year = {2025}, month = {2025 Aug 03}, abstract = {

Fluorescent proteins have transformed biological imaging, yet their limited photostability and brightness restrict their applications. We used deep learning-based de novo protein design methods to design binders to three bright, stable and cell-permeable dyes spanning the imaging spectrum: JF657 (far red), JF596 (orange-red) and JF494 (green). We obtain highly specific dye-binding proteins with low nanomolar affinities for the intended target; a crystal structure of one binder confirms close resemblance to the design model. Simultaneous labeling of mammalian cells expressing three dye-specific binders at different subcellular compartments demonstrates the utility in multiplex imaging. We further expand the functionality of the binder by incorporating an active site that carries out nucleophilic aromatic substitution to form a covalent linkage with the dye, and develop split versions which reconstitute fluorescence at subcellular locations where both halves are present towards monitoring in-cell protein interactions and chemically induced dimerization. Our designed high affinity and specificity dye binders open up new opportunities for multiplexed biological imaging.

}, doi = {10.1101/2025.08.03.668343}, url = {https://www.biorxiv.org/content/early/2025/08/03/2025.08.03.668343}, author = {Tran, Long and Sharma, Shajesh and Klein, Steffen and Jurgens, David and Decarreau, Justin and Liu, Bingxu and Wang, Yujia and Bera, Asim K. and Kang, Alex and Woods, Jon and Joyce, Emily and Vafeados, Dionne K and Roullier, Nicole and Chen, Wei and Lee, Gyu Rie and Mahamid, Julia and Lavis, Luke D. and An, Linna and Baker, David} } @article {70047, title = {Distributed control circuits across a brain-and-cord connectome}, journal = {bioRxiv}, year = {2025}, month = {2025 Aug 02}, abstract = {

Just as genomes revolutionized molecular genetics, connectomes (maps of neurons and synapses) are transforming neuroscience. To date, the only species with complete connectomes are worms\ and sea squirts\ (103-104 synapses). By contrast, the fruit fly is more complex (108 synaptic connections), with a brain that supports learning and spatial memory and an intricate ventral nerve cord analogous to the vertebrate spinal cord. Here we report the first adult fly connectome that unites the brain and ventral nerve cord, and we leverage this resource to investigate principles of neural control. We show that effector cells (motor neurons, endocrine cells and efferent neurons targeting the viscera) are primarily influenced by local sensory cells in the same body part, forming local feedback loops. These local loops are linked by long-range circuits involving ascending and descending neurons organized into behavior-centric modules. Single ascending and descending neurons are often positioned to influence the voluntary movements of multiple body parts, together with endocrine cells or visceral organs that support those movements. Brain regions involved in learning and navigation supervise these circuits. These results reveal an architecture that is distributed, parallelized and embodied (tightly connected to effectors), reminiscent of distributed control architectures in engineered systems.

}, doi = {10.1101/2025.07.31.667571}, url = {https://www.biorxiv.org/content/early/2025/08/02/2025.07.31.667571}, author = {Bates, Alexander Shakeel and Phelps, Jasper S. and Kim, Minsu and Yang, Helen H. and Matsliah, Arie and Ajabi, Zaki and Perlman, Eric and Delgado, Kevin M. and Osman, Mohammed Abdal Monium and Salmon, Christopher K. and Gager, Jay and Silverman, Benjamin and Renauld, Sophia and Collie, Matthew F. and Fan, Jingxuan and Pacheco, Diego A. and Zhao, Yunzhi and Patel, Janki and Zhang, Wenyi and Serratosa Capdevilla, Laia and Roberts, Ruair{\'\i} J.V. and Munnelly, Eva J. and Griggs, Nina and Langley, Helen and Moya-Llamas, Borja and Maloney, Ryan T. and Yu, Szi-Chieh and Sterling, Amy R. and Sorek, Marissa and Kruk, Krzysztof and Serafetinidis, Nikitas and Dhawan, Serene and St{\"u}rner, Tomke and Klemm, Finja and Brooks, Paul and Lesser, Ellen and Jones, Jessica M. and Pierce-Lundgren, Sara E. and Lee, Su-Yee and Luo, Yichen and Cook, Andrew P. and McKim, Theresa H. and Kophs, Emily C. and Falt, Tjalda and Negr{\'o}n Morales, Alexa M. and Burke, Austin and Hebditch, James and Willie, Kyle P. and Willie, Ryan and Popovych, Sergiy and Kemnitz, Nico and Ih, Dodam and Lee, Kisuk and Lu, Ran and Halageri, Akhilesh and Bae, J. Alexander and Jourdan, Ben and Schwartzman, Gregory and Demarest, Damian D. and Behnke, Emily and Bland, Doug and Kristiansen, Anne and Skelton, Jaime and Stocks, Tom and Garner, Dustin and Salman, Farzaan and Daly, Kevin C. and Hernandez, Anthony and Kumar, Sandeep and The BANC-FlyWire Consortium and Dorkenwald, Sven and Collman, Forrest and Suver, Marie P. and Fenk, Lisa M. and Pankratz, Michael J. and Jefferis, Gregory S.X.E. and Eichler, Katharina and Seeds, Andrew M. and Hampel, Stefanie and Agrawal, Sweta and Zandawala, Meet and Macrina, Thomas and Adjavon, Diane-Yayra and Funke, Jan and Tuthill, John C. and Azevedo, Anthony and Seung, H. Sebastian and de Bivort, Benjamin L. and Murthy, Mala and Drugowitsch, Jan and Wilson, Rachel I. and Lee, Wei-Chung Allen} } @article {69745, title = {Encoding of cerebellar dentate neuron activity during visual attention in rhesus macaques.}, journal = {Elife}, volume = {13}, year = {2025}, month = {2025 Jan 16}, abstract = {

The role of cerebellum in controlling eye movements is well established, but its contribution to more complex forms of visual behavior has remained elusive. To study cerebellar activity during visual attention we recorded extracellular activity of dentate nucleus (DN) neurons in two non-human primates (NHPs). NHPs were trained to read the direction indicated by a peripheral visual stimulus while maintaining fixation at the center, and report the direction of the cue by performing a saccadic eye movement into the same direction following a delay. We found that single-unit DN neurons modulated spiking activity over the entire time course of the task, and that their activity often bridged temporally separated intra-trial events, yet in a heterogeneous manner. To better understand the heterogeneous relationship between task structure, behavioral performance, and neural dynamics, we constructed a behavioral, an encoding, and a decoding model. Both NHPs showed different behavioral strategies, which influenced the performance. Activity of the DN neurons reflected the unique strategies, with the direction of the visual stimulus frequently being encoded long before an upcoming saccade. Moreover, the latency of the ramping activity of DN neurons following presentation of the visual stimulus was shorter in the better performing NHP. Labeling with the retrograde tracer Cholera Toxin B in the recording location in the DN indicated that these neurons predominantly receive inputs from Purkinje cells in the D1 and D2 zones of the lateral cerebellum as well as neurons of the principal olive and medial pons, all regions known to connect with neurons in the prefrontal cortex contributing to planning of saccades. Together, our results highlight that DN neurons can dynamically modulate their activity during a visual attention task, comprising not only sensorimotor but also cognitive attentional components.

}, keywords = {Animals, Attention, Cerebellar Nuclei, Macaca mulatta, Male, Neurons, Photic Stimulation, Saccades, Visual Perception}, issn = {2050-084X}, doi = {10.7554/eLife.99696}, author = {Flierman, Nico A and Koay, Sue Ann and van Hoogstraten, Willem S and Ruigrok, Tom J H and Roelfsema, Pieter and Badura, Aleksandra and De Zeeuw, Chris I} } @article {69840, title = {EPSILON: A method for pulse-chase labeling to probe synaptic AMPAR exocytosis during memory formation.}, journal = {Nat Neurosci}, year = {2025}, month = {2025 Mar 31}, abstract = {

A tool to map changes in synaptic strength during a defined time window could provide powerful insights into the mechanisms of learning and memory. Here we developed a technique, Extracellular Protein Surface Labeling in Neurons (EPSILON), to map α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor (AMPAR) exocytosis in vivo by sequential pulse-chase labeling of surface AMPARs with membrane-impermeable dyes. This approach yields synaptic-resolution maps of AMPAR exocytosis, a proxy for synaptic potentiation, in genetically targeted neurons during memory formation. In mice undergoing contextual fear conditioning, we investigated the relationship between synapse-level AMPAR exocytosis in CA1 pyramidal neurons and cell-level expression of the immediate early gene product cFos, a frequently used marker of engram neurons. We observed a strong correlation between AMPAR exocytosis and cFos expression, suggesting a synaptic mechanism for the association of cFos expression with memory engrams. The EPSILON technique is a useful tool for mapping synaptic plasticity and may be extended to investigate trafficking of other transmembrane proteins.

}, issn = {1546-1726}, doi = {10.1038/s41593-025-01922-5}, url = {https://doi.org/10.1038/s41593-025-01922-5}, author = {Kim, Doyeon and Park, Pojeong and Li, Xiuyuan and Wong-Campos, J David and Tian, He and Moult, Eric M and Grimm, Jonathan B and Lavis, Luke D and Cohen, Adam E} } @article {70178, title = {ER-to-Golgi trafficking is a nutrient-sensitive checkpoint linking glucose starvation to cell surface remodeling}, journal = {bioRxiv}, year = {2025}, month = {2025 October 31}, abstract = {

Cancer cells adapt to nutrient stress by remodeling the repertoire of proteins on their surface, enabling survival and progression under starvation conditions. However, the molecular mechanisms by which nutrient cues reshape the cell surface proteome to influence cell behavior remain largely unresolved. Here, we show that acute glucose starvation, but not amino acid deprivation or mTOR inhibition, selectively impairs ER-to-Golgi export of specific cargoes, such as E-cadherin, in a SEC24C-dependent manner. Quantitative cell surface proteomics reveal that glucose deprivation remodels the cell surface proteome, notably reducing surface expression of key adhesion molecules. This nutrient-sensitive reprogramming enhances cell migration in vitro and promotes metastasis in vivo. Mechanistically, we show that AMPK and ULK1 signaling orchestrate this process independent of autophagy, with ULK1-mediated phosphorylation of SEC31A driving SEC24C-dependent COPII reorganization. These findings establish ER-to-Golgi trafficking as a nutrient-sensitive regulatory node that links metabolic stress to cell surface remodeling and metastatic potential.

}, doi = {10.1101/2025.10.31.685804}, url = {https://www.biorxiv.org/content/early/2025/11/01/2025.10.31.685804}, author = {Joo, Joung Hyuck and Kasberg, William and Douglas, Spencer and Udoh, Uwemedimo and Carisey, Alexandre and Messing, James and Wang, Yong-Dong and Narina, Shilpa and Pruett-Miller, Shondra M. and Labelle, Myriam and Lippincott-Schwartz, Jennifer and Chang, Chi-Lun and Kundu, Mondira} } @article {70120, title = {Establishing a continuum of cell types in the visual cortex}, journal = {bioRxiv}, year = {2025}, month = {2025 Sep 22}, abstract = {

The mammalian cerebral cortex is composed of neurons whose properties vary in a continuous fashion rather than falling into discrete cell types. In the mouse visual cortex, excitatory neurons in layer 2 and 3 (L2/3) form such a continuum along cortical depth, patterned by the graded expression of hundreds of genes. Here we sought to understand how this continuum develops and contributes to cortical wiring. Using single-nucleus multiomics (RNA- and ATAC-Seq) and spatial transcriptomics, we show that the L2/3 continuum is established in two phases. During the first postnatal week, a genetically hardwired program establishes a primitive continuum of cell identities spanning the depth of L2/3. The second program, promoted by visual experience, is later superimposed upon the preexisting continuum. This second phase is driven by activity-regulated transcription factors that drive the L2/3 depth-dependent expression of genes linked to synaptic function and plasticity. We show that neurons at different positions along the L2/3 continuum project preferentially to distinct higher visual areas and that visual deprivation disrupts targeting to some higher visual areas while sparing others. Thus, cortical continua emerge through a stepwise process in which genetic programs and sensory experience specify neuronal identity and sculpt intracortical wiring specificity.

}, doi = {10.1101/2025.09.22.677893}, url = {https://www.biorxiv.org/content/early/2025/09/22/2025.09.22.677893}, author = {Yoo, Juyoun and Xie, Fangming and Butrus, Salwan and Xu, Runzhe and Tan, Zhiqun and Gorzek, Ryan and Mirshahidi, Parmis and Tring, Elaine and Suresh, Sanjana and Kim, Jinho and Fleishman, Greg and Tan, Liming and Ringach, Dario and Trachtenberg, Joshua and Xu, Xiangmin and Zipursky, S. Lawrence and Shekhar, Karthik and Jain, Saumya} } @article {69865, title = {FilaBuster: A strategy for rapid, specific, and spatiotemporally controlled intermediate filament disassembly}, journal = {bioRxiv}, year = {2025}, month = {2025 Apr 20}, abstract = {

Intermediate filaments (IFs) play key roles in cellular mechanics, signaling, and organization, but tools for their rapid, selective disassembly remain limited. Here, we introduce FilaBuster, a photochemical approach for efficient and spatiotemporally controlled IF disassembly in living cells. FilaBuster uses a three-step strategy: (1) targeting HaloTag to IFs, (2) labeling with a covalent photosensitizer ligand, and (3) light-induced generation of localized reactive oxygen species to trigger filament disassembly. This modular strategy applies broadly across IF subtypes{\textemdash}including vimentin, GFAP, desmin, peripherin, and keratin 18{\textemdash}and is compatible with diverse dyes and imaging platforms. Using vimentin IFs as a model system, we establish a baseline implementation in which vimentin-HaloTag labeled with a photosensitizer HaloTag ligand triggers rapid and specific IF disassembly upon light activation. We then refine this approach by (i) expanding targeting strategies to include a vimentin nanobody-HaloTag fusion, (ii) broadening the range of effective photosensitizers, and (iii) optimizing irradiation parameters to enable precise spatial control over filament disassembly. Together, these findings position FilaBuster as a robust platform for acute, selective, and spatiotemporally precise disassembly of IF networks, enabling new investigations into their structural and functional roles in cell physiology and disease.

}, doi = {10.1101/2025.04.20.649718}, url = {https://www.biorxiv.org/content/early/2025/04/21/2025.04.20.649718}, author = {Moore, Andrew S. and Krug, Tommy and Hansen, Simon B. and Ludlow, Alexander V. and Grimm, Jonathan B. and Ayala, Anthony X. and Plutkis, Sarah E. and Wang, Nan and Goldman, Robert D. and Medalia, Ohad and Lavis, Luke D. and Weitz, David A. and Lippincott-Schwartz, Jennifer} } @article {70031, title = {Fishexplorer: A multimodal cellular atlas platform for neuronal circuit dissection in larval zebrafish}, journal = {bioRxiv}, year = {2025}, month = {2025 Jul 14}, abstract = {

Understanding how neural circuits give rise to behavior requires comprehensive knowledge of neuronal morphology, connectivity, and function. Atlas platforms play a critical role in enabling the visualization, exploration, and dissemination of such information. Here, we present FishExplorer, an interactive and expandable community platform designed to integrate and analyze multimodal brain data from larval zebrafish. FishExplorer supports datasets acquired through light microscopy (LM), electron microscopy (EM), and X-ray imaging, all co-registered within a unified spatial coordinate system which enables seamless comparison of neuronal morphologies and synaptic connections. To further assist circuit analysis, FishExplorer includes a suite of tools for querying and visualizing connectivity at the whole-brain scale. By integrating data from recent large-scale EM reconstructions (presented in companion studies), FishExplorer enables researchers to validate circuit models, explore wiring principles, and generate new hypotheses. As a continuously evolving resource, FishExplorer is designed to facilitate collaborative discovery and serve the growing needs of the teleost neuroscience community.

}, doi = {10.1101/2025.07.14.664689}, url = {https://www.biorxiv.org/content/early/2025/07/15/2025.07.14.664689}, author = {Vohra, Sumit K. and Eberle, Maren and Boulanger-Weill, Jonathan and Petkova, Mariela D. and Schuhknecht, Gregor F. P. and Herrera, Kristian J. and K{\"a}mpf, Florian and Ruetten, Virginia M. S. and Lichtman, Jeff W. and Engert, Florian and Randlett, Owen and Bahl, Armin and Isoe, Yasuko and Hege, Hans-Christian and Baum, Daniel} } @article {70200, title = {Form and function of actin impacts actin health and aging}, journal = {bioRxiv}, year = {2025}, month = {2025 Nov 23 }, abstract = {

The actin cytoskeleton is a fundamental and highly conserved structure that functions in diverse cellular processes, yet its direct contribution to organismal aging remains unclear. Here, we systematically interrogated how genetic and pharmacologic perturbations of actin structure and function influence lifespan and various hallmarks of aging in Caenorhabditis elegans. Whole-animal and tissue-specific knockdown of actin and key actin-binding proteins (ABPs) - arx-2 (Arp2/3), unc-60 (cofilin), and lev-11 (tropomyosin) - led to premature disruption of filament organization, reduced lifespan, and tissue-specific physiological defects. Bulk and single-nucleus RNA-sequencing revealed that ABP knockdowns elicited a strongly {\textquotedblleft}aged{\textquotedblright} transcriptome. Actin dysfunction broadly exacerbated many age-associated phenotypes, including mitochondrial dysfunction, lipid dysregulation, loss of proteostasis, impaired autophagy, and intestinal barrier failure. Pharmacological destabilization with Latrunculin A mirrored genetic knockdowns, while mild stabilization with Jasplakinolide modestly extended lifespan, emphasizing that optimal and finely-tuned actin function is critical for healthy aging. Finally, analysis of human genome-wide association data revealed that common ACTB polymorphisms correlate with differences in age-related decline in gait speed, suggesting evolutionary conservation of actin{\textquoteright}s role in healthy aging. Taken together, our results provide a comprehensive and publicly accessible resource that maps, for the first time, how actin integrity intersects with diverse aging pathways across tissues and scales. This descriptive framework is intended to enable future mechanistic discovery by offering a deep, unbiased dataset that can be integrated with emerging studies to define how actin dynamics contribute to aging.

}, doi = {10.1101/2025.11.22.689949}, url = {https://www.biorxiv.org/content/early/2025/11/23/2025.11.22.689949}, author = {Averbukh, Maxim and Nelson, Hunter and Wang, Tiffany and Lange, Caitlin M and Alcala, Athena and Nair, Tripti and Kim, Juri and Suh, Daniella and Vega, Matthew and Gao, Shihong Max and Dutta, Naibedya and Aviles Barahona, Rebecca and Coakley, Aeowynn J and Gerke, Joe A. and Odron, Sofia F. and Clarke, Jacqueline G. and Singh, Navjot and Nickel, Matthew S. and Peterson, Jacob C. and Lam-Truong, Hannah L. and Moaddeli, Darius and Castro Torres, Toni and Oorloff, Maria and Peruch, Melissa and Mehta, Hemal and Thorwald, Max A. and Curran, Sean P. and Wang, Meng C and Johnson, Tyler A. and Crimmins, Eileen M. and Arpawong, Thalida Em and Kumsta, Caroline and Garcia, Gilberto and Higuchi-Sanabria, Ryo} } @article {69838, title = {Fourier-Based 3D Multistage Transformer for Aberration Correction in Multicellular Specimens}, journal = {Research Square}, year = {2025}, month = {2025 Apr 02}, abstract = {

High-resolution tissue imaging is often compromised by sample-induced optical aberrations that degrade resolution and contrast. While wavefront sensor-based adaptive optics (AO) can measure these aberrations, such hardware solutions are typically complex, expensive to implement, and slow when serially mapping spatially varying aberrations across large fields of view. Here, we introduce AOViFT (Adaptive Optical Vision Fourier Transformer)---a machine learning-based aberration sensing framework built around a 3D multistage Vision Transformer that operates on Fourier domain embeddings. AOViFT infers aberrations and restores diffraction-limited performance in puncta-labeled specimens with substantially reduced computational cost, training time, and memory footprint compared to conventional architectures or real-space networks. We validated AOViFT on live gene-edited zebrafish embryos, demonstrating its ability to correct spatially varying aberrations using either a deformable mirror or post-acquisition deconvolution. By eliminating the need for the guide star and wavefront sensing hardware and simplifying the experimental workflow, AOViFT lowers technical barriers for high-resolution volumetric microscopy across diverse biological samples.

}, doi = {10.21203/rs.3.rs-6273247/v1}, url = {https://doi.org/10.21203/rs.3.rs-6273247/v1}, author = {Thayer Alshaabi and Daniel Milkie and Gaoxiang Liu and Cyna Shirazinejad and Jason Hong and Kemal Achour and Frederik G{\"o}rlitz and Ana Milunovic-Jevtic and Cat Simmons and Ibrahim Abuzahriyeh and Erin Hong and Samara Williams and Nathanael Harrison and Evan Huang and Eun Bae and Alison Killilea and David Drubin and Ian Swinburne and Srigokul Upadhyayula and Eric Betzig} } @article {70132, title = {Fourier-based three-dimensional multistage transformer for aberration correction in multicellular specimens.}, journal = {Nat Methods}, year = {2025}, month = {2025 Oct 01}, abstract = {

High-resolution tissue imaging is often compromised by sample-induced optical aberrations that degrade resolution and contrast. Although wavefront sensor-based adaptive optics (AO) can measure these aberrations, such hardware solutions are typically complex, expensive to implement and slow when serially mapping spatially varying aberrations across large fields of view. Here we introduce AOVIFT (adaptive optical vision Fourier transformer)-a machine learning-based aberration sensing framework built around a three-dimensional multistage vision transformer that operates on Fourier domain embeddings. AOVIFT infers aberrations and restores diffraction-limited performance in puncta-labeled specimens with substantially reduced computational cost, training time and memory footprint compared to conventional architectures or real-space networks. We validated AOVIFT on live gene-edited zebrafish embryos, demonstrating its ability to correct spatially varying aberrations using either a deformable mirror or postacquisition deconvolution. By eliminating the need for the guide star and wavefront sensing hardware and simplifying the experimental workflow, AOVIFT lowers technical barriers for high-resolution volumetric microscopy across diverse biological samples.

Preprint: https://doi.org/10.48550/arXiv.2503.12593

}, issn = {1548-7105}, doi = {10.1038/s41592-025-02844-7}, url = {https://doi.org/10.1038/s41592-025-02844-7}, author = {Alshaabi, Thayer and Milkie, Daniel E and Liu, Gaoxiang and Shirazinejad, Cyna and Hong, Jason L and Achour, Kemal and G{\"o}rlitz, Frederik and Milunovic-Jevtic, Ana and Simmons, Cat and Abuzahriyeh, Ibrahim S and Hong, Erin and Williams, Samara Erin and Harrison, Nathanael and Huang, Evan and Bae, Eun Seok and Killilea, Alison N and Swinburne, Ian A and Drubin, David G and Upadhyayula, Srigokul and Betzig, Eric} } @article {70101, title = {Functional synapses between neurons and small cell lung cancer.}, journal = {Nature}, year = {2025}, month = {2025 Sep 10}, abstract = {

Small\ cell lung cancer (SCLC) is a highly aggressive type of lung cancer, characterized by rapid proliferation, early metastatic spread, frequent early relapse and a high mortality rate. Recent evidence has suggested that innervation has an important role in the development and progression of several types of cancer. Cancer-to-neuron synapses have been reported in gliomas, but whether peripheral tumours can form such structures is unknown. Here we show that SCLC cells can form functional synapses and receive synaptic transmission. Using in vivo insertional mutagenesis screening in conjunction with cross-species genomic and transcriptomic validation, we identified neuronal, synaptic and glutamatergic signalling gene sets in mouse and human SCLC. Further experiments revealed the ability of SCLC cells to form synaptic structures with neurons in vitro and in vivo. Electrophysiology and optogenetic experiments confirmed that cancer cells can receive NMDA receptor- and GABA receptor-mediated synaptic inputs. Fitting with a potential oncogenic role of neuron-SCLC interactions, we showed that SCLC cells derive a proliferation advantage when co-cultured with vagal sensory or cortical neurons. Moreover, inhibition of glutamate signalling had therapeutic efficacy in an autochthonous mouse model of SCLC. Therefore, following malignant transformation, SCLC cells seem to hijack synaptic signalling to promote tumour growth, thereby exposing a new route for therapeutic intervention.

}, issn = {1476-4687}, doi = {10.1038/s41586-025-09434-9}, url = {https://doi.org/10.1038/s41586-025-09434-9}, author = {Sakthivelu, Vignesh and Schmitt, Anna and Odenthal, Franka and Ndoci, Kristiano and Touet, Marian and Shaib, Ali H and Chihab, Abdulla and Wani, Gulzar A and Nieper, Pascal and Hartmann, Griffin G and Pintelon, Isabel and Kisis, Ilmars and Boecker, Maike and Eckert, Naja M and Iannicelli Caiaffa, Manoela and Ibruli, Olta and Weber, Julia and Maresch, Roman and Bebber, Christina M and Chitsaz, Ali and L{\"u}tz, Anna and Kim Alves Carpinteiro, Mira and Morris, Kaylee M and Franchino, Camilla A and Benz, Jonas and P{\'e}rez-Revuelta, Laura and Soriano-Campos, Jorge A and Huetzen, Maxim A and Goergens, Jonas and Jevtic, Milica and Jahn-Kelleter, Hannah M and Zempel, Hans and Placzek, Aleksandra and Hennrich, Alexandru A and Conzelmann, Karl-Klaus and Tumbrink, Hannah L and Hunold, Pascal and Isensee, Joerg and Werr, Lisa and Gaedke, Felix and Schauss, Astrid and Min{\`e}re, Marielle and M{\"u}ller, Marie and Fenselau, Henning and Liu, Yin and Heimsoeth, Alena and G{\"u}lc{\"u}ler Balta, G{\"u}lce S and Walczak, Henning and Frezza, Christian and Jachimowicz, Ron D and George, Julie and Schmiel, Marcel and Br{\"a}gelmann, Johannes and Hucho, Tim and von Karstedt, Silvia and Peifer, Martin and Annibaldi, Alessandro and H{\"a}nsel-Hertsch, Robert and Persigehl, Thorsten and Gr{\"u}ll, Holger and Sos, Martin L and Reifenberger, Guido and Fischer, Matthias and Adriaensen, Dirk and B{\"u}ttner, Reinhard and Sage, Julien and Brouns, Inge and Rad, Roland and Thomas, Roman K and Anst{\"o}tz, Max and Rizzoli, Silvio O and Bergami, Matteo and Motori, Elisa and Reinhardt, Hans Christian and Beleggia, Filippo} } @article {70007, title = {A genetic driver of epileptic encephalopathy impairs gating of synaptic glycolysis}, journal = {bioRxiv}, year = {2025}, month = {2025 Jun 19}, abstract = {

The brain is a disproportionately large consumer of fuel, estimated to expend \~20\% of the whole-body energy budget, and therefore it is critical to adequately control brain fuel expenditures while satisfying its on-demand needs for continued function. The brain is also metabolically vulnerable as the inability to adequately fuel cellular processes that support information transfer between cells leads to rapid neurological impairment. We show here that a genetic driver of early onset epileptic encephalopathy (EOEE), SLC13A5, a Na+/citrate cotransporter (NaCT), is critical for gating the activation of local presynaptic glycolysis. We show that SLC13A5 is in part localized to a presynaptic pool of membrane-bound organelles and acts to transiently clear axonal citrate during electrical activity, in turn activating phosphofructokinase 1. We show that loss of SLC13A5 or mistargeting to the plasma membrane results in suppressed glycolytic gating, activity dependent presynaptic bioenergetic deficits and synapse dysfunction.

}, doi = {10.1101/2025.06.17.660213}, url = {https://www.biorxiv.org/content/early/2025/06/19/2025.06.17.660213}, author = {Koshenov, Zhanat and Kokotos, Alexandros C. and Benedetti, Lorena and Schwartz, Jennifer Lippincott and Ryan, Timothy A.} } @article {69833, title = {Glutamate indicators with increased sensitivity and tailored deactivation rates}, journal = {Nat Methods}, year = {2025}, month = {2025 Dec 23}, abstract = {

Understanding how neurons integrate signals from thousands of input synapses requires methods to monitor neurotransmission across many sites simultaneously. The fluorescent protein glutamate indicator iGluSnFR enables visualization of synaptic signaling, but the sensitivity, scale and speed of such measurements are limited by existing variants. Here we developed two highly sensitive fourth-generation iGluSnFR variants with fast activation and tailored deactivation rates: iGluSnFR4f for tracking rapid dynamics, and iGluSnFR4s for recording from large populations of synapses. These indicators detect glutamate with high spatial specificity and single-vesicle sensitivity in vivo. We used them to record natural patterns of synaptic transmission across multiple experimental contexts in mice, including two-photon imaging in cortical layers 1{\textendash}4 and hippocampal CA1, and photometry in the midbrain. The iGluSnFR4 variants extend the speed, sensitivity and scalability of glutamate imaging, enabling direct observation of information flow through neural networks in the intact brain.

}, doi = {10.1038/s41592-025-02965-z}, url = {https://doi.org/10.1038/s41592-025-02965-z}, author = {Aggarwal, Abhi and Negrean, Adrian and Chen, Yang and Iyer, Rishyashring and Reep, Daniel and Liu, Anyi and Palutla, Anirudh and Xie, Michael E. and MacLennan, Bryan J. and Hagihara, Kenta M. and Kinsey, Lucas W. and Sun, Julianna L. and Yao, Pantong and Zheng, Jihong and Tsang, Arthur and Tsegaye, Getahun and Zhang, Yonghai and Patel, Ronak H. and Arthur, Benjamin J. and Hiblot, Julien and Leippe, Philipp and Tarnawski, Miroslaw and Marvin, Jonathan S. and Vevea, Jason D. and Turaga, Srinivas C. and Tebo, Alison G. and Carandini, Matteo and Rossi, L. F. and Kleinfeld, David and Konnerth, Arthur and Svoboda, Karel and Turner, Glenn C. and Hasseman, Jeremy and Podgorski, Kaspar} } @article {69856, title = {Glutamate indicators with increased sensitivity and tailored deactivation rates}, journal = {Research Square}, year = {2025}, month = {2025 Apr 8}, abstract = {

Identifying the input-output operations of neurons requires measurements of synaptic transmission simultaneously at many of a neuron{\textquoteright}s thousands of inputs in the intact brain. To facilitate this goal, we engineered and screened 3365 variants of the fluorescent protein glutamate indicator iGluSnFR3 in neuron culture, and selected variants in the mouse visual cortex. Two variants have high sensitivity, fast activation (\< 2 ms) and deactivation times tailored for recording large populations of synapses (iGluSnFR4s, 153 ms) or rapid dynamics (iGluSnFR4f, 26 ms). By imaging action-potential evoked signals on axons and visually-evoked signals on dendritic spines, we show that iGluSnFR4s/4f primarily detect local synaptic glutamate with single-vesicle sensitivity. The indicators detect a wide range of naturalistic synaptic transmission, including in the vibrissal cortex layer 4 and in hippocampal CA1 dendrites. iGluSnFR4 increases the sensitivity and scale (4s) or speed (4f) of tracking information flow in neural networks in vivo.

}, doi = {10.21203/rs.3.rs-6257403/v1}, url = {https://doi.org/10.21203/rs.3.rs-6257403/v1}, author = {Podgorski, Kaspar and Aggarwal, Abhi and Negrean, Adrian and Chen, Yang and Iyer, Rishyashring and Reep, Daniel and Liu, Anyi and Palutla, Anirudh and Xie, Michael and Maclennan, Bryan and Hagihara, Kenta and Kinsey, Lucas and Sun, Julianna and Yao, Pantong and Zheng, Jihong and Tsang, Arthur and Tsegaye, Getahun and Zhang, Yonghai and Patel, Ronak and Hasseman, Jeremy} } @article {69954, title = {High-resolution imaging of RNA and proteins in thick tissues using cycleHCR.}, journal = {Nat Rev Genet}, year = {2025}, month = {2025 May 28}, issn = {1471-0064}, doi = {10.1038/s41576-025-00860-z}, url = {https://doi.org/10.1038/s41576-025-00860-z}, author = {Kim, Jun and Liu, Zhe J} } @article {69825, title = {HortaCloud: An open and collaborative platform for whole brain neuronal reconstructions}, journal = {bioRxiv}, year = {2025}, month = {2025 Mar 13}, abstract = {

HortaCloud is a cloud-based, open-source platform designed to facilitate the collaborative reconstruction of long-range projection neurons from whole-brain light microscopy data. By providing virtual environments directly within the cloud, it eliminates the need for costly and time-consuming data downloads, allowing researchers to work efficiently with terabyte- scale volumetric datasets. Standardization of computational resources in the cloud make deployment easier and more predictable. The pay-as-you-go cloud model reduces adoption barriers by eliminating upfront investments in expensive hardware. Finally, HortaCloud{\textquoteright}s decentralized architecture enables global collaboration between researchers and between institutions.

}, doi = {10.1101/2025.03.13.642887}, url = {https://www.biorxiv.org/content/early/2025/03/14/2025.03.13.642887}, author = {Rokicki, Konrad and Schauder, David and Olbris, Donald J. and Goina, Cristian and Clements, Jody and Edson, Patrick and Kawase, Takashi and Svirskas, Robert and Arshadi, Cameron and Feng, David and Chandrashekar, Jayaram and Ferreira, Tiago A. and MouseLight Project Team} } @article {69831, title = {iGABASnFR2: Improved genetically encoded protein sensors of GABA}, journal = {bioRxiv}, year = {2025}, month = {2025 Mar 25}, abstract = {

Monitoring GABAergic inhibition in the nervous system has been enabled by development of an intensiometric molecular sensor that directly detects GABA. However the first generation iGABASnFR exhibits low signal-to-noise and suboptimal kinetics, making in vivo experiments challenging. To improve sensor performance, we targeted several sites in the protein for near-saturation mutagenesis, and evaluated the resulting sensor variants in a high throughput screening system using evoked synaptic release in primary cultured neurons. This identified a sensor variant, iGABASnFR2, with 4.2-fold improved sensitivity and 20\% faster kinetics, and binding affinity that remained in a range sensitive to changes in GABA concentration at synapses. We also identified sensors with an inverted response, decreasing fluorescence intensity upon GABA binding. We termed the best such negative-going sensor iGABASnFR2n, which can be used to corroborate observations with the positive-going sensor. These improvements yielded a qualitative enhancement of in vivo performance, enabling us to make the first measurements of direction selective GABA release in the retina and confirm a longstanding hypothesis for how sensitivity to motion arises in the visual system.

}, doi = {10.1101/2025.03.25.644953}, url = {https://www.biorxiv.org/content/early/2025/03/26/2025.03.25.644953}, author = {Kolb, Ilya and Hasseman, Jeremy P. and Matsumoto, Akihiro and Arthur, Benjamin J. and Zhang, Yan and Tsang, Arthur and Reep, Daniel and Tsegaye, Getahun and Zheng, Jihong and Patel, Ronak and Looger, Loren L. and Marvin, Jonathan S. and Korff, Wyatt L. and Yonehara, Keisuke and Turner, Glenn C.} } @article {70182, title = {iGlucoSnFR2: A genetically encoded fluorescent sensor for measuring intracellular or extracellular glucose in vivo in mouse brain.}, journal = {Sci Adv}, volume = {11}, year = {2025}, month = {2025 Nov 14}, pages = {eadz3889}, abstract = {

Continuous glucose monitors have proven invaluable for monitoring blood glucose levels for diabetics, but they are of limited use for observing glucose dynamics at the cellular (or subcellular) level. We have developed a second generation, genetically encoded intensity-based glucose sensing fluorescent reporter (iGlucoSnFR2). We show that when it is targeted to the cytosol, it reports intracellular glucose consumption and gluconeogenesis in cell culture, along with efflux from the endoplasmic reticulum. It outperforms the original iGlucoSnFR in vivo when observed by fiber photometry in mouse brain and reports transient increase in glucose concentration when stimulated by noradrenaline or electrical stimulation. Last, we demonstrate that membrane localized iGlucoSnFR2 can be calibrated in vivo to indicate absolute changes in extracellular glucose concentration in awake mice. We anticipate iGlucoSnFR2 facilitating previously unobservable measurements of glucose dynamics with high spatial and temporal resolution in living mammals and other experimental organisms.

}, issn = {2375-2548}, doi = {10.1126/sciadv.adz3889}, url = {https://doi.org/10.1126/sciadv.adz3889}, author = {Marvin, Jonathan S and M{\"a}chler, Philipp and Meng, Chengbo and Ates, Tayfun and Patel, Ronak H and Adhikari, Raghabendra and Makurath, Monika A and Ku, Zaneta and Feliciano, Daniel and Atasoy, Deniz and Cui, Guohong and Kleinfeld, David and Brown, Timothy A} } @article {70084, title = {Imaging cellular activity simultaneously across all organs of a vertebrate reveals body-wide circuits}, journal = {bioRxiv}, year = {2025}, month = {2025 August 22}, abstract = {

All cells in an animal collectively ensure, moment-to-moment, the survival of the whole organism in the face of environmental stressors1,2. Physiology seeks to elucidate the intricate network of interactions that sustain life, which often span multiple organs, cell types, and timescales, but a major challenge lies in the inability to simultaneously record time-varying cellular activity throughout the entire body.We developed WHOLISTIC, a method to image second-timescale, time-varying intracellular dynamics across cell-types of the vertebrate body. By advancing and integrating volumetric fluorescence microscopy, machine learning, and pancellular transgenic expression of calcium sensors in transparent young Danio rerio (zebrafish) and adult Danionella, the method enables real-time recording of cellular dynamics across the organism. Calcium is a universal intracellular messenger, with a large array of cellular processes depending on changes in calcium concentration across varying time-scales, making it an ideal proxy of cellular activity3.Using this platform to screen the dynamics of all cells in the body, we discovered unexpected responses of specific cell types to stimuli, such as chondrocyte reactions to cold, meningeal responses to ketamine, and state-dependent activity, such as oscillatory ependymal-cell activity during periods of extended motor quiescence. At the organ scale, the method uncovered pulsating traveling waves along the kidney nephron. At the multi-organ scale, we uncovered muscle synergies and independencies, as well as muscle-organ interactions. Integration with optogenetics allowed us to all-optically determine the causal direction of brain-body interactions. At the whole-organism scale, the method captured the rapid brainstem-controlled redistribution of blood flow across the body.Finally, we advanced Whole-Body Expansion Microscopy4 to provide ground-truth molecular and ultrastructural anatomical context, explaining the spatiotemporal structure of activity captured by WHOLISTIC. Together, these innovations establish a new paradigm for systems biology, bridging cellular and organismal physiology, with broad implications for both fundamental research and drug discovery.

}, doi = {10.1101/2025.08.20.670374}, url = {https://www.biorxiv.org/content/early/2025/08/22/2025.08.20.670374.1}, author = {Ruetten, Virginia M. S. and Zheng, Wei and Siwanowicz, Igor and Mensh, Brett D. and Eddison, Mark and Hu, Amy and Chi, Yunfeng and Lemire, Andrew L. and Guo, Caiying and Kadobianskyi, Mykola and Renz, Marc and Lelek-Greskovic, Sara and He, Yisheng and Close, Kari and Ihrke, Gudrun and Dev, Aparna and Petruncio, Alyson and Wan, Yinan and Engert, Florian and Fishman, Mark C. and Judkewitz, Benjamin and Rubinov, Mikail and Keller, Philipp J. and Satou, Chie and Yu, Guoqiang and Tillberg, Paul W. and Sahani, Maneesh and Ahrens, Misha B.} } @article {70006, title = {Large-scale high-density brain-wide neural recording in nonhuman primates.}, journal = {Nat Neurosci}, year = {2025}, month = {2025 Jun 23}, abstract = {

High-density silicon probes have transformed neuroscience by enabling large-scale neural recordings at single-cell resolution. However, existing technologies have provided limited functionality in nonhuman primates (NHPs) such as macaques. In the present report, we describe the design, fabrication and performance of Neuropixels 1.0 NHP, a high-channel electrode array designed to enable large-scale acute recording throughout large animal brains. The probe features 4,416 recording sites distributed along a 45-mm shank. Experimenters can programmably select 384 recording channels, enabling simultaneous multi-area recording from thousands of neurons with single or multiple probes. This technology substantially increases scalability and recording access relative to existing technologies and enables new classes of experiments that involve electrophysiological mapping of brain areas at single-neuron and single-spike resolution, measurement of spike-spike correlations between cells and simultaneous brain-wide recordings at scale.

Preprint: https://doi.org/10.1101/2023.02.01.526664\ 

}, issn = {1546-1726}, doi = {10.1038/s41593-025-01976-5}, url = {https://doi.org/10.1038/s41593-025-01976-5}, author = {Trautmann, Eric M and Hesse, Janis K and Stine, Gabriel M and Xia, Ruobing and Zhu, Shude and O{\textquoteright}Shea, Daniel J and Karsh, Bill and Colonell, Jennifer and Lanfranchi, Frank F and Vyas, Saurabh and Zimnik, Andrew and Amematsro, Elom and Steinemann, Natalie A and Wagenaar, Daniel A and Pachitariu, Marius and Andrei, Alexandru and Lopez, Carolina Mora and O{\textquoteright}Callaghan, John and Putzeys, Jan and Raducanu, Bogdan C and Welkenhuysen, Marleen and Churchland, Mark and Moore, Tirin and Shadlen, Michael and Shenoy, Krishna and Tsao, Doris and Dutta, Barundeb and Harris, Timothy} } @article {68938, title = {Learning produces an orthogonalized state machine in the hippocampus.}, journal = {Nature}, volume = {640}, year = {2025}, month = {2025 February 12}, abstract = {

Cognitive maps confer animals with flexible intelligence by representing spatial, temporal and abstract relationships that can be used to shape thought, planning and behaviour. Cognitive maps have been observed in the hippocampus1, but their algorithmic form and learning mechanisms remain obscure. Here we used large-scale, longitudinal two-photon calcium imaging to record activity from thousands of neurons in the CA1 region of the hippocampus while mice learned to efficiently collect rewards from two subtly different linear tracks in virtual reality. Throughout learning, both animal behaviour and hippocampal neural activity progressed through multiple stages, gradually revealing improved task representation that mirrored improved behavioural efficiency. The learning process involved progressive decorrelations in initially similar hippocampal neural activity within and across tracks, ultimately resulting in orthogonalized representations resembling a state machine capturing the inherent structure of the task. This decorrelation process was driven by individual neurons acquiring task-state-specific responses (that is, {\textquoteright}state cells{\textquoteright}). Although various standard artificial neural networks did not naturally capture these dynamics, the clone-structured causal graph, a hidden Markov model variant, uniquely reproduced both the final orthogonalized states and the learning trajectory seen in animals. The observed cellular and population dynamics constrain the mechanisms underlying cognitive map formation in the hippocampus, pointing to hidden state inference as a fundamental computational principle, with implications for both biological and artificial intelligence.

}, doi = {10.1038/s41586-024-08548-w}, url = {https://doi.org/10.1038/s41586-024-08548-w}, author = {Sun, Weinan and Winnubst, Johan and Natrajan, Maanasa and Lai, Chongxi and Kajikawa, Koichiro and Michaelos, Michalis and Gattoni, Rachel and Stringer, Carsen and Flickinger, Daniel and Fitzgerald, James E and Spruston, Nelson} } @article {69656, title = {Leukocytes use endothelial membrane tunnels to extravasate the vasculature}, journal = {Cell Rep}, year = {2025}, month = {2025 Sep 04}, abstract = {

Upon inflammation, leukocytes extravasate through endothelial cells. When they extravasate, it is generally accepted that neighboring endothelial cells disconnect. Careful examination of endothelial junctions showed a partial membrane overlap beyond VE-cadherin distribution. These overlaps are regulated by actin polymerization and, although marked by, do not require PECAM-1, nor VE-cadherin. Neutrophils prefer wider membrane overlaps as exit sites. Detailed 3D analysis of neutrophil transmigration in real time at high spatiotemporal resolution revealed that overlapping endothelial membranes form a tunnel during neutrophil transmigration. These tunnels are formed by the neutrophil lifting the membrane of the upper endothelial cell while indenting and crawling over the membrane of the underlying endothelial cell. Our work shows that endothelial cells do not simply retract upon the passage of neutrophils but provide membrane tunnels, allowing neutrophils to extravasate. This discovery defines the 3D multicellular architecture in which the paracellular transmigration of neutrophils occurs.

}, doi = {10.1016/j.celrep.2025.116242}, url = {https://doi.org/10.1016/j.celrep.2025.116242}, author = {van der Meer, Werner and van Steen, Abraham C.I. and Mahlandt, Eike and Rolas, Loic and Wang, Haitao and Arts, Janine and Kempers, Lanette and Gr{\"o}nloh, Max and Schoon, Rianne M. and Driessen, Amber and van Rijssel, Jos and Klaassen, Ingeborg and Schlingemann, Reinier O. and Manavski, Yosif and Hoogenboezem, Mark and Boon, Reinier A. and Khuon, Satya and Wait, Eric and Heddleston, John and Chew, Teng-Leong and Nolte, Martijn A and Nourshargh, Sussan and Goedhart, Joachim and van Buul, Jaap D} } @article {69773, title = {Live imaging of the extracellular matrix with a glycan-binding fluorophore.}, journal = {Nat Methods}, year = {2025}, month = {2025 Feb 06}, abstract = {

All multicellular systems produce and dynamically regulate extracellular matrices (ECMs) that play essential roles in both biochemical and mechanical signaling. Though the spatial arrangement of these extracellular assemblies is critical to their biological functions, visualization of ECM structure is challenging, in part because the biomolecules that compose the ECM are difficult to fluorescently label individually and collectively. Here, we present a cell-impermeable small-molecule fluorophore, termed Rhobo6, that turns on and red shifts upon reversible binding to glycans. Given that most ECM components are densely glycosylated, the dye enables wash-free visualization of ECM, in systems ranging from in vitro substrates to in vivo mouse mammary tumors. Relative to existing techniques, Rhobo6 provides a broad substrate profile, superior tissue penetration, non-perturbative labeling, and negligible photobleaching. This work establishes a straightforward method for imaging the distribution of ECM in live tissues and organisms, lowering barriers for investigation of extracellular biology.

}, issn = {1548-7105}, doi = {10.1038/s41592-024-02590-2}, url = {https://doi.org/10.1038/s41592-024-02590-2}, author = {Fiore, Antonio and Yu, Guoqiang and Northey, Jason J and Patel, Ronak and Ravenscroft, Thomas A. and Ikegami, Richard and Kolkman, W and Pratik Kumar and Dilan, Tanya L and Ruetten, Virginia M.S. and Ahrens, Misha B and Shroff, Hari and Wang, Shaohe and Weaver, Valarie M and Pedram, Kayvon} } @article {69945, title = {Macrophages release neuraminidase and cleaved calreticulin for programmed cell removal.}, journal = {Proc Natl Acad Sci U S A}, volume = {122}, year = {2025}, month = {2025 May 27}, pages = {e2426644122}, abstract = {

Calreticulin (CALR) is primarily an endoplasmic reticulum chaperone protein that also plays a key role in facilitating programmed cell removal (PrCR) by acting as an "eat-me" signal for macrophages, directing their recognition and engulfment of dying, diseased, or unwanted cells. Recent findings have demonstrated that macrophages can transfer their own CALR onto exposed asialoglycans on target cells, marking them for PrCR. Despite the critical role CALR plays in this process, the molecular mechanisms behind its secretion by macrophages and the formation of binding sites on target cells remain unclear. Our findings show that CALR undergoes C-terminal cleavage upon secretion, producing a truncated form that functions as the active eat-me signal detectable on target cells. We identify cathepsins as potential proteases involved in this cleavage process. Furthermore, we demonstrate that macrophages release neuraminidases, which modify the surface of target cells and facilitate CALR binding. These insights reveal a coordinated mechanism through which lipopolysaccharide (LPS)-activated macrophages regulate CALR cleavage and neuraminidase activity to mark target cells for PrCR. How they recognize the cells to be targeted remains unknown.

}, keywords = {Animals, Apoptosis, Calreticulin, Humans, Lipopolysaccharides, Macrophages, Mice, Neuraminidase}, issn = {1091-6490}, doi = {10.1073/pnas.2426644122}, url = {https://doi.org/10.1073/pnas.2426644122}, author = {Banuelos, Allison and Baez, Michelle and Zhang, Allison and Y{\i}lmaz, Leyla and Kasberg, William and Volk, Regan and Georgeos, Nardin and Koren-Sedova, Elle and Le, Uyen and Burden, Andrew T and Marjon, Kristopher D and Lippincott-Schwartz, Jennifer and Zaro, Balyn W and Weissman, Irving L} } @article {70050, title = {Michael Patrick Sheetz, 1946{\textendash}2025, a devotee of and major contributor to membrane and cytoskeletal biology}, journal = {Molecular Biology of the Cell}, volume = {36}, year = {2025}, month = {2025 Jul 31}, pages = {fe1}, abstract = {

Michael P. Sheetz (1946{\textendash}2025) advanced the field of mechanobiology through his creative experiments, new methodologies, and keen insights. His research touched many fields of cell biology, including membrane biophysics, motor proteins, the cytoskeleton, cell migration, and cellular senescence. In addition to his research, Sheetz was a leader who built vibrant academic departments and institutes and advanced the careers of many trainees.

}, doi = {10.1091/mbc.E25-05-0208}, url = {https://doi.org/10.1091/mbc.E25-05-0208}, author = {Kenney, Linda J. and Vale, Ronald D. and Spudich, James A.} } @article {70039, title = {MicroSplit: Semantic Unmixing of Fluorescent Microscopy Data}, journal = {bioRxiv}, year = {2025}, month = {2025 Jul 21}, abstract = {

Fluorescence microscopy, a key driver for progress in the life sciences, faces limitations due to the microscope{\textquoteright}s optics, fluorophore chemistry, and photon exposure limits, necessitating trade-offs in imaging speed, resolution, and depth. Here, we introduce MicroSplit, a computational multiplexing technique based on deep learning that allows multiple cellular structures to be imaged in a single fluorescent channel and then unmixed computationally, allowing faster imaging and reduced photon exposure. We show that MicroSplit efficiently separates up to four superimposed noisy structures into distinct denoised fluorescent image channels. Furthermore, using Variational Splitting Encoder-Decoder (VSE) networks, our approach can sample diverse predictions from a trained posterior of solutions. The diversity of these samples scales with the uncertainty in a given input, allowing us to estimate the true prediction errors by computing the variability between posterior samples. We demonstrate the robustness of MicroSplit across various datasets and noise levels and show its utility to image more, image faster, and improve downstream analysis. We provide MicroSplit along with all associated training and evaluation datasets as open resources, enabling life scientists to benefit from the potential of computational multiplexing and accelerate the pace of scientific discovery.

}, doi = {10.1101/2025.02.10.637323}, url = {https://www.biorxiv.org/content/early/2025/07/21/2025.02.10.637323}, author = {Ashesh, Ashesh and Carrara, Federico and Zubarev, Igor and Galinova, Vera and Croft, Melisande and Pezzotti, Melissa and Gong, Daozheng and Casagrande, Francesca and Colombo, Elisa and Giussani, Stefania and Restelli, Elena and Cammarota, Eugenia and Battagliotti, Juan Manuel and Klena, Nikolai and Di Sante, Moises and Adhikari, Raghabendra and Feliciano, Daniel and Pigino, Gaia and Taverna, Elena and Harschnitz, Oliver and Maghelli, Nicola and Scherer, Norbert and Dalle Nogare, Damian Edward and Deschamps, Joran and Pasqualini, Francesco and Jug, Florian} } @article {69862, title = {Mitochondrial complexity is regulated at ER-mitochondria contact sites via PDZD8-FKBP8 tethering.}, journal = {Nat Commun}, volume = {16}, year = {2025}, month = {2025 Apr 17}, pages = {3401}, abstract = {

Mitochondria-ER membrane contact sites (MERCS) represent a fundamental ultrastructural feature underlying unique biochemistry and physiology in eukaryotic cells. The ER protein PDZD8 is required for the formation of MERCS in many cell types, however, its tethering partner on the outer mitochondrial membrane (OMM) is currently unknown. Here we identify the OMM protein FKBP8 as the tethering partner of PDZD8 using a combination of unbiased proximity proteomics, CRISPR-Cas9 endogenous protein tagging, Cryo-electron tomography, and correlative light-electron microscopy. Single molecule tracking reveals highly dynamic diffusion properties of PDZD8 along the ER membrane with significant pauses and captures at MERCS. Overexpression of FKBP8 is sufficient to narrow the ER-OMM distance, whereas independent versus combined deletions of these two proteins demonstrate their interdependence for MERCS formation. Furthermore, PDZD8 enhances mitochondrial complexity in a FKBP8-dependent manner. Our results identify a novel ER-mitochondria tethering complex that regulates mitochondrial morphology in mammalian cells.

Preprint:\ 10.1101/2025.02.22.639343

}, keywords = {Animals, CRISPR-Cas Systems, Cryoelectron Microscopy, Endoplasmic Reticulum, HEK293 Cells, Hela Cells, Humans, Membrane Proteins, Mitochondria, Mitochondrial Membranes, Proteomics, Tacrolimus Binding Proteins}, issn = {2041-1723}, doi = {10.1038/s41467-025-58538-3}, url = {https://doi.org/10.1038/s41467-025-58538-3}, author = {Nakamura, Koki and Aoyama-Ishiwatari, Saeko and Nagao, Takahiro and Paaran, Mohammadreza and Obara, Christopher J and Sakurai-Saito, Yui and Johnston, Jake and Du, Yudan and Suga, Shogo and Tsuboi, Masafumi and Nakakido, Makoto and Tsumoto, Kouhei and Kishi, Yusuke and Gotoh, Yukiko and Kwak, Chulhwan and Rhee, Hyun-Woo and Seo, Jeong Kon and Kosako, Hidetaka and Potter, Clint and Carragher, Bridget and Lippincott-Schwartz, Jennifer and Polleux, Franck and Hirabayashi, Yusuke} } @article {69747, title = {Molecular determinants of optical modulation in ssDNA{\textendash}carbon nanotube biosensors}, journal = {ACS Nano}, year = {2025}, month = {01/2025}, abstract = {

Most traditional optical biosensors operate through molecular recognition, where ligand binding causes conformational changes that lead to optical perturbations in the emitting motif. Optical sensors developed from single-stranded DNA-functionalized single-walled carbon nanotubes (ssDNA{\textendash}SWCNTs) have started to make useful contributions to biological research. However, the mechanisms underlying their function have remained poorly understood. In this study, we combine experimental and computational approaches to show that ligand binding alone is not sufficient for optical modulation in this class of synthetic biosensors. Instead, the optical response that occurs after ligand binding is highly dependent on the chemical properties of the ligands, resembling mechanisms seen in activity-based biosensors. Specifically, we show that in ssDNA{\textendash}SWCNT catecholamine sensors, the optical response correlates positively with the electron density on the aryl motif, even among ligands with similar ligand binding affinities. Importantly, despite the strong correlations with electrochemical properties, we find that catechol oxidation itself is not necessary to drive the sensor optical response. We discuss how these findings could serve as a framework for tuning the performance of existing sensors and guiding the development of new biosensors of this class.

}, doi = {10.1021/acsnano.4c13814}, url = {https://doi.org/10.1021/acsnano.4c13814}, author = {Krasley, Andrew T. and Chakraborty, Sayantani and Vukovi{\'c}, Lela and Beyene, Abraham G.} } @article {69994, title = {A multimodal adaptive optical microscope for in vivo imaging from molecules to organisms}, journal = {bioRxiv}, year = {2025}, month = {2025 Jun 16}, abstract = {

Understanding biological systems requires observing features and processes across vast spatial and temporal scales, spanning nanometers to centimeters and milliseconds to days, often using multiple imaging modalities within complex native microenvironments. Yet, achieving this comprehensive view is challenging because microscopes optimized for specific tasks typically lack versatility due to inherent optical and sample handling trade-offs, and frequently suffer performance degradation from sample-induced optical aberrations in multicellular contexts. Here, we present MOSAIC, a reconfigurable microscope that integrates multiple advanced imaging techniques including light-sheet, label-free, super-resolution, and multi-photon, all equipped with adaptive optics. MOSAIC enables non-invasive imaging of subcellular dynamics in both cultured cells and live multicellular organisms, nanoscale mapping of molecular architectures across millimeter-scale expanded tissues, and structural/functional neural imaging within live mice. MOSAIC facilitates correlative studies across biological scales within the same specimen, providing an integrated platform for broad biological investigation.

Preprint:\ https://www.biorxiv.org/content/early/2025/06/13/2025.06.02.657494

}, doi = {10.1101/2025.06.02.657494}, url = {https://www.biorxiv.org/content/early/2025/06/13/2025.06.02.657494}, author = {Fu, Tian-Ming and Liu, Gaoxiang and Milkie, Daniel E. and Ruan, Xiongtao and G{\"o}rlitz, Frederik and Shi, Yu and Ferro, Valentina and Divekar, Nikita S. and Wang, Wei and York, Harrison M. and Kilic, Velat and Mueller, Matthew and Liang, Yajie and Daugird, Timothy A. and Gacha-Garay, Maria J. and Larkin, Kathryn A. and Adikes, Rebecca C. and Harrison, Nathaniel and Shirazinejad, Cyna and Williams, Samara and Nourse, Jamison L. and Sheu, Shu-Hsien and Gao, Liang and Li, Tongchao and Mondal, Chandrani and Achour, Kemal and Hercule, Wilmene and Stabley, Daniel and Emmerich, Kevin and Dong, Peng and Drubin, David and Liu, Zhe J. and Clapham, David and Mumm, Jeff S. and Koyama, Minoru and Killilea, Alison and Bravo-Cordero, Jose Javier and Keene, C. Dirk and Luo, Liqun and Kirchhausen, Tom and Pathak, Medha M. and Arumugam, Senthil and Nunez, James K. and Gao, Ruixuan and Matus, David Q. and Martin, Benjamin L. and Swinburne, Ian A. and Betzig, Eric and Legant, Wesley R. and Upadhyayula, Srigokul} } @article {70191, title = {Multimodal cell lineage reconstruction in the hindbrain reveals a link between progenitor origin and activity patterning}, journal = {bioRxiv}, year = {2025}, month = {2025 Nov 14}, abstract = {

The vertebrate brain develops through a precisely orchestrated sequence of events, where neural progenitors generate neuronal diversity. However, the importance of stem cell origin in the acquisition of functional characteristics and the assembly into circuits are largely unknown. Here, by multimodal cell lineage reconstruction we reveal a link between progenitor origin and excitatory and inhibitory activity in the hindbrain. By combining progenitor intersectional fate mapping with newly developed computational tools and CRISPR/Cas9-based loss-of-function experiments, we discovered the differential temporal contribution of distinct progenitor pools {\textendash}defined by proneural gene expression{\textendash} to both glutamatergic and GABAergic neurons. Whereas neurog1-progenitors contribute to both lineages in early embryonic stages, later, other progenitor pools either also assume this role or take over. Using calcium imaging in combination with genetic perturbations, we unveiled that spatiotemporally defined stem cell origin determines specific neuronal activity features. Overall, we provide in vivo evidence for a link between cell ontogeny and functionality in the vertebrate hindbrain.For more information or any specific tool, you can contact us.

}, doi = {10.1101/2025.11.14.688499}, url = {https://www.biorxiv.org/content/early/2025/11/14/2025.11.14.688499}, author = {Blanc, Matthias and Meister, Lydvina and Lemon, William C. and Fiuza, Ulla-Maj and Keller, Philipp J. and Espinosa-Medina, Isabel and Pujades, Cristina} } @article {69960, title = {Myocardium and endocardium of the early mammalian heart tube arise from independent multipotent lineages specified at the primitive streak}, journal = {Dev Cell}, year = {2025}, month = {2025 May 20}, abstract = {

The formation of the primitive heart tube from cardiomyocytes and endocardial cells is a key event in mammalian development. Previous studies suggested that cardiomyocytes and endocardial cells segregate from a shared cardiac progenitor around the onset of gastrulation, yet their lineage relationship with other mesodermal tissues remains unclear. Using retrospective and prospective clonal analyses in mouse embryos, we traced cardiomyocyte and endocardial progenitors from the primitive streak to the heart tube. Our results identify two independent mesodermal populations specified around gastrulation onset. While each of these populations is unipotent in producing cardiomyocytes or endocardium, they retain multipotency and contribute to different subsets of non-cardiac mesoderm. Nonetheless, live imaging identifies simultaneous ingression and intermingling of these two mesodermal lineages in the primitive streak, showing their coordinated specification and migration. The proposed model for cardiac progenitor specification will help understanding the origins of congenital heart diseases and designing tissue engineering strategies.

}, issn = {1878-1551}, doi = {10.1016/j.devcel.2025.05.002}, url = {https://doi.org/10.1016/j.devcel.2025.05.002}, author = {Sendra, Miquel and McDole, Katie and de Dios Hourcade, Juan and Temi{\~n}o, Susana and Raiola, Morena and Guignard, Leo and Keller, Philipp J and Dom{\'\i}nguez, Jorge N and Torres, Miguel} } @article {70156, title = {Natural evolution of intermale sexual behavior by multiple pheromone switches among Drosophila species}, journal = {bioRxiv}, year = {2025}, month = {2025 Oct 14}, abstract = {

We have identified a Drosophila species in which males exhibit spontaneous, elaborate, and robust intermale sexual behavior. Males of D. santomea, a West African island endemic, distinguish conspecific sexes but court males and females promiscuously and seldom attack. Elevated intermale courtship derives from at least three changes in two separate pheromone systems. In males, the sexually monomorphic cuticular pheromone 7-tricosene promotes rather than inhibits courtship and the courtship-inhibiting olfactory pheromone cVA is reduced 84-92\% compared to close relatives, including the sibling species D. yakuba. The third change is surprisingly in D. santomea females, where cVA suppresses rather than promotes sexual receptivity. The female cVA switch and male cVA reduction may have co-evolved to maintain efficient intraspecific mating in D. santomea but prevent sympatric hybridization with D. yakuba, or to reduce intraspecific aggression. We find that high intermale courtship and low cVA also co-occur and appear selectively derived in a distant monomorphic species D. persimilis, implying pheromonal and behavioral convergence in the two recently speciated taxa. The data suggest that sequential changes in the behavioral valence and levels of pheromones explain the recent evolutionary emergence of intermale sexual behavior in Drosophila.

}, doi = {10.1101/2025.10.14.682417}, url = {https://www.biorxiv.org/content/early/2025/10/15/2025.10.14.682417}, author = {Ouadah, Youcef and Naragon, Thomas H. and Smihula, Hayley and Behrman, Emily L. and Khallaf, Mohammed A. and Ding, Yun and Stern, David L. and Parker, Joseph and Anderson, David J.} } @article {70162, title = {Networks of sexually dimorphic neurons that regulate social behaviors in Drosophila}, journal = {bioRxiv}, year = {2025}, month = {2025 Oct 21}, abstract = {

Neural mechanisms underlying sexually dimorphic social behaviors remain enigmatic in most species. In Drosophila, sexually dimorphic P1/pC1x neurons have been described as a site of sensory integration that regulates mating and aggressive behaviors. We show that the male P1/pC1x population forms a highly intertwined network with male-specific mAL and aSP-a neurons that is poised to regulate male behavior. The 48 P1/pC1x cell types exhibit heterogeneous synaptic connections with a subset receiving strong input from identified sensory pathways. We also describe circuit motifs by which P1 and sexually dimorphic aIPg neurons co-regulate social behaviors. Genetic driver lines for these cell types were generated and used to discover distinct roles for P1/pC1x cell types in promoting social acoustic signaling and male-male interactions. Our results reveal unexpected diversity in the connectivity and behavioral roles of the P1/pC1x cell types and provide essential genetic tools for interrogating their neurophysiological and behavioral functions.

}, doi = {10.1101/2025.10.21.683766}, url = {https://www.biorxiv.org/content/early/2025/10/22/2025.10.21.683766}, author = {Rubin, Gerald M. and Managan, Claire and Dreher, Marisa and Kim, Elizabeth and Miller, Scott and Boone, Kaitlyn and Robie, Alice and Taylor, Adam L. and Branson, Kristin and Schretter, Catherine E. and Otopalik, Adriane G.} } @article {69946, title = {Neuronal growth patterns and synapse formation are mediated by distinct activity-dependent mechanisms.}, journal = {Sci Rep}, volume = {15}, year = {2025}, month = {2025 May 19}, pages = {17338}, abstract = {

All brain functions in animals rely upon neuronal connectivity that is established during early development. Although the activity-dependent mechanisms are deemed important for brain development and adult synaptic plasticity, the precise cellular and molecular mechanisms remain however, largely unknown. This lack of fundamental knowledge regarding developmental neuronal assembly owes its existence to the complexity of the mammalian brain as cell-cell interactions between individual neurons cannot be investigated directly. Here, we used individually identified synaptic partners from Lymnaea stagnalis to interrogate the role of neuronal activity patterns over an extended time period during various growth time points and synaptogenesis. Using intracellular recordings, microelectrode arrays, and time-lapse imaging, we identified unique patterns of activity throughout neurite outgrowth and synapse formation. Perturbation of voltage-gated Ca channels compromised neuronal growth patterns which also invoked a protein kinase A mediated pathway. Our findings underscore the importance of unique activity patterns in regulating neuronal growth, neurite branching, and synapse formation, and identify the underlying cellular and molecular mechanisms.

}, keywords = {Animals, Cyclic AMP-Dependent Protein Kinases, Lymnaea, Neurites, Neurogenesis, Neuronal Outgrowth, Neuronal Plasticity, Neurons, Synapses}, issn = {2045-2322}, doi = {10.1038/s41598-025-00806-9}, url = {https://doi.org/10.1038/s41598-025-00806-9}, author = {Yacoub, Matthew and Iqbal, Fahad and Khan, Zainab and Syeda, Atika and Lijnse, Thomas and Syed, Naweed I} } @article {69774, title = {Neuropixels Opto: Combining high-resolution electrophysiology and optogenetics}, journal = {bioRxiv}, year = {2025}, month = {2025 Feb 06}, abstract = {

High-resolution extracellular electrophysiology is the gold standard for recording spikes from distributed neural populations, and is especially powerful when combined with optogenetics for manipulation of specific cell types with high temporal resolution. We integrated these approaches into prototype Neuropixels Opto probes, which combine electronic and photonic circuits. These devices pack 960 electrical recording sites and two sets of 14 light emitters onto a 1 cm shank, allowing spatially addressable optogenetic stimulation with blue and red light. In mouse cortex, Neuropixels Opto probes delivered high-quality recordings together with spatially addressable optogenetics, differentially activating or silencing neurons at distinct cortical depths. In mouse striatum and other deep structures, Neuropixels Opto probes delivered efficient optotagging, facilitating the identification of two cell types in parallel. Neuropixels Opto probes represent an unprecedented tool for recording, identifying, and manipulating neuronal populations.

}, doi = {10.1101/2025.02.04.636286}, url = {https://www.biorxiv.org/content/early/2025/02/06/2025.02.04.636286}, author = {Lakunina, Anna and Socha, Karolina Z and Ladd, Alexander and Bowen, Anna J and Chen, Susu and Colonell, Jennifer and Doshi, Anjal and Karsh, Bill and Krumin, Michael and Kulik, Pavel and Li, Anna and Neutens, Pieter and O{\textquoteright}Callaghan, John and Olsen, Meghan and Putzeys, Jan and Tilmans, Harrie AC and Ye, Zhiwen and Welkenhuysen, Marleen and H{\"a}usser, Michael and Koch, Christof and Ting, Jonathan T. and Neuropixels Opto Consortium and Dutta, Barun and Harris, Timothy D and Steinmetz, Nicholas A and Svoboda, Karel and Siegle, Joshua H and Carandini, Matteo} } @article {70169, title = {Optimizing multifunctional fluorescent ligands for intracellular labeling.}, journal = {Proc Natl Acad Sci USA}, volume = {122}, year = {2025}, month = {2025 Nov 04}, pages = {e2510046122}, abstract = {

Enzyme-based self-labeling tags enable the covalent attachment of synthetic molecules to proteins inside living cells. A frontier of this field is designing cell-permeable multifunctional ligands that contain fluorophores in combination with affinity tags or pharmacological agents. This is challenging since attachment of additional chemical moieties onto fluorescent ligands can adversely affect membrane permeability. To address this problem, we examined the chemical properties of rhodamine-based self-labeling tag ligands through the lens of medicinal chemistry. We found that the lactone-zwitterion equilibrium constant () of rhodamines inversely correlates with their distribution coefficients (log), suggesting that ligands based on dyes exhibiting low and high log values, such as Si-rhodamines, would efficiently enter cells. We designed cell-permeable multifunctional HaloTag ligands with a biotin moiety to purify mitochondria or a JQ1 appendage to translocate BRD4 within the nucleus. We found that translocation of BRD4 to constitutive heterochromatin in cells leads to apparent increases in transcriptional activity. These fluorescent reagents enable affinity capture and translocation of intracellular proteins in living cells, and our general design concepts will facilitate the design of multifunctional chemical tools for biology.

Preprint:\ https://doi.org/10.1101/2022.07.02.498544
Preprint:\ https://doi.org/10.32388/0xcyuc

}, keywords = {Bromodomain Containing Proteins, Cell Cycle Proteins, Fluorescent Dyes, Humans, Ligands, Mitochondria, Nuclear Proteins, Rhodamines, Staining and Labeling, Transcription Factors}, issn = {1091-6490}, doi = {10.1073/pnas.2510046122}, url = {https://doi.org/10.1073/pnas.2510046122}, author = {Kumar, Pratik and Vevea, Jason D and Tkachuk, Ariana N and Campbell, Kirby R and Watson, Emma T and Ayala, Anthony X and Grimm, Jonathan B and Chapman, Edwin R and Solecki, David J and Lavis, Luke D} } @article {69756, title = {Periodic ER-plasma membrane junctions support long-range Ca signal integration in dendrites.}, journal = {Cell}, volume = {188}, year = {2025}, month = {01/2025}, pages = {484-500.e22}, abstract = {

Neuronal dendrites must relay synaptic inputs over long distances, but the mechanisms by which activity-evoked intracellular signals propagate over macroscopic distances remain unclear. Here, we discovered a system of periodically arranged endoplasmic reticulum-plasma membrane (ER-PM) junctions tiling the plasma membrane of dendrites at \~{}1\ μm intervals, interlinked by a meshwork of ER tubules patterned in a ladder-like array. Populated with Junctophilin-linked plasma membrane voltage-gated Ca channels and ER Ca-release channels (ryanodine receptors), ER-PM junctions are hubs for ER-PM crosstalk, fine-tuning of Ca homeostasis, and local activation of the Ca/calmodulin-dependent protein kinase II. Local spine stimulation activates the Ca modulatory machinery, facilitating signal transmission and ryanodine-receptor-dependent Ca release at ER-PM junctions over 20\ μm away. Thus, interconnected ER-PM junctions support signal propagation and Ca release from the spine-adjacent ER. The capacity of this subcellular architecture to modify both local and distant membrane-proximal biochemistry potentially contributes to dendritic computations.

}, keywords = {Animals, Calcium, Calcium Signaling, Calcium-Calmodulin-Dependent Protein Kinase Type 2, Cell Membrane, Dendrites, Endoplasmic Reticulum, Hippocampus, Humans, Male, Mice, Rats, Ryanodine Receptor Calcium Release Channel}, issn = {1097-4172}, doi = {10.1016/j.cell.2024.11.029}, author = {Benedetti, Lorena and Fan, Ruolin and Weigel, Aubrey V and Moore, Andrew S and Houlihan, Patrick R and Kittisopikul, Mark and Park, Grace and Petruncio, Alyson and Hubbard, Philip M and Pang, Song and Xu, C Shan and Hess, Harald F and Saalfeld, Stephan and Rangaraju, Vidhya and Clapham, David E and De Camilli, Pietro and Ryan, Timothy A and Lippincott-Schwartz, Jennifer} } @article {70119, title = {A priming circuit controls the olfactory response and memory in Drosophila}, journal = {eLife}, year = {2025}, month = {2025 Sep 23}, abstract = {

Priming is a process by which exposure to a stimulus affects the response to a subsequent stimulus in humans. In this study we found that in\ Drosophila, a prior encounter with an aversive stimulus results in enhanced preference for a following novel odor, while an appetitive stimulus leads to reduced preference for a new odor. This priming behavior of flies relies on the well-studied olfactory memory circuits including Kenyon cells (KC), dopaminergic neurons (DANs), and mushroom body output neurons (MBONs). Aversive stimulus results in increased odor responses in reward DANs that innervate the γ4 lobe of the mushroom body (MB) and decreased odor responses in a γ4γ5-innvervating repulsive MBON. We concluded that these neurons are required for the priming effects in flies. These results characterized the newly found priming behavior in flies and demonstrated the sheer influence of unconditioned stimulus on odor perception during associative learning.

}, doi = {10.7554/elife.107736.1}, url = {http://dx.doi.org/10.7554/eLife.107736.1}, author = {Yang, He and Jiang, Yang and Kunes, Samuel} } @article {70080, title = {RABGAP1 is a sensor that facilitates the sorting and processing of amyloid precursor protein}, journal = {The EMBO Journal}, year = {2025}, month = {2026 Aug 26}, pages = {1-20}, abstract = {

A hallmark of Alzheimer{\textquoteright}s disease (AD) is the accumulation of extracellular amyloid-β plaques in the brain. Amyloid-β is a 40{\textendash}42 amino acid peptide generated by proteolytic processing of amyloid precursor protein (APP) via membrane-bound proteases. APP is a transmembrane protein, and its trafficking to sites of proteolysis represents a rate-limiting step in AD progression. Although APP processing has been well-studied, its trafficking itinerary and machinery remain incompletely understood. To address this, we performed an unbiased interaction screen for interactors of the APP cytosolic tail. We identified previously characterised APP binders as well as novel interactors, including RABGAP1. We demonstrated that RABGAP1 partially co-localises with APP and directly interacts with a YENPTY motif in the APP cytosolic tail. Depletion or overexpression of RABGAP1 caused mistrafficking and misprocessing of endogenous APP in human and rodent neurons. This effect is dependent on the GAP activity of RABGAP1, demonstrating that RABGAP1 affects the trafficking of APP by modulating RAB activity on endosomal subdomains. This novel trafficking mechanism has implications for other NPXY cargoes and presents a possible therapeutic avenue to explore.

}, keywords = {Alzheimer{\textquoteright}s Disease, APP, RABGAP1, Trafficking}, doi = {10.1038/s44318-025-00530-0}, url = {https://www.embopress.org/doi/abs/10.1038/s44318-025-00530-0}, author = {Eden, Jessica and Kaufman, Jonathan G G and Pereira, Concei{\c c}{\~a}o and Fox, Eleanor and Cattin-Ortol{\'a}, Jerome and Benedetti, Lorena and Nieuwenhuis, Bart and Owen, David J and Lippincott-Schwartz, Jennifer and Munro, Sean and Gershlick, David C} } @article {69781, title = {Recovery of plasma membrane tension after a hyperosmotic shock.}, journal = {Mol Biol Cell}, year = {2025}, month = {2025 Feb 19}, pages = {mbcE24100436}, abstract = {

Maintaining proper tension is critical for the organization and function of the plasma membrane. To study the mechanisms by which yeast restores normal plasma membrane tension, we used a microfluidics device to expose yeast to hyperosmotic conditions, which reduced cell volume and caused a \~{}20\% drop in cell surface area. The resulting low tension plasma membrane exhibited large clusters of negatively-charged glycerophospholipids together with nutrient transporters, suggesting phase segregation of the membrane. We found that endocytosis was blocked by the phase segregation and thus was not involved in removing excess membrane. In contrast, rapid recovery of plasma membrane tension was dependent on 1) eisosome morphology changes that were able to absorb most of the excess surface area and 2) lipid transport from the plasma membrane to the endoplasmic reticulum, where lipids were shunted into newly formed lipid droplets.

}, issn = {1939-4586}, doi = {10.1091/mbc.E24-10-0436}, author = {Phan, Jasmine and Silva, Malan and Kohlmeyer, Robin and Ruethemann, Romy and Gay, Lincoln and Jorgensen, Erik and Babst, Markus} } @article {70198, title = {Redefining colocalization analysis with a novel Phasor Mixing Coefficient.}, journal = {J Cell Sci}, year = {2025}, month = {2025 Nov 24}, abstract = {

The first step to probing any potential interaction between two biomolecules is to determine their spatial association. In other words, if two biomolecules localize similarly within a cell, then it is plausible they could interact. Traditionally, this is quantified through various colocalization metrics. These measures infer this association by estimating the degree to which fluorescent signals from each biomolecule overlap or correlate. However, these metrics are, at best, proxies, and they depend strongly on various experimental choices. Alternatively, here we define a new strategy which leverages multispectral imaging and phasor analysis, termed the Phasor Mixing Coefficient (PMC). PMC measures the precise mixing of fluorescent signals in each pixel. We demonstrate how PMC captures complex biological subtlety by offering two distinct values, a global measure of overall color mixing and the homogeneity thereof. We additionally show that PMC exhibits less sensitivity to signal-to-noise ratio, intensity threshold, and background signal compared to canonical methods. Moreover, this method provides a means to visualize color mixing at each pixel. We show that PMC offers users a nuanced and robust metric to quantify biological association.

}, issn = {1477-9137}, doi = {10.1242/jcs.264388}, author = {Puls, Owen F and Aaron, Jesse S and Quarles, Ellen K and Khuon, Satya and Eisenman, Leanna R and Kamaid, Andres and Malacrida, Leonel and Chew, Teng-Leong} } @article {70203, title = {Redefining colocalization analysis with a novel Phasor Mixing Coefficient.}, journal = {J Cell Sci}, year = {2025}, month = {2025 Nov 24}, abstract = {

The first step to probing any potential interaction between two biomolecules is to determine their spatial association. In other words, if two biomolecules localize similarly within a cell, then it is plausible they could interact. Traditionally, this is quantified through various colocalization metrics. These measures infer this association by estimating the degree to which fluorescent signals from each biomolecule overlap or correlate. However, these metrics are, at best, proxies, and they depend strongly on various experimental choices. Alternatively, here we define a new strategy which leverages multispectral imaging and phasor analysis, termed the Phasor Mixing Coefficient (PMC). PMC measures the precise mixing of fluorescent signals in each pixel. We demonstrate how PMC captures complex biological subtlety by offering two distinct values, a global measure of overall color mixing and the homogeneity thereof. We additionally show that PMC exhibits less sensitivity to signal-to-noise ratio, intensity threshold, and background signal compared to canonical methods. Moreover, this method provides a means to visualize color mixing at each pixel. We show that PMC offers users a nuanced and robust metric to quantify biological association.

}, issn = {1477-9137}, doi = {10.1242/jcs.264388}, url = {https://doi.org/10.1242/jcs.264388}, author = {Puls, Owen F and Aaron, Jesse S and Quarles, Ellen K and Khuon, Satya and Eisenman, Leanna R and Kamaid, Andres and Malacrida, Leonel and Chew, Teng-Leong} } @article {69993, title = {RubyACRs enable red-shifted optogenetic inhibition in freely behaving Drosophila}, journal = {eLife}, year = {2025}, month = {2025 Sep 05}, abstract = {

Optogenetic activators with red-shifted excitation spectra, such as Chrimson, have significantly advanced Drosophila neuroscience. However, until recently, available optogenetic inhibitors required shorter activation wavelengths, which don{\textquoteright}t penetrate tissue as effectively and are stronger visual stimuli to the animal, potentially confounding behavioral results. Here, we assess the efficacy of two newly identified anion-conducting channelrhodopsins with spectral sensitivities similar to Chrimson: A1ACR and HfACR (RubyACRs). Electrophysiology and functional imaging confirmed that RubyACRs effectively hyperpolarize neurons, with stronger and faster effects than the widely used inhibitor GtACR1. Activation of RubyACRs led to circuit-specific behavioral changes in three different neuronal groups. In glutamatergic motor neurons, activating RubyACRs suppressed adult locomotor activity. In PPL1-γ1pedc dopaminergic neurons, pairing odors with RubyACR activation during learning produced odor responses consistent with synaptic silencing. Finally, activation of RubyACRs in the pIP10 neuron suppressed pulse song during courtship. Together, these results demonstrate that RubyACRs are effective and reliable tools for neuronal inhibition in Drosophila, expanding the optogenetic toolkit for circuit dissection in freely behaving animals.

Preprint: https://www.biorxiv.org/content/early/2025/06/15/2025.06.13.659144

}, doi = {10.7554/eLife.108210.1}, url = {https://doi.org/10.7554/eLife.108210.1}, author = {Bushey, Daniel and Shiozaki, Hiroshi and Shuai, Yichun and Zheng, Jihong and Jayaraman, Vivek and Hasseman, Jeremy P. and Kolb, Ilya and GENIE Project Team and Turner, Glenn C.} } @article {69712, title = {Scalable image-based visualization and alignment of spatial transcriptomics datasets}, journal = {Cell Syst}, year = {2025}, month = {2025 Apr 17}, abstract = {

We present the "spatial transcriptomics imaging framework" (STIM), an imaging-based computational framework focused on visualizing and aligning high-throughput spatial sequencing datasets. STIM is built on the powerful, scalable ImgLib2 and BigDataViewer (BDV) image data frameworks and thus enables novel development or transfer of existing computer vision techniques to the sequencing domain characterized by datasets with irregular measurement-spacing and arbitrary spatial resolution, such as spatial transcriptomics data generated by multiplexed targeted hybridization or spatial sequencing technologies. We illustrate STIM{\textquoteright}s capabilities by representing, interactively visualizing, 3D rendering, automatically registering, and segmenting publicly available spatial sequencing data from 13 serial sections of mouse brain tissue and from 19 sections of a human metastatic lymph node. We demonstrate that the simplest alignment mode of STIM achieves human-level accuracy.

Preprint:\ www.biorxiv.org/content/early/2024/10/07/2021.12.07.471629

}, doi = {10.1016/j.cels.2025.101264}, url = {https://doi.org/10.1016/j.cels.2025.101264}, author = {Preibisch, Stephan and Innerberger, Michael and Le{\'o}n-Peri{\~n}{\'a}n, Daniel and Karaiskos, Nikos and Rajewsky, Nikolaus} } @article {70155, title = {Sexual dimorphism in the complete connectome of the Drosophila male central nervous system}, journal = {bioRxiv}, year = {2025}, month = {2025 Oct 09}, abstract = {

Sex differences in behaviour exist across the animal kingdom, typically under strong genetic regulation. In Drosophila, previous work has shown that fruitless and doublesex transcription factors identify neurons driving sexually dimorphic behaviour. However, the organisation of dimorphic neurons into functional circuits remains unclear.We now present the connectome of the entire Drosophila male central nervous system. This contains 166,691 neurons spanning the brain and ventral nerve cord, fully proofread and comprehensively annotated including fruitless and doublesex expression and 11,691 cell types. By comparison with a previous female brain connectome, we provide the first comprehensive description of the differences between male and female brains to synaptic resolution. Of 7,319 cross-matched cell types in the central brain, 114 are dimorphic with an additional 262 male- and 69 female-specific (totalling 4.8\% of neurons in males and 2.4\% in females).This resource enables analysis of full sensory-to-motor circuits underlying complex behaviours as well as the impact of dimorphic elements. Sex-specific and dimorphic neurons are concentrated in higher brain centres while the sensory and motor periphery are largely isomorphic. Within higher centres, male-specific connections are organised into hotspots defined by male-specific neurons or the presence of male-specific arbours on neurons that are otherwise similar between sexes. Numerous circuit switches reroute sensory information to form conserved, antagonistic circuits controlling opposing behaviours.

}, doi = {10.1101/2025.10.09.680999}, url = {https://www.biorxiv.org/content/early/2025/10/09/2025.10.09.680999}, author = {Berg, Stuart and Beckett, Isabella R and Costa, Marta and Schlegel, Philipp and Januszewski, Micha{\l} and Marin, Elizabeth C and Nern, Aljoscha and Preibisch, Stephan and Qiu, Wei and Takemura, Shin-ya and Fragniere, Alexandra MC and Champion, Andrew S and Adjavon, Diane-Yayra and Cook, Michael and Gkantia, Marina and Hayworth, Kenneth J and Huang, Gary B and Katz, William T and K{\"a}mpf, Florian and Lu, Zhiyuan and Ordish, Christopher and Paterson, Tyler and St{\"u}rner, Tomke and Trautman, Eric T and Whittle, Catherine R and Burnett, Laura E and Hoeller, Judith and Li, Feng and Loesche, Frank and Morris, Billy J and Pietzsch, Tobias and Pleijzier, Markus W and Silva, Valeria and Yin, Yijie and Ali, Iris and Badalamente, Griffin and Bates, Alexander Shakeel and Bogovic, John and Brooks, Paul and Cachero, Sebastian and Canino, Brandon S and Chaisrisawatsuk, Bhumpanya and Clements, Jody and Crowe, Arthur and de Haan Vicente, In{\^e}s and Dempsey, Georgia and Don{\`a}, Erika and dos Santos, M{\'a}rcia and Dreher, Marisa and Dunne, Christopher R and Eichler, Katharina and Finley-May, Samantha and Flynn, Miriam A and Hameed, Imran and Hopkins, Gary Patrick and Hubbard, Philip M and Kiassat, Ladann and Kovalyak, Julie and Lauchie, Shirley A and Leonard, Meghan and Lohff, Alanna and Longden, Kit D and Maldonado, Charli A and Mitletton, Myrto and Moitra, Ilina and Moon, Sung Soo and Mooney, Caroline and Munnelly, Eva J and Okeoma, Nneoma and Olbris, Donald J and Pai, Anika and Patel, Birava and Phillips, Emily M and Plaza, Stephen M and Richards, Alana and Rivas Salinas, Jennifer and Roberts, Ruair{\'\i} Jv and Rogers, Edward M and Scott, Ashley L and Scuderi, Louis A and Seenivasan, Pavithraa and Serratosa Capdevila, Laia and Smith, Claire and Svirskas, Rob and Takemura, Satoko and Tastekin, Ibrahim and Thomson, Alexander and Umayam, Lowell and Walsh, John J and Whittome, Holly and Xu, C Shan and Yakal, Emily A and Yang, Tansy and Zhao, Arthur and George, Reed and Jain, Viren and Jayaraman, Vivek and Korff, Wyatt and Meissner, Geoffrey W and Romani, Sandro and Funke, Jan and Knecht, Christopher and Saalfeld, Stephan and Scheffer, Louis K and Waddell, Scott and Card, Gwyneth M and Ribeiro, Carlos and Reiser, Michael B and Hess, Harald F and Rubin, Gerald M and Jefferis, Gregory Sxe} } @article {70181, title = {Single-cell synaptome mapping of endogenous protein subpopulations in mammalian brain.}, journal = {Nat Commun}, volume = {16}, year = {2025}, month = {2025 Nov 11}, pages = {9705}, abstract = {

Different spatial or temporal protein populations, such as cell-surface/intracellular or pre-existing/nascent subpopulations, determine the basal and activity-induced functions of individual synapses within a neuron in vivo. Here, we developed a simple and generalizable platform to image different spatial and temporal subpopulations of endogenous proteins at thousands of synapses in single neurons in the mammalian brain. The platform is based on the development, improvement and integration of CRISPR-Cas9-mediated protein labeling methods, chemical tag labeling techniques, and a semi-automatic analytical pipeline. The combined platform enables whole-cell mapping of total, cell-surface, intracellular, pre-existing, nascent or nascent-and-surface populations of endogenous proteins, such as receptor, scaffold and signaling proteins, at thousands of synapses in individual neurons in living or fixed mouse brain. Our single-cell "synaptome" mapping of endogenous protein subpopulations comprehensively visualizes the spatial representation of synapse diversity in protein localization, trafficking and turnover, providing valuable insights into single-cell organization and computations in the brain.

}, keywords = {Animals, Brain, CRISPR-Cas Systems, Mice, Mice, Inbred C57BL, Neurons, Single-Cell Analysis, Synapses}, issn = {2041-1723}, doi = {10.1038/s41467-025-65813-w}, url = {https://doi.org/10.1038/s41467-025-65813-w}, author = {Uchigashima, Motokazu and Iguchi, Risa and Fujii, Kazuma and Shiku, Kaito and Kumar, Pratik and Liu, Xinyi and Isogai, Mari and Hoshino, Chiaki and Abe, Manabu and Nozumi, Motohiro and Okamura, Yosuke and Igarashi, Michihiro and Sakimura, Kenji and Bise, Ryoma and Lavis, Luke D and Mikuni, Takayasu} } @article {68799, title = {Single-cell type analysis of wing premotor circuits in the ventral nerve cord of Drosophila melanogaster}, journal = {eLife}, year = {2025}, month = {2025 May 06}, abstract = {

To perform most behaviors, animals must send commands from higher-order processing centers in the brain to premotor circuits that reside in ganglia distinct from the brain, such as the mammalian spinal cord or insect ventral nerve cord. How these circuits are functionally organized to generate the great diversity of animal behavior remains unclear. An important first step in unraveling the organization of premotor circuits is to identify their constituent cell types and create tools to monitor and manipulate these with high specificity to assess their functions. This is possible in the tractable ventral nerve cord of the fly. To generate such a toolkit, we used a combinatorial genetic technique (split-GAL4) to create 195 sparse transgenic driver lines targeting 196 individual cell types in the ventral nerve cord. These included wing and haltere motoneurons, modulatory neurons, and interneurons. Using a combination of behavioral, developmental, and anatomical analyses, we systematically characterized the cell types targeted in our collection. In addition, we identified correspondences between the cells in this collection and a recent connectomic data set of the ventral nerve cord. Taken together, the resources and results presented here form a powerful toolkit for future investigations of neuronal circuits and connectivity of premotor circuits while linking them to behavioral outputs.

}, doi = {10.7554/eLife.106548.1}, url = {https://doi.org/10.7554/eLife.106548.1}, author = {Erica Ehrhardt and Samuel C Whitehead and Shigehiro Namiki and Ryo Minegishi and Igor Siwanowicz and Kai Feng and Hideo Otsuna and FlyLight Project Team and Geoffrey W Meissner and David Stern and Jim Truman and David Shepherd and Michael H. Dickinson and Kei Ito and Barry J Dickson and Itai Cohen and Gwyneth M Card and Wyatt Korff} } @article {69995, title = {Skin health and biological aging.}, journal = {Nat Aging}, year = {2025}, month = {2025 Jun 17}, abstract = {

Accumulating evidence indicates that biological aging can be accelerated by environmental exposures, collectively called the {\textquoteright}exposome{\textquoteright}. The skin, as the largest and most exposed organ, can be viewed as a {\textquoteright}window{\textquoteright} for the deep exploration of the exposome and its effects on systemic aging. The complex interplay across hallmarks of aging in the skin and systemic biological aging suggests that physiological processes associated with skin aging influence, and are influenced by, systemic hallmarks of aging. This bidirectional relationship provides potential avenues for the prevention of accelerated biological aging and the identification of therapeutic targets. We provide a review of the interactions between skin exposure, aging hallmarks in the skin and associated systemic changes, and their implications in treatment and disease. We also discuss key questions that need to be addressed to maintain skin and overall health, highlighting the need for the development of precise biomarkers and advanced skin models.

}, issn = {2662-8465}, doi = {10.1038/s43587-025-00901-6}, url = {https://doi.org/10.1038/s43587-025-00901-6}, author = {Furman, David and Auwerx, Johan and Bulteau, Anne-Laure and Church, George and Couturaud, Virginie and Crabbe, Laure and Davies, Kelvin J A and Decottignies, Anabelle and Gladyshev, Vadim N and Kennedy, Brian K and Neretti, Nicola and Nizard, Carine and Pays, Karl and Robinton, Daisy and Sebastiano, Vittorio and Watson, Rachel E B and Wang, Meng C and Woltjen, Knut} } @article {70004, title = {SLAy-ing oversplitting errors in high-density electrophysiology spike sorting}, journal = {bioRxiv}, year = {2025}, month = {2025 Jun 25}, abstract = {

The growing channel count of silicon probes has substantially increased the number of neurons recorded in electrophysiology (ephys) experiments, rendering traditional manual spike sorting impractical. Instead, modern ephys recordings are processed with automated methods that use waveform template matching to isolate putative single neurons. While scalable, automated methods are subject to assumptions that often fail to account for biophysical changes in action potential waveforms, leading to systematic errors. Consequently, manual curation of these errors, which is both time-consuming and lacks reproducibility, remains necessary. To improve efficiency and reproducibility in the spike-sorting pipeline, we introduce here the Spike-sorting Lapse Amelioration System (SLAy), an algorithm that automatically merges oversplit spike clusters. SLAy employs two novel metrics: (1) a waveform similarity metric that uses a neural network to obtain spatially informed, time-shift invariant low-dimensional waveform representations, and (2) a cross-correlogram significance metric based on the earth-movers distance between the observed and null cross-correlograms. We demonstrate that SLAy achieves 85\% agreement with human curators across a diverse set of animal models, brain regions, and probe geometries. To illustrate the impact of spike sorting errors on downstream analyses, we develop a new burst-detection algorithm and show that SLAy fixes spike sorting errors that preclude the accurate detection of bursts in neural data. SLAy leverages GPU parallelization and multithreading for computational efficiency, and is compatible with Phy and NeuroData Without Borders, making it a practical and flexible solution for large-scale ephys data analysis.

}, doi = {10.1101/2025.06.20.660590}, url = {https://www.biorxiv.org/content/early/2025/06/25/2025.06.20.660590}, author = {Koukuntla, Sai and DeWeese, Tate and Cheng, Alexandra and Mildren, Robyn and Lawrence, Aamna and Graves, Austin R and Colonell, Jennifer and Harris, Timothy D and Charles, Adam S} } @article {70005, title = {Sleep need-dependent plasticity of a thalamic circuit promotes homeostatic recovery sleep.}, journal = {Science}, volume = {388}, year = {2025}, month = {2025 Jun 19}, pages = {eadm8203}, abstract = {

Prolonged wakefulness leads to persistent, deep recovery sleep (RS). However, the neuronal circuits that mediate this process remain elusive. From a circuit screen in mice, we identified a group of thalamic nucleus reuniens (RE) neurons activated during sleep deprivation (SD) and required for sleep homeostasis. Optogenetic activation of RE neurons leads to an unusual phenotype: presleep behaviors (grooming and nest organizing) followed by prolonged, intense sleep that resembles RS. Inhibiting RE activity during SD impairs subsequent RS, which suggests that these neurons signal sleep need. RE neurons act upstream of sleep-promoting zona incerta cells, and SD triggers plasticity of this circuit to strengthen their connectivity. These findings reveal a circuit mechanism by which sleep need transforms the functional coupling of a sleep circuit to promote persistent, deep sleep.

}, keywords = {Animals, Homeostasis, Male, Mice, Mice, Inbred C57BL, Midline Thalamic Nuclei, Neuronal Plasticity, Neurons, Optogenetics, Sleep Deprivation, Sleep, Slow-Wave, Wakefulness}, issn = {1095-9203}, doi = {10.1126/science.adm8203}, url = {https://doi.org/10.1126/science.adm8203}, author = {Lee, Sang Soo and Liu, Qiang and Cheng, Alexandra H R and Kim, Dong Won and Boudreau, Daphne M and Mehta, Anuradha and Keles, Mehmet F and Fejfer, Rafal and Palmer, Isabelle and Park, Kristen H and M{\"u}nzberg, Heike and Harris, Timothy D and Graves, Austin R and Blackshaw, Seth and Wu, Mark N} } @article {69984, title = {Spatial reasoning via recurrent neural dynamics in mouse retrosplenial cortex}, journal = {Nat Neurosci}, volume = {28}, year = {2025}, month = {2025 Jun 06}, pages = {1293-1299}, abstract = {

From visual perception to language, sensory stimuli change their meaning depending on previous experience. Recurrent neural dynamics can interpret stimuli based on externally cued context, but it is unknown whether they can compute and employ internal hypotheses to resolve ambiguities. Here we show that mouse retrosplenial cortex (RSC) can form several hypotheses over time and perform spatial reasoning through recurrent dynamics. In our task, mice navigated using ambiguous landmarks that are identified through their mutual spatial relationship, requiring sequential refinement of hypotheses. Neurons in RSC and in artificial neural networks encoded mixtures of hypotheses, location and sensory information, and were constrained by robust low-dimensional dynamics. RSC encoded hypotheses as locations in activity space with divergent trajectories for identical sensory inputs, enabling their correct interpretation. Our results indicate that interactions between internal hypotheses and external sensory data in recurrent circuits can provide a substrate for complex sequential cognitive reasoning.

Preprint: https://doi.org/10.1101/2022.04.12.488024v1

}, keywords = {Animals, Cerebral Cortex, Gyrus Cinguli, Male, Mice, Mice, Inbred C57BL, Models, Neurological, Neural Networks, Computer, Neurons, Photic Stimulation, Space Perception}, issn = {1546-1726}, doi = {10.1038/s41593-025-01944-z}, url = {https://doi.org/10.1038/s41593-025-01944-z}, author = {Voigts, Jakob and Kanitscheider, Ingmar and Miller, Nicholas J and Toloza, Enrique H S and Newman, Jonathan P and Fiete, Ila R and Harnett, Mark T} } @article {69490, title = {Spatial transcriptomics reveals human cortical layer and area specification}, journal = {Nature}, year = {2025}, month = {2025 May 14}, abstract = {

The human cerebral cortex, pivotal for advanced cognitive functions, is composed of six distinct layers and dozens of functionally specialized areas. The layers and areas are distinguished both molecularly, by diverse neuronal and glial cell subtypes, and structurally, through intricate spatial organization3,4. While single-cell transcriptomics studies have advanced molecular characterization of human cortical development, a critical gap exists due to the loss of spatial context during cell dissociation. Here, we utilized multiplexed error-robust fluorescence in situ hybridization (MERFISH)9, augmented with deep-learning-based cell segmentation, to examine the molecular, cellular, and cytoarchitectural development of human fetal cortex with spatially resolved single-cell resolution. Our extensive spatial atlas, encompassing 16 million single cells, spans eight cortical areas across four time points in the second and third trimesters. We uncovered an early establishment of the six-layer structure, identifiable in the laminar distribution of excitatory neuronal subtypes by mid-gestation, long before the emergence of cytoarchitectural layers. Notably, while anterior-posterior gradients of neuronal subtypes were generally observed in most cortical areas, a striking exception was the sharp molecular border between primary (V1) and secondary visual cortices (V2) at gestational week 20. Here we discovered an abrupt binary shift in neuronal subtype specification at the earliest stages, challenging the notion that continuous morphogen gradients dictate mid-gestation cortical arealization. Moreover, integrating single-nuclei RNA-sequencing and in situ whole transcriptomics revealed an early upregulation of synaptogenesis in V1-specific Layer 4 neurons, suggesting a role of synaptogenesis in this discrete border formation. Collectively, our findings underscore the crucial role of spatial relationships in determining the molecular specification of cortical layers and areas. This work not only provides a valuable resource for the field, but also establishes a spatially resolved single-cell analysis paradigm that paves the way for a comprehensive developmental atlas of the human brain.

Preprint:\ https://www.biorxiv.org/content/early/2024/06/10/2024.06.05.597673

}, doi = {10.1038/s41586-025-09010-1}, url = {https://doi.org/10.1038/s41586-025-09010-1}, author = {Qian, Xuyu and Coleman, Kyle and Jiang, Shunzhou and Kriz, Andrea J. and Marciano, Jack H. and Luo, Chunyu and Cai, Chunhui and Manam, Monica Devi and Caglayan, Emre and Otani, Aoi and Ghosh, Urmi and Shao, Diane D. and Andersen, Rebecca E. and Neil, Jennifer E. and Johnson, Robert and LeFevre, Alexandra and Hecht, Jonathan L. and Miller, Michael B. and Sun, Liang and Stringer, Carsen and Li, Mingyao and Walsh, Christopher A.} } @article {69721, title = {A split-GAL4 driver line resource for Drosophila neuron types}, year = {2025}, month = {01/2025}, doi = {10.7554/elife.98405.3}, url = {http://dx.doi.org/10.7554/eLife.98405.3}, author = {Meissner, Geoffrey W and Vannan, Allison and Jeter, Jennifer and Close, Kari and Depasquale, Gina M and Dorman, Zachary and Forster, Kaitlyn and Beringer, Jaye Anne and Gibney, Theresa V and Hausenfluck, Joanna H and He, Yisheng and Henderson, Kristin and Johnson, Lauren and Johnston, Rebecca M and Ihrke, Gudrun and Iyer, Nirmala and Lazarus, Rachel and Lee, Kelley and Li, Hsing-Hsi and Liaw, Hua-Peng and Melton, Brian and Miller, Scott and Motaher, Reeham and Novak, Alexandra and Ogundeyi, Omotara and Petruncio, Alyson and Price, Jacquelyn and Protopapas, Sophia and Tae, Susana and Taylor, Jennifer and Vorimo, Rebecca and Yarbrough, Brianna and Zeng, Kevin Xiankun and Zugates, Christopher T and Dionne, Heather and Angstadt, Claire and Ashley, Kelly and Cavallaro, Amanda and Dang, Tam and Gonzalez, Guillermo A and Hibbard, Karen L and Huang, Cuizhen and Kao, Jui-Chun and Laverty, Todd and Mercer, Monti and Perez, Brenda and Pitts, Scarlett and Ruiz, Danielle and Vallanadu, Viruthika and Zheng, Grace Zhiyu and Goina, Cristian and Otsuna, Hideo and Rokicki, Konrad and Svirskas, Robert R and Cheong, Han Sj and Dolan, Michael-John and Ehrhardt, Erica and Feng, Kai and El Galfi, Basel and Goldammer, Jens and Huston, Stephen J and Hu, Nan and Ito, Masayoshi and McKellar, Claire and minegishi, ryo and Namiki, Shigehiro and Nern, Aljoscha and Schretter, Catherine E and Sterne, Gabriella R and Venkatasubramanian, Lalanti and Wang, Kaiyu and Wolff, Tanya and Wu, Ming and George, Reed and Malkesman, Oz and Aso, Yoshinori and Card, Gwyneth M and Dickson, Barry J and Korff, Wyatt and Ito, Kei and Truman, James W and Zlatic, Marta and Rubin, Gerald M} } @article {70195, title = {Structural dynamics and neural representation of wing deformation.}, journal = {Proc Natl Acad Sci USA}, volume = {122}, year = {2025}, month = {2025 Nov 18}, pages = {e2518032122}, abstract = {

Locomotor control is facilitated by mechanosensory inputs that report how the body interacts with a physical medium. Effective representation of compliant wing deformations is particularly challenging due to the many degrees of freedom. Structural configurations can constrain the stimulus space, and strategic placement of sensors can simplify computation. Here, we measured and modeled wing displacement fields and characterized spatiotemporal encoding of the wing mechanosensors. Our data show how dragonfly wing architecture prescribes deformation modes consistent across models and measurements. We found that the wing{\textquoteright}s state under normal flapping conditions is detected by the spike timing of few sensors, with additional sensors recruited under perturbation. The functional integration of wing biomechanics and sensor placement enables a straightforward solution for information transfer.

}, keywords = {Animals, Biomechanical Phenomena, Flight, Animal, Models, Biological, Odonata, Wings, Animal}, issn = {1091-6490}, doi = {10.1073/pnas.2518032122}, url = {https://doi.org/10.1073/pnas.2518032122}, author = {Yarger, Alexandra M and Maeda, Masateru and Siwanowicz, Igor and Kajiyama, Haruhiro and Walker, Simon M and Bomphrey, Richard J and Lin, Huai-Ti} } @article {69771, title = {Symposia report of The Annual Biological Sciences Section Meeting of the Gerontological Society of America 2023, Tampa, Florida.}, journal = {J Gerontol A Biol Sci Med Sci}, year = {2025}, month = {2025 Feb 11}, abstract = {

The aging process is universal, and it is characterized by a progressive deterioration and decrease in physiological function leading to decline on the organismal level. Nevertheless, a number of genetic and non-genetic interventions have been described, which successfully extend healthspan and lifespan in different species. Furthermore, a number of clinical trials have been evaluating the feasibility of different interventions to promote human health. The goal of the annual Biological Sciences Section of the Gerontological Society of America meeting was to share current knowledge of different topics in aging research and provide a vision of the future of aging research. The meeting gathered international experts in diverse areas of aging research including basic biology, demography, and clinical and translational studies. Specific topics included metabolism, inflammaging, epigenetic clocks, frailty, senescence, neuroscience, stem cells, reproductive aging, inter-organelle crosstalk, comparative transcriptomics of longevity, circadian clock, metabolomics, and biodemography.

}, issn = {1758-535X}, doi = {10.1093/gerona/glaf026}, url = {https://doi.org/10.1093/gerona/glaf026}, author = {Rogina, Blanka and Anderson, Rozalyn and Lebrasseur, Nathan and Curan, Sean and Yousefzadeh, Matthew J and Ghosh, Bhaswati and Duque, Gustavo and Howlett, Susan and Austad, Steven and Demuth, Ilja and Gerstorf, Denis and Korfhage, Justin and Lombard, David B and Abadir, Peter and Christensen, Kaare and Carey, James R and Alberts, Susan C and Campos, Fernando and Palavicini, Juan Pablo and Palmer, Allyson and Bell, Justice and Basisty, Nathan and de Cabo, Rafael and Gomes, Ana and Dixit, Vishwa Deep and Sen, Payel and Baur, Joseph A and Imai, Shin-Ichiro and Li, Xiaoling and Valdez, Gregorio and Orr, Miranda E and Pletcher, Scott and Andersen, Julie and Jones, Leanne and Castillo-Azofeida, David and Bonaguidi, Michael and Suh, Yousin and Duncan, Francesca E and Murray, Anna and Burkewitz, Kristopher and Henne, Mike and Zhou, Kai and Bouhrara, Mustapha and Benjamini, Dan and Kolind, Shannon and Walker, Keenan A and Reiter, David A and Dean Iii, Doug and Gorbunova, Vera and Gladyshev, Vadim N and Palovics, Robert and Niedernhofer, Laura J and Fan, Rong and Bueckle, Andreas D and Hurley, Jennifer and Esser, Karyn A and Kapahi, Pankaj and Sato, Shogo and Jiang, Nisi and Ashiqueali, Sarah A and Diaz, Jose and Mishra, Sidharth Prasad and Ralmundo, Nuno and Banarjee, Reema and Allsopp, Richard and Reynolds, Lindsay M and Zhang, Bohan and Sebastiani, Paola and Monti, Stefano and Schork, Nicholas and Rappaport, Noa} } @article {70085, title = {Time-deterministic cryo-optical microscopy.}, journal = {Light Sci Appl}, volume = {14}, year = {2025}, month = {2025 Aug 23}, pages = {275}, abstract = {

Fluorescence microscopy enables the visualization of cellular morphology, molecular distribution, ion distribution, and their dynamic behaviors during biological processes. Enhancing the signal-to-noise ratio (SNR) in fluorescence imaging improves the quantification accuracy and spatial resolution; however, achieving high SNR at fast image acquisition rates, which is often required to observe cellular dynamics, still remains a challenge. In this study, we developed a technique to rapidly freeze biological cells in milliseconds during optical microscopy observation. Compared to chemical fixation, rapid freezing provides rapid immobilization of samples while more effectively preserving the morphology and conditions of cells. This technique combines the advantages of both live-cell and cryofixation microscopy, i.e., temporal dynamics and high SNR snapshots of selected moments, and is demonstrated by fluorescence and Raman microscopy with high spatial resolution and quantification under low temperature conditions. Furthermore, we also demonstrated that intracellular calcium dynamics can be frozen rapidly and visualized using fluorescent ion indicators, suggesting that ion distribution and conformation of the probe molecules can be fixed both spatially and temporally. These results confirmed that our technique can time-deterministically suspend and visualize cellular dynamics while preserving molecular and ionic states, indicating the potential to provide detailed insights into sample dynamics with improved spatial resolution and temporal accuracy in observations.

}, issn = {2047-7538}, doi = {10.1038/s41377-025-01941-8}, url = {https://doi.org/10.1038/s41377-025-01941-8}, author = {Tsuji, Kosuke and Yamanaka, Masahito and Kumamoto, Yasuaki and Tamura, Shoko and Miyamura, Wakana and Kubo, Toshiki and Mizushima, Kenta and Kono, Kakeru and Hirano, Hanae and Shiozaki, Momoko and Zhao, Xiaowei and Xi, Heqi and Sugiura, Kazunori and Fukushima, Shun-Ichi and Kunimoto, Takumi and Tanabe, Yoshino and Nishida, Kentaro and Mochizuki, Kentaro and Harada, Yoshinori and Smith, Nicholas I and Heintzmann, Rainer and Yu, Zhiheng and Wang, Meng C and Nagai, Takeharu and Tanaka, Hideo and Fujita, Katsumasa} } @article {70053, title = {Transgene-free generation of mouse post-gastrulation whole embryo models solely from naive ESCs and iPSCs.}, journal = {Cell Stem Cell}, year = {2025}, month = {2025 Aug 06}, abstract = {

The generation of post-gastrulation stem cell-derived mouse embryo models (SEMs) exclusively from naive embryonic stem cells (nESCs) has underscored their ability to give rise to embryonic and extra-embryonic lineages. However, existing protocols for mouse SEMs rely on the separate induction of extra-embryonic lineages and on ectopic expression of transcription factors to induce nESC differentiation into trophectoderm (TE) or primitive endoderm (PrE). Here, we demonstrate that mouse nESCs and naive induced pluripotent stem cells (niPSCs) can be simultaneously co-induced, via signaling pathway modulation, to generate PrE and TE extra-embryonic cells that self-organize into embryonic day (E) 8.5-E8.75 transgene-free (TF) SEMs. We also devised an alternative condition (AC) naive media that in vitro stabilizes TF-SEM-competent OCT4+/NANOG+ nESC colonies that co-express antagonistic CDX2 and/or GATA6 extra-embryonic fate master regulators and self-renew while remaining poised for TE and PrE differentiation, respectively. These findings improve mouse SEM strategies and shed light on amplifying an inherent and dormant extra-embryonic plasticity of mouse naive pluripotent cells in vitro.

}, issn = {1875-9777}, doi = {10.1016/j.stem.2025.07.005}, url = {https://doi.org/10.1016/j.stem.2025.07.005}, author = {Yilmaz, Alperen and Gurhan, Gulben and Comar, Mehmet-Yunus and Viukov, Sergey and Serfaty, Inbal and Gayretli, Mert and Golenchenko, Sergey and Lokshtanov, Dmitry and Ashouokhi, Shahd and Polanco, Angel and Berlad, Idan and Ha, Tae-Won and Aguilera-Castrejon, Alejandro and Tarazi, Shadi and Cohen, Marina and Livnat, Nir and Kumar, Komal and Cholakkal, Hisham and Levy, Nathan and Yosef, Nir and Khatib, Nizar and Kakun, Reli Rachel and Kedmi, Merav and Nachman, Inbal Bolocan and Keren-Shaul, Hadas and Addadi, Yoseph and Orenbuch, Ayelet-Hashahar and Korovin, Karina and Molchadsky, Alina and Hochedlinger, Konrad and Gafni, Ohad and Maza, Itay and Novershtern, Noa and Oldak, Bernardo and Hanna, Jacob H} } @article {70133, title = {Ultra-high-density Neuropixels probes improve detection and identification in neuronal recordings.}, journal = {Neuron}, volume = {113}, year = {2025}, month = {2025 Dec 3}, abstract = {

To understand the neural basis of behavior, it is essential to sensitively and accurately measure neural activity at single-neuron and single-spike resolution. Extracellular electrophysiology delivers this, but it has biases in the neurons it detects and it imperfectly resolves their action potentials. To minimize these limitations, we developed a silicon probe with much smaller and denser recording sites than previous designs, called Neuropixels Ultra (NP Ultra). Using NP Ultra, neuronal yield in mouse visual cortex recordings increased by more than 2-fold. With ultra-high spatial resolution, we discovered that a feature of extracellular waveforms, the spatial extent or "footprint," distinguished axonal from somatic recordings. In addition, three genetically identified cortical cell types could be discriminated from one another with \~{}80\% accuracy and from other neurons with \~{}85\% accuracy. NP Ultra improves yield, detection of subcellular compartments, and cell type identification to enable a more powerful dissection of neural circuit activity during behavior.

bioRxiv preprint: doi.org/10.1101/2023.08.23.554527

}, issn = {1097-4199}, doi = {10.1016/j.neuron.2025.08.030}, url = {https://doi.org/10.1016/j.neuron.2025.08.030}, author = {Ye, Zhiwen and Shelton, Andrew M and Shaker, Jordan R and Boussard, Julien and Colonell, Jennifer and Birman, Daniel and Manavi, Sahar and Chen, Susu and Windolf, Charlie and Hurwitz, Cole and Yu, Han and Namima, Tomoyuki and Pedraja, Federico and Weiss, Shahaf and Raducanu, Bogdan C and Ness, Torbj{\o}rn V and Jia, Xiaoxuan and Mastroberardino, Giulia and Rossi, L Federico and Carandini, Matteo and H{\"a}usser, Michael and Einevoll, Gaute T and Laurent, Gilles and Sawtell, Nathaniel B and Bair, Wyeth and Pasupathy, Anitha and Mora Lopez, Carolina and Dutta, Barundeb and Paninski, Liam and Siegle, Joshua H and Koch, Christof and Olsen, Shawn R and Harris, Timothy D and Steinmetz, Nicholas A} } @article {69790, title = {An updated catalogue of split-GAL4 driver lines for descending neurons in Drosophila melanogaster}, journal = {bioRxiv}, year = {2025}, month = {2025 Feb 24}, abstract = {

Descending neurons (DNs) occupy a key position in the sensorimotor hierarchy, conveying signals from the brain to the rest of the body below the neck. In Drosophila melanogaster flies, approximately 480 DN cell types have been described from electron-microscopy image datasets. Genetic access to these cell types is crucial for further investigation of their role in generating behaviour. We previously conducted the first large-scale survey of Drosophila melanogaster DNs, describing 98 unique cell types from light microscopy and generating cell-type-specific split-Gal4 driver lines for 65 of them. Here, we extend our previous work, describing the morphology of 137 additional DN types from light microscopy, bringing the total number DN types identified in light microscopy datasets to 235, or nearly 50\%. In addition, we produced 500 new sparse split-Gal4 driver lines and compiled a list of previously published DN lines from the literature for a combined list of 738 split-Gal4 driver lines targeting 171 DN types.

}, doi = {10.1101/2025.02.22.639679}, url = {https://www.biorxiv.org/content/early/2025/02/24/2025.02.22.639679}, author = {Zung, Jessica L. and Namiki, Shigehiro and Meissner, Geoffrey W. and Costa, Marta and Eichler, Katharina and St{\"u}rner, Tomke and Jefferis, Gregory S.X.E. and Managan, Claire and FlyLight Project Team and Korff, Wyatt and Card, Gwyneth M.} } @article {70123, title = {An updated catalogue of split-GAL4 driver lines for descending neurons in Drosophila melanogaster}, journal = {eLife}, year = {2025}, month = {2025 Sept 19}, abstract = {

Descending neurons (DNs) occupy a key position in the sensorimotor hierarchy, conveying signals from the brain to the rest of the body below the neck. In\ Drosophila melanogaster\ flies, approximately 480 DN cell types have been described from electron-microscopy image datasets. Genetic access to these cell types is crucial for further investigation of their role in generating behaviour. We previously conducted the first large-scale survey of\ Drosophila melanogaster\ DNs, describing 98 unique cell types from light microscopy and generating cell-type-specific split-Gal4 driver lines for 65 of them. Here, we extend our previous work, describing the morphology of 146 additional DN types from light microscopy, bringing the total number DN types identified in light microscopy datasets to 244, or roughly 50\% of all DN types. In addition, we produced 500 new sparse split-Gal4 driver lines and compiled a list of previously published DN lines from the literature for a combined list of 806 split-Gal4 driver lines targeting 190 DN types.

}, doi = {10.7554/elife.107450.1}, url = {http://dx.doi.org/10.7554/eLife.107450.1}, author = {Zung, Jessica L and Namiki, Shigehiro and Meissner, Geoffrey W and Cheong, Han Sj and Costa, Marta and Eichler, Katharina and St{\"u}rner, Tomke and Jefferis, Gregory Sxe and Managan, Claire and Korff, Wyatt and Card, Gwyneth M} } @article {69614, title = {Voltage imaging reveals circuit computations in the raphe underlying serotonin-mediated motor vigor learning}, journal = {Neuron}, year = {2025}, month = {2025 Jun 06}, abstract = {

As animals adapt to new situations, neuromodulation is a potent way to alter behavior, yet mechanisms by which neuromodulatory nuclei compute during behavior are underexplored. The serotonergic raphe supports motor learning in larval zebrafish by visually detecting distance traveled during swims, encoding action effectiveness, and modulating motor vigor. We found that swimming opens a gate for visual input to cause spiking in serotonergic neurons, enabling encoding of action outcomes and filtering out learning-irrelevant visual signals. Using light-sheet microscopy, voltage sensors, and neurotransmitter/modulator sensors, we tracked millisecond-timescale neuronal input-output computations during behavior. Swim commands initially inhibited serotonergic neurons via GABA, closing the gate to spiking. Immediately after, the gate briefly opened: voltage increased consistent with post-inhibitory rebound, allowing swim-induced visual motion to evoke firing through glutamate, triggering serotonin secretion and modulating motor vigor. Ablating GABAergic neurons impaired raphe coding and motor learning. Thus, serotonergic neuromodulation arises from action-outcome coincidence detection within the raphe, suggesting the existence of similarly fast and precise circuit computations across neuromodulatory nuclei.

Preprint:\ https://doi.org/10.1101/2024.09.15.613083

}, doi = {10.1016/j.neuron.2025.05.017}, url = {https://doi.org/10.1016/j.neuron.2025.05.017}, author = {Kawashima, Takashi and Wei, Ziqiang and Haruvi, Ravid and Shainer, Inbal and Narayan, Sujatha and Baier, Herwig and Ahrens, Misha B} } @article {69792, title = {Volumetric imaging of the 3D orientation of cellular structures with a polarized fluorescence light-sheet microscope.}, journal = {Proc Natl Acad Sci }, volume = {122}, year = {2025}, month = {2025 Feb 25}, pages = {e2406679122}, abstract = {

Polarized fluorescence microscopy is a valuable tool for measuring molecular orientations in biological samples, but techniques for recovering three-dimensional orientations and positions of fluorescent ensembles are limited. We report a polarized dual-view light-sheet system for determining the diffraction-limited three-dimensional distribution of the orientations and positions of ensembles of fluorescent dipoles that label biological structures. We share a set of visualization, histogram, and profiling tools for interpreting these positions and orientations. We model the distributions based on the polarization-dependent efficiency of excitation and detection of emitted fluorescence, using coarse-grained representations we call orientation distribution functions (ODFs). We apply ODFs to create physics-informed models of image formation with spatio-angular point-spread and transfer functions. We use theory and experiment to conclude that light-sheet tilting is a necessary part of our design for recovering all three-dimensional orientations. We use our system to extend known two-dimensional results to three dimensions in FM1-43-labeled giant unilamellar vesicles, fast-scarlet-labeled cellulose in xylem cells, and phalloidin-labeled actin in U2OS cells. Additionally, we observe phalloidin-labeled actin in mouse fibroblasts grown on grids of labeled nanowires and identify correlations between local actin alignment and global cell-scale orientation, indicating cellular coordination across length scales.

}, keywords = {Actins, Animals, Humans, Imaging, Three-Dimensional, Mice, Microscopy, Fluorescence, Microscopy, Polarization, Xylem}, issn = {1091-6490}, doi = {10.1073/pnas.2406679122}, author = {Chandler, Talon and Guo, Min and Su, Yijun and Chen, Jiji and Wu, Yicong and Liu, Junyu and Agashe, Atharva and Fischer, Robert S and Mehta, Shalin B and Kumar, Abhishek and Baskin, Tobias I and Jaumouill{\'e}, Valentin and Liu, Huafeng and Swaminathan, Vinay and Nain, Amrinder S and Oldenbourg, Rudolf and La Riviere, Patrick J and Shroff, Hari} } @article {69642, title = {All-optical reporting of inhibitory receptor driving force in the nervous system}, journal = {Nat Commun}, volume = {15}, year = {2024}, month = {2024 Oct 16}, pages = {8913}, abstract = {

Ionic driving forces provide the net electromotive force for ion movement across receptors, channels, and transporters, and are a fundamental property of all cells. In the nervous system, fast synaptic inhibition is mediated by chloride permeable GABA and glycine receptors, and single-cell intracellular recordings have been the only method for estimating driving forces across these receptors (DF). Here we present a tool for quantifying inhibitory receptor driving force named ORCHID: all-Optical Reporting of CHloride Ion Driving force. We demonstrate ORCHID{\textquoteright}s ability to provide accurate, high-throughput measurements of resting and dynamic DF from genetically targeted cell types over multiple timescales. ORCHID confirms theoretical predictions about the biophysical mechanisms that establish DF, reveals differences in DF between neurons and astrocytes, and affords the first in vivo measurements of intact DF. This work extends our understanding of inhibitory synaptic transmission and demonstrates the potential for all-optical methods to assess ionic driving forces.

}, keywords = {Animals, Astrocytes, Chlorides, Humans, Male, Mice, Neurons, Receptors, GABA-A, Receptors, Glycine, Synaptic Transmission}, issn = {2041-1723}, doi = {10.1038/s41467-024-53074-y}, url = {https://doi.org/10.1038/s41467-024-53074-y}, author = {Selfe, Joshua S and Steyn, Teresa J S and Shorer, Eran F and Burman, Richard J and D{\"u}sterwald, Kira M and Kraitzick, Ariel Z and Abdelfattah, Ahmed S and Schreiter, Eric R and Newey, Sarah E and Akerman, Colin J and Raimondo, Joseph V} } @article {69455, title = {Analysis of developmental gene expression using smFISH and in silico staging of C. elegans embryos}, journal = {bioRxiv}, year = {2024}, month = {2024 May 16}, abstract = {

Regulation of transcription during embryogenesis is key to development and differentiation. To study transcript expression throughout Caenorhabditis elegans embryogenesis at single-molecule resolution, we developed a high-throughput single-molecule fluorescence in situ hybridization (smFISH) method that relies on computational methods to developmentally stage embryos and quantify individual mRNA molecules in single embryos. We applied our system to sdc-2, a zygotically transcribed gene essential for hermaphrodite development and dosage compensation. We found that sdc-2 is rapidly activated during early embryogenesis by increasing both the number of mRNAs produced per transcription site and the frequency of sites engaged in transcription. Knockdown of sdc-2 and dpy-27, a subunit of the dosage compensation complex (DCC), increased the number of active transcription sites for the X chromosomal gene dpy-23 but not the autosomal gene mdh-1, suggesting that the DCC reduces the frequency of dpy-23 transcription. The temporal resolution from in silico staging of embryos showed that the deletion of a single DCC recruitment element near the dpy-23 gene causes higher dpy-23 mRNA expression after the start of dosage compensation, which could not be resolved using mRNAseq from mixed-stage embryos. In summary, we have established a computational approach to quantify temporal regulation of transcription throughout C. elegans embryogenesis and demonstrated its potential to provide new insights into developmental gene regulation.

}, doi = {10.1101/2024.05.15.594414}, url = {https://www.biorxiv.org/content/early/2024/05/16/2024.05.15.594414}, author = {Breimann, Laura and Bahry, Ella and Zouinkhi, Marwan and Kolyvanov, Klim and Street, Lena Annika and Preibisch, Stephan and Ercan, Sevin{\c c}} } @article {69713, title = {Analysis of developmental gene expression using smFISH and in silico staging of C. elegans embryos}, journal = {bioRxiv}, year = {2024}, month = {05/2024}, abstract = {

Regulation of transcription during embryogenesis is key to development and differentiation. To study transcript expression throughout Caenorhabditis elegans embryogenesis at single-molecule resolution, we developed a high-throughput single-molecule fluorescence in situ hybridization (smFISH) method that relies on computational methods to developmentally stage embryos and quantify individual mRNA molecules in single embryos. We applied our system to sdc-2, a zygotically transcribed gene essential for hermaphrodite development and dosage compensation. We found that sdc-2 is rapidly activated during early embryogenesis by increasing both the number of mRNAs produced per transcription site and the frequency of sites engaged in transcription. Knockdown of sdc-2 and dpy-27, a subunit of the dosage compensation complex (DCC), increased the number of active transcription sites for the X chromosomal gene dpy-23 but not the autosomal gene mdh-1, suggesting that the DCC reduces the frequency of dpy-23 transcription. The temporal resolution from in silico staging of embryos showed that the deletion of a single DCC recruitment element near the dpy-23 gene causes higher dpy-23 mRNA expression after the start of dosage compensation, which could not be resolved using mRNAseq from mixed-stage embryos. In summary, we have established a computational approach to quantify temporal regulation of transcription throughout C. elegans embryogenesis and demonstrated its potential to provide new insights into developmental gene regulation.Competing Interest StatementThe authors have declared no competing interest.

}, doi = {10.1101/2024.05.15.594414}, url = {https://www.biorxiv.org/content/early/2024/05/16/2024.05.15.594414}, author = {Breimann, Laura and Bahry, Ella and Zouinkhi, Marwan and Kolyvanov, Klim and Street, Lena Annika and Preibisch, Stephan and Ercan, Sevin{\c c}} } @article {69284, title = {Analysis of meiotic recombination in Drosophila simulans shows heterozygous inversions do not cause an interchromosomal effect}, journal = {Genetics}, year = {2024}, month = {2024 May 19}, abstract = {

Chromosome inversions are of unique importance in the evolution of genomes and species because when heterozygous with a standard arrangement chromosome, they suppress meiotic crossovers within the inversion. In Drosophila species, heterozygous inversions also cause the interchromosomal effect, whereby the presence of a heterozygous inversion induces a dramatic increase in crossover frequencies in the remainder of the genome within a single meiosis. To date, the interchromosomal effect has been studied exclusively in species that also have high frequencies of inversions in wild populations. We took advantage of a recently developed approach for generating inversions in Drosophila simulans, a species that does not have inversions in wild populations, to ask if there is an interchromosomal effect. We used the existing chromosome 3R balancer and generated a new chromosome 2L balancer to assay for the interchromosomal effect genetically and cytologically. We found no evidence of an interchromosomal effect in D. simulans. To gain insight into the underlying mechanistic reasons, we qualitatively analyzed the relationship between meiotic double-strand break formation and synaptonemal complex assembly. We find that the synaptonemal complex is assembled prior to double-strand break formation as in D. melanogaster; however, we show that the synaptonemal complex is assembled prior to localization of the oocyte determination factor Orb, whereas in D. melanogaster, synaptonemal complex formation does not begin until Orb is localized. Together, our data show heterozygous inversions in D. simulans do not induce an interchromosomal effect and that there are differences in the developmental programming of the early stages of meiosis.

}, doi = {10.1093/genetics/iyae084}, author = {Bowen Man and Elizabeth Kim and Alekhya Vadlakonda and David L Stern and Nicole Crown} } @article {69657, title = {Anti-seizure effects of norepinephrine-induced free fatty acid release.}, journal = {Cell Metab}, year = {2024}, month = {2024 Oct 29}, abstract = {

The brain{\textquoteright}s ability to rapidly transition between sleep, quiet wakefulness, and states of high vigilance is remarkable. Cerebral norepinephrine (NE) plays a key role in promoting wakefulness, but how does the brain avoid neuronal hyperexcitability upon arousal? Here, we show that NE exposure results in the generation of free fatty acids (FFAs) within the plasma membrane from both astrocytes and neurons. In turn, FFAs dampen excitability by differentially modulating the activity of astrocytic and neuronal Na, K, ATPase. Direct application of FFA to the occipital cortex in awake, behaving mice dampened visual-evoked potential (VEP). Conversely, blocking FFA production via local application of a lipase inhibitor heightened VEP and triggered seizure-like activity. These results suggest that FFA release is a crucial step in NE signaling that safeguards against hyperexcitability. Targeting lipid-signaling pathways may offer a novel therapeutic approach for seizure prevention.

}, issn = {1932-7420}, doi = {10.1016/j.cmet.2024.10.011}, url = {https://doi.org/10.1016/j.cmet.2024.10.011}, author = {Li, Baoman and Sun, Qian and Ding, Fengfei and Xu, Qiwu and Kang, Ning and Xue, Yang and Ladron-de-Guevara, Antonio and Hirase, Hajime and Weikop, Pia and Gong, Sheng and Nathan, Smith and Nedergaard, Maiken} } @article {69641, title = {Assembloid model to study loop circuits of the human nervous system}, journal = {bioRxiv}, year = {2024}, month = {2024 Oct 14}, abstract = {

Neural circuits connecting the cerebral cortex, the basal ganglia and the thalamus are fundamental networks for sensorimotor processing and their dysfunction has been consistently implicated in neuropsychiatric disorders1-9. These recursive, loop circuits have been investigated in animal models and by clinical neuroimaging, however, direct functional access to developing human neurons forming these networks has been limited. Here, we use human pluripotent stem cells to reconstruct an in vitro cortico-striatal-thalamic-cortical circuit by creating a four-part loop assembloid. More specifically, we generate regionalized neural organoids that resemble the key elements of the cortico-striatal-thalamic-cortical circuit, and functionally integrate them into loop assembloids using custom 3D-printed biocompatible wells. Volumetric and mesoscale calcium imaging, as well as extracellular recordings from individual parts of these assembloids reveal the emergence of synchronized patterns of neuronal activity. In addition, a multi{\textendash}step rabies retrograde tracing approach demonstrate the formation of neuronal connectivity across the network in loop assembloids. Lastly, we apply this system to study heterozygous loss of ASH1L gene associated with autism spectrum disorder and Tourette syndrome and discover aberrant synchronized activity in disease model assembloids. Taken together, this human multi-cellular platform will facilitate functional investigations of the cortico-striatal-thalamic-cortical circuit in the context of early human development and in disease conditions.

}, doi = {10.1101/2024.10.13.617729}, url = {https://doi.org/10.1101/2024.10.13.617729}, author = {Miura, Yuki and Kim, Ji-il and Jurjut, Ovidiu and Kelley, Kevin W. and Yang, Xiao and Chen, Xiaoyu and Thete, Mayuri Vijay and Revah, Omer and Cui, Bianxiao and Pachitariu, Marius and Pasca, Sergiu P.} } @article {69454, title = {Astrocyte Calcium Signaling}, journal = {Cold Spring Harb Perspect Biol}, year = {2024}, month = {2024 May 20}, abstract = {

Astrocytes are predominant glial cells that tile the central nervous system and participate in well-established functional and morphological interactions with neurons, blood vessels, and other glia. These ubiquitous cells display rich intracellular Ca signaling, which has now been studied for over 30 years. In this review, we provide a summary and perspective of recent progress concerning the study of astrocyte intracellular Ca signaling as well as discussion of its potential functions. Progress has occurred in the areas of imaging, silencing, activating, and analyzing astrocyte Ca signals. These insights have collectively permitted exploration of the relationships of astrocyte Ca signals to neural circuit function and behavior in a variety of species. We summarize these aspects along with a framework for mechanistically interpreting behavioral studies to identify directly causal effects. We finish by providing a perspective on new avenues of research concerning astrocyte Ca signaling.

}, issn = {1943-0264}, doi = {10.1101/cshperspect.a041353}, author = {Ahrens, Misha B and Khakh, Baljit S and Poskanzer, Kira E} } @article {69147, title = {Believing is seeing - the deceptive influence of bias in quantitative microscopy.}, journal = {Journal of Cell Science}, volume = {137}, year = {2024}, month = {2024 Jan 10}, abstract = {

The visual allure of microscopy makes it an intuitively powerful research tool. Intuition, however, can easily obscure or distort the reality of the information contained in an image. Common cognitive biases, combined with institutional pressures that reward positive research results, can quickly skew a microscopy project towards upholding, rather than rigorously challenging, a hypothesis. The impact of these biases on a variety of research topics is well known. What might be less appreciated are the many forms in which bias can permeate a microscopy experiment. Even well-intentioned researchers are susceptible to bias, which must therefore be actively recognized to be mitigated. Importantly, although image quantification has increasingly become an expectation, ostensibly to confront subtle biases, it is not a guarantee against bias and cannot alone shield an experiment from cognitive distortions. Here, we provide illustrative examples of the insidiously pervasive nature of bias in microscopy experiments - from initial experimental design to image acquisition, analysis and data interpretation. We then provide suggestions that can serve as guard rails against bias.

}, keywords = {Bias, Humans, Microscopy, Research Personnel}, issn = {1477-9137}, doi = {10.1242/jcs.261567}, author = {Lee, Rachel M and Eisenman, Leanna R and Khuon, Satya and Aaron, Jesse S and Chew, Teng-Leong} } @article {69445, title = {Building momentum through networks: Bioimaging across the Americas}, journal = {Journal of Microscopy}, volume = {n/a}, year = {2024}, month = {2024 May 15}, abstract = {

In September 2023, the two largest bioimaging networks in the Americas, Latin America Bioimaging (LABI) and BioImaging North America (BINA), came together during a 1-week meeting in Mexico. This meeting provided opportunities for participants to interact closely with decision-makers from imaging core facilities across the Americas. The meeting was held in a hybrid format and attended in-person by imaging scientists from across the Americas, including Canada, the United States, Mexico, Colombia, Peru, Argentina, Chile, Brazil and Uruguay. The aims of the meeting were to discuss progress achieved over the past year, to foster networking and collaborative efforts among members of both communities, to bring together key members of the international imaging community to promote the exchange of experience and expertise, to engage with industry partners, and to establish future directions within each individual network, as well as common goals. This meeting report summarises the discussions exchanged, the achievements shared, and the goals set during the LABIxBINA2023: Bioimaging across the Americas meeting.

}, keywords = {bioimaging networks, BioImaging North America, community-building, imaging scientists, Latin America Bioimaging}, doi = {https://doi.org/10.1111/jmi.13318}, url = {https://onlinelibrary.wiley.com/doi/abs/10.1111/jmi.13318}, author = {De Niz, Mariana and Escobedo Garc{\'\i}a, Rodrigo and Ter{\'a}n Ramirez, Celina and Pakowski, Ysa and Abonza, Yuriney and Bialy, Nikki and Orr, Vanessa L. and Olivera, Andres and Abonza, Victor and Alleva, Karina and Allodi, Silvana and Almeida, Michael F. and Becerril Cuevas, Alexis Ricardo and Bonnet, Frederic and Burgos Solorio, Armando and Chew, Teng-Leong and Chiabrando, Gustavo and Cimini, Beth and Cleret-Buhot, Aur{\'e}lie and Contreras Jim{\'e}nez, Gast{\'o}n and Daza, Laura and De S{\'a}, Vanessa and De Val, Natalia and Delgado-{\'A}lvarez, Diego L. and Eliceiri, Kevin and Fiolka, Reto and Grecco, Hernan and Hanein, Dorit and Hern{\'a}ndez Herrera, Pa{\'u}l and Hockberger, Phil and Hernandez, Haydee O. and Hernandez Guadarrama, Yael and Itano, Michelle and Jacobs, Caron A. and Jim{\'e}nez-Garc{\'\i}a, Luis F. and Jim{\'e}nez Sabinina, Vilma and Kamaid, Andres and Keppler, Antje and Kumar, Abhishek and Lacoste, Judith and Lovy, Alenka and Luby-Phelps, Kate and Mahadevan-Jansen, Anita and Malacrida, Leonel and Mehta, Shalin B. and Miller, Caroline and Miranda, Kildare and Moore, Joshua A. and North, Alison and O{\textquoteright}Toole, Peter and Olivares Urbano, Mariana and Pietrasanta, L{\'\i}a I. and Portugal, Rodrigo V. and Rossi, Andr{\'e}s H. and Sanchez Contreras, Jonathan and Strambio-De-Castilla, Caterina and Soldevila, Gloria and Vale, Bruno and Vazquez, Diana and Wood, Chris and Brown, Claire M. and Guerrero, Adan} } @article {69289, title = {Carbon Nanomaterial Fluorescent Probes and Their Biological Applications}, journal = {Chemical Reviews}, year = {2024}, month = {2024 Mar 13}, abstract = {

Fluorescent carbon nanomaterials have broadly useful chemical and photophysical attributes that are conducive to applications in biology. In this review, we focus on materials whose photophysics allow for the use of these materials in biomedical and environmental applications, with emphasis on imaging, biosensing, and cargo delivery. The review focuses primarily on graphitic carbon nanomaterials including graphene and its derivatives, carbon nanotubes, as well as carbon dots and carbon nanohoops. Recent advances in and future prospects of these fields are discussed at depth, and where appropriate, references to reviews pertaining to older literature are provided.

}, doi = {10.1021/acs.chemrev.3c00581}, url = {https://doi.org/10.1021/acs.chemrev.3c00581}, author = {Krasley, Andrew T. and Li, Eugene and Galeana, Jesus M. and Bulumulla, Chandima and Beyene, Abraham G. and Demirer, Gozde S.} } @article {69251, title = {The cell-type-specific spatial organization of the anterior thalamic nuclei of the mouse brain.}, journal = {Cell Reports}, volume = {43}, year = {2024}, month = {2024 Feb 29}, pages = {113842}, abstract = {

Understanding the cell-type composition and spatial organization of brain regions is crucial for interpreting brain computation and function. In the thalamus, the anterior thalamic nuclei (ATN) are involved in a wide variety of functions, yet the cell-type composition of the ATN remains unmapped at a single-cell and spatial resolution. Combining single-cell RNA sequencing, spatial transcriptomics, and multiplexed fluorescent in situ hybridization, we identify three discrete excitatory cell-type clusters that correspond to the known nuclei of the ATN and uncover marker genes, molecular pathways, and putative functions of these cell types. We further illustrate graded spatial variation along the dorsomedial-ventrolateral axis for all individual nuclei of the ATN and additionally demonstrate that the anteroventral nucleus exhibits spatially covarying protein products and long-range inputs. Collectively, our study reveals discrete and continuous cell-type organizational principles of the ATN, which will help to guide and interpret experiments on ATN computation and function.

}, issn = {2211-1247}, doi = {10.1016/j.celrep.2024.113842}, author = {Kapustina, Margarita and Zhang, Angela A and Tsai, Jennifer Y J and Bristow, Brianna N and Kraus, Larissa and Sullivan, Kaitlin E and Erwin, Sarah R and Wang, Lihua and Stach, Tara R and Clements, Jody and Lemire, Andrew L and Cembrowski, Mark S} } @article {69548, title = {Cohesin prevents cross-domain gene coactivation.}, journal = {Nat Genet}, year = {2024}, month = {2024 Jul 24}, abstract = {

The contrast between the disruption of genome topology after cohesin loss and the lack of downstream gene expression changes instigates intense debates regarding the structure-function relationship between genome and gene regulation. Here, by analyzing transcriptome and chromatin accessibility at the single-cell level, we discover that, instead of dictating population-wide gene expression levels, cohesin supplies a general function to neutralize stochastic coexpression tendencies of cis-linked genes in single cells. Notably, cohesin loss induces widespread gene coactivation and chromatin co-opening tens of million bases apart in cis. Spatial genome and protein imaging reveals that cohesin prevents gene co-bursting along the chromosome and blocks spatial mixing of transcriptional hubs. Single-molecule imaging shows that cohesin confines the exploration of diverse enhancer and core promoter binding transcriptional regulators. Together, these results support that cohesin arranges nuclear topology to control gene coexpression in single cells.

}, issn = {1546-1718}, doi = {10.1038/s41588-024-01852-1}, url = {https://doi.org/10.1038/s41588-024-01852-1}, author = {Dong, Peng and Zhang, Shu and Gandin, Valentina and Xie, Liangqi and Wang, Lihua and Lemire, Andrew L and Li, Wenhong and Otsuna, Hideo and Kawase, Takashi and Lander, Arthur D and Chang, Howard Y and Liu, Zhe J} } @article {69489, title = {Comparative connectomics of the descending and ascending neurons of the Drosophila nervous system: stereotypy and sexual dimorphism}, journal = {bioRxiv}, year = {2024}, month = {2024 Jun 06}, abstract = {

In most complex nervous systems there is a clear anatomical separation between the nerve cord, which contains most of the final motor outputs necessary for behaviour, and the brain. In insects, the neck connective is both a physical and information bottleneck connecting the brain and the ventral nerve cord (VNC, spinal cord analogue) and comprises diverse populations of descending (DN), ascending (AN) and sensory ascending neurons, which are crucial for sensorimotor signalling and control.Integrating three separate EM datasets, we now provide a complete connectomic description of the ascending and descending neurons of the female nervous system of Drosophila and compare them with neurons of the male nerve cord. Proofread neuronal reconstructions have been matched across hemispheres, datasets and sexes. Crucially, we have also matched 51\% of DN cell types to light level data defining specific driver lines as well as classifying all ascending populations.We use these results to reveal the general architecture, tracts, neuropil innervation and connectivity of neck connective neurons. We observe connected chains of descending and ascending neurons spanning the neck, which may subserve motor sequences. We provide a complete description of sexually dimorphic DN and AN populations, with detailed analysis of circuits implicated in sex-related behaviours, including female ovipositor extrusion (DNp13), male courtship (DNa12/aSP22) and song production (AN hemilineage 08B). Our work represents the first EM-level circuit analyses spanning the entire central nervous system of an adult animal.

}, doi = {10.1101/2024.06.04.596633}, url = {https://www.biorxiv.org/content/early/2024/06/06/2024.06.04.596633}, author = {St{\"u}rner, Tomke and Brooks, Paul and Serratosa Capdevila, Laia and Morris, Billy J. and Javier, Alexandre and Fang, Siqi and Gkantia, Marina and Cachero, Sebastian and Beckett, Isabella R. and Champion, Andrew S. and Moitra, Ilina and Richards, Alana and Klemm, Finja and Kugel, Leonie and Namiki, Shigehiro and Cheong, Han S.J. and Kovalyak, Julie and Tenshaw, Emily and Parekh, Ruchi and Schlegel, Philipp and Phelps, Jasper S. and Mark, Brandon and Dorkenwald, Sven and Bates, Alexander S. and Matsliah, Arie and Yu, Szi-Chieh and McKellar, Claire E. and Sterling, Amy and Seung, Sebastian and Murthy, Mala and Tuthill, John and Lee, Wei-Chung A. and Card, Gwyneth M. and Costa, Marta and Jefferis, Gregory S.X.E. and Eichler, Katharina} } @article {69373, title = {Connectome-driven neural inventory of a complete visual system}, journal = {Nature}, year = {2024}, month = {2025 Mar 26}, abstract = {

Vision provides animals with detailed information about their surroundings, conveying diverse features such as color, form, and movement across the visual scene. Computing these parallel spatial features requires a large and diverse network of neurons, such that in animals as distant as flies and humans, visual regions comprise half the brain{\textquoteright}s volume. These visual brain regions often reveal remarkable structure-function relationships, with neurons organized along spatial maps with shapes that directly relate to their roles in visual processing. To unravel the stunning diversity of a complex visual system, a careful mapping of the neural architecture matched to tools for targeted exploration of that circuitry is essential. Here, we report a new connectome of the right optic lobe from a male Drosophila central nervous system FIB-SEM volume and a comprehensive inventory of the fly{\textquoteright}s visual neurons. We developed a computational framework to quantify the anatomy of visual neurons, establishing a basis for interpreting how their shapes relate to spatial vision. By integrating this analysis with connectivity information, neurotransmitter identity, and expert curation, we classified the 53,000 neurons into 727 types, about half of which are systematically described and named for the first time. Finally, we share an extensive collection of split-GAL4 lines matched to our neuron type catalog. Together, this comprehensive set of tools and data unlock new possibilities for systematic investigations of vision in Drosophila, a foundation for a deeper understanding of sensory processing.

}, doi = {10.1038/s41586-025-08746-0}, url = {https://doi.org/10.1038/s41586-025-08746-0}, author = {Aljoscha Nern and Frank Loesche and Shin-ya Takemura and Laura E Burnett and Marisa Dreher and Eyal Gruntman and Judith Hoeller and Gary B Huang and Michal Januszewski and Nathan C Klapoetke and Sanna Koskela and Kit D Longden and Zhiyuan Lu and Stephan Preibisch and Wei Qiu and Edward M Rogers and Pavithraa Seenivasan and Arthur Zhao and John Bogovic and Brandon S Canino and Jody Clements and Michael Cook and Samantha Finley-May and Miriam A Flynn and Imran Hameed and Kenneth J Hayworth and Gary Patrick Hopkins and Philip M Hubbard and William T Katz and Julie Kovalyak and Shirley A Lauchie and Meghan Leonard and Alanna Lohff and Charli A Maldonado and Caroline Mooney and Nneoma Okeoma and Donald J Olbris and Christopher Ordish and Tyler Paterson and Emily M Phillips and Tobias Pietzsch and Jennifer Rivas Salinas and Patricia K Rivlin and Ashley L Scott and Louis A Scuderi and Satoko Takemura and Iris Talebi and Alexander Thomson and Eric T Trautman and Lowell Umayam and Claire Walsh and John J Walsh and C Shan Xu and Emily A Yakal and Tansy Yang and Ting Zhao and Jan Funke and Reed George and Harald F Hess and Gregory S X E Jefferis and Christopher Knecht and Wyatt Korff and Stephen M Plaza and Sandro Romani and Stephan Saalfeld and Louis K Scheffer and Stuart Berg and Gerald M Rubin and Michael B Reiser} } @article {69138, title = {Connectomic reconstruction predicts visual features used for navigation.}, journal = {Nature}, volume = {634}, year = {2024}, month = {2024 Oct 02}, pages = {181-190}, abstract = {

Many animals use visual information to navigate, but how such information is encoded and integrated by the navigation system remains incompletely understood. In Drosophila melanogaster, EPG neurons in the central complex compute the heading direction by integrating visual input from ER neurons, which are part of the anterior visual pathway (AVP). Here we densely reconstruct all neurons in the AVP using electron-microscopy data. The AVP comprises four neuropils, sequentially linked by three major classes of neurons: MeTu neurons, which connect the medulla in the optic lobe to the small unit of the anterior optic tubercle (AOTUsu) in the central brain; TuBu neurons, which connect the AOTUsu to the bulb neuropil; and ER neurons, which connect the bulb to the EPG neurons. On the basis of morphologies, connectivity between neural classes and the locations of synapses, we identify distinct information channels that originate from four types of MeTu neurons, and we further divide these into ten subtypes according to the presynaptic connections in the medulla and the postsynaptic connections in the AOTUsu. Using the connectivity of the entire AVP and the dendritic fields of the MeTu neurons in the optic lobes, we infer potential visual features and the visual area from which any ER neuron receives input. We confirm some of these predictions physiologically. These results provide a strong foundation for understanding how distinct sensory features can be extracted and transformed across multiple processing stages to construct higher-order cognitive representations.

}, doi = {10.1038/s41586-024-07967-z}, url = {https://doi.org/10.1038/s41586-024-07967-z}, author = {Garner, Dustin and Kind, Emil and Yuet Ha Lai, Jennifer and Nern, Aljoscha and Zhao, Arthur and Houghton, Lucy and Sancer, Gizem and Wolff, Tanya and Rubin, Gerald M and Wernet, Mathias F and Kim, Sung Soo} } @article {69663, title = {A consensus definition for deep layer 6 excitatory neurons in mouse neocortex}, journal = {bioRxiv}, year = {2024}, month = {2024 Nov 05}, abstract = {

To understand neocortical function, we must first define its cell types. Recent studies indicate that neurons in the deepest cortical layer play roles in mediating thalamocortical interactions and modulating brain state and are implicated in neuropsychiatric disease. However, understanding the functions of deep layer 6 (L6b) neurons has been hampered by the lack of agreed upon definitions for these cell types. We compared commonly used methods for defining L6b neurons, including molecular, transcriptional and morphological approaches as well as transgenic mouse lines, and identified a core population of L6b neurons. This population does not innervate sensory thalamus, unlike layer 6 corticothalamic neurons (L6CThNs) in more superficial layer 6. Rather, single L6b neurons project ipsilaterally between cortical areas. Although L6b neurons undergo early developmental changes, we found that their intrinsic electrophysiological properties were stable after the first postnatal week. Our results provide a consensus definition for L6b neurons, enabling comparisons across studies.

}, doi = {10.1101/2024.11.04.621933}, url = {https://doi.org/10.1101/2024.11.04.621933}, author = {Kim, Su-Jeong and Babola, Travis A. and Lee, Kihwan and Matney, Chanel J. and Spiegel, Alina C. and Liew, Michael H. and Schulteis, Eva M. and Coye, Austin E. and Proskurin, Mikhail and Kang, Hyunwook and Kim, Julia A. and Chevee, Maxime and Lee, Kiwoong and Kanold, Patrick O. and Goff, Loyal A. and Kim, Juhyun and Brown, Solange P} } @article {69588, title = {Coordinated cross-brain activity during accumulation of sensory evidence and decision commitment}, journal = {bioRxiv}, year = {2024}, month = {2024 Aug 25}, abstract = {

Cognition is produced by the continuous interactions between many regions across the brain, but has typically been studied one brain region at a time. How signals in different regions coordinate to achieve a single coherent action remains unclear. Here, we address this question by characterizing the simultaneous interactions between up to 20 brain regions across the brain (10 targeted regions per hemisphere), of rats performing the {\textquotedblleft}Poisson Clicks{\textquotedblright} task, a decision-making task that demands the gradual accumulation of momentary evidence. Using 8 Neuropixels probes in each animal, we recorded simultaneously in prefrontal cortex, striatum, motor cortex, hippocampus, amygdala, and thalamus. To assess decision-related interactions between regions, we quantified correlations of each region{\textquoteright}s {\textquotedblleft}decision variable{\textquotedblright}: moment-to-moment co-fluctuations along the axis in neural state space that best predicts the upcoming choice. This revealed a network of strongly correlated brain regions that include the dorsomedial frontal cortex (dmFC), anterior dorsal striatum (ADS), and primary motor cortex (M1), whose decision variables also led the rest of the brain. If coordinated activity within this subnetwork reflects an ongoing evidence accumulation process, these correlations should cease at the time of decision commitment. We therefore compared correlations before versus after {\textquotedblleft}nTc{\textquotedblright}, a recently reported estimator for the time of internal decision commitment. We found that correlations in the decision variables between different brain regions decayed to near-zero after nTc. Additionally, we found that choice-predictive activity steadily increased over time before nTc, but abruptly stopped growing at nTc, consistent with an evidence accumulation process that has stopped evolving at that time. Assessing nTc from the activity of individual regions revealed that nTc could be reliably detected earlier in M1 than other regions. These results show that evidence accumulation involves coordination within a network of frontal cortical and striatal regions, and suggests that termination of this process may initiate in M1.

}, doi = {10.1101/2024.08.21.609044}, url = {https://www.doi.org/10.1101/2024.08.21.609044}, author = {Bondy, Adrian G. and Charlton, Julie A. and Luo, Thomas Zhihao and Kopec, Charles D. and Stagnaro, Wynne M. and Venditto, Sarah Jo C. and Lynch, Laura and Janarthanan, Sanjeev and Oline, Stefan N. and Harris, Timothy D. and Brody, Carlos D.} } @article {69442, title = {Coumarin as a general switching auxiliary to prepare photochromic and spontaneously blinking fluorophores}, journal = {bioRxiv}, year = {2024}, month = {2024 May 12}, abstract = {

Single-molecule localization microscopy (SMLM) uses activatable or switchable fluorophores to create non-diffraction limited maps of molecular location in biological samples. Despite the utility of this imaging technique, the portfolio of appropriate labels for SMLM remains limited. Here, we describe a general strategy for the construction of {\textquotedblleft}glitter bomb{\textquotedblright} labels by simply combining rhodamine and coumarin dyes though an amide bond. Condensation of the ortho-carboxyl group on the pendant phenyl ring of rhodamine dyes with a 7-aminocoumarin yields photochromic or spontaneously blinking fluorophores depending on the parent rhodamine structure. We apply this strategy to prepare labels useful super-resolution experiments in fixed cells using different attachment techniques. This general glitter bomb strategy should lead to improved labels for SMLM, ultimately enabling the creation of detailed molecular maps in biological samples.

}, doi = {10.1101/2024.05.12.593749}, url = {https://www.biorxiv.org/content/early/2024/05/12/2024.05.12.593749}, author = {Jradi, Fadi M. and English, Brian P. and Brown, Timothy A. and Aaron, Jesse and Khuon, Satya and Galbraith, James A. and Galbraith, Catherine G. and Lavis, Luke D.} } @article {69602, title = {Data Release: High-Resolution Imaging and Segmentation of P7 Mouse Tissue Microarchitecture Using FIB-SEM and Machine Learning}, journal = {bioRxiv}, year = {2024}, month = {2024 Sep 07}, abstract = {

This report presents a comprehensive data release exploring the tissue microarchitecture of P7 aged mice using Focused Ion Beam Scanning Electron Microscopy (FIB-SEM) combined with machine learning-based segmentations of nuclei. The study includes high-resolution 3D volumes and nucleus segmentations for seven vital tissues{\textemdash}pancreas, liver, kidney, heart, thymus, hippocampus, and skin{\textemdash}from a single mouse. The detailed datasets are openly accessible on OpenOrganelle.org, providing a valuable resource for the scientific community to support further research and collaboration.

}, doi = {10.1101/2024.09.05.611438}, url = {https://www.doi.org/10.1101/2024.09.05.611438}, author = {Ackerman, David and Avetissian, Emma and Bleck, Christopher K. E. and Bogovic, John A. and Innerberger, Michael and Korff, Wyatt and Li, Wei-Ping and Lu, Zhiyuan and Petruncio, Alyson and Preibisch, Stephan and Qiu, Wei and Jeff L Rhoades and Saalfeld, Stephan and Silva, Malan and Trautman, Eric T. and Vorimo, Rebecca and Weigel, Aubrey and Yu, Zhiheng and Zubov, Yurii and ,} } @article {69693, title = {Development of a novel methodology for ascertaining scientific opinion and extent of agreement.}, journal = {PLoS One}, volume = {19}, year = {2024}, month = {2024 Dec 06}, pages = {e0313541}, abstract = {

We take up the challenge of developing an international network with capacity to survey the world{\textquoteright}s scientists on an ongoing basis, providing rich datasets regarding the opinions of scientists and scientific sub-communities, both at a time and also over time. The novel methodology employed sees local coordinators, at each institution in the network, sending survey invitation emails internally to scientists at their home institution. The emails link to a {\textquoteright}10 second survey{\textquoteright}, where the participant is presented with a single statement to consider, and a standard five-point Likert scale. In June 2023, a group of 30 philosophers and social scientists invited 20,085 scientists across 30 institutions in 12 countries to participate, gathering 6,807 responses to the statement Science has put it beyond reasonable doubt that COVID-19 is caused by a virus. The study demonstrates that it is possible to establish a global network to quickly ascertain scientific opinion on a large international scale, with high response rate, low opt-out rate, and in a way that allows for significant (perhaps indefinite) repeatability. Measuring scientific opinion in this new way would be a valuable complement to currently available approaches, potentially informing policy decisions and public understanding across diverse fields.

}, keywords = {COVID-19, Humans, Public Opinion, SARS-CoV-2, Science, Surveys and Questionnaires}, issn = {1932-6203}, doi = {10.1371/journal.pone.0313541}, author = {Vickers, Peter and Adamo, Ludovica and Alfano, Mark and Clark, Cory and Cresto, Eleonora and Cui, He and Dang, Haixin and Dells{\'e}n, Finnur and Dupin, Nathalie and Gradowski, Laura and Graf, Simon and Guevara, Aline and Hallap, Mark and Hamilton, Jesse and Hardey, Mariann and Helm, Paula and Landrum, Asheley and Levy, Neil and Machery, Edouard and Mills, Sarah and Muller, Se{\'a}n and Sheppard, Joanne and N K, Shinod and Slater, Matthew and Stegenga, Jacob and Strandin, Henning and Stuart, Michael T and Sweet, David and Tasdan, Ufuk and Taylor, Henry and Towler, Owen and Tulodziecki, Dana and Tworek, Heidi and Wallbank, Rebecca and Wiltsche, Harald and Mitchell Finnigan, Samantha} } @article {69710, title = {Discovery of neuronal cell types by pairing whole cell reconstructions with RNA expression profiles}, journal = {bioRxiv}, year = {2024}, month = {12/2024}, abstract = {

Effective classification of neuronal cell types requires both molecular and morphological descriptors to be collected in situ at single cell resolution. However, current spatial transcriptomics techniques are not compatible with imaging workflows that successfully reconstruct the morphology of complete axonal projections. Here, we introduce a new methodology that combines tissue clearing, submicron whole-brain two photon imaging, and Expansion-Assisted Iterative Fluorescence In Situ Hybridization (EASI-FISH) to assign molecular identities to fully reconstructed neurons in the mouse brain, which we call morphoFISH. We used morphoFISH to molecularly identify a previously unknown population of cingulate neurons projecting ipsilaterally to the dorsal striatum and contralaterally to higher-order thalamus. By pairing whole-brain morphometry, improved techniques for nucleic acid preservation and spatial gene expression, morphoFISH offers a quantitative solution for discovery of multimodal cell types and complements existing techniques for characterization of increasingly fine-grained cellular heterogeneity in brain circuits.Competing Interest StatementThe authors have declared no competing interest.

}, doi = {10.1101/2024.12.30.630829}, url = {https://www.biorxiv.org/content/early/2024/12/31/2024.12.30.630829}, author = {The MouseLight Project Team and Ferreira, Tiago A. and Eddison, Mark and Copeland, Monique and Lay, Mary and Tenshaw, Emily and Weldon, Monet and Schauder, David and Olbris, Donald J. and Rokicki, Konrad and Spruston, Nelson and Tillberg, Paul W. and Korff, Wyatt and Dudman, Joshua T.} } @article {69633, title = {A Drosophila computational brain model reveals sensorimotor processing.}, journal = {Nature}, volume = {634}, year = {2024}, month = {2024 Oct 02}, pages = {210-219}, abstract = {

The recent assembly of the adult Drosophila melanogaster central brain connectome, containing more than 125,000 neurons and 50 million synaptic connections, provides a template for examining sensory processing throughout the brain. Here we create a leaky integrate-and-fire computational model of the entire Drosophila brain, on the basis of neural connectivity and neurotransmitter identity, to study circuit properties of feeding and grooming behaviours. We show that activation of sugar-sensing or water-sensing gustatory neurons in the computational model accurately predicts neurons that respond to tastes and are required for feeding initiation. In addition, using the model to activate neurons in the feeding region of the Drosophila brain predicts those that elicit motor neuron firing-a testable hypothesis that we validate by optogenetic activation and behavioural studies. Activating different classes of gustatory neurons in the model makes accurate predictions of how several taste modalities interact, providing circuit-level insight into aversive and appetitive taste processing. Additionally, we applied this model to mechanosensory circuits and found that computational activation of mechanosensory neurons predicts activation of a small set of neurons comprising the antennal grooming circuit, and accurately describes the circuit response upon activation of different mechanosensory subtypes. Our results demonstrate that modelling brain circuits using only synapse-level connectivity and predicted neurotransmitter identity generates experimentally testable hypotheses and can describe complete sensorimotor transformations.

}, keywords = {Animals, Brain, Computer Simulation, Connectome, Drosophila melanogaster, Feeding Behavior, Female, Grooming, Male, Models, Neurological, Motor Neurons, Optogenetics, Synapses, Taste}, issn = {1476-4687}, doi = {10.1038/s41586-024-07763-9}, url = {https://doi.org/10.1038/s41586-024-07763-9}, author = {Shiu, Philip K and Sterne, Gabriella R and Spiller, Nico and Franconville, Romain and Sandoval, Andrea and Zhou, Joie and Simha, Neha and Kang, Chan Hyuk and Yu, Seongbong and Kim, Jinseop S and Dorkenwald, Sven and Matsliah, Arie and Schlegel, Philipp and Yu, Szi-Chieh and McKellar, Claire E and Sterling, Amy and Costa, Marta and Eichler, Katharina and Bates, Alexander Shakeel and Eckstein, Nils and Funke, Jan and Jefferis, Gregory S X E and Murthy, Mala and Bidaye, Salil S and Hampel, Stefanie and Seeds, Andrew M and Scott, Kristin} } @article {68830, title = {Dynamic 1D Search and Processive Nucleosome Translocations by RSC and ISW2 Chromatin Remodelers}, journal = {eLife}, year = {2024}, month = {2024 Mar 18}, abstract = {

Eukaryotic gene expression is linked to chromatin structure and nucleosome positioning by ATP-dependent chromatin remodelers that establish and maintain nucleosome-depleted regions (NDRs) near transcription start sites. Conserved yeast RSC and ISW2 remodelers exert antagonistic effects on nucleosomes flanking NDRs, but the temporal dynamics of remodeler search, engagement, and directional nucleosome mobilization for promoter accessibility are unknown. Using optical tweezers and two-color single-particle imaging, we investigated the Brownian diffusion of RSC and ISW2 on free DNA and sparse nucleosome arrays. RSC and ISW2 rapidly scan DNA by one-dimensional hopping and sliding, respectively, with dynamic collisions between remodelers followed by recoil or apparent co-diffusion. Static nucleosomes block remodeler diffusion resulting in remodeler recoil or sequestration. Remarkably, both RSC and ISW2 use ATP hydrolysis to translocate mono-nucleosomes processively at ~30 bp/s on extended linear DNA under tension. Processivity and opposing push-pull directionalities of nucleosome translocation shown by RSC and ISW2 shape the distinctive landscape of promoter chromatin.

}, doi = {10.7554/eLife.91433}, url = {https://doi.org/10.7554/eLife.91433}, author = {Jee Min Kim and Claudia C. Carcamo and Sina Jazani and Zepei Xie and Xinyu A. Feng and Matthew Poyton and Katie L. Holland and Jonathan B. Grimm and Luke D. Lavis and Taekjip Ha and Carl Wu} } @article {69544, title = {Encoding of cerebellar dentate neuronal activity during visual attention in rhesus macaques}, journal = {bioRxiv}, year = {2024}, month = {2024 Jul 20}, abstract = {

The role of cerebellum in controlling eye movements is well established, but its contribution to more complex forms of visual behavior has remained elusive. To study cerebellar activity during visual attention we recorded extracellular activity of dentate nucleus (DN) neurons in two non-human primates (NHPs). NHPs were trained to read the direction indicated by a peripheral visual stimulus while maintaining fixation at the center, and report the direction of the cue by performing a saccadic eye movement into the same direction following a delay. We found that single unit DN neurons modulated spiking activity over the entire time-course of the task, and that their activity often bridged temporally separated intra-trial events, yet in a heterogeneous manner. To better understand the heterogeneous relationship between task structure, behavioral performance and neural dynamics, we constructed a behavioral, an encoding and a decoding model. Both NHPs showed different behavioral strategies, which influenced the performance. Activity of the DN neurons reflected the unique strategies, with the direction of the visual stimulus frequently being encoded long before an upcoming saccade. Retrograde labeling of the recording location indicated that these neurons receive predominantly inputs from Purkinje cells in the lateral cerebellum as well as neurons of the principal olive and medial pons, all regions known to connect with neurons in the prefrontal cortex contributing to planning of saccades. Together, our results highlight that DN neurons can dynamically modulate their activity during a visual attention task, comprising not only sensorimotor but also cognitive attentional components.

}, doi = {10.1101/2024.07.18.604119}, url = {https://www.doi.org/10.1101/2024.07.18.604119}, author = {Flierman, Nico A. and Koay, Sue Ann and van Hoogstraten, Willem S. and Ruigrok, Tom J.H. and Roelfsema, Pieter R. and Badura, Aleksandra and De Zeeuw, Chris I.} } @article {69145, title = {Epigenetic priming of embryonic lineages in the mammalian epiblast}, journal = {bioRxiv}, year = {2024}, month = {2024 Jan 11}, abstract = {

Understanding the diversification of mammalian cell lineages is an essential to embryonic development, organ regeneration and tissue engineering. Shortly after implantation in the uterus, the pluripotent cells of the mammalian epiblast generate the three germ layers: ectoderm, mesoderm and endoderm1. Although clonal analyses suggest early specification of epiblast cells towards particular cell lineages2{\textendash}4, single-cell transcriptomes do not identify lineage-specific markers in the epiblast5{\textendash}11\ and thus, the molecular regulation of such specification remains unknow. Here, we studied the epigenetic landscape of single epiblast cells, which revealed lineage priming towards endoderm, ectoderm or mesoderm. Unexpectedly, epiblast cells with mesodermal priming show a strong signature for the endothelial/endocardial fate, suggesting early specification of this lineage aside from other mesoderm. Through clonal analysis and live imaging, we show that endothelial precursors show early lineage divergence from the rest of mesodermal derivatives. In particular, cardiomyocytes and endocardial cells show limited lineage relationship, despite being temporally and spatially co-recruited during gastrulation. Furthermore, analysing the live tracks of single cells through unsupervised classification of cell migratory activity, we found early behavioral divergence of endothelial precursors shortly after the onset of mesoderm migration towards the cardiogenic area. These results provide a new model for the phenotypically silent specification of mammalian cell lineages in pluripotent cells of the epiblast and modify current knowledge on the sequence and timing of cardiovascular lineages diversification.

}, doi = {10.1101/2024.01.11.575188}, url = {https://doi.org/10.1101/2024.01.11.575188}, author = {Miquel Sendra and Katie McDole and Daniel Jimenez-Carretero and Juan de Dios Hourcade and Susana Temi{\~n}o and L{\'e}o Guignard and Philipp J Keller and F{\'a}tima S{\'a}nchez-Cabo and Jorge N. Dom{\'\i}nguez and Miguel Torres} } @article {69154, title = {Failure to mate enhances investment in behaviors that may promote mating reward and impairs the ability to cope with stressors via a subpopulation of Neuropeptide F receptor neurons.}, journal = {PLoS Genetics}, volume = {20}, year = {2024}, month = {2024 Jan 18}, pages = {e1011054}, abstract = {

Living in dynamic environments such as the social domain, where interaction with others determines the reproductive success of individuals, requires the ability to recognize opportunities to obtain natural rewards and cope with challenges that are associated with achieving them. As such, actions that promote survival and reproduction are reinforced by the brain reward system, whereas coping with the challenges associated with obtaining these rewards is mediated by stress-response pathways, the activation of which can impair health and shorten lifespan. While much research has been devoted to understanding mechanisms underlying the way by which natural rewards are processed by the reward system, less attention has been given to the consequences of failure to obtain a desirable reward. As a model system to study the impact of failure to obtain a natural reward, we used the well-established courtship suppression paradigm in Drosophila melanogaster as means to induce repeated failures to obtain sexual reward in male flies. We discovered that beyond the known reduction in courtship actions caused by interaction with non-receptive females, repeated failures to mate induce a stress response characterized by persistent motivation to obtain the sexual reward, reduced male-male social interaction, and enhanced aggression. This frustrative-like state caused by the conflict between high motivation to obtain sexual reward and the inability to fulfill their mating drive impairs the capacity of rejected males to tolerate stressors such as starvation and oxidative stress. We further show that sensitivity to starvation and enhanced social arousal is mediated by the disinhibition of a small population of neurons that express receptors for the fly homologue of neuropeptide Y. Our findings demonstrate for the first time the existence of social stress in flies and offers a framework to study mechanisms underlying the crosstalk between reward, stress, and reproduction in a simple nervous system that is highly amenable to genetic manipulation.

}, keywords = {Animals, Drosophila melanogaster, Female, Humans, Male, Neurons, Neuropeptides, Reproduction, Reward, Sexual Behavior, Animal}, issn = {1553-7404}, doi = {10.1371/journal.pgen.1011054}, author = {Ryvkin, Julia and Omesi, Liora and Kim, Yong-Kyu and Levi, Mali and Pozeilov, Hadar and Barak-Buchris, Lital and Agranovich, Bella and Abramovich, Ifat and Gottlieb, Eyal and Jacob, Avi and N{\"a}ssel, Dick R and Heberlein, Ulrike and Shohat-Ophir, Galit} } @article {69649, title = {FITM2 deficiency results in ER lipid accumulation, ER stress, and reduced apolipoprotein B lipidation and VLDL triglyceride secretion in vitro and in mouse liver.}, journal = {Mol Metab}, year = {2024}, month = {2024 Oct 18}, pages = {102048}, abstract = {

OBJECTIVES: Triglycerides (TGs) associate with apolipoprotein B100 (apoB100) to form very low density lipoproteins (VLDLs) in the liver. The repertoire of factors that facilitate this association is incompletely understood. FITM2, an integral endoplasmic reticulum (ER) protein, was originally discovered as a factor participating in cytosolic lipid droplet (LD) biogenesis in tissues that do not form VLDL. We hypothesized that in the liver, in addition to promoting cytosolic LD formation, FITM2 would also transfer TG from its site of synthesis in the ER membrane to nascent VLDL particles within the ER lumen.

METHODS: Experiments were conducted using a rat hepatic cell line (McArdle-RH7777, or McA cells), an established model of mammalian lipoprotein metabolism, and mice. FITM2 expression was reduced using siRNA in cells and by liver specific cre-recombinase mediated deletion of the Fitm2 gene in mice. Effects of FITM2 deficiency on VLDL assembly and secretion in\ vitro and in\ vivo were measured by multiple methods, including density gradient ultracentrifugation, chromatography, mass spectrometry, stimulated Raman scattering (SRS) microscopy, sub-cellular fractionation, immunoprecipitation, immunofluorescence, and electron microscopy.

MAIN FINDINGS: 1) FITM2-deficient hepatic cells in\ vitro and in\ vivo secrete TG-depleted VLDL particles, but the number of particles is unchanged compared to controls; 2) FITM2 deficiency in mice on a high fat diet (HFD) results in decreased plasma TG levels. The number of apoB100-containing lipoproteins remains similar, but shift from VLDL to low density lipoprotein (LDL) density; 3) Both in\ vitro and in\ vivo, when TG synthesis is stimulated and FITM2 is deficient, TG accumulates in the ER, and despite its availability this pool is unable to fully lipidate apoB100 particles; 4) FITM2 deficiency disrupts ER morphology and results in ER stress.

PRINCIPAL CONCLUSIONS: The results suggest that FITM2 contributes to VLDL lipidation, especially when newly synthesized hepatic TG is in abundance. In addition to its fundamental importance in VLDL assembly, the results also suggest that under dysmetabolic conditions, FITM2 may be an important factor in the partitioning of TG between cytosolic LDs and VLDL particles.

}, issn = {2212-8778}, doi = {10.1016/j.molmet.2024.102048}, url = {https://doi.org/10.1016/j.molmet.2024.102048}, author = {Wang, Haizhen and Nikain, Cyrus and Fortounas, Konstantinos I and Amengual, Jaime and Tufanli, Ozlem and La Forest, Maxwell and Yu, Yong and Wang, Meng C and Watts, Russell and Lehner, Richard and Qiu, Yunping and Cai, Min and Kurland, Irwin J and Goldberg, Ira J and Rajan, Sujith and Hussain, M Mahmood and Brodsky, Jeffrey L and Fisher, Edward A} } @article {69108, title = {A fluorogenic complementation tool kit for interrogating lipid droplet-organelle interaction}, journal = {J. Cell Biol.}, volume = {223}, year = {2024}, month = {2024 Jul 01}, pages = {e202311126}, abstract = {

Contact sites between lipid droplets and other organelles are essential for cellular lipid and energy homeostasis upon metabolic demands. Detection of these contact sites at the nanometer scale over time in living cells is challenging. We developed a tool kit for detecting contact sites based on fluorogen-activated bimolecular complementation at CONtact sites, FABCON, using a reversible, low-affinity split fluorescent protein, splitFAST. FABCON labels contact sites with minimal perturbation to organelle interaction. Via FABCON, we quantitatively demonstrated that endoplasmic reticulum (ER)- and mitochondria (mito)-lipid droplet contact sites are dynamic foci in distinct metabolic conditions, such as during lipid droplet biogenesis and consumption. An automated analysis pipeline further classified individual contact sites into distinct subgroups based on size, likely reflecting differential regulation and function. Moreover, FABCON is generalizable to visualize a repertoire of organelle contact sites including ER-mito. Altogether, FABCON reveals insights into the dynamic regulation of lipid droplet-organelle contact sites and generates new hypotheses for further mechanistical interrogation during metabolic regulation.

}, doi = {10.1083/jcb.202311126 }, url = {https://doi.org/10.1083/jcb.202311126 }, author = {Xiao Li and Rico Gamuyao and Ming-Lun Wu and Woo Jung Cho and Nathan B. Kurtz and Sharon V. King and R.A. Petersen and Daniel R. Stabley and Caleb Lindow and Leslie Climer and Abbas Shirinifard and Francesca Ferrara and Robert E. Throm and Camenzind G. Robinson and Alex Carisey and Alison G. Tebo and Chi-Lun Chang} } @article {69658, title = {The Fly Disco: Hardware and software for optogenetics and fine-grained fly behavior analysis}, journal = {bioRxiv}, year = {2024}, month = {2024 Nov 04}, abstract = {

In the fruit fly, Drosophila melanogaster, connectome data and genetic tools provide a unique opportunity to study complex behaviors including navigation, mating, aggression, and grooming in an organism with a tractable nervous system of 140,000 neurons. Here we present the Fly Disco, a flexible system for high quality video collection, optogenetic manipulation, and fine-grained behavioral analysis of freely walking and socializing fruit fly groups. The data collection hardware and software automates the collection of videos synced to programmable optogenetic stimuli. Key pipeline features include behavioral analysis based on trajectories of 21 keypoints and optogenetic-specific summary statistics and data visualization. We created the multifly dataset for pose estimation that includes 9701 examples enriched in complex behaviors. All hardware designs, software, and the multifly dataset are freely available.

}, doi = {10.1101/2024.11.04.621948}, url = {https://doi.org/10.1101/2024.11.04.621948}, author = {Robie, Alice A and Taylor, Adam L and Schretter, Catherine E and Kabra, Mayank and Branson, Kristin} } @article {69320, title = {Focal adhesions contain three specialized actin nanoscale layers}, journal = {Nature Communications}, volume = {15}, year = {2024}, month = {2024 Mar 21}, abstract = {

Focal adhesions (FAs) connect inner workings of cell to the extracellular matrix to control cell adhesion, migration and mechanosensing. Previous studies demonstrated that FAs contain three vertical layers, which connect extracellular matrix to the cytoskeleton. By using super-resolution iPALM microscopy, we identify two additional nanoscale layers within FAs, specified by actin filaments bound to tropomyosin isoforms Tpm1.6 and Tpm3.2. The Tpm1.6-actin filaments, beneath the previously identified α-actinin cross-linked actin filaments, appear critical for adhesion maturation and controlled cell motility, whereas the adjacent Tpm3.2-actin filament layer beneath seems to facilitate adhesion disassembly. Mechanistically, Tpm3.2 stabilizes ACF-7/MACF1 and KANK-family proteins at adhesions, and hence targets microtubule plus-ends to FAs to catalyse their disassembly. Tpm3.2 depletion leads to disorganized microtubule network, abnormally stable FAs, and defects in tail retraction during migration. Thus, FAs are composed of distinct actin filament layers, and each may have specific roles in coupling adhesions to the cytoskeleton, or in controlling adhesion dynamics.

}, doi = {10.1038/s41467-024-46868-7}, url = {https://www.nature.com/articles/s41467-024-46868-7https://www.nature.com/articles/s41467-024-46868-7.pdfhttps://www.nature.com/articles/s41467-024-46868-7.pdfhttps://www.nature.com/articles/s41467-024-46868-7}, author = {Kumari, Reena and Ven, Katharina and Chastney, Megan and Kokate, Shrikant B. and Per{\"a}nen, Johan and Aaron, Jesse and Kogan, Konstantin and Almeida-Souza, Leonardo and Kremneva, Elena and Poincloux, Renaud and Chew, Teng-Leong and Gunning, Peter W. and Ivaska, Johanna and Lappalainen, Pekka} } @article {69680, title = {Global Neuron Shape Reasoning with Point Affinity Transformers}, journal = {bioRxiv}, year = {2024}, month = {2024 Nov 24}, abstract = {

Connectomics is a subfield of neuroscience that aims to map the brain{\textquoteright}s intricate wiring diagram. Accurate neuron segmentation from microscopy volumes is essential for automating connectome reconstruction. However, current state-of-the-art algorithms use image-based convolutional neural networks that are limited to local neuron shape context. Thus, we introduce a new framework that reasons over global neuron shape with a novel point affinity transformer. Our framework embeds a (multi-)neuron point cloud into a fixed-length feature set from which we can decode any point pair affinities, enabling clustering neuron point clouds for automatic proofreading. We also show that the learned feature set can easily be mapped to a contrastive embedding space that enables neuron type classification using a simple KNN classifier. Our approach excels in two demanding connectomics tasks: proofreading segmentation errors and classifying neuron types. Evaluated on three benchmark datasets derived from state-of-the-art connectomes, our method outperforms point transformers, graph neural networks, and unsupervised clustering baselines.

}, doi = {10.1101/2024.11.24.625067}, url = {https://doi.org/10.1101/2024.11.24.625067}, author = {Troidl, Jakob and Knittel, Johannes and Li, Wanhua and Zhan, Fangneng and Pfister, Hanspeter and Turaga, Srinivas} } @article {69254, title = {Haploidy-linked cell proliferation defects limit larval growth in Zebrafish}, journal = {Open Biol.}, volume = {14}, year = {2024}, month = {2024 Oct 09}, pages = {240126}, abstract = {

Haploid larvae in non-mammalian vertebrates are lethal, with characteristic organ growth retardation collectively called {\textquoteright}haploid syndrome{\textquoteright}. In contrast to mammals, whose haploid intolerance is attributed to imprinting misregulation, the cellular principle of haploidy-linked defects in non-mammalian vertebrates remains unknown. Here, we investigated cellular defects that disrupt the ontogeny of gynogenetic haploid zebrafish larvae. Unlike diploid control larvae, haploid larvae manifested unscheduled cell death at the organogenesis stage, attributed to haploidy-linked p53 upregulation. Moreover, we found that haploid larvae specifically suffered the gradual aggravation of mitotic spindle monopolarization during 1-3 days post-fertilization, causing spindle assembly checkpoint-mediated mitotic arrest throughout the entire body. High-resolution imaging revealed that this mitotic defect accompanied the haploidy-linked centrosome loss occurring concomitantly with the gradual decrease in larval cell size. Either resolution of mitotic arrest or depletion of p53 partially improved organ growth in haploid larvae. Based on these results, we propose that haploidy-linked mitotic defects and cell death are parts of critical cellular causes shared among vertebrates that limit the larval growth in the haploid state, contributing to an evolutionary constraint on allowable ploidy status in the vertebrate life cycle.

}, doi = {10.1098/rsob.240126}, url = {https://doi.org/10.1098/rsob.240126}, author = {Kan Yaguchi and Daiki Saito and Triveni Menon and Akira Matsura and Takeomi Mizutani and Tomoya Kotani and Sreelaja Nair and Ryota Uehara} } @article {69250, title = {High-Performance Genetically Encoded Green Fluorescent Biosensors for Intracellular l-Lactate.}, journal = {ACS Central Science}, volume = {10}, year = {2024}, month = {2024 Feb 28}, pages = {402-416}, abstract = {

l-Lactate is a monocarboxylate produced during the process of cellular glycolysis and has long generally been considered a waste product. However, studies in recent decades have provided new perspectives on the physiological roles of l-lactate as a major energy substrate and a signaling molecule. To enable further investigations of the physiological roles of l-lactate, we have developed a series of high-performance (Δ/ = 15 to 30 ), intensiometric, genetically encoded green fluorescent protein (GFP)-based intracellular l-lactate biosensors with a range of affinities. We evaluated these biosensors in cultured cells and demonstrated their application in an preparation of brain tissue. Using these biosensors, we were able to detect glycolytic oscillations, which we analyzed and mathematically modeled.

}, issn = {2374-7943}, doi = {10.1021/acscentsci.3c01250}, author = {Hario, Saaya and Le, Giang N T and Sugimoto, Hikaru and Takahashi-Yamashiro, Kei and Nishinami, Suguru and Toda, Hirofumi and Li, Selene and Marvin, Jonathan S and Kuroda, Shinya and Drobizhev, Mikhail and Terai, Takuya and Nasu, Yusuke and Campbell, Robert E} } @article {68831, title = {Host ZCCHC3 blocks HIV-1 infection and production by a dual mechanism}, journal = {iScience}, year = {2024}, month = {05/2024}, abstract = {

Most mammalian cells prevent viral infection and proliferation by expressing various restriction factors and sensors that activate the immune system. While anti-human immunodeficiency virus type 1 (HIV-1) host restriction factors have been identified, most of them are antagonized by viral proteins. This has severely hindered their development in anti-HIV-1 therapy. Here, we describe CCHC-type zinc-finger-containing protein 3 (ZCCHC3) as a novel anti-HIV-1 factor that is not antagonized by viral proteins. ZCCHC3 suppresses production of HIV-1 and other retroviruses. We show that ZCCHC3 acts by binding to Gag nucleocapsid protein via zinc-finger motifs. This prevents interaction between the Gag nucleocapsid protein and viral genome and results in production of genome-deficient virions. ZCCHC3 also binds to the long terminal repeat on the viral genome via the middle-folded domain, sequestering the viral genome to P-bodies, which leads to decreased viral replication and production. Such a dual antiviral mechanism is distinct from that of any other known host restriction factors. Therefore, ZCCHC3 is a novel potential target in anti-HIV-1 therapy.

}, doi = {10.1101/2023.06.14.544911}, url = {https://doi.org/10.1101/2023.06.14.544911}, author = {Binbin Yi and Yuri L Tanaka and Hidetaka Kosako and Erika P Butlertanaka and Prabuddha Sengupta and Jennifer Lippincott-Schwartz and Akatsuki Saito and Shige H. Yoshimura} } @article {68969, title = {iATPSnFR2: a high dynamic range fluorescent sensor for monitoring intracellular ATP}, journal = {Proc Natl Acad Sci U S A}, year = {2024}, month = {2024 May 21}, abstract = {

We developed a significantly improved genetically encoded quantitative adenosine triphosphate (ATP) sensor to provide real-time dynamics of ATP levels in subcellular compartments. iATPSnFR2 is a variant of iATPSnFR1, a previously developed sensor that has circularly permuted super-folder GFP inserted between the ATP-binding helices of the\ ε-subunit of a bacterial F0-F1\ ATPase. Optimizing the linkers joining the two domains resulted in a \~{} 5-6 fold improvement in the dynamic range compared to the previous generation sensor, with excellent discrimination against other analytes and affinity variants varying from 4 μM to 500 μM. A chimeric version of this sensor fused to either the HaloTag protein or a suitably spectrally separated fluorescent protein, provides a ratiometric readout allowing comparisons of ATP across cellular regions. Subcellular targeting of the sensor to nerve terminals reveals previously uncharacterized single synapse metabolic signatures, while targeting to the mitochondrial matrix allowed direct quantitative probing of oxidative phosphorylation dynamics.

}, doi = {10.1073/pnas.2314604121 }, author = {Jonathan S. Marvin and Alexandros C. Kokotos and Mukesh Kumar and Camila Pulido and Ariana N. Tkachuk and Jocelyn Shuxin Yao and Timothy A. Brown and Timothy A. Ryan} } @article {69142, title = {Image processing tools for petabyte-scale light sheet microscopy data.}, journal = {Nat. Methods}, year = {2024}, month = {2024 Oct 17}, abstract = {

Light sheet microscopy is a powerful technique for high-speed three-dimensional imaging of subcellular dynamics and large biological specimens. However, it often generates datasets ranging from hundreds of gigabytes to petabytes in size for a single experiment. Conventional computational tools process such images far slower than the time to acquire them and often fail outright due to memory limitations. To address these challenges, we present PetaKit5D, a scalable software solution for efficient petabyte-scale light sheet image processing. This software incorporates a suite of commonly used processing tools that are optimized for memory and performance. Notable advancements include rapid image readers and writers, fast and memory-efficient geometric transformations, high-performance Richardson-Lucy deconvolution and scalable Zarr-based stitching. These features outperform state-of-the-art methods by over one order of magnitude, enabling the processing of petabyte-scale image data at the full teravoxel rates of modern imaging cameras. The software opens new avenues for biological discoveries through large-scale imaging experiments.

}, doi = {10.1038/s41592-024-02475-4}, url = {https://doi.org/10.1038/s41592-024-02475-4}, author = {Xiongtao Ruan and Matthew Mueller and Gaoxiang Liu and Frederik G{\"o}rlitz and Tian-Ming Fu and Daniel E. Milkie and Joshua Lillvis and Alison Killilea and Eric Betzig and Srigokul Upadhyayula} } @article {69440, title = {Imaging the extracellular matrix in live tissues and organisms with a glycan-binding fluorophore}, journal = {bioRxiv}, year = {2024}, month = {2024 May 10}, abstract = {

All multicellular systems produce and dynamically regulate extracellular matrices (ECM) that play important roles in both biochemical and mechanical signaling. Though the spatial arrangement of these extracellular assemblies is critical to their biological functions, visualization of ECM structure is challenging, in part because the biomolecules that compose the ECM are difficult to fluorescently label individually and collectively. Here, we present a cell-impermeable small molecule fluorophore, termed Rhobo6, that turns on and red shifts upon reversible binding to glycans. Given that most ECM components are densely glycosylated, the dye enables wash-free visualization of ECM, in systems ranging from in vitro substrates to in vivo mouse mammary tumors. Relative to existing techniques, Rhobo6 provides a broad substrate profile, superior tissue penetration, nonperturbative labeling, and negligible photobleaching. This work establishes a straightforward method for imaging the distribution of ECM in live tissues and organisms, lowering barriers for investigation of extracellular biology.

}, doi = {10.1101/2024.05.09.593460}, url = {https://www.biorxiv.org/content/early/2024/05/10/2024.05.09.593460}, author = {Fiore, Antonio and Yu, Guoqiang and Northey, Jason J. and Patel, Ronak and Ravenscroft, Thomas A. and Ikegami, Richard and Kolkman, Wiert and Kumar, Pratik and Grimm, Jonathan B. and Dilan, Tanya L. and Ruetten, Virginia M.S. and Ahrens, Misha B. and Shroff, Hari and Lavis, Luke D. and Wang, Shaohe and Weaver, Valerie M. and Pedram, Kayvon} } @article {69215, title = {Integrating across behaviors and timescales to understand the neural control of movement.}, journal = {Current Opinion in Neurobiology}, volume = {85}, year = {2024}, month = {2024 Feb 13}, pages = {102843}, abstract = {

The nervous system evolved to enable navigation throughout the environment in the pursuit of resources. Evolutionarily newer structures allowed increasingly complex adaptations but necessarily added redundancy. A dominant view of movement neuroscientists is that there is a one-to-one mapping between brain region and function. However, recent experimental data is hard to reconcile with the most conservative interpretation of this framework, suggesting a degree of functional redundancy during the performance of well-learned, constrained behaviors. This apparent redundancy likely stems from the bidirectional interactions between the various cortical and subcortical structures involved in motor control. We posit that these bidirectional connections enable flexible interactions across structures that change depending upon behavioral demands, such as during acquisition, execution or adaptation of a skill. Observing the system across both multiple actions and behavioral timescales can help isolate the functional contributions of individual structures, leading to an integrated understanding of the neural control of movement.

}, issn = {1873-6882}, doi = {10.1016/j.conb.2024.102843}, author = {Gmaz, Jimmie M and Keller, Jason A and Dudman, Joshua T and Gallego, Juan A} } @article {69643, title = {IRE1α silences dsRNA to prevent taxane-induced pyroptosis in triple-negative breast cancer.}, journal = {Cell}, year = {2024}, month = {2024 Oct 14}, abstract = {

Chemotherapy is often combined with immune checkpoint inhibitor (ICIs) to enhance immunotherapy responses. Despite the approval of chemo-immunotherapy in multiple human cancers, many immunologically cold tumors remain unresponsive. The mechanisms determining the immunogenicity of chemotherapy are elusive. Here, we identify the ER stress sensor IRE1α as a critical checkpoint that restricts the immunostimulatory effects of taxane chemotherapy and prevents the innate immune recognition of immunologically cold triple-negative breast cancer (TNBC). IRE1α RNase silences taxane-induced double-stranded RNA (dsRNA) through regulated IRE1-dependent decay (RIDD) to prevent NLRP3 inflammasome-dependent pyroptosis. Inhibition of IRE1α in Trp53 TNBC allows taxane to induce extensive dsRNAs that are sensed by ZBP1, which in turn activates NLRP3-GSDMD-mediated pyroptosis. Consequently, IRE1α RNase inhibitor plus taxane converts PD-L1-negative, ICI-unresponsive TNBC tumors into PD-L1 immunogenic tumors that are hyper-sensitive to ICI. We reveal IRE1α as a cancer cell defense mechanism that prevents taxane-induced danger signal accumulation and pyroptotic cell death.

}, issn = {1097-4172}, doi = {10.1016/j.cell.2024.09.032}, url = {https://doi.org/10.1016/j.cell.2024.09.032}, author = {Xu, Longyong and Peng, Fanglue and Luo, Qin and Ding, Yao and Yuan, Fei and Zheng, Liting and He, Wei and Zhang, Sophie S and Fu, Xin and Liu, Jin and Mutlu, Ayse Sena and Wang, Shuyue and Nehring, Ralf Bernd and Li, Xingyu and Tang, Qianzi and Li, Catherine and Lv, Xiangdong and Dobrolecki, Lacey E and Zhang, Weijie and Han, Dong and Zhao, Na and Jaehnig, Eric and Wang, Jingyi and Wu, Weiche and Graham, Davis A and Li, Yumei and Chen, Rui and Peng, Weiyi and Chen, Yiwen and Catic, Andre and Zhang, Zhibin and Zhang, Bing and Mustoe, Anthony M and Koong, Albert C and Miles, George and Lewis, Michael T and Wang, Meng C and Rosenberg, Susan M and O{\textquoteright}Malley, Bert W and Westbrook, Thomas F and Xu, Han and Zhang, Xiang H-F and Osborne, C Kent and Li, Jin Billy and Ellis, Matthew J and Rimawi, Mothaffar F and Rosen, Jeffrey M and Chen, Xi} } @article {69372, title = {Leptin Activated Hypothalamic BNC2 Neurons Acutely Suppress Food Intake}, journal = {bioRxiv}, year = {2024}, month = {12 Apr 2024}, abstract = {

Leptin is an adipose tissue hormone that maintains homeostatic control of adipose tissue mass by regulating the activity of specific neural populations controlling appetite and metabolism1. Leptin regulates food intake by inhibiting orexigenic agouti-related protein (AGRP) neurons and activating anorexigenic pro-opiomelanocortin (POMC) neurons2. However, while AGRP neurons regulate food intake on a rapid time scale, acute activation of POMC neurons has only a minimal effect3{\textendash}5. This has raised the possibility that there is a heretofore unidentified leptin-regulated neural population that suppresses appetite on a rapid time scale. Here, we report the discovery of a novel population of leptin-target neurons expressing basonuclin 2 (Bnc2) that acutely suppress appetite by directly inhibiting AGRP neurons. Opposite to the effect of AGRP activation, BNC2 neuronal activation elicited a place preference indicative of positive valence in hungry but not fed mice. The activity of BNC2 neurons is finely tuned by leptin, sensory food cues, and nutritional status. Finally, deleting leptin receptors in BNC2 neurons caused marked hyperphagia and obesity, similar to that observed in a leptin receptor knockout in AGRP neurons. These data indicate that BNC2-expressing neurons are a key component of the neural circuit that maintains energy balance, thus filling an important gap in our understanding of the regulation of food intake and leptin action.

}, doi = {10.1101/2024.01.25.577315}, url = {https://www.biorxiv.org/content/early/2024/04/12/2024.01.25.577315}, author = {Han L. Tan and Luping Yin and Yuqi Tan and Jessica Ivanov and Kaja Plucinska and Anoj Ilanges and Brian R. Herb and Putianqi Wang and Christin Kosse and Paul Cohen and Dayu Lin and Jeffrey M. Friedman} } @article {68956, title = {Magnetic voluntary head-fixation in transgenic rats enables lifetime imaging of hippocampal neurons}, journal = {Nat. Commun.}, year = {2024}, month = {2024 May 16}, abstract = {

The precise neural mechanisms within the brain that contribute to the remarkable lifetime persistence of memory remain unknown. Existing techniques to record neurons in animals are either unsuitable for longitudinal recording from the same cells or make it difficult for animals to express their full naturalistic behavioral repertoire. We present a magnetic voluntary head-fixation system that provides stable optical access to the brain during complex behavior. Compared to previous systems that used mechanical restraint, there are no moving parts and animals can engage and disengage entirely at will. This system is failsafe, easy for animals to use and reliable enough to allow long-term experiments to be routinely performed. Together with a novel two-photon fluorescence collection scheme that increases two-photon signal and a transgenic rat line that stably expresses the calcium sensor GCaMP6f in dorsal CA1, we are able to track and record activity from the same hippocampal neurons, during behavior, over a large fraction of animals{\textquoteright} lives.

}, doi = {10.1038/s41467-024-48505-9}, author = {P. D. Rich and S. Y. Thiberge and B. B. Scott and C. Guo and D. G. Tervo and C. D. Brody and A. Y. Karpova and N. D. Daw and D. W. Tank} } @article {69171, title = {Mapping of multiple neurotransmitter receptor subtypes and distinct protein complexes to the connectome.}, journal = {Neuron}, year = {2024}, month = {2024 Jan 24}, abstract = {

Neurons express various combinations of neurotransmitter receptor (NR) subunits and receive inputs from multiple neuron types expressing different neurotransmitters. Localizing NR subunits to specific synaptic inputs has been challenging. Here, we use epitope-tagged endogenous NR subunits, expansion light-sheet microscopy, and electron microscopy (EM) connectomics to molecularly characterize synapses in Drosophila. We show that in directionally selective motion-sensitive neurons, different multiple NRs elaborated a highly stereotyped molecular topography with NR localized to specific domains receiving cell-type-specific inputs. Developmental studies suggested that NRs or complexes of them with other membrane proteins determine patterns of synaptic inputs. In support of this model, we identify a transmembrane protein selectively associated with a subset of spatially restricted synapses and demonstrate its requirement for synapse formation through genetic analysis. We propose that mechanisms that regulate the precise spatial distribution of NRs provide a molecular cartography specifying the patterns of synaptic connections onto dendrites.

}, issn = {1097-4199}, doi = {10.1016/j.neuron.2023.12.014}, author = {Sanfilippo, Piero and Kim, Alexander J and Bhukel, Anuradha and Yoo, Juyoun and Mirshahidi, Pegah S and Pandey, Vijaya and Bevir, Harry and Yuen, Ashley and Mirshahidi, Parmis S and Guo, Peiyi and Li, Hong-Sheng and Wohlschlegel, James A and Aso, Yoshinori and Zipursky, S Lawrence} } @article {69247, title = {MarShie: a clearing protocol for 3D analysis of single cells throughout the bone marrow at subcellular resolution.}, journal = {Nature Communincations}, volume = {15}, year = {2024}, month = {2024 Feb 26}, pages = {1764}, abstract = {

Analyzing immune cell interactions in the bone marrow is vital for understanding hematopoiesis and bone homeostasis. Three-dimensional analysis of the complete, intact bone marrow within the cortex of whole long bones remains a challenge, especially at subcellular resolution. We present a method that stabilizes the marrow and provides subcellular resolution of fluorescent signals throughout the murine femur, enabling identification and spatial characterization of hematopoietic and stromal cell subsets. By combining a pre-processing algorithm for stripe artifact removal with a machine-learning approach, we demonstrate reliable cell segmentation down to the deepest bone marrow regions. This reveals age-related changes in the marrow. It highlights the interaction between CXCR1 cells and the vascular system in homeostasis, in contrast to other myeloid cell types, and reveals their spatial characteristics after injury. The broad applicability of this method will contribute to a better understanding of bone marrow biology.

}, keywords = {Animals, Bone Marrow, Bone Marrow Cells, Hematopoiesis, Hematopoietic Stem Cells, Mice, Stromal Cells}, issn = {2041-1723}, doi = {10.1038/s41467-024-45827-6}, author = {Mertens, Till Fabian and Liebheit, Alina Tabea and Ehl, Johanna and K{\"o}hler, Ralf and Rakhymzhan, Asylkhan and Woehler, Andrew and Katth{\"a}n, Lukas and Ebel, Gernot and Liublin, Wjatscheslaw and Kasapi, Ana and Triantafyllopoulou, Antigoni and Schulz, Tim Julius and Niesner, Raluca Aura and Hauser, Anja Erika} } @article {69484, title = {Mechanosensory representation of wing deformations}, journal = {bioRxiv}, year = {2024}, month = {2024 Jun 02}, abstract = {

Efficient representation of structural deformations is crucial for monitoring the instantaneous state of biological structures. Insects{\textquoteright} ability to encode wing deformations during flight demonstrates a general morphological computing principle applicable across sensory systems in nature as well as engineered systems. To characterize how relevant features are encoded, we measured and modelled displacement and strain across dragonfly wing surfaces in tethered and free flight. Functional interpretations were supported by neuroanatomical maps, and ablation and perturbation experiments. We find that signal redundancy is reduced by non-random sensor distributions and that morphology limits the stimulus space such that sensory systems can monitor natural states with few sensors. Deviations from the natural states are detected by a flexible population of additional sensors with many distinguishable activation patterns.

}, doi = {10.1101/2024.06.02.596338}, url = {https://www.biorxiv.org/content/early/2024/06/02/2024.06.02.596338}, author = {Yarger, Alexandra M and Maeda, Masateru and Siwanowicz, Igor and Kajiyama, Haruhiro and Walker, Simon M and Bomphrey, Richard J and Lin, Huai-Ti} } @article {69622, title = {A modular chemigenetic calcium indicator for multiplexed in vivo functional imaging.}, journal = {Nat Methods}, year = {2024}, month = {2024 Sep 20}, abstract = {

Genetically encoded fluorescent calcium indicators allow cellular-resolution recording of physiology. However, bright, genetically targetable indicators that can be multiplexed with existing tools in vivo are needed for simultaneous imaging of multiple signals. Here we describe WHaloCaMP, a modular chemigenetic calcium indicator built from bright dye-ligands and protein sensor domains. Fluorescence change in WHaloCaMP results from reversible quenching of the bound dye via a strategically placed tryptophan. WHaloCaMP is compatible with rhodamine dye-ligands that fluoresce from green to near-infrared, including several that efficiently label the brain in animals. When bound to a near-infrared dye-ligand, WHaloCaMP shows a 7{\texttimes} increase in fluorescence intensity and a 2.1-ns increase in fluorescence lifetime upon calcium binding. We use WHaloCaMP1a to image Ca responses in vivo in flies and mice, to perform three-color multiplexed functional imaging of hundreds of neurons and astrocytes in zebrafish larvae and to quantify Ca concentration using fluorescence lifetime imaging microscopy (FLIM).

}, issn = {1548-7105}, doi = {10.1038/s41592-024-02411-6}, url = {https://doi.org/10.1038/s41592-024-02411-6}, author = {Farrants, Helen and Shuai, Yichun and Lemon, William C and Monroy Hernandez, Christian and Zhang, Deng and Yang, Shang and Patel, Ronak and Qiao, Guanda and Frei, Michelle S and Plutkis, Sarah E and Grimm, Jonathan B and Hanson, Timothy L and Tomaska, Filip and Turner, Glenn C and Stringer, Carsen and Keller, Philipp J and Beyene, Abraham G and Chen, Yao and Liang, Yajie and Lavis, Luke D and Schreiter, Eric R} } @article {69301, title = {Motor neurons generate pose-targeted movements via proprioceptive sculpting.}, journal = {Nature}, year = {2024}, month = {2024 Mar 20}, abstract = {

Motor neurons are the final common pathway through which the brain controls movement of the body, forming the basic elements from which all movement is composed. Yet how a single motor neuron contributes to control during natural movement remains unclear. Here we anatomically and functionally characterize the individual roles of the motor neurons that control head movement in the fly, Drosophila melanogaster. Counterintuitively, we find that activity in a single motor neuron rotates the head in different directions, depending on the starting posture of the head, such that the head converges towards a pose determined by the identity of the stimulated motor neuron. A feedback model predicts that this convergent behaviour results from motor neuron drive interacting with proprioceptive feedback. We identify and genetically suppress a single class of proprioceptive neuron that changes the motor neuron-induced convergence as predicted by the feedback model. These data suggest a framework for how the brain controls movements: instead of directly generating movement in a given direction by activating a fixed set of motor neurons, the brain controls movements by adding bias to a continuing proprioceptive-motor loop.

}, issn = {1476-4687}, doi = {10.1038/s41586-024-07222-5}, author = {Gorko, Benjamin and Siwanowicz, Igor and Close, Kari and Christoforou, Christina and Hibbard, Karen L and Kabra, Mayank and Lee, Allen and Park, Jin-Yong and Li, Si Ying and Chen, Alex B and Namiki, Shigehiro and Chen, Chenghao and Tuthill, John C and Bock, Davi D and Rouault, Herv{\'e} and Branson, Kristin and Ihrke, Gudrun and Huston, Stephen J} } @article {69172, title = {A neural circuit architecture for rapid learning in goal-directed navigation}, journal = {Neuron}, volume = {112}, year = {2024}, month = {2024 May 23}, pages = {2581-2599.e23}, abstract = {

Anchoring goals to spatial representations enables flexible navigation but is challenging in novel environments when both representations must be acquired simultaneously. We propose a framework for how Drosophila uses internal representations of head direction (HD) to build goal representations upon selective thermal reinforcement. We show that flies use stochastically generated fixations and directed saccades to express heading preferences in an operant visual learning paradigm and that HD neurons are required to modify these preferences based on reinforcement. We used a symmetric visual setting to expose how flies{\textquoteright} HD and goal representations co-evolve and how the reliability of these interacting representations impacts behavior. Finally, we describe how rapid learning of new goal headings may rest on a behavioral policy whose parameters are flexible but whose form is genetically encoded in circuit architecture. Such evolutionarily structured architectures, which enable rapidly adaptive behavior driven by internal representations, may be relevant across species.

}, doi = {https://doi.org/10.1016/j.neuron.2024.04.036}, url = {https://doi.org/10.1016/j.neuron.2024.04.036}, author = {Chuntao Dan and Brad K. Hulse and Ramya Kappagantula and Vivek Jayaraman and Ann M. Hermundstad} } @article {69045, title = {Neural-circuit basis of song preference learning in fruit flies}, journal = {iScience}, volume = {27}, year = {2024}, month = {2024 Jul 19}, abstract = {

As observed in human language learning and song learning in birds, the fruit fly Drosophila melanogaster changes its auditory behaviors according to prior sound experiences. This phenomenon, known as song preference learning in flies, requires GABAergic input to pC1 neurons in the brain, with these neurons playing a key role in mating behavior. The neural circuit basis of this GABAergic input, however, is not known. Here, we find that GABAergic neurons expressing the sex-determination gene doublesex are necessary for song preference learning. In the brain, only four doublesex-expressing GABAergic neurons exist per hemibrain, identified as pCd-2 neurons. pCd-2 neurons directly, and in many cases mutually, connect with pC1 neurons, suggesting the existence of reciprocal circuits between them. Moreover, GABAergic and dopaminergic inputs to doublesex-expressing GABAergic neurons are necessary for song preference learning. Together, this study provides a neural circuit model that underlies experience-dependent auditory plasticity at a single-cell resolution.

}, doi = {10.1016/j.isci.2024.110266}, url = {https://doi.org/10.1016/j.isci.2024.110266}, author = {Keisuke Imoto and Yuki Ishikawa and Yoshinori Aso and Jan Funke and Ryoya Tanaka and Azusa Kamikouchi} } @article {68889, title = {Neuronal wiring diagram of an adult brain.}, journal = {Nature}, volume = {634}, year = {2024}, month = {2024 Oct 02}, pages = {124-138}, abstract = {

Connections between neurons can be mapped by acquiring and analysing electron microscopic brain images. In recent years, this approach has been applied to chunks of brains to reconstruct local connectivity maps that are highly informative, but nevertheless inadequate for understanding brain function more globally. Here we present a neuronal wiring diagram of a whole brain containing 5 {\texttimes} 107 chemical synapses between 139,255 neurons reconstructed from an adult female Drosophila melanogaster. The resource also incorporates annotations of cell classes and types, nerves, hemilineages and predictions of neurotransmitter identities. Data products are available for download, programmatic access and interactive browsing and have been made interoperable with other fly data resources. We derive a projectome-a map of projections between regions-from the connectome and report on tracing of synaptic pathways and the analysis of information flow from inputs (sensory and ascending neurons) to outputs (motor, endocrine and descending neurons) across both hemispheres and between the central brain and the optic lobes. Tracing from a subset of photoreceptors to descending motor pathways illustrates how structure can uncover putative circuit mechanisms underlying sensorimotor behaviours. The technologies and open ecosystem reported here set the stage for future large-scale connectome projects in other species.

}, doi = {10.1038/s41586-024-07558-y}, url = {https://doi.org/10.1038/s41586-024-07558-y}, author = {Dorkenwald, Sven and Matsliah, Arie and Sterling, Amy R and Schlegel, Philipp and Yu, Szi-Chieh and McKellar, Claire E and Lin, Albert and Costa, Marta and Eichler, Katharina and Yin, Yijie and Silversmith, Will and Schneider-Mizell, Casey and Jordan, Chris S and Brittain, Derrick and Halageri, Akhilesh and Kuehner, Kai and Ogedengbe, Oluwaseun and Morey, Ryan and Gager, Jay and Kruk, Krzysztof and Perlman, Eric and Yang, Runzhe and Deutsch, David and Bland, Doug and Sorek, Marissa and Lu, Ran and Macrina, Thomas and Lee, Kisuk and Bae, J Alexander and Mu, Shang and Nehoran, Barak and Mitchell, Eric and Popovych, Sergiy and Wu, Jingpeng and Jia, Zhen and Castro, Manuel and Kemnitz, Nico and Ih, Dodam and Bates, Alexander Shakeel and Eckstein, Nils and Funke, Jan and Collman, Forrest and Bock, Davi D and Jefferis, Gregory S X E and Seung, H Sebastian and Murthy, Mala} } @article {68258, title = {NeuronBridge: an intuitive web application for neuronal morphology search across large data sets}, journal = {BMC Bioinformatics}, volume = {25}, year = {2024}, month = {2024 Mar 15}, pages = {114}, abstract = {

Background

Neuroscience research in\ Drosophila\ is benefiting from large-scale connectomics efforts using electron microscopy (EM) to reveal all the neurons in a brain and their connections. To exploit this knowledge base, researchers relate a connectome{\textquoteright}s structure to neuronal function, often by studying individual neuron cell types. Vast libraries of fly driver lines expressing fluorescent reporter genes in sets of neurons have been created and imaged using confocal light microscopy (LM), enabling the targeting of neurons for experimentation. However, creating a fly line for driving gene expression within a single neuron found in an EM connectome remains a challenge, as it typically requires identifying a pair of driver lines where only the neuron of interest is expressed in both. This task and other emerging scientific workflows require finding similar neurons across large data sets imaged using different modalities.

Results

Here, we present NeuronBridge, a web application for easily and rapidly finding putative morphological matches between large data sets of neurons imaged using different modalities. We describe the functionality and construction of the NeuronBridge service, including its user-friendly graphical user interface (GUI), extensible data model, serverless cloud architecture, and massively parallel image search engine.

Conclusions

NeuronBridge fills a critical gap in the\ Drosophila\ research workflow and is used by hundreds of neuroscience researchers around the world. We offer our software code, open APIs, and processed data sets for integration and reuse, and provide the application as a service at\ http://neuronbridge.janelia.org.

}, doi = {10.1186/s12859-024-05732-7}, url = {https://bmcbioinformatics.biomedcentral.com/articles/10.1186/s12859-024-05732-7}, author = {Jody Clements and Cristian Goina and Philip M. Hubbard and Takashi Kawase and Donald J. Olbris and Hideo Otsuna and Robert Svirskas and Konrad Rokicki} } @article {69325, title = {Patch-walking: Coordinated multi-pipette patch clamp for efficiently finding synaptic connections}, journal = {eLife}, volume = {13}, year = {2024}, month = {2024 Nov 18}, pages = {RP97399}, abstract = {

Significant technical challenges exist when measuring synaptic connections between neurons in living brain tissue. The patch clamping technique, when used to probe for synaptic connections, is manually laborious and time-consuming. To improve its efficiency, we pursued another approach: instead of retracting all patch clamping electrodes after each recording attempt, we cleaned just one of them and reused it to obtain another recording while maintaining the others. With one new patch clamp recording attempt, many new connections can be probed. By placing one pipette in front of the others in this way, one can {\textquoteright}walk{\textquoteright} across the mouse brain slice, termed {\textquoteright}patch-walking.{\textquoteright} We performed 136 patch clamp attempts for two pipettes, achieving 71 successful whole cell recordings (52.2\%). Of these, we probed 29 pairs (i.e. 58 bidirectional probed connections) averaging 91 μm intersomatic distance, finding three connections. Patch-walking yields 80-92\% more probed connections, for experiments with 10-100 cells than the traditional synaptic connection searching method.

}, doi = {10.7554/elife.97399}, url = {https://doi.org/10.7554/elife.97399}, author = {Mighten C. Yip and Mercedes M. Gonzalez and Colby F. Lewallen and Corey R. Landry and Ilya Kolb and Bo Yang and William M. Stoy and Ming-fai Fong and Matthew JM Rowan and Edward S. Boyden and Craig R. Forest} } @article {69704, title = {Periodic ER-plasma membrane junctions support long-range Ca signal integration in dendrites.}, journal = {Cell}, year = {2024}, month = {2024 Dec 18}, abstract = {

Neuronal dendrites must relay synaptic inputs over long distances, but the mechanisms by which activity-evoked intracellular signals propagate over macroscopic distances remain unclear. Here, we discovered a system of periodically arranged endoplasmic reticulum-plasma membrane (ER-PM) junctions tiling the plasma membrane of dendrites at \~{}1\ μm intervals, interlinked by a meshwork of ER tubules patterned in a ladder-like array. Populated with Junctophilin-linked plasma membrane voltage-gated Ca channels and ER Ca-release channels (ryanodine receptors), ER-PM junctions are hubs for ER-PM crosstalk, fine-tuning of Ca homeostasis, and local activation of the Ca/calmodulin-dependent protein kinase II. Local spine stimulation activates the Ca modulatory machinery, facilitating signal transmission and ryanodine-receptor-dependent Ca release at ER-PM junctions over 20\ μm away. Thus, interconnected ER-PM junctions support signal propagation and Ca release from the spine-adjacent ER. The capacity of this subcellular architecture to modify both local and distant membrane-proximal biochemistry potentially contributes to dendritic computations.

}, issn = {1097-4172}, doi = {10.1016/j.cell.2024.11.029}, author = {Benedetti, Lorena and Fan, Ruolin and Weigel, Aubrey V and Moore, Andrew S and Houlihan, Patrick R and Kittisopikul, Mark and Park, Grace and Petruncio, Alyson and Hubbard, Philip M and Pang, Song and Xu, C Shan and Hess, Harald F and Saalfeld, Stephan and Rangaraju, Vidhya and Clapham, David E and De Camilli, Pietro and Ryan, Timothy A and Lippincott-Schwartz, Jennifer} } @article {69139, title = {Petascale pipeline for precise alignment of images from serial section electron microscopy.}, journal = {Nature Communications}, volume = {15}, year = {2024}, month = {2024 Jan 04}, pages = {289}, abstract = {

The reconstruction of neural circuits from serial section electron microscopy (ssEM) images is being accelerated by automatic image segmentation methods. Segmentation accuracy is often limited by the preceding step of aligning 2D section images to create a 3D image stack. Precise and robust alignment in the presence of image artifacts is challenging, especially as datasets are attaining the petascale. We present a computational pipeline for aligning ssEM images with several key elements. Self-supervised convolutional nets are trained via metric learning to encode and align image pairs, and they are used to initialize iterative fine-tuning of alignment. A procedure called vector voting increases robustness to image artifacts or missing image data. For speedup the series is divided into blocks that are distributed to computational workers for alignment. The blocks are aligned to each other by composing transformations with decay, which achieves a global alignment without resorting to a time-consuming global optimization. We apply our pipeline to a whole fly brain dataset, and show improved accuracy relative to prior state of the art. We also demonstrate that our pipeline scales to a cubic millimeter of mouse visual cortex. Our pipeline is publicly available through two open source Python packages.

}, keywords = {Animals, Brain, Image Processing, Computer-Assisted, Imaging, Three-Dimensional, Mice, Microscopy, Electron}, issn = {2041-1723}, doi = {10.1038/s41467-023-44354-0}, author = {Popovych, Sergiy and Macrina, Thomas and Kemnitz, Nico and Castro, Manuel and Nehoran, Barak and Jia, Zhen and Bae, J Alexander and Mitchell, Eric and Mu, Shang and Trautman, Eric T and Saalfeld, Stephan and Li, Kai and Seung, H Sebastian} } @article {69128, title = {Phase diversity-based wavefront sensing for fluorescence microscopy.}, journal = {Optica}, volume = {11}, year = {2024}, month = {2024 Jun 10}, pages = {806-820}, abstract = {

Fluorescence microscopy is an invaluable tool in biology, yet its performance is compromised when the wavefront of light is distorted due to optical imperfections or the refractile nature of the sample. Such optical aberrations can dramatically lower the information content of images by degrading image contrast, resolution, and signal. Adaptive optics (AO) methods can sense and subsequently cancel the aberrated wavefront, but are too complex, inefficient, slow, or expensive for routine adoption by most labs. Here we introduce a rapid, sensitive, and robust wavefront sensing scheme based on phase diversity, a method successfully deployed in astronomy but underused in microscopy. Our method enables accurate wavefront sensing to less than λ/35 root mean square (RMS) error with few measurements, and AO with no additional hardware besides a corrective element. After validating the method with simulations, we demonstrate calibration of a deformable mirror \> 100-fold faster than comparable methods (corresponding to wavefront sensing on the ~100 ms scale), and sensing and subsequent correction of severe aberrations (RMS wavefront distortion exceeding λ/2), restoring diffraction-limited imaging on extended biological samples.

}, doi = {10.1364/OPTICA.518559}, url = {https://doi.org/10.1364/OPTICA.518559}, author = {Johnson, Courtney and Guo, Min and Schneider, Magdalena C and Su, Yijun and Khuon, Satya and Reiser, Nikolaj and Wu, Yicong and Riviere, Patrick La and Shroff, Hari} } @article {69404, title = {Recognising the importance and impact of Imaging Scientists: Global guidelines for establishing career paths within core facilities}, journal = {J Microsc}, year = {2024}, month = {2024 May 01}, abstract = {

In the dynamic landscape of scientific research, imaging core facilities are vital hubs propelling collaboration and innovation at the technology development and dissemination frontier. Here, we present a collaborative effort led by Global BioImaging (GBI), introducing international recommendations geared towards elevating the careers of Imaging Scientists in core facilities. Despite the critical role of Imaging Scientists in modern research ecosystems, challenges persist in recognising their value, aligning performance metrics and providing avenues for career progression and job security. The challenges encompass a mismatch between classic academic career paths and service-oriented roles, resulting in a lack of understanding regarding the value and impact of Imaging Scientists and core facilities and how to evaluate them properly. They further include challenges around sustainability, dedicated training opportunities and the recruitment and retention of talent. Structured across these interrelated sections, the recommendations within this publication aim to propose globally applicable solutions to navigate these challenges. These recommendations apply equally to colleagues working in other core facilities and research institutions through which access to technologies is facilitated and supported. This publication emphasises the pivotal role of Imaging Scientists in advancing research programs and presents a blueprint for fostering their career progression within institutions all around the world.

}, issn = {1365-2818}, doi = {10.1111/jmi.13307}, author = {Wright, Graham D and Thompson, Kerry A and Reis, Yara and Bischof, Johanna and Hockberger, Philip Edward and Itano, Michelle S and Yen, Lisa and Adelodun, Stephen Taiye and Bialy, Nikki and Brown, Claire M and Chaabane, Linda and Chew, Teng-Leong and Chitty, Andrew I and Cordeli{\`e}res, Fabrice P and De Niz, Mariana and Ellenberg, Jan and Engelbrecht, Lize and Fabian-Morales, Eunice and Fazeli, Elnaz and Fernandez-Rodriguez, Julia and Ferrando-May, Elisa and Fletcher, Georgina and Galloway, Graham John and Guerrero, Adan and Guimar{\~a}es, Jander Matos and Jacobs, Caron A and Jayasinghe, Sachintha and Kable, Eleanor and Kitten, Gregory T and Komoto, Shinya and Ma, Xiaoxiao and Marques, J{\'e}ssica Ara{\'u}jo and Millis, Bryan A and Miranda, Kildare and JohnO{\textquoteright}Toole, Peter and Olatunji, Sunday Yinka and Paina, Federica and Pollak, Cora Noemi and Prats, Clara and Pylv{\"a}n{\"a}inen, Joanna W and Rahmoon, Mai Atef and Reiche, Michael A and Riches, James Douglas and Rossi, Andres Hugo and Salamero, Jean and Thiriet, Caroline and Terjung, Stefan and Vasconcelos, Aldenora Dos Santos and Keppler, Antje} } @article {69248, title = {Recommendations for accelerating open preprint peer review to improve the culture of science}, journal = {PLOS Biology}, volume = {22}, year = {2024}, month = {2024 Feb 29}, pages = {e3002502}, abstract = {

Peer review is an important part of the scientific process, but traditional peer review at journals is coming under increased scrutiny for its inefficiency and lack of transparency. As preprints become more widely used and accepted, they raise the possibility of rethinking the peer-review process. Preprints are enabling new forms of peer review that have the potential to be more thorough, inclusive, and collegial than traditional journal peer review, and to thus fundamentally shift the culture of peer review toward constructive collaboration. In this Consensus View, we make a call to action to stakeholders in the community to accelerate the growing momentum of preprint sharing and provide recommendations to empower researchers to provide open and constructive peer review for preprints.

}, doi = {10.1371/journal.pbio.300250210.1371/journal.pbio.3002502.g001}, url = {https://dx.plos.org/10.1371/journal.pbio.3002502https://dx.plos.org/10.1371/journal.pbio.3002502https://dx.plos.org/10.1371/journal.pbio.3002502.g001}, author = {Avissar-Whiting, Michele and Belliard, Fr{\'e}d{\'e}rique and Bertozzi, Stefano M. and Brand, Amy and Brown, Katherine and Cl{\'e}ment-Stoneham, G{\'e}raldine and Dawson, Stephanie and Dey, Gautam and Ecer, Daniel and Edmunds, Scott C. and Farley, Ashley and Fischer, Tara D. and Franko, Maryrose and Fraser, James S. and Funk, Kathryn and Ganier, Clarisse and Harrison, Melissa and Hatch, Anna and Hazlett, Haley and Hindle, Samantha and Hook, Daniel W. and Hurst, Phil and Kamoun, Sophien and Kiley, Robert and Lacy, Michael M. and LaFlamme, Marcel and Lawrence, Rebecca and Lemberger, Thomas and Leptin, Maria and Lumb, Elliott and MacCallum, Catriona J. and Marcum, Christopher Steven and Marinello, Gabriele and Mendon{\c c}a, Alex and Monaco, Sara and Neves, Kleber and Pattinson, Damian and Polka, Jessica K. and Puebla, Iratxe and Rittman, Martyn and Royle, Stephen J. and Saderi, Daniela and Sever, Richard and Shearer, Kathleen and Spiro, John E. and Stern, Bodo and Taraborelli, Dario and Vale, Ron and Vasquez, Claudia G. and Waltman, Ludo and Watt, Fiona M. and Weinberg, Zara Y. and Williams, Mark} } @article {69231, title = {Recording physiological history of cells with chemical labeling.}, journal = {Science}, volume = {383}, year = {2024}, month = {2024 Feb 23}, pages = {890-897}, abstract = {

Recordings of the physiological history of cells provide insights into biological processes, yet obtaining such recordings is a challenge. To address this, we introduce a method to record transient cellular events for later analysis. We designed proteins that become labeled in the presence of both a specific cellular activity and a fluorescent substrate. The recording period is set by the presence of the substrate, whereas the cellular activity controls the degree of the labeling. The use of distinguishable substrates enabled the recording of successive periods of activity. We recorded protein-protein interactions, G protein-coupled receptor activation, and increases in intracellular calcium. Recordings of elevated calcium levels allowed selections of cells from heterogeneous populations for transcriptomic analysis and tracking of neuronal activities in flies and zebrafish.

}, keywords = {Animals, Calcium, Cell Physiological Phenomena, Cells, Coloring Agents, Gene Expression Profiling, Protein Interaction Domains and Motifs, Staining and Labeling, Zebrafish}, issn = {1095-9203}, doi = {10.1126/science.adg0812}, author = {Huppertz, Magnus-Carsten and Wilhelm, Jonas and Grenier, Vincent and Schneider, Martin W and Falt, Tjalda and Porzberg, Nicola and Hausmann, David and Hoffmann, Dirk C and Hai, Ling and Tarnawski, Miroslaw and Pino, Gabriela and Slanchev, Krasimir and Kolb, Ilya and Acuna, Claudio and Fenk, Lisa M and Baier, Herwig and Hiblot, Julien and Johnsson, Kai} } @article {69473, title = {SciJava Ops: An Improved Algorithms Framework for Fiji and Beyond}, journal = {Front. Bioinform.}, volume = {4}, year = {2024}, month = {2024 Sep 26}, abstract = {

Decades of iteration on scientific imaging hardware and software has yielded an explosion in not only the size, complexity, and heterogeneity of image datasets but also in the tooling used to analyze this data. This wealth of image analysis tools, spanning different programming languages, frameworks, and data structures, is itself a problem for data analysts who must adapt to new technologies and integrate established routines to solve increasingly complex problems. While many {\textquotedblleft}bridge{\textquotedblright} layers exist to unify pairs of popular tools, there exists a need for a general solution to unify new and existing toolkits. The SciJava Ops library presented here addresses this need through two novel principles. Algorithm implementations are declared as plugins called Ops, providing a uniform interface regardless of the toolkit they came from. Users express their needs declaratively to the Op environment, which can then find and adapt available Ops on demand. By using these principles instead of direct function calls, users can write streamlined workflows while avoiding the translation boilerplate of bridge layers. Developers can easily extend SciJava Ops to introduce new libraries and more efficient, specialized algorithm implementations, even immediately benefitting existing workflows. We provide several use cases showing both user and developer benefits, as well as benchmarking data to quantify the negligible impact on overall analysis performance. We have initially deployed SciJava Ops on the Fiji platform, however it would be suitable for integration with additional analysis platforms in the future.

}, doi = {10.3389/fbinf.2024.1435733}, url = {https://doi.org/10.3389/fbinf.2024.1435733 }, author = {Gabriel J. Selzer and Curtis T. Rueden and Mark C. Hiner and Edward L. Evans III and David Kolb and Marcel Wiedenmann and Christian Birkhold and Tim-Oliver Buchholz and Stefan Helfrich and Brian Northan and Alison Walter and Johannes Schindelin and Tobias Pietzsch and Stephan Saalfeld and Michael R. Berthold and Kevin W. Eliceiri} } @article {69296, title = {Social state gates vision using three circuit mechanisms in Drosophila}, journal = {Nature}, year = {2024}, month = {2024 Nov 20}, abstract = {

Animals are often bombarded with visual information and must prioritize specific visual features based on their current needs. The neuronal circuits that detect and relay visual features have been well studied. Much less is known about how an animal adjusts its visual attention as its goals or environmental conditions change. During social behaviours, flies need to focus on nearby flies. Here we study how the flow of visual information is altered when female Drosophila enter an aggressive state. From the connectome, we identify three state-dependent circuit motifs poised to modify the response of an aggressive female to fly-sized visual objects: convergence of excitatory inputs from neurons conveying select visual features and internal state; dendritic disinhibition of select visual feature detectors; and a switch that toggles between two visual feature detectors. Using cell-type-specific genetic tools, together with behavioural and neurophysiological analyses, we show that each of these circuit motifs is used during female aggression. We reveal that features of this same switch operate in male Drosophila during courtship pursuit, suggesting that disparate social behaviours may share circuit mechanisms. Our study provides a compelling example of using the connectome to infer circuit mechanisms that underlie dynamic processing of sensory signals.

}, doi = {10.1038/s41586-024-08255-6}, url = {https://doi.org/10.1038/s41586-024-08255-6}, author = {Catherine E. Schretter and Tom Hindmarsh Sten and Nathan Klapoetke and Mei Shao and Aljoscha Nern and Marisa Dreher and Daniel Bushey and Alice A. Robie and Adam L. Taylor and Kristin M. Branson and Adriane Otopalik and Vanessa Ruta and Gerald M. Rubin} } @article {69146, title = {Song Torrent: A modular, open-source 96-chamber audio and video recording apparatus with optogenetic activation and inactivation capabilities for Drosophila}, journal = {bioRxiv}, year = {2024}, month = {2024 Jan 10}, abstract = {

Background

Results

Implications

}, doi = {10.1101/2024.01.09.574712}, url = {https://doi.org/10.1101/2024.01.09.574712}, author = {Steve Sawtelle and Lakshmi Narayan and Yun Ding and Elizabeth Kim and Emily L. Behrman and Joshua L. Lillvis and Takashi Kawase and David L. Stern} } @article {69329, title = {A split-GAL4 driver line resource for Drosophila CNS cell types}, journal = {bioRxiv}, year = {2024}, month = {2024 Jan 10}, abstract = {

Techniques that enable precise manipulations of subsets of neurons in the fly central nervous system have greatly facilitated our understanding of the neural basis of behavior. Split-GAL4 driver lines allow specific targeting of cell types in Drosophila melanogaster and other species. We describe here a collection of 3060 lines targeting a range of cell types in the adult Drosophila central nervous system and 1373 lines characterized in third-instar larvae. These tools enable functional, transcriptomic, and proteomic studies based on precise anatomical targeting. NeuronBridge and other search tools relate light microscopy images of these split-GAL4 lines to connectomes reconstructed from electron microscopy images. The collections are the result of screening over 77,000 split hemidriver combinations. In addition to images and fly stocks for these well-characterized lines, we make available 300,000 new 3D images of other split-GAL4 lines.

}, doi = {10.1101/2024.01.09.574419}, url = {https://www.biorxiv.org/content/early/2024/04/01/2024.01.09.574419}, author = {Geoffrey W Meissner and Allison Vannan and Jennifer Jeter and Kari Close and Gina M DePasquale and Zachary Dorman and Kaitlyn Forster and Jaye Anne Beringer and Theresa V Gibney and Joanna H Hausenfluck and Yisheng He and Kristin Henderson and Lauren Johnson and Rebecca M Johnston and Gudrun Ihrke and Nirmala Iyer and Rachel Lazarus and Kelley Lee and Hsing-Hsi Li and Hua-Peng Liaw and Brian Melton and Scott Miller and Reeham Motaher and Alexandra Novak and Omotara Ogundeyi and Alyson Petruncio and Jacquelyn Price and Sophia Protopapas and Susana Tae and Jennifer Taylor and Rebecca Vorimo and Brianna Yarbrough and Kevin Xiankun Zeng and Christopher T Zugates and Heather Dionne and Claire Angstadt and Kelly Ashley and Amanda Cavallaro and Tam Dang and Guillermo A Gonzalez III and Karen L Hibbard and Cuizhen Huang and Jui-Chun Kao and Todd Laverty and Monti Mercer and Brenda Perez and Scarlett Pitts and Danielle Ruiz and Viruthika Vallanadu and Grace Zhiyu Zheng and Cristian Goina and Hideo Otsuna and Konrad Rokicki and Robert R Svirskas and Han SJ Cheong and Michael-John Dolan and Erica Ehrhardt and Kai Feng and Basel El Galfi and Jens Goldammer and Stephen J Huston and Nan Hu and Masayoshi Ito and Claire McKellar and Ryo Minegishi and Shigehiro Namiki and Aljoscha Nern and Catherine E Schretter and Gabriella R Sterne and Lalanti Venkatasubramanian and Kaiyu Wang and Tanya Wolff and Ming Wu and Reed George and Oz Malkesman and Yoshinori Aso and Gwyneth M Card and Barry J Dickson and Wyatt Korff and Kei Ito and James W Truman and Marta Zlatic and Gerald M Rubin and FlyLight Project Team} } @article {69109, title = {Structure, interaction, and nervous connectivity of beta cell primary cilia}, journal = {Nat Commun}, volume = {15}, year = {2024}, month = {2024 Oct 24}, pages = {9168}, abstract = {

Primary cilia are sensory organelles present in many cell types, partaking in various signaling processes. Primary cilia of pancreatic beta cells play pivotal roles in paracrine signaling and their dysfunction is linked to diabetes. Yet, the structural basis for their functions is unclear. We present three-dimensional reconstructions of beta cell primary cilia by electron and expansion microscopy. These cilia are spatially confined within deep ciliary pockets or narrow spaces between cells, lack motility components and display an unstructured axoneme organization. Furthermore, we observe a plethora of beta cell cilia-cilia and cilia-cell interactions with other islet and non-islet cells. Most remarkably, we have identified and characterized axo-ciliary synapses between beta cell cilia and the cholinergic islet innervation. These findings highlight the beta cell cilia{\textquoteright}s role in islet connectivity, pointing at their function in integrating islet intrinsic and extrinsic signals and contribute to understanding their significance in health and diabetes.

}, doi = {10.1038/s41467-024-53348-5}, url = {https://doi.org/10.1038/s41467-024-53348-5}, author = {Andreas M{\"u}ller and Nikolai Klena and Song Pang and Leticia Elizabeth Galicia Garcia and Davud Sulaymankhil and Oleksandra Topcheva and Monika Seliskar and Hassan Mziaut and Eyke Sch{\"o}niger and Daniela Friedland and Nicole Kipke and Susanne Kretschmar and Carla M{\"u}nster and J{\"u}rgen Weitz and Marius Distler and Thomas Kurth and Deborah Schmidt and Harald F. Hess and C. Shan Xu and Gaia Pigino and Michele Solimena} } @article {69282, title = {Three-dimensional spatio-angular fluorescence microscopy with a polarized dual-view inverted selective-plane illumination microscope (pol-diSPIM)}, journal = {bioRxiv}, year = {2024}, month = {2024 Mar 12}, abstract = {

Polarized fluorescence microscopy is a valuable tool for measuring molecular orientations, but techniques for recovering three-dimensional orientations and positions of fluorescent ensembles are limited. We report a polarized dual-view light-sheet system for determining the three-dimensional orientations and diffraction-limited positions of ensembles of fluorescent dipoles that label biological structures, and we share a set of visualization, histogram, and profiling tools for interpreting these positions and orientations. We model our samples, their excitation, and their detection using coarse-grained representations we call orientation distribution functions (ODFs). We apply ODFs to create physics-informed models of image formation with spatio-angular point-spread and transfer functions. We use theory and experiment to conclude that light-sheet tilting is a necessary part of our design for recovering all three-dimensional orientations. We use our system to extend known two-dimensional results to three dimensions in FM1-43-labelled giant unilamellar vesicles, fast-scarlet-labelled cellulose in xylem cells, and phalloidin-labelled actin in U2OS cells. Additionally, we observe phalloidin-labelled actin in mouse fibroblasts grown on grids of labelled nanowires and identify correlations between local actin alignment and global cell-scale orientation, indicating cellular coordination across length scales.Competing Interest StatementH.S., A.K., S.M., P.L.R., R.O., Y.W., and T.C. hold US Patent $\#$11428632.

}, doi = {10.1101/2024.03.09.584243}, url = {https://www.biorxiv.org/content/early/2024/03/12/2024.03.09.584243}, author = {Talon Chandler and Min Guo and Yijun Su and Jiji Chen and Yicong Wu and Junyu Liu and Atharva Agashe and Robert S. Fischer and Shalin B. Mehta and Abhishek Kumar and Tobias I. Baskin and Valentin Jamouille and Huafeng Liu and Vinay Swaminathan and Amrinder Nain and Rudolf Oldenbourg and Patrick La Riviere and Hari Shroff} } @article {69674, title = {Tracking fructose 1,6-bisphosphate dynamics in liver cancer cells using a fluorescent biosensor}, journal = {iScience}, volume = {27}, year = {2024}, month = {2024 Nov 20}, pages = {111336}, abstract = {

Summary HYlight is a genetically encoded fluorescent biosensor that ratiometrically monitors fructose 1,6-bisphosphate (FBP), a key glycolytic metabolite. Given the role of glucose in liver cancer metabolism, we expressed HYlight in human liver cancer cells and primary mouse hepatocytes. Through in\ vitro, in silico, and in cellulo experiments, we showed HYlight{\textquoteright}s ability to monitor FBP changes linked to glycolysis, not gluconeogenesis. HYlight{\textquoteright}s affinity for FBP was \~{}1\ μM and stable within physiological pH range. HYlight demonstrated weak binding to dihydroxyacetone phosphate, and its ratiometric response was influenced by both ionic strength and phosphate. Therefore, simulating cytosolic conditions in\ vitro was necessary to establish a reliable correlation between HYlight{\textquoteright}s cellular responses and FBP concentrations. FBP concentrations were found to be in the lower micromolar range, far lower than previous millimolar estimates. Altogether, this biosensor approach offers real-time monitoring of FBP concentrations at single-cell resolution, making it an invaluable tool for the understanding of cancer metabolism.

}, keywords = {biochemistry methods, cancer, metabolomics}, issn = {2589-0042}, doi = {https://doi.org/10.1016/j.isci.2024.111336}, url = {https://www.sciencedirect.com/science/article/pii/S2589004224025616}, author = {Israel P{\'e}rez-Ch{\'a}vez and John N. Koberstein and Julia Malo Pueyo and Eduardo H. Gilglioni and Didier Vertommen and Nicolas Baeyens and Daria Ezerina and Esteban N. Gurzov and Joris Messens} } @article {69144, title = {Transforming chemigenetic bimolecular fluorescence complementation systems into chemical dimerizers using chemistry.}, journal = {bioRxiv}, year = {2024}, month = {2024 Jan 01}, abstract = {

Chemigenetic tags are versatile labels for fluorescence microscopy that combine some of the advantages of genetically encoded tags with small molecule fluorophores. The Fluorescence Activating and absorbance Shifting Tags (FASTs) bind a series of highly fluorogenic and cell-permeable chromophores. Furthermore, FASTs can be used in complementation-based systems for detecting or inducing protein-protein interactions, depending on the exact FAST protein variant chosen. In this study, we systematically explore substitution patterns on FAST fluorogens and generate a series of fluorogens that bind to FAST variants, thereby activating their fluorescence. This effort led to the discovery of a novel fluorogen with superior properties, as well as a fluorogen that transforms splitFAST systems into a fluorogenic dimerizer, eliminating the need for additional protein engineering.

}, doi = {10.1101/2023.12.30.573644}, url = {https://doi.org/10.1101/2023.12.30.573644}, author = {Pratik Kumar and Alina Gutu and Amelia Waring and Timothy A. Brown and Luke D. Lavis and Alison G. Tebo} } @article {69662, title = {Type IV pili-associated secretion of a biofilm matrix protein from Clostridium perfringens that forms intermolecular isopeptide bonds}, journal = {bioRxiv}, year = {2024}, month = {2024 Nov 04}, abstract = {

Clostridium perfringens is a Gram-positive anaerobic spore-forming bacterial pathogen of humans and animals. C. perfringens also produces type IV pili (T4P) and has two complete sets of T4P-associated genes, one of which has been shown to produce surface pili needed for cell adherence. One hypothesis about the role of the other set of T4P genes is that they could comprise a system analogous to the type II secretion systems (TTSS) found in Gram-negative bacteria, which is used to export folded proteins from the periplasm through the outer membrane to the extracellular environment. Gram-positive bacteria have a similar secretion barrier in the thick peptidoglycan (PG) layer, which blocks secretion of folded proteins \>25 kD. To determine if the T4P-associated genes comprise a Gram-positive TTSS, the secretome of mutants lacking type IV pilins were examined and a single protein, a von Willebrand A domain containing protein BsaC (CPE0517) was identified as being dependent on PilA3 for secretion. BsaC is in an operon with a signal peptidase and two putative biofilm matrix proteins with homology to Bacillus subtilis TasA. One of these proteins, BsaA, was shown by another group to produce high mol wt oligomers. We analyzed BsaA monomer interactions with de novo modeling, which projected that the monomers formed isopeptide bonds as part of a donor strand exchange process. Mutations in residues predicted to form the isopeptide bonds led to loss of oligomerization, supporting the predicted bond formation process. Phylogenetic analysis showed the BsaA family of proteins are widespread among bacteria and archaea but only a subset are predicted to form isopeptide bonds.

}, doi = {10.1101/2024.11.04.621531}, url = {https://doi.org/10.1101/2024.11.04.621531}, author = {Kivimaki, Sarah E. and Dempsey, Samantha and Tani, Julia M. and Camper, Colette and Hicklin, Ian K. and Blaby-Haas, Crysten E. and Brown, Anne M and Melville, Stephen} } @article {69350, title = {Ultra-high density electrodes improve detection, yield, and cell type identification in neuronal recordings}, journal = {bioRxiv}, year = {2024}, month = {2024 Apr 10}, abstract = {

To understand the neural basis of behavior, it is essential to sensitively and accurately measure neural activity at single neuron and single spike resolution. Extracellular electrophysiology delivers this, but it has biases in the neurons it detects and it imperfectly resolves their action potentials. To minimize these limitations, we developed a silicon probe with much smaller and denser recording sites than previous designs, called Neuropixels Ultra (NP Ultra). This device samples neuronal activity at ultra-high spatial density ( 10 times higher than previous probes) with low noise levels, while trading off recording span. NP Ultra is effectively an implantable voltage-sensing camera that captures a planar image of a neuron{\textquoteright}s electrical field. We use a spike sorting algorithm optimized for these probes to demonstrate that the yield of visually-responsive neurons in recordings from mouse visual cortex improves up to 3-fold. We show that NP Ultra can record from small neuronal structures including axons and dendrites. Recordings across multiple brain regions and four species revealed a subset of extracellular action potentials with unexpectedly small spatial spread and axon-like features. We share a large-scale dataset of these brain-wide recordings in mice as a resource for studies of neuronal biophysics. Finally, using ground-truth identification of three major inhibitory cortical cell types, we found that these cell types were discriminable with approximately 75\% success, a significant improvement over lower-resolution recordings. NP Ultra improves spike sorting performance, detection of subcellular compartments, and cell type classification to enable more powerful dissection of neural circuit activity during behavior.

}, doi = {10.1101/2023.08.23.554527}, url = {https://www.biorxiv.org/content/early/2024/04/10/2023.08.23.554527}, author = {Zhiwen Ye and Andrew M Shelton and Jordan R Shaker and Julien M Boussard and Jennifer Colonell and Daniel Birman and Sahar Manavi and Susu Chen and Charlie Windolf and Cole Hurwitz and Tomoyuki Namima and Frederico Pedraja and Shahaf Weiss and Bogdan Raducanu and Torbj{\o}rn Ness and Xiaoxuan Jia and Giulia Mastroberardino and L. Federico Rossi and Matteo Carandini and Michael Hausser and Gaute T Einevoll and Gilles Laurent and Nathaniel B Sawtell and Wyeth Bair and Anitha Pasupathy and Carolina Mora-Lopez and Barun Dutta and Liam Paninski and Joshua H Siegle and Christof Koch and Shawn R Olsen and Timothy D Harris and Nicholas A Steinmetz} } @article {69459, title = {Ultrastructural differences impact cilia shape and external exposure across cell classes in the visual cortex}, journal = {Curr Biol}, year = {2024}, month = {2024 May 13}, abstract = {

A primary cilium is a membrane-bound extension from the cell surface that contains receptors for perceiving and transmitting signals that modulate cell state and activity. Primary cilia in the brain are less accessible than cilia on cultured cells or epithelial tissues because in the brain they protrude into a deep, dense network of glial and neuronal processes. Here, we investigated cilia frequency, internal structure, shape, and position in large, high-resolution transmission electron microscopy volumes of mouse primary visual cortex. Cilia extended from the cell bodies of nearly all excitatory and inhibitory neurons, astrocytes, and oligodendrocyte precursor cells (OPCs) but were absent from oligodendrocytes and microglia. Ultrastructural comparisons revealed that the base of the cilium and the microtubule organization differed between neurons and glia. Investigating cilia-proximal features revealed that many cilia were directly adjacent to synapses, suggesting that cilia are poised to encounter locally released signaling molecules. Our analysis indicated that synapse proximity is likely due to random encounters in the neuropil, with no evidence that cilia modulate synapse activity as would be expected in tetrapartite synapses. The observed cell class differences in proximity to synapses were largely due to differences in external cilia length. Many key structural features that differed between neuronal and glial cilia influenced both cilium placement and shape and, thus, exposure to processes and synapses outside the cilium. Together, the ultrastructure both within and around neuronal and glial cilia suggest differences in cilia formation and function across cell types in the brain.

}, issn = {1879-0445}, doi = {10.1016/j.cub.2024.04.043}, author = {Ott, Carolyn M and Torres, Russel and Kuan, Tung-Sheng and Kuan, Aaron and Buchanan, JoAnn and Elabbady, Leila and Seshamani, Sharmishtaa and Bodor, Agnes L and Collman, Forrest and Bock, Davi D and Lee, Wei Chung and da Costa, Nuno Ma{\c c}arico and Lippincott-Schwartz, Jennifer} } @inbook {69316, title = {Visualization of Glutamatergic Neurotransmission in Diverse Model Organisms with Genetically Encoded Indicators}, booktitle = {New Technologies for Glutamate Interaction: Neurons and Glia}, volume = {2780}, number = {Neuromethods}, year = {2024}, pages = {3{\textendash}34}, publisher = {Humana}, organization = {Humana}, address = {New York, NY}, abstract = {

Glutamate is the principal excitatory neurotransmitter, and occasionally subserves inhibitory roles, in the vertebrate nervous system. Glutamatergic synapses are dense in the vertebrate brain, at \textasciitilde1/μm3. Glutamate is released from and onto diverse components of the nervous system, including neurons, glia, and other cells. Methods for glutamate detection are critically important for understanding the function of synapses and neural circuits in normal physiology, development, and disease. Here we describe the development, optimization, and deployment of genetically encoded fluorescent glutamate indicators. We review the theoretical considerations governing glutamate sensor properties from first principles of synapse biology, microscopy, and protein structure-function relationships. We provide case studies of the state-of-the-art iGluSnFR glutamate sensor, encompassing design and optimization, mechanism of action, in vivo imaging, data analysis, and future directions. We include detailed protocols for iGluSnFR imaging in common preparations (bacteria, cell culture, and brain slices) and model organisms (worm, fly, fish, rodent).

}, isbn = {978-1-0716-3742-5}, doi = {10.1007/978-1-0716-3742-5_1}, url = {https://doi.org/10.1007/978-1-0716-3742-5_1}, author = {Aggarwal, Abhi and Chan, Joyce and Waring, Amelia K. and Negrean, Adrian and Marvin, Jonathan S. and Podgorski, Kaspar and Looger, Loren L.}, editor = {Kukley, Maria} } @article {68519, title = {ACC neural ensemble dynamics are structured by strategy prevalence}, journal = {eLife}, year = {2023}, month = {2023 Nov 22}, abstract = {

Medial frontal cortical areas are thought to play a critical role in the brain{\textquoteright}s ability to flexibly deploy strategies that are effective in complex settings. Still, the specific circuit computations that underpin this foundational aspect of intelligence remain unclear. Here, by examining neural ensemble activity in rats that sample different strategies in a self-guided search for latent task structure, we demonstrate a robust tracking of individual strategy prevalence in the anterior cingulate cortex (ACC), especially in an area homologous to primate area 32D. Prevalence encoding in the ACC is wide-scale, independent of reward delivery, and persists through a substantial ensemble reorganization that tags ACC representations with contextual content. Our findings argue that ACC ensemble dynamics is structured by a summary statistic of recent behavioral choices, raising the possibility that ACC plays a role in estimating - through statistical learning - which actions promote the occurrence of events in the environment.

}, doi = {10.7554/eLife.84897}, url = {https://doi.org/10.7554/eLife.84897}, author = {Mikhail Proskurin and Maxim Manakov and Alla Y. Karpova} } @article {69137, title = {Anatomically distributed neural representations of instincts in the hypothalamus.}, journal = {bioRxiv}, year = {2023}, month = {2023 Dec 07}, abstract = {

Artificial activation of anatomically localized, genetically defined hypothalamic neuron populations is known to trigger distinct innate behaviors, suggesting a hypothalamic nucleus-centered organization of behavior control. To assess whether the encoding of behavior is similarly anatomically confined, we performed simultaneous neuron recordings across twenty hypothalamic regions in freely moving animals. Here we show that distinct but anatomically distributed neuron ensembles encode the social and fear behavior classes, primarily through mixed selectivity. While behavior class-encoding ensembles were spatially distributed, individual ensembles exhibited strong localization bias. Encoding models identified that behavior actions, but not motion-related variables, explained a large fraction of hypothalamic neuron activity variance. These results identify unexpected complexity in the hypothalamic encoding of instincts and provide a foundation for understanding the role of distributed neural representations in the expression of behaviors driven by hardwired circuits.

}, doi = {10.1101/2023.11.21.568163}, url = {https://doi.org/10.1101/2023.11.21.568163}, author = {Stagkourakis, Stefanos and Spigolon, Giada and Marks, Markus and Feyder, Michael and Kim, Joseph and Perona, Pietro and Pachitariu, Marius and Anderson, David J} } @article {68807, title = {Architecture and dynamics of a desmosome-endoplasmic reticulum complex.}, journal = {Nature Cell Biology}, volume = {25}, year = {2023}, month = {2023 Jun 16}, pages = {823-835}, abstract = {

The endoplasmic reticulum (ER) forms a dynamic network that contacts other cellular membranes to regulate stress responses, calcium signalling and lipid transfer. Here, using high-resolution volume electron microscopy, we find that the ER forms a previously unknown association with keratin intermediate filaments and desmosomal cell-cell junctions. Peripheral ER assembles into mirror image-like arrangements at desmosomes and exhibits nanometre proximity to keratin filaments and the desmosome cytoplasmic plaque. ER tubules exhibit stable associations with desmosomes, and perturbation of desmosomes or keratin filaments alters ER organization, mobility and expression of ER stress transcripts. These findings indicate that desmosomes and the keratin cytoskeleton regulate the distribution, function and dynamics of the ER network. Overall, this study reveals a previously unknown subcellular architecture defined by the structural integration of ER tubules with an epithelial intercellular junction.

}, keywords = {Cytoskeleton, Desmosomes, Endoplasmic Reticulum, Intermediate Filaments, Keratins}, issn = {1476-4679}, doi = {10.1038/s41556-023-01154-4}, author = {Bharathan, Navaneetha Krishnan and Giang, William and Hoffman, Coryn L and Aaron, Jesse S and Khuon, Satya and Chew, Teng-Leong and Preibisch, Stephan and Trautman, Eric T and Heinrich, Larissa and Bogovic, John and Bennett, Davis and Ackerman, David and Park, Woohyun and Petruncio, Alyson and Weigel, Aubrey V and Saalfeld, Stephan and Wayne Vogl, A and Stahley, Sara N and Kowalczyk, Andrew P} } @article {68587, title = {Automated reconstruction of whole-embryo cell lineages by learning from sparse annotations.}, journal = {Nature Biotechnology}, volume = {41}, year = {2023}, month = {2023 Jan 01}, pages = {44-49}, abstract = {

We present a method to automatically identify and track nuclei in time-lapse microscopy recordings of entire developing embryos. The method combines deep learning and global optimization. On a mouse dataset, it reconstructs 75.8\% of cell lineages spanning 1 h, as compared to 31.8\% for the competing method. Our approach improves understanding of where and when cell fate decisions are made in developing embryos, tissues, and organs.

}, keywords = {Animals, Blastocyst, Cell Lineage, Embryo, Mammalian, Mice, Microscopy}, issn = {1546-1696}, doi = {10.1038/s41587-022-01427-7}, author = {Malin-Mayor, Caroline and Hirsch, Peter and Guignard, Leo and McDole, Katie and Wan, Yinan and Lemon, William C and Kainmueller, Dagmar and Keller, Philipp J and Preibisch, Stephan and Funke, Jan} } @article {68709, title = {BigNeuron: a resource to benchmark and predict performance of algorithms for automated tracing of neurons in light microscopy datasets.}, journal = {Nature Methods}, volume = {20}, year = {2023}, month = {2023 Jun 01}, abstract = {

BigNeuron is an open community bench-testing platform with the goal of setting open standards for accurate and fast automatic neuron tracing. We gathered a diverse set of image volumes across several species that is representative of the data obtained in many neuroscience laboratories interested in neuron tracing. Here, we report generated gold standard manual annotations for a subset of the available imaging datasets and quantified tracing quality for 35 automatic tracing algorithms. The goal of generating such a hand-curated diverse dataset is to advance the development of tracing algorithms and enable generalizable benchmarking. Together with image quality features, we pooled the data in an interactive web application that enables users and developers to perform principal component analysis, t-distributed stochastic neighbor embedding, correlation and clustering, visualization of imaging and tracing data, and benchmarking of automatic tracing algorithms in user-defined data subsets. The image quality metrics explain most of the variance in the data, followed by neuromorphological features related to neuron size. We observed that diverse algorithms can provide complementary information to obtain accurate results and developed a method to iteratively combine methods and generate consensus reconstructions. The consensus trees obtained provide estimates of the neuron structure ground truth that typically outperform single algorithms in noisy datasets. However, specific algorithms may outperform the consensus tree strategy in specific imaging conditions. Finally, to aid users in predicting the most accurate automatic tracing results without manual annotations for comparison, we used support vector machine regression to predict reconstruction quality given an image volume and a set of automatic tracings.

}, issn = {1548-7105}, doi = {10.1038/s41592-023-01848-5}, author = {Manubens-Gil, Linus and Zhou, Zhi and Chen, Hanbo and Ramanathan, Arvind and Liu, Xiaoxiao and Liu, Yufeng and Bria, Alessandro and Gillette, Todd and Ruan, Zongcai and Yang, Jian and Radojevi{\'c}, Miroslav and Zhao, Ting and Cheng, Li and Qu, Lei and Liu, Siqi and Bouchard, Kristofer E and Gu, Lin and Cai, Weidong and Ji, Shuiwang and Roysam, Badrinath and Wang, Ching-Wei and Yu, Hongchuan and Sironi, Amos and Iascone, Daniel Maxim and Zhou, Jie and Bas, Erhan and Conde-Sousa, Eduardo and Aguiar, Paulo and Li, Xiang and Li, Yujie and Nanda, Sumit and Wang, Yuan and Muresan, Leila and Fua, Pascal and Ye, Bing and He, Hai-Yan and Staiger, Jochen F and Peter, Manuel and Cox, Daniel N and Simonneau, Michel and Oberlaender, Marcel and Jefferis, Gregory and Ito, Kei and Gonzalez-Bellido, Paloma and Kim, Jinhyun and Rubel, Edwin and Cline, Hollis T and Zeng, Hongkui and Nern, Aljoscha and Chiang, Ann-Shyn and Yao, Jianhua and Roskams, Jane and Livesey, Rick and Stevens, Janine and Liu, Tianming and Dang, Chinh and Guo, Yike and Zhong, Ning and Tourassi, Georgia and Hill, Sean and Hawrylycz, Michael and Koch, Christof and Meijering, Erik and Ascoli, Giorgio A and Peng, Hanchuan} } @article {68933, title = {Biomechanical origins of proprioceptor feature selectivity and topographic maps in the Drosophila leg.}, journal = {Neuron}, year = {2023}, month = {2023 Aug 04}, abstract = {

Our ability to sense and move our bodies relies on proprioceptors, sensory neurons that detect mechanical forces within the body. Different subtypes of proprioceptors detect different kinematic features, such as joint position, movement, and vibration, but the mechanisms that underlie proprioceptor feature selectivity remain poorly understood. Using single-nucleus RNA sequencing (RNA-seq), we found that proprioceptor subtypes in the Drosophila leg lack differential expression of mechanosensitive ion channels. However, anatomical reconstruction of the proprioceptors and connected tendons revealed major biomechanical differences between subtypes. We built a model of the proprioceptors and tendons that identified a biomechanical mechanism for joint angle selectivity and predicted the existence of a topographic map of joint angle, which we confirmed using calcium imaging. Our findings suggest that biomechanical specialization is a key determinant of proprioceptor feature selectivity in Drosophila. More broadly, the discovery of proprioceptive maps reveals common organizational principles between proprioception and other topographically organized sensory systems.

}, issn = {1097-4199}, doi = {10.1016/j.neuron.2023.07.009}, author = {Mamiya, Akira and Sustar, Anne and Siwanowicz, Igor and Qi, Yanyan and Lu, Tzu-Chiao and Gurung, Pralaksha and Chen, Chenghao and Phelps, Jasper S and Kuan, Aaron T and Pacureanu, Alexandra and Lee, Wei-Chung Allen and Li, Hongjie and Mhatre, Natasha and Tuthill, John C} } @article {68964, title = {Brain wiring determinants uncovered by integrating connectomes and transcriptomes.}, journal = {Current Biology}, volume = {33}, year = {2023}, month = {2023 Aug 23}, pages = {3998-3998}, abstract = {

Advances in brain connectomics have demonstrated the extraordinary complexity of neural circuits. Developing neurons encounter the axons and dendrites of many different neuron types and form synapses with only a subset of them. During circuit assembly, neurons express cell-type-specific repertoires comprising many cell adhesion molecules (CAMs) that can mediate interactions between developing neurites. Many CAM families have been shown to contribute to brain wiring in different ways. It has been challenging, however, to identify receptor-ligand pairs directly matching neurons with their synaptic targets. Here, we integrated the synapse-level connectome of the neural circuit with the developmental expression patterns and binding specificities of CAMs on pre- and postsynaptic neurons in the Drosophila visual system. To overcome the complexity of neural circuits, we focus on pairs of genetically related neurons that make differential wiring choices. In the motion detection circuit, closely related subtypes of T4/T5 neurons choose between alternative synaptic targets in adjacent layers of neuropil. This choice correlates with the matching expression in synaptic partners of different receptor-ligand pairs of the Beat and Side families of CAMs. Genetic analysis demonstrated that presynaptic Side-II and postsynaptic Beat-VI restrict synaptic partners to the same layer. Removal of this receptor-ligand pair disrupts layers and leads to inappropriate targeting of presynaptic sites and postsynaptic dendrites. We propose that different Side/Beat receptor-ligand pairs collaborate with other recognition molecules to determine wiring specificities in the fly brain. Combining transcriptomes, connectomes, and protein interactome maps allow unbiased identification of determinants of brain wiring.

}, issn = {1879-0445}, doi = {10.1016/j.cub.2023.08.020}, author = {Yoo, Juyoun and Dombrovski, Mark and Mirshahidi, Parmis and Nern, Aljoscha and LoCascio, Samuel A and Zipursky, S Lawrence and Kurmangaliyev, Yerbol Z} } @article {68257, title = {Characterization, Comparison, and Optimization of Lattice Light Sheets}, journal = {Science Advances}, year = {2023}, month = {2023 Mar 31}, abstract = {

Lattice light sheet microscopy excels at the non-invasive imaging of three-dimensional (3D) dynamic processes at high spatiotemporal resolution within cells and developing embryos. Recently, several papers have called into question the performance of lattice light sheets relative to the Gaussian sheets most common in light sheet microscopy. Here we undertake a comprehensive theoretical and experimental analysis of various forms of light sheet microscopy which both demonstrates and explains why lattice light sheets provide significant improvements in resolution and photobleaching reduction. The analysis provides a procedure to select the correct light sheet for a desired experiment and specifies the processing that maximizes the use of all fluorescence generated within the light sheet excitation envelope for optimal resolution while minimizing image artifacts and photodamage. Development of a new type of {\textquotedblleft}harmonic balanced{\textquotedblright} lattice light sheet is shown to improve performance at all spatial frequencies within its 3D resolution limits and maintains this performance over lengthened propagation distances allowing for expanded fields of view.

}, doi = {10.1126/sciadv.ade6623}, url = {https://doi.org/10.1126/sciadv.ade6623}, author = {Gaoxiang Liu and Xiongtao Ruan and Daniel E. Milkie and Frederik G{\"o}rlitz and Matthew Mueller and Wilmene Hercule and Alison Kililea and Eric Betzig and Srigokul Upadhyayula} } @article {69018, title = {Complete human day 14 post-implantation embryo models from na{\"\i}ve ES cells}, journal = {Nature}, year = {2023}, month = {09/2023}, abstract = {

The ability to study human post-implantation development remains limited due to ethical and technical challenges associated with intrauterine development after implantation1. Embryo-like models with spatially organized morphogenesis of all defining embryonic and extra-embryonic tissues of the post-implantation human conceptus (i.e., embryonic disk, bilaminar disk, yolk- and chorionic sacs, surrounding trophoblasts) remain lacking2. Mouse na{\"\i}ve embryonic stem cells (ESCs) have recently been shown to give rise to embryonic and extra-embryonic stem cells capable of self-assembling into post-gastrulation mouse Structured Stem cell-based Embryo Models with spatially organized morphogenesis (SEMs)3. Here, we extend these findings to humans, while using only genetically unmodified human na{\"\i}ve ESCs (in HENSM conditions)4. Such human fully integrated SEMs recapitulate the organization of nearly all known lineages and compartments of post-implantation human embryos including epiblast, hypoblast, extra-embryonic mesoderm, and trophoblast surrounding the latter layers. These human complete SEMs demonstrated developmental growth dynamics that resemble key hallmarks of post-implantation stage embryogenesis up to 13-14 days post-fertilization (dpf) (Carnegie stage 6a). This includes embryonic disk and bilaminar disk formation, epiblast lumenogenesis, polarized amniogenesis, anterior-posterior symmetry breaking, PGC specification, polarized yolk sac with visceral and parietal endoderm, extra-embryonic mesoderm expansion that defines a chorionic cavity and a connecting stalk, a trophoblast surrounding compartment demonstrating syncytium and lacunae formation. This SEM platform may enable the experimental interrogation of previously inaccessible windows of human early post-implantation up to peri-gastrulation development.

}, issn = {0028-0836}, doi = {10.1038/s41586-023-06604-5}, url = {https://www.nature.com/articles/s41586-023-06604-5https://www.nature.com/articles/s41586-023-06604-5.pdfhttps://www.nature.com/articles/s41586-023-06604-5.pdfhttps://www.nature.com/articles/s41586-023-06604-5}, author = {Oldak, Bernardo and Wildschutz, Emilie and Bondarenko, Vladyslav and Comar, Mehmet-Yunus and Zhao, Cheng and Aguilera-Castrejon, Alejandro and Tarazi, Shadi and Viukov, Sergey and Pham, Thi Xuan Ai and Ashouokhi, Shahd and Lokshtanov, Dmitry and Roncato, Francesco and Ariel, Eitan and Rose, Max and Livnat, Nir and Shani, Tom and Joubran, Carine and Cohen, Roni and Addadi, Yoseph and Chemla, Muriel and Kedmi, Merav and Keren-Shaul, Hadas and Pasque, Vincent and Petropoulos, Sophie and Lanner, Fredrik and Novershtern, Noa and Hanna, Jacob H.} } @article {69039, title = {A comprehensive neuroanatomical survey of the Drosophila Lobula Plate Tangential Neurons with predictions for their optic flow sensitivity.}, journal = {bioRxiv}, year = {2023}, month = {2023 Oct 17}, abstract = {

Flying insects exhibit remarkable navigational abilities controlled by their compact nervous systems. Optic flow, the pattern of changes in the visual scene induced by locomotion, is a crucial sensory cue for robust self-motion estimation, especially during rapid flight. Neurons that respond to specific, large-field optic flow patterns have been studied for decades, primarily in large flies, such as houseflies, blowflies, and hover flies. The best-known optic-flow sensitive neurons are the large tangential cells of the dipteran lobula plate, whose visual-motion responses, and to a lesser extent, their morphology, have been explored using single-neuron neurophysiology. Most of these studies have focused on the large, Horizontal and Vertical System neurons, yet the lobula plate houses a much larger set of {\textquoteright}optic-flow{\textquoteright} sensitive neurons, many of which have been challenging to unambiguously identify or to reliably target for functional studies. Here we report the comprehensive reconstruction and identification of the Lobula Plate Tangential Neurons in an Electron Microscopy (EM) volume of a whole Drosophila brain. This catalog of 58 LPT neurons (per brain hemisphere) contains many neurons that are described here for the first time and provides a basis for systematic investigation of the circuitry linking self-motion to locomotion control. Leveraging computational anatomy methods, we estimated the visual motion receptive fields of these neurons and compared their tuning to the visual consequence of body rotations and translational movements. We also matched these neurons, in most cases on a one-for-one basis, to stochastically labeled cells in genetic driver lines, to the mirror-symmetric neurons in the same EM brain volume, and to neurons in an additional EM data set. Using cell matches across data sets, we analyzed the integration of optic flow patterns by neurons downstream of the LPTs and find that most central brain neurons establish sharper selectivity for global optic flow patterns than their input neurons. Furthermore, we found that self-motion information extracted from optic flow is processed in distinct regions of the central brain, pointing to diverse foci for the generation of visual behaviors.

}, doi = {10.1101/2023.10.16.562634}, url = {https://doi.org/10.1101/2023.10.16.562634}, author = {Arthur Zhao and Aljoscha Nern and Sanna Koskela and Marisa Dreher and Mert Erginkaya and Connor W Laughland and Henrique DF Ludwig and Alex G Thomson and Judith Hoeller and Ruchi Parekh and Sandro Romani and Davi D Bock and Eugenia Chiappe and Michael B Reiser} } @article {68800, title = {A Connectome of the Male Drosophila Ventral Nerve Cord}, journal = {bioRxiv}, year = {2023}, month = {2023 Jun 06}, abstract = {

Animal behavior is principally expressed through neural control of muscles. Therefore understanding how the brain controls behavior requires mapping neuronal circuits all the way to motor neurons. We have previously established technology to collect large-volume electron microscopy data sets of neural tissue and fully reconstruct the morphology of the neurons and their chemical synaptic connections throughout the volume. Using these tools we generated a dense wiring diagram, or connectome, for a large portion of the\ Drosophila\ central brain. However, in most animals, including the fly, the majority of motor neurons are located outside the brain in a neural center closer to the body, i.e. the mammalian spinal cord or insect ventral nerve cord (VNC). In this paper, we extend our effort to map full neural circuits for behavior by generating a connectome of the VNC of a male fly.

}, doi = {10.1101/2023.06.05.543757}, url = {https://doi.org/10.1101/2023.06.05.543757}, author = {Shin-ya Takemura and Kenneth J Hayworth and Gary B Huang and Michal Januszewski and Zhiyuan Lu and Elizabeth C Marin and Stephan Preibisch and C Shan Xu and John Bogovic and Andrew S Champion and Han S J Cheong and Marta Costa and Katharina Eichler and William Katz and Christopher Knecht and Feng Li and Billy J Morris and Christopher Ordish and Patricia K Rivlin and Philipp Schlegel and Kazunori Shinomiya and Tomke Sturner and Ting Zhao and Griffin Badalamente and Dennis Bailey and Paul Brooks and Brandon S Canino and Jody Clements and Michael Cook and Octave Duclos and Christopher R Dunne and Kelli Fairbanks and Siqi Fang and Samantha Finley-May and Audrey Francis and Reed George and Marina Gkantia and Kyle Harrington and Gary Patrick Hopkins and Joseph Hsu and Philip M Hubbard and Alexandre Javier and Dagmar Kainmueller and Wyatt Korff and Julie Kovalyak and Dominik Krzeminski and Shirley A Lauchie and Alanna Lohff and Charli Maldonado and Emily A Manley and Caroline Mooney and Erika Neace and Matthew Nichols and Omotara Ogundeyi and Nneoma Okeoma and Tyler Paterson and Elliott Phillips and Emily M Phillips and Caitlin Ribeiro and Sean M Ryan and Jon Thomson Rymer and Anne K Scott and Ashley L Scott and David Shepherd and Aya Shinomiya and Claire Smith and Alia Suleiman and Satoko Takemura and Iris Talebi and Imaan F M Tamimi and Eric T Trautman and Lowell Umayam and John J Walsh and Tansy Yang and Gerald M Rubin and Louis K Scheffer and Jan Funke and Stephan Saalfeld and Harald F Hess and Stephen M Plaza and Gwyneth M Card and Gregory S X E Jefferis and Stuart Berg} } @article {68657, title = {Cristae formation is a mechanical buckling event controlled by the inner membrane lipidome}, journal = {bioRxiv}, year = {2023}, month = {2023 Mar 15}, abstract = {

The inner mitochondrial membrane (IMM) is the site of bulk ATP generation in cells and has a broadly conserved lipid composition enriched in unsaturated phospholipids and cardiolipin (CL). While proteins that shape the IMM and its characteristic cristae membranes (CM) have been defined, specific mechanisms by which mitochondrial lipids dictate its structure and function have yet to be elucidated. Here we combine experimental lipidome dissection with multi-scale modeling to investigate how lipid interactions shape CM morphology and ATP generation. When modulating fatty acid unsaturation in engineered yeast strains, we observed that loss of di-unsaturated phospholipids (PLs) led to a breakpoint in IMM topology and respiratory capacity. We found that PL unsaturation modulates the organization of ATP synthases that shape cristae ridges. Based on molecular modeling of mitochondrial-specific membrane adaptations, we hypothesized that conical lipids like CL buffer against the effects of saturation on the IMM. In cells, we discovered that loss of CL collapses the IMM at intermediate levels of PL saturation, an effect that is independent of ATP synthase oligomerization. To explain this interaction, we employed a continuum modeling approach, finding that lipid and protein-mediated curvatures are predicted to act in concert to form curved membranes in the IMM. The model highlighted a snapthrough instability in cristae tubule formation, which could drive IMM collapse upon small changes in composition. The interaction between CL and di-unsaturated PLs suggests that growth conditions that alter the fatty acid pool, such as oxygen availability, could define CL function. While loss of CL only has a minimal phenotype under standard laboratory conditions, we show that its synthesis is essential under microaerobic conditions that better mimic natural yeast fermentation. Lipid and protein-mediated mechanisms of curvature generation can thus act together to support mitochondrial architecture under changing environments.

}, doi = {10.1101/2023.03.13.532310}, url = {https://doi.org/10.1101/2023.03.13.532310}, author = {Kailash Venkatraman and Christopher T Lee and Guadalupe C. Garcia and Arijit Mahapatra and Guy Perkins and Keun-Young Kim and Hilda Amalia Pasolli and Sebastien Phan and Jennifer Lippincott-Schwartz and Mark Ellisman and Padmini Rangamani and Itay Budin} } @article {68467, title = {De novo protein identification in mammalian sperm using high-resolution in situ cryo-electron tomography}, journal = {Cell}, volume = {186}, year = {2023}, month = {2023 Nov 09}, pages = {5041-5053.e19}, abstract = {

Understanding molecular mechanisms of cellular pathways requires knowledge of the identities of participating proteins, their cellular localization and their 3D structures. Contemporary workflows typically require multiple techniques to identify target proteins, track their localization using fluorescence microscopy, followed by\ in vitro\ structure determination. To identify mammal-specific sperm proteins and understand their functions, we developed a visual proteomics workflow to directly address these challenges. Our\ in situ\ cryo-electron tomography and subtomogram averaging provided 6.0 {\r A} resolution reconstructions of axonemal microtubules and their associated proteins. The well-resolved secondary and tertiary structures allowed us to computationally match, in an unbiased manner, novel densities in our 3D reconstruction maps with 21,615 AlphaFold2-predicted protein models of the mouse proteome. We identified Tektin 5, CCDC105 and SPACA9 as novel microtubule inner proteins that form an extensive network crosslinking the lumen of microtubule and existing proteins. Additional biochemical and mass spectrometry analyses helped validate potential candidates. The novel axonemal sperm structures identified by this approach form an extensive interaction network within the lumen of microtubules, suggesting they have a role in the mechanical and elastic properties of the microtubule filaments required for the vigorous beating motions of flagella.

}, doi = {10.1016/j.cell.2023.09.017}, url = {https://doi.org/10.1016/j.cell.2023.09.017}, author = {Zhen Chen and Momoko Shiozaki and Kelsey M. Haas and Shumei Zhao and Caiying Guo and Benjamin J. Polacco and Zhiheng Yu and Nevan J. Krogan and Robyn M. Kaake and Ronald D. Vale and David A. Agard} } @article {68664, title = {Differential Encoding of Trace and Delay Fear Memory in the Entorhinal Cortex.}, journal = {Experimental Neurobiology}, volume = {32}, year = {2023}, month = {2023 Feb 28}, pages = {20-30}, abstract = {

Trace fear conditioning is characterized by a stimulus-free trace interval (TI) between the conditioned stimulus (CS) and the unconditioned stimulus (US), which requires an array of brain structures to support the formation and storage of associative memory. The entorhinal cortex (EC) has been proposed to provide essential neural code for resolving temporal discontinuity in conjunction with the hippocampus. However, how the CS and TI are encoded at the neuronal level in the EC is not clear. In Exp. 1, we tested the effect of bilateral pre-training electrolytic lesions of EC on trace vs. delay fear conditioning using rats as subjects. We found that the lesions impaired the acquisition of trace but not delay fear conditioning confirming that EC is a critical brain area for trace fear memory formation. In Exp. 2, single-unit activities from EC were recorded during the pre-training baseline and post-training retention sessions following trace or delay conditioning. The recording results showed that a significant proportion of the EC neurons modulated their firing during TI after the trace conditioning, but not after the delay fear conditioning. Further analysis revealed that the majority of modulated units decreased the firing rate during the TI or the CS. Taken together, these results suggest that EC critically contributes to trace fear conditioning by modulating neuronal activity during the TI to facilitate the association between the CS and US across a temporal gap.

}, issn = {1226-2560}, doi = {10.5607/en22042}, author = {Kong, Mi-Seon and Kim, Namsoo and Jo, Kyeong Im and Kim, Sung-Phil and Choi, June-Seek} } @article {67959, title = {Dimensionality reduction of calcium-imaged neuronal population activity}, journal = {Nature Computational Science}, year = {2023}, month = {2023 Jan 01}, abstract = {

Calcium imaging has been widely adopted for its ability to record from large neuronal populations. To summarize the time course of neural activity, dimensionality reduction methods, which have been applied extensively to population spiking activity, may be particularly useful. However, it is unclear if the dimensionality reduction methods applied to spiking activity are appropriate for calcium imaging. We thus carried out a systematic study of design choices based on standard dimensionality reduction methods. We also developed a novel method to perform deconvolution and dimensionality reduction simultaneously (termed CILDS). CILDS most accurately recovered the single-trial, low-dimensional time courses from calcium imaging that would have been recovered from spiking activity. CILDS also outperformed the other methods on calcium imaging recordings from larval zebrafish and mice. More broadly, this study represents a foundation for summarizing calcium imaging recordings of large neuronal populations using dimensionality reduction in diverse experimental settings.

}, doi = {10.1038/s43588-022-00390-2}, url = {https://doi.org/10.1038/s43588-022-00390-2}, author = {Tze Hui Koh and William E. Bishop and Takashi Kawashima and Brian B. Jeon and Ranjani Srinivasan and Sandra J. Kuhlman and Misha B. Ahrens and Steven M. Chase and Byron M. Yu} } @article {68634, title = {Distinct mobility patterns of BRCA2 molecules at DNA damage sites}, journal = {Nucleic Acids Res.}, year = {2023}, month = {2024 Jul 02}, pages = {gkae559}, abstract = {

BRCA2 is an essential tumor suppressor protein involved in promoting faithful repair of DNA lesions. The activity of BRCA2 needs to be tuned precisely to be active when and where it is needed. Here, we quantified the spatio-temporal dynamics of BRCA2 in living cells using aberration-corrected multifocal microscopy (acMFM). Using multicolor imaging to identify DNA damage sites, we were able to quantify its dynamic motion patterns in the nucleus and at DNA damage sites. While a large fraction of BRCA2 molecules localized near DNA damage sites appear immobile, an additional fraction of molecules exhibits subdiffusive motion, providing a potential mechanism to retain an increased number of molecules at DNA lesions. Super-resolution microscopy revealed inhomogeneous localization of BRCA2 relative to other DNA repair factors at sites of DNA damage. This suggests the presence of multiple nanoscale compartments in the chromatin surrounding the DNA lesion, which could play an important role in the contribution of BRCA2 to the regulation of the repair process.

}, doi = {10.1093/nar/gkae559 }, url = {https://doi.org/10.1093/nar/gkae559 }, author = {Maarten W. Paul and Jesse Aaron and Eric Wait and Romano M. van Genderen and Ihor Smal and Teng-Leong Chew and Roland Kanaar and Claire Wyman} } @article {69091, title = {Distributed representations of innate behaviors in the hypothalamus do not predict specialized functional centers.}, journal = {bioRxiv}, year = {2023}, month = {2023 Nov 21}, abstract = {

Survival behaviors are orchestrated by hardwired circuits located in deep subcortical brain regions, most prominently the hypothalamus. Artificial activation of spatially localized, genetically defined hypothalamic cell populations is known to trigger distinct behaviors, suggesting a nucleus-centered organization of behavioral control. However, no study has investigated the hypothalamic representation of innate behaviors using unbiased, large-scale single neuron recordings. Here, using custom silicon probes, we performed recordings across the rostro-caudal extent of the medial hypothalamus in freely moving animals engaged in a diverse array of social and predator defense ({\textquotedblleft}fear{\textquotedblright}) behaviors. Nucleus-averaged activity revealed spatially distributed generic {\textquotedblleft}ignition signals{\textquotedblright} that occurred at the onset of each behavior, and did not identify sparse, nucleus-specific behavioral representations. Single-unit analysis revealed that social and fear behavior classes are encoded by activity in distinct sets of spatially distributed neuronal ensembles spanning the entire hypothalamic rostro-caudal axis. Individual ensemble membership, however, was drawn from neurons in 3-4 adjacent nuclei. Mixed selectivity was identified as the most prevalent mode of behavior representation by individual hypothalamic neurons. Encoding models indicated that a significant fraction of the variance in single neuron activity is explained by behavior. This work reveals that innate behaviors are encoded in the hypothalamus by activity in spatially distributed neural ensembles that each span multiple neighboring nuclei, complementing the prevailing view of hypothalamic behavioral control by single nucleus-restricted cell types derived from perturbational studies.

}, doi = {10.1101/2023.11.21.568163}, url = {https://doi.org/10.1101/2023.11.21.568163}, author = {Stefanos Stagkourakis and Giada Spigolon and Markus Marks and Michael Feyder and Joseph Kim and Pietro Perona and Marius Pachitariu and David J. Anderson} } @article {68786, title = {Distributing task-related neural activity across a cortical network through task-independent connections.}, journal = {Nature Communications}, volume = {14}, year = {2023}, month = {2023 May 18}, pages = {2851}, abstract = {

Task-related neural activity is widespread across populations of neurons during goal-directed behaviors. However, little is known about the synaptic reorganization and circuit mechanisms that lead to broad activity changes. Here we trained a subset of neurons in a spiking network with strong synaptic interactions to reproduce the activity of neurons in the motor cortex during a decision-making task. Task-related activity, resembling the neural data, emerged across the network, even in the untrained neurons. Analysis of trained networks showed that strong untrained synapses, which were independent of the task and determined the dynamical state of the network, mediated the spread of task-related activity. Optogenetic perturbations suggest that the motor cortex is strongly-coupled, supporting the applicability of the mechanism to cortical networks. Our results reveal a cortical mechanism that facilitates distributed representations of task-variables by spreading the activity from a subset of plastic neurons to the entire network through task-independent strong synapses.

}, keywords = {Action Potentials, Models, Neurological, Neurons, Synapses}, issn = {2041-1723}, doi = {10.1038/s41467-023-38529-y}, author = {Kim, Christopher M and Finkelstein, Arseny and Chow, Carson C and Svoboda, Karel and Darshan, Ran} } @article {68654, title = {Drosophila antennae are dispensable for gravity orientation}, journal = {bioRxiv}, year = {2023}, month = {2023 Mar 10}, abstract = {

The nearly constant downward force of gravity has powerfully shaped the behaviors of many organisms [1]. Walking flies readily orient against gravity in a behavior termed negative gravitaxis. In\ Drosophila\ this behavior is studied by observing the position of flies in vials [2{\textendash}4] or simple mazes [5{\textendash}9]. These assays have been used to conduct forward-genetic screens [5,\ 6,\ 8] and as simple tests of locomotion deficits [10{\textendash}12]. Despite this long history of investigation, the sensory basis of gravitaxis is largely unknown [1]. Recent studies have implicated the antennae as a major mechanosensory input [3,\ 4], but many details remain unclear. Fly orientation behavior is expected to depend on the direction and amplitude of the gravitational pull, but little is known about the sensitivity of flies to these features of the environment. Here we directly measure the gravity-dependent orientation behavior of flies walking on an adjustable tilted platform, that is inspired by previous insect studies [13{\textendash}16]. In this arena, flies can freely orient with respect to gravity. Our findings indicate that flies are exquisitely sensitive to the direction of gravity{\textquoteright}s pull. Surprisingly, this orientation behavior does not require antennal mechanosensory input, suggesting that other sensory structures must be involved.

}, doi = {10.1101/2023.03.08.531317}, url = {https://doi.org/10.1101/2023.03.08.531317}, author = {Nikolay Kladt and Michael B. Reiser} } @article {69021, title = {Extracellular glutamate and GABA transients at the transition from interictal spiking to seizures.}, journal = {Brain}, year = {2023}, month = {2023 Oct 03}, abstract = {

Focal epilepsy is associated with intermittent brief population discharges (interictal spikes), which resemble sentinel spikes that often occur at the onset of seizures. Why interictal spikes self-terminate whilst seizures persist and propagate is incompletely understood. We used fluorescent glutamate and GABA sensors in an awake rodent model of neocortical seizures to resolve the spatiotemporal evolution of both neurotransmitters in the extracellular space. Interictal spikes were accompanied by brief glutamate transients which were maximal at the initiation site and rapidly propagated centrifugally. GABA transients lasted longer than glutamate transients and were maximal \~{}1.5 mm from the focus where they propagated centripetally. Prior to seizure initiation GABA transients were attenuated, whilst glutamate transients increased, consistent with a progressive failure of local inhibitory restraint. As seizures increased in frequency, there was a gradual increase in the spatial extent of spike-associated glutamate transients associated with interictal spikes. Neurotransmitter imaging thus reveals a progressive collapse of an annulus of feed-forward GABA release, allowing seizures to escape from local inhibitory restraint.

}, issn = {1460-2156}, doi = {10.1093/brain/awad336}, author = {Shimoda, Yoshiteru and Leite, Marco and Graham, Robert T and Marvin, Jonathan S and Hasseman, Jeremy and Kolb, Ilya and Looger, Loren L and Magloire, Vincent and Kullmann, Dimitri M} } @article {68665, title = {Fast and sensitive GCaMP calcium indicators for imaging neural populations.}, journal = {Nature}, year = {2023}, month = {2023 Mar 15}, abstract = {

Calcium imaging with protein-based indicators is widely used to follow neural activity in intact nervous systems, but current protein sensors report neural activity at timescales much slower than electrical signalling and are limited by trade-offs between sensitivity and kinetics. Here we used large-scale screening and structure-guided mutagenesis to develop and optimize several fast and sensitive GCaMP-type indicators. The resulting {\textquoteright}jGCaMP8{\textquoteright} sensors, based on the calcium-binding protein calmodulin and a fragment of endothelial nitric oxide synthase, have ultra-fast kinetics (half-rise times of 2 ms) and the highest sensitivity for neural activity reported for a protein-based calcium sensor. jGCaMP8 sensors will allow tracking of large populations of neurons on timescales relevant to neural computation.

}, issn = {1476-4687}, doi = {10.1038/s41586-023-05828-9}, author = {Zhang, Yan and Rozsa, Marton and Liang, Yajie and Bushey, Daniel and Wei, Ziqiang and Zheng, Jihong and Reep, Daniel and Broussard, Gerard Joey and Tsang, Arthur and Tsegaye, Getahun and Narayan, Sujatha and Obara, Christopher J and Lim, Jing-Xuan and Patel, Ronak and Zhang, Rongwei and Ahrens, Misha B and Turner, Glenn C and Wang, Samuel S-H and Korff, Wyatt L and Schreiter, Eric R and Svoboda, Karel and Hasseman, Jeremy P and Kolb, Ilya and Looger, Loren L} } @article {68521, title = {From primordial clocks to circadian oscillators}, journal = {Nature}, year = {2023}, month = {2023 Apr 01}, abstract = {

Circadian rhythms play an essential role in many biological processes and surprisingly only three prokaryotic proteins are required to constitute a true post-translational circadian oscillator. The evolutionary history of the three Kai proteins indicates that KaiC is the oldest member and central component of the clock, with subsequent additions of KaiB and KaiA to regulate its phosphorylation state for time synchronization. The canonical KaiABC system in cyanobacteria is well understood, but little is known about more ancient systems that possess just KaiBC, except for reports that they might exhibit a basic, hourglass-like timekeeping mechanism. Here, we investigate the primordial circadian clock in\ Rhodobacter sphaeroides\ (RS) that contains only KaiBC to elucidate its inner workings despite the missing KaiA. Using a combination X-ray crystallography and cryo-EM we find a novel dodecameric fold for KaiCRS\ where two hexamers are held together by a coiled-coil bundle of 12 helices. This interaction is formed by the C-terminal extension of KaiCRS\ and serves as an ancient regulatory moiety later superseded by KaiA. A coiled-coil register shift between daytime- and nighttime-conformations is connected to the phosphorylation sites through a long-range allosteric network that spans over 160 {\r A}. Our kinetic data identify the difference in ATP-to-ADP ratio between day and night as the environmental cue that drives the clock and further unravels mechanistic details that shed light on the evolution of self-sustained oscillators.

}, doi = {10.1038/s41586-023-05836-9}, url = {https://doi.org/10.1038/s41586-023-05836-9}, author = {Warintra Pitsawong and Ricardo A. P. P{\'a}dua and Timothy Grant and Marc Hoemberger and Renee Otten and Niels Bradshaw and Nikolaus Grigorieff and Dorothee Kern} } @article {68772, title = {Glutamate indicators with improved activation kinetics and localization for imaging synaptic transmission.}, journal = {Nature Methods}, volume = {20}, year = {2023}, month = {2023 Jun 01}, abstract = {

The fluorescent glutamate indicator iGluSnFR enables imaging of neurotransmission with genetic and molecular specificity. However, existing iGluSnFR variants exhibit low in vivo signal-to-noise ratios, saturating activation kinetics and exclusion from postsynaptic densities. Using a multiassay screen in bacteria, soluble protein and cultured neurons, we generated variants with improved signal-to-noise ratios and kinetics. We developed surface display constructs that improve iGluSnFR{\textquoteright}s nanoscopic localization to postsynapses. The resulting indicator iGluSnFR3 exhibits rapid nonsaturating activation kinetics and reports synaptic glutamate release with decreased saturation and increased specificity versus extrasynaptic signals in cultured neurons. Simultaneous imaging and electrophysiology at individual boutons in mouse visual cortex showed that iGluSnFR3 transients report single action potentials with high specificity. In vibrissal sensory cortex layer 4, we used iGluSnFR3 to characterize distinct patterns of touch-evoked feedforward input from thalamocortical boutons and both feedforward and recurrent input onto L4 cortical neuron dendritic spines.

}, issn = {1548-7105}, doi = {10.1038/s41592-023-01863-6}, author = {Aggarwal, Abhi and Liu, Rui and Chen, Yang and Ralowicz, Amelia J and Bergerson, Samuel J and Tomaska, Filip and Mohar, Boaz and Hanson, Timothy L and Hasseman, Jeremy P and Reep, Daniel and Tsegaye, Getahun and Yao, Pantong and Ji, Xiang and Kloos, Marinus and Walpita, Deepika and Patel, Ronak and Mohr, Manuel A and Tillberg, Paul W and Looger, Loren L and Marvin, Jonathan S and Hoppa, Michael B and Konnerth, Arthur and Kleinfeld, David and Schreiter, Eric R and Podgorski, Kaspar} } @article {68957, title = {Hydrophobic interactions dominate the recognition of a KRAS G12V neoantigen.}, journal = {Nature Communications}, volume = {14}, year = {2023}, month = {2023 Aug 21}, pages = {5063}, abstract = {

Specificity remains a major challenge to current therapeutic strategies for cancer. Mutation associated neoantigens (MANAs) are products of genetic alterations, making them highly specific therapeutic targets. MANAs are HLA-presented (pHLA) peptides derived from intracellular mutant proteins that are otherwise inaccessible to antibody-based therapeutics. Here, we describe the cryo-EM structure of an antibody-MANA pHLA complex. Specifically, we determine a TCR mimic (TCRm) antibody bound to its MANA target, the KRAS peptide presented by HLA-A*03:01. Hydrophobic residues appear to account for the specificity of the mutant G12V residue. We also determine the structure of the wild-type G12 peptide bound to HLA-A*03:01, using X-ray crystallography. Based on these structures, we perform screens to validate the key residues required for peptide specificity. These experiments led us to a model for discrimination between the mutant and the wild-type peptides presented on HLA-A*03:01 based exclusively on hydrophobic interactions.

}, keywords = {Antibodies, HLA-A Antigens, Hydrophobic and Hydrophilic Interactions, Proto-Oncogene Proteins p21(ras), Recognition, Psychology}, issn = {2041-1723}, doi = {10.1038/s41467-023-40821-w}, author = {Wright, Katharine M and DiNapoli, Sarah R and Miller, Michelle S and Aitana Azurmendi, P and Zhao, Xiaowei and Yu, Zhiheng and Chakrabarti, Mayukh and Shi, WuXian and Douglass, Jacqueline and Hwang, Michael S and Hsiue, Emily Han-Chung and Mog, Brian J and Pearlman, Alexander H and Paul, Suman and Konig, Maximilian F and Pardoll, Drew M and Bettegowda, Chetan and Papadopoulos, Nickolas and Kinzler, Kenneth W and Vogelstein, Bert and Zhou, Shibin and Gabelli, Sandra B} } @article {68697, title = {Identifying determinants of synaptic specificity by integrating connectomes and transcriptomes}, journal = {bioRxiv}, year = {2023}, month = {2023 Apr 04}, abstract = {

How do developing neurons select their synaptic partners? To identify molecules matching synaptic partners, we integrated the synapse-level connectome of neural circuits with the developmental expression patterns and binding specificities of cell adhesion molecules (CAMs) on pre- and postsynaptic neurons. We focused on parallel synaptic pathways in the Drosophila visual system, in which closely related neurons form synapses onto closely related target neurons. We show that the choice of synaptic partners correlates with the matching expression of receptor-ligand pairs of Beat and Side proteins of the immunoglobulin superfamily (IgSF) CAMs. Genetic analysis demonstrates that these proteins determine the choice between alternative synaptic targets. Combining transcriptomes, connectomes, and protein interactome maps provides a framework to uncover the molecular logic of synaptic connectivity.

}, doi = {10.1101/2023.04.03.534791}, url = {https://doi.org/10.1101/2023.04.03.534791}, author = {Juyoun Yoo and Mark Dombrovski and Parmis Mirshahidi and Aljoscha Nern and Samuel A. LoCascio and S. Lawrence Zipursky and Yerbol Z. Kurmangaliyev} } @article {68832, title = {The Janelia Atalanta plasmids provide a simple and efficient CRISPR/Cas9-mediated homology directed repair platform for Drosophila}, journal = {bioRxiv}, year = {2023}, month = {2023 Jun 17}, abstract = {

Homology-directed repair (HDR) is a powerful tool for modifying genomes in precise ways to address many biological questions. Use of Clustered Regularly Interspersed Short Palindromic Repeats (CRISPR)-Cas9 induced targeted DNA double-strand breakage has substantially simplified use of homology-directed repair to introduce specific perturbations in\ Drosophila, but existing platforms for CRISPR-Cas9-mediated HDR in\ Drosophila\ involve multiple cloning steps and have low efficiency. To simplify cloning of HDR plasmids, we designed a new plasmid platform, the Janelia Atalanta (pJAT) series, that exploits recent advances in dsDNA synthesis to facilitate Gateway cloning of gRNA sequences and homology arms in one step. Surprisingly, the pJAT plasmids yielded considerably higher HDR efficiency (approximately 25\%) than we have observed with other approaches. pJAT plasmids work in multiple\ Drosophila\ species and exhibited such high efficiency that previously impossible experiments in\ Drosophila, such as driving targeted chromosomal inversions, were made possible. We provide pJAT plasmids for a range of commonly performed experiments including targeted insertional mutagenesis, insertion of phiC31-mediated attP landing sites, generation of strains carrying a germ-line source of Cas9, and induction of chromosomal rearrangements. We also provide {\textquotedblleft}empty{\textquotedblright} pJAT plasmids with multiple cloning sites to simplify construction of plasmids with new functionality. The pJAT platform is generic and may facilitate improved efficiency CRISPR-Cas9 HDR in a wide range of model and non-model organisms.

}, doi = {10.1101/2023.06.17.545412}, url = {https://doi.org/10.1101/2023.06.17.545412}, author = {David L. Stern and Elizabeth Kim and Emily L. Behrman} } @article {69053, title = {Lactate biosensors for spectrally and spatially multiplexed fluorescence imaging.}, journal = {Nature Communications}, volume = {14}, year = {2023}, month = {2023 Oct 27}, pages = {6598}, abstract = {

L-Lactate is increasingly appreciated as a key metabolite and signaling molecule in mammals. However, investigations of the inter- and intra-cellular dynamics of L-lactate are currently hampered by the limited selection and performance of L-lactate-specific genetically encoded biosensors. Here we now report a spectrally and functionally orthogonal pair of high-performance genetically encoded biosensors: a green fluorescent extracellular L-lactate biosensor, designated eLACCO2.1, and a red fluorescent intracellular L-lactate biosensor, designated R-iLACCO1. eLACCO2.1 exhibits excellent membrane localization and robust fluorescence response. To the best of our knowledge, R-iLACCO1 and its affinity variants exhibit larger fluorescence responses than any previously reported intracellular L-lactate biosensor. We demonstrate spectrally and spatially multiplexed imaging of L-lactate dynamics by coexpression of eLACCO2.1 and R-iLACCO1 in cultured cells, and in vivo imaging of extracellular and intracellular L-lactate dynamics in mice.

}, keywords = {Animals, Biosensing Techniques, Cells, Cultured, Fluorescence Resonance Energy Transfer, Lactic Acid, Mammals, Mice, Optical Imaging}, issn = {2041-1723}, doi = {10.1038/s41467-023-42230-5}, author = {Nasu, Yusuke and Aggarwal, Abhi and Le, Giang N T and Vo, Camilla Trang and Kambe, Yuki and Wang, Xinxing and Beinlich, Felix R M and Lee, Ashley Bomin and Ram, Tina R and Wang, Fangying and Gorzo, Kelsea A and Kamijo, Yuki and Boisvert, Marc and Nishinami, Suguru and Kawamura, Genki and Ozawa, Takeaki and Toda, Hirofumi and Gordon, Grant R and Ge, Shaoyu and Hirase, Hajime and Nedergaard, Maiken and Paquet, Marie-Eve and Drobizhev, Mikhail and Podgorski, Kaspar and Campbell, Robert E} } @article {68607, title = {Large-scale brain-wide neural recording in nonhuman primates}, journal = {bioRxiv}, year = {2023}, month = {2023 Feb 04}, abstract = {

High-density, integrated silicon electrodes have begun to transform systems neuroscience, by enabling large-scale neural population recordings with single cell resolution. Existing technologies, however, have provided limited functionality in nonhuman primate species such as macaques, which offer close models of human cognition and behavior. Here, we report the design, fabrication, and performance of Neuropixels 1.0-NHP, a high channel count linear electrode array designed to enable large-scale simultaneous recording in superficial and deep structures within the macaque or other large animal brain. These devices were fabricated in two versions: 4416 electrodes along a 45 mm shank, and 2496 along a 25 mm shank. For both versions, users can programmably select 384 channels, enabling simultaneous multi-area recording with a single probe. We demonstrate recording from over 3000 single neurons within a session, and simultaneous recordings from over 1000 neurons using multiple probes. This technology represents a significant increase in recording access and scalability relative to existing technologies, and enables new classes of experiments involving fine-grained electrophysiological characterization of brain areas, functional connectivity between cells, and simultaneous brain-wide recording at scale.

}, doi = {10.1101/2023.02.01.526664}, url = {https://doi.org/10.1101/2023.02.01.526664}, author = {Eric M. Trautmann and Janis K. Hesse and Gabriel M. Stine and Ruobing Xia and Shude Zhu and Daniel J. O{\textquoteright}Shea and Bill Karsh and Jennifer Colonell and Frank F. Lanfranchi and Saurabh Vyas and Andrew Zimnik and Natalie A. Steinmann and Daniel A. Wagenaar and Alexandru Andrei and Carolina Mora Lopez and John O{\textquoteright}Callaghan and Jan Putzeys and Bogdan C. Raducanu and Marleen Welkenhuysen and Mark Churchland and Tirin Moore and Michael Shadlen and Krishna Shenoy and Doris Tsao and Barundeb Dutta and Timothy Harris} } @article {68774, title = {A leaky integrate-and-fire computational model based on the connectome of the entire adult Drosophila brain reveals insights into sensorimotor processing}, journal = {bioRxiv}, year = {2023}, month = {2023 May 02}, abstract = {

The forthcoming assembly of the adult\ Drosophila melanogaster\ central brain connectome, containing over 125,000 neurons and 50 million synaptic connections, provides a template for examining sensory processing throughout the brain. Here, we create a leaky integrate-and-fire computational model of the entire\ Drosophila\ brain, based on neural connectivity and neurotransmitter identity, to study circuit properties of feeding and grooming behaviors. We show that activation of sugar-sensing or water-sensing gustatory neurons in the computational model accurately predicts neurons that respond to tastes and are required for feeding initiation. Computational activation of neurons in the feeding region of the\ Drosophila\ brain predicts those that elicit motor neuron firing, a testable hypothesis that we validate by optogenetic activation and behavioral studies. Moreover, computational activation of different classes of gustatory neurons makes accurate predictions of how multiple taste modalities interact, providing circuit-level insight into aversive and appetitive taste processing. Our computational model predicts that the sugar and water pathways form a partially shared appetitive feeding initiation pathway, which our calcium imaging and behavioral experiments confirm. Additionally, we applied this model to mechanosensory circuits and found that computational activation of mechanosensory neurons predicts activation of a small set of neurons comprising the antennal grooming circuit that do not overlap with gustatory circuits, and accurately describes the circuit response upon activation of different mechanosensory subtypes. Our results demonstrate that modeling brain circuits purely from connectivity and predicted neurotransmitter identity generates experimentally testable hypotheses and can accurately describe complete sensorimotor transformations.

}, doi = {10.1101/2023.05.02.539144}, url = {https://doi.org/10.1101/2023.05.02.539144}, author = {Philip K. Shiu and Gabriella R. Sterne and Nico Spiller and Romain Franconville and Andrea Sandoval and Joie Zhou and Neha Simha and Chan Hyuk Kang and Seongbong Yu and Jinseop S. Kim and Sven Dorkenwald and Arie Matsliah and Philipp Schlegel and Szi-chieh Yu and Claire E. McKellar and Amy Sterling and Marta Costa and Katharina Eichler and Gregory S.X.E. Jefferis and Mala Murthy and Alexander Shakeel Bates and Nils Eckstein and Jan Funke and Salil S. Bidaye and Stefanie Hampel and Andrew M. Seeds and Kristin Scott} } @article {69009, title = {Low-latency extracellular spike assignment for high-density electrodes at single-neuron resolution}, journal = {bioRxiv}, year = {2023}, month = {2023 Sep 15}, abstract = {

Real-time neural signal processing is essential for brain-machine interfaces and closed-loop neuronal perturbations. However, most existing applications sacrifice cell-specific identity and temporal spiking information for speed. We developed a hybrid hardware-software system that utilizes a Field Programmable Gate Array (FPGA) chip to acquire and process data in parallel, enabling individual spikes from many simultaneously recorded neurons to be assigned single-neuron identities with 1-millisecond latency. The FPGA assigns labels, validated with ground-truth data, by comparing multichannel spike waveforms from tetrode or silicon probe recordings to a spike-sorted model generated offline in software. This platform allowed us to rapidly inactivate a region in vivo based on spikes from an upstream neuron before these spikes could excite the downstream region. Furthermore, we could decode animal location within 3 ms using data from a population of individual hippocampal neurons. These results demonstrate our system{\textquoteright}s suitability for a broad spectrum of research and clinical applications.

}, doi = {10.1101/2023.09.14.557854}, url = {https://doi.org/10.1101/2023.09.14.557854}, author = {Chongxi Lai and Dohoung Kim and Brian Lustig and Shinsuke Tanaka and Brian Barbarits and Lakshmi Narayan and Jennifer Colonell and Ole Paulsen and Albert K. Lee and Timothy D. Harris} } @article {68791, title = {Mapping memories: pulse-chase labeling reveals AMPA receptor dynamics during memory formation.}, journal = {bioRxiv}, year = {2023}, month = {2023 May 29}, abstract = {

A tool to map changes in synaptic strength during a defined time window could provide powerful insights into the mechanisms governing learning and memory. We developed a technique, Extracellular Protein Surface Labeling in Neurons (EPSILON), to map α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor (AMPAR) insertion\ in vivo\ by pulse-chase labeling of surface AMPARs with membrane-impermeable dyes. This approach allows for single-synapse resolution maps of plasticity in genetically targeted neurons during memory formation. We investigated the relationship between synapse-level and cell-level memory encodings by mapping synaptic plasticity and cFos expression in hippocampal CA1 pyramidal cells upon contextual fear conditioning (CFC). We observed a strong correlation between synaptic plasticity and cFos expression, suggesting a synaptic mechanism for the association of cFos expression with memory engrams. The EPSILON technique is a useful tool for mapping synaptic plasticity and may be extended to investigate trafficking of other transmembrane proteins.

}, doi = {10.1101/2023.05.26.541296}, url = {https://doi.org/10.1101/2023.05.26.541296}, author = {Doyeon Kim and Pojeong Park and Xiuyuan Li and J. David Wong Campos and He Tian and Eric M. Moult and Jonathan B. Grimm and Luke Lavis and Adam E. Cohen} } @article {68773, title = {Meta-learning in head fixed mice navigating in virtual reality: A Behavioral Analysis}, journal = {bioRxiv}, year = {2023}, month = {2023 May 02}, abstract = {

Animals can learn general task structures and use them to solve new problems with novel sensory specifics. This capacity of {\textquoteleft}learning to learn{\textquoteright}, or meta-learning, is difficult to achieve in artificial systems, and the mechanisms by which it is achieved in animals are unknown. As a step toward enabling mechanistic studies, we developed a behavioral paradigm that demonstrates meta-learning in head-fixed mice. We trained mice to perform a two-alternative forced-choice task in virtual reality (VR), and successively changed the visual cues that signaled reward location. Mice showed increased learning speed in both cue generalization and serial reversal tasks. During reversal learning, behavior exhibited sharp transitions, with the transition occurring earlier in each successive reversal. Analysis of motor patterns revealed that animals utilized similar motor programs to execute the same actions in response to different cues but modified the motor programs during reversal learning. Our study demonstrates that mice can perform meta-learning tasks in VR, thus opening up opportunities for future mechanistic studies.

}, doi = {10.1101/2023.05.01.538936}, url = {https://doi.org/10.1101/2023.05.01.538936}, author = {Xinyu Zhao and Rachel Gattoni and Andrea Kozlosky and Angela Jacobs and Colin Morrow and Sarah Lindo and Nelson Spruston} } @article {68836, title = {Method To Diversify Cyanine Chromophore Functionality Enables Improved Biomolecule Tracking and Intracellular Imaging.}, journal = {Journal of the American Chemical Society}, volume = {145}, year = {2023}, month = {2023 Jun 27}, pages = {14647{\textendash}14659}, abstract = {

Heptamethine indocyanines are invaluable probes for near-infrared (NIR) imaging. Despite broad use, there are only a few synthetic methods to assemble these molecules, and each has significant limitations. Here, we report the use of pyridinium benzoxazole (PyBox) salts as heptamethine indocyanine precursors. This method is high yielding, simple to implement, and provides access to previously unknown chromophore functionality. We applied this method to create molecules to address two outstanding objectives in NIR fluorescence imaging. First, we used an iterative approach to develop molecules for protein-targeted tumor imaging. When compared to common NIR fluorophores, the optimized probe increases the tumor specificity of monoclonal antibody (mAb) and nanobody conjugates. Second, we developed cyclizing heptamethine indocyanines with the goal of improving cellular uptake and fluorogenic properties. By modifying both the electrophilic and nucleophilic components, we demonstrate that the solvent sensitivity of the ring-open/ring-closed equilibrium can be modified over a wide range. We then show that a chloroalkane derivative of a compound with tuned cyclization properties undergoes particularly efficient no-wash live cell imaging using organelle-targeted HaloTag self-labeling proteins. Overall, the chemistry reported here broadens the scope of accessible chromophore functionality, and, in turn, enables the discovery of NIR probes with promising properties for advanced imaging applications.

}, issn = {1520-5126}, doi = {10.1021/jacs.3c01765}, author = {Usama, Syed Muhammad and Marker, Sierra C and Li, Dong-Hao and Caldwell, Donald R and Stroet, Marcus and Patel, Nimit L and Tebo, Alison G and Hernot, Sophie and Kalen, Joseph D and Schnermann, Martin} } @article {68469, title = {Multimodal mapping of cell types and projections in the central nucleus of the amygdala}, journal = {eLife}, year = {2023}, month = {2023 Jan 20}, abstract = {

The central nucleus of the amygdala (CEA) is a brain region that integrates external and internal sensory information and executes innate and adaptive behaviors through distinct output pathways. Despite its complex functions, the diversity of molecularly defined neuronal types in the CEA and their contributions to major axonal projection targets have not been examined systematically. Here, we performed single-cell RNA-sequencing (scRNA-Seq) to classify molecularly defined cell types in the CEA and identified marker-genes to map the location of these neuronal types using\ expansion\ assisted iterative fluorescence in\ situ\ hybridization (EASI-FISH). We developed new methods to integrate EASI-FISH with 5-plex retrograde axonal labeling to determine the spatial, morphological, and connectivity properties of \~{}30,000 molecularly defined CEA neurons. Our study revealed spatio-molecular organization of the CEA, with medial and lateral CEA associated with distinct cell families. We also found a long-range axon projection network from the CEA, where target regions receive inputs from multiple molecularly defined cell types. Axon collateralization was found primarily among projections to hindbrain targets, which are distinct from forebrain projections. This resource reports marker-gene combinations for molecularly defined cell types and axon-projection types, which will be useful for selective interrogation of these neuronal populations to study their contributions to the diverse functions of the CEA.

}, doi = {10.7554/eLife.84262}, url = {https://doi.org/10.7554/eLife.84262}, author = {Yuhan Wang and Sabine Krabbe and Mark Eddison and Fredrick E. Henry and Greg Fleishman and Andrew L. Lemire and Lihua Wang and Wyatt Korff and Paul W. Tillberg and Andreas L{\"u}thi and Scott M. Sternson} } @article {69058, title = {Nanometer-scale views of visual cortex reveal anatomical features of primary cilia poised to detect synaptic spillover}, journal = {bioRxiv}, year = {2023}, month = {2023 Nov 01}, abstract = {

A primary cilium is a thin membrane-bound extension off a cell surface that contains receptors for perceiving and transmitting signals that modulate cell state and activity. While many cell types have a primary cilium, little is known about primary cilia in the brain, where they are less accessible than cilia on cultured cells or epithelial tissues and protrude from cell bodies into a deep, dense network of glial and neuronal processes. Here, we investigated cilia frequency, internal structure, shape, and position in large, high-resolution transmission electron microscopy volumes of mouse primary visual cortex. Cilia extended from the cell bodies of nearly all excitatory and inhibitory neurons, astrocytes, and oligodendrocyte precursor cells (OPCs), but were absent from oligodendrocytes and microglia. Structural comparisons revealed that the membrane structure at the base of the cilium and the microtubule organization differed between neurons and glia. OPC cilia were distinct in that they were the shortest and contained pervasive internal vesicles only occasionally observed in neuron and astrocyte cilia. Investigating cilia-proximal features revealed that many cilia were directly adjacent to synapses, suggesting cilia are well poised to encounter locally released signaling molecules. The internal anatomy, including microtubule changes and centriole location, defined key structural features including cilium placement and shape. Together, the anatomical insights both within and around neuron and glia cilia provide new insights into cilia formation and function across cell types in the brain.

}, doi = {10.1101/2023.10.31.564838}, url = {https://doi.org/10.1101/2023.10.31.564838}, author = {Carolyn M Ott and Russel Torres and Tung-Sheng Kuan and Aaron T Kuan and JoAnn Buchanan and Leila Elabbady and Sharmishtaa Seshamani and Agnes L Bodor and Forrest C Collman and Davi D Bock and Wei-Chung Allen Lee and Nuno Macarico da Costa and Jennifer Lippincott-Schwartz} } @article {68785, title = {Nanoparticle-based targeting of microglia improves the neural regeneration enhancing effects of immunosuppression in the zebrafish retina.}, journal = {Communications Biology}, volume = {6}, year = {2023}, month = {2023 May 18}, pages = {534}, abstract = {

Retinal M{\"u}ller glia function as injury-induced stem-like cells in zebrafish but not mammals. However, insights gleaned from zebrafish have been applied to stimulate nascent regenerative responses in the mammalian retina. For instance, microglia/macrophages regulate M{\"u}ller glia stem cell activity in the chick, zebrafish, and mouse. We previously showed that post-injury immunosuppression by the glucocorticoid dexamethasone accelerated retinal regeneration kinetics in zebrafish. Similarly, microglia ablation enhances regenerative outcomes in the mouse retina. Targeted immunomodulation of microglia reactivity may therefore enhance the regenerative potential of M{\"u}ller glia for therapeutic purposes. Here, we investigated potential mechanisms by which post-injury dexamethasone accelerates retinal regeneration kinetics, and the effects of dendrimer-based targeting of dexamethasone to reactive microglia. Intravital time-lapse imaging revealed that post-injury dexamethasone inhibited microglia reactivity. The dendrimer-conjugated formulation: (1) decreased dexamethasone-associated systemic toxicity, (2) targeted dexamethasone to reactive microglia, and (3) improved the regeneration enhancing effects of immunosuppression by increasing stem/progenitor proliferation rates. Lastly, we show that the gene rnf2 is required for the enhanced regeneration effect of D-Dex. These data support the use of dendrimer-based targeting of reactive immune cells to reduce toxicity and enhance the regeneration promoting effects of immunosuppressants in the retina.

}, keywords = {Animals, Dendrimers, Dexamethasone, Immunosuppression Therapy, Mammals, Mice, Microglia, Retina, Zebrafish}, issn = {2399-3642}, doi = {10.1038/s42003-023-04898-9}, author = {Emmerich, Kevin and White, David T and Kambhampati, Siva P and Casado, Grace L and Fu, Tian-Ming and Chunawala, Zeeshaan and Sahoo, Arpan and Nimmagadda, Saumya and Krishnan, Nimisha and Saxena, Meera T and Walker, Steven L and Betzig, Eric and Kannan, Rangaramanujam M and Mumm, Jeff S} } @article {68635, title = {OME-Zarr: a cloud-optimized bioimaging file format with international community support.}, journal = {Histochemistry and Cell Biology}, volume = {160}, year = {2023}, month = {2023 Feb 25}, pages = {223-251}, abstract = {

A growing community is constructing a next-generation file format (NGFF) for bioimaging to overcome problems of scalability and heterogeneity. Organized by the Open Microscopy Environment (OME), individuals and institutes across diverse modalities facing these problems have designed a format specification process (OME-NGFF) to address these needs. This paper brings together a wide range of those community members to describe the format itself {\textendash} OME-Zarr {\textendash} along with tools and data resources available today to increase FAIR access and remove barriers in the scientific process. The current momentum offers an opportunity to unify a key component of the bioimaging domain {\textemdash} the file format that underlies so many personal, institutional, and global data management and analysis tasks.

}, doi = {10.1007/s00418-023-02209-1}, url = {https://doi.org/10.1007/s00418-023-02209-1}, author = {Josh Moore and Daniela Basurto-Lozada and S{\'e}bastien Besson and John Bogovic and Eva M. Brown and Jean-Marie Burel and Gustavo de Medeiros and Erin E. Diel and David Gault and Satrajit S. Ghosh and Ilan Gold and Yaroslav O. Halchenko and Matthew Hartley and Dave Horsfall and Mark S. Keller and Mark Kittisopikul and Gabor Kovacs and Ayb{\"u}ke K{\"u}pc{\"u} Yolda{\c s} and Albane le Tournoulx de la Villegeorges and Tong Li and Prisca Liberali and Melissa Linkert and Dominik Lindner and Joel L{\"u}thi and Jeremy Maitin-Shepard and Trevor Manz and Matthew McCormick and Khaled Mohamed and William Moore and Bugra {\"O}zdemir and Constantin Pape and Lucas Pelkmans and Martin Prete and Tobias Pietzsch and Stephan Preibisch and Norman Rzepka and David R. Stirling and Jonathan Striebel and Christian Tischer and Daniel Toloudis and Petr Walczysko and Alan M. Watson and Frances Wong and Kevin A. Yamauchi and Omer Bayraktar and Muzlifah Haniffa and Stephan Saalfeld and Jason R. Swedlow} } @article {68675, title = {Origin of wiring specificity in an olfactory map revealed by neuron type-specific, time-lapse imaging of dendrite targeting.}, journal = {eLife}, volume = {12}, year = {2023}, month = {2023 Mar 28}, abstract = {

How does wiring specificity of neural maps emerge during development? Formation of the adult olfactory glomerular map begins with patterning of projection neuron (PN) dendrites at the early pupal stage. To better understand the origin of wiring specificity of this map, we created genetic tools to systematically characterize dendrite patterning across development at PN type-specific resolution. We find that PNs use lineage and birth order combinatorially to build the initial dendritic map. Specifically, birth order directs dendrite targeting in rotating and binary manners for PNs of the anterodorsal and lateral lineages, respectively. Two-photon- and adaptive optical lattice light-sheet microscope-based time-lapse imaging reveals that PN dendrites initiate active targeting with direction-dependent branch stabilization on the timescale of seconds. Moreover, PNs that are used in both the larval and adult olfactory circuits prune their larval-specific dendrites and re-extend new dendrites simultaneously to facilitate timely olfactory map organization. Our work highlights the power and necessity of type-specific neuronal access and time-lapse imaging in identifying wiring mechanisms that underlie complex patterns of functional neural maps.

}, issn = {2050-084X}, doi = {10.7554/eLife.85521}, author = {Wong, Kenneth Kin Lam and Li, Tongchao and Fu, Tian-Ming and Liu, Gaoxiang and Lyu, Cheng and Kohani, Sayeh and Xie, Qijing and Luginbuhl, David J and Upadhyayula, Srigokul and Betzig, Eric and Luo, Liqun} } @article {68631, title = {Phase separation of Hippo signalling complexes.}, journal = {EMBO Journal}, volume = {42}, year = {2023}, month = {2023 Feb 20}, pages = {e112863}, abstract = {

The Hippo pathway was originally discovered to control tissue growth in Drosophila and includes the Hippo kinase (Hpo; MST1/2 in mammals), scaffold protein Salvador (Sav; SAV1 in mammals) and the Warts kinase (Wts; LATS1/2 in mammals). The Hpo kinase is activated by binding to Crumbs-Expanded (Crb-Ex) and/or Merlin-Kibra (Mer-Kib) proteins at the apical domain of epithelial cells. Here we show that activation of Hpo also involves the formation of supramolecular complexes with properties of a biomolecular condensate, including concentration dependence and\ sensitivity to starvation, macromolecular crowding, or 1,6-hexanediol treatment. Overexpressing Ex or Kib induces formation of micron-scale Hpo condensates in the cytoplasm, rather than at the apical membrane. Several Hippo pathway components contain unstructured low-complexity domains and purified Hpo-Sav complexes undergo phase separation in\ vitro. Formation of Hpo condensates is conserved in human cells. We propose that apical Hpo kinase activation occurs in phase separated "signalosomes" induced by clustering of upstream pathway components.

}, issn = {1460-2075}, doi = {10.15252/embj.2022112863}, author = {Bonello, Teresa T and Cai, Danfeng and Fletcher, Georgina C and Wiengartner, Kyler and Pengilly, Victoria and Lange, Kimberly S and Liu, Zhe and Lippincott-Schwartz, Jennifer and Kavran, Jennifer M and Thompson, Barry J} } @article {68917, title = {Pinpoint: trajectory planning for multi-probe electrophysiology and injections in an interactive web-based 3D environment}, journal = {bioRxiv}, year = {2023}, month = {2023 Jul 15}, abstract = {

Targeting deep brain structures during electrophysiology and injections requires intensive training and expertise. Even with experience, researchers often can{\textquoteright}t be certain that a probe is placed precisely in a target location and this complexity scales with the number of simultaneous probes used in an experiment. Here, we present Pinpoint, open-source software that allows for interactive exploration of stereotaxic insertion plans. Once an insertion plan is created, Pinpoint allows users to save these online and share them with collaborators. 3D modeling tools allow users to explore their insertions alongside rig and implant hardware and ensure plans are physically possible. Probes in Pinpoint can be linked to electronic micro-manipulators allowing real-time visualization of current brain region targets alongside neural data. In addition, Pinpoint can control manipulators to automate and parallelize the insertion process. Compared to previously available software, Pinpoint{\textquoteright}s easy access through web browsers, extensive features, and real-time experiment integration enable more efficient and reproducible recordings.

}, doi = {10.1101/2023.07.14.548952}, url = {https://doi.org/10.1101/2023.07.14.548952}, author = {Daniel Birman and Kenneth J. Yang and Steven J. West and Bill Karsh and Yoni Browning and the International Brain Laboratory and Joshua H. Siegle and Nicholas A. Steinmetz} } @article {68770, title = {The power of peer networking for improving STEM faculty job applications: a successful pilot programme.}, journal = {Proceedings. Biological Sciences}, volume = {290}, year = {2023}, month = {2023 Apr 26}, pages = {20230124}, abstract = {

To attain a faculty position, postdoctoral fellows submit job applications that require considerable time and effort to produce. Although mentors and colleagues review these applications, postdocs rarely receive iterative feedback from reviewers with the breadth of expertise typically found on an academic search committee. To address this gap, we describe an international peer-reviewing programme for postdocs across disciplines to receive reciprocal, iterative feedback on faculty applications. A participant survey revealed that nearly all participants would recommend the programme to others. Furthermore, our programme was more likely to attract postdocs who struggled to find mentoring, possibly because of their identity as a woman or member of an underrepresented population in STEM or because they changed fields. Between 2018 and 2021, our programme provided nearly 150 early career academics with a diverse and supportive community of peer mentors during the difficult search for a faculty position and continues to do so today. As the transition from postdoc to faculty represents the largest {\textquoteright}leak{\textquoteright} in the academic pipeline, implementation of similar programmes by universities or professional societies would provide psycho-social support necessary to prevent attrition of individuals from underrepresented populations as well as increase the chances of success for early career academics in their search for independence.

}, keywords = {Faculty, Female, Humans, Mentoring, Mentors, Peer Group, Pilot Projects}, issn = {1471-2954}, doi = {10.1098/rspb.2023.0124}, author = {Guardia, Carlos M and Kane, Erin and Tebo, Alison G and Sanders, Anna A W M and Kaya, Devrim and Grogan, Kathleen E} } @article {68466, title = {A scalable implementation of the recursive least-squares algorithm for training spiking neural networks}, journal = {Frontiers in Neuroinformatics}, year = {2023}, month = {2023 Jun 27}, abstract = {

Training spiking recurrent neural networks on neuronal recordings or behavioral tasks has become a prominent tool to study computations in the brain. With an increasing size and complexity of neural recordings, there is a need for fast algorithms that can scale to large datasets. We present optimized CPU and GPU implementations of the recursive least-squares algorithm in spiking neural networks. The GPU implementation allows training networks to reproduce neural activity of an order of millions neurons at order of magnitude times faster than the CPU implementation. We demonstrate this by applying our algorithm to reproduce the activity of\ \>\ 66, 000 recorded neurons of a mouse performing a decision-making task. The fast implementation enables efficient training of large-scale spiking models, thus allowing for\ in-silico\ study of the dynamics and connectivity underlying multi-area computations.

}, doi = {10.3389/fninf.2023.1099510}, url = {https://doi.org/10.3389/fninf.2023.1099510}, author = {Benjamin J. Arthur and Christopher M. Kim and Susu Chen and Stephan Preibisch and Ran Darshan} } @article {68629, title = {A searchable image resource of Drosophila GAL4-driver expression patterns with single neuron resolution.}, journal = {eLife}, volume = {12}, year = {2023}, month = {2023 Feb 23}, abstract = {

Precise, repeatable genetic access to specific neurons via GAL4/UAS and related methods is a key advantage of Drosophila neuroscience. Neuronal targeting is typically documented using light microscopy of full GAL4 expression patterns, which generally lack the single-cell resolution required for reliable cell type identification. Here we use stochastic GAL4 labeling with the MultiColor FlpOut approach to generate cellular resolution confocal images at large scale. We are releasing aligned images of 74,000 such adult central nervous systems. An anticipated use of this resource is to bridge the gap between neurons identified by electron or light microscopy. Identifying individual neurons that make up each GAL4 expression pattern improves the prediction of split-GAL4 combinations targeting particular neurons. To this end we have made the images searchable on the NeuronBridge website. We demonstrate the potential of NeuronBridge to rapidly and effectively identify neuron matches based on morphology across imaging modalities and datasets.

}, issn = {2050-084X}, doi = {10.7554/eLife.80660}, author = {Meissner, Geoffrey Wilson and Nern, Aljoscha and Dorman, Zachary and Depasquale, Gina M and Forster, Kaitlyn and Gibney, Theresa and Hausenfluck, Joanna H and He, Yisheng and Iyer, Nirmala A and Jeter, Jennifer and Johnson, Lauren and Johnston, Rebecca M and Lee, Kelley and Melton, Brian and Yarbrough, Brianna and Zugates, Christopher T and Clements, Jody and Goina, Cristian and Otsuna, Hideo and Rokicki, Konrad and Svirskas, Robert R and Aso, Yoshinori and Card, Gwyneth M and Dickson, Barry J and Ehrhardt, Erica and Goldammer, Jens and Ito, Masayoshi and Kainmueller, Dagmar and Korff, Wyatt and Mais, Lisa and minegishi, ryo and Namiki, Shigehiro and Rubin, Gerald M and Sterne, Gabriella R and Wolff, Tanya and Malkesman, Oz} } @article {68646, title = {Selective Serotonin Reuptake Inhibitors within Cells: Temporal Resolution in Cytoplasm, Endoplasmic Reticulum, and Membrane.}, journal = {Journal of Neuroscience}, year = {2023}, month = {2023 Mar 02}, abstract = {

Selective serotonin reuptake inhibitors (SSRIs) are the most prescribed treatment for individuals experiencing major depressive disorder (MDD). The therapeutic mechanisms that take place before, during, or after SSRIs bind the serotonin transporter (SERT) are poorly understood, partially because no studies exist of the cellular and subcellular pharmacokinetic properties of SSRIs in living cells. We studied escitalopram and fluoxetine using new intensity-based drug-sensing fluorescent reporters ("iDrugSnFRs") targeted to the plasma membrane (PM), cytoplasm, or endoplasmic reticulum (ER) of cultured neurons and mammalian cell lines. We also employed chemical detection of drug within cells and phospholipid membranes. The drugs attain equilibrium in neuronal cytoplasm and ER, at approximately the same concentration as the externally applied solution, with time constants of a few s (escitalopram) or 200-300 s (fluoxetine). Simultaneously, the drugs accumulate within lipid membranes by >= 18-fold (escitalopram) or 180-fold (fluoxetine), and possibly by much larger factors. Both drugs leave cytoplasm, lumen, and membranes just as quickly during washout. We synthesized membrane-impermeant quaternary amine derivatives of the two SSRIs. The quaternary derivatives are substantially excluded from membrane, cytoplasm, and ER for \> 2.4 h. They inhibit SERT transport-associated currents 6- or 11-fold less potently than the SSRIs (escitalopram or fluoxetine derivative, respectively), providing useful probes for distinguishing compartmentalized SSRI effects. Although our measurements are orders of magnitude faster than the "therapeutic lag" of SSRIs, these data suggest that SSRI-SERT interactions within organelles or membranes may play roles during either the therapeutic effects or the "antidepressant discontinuation syndrome".Selective serotonin reuptake inhibitors stabilize mood in several disorders. In general, these drugs bind to the serotonin (5-hydroxytryptamine) transporter (SERT), which clears serotonin from CNS and peripheral tissues. SERT ligands are effective and relatively safe; primary care practitioners often prescribe them. However, they have several side effects and require 2 to 6 weeks of continuous administration until they act effectively. How they work remains perplexing, contrasting with earlier assumptions that the therapeutic mechanism involves SERT inhibition followed by increased extracellular serotonin levels. This study establishes that two SERT ligands, fluoxetine and escitalopram, enter neurons within minutes, while simultaneously accumulating in many membranes. Such knowledge will motivate future research, hopefully revealing where and how SERT ligands "engage" their therapeutic target(s).

}, issn = {1529-2401}, doi = {10.1523/JNEUROSCI.1519-22.2022}, author = {Nichols, Aaron L and Blumenfeld, Zack and Luebbert, Laura and Knox, Hailey J and Muthusamy, Anand K and Marvin, Jonathan S and Kim, Charlene H and Grant, Stephen N and Walton, David P and Cohen, Bruce N and Hammar, Rebekkah and Looger, Loren and Artursson, Per and Dougherty, Dennis A and Lester, Henry A} } @article {68690, title = {Sensitivity optimization of a rhodopsin-based fluorescent voltage indicator}, journal = {Neuron}, volume = {111}, year = {2023}, month = {2023 May 17}, pages = {1547-1563}, abstract = {

The ability to optically image cellular transmembrane voltages at millisecond-timescale resolutions can offer unprecedented insight into the function of living brains in behaving animals. Here, we present a\ point mutation\ that increases the sensitivity of Ace2 opsin-based voltage indicators. We use the mutation to develop Voltron2, an improved chemigeneic voltage indicator that has a 65\% higher sensitivity to single APs and 3-fold higher sensitivity to subthreshold potentials than Voltron. Voltron2 retained the sub-millisecond kinetics and photostability of its predecessor, although with lower baseline fluorescence. In multiple\ in\ vitro\ and\ in\ vivo\ comparisons with its predecessor across multiple species, we found Voltron2 to be more sensitive to APs and subthreshold fluctuations. Finally, we used Voltron2 to study and evaluate the possible mechanisms of\ interneuron\ synchronization in the mouse hippocampus. Overall, we have discovered a generalizable mutation that significantly increases the sensitivity of Ace2 rhodopsin-based sensors, improving their voltage reporting capability.

}, issn = {08966273}, doi = {10.1016/j.neuron.2023.03.009}, url = {https://linkinghub.elsevier.com/retrieve/pii/S0896627323002052}, author = {Abdelfattah, Ahmed S. and Zheng, Jihong and Singh, Amrita and Huang, Yi-Chieh and Reep, Daniel and Tsegaye, Getahun and Tsang, Arthur and Arthur, Benjamin J. and Rehorova, Monika and Olson, Carl V.L. and Shuai, Yichun and Zhang, Lixia and Fu, Tian-Ming and Milkie, Daniel E. and Moya, Maria V. and Weber, Timothy D. and Lemire, Andrew L. and Baker, Christopher A. and Falco, Natalie and Zheng, Qinsi and Grimm, Jonathan B. and Yip, Mighten C. and Walpita, Deepika and Chase, Martin and Campagnola, Luke and Murphy, Gabe J. and Wong, Allan M. and Forest, Craig R. and Mertz, Jerome and Economo, Michael N. and Turner, Glenn C. and Koyama, Minoru and Lin, Bei-Jung and Betzig, Eric and Novak, Ondrej and Lavis, Luke D. and Svoboda, Karel and Korff, Wyatt and Chen, Tsai-Wen and Schreiter, Eric R. and Hasseman, Jeremy P. and Kolb, Ilya} } @article {68695, title = {Sequence and Structural Motifs Controlling the Broad Substrate Specificity of the Mycobacterial Hormone-Sensitive Lipase LipN}, journal = {ACS Omega}, volume = {8}, year = {2023}, month = {2023 Mar 30}, pages = {13252 - 13264}, abstract = {

Mycobacterium tuberculosis\ has a complex life cycle transitioning between active and dormant growth states depending on environmental conditions. LipN (Rv2970c) is a conserved mycobacterial serine hydrolase with regulated catalytic activity at the interface between active and dormant growth conditions. LipN also catalyzes the xenobiotic degradation of a tertiary ester substrate and contains multiple conserved motifs connected with the ability to catalyze the hydrolysis of difficult tertiary ester substrates. Herein, we expanded a library of fluorogenic ester substrates to include more tertiary and constrained esters and screened 33 fluorogenic substrates for activation by LipN, identifying its unique substrate signature. LipN preferred short, unbranched ester substrates, but had its second highest activity against a heteroaromatic five-membered oxazole ester. Oxazole esters are present in multiple mycobacterial serine hydrolase inhibitors but have not been tested widely as ester substrates. Combined structural modeling, kinetic measurements, and substitutional analysis of LipN showcased a fairly rigid binding pocket preorganized for catalysis of short ester substrates. Substitution of diverse amino acids across the binding pocket significantly impacted the folded stability and catalytic activity of LipN with two conserved motifs (HGGGW and GDSAG) playing interconnected, multidimensional roles in regulating its substrate specificity. Together this detailed substrate specificity profile of LipN illustrates the complex interplay between structure and function in mycobacterial hormone-sensitive lipase homologues and indicates oxazole esters as promising inhibitor and substrate scaffolds for mycobacterial hydrolases.

}, issn = {2470-1343}, doi = {10.1021/acsomega.3c0053410.1021/acsomega.3c00534.s001}, url = {https://pubs.acs.org/doi/10.1021/acsomega.3c00534}, author = {Schemenauer, Daniel E. and Pool, Emily H. and Raynor, Stephanie N. and Ruiz, Gabriela P. and Goehring, Leah M. and Koelper, Andrew J. and Wilson, Madeleine A. and Durand, Anthony J. and Kourtoglou, Elexi C. and Larsen, Erik M. and Lavis, Luke D. and Esteb, John J. and Hoops, Geoffrey C. and Johnson, R. Jeremy} } @article {68838, title = {Small-field visual projection neurons detect translational optic flow and support walking control}, journal = {bioRxiv}, year = {2023}, month = {2023 Jun 22}, abstract = {

Animals rely on visual motion for navigating the world, and research in flies has clarified how neural circuits extract information from moving visual scenes. However, the major pathways connecting these patterns of optic flow to behavior remain poorly understood. Using a high-throughput quantitative assay of visually guided behaviors and genetic neuronal silencing, we discovered a region in\ Drosophila{\textquoteright}s protocerebrum critical for visual motion following. We used neuronal silencing, calcium imaging, and optogenetics to identify a single cell type, LPC1, that innervates this region, detects translational optic flow, and plays a key role in regulating forward walking. Moreover, the population of LPC1s can estimate the travelling direction, such as when gaze direction diverges from body heading. By linking specific cell types and their visual computations to specific behaviors, our findings establish a foundation for understanding how the nervous system uses vision to guide navigation.

}, doi = {10.1101/2023.06.21.546024}, url = {https://doi.org/10.1101/2023.06.21.546024}, author = {Mathew D. Isaacson and Jessica L. M. Eliason and Aljoscha Nern and Edward M. Rogers and Gus K. Lott and Tanya Tabachnik and William J. Rowell and Austin W. Edwards and Wyatt L. Korff and Gerald M. Rubin and Kristin Branson and Michael B. Reiser} } @article {68666, title = {Social association predicts immunological similarity in rewilded mice}, journal = {bioRxiv}, year = {2023}, month = {2023 Mar 16}, abstract = {

Environmental influences on immune phenotypes are well-documented, but our understanding of which elements of the environment affect immune systems, and how, remains vague. Behaviors, including socializing with others, are central to an individual{\textquoteright}s interaction with its environment. We tracked behavior of rewilded laboratory mice of three inbred strains in outdoor enclosures and examined contributions of behavior, including social associations, to immune phenotypes. We found that the more associated two individuals were, the more similar their immune phenotypes were. Social association was particularly predictive of similar memory T and B cell profiles and was more influential than sibling relationships or worm infection status. These results highlight the importance of social networks for immune phenotype and reveal important immunological correlates of social life.

}, doi = {10.1101/2023.03.15.532825}, url = {https://doi.org/10.1101/2023.03.15.532825}, author = {A. E. Downie and O. Oyesola and R. S. Barre and Q. Caudron and Y.-H. Chen and E. J. Dennis and R. Garnier and K. Kiwanuka and A. Menezes and D. J. Navarrete and O. Mondrag{\'o}n-Palomino and J. B. Saunders and C. K. Tokita and K. Zaldana and K. Cadwell and P. Loke and A. L. Graham} } @article {69125, title = {Spatiotemporal-social association predicts immunological similarity in rewilded mice.}, journal = {Science Advances}, volume = {9}, year = {2023}, month = {2023 Dec 22}, pages = {eadh8310}, abstract = {

Environmental influences on immune phenotypes are well-documented, but our understanding of which elements of the environment affect immune systems, and how, remains vague. Behaviors, including socializing with others, are central to an individual{\textquoteright}s interaction with its environment. We therefore tracked behavior of rewilded laboratory mice of three inbred strains in outdoor enclosures and examined contributions of behavior, including associations measured from spatiotemporal co-occurrences, to immune phenotypes. We found extensive variation in individual and social behavior among and within mouse strains upon rewilding. In addition, we found that the more associated two individuals were, the more similar their immune phenotypes were. Spatiotemporal association was particularly predictive of similar memory T and B cell profiles and was more influential than sibling relationships or shared infection status. These results highlight the importance of shared spatiotemporal activity patterns and/or social networks for immune phenotype and suggest potential immunological correlates of social life.

}, keywords = {Animals, Immune System, Mice, Phenotype, Social Behavior}, issn = {2375-2548}, doi = {10.1126/sciadv.adh8310}, author = {Downie, Alexander E and Oyesola, Oyebola and Barre, Ramya S and Caudron, Quentin and Chen, Ying-Han and Dennis, Emily J and Garnier, Romain and Kiwanuka, Kasalina and Menezes, Arthur and Navarrete, Daniel J and Mondrag{\'o}n-Palomino, Octavio and Saunders, Jesse B and Tokita, Christopher K and Zaldana, Kimberly and Cadwell, Ken and Loke, P{\textquoteright}ng and Graham, Andrea L} } @article {68633, title = {Specialized actin nanoscale layers control focal adhesion turnover}, journal = {bioRxiv}, year = {2023}, month = {2023 Feb 15}, abstract = {

Focal adhesions (FAs) connect inner workings of the cell to the extracellular matrix to control cell adhesion, migration, and mechanosensing1,2. Previous studies demonstrated that FAs contain three vertical layers, which connect extracellular matrix to the cytoskeleton3,4,5. However, cellular processes rely on precisely-regulated FA turnover, but the molecular machineries that control FA assembly and disassembly have remained elusive. By using super-resolution iPALM microscopy, we identified two unprecedented nanoscale layers within FAs, specified by actin filaments bound to tropomyosin isoforms Tpm1.6 and Tpm3.2. The Tpm1.6-actin filaments beneath the previously identified {\textquoteleft}actin-regulatory layer{\textquoteright} are critical for adhesion maturation and controlled cell motility, whereas the Tpm3.2-actin filament layer towards the bottom of FA facilitates adhesion disassembly. Mechanistically, Tpm3.2 stabilizes KANK-family proteins at adhesions, and hence targets microtubule plus-ends to FAs to catalyse their disassembly. Loss of Tpm3.2 leads to disorganized microtubule network, abnormally stable FAs, and defects in tail retraction during cell migration. Thus, FAs are composed of at least three distinct actin filament layers, each having specific roles in coupling of adhesion to the cytoskeleton, or in controlling adhesion dynamics. In a broader context, these findings demonstrate how distinct actin filament populations can co-exist and perform specific functions within a defined cellular compartment.

}, doi = {10.1101/2023.02.15.528622}, url = {https://doi.org/10.1101/2023.02.15.528622}, author = {Reena Kumari and Katharina Ven and Megan Chastney and Johan Per{\"a}nen and Jesse Aaron and Leonardo Almeida-Souza and Elena Kremneva and Renaud Poincloux and Teng-Leong Chew and Peter W. Gunning and Johanna Ivaska and Pekka Lappalainen} } @article {68566, title = {Synaptic gradients transform object location to action.}, journal = {Nature}, volume = {613}, year = {2023}, month = {2023 Jan 04}, pages = {534-42}, abstract = {

To survive, animals must convert sensory information into appropriate behaviours. Vision is a common sense for locating ethologically relevant stimuli and guiding motor responses. How circuitry converts object location in retinal coordinates to movement direction in body coordinates remains largely unknown. Here we show through behaviour, physiology, anatomy and connectomics in Drosophila that visuomotor transformation occurs by conversion of topographic maps formed by the dendrites of feature-detecting visual projection neurons (VPNs) into synaptic weight gradients of VPN outputs onto central brain neurons. We demonstrate how this gradient motif transforms the anteroposterior location of a visual looming stimulus into the fly{\textquoteright}s directional escape. Specifically, we discover that two neurons postsynaptic to a looming-responsive VPN type promote opposite takeoff directions. Opposite synaptic weight gradients onto these neurons from looming VPNs in different visual field regions convert localized looming threats into correctly oriented escapes. For a second looming-responsive VPN type, we demonstrate graded responses along the dorsoventral axis. We show that this synaptic gradient motif generalizes across all 20 primary VPN cell types and most often arises without VPN axon topography. Synaptic gradients may thus be a general mechanism for conveying spatial features of sensory information into directed motor outputs.

}, issn = {1476-4687}, doi = {10.1038/s41586-022-05562-8}, author = {Dombrovski, Mark and Peek, Martin Y and Park, Jin-Yong and Vaccari, Andrea and Sumathipala, Marissa and Morrow, Carmen and Breads, Patrick and Zhao, Arthur and Kurmangaliyev, Yerbol Z and Sanfilippo, Piero and Rehan, Aadil and Polsky, Jason and Alghailani, Shada and Tenshaw, Emily and Namiki, Shigehiro and Zipursky, S Lawrence and Card, Gwyneth M} } @article {68801, title = {Systematic annotation of a complete adult male Drosophila nerve cord connectome reveals principles of functional organisation}, journal = {bioRxiv}, year = {2023}, month = {2023 Jun 06}, abstract = {

Our companion paper (Takemura et al., 2023) introduces the first completely proofread connectome of the nerve cord of an animal that can walk or fly. The base connectome consists of neuronal morphologies and the connections between them. However, in order to efficiently navigate and understand this connectome, it is crucial to have a system of annotations that systematically categorises and names neurons, linking them to the existing literature. In this paper we describe the comprehensive annotation of the VNC connectome, first by a system of hierarchical coarse annotations, then by grouping left-right and serially homologous neurons and eventually by defining systematic cell types for the intrinsic interneurons and sensory neurons of the VNC; descending and motor neurons are typed in (Cheong et al., 2023). We assign a sensory modality to over 5000 sensory neurons, cluster them by connectivity, and identify serially homologous cell types and a layered organisation likely corresponding to peripheral topography. We identify the developmental neuroblast of origin of the large majority of VNC neurons and confirm that (in most cases) all secondary neurons of each hemilineage express a single neurotransmitter. Neuroblast hemilineages are serially repeated along the segments of the nerve cord and generally exhibit consistent hemilineage-to-hemilineage connectivity across neuromeres, supporting the idea that hemilineages are a major organisational feature of the VNC. We also find that more than a third of individual neurons belong to serially homologous cell types, which were crucial for identifying motor neurons and sensory neurons across leg neuropils. Categorising interneurons by their neuropil innervation patterns provides an additional organisation axis. Over half of the intrinsic neurons of the VNC appear dedicated to the legs, with the majority restricted to single leg neuropils; in contrast, inhibitory interneurons connecting different leg neuropils, especially those crossing the midline, appear rarer than anticipated by standard models of locomotor circuitry. Our annotations are being released as part of the neuprint.janelia.org web application and also serve as the basis of programmatic analysis of the connectome through dedicated tools that we describe in this paper.

}, doi = {10.1101/2023.06.05.543407}, url = {https://doi.org/10.1101/2023.06.05.543407}, author = {Elizabeth C Marin and Billy J Morris and Tomke Stuerner and Andrew S Champion and Dominik Krzeminski and Griffin Badalamente and Marina Gkantia and Imaan F M Tamimi and Siqi Fang and Sung Soo Moon and Han S J Cheong and Feng Li and Philipp Schlegel and Stuart Berg and FlyEM Project Team and Gwyneth M Card and Marta Costa and David Shepherd and Gregory S X E Jefferis} } @article {68606, title = {TEMPO enables sequential genetic labeling and manipulation of vertebrate cell lineages.}, journal = {Neuron}, volume = {111}, year = {2023}, month = {2023 Feb 01}, pages = {345-361.e10}, abstract = {

During development, regulatory factors appear in a precise order to determine cell fates over time. Consequently, to investigate complex tissue development, it is necessary to visualize and manipulate cell lineages with temporal control. Current strategies for tracing vertebrate cell lineages lack genetic access to sequentially produced cells. Here, we present TEMPO (Temporal Encoding and Manipulation in a Predefined Order), an imaging-readable genetic tool allowing differential labeling and manipulation of consecutive cell generations in vertebrates. TEMPO is based on CRISPR and powered by a cascade of gRNAs that drive orderly activation and inactivation of reporters and/or effectors. Using TEMPO to visualize zebrafish and mouse neurogenesis, we recapitulated birth-order-dependent neuronal fates. Temporally manipulating cell-cycle regulators in mouse cortex progenitors altered the proportion and distribution of neurons and glia, revealing the effects of temporal gene perturbation on serial cell fates. Thus, TEMPO enables sequential manipulation of molecular factors, crucial to study cell-type specification.

}, keywords = {Animals, Cell Differentiation, Cell Lineage, Gene Expression Regulation, Developmental, Mice, Neurogenesis, Neuroglia, Neurons, Zebrafish}, issn = {1097-4199}, doi = {10.1016/j.neuron.2022.10.035}, author = {Espinosa-Medina, Isabel and Feliciano, Daniel and Belmonte-Mateos, Carla and Linda Miyares, Rosa and Garcia-Marques, Jorge and Foster, Benjamin and Lindo, Sarah and Pujades, Cristina and Koyama, Minoru and Lee, Tzumin} } @article {68884, title = {Ten-fold Robust Expansion Microscopy.}, journal = {Bio-Protocol}, volume = {13}, year = {2023}, month = {2023 Jun 20}, pages = {e4698}, abstract = {

Expansion microscopy (ExM) is a powerful technique to overcome the diffraction limit of light microscopy that can be applied in both tissues and cells. In ExM, samples are embedded in a swellable polymer gel to physically expand the sample and isotropically increase resolution in x, y, and z. By systematic exploration of the ExM recipe space, we developed a novel ExM method termed Ten-fold Robust Expansion Microscopy (TREx) that, as the original ExM method, requires no specialized equipment or procedures. TREx enables ten-fold expansion of both thick mouse brain tissue sections and cultured human cells, can be handled easily, and enables high-resolution subcellular imaging with a single expansion step. Furthermore, TREx can provide ultrastructural context to subcellular protein localization by combining antibody-stained samples with off-the-shelf small molecule stains for both total protein and membranes.

}, issn = {2331-8325}, doi = {10.21769/BioProtoc.4698}, author = {Damstra, Hugo G J and Mohar, Boaz and Eddison, Mark and Akhmanova, Anna and Kapitein, Lukas C and Tillberg, Paul W} } @article {68999, title = {Theta oscillations as a substrate for medial prefrontal-hippocampal assembly interactions.}, journal = {Cell Reports}, volume = {42}, year = {2023}, month = {2023 Aug 24}, pages = {113015}, abstract = {

The execution of cognitive functions requires coordinated circuit activity across different brain areas that involves the associated firing of neuronal assemblies. Here, we tested the circuit mechanism behind assembly interactions between the hippocampus and the medial prefrontal cortex (mPFC) of adult rats by recording neuronal populations during a rule-switching task. We identified functionally coupled CA1-mPFC cells that synchronized their activity beyond that expected from common spatial coding or oscillatory firing. When such cell pairs fired together, the mPFC cell strongly phase locked to CA1 theta oscillations and maintained consistent theta firing phases, independent of the theta timing of their CA1 counterpart. These functionally connected CA1-mPFC cells formed interconnected assemblies. While firing together with their CA1 assembly partners, mPFC cells fired along specific theta sequences. Our results suggest that upregulated theta oscillatory firing of mPFC cells can signal transient interactions with specific CA1 assemblies, thus enabling distributed computations.

}, issn = {2211-1247}, doi = {10.1016/j.celrep.2023.113015}, author = {Nardin, Michele and Kaefer, Karola and Stella, Federico and Csicsvari, Jozsef} } @article {68667, title = {Three-dimensional reconstructions of mechanosensory end organs suggest a unifying mechanism underlying dynamic, light touch}, journal = {Neuron}, year = {2023}, month = {2023 Oct 18}, abstract = {

Specialized mechanosensory end organs within mammalian skin{\textemdash}hair follicle-associated lanceolate complexes, Meissner corpuscles, and Pacinian corpuscles{\textemdash}enable our perception of light, dynamic touch1. In each of these end organs, fast-conducting mechanically sensitive neurons, called Aβ low-threshold mechanoreceptors (Aβ LTMRs), associate with resident glial cells, known as terminal Schwann cells (TSCs) or lamellar cells, to form complex axon ending structures. Lanceolate-forming and corpuscle-innervating Aβ LTMRs share a low threshold for mechanical activation, a rapidly adapting (RA) response to force indentation, and high sensitivity to dynamic stimuli1{\textendash}6. How mechanical stimuli lead to activation of the requisite mechanotransduction channel Piezo27{\textendash}15\ and Aβ RA-LTMR excitation across the morphologically dissimilar mechanosensory end organ structures is not understood. Here, we report the precise subcellular distribution of Piezo2 and high-resolution, isotropic 3D reconstructions of all three end organs formed by Aβ RA-LTMRs determined by large volume enhanced Focused Ion Beam Scanning Electron Microscopy (FIB-SEM) imaging. We found that within each end organ, Piezo2 is enriched along the sensory axon membrane and is minimally or not expressed in TSCs and lamellar cells. We also observed a large number of small cytoplasmic protrusions enriched along the Aβ RA-LTMR axon terminals associated with hair follicles, Meissner corpuscles, and Pacinian corpuscles. These axon protrusions reside within close proximity to axonal Piezo2, occasionally contain the channel, and often form adherens junctions with nearby non-neuronal cells. Our findings support a unified model for Aβ RA-LTMR activation in which axon protrusions anchor Aβ RA-LTMR axon terminals to specialized end organ cells, enabling mechanical stimuli to stretch the axon in hundreds to thousands of sites across an individual end organ and leading to activation of proximal Piezo2 channels and excitation of the neuron.

}, doi = {10.1016/j.neuron.2023.08.023}, url = {https://doi.org/10.1016/j.neuron.2023.08.023}, author = {Annie Handler and Qiyu Zhang and Song Pang and Tri M. Nguyen and Michael Iskols and Michael Nolan-Tamariz and Stuart Cattel and Rebecca Plumb and Brianna Sanchez and Karyl Ashjian and Aria Shotland and Bartianna Brown and Madiha Kabeer and Josef Turecek and Genelle Rankin and Wangchu Xiang and Elisa C. Pavarino and Nusrat Africawala and Celine Santiago and Wei-Chung Allen Lee and C. Shan Xu and David D. Ginty} } @article {68958, title = {Towards Generalizable Organelle Segmentation in Volume Electron Microscopy.}, journal = {Microscopy and Microanalysis}, volume = {29}, year = {2023}, month = {2023 Jul 22}, pages = {975}, keywords = {Organelles, Volume Electron Microscopy}, issn = {1435-8115}, doi = {10.1093/micmic/ozad067.487}, author = {Heinrich, Larissa and Patton, Will and Bennett, Davis and Ackerman, David and Park, Grace and Bogovic, John A and Eckstein, Nils and Petruncio, Alyson and Clements, Jody and Pang, Song and Shan Xu, C and Funke, Jan and Korff, Wyatt and Hess, Harald and Lippincott-Schwartz, Jennifer and Saalfeld, Stephan and Weigel, Aubrey} } @article {68968, title = {Ultra-high density electrodes improve detection, yield, and cell type specificity of brain recordings.}, journal = {bioRxiv}, year = {2023}, month = {2023 Aug 25}, abstract = {

To study the neural basis of behavior, we require methods to sensitively and accurately measure neural activity at single neuron and single spike resolution. Extracellular electrophysiology is a principal method for achieving this, but it has biases in the neurons it detects and it imperfectly resolves their action potentials. To overcome these limitations, we developed a silicon probe with significantly smaller and denser recording sites than previous designs, called Neuropixels Ultra (NP Ultra). This device measures neuronal activity at ultra-high densities (\>1300 sites per mm, 10 times higher than previous probes), with 6 {\textmu}m center-to-center spacing and low noise. This device effectively comprises an implantable voltage-sensing camera that captures a planar image of a neuron{\textquoteright}s electrical field. We introduce a new spike sorting algorithm optimized for these probes and use it to find that the yield of visually-responsive neurons in recordings from mouse visual cortex improves \~{}3-fold. Recordings across multiple brain regions and four species revealed a subset of unexpectedly small extracellular action potentials not previously reported. Further experiments determined that, in visual cortex, these do not correspond to major subclasses of interneurons and instead likely reflect recordings from axons. Finally, using ground-truth identification of cortical inhibitory cell types with optotagging, we found that cell type was discriminable with approximately 75\% success among three types, a significant improvement over lower-resolution recordings. NP Ultra improves spike sorting performance, sampling bias, and cell type classification.

}, doi = {10.1101/2023.08.23.554527}, author = {Ye, Zhiwen and Shelton, Andrew M and Shaker, Jordan R and Boussard, Julien and Colonell, Jennifer and Manavi, Sahar and Chen, Susu and Windolf, Charlie and Hurwitz, Cole and Namima, Tomoyuki and Pedraja, Federico and Weiss, Shahaf and Raducanu, Bogdan and Ness, Torbj{\o}rn V and Einevoll, Gaute T and Laurent, Gilles and Sawtell, Nathaniel B and Bair, Wyeth and Pasupathy, Anitha and Mora Lopez, Carolina and Dutta, Barun and Paninski, Liam and Siegle, Joshua H and Koch, Christof and Olsen, Shawn R and Harris, Timothy D and Steinmetz, Nicholas A} } @article {69074, title = {Vagal sensory neurons mediate the Bezold-Jarisch reflex and induce syncope.}, journal = {Nature}, volume = {623}, year = {2023}, month = {2023 Nov 01}, pages = {387-396}, abstract = {

Visceral sensory pathways mediate homeostatic reflexes, the dysfunction of which leads to many neurological disorders. The Bezold-Jarisch reflex (BJR), first described in 1867, is a cardioinhibitory reflex that is speculated to be mediated by vagal sensory neurons (VSNs) that also triggers syncope. However, the molecular identity, anatomical organization, physiological characteristics and behavioural influence of cardiac VSNs remain mostly unknown. Here we leveraged single-cell RNA-sequencing data and HYBRiD tissue clearing to show that VSNs that express neuropeptide Y receptor Y2 (NPY2R) predominately connect the heart ventricular wall to the area postrema. Optogenetic activation of NPY2R VSNs elicits the classic triad of BJR responses-hypotension, bradycardia and suppressed respiration-and causes an animal to faint. Photostimulation during high-resolution echocardiography and laser Doppler flowmetry with behavioural observation revealed a range of phenotypes reflected in clinical syncope, including reduced cardiac output, cerebral hypoperfusion, pupil dilation and eye-roll. Large-scale Neuropixels brain recordings and machine-learning-based modelling showed that this manipulation causes the suppression of activity across a large distributed neuronal population that is not explained by changes in spontaneous behavioural movements. Additionally, bidirectional manipulation of the periventricular zone had a push-pull effect, with inhibition leading to longer syncope periods and activation inducing arousal. Finally, ablating NPY2R VSNs specifically abolished the BJR. Combined, these results demonstrate a genetically defined cardiac reflex that recapitulates characteristics of human syncope at physiological, behavioural and neural network levels.

}, keywords = {Area Postrema, Bradycardia, Cardiac Output, Low, Echocardiography, Heart, Heart Rate, Humans, Hypotension, Laser-Doppler Flowmetry, Nerve Net, Reflex, Sensory Receptor Cells, Single-Cell Gene Expression Analysis, Syncope, vagus nerve}, issn = {1476-4687}, doi = {10.1038/s41586-023-06680-7}, author = {Lovelace, Jonathan W and Ma, Jingrui and Yadav, Saurabh and Chhabria, Karishma and Shen, Hanbing and Pang, Zhengyuan and Qi, Tianbo and Sehgal, Ruchi and Zhang, Yunxiao and Bali, Tushar and Vaissiere, Thomas and Tan, Shawn and Liu, Yuejia and Rumbaugh, Gavin and Ye, Li and Kleinfeld, David and Stringer, Carsen and Augustine, Vineet} } @article {68792, title = {Voltage dynamics of dendritic integration and back-propagation in vivo}, journal = {bioRxiv}, year = {2023}, month = {2023 May 26}, abstract = {

Neurons integrate synaptic inputs within their dendrites and produce spiking outputs, which then propagate down the axon and back into the dendrites where they contribute to plasticity. Mapping the voltage dynamics in dendritic arbors of live animals is crucial for understanding neuronal computation and plasticity rules. Here we combine patterned channelrhodopsin activation with dual-plane structured illumination voltage imaging, for simultaneous perturbation and monitoring of dendritic and somatic voltage in Layer 2/3 pyramidal neurons in anesthetized and awake mice. We examined the integration of synaptic inputs and compared the dynamics of optogenetically evoked, spontaneous, and sensory-evoked back-propagating action potentials (bAPs). Our measurements revealed a broadly shared membrane voltage throughout the dendritic arbor, and few signatures of electrical compartmentalization among synaptic inputs. However, we observed spike rate acceleration-dependent propagation of bAPs into distal dendrites. We propose that this dendritic filtering of bAPs may play a critical role in activity-dependent plasticity.

}, doi = {10.1101/2023.05.25.542363}, url = {https://doi.org/10.1101/2023.05.25.542363}, author = {J. David Wong-Campos and Pojeong Park and Hunter Davis and Yitong Qi and He Tian and Daniel G. Itkis and Doyeon Kim and Sarah E. Plutkis and Luke Lavis and Adam E. Cohen} } @article {68104, title = {Volume-transmitted GABA waves pace epileptiform rhythms in the hippocampal network}, journal = {Current Biology}, volume = {33}, year = {2023}, month = {2023 Apr 10}, pages = {1249}, abstract = {

Mechanisms that entrain and drive rhythmic epileptiform discharges remain debated. Traditionally, this quest has been focusing on interneuronal networks driven by GABAergic connections that activate synaptic or extrasynaptic receptors. However, synchronised interneuronal discharges could also trigger a transient elevation of extracellular GABA across the tissue volume, thus raising tonic GABAA\ receptor conductance (Gtonic) in multiple cells. Here, we use patch-clamp GABA {\textquoteleft}sniffer{\textquoteright} and optical GABA sensor to show that periodic epileptiform discharges are preceded by region-wide, rising waves of extracellular GABA. Neural network simulations that incorporate volume-transmitted GABA signals point to mechanistic principles underpinning this relationship. We validate this hypothesis using simultaneous patch-clamp recordings from multiple nerve cells, selective optogenetic stimulation of fast-spiking interneurons. Critically, we manipulate GABA uptake to suppress extracellular GABA waves but not synaptic GABAergic transmission, which shows a clear effect on rhythm generation. Our findings thus unveil a key role of extrasynaptic, volume-transmitted GABA actions in pacing regenerative rhythmic activity in brain networks.

}, doi = {10.1016/j.cub.2023.02.051}, url = {https://doi.org/10.1016/j.cub.2023.02.051}, author = {Vincent Magloire and Leonid P. Savtchenko and Sergyi Sylantyev and Thomas P. Jensen and Nicholas Cole and Jonathan S. Marvin and Loren L. Looger and Dimitri M. Kullmann and Matthew C. Walker and Ivan Pavlov and Dmitri A. Rusakov} } @article {67946, title = {2,7-Diaminobenzopyrylium Dyes Are Live-Cell Mitochondrial Stains2,7-Diaminobenzopyrylium Dyes Are Live-Cell Mitochondrial Stains}, journal = {ACS Bio \& Med Chem Au}, volume = {2}, year = {2022}, month = {2022 Jun 15}, pages = {307-12}, abstract = {

Small-molecule fluorescent stains enable the imaging of cellular structures without the need for genetic manipulation. Here, we introduce 2,7-diaminobenzopyrylium (DAB) dyes as live-cell mitochondrial stains excited with violet light. This amalgam of the coumarin and rhodamine fluorophore structures yields dyes with high photostability and tunable spectral properties.

}, issn = {2694-2437}, doi = {10.1021/acsbiomedchemau.1c00068}, url = {https://pubs.acs.org/doi/10.1021/acsbiomedchemau.1c00068}, author = {Banala, Sambashiva and Tkachuk, Ariana N. and Patel, Ronak and Kumar, Pratik and Brown, Timothy A. and Lavis, Luke D.} } @article {68045, title = {Actin nano-architecture of phagocytic podosomes}, journal = {Nature Communications}, volume = {13}, year = {2022}, month = {2022 Jul 27}, pages = {4363}, abstract = {

Podosomes are actin-enriched adhesion structures important for multiple cellular processes, including migration, bone remodeling, and phagocytosis. Here, we characterized the structure and organization of phagocytic podosomes using interferometric photoactivated localization microscopy (iPALM), a super-resolution microscopy technique capable of 15-20 nm resolution, together with structured illumination microscopy (SIM) and localization-based superresolution microscopy. Phagocytic podosomes were observed during frustrated phagocytosis, a model in which cells attempt to engulf micro-patterned IgG antibodies. For circular patterns, this resulted in regular arrays of podosomes with well-defined geometry. Using persistent homology, we developed a pipeline for semi-automatic identification and measurement of podosome features. These studies revealed an "hourglass" shape of the podosome actin core, a protruding "knob" at the bottom of the core, and two actin networks extending from the core. Additionally, the distributions of paxillin, talin, myosin II, α-actinin, cortactin, and microtubules relative to actin were characterized.

}, doi = {10.1101/2022.05.04.490675}, author = {J. Cody Herron and Shiqiong Hu and Takashi Watanabe and Ana T. Nogueira and Bei Liu and Megan Kern and Jesse Aaron and Aaron Taylor and Michael Pablo and Teng-Leong Chew and Timothy C. Elston and Klaus M. Hahn} } @article {68023, title = {Allosteric interactions prime androgen receptor dimerization and activation.}, journal = {Molecular Cell}, volume = {82}, year = {2022}, month = {2022 Jun 02}, pages = {2021-31}, abstract = {

The androgen receptor (AR) is a nuclear receptor that governs gene expression programs required for prostate development and male phenotype maintenance. Advanced prostate cancers display AR hyperactivation and transcriptome expansion, in part, through AR amplification and interaction with oncoprotein cofactors. Despite its biological importance, how AR domains and cofactors cooperate to bind DNA has remained elusive. Using single-particle cryo-electron microscopy, we isolated three conformations of AR bound to DNA, showing that AR forms a non-obligate dimer, with the buried dimer interface utilized by ancestral steroid receptors repurposed to facilitate cooperative DNA binding. We identify novel allosteric surfaces which are compromised in androgen insensitivity syndrome and reinforced by AR{\textquoteright}s oncoprotein cofactor, ERG, and by DNA-binding motifs. Finally, we present evidence that this plastic dimer interface may have been adopted for transactivation at the expense of DNA binding. Our work highlights how fine-tuning AR{\textquoteright}s cooperative interactions translate to consequences in development and disease.

}, issn = {1097-4164}, doi = {10.1016/j.molcel.2022.03.035}, author = {Wasmuth, Elizabeth V and Broeck, Arnaud Vanden and LaClair, Justin R and Hoover, Elizabeth A and Lawrence, Kayla E and Paknejad, Navid and Pappas, Kyrie and Matthies, Doreen and Wang, Biran and Feng, Weiran and Watson, Philip A and Zinder, John C and Karthaus, Wouter R and de la Cruz, M Jason and Hite, Richard K and Manova-Todorova, Katia and Yu, Zhiheng and Weintraub, Susan T and Klinge, Sebastian and Sawyers, Charles L} } @article {68193, title = {Architecture and dynamics of a novel desmosome-endoplasmic reticulum organelle}, journal = {bioRxiv}, year = {2022}, month = {2022 Jul 08}, abstract = {

The endoplasmic reticulum (ER) forms a dynamic network that contacts other cellular membranes to regulate stress responses, calcium signaling, and lipid transfer. Using high-resolution volume electron microscopy, we find that the ER forms a previously unknown association with keratin intermediate filaments and desmosomal cell-cell junctions. Peripheral ER assembles into mirror image-like arrangements at desmosomes and exhibits nanometer proximity to keratin filaments and the desmosome cytoplasmic plaque. ER tubules exhibit stable associations with desmosomes, and perturbation of desmosomes or keratin filaments alters ER organization and mobility. These findings indicate that desmosomes and the keratin cytoskeleton pattern the distribution of the ER network. Overall, this study reveals a previously unknown subcellular architecture defined by the structural integration of ER tubules with an epithelial intercellular junction.

}, doi = {10.1101/2022.07.07.499185}, url = {https://doi.org/10.1101/2022.07.07.499185}, author = {Navaneetha Krishnan Bharathan and William Giang and Jesse S. Aaron and Satya Khuon and Teng-Leong Chew and Stephan Preibisch and Eric T. Trautman and Larissa Heinrich and John Bogovic and Davis Bennett and David Ackerman and Woohyun Park and Alyson Petruncio and Aubrey V. Weigel and Stephan Saalfeld and COSEM Project Team and A. Wayne Vogl and Sara N. Stahley and Andrew P. Kowalczyk} } @article {68313, title = {Automated reconstruction of whole-embryo cell lineages by learning from sparse annotations.}, journal = {Nature Biotechnology}, year = {2022}, month = {2022 Sep 05}, abstract = {

We present a method to automatically identify and track nuclei in time-lapse microscopy recordings of entire developing embryos. The method combines deep learning and global optimization. On a mouse dataset, it reconstructs 75.8\% of cell lineages spanning 1 h, as compared to 31.8\% for the competing method. Our approach improves understanding of where and when cell fate decisions are made in developing embryos, tissues, and organs.

}, issn = {1546-1696}, doi = {10.1038/s41587-022-01427-7}, author = {Malin-Mayor, Caroline and Hirsch, Peter and Guignard, Leo and McDole, Katie and Wan, Yinan and Lemon, William C and Kainmueller, Dagmar and Keller, Philipp J and Preibisch, Stephan and Funke, Jan} } @article {69715, title = {Best practice standards for circular RNA research}, journal = {Nature Methods}, volume = {19}, year = {2022}, month = {05/2022}, pages = {1208 - 1220}, abstract = {

Circular RNAs (circRNAs) are formed in all domains of life and via different mechanisms. There has been an explosion in the number of circRNA papers in recent years; however, as a relatively young field, circRNA biology has an urgent need for common experimental standards for isolating, analyzing, expressing and depleting circRNAs. Here we propose a set of guidelines for circRNA studies based on the authors{\textquoteright} experience. This Perspective will specifically address the major class of circRNAs in Eukarya that are generated by a spliceosome-catalyzed back-splicing event. We hope that the implementation of best practice principles for circRNA research will help move the field forward and allow a better functional understanding of this fascinating group of RNAs.

}, issn = {1548-7091}, doi = {10.1038/s41592-022-01487-2}, url = {https://www.nature.com/articles/s41592-022-01487-2https://www.nature.com/articles/s41592-022-01487-2.pdfhttps://www.nature.com/articles/s41592-022-01487-2.pdfhttps://www.nature.com/articles/s41592-022-01487-2}, author = {Nielsen, Anne F. and Bindereif, Albrecht and Bozzoni, Irene and Hanan, Mor and Hansen, Thomas B. and Irimia, Manuel and Kadener, Sebastian and Kristensen, Lasse S. and Legnini, Ivano and Morlando, Mariangela and Jarlstad Olesen, Morten T. and Pasterkamp, R. Jeroen and Preibisch, Stephan and Rajewsky, Nikolaus and Suenkel, Christin and Kjems, J{\o}rgen} } @article {68274, title = {Biomechanical origins of proprioceptive maps in the Drosophila leg}, journal = {bioRxiv}, year = {2022}, month = {2022 Aug 09}, abstract = {

Proprioception, the sense of body position and movement, is essential for effective motor control. Because proprioceptive sensory neurons are embedded in complex and dynamic tissues, it has been challenging to understand how they sense and encode mechanical stimuli. Here, we find that proprioceptor neurons in the\ Drosophila\ femur are organized into functional groups that are biomechanically specialized to detect features of tibia joint kinematics. The dendrites of position and vibration-tuned proprioceptors receive distinct mechanical signals via the\ arculum, an elegant mechanical structure that decomposes movement of the tibia joint into orthogonal components. The cell bodies of position-tuned proprioceptors form a\ goniotopic\ map of joint angle, whereas the dendrites of vibration-tuned proprioceptors form a\ tonotopic\ map of tibia vibration frequency. Our findings reveal biomechanical mechanisms that underlie proprioceptor feature selectivity and identify common organizational principles between proprioception and other topographically organized sensory systems.

}, doi = {10.1101/2022.08.08.503192}, url = {https://doi.org/10.1101/2022.08.08.503192}, author = {Akira Mamiya and Pralaksha Gurung and Igor Siwanowicz and Anne Sustar and Chenghao Chen and Jasper S. Phelps and Aaron T. Kuan and Alexandra Pacureanu and Wei-Chung Allen Lee and Natasha Mhatre and John C. Tuthill} } @article {67992, title = {Bromodomains regulate dynamic targeting of the PBAF chromatin remodeling complex to chromatin hubs.}, journal = {Biophysical Journal}, volume = {121}, year = {2022}, month = {2022 May 3}, pages = {1738-1752}, abstract = {

Chromatin remodelers actively target arrays of acetylated nucleosomes at select enhancers and promoters to facilitate or shut down the repeated recruitment of RNA Pol II during transcriptional bursting. It is poorly understood how chromatin remodelers such as PBAF dynamically target different chromatin states inside a live cell. Our live-cell single molecule fluorescence microscopy study reveals chromatin hubs throughout the nucleus where PBAF rapidly cycles on and off the genome. Deletion of PBAF{\textquoteright}s bromodomains impairs targeting and stable engagement of chromatin in hubs. Dual color imaging reveals that PBAF targets both euchromatic and heterochromatic hubs with distinct genome binding kinetic profiles that mimic chromatin stability. Removal of PBAF{\textquoteright}s bromodomains stabilizes H3.3 binding within chromatin indicating that bromodomains may play a direct role in remodeling of the nucleosome. Our data suggests that PBAF{\textquoteright}s dynamic bromodomain mediated engagement of a nucleosome may reflect the chromatin remodeling potential of differentially bound chromatin states.

}, issn = {1542-0086}, doi = {10.1016/j.bpj.2022.03.027}, author = {Kenworthy, Charles A and Haque, Nayem and Liou, Shu-Hao and Chandris, Panagiotis and Wong, Vincent and Dziuba, Patrycja and Lavis, Luke D and Liu, Wei-Li and Singer, Robert H and Coleman, Robert A} } @article {68520, title = {The connectome of an insect brain}, journal = {bioRxiv}, year = {2022}, month = {2022 Nov 28}, abstract = {

Brains contain networks of interconnected neurons, so knowing the network architecture is essential for understanding brain function. We therefore mapped the synaptic-resolution connectome of an insect brain (Drosophila\ larva) with rich behavior, including learning, value-computation, and action-selection, comprising 3,013 neurons and 544,000 synapses. We characterized neuron-types, hubs, feedforward and feedback pathways, and cross-hemisphere and brain-nerve cord interactions. We found pervasive multisensory and interhemispheric integration, highly recurrent architecture, abundant feedback from descending neurons, and multiple novel circuit motifs. The brain{\textquoteright}s most recurrent circuits comprised the input and output neurons of the learning center. Some structural features, including multilayer shortcuts and nested recurrent loops, resembled powerful machine learning architectures. The identified brain architecture provides a basis for future experimental and theoretical studies of neural circuits.

}, doi = {10.1101/2022.11.28.516756}, url = {https://doi.org/10.1101/2022.11.28.516756}, author = {Michael Winding and Benjamin D. Pedigo and Christopher L. Barnes and Heather G. Patsolic and Youngser Park and Tom Kazimiers and Akira Fushiki and Ingrid V. Andrade and Avinash Khandelwal and Javier Valdes-Aleman and Feng Li and Nadine Randel and Elizabeth Barsotti and Ana Correia and Richard D. Fetter and Volker Hartenstein and Carey E. Priebe and Joshua T. Vogelstein and Albert Cardona and Marta Zlatic} } @article {67966, title = {The Consistency of Gastropod Identified Neurons Distinguishes Intra-Individual Plasticity From Inter-Individual Variability in Neural Circuits.}, journal = {Frontiers in Behavioral Neuroscience}, volume = {16}, year = {2022}, month = {2022 Mar 03}, pages = {855235}, abstract = {

Gastropod mollusks are known for their large, individually identifiable neurons, which are amenable to long-term intracellular recordings that can be repeated from animal to animal. The constancy of individual neurons can help distinguish state-dependent or temporal variation within an individual from actual variability between individual animals. Investigations into the circuitry underlying rhythmic swimming movements of the gastropod species, and have uncovered intra- and inter-individual variability in synaptic connectivity and serotonergic neuromodulation. has a reliably evoked escape swim behavior that is produced by a central pattern generator (CPG) composed of a small number of identifiable neurons. There is apparent individual variability in some of the connections between neurons that is inconsequential for the production of the swim behavior under normal conditions, but determines whether that individual can swim following a neural lesion. Serotonergic neuromodulation of synaptic strength intrinsic to the CPG creates neural circuit plasticity within an individual and contributes to reorganization of the network during recovery from injury and during learning. In , variability over time in the modulatory actions of serotonin and in expression of serotonin receptor genes in an identified neuron directly reflects variation in swimming behavior. Tracking behavior and electrophysiology over hours to days was necessary to identify the functional consequences of these intra-individual, time-dependent variations. This work demonstrates the importance of unambiguous neuron identification, properly assessing the animal and network states, and tracking behavior and physiology over time to distinguish plasticity within the same animal at different times from variability across individual animals.

}, issn = {1662-5153}, doi = {10.3389/fnbeh.2022.855235}, author = {Tamvacakis, Arianna N and Lillvis, Joshua L and Sakurai, Akira and Katz, Paul S} } @article {68593, title = {Desmosomal connectomics of all somatic muscles in an annelid larva.}, journal = {eLife}, volume = {11}, year = {2022}, month = {2022 Dec 20}, abstract = {

Cells form networks in animal tissues through synaptic, chemical, and adhesive links. Invertebrate muscle cells often connect to other cells through desmosomes, adhesive junctions anchored by intermediate filaments. To study desmosomal networks, we skeletonised 853 muscle cells and their desmosomal partners in volume electron microscopy data covering an entire larva of the annelid . Muscle cells adhere to each other, to epithelial, glial, ciliated, and bristle-producing cells and to the basal lamina, forming a desmosomal connectome of over 2000 cells. The aciculae - chitin rods that form an endoskeleton in the segmental appendages - are highly connected hubs in this network. This agrees with the many degrees of freedom of their movement, as revealed by video microscopy. Mapping motoneuron synapses to the desmosomal connectome allowed us to infer the extent of tissue influenced by motoneurons. Our work shows how cellular-level maps of synaptic and adherent force networks can elucidate body mechanics.

}, keywords = {Animals, Annelida, Connectome, Larva, Muscles, Polychaeta}, issn = {2050-084X}, doi = {10.7554/eLife.71231}, author = {Jasek, Sanja and Veraszt{\'o}, Csaba and Brodrick, Emelie and Shahidi, R{\'e}za and Kazimiers, Tom and Kerbl, Alexandra and J{\'e}kely, G{\'a}sp{\'a}r} } @article {68296, title = {Disorder and the Neural Representation of Complex Odors.}, journal = {Frontiers in Computational Neuroscience}, volume = {16}, year = {2022}, month = {2022 Aug 08}, pages = {917786}, abstract = {

Animals smelling in the real world use a small number of receptors to sense a vast number of natural molecular mixtures, and proceed to learn arbitrary associations between odors and valences. Here, we propose how the architecture of olfactory circuits leverages disorder, diffuse sensing and redundancy in representation to meet these immense complementary challenges. First, the diffuse and disordered binding of receptors to many molecules compresses a vast but sparsely-structured odor space into a small receptor space, yielding an odor code that preserves similarity in a precise sense. Introducing any order/structure in the sensing degrades similarity preservation. Next, lateral interactions further reduce the correlation present in the low-dimensional receptor code. Finally, expansive disordered projections from the periphery to the central brain reconfigure the densely packed information into a high-dimensional representation, which contains multiple redundant subsets from which downstream neurons can learn flexible associations and valences. Moreover, introducing any order in the expansive projections degrades the ability to recall the learned associations in the presence of noise. We test our theory empirically using data from . Our theory suggests that the neural processing of sparse but high-dimensional olfactory information differs from the other senses in its fundamental use of disorder.

}, issn = {1662-5188}, doi = {10.3389/fncom.2022.917786}, author = {Krishnamurthy, Kamesh and Hermundstad, Ann M and Mora, Thierry and Walczak, Aleksandra M and Balasubramanian, Vijay} } @article {68166, title = {Distributing task-related neural activity across a cortical network through task-independent connections}, journal = {bioRxiv}, year = {2022}, month = {2022 Jun 18}, abstract = {

Task-related neural activity is widespread across populations of neurons during goal-directed behaviors. However, little is known about the synaptic reorganization and circuit mechanisms that lead to broad activity changes. Here we trained a limited subset of neurons in a spiking network with strong synaptic interactions to reproduce the activity of neurons in the motor cortex during a decision-making task. We found that task-related activity, resembling the neural data, emerged across the network, even in the untrained neurons. Analysis of trained networks showed that strong untrained synapses, which were independent of the task and determined the dynamical state of the network, mediated the spread of task-related activity. Optogenetic perturbations suggest that the motor cortex is strongly-coupled, supporting the applicability of the mechanism to cortical networks. Our results reveal a cortical mechanism that facilitates distributed representations of task-variables by spreading the activity from a subset of plastic neurons to the entire network through task-independent strong synapses.

}, doi = {10.1101/2022.06.17.496618}, url = {https://doi.org/10.1101/2022.06.17.496618}, author = {Christopher M. Kim and Arseny Finkelstein and Carson C. Chow and Karel Svoboda and Ran Darshan} } @article {68570, title = {Eye structure shapes neuron function in Drosophila motion vision}, journal = {bioRxiv}, year = {2022}, month = {2022 Dec 15}, abstract = {

Many animals rely on vision to navigate through their environment. The pattern of changes in the visual scene induced by self-motion is the\ optic flow1, which is first estimated in local patches by directionally selective (DS) neurons2{\textendash}4. But how should the arrays of DS neurons, each responsive to motion in a preferred direction at a specific retinal position, be organized to support robust decoding of optic flow by downstream circuits? Understanding this global organization is challenging because it requires mapping fine, local features of neurons across the animal{\textquoteright}s field of view3. In\ Drosophila, the asymmetric dendrites of the T4 and T5 DS neurons establish their preferred direction, making it possible to predict DS responses from anatomy4,5. Here we report that the preferred directions of fly DS neurons vary at different retinal positions and show that this spatial variation is established by the anatomy of the compound eye. To estimate the preferred directions across the visual field, we reconstructed hundreds of T4 neurons in a full brain EM volume6\ and discovered unexpectedly stereotypical dendritic arborizations that are independent of location. We then used whole-head μCT scans to map the viewing directions of all compound eye facets and found a non-uniform sampling of visual space that explains the spatial variation in preferred directions. Our findings show that the organization of preferred directions in the fly is largely determined by the compound eye, exposing an intimate and unexpected connection between the peripheral structure of the eye, functional properties of neurons deep in the brain, and the control of body movements.

}, doi = {10.1101/2022.12.14.520178}, url = {https://doi.org/10.1101/2022.12.14.520178}, author = {Arthur Zhao and Eyal Gruntman and Aljoscha Nern and Nirmala A. Iyer and Edward M. Rogers and Sanna Koskela and Igor Siwanowicz and Marisa Dreher and Miriam A. Flynn and Connor W. Laughland and Henrique D.F. Ludwig and Alex G. Thomson and Cullen P. Moran and Bruck Gezahegn and Davi D. Bock and Michael B. Reiser} } @article {67848, title = {Fluorescence activation mechanism and imaging of drug permeation with new sensors for smoking-cessation ligands.}, journal = {eLife}, volume = {11}, year = {2022}, month = {2022 Jan 04}, abstract = {

Nicotinic partial agonists provide an accepted aid for smoking cessation and thus contribute to decreasing tobacco-related disease. Improved drugs constitute a continued area of study. However, there remains no reductionist method to examine the cellular and subcellular pharmacokinetic properties of these compounds in living cells. Here, we developed new intensity-based drug sensing fluorescent reporters ({\textquoteright}iDrugSnFRs{\textquoteright}) for the nicotinic partial agonists dianicline, cytisine, and two cytisine derivatives - 10-fluorocytisine and 9-bromo-10-ethylcytisine. We report the first atomic-scale structures of liganded periplasmic binding protein-based biosensors, accelerating development of iDrugSnFRs and also explaining the activation mechanism. The nicotinic iDrugSnFRs detect their drug partners in solution, as well as at the plasma membrane (PM) and in the endoplasmic reticulum (ER) of cell lines and mouse hippocampal neurons. At the PM, the speed of solution changes limits the growth and decay rates of the fluorescence response in almost all cases. In contrast, we found that rates of membrane crossing differ among these nicotinic drugs by \> 30 fold. The new nicotinic iDrugSnFRs provide insight into the real-time pharmacokinetic properties of nicotinic agonists and provide a methodology whereby iDrugSnFRs can inform both pharmaceutical neuroscience and addiction neuroscience.

}, issn = {2050-084X}, doi = {10.7554/eLife.74648}, author = {Nichols, Aaron L and Blumenfeld, Zack and Fan, ChengCheng and Luebbert, Laura and Blom, Annet Em and Cohen, Bruce N and Marvin, Jonathan S and Borden, Philip M and Kim, Charlene H and Muthusamy, Anand K and Shivange, Amol V and Knox, Hailey J and Campello, Hugo Rego and Wang, Jonathan H and Dougherty, Dennis A and Looger, Loren L and Gallagher, Timothy and Rees, Douglas C and Lester, Henry A} } @article {68422, title = {Fly Cell Atlas: A single-nucleus transcriptomic atlas of the adult fruit fly.}, journal = {Science}, volume = {375}, year = {2022}, month = {03/2022}, pages = {eabk2432}, abstract = {

For more than 100 years, the fruit fly has been one of the most studied model organisms. Here, we present a single-cell atlas of the adult fly, Tabula , that includes 580,000 nuclei from 15 individually dissected sexed tissues as well as the entire head and body, annotated to \>250 distinct cell types. We provide an in-depth analysis of cell type-related gene signatures and transcription factor markers, as well as sexual dimorphism, across the whole animal. Analysis of common cell types between tissues, such as blood and muscle cells, reveals rare cell types and tissue-specific subtypes. This atlas provides a valuable resource for the community and serves as a reference to study genetic perturbations and disease models at single-cell resolution.

}, keywords = {Animals, Cell Nucleus, Databases, Genetic, Drosophila melanogaster, Drosophila Proteins, Female, Gene Expression Regulation, Gene Regulatory Networks, Genes, Insect, Male, RNA-Seq, Sex Characteristics, Single-Cell Analysis, Transcription Factors, Transcriptome}, issn = {1095-9203}, doi = {10.1126/science.abk2432}, author = {Li, Hongjie and Janssens, Jasper and De Waegeneer, Maxime and Kolluru, Sai Saroja and Davie, Kristofer and Gardeux, Vincent and Saelens, Wouter and David, Fabrice P A and Brbi{\'c}, Maria and Spanier, Katina and Leskovec, Jure and McLaughlin, Colleen N and Xie, Qijing and Jones, Robert C and Brueckner, Katja and Shim, Jiwon and Tattikota, Sudhir Gopal and Schnorrer, Frank and Rust, Katja and Nystul, Todd G and Carvalho-Santos, Zita and Ribeiro, Carlos and Pal, Soumitra and Mahadevaraju, Sharvani and Przytycka, Teresa M and Allen, Aaron M and Goodwin, Stephen F and Berry, Cameron W and Fuller, Margaret T and White-Cooper, Helen and Matunis, Erika L and DiNardo, Stephen and Galenza, Anthony and O{\textquoteright}Brien, Lucy Erin and Dow, Julian A T and Jasper, Heinrich and Oliver, Brian and Perrimon, Norbert and Deplancke, Bart and Quake, Stephen R and Luo, Liqun and Aerts, Stein and Agarwal, Devika and Ahmed-Braimah, Yasir and Arbeitman, Michelle and Ariss, Majd M and Augsburger, Jordan and Ayush, Kumar and Baker, Catherine C and Banisch, Torsten and Birker, Katja and Bodmer, Rolf and Bolival, Benjamin and Brantley, Susanna E and Brill, Julie A and Brown, Nora C and Buehner, Norene A and Cai, Xiaoyu Tracy and Cardoso-Figueiredo, Rita and Casares, Fernando and Chang, Amy and Clandinin, Thomas R and Crasta, Sheela and Desplan, Claude and Detweiler, Angela M and Dhakan, Darshan B and Don{\`a}, Erika and Engert, Stefanie and Floc{\textquoteright}hlay, Swann and George, Nancy and Gonz{\'a}lez-Segarra, Amanda J and Groves, Andrew K and Gumbin, Samantha and Guo, Yanmeng and Harris, Devon E and Heifetz, Yael and Holtz, Stephen L and Horns, Felix and Hudry, Bruno and Hung, Ruei-Jiun and Jan, Yuh Nung and Jaszczak, Jacob S and Jefferis, Gregory S X E and Karkanias, Jim and Karr, Timothy L and Katheder, Nadja Sandra and Kezos, James and Kim, Anna A and Kim, Seung K and Kockel, Lutz and Konstantinides, Nikolaos and Kornberg, Thomas B and Krause, Henry M and Labott, Andrew Thomas and Laturney, Meghan and Lehmann, Ruth and Leinwand, Sarah and Li, Jiefu and Li, Joshua Shing Shun and Li, Kai and Li, Ke and Li, Liying and Li, Tun and Litovchenko, Maria and Liu, Han-Hsuan and Liu, Yifang and Lu, Tzu-Chiao and Manning, Jonathan and Mase, Anjeli and Matera-Vatnick, Mikaela and Matias, Neuza Reis and McDonough-Goldstein, Caitlin E and McGeever, Aaron and McLachlan, Alex D and Moreno-Roman, Paola and Neff, Norma and Neville, Megan and Ngo, Sang and Nielsen, Tanja and O{\textquoteright}Brien, Caitlin E and Osumi-Sutherland, David and Ozel, Mehmet Neset and Papatheodorou, Irene and Petkovic, Maja and Pilgrim, Clare and Pisco, Angela Oliveira and Reisenman, Carolina and Sanders, Erin Nicole and Dos Santos, Gilberto and Scott, Kristin and Sherlekar, Aparna and Shiu, Philip and Sims, David and Sit, Rene V and Slaidina, Maija and Smith, Harold E and Sterne, Gabriella and Su, Yu-Han and Sutton, Daniel and Tamayo, Marco and Tan, Michelle and Tastekin, Ibrahim and Treiber, Christoph and Vacek, David and Vogler, Georg and Waddell, Scott and Wang, Wanpeng and Wilson, Rachel I and Wolfner, Mariana F and Wong, Yiu-Cheung E and Xie, Anthony and Xu, Jun and Yamamoto, Shinya and Yan, Jia and Yao, Zepeng and Yoda, Kazuki and Zhu, Ruijun and Zinzen, Robert P} } @article {67952, title = {A functionally ordered visual feature map in the Drosophila brain.}, journal = {Neuron}, volume = {110}, year = {2022}, month = {2022 May 18}, pages = {1700}, abstract = {

Topographic maps, the systematic spatial ordering of neurons by response tuning, are common across species. In Drosophila, the lobula columnar (LC) neuron types project from the optic lobe to the central brain, where each forms a glomerulus in a distinct position. However, the advantages of this glomerular arrangement are unclear. Here, we examine the functional and spatial relationships of 10 glomeruli using single-neuron calcium imaging. We discover novel detectors for objects smaller than the lens resolution (LC18) and for complex line motion (LC25). We find that glomeruli are spatially clustered by selectivity for looming versus drifting object motion and ordered by size tuning to form a topographic visual feature map. Furthermore, connectome analysis shows that downstream neurons integrate from sparse subsets of possible glomeruli combinations, which are biased for glomeruli encoding similar features. LC neurons are thus an explicit example of distinct feature detectors topographically organized to facilitate downstream circuit integration.

}, issn = {1097-4199}, doi = {10.1016/j.neuron.2022.02.013}, author = {Klapoetke, Nathan C and Nern, Aljoscha and Rogers, Edward M and Rubin, Gerald M and Reiser, Michael B and Card, Gwyneth M} } @article {68414, title = {High-throughput automated methods for classical and operant conditioning of larvae.}, journal = {eLife}, volume = {11}, year = {2022}, month = {2022 Oct 28}, abstract = {

Learning which stimuli (classical conditioning) or which actions (operant conditioning) predict rewards or punishments can improve chances of survival. However, the circuit mechanisms that underlie distinct types of associative learning are still not fully understood. Automated, high-throughput paradigms for studying different types of associative learning, combined with manipulation of specific neurons in freely behaving animals, can help advance this field. The Drosophila melanogaster larva is a tractable model system for studying the circuit basis of behaviour, but many forms of associative learning have not yet been demonstrated in this animal. Here, we developed a high-throughput (i. e. multi-larva) training system that combines real-time behaviour detection of freely moving larvae with targeted opto- and thermogenetic stimulation of tracked animals. Both stimuli are controlled in either open- or closed-loop, and delivered with high temporal and spatial precision. Using this tracker, we show for the first time that Drosophila larvae can perform classical conditioning with no overlap between sensory stimuli (i. e. trace conditioning). We also demonstrate that larvae are capable of operant conditioning by inducing a bend direction preference through optogenetic activation of reward-encoding serotonergic neurons. Our results extend the known associative learning capacities of Drosophila larvae. Our automated training rig will facilitate the study of many different forms of associative learning and the identification of the neural circuits that underpin them.

}, issn = {2050-084X}, doi = {10.7554/eLife.70015}, author = {Croteau-Chonka, Elise C and Clayton, Michael S and Venkatasubramanian, Lalanti and Harris, Samuel N and Jones, Benjamin M W and Narayan, Lakshmi and Winding, Michael and Masson, Jean-Baptiste and Zlatic, Marta and Klein, Kristina T} } @article {67931, title = {Identifying Inhibitors of -1 Programmed Ribosomal Frameshifting in a Broad Spectrum of Coronaviruses.}, journal = {Viruses}, volume = {14}, year = {2022}, month = {2022 Jan 18}, abstract = {

Recurrent outbreaks of novel zoonotic coronavirus (CoV) diseases in recent years have highlighted the importance of developing therapeutics with broad-spectrum activity against CoVs. Because all CoVs use -1 programmed ribosomal frameshifting (-1 PRF) to control expression of key viral proteins, the frameshift signal in viral mRNA that stimulates -1 PRF provides a promising potential target for such therapeutics. To test the viability of this strategy, we explored whether small-molecule inhibitors of -1 PRF in SARS-CoV-2 also inhibited -1 PRF in a range of bat CoVs-the most likely source of future zoonoses. Six inhibitors identified in new and previous screens against SARS-CoV-2 were evaluated against the frameshift signals from a panel of representative bat CoVs as well as MERS-CoV. Some drugs had strong activity against subsets of these CoV-derived frameshift signals, while having limited to no effect on -1 PRF caused by frameshift signals from other viruses used as negative controls. Notably, the serine protease inhibitor nafamostat suppressed -1 PRF significantly for multiple CoV-derived frameshift signals. These results suggest it is possible to find small-molecule ligands that inhibit -1 PRF specifically in a broad spectrum of CoVs, establishing frameshift signals as a viable target for developing pan-coronaviral therapeutics.

}, issn = {1999-4915}, doi = {10.3390/v14020177}, author = {Munshi, Sneha and Neupane, Krishna and Ileperuma, Sandaru M and Halma, Matthew T J and Kelly, Jamie A and Halpern, Clarissa F and Dinman, Jonathan D and Loerch, Sarah and Woodside, Michael T} } @article {68515, title = {Ketamine triggers a switch in excitatory neuronal activity across neocortex.}, journal = {Nature Neuroscience}, year = {2022}, month = {2022 Nov 24}, abstract = {

The brain can become transiently disconnected from the environment while maintaining vivid, internally generated experiences. This so-called {\textquoteright}dissociated state{\textquoteright} can occur in pathological conditions and under the influence of psychedelics or the anesthetic ketamine (KET). The cellular and circuit mechanisms producing the dissociative state remain poorly understood. We show in mice that KET causes spontaneously active neurons to become suppressed while previously silent neurons become spontaneously activated. This switch occurs in all cortical layers and different cortical regions, is induced by both systemic and cortical application of KET and is mediated by suppression of parvalbumin and somatostatin interneuron activity and inhibition of NMDA receptors and HCN channels. Combined, our results reveal two largely non-overlapping cortical neuronal populations-one engaged in wakefulness, the other contributing to the KET-induced brain state-and may lay the foundation for understanding how the brain might become disconnected from the surrounding environment while maintaining internal subjective experiences.

}, issn = {1546-1726}, doi = {10.1038/s41593-022-01203-5}, author = {Cichon, Joseph and Wasilczuk, Andrzej Z and Looger, Loren L and Contreras, Diego and Kelz, Max B and Proekt, Alex} } @article {68420, title = {KIRCD8 T cells suppress pathogenic T cells and are active in autoimmune diseases and COVID-19.}, journal = {Science}, volume = {376}, year = {2022}, month = {04/2022}, pages = {eabi9591}, abstract = {

In this work, we find that CD8 T cells expressing inhibitory killer cell immunoglobulin-like receptors (KIRs) are the human equivalent of Ly49CD8 regulatory T cells in mice and are increased in the blood and inflamed tissues of patients with a variety of autoimmune diseases. Moreover, these CD8 T cells efficiently eliminated pathogenic gliadin-specific CD4 T cells from the leukocytes of celiac disease patients in vitro. We also find elevated levels of KIRCD8 T cells, but not CD4 regulatory T cells, in COVID-19 patients, correlating with disease severity and vasculitis. Selective ablation of Ly49CD8 T cells in virus-infected mice led to autoimmunity after infection. Our results indicate that in both species, these regulatory CD8 T cells act specifically to suppress pathogenic T cells in autoimmune and infectious diseases.

}, keywords = {Animals, Autoimmune Diseases, CD8-Positive T-Lymphocytes, COVID-19, Humans, Mice, Receptors, KIR, T-Lymphocytes, Regulatory}, issn = {1095-9203}, doi = {10.1126/science.abi9591}, author = {Li, Jing and Zaslavsky, Maxim and Su, Yapeng and Guo, Jing and Sikora, Michael J and van Unen, Vincent and Christophersen, Asbj{\o}rn and Chiou, Shin-Heng and Chen, Liang and Li, Jiefu and Ji, Xuhuai and Wilhelmy, Julie and McSween, Alana M and Palanski, Brad A and Mallajosyula, Venkata Vamsee Aditya and Bracey, Nathan A and Dhondalay, Gopal Krishna R and Bhamidipati, Kartik and Pai, Joy and Kipp, Lucas B and Dunn, Jeffrey E and Hauser, Stephen L and Oksenberg, Jorge R and Satpathy, Ansuman T and Robinson, William H and Dekker, Cornelia L and Steinmetz, Lars M and Khosla, Chaitan and Utz, Paul J and Sollid, Ludvig M and Chien, Yueh-Hsiu and Heath, James R and Fernandez-Becker, Nielsen Q and Nadeau, Kari C and Saligrama, Naresha and Davis, Mark M} } @article {67964, title = {Many dissimilar protein domains switch between α-helix and β-sheet folds}, journal = {Nature Communications}, volume = {13}, year = {2022}, month = {2022 Jul01}, abstract = {

Hundreds of millions of structured proteins sustain life through chemical interactions and catalytic reactions1. Though dynamic, these proteins are assumed to be built upon fixed scaffolds of secondary structure, α-helices and β-sheets. Experimentally determined structures of over \>58,000 non-redundant proteins support this assumption, though it has recently been challenged by \~{}100 fold-switching proteins2. These {\textquotedblleft}metamorphic3{\textquotedblright} proteins, though ostensibly rare, raise the question of how many uncharacterized proteins have shapeshifting{\textendash}rather than fixed{\textendash}secondary structures. To address this question, we developed a comparative sequence-based approach that predicts fold-switching proteins from differences in secondary structure propensity. We applied this approach to the universally conserved NusG transcription factor family of \~{}15,000 proteins, one of which has a 50-residue regulatory subunit experimentally shown to switch between α-helical and β-sheet folds4. Our approach predicted that 25\% of the sequences in this family undergo similar α-helix ⇌ β-sheet transitions, a frequency two orders of magnitude larger than previously observed. Our predictions evade state-of-the-art computational methods but were confirmed experimentally by circular dichroism and nuclear magnetic resonance spectroscopy for all 10 assiduously chosen dissimilar variants. These results suggest that fold switching is a pervasive mechanism of transcriptional regulation in all kingdoms of life and imply that numerous uncharacterized proteins may also switch folds.

}, doi = {10.1101/2021.06.10.447921}, url = {https://doi.org/10.1101/2021.06.10.447921}, author = {Lauren L. Porter and Allen K. Kim and Swechha Rimal and Loren L. Looger and Ananya Majumdar and Brett D. Mensh and Mary Starich} } @article {68299, title = {Many sequence-diverse domains switch between alpha-helix and beta-sheet folds}, journal = {Biophysical Journal}, volume = {121}, year = {2022}, month = {2022 Feb 01}, pages = {156a}, abstract = {

The protein folding paradigm asserts that the three-dimensional structure of a protein is determined by its amino acid sequence. Here we show that a substantial population of proteins from the NusG superfamily of transcription factors do not adhere to this paradigm. Previous work demonstrated that one member of this superfamily has a regulatory domain that completely switches between α-helical and β-sheet folds, but the pervasiveness of this fold-switching mechanism is uncertain. To address this question, we developed a sequence-based predictor, which revealed that thousands of proteins from this superfamily switch folds. Circular dichroism and nuclear magnetic resonance spectroscopies of 10 sequence-diverse variants confirmed our predictions. By contrast, state-of-the-art methods based on the protein folding paradigm assume that related sequences adopt the same fold and thus predicted that the regulatory domains of all variants adopt only the β-sheet fold. Removal of this bias revealed that residue-residue contacts from both α-helical and β-sheet folds are conserved in a large subpopulation of fold-switching domains, poising them to assume disparate conformations. Our results suggest that fold switching is a pervasive mechanism of transcriptional regulation in all kingdoms of life and indicate that expanding the protein folding paradigm may reveal the involvement of fold-switching proteins in diverse biological processes.

}, issn = {00063495}, doi = {10.1016/j.bpj.2021.11.1945}, url = {https://linkinghub.elsevier.com/retrieve/pii/S0006349521029209}, author = {Porter, Lauren L. and Kim, Allen and Looger, Loren and Majumdar, Ananya K. and Starich, Mary} } @article {67929, title = {Melding Synthetic Molecules and Genetically Encoded Proteins to Forge New Tools for Neuroscience.}, journal = {Annual Review of Neuroscience}, year = {2022}, month = {2022 Feb 28}, abstract = {

Unraveling the complexity of the brain requires sophisticated methods to probe and perturb neurobiological processes with high spatiotemporal control. The field of chemical biology has produced general strategies to combine the molecular specificity of small-molecule tools with the cellular specificity of genetically encoded reagents. Here, we survey the application, refinement, and extension of these hybrid small-molecule:protein methods to problems in neuroscience, which yields powerful reagents to precisely measure and manipulate neural systems. Expected final online publication date for the , Volume 45 is July 2022. Please see http://www.annualreviews.org/page/journal/pubdates for revised estimates.

}, issn = {1545-4126}, doi = {10.1146/annurev-neuro-110520-030031}, author = {Kumar, Pratik and Lavis, Luke D} } @article {67947, title = {Melding Synthetic Molecules and Genetically Encoded Proteins to Forge New Tools for Neuroscience.}, journal = {Annual Review Neuroscience}, year = {2022}, month = {2022 Feb 28}, abstract = {

Unraveling the complexity of the brain requires sophisticated methods to probe and perturb neurobiological processes with high spatiotemporal control. The field of chemical biology has produced general strategies to combine the molecular specificity of small-molecule tools with the cellular specificity of genetically encoded reagents. Here, we survey the application, refinement, and extension of these hybrid small-molecule:protein methods to problems in neuroscience, which yields powerful reagents to precisely measure and manipulate neural systems. Expected final online publication date for the , Volume 45 is July 2022. Please see http://www.annualreviews.org/page/journal/pubdates for revised estimates.

}, issn = {1545-4126}, doi = {10.1146/annurev-neuro-110520-030031}, author = {Kumar, Pratik and Lavis, Luke D} } @article {68223, title = {Melding Synthetic Molecules and Genetically Encoded Proteins to Forge New Tools for Neuroscience.}, journal = {Annual Review Neuroscience}, volume = {45}, year = {2022}, month = {2022 Jul 08}, pages = {131-150}, abstract = {

Unraveling the complexity of the brain requires sophisticated methods to probe and perturb neurobiological processes with high spatiotemporal control. The field of chemical biology has produced general strategies to combine the molecular specificity of small-molecule tools with the cellular specificity of genetically encoded reagents. Here, we survey the application, refinement, and extension of these hybrid small-molecule:protein methods to problems in neuroscience, which yields powerful reagents to precisely measure and manipulate neural systems.

}, keywords = {Brain, Neurosciences}, issn = {1545-4126}, doi = {10.1146/annurev-neuro-110520-030031}, author = {Kumar, Pratik and Lavis, Luke D} } @article {68472, title = {Microbial models of development: Inspiration for engineering self-assembled synthetic multicellularity.}, journal = {Semin Cell Dev Biol}, year = {2022}, month = {05/2022}, abstract = {

While the field of synthetic developmental biology has traditionally focused on the study of the rich developmental processes seen in metazoan systems, an attractive alternate source of inspiration comes from microbial developmental models. Microbes face unique lifestyle challenges when forming emergent multicellular collectives. As a result, the solutions they employ can inspire the design of novel multicellular systems. In this review, we dissect the strategies employed in multicellular development by two model microbial systems: the cellular slime mold Dictyostelium discoideum and the biofilm-forming bacterium Bacillus subtilis. Both microbes face similar challenges but often have different solutions, both from metazoan systems and from each other, to create emergent multicellularity. These challenges include assembling and sustaining a critical mass of participating individuals to support development, regulating entry into development, and assigning cell fates. The mechanisms these microbial systems exploit to robustly coordinate development under a wide range of conditions offer inspiration for a new toolbox of solutions to the synthetic development community. Additionally, recreating these phenomena synthetically offers a pathway to understanding the key principles underlying how these behaviors are be coordinated naturally.

}, issn = {1096-3634}, doi = {10.1016/j.semcdb.2022.04.014}, author = {Ricci-Tam, Chiara and Kuipa, Sophia and Kostman, Maya Peters and Aronson, Mark S and Sgro, Allyson E} } @article {68357, title = {Molecularly defined circuits for cardiovascular and cardiopulmonary control}, journal = {Nature}, volume = {606}, year = {2022}, month = {06/2022}, pages = {739 - 746}, abstract = {

The sympathetic and parasympathetic nervous systems regulate the activities of internal organs1, but the molecular and functional diversity of their constituent neurons and circuits remains largely unknown. Here we use retrograde neuronal tracing, single-cell RNA sequencing, optogenetics and physiological experiments to dissect the cardiac parasympathetic control circuit in mice. We show that cardiac-innervating neurons in the brainstem nucleus ambiguus (Amb) are comprised of two molecularly, anatomically and functionally distinct subtypes. The first, which we call ambiguus cardiovascular (ACV) neurons (approximately 35 neurons per Amb), define the classical cardiac parasympathetic circuit. They selectively innervate a subset of cardiac parasympathetic ganglion neurons and mediate the baroreceptor reflex, slowing heart rate and atrioventricular node conduction in response to increased blood pressure. The other, ambiguus cardiopulmonary (ACP) neurons (approximately 15 neurons per Amb) innervate cardiac ganglion neurons intermingled with and functionally indistinguishable from those innervated by ACV neurons. ACP neurons also innervate most or all lung parasympathetic ganglion neurons{\textemdash}clonal labelling shows that individual ACP neurons innervate both organs. ACP neurons mediate the dive reflex, the simultaneous bradycardia and bronchoconstriction that follows water immersion. Thus, parasympathetic control of the heart is organized into two parallel circuits, one that selectively controls cardiac function (ACV circuit) and another that coordinates cardiac and pulmonary function (ACP circuit). This new understanding of cardiac control has implications for treating cardiac and pulmonary diseases and for elucidating the control and coordination circuits of other organs.

}, issn = {0028-0836}, doi = {10.1038/s41586-022-04760-8}, url = {https://www.nature.com/articles/s41586-022-04760-8}, author = {Veerakumar, Avin and Yung, Andrea R. and Liu, Yin and Krasnow, Mark A.} } @article {68194, title = {Multifunctional fluorophores for live-cell imaging and affinity capture of proteins}, journal = {bioRxiv}, year = {2022}, month = {2022 Jul 03}, abstract = {

The development of enzyme-based self-labeling tags allow the labeling of proteins in living cells with synthetic small-molecules. Use of a fluorophore-containing ligand enables the visualization of protein location inside cells using fluorescence microscopy. Alternatively, deployment of a biotin-containing ligand allows purification of tagged protein using affinity resins. Despite these various applications of self-labeling tags, most ligands serve a single purpose. Here, we describe self-labeling tag ligands that allow both visualization and subsequent capture of a protein. A key design principle is exploiting the chemical properties and size of a rhodamine fluorophore to optimize cell-permeability of the ligand and the capture efficiency of the biotin conjugate. This work generates useful {\textquotedblleft}multifunctional{\textquotedblright} fluorophores with generalizable design principles that will allow the construction of new tools for biology.

}, doi = {10.1101/2022.07.02.498544}, url = {https://doi.org/10.1101/2022.07.02.498544}, author = {Kumar, Pratik and Jason D. Vevea and Edwin R. Chapman and Luke D. Lavis} } @article {67895, title = {A neural circuit linking learning and sleep in Drosophila long-term memory.}, journal = {Nature Communications}, volume = {13}, year = {2022}, month = {2022 Feb 01}, pages = {609}, abstract = {

Animals retain some but not all experiences in long-term memory (LTM). Sleep supports LTM retention across animal species. It is well established that learning experiences enhance post-learning sleep. However, the underlying mechanisms of how learning mediates sleep for memory retention are not clear. Drosophila males display increased amounts of sleep after courtship learning. Courtship learning depends on Mushroom Body (MB) neurons, and post-learning sleep is mediated by the sleep-promoting ventral Fan-Shaped Body neurons (vFBs). We show that post-learning sleep is regulated by two opposing output neurons\ (MBONs) from the MB, which encode a measure of learning. Excitatory MBONs-γ2α{\textquoteright}1 becomes increasingly active upon increasing time of learning, whereas inhibitory MBONs-β{\textquoteright}2mp is activated only by a short learning experience. These MB outputs are integrated by SFS neurons, which excite vFBs to promote sleep after prolonged but not short training. This circuit may ensure that only longer or more intense learning experiences induce sleep and are thereby consolidated into LTM.

}, issn = {2041-1723}, doi = {10.1038/s41467-022-28256-1}, author = {Lei, Zhengchang and Henderson, Kristin and Keleman, Krystyna} } @article {68148, title = {Neural circuit pathology driven by Shank3 mutation disrupts social behaviors.}, journal = {Cell Reports}, volume = {39}, year = {2022}, month = {2022 Jun 07}, pages = {110906}, abstract = {

Dysfunctional sociability is a core symptom in autism spectrum disorder (ASD) that may arise from neural-network dysconnectivity between multiple brain regions. However, pathogenic neural-network mechanisms underlying social dysfunction are largely unknown. Here, we demonstrate that circuit-selective mutation (ctMUT) of ASD-risk Shank3 gene within a unidirectional projection from the prefrontal cortex to the basolateral amygdala alters spine morphology and excitatory-inhibitory balance of the circuit. Shank3 ctMUT mice show reduced sociability as well as elevated neural activity and its amplitude variability, which is consistent with the neuroimaging results from human ASD patients. Moreover, the circuit hyper-activity disrupts the temporal correlation of socially tuned neurons to the events of social interactions. Finally, optogenetic circuit activation in wild-type mice partially recapitulates the reduced sociability of Shank3 ctMUT mice, while circuit inhibition in Shank3 ctMUT mice partially rescues social behavior. Collectively, these results highlight a circuit-level pathogenic mechanism of Shank3 mutation that drives social dysfunction.

}, keywords = {Animals, Autism Spectrum Disorder, Disease Models, Animal, Humans, Mice, Microfilament Proteins, Mutation, Nerve Tissue Proteins, Optogenetics, Prefrontal Cortex, Social Behavior}, issn = {2211-1247}, doi = {10.1016/j.celrep.2022.110906}, author = {Kim, Sunwhi and Kim, Yong-Eun and Song, Inuk and Ujihara, Yusuke and Kim, Namsoo and Jiang, Yong-Hui and Yin, Henry H and Lee, Tae-Ho and Kim, Il Hwan} } @article {68501, title = {Neuronal cell types, projections, and spatial organization of the central amygdala}, journal = {iScience}, volume = {25}, year = {2022}, month = {2022 Dec 22}, pages = {105497}, abstract = {

The central amygdala (CEA) has been richly studied for interpreting function and behavior according to specific cell types and circuits. Such work has typically defined molecular cell types by classical inhibitory marker genes; consequently, whether marker-gene-defined cell types exhaustively cover the CEA and co-vary with connectivity remains unresolved. Here, we combined single-cell RNA sequencing, multiplexed fluorescent\ in situ\ hybridization, immunohistochemistry, and long-range projection mapping to derive a {\textquotedblleft}bottom-up{\textquotedblright} understanding of CEA cell types. In doing so, we identify two major cell types, encompassing one-third of all CEA neurons, that have gone unresolved in previous studies. In spatially mapping these novel types, we identify a non-canonical CEA subdomain associated with\ Nr2f2\ expression and uncover an\ Isl1-expressing medial cell type that accounts for many long-range CEA projections. Our results reveal new CEA organizational principles across cell types and spatial scales and provide a framework for future work examining cell-type-specific behavior and function.

}, issn = {25890042}, doi = {10.1016/j.isci.2022.105497}, url = {https://linkinghub.elsevier.com/retrieve/pii/S2589004222017692}, author = {O{\textquoteright}Leary, Timothy P. and Kendrick, Rennie M. and Bristow, Brianna N. and Sullivan, Kaitlin E. and Wang, Lihua and Clements, Jody and Lemire, Andrew L. and Cembrowski, Mark S.} } @article {68012, title = {Nuclear lamin isoforms differentially contribute to LINC complex-dependent nucleocytoskeletal coupling and whole-cell mechanics.}, journal = {Proceedings of the National Academy of Sciences of the U.S.A.}, volume = {119}, year = {2022}, month = {2022 Apr 19}, pages = {e2121816119}, abstract = {

SignificanceInteractions between the cell nucleus and cytoskeleton regulate cell mechanics and are facilitated by the interplay between the nuclear lamina and linker of nucleoskeleton and cytoskeleton (LINC) complexes. To date, the specific contribution of the four lamin isoforms to nucleocytoskeletal connectivity and whole-cell mechanics remains unknown. We discover that A- and B-type lamins distinctively interact with LINC complexes that bind F-actin and vimentin filaments to differentially modulate cortical stiffness, cytoplasmic stiffness, and contractility of mouse embryonic fibroblasts (MEFs). We propose and experimentally verify an integrated lamin-LINC complex-cytoskeleton model that explains cellular mechanical phenotypes in lamin-deficient MEFs. Our findings uncover potential mechanisms for cellular defects in human laminopathies and many cancers associated with mutations or modifications in lamin isoforms.

}, keywords = {Animals, Cell Nucleus, Cytoskeleton, Fibroblasts, Lamin Type A, Lamin Type B, Mice, Nuclear Lamina, Nuclear Matrix, Protein Isoforms}, issn = {1091-6490}, doi = {10.1073/pnas.2121816119}, author = {Vahabikashi, Amir and Sivagurunathan, Suganya and Nicdao, Fiona Ann Sadsad and Han, Yu Long and Park, Chan Young and Kittisopikul, Mark and Wong, Xianrong and Tran, Joseph R and Gundersen, Gregg G and Reddy, Karen L and Luxton, G W Gant and Guo, Ming and Fredberg, Jeffrey J and Zheng, Yixian and Adam, Stephen A and Goldman, Robert D} } @article {68512, title = {Oligodendrocyte precursor cells ingest axons in the mouse neocortex.}, journal = {Proceedings of the National Academies of Science of the U.S.A.}, volume = {119}, year = {2022}, month = {2022 Nov 29}, pages = {e2202580119}, abstract = {

Neurons in the developing brain undergo extensive structural refinement as nascent circuits adopt their mature form. This physical transformation of neurons is facilitated by the engulfment and degradation of axonal branches and synapses by surrounding glial cells, including microglia and astrocytes. However, the small size of phagocytic organelles and the complex, highly ramified morphology of glia have made it difficult to define the contribution of these and other glial cell types to this crucial process. Here, we used large-scale, serial section transmission\ electron microscopy (TEM) with computational volume segmentation to reconstruct the complete 3D morphologies of distinct glial types in the mouse visual cortex, providing unprecedented resolution of their morphology and composition. Unexpectedly, we discovered that the fine processes of oligodendrocyte precursor cells (OPCs), a population of abundant, highly dynamic glial progenitors, frequently surrounded small branches of axons. Numerous phagosomes and phagolysosomes (PLs) containing fragments of axons and vesicular structures were present inside their processes, suggesting that OPCs engage in axon pruning. Single-nucleus RNA sequencing from the developing mouse cortex revealed that OPCs express key phagocytic genes at this stage, as well as neuronal transcripts, consistent with active axon engulfment. Although microglia are thought to be responsible for the majority of synaptic pruning and structural refinement, PLs were ten times more abundant in OPCs than in microglia at this stage, and these structures were markedly less abundant in newly generated oligodendrocytes, suggesting that OPCs contribute substantially to the refinement of neuronal circuits during cortical development.

}, keywords = {Animals, Axons, Mice, Neocortex, Neurons, Oligodendrocyte Precursor Cells, Oligodendroglia}, issn = {1091-6490}, doi = {10.1073/pnas.2202580119}, author = {Buchanan, JoAnn and Elabbady, Leila and Collman, Forrest and Jorstad, Nikolas L and Bakken, Trygve E and Ott, Carolyn and Glatzer, Jenna and Bleckert, Adam A and Bodor, Agnes L and Brittain, Derrick and Bumbarger, Daniel J and Mahalingam, Gayathri and Seshamani, Sharmishtaa and Schneider-Mizell, Casey and Takeno, Marc M and Torres, Russel and Yin, Wenjing and Hodge, Rebecca D and Castro, Manuel and Dorkenwald, Sven and Ih, Dodam and Jordan, Chris S and Kemnitz, Nico and Lee, Kisuk and Lu, Ran and Macrina, Thomas and Mu, Shang and Popovych, Sergiy and Silversmith, William M and Tartavull, Ignacio and Turner, Nicholas L and Wilson, Alyssa M and Wong, William and Wu, Jingpeng and Zlateski, Aleksandar and Zung, Jonathan and Lippincott-Schwartz, Jennifer and Lein, Ed S and Seung, H Sebastian and Bergles, Dwight E and Reid, R Clay and da Costa, Nuno Ma{\c c}arico} } @article {67873, title = {Open-source, Python-based, hardware and software for controlling behavioural neuroscience experiments.}, journal = {eLife}, volume = {11}, year = {2022}, month = {2022 Jan 19}, abstract = {

Laboratory behavioural tasks are an essential research tool. As questions asked of behaviour and brain activity become more sophisticated, the ability to specify and run richly structured tasks becomes more important. An increasing focus on reproducibility also necessitates accurate communication of task logic to other researchers. To these ends, we developed pyControl, a system of open-source hardware and software for controlling behavioural experiments comprising a simple yet flexible Python-based syntax for specifying tasks as extended state machines, hardware modules for building behavioural setups, and a graphical user interface designed for efficiently running high-throughput experiments on many setups in parallel, all with extensive online documentation. These tools make it quicker, easier, and cheaper to implement rich behavioural tasks at scale. As important, pyControl facilitates communication and reproducibility of behavioural experiments through a highly readable task definition syntax and self-documenting features. Here, we outline the system{\textquoteright}s design and rationale, present validation experiments characterising system performance, and demonstrate example applications in freely moving and head-fixed mouse behaviour.

}, issn = {2050-084X}, doi = {10.7554/eLife.67846}, author = {Akam, Thomas and Lustig, Andy and Rowland, James M and Kapanaiah, Sampath Kt and Esteve-Agraz, Joan and Panniello, Mariangela and M{\'a}rquez, Cristina and Kohl, Michael M and K{\"a}tzel, Dennis and Costa, Rui M and Walton, Mark E} } @article {67994, title = {Petascale pipeline for precise alignment of images from serial section electron microscopy.}, journal = {bioRxiv}, year = {2022}, month = {2022 Mar 27}, abstract = {

The reconstruction of neural circuits from serial section electron microscopy (ssEM) images is being accelerated by automatic image segmentation methods. Segmentation accuracy is often limited by the preceding step of aligning 2D section images to create a 3D image stack. Precise and robust alignment in the presence of image artifacts is challenging, especially as datasets are attaining the petascale. We present a computational pipeline for aligning ssEM images with several key elements. Self-supervised convolutional nets are trained via metric learning to encode and align image pairs, and they are used to initialize iterative fine-tuning of alignment. A procedure called vector voting increases robustness to image artifacts or missing image data. For speedup the series is divided into blocks that are distributed to computational workers for alignment. The blocks are aligned to each other by composing transformations with decay, which achieves a global alignment without resorting to a time-consuming global optimization. We apply our pipeline to a whole fly brain dataset, and show improved accuracy relative to prior state of the art. We also demonstrate that our pipeline scales to a cubic millimeter of mouse visual cortex. Our pipeline is publicly available through two open source Python packages.

}, doi = {10.1101/2022.03.25.485816}, url = {https://doi.org/10.1101/2022.03.25.485816}, author = {Sergiy Popovych and Thomas Macrina and Nico Kemnitz and Manuel Castro and Barak Nehoran and Zhen Jia and J. Alexander Bae and Eric Mitchell and Shang Mu and Eric T. Trautman and Stephan Saalfeld and Kai Li and Sebastian Seung} } @article {67892, title = {A population of descending neurons that regulates the flight motor of Drosophila.}, journal = {Current Biology}, volume = {32}, year = {2022}, month = {2022 Mar 14}, pages = {1189-1196}, abstract = {

Similar to many insect species, Drosophila melanogaster is capable of maintaining a stable flight trajectory for periods lasting up to several hours. Because aerodynamic torque is roughly proportional to the fifth power of wing length, even small asymmetries in wing size require the maintenance of subtle bilateral differences in flapping motion to maintain a stable path. Flies can even fly straight after losing half of a wing, a feat they accomplish via very large, sustained kinematic changes to both the damaged and intact wings. Thus, the neural network responsible for stable flight must be capable of sustaining fine-scaled control over wing motion across a large dynamic range. In this study, we describe an unusual type of descending neuron (DNg02) that projects directly from visual output regions of the brain to the dorsal flight neuropil of the ventral nerve cord. Unlike many descending neurons, which exist as single bilateral pairs with unique morphology, there is a population of at least 15 DNg02 cell pairs with nearly identical shape. By optogenetically activating different numbers of DNg02 cells, we demonstrate that these neurons regulate wingbeat amplitude over a wide dynamic range via a population code. Using two-photon functional imaging, we show that DNg02 cells are responsive to visual motion during flight in a manner that would make them well suited to continuously regulate bilateral changes in wing kinematics. Collectively, we have identified a critical set of descending neurons that provides the sensitivity and dynamic range required for flight control.

}, issn = {1879-0445}, doi = {10.1016/j.cub.2022.01.008}, author = {Namiki, Shigehiro and Ros, Ivo G and Morrow, Carmen and Rowell, William J and Card, Gwyneth M and Korff, Wyatt and Dickinson, Michael H} } @article {68983, title = {Post-gastrulation synthetic embryos generated ex utero from mouse naive ESCs}, journal = {Cell}, volume = {185}, year = {2022}, month = {09/2022}, pages = {3290-3306.e25}, abstract = {

Summary In\ vitro cultured stem cells with distinct developmental capacities can contribute to embryonic or extraembryonic tissues after microinjection into pre-implantation mammalian embryos. However, whether cultured stem cells can independently give rise to entire gastrulating embryo-like structures with embryonic and extraembryonic compartments remains unknown. Here, we adapt a recently established platform for prolonged ex utero growth of natural embryos to generate mouse post-gastrulation synthetic whole embryo models (sEmbryos), with both embryonic and extraembryonic compartments, starting solely from naive ESCs. This was achieved by co-aggregating non-transduced ESCs, with naive ESCs transiently expressing Cdx2 or Gata4 to promote their priming toward trophectoderm and primitive endoderm lineages, respectively. sEmbryos adequately accomplish gastrulation, advance through key developmental milestones, and develop organ progenitors within complex extraembryonic compartments similar to E8.5 stage mouse embryos. Our findings highlight the plastic potential of naive pluripotent cells to self-organize and functionally reconstitute and model the entire mammalian embryo beyond gastrulation.

}, keywords = {embryogenesis, embryoids, ESCs, ex utero, extra-embryonic Tissues, iPSCs, naive pluripotency, PGCs, synthetic embryos}, issn = {0092-8674}, doi = {https://doi.org/10.1016/j.cell.2022.07.028}, url = {https://www.sciencedirect.com/science/article/pii/S0092867422009813}, author = {Shadi Tarazi and Alejandro Aguilera-Castrejon and Carine Joubran and Nadir Ghanem and Shahd Ashouokhi and Francesco Roncato and Emilie Wildschutz and Montaser Haddad and Bernardo Oldak and Elidet Gomez-Cesar and Nir Livnat and Sergey Viukov and Dmitry Lokshtanov and Segev Naveh-Tassa and Max Rose and Suhair Hanna and Calanit Raanan and Ori Brenner and Merav Kedmi and Hadas Keren-Shaul and Tsvee Lapidot and Itay Maza and Noa Novershtern and Jacob H. Hanna} } @article {67989, title = {Quantifying Molecular Dynamics within Complex Cellular Morphologies using LLSM-FRAP.}, journal = {Small Methods}, year = {2022}, month = {2022 Jun 01}, pages = {e2200149}, abstract = {

Quantifying molecular dynamics within the context of complex cellular morphologies is essential toward understanding the inner workings and function of cells. Fluorescence recovery after photobleaching (FRAP) is one of the most broadly applied techniques to measure the reaction diffusion dynamics of molecules in living cells. FRAP measurements typically restrict themselves to single-plane image acquisition within a subcellular-sized region of interest due to the limited temporal resolution and undesirable photobleaching induced by 3D fluorescence confocal or widefield microscopy. Here, an experimental and computational pipeline combining lattice light sheet microscopy, FRAP, and numerical simulations, offering rapid and minimally invasive quantification of molecular dynamics with respect to 3D cell morphology is presented. Having the opportunity to accurately measure and interpret the dynamics of molecules in 3D with respect to cell morphology has the potential to reveal unprecedented insights into the function of living cells.

}, issn = {2366-9608}, doi = {10.1002/smtd.202200149}, author = {Colin-York, Huw and Heddleston, John and Wait, Eric and Karedla, Narain and DeSantis, Michael and Khuon, Satya and Chew, Teng-Leong and Sbalzarini, Ivo F and Fritzsche, Marco} } @article {67849, title = {Rapid reconstruction of neural circuits using tissue expansion and lattice light sheet microscopy}, journal = {eLife}, year = {2022}, month = {2022 Oct 26}, abstract = {

Electron microscopy (EM) allows for the reconstruction of dense neuronal connectomes but suffers from low throughput, limiting its application to small numbers of reference specimens. We developed a protocol and analysis pipeline using tissue expansion and lattice light-sheet microscopy (ExLLSM) to rapidly reconstruct selected circuits across many samples with single synapse resolution and molecular contrast. We validate this approach in\ Drosophila, demonstrating that it yields synaptic counts similar to those obtained by EM, can be used to compare counts across sex and experience, and to correlate structural connectivity with functional connectivity. This approach fills a critical methodological gap in studying variability in the structure and function of neural circuits across individuals within and between species.

}, doi = {10.7554/eLife.81248}, url = {https://doi.org/10.7554/eLife.81248}, author = {Joshua L. Lillvis and Hideo Otsuna and Xiaoyu Ding and Igor Pisarev and Takashi Kawase and Jennifer Colonell and Konrad Rokicki and Cristian Goina and Ruixuan Gao and Amy Hu and Kaiyu Wang and John Bogovic and Daniel E. Milkie and Edward S. Boyden and Stephan Saalfeld and Paul W. Tillberg and Barry J. Dickson} } @article {68079, title = {Rescue of behavioral and electrophysiological phenotypes in a Pitt-Hopkins syndrome mouse model by genetic restoration of expression.}, journal = {eLife}, volume = {11}, year = {2022}, month = {2022 May 10}, abstract = {

Pitt-Hopkins syndrome (PTHS) is a neurodevelopmental disorder caused by monoallelic mutation or deletion in the () gene. Individuals with PTHS typically present in the first year of life with developmental delay and exhibit intellectual disability, lack of speech, and motor incoordination. There are no effective treatments available for PTHS, but the root cause of the disorder, haploinsufficiency, suggests that it could be treated by normalizing gene expression. Here, we performed proof-of-concept viral gene therapy experiments using a conditional mouse model of PTHS and found that postnatally reinstating expression in neurons improved anxiety-like behavior, activity levels, innate behaviors, and memory. Postnatal reinstatement also partially corrected EEG abnormalities, which we characterized here for the first time, and the expression of key TCF4-regulated genes. Our results support a genetic normalization approach as a treatment strategy for PTHS, and possibly other TCF4-linked disorders.

}, keywords = {Animals, Disease Models, Animal, Facies, Hyperventilation, Intellectual Disability, Mice, Phenotype, Transcription Factor 4}, issn = {2050-084X}, doi = {10.7554/eLife.72290}, author = {Kim, Hyojin and Gao, Eric B and Draper, Adam and Berens, Noah C and Vihma, Hanna and Zhang, Xinyuan and Higashi-Howard, Alexandra and Ritola, Kimberly D and Simon, Jeremy M and Kennedy, Andrew J and Philpot, Benjamin D} } @article {65821, title = {Robotic Multi-Probe-Single-Actuator Inchworm Neural Microdrive}, journal = {eLife}, year = {2022}, month = {2022 Nov 10}, abstract = {

Electrophysiology is one of the major experimental techniques used in neuroscience. The favorable spatial and temporal resolution as well as the increasingly larger site counts of brain recording electrodes contribute to the popularity and importance of electrophysiology in neuroscience. Such electrodes are typically mechanically placed in the brain to perform acute or chronic freely moving animal measurements. The micro positioners currently used for such tasks employ a single translator per independent probe being placed into the targeted brain region, leading to significant size and weight restrictions. To overcome this limitation, we have developed a miniature robotic multi-probe neural microdrive that utilizes novel phase-change-material-filled resistive heater micro-grippers. The microscopic dimensions, gentle gripping action, independent electronic actuation control, and high packing density of the grippers allow for micrometer-precision independent positioning of multiple arbitrarily shaped parallel neural electrodes with only a single piezo actuator in an inchworm motor configuration. This multi-probe-single-actuator design allows for significant size and weight reduction, as well as remote control and potential automation of the microdrive. We demonstrate accurate placement of multiple independent recording electrodes into the CA1 region of the rat hippocampus\ in vivo\ in acute and chronic settings. Thus, our robotic neural microdrive technology is applicable towards basic neuroscience and clinical studies, as well as other multi-probe or multi-sensor micro-positioning applications.

}, doi = {10.7554/eLife.71876}, author = {Smith, R.D. and Kolb, Ilya and Tanaka, Shinsuke and Lee, A.K. and Harris, T.D. and Barbic, M.} } @article {68559, title = {Robust cell identity specifications through transitions in the collective state of growing developmental systems}, journal = {Current Opinion in Systems Biology}, volume = {31}, year = {2022}, month = {2022 Nov 08}, pages = {100437}, abstract = {

Mammalian development is characterized with transitions from homogeneous populations of precursor to heterogeneous population of specified cells. We review here the main dynamical mechanisms through which such transitions are conceptualized, and discuss that the differentiation timing, robust cell-type proportions and recovery upon perturbation are emergent property of proliferating and communicating cell populations. We argue that studying developmental systems using transitions in collective system states is necessary to describe observed experimental features, and propose additionally the basis of a novel analytical method to deduce the relationship between single-cell dynamics and the collective, symmetry-broken states in cellular populations.

}, issn = {24523100}, doi = {10.1016/j.coisb.2022.100437}, url = {https://linkinghub.elsevier.com/retrieve/pii/S2452310022000233}, author = {Stanoev, Angel and Koseska, Aneta} } @article {67774, title = {RS-FISH: Precise, interactive, fast, and scalable FISH spot detection}, journal = {Nature Methods}, volume = {19}, year = {2022}, month = {2022 Nov 17}, pages = {1563-7}, abstract = {

Fluorescent in-situ hybridization (FISH)-based methods are powerful tools to study molecular processes with subcellular resolution, relying on accurate identification and localization of diffraction-limited spots in microscopy images. We developed the Radial Symmetry-FISH (RS-FISH) software that accurately, robustly, and quickly detects single-molecule spots in two and three dimensions, making it applicable to several key assays, including single-molecule FISH (smFISH), spatial transcriptomics, and spatial genomics. RS-FISH allows interactive parameter tuning and scales to large sets of images as well as tera-byte sized image volumes such as entire brain scans using straight-forward distributed processing on workstations, clusters, and in the cloud.

}, doi = {10.1038/s41592-022-01669-y}, author = {Ella Bahry and Laura Breimann and Marwan Zouinkhi and Leo Epstein and Klim Kolyvanov and Xi Long and Kyle I S Harrington and Timoth{\'e}e Lionnet and Stephan Preibisch} } @article {68251, title = {A scalable and modular automated pipeline for stitching of large electron microscopy datasets.}, journal = {eLife}, volume = {11}, year = {2022}, month = {2022 Jul 26}, abstract = {

Serial-section electronmicroscopy (ssEM) is themethod of choice for studyingmacroscopic biological samples at extremely high resolution in three dimensions. In the nervous system, nanometer-scale images are necessary to reconstruct dense neural wiring diagrams in the brain, so called connectomes. In order to use this data, consisting of up to 10 individual EM images, it must be assembled into a volume, requiring seamless 2D stitching from each physical section followed by 3D alignment of the stitched sections. The high throughput of ssEM necessitates 2D stitching to be done at the pace of imaging, which currently produces tens of terabytes per day. To achieve this, we present a modular volume assembly software pipeline ASAP (Assembly Stitching and Alignment Pipeline) that is scalable to datasets containing petabytes of data and parallelized to work in a distributed computational environment. The pipeline is built on top of the Render (27) services used in the volume assembly of the brain of adult Drosophilamelanogaster (30). It achieves high throughput by operating on themeta-data and transformations of each image stored in a database, thus eliminating the need to render intermediate output. ASAP ismodular, allowing for easy incorporation of new algorithms without significant changes in the workflow. The entire software pipeline includes a complete set of tools for stitching, automated quality control, 3D section alignment, and final rendering of the assembled volume to disk. ASAP has been deployed for continuous stitching of several large-scale datasets of the mouse visual cortex and human brain samples including one cubic millimeter of mouse visual cortex (28; 8) at speeds that exceed imaging. The pipeline also has multi-channel processing capabilities and can be applied to fluorescence and multi-modal datasets like array tomography.

}, issn = {2050-084X}, doi = {10.7554/eLife.76534}, author = {Mahalingam, Gayathri and Torres, Russel and Kapner, Daniel and Trautman, Eric T and Fliss, Tim and Seshamani, Shamishtaa and Perlman, Eric and Young, Rob and Kinn, Samuel and Buchanan, JoAnn and Takeno, Marc M and Yin, Wenjing and Bumbarger, Daniel J and Gwinn, Ryder P and Nyhus, Julie and Lein, Ed and Smith, Steven J and Reid, R Clay and Khairy, Khaled A and Saalfeld, Stephan and Collman, Forrest and Macarico da Costa, Nuno} } @article {68278, title = {Selective Serotonin Reuptake Inhibitors Within Cells: Temporal Resolution in Cytoplasm, Endoplasmic Reticulum, and Membrane}, journal = {bioRxiv}, year = {2022}, month = {2022 Aug 12}, abstract = {

Selective serotonin reuptake inhibitors (SSRIs) are the most prescribed treatment for individuals experiencing major depressive disorder (MDD). The therapeutic mechanisms that take place before, during, or after SSRIs bind the serotonin transporter (SERT) are poorly understood, partially because no studies exist of the cellular and subcellular pharmacokinetic properties of SSRIs in living cells. We studied escitalopram and fluoxetine using new intensity- based drug-sensing fluorescent reporters ({\textquotedblleft}iDrugSnFRs{\textquotedblright}) targeted to the plasma membrane (PM), cytoplasm, or endoplasmic reticulum (ER) of cultured neurons and mammalian cell lines. We also employed chemical detection of drug within cells and phospholipid membranes. The drugs attain equilibrium in neuronal cytoplasm and ER, at approximately the same concentration as the externally applied solution, with time constants of a few s (escitalopram) or 200-300 s (fluoxetine). Simultaneously, the drugs accumulate within lipid membranes by >= 18-fold (escitalopram) or 180-fold (fluoxetine), and possibly by much larger factors. Both drugs leave cytoplasm, lumen, and membranes just as quickly during washout. We synthesized membrane-impermeant quaternary amine derivatives of the two SSRIs. The quaternary derivatives are substantially excluded from membrane, cytoplasm, and ER for \> 2.4 h. They inhibit SERT transport-associated currents 6- or 11-fold less potently than the SSRIs (escitalopram or fluoxetine derivative, respectively), providing useful probes for distinguishing compartmentalized SSRI effects. Although our measurements are orders of magnitude faster than the {\textquotedblleft}therapeutic lag{\textquotedblright} of SSRIs, these data suggest that SSRI-SERT interactions within organelles or membranes may play roles during either the therapeutic effects or the {\textquotedblleft}antidepressant discontinuation syndrome{\textquotedblright}.

}, doi = {10.1101/2022.08.09.502705}, url = {https://doi.org/10.1101/2022.08.09.502705}, author = {Aaron L. Nichols and Zack Blumenfeld and Laura Luebbert and Hailey J. Knox and Anand K. Muthusamy and Jonathan S. Marvin and Charlene H. Kim and Stephen N. Grant and David P. Walton and Bruce N. Cohen and Rebekkah Hammar and Loren L. Looger and Per Artursson and Dennis A. Dougherty and Henry A. Lester} } @article {68312, title = {A sensitive and specific genetically encoded potassium ion biosensor for in vivo applications across the tree of life.}, journal = {PLoS Biology}, volume = {20}, year = {2022}, month = {2022 Sep 06}, pages = {e3001772}, abstract = {

Potassium ion (K+) plays a critical role as an essential electrolyte in all biological systems. Genetically encoded fluorescent K+ biosensors are promising tools to further improve our understanding of K+-dependent processes under normal and pathological conditions. Here, we report the crystal structure of a previously reported genetically encoded fluorescent K+ biosensor, GINKO1, in the K+-bound state. Using structure-guided optimization and directed evolution, we have engineered an improved K+ biosensor, designated GINKO2, with higher sensitivity and specificity. We have demonstrated the utility of GINKO2 for in vivo detection and imaging of K+ dynamics in multiple model organisms, including bacteria, plants, and mice.

}, issn = {1545-7885}, doi = {10.1371/journal.pbio.3001772}, author = {Wu, Sheng-Yi and Wen, Yurong and Serre, Nelson B C and Laursen, Cathrine Charlotte Heiede and Dietz, Andrea Grost{\o}l and Taylor, Brian R and Drobizhev, Mikhail and Molina, Rosana S and Abhi Aggarwal and Rancic, Vladimir and Becker, Michael and Ballanyi, Klaus and Podgorski, Kaspar and Hirase, Hajime and Nedergaard, Maiken and Fendrych, Maty{\'a}{\v s} and Lemieux, M Joanne and Eberl, Daniel F and Kay, Alan R and Campbell, Robert E and Shen, Yi} } @article {67810, title = {A serotonergic axon-cilium synapse drives nuclear signaling to maintain chromatin accessibility}, journal = {Cell}, volume = {185}, year = {2022}, month = {2022 Sep 01}, pages = {3390-3407}, abstract = {

Chemical synapses between axons and dendrites mediate much of the brain{\textquoteright}s intercellular communication. Here we describe a new kind of synapse {\textendash} the axo-ciliary synapse - between axons and primary cilia. By employing enhanced focused ion beam {\textendash} scanning electron microscopy on samples with optimally preserved ultrastructure, we discovered synapses between the serotonergic axons arising from the brainstem, and the primary cilia of hippocampal CA1 pyramidal neurons. Functionally, these cilia are enriched in a ciliary-restricted serotonin receptor, 5-hydroxytryptamine receptor 6 (HTR6), whose mutation is associated with learning and memory defects. Using a newly developed cilia-targeted serotonin sensor, we show that optogenetic stimulation of serotonergic axons results in serotonin release onto cilia. Ciliary HTR6 stimulation activates a non-canonical Gαq/11-RhoA pathway. Ablation of this pathway results in nuclear actin and chromatin accessibility changes in CA1 pyramidal neurons. Axo-ciliary synapses serve as a distinct mechanism for neuromodulators to program neuron transcription through privileged access to the nuclear compartment.

}, doi = {10.1016/j.cell.2022.07.026}, author = {Shu-Hsien Sheu and Srigokul Upadhyayula and Vincent Dupuy and Song Pang and Andrew L. Lemire and Deepika Walpita and H. Amalia Pasolli and Fei Deng and Jinxia Wan and Lihua Wang and Justin Houser and Silvia Sanchez-Martinez and Sebastian E. Brauchi and Sambashiva Banala and Melanie Freeman and C. Shan Xu and Tom Kirchhausen and Harald F. Hess and Luke Lavis and Yu-Long Li and S{\'e}verine Chaumont-Dubel and David E. Clapham} } @article {68297, title = {A single-cell transcriptomic atlas of complete insect nervous systems across multiple life stages.}, journal = {Neural Development}, volume = {17}, year = {2022}, month = {2022 Aug 24}, pages = {8}, abstract = {

Molecular profiles of neurons influence neural development and function but bridging the gap between genes, circuits, and behavior has been very difficult. Here we used single cell RNAseq to generate a complete gene expression atlas of the Drosophila larval central nervous system composed of 131,077 single cells across three developmental stages (1\ h, 24\ h and 48\ h after hatching). We identify 67 distinct cell clusters based on the patterns of gene expression. These include 31 functional mature larval neuron clusters, 1 ring gland cluster, 8 glial clusters, 6 neural precursor clusters, and 13 developing immature adult neuron clusters. Some clusters are present across all stages of larval development, while others are stage specific (such as developing adult neurons). We identify genes that are differentially expressed in each cluster, as well as genes that are differentially expressed at distinct stages of larval life. These differentially expressed genes provide promising candidates for regulating the function of specific neuronal and glial types in the larval nervous system, or the specification and differentiation of adult neurons. The cell transcriptome Atlas of the Drosophila larval nervous system is a valuable resource for developmental biology and systems neuroscience and provides a basis for elucidating how genes regulate neural development and function.

}, keywords = {Animals, Drosophila, Gene Expression Regulation, Developmental, Larva, Neuroglia, Neurons, Transcriptome}, issn = {1749-8104}, doi = {10.1186/s13064-022-00164-6}, author = {Corrales, Marc and Cocanougher, Benjamin T and Kohn, Andrea B and Wittenbach, Jason D and Long, Xi S and Lemire, Andrew and Cardona, Albert and Singer, Robert H and Moroz, Leonid L and Zlatic, Marta} } @article {68514, title = {SMP30-mediated synthesis of vitamin C activates the liver PPARα/FGF21 axis to regulate thermogenesis in mice.}, journal = {Experimental and Molecular Medicine}, volume = {54}, year = {2022}, month = {2022 Nov 25}, pages = {2036-2046}, abstract = {

The vitamin-C-synthesizing enzyme senescent marker protein 30 (SMP30) is a cold resistance gene in Drosophila, and vitamin C concentration increases in brown adipose tissue post-cold exposure. However, the roles of SMP30 in thermogenesis are unknown. Here, we tested the molecular mechanism of thermogenesis using wild-type (WT) and vitamin C-deficient SMP30-knockout (KO) mice. SMP30-KO mice gained more weight than WT mice without a change in food intake in response to short-term high-fat diet feeding. Indirect calorimetry and cold-challenge experiments indicated that energy expenditure is lower in SMP30-KO mice, which is associated with decreased thermogenesis in adipose tissues. Therefore, SMP30-KO mice do not lose weight during cold exposure, whereas WT mice lose weight markedly. Mechanistically, the levels of serum FGF21 were notably lower in SMP30-KO mice, and vitamin C supplementation in SMP30-KO mice recovered FGF21 expression and thermogenesis, with a marked reduction in body weight during cold exposure. Further experiments revealed that vitamin C activates PPARα to upregulate FGF21. Our findings demonstrate that SMP30-mediated synthesis of vitamin C activates the PPARα/FGF21 axis, contributing to the maintenance of thermogenesis in mice.

}, issn = {2092-6413}, doi = {10.1038/s12276-022-00888-9}, author = {Lee, Bonggi and An, Hye Jin and Kim, Dae Hyun and Lee, Min-Kyeong and Jeong, Hyeon Hak and Chung, Ki Wung and Go, Younghoon and Seo, Arnold Y and Kim, Il Yong and Seong, Je Kyung and Yu, Byung Pal and Lee, Jaewon and Im, Eunok and Lee, In-Kyu and Lee, Myung-Shik and Yamada, Ken-Ichi and Chung, Hae Young} } @article {68408, title = {Structure of the OMEGA nickase IsrB in complex with ωRNA and target DNA.}, journal = {Nature}, volume = {610}, year = {2022}, month = {2022 Oct 12}, pages = {575-581}, abstract = {

RNA-guided systems, such as CRISPR-Cas, combine programmable substrate recognition with enzymatic function, a combination that has been used advantageously to develop powerful molecular technologies. Structural studies of these systems have illuminated how the RNA and protein jointly recognize and cleave their substrates, guiding rational engineering for further technology development. Recent work identified a new class of RNA-guided systems, termed OMEGA, which include IscB, the likely ancestor of Cas9, and the nickase IsrB, a homologue of IscB lacking the HNH nuclease domain. IsrB consists of only around 350 amino acids, but its small size is counterbalanced by a relatively large RNA guide (roughly 300-nt ωRNA). Here, we report the cryogenic-electron microscopy structure of Desulfovirgula thermocuniculi IsrB (DtIsrB) in complex with its cognate ωRNA and a target DNA. We find the overall structure of the IsrB protein shares a common scaffold with Cas9. In contrast to Cas9, however, which uses a recognition (REC) lobe to facilitate target selection, IsrB relies on its ωRNA, part of which forms an intricate ternary structure positioned analogously to REC. Structural analyses of IsrB and its ωRNA as well as comparisons to other RNA-guided systems highlight the functional interplay between protein and RNA, advancing our understanding of the biology and evolution of these diverse systems.

}, keywords = {CRISPR-Associated Proteins, CRISPR-Cas Systems, Cryoelectron Microscopy, Deoxyribonuclease I, DNA, RNA, Guide}, issn = {1476-4687}, doi = {10.1038/s41586-022-05324-6}, author = {Hirano, Seiichi and Kappel, Kalli and Altae-Tran, Han and Faure, Guilhem and Wilkinson, Max E and Kannan, Soumya and Demircioglu, F Esra and Yan, Rui and Shiozaki, Momoko and Yu, Zhiheng and Makarova, Kira S and Koonin, Eugene V and Macrae, Rhiannon K and Zhang, Feng} } @article {68222, title = {Structured sampling of olfactory input by the fly mushroom body.}, journal = {Current Biology}, volume = {32}, year = {2022}, month = {2022 Aug 08}, pages = {3334-3349.e6}, abstract = {

Associative memory formation and recall in the fruit fly Drosophila melanogaster is subserved by the mushroom body (MB). Upon arrival in the MB, sensory information undergoes a profound transformation from broadly tuned and stereotyped odorant responses in the olfactory projection neuron (PN) layer to narrowly tuned and nonstereotyped responses in the Kenyon cells (KCs). Theory and experiment suggest that this transformation is implemented by random connectivity between KCs and PNs. However, this hypothesis has been challenging to test, given the difficulty of mapping synaptic connections between large numbers of brain-spanning neurons. Here, we used a recent whole-brain electron microscopy volume of the adult fruit fly to map PN-to-KC connectivity at synaptic resolution. The PN-KC connectome revealed unexpected structure, with preponderantly food-responsive PN types converging at above-chance levels on downstream KCs. Axons of the overconvergent PN types tended to arborize near one another in the MB main calyx, making local KC dendrites more likely to receive input from those types. Overconvergent PN types preferentially co-arborize and connect with dendrites of αβ and α{\textquoteright}β{\textquoteright} KC subtypes. Computational simulation of the observed network showed degraded discrimination performance compared with a random network, except when all signal flowed through the overconvergent, primarily food-responsive PN types. Additional theory and experiment will be needed to fully characterize the impact of the observed non-random network structure on associative memory formation and recall.

}, issn = {1879-0445}, doi = {10.1016/j.cub.2022.06.031}, author = {Zheng, Zhihao and Li, Feng and Fisher, Corey and Ali, Iqbal J and Sharifi, Nadiya and Calle-Schuler, Steven and Hsu, Joseph and Masoodpanah, Najla and Kmecova, Lucia and Kazimiers, Tom and Perlman, Eric and Nichols, Matthew and Li, Peter H and Jain, Viren and Bock, Davi D} } @article {68987, title = {SUMOylation of linker histone H1 drives chromatin condensation and restriction of embryonic cell fate identity}, journal = {Molecular Cell}, volume = {82}, year = {2022}, month = {01/2022}, pages = {106-122.e9}, abstract = {

Summary The fidelity of the early embryonic program is underlined by tight regulation of the chromatin. Yet, how the chromatin is organized to prohibit the reversal of the developmental program remains unclear. Specifically, the totipotency-to-pluripotency transition marks one of the most dramatic events to the chromatin, and yet, the nature of histone alterations underlying this process is incompletely characterized. Here, we show that linker histone H1 is post-translationally modulated by SUMO2/3, which facilitates its fixation onto ultra-condensed heterochromatin in embryonic stem cells (ESCs). Upon SUMOylation depletion, the chromatin becomes de-compacted and H1 is evicted, leading to totipotency reactivation. Furthermore, we show that H1 and SUMO2/3 jointly mediate the repression of totipotent elements. Lastly, we demonstrate that preventing SUMOylation on H1 abrogates its ability to repress the totipotency program in ESCs. Collectively, our findings unravel a critical role for SUMOylation of H1 in facilitating chromatin repression and desolation of the totipotent identity.

}, issn = {1097-2765}, doi = {https://doi.org/10.1016/j.molcel.2021.11.011}, url = {https://www.sciencedirect.com/science/article/pii/S1097276521009904}, author = {Daoud Sheban and Tom Shani and Roey Maor and Alejandro Aguilera-Castrejon and Nofar Mor and Bernardo Oldak and Merav D. Shmueli and Avital Eisenberg-Lerner and Jonathan Bayerl and Jakob Hebert and Sergey Viukov and Guoyun Chen and Assaf Kacen and Vladislav Krupalnik and Valeriya Chugaeva and Shadi Tarazi and Alejandra Rodr{\'\i}guez-delaRosa and Mirie Zerbib and Adi Ulman and Solaiman Masarwi and Meital Kupervaser and Yishai Levin and Efrat Shema and Yael David and Noa Novershtern and Jacob H. Hanna and Yifat Merbl} } @article {68078, title = {Super-resolution microscopy reveals actomyosin dynamics in medioapical arrays.}, journal = {Molecular Biology of the Cell}, year = {2022}, month = {2022 May 11}, pages = {mbcE21110537}, abstract = {

Arrays of actin filaments (F-actin) near the apical surface of epithelial cells (medioapical arrays) contribute to apical constriction and morphogenesis throughout phylogeny. Here, super-resolution approaches (grazing incidence structured illumination, GI-SIM and lattice light sheet, LLSM) microscopy resolve individual, fluorescently labeled F-actin and bipolar myosin filaments that drive amnioserosa cell shape changes during dorsal closure in . In expanded cells, F-actin and myosin form loose, apically domed meshworks at the plasma membrane. The arrays condense as cells contract, drawing the domes into the plane of the junctional belts. As condensation continues, individual filaments are no longer uniformly apparent. As cells expand, arrays of actomyosin are again resolved - some F-actin turnover likely occurs, but a large fraction of existing filaments rearrange. In morphologically isotropic cells, actin filaments are randomly oriented and during contraction, are drawn together but remain essentially randomly oriented. In anisotropic cells, largely parallel actin filaments are drawn closer to one another. Our images offer unparalleled resolution of F-actin in embryonic tissue show that medioapical arrays are tightly apposed to the plasma membrane, are continuous with meshworks of lamellar F-actin and thereby constitute modified cell cortex. In concert with other tagged array components, super-resolution imaging of live specimens will offer new understanding of cortical architecture and function. [Media: see text] [Media: see text] [Media: see text] [Media: see text] [Media: see text] [Media: see text] [Media: see text] [Media: see text] [Media: see text] [Media: see text].

}, issn = {1939-4586}, doi = {10.1091/mbc.E21-11-0537}, author = {Moore, Regan P and Fogerson, Stephanie M and Tulu, U Serdar and Yu, Jason W and Cox, Amanda H and Sican, Melissa A and Li, Dong and Legant, Wesley R and Weigel, Aubrey V and Crawford, Janice M and Betzig, Eric and Kiehart, Daniel P} } @article {68328, title = {Superresolution microscopy reveals actomyosin dynamics in medioapical arrays.}, journal = {Molecular Biology of the Cell}, volume = {33}, year = {2022}, month = {2022 Sep 15}, pages = {ar94}, abstract = {

Arrays of actin filaments (F-actin) near the apical surface of epithelial cells (medioapical arrays) contribute to apical constriction and morphogenesis throughout phylogeny. Here, superresolution approaches (grazing incidence structured illumination, GI-SIM, and lattice light sheet, LLSM) microscopy resolve individual, fluorescently labeled F-actin and bipolar myosin filaments that drive amnioserosa cell shape changes during dorsal closure in . In expanded cells, F-actin and myosin form loose, apically domed meshworks at the plasma membrane. The arrays condense as cells contract, drawing the domes into the plane of the junctional belts. As condensation continues, individual filaments are no longer uniformly apparent. As cells expand, arrays of actomyosin are again resolved-some F-actin turnover likely occurs, but a large fraction of existing filaments rearrange. In morphologically isotropic cells, actin filaments are randomly oriented and during contraction are drawn together but remain essentially randomly oriented. In anisotropic cells, largely parallel actin filaments are drawn closer to one another. Our images offer unparalleled resolution of F-actin in embryonic tissue, show that medioapical arrays are tightly apposed to the plasma membrane and are continuous with meshworks of lamellar F-actin. Medioapical arrays thereby constitute modified cell cortex. In concert with other tagged array components, superresolution imaging of live specimens will offer new understanding of cortical architecture and function.

}, keywords = {Actin Cytoskeleton, Actins, Actomyosin, Animals, Drosophila, Microscopy, Myosins}, issn = {1939-4586}, doi = {10.1091/mbc.E21-11-0537}, author = {Moore, Regan P and Fogerson, Stephanie M and Tulu, U Serdar and Yu, Jason W and Cox, Amanda H and Sican, Melissa A and Li, Dong and Legant, Wesley R and Weigel, Aubrey V and Crawford, Janice M and Betzig, Eric and Kiehart, Daniel P} } @article {68145, title = {Targeting LIPA independent of its lipase activity is a therapeutic strategy in solid tumors via induction of endoplasmic reticulum stress.}, journal = {Nature Cancer}, volume = {3}, year = {2022}, month = {2022 Jun 02}, pages = {866-884}, abstract = {

Triple-negative breast cancer (TNBC) has a poor clinical outcome, due to a lack of actionable therapeutic targets. Herein we define lysosomal acid lipase A (LIPA) as a viable molecular target in TNBC and identify a stereospecific small molecule (ERX-41) that binds LIPA. ERX-41 induces endoplasmic reticulum (ER) stress resulting in cell death, and this effect is on target as evidenced by specific LIPA mutations providing resistance. Importantly, we demonstrate that ERX-41 activity is independent of LIPA lipase function but dependent on its ER localization. Mechanistically, ERX-41 binding of LIPA decreases expression of multiple ER-resident proteins involved in protein folding. This targeted vulnerability has a large therapeutic window, with no adverse effects either on normal mammary epithelial cells or in mice. Our study implicates a targeted strategy for solid tumors, including breast, brain, pancreatic and ovarian, whereby small, orally bioavailable molecules targeting LIPA block protein folding, induce ER stress and result in tumor cell death.

}, issn = {2662-1347}, doi = {10.1038/s43018-022-00389-8}, author = {Liu, Xihui and Viswanadhapalli, Suryavathi and Kumar, Shourya and Lee, Tae-Kyung and Moore, Andrew and Ma, Shihong and Chen, Liping and Hsieh, Michael and Li, Mengxing and Sareddy, Gangadhara R and Parra, Karla and Blatt, Eliot B and Reese, Tanner C and Zhao, Yuting and Chang, Annabel and Yan, Hui and Xu, Zhenming and Pratap, Uday P and Liu, Zexuan and Roggero, Carlos M and Tan, Zhenqiu and Weintraub, Susan T and Peng, Yan and Tekmal, Rajeshwar R and Arteaga, Carlos L and Lippincott-Schwartz, Jennifer and Vadlamudi, Ratna K and Ahn, Jung-Mo and Raj, Ganesh V} } @article {68518, title = {TEMPO enables sequential genetic labeling and manipulation of vertebrate cell lineages.}, journal = {Neuron}, year = {2022}, month = {2022 Nov 18}, abstract = {

During development, regulatory factors appear in a precise order to determine cell fates over time. Consequently, to investigate complex tissue development, it is necessary to visualize and manipulate cell lineages with temporal control. Current strategies for tracing vertebrate cell lineages lack genetic access to sequentially produced cells. Here, we present TEMPO (Temporal Encoding and Manipulation in a Predefined Order), an imaging-readable genetic tool allowing differential labeling and manipulation of consecutive cell generations in vertebrates. TEMPO is based on CRISPR and powered by a cascade of gRNAs that drive orderly activation and inactivation of reporters and/or effectors. Using TEMPO to visualize zebrafish and mouse neurogenesis, we recapitulated birth-order-dependent neuronal fates. Temporally manipulating cell-cycle regulators in mouse cortex progenitors altered the proportion and distribution of neurons and glia, revealing the effects of temporal gene perturbation on serial cell fates. Thus, TEMPO enables sequential manipulation of molecular factors, crucial to study cell-type specification.

}, issn = {1097-4199}, doi = {10.1016/j.neuron.2022.10.035}, author = {Espinosa-Medina, Isabel and Feliciano, Daniel and Belmonte-Mateos, Carla and Linda Miyares, Rosa and Garcia-Marques, Jorge and Foster, Benjamin and Lindo, Sarah and Pujades, Cristina and Koyama, Minoru and Lee, Tzumin} } @article {67928, title = {Three Mutations Convert the Selectivity of a Protein Sensor From Nicotinic Agonists to S-Methadone For Use in Cells, Organelles, and Biofluids.}, journal = {Journal of the American Chemical Society}, volume = {144}, year = {2022}, month = {2022 May 18}, abstract = {

We report a reagentless, intensity-based S-methadone fluorescent sensor, iS-methadoneSnFR, consisting of a circularly permuted GFP inserted within the sequence of a mutated bacterial periplasmic binding protein (PBP). We used directed evolution to convert a previously reported nicotine-binding PBP to a selective S-methadone-binding sensor, via three mutations in the PBP{\textquoteright}s second shell and hinge regions. iS-methadoneSnFR displays sensitivity across the pharmacologically relevant range and selectivity against endogenous analytes and other opioids. Robust iS-methadoneSnFR responses in human sweat and saliva and mouse serum enable diagnostic uses. Genetic encoding and imaging in mammalian demonstrated the acid trapping of S-methadone in the Golgi apparatus where opioid receptors can signal. This work shows a straightforward strategy in adapting existing PBPs to serve real-time applications ranging from subcellular to personal pharmacokinetics.

}, doi = {10.1101/2022.02.24.481226}, author = {Anand K. Muthusamy and Charlene H. Kim and Scott C. Virgil and Hailey J. Knox and Jonathan S. Marvin and Aaron L. Nichols and Bruce N. Cohen and Dennis A. Dougherty and Loren L. Looger and Henry A. Lester} } @article {68419, title = {Transcription factor Acj6 controls dendrite targeting via a combinatorial cell-surface code.}, journal = {Neuron}, volume = {110}, year = {2022}, month = {07/2022}, pages = {2299-2314.e8}, abstract = {

Transcription factors specify the fate and connectivity of developing neurons. We investigate how a lineage-specific transcription factor, Acj6, controls the precise dendrite targeting of Drosophila olfactory projection neurons (PNs) by regulating the expression of cell-surface proteins. Quantitative cell-surface proteomic profiling of wild-type and acj6 mutant PNs in intact developing brains, and a proteome-informed genetic screen identified PN surface proteins that execute Acj6-regulated wiring decisions. These include canonical cell adhesion molecules and proteins previously not associated with wiring, such as Piezo, whose mechanosensitive ion channel activity is dispensable for its function in PN dendrite targeting. Comprehensive genetic analyses revealed that Acj6 employs unique sets of cell-surface proteins in different PN types for dendrite targeting. Combined expression of Acj6 wiring executors rescued acj6 mutant phenotypes with higher efficacy and breadth than expression of individual executors. Thus, Acj6 controls wiring specificity of different neuron types by specifying distinct combinatorial expression of cell-surface executors.

}, keywords = {Animals, Dendrites, Drosophila, Drosophila Proteins, Ion Channels, Membrane Proteins, Nerve Tissue Proteins, Olfactory Pathways, Olfactory Receptor Neurons, POU Domain Factors, Proteomics, Transcription Factors}, issn = {1097-4199}, doi = {10.1016/j.neuron.2022.04.026}, author = {Xie, Qijing and Li, Jiefu and Li, Hongjie and Udeshi, Namrata D and Svinkina, Tanya and Orlin, Daniel and Kohani, Sayeh and Guajardo, Ricardo and Mani, D R and Xu, Chuanyun and Li, Tongchao and Han, Shuo and Wei, Wei and Shuster, S Andrew and Luginbuhl, David J and Quake, Stephen R and Murthy, Swetha E and Ting, Alice Y and Carr, Steven A and Luo, Liqun} } @article {68812, title = {A viral toolbox for conditional and transneuronal gene expression in zebrafish}, journal = {eLife}, volume = {11}, year = {2022}, month = {07/2022}, pages = {e77153}, abstract = {

The zebrafish is an important model in systems neuroscience but viral tools to dissect the structure and function of neuronal circuitry are not established. We developed methods for efficient gene transfer and retrograde tracing in adult and larval zebrafish by herpes simplex viruses (HSV1). HSV1 was combined with the Gal4/UAS system to target cell types with high spatial, temporal, and molecular specificity. We also established methods for efficient transneuronal tracing by modified rabies viruses in zebrafish. We demonstrate that HSV1 and rabies viruses can be used to visualize and manipulate genetically or anatomically identified neurons within and across different brain areas of adult and larval zebrafish. An expandable library of viruses is provided to express fluorescent proteins, calcium indicators, optogenetic probes, toxins and other molecular tools. This toolbox creates new opportunities to interrogate neuronal circuits in zebrafish through combinations of genetic and viral approaches.

}, keywords = {Gene Transfer, Imaging, neuronal circuit, viral vector}, issn = {2050-084X}, doi = {10.7554/eLife.77153}, url = {https://doi.org/10.7554/eLife.77153}, author = {Satou, Chie and Neve, Rachael L and Oyibo, Hassana K and Zmarz, Pawel and Huang, Kuo-Hua and Arn Bouldoires, Estelle and Mori, Takuma and Higashijima, Shin-Ichi and Keller, Georg B and Friedrich, Rainer W}, editor = {Bagnall, Martha W and Stainier, Didier YR and Jin, Lei} } @article {66029, title = {Visualizing cellular and tissue ultrastructure using Ten-fold Robust Expansion Microscopy (TREx)}, journal = {eLife}, year = {2022}, month = {2022 Feb 18}, abstract = {

Expansion microscopy (ExM) is a powerful technique to overcome the diffraction limit of light microscopy that can be applied in both tissues and cells. In ExM, samples are embedded in a swellable polymer gel to physically expand the sample and isotropically increase resolution in x, y and z. The maximum resolution increase is limited by the expansion factor of the polymer gel, which is four-fold for the original ExM protocol. Variations on the original ExM method have been reported that allow for greater expansion factors, for example using iterative expansion, but at the cost of ease of adoption or versatility. Here, we systematically explore the ExM recipe space and present a novel method termed Ten-fold Robust Expansion Microscopy (TREx) that, like the original ExM method, requires no specialized equipment or procedures to carry out. We demonstrate that TREx gels expand ten-fold, can be handled easily, and can be applied to both thick tissue sections and cells enabling high-resolution subcellular imaging in a single expansion step. We show that applying TREx on antibody-stained samples can be combined with off-the-shelf small molecule stains for both total protein and membranes to provide ultrastructural context to subcellular protein localization.

}, doi = {https://doi.org/10.1101/2021.02.03.428837}, author = {Hugo G.J. Damstra and Boaz Mohar and Mark Eddison and Anna Akhmanova and Lukas C. Kapitein and Paul W. Tillberg} } @article {67894, title = {Visualizing Synaptic Dopamine Efflux with a 2D Nanofilm.}, journal = {eLife}, year = {2022}, month = {2022 Jul 04}, abstract = {

Chemical neurotransmission constitutes one of the fundamental modalities of communication between neurons. Monitoring release of these chemicals has traditionally been difficult to carry out at spatial and temporal scales relevant to neuron function. To understand chemical neurotransmission more fully, we need to improve the spatial and temporal resolutions of measurements for neurotransmitter release. To address this, we engineered a chemi-sensitive, two-dimensional nanofilm that facilitates subcellular visualization of the release and diffusion of the neurochemical dopamine with synaptic resolution, quantal sensitivity, and simultaneously from hundreds of release sites. Using this technology, we were able to monitor the spatiotemporal dynamics of dopamine release in dendritic processes, a poorly understood phenomenon. We found that dopamine release is broadcast from a subset of dendritic processes as hotspots that have a mean spatial spread of ≈3.2 {\textmu}m (full width at half maximum) and are observed with a mean spatial frequency of 1 hotspot per ≈7.5 {\textmu}m of dendritic length. Major dendrites of dopamine neurons and fine dendritic processes, as well as dendritic arbors and dendrites with no apparent varicose morphology participated in dopamine release. Remarkably, these release hotspots colocalized with Bassoon, suggesting that Bassoon may contribute to organizing active zones in dendrites, similar to its role in axon terminals.

}, doi = {10.7554/eLife.78773}, url = {https://doi.org/10.7554/eLife.78773}, author = {Chandima Bulumulla and Andrew T. Krasley and Deepika Walpita and Abraham G. Beyene} } @article {67890, title = {Voltage imaging identifies spinal circuits that modulate locomotor adaptation in zebrafish.}, journal = {Neuron}, year = {2022}, month = {2022 Jan 24}, abstract = {

Motor systems must continuously adapt their output to maintain a desired trajectory. While the spinal circuits underlying rhythmic locomotion are well described, little is known about how the network modulates its output strength. A major challenge has been the difficulty of recording from spinal neurons during behavior. Here, we use voltage imaging to map the membrane potential of large populations of glutamatergic neurons throughout the spinal cord of the larval zebrafish during fictive swimming in a virtual environment. We characterized a previously undescribed subpopulation of tonic-spiking ventral V3 neurons whose spike rate correlated with swimming strength and bout length. Optogenetic activation of V3 neurons led to stronger swimming and longer bouts but did not affect tail beat frequency. Genetic ablation of V3 neurons led to reduced locomotor adaptation. The power of voltage imaging allowed us to identify V3 neurons as a critical driver of locomotor adaptation in zebrafish.

}, issn = {1097-4199}, doi = {10.1016/j.neuron.2022.01.001}, author = {B{\"o}hm, Urs L and Kimura, Yukiko and Kawashima, Takashi and Ahrens, Misha B and Higashijima, Shin-Ichi and Engert, Florian and Cohen, Adam E} } @article {67991, title = {Voltage imaging identifies spinal circuits that modulate locomotor adaptation in zebrafish.}, journal = {Neuron}, volume = {110}, year = {2022}, month = {2022 Apr 06}, pages = {1211-1222.e4}, abstract = {

Motor systems must continuously adapt their output to maintain a desired trajectory. While the spinal circuits underlying rhythmic locomotion are well described, little is known about how the network modulates its output strength. A major challenge has been the difficulty of recording from spinal neurons during behavior. Here, we use voltage imaging to map the membrane potential of large populations of glutamatergic neurons throughout the spinal cord of the larval zebrafish during fictive swimming in a virtual environment. We characterized a previously undescribed subpopulation of tonic-spiking ventral V3 neurons whose spike rate correlated with swimming strength and bout length. Optogenetic activation of V3 neurons led to stronger swimming and longer bouts but did not affect tail beat frequency. Genetic ablation of V3 neurons led to reduced locomotor adaptation. The power of voltage imaging allowed us to identify V3 neurons as a critical driver of locomotor adaptation in zebrafish.

}, issn = {1097-4199}, doi = {10.1016/j.neuron.2022.01.001}, author = {B{\"o}hm, Urs L and Kimura, Yukiko and Kawashima, Takashi and Ahrens, Misha B and Higashijima, Shin-Ichi and Engert, Florian and Cohen, Adam E} } @article {68887, title = {Volume electron microscopy.}, journal = {Nature Reviews: Methods Primers}, volume = {2}, year = {2022}, month = {2022 Jul 07}, pages = {51}, abstract = {

Life exists in three dimensions, but until the turn of the century most electron microscopy methods provided only 2D image data. Recently, electron microscopy techniques capable of delving deep into the structure of cells and tissues have emerged, collectively called volume electron microscopy (vEM). Developments in vEM have been dubbed a quiet revolution as the field evolved from established transmission and scanning electron microscopy techniques, so early publications largely focused on the bioscience applications rather than the underlying technological breakthroughs. However, with an explosion in the uptake of vEM across the biosciences and fast-paced advances in volume, resolution, throughput and ease of use, it is timely to introduce the field to new audiences. In this Primer, we introduce the different vEM imaging modalities, the specialized sample processing and image analysis pipelines that accompany each modality and the types of information revealed in the data. We showcase key applications in the biosciences where vEM has helped make breakthrough discoveries and consider limitations and future directions. We aim to show new users how vEM can support discovery science in their own research fields and inspire broader uptake of the technology, finally allowing its full adoption into mainstream biological imaging.

}, issn = {2662-8449}, doi = {10.1038/s43586-022-00131-9}, author = {Peddie, Christopher J and Genoud, Christel and Kreshuk, Anna and Meechan, Kimberly and Micheva, Kristina D and Narayan, Kedar and Pape, Constantin and Parton, Robert G and Schieber, Nicole L and Schwab, Yannick and Titze, Benjamin and Verkade, Paul and Aubrey, Aubrey and Collinson, Lucy M} } @article {67987, title = {When light meets biology - how the specimen affects quantitative microscopy.}, journal = {Journal of Cell Science}, volume = {135}, year = {2022}, month = {2022 Mar 15}, abstract = {

Fluorescence microscopy images should not be treated as perfect representations of biology. Many factors within the biospecimen itself can drastically affect quantitative microscopy data. Whereas some sample-specific considerations, such as photobleaching and autofluorescence, are more commonly discussed, a holistic discussion of sample-related issues (which includes less-routine topics such as quenching, scattering and biological anisotropy) is required to appropriately guide life scientists through the subtleties inherent to bioimaging. Here, we consider how the interplay between light and a sample can cause common experimental pitfalls and unanticipated errors when drawing biological conclusions. Although some of these discrepancies can be minimized or controlled for, others require more pragmatic considerations when interpreting image data. Ultimately, the power lies in the hands of the experimenter. The goal of this Review is therefore to survey how biological samples can skew quantification and interpretation of microscopy data. Furthermore, we offer a perspective on how to manage many of these potential pitfalls.

}, issn = {1477-9137}, doi = {10.1242/jcs.259656}, author = {Reiche, Michael A and Aaron, Jesse S and Boehm, Ulrike and DeSantis, Michael C and Hobson, Chad M and Khuon, Satya and Lee, Rachel M and Chew, Teng-Leong} } @article {65905, title = {3D FIB-SEM reconstruction of microtubule-organelle interaction in whole primary mouse β cells.}, journal = {Journal of Cell Biology}, volume = {220}, year = {2021}, month = {2021 Feb 01}, abstract = {

Microtubules play a major role in intracellular trafficking of vesicles in endocrine cells. Detailed knowledge of microtubule organization and their relation to other cell constituents is crucial for understanding cell function. However, their role in insulin transport and secretion is under debate. Here, we use FIB-SEM to image islet β cells in their entirety with unprecedented resolution. We reconstruct mitochondria, Golgi apparati, centrioles, insulin secretory granules, and microtubules of seven β cells, and generate a comprehensive spatial map of microtubule-organelle interactions. We find that microtubules form nonradial networks that are predominantly not connected to either centrioles or endomembranes. Microtubule number and length, but not microtubule polymer density, vary with glucose stimulation. Furthermore, insulin secretory granules are enriched near the plasma membrane, where they associate with microtubules. In summary, we provide the first 3D reconstructions of complete microtubule networks in primary mammalian cells together with evidence regarding their importance for insulin secretory granule positioning and thus their supportive role in insulin secretion.

}, issn = {1540-8140}, doi = {10.1083/jcb.202010039}, author = {M{\"u}ller, Andreas and Schmidt, Deborah and Xu, C Shan and Pang, Song and D{\textquoteright}Costa, Joyson Verner and Kretschmar, Susanne and M{\"u}nster, Carla and Kurth, Thomas and Jug, Florian and Weigert, Martin and Hess, Harald F and Solimena, Michele} } @article {67298, title = {Adhesion-mediated mechanosignaling forces mitohormesis.}, journal = {Cell Metabolism}, volume = {33}, year = {2021}, month = {2021 July 6}, pages = {1322}, abstract = {

Mitochondria control eukaryotic cell fate by producing the energy needed to support life and the signals required to execute programed cell death. The biochemical milieu is known to affect mitochondrial function and contribute to the dysfunctional mitochondrial phenotypes implicated in cancer and the morbidities of aging. However, the physical characteristics of the extracellular matrix are also altered in cancerous and aging tissues. Here, we demonstrate that cells sense the physical properties of the extracellular matrix and activate a mitochondrial stress response that adaptively tunes mitochondrial function via solute carrier family 9 member A1-dependent ion exchange and heat shock factor 1-dependent transcription. Overall, our data indicate that adhesion-mediated mechanosignaling may play an unappreciated role in the altered mitochondrial functions observed in aging and cancer.

}, issn = {1932-7420}, doi = {10.1016/j.cmet.2021.04.017}, author = {Tharp, Kevin M and Higuchi-Sanabria, Ryo and Timblin, Greg A and Ford, Breanna and Garzon-Coral, Carlos and Schneider, Catherine and Muncie, Jonathon M and Stashko, Connor and Daniele, Joseph R and Moore, Andrew S and Frankino, Phillip A and Homentcovschi, Stefan and Manoli, Sagar S and Shao, Hao and Richards, Alicia L and Chen, Kuei-Ho and Hoeve, Johanna Ten and Ku, Gregory M and Hellerstein, Marc and Nomura, Daniel K and Saijo, Karou and Gestwicki, Jason and Dunn, Alexander R and Krogan, Nevan J and Swaney, Danielle L and Dillin, Andrew and Weaver, Valerie M} } @article {67314, title = {Alpha-1 adrenergic receptor antagonists to prevent hyperinflammation and death from lower respiratory tract infection.}, journal = {eLife}, volume = {10}, year = {2021}, month = {2021 Jun 11}, abstract = {

In severe viral pneumonia, including Coronavirus disease 2019 (COVID-19), the viral replication phase is often followed by hyperinflammation, which can lead to acute respiratory distress syndrome, multi-organ failure, and death. We previously demonstrated that alpha-1 adrenergic receptor (⍺-AR) antagonists can prevent hyperinflammation and death in mice. Here, we conducted retrospective analyses in two cohorts of patients with acute respiratory distress (ARD, n = 18,547) and three cohorts with pneumonia (n = 400,907). Federated across two ARD cohorts, we find that patients exposed to ⍺-AR antagonists, as compared to unexposed patients, had a 34\% relative risk reduction for mechanical ventilation and death (OR = 0.70, p = 0.021). We replicated these methods on three pneumonia cohorts, all with similar effects on both outcomes. All results were robust to sensitivity analyses. These results highlight the urgent need for prospective trials testing whether prophylactic use of ⍺-AR antagonists ameliorates lower respiratory tract infection-associated hyperinflammation and death, as observed in COVID-19.

}, issn = {2050-084X}, doi = {10.7554/eLife.61700}, author = {Koenecke, Allison and Powell, Michael and Xiong, Ruoxuan and Shen, Zhu and Fischer, Nicole and Huq, Sakibul and Khalafallah, Adham M and Trevisan, Marco and Sparen, P{\"a}r and Carrero, Juan J and Nishimura, Akihiko and Caffo, Brian and Stuart, Elizabeth A and Bai, Renyuan and Staedtke, Verena and Thomas, David L and Papadopoulos, Nickolas and Kinzler, Ken W and Vogelstein, Bert and Zhou, Shibin and Bettegowda, Chetan and Konig, Maximilian F and Mensh, Brett D and Vogelstein, Joshua T and Athey, Susan} } @article {67235, title = {The anterior cingulate cortex directs exploration of alternative strategies.}, journal = {Neuron}, volume = {109}, year = {2021}, month = {2021 Jun 2}, pages = {1876-87}, abstract = {

The ability to adjust one{\textquoteright}s behavioral strategy in complex environments is at the core of cognition. Doing so efficiently requires monitoring the reliability of the ongoing strategy and, when appropriate, switching away from it to evaluate alternatives. Studies in humans and non-human primates have uncovered signals in the anterior cingulate cortex (ACC) that reflect the pressure to switch away from the ongoing strategy, whereas other ACC signals relate to the pursuit of alternatives. However, whether these signals underlie computations that actually underpin strategy switching or merely reflect tracking of related variables remains unclear. Here we provide causal evidence that the rodent ACC actively arbitrates between persisting with the ongoing behavioral strategy and temporarily switching away to re-evaluate alternatives. Furthermore, by individually perturbing distinct output pathways, we establish that the two associated computations-determining whether to switch strategy and committing to the pursuit of a specific alternative-are segregated in the ACC microcircuitry.

}, issn = {1097-4199}, doi = {10.1016/j.neuron.2021.03.028}, author = {Tervo, D Gowanlock R and Kuleshova, Elena and Manakov, Maxim and Proskurin, Mikhail and Karlsson, Mattias and Lustig, Andy and Behnam, Reza and Karpova, Alla Y} } @article {65832, title = {Automatic Detection of Synaptic Partners in a Whole-Brain Drosophila EM Dataset}, journal = {Nature Methods}, volume = {18}, year = {2021}, month = {2021 Jul 1}, pages = {771-4}, abstract = {

The study of neural circuits requires the reconstruction of neurons and the identification of synaptic connections between them. To scale the reconstruction to the size of whole-brain datasets, semi-automatic methods are needed to solve those tasks. Here, we present an automatic method for synaptic partner identification in insect brains, which uses convolutional neural networks to identify post-synaptic sites and their pre-synaptic partners. The networks can be trained from human generated point annotations alone and requires only simple post-processing to obtain final predictions. We used our method to extract 244 million putative synaptic partners in the fifty-teravoxel full adult fly brain (FAFB) electron microscopy (EM) dataset and evaluated its accuracy on 146,643 synapses from 702 neurons with a total cable length of 312 mm in four different brain regions. The predicted synaptic connections can be used together with a neuron segmentation to infer a connectivity graph with high accuracy: 96\% of edges between connected neurons are correctly classified as weakly connected (less than five synapses) and strongly connected (at least five synapses). Our synaptic partner predictions for the FAFB dataset are publicly available, together with a query library allowing automatic retrieval of up- and downstream neurons.

}, doi = {10.1038/s41592-021-01183-7}, author = {Buhmann, Julia and Sheridan, Arlo and Gerhard, Stephan and Krause, Renate and Nguyen, Tri and Heinrich, Larissa and Schlegel, Philipp and Lee, Wei-Chung Allen and Wilson, Rachel and Saalfeld, Stephan and Jefferis, Gregory and Bock, Davi and Turaga, Srinivas and Cook, Matthew and Funke, Jan} } @article {66902, title = {Brain microvasculature has a common topology with local differences in geometry that match metabolic load.}, journal = {Neuron}, volume = {109}, year = {2021}, month = {2021 April 01}, pages = {1168}, abstract = {

The microvasculature underlies the supply networks that support neuronal activity within heterogeneous brain regions. What are common versus heterogeneous aspects of the connectivity, density, and orientation of capillary networks? To address this, we imaged, reconstructed, and analyzed the microvasculature connectome in whole adult mice brains with sub-micrometer resolution. Graph analysis revealed common network topology across the brain that leads to a shared structural robustness against the rarefaction of vessels. Geometrical analysis, based on anatomically accurate reconstructions, uncovered a scaling law that links length density, i.e., the length of vessel per volume, with tissue-to-vessel distances. We then derive a formula that connects regional differences in metabolism to differences in length density and, further, predicts a common value of maximum tissue oxygen tension across the brain. Last, the orientation of capillaries is weakly anisotropic with the exception of a few strongly anisotropic regions; this variation can impact the interpretation of fMRI data.

}, issn = {1097-4199}, doi = {10.1016/j.neuron.2021.02.006}, author = {Ji, Xiang and Ferreira, Tiago and Friedman, Beth and Liu, Rui and Liechty, Hannah and Bas, Erhan and Chandrashekar, Jayaram and Kleinfeld, David} } @article {67696, title = {Campylobacter jejuni Triggers Signaling through Host Cell Focal Adhesions To Inhibit Cell Motility.}, journal = {mBio}, year = {2021}, month = {2021 Aug 24}, pages = {e0149421}, abstract = {

Campylobacter jejuni is a major foodborne pathogen that exploits the focal adhesions of intestinal cells to promote invasion and cause severe gastritis. Focal adhesions are multiprotein complexes involved in bidirectional signaling between the actin cytoskeleton and the extracellular matrix. We investigated the dynamics of focal adhesion structure and function in C. jejuni-infected cells using a comprehensive set of approaches, including confocal microscopy of live and fixed cells, immunoblotting, and superresolution interferometric photoactivated localization microscopy (iPALM). We found that C. jejuni infection of epithelial cells results in increased focal adhesion size and altered topology. These changes resulted in a persistent modulatory effect on the host cell focal adhesion, evidenced by an increase in cell adhesion strength, a decrease in individual cell motility, and a reduction in collective cell migration. We discovered that C. jejuni infection causes an increase in phosphorylation of paxillin and an alteration of paxillin turnover at the focal adhesion, which together represent a potential mechanistic basis for altered cell motility. Finally, we observed that infection of epithelial cells with the C. jejuni wild-type strain in the presence of a protein synthesis inhibitor, a C. jejuni CadF and FlpA fibronectin-binding protein mutant, or a C. jejuni flagellar export mutant blunts paxillin phosphorylation and partially reestablishes individual host cell motility and collective cell migration. These findings provide a potential mechanism for the restricted intestinal repair observed in C. jejuni-infected animals and raise the possibility that bacteria targeting extracellular matrix components can alter cell behavior after binding and internalization by manipulating focal adhesions. Campylobacter jejuni is a major foodborne pathogen that causes severe gastritis. We investigated the dynamics of focal adhesion structure and function in C. jejuni-infected epithelial cells. Focal adhesions act as signaling complexes that connect the extracellular matrix to the intracellular cytoskeleton. The key findings of this study show that C. jejuni changes the structure (size and position), composition, and function of cellular focal adhesions using a combination of virulence factors. Mechanistically, we found that the changes in focal adhesion dynamics are dependent upon the activation of host cell signaling pathways, which affect the assembly and disassembly of cellular proteins from the focal adhesion. To summarize, we have identified a new cellular phenotype in C. jejuni-infected cells that may be responsible for the restricted intestinal repair observed in C. jejuni-infected animals.

}, issn = {2150-7511}, doi = {10.1128/mBio.01494-21}, author = {Klappenbach, Courtney M and Negretti, Nicholas M and Aaron, Jesse and Chew, Teng-Leong and Konkel, Michael E} } @article {67711, title = {Campylobacter jejuni Triggers Signaling through Host Cell Focal Adhesions To Inhibit Cell Motility.}, journal = {mBio}, volume = {12}, year = {2021}, month = {2021 Aug 24}, pages = {e0149421}, abstract = {

Campylobacter jejuni is a major foodborne pathogen that exploits the focal adhesions of intestinal cells to promote invasion and cause severe gastritis. Focal adhesions are multiprotein complexes involved in bidirectional signaling between the actin cytoskeleton and the extracellular matrix. We investigated the dynamics of focal adhesion structure and function in C. jejuni-infected cells using a comprehensive set of approaches, including confocal microscopy of live and fixed cells, immunoblotting, and superresolution interferometric photoactivated localization microscopy (iPALM). We found that C. jejuni infection of epithelial cells results in increased focal adhesion size and altered topology. These changes resulted in a persistent modulatory effect on the host cell focal adhesion, evidenced by an increase in cell adhesion strength, a decrease in individual cell motility, and a reduction in collective cell migration. We discovered that C. jejuni infection causes an increase in phosphorylation of paxillin and an alteration of paxillin turnover at the focal adhesion, which together represent a potential mechanistic basis for altered cell motility. Finally, we observed that infection of epithelial cells with the C. jejuni wild-type strain in the presence of a protein synthesis inhibitor, a C. jejuni CadF and FlpA fibronectin-binding protein mutant, or a C. jejuni flagellar export mutant blunts paxillin phosphorylation and partially reestablishes individual host cell motility and collective cell migration. These findings provide a potential mechanism for the restricted intestinal repair observed in C. jejuni-infected animals and raise the possibility that bacteria targeting extracellular matrix components can alter cell behavior after binding and internalization by manipulating focal adhesions. Campylobacter jejuni is a major foodborne pathogen that causes severe gastritis. We investigated the dynamics of focal adhesion structure and function in C. jejuni-infected epithelial cells. Focal adhesions act as signaling complexes that connect the extracellular matrix to the intracellular cytoskeleton. The key findings of this study show that C. jejuni changes the structure (size and position), composition, and function of cellular focal adhesions using a combination of virulence factors. Mechanistically, we found that the changes in focal adhesion dynamics are dependent upon the activation of host cell signaling pathways, which affect the assembly and disassembly of cellular proteins from the focal adhesion. To summarize, we have identified a new cellular phenotype in C. jejuni-infected cells that may be responsible for the restricted intestinal repair observed in C. jejuni-infected animals.

}, issn = {2150-7511}, doi = {10.1128/mBio.01494-21}, author = {Klappenbach, Courtney M and Negretti, Nicholas M and Aaron, Jesse and Chew, Teng-Leong and Konkel, Michael E} } @article {65971, title = {Characterization of a common progenitor pool of the epicardium and myocardium.}, journal = {Science}, year = {2021}, month = {2021 Jan 07}, abstract = {

The mammalian heart is derived from multiple cell lineages; however, our understanding of when and how the diverse cardiac cell types arise is limited. We mapped the origin of the embryonic mouse heart at single-cell resolution using a combination of transcriptomic, imaging, and genetic lineage labeling approaches. This provided a transcriptional and anatomic definition of cardiac progenitor types. Furthermore, it revealed a cardiac progenitor pool that is anatomically and transcriptionally distinct from currently known cardiac progenitors. Besides contributing to cardiomyocytes, these cells also represent the earliest progenitor of the epicardium, a source of trophic factors and cells during cardiac development and injury. This study provides detailed insights into the formation of early cardiac cell types, with particular relevance to the development of cell-based cardiac regenerative therapies.

}, issn = {1095-9203}, doi = {10.1126/science.abb2986}, author = {Tyser, Richard C V and Ibarra-Soria, Ximena and McDole, Katie and A Jayaram, Satish and Godwin, Jonathan and van den Brand, Teun A H and Miranda, Antonio M A and Scialdone, Antonio and Keller, Philipp J and Marioni, John C and Srinivas, Shankar} } @article {67781, title = {Cre-Dependent Anterograde Transsynaptic Labeling and Functional Imaging in Zebrafish Using VSV With Reduced Cytotoxicity.}, journal = {Frontiers in Neuroanatomy}, volume = {15}, year = {2021}, month = {2021 Oct 06}, pages = {758350}, abstract = {

The small size and translucency of larval zebrafish () have made it a unique experimental system to investigate whole-brain neural circuit structure and function. Still, the connectivity patterns between most neuronal types remain mostly unknown. This gap in knowledge underscores the critical need for effective neural circuit mapping tools, especially ones that can integrate structural and functional analyses. To address this, we previously developed a vesicular stomatitis virus (VSV) based approach called Tracer with Restricted Anterograde Spread (TRAS). TRAS utilizes lentivirus to complement replication-incompetent VSV (VSVΔG) to allow restricted (monosynaptic) anterograde labeling from projection neurons to their target cells in the brain. Here, we report the second generation of TRAS (TRAS-M51R), which utilizes a mutant variant of VSVΔG [VSV(M51R)ΔG] with reduced cytotoxicity. Within the primary visual pathway, we found that TRAS-M51R significantly improved long-term viability of transsynaptic labeling (compared to TRAS) while maintaining anterograde spread activity. By using Cre-expressing VSV(M51R)ΔG, TRAS-M51R could selectively label excitatory ( positive) and inhibitory ( positive) retinorecipient neurons. We further show that these labeled excitatory and inhibitory retinorecipient neurons retained neuronal excitability upon visual stimulation at 5-8 days post fertilization (2-5 days post-infection). Together, these findings show that TRAS-M51R is suitable for neural circuit studies that integrate structural connectivity, cell-type identity, and neurophysiology.

}, issn = {1662-5129}, doi = {10.3389/fnana.2021.758350}, author = {Kler, Stanislav and Ma, Manxiu and Narayan, Sujatha and Ahrens, Misha B and Pan, Y Albert} } @article {67803, title = {De novo endocytic clathrin coats develop curvature at early stages of their formation.}, journal = {Development Cell}, volume = {56}, year = {2021}, month = {2021 Nov 22}, pages = {3146-3159.e5}, abstract = {

Sculpting a flat patch of membrane into an endocytic vesicle requires curvature generation on the cell surface, which is the primary function of the endocytosis machinery. Using super-resolved live cell fluorescence imaging, we demonstrate that curvature generation by individual clathrin-coated pits can be detected in real time within cultured cells and tissues of developing organisms. Our analyses demonstrate that the footprint of clathrin coats increases monotonically during the formation of pits at different levels of plasma membrane tension. These findings are only compatible with models that predict curvature generation at the early stages of endocytic clathrin pit formation. We also found that CALM adaptors associated with clathrin plaques form clusters, whereas AP2 distribution is more homogenous. Considering the curvature sensing and driving roles of CALM, we propose that CALM clusters may increase the strain on clathrin lattices locally, eventually giving rise to rupture and subsequent pit completion at the edges of plaques.

}, issn = {1878-1551}, doi = {10.1016/j.devcel.2021.10.019}, author = {Willy, Nathan M and Ferguson, Joshua P and Akatay, Ata and Huber, Scott and Djakbarova, Umidahan and Silahli, Salih and Cakez, Cemal and Hasan, Farah and Chang, Henry C and Travesset, Alex and Li, Siyu and Zandi, Roya and Li, Dong and Betzig, Eric and Cocucci, Emanuele and Kural, Comert} } @article {67226, title = {Deep learning enables fast and dense single-molecule localization with high accuracy}, journal = {Nature Methods}, volume = {18}, year = {2021}, month = {2021 Sep 03}, abstract = {

Single-molecule localization microscopy (SMLM) has had remarkable success in imaging cellular structures with nanometer resolution, but the need for activating only single isolated emitters limits imaging speed and labeling density. Here, we overcome this major limitation using deep learning. We developed DECODE, a computational tool that can localize single emitters at high density in 3D with highest accuracy for a large range of imaging modalities and conditions. In a public software benchmark competition, it outperformed all other fitters on 12 out of 12 data-sets when comparing both detection accuracy and localization error, often by a substantial margin. DECODE allowed us to take live-cell SMLM data with reduced light exposure in just 3 seconds and to image microtubules at ultra-high labeling density. Packaged for simple installation and use, DECODE will enable many labs to reduce imaging times and increase localization density in SMLM.Competing Interest StatementThe authors have declared no competing interest.

}, doi = {10.1101/2020.10.26.355164}, url = {https://www.biorxiv.org/content/early/2020/10/26/2020.10.26.355164}, author = {Speiser, Artur and M{\"u}ller, Lucas-Raphael and Matti, Ulf and Obara, Christopher J. and Legant, Wesley R. and Kreshuk, Anna and Macke, Jakob H. and Ries, Jonas and Turaga, Srinivas C.} } @article {65970, title = {Direct detection of SARS-CoV-2 RNA using high-contrast pH-sensitive dyes.}, journal = {Journal of Biomolecular Techniques}, volume = {32}, year = {2021}, month = {2021 Sep 01}, pages = {121-133}, abstract = {

The worldwide COVID-19 pandemic has had devastating effects on health, healthcare infrastructure, social structure, and economics. One of the limiting factors in containing the spread of this virus has been the lack of widespread availability of fast, inexpensive, and reliable methods for testing of individuals. Frequent screening for infected and often asymptomatic people is a cornerstone of pandemic management plans. Here, we introduce two pH sensitive {\textquoteleft}LAMPshade{\textquoteright} dyes as novel readouts in an isothermal RT- LAMP amplification assay for SARS-CoV-2 RNA. The resulting JaneliaLAMP (jLAMP) assay is robust, simple, inexpensive, has low technical requirements and we describe its use and performance in direct testing of contrived and clinical samples without RNA extraction.

}, doi = {https://doi.org/10.1101/2020.12.26.20248878}, author = {Timothy A. Brown and Katherine S. Schaefer and Arthur Tsang and Hyun Ah Yi and Jonathan B. Grimm and Andrew L. Lemire and Fadi M. Jradi and Charles Kim and Kevin McGowan and Kimberly Ritola and Derek T. Armstrong and Heba H. Mostafa and Wyatt Korff and Ronald D. Vale and Luke D. Lavis} } @article {67735, title = {A dominant-negative SOX18 mutant disrupts multiple regulatory layers essential to transcription factor activity.}, journal = {Nucleic Acids Research}, year = {2021}, month = {2021 Sep 27}, abstract = {

Few genetically dominant mutations involved in human disease have been fully explained at the molecular level. In cases where the mutant gene encodes a transcription factor, the dominant-negative mode of action of the mutant protein is particularly poorly understood. Here, we studied the genome-wide mechanism underlying a dominant-negative form of the SOX18 transcription factor (SOX18RaOp) responsible for both the classical mouse mutant Ragged Opossum and the human genetic disorder Hypotrichosis-lymphedema-telangiectasia-renal defect syndrome. Combining three single-molecule imaging assays in living cells together with genomics and proteomics analysis, we found that SOX18RaOp disrupts the system through an accumulation of molecular interferences which impair several functional properties of the wild-type SOX18 protein, including its target gene selection process. The dominant-negative effect is further amplified by poisoning the interactome of its wild-type counterpart, which perturbs regulatory nodes such as SOX7 and MEF2C. Our findings explain in unprecedented detail the multi-layered process that underpins the molecular aetiology of dominant-negative transcription factor function.

}, issn = {1362-4962}, doi = {10.1093/nar/gkab820}, author = {McCann, Alex J and Lou, Jieqiong and Moustaqil, Mehdi and Graus, Matthew S and Blum, Ailisa and Fontaine, Frank and Liu, Hui and Luu, Winnie and Rudolffi-Soto, Paulina and Koopman, Peter and Sierecki, Emma and Gambin, Yann and Meunier, Fr{\'e}d{\'e}ric A and Liu, Zhe and Hinde, Elizabeth and Francois, Mathias} } @article {67746, title = {A dominant-negative SOX18 mutant disrupts multiple regulatory layers essential to transcription factor activity.}, journal = {Nucleic Acids Research}, year = {2021}, month = {2021 Sep 27}, abstract = {

Few genetically dominant mutations involved in human disease have been fully explained at the molecular level. In cases where the mutant gene encodes a transcription factor, the dominant-negative mode of action of the mutant protein is particularly poorly understood. Here, we studied the genome-wide mechanism underlying a dominant-negative form of the SOX18 transcription factor (SOX18RaOp) responsible for both the classical mouse mutant Ragged Opossum and the human genetic disorder Hypotrichosis-lymphedema-telangiectasia-renal defect syndrome. Combining three single-molecule imaging assays in living cells together with genomics and proteomics analysis, we found that SOX18RaOp disrupts the system through an accumulation of molecular interferences which impair several functional properties of the wild-type SOX18 protein, including its target gene selection process. The dominant-negative effect is further amplified by poisoning the interactome of its wild-type counterpart, which perturbs regulatory nodes such as SOX7 and MEF2C. Our findings explain in unprecedented detail the multi-layered process that underpins the molecular aetiology of dominant-negative transcription factor function.

}, issn = {1362-4962}, doi = {10.1093/nar/gkab820}, author = {McCann, Alex J and Lou, Jieqiong and Moustaqil, Mehdi and Graus, Matthew S and Blum, Ailisa and Fontaine, Frank and Liu, Hui and Luu, Winnie and Rudolffi-Soto, Paulina and Koopman, Peter and Sierecki, Emma and Gambin, Yann and Meunier, Fr{\'e}d{\'e}ric A and Liu, Zhe and Hinde, Elizabeth and Francois, Mathias} } @article {67804, title = {Drosophila {\ss}-Spectrin is required in polarized ensheathing glia that form a diffusion-barrier around the neuropil.}, journal = {Nature Communications}, volume = {12}, year = {2021}, month = {2021 Nov 04}, pages = {6357}, abstract = {

In the central nervous system (CNS), functional tasks are often allocated to distinct compartments. This is also evident in the Drosophila CNS where synapses and dendrites are clustered in distinct neuropil regions. The neuropil is separated from neuronal cell bodies by ensheathing glia, which as we show using dye injection experiments, contribute to the formation of an internal diffusion barrier. We find that ensheathing glia are polarized with a basolateral plasma membrane rich in phosphatidylinositol-(3,4,5)-triphosphate (PIP) and the Na/K-ATPase Nervana2 (Nrv2) that abuts an extracellular matrix formed at neuropil-cortex interface. The apical plasma membrane is facing the neuropil and is rich in phosphatidylinositol-(4,5)-bisphosphate (PIP) that is supported by a sub-membranous {\ss}-Spectrin cytoskeleton. {\ss}-spectrin mutant larvae affect ensheathing glial cell polarity with delocalized PIP and Nrv2 and exhibit an abnormal locomotion which is similarly shown by ensheathing glia ablated larvae. Thus, polarized glia compartmentalizes the brain and is essential for proper nervous system function.

}, issn = {2041-1723}, doi = {10.1038/s41467-021-26462-x}, author = {Pogodalla, Nicole and Kranenburg, Holger and Rey, Simone and Rodrigues, Silke and Cardona, Albert and Kl{\"a}mbt, Christian} } @article {67815, title = {EASI-FISH for thick tissue defines lateral hypothalamus spatio-molecular organization.}, journal = {Cell}, volume = {184}, year = {2021}, month = {2021 Dec 01}, pages = {6361}, abstract = {

Determining the spatial organization and morphological characteristics of molecularly defined cell types is a major bottleneck for characterizing the architecture underpinning brain function. We developed Expansion-Assisted Iterative Fluorescence In Situ Hybridization (EASI-FISH) to survey gene expression in brain tissue, as well as a turnkey computational pipeline to rapidly process large EASI-FISH image datasets. EASI-FISH was optimized for thick brain sections (300\ μm) to facilitate reconstruction of spatio-molecular domains that generalize across brains. Using the EASI-FISH pipeline, we investigated the spatial distribution of dozens of molecularly defined cell types in the lateral hypothalamic area (LHA), a brain region with poorly defined anatomical organization. Mapping cell types in the LHA revealed nine spatially and molecularly defined subregions. EASI-FISH also facilitates iterative reanalysis of scRNA-seq datasets to determine marker-genes that further dissociated spatial and morphological heterogeneity. The EASI-FISH pipeline democratizes mapping molecularly defined cell types, enabling discoveries about brain organization.

}, issn = {1097-4172}, doi = {10.1016/j.cell.2021.11.024}, author = {Wang, Yuhan and Eddison, Mark and Fleishman, Greg and Weigert, Martin and Xu, Shengjin and Wang, Tim and Rokicki, Konrad and Goina, Cristian and Henry, Fredrick E and Lemire, Andrew L and Schmidt, Uwe and Yang, Hui and Svoboda, Karel and Myers, Eugene W and Saalfeld, Stephan and Korff, Wyatt and Sternson, Scott M and Tillberg, Paul W} } @article {67806, title = {ecDNA hubs drive cooperative intermolecular oncogene expression.}, journal = {Nature}, volume = {600}, year = {2021}, month = {2021 Dec 01}, pages = {731-6}, abstract = {

Extrachromosomal DNA (ecDNA) is prevalent in human cancers and mediates high expression of oncogenes through gene amplification and altered gene regulation. Gene induction typically involves cis-regulatory elements that contact and activate genes on the same chromosome. Here we show that ecDNA hubs-clusters of around 10-100 ecDNAs within the nucleus-enable intermolecular enhancer-gene interactions to promote oncogene overexpression. ecDNAs that encode multiple distinct oncogenes form hubs in diverse cancer cell types and primary tumours. Each ecDNA is more likely to transcribe the oncogene when spatially clustered with additional ecDNAs. ecDNA hubs are tethered by the bromodomain and extraterminal domain (BET) protein BRD4 in a MYC-amplified colorectal cancer cell line. The BET inhibitor JQ1 disperses ecDNA hubs and preferentially inhibits ecDNA-derived-oncogene transcription. The BRD4-bound PVT1 promoter is ectopically fused to MYC and duplicated in ecDNA, receiving promiscuous enhancer input to drive potent expression of MYC. Furthermore, the PVT1 promoter on an exogenous episome suffices to mediate gene activation in trans by ecDNA hubs in a JQ1-sensitive manner. Systematic silencing of ecDNA enhancers by CRISPR interference reveals intermolecular enhancer-gene activation among multiple oncogene loci that are amplified on distinct ecDNAs. Thus, protein-tethered ecDNA hubs enable intermolecular transcriptional regulation and may serve as units of oncogene function and cooperative evolution and as potential targets for cancer therapy.

}, issn = {1476-4687}, doi = {10.1038/s41586-021-04116-8}, author = {Hung, King L and Yost, Kathryn E and Xie, Liangqi and Shi, Quanming and Helmsauer, Konstantin and Luebeck, Jens and Sch{\"o}pflin, Robert and Lange, Joshua T and Chamorro Gonz{\'a}lez, Roc{\'\i}o and Weiser, Natasha E and Chen, Celine and Valieva, Maria E and Wong, Ivy Tsz-Lo and Wu, Sihan and Dehkordi, Siavash R and Duffy, Connor V and Kraft, Katerina and Tang, Jun and Belk, Julia A and Rose, John C and Corces, M Ryan and Granja, Jeffrey M and Li, Rui and Rajkumar, Utkrisht and Friedlein, Jordan and Bagchi, Anindya and Satpathy, Ansuman T and Tjian, Robert and Mundlos, Stefan and Bafna, Vineet and Henssen, Anton G and Mischel, Paul S and Liu, Zhe and Chang, Howard Y} } @article {67721, title = {Electrode pooling can boost the yield of extracellular recordings with switchable silicon probes.}, journal = {Nature Communications}, volume = {12}, year = {2021}, month = {2021 Sep 02}, pages = {5245}, abstract = {

State-of-the-art silicon probes for electrical recording from neurons have thousands of recording sites. However, due to volume limitations there are typically many fewer wires carrying signals off the probe, which restricts the number of channels that can be recorded simultaneously. To overcome this fundamental constraint, we propose a method called electrode pooling that uses a single wire to serve many recording sites through a set of controllable switches. Here we present the framework behind this method and an experimental strategy to support it. We then demonstrate its feasibility by implementing electrode pooling on the Neuropixels 1.0 electrode array and characterizing its effect on signal and noise. Finally we use simulations to explore the conditions under which electrode pooling saves wires without compromising the content of the recordings. We make recommendations on the design of future devices to take advantage of this strategy.

}, keywords = {Action Potentials, Animals, Brain, Electrodes, Implanted, Electrophysiology, Equipment Design, Extracellular Space, Mice, Nerve Net, Neurons, Signal Processing, Computer-Assisted, Silicon}, issn = {2041-1723}, doi = {10.1038/s41467-021-25443-4}, author = {Lee, Kyu Hyun and Ni, Yu-Li and Colonell, Jennifer and Karsh, Bill and Putzeys, Jan and Pachitariu, Marius and Harris, Timothy D and Meister, Markus} } @article {66006, title = {Endothelial junctional membrane protrusions serve as hotspots for neutrophil transmigration.}, journal = {bioRxiv}, year = {2021}, month = {2021 Jan 21}, abstract = {

Upon inflammation, leukocytes rapidly transmigrate across the endothelium to enter the inflamed tissue. Evidence accumulates that leukocytes use preferred exit sites, though it is not yet clear how these hotspots in the endothelium are defined and how they are recognized by the leukocyte. Using lattice light sheet microscopy, we discovered that leukocytes prefer endothelial membrane protrusions at cell junctions for transmigration. Phenotypically, these junctional membrane protrusions are present in an asymmetric manner, meaning that one endothelial cell shows the protrusion and the adjacent one does not. Consequently, leukocytes cross the junction by migrating underneath the protruding endothelial cell. These protrusions depend on Rac1 activity and by using a photo-activatable Rac1 probe, we could artificially generate local exit-sites for leukocytes. Overall, we have discovered a new mechanism that uses local induced junctional membrane protrusions to facilitate/steer the leukocyte escape/exit from inflamed vessel walls.

}, doi = {https://doi.org/10.1101/2021.01.18.427135}, author = {Janine J.G. Arts and Eike K. Mahlandt and Max L.B. Gr{\"o}nloh and Lilian Schimmel and Ivar Noordstra and Abraham C.I. van Steen and Simon Tol and Jos van Rijssel and Martijn A. Nolte and Marten Postma and Satya Khuon and John M. Heddleston and Eric Wait and Teng-Leong Chew and Mark Winter and Eloi Montanez and Joachim Goedhart and Jaap D. van Buul} } @article {67710, title = {Endothelial junctional membrane protrusions serve as hotspots for neutrophil transmigration.}, journal = {eLife}, volume = {10}, year = {2021}, month = {2021 Aug 25}, abstract = {

Upon inflammation, leukocytes rapidly transmigrate across the endothelium to enter the inflamed tissue. Evidence accumulates that leukocytes use preferred exit sites, though it is not yet clear how these hotspots in the endothelium are defined and how they are recognized by the leukocyte. Using lattice light sheet microscopy, we discovered that leukocytes prefer endothelial membrane protrusions at cell junctions for transmigration. Phenotypically, these junctional membrane protrusions are present in an asymmetric manner, meaning that one endothelial cell shows the protrusion and the adjacent one does not. Consequently, leukocytes cross the junction by migrating underneath the protruding endothelial cell. These protrusions depend on Rac1 activity and by using a photo-activatable Rac1 probe, we could artificially generate local exit-sites for leukocytes. Overall, we have discovered a new mechanism that uses local induced junctional membrane protrusions to facilitate/steer the leukocyte escape/exit from inflamed vessel walls.

}, issn = {2050-084X}, doi = {10.7554/eLife.66074}, author = {Arts, Janine Jg and Mahlandt, Eike K and Gr{\"o}nloh, Max and Schimmel, Lilian and Noordstra, Ivar and Gordon, Emma and van Steen, Abraham Ci and Tol, Simon and Walzog, Barbara and van Rijssel, Jos and Nolte, Martijn A and Postma, Marten and Khuon, Satya and Heddleston, John M and Wait, Eric and Chew, Teng Leong and Winter, Mark and Montanez, Eloi and Goedhart, Joachim and van Buul, Jaap D} } @article {67233, title = {ER-to-Golgi protein delivery through an interwoven, tubular network extending from ER.}, journal = {Cell}, volume = {184}, year = {2021}, month = {2021 Apr 29}, pages = {2412}, abstract = {

Cellular versatility depends on accurate trafficking of diverse proteins to their organellar destinations. For the secretory pathway (followed by approximately 30\% of all proteins), the physical nature of the vessel conducting the first portage (endoplasmic reticulum [ER] to Golgi apparatus) is unclear. We provide a dynamic 3D view of early secretory compartments in mammalian cells with isotropic resolution and precise protein localization using whole-cell, focused ion beam scanning electron microscopy with cryo-structured illumination microscopy and live-cell synchronized cargo release approaches. Rather than vesicles alone, the ER spawns an elaborate, interwoven tubular network of contiguous lipid bilayers (ER exit site) for protein export. This receptacle is capable of extending microns along microtubules while still connected to the ER by a thin neck. COPII localizes to this neck region and dynamically regulates cargo entry from the ER, while COPI acts more distally, escorting the detached, accelerating tubular entity on its way to joining the Golgi apparatus through microtubule-directed movement.

}, issn = {1097-4172}, doi = {10.1016/j.cell.2021.03.035}, author = {Weigel, Aubrey V and Chang, Chi-Lun and Shtengel, Gleb and Xu, C Shan and Hoffman, David P and Freeman, Melanie and Iyer, Nirmala and Aaron, Jesse and Khuon, Satya and Bogovic, John and Qiu, Wei and Hess, Harald F and Lippincott-Schwartz, Jennifer} } @article {66894, title = {Evaluation of multi-color genetically encoded Ca indicators in filamentous fungi.}, journal = {Fungal Genetics and Biology}, year = {2021}, month = {2021 Feb 16}, pages = {103540}, abstract = {

Genetically encoded Ca indicators (GECIs) enable long-term monitoring of cellular and subcellular dynamics of this second messenger in response to environmental and developmental cues without relying on exogenous dyes. Continued development and optimization in GECIs, combined with advances in gene manipulation, offer new opportunities for investigating the mechanism of Ca signaling in fungi, ranging from documenting Ca signatures under diverse conditions and genetic backgrounds to evaluating how changes in Ca signature impact calcium-binding proteins and subsequent cellular changes. Here, we attempted to express multi-color (green, yellow, blue, cyan, and red) circularly permuted fluorescent protein (FP)-based Ca indicators driven by multiple fungal promoters in Fusarium oxysporum, F. graminearum, and Neurospora crassa. Several variants were successfully expressed, with GCaMP5G driven by the Magnaporthe oryzae ribosomal protein 27 (P) and F. verticillioides elongation factor-1α (P) gene promoters being optimal for F. graminearum and F. oxysporum, respectively. Transformants expressing GCaMP5G were compared with those expressing YC3.60, a ratiometric Cameleon Ca indicator. Wild-type and three Ca signaling mutants of F. graminearum expressing GCaMP5G exhibited improved signal-to-noise and increased temporal and spatial resolution and are also more amenable to studies involving multiple FPs compared to strains expressing YC3.60.

}, issn = {1096-0937}, doi = {10.1016/j.fgb.2021.103540}, author = {Kim, Hye-Seon and Kim, Jung-Eun and Hwangbo, Aram and Akerboom, Jasper and Looger, Loren L and Duncan, Randall and Son, Hokyoung and Czymmek, Kirk J and Kang, Seogchan} } @article {68989, title = {Ex utero mouse embryogenesis from pre-gastrulation to late organogenesis.}, journal = {Nature}, volume = {593}, year = {2021}, month = {05/2021}, pages = {119-124}, abstract = {

The mammalian body plan is established shortly after the embryo implants into the maternal uterus, and our understanding of post-implantation developmental processes remains limited. Although pre- and peri-implantation mouse embryos are routinely cultured in vitro, approaches for the robust culture of post-implantation embryos from egg cylinder stages until advanced organogenesis remain to be established. Here we present highly effective platforms for the ex utero culture of post-implantation mouse embryos, which enable the appropriate development of embryos from before gastrulation (embryonic day (E) 5.5) until the hindlimb formation stage (E11). Late gastrulating embryos (E7.5) are grown in three-dimensional rotating bottles, whereas extended culture from pre-gastrulation stages (E5.5 or E6.5) requires a combination of static and rotating bottle culture platforms. Histological, molecular and single-cell RNA sequencing analyses confirm that the ex utero cultured embryos recapitulate in utero development precisely. This culture system is amenable to the introduction of a variety of embryonic perturbations and micro-manipulations, the results of which can be followed ex utero for up to six days. The establishment of a system for robustly growing normal mouse embryos ex utero from pre-gastrulation to advanced organogenesis represents a valuable tool for investigating embryogenesis, as it eliminates the uterine barrier and allows researchers to mechanistically interrogate post-implantation morphogenesis and artificial embryogenesis\ in mammals.

}, keywords = {Animals, Embryo Culture Techniques, Embryo, Mammalian, Embryonic Development, Female, Gastrulation, In Vitro Techniques, Male, Mice, Organogenesis, Time Factors, Uterus}, issn = {1476-4687}, doi = {10.1038/s41586-021-03416-3}, author = {Aguilera-Castrejon, Alejandro and Oldak, Bernardo and Shani, Tom and Ghanem, Nadir and Itzkovich, Chen and Slomovich, Sharon and Tarazi, Shadi and Bayerl, Jonathan and Chugaeva, Valeriya and Ayyash, Muneef and Ashouokhi, Shahd and Sheban, Daoud and Livnat, Nir and Lasman, Lior and Viukov, Sergey and Zerbib, Mirie and Addadi, Yoseph and Rais, Yoach and Cheng, Saifeng and Stelzer, Yonatan and Keren-Shaul, Hadas and Shlomo, Raanan and Massarwa, Rada and Novershtern, Noa and Maza, Itay and Hanna, Jacob H} } @article {67243, title = {Expansion-Assisted Iterative-FISH defines lateral hypothalamus spatio-molecular organization}, journal = {bioRxiv}, year = {2021}, month = {2021 Mar 8}, abstract = {

Determining the spatial organization and morphological characteristics of molecularly defined cell types is a major bottleneck for characterizing the architecture underpinning brain function. We developed\ Expansion-Assisted Iterative\ Fluorescence\ In\ Situ\ Hybridization (EASI-FISH) to survey gene expression in brain tissue, as well as a turnkey computational pipeline to rapidly process large EASI-FISH image datasets. EASI-FISH was optimized for thick brain sections (300 {\textmu}m) to facilitate reconstruction of spatio-molecular domains that generalize across brains. Using the EASI-FISH pipeline, we investigated the spatial distribution of dozens of molecularly defined cell types in the lateral hypothalamic area (LHA), a brain region with poorly defined anatomical organization. Mapping cell types in the LHA revealed nine novel spatially and molecularly defined subregions. EASI-FISH also facilitates iterative re-analysis of scRNA-Seq datasets to determine marker-genes that further dissociated spatial and morphological heterogeneity. The EASI-FISH pipeline democratizes mapping molecularly defined cell types, enabling discoveries about brain organization.

}, doi = {10.1101/2021.03.08.434304}, url = {https://doi.org/10.1101/2021.03.08.434304}, author = {Yuhan Wang and Mark Eddison and Greg Fleishman and Martin Weigert and Shengjin Xu and Frederick E. Henry and Tim Wang and Andrew L. Lemire and Uwe Schmidt and Hui Yang and Konrad Rokicki and Cristian Goina and Karel Svoboda and Eugene W. Myers and Stephan Saalfeld and Wyatt Korff and Scott M. Sternson and Paul W. Tillberg} } @article {67199, title = {Flexible scaling and persistence of social vocal communication.}, journal = {Nature}, volume = {593}, year = {2021}, month = {2021 May 01}, pages = {108-13}, abstract = {

Innate vocal sounds such as laughing, screaming or crying convey one{\textquoteright}s feelings to others. In many species, including humans, scaling the amplitude and duration of vocalizations is essential for effective social communication. In mice, female scent triggers male mice to emit innate courtship ultrasonic vocalizations (USVs). However, whether mice flexibly scale their vocalizations and how neural circuits are structured to generate flexibility remain largely unknown. Here we identify mouse neurons from the lateral preoptic area (LPOA) that express oestrogen receptor 1 (LPOA neurons) and, when activated, elicit the complete repertoire of USV syllables emitted during natural courtship. Neural anatomy and functional data reveal a two-step, di-synaptic circuit motif in which primary long-range inhibitory LPOA neurons relieve a clamp of local periaqueductal grey (PAG) inhibition, enabling excitatory PAG USV-gating neurons to trigger vocalizations. We find that social context shapes a wide range of USV amplitudes and bout durations. This variability is absent when PAG neurons are stimulated directly; PAG-evoked vocalizations are time-locked to neural activity and stereotypically loud. By contrast, increasing the activity of LPOA neurons scales the amplitude of vocalizations, and delaying the recovery of the inhibition clamp prolongs USV bouts. Thus, the LPOA disinhibition motif contributes to flexible loudness and the duration and persistence of bouts, which are key aspects of effective vocal social communication.

}, issn = {1476-4687}, doi = {10.1038/s41586-021-03403-8}, author = {Chen, Jingyi and Markowitz, Jeffrey E and Lilascharoen, Varoth and Taylor, Sandra and Sheurpukdi, Pete and Keller, Jason A and Jensen, Jennifer R and Lim, Byung Kook and Datta, Sandeep Robert and Stowers, Lisa} } @article {67805, title = {Functionally distinct roles for eEF2K in the control of ribosome availability and p-body abundance.}, journal = {Nature Communications}, volume = {12}, year = {2021}, month = {2021 Nov 23}, pages = {6789}, abstract = {

Processing bodies (p-bodies) are a prototypical phase-separated RNA-containing granule. Their abundance is highly dynamic and has been linked to translation. Yet, the molecular mechanisms responsible for coordinate control of the two processes are unclear. Here, we uncover key roles for eEF2 kinase (eEF2K) in the control of ribosome availability and p-body abundance. eEF2K acts on a sole known substrate, eEF2, to inhibit translation. We find that the eEF2K agonist nelfinavir abolishes p-bodies in sensory neurons and impairs translation. To probe the latter, we used cryo-electron microscopy. Nelfinavir stabilizes vacant 80S ribosomes. They contain SERBP1 in place of mRNA and eEF2 in the acceptor site. Phosphorylated eEF2 associates with inactive ribosomes that resist splitting in vitro. Collectively, the data suggest that eEF2K defines a population of inactive ribosomes resistant to recycling and protected from degradation. Thus, eEF2K activity is central to both p-body abundance and ribosome availability in sensory neurons.

}, issn = {2041-1723}, doi = {10.1038/s41467-021-27160-4}, author = {Smith, Patrick R and Loerch, Sarah and Kunder, Nikesh and Stanowick, Alexander D and Lou, Tzu-Fang and Campbell, Zachary T} } @article {67816, title = {A genetically encoded fluorescent biosensor for extracellular L-lactate.}, journal = {Nature Communications}, volume = {12}, year = {2021}, month = {2021 Dec 06}, pages = {7058}, abstract = {

L-Lactate, traditionally considered a metabolic waste product, is increasingly recognized as an important intercellular energy currency in mammals. To enable investigations of the emerging roles of intercellular shuttling of L-lactate, we now report an intensiometric green fluorescent genetically encoded biosensor for extracellular L-lactate. This biosensor, designated eLACCO1.1, enables cellular resolution imaging of extracellular L-lactate in cultured mammalian cells and brain tissue.

}, issn = {2041-1723}, doi = {10.1038/s41467-021-27332-2}, author = {Nasu, Yusuke and Murphy-Royal, Ciaran and Wen, Yurong and Haidey, Jordan N and Molina, Rosana S and Abhi Aggarwal and Zhang, Shuce and Kamijo, Yuki and Paquet, Marie-Eve and Podgorski, Kaspar and Drobizhev, Mikhail and Bains, Jaideep S and Lemieux, M Joanne and Gordon, Grant R and Campbell, Robert E} } @article {67211, title = {High-fidelity estimates of spikes and subthreshold waveforms from 1-photon voltage imaging in~vivo.}, journal = {Cell Reports}, volume = {35}, year = {2021}, month = {2021 Apr 06}, pages = {108954}, abstract = {

The ability to probe the membrane potential of multiple genetically defined neurons simultaneously would have a profound impact on neuroscience research. Genetically encoded voltage indicators are a promising tool for this purpose, and recent developments have achieved a high signal-to-noise ratio in\ vivo with 1-photon fluorescence imaging. However, these recordings exhibit several sources of noise and signal extraction remains a challenge. We present an improved signal extraction pipeline, spike-guided penalized matrix decomposition-nonnegative matrix factorization (SGPMD-NMF), which resolves supra- and subthreshold voltages in\ vivo. The method incorporates biophysical and optical constraints. We validate the pipeline with simultaneous patch-clamp and optical recordings from mouse layer 1 in\ vivo and with simulated and composite datasets with realistic noise. We demonstrate applications to mouse hippocampus expressing paQuasAr3-s or SomArchon1, mouse cortex expressing SomArchon1 or Voltron, and zebrafish spines expressing zArchon1.

}, issn = {2211-1247}, doi = {10.1016/j.celrep.2021.108954}, author = {Xie, Michael E and Adam, Yoav and Fan, Linlin Z and B{\"o}hm, Urs L and Kinsella, Ian and Zhou, Ding and Rozsa, Marton and Singh, Amrita and Svoboda, Karel and Paninski, Liam and Cohen, Adam E} } @article {65999, title = {A high-throughput predictive method for sequence-similar fold switchers.}, journal = {Biopolymers}, volume = {112}, year = {2021}, month = {2021 Oct 01}, pages = {e23416}, abstract = {

Although most experimentally characterized proteins with similar sequences assume the same folds and perform similar functions, an increasing number of exceptions is emerging. One class of exceptions comprises sequence-similar fold switchers, whose secondary structures shift from α-helix \<-\> β-sheet through a small number of mutations, a sequence insertion, or a deletion. Predictive methods for identifying sequence-similar fold switchers are desirable because some are associated with disease and/or can perform different functions in cells. Here, we use homology-based secondary structure predictions to identify sequence-similar fold switchers from their amino acid sequences alone. To do this, we predicted the secondary structures of sequence-similar fold switchers using three different homology-based secondary structure predictors: PSIPRED, JPred4, and SPIDER3. We found that α-helix \<-\> β-strand prediction discrepancies from JPred4 discriminated between the different conformations of sequence-similar fold switchers with high statistical significance (P \< 1.8*10 ). Thus, we used these discrepancies as a classifier and found that they can often robustly discriminate between sequence-similar fold switchers and sequence-similar proteins that maintain the same folds (Matthews Correlation Coefficient of 0.82). We found that JPred4 is a more robust predictor of sequence-similar fold switchers because of (a) the curated sequence database it uses to produce multiple sequence alignments and (b) its use of sequence profiles based on Hidden Markov Models. Our results indicate that inconsistencies between JPred4 secondary structure predictions can be used to identify some sequence-similar fold switchers from their sequences alone. Thus, the negative information from inconsistent secondary structure predictions can potentially be leveraged to identify sequence-similar fold switchers from the broad base of genomic sequences.

}, issn = {1097-0282}, doi = {10.1002/bip.23416}, author = {Kim, Allen K and Looger, Loren L and Porter, Lauren L} } @article {67263, title = {Hunger or thirst state uncertainty is resolved by outcome evaluation in medial prefrontal cortex to guide decision-making.}, journal = {Nature Neuroscience}, volume = {24}, year = {2021}, month = {2021 Jul 01}, pages = {907-912}, abstract = {

Physiological need states direct decision-making toward re-establishing homeostasis. Using a two-alternative forced choice task for mice that models elements of human decisions, we found that varying hunger and thirst states caused need-inappropriate choices, such as food seeking when thirsty. These results show limits on interoceptive knowledge of hunger and thirst states to guide decision-making. Instead, need states were identified after food and water consumption by outcome evaluation, which depended on the medial prefrontal cortex.

}, issn = {1546-1726}, doi = {10.1038/s41593-021-00850-4}, author = {Eiselt, Anne-Kathrin and Chen, Susu and Chen, Jim and Arnold, Jon and Kim, Tahnbee and Pachitariu, Marius and Sternson, Scott M} } @article {66002, title = {Image-based pooled whole-genome CRISPRi screening for subcellular phenotypes.}, journal = {Journal of Cell Biology}, volume = {220}, year = {2021}, month = {2021 Feb 01}, abstract = {

Genome-wide CRISPR screens have transformed our ability to systematically interrogate human gene function, but are currently limited to a subset of cellular phenotypes. We report a novel pooled screening approach for a wider range of cellular and subtle subcellular phenotypes. Machine learning and convolutional neural network models are trained on the subcellular phenotype to be queried. Genome-wide screening then utilizes cells stably expressing dCas9-KRAB (CRISPRi), photoactivatable fluorescent protein (PA-mCherry), and a lentiviral guide RNA (gRNA) pool. Cells are screened by using microscopy and classified by artificial intelligence (AI) algorithms, which precisely identify the genetically altered phenotype. Cells with the phenotype of interest are photoactivated and isolated via flow cytometry, and the gRNAs are identified by sequencing. A proof-of-concept screen accurately identified PINK1 as essential for Parkin recruitment to mitochondria. A genome-wide screen identified factors mediating TFEB relocation from the nucleus to the cytosol upon prolonged starvation. Twenty-one of the 64 hits called by the neural network model were independently validated, revealing new effectors of TFEB subcellular localization. This approach, AI-photoswitchable screening (AI-PS), offers a novel screening platform capable of classifying a broad range of mammalian subcellular morphologies, an approach largely unattainable with current methodologies at genome-wide scale.

}, issn = {1540-8140}, doi = {10.1083/jcb.202006180}, author = {Kanfer, Gil and Sarraf, Shireen A and Maman, Yaakov and Baldwin, Heather and Dominguez-Martin, Eunice and Johnson, Kory R and Ward, Michael E and Kampmann, Martin and Lippincott-Schwartz, Jennifer and Youle, Richard J} } @article {67871, title = {Image-based representation of massive spatial transcriptomics datasets.}, journal = {bioRxiv}, year = {2021}, month = {2021 Dec 07}, abstract = {

We present STIM, an imaging-based computational framework for exploring, visualizing, and processing high-throughput spatial sequencing datasets. STIM is built on the powerful ImgLib2, N5 and BigDataViewer (BDV) frameworks enabling transfer of computer vision techniques to datasets with irregular measurement-spacing and arbitrary spatial resolution, such as spatial transcriptomics data generated by multiplexed targeted hybridization or spatial sequencing technologies. We illustrate STIM{\textquoteright}s capabilities by representing, visualizing, and automatically registering publicly available spatial sequencing data from 14 serial sections of mouse brain tissue.

}, doi = {10.1101/2021.12.07.471629}, url = {https://doi.org/10.1101/2021.12.07.471629}, author = {Stephan Preibisch and Nikos Karaiskos and Nikolaus Rajewsky} } @article {67670, title = {Imaging Africa: a strategic approach to optical microscopy training in Africa.}, journal = {Nature Methods}, volume = {18}, year = {2021}, month = {2021 Aug 20}, pages = {847-855}, issn = {1548-7105}, doi = {10.1038/s41592-021-01227-y}, author = {Reiche, Michael A and Warner, Digby F and Aaron, Jesse and Khuon, Satya and Fletcher, Daniel A and Hahn, Klaus and Rogers, Kelly L and Mhlanga, Musa and Koch, Anastasia and Quaye, Wendye and Chew, Teng-Leong} } @article {67627, title = {In~vivo glucose imaging in multiple model organisms with an engineered single-wavelength sensor.}, journal = {Cell Reports}, volume = {35}, year = {2021}, month = {2021 Jun 22}, pages = {109284}, abstract = {

Glucose is arguably the most important molecule in metabolism, and its dysregulation underlies diabetes. We describe a family of single-wavelength genetically encoded glucose sensors with a high signal-to-noise ratio, fast kinetics, and affinities varying over four orders of magnitude (1\ μM to 10\ mM). The sensors allow mechanistic characterization of glucose transporters expressed in cultured cells with high spatial and temporal resolution. Imaging of neuron/glia co-cultures revealed \~{}3-fold faster glucose changes in astrocytes. In larval Drosophila central nervous system explants, intracellular neuronal glucose fluxes suggested a rostro-caudal transport pathway in the ventral nerve cord neuropil. In zebrafish, expected glucose-related physiological sequelae of insulin and epinephrine treatments were directly visualized. Additionally, spontaneous muscle twitches induced glucose uptake in muscle, and sensory and pharmacological perturbations produced large changes in the brain. These sensors will enable rapid, high-resolution imaging of glucose influx, efflux, and metabolism in behaving animals.

}, issn = {2211-1247}, doi = {10.1016/j.celrep.2021.109284}, author = {Keller, Jacob P and Marvin, Jonathan S and Lacin, Haluk and Lemon, William C and Shea, Jamien and Kim, Soomin and Lee, Richard T and Koyama, Minoru and Keller, Philipp J and Looger, Loren L} } @article {67656, title = {Intercellular Arc Signaling Regulates Vasodilation.}, journal = {Journal of Neuroscience}, year = {2021}, month = {2021 Jul 29}, abstract = {

Injury responses require communication between different cell types in the skin. Sensory neurons contribute to inflammation and can secrete signaling molecules that affect non-neuronal cells. Despite the pervasive role of translational regulation in nociception, the contribution of activity-dependent protein synthesis to inflammation is not well understood. To address this problem, we examined the landscape of nascent translation in murine dorsal root ganglion (DRG) neurons treated with inflammatory mediators using ribosome profiling. We identified the activity-dependent gene, Arc, as a target of translation and Inflammatory cues promote local translation of Arc in the skin. Arc-deficient male mice display exaggerated paw temperatures and vasodilation in response to an inflammatory challenge. Since Arc has recently been shown to be released from neurons in extracellular vesicles (EVs), we hypothesized that intercellular Arc signaling regulates the inflammatory response in skin. We found that the excessive thermal responses and vasodilation observed in Arc defective mice are rescued by injection of Arc-containing EVs into the skin. Our findings suggest that activity-dependent production of Arc in afferent fibers regulates neurogenic inflammation potentially through intercellular signaling.Nociceptors play prominent roles in pain and inflammation. We examined rapid changes in the landscape of nascent translation in cultured dorsal root ganglia (DRGs) treated with a combination of inflammatory mediators using ribosome profiling. We identified several hundred transcripts subject to rapid preferential translation. Among them is the immediate early gene (IEG) Arc. We provide evidence that Arc is translated in afferent fibers in the skin. Arc-deficient mice display several signs of exaggerated inflammation which is normalized on injection of Arc containing extracellular vesicles (EVs). Our work suggests that noxious cues can trigger Arc production by nociceptors which in turn constrains neurogenic inflammation in the skin.

}, issn = {1529-2401}, doi = {10.1523/JNEUROSCI.0440-21.2021}, author = {de la Pe{\~n}a, June Bryan and Barragan-Iglesias, Paulino and Lou, Tzu-Fang and Kunder, Nikesh and Loerch, Sarah and Shukla, Tarjani and Basavarajappa, Lokesh and Song, Jane and James, Dominique N and Megat, Salim and Moy, Jamie K and Wanghzou, Andi and Ray, Pradipta R and Hoyt, Kenneth and Steward, Oswald and Price, Theodore J and Shepherd, Jason and Campbell, Zachary T} } @article {67672, title = {jumps with greater velocity and acceleration than previously reported.}, journal = {MicroPublication Biology}, volume = {2021}, year = {2021}, month = {2021 Aug 02}, abstract = {

Infective juveniles of the insect-parastic nematode canjump greater than 6 times their height, a striking evolved novelty in some species of this genus. Using high-speed videography, we observed the kinematics of spontaneousjumping behavior. Our analysis places a lower bound on the velocity and acceleration of these worms.

}, issn = {2578-9430}, doi = {10.17912/micropub.biology.000435}, author = {Dillman, Adler R and Korff, Wyatt and Dickinson, Michael H and Sternberg, Paul W} } @article {66021, title = {The level of oncogenic Ras determines the malignant transformation of Lkb1 mutant tissue in vivo.}, journal = {Communications Biology}, volume = {4}, year = {2021}, month = {2021 Jan 29}, pages = {142}, abstract = {

The genetic and metabolic heterogeneity of RAS-driven cancers has confounded therapeutic strategies in the clinic. To address this, rapid and genetically tractable animal models are needed that recapitulate the heterogeneity of RAS-driven cancers in vivo. Here, we generate a Drosophila melanogaster model of Ras/Lkb1 mutant carcinoma. We show that low-level expression of oncogenic Ras (Ras) promotes the survival of Lkb1 mutant tissue, but results in autonomous cell cycle arrest and non-autonomous overgrowth of wild-type tissue. In contrast, high-level expression of oncogenic Ras (Ras) transforms Lkb1 mutant tissue resulting in lethal malignant tumors. Using simultaneous multiview light-sheet microcopy, we have characterized invasion phenotypes of Ras/Lkb1 tumors in living larvae. Our molecular analysis reveals sustained activation of the AMPK pathway in malignant Ras/Lkb1 tumors, and demonstrate the genetic and pharmacologic dependence of these tumors on CaMK-activated Ampk. We further show that LKB1 mutant human lung adenocarcinoma patients with high levels of oncogenic KRAS exhibit worse overall survival and increased AMPK activation. Our results suggest that high levels of oncogenic KRAS is a driving event in the malignant transformation of LKB1 mutant tissue, and uncovers a vulnerability that may be used to target this aggressive genetic subset of RAS-driven tumors.

}, issn = {2399-3642}, doi = {10.1038/s42003-021-01663-8}, author = {Rackley, Briana and Seong, Chang-Soo and Kiely, Evan and Parker, Rebecca E and Rupji, Manali and Dwivedi, Bhakti and Heddleston, John M and Giang, William and Anthony, Neil and Chew, Teng-Leong and Gilbert-Ross, Melissa} } @article {67271, title = {Lupus susceptibility region containing CDKN1B rs34330 mechanistically influences expression and function of multiple target genes, also linked to proliferation and apoptosis.}, journal = {Arthritis Rheumatology}, volume = {73}, year = {2021}, month = {2021 Dec 01}, pages = {2303-13}, abstract = {

OBJECTIVE: A recent genome-wide association study (GWAS) reported a significant genetic association between rs34330 of cyclin-dependent kinase inhibitor 1B (CDKN1B) and risk of systemic lupus erythematosus (SLE) in Han Chinese. This study aims to validate the reported association and elucidate the biochemical mechanisms underlying the variant{\textquoteright}s effect.

METHODS: We performed allelic association with SLE followed by meta-analysis across 11 independent cohorts (n=28,872). We applied in silico bioinformatics and experimental validation in SLE-relevant cell lines to determine the functional consequences of rs34330.

RESULTS: We replicated genetic association between SLE and rs34330 (P =5.29x10 , OR (95\% CI)=0.84 (0.81-0.87)). Follow-up bioinformatics and eQTL analysis suggest that rs34330 is located in active chromatin and potentially regulates several target genes. Using luciferase and ChIP-qPCR, we demonstrated substantial allele-specific promoter and enhancer activity, and allele-specific binding of three histone marks (H3K27ac, H3K4me3, H3K4me1), RNA pol II, CTCF, and a critical immune transcription factor (IRF-1). Chromosome conformation capture (3C) detected long-range chromatin interactions between rs34330 and the promoters of neighboring genes APOLD1 and DDX47, and effects on CDKN1B and the other target genes were directly validated by CRISPR-based genome editing. Finally, CRISPR-dCas9-based epigenetic activation/silencing confirmed these results. Gene-edited cell lines also showed higher levels of proliferation and apoptosis.

CONCLUSION: Collectively, these findings suggest a mechanism whereby the rs34330 risk allele (C) influences the presence of histone marks, RNA pol II, and the IRF-1 transcription factor to regulate expression of several target genes linked to proliferation and apoptosis, which potentially underlie the association of rs34330 with SLE.

}, issn = {2326-5205}, doi = {10.1002/art.41799}, author = {Singh, Bhupinder and Maiti, Guru P and Zhou, Xujie and Fazel-Najafabadi, Mehdi and Bae, Sang-Cheol and Sun, Celi and Terao, Chikashi and Okada, Yukinori and Chua, Kek Heng and Kochi, Yuta and Guthridge, Joel M and Zhang, Hong and Weirauch, Matthew and James, Judith A and Harley, John B and Varshney, Gaurav K and Looger, Loren L and Nath, Swapan K} } @article {67695, title = {Method for Rapid Enzymatic Cleaning for Reuse of Patch Clamp Pipettes: Increasing Throughput by Eliminating Manual Pipette Replacement between Patch Clamp Attempts.}, journal = {Bio Protocol}, volume = {11}, year = {2021}, month = {2021 Jul 20}, pages = {e4085}, abstract = {

The whole-cell patch-clamp method is a gold standard for single-cell analysis of electrical activity, cellular morphology, and gene expression. Prior to our discovery that patch-clamp pipettes could be cleaned and reused, experimental throughput and automation were limited by the need to replace pipettes manually after each experiment. This article presents an optimized protocol for pipette cleaning, which enables it to be performed quickly (\< 30 s), resulting in a high yield of whole-cell recording success rate (\> 90\%) for over 100 reuses of a single pipette. For most patch-clamp experiments (\< 30 whole-cell recordings per day), this method enables a single pipette to be used for an entire day of experiments. In addition, we describe easily implementable hardware and software as well as troubleshooting tips to help other labs implement this method in their own experiments. Pipette cleaning enables patch-clamp experiments to be performed with higher throughput, whether manually or in an automated fashion, by eliminating the tedious and skillful task of replacing pipettes. From our experience with numerous electrophysiology laboratories, pipette cleaning can be integrated into existing patch-clamp setups in approximately one day using the hardware and software described in this article. Graphic abstract: Rapid enzymatic cleaning for reuse of patch-clamp pipettes.

}, issn = {2331-8325}, doi = {10.21769/BioProtoc.4085}, author = {Landry, Corey R and Yip, Mighten C and Kolb, Ilya and Stoy, William A and Gonzalez, Mercedes M and Forest, Craig R} } @article {67305, title = {Micro-Meta App: an interactive software tool to facilitate the collection of microscopy metadata based on community-driven specifications}, journal = {bioRxiv}, year = {2021}, month = {2021 May 31}, abstract = {

For the information content of microscopy images to be appropriately interpreted, reproduced, and meet FAIR (Findable Accessible Interoperable and Reusable) principles, they should be accompanied by detailed descriptions of microscope hardware, image acquisition settings, image pixel and dimensional structure, and instrument performance. Nonetheless, the thorough documentation of imaging experiments is significantly impaired by the lack of community-sanctioned easy-to-use software tools to facilitate the extraction and collection of relevant microscopy metadata. Here we present\ Micro-Meta App, an intuitive open-source software designed to tackle these issues that was developed in the context of nascent global bioimaging community organizations, including\ BioImaging\ North\ America (BINA) and\ QUAlity Assessment and\ REProducibility in\ Light\ Microscopy (QUAREP-LiMi), whose goal is to improve reproducibility, data quality and sharing value for imaging experiments. The App provides a user-friendly interface for building comprehensive descriptions of the conditions utilized to produce individual microscopy datasets as specified by the recently proposed 4DN-BINA-OME tiered-system of Microscopy Metadata model. To achieve this goal the App provides a visual guide for a microscope-user to: 1) interactively build diagrammatic representations of hardware configurations of given microscopes that can be easily reused and shared with colleagues needing to document similar instruments. 2) Automatically extracts relevant metadata from image files and facilitates the collection of missing image acquisition settings and calibration metrics associated with a given experiment. 3) Output all collected Microscopy Metadata to interoperable files that can be used for documenting imaging experiments and shared with the community. In addition to significantly lowering the burden of quality assurance, the visual nature of Micro-Meta App makes it particularly suited for training users that have limited knowledge of the intricacies of light microscopy experiments. To ensure wide-adoption by microscope-users with different needs Micro-Meta App closely interoperates with\ MethodsJ2\ and\ OMERO.mde, two complementary tools described in parallel manuscripts.

}, author = {Alex Rigano and Shannon Ehmsen and Serkan Utku Ozturk and Joel Ryan and Alexander Balashov and Mathias Hammer and Koray Kirli and Karl Bellve and Ulrike Boehm and Claire M. Brown and James J. Chambers and Robert A. Coleman and Andrea Cosolo and Orestis Faklaris and Kevin Fogarty and Thomas Guilbert and Anna B. Hamacher and Michelle S. Itano and Daniel P. Keeley and Susanne Kunis and Judith Lacoste and Alex Laude and Willa Ma and Marco Marcello and Paula Montero-Llopis and Glyn Nelson and Roland Nitschke and Jaime A. Pimentel and Stefanie Weidtkamp-Peters and Peter J. Park and Burak Alver and David Grunwald and Caterina Strambio-De-Castillia} } @article {67811, title = {Micro-Meta App: an interactive tool for collecting microscopy metadata based on community specifications.}, journal = {Nature Methods}, volume = {18}, year = {2021}, month = {2021 Dec 03}, pages = {1489-1495}, abstract = {

For quality, interpretation, reproducibility and sharing value, microscopy images should be accompanied by detailed descriptions of the conditions that were used to produce them. Micro-Meta App is an intuitive, highly interoperable, open-source software tool that was developed in the context of the 4D Nucleome (4DN) consortium and is designed to facilitate the extraction and collection of relevant microscopy metadata as specified by the recent 4DN-BINA-OME tiered-system of Microscopy Metadata specifications. In addition to substantially lowering the burden of quality assurance, the visual nature of Micro-Meta App makes it particularly suited for training purposes.

}, issn = {1548-7105}, doi = {10.1038/s41592-021-01315-z}, author = {Rigano, Alessandro and Ehmsen, Shannon and {\"O}zt{\"u}rk, Serkan Utku and Ryan, Joel and Balashov, Alexander and Hammer, Mathias and Kirli, Koray and Boehm, Ulrike and Brown, Claire M and Bellve, Karl and Chambers, James J and Cosolo, Andrea and Coleman, Robert A and Faklaris, Orestis and Fogarty, Kevin E and Guilbert, Thomas and Hamacher, Anna B and Itano, Michelle S and Keeley, Daniel P and Kunis, Susanne and Lacoste, Judith and Laude, Alex and Ma, Willa Y and Marcello, Marco and Montero-Llopis, Paula and Nelson, Glyn and Nitschke, Roland and Pimentel, Jaime A and Weidtkamp-Peters, Stefanie and Park, Peter J and Alver, Burak H and Grunwald, David and Strambio-De-Castillia, Caterina} } @article {68358, title = {Molecular, anatomical, and functional organization of lung interoceptors}, journal = {bioRxiv}, year = {2021}, month = {11/2021}, abstract = {

Interoceptors, sensory neurons that monitor internal organs and states, are essential for physiological homeostasis and generating internal perceptions. Here we describe a comprehensive transcriptomic atlas of interoceptors of the mouse lung, defining 10 molecular subtypes that differ in developmental origin, myelination, receptive fields, terminal morphologies, and cell contacts. Each subtype expresses a unique but overlapping combination of sensory receptors that detect diverse physiological and pathological stimuli, and each can signal to distinct sets of lung cells including immune cells, forming a local neuroimmune interaction network. Functional interrogation of two mechanosensory subtypes reveals exquisitely-specific homeostatic roles in breathing, one regulating inspiratory time and the other inspiratory flow. The results suggest that lung interoceptors encode diverse and dynamic sensory information rivaling that of canonical exteroceptors, and this information is used to drive myriad local cellular interactions and enable precision control of breathing, while providing only vague perceptions of organ states.Competing Interest StatementThe authors have declared no competing interest.

}, doi = {10.1101/2021.11.10.468116}, url = {https://www.biorxiv.org/content/early/2021/11/13/2021.11.10.468116}, author = {Liu, Yin and Diaz de Arce, Alex J. and Krasnow, Mark A.} } @article {67648, title = {Molecular characterization of projection neuron subtypes in the mouse olfactory bulb.}, journal = {eLife}, volume = {10}, year = {2021}, month = {2021 Jul 22}, abstract = {

Projection neurons (PNs) in the mammalian olfactory bulb (OB) receive input from the nose and project to diverse cortical and subcortical areas. Morphological and physiological studies have highlighted functional heterogeneity, yet no molecular markers have been described that delineate PN subtypes. Here, we used viral injections into olfactory cortex and fluorescent nucleus sorting to enrich PNs for high-throughput single nucleus and bulk RNA deep sequencing. Transcriptome analysis and RNA hybridization identified distinct mitral and tufted cell populations with characteristic transcription factor network topology, cell adhesion and excitability-related gene expression. Finally, we describe a new computational approach for integrating bulk and snRNA-seq data, and provide evidence that different mitral cell populations preferentially project to different target regions. Together, we have identified potential molecular and gene regulatory mechanisms underlying PN diversity and provide new molecular entry points into studying the diverse functional roles of mitral and tufted cell subtypes.

}, issn = {2050-084X}, doi = {10.7554/eLife.65445}, author = {Zeppilli, Sara and Ackels, Tobias and Attey, Robin and Klimpert, Nell and Kimberly Ritola, Dr and Boeing, Stefan and Crombach, Anton and Schaefer, Andreas T and Fleischmann, Alexander} } @article {69336, title = {Molecular profiling of single neurons of known identity in two ganglia from the crab Cancer borealis}, journal = {Proceedings of the National Academy of Sciences}, volume = {116}, year = {2021}, month = {2021 Feb 12}, pages = {26980 - 26990}, abstract = {

Understanding circuit organization depends on identification of cell types. Recent advances in transcriptional profiling methods have enabled classification of cell types by their gene expression. While exceptionally powerful and high throughput, the ground-truth validation of these methods is difficult: If cell type is unknown, how does one assess whether a given analysis accurately captures neuronal identity? To shed light on the capabilities and limitations of solely using transcriptional profiling for cell-type classification, we performed 2 forms of transcriptional profiling{\textemdash}RNA-seq and quantitative RT-PCR, in single, unambiguously identified neurons from 2 small crustacean neuronal networks: The stomatogastric and cardiac ganglia. We then combined our knowledge of cell type with unbiased clustering analyses and supervised machine learning to determine how accurately functionally defined neuron types can be classified by expression profile alone. The results demonstrate that expression profile is able to capture neuronal identity most accurately when combined with multimodal information that allows for post hoc grouping, so analysis can proceed from a supervised perspective. Solely unsupervised clustering can lead to misidentification and an inability to distinguish between 2 or more cell types. Therefore, this study supports the general utility of cell identification by transcriptional profiling, but adds a caution: It is difficult or impossible to know under what conditions transcriptional profiling alone is capable of assigning cell identity. Only by combining multiple modalities of information such as physiology, morphology, or innervation target can neuronal identity be unambiguously determined.

}, issn = {0027-8424}, doi = {10.1073/pnas.1911413116}, url = {https://pnas.org/doi/full/10.1073/pnas.1911413116https://pnas.org/doi/pdf/10.1073/pnas.1911413116http://www.pnas.org/syndication/doi/10.1073/pnas.1911413116}, author = {Northcutt, Adam J. and Kick, Daniel R. and Otopalik, Adriane G. and Goetz, Benjamin M. and Harris, Rayna M. and Santin, Joseph M. and Hofmann, Hans A. and Marder, Eve and Schulz, David J.} } @article {65969, title = {Neuromolecular and behavioral effects of ethanol deprivation in Drosophila}, journal = {bioRxiv}, year = {2021}, month = {2021 Jan 04}, abstract = {

Alcohol use disorder (AUD) is characterized by loss of control in limiting alcohol intake. This may involve intermittent periods of abstinence followed by alcohol seeking and, consequently, relapse. However, little is understood of the molecular mechanisms underlying the impact of alcohol deprivation on behavior. Using a new\ Drosophila melanogaster\ repeated intermittent alcohol exposure model, we sought to identify how ethanol deprivation alters spontaneous behavior, determine the associated neural structures, and reveal correlated changes in brain gene expression. We found that repeated intermittent ethanol-odor exposures followed by ethanol-deprivation dynamically induces behaviors associated with a negative affect state. Although behavioral states broadly mapped to many brain regions, persistent changes in social behaviors mapped to the mushroom body and surrounding neuropil. This occurred concurrently with changes in expression of genes associated with sensory responses, neural plasticity, and immunity. Like social behaviors, immune response genes were upregulated following three-day repeated intermittent ethanol-odor exposures and persisted with one or two days of ethanol-deprivation, suggesting an enduring change in molecular function. Our study provides a framework for identifying how ethanol deprivation alters behavior with correlated underlying circuit and molecular changes.

}, doi = {https://doi.org/10.1101/2021.01.02.425101}, author = {Natalie M. D{\textquoteright}Silva and Katie S. McCullar and Ashley M. Conard and Tyler Blackwater and Reza Azanchi and Ulrike Heberlein and Erica Larschan and Karla R. Kaun} } @article {67232, title = {Neuropixels 2.0: A miniaturized high-density probe for stable, long-term brain recordings.}, journal = {Science}, volume = {372}, year = {2021}, month = {2021 Apr 16}, abstract = {

Measuring the dynamics of neural processing across time scales requires following the spiking of thousands of individual neurons over milliseconds and months. To address this need, we introduce the Neuropixels 2.0 probe together with newly designed analysis algorithms. The probe has more than 5000 sites and is miniaturized to facilitate chronic implants in small mammals and recording during unrestrained behavior. High-quality recordings over long time scales were reliably obtained in mice and rats in six laboratories. Improved site density and arrangement combined with newly created data processing methods enable automatic post hoc correction for brain movements, allowing recording from the same neurons for more than 2 months. These probes and algorithms enable stable recordings from thousands of sites during free behavior, even in small animals such as mice.

}, issn = {1095-9203}, doi = {10.1126/science.abf4588}, author = {Steinmetz, Nicholas A and Ayd{\i}n, {\c C}a{\u g}atay and Lebedeva, Anna and Okun, Michael and Pachitariu, Marius and Bauza, Marius and Beau, Maxime and Bhagat, Jai and B{\"o}hm, Claudia and Broux, Martijn and Chen, Susu and Colonell, Jennifer and Gardner, Richard J and Karsh, Bill and Kloosterman, Fabian and Kostadinov, Dimitar and Mora-Lopez, Carolina and O{\textquoteright}Callaghan, John and Park, Junchol and Putzeys, Jan and Sauerbrei, Britton and van Daal, Rik J J and Vollan, Abraham Z and Wang, Shiwei and Welkenhuysen, Marleen and Ye, Zhiwen and Dudman, Joshua T and Dutta, Barundeb and Hantman, Adam W and Harris, Kenneth D and Lee, Albert K and Moser, Edvard I and O{\textquoteright}Keefe, John and Renart, Alfonso and Svoboda, Karel and H{\"a}usser, Michael and Haesler, Sebastian and Carandini, Matteo and Harris, Timothy D} } @article {67826, title = {Nitrite reductase activity within an antiparallel de novo scaffold.}, journal = {JBIC Journal of Biological Inorganic Chemistry}, volume = {26}, year = {2021}, month = {09/2021}, pages = {855 - 862}, abstract = {

Copper nitrite reductase (CuNiR) is a copper enzyme that converts nitrite to nitric oxide and is an important part of the global nitrogen cycle in bacteria. The relatively simple CuHis3\ binding site of the CuNiR active site has made it an enticing target for small molecule modeling and de novo protein design studies. We have previously reported symmetric CuNiR models within parallel three stranded coiled coil systems, with activities that span a range of three orders of magnitude. In this report, we investigate the same CuHis3\ binding site within an antiparallel three helical bundle scaffold, which allows the design of asymmetric constructs. We determine that a simple CuHis3\ binding site can be designed within this scaffold with enhanced activity relative to the comparable construct in parallel coiled coils. Incorporating more complex designs or repositioning this binding site can decrease this activity as much as 15 times. Comparing these constructs, we reaffirm a previous result in which a blue shift in the 1s to 4p transition energy determined by Cu(I) X-ray absorption spectroscopy is correlated with an enhanced activity within imidazole-based constructs. With this step and recent successful electron transfer site designs within this scaffold, we are one step closer to a fully functional de novo designed nitrite reductase.

}, issn = {0949-8257}, doi = {10.1007/s00775-021-01889-1}, url = {https://link.springer.com/10.1007/s00775-021-01889-1}, author = {Koebke, Karl J. and Tebo, Alison G. and Manickas, Elizabeth C. and Deb, Aniruddha and Penner-Hahn, James E. and Pecoraro, Vincent L.} } @article {67181, title = {A novel family of secreted insect proteins linked to plant gall development.}, journal = {Current Biology}, volume = {31}, year = {2021}, month = {2021 May 10}, pages = {1836}, abstract = {

In an elaborate form of inter-species exploitation, many insects hijack plant development to induce novel plant organs called galls that provide the insect with a source of nutrition and a temporary home. Galls result from dramatic reprogramming of plant cell biology driven by insect molecules, but the roles of specific insect molecules in gall development have not yet been determined. Here, we study the aphid Hormaphis cornu, which makes distinctive "cone" galls on leaves of witch hazel Hamamelis virginiana. We found that derived genetic variants in the aphid gene determinant of gall color (dgc) are associated with strong downregulation of dgc transcription in aphid salivary glands, upregulation in galls of seven genes involved in anthocyanin synthesis, and deposition of two red anthocyanins in galls. We hypothesize that aphids inject DGC protein into galls and that this results in differential expression of a small number of plant genes. dgc is a member of a large, diverse family of novel predicted secreted proteins characterized by a pair of widely spaced cysteine-tyrosine-cysteine (CYC) residues, which we named BICYCLE proteins. bicycle genes are most strongly expressed in the salivary glands specifically of galling aphid generations, suggesting that they may regulate many aspects of gall development. bicycle genes have experienced unusually frequent diversifying selection, consistent with their potential role controlling gall development in a molecular arms race between aphids and their host plants.

}, issn = {1879-0445}, doi = {10.1016/j.cub.2021.01.104}, author = {Korgaonkar, Aishwarya and Han, Clair and Lemire, Andrew L and Siwanowicz, Igor and Bennouna, Djawed and Kopec, Rachel E and Andolfatto, Peter and Shigenobu, Shuji and Stern, David L} } @article {67794, title = {The organization and development of cortical interneuron presynaptic circuits are area specific.}, journal = {Cell Reports}, volume = {37}, year = {2021}, month = {2021 Nov 09}, pages = {109993}, abstract = {

Parvalbumin and somatostatin inhibitory interneurons gate information flow in discrete cortical areas that compute sensory and cognitive functions. Despite the considerable differences between areas, individual interneuron subtypes are genetically invariant and are thought to form canonical circuits regardless of which area they are embedded in. Here, we investigate whether this is achieved through selective and systematic variations in their afferent connectivity during development. To this end, we examined the development of their inputs within distinct cortical areas. We find that interneuron afferents show little evidence of being globally stereotyped. Rather, each subtype displays characteristic regional connectivity and distinct developmental dynamics by which this connectivity is achieved. Moreover, afferents dynamically regulated during development are disrupted by early sensory deprivation and in a model of fragile X syndrome. These data provide a comprehensive map of interneuron afferents across cortical areas and reveal the logic by which these circuits are established during development.

}, issn = {2211-1247}, doi = {10.1016/j.celrep.2021.109993}, author = {Pouchelon, Gabrielle and Dwivedi, Deepanjali and Bollmann, Yannick and Agba, Chimuanya K and Xu, Qing and Mirow, Andrea M C and Kim, Sehyun and Qiu, Yanjie and Sevier, Elaine and Ritola, Kimberly D and Cossart, Rosa and Fishell, Gord} } @article {67702, title = {Pervasive fold switching in a ubiquitous protein superfamily.}, journal = {bioRxiv}, year = {2021}, month = {2021 Jun 11}, abstract = {

Fold-switching proteins challenge the one-sequence-one-structure paradigm by adopting multiple stable folds. Nevertheless, it is uncertain whether fold switchers are naturally pervasive or rare exceptions to the well-established rule. To address this question, we developed a predictive method and applied it to the NusG superfamily of \>15,000 transcription factors. We predicted that a substantial population (25\%) of the proteins in this family switch folds. Circular dichroism and nuclear magnetic resonance spectroscopies of 10 sequence-diverse variants confirmed our predictions. Thus, we leveraged family-wide predictions to determine both conserved contacts and taxonomic distributions of fold-switching proteins. Our results indicate that fold switching is pervasive in the NusG superfamily and that the single-fold paradigm significantly biases structure-prediction strategies.

}, doi = {10.1101/2021.06.10.447921}, author = {Lauren L. Porter and Allen K. Kim and Loren L. Looger and Anaya Majumdar and Mary Starich} } @article {67825, title = {The power of peer networking for improving STEM faculty job applications: a successful pilot program}, journal = {bioRxiv}, year = {2021}, month = {10/2021}, abstract = {

In order to successfully obtain a faculty position, postdoctoral fellows or {\textquoteleft}postdocs{\textquoteright}, must submit an application which requires considerable time and effort to produce. These job applications are often reviewed by mentors and colleagues, but rarely are postdocs offered the opportunity to solicit feedback multiple times from reviewers with the same breadth of expertise often found on an academic search committee. To address this gap, this manuscript describes an international peer reviewing program for small groups of postdocs with a broad range of expertise to reciprocally and iteratively provide feedback to each other on their application materials. Over 145 postdocs have participated, often multiple times, over three years. A survey of participants in this program revealed that nearly all participants would recommend participation in such a program to other faculty applicants. Furthermore, this program was more likely to attract participants who struggled to find mentoring and support elsewhere, either because they changed fields or because of their identity as a woman or member of an underrepresented population in STEM. Participation in programs like this one could provide early career academics like postdocs with a diverse and supportive community of peer mentors during the difficult search for a faculty position. Such psychosocial support and encouragement has been shown to prevent attrition of individuals from these populations and programs like this one target the largest {\textquoteleft}leak{\textquoteright} in the pipeline, that of postdoc to faculty. Implementation of similar peer reviewing programs by universities or professional scientific societies could provide a valuable mechanism of support and increased chances of success for early-career academics in their search for independence.Competing Interest StatementThe authors have declared no competing interest.

}, doi = {10.1101/2021.10.16.464662}, url = {https://www.biorxiv.org/content/early/2021/10/18/2021.10.16.464662}, author = {Guardia, Carlos M. and Kane, Erin and Tebo, Alison G. and Sanders, Anna A.W.M. and Kaya, Devrim and Grogan, Kathleen E.} } @article {66000, title = {Precision Mapping of COVID-19 Vulnerable Locales by Epidemiological and Socioeconomic Risk Factors, Developed Using South Korean Data.}, journal = {International Journal of Environmental Research and Public Health}, volume = {18}, year = {2021}, month = {2021 Jan 12}, abstract = {

COVID-19 has severely impacted socioeconomically disadvantaged populations. To support pandemic control strategies, geographically weighted negative binomial regression (GWNBR) mapped COVID-19 risk related to epidemiological and socioeconomic risk factors using South Korean incidence data (January 20, 2020 to July 1, 2020). We constructed COVID-19-specific socioeconomic and epidemiological themes using established social theoretical frameworks and created composite indexes through principal component analysis. The risk of COVID-19 increased with higher area morbidity, risky health behaviours, crowding, and population mobility, and with lower social distancing, healthcare access, and education. Falling COVID-19 risks and spatial shifts over three consecutive time periods reflected effective public health interventions. This study provides a globally replicable methodological framework and precision mapping for COVID-19 and future pandemics.

}, issn = {1660-4601}, doi = {10.3390/ijerph18020604}, author = {Weinstein, Bayarmagnai and da Silva, Alan R and Kouzoukas, Dimitrios E and Bose, Tanima and Kim, Gwang-Jin and Correa, Paola A and Pondugula, Santhi and Lee, YoonJung and Kim, Jihoo and Carpenter, David O} } @article {68990, title = {Principles of signaling pathway modulation for enhancing human naive pluripotency induction.}, journal = {Cell Stem Cell}, volume = {28}, year = {2021}, month = {09/2021}, pages = {1549-1565.e12}, abstract = {

Isolating human MEK/ERK signaling-independent pluripotent stem cells (PSCs) with naive pluripotency characteristics while maintaining differentiation competence and (epi)genetic integrity remains challenging. Here, we engineer reporter systems that allow the screening for defined conditions that induce molecular and functional features of human naive pluripotency. Synergistic inhibition of WNT/β-CATENIN, protein kinase C (PKC), and SRC signaling consolidates the induction of teratoma-competent naive human PSCs, with the capacity to differentiate into trophoblast stem cells (TSCs) and extraembryonic naive endodermal (nEND) cells in\ vitro. Divergent signaling and transcriptional requirements for boosting naive pluripotency were found between mouse and human. P53 depletion in naive hPSCs increased their contribution to mouse-human cross-species chimeric embryos upon priming and differentiation. Finally, MEK/ERK inhibition can be substituted with the inhibition of NOTCH/RBPj, which induces alternative naive-like hPSCs with a diminished risk for deleterious global DNA hypomethylation. Our findings set a framework for defining the signaling foundations of human naive pluripotency.

}, keywords = {Animals, Cell Differentiation, Embryo, Mammalian, Humans, Mice, Pluripotent Stem Cells, Signal Transduction, Trophoblasts}, issn = {1875-9777}, doi = {10.1016/j.stem.2021.04.001}, author = {Bayerl, Jonathan and Ayyash, Muneef and Shani, Tom and Manor, Yair Shlomo and Gafni, Ohad and Massarwa, Rada and Kalma, Yael and Aguilera-Castrejon, Alejandro and Zerbib, Mirie and Amir, Hadar and Sheban, Daoud and Geula, Shay and Mor, Nofar and Weinberger, Leehee and Naveh Tassa, Segev and Krupalnik, Vladislav and Oldak, Bernardo and Livnat, Nir and Tarazi, Shadi and Tawil, Shadi and Wildschutz, Emilie and Ashouokhi, Shahd and Lasman, Lior and Rotter, Varda and Hanna, Suhair and Ben-Yosef, Dalit and Novershtern, Noa and Viukov, Sergey and Hanna, Jacob H} } @article {66900, title = {pyControl: Open source, Python based, hardware and software for controlling behavioural neuroscience experiments}, journal = {bioRxiv}, year = {2021}, month = {2021 Feb 23}, abstract = {

Laboratory behavioural tasks are an essential research tool. As questions asked of behaviour and brain activity become more sophisticated, the ability to specify and run richly structured tasks becomes more important. An increasing focus on reproducibility also necessitates accurate communication of task logic to other researchers. To these ends we developed pyControl, a system of open source hardware and software for controlling behavioural experiments comprising; a simple yet flexible Python-based syntax for specifying tasks as extended state machines, hardware modules for building behavioural setups, and a graphical user interface designed for efficiently running high throughput experiments on many setups in parallel, all with extensive online documentation. These tools make it quicker, easier and cheaper to implement rich behavioural tasks at scale. As important, pyControl facilitates communication and reproducibility of behavioural experiments through a highly readable task definition syntax and self-documenting features.

}, doi = {https://doi.org/10.1101/2021.02.22.432227}, author = {Thomas Akam and Andy Lustig and James Rowland and Sampath K.T. Kapanaiah and Joan Esteve-Agraz and Mariangela Panniello and Cristina Marquez and Michael Kohl and Dennis K{\"a}tzel and Rui M. Costa and Mark Walton} } @article {67295, title = {QUAREP-LiMi: a community endeavor to advance quality assessment and reproducibility in light microscopy.}, journal = {Nature Methods}, year = {2021}, month = {2021 May 21}, issn = {1548-7105}, doi = {10.1038/s41592-021-01162-y}, author = {Ulrike Boehm and Nelson, Glyn and Brown, Claire M and Bagley, Steve and Bajcsy, Peter and Bischof, Johanna and Dauphin, Aurelien and Dobbie, Ian M and Eriksson, John E and Faklaris, Orestis and Fernandez-Rodriguez, Julia and Ferrand, Alexia and Gelman, Laurent and Gheisari, Ali and Hartmann, Hella and Kukat, Christian and Laude, Alex and Mitkovski, Miso and Munck, Sebastian and North, Alison J and Rasse, Tobias M and Resch-Genger, Ute and Schuetz, Lucas C and Seitz, Arne and Strambio-De-Castillia, Caterina and Swedlow, Jason R and Nitschke, Roland} } @article {66009, title = {QUAREP-LiMi: A community-driven initiative to establish guidelines for quality assessment and reproducibility for instruments and images in light microscopy}, journal = {Journal of Microscopy}, volume = {284}, year = {2021}, month = {2021 Oct 01}, pages = {56-73}, abstract = {

In April 2020, the QUality Assessment and REProducibility for Instruments and Images in Light Microscopy (QUAREP-LiMi) initiative was formed. This initiative comprises imaging scientists from academia and industry who share a common interest in achieving a better understanding of the performance and limitations of microscopes and improved quality control (QC) in light microscopy. The ultimate goal of the QUAREP-LiMi initiative is to establish a set of common QC standards, guidelines, metadata models, and tools, including detailed protocols, with the ultimate aim of improving reproducible advances in scientific research. This White Paper 1) summarizes the major obstacles identified in the field that motivated the launch of the QUAREP-LiMi initiative; 2) identifies the urgent need to address these obstacles in a grassroots manner, through a community of stakeholders including, researchers, imaging scientists, bioimage analysts, bioimage informatics developers, corporate partners, funding agencies, standards organizations, scientific publishers, and observers of such; 3) outlines the current actions of the QUAREP-LiMi initiative, and 4) proposes future steps that can be taken to improve the dissemination and acceptance of the proposed guidelines to manage QC. To summarize, the principal goal of the QUAREP-LiMi initiative is to improve the overall quality and reproducibility of light microscope image data by introducing broadly accepted standard practices and accurately captured image data metrics.

}, url = {https://arxiv.org/abs/2101.09153}, author = {Glyn Nelson and Ulrike Boehm and Steve Bagley and Peter Bajcsy and Johanna Bischof and Claire M Brown and Aurelien Dauphin and Ian M Dobbie and John E Eriksson and Orestis Faklaris and Julia Fernandez-Rodriguez and Alexia Ferrand and Ali Gheisari and Hella Hartmann and Christian Kukat and Alex Laude and Miso Mitkovski and Sebastian Munck and Alison J North and Tobias M Rasse and Ute Resch-Genger and Lucas C Schuetz and Arne Seitz and Caterina Strambio-De-Castillia and Jason R Swedlow and Ioannis Alexopoulos and Karin Aumayr and Sergiy Avilov and Gert-Jan Bakker and Rodrigo R Bammann and Andrea Bassi and Hannes Beckert and Sebastian Beer and Yury Belyaev and Jakob Bierwagen and Konstantin A Birngruber and Manel Bosch and Juergen Breitlow and Lisa A Cameron and Joe Chalfoun and James J Chambers and Chieh-Li Chen and Eduardo Conde-Sousa and Alexander D Corbett and Fabrice P Cordelieres and Elaine Del Nery and Ralf Dietzel and Frank Eismann and Elnaz Fazeli and Andreas Felscher and Hans Fried and Nathalie Gaudreault and Wah Ing Goh and Thomas Guilbert and Roland Hadleigh and Peter Hemmerich and Gerhard A Holst and Michelle S Itano and Claudia B Jaffe and Helena K Jambor and Stuart C Jarvis and Antje Keppler and David Kirchenbuechler and Marcel Kirchner and Norio Kobayashi and Gabriel Krens and Susanne Kunis and Judith Lacoste and Marco Marcell and Gabriel G Martins and Daniel J Metcalf and Claire A Mitchell and Joshua Moore and Tobias Mueller and Michael S Nelson and Stephen Ogg and Shuichi Onami and Alexandra L Palmer and Perrine Paul-Gilloteaux and Jaime A Pimentel and Laure Plantard and Santosh Podder and Elton Rexhepaj and Arnaud Royon and Markku A Saari and Damien Schapman and Vincent Schoonderwoert and Britta Schroth-Diez and Stanley Schwartz and Michael Shaw and Martin Spitaler and Martin T Stoeckl and Damir Sudar and Jeremie Teillon and Stefan Terjung and Roland Thuenauer and Christian D Wilms and Graham D Wright and Roland Nitschke and Laurent Gelman} } @article {65833, title = {Reconstruction of motor control circuits in adult Drosophila using automated transmission electron microscopy}, journal = {Cell}, volume = {184}, year = {2021}, month = {2021 Feb 04}, abstract = {

Many animals use coordinated limb movements to interact with and navigate through the environment. To investigate circuit mechanisms underlying locomotor behavior, we used serial-section electron microscopy (EM) to map synaptic connectivity within a neuronal network that controls limb movements. We present a synapse-resolution EM dataset containing the ventral nerve cord (VNC) of an adult female Drosophila melanogaster. To generate this dataset, we developed GridTape, a technology that combines automated serial-section collection with automated high-throughput transmission EM. Using this dataset, we reconstructed 507 motor neurons, including all those that control the legs and wings. We show that a specific class of leg sensory neurons directly synapse onto the largest-caliber motor neuron axons on both sides of the body, representing a unique feedback pathway for fast limb control. We provide open access to the dataset and reconstructions registered to a standard atlas to permit matching of cells between EM and light microscopy data. We also provide GridTape instrumentation designs and software to make large-scale EM data acquisition more accessible and affordable to the scientific community.

}, doi = {10.1101/2020.01.10.902478}, author = {Maniates-Selvin, Jasper T. and Hildebrand, David Grant Colburn and Graham, Brett J. and Kuan, Aaron T. and Thomas, Logan A. and Nguyen, Tri and Buhmann, Julia and Azevedo, Anthony W. and Shanny, Brendan L. and Funke, Jan and Tuthill, John C. and Lee, Wei-Chung Allen} } @article {66904, title = {Recording Electrical Currents across the Plasma Membrane of Mammalian Sperm Cells.}, journal = {Journal of Visualized Experiments: JOVE}, year = {2021}, month = {2021 Feb 14}, abstract = {

Recording of the electrical activity from one of the smallest cells of a mammalian organism- a sperm cell- has been a challenging task for electrophysiologists for many decades. The method known as "spermatozoan patch clamp" was introduced in 2006. It has enabled the direct recording of ion channel activity in whole-cell and cell-attached configurations and has been instrumental in describing sperm cell physiology and the molecular identity of various calcium, potassium, sodium, chloride, and proton ion channels. However, recording from single spermatozoa requires advanced skills and training in electrophysiology. This detailed protocol summarizes the step-by-step procedure and highlights several {\textquoteright}tricks-of-the-trade{\textquoteright} in order to make it available to anyone who wishes to explore the fascinating physiology of the sperm cell. Specifically, the protocol describes recording from human and murine sperm cells but can be adapted to essentially any mammalian sperm cell of any species. The protocol covers important details of the application of this technique, such as isolation of sperm cells, selection of reagents and equipment, immobilization of the highly motile cells, formation of the tight (Gigaohm) seal between a recording electrode and the plasma membrane of the sperm cells, transition into the whole-spermatozoan mode (also known as break-in), and exemplary recordings of the sperm cell calcium ion channel, CatSper, from six mammalian species. The advantages and limitations of the sperm patch clamp method, as well as the most critical steps, are discussed.

}, issn = {1940-087X}, doi = {10.3791/62049}, author = {Liu, Boheng and Mundt, Nadine and Miller, Melissa and Clapham, David E and Kirichok, Yuriy and Lishko, Polina V} } @article {67296, title = {Regulation of coordinated muscular relaxation in Drosophila larvae by a pattern-regulating intersegmental circuit.}, journal = {Nature Communications}, volume = {12}, year = {2021}, month = {2021 May 19}, pages = {2943}, abstract = {

Typical patterned movements in animals are achieved through combinations of contraction and delayed relaxation of groups of muscles. However, how intersegmentally coordinated patterns of muscular relaxation are regulated by the neural circuits remains poorly understood. Here, we identify Canon, a class of higher-order premotor interneurons, that regulates muscular relaxation during backward locomotion of Drosophila larvae. Canon neurons are cholinergic interneurons present in each abdominal neuromere and show wave-like activity during fictive backward locomotion. Optogenetic activation of Canon neurons induces relaxation of body wall muscles, whereas inhibition of these neurons disrupts timely muscle relaxation. Canon neurons provide excitatory outputs to inhibitory premotor interneurons. Canon neurons also connect with each other to form an intersegmental circuit and regulate their own wave-like activities. Thus, our results demonstrate how coordinated muscle relaxation can be realized by an intersegmental circuit that regulates its own patterned activity and sequentially terminates motor activities along the anterior-posterior axis.

}, issn = {2041-1723}, doi = {10.1038/s41467-021-23273-y}, author = {Hiramoto, Atsuki and Jonaitis, Julius and Niki, Sawako and Kohsaka, Hiroshi and Fetter, Richard D and Cardona, Albert and Pulver, Stefan R and Nose, Akinao} } @article {68425, title = {Single-cell transcriptomes of developing and adult olfactory receptor neurons in Drosophila}, journal = {Elife}, volume = {10}, year = {2021}, month = {02/2021}, abstract = {

Recognition of environmental cues is essential for the survival of all organisms. Transcriptional changes occur to enable the generation and function of the neural circuits underlying sensory perception. To gain insight into these changes, we generated single-cell transcriptomes of olfactory- (ORNs), thermo-, and hygro-sensory neurons at an early developmental and adult stage using single-cell and single-nucleus RNA sequencing. We discovered that ORNs maintain expression of the same olfactory receptors across development. Using receptor expression and computational approaches, we matched transcriptomic clusters corresponding to anatomically and physiologically defined neuron types across multiple developmental stages. We found that cell-type-specific transcriptomes partly reflected axon trajectory choices in development and sensory modality in adults. We uncovered stage-specific genes that could regulate the wiring and sensory responses of distinct ORN types. Collectively, our data reveal transcriptomic features of sensory neuron biology and provide a resource for future studies of their development and physiology.

}, keywords = {Animals, Drosophila melanogaster, Drosophila Proteins, Female, Male, Olfactory Receptor Neurons, Sequence Analysis, RNA, Single-Cell Analysis, Smell, Transcriptome}, issn = {2050-084X}, doi = {10.7554/eLife.63856}, author = {McLaughlin, Colleen N and Brbi{\'c}, Maria and Xie, Qijing and Li, Tongchao and Horns, Felix and Kolluru, Sai Saroja and Kebschull, Justus M and Vacek, David and Xie, Anthony and Li, Jiefu and Jones, Robert C and Leskovec, Jure and Quake, Stephen R and Luo, Liqun and Li, Hongjie} } @article {67647, title = {Single-molecule imaging of chromatin remodelers reveals role of ATPase in promoting fast kinetics of target search and dissociation from chromatin.}, journal = {eLife}, volume = {10}, year = {2021}, month = {2021 Jul 27}, abstract = {

Conserved ATP-dependent chromatin remodelers establish and maintain genome-wide chromatin architectures of regulatory DNA during cellular lifespan, but the temporal interactions between remodelers and chromatin targets have been obscure. We performed live-cell single-molecule tracking for RSC, SWI/SNF, CHD1, ISW1, ISW2, and INO80 remodeling complexes in budding yeast and detected hyperkinetic behaviors for chromatin-bound molecules that frequently transition to the free state for all complexes. Chromatin-bound remodelers display notably higher diffusion than nucleosomal histones, and strikingly fast dissociation kinetics with 4-7 s mean residence times. These enhanced dynamics require ATP binding or hydrolysis by the catalytic ATPase, uncovering an additional function to its established role in nucleosome remodeling. Kinetic simulations show that multiple remodelers can repeatedly occupy the same promoter region on a timescale of minutes, implicating an unending {\textquoteright}tug-of-war{\textquoteright} that controls a temporally shifting window of accessibility for the transcription initiation machinery.

}, issn = {2050-084X}, doi = {10.7554/eLife.69387}, author = {Kim, Jee Min and Visanpattanasin, Pat and Jou, Vivian and Liu, Sheng and Tang, Xiaona and Zheng, Qinsi and Li, Kai Yu and Snedeker, Jonathan and Lavis, Luke D and Lionnet, Timoth{\'e}e and Wu, Carl} } @article {67872, title = {Small-molecule ligands can inhibit -1 programmed ribosomal frameshifting in a broad spectrum of coronaviruses.}, journal = {bioRxiv}, year = {2021}, month = {2021 Aug 06}, abstract = {

Recurrent outbreaks of novel zoonotic coronavirus (CoV) diseases since 2000 have high-lighted the importance of developing therapeutics with broad-spectrum activity against CoVs. Because all CoVs use -1 programmed ribosomal frameshifting (-1 PRF) to control expression of key viral proteins, the frameshift signal in viral mRNA that stimulates -1 PRF provides a promising potential target for such therapeutics. To test the viability of this strategy, we explored a group of 6 small-molecule ligands, evaluating their activity against the frameshift signals from a panel of representative bat CoVs{\textemdash}the most likely source of future zoonoses{\textemdash}as well as SARS-CoV-2 and MERS-CoV. We found that whereas some ligands had notable activity against only a few of the frameshift signals, the serine protease inhibitor nafamostat suppressed -1 PRF significantly in several of them, while having limited to no effect on -1 PRF caused by frameshift signals from other viruses used as negative controls. These results suggest it is possible to find small-molecule ligands that inhibit -1 PRF specifically in a broad spectrum of CoVs, establishing the frameshift signal as a viable target for developing pan-coronaviral therapeutics.

}, doi = {10.1101/2021.08.06.455424}, url = {https://doi.org/10.1101/2021.08.06.455424}, author = {Sneha Munshi and Krishna Neupane and Sandaru M. Ileperuma and Matthew T.J. Halma and Jamie A. Kelly and Clarissa F. Halpern and Jonathan D. Dinman and Sarah Loerch and Michael T. Woodside} } @article {67229, title = {SongExplorer: A deep learning workflow for discovery and segmentation of animal acoustic communication signals}, journal = {bioRxiv}, year = {2021}, month = {03/2021}, abstract = {

Many animals produce distinct sounds or substrate-borne vibrations, but these signals have proved challenging to segment with automated algorithms. We have developed SongExplorer, a web-browser based interface wrapped around a deep-learning algorithm that supports an interactive workflow for (1) discovery of animal sounds, (2) manual annotation, (3) supervised training of a deep convolutional neural network, and (4) automated segmentation of recordings. Raw data can be explored by simultaneously examining song events, both individually and in the context of the entire recording, watching synced video, and listening to song. We provide a simple way to visualize many song events from large datasets within an interactive low-dimensional visualization, which facilitates detection and correction of incorrectly labelled song events. The machine learning model we implemented displays higher accuracy than existing heuristic algorithms and similar accuracy as two expert human annotators. We show that SongExplorer allows rapid detection of all song types from new species and of novel song types in previously well-studied species.Competing Interest StatementThe authors have declared no competing interest.

}, doi = {10.1101/2021.03.26.437280}, url = {https://www.biorxiv.org/content/early/2021/03/26/2021.03.26.437280}, author = {Arthur, Benjamin J. and Ding, Yun and Sosale, Medhini and Khalif, Faduma and Kim, Elizabeth and Waddell, Peter and Turaga, Srinivas C. and Stern, David L.} } @article {67673, title = {Spatially patterned excitatory neuron subtypes and projections of the claustrum.}, journal = {eLife}, volume = {10}, year = {2021}, month = {2021 Aug 16}, abstract = {

The claustrum is a functionally and structurally complex brain region, whose very spatial extent remains debated. Histochemical-based approaches typically treat the claustrum as a relatively narrow anatomical region that primarily projects to the neocortex, whereas circuit-based approaches can suggest a broader claustrum region containing projections to the neocortex and other regions. Here, in the mouse, we took a bottom-up and cell-type-specific approach to complement and possibly unite these seemingly disparate conclusions. Using single-cell RNA-sequencing, we found that the claustrum comprises two excitatory neuron subtypes that are differentiable from the surrounding cortex. Multicolor retrograde tracing in conjunction with 12-channel multiplexed in situ hybridization revealed a core-shell spatial arrangement of these subtypes, as well as differential downstream targets. Thus, the claustrum comprises excitatory neuron subtypes with distinct molecular and projection properties, whose spatial patterns reflect the narrower and broader claustral extents debated in previous research. This subtype-specific heterogeneity likely shapes the functional complexity of the claustrum.

}, issn = {2050-084X}, doi = {10.7554/eLife.68967}, author = {Erwin, Sarah R and Bristow, Brianna N and Sullivan, Kaitlin E and Kendrick, Rennie M and Marriott, Brian and Wang, Lihua and Clements, Jody and Lemire, Andrew L and Jackson, Jesse and Cembrowski, Mark S} } @article {67675, title = {Structured patterns of activity in pulse-coupled oscillator networks with varied connectivity.}, journal = {PLoS One}, volume = {16}, year = {2021}, month = {2021 Aug 11}, pages = {e0256034}, abstract = {

Identifying coordinated activity within complex systems is essential to linking their structure and function. We study collective activity in networks of pulse-coupled oscillators that have variable network connectivity and integrate-and-fire dynamics. Starting from random initial conditions, we see the emergence of three broad classes of behaviors that differ in their collective spiking statistics. In the first class ("temporally-irregular"), all nodes have variable inter-spike intervals, and the resulting firing patterns are irregular. In the second ("temporally-regular"), the network generates a coherent, repeating pattern of activity in which all nodes fire with the same constant inter-spike interval. In the third ("chimeric"), subgroups of coherently-firing nodes coexist with temporally-irregular nodes. Chimera states have previously been observed in networks of oscillators; here, we find that the notions of temporally-regular and chimeric states encompass a much richer set of dynamical patterns than has yet been described. We also find that degree heterogeneity and connection density have a strong effect on the resulting state: in binomial random networks, high degree variance and intermediate connection density tend to produce temporally-irregular dynamics, while low degree variance and high connection density tend to produce temporally-regular dynamics. Chimera states arise with more frequency in networks with intermediate degree variance and either high or low connection densities. Finally, we demonstrate that a normalized compression distance, computed via the Lempel-Ziv complexity of nodal spike trains, can be used to distinguish these three classes of behavior even when the phase relationship between nodes is arbitrary.

}, issn = {1932-6203}, doi = {10.1371/journal.pone.0256034}, author = {Kadhim, Kyra L and Hermundstad, Ann M and Brown, Kevin S} } @article {66008, title = {Survey of spiking in the mouse visual system reveals functional hierarchy.}, journal = {Nature}, volume = {592(7852):86-92}, year = {2021}, month = {2021 Jan 20}, pages = {86-92}, abstract = {

The anatomy of the mammalian visual system, from the retina to the neocortex, is organized hierarchically. However, direct observation of cellular-level functional interactions across this hierarchy is lacking due to the challenge of simultaneously recording activity across numerous regions. Here we describe a large, open dataset-part of the Allen Brain Observatory-that surveys spiking from tens of thousands of units in six cortical and two thalamic regions in the brains of mice responding to a battery of visual stimuli. Using cross-correlation analysis, we reveal that the organization of inter-area functional connectivity during visual stimulation mirrors the anatomical hierarchy from the Allen Mouse Brain Connectivity Atlas. We find that four classical hierarchical measures-response latency, receptive-field size, phase-locking to drifting gratings and response decay timescale-are all correlated with the hierarchy. Moreover, recordings obtained during a visual task reveal that the correlation between neural activity and behavioural choice also increases along the hierarchy. Our study provides a foundation for understanding coding and signal propagation across hierarchically organized cortical and thalamic visual areas.

}, issn = {1476-4687}, doi = {10.1038/s41586-020-03171-x}, author = {Siegle, Joshua H and Jia, Xiaoxuan and Durand, Severine and Gale, Sam and Bennett, Corbett and Graddis, Nile and Heller, Greggory and Ramirez, Tamina K and Choi, Hannah and Luviano, Jennifer A and Groblewski, Peter A and Ahmed, Ruweida and Arkhipov, Anton and Bernard, Amy and Billeh, Yazan N and Brown, Dillan and Buice, Michael A and Cain, Nicolas and Caldejon, Shiella and Casal, Linzy and Cho, Andrew and Chvilicek, Maggie and Cox, Timothy C and Dai, Kael and Denman, Daniel J and de Vries, Saskia E J and Dietzman, Roald and Esposito, Luke and Farrell, Colin and Feng, David and Galbraith, John and Garrett, Marina and Gelfand, Emily C and Hancock, Nicole and Harris, Julie A and Howard, Robert and Hu, Brian and Hytnen, Ross and Iyer, Ramakrishnan and Jessett, Erika and Johnson, Katelyn and Kato, India and Kiggins, Justin and Lambert, Sophie and Lecoq, Jerome and Ledochowitsch, Peter and Lee, Jung Hoon and Leon, Arielle and Li, Yang and Liang, Elizabeth and Long, Fuhui and Mace, Kyla and Melchior, Jose and Millman, Daniel and Mollenkopf, Tyler and Nayan, Chelsea and Ng, Lydia and Ngo, Kiet and Nguyen, Thuyahn and Nicovich, Philip R and North, Kat and Ocker, Gabriel Koch and Ollerenshaw, Doug and Oliver, Michael and Pachitariu, Marius and Perkins, Jed and Reding, Melissa and Reid, David and Robertson, Miranda and Ronellenfitch, Kara and Seid, Sam and Slaughterbeck, Cliff and Stoecklin, Michelle and Sullivan, David and Sutton, Ben and Swapp, Jackie and Thompson, Carol and Turner, Kristen and Wakeman, Wayne and Whitesell, Jennifer D and Williams, Derric and Williford, Ali and Young, Rob and Zeng, Hongkui and Naylor, Sarah and Phillips, John W and Reid, R Clay and Mihalas, Stefan and Olsen, Shawn R and Koch, Christof} } @article {67843, title = {Synaptic targets of photoreceptors specialized to detect color and skylight polarization in .}, journal = {eLife}, volume = {10}, year = {2021}, month = {2021 Dec 16}, abstract = {

Color and polarization provide complementary information about the world and are detected by specialized photoreceptors. However, the downstream neural circuits that process these distinct modalities are incompletely understood in any animal. Using electron microscopy, we have systematically reconstructed the synaptic targets of the photoreceptors specialized to detect color and skylight polarization in Drosophila, and we have used light microscopy to confirm many of our findings. We identified known and novel downstream targets that are selective for different wavelengths or polarized light, and followed their projections to other areas in the optic lobes and the central brain. Our results revealed many synapses along the photoreceptor axons between brain regions, new pathways in the optic lobes, and spatially segregated projections to central brain regions. Strikingly, photoreceptors in the polarization-sensitive dorsal rim area target fewer cell types, and lack strong connections to the lobula, a neuropil involved in color processing. Our reconstruction identifies shared wiring and modality-specific specializations for color and polarization vision, and provides a comprehensive view of the first steps of the pathways processing color and polarized light inputs.

}, issn = {2050-084X}, doi = {10.7554/eLife.71858}, author = {Kind, Emil and Longden, Kit D and Nern, Aljoscha and Zhao, Arthur and Sancer, Gizem and Flynn, Miriam A and Laughland, Connor W and Gezahegn, Bruck and Ludwig, Henrique Df and Thomson, Alex G and Obrusnik, Tessa and Alarc{\'o}n, Paula G and Dionne, Heather and Bock, Davi D and Rubin, Gerald M and Reiser, Michael B and Wernet, Mathias F} } @article {67713, title = {Synchronous and opponent thermosensors use flexible cross-inhibition to orchestrate thermal homeostasis.}, journal = {Science Advances}, volume = {7}, year = {2021}, month = {2021 Aug 27}, abstract = {

Body temperature homeostasis is essential and reliant upon the integration of outputs from multiple classes of cooling- and warming-responsive cells. The computations that integrate these outputs are not understood. Here, we discover a set of warming cells (WCs) and show that the outputs of these WCs combine with previously described cooling cells (CCs) in a cross-inhibition computation to drive thermal homeostasis in larval WCs and CCs detect temperature changes using overlapping combinations of ionotropic receptors: Ir68a, Ir93a, and Ir25a for WCs and Ir21a, Ir93a, and Ir25a for CCs. WCs mediate avoidance to warming while cross-inhibiting avoidance to cooling, and CCs mediate avoidance to cooling while cross-inhibiting avoidance to warming. Ambient temperature-dependent regulation of the strength of WC- and CC-mediated cross-inhibition keeps larvae near their homeostatic set point. Using neurophysiology, quantitative behavioral analysis, and connectomics, we demonstrate how flexible integration between warming and cooling pathways can orchestrate homeostatic thermoregulation.

}, issn = {2375-2548}, doi = {10.1126/sciadv.abg6707}, author = {Hernandez-Nunez, Luis and Chen, Alicia and Budelli, Gonzalo and Berck, Matthew E and Richter, Vincent and Rist, Anna and Thum, Andreas S and Cardona, Albert and Klein, Mason and Garrity, Paul and Samuel, Aravinthan D T} } @article {68342, title = {TEMPO: A system to sequentially label and genetically manipulate vertebrate cell lineages}, journal = {bioRxiv}, year = {2021}, month = {10/2021}, abstract = {

During development, regulatory factors appear in a precise order to determine cell fates over time. To investigate complex tissue development, one should not just label cell lineages but further visualize and manipulate cells with temporal control. Current strategies for tracing vertebrate cell lineages lack genetic access to sequentially produced cells. Here we present TEMPO (Temporal Encoding and Manipulation in a Predefined Order), an imaging-readable genetic tool allowing differential labelling and manipulation of consecutive cell generations in vertebrates. TEMPO is based on CRISPR and powered by a cascade of gRNAs that drive orderly activation/inactivation of reporters/effectors. Using TEMPO to visualize zebrafish and mouse neurogenesis, we recapitulated birth-order-dependent neuronal fates. Temporally manipulating cell-cycle regulators in mouse cortex progenitors altered the proportion and distribution of neurons and glia, revealing the effects of temporal gene perturbation on serial cell fates. Thus, TEMPO enables sequential manipulation of molecular factors, crucial to study cell-type specification.One-Sentence Summary Gaining sequential genetic access to vertebrate cell lineages.Competing Interest StatementThe authors have declared no competing interest.

}, doi = {10.1101/2021.10.27.466134}, url = {https://www.biorxiv.org/content/early/2021/10/28/2021.10.27.466134}, author = {Espinosa-Medina, Isabel and Feliciano, Daniel and Belmonte-Mateos, Carla and Garcia-Marques, Jorge and Foster, Benjamin and Miyares, Rosa Linda and Pujades, Cristina and Koyama, Minoru and Lee, Tzumin} } @article {68426, title = {Temporal evolution of single-cell transcriptomes of olfactory projection neurons.}, journal = {Elife}, volume = {10}, year = {2021}, month = {01/2021}, abstract = {

Neurons undergo substantial morphological and functional changes during development to form precise synaptic connections and acquire specific physiological properties. What are the underlying transcriptomic bases? Here, we obtained the single-cell transcriptomes of olfactory projection neurons (PNs) at four developmental stages. We decoded the identity of 21 transcriptomic clusters corresponding to 20 PN types and developed methods to match transcriptomic clusters representing the same PN type across development. We discovered that PN transcriptomes reflect unique biological processes unfolding at each stage-neurite growth and pruning during metamorphosis at an early pupal stage; peaked transcriptomic diversity during olfactory circuit assembly at mid-pupal stages; and neuronal signaling in adults. At early developmental stages, PN types with adjacent birth order share similar transcriptomes. Together, our work reveals principles of cellular diversity during brain development and provides a resource for future studies of neural development in PNs and other neuronal types.

}, keywords = {Animals, Drosophila melanogaster, Neurites, Olfactory Nerve, Single-Cell Analysis, Time Factors, Transcriptome}, issn = {2050-084X}, doi = {10.7554/eLife.63450}, author = {Xie, Qijing and Brbi{\'c}, Maria and Horns, Felix and Kolluru, Sai Saroja and Jones, Robert C and Li, Jiefu and Reddy, Anay R and Xie, Anthony and Kohani, Sayeh and Li, Zhuoran and McLaughlin, Colleen N and Li, Tongchao and Xu, Chuanyun and Vacek, David and Luginbuhl, David J and Leskovec, Jure and Quake, Stephen R and Luo, Liqun and Li, Hongjie} } @article {67845, title = {Unraveling Single-Particle Trajectories Confined in Tubular Networks}, journal = {arXiv}, year = {2021}, month = {2021 Dec 11}, abstract = {

The analysis of single particle trajectories plays an important role in elucidating dynamics within complex environments such as those found in living cells. However, the characterization of intracellular particle motion is often confounded by confinement of the particles within non-trivial subcellular geometries. Here, we focus specifically on the case of particles undergoing Brownian motion within a tubular network, as found in some cellular organelles. An unraveling algorithm is developed to uncouple particle motion from the confining network structure, allowing for an accurate extraction of the diffusion coefficient, as well as differentiating between Brownian and fractional Brownian dynamics. We validate the algorithm with simulated trajectories and then highlight its application to an example system: analyzing the motion of membrane proteins confined in the tubules of the peripheral endoplasmic reticulum in mammalian cells. We show that these proteins undergo diffusive motion with a well-characterized diffusivity. Our algorithm provides a generally applicable approach for disentangling geometric morphology and particle dynamics in networked architectures.

}, url = {https://arxiv.org/abs/2112.05884}, author = {Yunhao Sun and Zexi Yu and Christopher Obara and Keshav Mittal and Jennifer Lippincott-Schwarz and Elena F Koslover} } @article {67750, title = {Whole-cell organelle segmentation in volume electron microscopy.}, journal = {Nature}, volume = {599}, year = {2021}, month = {2021 Nov 01}, pages = {141-46}, abstract = {

Cells contain hundreds of organelles and macromolecular assemblies. Obtaining a complete understanding of their intricate organization requires the nanometre-level, three-dimensional reconstruction of whole cells, which is only feasible with robust and scalable automatic methods. Here, to support the development of such methods, we annotated up to 35 different cellular organelle classes-ranging from endoplasmic reticulum to microtubules to ribosomes-in diverse sample volumes from multiple cell types imaged at a near-isotropic resolution of 4 nm per voxel with focused ion beam scanning electron microscopy (FIB-SEM). We trained deep learning architectures to segment these structures in 4 nm and 8 nm per voxel FIB-SEM volumes, validated their performance and showed that automatic reconstructions can be used to directly quantify previously inaccessible metrics including spatial interactions between cellular components. We also show that such reconstructions can be used to automatically register light and electron microscopy images for correlative studies. We have created an open data and open-source web repository, {\textquoteright}OpenOrganelle{\textquoteright}, to share the data, computer code and trained models, which will enable scientists everywhere to query and further improve automatic reconstruction of these datasets.

}, issn = {1476-4687}, doi = {10.1038/s41586-021-03977-3}, author = {Heinrich, Larissa and Bennett, Davis and Ackerman, David and Park, Woohyun and Bogovic, John and Eckstein, Nils and Petruncio, Alyson and Clements, Jody and Pang, Song and Xu, C Shan and Funke, Jan and Korff, Wyatt and Hess, Harald F and Lippincott-Schwartz, Jennifer and Saalfeld, Stephan and Weigel, Aubrey V} } @article {67646, title = {YAP1 nuclear efflux and transcriptional reprograming follow membrane diminution upon VSV-G-induced cell fusion.}, journal = {Nature Communications}, volume = {12}, year = {2021}, month = {2021 Jul 23}, pages = {4502}, abstract = {

Cells in many tissues, such as bone, muscle, and placenta, fuse into syncytia to acquire new functions and transcriptional programs. While it is known that fused cells are specialized, it is unclear whether cell-fusion itself contributes to programmatic-changes that generate the new cellular state. Here, we address this by employing a fusogen-mediated, cell-fusion system to create syncytia from undifferentiated cells.\ RNA-Seq analysis reveals VSV-G-induced cell fusion precedes transcriptional changes. To gain mechanistic insights, we measure the plasma membrane surface area after cell-fusion and observe it diminishes through increases in endocytosis. Consequently, glucose transporters internalize, and cytoplasmic glucose and ATP transiently decrease. This reduced energetic state activates AMPK, which inhibits YAP1, causing transcriptional-reprogramming and cell-cycle arrest. Impairing either endocytosis or AMPK activity prevents YAP1 inhibition and cell-cycle arrest after fusion. Together, these data demonstrate plasma membrane diminishment upon cell-fusion causes transient nutrient stress that may promote transcriptional-reprogramming independent from extrinsic cues.

}, issn = {2041-1723}, doi = {10.1038/s41467-021-24708-2}, author = {Feliciano, Daniel and Ott, Carolyn M and Isabel Espinosa Medina and Weigel, Aubrey V and Benedetti, Lorena and Milano, Kristin M and Tang, Zhonghua and Lee, Tzumin and Kliman, Harvey J and Guller, Seth M and Lippincott-Schwartz, Jennifer} } @article {65368, title = {3D ATAC-PALM: super-resolution imaging of the accessible genome.}, journal = {Nature Methods}, volume = {17}, year = {2020}, month = {2020 Apr 01}, pages = {430-6}, abstract = {

To image the accessible genome at nanometer scale in situ, we developed three-dimensional assay for transposase-accessible chromatin-photoactivated localization microscopy (3D ATAC-PALM) that integrates an assay for transposase-accessible chromatin with visualization, PALM super-resolution imaging and lattice light-sheet microscopy. Multiplexed with oligopaint DNA{\textendash}fluorescence in situ hybridization (FISH), RNA{\textendash}FISH and protein fluorescence, 3D ATAC-PALM connected microscopy and genomic data, revealing spatially segregated accessible chromatin domains (ACDs) that enclose active chromatin and transcribed genes. Using these methods to analyze genetically perturbed cells, we demonstrated that genome architectural protein CTCF prevents excessive clustering of accessible chromatin and decompacts ACDs. These results highlight 3D ATAC-PALM as a useful tool to probe the structure and organizing mechanism of the genome.

}, issn = {1548-7091}, doi = {10.1038/s41592-020-0775-2}, url = {http://www.nature.com/articles/s41592-020-0775-2}, author = {Xie, Liangqi and Dong, Peng and Chen, Xingqi and Hsieh, Tsung-Han S. and Banala, Sambashiva and De Marzio, Margherita and English, Brian P. and Qi, Yifeng and Jung, Seol Kyoung and Kieffer-Kwon, Kyong-Rim and Legant, Wesley R. and Hansen, Anders S. and Schulmann, Anton and Casellas, Rafael and Zhang, Bin and Betzig, Eric and Lavis, Luke D. and Chang, Howard Y. and Tjian, Robert and Liu, Zhe} } @article {68841, title = {Accelerated EM Connectome Reconstruction using 3D Visualization and Segmentation Graphs}, journal = {bioRxiv}, year = {2020}, month = {2020 Jan 17}, abstract = {

Recent advances in automatic image segmentation and synapse prediction in electron microscopy (EM) datasets of the brain enable more efficient reconstruction of neural connectivity. In these datasets, a single neuron can span thousands of images containing complex tree-like arbors with thousands of synapses. While image segmentation algorithms excel within narrow fields of views, the algorithms sometimes struggle to correctly segment large neurons, which require large context given their size and complexity. Conversely, humans are comparatively good at reasoning with large objects. In this paper, we introduce several semi-automated strategies that combine 3D visualization and machine guidance to accelerate connectome reconstruction. In particular, we introduce a strategy to quickly correct a segmentation through merging and\ cleaving, or splitting a segment along supervoxel boundaries, with both operations driven by affinity scores in the underlying segmentation. We deploy these algorithms as streamlined workflows in a tool called\ Neu3\ and demonstrate superior performance compared to prior work, thus enabling efficient reconstruction of much larger datasets. The insights into proofreading from our work clarify the trade-offs to consider when tuning the parameters of image segmentation algorithms.

}, doi = {10.1101/2020.01.17.909572}, url = {https://doi.org/10.1101/2020.01.17.909572}, author = {Philip M. Hubbard and Stuart Berg and Ting Zhao and Donald J. Olbris and Lowell Umayam and Jeremy Maitin-Shepard and Michal Januszewski and William T. Katz and Erika R. Neace and Stephen M. Plaza} } @article {65615, title = {Arousal modulates retinal output.}, journal = {Neuron}, volume = {107}, year = {2020}, month = {2020 Aug 01}, pages = {487}, abstract = {

At various stages of the visual system, visual responses are modulated by arousal. Here, we find that in mice this modulation operates as early as in the first synapse from the retina and even in retinal axons. To measure retinal activity in the awake, intact brain, we imaged the synaptic boutons of retinal axons in the superior colliculus. Their activity depended not only on vision but also on running speed and pupil size, regardless of retinal illumination. Arousal typically reduced their visual responses and selectivity for direction and orientation. Recordings from retinal axons in the optic tract revealed that arousal modulates the firing of some retinal ganglion cells. Arousal had similar effects postsynaptically in colliculus neurons, independent of activity in the other main source of visual inputs to the colliculus, the primary visual cortex. These results indicate that arousal modulates activity at every stage of the mouse visual system.

}, issn = {1097-4199}, doi = {10.1016/j.neuron.2020.04.026}, author = {Schr{\"o}der, Sylvia and Steinmetz, Nicholas A and Krumin, Michael and Pachitariu, Marius and Rizzi, Matteo and Lagnado, Leon and Harris, Kenneth D and Carandini, Matteo} } @article {65240, title = {Aurora B functions at the apical surface after specialized cytokinesis during morphogenesis in C. elegans.}, journal = {Development}, volume = {147}, year = {2020}, month = {2020 Jan}, pages = {1-16}, abstract = {

While cytokinesis has been intensely studied, the way it is executed during development is not well understood, despite a long-standing appreciation that various aspects of cytokinesis vary across cell and tissue types. To address this, we investigated cytokinesis during the invariant embryonic divisions and found several reproducibly altered parameters at different stages. During early divisions, furrow ingression asymmetry and midbody inheritance is consistent, suggesting specific regulation of these events. During morphogenesis, we found several unexpected alterations to cytokinesis including apical midbody migration in polarizing epithelial cells of the gut, pharynx and sensory neurons. Aurora B kinase, which is essential for several aspects of cytokinesis, remains apically localized in each of these tissues after internalization of midbody ring components. Aurora B inactivation disrupts cytokinesis and causes defects in apical structures, even if inactivated post-mitotically. Therefore, cytokinesis is implemented in a specialized way during epithelial polarization and Aurora B has a new role in the formation of the apical surface.

}, issn = {1477-9129}, doi = {10.1242/dev.181099}, author = {Bai, Xiaofei and Melesse, Michael and Sorensen Turpin, Christopher G and Sloan, Dillon and Chen, Chin-Yi and Wang, Wen-Cheng and Lee, Po-Yi and Simmons, James R and Nebenfuehr, Benjamin and Mitchell, Diana and Klebanow, Lindsey R and Mattson, Nicholas and Betzig, Eric and Chen, Bi-Chang and Cheerambathur, Dhanya and Bembenek, Joshua N} } @article {65311, title = {Behavioral features of motivated response to alcohol in Drosophila.}, journal = {bioRxiv}, year = {2020}, month = {2020 Feb 17}, abstract = {

Animals avoid predators and find the best food and mates by learning from the consequences of their behavior. However, reinforcers are not always uniquely appetitive or aversive but can have complex properties. Most intoxicating substances fall within this category; provoking aversive sensory and physiological reactions while simultaneously inducing overwhelming appetitive properties. Here we describe the subtle behavioral features associated with continued seeking for alcohol despite aversive consequences. We developed an automated runway apparatus to measure how Drosophila respond to consecutive exposures of a volatilized substance. Behavior within this Behavioral Expression of Ethanol Reinforcement Runway (BEER Run) demonstrated a defined shift from aversive to appetitive responses to volatilized ethanol. Behavioral metrics attained by combining computer vision and machine learning methods, reveal that a subset of 9 classified behaviors and component behavioral features associate with this shift. We propose this combination of 9 be

}, author = {Catalano, Jamie L. and Mei, Nicholas and Azanchi, Reza and Song, Sophia and Blackwater, Tyler and Heberlein, Ulrike and Kaun, Karla R.} } @article {65650, title = {Bright and high-performance genetically encoded Ca indicator based on mNeonGreen fluorescent protein.}, journal = {ACS Sensors}, year = {2020}, month = {2020 Jul 08}, abstract = {

Genetically encodable calcium ion (Ca) indicators (GECIs) based on green fluorescent proteins (GFP) are powerful tools for imaging of cell signaling and neural activity in model organisms. Following almost 2 decades of steady improvements in the GFP-based GCaMP series of GECIs, the performance of the most recent generation (i.e., jGCaMP7) may have reached its practical limit due to the inherent properties of GFP. In an effort to sustain the steady progression toward ever-improved GECIs, we undertook the development of a new GECI based on the bright monomeric GFP, mNeonGreen (mNG). The resulting indicator, mNG-GECO1, is 60\% brighter than GCaMP6s in vitro and provides comparable performance as demonstrated by imaging Ca dynamics in cultured cells, primary neurons, and in vivo in larval zebrafish. These results suggest that mNG-GECO1 is a promising next-generation GECI that could inherit the mantle of GCaMP and allow the steady improvement of GECIs to continue for generations to come.

}, issn = {2379-3694}, doi = {10.1021/acssensors.0c00279}, author = {Zarowny, Landon and Abhi Aggarwal and Rutten, Virginia M S and Kolb, Ilya and Patel, Ronak and Huang, Hsin-Yi and Chang, Yu-Fen and Phan, Tiffany and Kanyo, Richard and Ahrens, Misha B and Allison, W Ted and Podgorski, Kaspar and Campbell, Robert E} } @article {66015, title = {Bump-and-Hole Engineering Identifies Specific Substrates of Glycosyltransferases in Living Cells}, journal = {Molecular Cell}, volume = {78}, year = {2020}, month = {Jan-06-2020}, pages = {824 - 834.e15}, abstract = {

Studying posttranslational modifications classically relies on experimental strategies that oversimplify the complex biosynthetic machineries of living cells. Protein glycosylation contributes to essential biological processes, but correlating glycan structure, underlying protein, and disease-relevant biosynthetic regulation is currently elusive. Here, we engineer living cells to tag glycans with editable chemical functionalities while providing information on biosynthesis, physiological context, and glycan fine structure. We introduce a non-natural substrate biosynthetic pathway and use engineered glycosyltransferases to incorporate chemically tagged sugars into the cell surface glycome of the living cell. We apply the strategy to a particularly redundant yet disease-relevant human glycosyltransferase family, the polypeptide\ N-acetylgalactosaminyl transferases. This approach bestows a gain-of-chemical-functionality modification on cells, where the products of individual glycosyltransferases can be selectively characterized or manipulated to understand glycan contribution to major physiological processes.

}, issn = {10972765}, doi = {10.1016/j.molcel.2020.03.030}, url = {https://www.sciencedirect.com/science/article/pii/S1097276520301982?via\%3Dihub}, author = {Schumann, Benjamin and Malaker, Stacy Alyse and Wisnovsky, Simon Peter and Debets, Marjoke Froukje and Agbay, Anthony John and Fernandez, Daniel and Wagner, Lauren Jan Sarbo and Lin, Liang and Li, Zhen and Choi, Junwon and Fox, Douglas Michael and Peh, Jessie and Gray, Melissa Anne and Pedram, Kayvon and Kohler, Jennifer Jean and Mrksich, Milan and Bertozzi, Carolyn Ruth} } @article {68429, title = {Cell-Surface Proteomic Profiling in the Fly Brain Uncovers Wiring Regulators.}, journal = {Cell}, volume = {180}, year = {2020}, month = {01/2020}, pages = {373-386.e15}, abstract = {

Molecular interactions at the cellular interface mediate organized assembly of single cells into tissues and, thus, govern the development and physiology of multicellular organisms. Here, we developed a cell-type-specific, spatiotemporally resolved approach to profile cell-surface proteomes in intact tissues. Quantitative profiling of cell-surface proteomes of Drosophila olfactory projection neurons (PNs) in pupae and adults revealed global downregulation of wiring molecules and upregulation of synaptic molecules in the transition from developing to mature PNs. A proteome-instructed in\ vivo screen identified 20 cell-surface molecules regulating neural circuit assembly, many of which belong to evolutionarily conserved protein families not previously linked to neural development. Genetic analysis further revealed that the lipoprotein receptor LRP1 cell-autonomously controls PN dendrite targeting, contributing to the formation of a precise olfactory map. These findings highlight the power of temporally resolved in situ cell-surface proteomic profiling in discovering regulators of brain wiring.

}, keywords = {Animals, Axons, Brain, Dendrites, Drosophila melanogaster, Drosophila Proteins, Gene Expression Profiling, Gene Expression Regulation, Developmental, Membrane Proteins, Neurogenesis, Olfactory Nerve, Olfactory Pathways, Olfactory Receptor Neurons, Proteomics, Receptors, Lipoprotein, Smell}, issn = {1097-4172}, doi = {10.1016/j.cell.2019.12.029}, author = {Li, Jiefu and Han, Shuo and Li, Hongjie and Udeshi, Namrata D and Svinkina, Tanya and Mani, D R and Xu, Chuanyun and Guajardo, Ricardo and Xie, Qijing and Li, Tongchao and Luginbuhl, David J and Wu, Bing and McLaughlin, Colleen N and Xie, Anthony and Kaewsapsak, Pornchai and Quake, Stephen R and Carr, Steven A and Ting, Alice Y and Luo, Liqun} } @article {65363, title = {Characterization of the Genetic Architecture Underlying Eye Size Variation Within Drosophila melanogaster and Drosophila simulans.}, journal = {Genes|Genomes|Genetics}, volume = {10}, year = {2020}, month = {2020 Mar 01}, pages = {1005-18}, doi = {10.1534/g3.119.400877}, url = {http://g3journal.org/lookup/doi/10.1534/g3.119.400877https://syndication.highwire.org/content/doi/10.1534/g3.119.400877}, author = {Gaspar, Pedro and Arif, Saad and Sumner-Rooney, Lauren and Kittelmann, Maike and Bodey, Andrew J. and Stern, David L. and Nunes, Maria D. S. and McGregor, Alistair P.} } @article {65900, title = {Cis-regulatory variation in the shavenbaby gene underlies intraspecific phenotypic variation, mirroring interspecific divergence in the same trait.}, journal = {Evolution}, year = {2020}, month = {2020 Dec 14}, abstract = {

Despite considerable progress in recent decades in dissecting the genetic causes of natural morphological variation, there is limited understanding of how variation within species ultimately contributes to species differences. We have studied patterning of the non-sensory hairs, commonly known as "trichomes," on the dorsal cuticle of first-instar larvae of Drosophila. Most Drosophila species produce a dense lawn of dorsal trichomes, but a subset of these trichomes were lost in D. sechellia and D. ezoana due entirely to regulatory evolution of the shavenbaby (svb) gene. Here, we describe intraspecific variation in dorsal trichome patterns of first-instar larvae of D. virilis that is similar to the trichome pattern variation identified previously between species. We found that a single large effect QTL, which includes svb, explains most of the trichome number difference between two D. virilis strains and that svb expression correlates with the trichome difference between strains. This QTL does not explain the entire difference between strains, implying that additional loci contribute to variation in trichome numbers. Thus, the genetic architecture of intraspecific variation exhibits similarities and differences with interspecific variation that may reflect differences in long-term and short-term evolutionary processes.

}, issn = {1558-5646}, doi = {10.1111/evo.14142}, author = {Soverna, Ana Faigon and Rodriguez, Nahuel Cabrera and Korgaonkar, Aishwarya and Hasson, Esteban and Stern, David L and Frankel, Nicol{\'a}s} } @article {67225, title = {Comparative single-cell transcriptomics of complete insect nervous systems}, journal = {bioRxiv}, year = {2020}, month = {01/2020}, abstract = {

Molecular profiles of neurons influence information processing, but bridging the gap between genes, circuits, and behavior has been very difficult. Furthermore, the behavioral state of an animal continuously changes across development and as a result of sensory experience. How behavioral state influences molecular cell state is poorly understood. Here we present a complete atlas of the Drosophila larval central nervous system composed of over 200,000 single cells across four developmental stages. We develop polyseq, a python package, to perform cell-type analyses. We use single-molecule RNA-FISH to validate our scRNAseq findings. To investigate how internal state affects cell state, we optogentically altered internal state with high-throughput behavior protocols designed to mimic wasp sting and over activation of the memory system. We found nervous system-wide and neuron-specific gene expression changes. This resource is valuable for developmental biology and neuroscience, and it advances our understanding of how genes, neurons, and circuits generate behavior.

}, doi = {10.1101/785931}, url = {https://www.biorxiv.org/content/early/2020/01/16/785931}, author = {Cocanougher, Benjamin T. and Wittenbach, Jason D. and Long, Xi Salina and Kohn, Andrea B. and Norekian, Tigran P. and Yan, Jinyao and Colonell, Jennifer and Masson, Jean-Baptiste and Truman, James W. and Cardona, Albert and Turaga, Srinivas C. and Singer, Robert H. and Moroz, Leonid L. and Zlatic, Marta} } @article {68507, title = {A connectome and analysis of the adult Drosophila central brain.}, journal = {Elife}, volume = {9}, year = {2020}, month = {2020 Sep 07}, abstract = {

The neural circuits responsible for animal behavior remain largely unknown. We summarize new methods and present the circuitry of a large fraction of the brain of the fruit fly . Improved methods include new procedures to prepare, image, align, segment, find synapses in, and proofread such large data sets. We define cell types, refine computational compartments, and provide an exhaustive atlas of cell examples and types, many of them novel. We provide detailed circuits consisting of neurons and their chemical synapses for most of the central brain. We make the data public and simplify access, reducing the effort needed to answer circuit questions, and provide procedures linking the neurons defined by our analysis with genetic reagents. Biologically, we examine distributions of connection strengths, neural motifs on different scales, electrical consequences of compartmentalization, and evidence that maximizing packing density is an important criterion in the evolution of the fly{\textquoteright}s brain.

}, keywords = {Animals, Brain, Connectome, Drosophila melanogaster, Female, Male, Neurons, Synapses}, issn = {2050-084X}, doi = {10.7554/eLife.57443}, author = {Scheffer, Louis K and Xu, C Shan and Januszewski, Michal and Lu, Zhiyuan and Takemura, Shin-ya and Hayworth, Kenneth J and Huang, Gary B and Shinomiya, Kazunori and Maitlin-Shepard, Jeremy and Berg, Stuart and Clements, Jody and Hubbard, Philip M and Katz, William T and Umayam, Lowell and Zhao, Ting and Ackerman, David and Blakely, Tim and Bogovic, John and Dolafi, Tom and Kainmueller, Dagmar and Kawase, Takashi and Khairy, Khaled A and Leavitt, Laramie and Li, Peter H and Lindsey, Larry and Neubarth, Nicole and Olbris, Donald J and Otsuna, Hideo and Trautman, Eric T and Ito, Masayoshi and Bates, Alexander S and Goldammer, Jens and Wolff, Tanya and Svirskas, Robert and Schlegel, Philipp and Neace, Erika and Knecht, Christopher J and Alvarado, Chelsea X and Bailey, Dennis A and Ballinger, Samantha and Borycz, Jolanta A and Canino, Brandon S and Cheatham, Natasha and Cook, Michael and Dreher, Marisa and Duclos, Octave and Eubanks, Bryon and Fairbanks, Kelli and Finley, Samantha and Forknall, Nora and Francis, Audrey and Hopkins, Gary Patrick and Joyce, Emily M and Kim, SungJin and Kirk, Nicole A and Kovalyak, Julie and Lauchie, Shirley A and Lohff, Alanna and Maldonado, Charli and Manley, Emily A and McLin, Sari and Mooney, Caroline and Ndama, Miatta and Ogundeyi, Omotara and Okeoma, Nneoma and Ordish, Christopher and Padilla, Nicholas and Patrick, Christopher M and Paterson, Tyler and Phillips, Elliott E and Phillips, Emily M and Rampally, Neha and Ribeiro, Caitlin and Robertson, Madelaine K and Rymer, Jon Thomson and Ryan, Sean M and Sammons, Megan and Scott, Anne K and Scott, Ashley L and Shinomiya, Aya and Smith, Claire and Smith, Kelsey and Smith, Natalie L and Sobeski, Margaret A and Suleiman, Alia and Swift, Jackie and Takemura, Satoko and Talebi, Iris and Tarnogorska, Dorota and Tenshaw, Emily and Tokhi, Temour and Walsh, John J and Yang, Tansy and Horne, Jane Anne and Li, Feng and Parekh, Ruchi and Rivlin, Patricia K and Jayaraman, Vivek and Costa, Marta and Jefferis, Gregory Sxe and Ito, Kei and Saalfeld, Stephan and George, Reed and Meinertzhagen, Ian A and Rubin, Gerald M and Hess, Harald F and Jain, Viren and Plaza, Stephen M} } @article {65382, title = {Conservation and divergence of related neuronal lineages in the central brain.}, journal = {eLife}, volume = {9}, year = {2020}, month = {2020 Apr 07}, abstract = {

Wiring a complex brain requires many neurons with intricate cell specificity, generated by a limited number of neural stem cells. central brain lineages are a predetermined series of neurons, born in a specific order. To understand how lineage identity translates to neuron morphology, we mapped 18 central brain lineages. While we found large aggregate differences between lineages, we also discovered shared patterns of morphological diversification. Lineage identity plus Notch-mediated sister fate govern primary neuron trajectories, whereas temporal fate diversifies terminal elaborations. Further, morphological neuron types may arise repeatedly, interspersed with other types. Despite the complexity, related lineages produce similar neuron types in comparable temporal patterns. Different stem cells even yield two identical series of dopaminergic neuron types, but with unrelated sister neurons. Together, these phenomena suggest that straightforward rules drive incredible neuronal complexity, and that large changes in morphology can result from relatively simple fating mechanisms.

}, issn = {2050-084X}, doi = {10.7554/eLife.53518}, author = {Lee, Ying-Jou and Yang, Ching-Po and Miyares, Rosa L and Huang, Yu-Fen and He, Yisheng and Ren, Qingzhong and Chen, Hui-Min and Kawase, Takashi and Ito, Masayoshi and Otsuna, Hideo and Sugino, Ken and Aso, Yoshi and Ito, Kei and Lee, Tzumin} } @article {68991, title = {Context-dependent functional compensation between Ythdf mA reader proteins.}, journal = {Genes Dev}, volume = {34}, year = {2020}, month = {10/2020}, pages = {1373-1391}, abstract = {

The N6-methyladenosine (mA) modification is the most prevalent post-transcriptional mRNA modification, regulating mRNA decay and splicing. It plays a major role during normal development, differentiation, and disease progression. The modification is regulated by a set of writer, eraser, and reader proteins. The YTH domain family of proteins consists of three homologous mA-binding proteins, Ythdf1, Ythdf2, and Ythdf3, which were suggested to have different cellular functions. However, their sequence similarity and their tendency to bind the same targets suggest that they may have overlapping roles. We systematically knocked out (KO) the Mettl3 writer, each of the Ythdf readers, and the three readers together (triple-KO). We then estimated the effect in vivo in mouse gametogenesis, postnatal viability, and in vitro in mouse embryonic stem cells (mESCs). In gametogenesis, severity is increased as the deletion occurs earlier in the process, and Ythdf2 has a dominant role that cannot be compensated by Ythdf1 or Ythdf3, due to differences in readers{\textquoteright} expression pattern across different cell types, both in quantity and in spatial location. Knocking out the three readers together and systematically testing viable offspring genotypes revealed a redundancy in the readers{\textquoteright} role during early development that is gene dosage-dependent. Finally, in mESCs there is compensation between the three Ythdf reader proteins, since the resistance to differentiate and the significant effect on mRNA decay occur only in the triple-KO cells and not in the single KOs. Thus, we suggest a new model for the Ythdf readers function, in which there is profound dosage-dependent redundancy when all three readers are equivalently coexpressed in the same cell types.

}, keywords = {Animals, Cell Line, Dosage Compensation, Genetic, Embryonic Stem Cells, Fertility, Gametogenesis, Gene Deletion, Gene Expression Profiling, Gene Expression Regulation, Developmental, Methyltransferases, Mice, Mice, Knockout, RNA-Binding Proteins}, issn = {1549-5477}, doi = {10.1101/gad.340695.120}, author = {Lasman, Lior and Krupalnik, Vladislav and Viukov, Sergey and Mor, Nofar and Aguilera-Castrejon, Alejandro and Schneir, Dan and Bayerl, Jonathan and Mizrahi, Orel and Peles, Shani and Tawil, Shadi and Sathe, Shashank and Nachshon, Aharon and Shani, Tom and Zerbib, Mirie and Kilimnik, Itay and Aigner, Stefan and Shankar, Archana and Mueller, Jasmine R and Schwartz, Schraga and Stern-Ginossar, Noam and Yeo, Gene W and Geula, Shay and Novershtern, Noa and Hanna, Jacob H} } @article {65600, title = {The corazonin-PTTH neuronal axis controls systemic body growth by regulating basal ecdysteroid biosynthesis in Drosophila melanogaster.}, journal = {Current Biology}, volume = {30}, year = {2020}, month = {2020 Jun 8}, pages = {2156-65}, abstract = {

Steroid hormones play key roles in development, growth, and reproduction in various animal phyla [1]. The insect steroid hormone, ecdysteroid, coordinates growth and maturation, represented by molting and metamorphosis [2]. In Drosophila melanogaster, the prothoracicotropic hormone (PTTH)-producing neurons stimulate peak levels of ecdysteroid biosynthesis for maturation [3]. Additionally, recent studies on PTTH signaling indicated that basal levels of ecdysteroid negatively affect systemic growth prior to maturation [4-8]. However, it remains unclear how PTTH signaling is regulated for basal ecdysteroid biosynthesis. Here, we report that Corazonin (Crz)-producing neurons regulate basal ecdysteroid biosynthesis by affecting PTTH neurons. Crz belongs to gonadotropin-releasing hormone (GnRH) superfamily, implying an analogous role in growth and maturation [9]. Inhibition of Crz neuronal activity increased pupal size, whereas it hardly affected pupariation timing. This phenotype resulted from enhanced growth rate and a delay in ecdysteroid elevation during the mid-third instar larval (L3) stage. Interestingly, Crz receptor (CrzR) expression in PTTH neurons was higher during the mid- than the late-L3 stage. Silencing of CrzR in PTTH neurons increased pupal size, phenocopying the inhibition of Crz neuronal activity. When Crz neurons were optogenetically activated, a strong calcium response was observed in PTTH neurons during the mid-L3, but not the late-L3, stage. Furthermore, we found that octopamine neurons contact Crz neurons in the subesophageal zone (SEZ), transmitting signals for systemic growth. Together, our results suggest that the Crz-PTTH neuronal axis modulates ecdysteroid biosynthesis in response to octopamine, uncovering a regulatory neuroendocrine system in the developmental transition from growth to maturation.

}, issn = {1879-0445}, doi = {10.1016/j.cub.2020.03.050}, author = {Imura, Eisuke and Shimada-Niwa, Yuko and Nishimura, Takashi and H{\"u}ckesfeld, Sebastian and Schlegel, Philipp and Ohhara, Yuya and Kondo, Shu and Tanimoto, Hiromu and Cardona, Albert and Pankratz, Michael J and Niwa, Ryusuke} } @article {68479, title = {Core Competencies for Undergraduates in Bioengineering and Biomedical Engineering: Findings, Consequences, and Recommendations.}, journal = {Ann Biomed Eng}, volume = {48}, year = {2020}, month = {04/2020}, pages = {905-912}, abstract = {

This paper provides a synopsis of discussions related to biomedical engineering core curricula that occurred at the Fourth BME Education Summit held at Case Western Reserve University in Cleveland, Ohio in May 2019. This summit was organized by the Council of Chairs of Bioengineering and Biomedical Engineering, and participants included over 300 faculty members from 100+ accredited undergraduate programs. This discussion focused on six key questions: QI: Is there a core curriculum, and if so, what are its components? QII: How does our purported core curriculum prepare students for careers, particularly in industry? QIII: How does design distinguish BME/BIOE graduates from other engineers? QIV: What is the state of engineering analysis and systems-level modeling in BME/BIOE curricula? QV: What is the role of data science in BME/BIOE undergraduate education? QVI: What core experimental skills are required for BME/BIOE undergrads?\ s. Indeed, BME/BIOI core curricula exists and has matured to emphasize interdisciplinary topics such as physiology, instrumentation, mechanics, computer programming, and mathematical modeling. Departments demonstrate their own identities by highlighting discipline-specific sub-specialties. In addition to technical competence, Industry partners most highly value our students{\textquoteright} capacity for problem solving and communication. As such, BME/BIOE curricula includes open-ended projects that address unmet patient and clinician needs as primary methods to prepare graduates for careers in industry. Culminating senior design experiences distinguish BME/BIOE graduates through their development of client-centered engineering solutions to healthcare problems. Finally, the overall BME/BIOE curriculum is not stagnant-it is clear that data science will become an ever-important element of our students{\textquoteright} training and that new methods to enhance student engagement will be of pedagogical importance as we embark on the next decade.

}, keywords = {Biomedical Engineering, Curriculum, Data Science, Humans, Students, Universities}, issn = {1573-9686}, doi = {10.1007/s10439-020-02468-2}, author = {White, John A and Gaver, Donald P and Butera, Robert J and Choi, Bernard and Dunlop, Mary J and Grande-Allen, K Jane and Grosberg, Anna and Hitchcock, Robert W and Huang-Saad, Aileen Y and Kotche, Miiri and Kyle, Aaron M and Lerner, Amy L and Linehan, John H and Linsenmeier, Robert A and Miller, Michael I and Papin, Jason A and Setton, Lori and Sgro, Allyson and Smith, Michael L and Zaman, Muhammad and Lee, Abraham P} } @article {65281, title = {Correlative three-dimensional super-resolution and block-face electron microscopy of whole vitreously frozen cells.}, journal = {Science}, volume = {367}, year = {2020}, month = {2020 Jan 17}, abstract = {

Within cells, the spatial compartmentalization of thousands of distinct proteins serves a multitude of diverse biochemical needs. Correlative super-resolution (SR) fluorescence and electron microscopy (EM) can elucidate protein spatial relationships to global ultrastructure, but has suffered from tradeoffs of structure preservation, fluorescence retention, resolution, and field of view. We developed a platform for three-dimensional cryogenic SR and focused ion beam-milled block-face EM across entire vitreously frozen cells. The approach preserves ultrastructure while enabling independent SR and EM workflow optimization. We discovered unexpected protein-ultrastructure relationships in mammalian cells including intranuclear vesicles containing endoplasmic reticulum-associated proteins, web-like adhesions between cultured neurons, and chromatin domains subclassified on the basis of transcriptional activity. Our findings illustrate the value of a comprehensive multimodal view of ultrastructural variability across whole cells.

}, issn = {1095-9203}, doi = {10.1126/science.aaz5357}, author = {Hoffman, David P and Shtengel, Gleb and Xu, C Shan and Campbell, Kirby R and Freeman, Melanie and Wang, Lei and Milkie, Daniel E and Pasolli, H Amalia and Iyer, Nirmala and Bogovic, John A and Stabley, Daniel R and Shirinifard, Abbas and Pang, Song and Peale, David and Schaefer, Kathy and Pomp, Wim and Chang, Chi-Lun and Lippincott-Schwartz, Jennifer and Kirchhausen, Tom and Solecki, David J and Betzig, Eric and Hess, Harald F} } @article {65259, title = {Cortical pattern generation during dexterous movement is input-driven.}, journal = {Nature}, volume = {577}, year = {2020}, month = {2020 Jan 16}, pages = {386-91}, abstract = {

The motor cortex controls skilled arm movement by sending temporal patterns of activity to lower motor centres. Local cortical dynamics are thought to shape these patterns throughout movement execution. External inputs have been implicated in setting the initial state of the motor cortex, but they may also have a pattern-generating role. Here we dissect the contribution of local dynamics and inputs to cortical pattern generation during a prehension task in mice. Perturbing cortex to an aberrant state prevented movement initiation, but after the perturbation was released, cortex either bypassed the normal initial state and immediately generated the pattern that controls reaching or failed to generate this pattern. The difference in these two outcomes was probably a result of external inputs. We directly investigated the role of inputs by inactivating the thalamus; this perturbed cortical activity and disrupted limb kinematics at any stage of the movement. Activation of thalamocortical axon terminals at different frequencies disrupted cortical activity and arm movement in a graded manner. Simultaneous recordings revealed that both thalamic activity and the current state of cortex predicted changes in cortical activity. Thus, the pattern generator for dexterous arm movement is distributed across multiple, strongly interacting brain regions.

}, issn = {1476-4687}, doi = {10.1038/s41586-019-1869-9}, author = {Sauerbrei, Britton A and Guo, Jian-Zhong and Cohen, Jeremy D and Mischiati, Matteo and Guo, Wendy and Kabra, Mayank and Verma, Nakul and Mensh, Brett and Branson, Kristin and Hantman, Adam W} } @article {65725, title = {Dense neuronal reconstruction through X-ray holographic nano-tomography.}, journal = {Nature Neuroscience}, volume = {23}, year = {2020}, month = {2020 Dec -1}, pages = {1637-43}, abstract = {

Imaging neuronal networks provides a foundation for understanding the nervous system, but resolving dense nanometer-scale structures over large volumes remains challenging for light microscopy (LM) and electron microscopy (EM). Here we show that X-ray holographic nano-tomography (XNH) can image millimeter-scale volumes with sub-100-nm resolution, enabling reconstruction of dense wiring in Drosophila melanogaster and mouse nervous tissue. We performed correlative XNH and EM to reconstruct hundreds of cortical pyramidal cells and show that more superficial cells receive stronger synaptic inhibition on their apical dendrites. By combining multiple XNH scans, we imaged an adult Drosophila leg with sufficient resolution to comprehensively catalog mechanosensory neurons and trace individual motor axons from muscles to the central nervous system. To accelerate neuronal reconstructions, we trained a convolutional neural network to automatically segment neurons from XNH volumes. Thus, XNH bridges a key gap between LM and EM, providing a new avenue for neural circuit discovery.

}, issn = {1546-1726}, doi = {10.1038/s41593-020-0704-9}, author = {Kuan, Aaron T and Phelps, Jasper S and Thomas, Logan A and Nguyen, Tri M and Han, Julie and Chen, Chiao-Lin and Azevedo, Anthony W and Tuthill, John C and Funke, Jan and Cloetens, Peter and Pacureanu, Alexandra and Lee, Wei-Chung Allen} } @conference {65612, title = {Detecting the Starting Frame of Actions in Video}, booktitle = {2020 IEEE Winter Conference on Applications of Computer Vision (WACV)}, year = {2020}, month = {2020 May 14}, publisher = {IEEE}, organization = {IEEE}, address = {Snowmass Village, CO, USA}, abstract = {

In this work, we address the problem of precisely localizing key frames of an action, for example, the precise time that a pitcher releases a baseball, or the precise time that a crowd begins to applaud. Key frame localization is a largely overlooked and important action-recognition problem, for example in the field of neuroscience, in which we would like to understand the neural activity that produces the start of a bout of an action. To address this problem, we introduce a novel structured loss function that properly weights the types of errors that matter in such applications: it more heavily penalizes extra and missed action start detections over small misalignments. Our structured loss is based on the best matching between predicted and labeled action starts. We train recurrent neural networks (RNNs) to minimize differentiable approximations of this loss. To evaluate these methods, we introduce the Mouse Reach Dataset, a large, annotated video dataset of mice performing a sequence of actions. The dataset was collected and labeled by experts for the purpose of neuroscience research. On this dataset, we demonstrate that our method outperforms related approaches and baseline methods using an unstructured loss.

}, doi = {10.1109/WACV45572.202010.1109/WACV45572.2020.9093405}, url = {https://ieeexplore.ieee.org/xpl/mostRecentIssue.jsp?punumber=9087828}, author = {Kwak, Iljung S. and Guo, Jian-Zhong and Hantman, Adam and Branson, Kristin and Kriegman, David} } @article {65902, title = {Directed Evolution of a Selective and Sensitive Serotonin Sensor via Machine Learning.}, journal = {Cell}, volume = {183}, year = {2020}, month = {2020 Dec 23}, pages = {1986-2002.e26}, abstract = {

Serotonin plays a central role in cognition and is the target of most pharmaceuticals for psychiatric disorders. Existing drugs have limited efficacy; creation of improved versions will require better understanding of serotonergic circuitry, which has been hampered by our inability to monitor serotonin release and transport with high spatial and temporal resolution. We developed and applied a binding-pocket redesign strategy, guided by machine learning, to create a high-performance, soluble, fluorescent serotonin sensor (iSeroSnFR), enabling optical detection of millisecond-scale serotonin transients. We demonstrate that iSeroSnFR can be used to detect serotonin release in freely behaving mice during fear conditioning, social interaction, and sleep/wake transitions. We also developed a robust assay of serotonin transporter function and modulation by drugs. We expect that both machine-learning-guided binding-pocket redesign and iSeroSnFR will have broad utility for the development of other sensors and in\ vitro and in\ vivo serotonin detection, respectively.

}, issn = {1097-4172}, doi = {10.1016/j.cell.2020.11.040}, author = {Unger, Elizabeth K and Keller, Jacob P and Altermatt, Michael and Liang, Ruqiang and Matsui, Aya and Dong, Chunyang and Hon, Olivia J and Yao, Zi and Sun, Junqing and Banala, Samba and Flanigan, Meghan E and Jaffe, David A and Hartanto, Samantha and Carlen, Jane and Mizuno, Grace O and Borden, Phillip M and Shivange, Amol V and Cameron, Lindsay P and Sinning, Steffen and Underhill, Suzanne M and Olson, David E and Amara, Susan G and Temple Lang, Duncan and Rudnick, Gary and Marvin, Jonathan S and Lavis, Luke D and Lester, Henry A and Alvarez, Veronica A and Fisher, Andrew J and Prescher, Jennifer A and Kash, Thomas L and Yarov-Yarovoy, Vladimir and Gradinaru, Viviana and Looger, Loren L and Tian, Lin} } @article {66017, title = {An enzymatic toolkit for selective proteolysis, detection, and visualization of mucin-domain glycoproteins}, journal = {Proceedings of the National Academy of Sciences}, volume = {117}, year = {2020}, month = {Jan-09-2020}, pages = {21299 - 21307}, abstract = {

Densely O-glycosylated mucin domains are found in a broad range of cell surface and secreted proteins, where they play key physiological roles. In addition, alterations in mucin expression and glycosylation are common in a variety of human diseases, such as cancer, cystic fibrosis, and inflammatory bowel diseases. These correlations have been challenging to uncover and establish because tools that specifically probe mucin domains are lacking. Here, we present a panel of bacterial proteases that cleave mucin domains via distinct peptide- and glycan-based motifs, generating a diverse enzymatic toolkit for mucin-selective proteolysis. By mutating catalytic residues of two such enzymes, we engineered mucin-selective binding agents with retained glycoform preferences. StcEE447D\ is a pan-mucin stain derived from enterohemorrhagic\ Escherichia coli\ that is tolerant to a wide range of glycoforms. BT4244E575A\ derived from\ Bacteroides thetaiotaomicron\ is selective for truncated, asialylated core 1 structures commonly associated with malignant and premalignant tissues. We demonstrated that these catalytically inactive point mutants enable robust detection and visualization of mucin-domain glycoproteins by flow cytometry, Western blot, and immunohistochemistry. Application of our enzymatic toolkit to ascites fluid and tissue slices from patients with ovarian cancer facilitated characterization of patients based on differences in mucin cleavage and expression patterns.

}, issn = {0027-8424}, doi = {10.1073/pnas.2012196117}, url = {http://www.pnas.org/lookup/doi/10.1073/pnas.2012196117https://syndication.highwire.org/content/doi/10.1073/pnas.2012196117http://www.pnas.org/syndication/doi/10.1073/pnas.2012196117}, author = {Shon, D. Judy and Malaker, Stacy A. and Pedram, Kayvon and Yang, Emily and Krishnan, Venkatesh and Dorigo, Oliver and Bertozzi, Carolyn R.} } @article {65367, title = {ER membranes exhibit phase behavior at sites of organelle contact.}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {117}, year = {2020}, month = {2020 March 31}, pages = {7225-7235}, abstract = {

The endoplasmic reticulum (ER) is the site of synthesis of secretory and membrane proteins and contacts every organelle of the cell, exchanging lipids and metabolites in a highly regulated manner. How the ER spatially segregates its numerous and diverse functions, including positioning nanoscopic contact sites with other organelles, is unclear. We demonstrate that hypotonic swelling of cells converts the ER and other membrane-bound organelles into micrometer-scale large intracellular vesicles (LICVs) that retain luminal protein content and maintain contact sites with each other through localized organelle tethers. Upon cooling, ER-derived LICVs phase-partition into microscopic domains having different lipid-ordering characteristics, which is reversible upon warming. Ordered ER lipid domains mark contact sites with ER and mitochondria, lipid droplets, endosomes, or plasma membrane, whereas disordered ER lipid domains mark contact sites with lysosomes or peroxisomes. Tethering proteins concentrate at ER{\textendash}organelle contact sites, allowing time-dependent behavior of lipids and proteins to be studied at these sites. These findings demonstrate that LICVs provide a useful model system for studying the phase behavior and interactive properties of organelles in intact cells.

}, issn = {0027-8424}, doi = {10.1073/pnas.1910854117}, url = {http://www.pnas.org/lookup/doi/10.1073/pnas.1910854117}, author = {King, Christopher and Sengupta, Prabuddha and Seo, Arnold Y. and Lippincott-Schwartz, Jennifer} } @article {67704, title = {Extracellular glutamate and GABA transients at the transition from interictal spiking to seizures}, journal = {bioRxiv}, year = {2020}, month = {2020 Nov 15}, abstract = {

Focal epilepsy is associated with intermittent brief population discharges (interictal spikes), which resemble sentinel spikes that often occur at the onset of seizures. Why interictal spikes self-terminate whilst seizures persist and propagate is incompletely understood. Here we use fluorescent glutamate and GABA sensors in an awake rodent model of neocortical seizures to resolve the spatiotemporal evolution of both neurotransmitters in the extracellular space. Interictal spikes are accompanied by brief glutamate transients which are maximal at the initiation site and rapidly propagate centrifugally. GABA transients last longer than glutamate transients and are maximal ~1.5 mm from the focus where they propagate centripetally. At the transition to seizures, GABA transients are attenuated, whilst glutamate transients increase in spatial extent. The data imply that an annulus of feed-forward GABA release intermittently collapses, allowing seizures to escape from local inhibitory restraint.

}, doi = {10.1101/2020.11.13.381707}, author = {Yoshiteru Shimoda and Vincent Magloire and Jonathan S Marvin and Marco Leite and Loren L Looger and Dimitri M Kullmann} } @article {65300, title = {A fast genetically encoded fluorescent sensor for faithful in vivo acetylcholine detection in mice, fish, worms and flies.}, journal = {bioRxiv}, year = {2020}, month = {2020 Feb 8}, abstract = {

Here we design and optimize a genetically encoded fluorescent indicator, iAChSnFR, for the ubiquitous neurotransmitter acetylcholine, based on a bacterial periplasmic binding protein. iAChSnFR shows large fluorescence changes, rapid rise and decay kinetics, and insensitivity to most cholinergic drugs. iAChSnFR revealed large transients in a variety of slice and\ in vivo\ preparations in mouse, fish, fly and worm. iAChSnFR will be useful for the study of acetylcholine in all animals.

}, doi = {https://doi.org/10.1101/2020.02.07.939504}, author = {Borden, Philip and Zhang, Peng and Shivange, Amol V and Marvin, Jonathan S and Cichon, Joseph and Dan, Chuntao and Podgorski, Kaspar and Figueiredo, Antonio and Novak, Ondrej and Tanimoto, Masashi and Shigetomi, Eiji and Lobas, Mark A and Kim, Hyuntae and Zhu, Paula and Zhang, Yajun and Zheng, W. Sharon and Fan, ChengCheng and Wang, Guangfu and Xiang, Bowen and Gan, Li and Zhang, Guang-Xian and Guo, Kaiming and Lin, Li and Cai, Yuan and Yee, Andrew G. and Aggarwal, Aman and Ford, Christopher P. and Rees, Douglas C. and Dietrich, Dirk and Khakh, Baljit S. and Dittman, Jeremy S. and Gan, Wen-Biao and Koyama, Minoru and Jayaraman, Vivek and Cheer, Joseph F. and Lester, Henry A. and Zhu, J. Julius and Looger, Loren L.} } @article {65601, title = {Frontline science: dynamic cellular and subcellular features of migrating leukocytes revealed by in vivo lattice lightsheet microscopy.}, journal = {Journal of Leukocyte Biology}, year = {2020}, month = {2020 Apr 23}, abstract = {

Neutrophil and macrophage (Mϕ) migration underpin the inflammatory response. However, the fast velocity, multidirectional instantaneous movement, and plastic, ever-changing shape of phagocytes confound high-resolution intravital imaging. Lattice lightsheet microscopy (LLSM) captures highly dynamic cell morphology at exceptional spatiotemporal resolution. We demonstrate the first extensive application of LLSM to leukocytes in vivo, utilizing optically transparent zebrafish, leukocyte-specific reporter lines that highlighted subcellular structure, and a wounding assay for leukocyte migration. LLSM revealed details of migrating leukocyte morphology, and permitted intricate, volumetric interrogation of highly dynamic activities within their native physiological setting. Very thin, recurrent uropod extensions must now be considered a characteristic feature of migrating neutrophils. LLSM resolved trailing uropod extensions, demonstrating their surprising length, and permitting quantitative assessment of cytoskeletal contributions to their evanescent form. Imaging leukocytes in blood vessel microenvironments at LLSM{\textquoteright}s spatiotemporal resolution displayed blood-flow-induced neutrophil dynamics and demonstrated unexpected leukocyte-endothelial interactions such as leukocyte-induced endothelial deformation against the intravascular pressure. LLSM of phagocytosis and cell death provided subcellular insights and uncovered novel behaviors. Collectively, we provide high-resolution LLSM examples of leukocyte structures (filopodia lamellipodia, uropod extensions, vesicles), and activities (interstitial and intravascular migration, leukocyte rolling, phagocytosis, cell death, and cytoplasmic ballooning). Application of LLSM to intravital leukocyte imaging sets the stage for transformative studies into the cellular and subcellular complexities of phagocyte biology.

}, issn = {1938-3673}, doi = {10.1002/JLB.3HI0120-589R}, author = {Manley, Harriet R and Potter, David L and Heddleston, John M and Chew, Teng-Leong and Keightley, M Cristina and Lieschke, Graham J} } @article {68813, title = {Functional Diversity of Glycinergic Commissural Inhibitory Neurons in Larval Zebrafish}, journal = {Cell Reports}, volume = {30}, year = {2020}, month = {03/2020}, pages = {3036-3050.e4}, abstract = {

Summary Commissural inhibitory neurons in the spinal cord of aquatic vertebrates coordinate left-right body alternation during swimming. Their developmental origin, however, has been elusive. We investigate this by comparing the anatomy and function of two commissural inhibitory neuron types, dI6dmrt3a and V0d, derived from the pd6 and p0 progenitor domains, respectively. We find that both of these commissural neuron types have monosynaptic, inhibitory connections to neuronal populations active during fictive swimming, supporting their role in providing inhibition to the contralateral side. V0d neurons tend to fire during faster and stronger movements, while dI6dmrt3a neurons tend to fire more consistently during normal fictive swimming. Ablation of dI6dmrt3a neurons leads to an impairment of left-right alternating activity through abnormal co-activation of ventral root neurons on both sides of the spinal cord. Our results suggest that dI6dmrt3a and V0d commissural inhibitory neurons synergistically provide inhibition to the opposite side across different swimming behaviors.

}, keywords = {commissural inhibitory neurons, dI6~ neurons, glycinergic interneuons, mid cycle inhibition, spinal cord circuit, V0d neurons, Zebrafish}, issn = {2211-1247}, doi = {https://doi.org/10.1016/j.celrep.2020.02.015}, url = {https://www.sciencedirect.com/science/article/pii/S2211124720301698}, author = {Chie Satou and Takumi Sugioka and Yuto Uemura and Takashi Shimazaki and Pawel Zmarz and Yukiko Kimura and Shin-ichi Higashijima} } @article {65283, title = {Fundamental law of memory recall.}, journal = {Physical Review Letters}, volume = {124}, year = {2020}, month = {2020 Jan 10}, pages = {018101}, abstract = {

Human memory appears to be fragile and unpredictable. Free recall of random lists of words is a standard paradigm used to probe episodic memory. We proposed an associative search process that can be reduced to a deterministic walk on random graphs defined by the structure of memory representations. The corresponding graph model can be solved analytically, resulting in a novel parameter-free prediction for the average number of memory items recalled (R) out of M items in memory: R=sqrt[3πM/2]. This prediction was verified with a specially designed experimental protocol combining large-scale crowd-sourced free recall and recognition experiments with randomly assembled lists of words or common facts. Our results show that human memory can be described by universal laws derived from first principles.

}, issn = {1079-7114}, doi = {10.1103/PhysRevLett.124.018101}, author = {Naim, Michelangelo and Katkov, Mikhail and Romani, Sandro and Tsodyks, Misha} } @article {65378, title = {A general approach to engineer positive-going eFRET voltage indicators}, journal = {Nature Communications}, volume = {11}, year = {2020}, month = {2020 Jul 10}, abstract = {

We engineered electrochromic fluorescence resonance energy transfer (eFRET) genetically encoded voltage indicators (GEVIs) with {\textquotedblleft}positive-going{\textquotedblright} fluorescence response to membrane depolarization through rational manipulation of the native proton transport pathway in microbial rhodopsins. We transformed the state-of-the-art eFRET GEVI Voltron into Positron, with kinetics and sensitivity equivalent to Voltron but flipped fluorescence signal polarity. We further applied this general approach to GEVIs containing different voltage sensitive rhodopsin domains and various fluorescent dye and fluorescent protein reporters.

}, author = {Abdelfattah, Ahmed S and Valenti, Rosario and Zheng, Jihong and Wong, Allan and Podgorski, Kaspar and Koyama, Minoru and Kim, Douglas S. and Schreiter, Eric R and Project Team GENIE} } @article {65636, title = {Genome-wide kinetic properties of transcriptional bursting in mouse embryonic stem cells.}, journal = {Science Advances}, volume = {6}, year = {2020}, month = {2020 Jun 17}, pages = {eaaz6699}, abstract = {

Transcriptional bursting is the stochastic activation and inactivation of promoters, contributing to cell-to-cell heterogeneity in gene expression. However, the mechanism underlying the regulation of transcriptional bursting kinetics (burst size and frequency) in mammalian cells remains elusive. In this study, we performed single-cell RNA sequencing to analyze the intrinsic noise and mRNA levels for elucidating the transcriptional bursting kinetics in mouse embryonic stem cells. Informatics analyses and functional assays revealed that transcriptional bursting kinetics was regulated by a combination of promoter- and gene body-binding proteins, including the polycomb repressive complex 2 and transcription elongation factors. Furthermore, large-scale CRISPR-Cas9-based screening identified that the Akt/MAPK signaling pathway regulated bursting kinetics by modulating transcription elongation efficiency. These results uncovered the key molecular mechanisms underlying transcriptional bursting and cell-to-cell gene expression noise in mammalian cells.

}, issn = {2375-2548}, doi = {10.1126/sciadv.aaz6699}, author = {Ochiai, Hiroshi and Hayashi, Tetsutaro and Umeda, Mana and Yoshimura, Mika and Harada, Akihito and Shimizu, Yukiko and Nakano, Kenta and Saitoh, Noriko and Liu, Zhe and Yamamoto, Takashi and Okamura, Tadashi and Ohkawa, Yasuyuki and Kimura, Hiroshi and Nikaido, Itoshi} } @article {65657, title = {Growth cone-localized microtubule organizing center establishes microtubule orientation in dendrites.}, journal = {eLife}, volume = {9}, year = {2020}, month = {2020 Jul 13}, abstract = {

A polarized arrangement of neuronal microtubule arrays is the foundation of membrane trafficking and subcellular compartmentalization. Conserved among both invertebrates and vertebrates, axons contain exclusively {\textquoteright}plus-end-out{\textquoteright} microtubules while dendrites contain a high percentage of {\textquoteright}minus-end-out{\textquoteright} microtubules, the origins of which have been a mystery. Here we show that in the dendritic growth cone contains a non-centrosomal microtubule organizing center, which generates minus-end-out microtubules along outgrowing dendrites and plus-end-out microtubules in the growth cone. RAB-11-positive endosomes accumulate in this region and co-migrate with the microtubule nucleation complex γ-TuRC. The MTOC tracks the extending growth cone by kinesin-1/UNC-116-mediated endosome movements on distal plus-end-out microtubules and dynein clusters this advancing MTOC. Critically, perturbation of the function or localization of the MTOC causes reversed microtubule polarity in dendrites. These findings unveil the endosome-localized dendritic MTOC as a critical organelle for establishing axon-dendrite polarity.

}, issn = {2050-084X}, doi = {10.7554/eLife.56547}, author = {Liang, Xing and Kokes, Marcela and Fetter, Richard D and Sallee, Maria Danielle and Moore, Adrian W and Feldman, Jessica L and Shen, Kang} } @article {65972, title = {High throughput instrument to screen fluorescent proteins under two-photon excitation.}, journal = {Biomedical Optics Express}, volume = {11}, year = {2020}, month = {2020 Dec 01}, pages = {7192-7203}, abstract = {

Two-photon microscopy together with fluorescent proteins and fluorescent protein-based biosensors are commonly used tools in neuroscience. To enhance their experimental scope, it is important to optimize fluorescent proteins for two-photon excitation. Directed evolution of fluorescent proteins under one-photon excitation is common, but many one-photon properties do not correlate with two-photon properties. A simple system for expressing fluorescent protein mutants is colonies on an agar plate. The small focal volume of two-photon excitation makes creating a high throughput screen in this system a challenge for a conventional point-scanning approach. We present an instrument and accompanying software that solves this challenge by selectively scanning each colony based on a colony map captured under one-photon excitation. This instrument, called the GIZMO, can measure the two-photon excited fluorescence of 10,000 colonies in 7 hours. We show that the GIZMO can be used to evolve a fluorescent protein under two-photon excitation.

}, issn = {2156-7085}, doi = {10.1364/BOE.409353}, author = {Molina, Rosana S and King, Jonathan and Franklin, Jacob and Clack, Nathan and McRaven, Christopher and Goncharov, Vasily and Flickinger, Daniel and Svoboda, Karel and Drobizhev, Mikhail and Hughes, Thomas E} } @article {65377, title = {High-throughput cellular-resolution synaptic connectivity mapping in vivo with concurrent two-photon optogenetics and volumetric Ca2+ imaging}, journal = {bioRxiv}, year = {2020}, month = {2020 Feb 25}, abstract = {

The ability to measure synaptic connectivity and properties is essential for understanding neuronal circuits. However, existing methods that allow such measurements at cellular resolution are laborious and technically demanding. Here, we describe a system that allows such measurements in a high-throughput way by combining two-photon optogenetics and volumetric Ca2+\ imaging with whole-cell recording. We reveal a circuit motif for generating fast undulatory locomotion in zebrafish.

}, doi = {https://doi.org/10.1101/2020.02.21.959650}, author = {McRaven, Christopher and Tanese, Dimitrii and Zhang, Lixia and Yang, Chao-Tsung and Ahrens, Misha B and Emiliani, Valentina and Koyama, Minoru} } @article {65314, title = {Input connectivity reveals additional heterogeneity of dopaminergic reinforcement in Drosophila}, journal = {Current Biology}, volume = {30}, year = {2020}, month = {2020 Aug 17}, pages = {3200-11}, abstract = {

Different types of\ Drosophila\ dopaminergic neurons (DANs) reinforce memories of unique valence and provide state-dependent motivational control [1]. Prior studies suggest that the compartment architecture of the mushroom body (MB) is the relevant resolution for distinct DAN functions [2,\ 3]. Here we used a recent electron microscope volume of the fly brain [4] to reconstruct the fine anatomy of individual DANs within three MB compartments. We find the 20 DANs of the γ5 compartment, at least some of which provide reward teaching signals, can be clustered into 5 anatomical subtypes that innervate different regions within γ5. Reconstructing 821 upstream neurons reveals input selectivity, supporting the functional relevance of DAN sub-classification. Only one PAM-γ5 DAN subtype (γ5fb) receives direct recurrent input from γ5β{\textquoteright}2a mushroom body output neurons (MBONs) and behavioral experiments distinguish a role for these DANs in memory revaluation from those reinforcing sugar memory. Other DAN subtypes receive major, and potentially reinforcing, inputs from putative gustatory interneurons or lateral horn neurons, which can also relay indirect feedback from the MB. We similarly reconstructed the single aversively reinforcing PPL1-γ1pedc DAN. The γ1pedc DAN inputs are mostly different to those of γ5 DANs and are clustered onto distinct branches of its dendritic tree, presumably separating its established roles in aversive reinforcement and appetitive motivation [5,\ 6]. Additional tracing identified neurons that provide broad input to γ5, β{\textquoteright}2a and γ1pedc DANs suggesting that distributed DAN populations can be coordinately regulated. These connectomic and behavioral analyses therefore reveal additional complexity of dopaminergic reinforcement circuits between and within MB compartments.

}, author = {Otto, Nils and Pleijzier, Markus W. and Morgan, Isabel C. and Edmondson-Stait, Amelia J and Heinz, Konrad J. and Stark, Ildiko and Dempsey, Georgia and Ito, Masayoshi and Kapoor, Ishaan and Hsu, Joseph and Schlegel, Philipp M. and Bates, Alexander S. and Costa, Marta and Ito, Kei and Bock, Davi D. and Rubin, Gerald M. and Jefferis, Gregory S.X.E. and Waddell, Scott} } @article {65618, title = {jYCaMP: an optimized calcium indicator for two-photon imaging at fiber laser wavelengths.}, journal = {Nature Methods}, volume = {17}, year = {2020}, month = {2020 May 25}, pages = {694-97}, abstract = {

Femtosecond lasers at fixed wavelengths above 1,000 nm are powerful, stable and inexpensive, making them promising sources for two-photon microscopy. Biosensors optimized for these wavelengths are needed for both next-generation microscopes and affordable turn-key systems. Here we report jYCaMP1, a yellow variant of the calcium indicator jGCaMP7 that outperforms its parent in mice and flies at excitation wavelengths above 1,000 nm and enables improved two-color calcium imaging with red fluorescent protein-based indicators.

}, issn = {1548-7105}, doi = {10.1038/s41592-020-0835-7}, author = {Mohr, Manuel Alexander and Bushey, Daniel and Abhi Aggarwal and Marvin, Jonathan S and Kim, Jeong Jun and Marquez, Emiliano Jimenez and Liang, Yajie and Patel, Ronak and Macklin, John J and Lee, Chi-Yu and Tsang, Arthur and Tsegaye, Getahun and Ahrens, Allison M and Chen, Jerry L and Kim, Douglas S and Wong, Allan M and Looger, Loren L and Schreiter, Eric R and Podgorski, Kaspar} } @article {65581, title = {Live-cell single particle imaging reveals the role of RNA polymerase II in histone H2A.Z eviction.}, journal = {eLife}, volume = {9}, year = {2020}, month = {2020 Apr 27}, abstract = {

The H2A.Z histone variant, a genome-wide hallmark of permissive chromatin, is enriched near transcription start sites in all eukaryotes. H2A.Z is deposited by the SWR1 chromatin remodeler and evicted by unclear mechanisms. We tracked H2A.Z in living yeast at single-molecule resolution, and found that H2A.Z eviction is dependent on RNA Polymerase II (Pol II) and the Kin28/Cdk7 kinase, which phosphorylates Serine 5 of heptapeptide repeats on the carboxy-terminal domain of the largest Pol II subunit Rpb1. These findings link H2A.Z eviction to transcription initiation, promoter escape and early elongation activities of Pol II. Because passage of Pol II through +1 nucleosomes genome-wide would obligate H2A.Z turnover, we propose that global transcription at yeast promoters is responsible for eviction of H2A.Z. Such usage of yeast Pol II suggests a general mechanism coupling eukaryotic transcription to erasure of the H2A.Z epigenetic signal.

}, issn = {2050-084X}, doi = {10.7554/eLife.55667}, author = {Ranjan, Anand and Nguyen, Vu Q and Liu, Sheng and Wisniewski, Jan and Kim, Jee Min and Tang, Xiaona and Mizuguchi, Gaku and Elalaoui, Ejlal and Nickels, Timothy J and Jou, Vivian and English, Brian P and Zheng, Qinsi and Luk, Ed and Lavis, Luke D and Lionnet, Timoth{\'e}e and Wu, Carl} } @article {68359, title = {Meissner corpuscles and their spatially intermingled afferents underlie gentle touch perception}, journal = {Science}, volume = {368}, year = {2020}, month = {06/2020}, pages = {eabb2751}, abstract = {

The Meissner corpuscle, a mechanosensory end organ, was discovered more than 165 years ago and has since been found in the glabrous skin of all mammals, including that on human fingertips. Although prominently featured in textbooks, the function of the Meissner corpuscle is unknown. Neubarth et al. generated adult mice without Meissner corpuscles and used them to show that these corpuscles alone mediate behavioral responses to, and perception of, gentle forces (see the Perspective by Marshall and Patapoutian). Each Meissner corpuscle is innervated by two molecularly distinct, yet physiologically similar, mechanosensory neurons. These two neuronal subtypes are developmentally interdependent and their endings are intertwined within the corpuscle. Both Meissner mechanosensory neuron subtypes are homotypically tiled, ensuring uniform and complete coverage of the skin, yet their receptive fields are overlapping and offset with respect to each other. Science, this issue p. eabb2751; see also p. 1311 Light touch perception and fine sensorimotor control arise from spatially overlapping mechanoreceptors of the Meissner corpuscle. Meissner corpuscles are mechanosensory end organs that densely occupy mammalian glabrous skin. We generated mice that selectively lacked Meissner corpuscles and found them to be deficient in both perceiving the gentlest detectable forces acting on glabrous skin and fine sensorimotor control. We found that Meissner corpuscles are innervated by two mechanoreceptor subtypes that exhibit distinct responses to tactile stimuli. The anatomical receptive fields of these two mechanoreceptor subtypes homotypically tile glabrous skin in a manner that is offset with respect to one another. Electron microscopic analysis of the two Meissner afferents within the corpuscle supports a model in which the extent of lamellar cell wrappings of mechanoreceptor endings determines their force sensitivity thresholds and kinetic properties.

}, doi = {10.1126/science.abb2751}, url = {https://www.science.org/doi/abs/10.1126/science.abb2751}, author = {Nicole L. Neubarth and Alan J. Emanuel and Yin Liu and Mark W. Springel and Annie Handler and Qiyu Zhang and Brendan P. Lehnert and Chong Guo and Lauren L. Orefice and Amira Abdelaziz and Michelle M. DeLisle and Michael Iskols and Julia Rhyins and Soo J. Kim and Stuart J. Cattel and Wade Regehr and Christopher D. Harvey and Jan Drugowitsch and David D. Ginty} } @article {65826, title = {Metabolic determinants of cellular fitness dependent on mitochondrial reactive oxygen species.}, journal = {Science Advances}, volume = {6}, year = {2020}, month = {2020 Nov 04}, abstract = {

Mitochondria-derived reactive oxygen species (mROS) are required for the survival, proliferation, and metastasis of cancer cells. The mechanism by which mitochondrial metabolism regulates mROS levels to support cancer cells is not fully understood. To address this, we conducted a metabolism-focused CRISPR-Cas9 genetic screen and uncovered that loss of genes encoding subunits of mitochondrial complex I was deleterious in the presence of the mitochondria-targeted antioxidant mito-vitamin E (MVE). Genetic or pharmacologic inhibition of mitochondrial complex I in combination with the mitochondria-targeted antioxidants, MVE or MitoTEMPO, induced a robust integrated stress response (ISR) and markedly diminished cell survival and proliferation in vitro. This was not observed following inhibition of mitochondrial complex III. Administration of MitoTEMPO in combination with the mitochondrial complex I inhibitor phenformin decreased the leukemic burden in a mouse model of T cell acute lymphoblastic leukemia. Thus, mitochondrial complex I is a dominant metabolic determinant of mROS-dependent cellular fitness.

}, issn = {2375-2548}, doi = {10.1126/sciadv.abb7272}, author = {Kong, Hyewon and Reczek, Colleen R and McElroy, Gregory S and Steinert, Elizabeth M and Wang, Tim and Sabatini, David M and Chandel, Navdeep S} } @article {65738, title = {The mind of a mouse.}, journal = {Cell}, volume = {182}, year = {2020}, month = {2020 Sep 17}, pages = {1372-1376}, abstract = {

Large scientific projects in genomics and astronomy are influential not because they answer any single question but because they enable investigation of continuously arising new questions from the same data-rich sources. Advances in automated mapping of the brain{\textquoteright}s synaptic connections (connectomics) suggest that the complicated circuits underlying brain function are ripe for analysis. We discuss benefits of mapping a mouse brain at the level of synapses.

}, issn = {1097-4172}, doi = {10.1016/j.cell.2020.08.010}, author = {Abbott, Larry F and Bock, Davi D and Callaway, Edward M and Denk, Winfried and Dulac, Catherine and Fairhall, Adrienne L and Fiete, Ila and Harris, Kristen M and Helmstaedter, Moritz and Jain, Viren and Kasthuri, Narayanan and LeCun, Yann and Lichtman, Jeff W and Littlewood, Peter B and Luo, Liqun and Maunsell, John H R and Reid, R Clay and Rosen, Bruce R and Rubin, Gerald M and Sejnowski, Terrence J and Seung, H Sebastian and Svoboda, Karel and Tank, David W and Tsao, Doris and Van Essen, David C} } @article {65613, title = {mRNA stem-loops can pause the ribosome by hindering A-site tRNA binding.}, journal = {eLife}, volume = {9}, year = {2020}, month = {2020 May 19}, abstract = {

Although the elongating ribosome is an efficient helicase, certain mRNA stem-loop structures are known to impede ribosome movement along mRNA and stimulate programmed ribosome frameshifting via mechanisms that are not well understood. Using biochemical and single-molecule F{\"o}rster resonance energy transfer (smFRET) experiments, we studied how frameshift-inducing stem-loops from mRNA and the transcript of Human Immunodeficiency Virus (HIV) perturb translation elongation. We find that upon encountering the ribosome, the stem-loops strongly inhibit A-site tRNA binding and ribosome intersubunit rotation that accompanies translation elongation. Electron cryo-microscopy (cryo-EM) reveals that the HIV stem-loop docks into the A site of the ribosome. Our results suggest that mRNA stem-loops can transiently escape the ribosome helicase by binding to the A site. Thus, the stem-loops can modulate gene expression by sterically hindering tRNA binding and inhibiting translation elongation.

}, issn = {2050-084X}, doi = {10.7554/eLife.55799}, author = {Bao, Chen and Loerch, Sarah and Ling, Clarence and Korostelev, Andrei A and Grigorieff, Nikolaus and Ermolenko, Dmitri N} } @article {65791, title = {The mTORC1/S6K/PDCD4/eIF4A axis determines outcome of mitotic arrest.}, journal = {Cell Reports}, volume = {33}, year = {2020}, month = {2020 Oct 06}, pages = {108230}, abstract = {

mTOR is a serine/threonine kinase and a master regulator of cell growth and proliferation. Raptor, a scaffolding protein that recruits substrates to mTOR complex 1 (mTORC1), is known to be phosphorylated during mitosis, but the significance of this phosphorylation remains largely unknown. Here we show that raptor expression and mTORC1 activity are dramatically reduced in cells arrested in mitosis. Expression of a non-phosphorylatable raptor mutant reactivates mTORC1 and significantly reduces cytotoxicity of the mitotic poison Taxol. This effect is mediated via degradation of PDCD4, a tumor suppressor protein that inhibits eIF4A activity and is negatively regulated by the mTORC1/S6K pathway. Moreover, pharmacological inhibition of eIF4A is able to enhance the effects of Taxol and restore sensitivity in Taxol-resistant cancer cells. These findings indicate that the mTORC1/S6K/PDCD4/eIF4A axis has a pivotal role in the death versus slippage decision during mitotic arrest and may be exploited clinically to treat tumors resistant to anti-mitotic agents.

}, issn = {2211-1247}, doi = {10.1016/j.celrep.2020.108230}, author = {Moustafa-Kamal, Mohamed and Kucharski, Thomas J and El-Assaad, Wissal and Abbas, Yazan M and Gandin, Valentina and Nagar, Bhushan and Pelletier, Jerry and Topisirovic, Ivan and Teodoro, Jose G} } @article {65653, title = {Multifield and inverse-contrast switching of magnetocaloric high contrast ratio MRI labels.}, journal = {Magnetic Resonance in Medicine}, volume = {85}, year = {2020}, month = {2020 Jul 07}, pages = {506-17}, abstract = {

PURPOSE: Demonstrating multifield and inverse contrast switching of magnetocaloric high contrast ratio MRI labels that either have increasing or decreasing moment versus temperature slopes depending on the material at physiological temperatures and different MRI magnetic field strengths.

METHODS: Two iron-rhodium samples of different purity (99\% and 99.9\%) and a lanthanum-iron-silicon sample were obtained from commercial vendors. Temperature and magnetic field-dependent magnetic moment measurements of the samples were performed on a vibrating sample magnetometer. Temperature-dependent MRI of different iron-rhodium and lanthanum-iron-silicon samples were performed on 3 different MRI scanners at 1 Tesla (T), 4.7T, and 7T.

RESULTS: Sharp, first-order magnetic phase transition of each iron-rhodium sample at a physiologically relevant temperature (~37{\textdegree}C) but at different MRI magnetic fields (1T, 4.7T, and 7T, depending on the sample) showed clear image contrast changes in temperature-dependent MRI. Iron-rhodium and lanthanum-iron-silicon samples with sharp, first-order magnetic phase transitions at the same MRI field of 1T and physiological temperature of 37{\textdegree}C, but with positive and negative slope of magnetization versus temperature, respectively, showed clear inverse contrast image changes. Temperature-dependent MRI on individual microparticle samples of lanthanum-iron-silicon also showed sharp image contrast changes.

CONCLUSION: Magnetocaloric materials of different purity and composition were demonstrated to act as diverse high contrast ratio switchable MRI contrast agents. Thus, we show that a range of magnetocaloric materials can be optimized for unique image contrast response under MRI-appropriate conditions at physiological temperatures and be controllably switched in situ.

}, issn = {1522-2594}, doi = {10.1002/mrm.28400}, author = {Barbic, Mladen and Dodd, Stephen J and ElBidweihy, Hatem and Dilley, Neil R and Marcheschi, Barbara and Huston, Alan L and Morris, H Douglas and Koretsky, Alan P} } @article {65262, title = {Nanoscale subcellular architecture revealed by multicolor three-dimensional salvaged fluorescence imaging.}, journal = {Nature Methods}, volume = {17}, year = {2020}, month = {2020 Jan 06}, pages = {225-231}, abstract = {

Combining the molecular specificity of fluorescent probes with three-dimensional imaging at nanoscale resolution is critical for investigating the spatial organization and interactions of cellular organelles and protein complexes. We present a 4Pi single-molecule switching super-resolution microscope that enables ratiometric multicolor imaging of mammalian cells at 5-10-nm localization precision in three dimensions using {\textquoteright}salvaged fluorescence{\textquoteright}. Imaging two or three fluorophores simultaneously, we show fluorescence images that resolve the highly convoluted Golgi apparatus and the close contacts between the endoplasmic reticulum and the plasma membrane, structures that have traditionally been the imaging realm of electron microscopy. The salvaged fluorescence approach is equally applicable in most single-objective microscopes.

}, issn = {1548-7105}, doi = {10.1038/s41592-019-0676-4}, author = {Zhang, Yongdeng and Schroeder, Lena K and Lessard, Mark D and Kidd, Phylicia and Chung, Jeeyun and Song, Yuanbin and Benedetti, Lorena and Li, Yiming and Ries, Jonas and Grimm, Jonathan B and Lavis, Luke D and De Camilli, Pietro and Rothman, James E and Baddeley, David and Bewersdorf, Joerg} } @article {65604, title = {Nanoscopic visualization of restricted nonvolume cholinergic and monoaminergic transmission with genetically encoded sensors.}, journal = {Nano Letters}, volume = {20}, year = {2020}, month = {2020 Jun}, pages = {4073-83}, abstract = {

How neuromodulatory transmitters diffuse into the extracellular space remains an unsolved fundamental biological question, despite wide acceptance of the volume transmission model. Here, we report development of a method combining genetically encoded fluorescent sensors with high-resolution imaging and analysis algorithms which permits the first direct visualization of neuromodulatory transmitter diffusion at various neuronal and non-neuronal cells. Our analysis reveals that acetylcholine and monoamines diffuse at individual release sites with a spread length constant of \~{}0.75 μm. These transmitters employ varied numbers of release sites, and when spatially close-packed release sites coactivate they can spillover into larger subcellular areas. Our data indicate spatially restricted (i.e., nonvolume) neuromodulatory transmission to be a prominent intercellular communication mode, reshaping current thinking of control and precision of neuromodulation crucial for understanding behaviors and diseases.

}, issn = {1530-6992}, doi = {10.1021/acs.nanolett.9b04877}, author = {Zhu, Paula K and Zheng, W Sharon and Zhang, Peng and Jing, Miao and Borden, Philip M and Ali, Farhan and Guo, Kaiming and Feng, Jiesi and Marvin, Jonathan S and Wang, Yali and Wan, Jinxia and Gan, Li and Kwan, Alex C and Lin, Li and Looger, Loren L and Li, Yulong and Zhang, Yajun} } @article {65263, title = {The neuropeptide Drosulfakinin regulates social isolation-induced aggression in Drosophila.}, journal = {Journal of Experimental Biology}, volume = {223}, year = {2020}, month = {2020 Jan 03}, abstract = {

Social isolation strongly modulates behavior across the animal kingdom. We utilized the fruit fly to study social isolation-driven changes in animal behavior and gene expression in the brain. RNA-seq identified several head-expressed genes strongly responding to social isolation or enrichment. Of particular interest, social isolation downregulated expression of the gene encoding the neuropeptide (), the homologue of vertebrate cholecystokinin (CCK), which is critical for many mammalian social behaviors. knockdown significantly increased social isolation-induced aggression. Genetic activation or silencing of neurons each similarly increased isolation-driven aggression. Our results suggest a U-shaped dependence of social isolation-induced aggressive behavior on signaling, similar to the actions of many neuromodulators in other contexts.

}, issn = {1477-9145}, doi = {10.1242/jeb.207407}, author = {Agrawal, Pavan and Kao, Damian and Chung, Phuong and Looger, Loren L} } @article {65318, title = {Nicotine exposure and neuronal activity regulate Golgi membrane dispersal and distribution}, journal = {bioRxiv}, year = {2020}, month = {2020 Feb 26}, abstract = {

How nicotine exposure produces long-lasting changes that remodel neural circuits with addiction is unknown. Here, we report that long-term nicotine exposure alters the trafficking of α4β2-type nicotinic acetylcholine receptors (α4β2Rs) by dispersing and redistributing the Golgi apparatus. In cultured neurons, dispersed Golgi membranes were distributed throughout somata, dendrites and axons. Small, mobile vesicles in dendrites and axons lacked standard Golgi markers and were identified by other Golgi enzymes that modify glycans. Nicotine exposure increased levels of dispersed Golgi membranes, which required α4β2R expression. Similar nicotine-induced changes occurred\ in vivo\ at dopaminergic neurons at mouse nucleus accumbens terminals, consistent with these events contributing to nicotine{\textquoteright}s addictive effects. Characterization\ in vitro\ demonstrated that dispersal was reversible, that dispersed Golgi membranes were functional, and that membranes were heterogenous in size, with smaller vesicles emerging from larger {\textquotedblleft}ministacks{\textquotedblright}, similar to Golgi dispersal induced by nocadazole. Protocols that increased cultured neuronal synaptic excitability also increased Golgi dispersal, without the requirement of α4β2R expression. Our findings reveal novel activity- and nicotine-dependent changes in neuronal intracellular morphology. These changes regulate levels and location of dispersed Golgi membranes at dendrites and axons, which function in local trafficking at subdomains.

}, author = {Govind, Anitha P. and Jeyifous, Okunola and Russell, Theron A. and Vaasjo, Lee O. and Yi, Zola and Weigel, Aubrey V. and Newell, Luke and Koranda, Jessica L. and Singh, Karanveer and Valbuena, Fernando and Glick, Benjamin S. and Mukherjee, Jogeshwar and Lippincott-Schwartz, Jennifer and Zhuang, Xiaoxi and Green, William N.} } @article {65724, title = {Novel fluorescent ligands enable single-molecule localization microscopy of the dopamine transporter.}, journal = {ACS Chemical Neuroscience}, year = {2020}, month = {2020 Sep 14}, abstract = {

The dopamine transporter (DAT) is critical for spatiotemporal control of dopaminergic neurotransmission and the target for therapeutic agents, including ADHD medications, and abused substances, such as cocaine. Here, we develop new fluorescently labeled ligands that bind DAT with high affinity and enable single-molecule detection of the transporter. The cocaine analogue MFZ2-12 (1) was conjugated to novel rhodamine-based Janelia Fluorophores (JF549 and JF646). High affinity binding of the resulting ligands to DAT was demonstrated by potent inhibition of [3H]dopamine uptake in DAT transfected CAD cells and by competition radioligand binding experiments on rat striatal membranes. Visualization of binding was substantiated by confocal or TIRF microscopy revealing selective binding of the analogues to DAT transfected CAD cells. Single particle tracking experiments were performed with JF549-conjugated DG3-80 (3) and JF646-conjugated DG4-91 (4) on DAT transfected CAD cells enabling quantification and categorization of the dynamic behavior of DAT into four distinct motion classes (immobile, confined, Brownian, and directed). Finally, we show that the ligands can be used in direct stochastic optical reconstruction microscopy (dSTORM) experiments permitting further analyses of DAT distribution on the nanoscale. In summary, these novel fluorescent ligands are promising new tools for studying DAT localization and regulation with single-molecule resolution.

}, issn = {1948-7193}, doi = {10.1021/acschemneuro.0c00397}, author = {Guthrie, Daryl and Klein Herenbrink, Carmen and Lycas, Matthew and Ku, Therese and Bonifazi, Alessandro and DeVree, Brian and Mathiasen, Signe and Javitch, Jonathan and Grimm, Jonathan B and Lavis, Luke D and Gether, Ulrik and Newman, Amy Hauck} } @article {65758, title = {Nuclear crowding and nonlinear diffusion during interkinetic nuclear migration in the zebrafish retina.}, journal = {eLife}, volume = {9}, year = {2020}, month = {2020 Oct 06}, abstract = {

An important question in early neural development is the origin of stochastic nuclear movement between apical and basal surfaces of neuroepithelia during interkinetic nuclear migration. Tracking of nuclear subpopulations has shown evidence of diffusion - mean squared displacements growing linearly in time - and suggested crowding from cell division at the apical surface drives basalward motion. Yet, this hypothesis has not yet been tested, and the forces involved not quantified. We employ long-term, rapid light-sheet and two-photon imaging of early zebrafish retinogenesis to track entire populations of nuclei within the tissue. The time-varying concentration profiles show clear evidence of crowding as nuclei reach close-packing and are quantitatively described by a nonlinear diffusion model. Considerations of nuclear motion constrained inside the enveloping cell membrane show that concentration-dependent stochastic forces inside cells, compatible in magnitude to those found in cytoskeletal transport, can explain the observed magnitude of the diffusion constant.

}, issn = {2050-084X}, doi = {10.7554/eLife.58635}, author = {Azizi, Afnan and Herrmann, Anne and Wan, Yinan and Buse, Salvador Jrp and Keller, Philipp J and Goldstein, Raymond E and Harris, William A} } @article {65634, title = {Precision Calcium Imaging of Dense Neural Populations via a Cell-Body-Targeted Calcium Indicator.}, journal = {Neuron}, volume = {107}, year = {2020}, month = {2020 Aug 01}, pages = {470}, abstract = {

Methods for one-photon fluorescent imaging of calcium dynamics can capture the activity of hundreds of neurons across large fields of view at a low equipment complexity and cost. In contrast to two-photon methods, however, one-photon methods suffer from higher levels of crosstalk from neuropil, resulting in a decreased signal-to-noise ratio and artifactual correlations of neural activity. We address this problem by engineering cell-body-targeted variants of the fluorescent calcium indicators GCaMP6f and GCaMP7f. We screened fusions of GCaMP to natural, as well as artificial, peptides and identified fusions that localized GCaMP to within 50\ μm of the cell body of neurons in mice and larval zebrafish. One-photon imaging of soma-targeted GCaMP in dense neural circuits reported fewer artifactual spikes from neuropil, an increased signal-to-noise ratio, and decreased artifactual correlation across neurons. Thus, soma-targeting of fluorescent calcium indicators facilitates usage of simple, powerful, one-photon methods for imaging neural calcium dynamics.

}, issn = {1097-4199}, doi = {10.1016/j.neuron.2020.05.029}, author = {Shemesh, Or A and Linghu, Changyang and Piatkevich, Kiryl D and Goodwin, Daniel and Celiker, Orhan Tunc and Gritton, Howard J and Romano, Michael F and Gao, Ruixuan and Yu, Chih-Chieh Jay and Tseng, Hua-An and Bensussen, Seth and Narayan, Sujatha and Yang, Chao-Tsung and Freifeld, Limor and Siciliano, Cody A and Gupta, Ishan and Wang, Joyce and Pak, Nikita and Yoon, Young-Gyu and Ullmann, Jeremy F P and Guner-Ataman, Burcu and Noamany, Habiba and Sheinkopf, Zoe R and Park, Won Min and Asano, Shoh and Keating, Amy E and Trimmer, James S and Reimer, Jacob and Tolias, Andreas S and Bear, Mark F and Tye, Kay M and Han, Xue and Ahrens, Misha B and Boyden, Edward S} } @article {65721, title = {Preventing cytokine storm syndrome in COVID-19 using α-1 adrenergic receptor antagonists.}, journal = {The Journal of Clinical Investigatio}, volume = {130}, year = {2020}, month = {2020 Jul 01}, pages = {3345-47}, keywords = {Adrenergic alpha-1 Receptor Antagonists, Clinical Trials as Topic, Coronavirus Infections, Cytokine Release Syndrome, Humans, Pandemics, Pneumonia, Viral}, issn = {1558-8238}, doi = {10.1172/JCI139642}, author = {Konig, Maximilian F and Powell, Mike and Staedtke, Verena and Bai, Ren-Yuan and Thomas, David L and Fischer, Nicole and Huq, Sakibul and Khalafallah, Adham M and Koenecke, Allison and Xiong, Ruoxuan and Mensh, Brett and Papadopoulos, Nickolas and Kinzler, Kenneth W and Vogelstein, Bert and Vogelstein, Joshua T and Athey, Susan and Zhou, Shibin and Bettegowda, Chetan} } @article {65669, title = {A programmable sequence of reporters for lineage analysis.}, journal = {Nature Neuroscience}, volume = {23}, year = {2020}, month = {2020 Dec 01}, pages = {1618-28}, abstract = {

We present CLADES (cell lineage access driven by an edition sequence), a technology for cell lineage studies based on CRISPR-Cas9 techniques. CLADES relies on a system of genetic switches to activate and inactivate reporter genes in a predetermined order. Targeting CLADES to progenitor cells allows the progeny to inherit a sequential cascade of reporters, thereby coupling birth order to reporter expression. This system, which can also be temporally induced by heat shock, enables the temporal resolution of lineage development and can therefore be used to deconstruct an extended cell lineage by tracking the reporters expressed in the progeny. When targeted to the germ line, the same cascade progresses across animal generations, predominantly marking each generation with the corresponding combination of reporters. CLADES therefore offers an innovative strategy for making programmable cascades of genes that can be used for genetic manipulation or to record serial biological events.

}, issn = {1546-1726}, doi = {10.1038/s41593-020-0676-9}, author = {Garcia-Marques, Jorge and Isabel Espinosa Medina and Ku, Kai-Yuan and Yang, Ching-Po and Koyama, Minoru and Yu, Hung-Hsiang and Lee, Tzumin} } @article {65668, title = {Rational design of bioavailable photosensitizers for manipulation and imaging of biological systems.}, journal = {Cell Chemical Biology}, volume = {27}, year = {2020}, month = {2020 Aug 20}, pages = {1063-72}, abstract = {

Light-mediated chemical reactions are powerful methods for manipulating and interrogating biological systems. Photosensitizers, compounds that generate reactive oxygen species upon excitation with light, can be utilized for numerous biological experiments, but the repertoire of bioavailable photosensitizers is limited. Here, we describe the synthesis, characterization, and utility of two photosensitizers based upon the widely used rhodamine scaffold and demonstrate their efficacy for chromophore-assisted light inactivation, cell ablation in culture and in\ vivo, and photopolymerization of diaminobenzidine for electron microscopy. These chemical tools will facilitate a broad range of applications spanning from targeted destruction of proteins to high-resolution imaging.

}, issn = {2451-9448}, doi = {10.1016/j.chembiol.2020.07.001}, author = {Binns, Thomas C and Ayala, Anthony X and Grimm, Jonathan B and Tkachuk, Ariana N and Castillon, Guillaume A and Phan, Sebastien and Zhang, Lixia and Brown, Timothy A and Liu, Zhe and Adams, Stephen R and Ellisman, Mark H and Koyama, Minoru and Lavis, Luke D} } @article {68428, title = {Single-Cell Transcriptomes Reveal Diverse Regulatory Strategies for Olfactory Receptor Expression and Axon Targeting.}, journal = {Curr Biol}, volume = {30}, year = {2020}, month = {04/2020}, pages = {1189-1198.e5}, abstract = {

The regulatory mechanisms by which neurons coordinate their physiology and connectivity are not well understood. The Drosophila olfactory receptor neurons (ORNs) provide an excellent system to investigate this question. Each ORN type expresses a unique olfactory receptor, or a combination thereof, and sends their axons to a stereotyped glomerulus. Using single-cell RNA sequencing, we identified 33 transcriptomic clusters for ORNs and mapped 20 to their glomerular types, demonstrating that transcriptomic clusters correspond well with anatomically and physiologically defined ORN types. Each ORN type expresses hundreds of transcription factors. Transcriptome-instructed genetic analyses revealed that (1) one broadly expressed transcription factor (Acj6) only regulates olfactory receptor expression in one ORN type and only wiring specificity in another type, (2) one type-restricted transcription factor (Forkhead) only regulates receptor expression, and (3) another type-restricted transcription factor (Unplugged) regulates both events. Thus, ORNs utilize diverse strategies and complex regulatory networks to coordinate their physiology and connectivity.

}, keywords = {Animals, Axons, Drosophila melanogaster, Drosophila Proteins, Forkhead Transcription Factors, Homeodomain Proteins, Nerve Tissue Proteins, Olfactory Receptor Neurons, POU Domain Factors, Receptors, Odorant, Single-Cell Analysis, Smell, Transcriptome}, issn = {1879-0445}, doi = {10.1016/j.cub.2020.01.049}, author = {Li, Hongjie and Li, Tongchao and Horns, Felix and Li, Jiefu and Xie, Qijing and Xu, Chuanyun and Wu, Bing and Kebschull, Justus M and McLaughlin, Colleen N and Kolluru, Sai Saroja and Jones, Robert C and Vacek, David and Xie, Anthony and Luginbuhl, David J and Quake, Stephen R and Luo, Liqun} } @article {65635, title = {Structure of human GABA receptor in an inactive state.}, journal = {Nature}, volume = {584}, year = {2020}, month = {2020 Aug 01}, pages = {304-9}, abstract = {

The human GABA receptor-a member of the class C family of G-protein-coupled receptors (GPCRs)-mediates inhibitory neurotransmission and has been implicated in epilepsy, pain and addiction. A unique GPCR that is known to require heterodimerization for function, the GABA receptor has two subunits, GABA and GABA, that are structurally homologous but perform distinct and complementary functions. GABA recognizes orthosteric ligands, while GABA couples with G proteins. Each subunit is characterized by an extracellular Venus flytrap (VFT) module, a descending peptide linker, a seven-helix transmembrane domain and a cytoplasmic tail. Although the VFT heterodimer structure has been resolved, the structure of the full-length receptor and its transmembrane signalling mechanism remain unknown. Here we present a near full-length structure of the GABA receptor at atomic resolution, captured in an inactive state by cryo-electron microscopy. Our structure reveals several ligands that preassociate with the receptor, including two large endogenous phospholipids that are embedded within the transmembrane domains to maintain receptor integrity and modulate receptor function. We also identify a previously unknown heterodimer interface between transmembrane helices 3 and 5 of both subunits, which serves as a signature of the inactive conformation. A unique {\textquoteright}intersubunit latch{\textquoteright} within this transmembrane interface maintains the inactive state, and its disruption leads to constitutive receptor activity.

}, issn = {1476-4687}, doi = {10.1038/s41586-020-2452-0}, author = {Park, Jinseo and Fu, Ziao and Frangaj, Aurel and Liu, Jonathan and Mosyak, Lidia and Shen, Tong and Slavkovich, Vesna N and Ray, Kimberly M and Taura, Jaume and Cao, Baohua and Geng, Yong and Zuo, Hao and Kou, Yongjun and Grassucci, Robert and Chen, Shaoxia and Liu, Zheng and Lin, Xin and Williams, Justin P and Rice, William J and Eng, Edward T and Huang, Rick K and Soni, Rajesh K and Kloss, Brian and Yu, Zhiheng and Javitch, Jonathan A and Hendrickson, Wayne A and Slesinger, Paul A and Quick, Matthias and Graziano, Joseph and Yu, Hongtao and Fiehn, Oliver and Clarke, Oliver B and Frank, Joachim and Fan, Qing R} } @article {65855, title = {Super-resolution ophthalmoscopy: Virtually structured detection for resolution improvement in retinal imaging.}, journal = {Experimental Biology and Medicine (Maywood)}, year = {2020}, month = {2020 Nov 27}, pages = {1535370220970533}, abstract = {

Quantitative retinal imaging is essential for advanced study and clinical management of eye diseases. However, spatial resolution of retinal imaging has been limited due to available numerical aperture and optical aberration of the ocular optics. Structured illumination microscopy has been established to break the diffraction-limit resolution in conventional light microscopy. However, practical implementation of structured illumination microscopy for ophthalmoscopy of the retina is challenging due to inevitable eye movements that can produce phase artifacts. Recently, we have demonstrated the feasibility of using virtually structured detection as one alternative to structured illumination microscopy for super-resolution imaging. By providing the flexibility of digital compensation of eye movements, the virtually structured detection provides a feasible, phase-artifact-free strategy to achieve super-resolution ophthalmoscopy. In this article, we summarize the technical rationale of virtually structured detection, and its implementations for super-resolution imaging of freshly isolated retinas, intact animals, and awake human subjects.

}, issn = {1535-3699}, doi = {10.1177/1535370220970533}, author = {Yao, Xincheng and Lu, Rongwen and Wang, Benquan and Lu, Yiming and Kim, Tae-Hoon} } @article {65723, title = {A systematic nomenclature for the Drosophila ventral nerve cord.}, journal = {Neuron}, volume = {107}, year = {2020}, month = {2020 Sep 14}, pages = {1071-79}, abstract = {

Drosophila melanogaster is an established model for neuroscience research with relevance in biology and medicine. Until recently, research on the Drosophila brain was hindered by the lack of a complete and uniform nomenclature. Recognizing this, Ito et\ al. (2014) produced an authoritative nomenclature for the adult insect brain, using Drosophila as the reference. Here, we extend this nomenclature to the adult thoracic and abdominal neuromeres, the ventral nerve cord (VNC), to provide an anatomical description of this major component of the Drosophila nervous system. The VNC is the locus for the reception and integration of sensory information and involved in generating most of the locomotor actions that underlie fly behaviors. The aim is to create a nomenclature, definitions, and spatial boundaries for the Drosophila VNC that are consistent with other insects. The work establishes an anatomical framework that provides a powerful tool for analyzing the functional organization of the VNC.

}, issn = {1097-4199}, doi = {10.1016/j.neuron.2020.08.005}, author = {Court, Robert and Namiki, Shigehiro and Armstrong, J Douglas and Borner, Jana and Card, Gwyneth and Costa, Marta and Dickinson, Michael and Duch, Carsten and Korff, Wyatt and Mann, Richard and Merritt, David and Murphey, Rod K and Seeds, Andrew M and Shirangi, Troy and Simpson, Julie H and Truman, James W and Tuthill, John C and Williams, Darren W and Shepherd, David} } @article {65253, title = {Tissue clearing and its applications in neuroscience}, journal = {Nature Reviews Neuroscience}, year = {2020}, month = {2020 Feb 1}, abstract = {

State-of-the-art tissue-clearing methods provide subcellular-level optical access to intact tissues from individual organs and even to some entire mammals. When combined with light-sheet microscopy and automated approaches to image analysis, existing tissue-clearing methods can speed up and may reduce the cost of conventional histology by several orders of magnitude. In addition, tissue-clearing chemistry allows whole-organ antibody labelling, which can be applied even to thick human tissues. By combining the most powerful labelling, clearing, imaging and data-analysis tools, scientists are extracting structural and functional cellular and subcellular information on complex mammalian bodies and large human specimens at an accelerated pace. The rapid generation of terabyte-scale imaging data furthermore creates a high demand for efficient computational approaches that tackle challenges in large-scale data analysis and management. In this Review, we discuss how tissue-clearing methods could provide an unbiased, system-level view of mammalian bodies and human specimens and discuss future opportunities for the use of these methods in human neuroscience.

}, issn = {1471-003X}, doi = {10.1038/s41583-019-0250-1}, url = {http://www.nature.com/articles/s41583-019-0250-1}, author = {Ueda, Hiroki R. and Ert{\"u}rk, Ali and Chung, Kwanghun and Gradinaru, Viviana and Ch{\'e}dotal, Alain and Tomancak, Pavel and Keller, Philipp J.} } @article {65264, title = {Tissue clearing and its applications in~neuroscience.}, journal = {Nature Reviews Neuroscience}, volume = {21}, year = {2020}, month = {2020 Jan 02}, pages = {61-79}, abstract = {

State-of-the-art tissue-clearing methods provide subcellular-level optical access to intact tissues from individual organs and even to some entire mammals. When combined with light-sheet microscopy and automated approaches to image analysis, existing tissue-clearing methods can speed up and may reduce the cost of conventional histology by several orders of magnitude. In addition, tissue-clearing chemistry allows whole-organ antibody labelling, which can be applied even to thick human tissues. By combining the most powerful labelling, clearing, imaging and data-analysis tools, scientists are extracting structural and functional cellular and subcellular information on complex mammalian bodies and large human specimens at an accelerated pace. The rapid generation of terabyte-scale imaging data furthermore creates a high demand for efficient computational approaches that tackle challenges in large-scale data analysis and management. In this Review, we discuss how tissue-clearing methods could provide an unbiased, system-level view of mammalian bodies and human specimens and discuss future opportunities for the use of these methods in human neuroscience.

}, issn = {1471-0048}, doi = {10.1038/s41583-019-0250-1}, author = {Ueda, Hiroki R and Ert{\"u}rk, Ali and Chung, Kwanghun and Gradinaru, Viviana and Ch{\'e}dotal, Alain and Tomancak, Pavel and Keller, Philipp J} } @article {65603, title = {Ultrastructural visualization of 3D chromatin folding using volume electron microscopy and DNA in situ hybridization.}, journal = {Nature Communications}, volume = {11}, year = {2020}, month = {2020 May 01}, pages = {2120}, abstract = {

The human genome is extensively folded into 3-dimensional organization. However, the detailed 3D chromatin folding structures have not been fully visualized due to the lack of robust and ultra-resolution imaging capability. Here, we report the development of an electron microscopy method that combines serial block-face scanning electron microscopy with in situ hybridization (3D-EMISH) to visualize 3D chromatin folding at targeted genomic regions with ultra-resolution (5 {\texttimes} 5 {\texttimes} 30 nm in xyz dimensions) that is superior to the current super-resolution by fluorescence light microscopy. We apply 3D-EMISH to human lymphoblastoid cells at a 1.7 Mb segment of the genome and visualize a large number of distinctive 3D chromatin folding structures in ultra-resolution. We further quantitatively characterize the reconstituted chromatin folding structures by identifying sub-domains, and uncover a high level heterogeneity of chromatin folding ultrastructures in individual nuclei, suggestive of extensive dynamic fluidity in 3D chromatin states.

}, issn = {2041-1723}, doi = {10.1038/s41467-020-15987-2}, author = {Trzaskoma, Pawe{\l} and Ruszczycki, B{\l}a{\.z}ej and Lee, Byoungkoo and Pels, Katarzyna K and Krawczyk, Katarzyna and Bokota, Grzegorz and Szczepankiewicz, Andrzej A and Aaron, Jesse and Walczak, Agnieszka and {\'S}liwi{\'n}ska, Ma{\l}gorzata A and Magalska, Adriana and Kadlof, Michal and Wolny, Artur and Parteka, Zofia and Arabasz, Sebastian and Kiss-Arabasz, Magdalena and Plewczy{\'n}ski, Dariusz and Ruan, Yijun and Wilczy{\'n}ski, Grzegorz M} } @article {65599, title = {Whole-brain profiling of cells and circuits in mammals by tissue clearing and light-sheet microscopy.}, journal = {Neuron}, volume = {106}, year = {2020}, month = {2020 May 06}, pages = {369-387}, abstract = {

Tissue clearing and light-sheet microscopy have a 100-year-plus history, yet these fields have been combined only recently to facilitate novel experiments and measurements in neuroscience. Since tissue-clearing methods were first combined with modernized light-sheet microscopy a decade ago, the performance of both technologies has rapidly improved, broadening their applications. Here, we review the state of the art of tissue-clearing methods and light-sheet microscopy and discuss applications of these techniques in profiling cells and circuits in mice. We examine outstanding challenges and future opportunities for expanding these techniques to achieve brain-wide profiling of cells and circuits in primates and humans. Such integration will help provide a systems-level understanding of the physiology and pathology of our central nervous system.

}, issn = {1097-4199}, doi = {10.1016/j.neuron.2020.03.004}, author = {Ueda, Hiroki R and Dodt, Hans-Ulrich and Osten, Pavel and Economo, Michael N and Chandrashekar, Jayaram and Keller, Philipp J} } @article {65633, title = {Whole-brain profiling of cells and circuits in mammals by tissue clearing and light-sheet microscopy.}, journal = {Neuron}, volume = {106}, year = {2020}, month = {2020 May 06}, pages = {369-387}, abstract = {

Tissue clearing and light-sheet microscopy have a 100-year-plus history, yet these fields have been combined only recently to facilitate novel experiments and measurements in neuroscience. Since tissue-clearing methods were first combined with modernized light-sheet microscopy a decade ago, the performance of both technologies has rapidly improved, broadening their applications. Here, we review the state of the art of tissue-clearing methods and light-sheet microscopy and discuss applications of these techniques in profiling cells and circuits in mice. We examine outstanding challenges and future opportunities for expanding these techniques to achieve brain-wide profiling of cells and circuits in primates and humans. Such integration will help provide a systems-level understanding of the physiology and pathology of our central nervous system.

}, issn = {1097-4199}, doi = {10.1016/j.neuron.2020.03.004}, author = {Ueda, Hiroki R and Dodt, Hans-Ulrich and Osten, Pavel and Economo, Michael N and Chandrashekar, Jayaram and Keller, Philipp J} } @article {65656, title = {The wiring logic of an identified serotonergic neuron that spans sensory networks.}, journal = {Journal of Neuroscience}, volume = {40}, year = {2020}, month = {2020 Aug 12}, pages = {6309-27}, abstract = {

Serotonergic neurons project widely throughout the brain to modulate diverse physiological and behavioral processes. However, a single cell resolution understanding of the connectivity of serotonergic neurons is currently lacking. Using a whole-brain EM dataset of a female , we comprehensively determine the wiring logic of a broadly projecting serotonergic neuron (the "CSDn") that spans several olfactory regions. Within the antennal lobe, the CSDn differentially innervates each glomerulus, yet surprisingly, this variability reflects a diverse set of presynaptic partners, rather than glomerulus-specific differences in synaptic output, which is predominately to local interneurons. Moreover, the CSDn has distinct connectivity relationships with specific local interneuron subtypes, suggesting that the CSDn influences distinct aspects of local network processing. Across olfactory regions, the CSDn has different patterns of connectivity, even having different connectivity with individual projection neurons that also span these regions. Whereas the CSDn targets inhibitory local neurons in the antennal lobe, the CSDn has more distributed connectivity in the LH, preferentially synapsing with principal neuron types based on transmitter content. Lastly, we identify individual novel synaptic partners associated with other sensory domains that provide strong, top-down input to the CSDn. Taken together, our study reveals the complex connectivity of serotonergic neurons which combine the integration of local and extrinsic synaptic input in a nuanced, region-specific manner.All sensory systems receive serotonergic modulatory input. However, a comprehensive understanding of the synaptic connectivity of individual serotonergic neurons is lacking. In this study, we use a whole-brain EM microscopy dataset to comprehensively determine the wiring logic of a broadly projecting serotonergic neuron in the olfactory system of Collectively, our study demonstrates at a single cell level, the complex connectivity of serotonergic neurons within their target networks, identifies specific cell classes heavily targeted for serotonergic modulation in the olfactory system, and reveals novel extrinsic neurons that provide strong input to this serotonergic system outside of the context of olfaction. Elucidating the connectivity logic of individual modulatory neurons provides a ground plan for the seemingly heterogeneous effects of modulatory systems.

}, issn = {1529-2401}, doi = {10.1523/JNEUROSCI.0552-20.2020}, author = {Coates, Kaylynn E and Calle-Schuler, Steven A and Helmick, Levi M and Knotts, Victoria L and Martik, Brennah N and Salman, Farzaan and Warner, Lauren T and Valla, Sophia V and Bock, Davi D and Dacks, Andrew M} } @article {65296, title = {Wiring patterns from auditory sensory neurons to the escape and song-relay pathways in fruit flies.}, journal = {Journal of Comparative Neurology}, volume = {528}, year = {2020}, month = {2020 Aug 01}, pages = {2068}, abstract = {

Many animals rely on acoustic cues to decide what action to take next. Unraveling the wiring patterns of the auditory neural pathways is prerequisite for understanding such information processing. Here we reconstructed the first step of the auditory neural pathway in the fruit fly brain, from primary to secondary auditory neurons, at the resolution of transmission electron microscopy. By tracing axons of two major subgroups of auditory sensory neurons in fruit flies, low-frequency tuned Johnston{\textquoteright}s organ (JO)-B neurons and high-frequency tuned JO-A neurons, we observed extensive connections from JO-B neurons to the main second-order neurons in both the song-relay and escape pathways. In contrast, JO-A neurons connected strongly to a neuron in the escape pathway. Our findings suggest that heterogeneous JO neuronal populations could be recruited to modify escape behavior whereas only specific JO neurons contribute to courtship behavior. We also found that all JO neurons have postsynaptic sites at their axons. Presynaptic modulation at the output sites of JO neurons could affect information processing of the auditory neural pathway in flies. This article is protected by copyright. All rights reserved.

}, issn = {1096-9861}, doi = {10.1002/cne.24877}, author = {Kim, Hyunsoo and Horigome, Mihoko and Ishikawa, Yuki and Li, Feng and Lauritzen, J Scott and Card, Gwyneth and Bock, Davi D and Kamikouchi, Azusa} } @article {64890, title = {Amino acid signatures of HLA Class-I and II molecules are strongly associated with SLE susceptibility and autoantibody production in Eastern Asians.}, journal = {PLoS Genetics}, volume = {15}, year = {2019}, month = {2019 Apr 25}, pages = {e1008092}, abstract = {

Human leukocyte antigen (HLA) is a key genetic factor conferring risk of systemic lupus erythematosus (SLE), but precise independent localization of HLA effects is extremely challenging. As a result, the contribution of specific HLA alleles and amino-acid residues to the overall risk of SLE and to risk of specific autoantibodies are far from completely understood. Here, we dissected (a) overall SLE association signals across HLA, (b) HLA-peptide interaction, and (c) residue-autoantibody association. Classical alleles, SNPs, and amino-acid residues of eight HLA genes were imputed across 4,915 SLE cases and 13,513 controls from Eastern Asia. We performed association followed by conditional analysis across HLA, assessing both overall SLE risk and risk of autoantibody production. DR15 alleles HLA-DRB1*15:01 (P = 1.4x10-27, odds ratio (OR) = 1.57) and HLA-DQB1*06:02 (P = 7.4x10-23, OR = 1.55) formed the most significant haplotype (OR = 2.33). Conditioned protein-residue signals were stronger than allele signals and mapped predominantly to HLA-DRB1 residue 13 (P = 2.2x10-75) and its proxy position 11 (P = 1.1x10-67), followed by HLA-DRB1-37 (P = 4.5x10-24). After conditioning on HLA-DRB1, novel associations at HLA-A-70 (P = 1.4x10-8), HLA-DPB1-35 (P = 9.0x10-16), HLA-DQB1-37 (P = 2.7x10-14), and HLA-B-9 (P = 6.5x10-15) emerged. Together, these seven residues increased the proportion of explained heritability due to HLA to 2.6\%. Risk residues for both overall disease and hallmark autoantibodies (i.e., nRNP: DRB1-11, P = 2.0x10-14; DRB1-13, P = 2.9x10-13; DRB1-30, P = 3.9x10-14) localized to the peptide-binding groove of HLA-DRB1. Enrichment for specific amino-acid characteristics in the peptide-binding groove correlated with overall SLE risk and with autoantibody presence. Risk residues were in primarily negatively charged side-chains, in contrast with rheumatoid arthritis. We identified novel SLE signals in HLA Class I loci (HLA-A, HLA-B), and localized primary Class II signals to five residues in HLA-DRB1, HLA-DPB1, and HLA-DQB1. These findings provide insights about the mechanisms by which the risk residues interact with each other to produce autoantibodies and are involved in SLE pathophysiology.

}, issn = {1553-7404}, doi = {10.1371/journal.pgen.1008092}, author = {Molineros, Julio E and Looger, Loren L and Kim, Kwangwoo and Okada, Yukinori and Terao, Chikashi and Sun, Celi and Zhou, Xu-Jie and Raj, Prithvi and Kochi, Yuta and Suzuki, Akari and Akizuki, Shuji and Nakabo, Shuichiro and Bang, So-Young and Lee, Hye-Soon and Kang, Young Mo and Suh, Chang-Hee and Chung, Won Tae and Park, Yong-Beom and Choe, Jung-Yoon and Shim, Seung-Cheol and Lee, Shin-Seok and Zuo, Xiaoxia and Yamamoto, Kazuhiko and Li, Quan-Zhen and Shen, Nan and Porter, Lauren L and Harley, John B and Chua, Kek Heng and Zhang, Hong and Wakeland, Edward K and Tsao, Betty P and Bae, Sang-Cheol and Nath, Swapan K} } @article {65198, title = {A bidirectional network for appetite control in larval zebrafish.}, journal = {Elife}, volume = {8}, year = {2019}, month = {2019 Oct 18}, abstract = {

Medial and lateral hypothalamic loci are known to suppress and enhance appetite, respectively, but the dynamics and functional significance of their interaction have yet to be explored. Here we report that, in larval zebrafish, primarily serotonergic neurons of the ventromedial caudal hypothalamus (cH) become increasingly active during food deprivation, whereas activity in the lateral hypothalamus (LH) is reduced. Exposure to food sensory and consummatory cues reverses the activity patterns of these two nuclei, consistent with their representation of opposing internal hunger states. Baseline activity is restored as food-deprived animals return to satiety via voracious feeding. The antagonistic relationship and functional importance of cH and LH activity patterns were confirmed by targeted stimulation and ablation of cH neurons. Collectively, the data allow us to propose a model in which these hypothalamic nuclei regulate different phases of hunger and satiety and coordinate energy balance via antagonistic control of distinct behavioral outputs.

}, issn = {2050-084X}, doi = {10.7554/eLife.43775}, author = {Wee, Caroline Lei and Song, Erin Yue and Johnson, Robert Evan and Ailani, Deepak and Randlett, Owen and Kim, Ji-Yoon and Nikitchenko, Maxim and Bahl, Armin and Yang, Chao-Tsung and Ahrens, Misha B and Kawakami, Koichi and Engert, Florian and Kunes, Sam} } @article {65079, title = {BigStitcher: reconstructing high-resolution image datasets of cleared and expanded samples.}, journal = {Nature Methods}, volume = {16}, year = {2019}, month = {2019 Sep}, pages = {870-74}, abstract = {

Light-sheet imaging of cleared and expanded samples creates terabyte-sized datasets that consist of many unaligned three-dimensional image tiles, which must be reconstructed before analysis. We developed the BigStitcher software to address this challenge. BigStitcher enables interactive visualization, fast and precise alignment, spatially resolved quality estimation, real-time fusion and deconvolution of dual-illumination, multitile, multiview datasets. The software also compensates for optical effects, thereby improving accuracy and enabling subsequent biological analysis.

}, issn = {1548-7091}, doi = {10.1038/s41592-019-0501-0}, url = {http://www.nature.com/articles/s41592-019-0501-0http://www.nature.com/articles/s41592-019-0501-0.pdfhttp://www.nature.com/articles/s41592-019-0501-0.pdfhttp://www.nature.com/articles/s41592-019-0501-0}, author = {H{\"o}rl, David and Rojas Rusak, Fabio and Preusser, Friedrich and Tillberg, Paul and Randel, Nadine and Chhetri, Raghav K. and Cardona, Albert and Keller, Philipp J. and Harz, Hartmann and Leonhardt, Heinrich and Treier, Mathias and Preibisch, Stephan} } @article {65242, title = {Biosensors show the pharmacokinetics of S-Ketamine in the endoplasmic reticulum.}, journal = {Frontiers in Cellular Neuroscience}, volume = {13}, year = {2019}, month = {2019 Nov 12}, pages = {499}, abstract = {

The target for the "rapid" (\<24 h) antidepressant effects of S-ketamine is unknown, vitiating programs to rationally develop more effective rapid antidepressants. To describe a drug{\textquoteright}s target, one must first understand the compartments entered by the drug, at all levels-the organ, the cell, and the organelle. We have, therefore, developed molecular tools to measure the subcellular, organellar pharmacokinetics of S-ketamine. The tools are genetically encoded intensity-based S-ketamine-sensing fluorescent reporters, iSKetSnFR1 and iSKetSnFR2. In solution, these biosensors respond to S-ketamine with a sensitivity, S-slope = delta(F/F)/(delta[S-ketamine]) of 0.23 and 1.9/μM, respectively. The iSKetSnFR2 construct allows measurements at \<0.3 μM S-ketamine. The iSKetSnFR1 and iSKetSnFR2 biosensors display \>100-fold selectivity over other ligands tested, including R-ketamine. We targeted each of the sensors to either the plasma membrane (PM) or the endoplasmic reticulum (ER). Measurements on these biosensors expressed in Neuro2a cells and in human dopaminergic neurons differentiated from induced pluripotent stem cells (iPSCs) show that S-ketamine enters the ER within a few seconds after appearing in the external solution near the PM, then leaves as rapidly after S-ketamine is removed from the extracellular solution. In cells, S-slopes for the ER and PM-targeted sensors differ by \<2-fold, indicating that the ER [S-ketamine] is less than 2-fold different from the extracellular [S-ketamine]. Organelles represent potential compartments for the engagement of S-ketamine with its antidepressant target, and potential S-ketamine targets include organellar ion channels, receptors, and transporters.

}, issn = {1662-5102}, doi = {10.3389/fncel.2019.00499}, author = {Bera, Kallol and Kamajaya, Aron and Shivange, Amol V and Muthusamy, Anand K and Nichols, Aaron L and Borden, Philip M and Grant, Stephen and Jeon, Janice and Lin, Elaine and Bishara, Ishak and Chin, Theodore M and Cohen, Bruce N and Kim, Charlene H and Unger, Elizabeth K and Tian, Lin and Marvin, Jonathan S and Looger, Loren L and Lester, Henry A} } @article {65081, title = {Bright and photostable chemigenetic indicators for extended in vivo voltage imaging.}, journal = {Science}, volume = {365}, year = {2019}, month = {2019 Aug 13}, pages = {699-704}, abstract = {

Imaging changes in membrane potential using genetically encoded fluorescent voltage indicators (GEVIs) has great potential for monitoring neuronal activity with high spatial and temporal resolution. Brightness and photostability of fluorescent proteins and rhodopsins have limited the utility of existing GEVIs. We engineered a novel GEVI, "Voltron", that utilizes bright and photostable synthetic dyes instead of protein-based fluorophores, extending the combined duration of imaging and number of neurons imaged simultaneously by more than tenfold relative to existing GEVIs. We used Voltron for in vivo voltage imaging in mice, zebrafish, and fruit flies. In mouse cortex, Voltron allowed single-trial recording of spikes and subthreshold voltage signals from dozens of neurons simultaneously, over 15 min of continuous imaging. In larval zebrafish, Voltron enabled the precise correlation of spike timing with behavior.

}, issn = {1095-9203}, doi = {10.1126/science.aav6416}, author = {Abdelfattah, Ahmed S and Kawashima, Takashi and Singh, Amrita and Novak, Ondrej and Liu, Hui and Shuai, Yichun and Huang, Yi-Chieh and Campagnola, Luke and Seeman, Stephanie C and Yu, Jianing and Zheng, Jihong and Grimm, Jonathan B and Patel, Ronak and Friedrich, Johannes and Mensh, Brett D and Paninski, Liam and Macklin, John J and Murphy, Gabe J and Podgorski, Kaspar and Lin, Bei-Jung and Chen, Tsai-Wen and Turner, Glenn C and Liu, Zhe and Koyama, Minoru and Svoboda, Karel and Ahrens, Misha B and Lavis, Luke D and Schreiter, Eric R} } @article {67280, title = {CaImAn an open source tool for scalable calcium imaging data analysis}, journal = {eLife}, volume = {8}, year = {2019}, month = {01/2019}, abstract = {

Advances in fluorescence microscopy enable monitoring larger brain areas in-vivo with finer time resolution. The resulting data rates require reproducible analysis pipelines that are reliable, fully automated, and scalable to datasets generated over the course of months. We present CaImAn, an open-source library for calcium imaging data analysis. CaImAn\ provides automatic and scalable methods to address problems common to pre-processing, including motion correction, neural activity identification, and registration across different sessions of data collection. It does this while requiring minimal user intervention, with good scalability on computers ranging from laptops to high-performance computing clusters. CaImAn\ is suitable for two-photon and one-photon imaging, and also enables real-time analysis on streaming data. To benchmark the performance of CaImAn\ we collected and combined a corpus of manual annotations from multiple labelers on nine mouse two-photon datasets. We demonstrate that CaImAn\ achieves near-human performance in detecting locations of active neurons

}, doi = {10.7554/eLife.38173}, url = {https://elifesciences.org/articles/38173}, author = {Giovannucci, Andrea and Friedrich, Johannes and Gunn, Pat and Kalfon, J{\'e}r{\'e}mie and Brown, Brandon L and Koay, Sue Ann and Taxidis, Jiannis and Najafi, Farzaneh and Gauthier, Jeffrey L and Zhou, Pengcheng and Khakh, Baljit S and Tank, David W and Chklovskii, Dmitri B and Pnevmatikakis, Eftychios A} } @article {65137, title = {Can one concurrently record electrical spikes from every neuron in a mammalian brain?}, journal = {Neuron}, volume = {103}, year = {2019}, month = {2019 Sep 25}, pages = {1005}, abstract = {

The classic approach to measure the spiking response of neurons involves the use of metal electrodes to record extracellular potentials. Starting over 60 years ago with a single recording site, this technology now extends to ever larger numbers and densities of sites. We argue, based on the mechanical and electrical properties of existing materials, estimates of signal-to-noise ratios, assumptions regarding extracellular space in the brain, and estimates of heat generation by the electronic interface, that it should be possible to fabricate rigid electrodes to concurrently record from essentially every neuron in the cortical mantle. This will involve fabrication with existing yet nontraditional materials and procedures. We further emphasize the need to advance materials for improved flexible electrodes as an essential advance to record from neurons in brainstem and spinal cord in moving animals.

}, issn = {08966273}, doi = {10.1016/j.neuron.2019.08.011}, author = {Kleinfeld, David and Luan, Lan and Mitra, Partha P. and Robinson, Jacob T. and Sarpeshkar, Rahul and Shepard, Kenneth and Xie, Chong and Harris, Timothy D.} } @article {65058, title = {CHP1 regulates compartmentalized glycerolipid synthesis by activating GPAT4.}, journal = {Molecular Cell}, volume = {74}, year = {2019}, month = {2019 Apr 4}, pages = {45-58.e7}, abstract = {

Cells require a constant supply of fatty acids to survive and proliferate. Fatty acids incorporate into membrane and storage glycerolipids through a series of endoplasmic reticulum (ER) enzymes, but how these enzymes are regulated is not well understood. Here, using a combination of CRISPR-based genetic screens and unbiased lipidomics, we identified calcineurin B homologous protein 1 (CHP1) as a major regulator of ER glycerolipid synthesis. Loss of\ CHP1 severely reduces fatty acid incorporation and storage in mammalian cells and invertebrates. Mechanistically, CHP1 binds and activates GPAT4, which catalyzes the initial rate-limiting step in glycerolipid synthesis. GPAT4 activity requires CHP1 to be N-myristoylated, forming a key molecular interface between the two proteins. Interestingly, upon CHP1 loss, the peroxisomal enzyme, GNPAT, partially compensates for the loss of ER lipid synthesis, enabling cell proliferation. Thus, our work identifies a conserved regulator of glycerolipid metabolism and reveals plasticity in lipid synthesis of proliferating cells.

}, keywords = {3T3 Cells, Acyltransferases, Animals, Caenorhabditis elegans, Caenorhabditis elegans Proteins, Calcium-Binding Proteins, Cell Proliferation, Drosophila melanogaster, Drosophila Proteins, Endoplasmic Reticulum, Enzyme Activation, Gene Expression Regulation, Enzymologic, Glycerides, Glycerol-3-Phosphate O-Acyltransferase, HEK293 Cells, Hela Cells, Hep G2 Cells, Humans, Jurkat Cells, Lipogenesis, Mice, Palmitic Acid, Protein Binding}, issn = {1097-4164}, doi = {10.1016/j.molcel.2019.01.037}, author = {Zhu, Xiphias Ge and Nicholson Puthenveedu, Shirony and Shen, Yihui and La, Konnor and Ozlu, Can and Wang, Tim and Klompstra, Diana and Gultekin, Yetis and Chi, Jingyi and Fidelin, Justine and Peng, Tao and Molina, Henrik and Hang, Howard C and Min, Wei and Birsoy, K{\i}van{\c c}} } @article {64814, title = {Chronology-based architecture of descending circuits that underlie the development of locomotor repertoire after birth.}, journal = {eLife}, volume = {8}, year = {2019}, month = {2019 Feb 25}, abstract = {

The emergence of new and increasingly sophisticated behaviors after birth is accompanied by dramatic increase of newly established synaptic connections in the nervous system. Little is known, however, of how nascent connections are organized to support such new behaviors alongside existing ones. To understand this, in the larval zebrafish we examined the development of spinal pathways from hindbrain V2a neurons and the role of these pathways in the development of locomotion. We found that new projections are continually layered laterally to existing neuropil, and give rise to distinct pathways that function in parallel to existing pathways. Across these chronologically layered pathways, the connectivity patterns and biophysical properties vary systematically to support a behavioral repertoire with a wide range of kinematics and dynamics. Such layering of new parallel circuits equipped with systematically changing properties may be central to the postnatal diversification and increasing sophistication of an animal{\textquoteright}s behavioral repertoire.

}, issn = {2050-084X}, doi = {10.7554/eLife.42135}, author = {Pujala, Avinash and Koyama, Minoru} } @article {65379, title = {CLADES: a programmable sequence of reporters for lineage analysis}, journal = {bioRxiv}, year = {2019}, month = {2019 May 30}, abstract = {

We present CLADES (Cell Lineage Access Driven by an Edition Sequence), a technology for cell lineage studies based on CRISPR/Cas9. CLADES relies on a system of genetic switches to activate and inactivate reporter genes in a pre-determined order. Targeting CLADES to progenitor cells allows the progeny to inherit a sequential cascade of reporters, coupling birth order with reporter expression. This gives us temporal resolution of lineage development that can be used to deconstruct an extended cell lineage by tracking the reporters expressed in the progeny. When targeted to the germ line, the same cascade progresses across animal generations, marking each generation with the corresponding combination of reporters. CLADES thus offers an innovative strategy for making programmable cascades of genes that can be used for genetic manipulation or to record serial biological events.

}, doi = {https://doi.org/10.1101/655308}, author = {Garcia-Marques, Jorge and Yang, Ching-Po and Isabel Espinosa Medina and Koyama, Minoru and Lee, Tzumin} } @article {65082, title = {Comparing 3D ultrastructure of presynaptic and postsynaptic mitochondria.}, journal = {Biology Open}, volume = {8}, year = {2019}, month = {2019 Aug 01}, abstract = {

Serial-section electron microscopy such as FIB-SEM (focused ion beam scanning electron microscopy) has become an important tool for neuroscientists to trace the trajectories and global architecture of neural circuits in the brain, as well as to visualize the 3D ultrastructure of cellular organelles in neurons. In this study, we examined 3D features of mitochondria in electron microscope images generated from serial sections of four regions of mouse brains: nucleus accumbens (NA), hippocampal CA1, somatosensory cortex and dorsal cochlear nucleus (DCN). We compared mitochondria in the presynaptic terminals to those in the postsynaptic/dendritic compartments, and we focused on the shape and size of mitochondria. A common feature of mitochondria among the four brain regions is that presynaptic mitochondria generally are small and short, and most of them do not extend beyond presynaptic terminals. In contrast, the majority of postsynaptic/dendritic mitochondria are large and many of them spread through significant portions of the dendrites. Comparing among the brain areas, the cerebral cortex and DCN have even larger postsynaptic/dendritic mitochondria than the NA and CA1. Our analysis reveals that mitochondria in neurons are differentially sized and arranged according to their subcellular locations, suggesting a spatial organizing principle of mitochondria at the synapse.

}, issn = {2046-6390}, doi = {10.1242/bio.044834}, author = {Delgado, Thomas and Petralia, Ronald S and Freeman, David W and Sedlacek, Miloslav and Wang, Ya-Xian and Brenowitz, Stephan D and Sheu, Shu-Hsien and Gu, Jeffrey W and Kapogiannis, Dimitrios and Mattson, Mark P and Yao, Pamela J} } @article {64831, title = {Cytoskeletal actin patterns shape mast cell activation.}, journal = {Communications Biology}, volume = {2}, year = {2019}, month = {2019}, pages = {93}, abstract = {

Activation of immune cells relies on a dynamic actin cytoskeleton. Despite detailed knowledge of molecular actin assembly, the exact processes governing actin organization during activation remain elusive. Using advanced microscopy, we here show that Rat Basophilic Leukemia (RBL) cells, a model mast cell line, employ an orchestrated series of reorganization events within the cortical actin network during activation. In response to IgE antigen-stimulation of FCε receptors (FCεR) at the RBL cell surface, we observed symmetry breaking of the F-actin network and subsequent rapid disassembly of the actin cortex. This was followed by a reassembly process that may be driven by the coordinated transformation of distinct nanoscale F-actin architectures, reminiscent of self-organizing actin patterns. Actin patterns co-localized with zones of Arp2/3 nucleation, while network reassembly was accompanied by myosin-II activity. Strikingly, cortical actin disassembly coincided with zones of granule secretion, suggesting that cytoskeletal actin patterns contribute to orchestrate RBL cell activation.

}, issn = {2399-3642}, doi = {10.1038/s42003-019-0322-9}, author = {Colin-York, Huw and Li, Dong and Korobchevskaya, Kseniya and Chang, Veronica T and Betzig, Eric and Eggeling, Christian and Fritzsche, Marco} } @article {64852, title = {Cytoskeletal control of antigen-dependent T cell activation.}, journal = {Cell Reports}, volume = {26}, year = {2019}, month = {2019 Mar 19}, pages = {3369-3379.e5}, abstract = {

Cytoskeletal actin dynamics is essential for T\ cell activation. Here, we show evidence that the binding kinetics of the antigen engaging the T\ cell receptor influences the nanoscale actin organization and mechanics of the immune synapse. Using an engineered T\ cell system expressing a specific T\ cell receptor and stimulated by a range of antigens, we found that the peak force experienced by the T\ cell receptor during activation was independent of the unbinding kinetics of the stimulating antigen. Conversely, quantification of the actin retrograde flow velocity at the synapse revealed a striking dependence on the antigen unbinding kinetics. These findings suggest that the dynamics of the actin cytoskeleton actively adjusted to normalize the force experienced by the T\ cell receptor in an antigen-specific manner. Consequently, tuning actin dynamics in response to antigen kinetics may thus\ be a mechanism that allows T\ cells to adjust the lengthscale and timescale of T\ cell receptor signaling.

}, issn = {2211-1247}, doi = {10.1016/j.celrep.2019.02.074}, author = {Colin-York, Huw and Javanmardi, Yousef and Skamrahl, Mark and Kumari, Sudha and Chang, Veronica T and Khuon, Satya and Taylor, Aaron and Chew, Teng-Leong and Betzig, Eric and Moeendarbary, Emad and Cerundolo, Vincenzo and Eggeling, Christian and Fritzsche, Marco} } @article {64918, title = {De novo design of tunable, pH-driven conformational changes.}, journal = {Science}, volume = {364}, year = {2019}, month = {2019 May 17}, pages = {658-64}, abstract = {

The ability of naturally occurring proteins to change conformation in response to environmental changes is critical to biological function. Although there have been advances in the de novo design of stable proteins with a single, deep free-energy minimum, the design of conformational switches remains challenging. We present a general strategy to design pH-responsive protein conformational changes by precisely preorganizing histidine residues in buried hydrogen-bond networks. We design homotrimers and heterodimers that are stable above pH 6.5 but undergo cooperative, large-scale conformational changes when the pH is lowered and electrostatic and steric repulsion builds up as the network histidine residues become protonated. The transition pH and cooperativity can be controlled through the number of histidine-containing networks and the strength of the surrounding hydrophobic interactions. Upon disassembly, the designed proteins disrupt lipid membranes both in vitro and after being endocytosed in mammalian cells. Our results demonstrate that environmentally triggered conformational changes can now be programmed by de novo protein design.

}, doi = {10.1126/science.aav7897}, author = {Boyken, Scott E and Benhaim, Mark A and Busch, Florian and Jia, Mengxuan and Back, Matthew J and Choi, Heejun and Klima, Jason C and Chen, Zibo and Walkey, Carl and Mileant, Alexander and Sahasrabuddhe, Aniruddha and Wei, Kathy Y and Hodge, Edgar A and Byron, Sarah and Quijano-Rubio, Alfredo and Sankaran, Banumathi and King, Neil P and Lippincott-Schwartz, Jennifer and Wysocki, Vicki H and et al} } @article {64546, title = {Determining the pharmacokinetics of nicotinic drugs in the endoplasmic reticulum using biosensors.}, journal = {The Journal of General Physiology}, year = {2019}, month = {2019 Feb 04}, abstract = {

Nicotine dependence is thought to arise in part because nicotine permeates into the endoplasmic reticulum (ER), where it binds to nicotinic receptors (nAChRs) and begins an "inside-out" pathway that leads to up-regulation of nAChRs on the plasma membrane. However, the dynamics of nicotine entry into the ER are unquantified. Here, we develop a family of genetically encoded fluorescent biosensors for nicotine, termed iNicSnFRs. The iNicSnFRs are fusions between two proteins: a circularly permutated GFP and a periplasmic choline-/betaine-binding protein engineered to bind nicotine. The biosensors iNicSnFR3a and iNicSnFR3b respond to nicotine by increasing fluorescence at [nicotine] \<1 {\textmu}M, the concentration in the plasma and cerebrospinal fluid of a smoker. We target iNicSnFR3 biosensors either to the plasma membrane or to the ER and measure nicotine kinetics in HeLa, SH-SY5Y, N2a, and HEK293 cell lines, as well as mouse hippocampal neurons and human stem cell-derived dopaminergic neurons. In all cell types, we find that nicotine equilibrates in the ER within 10 s (possibly within 1 s) of extracellular application and leaves as rapidly after removal from the extracellular solution. The [nicotine] in the ER is within twofold of the extracellular value. We use these data to run combined pharmacokinetic and pharmacodynamic simulations of human smoking. In the ER, the inside-out pathway begins when nicotine becomes a stabilizing pharmacological chaperone for some nAChR subtypes, even at concentrations as low as \~{}10 nM. Such concentrations would persist during the 12 h of a typical smoker{\textquoteright}s day, continually activating the inside-out pathway by \>75\%. Reducing nicotine intake by 10-fold decreases activation to \~{}20\%. iNicSnFR3a and iNicSnFR3b also sense the smoking cessation drug varenicline, revealing that varenicline also permeates into the ER within seconds. Our iNicSnFRs enable optical subcellular pharmacokinetics for nicotine and varenicline during an early event in the inside-out pathway.

}, issn = {1540-7748}, doi = {10.1085/jgp.201812201}, author = {Shivange, Amol V and Borden, Philip M and Muthusamy, Anand K and Nichols, Aaron L and Bera, Kallol and Bao, Huan and Bishara, Ishak and Jeon, Janice and Mulcahy, Matthew J and Cohen, Bruce and O{\textquoteright}Riordan, Saidhbhe L and Kim, Charlene and Dougherty, Dennis A and Chapman, Edwin R and Marvin, Jonathan and Looger, Loren and Lester, Henry A} } @article {64783, title = {Determining the pharmacokinetics of nicotinic drugs in the endoplasmic reticulum using biosensors.}, journal = {The Journal of General Physiology}, volume = {151}, year = {2019}, month = {2019 Feb 04}, pages = {738-57}, abstract = {

Nicotine dependence is thought to arise in part because nicotine permeates into the endoplasmic reticulum (ER), where it binds to nicotinic receptors (nAChRs) and begins an "inside-out" pathway that leads to up-regulation of nAChRs on the plasma membrane. However, the dynamics of nicotine entry into the ER are unquantified. Here, we develop a family of genetically encoded fluorescent biosensors for nicotine, termed iNicSnFRs. The iNicSnFRs are fusions between two proteins: a circularly permutated GFP and a periplasmic choline-/betaine-binding protein engineered to bind nicotine. The biosensors iNicSnFR3a and iNicSnFR3b respond to nicotine by increasing fluorescence at [nicotine] \<1 {\textmu}M, the concentration in the plasma and cerebrospinal fluid of a smoker. We target iNicSnFR3 biosensors either to the plasma membrane or to the ER and measure nicotine kinetics in HeLa, SH-SY5Y, N2a, and HEK293 cell lines, as well as mouse hippocampal neurons and human stem cell-derived dopaminergic neurons. In all cell types, we find that nicotine equilibrates in the ER within 10 s (possibly within 1 s) of extracellular application and leaves as rapidly after removal from the extracellular solution. The [nicotine] in the ER is within twofold of the extracellular value. We use these data to run combined pharmacokinetic and pharmacodynamic simulations of human smoking. In the ER, the inside-out pathway begins when nicotine becomes a stabilizing pharmacological chaperone for some nAChR subtypes, even at concentrations as low as \~{}10 nM. Such concentrations would persist during the 12 h of a typical smoker{\textquoteright}s day, continually activating the inside-out pathway by \>75\%. Reducing nicotine intake by 10-fold decreases activation to \~{}20\%. iNicSnFR3a and iNicSnFR3b also sense the smoking cessation drug varenicline, revealing that varenicline also permeates into the ER within seconds. Our iNicSnFRs enable optical subcellular pharmacokinetics for nicotine and varenicline during an early event in the inside-out pathway.

}, issn = {1540-7748}, doi = {10.1085/jgp.201812201}, author = {Shivange, Amol V and Borden, Philip M and Muthusamy, Anand K and Nichols, Aaron L and Bera, Kallol and Bao, Huan and Bishara, Ishak and Jeon, Janice and Mulcahy, Matthew J and Cohen, Bruce and O{\textquoteright}Riordan, Saidhbhe L and Kim, Charlene and Dougherty, Dennis A and Chapman, Edwin R and Marvin, Jonathan and Looger, Loren and Lester, Henry A} } @article {68993, title = {Deterministic Somatic Cell Reprogramming Involves Continuous Transcriptional Changes Governed by Myc and Epigenetic-Driven Modules.}, journal = {Cell Stem Cell}, volume = {24}, year = {2019}, month = {02/2019}, pages = {328-341.e9}, abstract = {

The epigenetic dynamics of induced pluripotent stem cell (iPSC) reprogramming in correctly reprogrammed cells at high resolution and throughout the entire process remain largely undefined. Here, we characterize conversion of mouse fibroblasts into iPSCs using Gatad2a-Mbd3/NuRD-depleted and highly efficient reprogramming systems. Unbiased high-resolution profiling of dynamic changes in levels of gene expression, chromatin engagement, DNA accessibility, and DNA methylation were obtained. We identified two distinct and synergistic transcriptional modules that dominate successful reprogramming, which are associated with cell identity and biosynthetic genes. The pluripotency module is governed by dynamic alterations in epigenetic modifications to promoters and binding by Oct4, Sox2, and Klf4, but not Myc. Early DNA demethylation at certain enhancers prospectively marks cells fated to reprogram. Myc activity drives expression of the essential biosynthetic module and is associated with optimized changes in tRNA codon usage. Our functional validations highlight interweaved epigenetic- and Myc-governed essential reconfigurations that rapidly commission and propel deterministic reprogramming toward naive pluripotency.

}, keywords = {Animals, Cell Lineage, Cellular Reprogramming, Chromatin, Demethylation, Epigenesis, Genetic, Humans, Induced Pluripotent Stem Cells, Kruppel-Like Factor 4, Mice, Protein Binding, Proto-Oncogene Proteins c-myc, RNA, Transfer, Transcription Factors, Transcription, Genetic}, issn = {1875-9777}, doi = {10.1016/j.stem.2018.11.014}, author = {Zviran, Asaf and Mor, Nofar and Rais, Yoach and Gingold, Hila and Peles, Shani and Chomsky, Elad and Viukov, Sergey and Buenrostro, Jason D and Scognamiglio, Roberta and Weinberger, Leehee and Manor, Yair S and Krupalnik, Vladislav and Zerbib, Mirie and Hezroni, Hadas and Jaitin, Diego Adhemar and Larastiaso, David and Gilad, Shlomit and Benjamin, Sima and Gafni, Ohad and Mousa, Awni and Ayyash, Muneef and Sheban, Daoud and Bayerl, Jonathan and Aguilera-Castrejon, Alejandro and Massarwa, Rada and Maza, Itay and Hanna, Suhair and Stelzer, Yonatan and Ulitsky, Igor and Greenleaf, William J and Tanay, Amos and Trumpp, Andreas and Amit, Ido and Pilpel, Yitzhak and Novershtern, Noa and Hanna, Jacob H} } @article {65043, title = {Direct wavefront sensing enables functional imaging of infragranular axons and spines.}, journal = {Nature Methods}, volume = {16}, year = {2019}, month = {2019 Jul}, pages = {615-618}, abstract = {

We advance two-photon microscopy for near-diffraction-limited imaging up to 850 {\textmu}m below the pia in awake mice. Our approach combines direct wavefront sensing of light from a guidestar (formed by descanned fluorescence from Cy5.5-conjugated dextran in brain microvessels) with adaptive optics to compensate for tissue-induced aberrations in the wavefront. We achieve high signal-to-noise ratios in recordings of glutamate release from thalamocortical axons and calcium transients in spines of layer 5b basal dendrites during active tactile sensing.

}, issn = {1548-7105}, doi = {10.1038/s41592-019-0434-7}, author = {Liu, Rui and Li, Zengyi and Marvin, Jonathan S and Kleinfeld, David} } @article {64542, title = {DVID: Distributed versioned image-oriented dataservice.}, journal = {Frontiers in Neural Circuits}, volume = {13}, year = {2019}, month = {2019 Feb 05}, abstract = {

Open-source software development has skyrocketed in part due to community tools like github.com, which allows publication of code as well as the ability to create branches and push accepted modifications back to the original repository. As the number and size of EM-based datasets increases, the connectomics community faces similar issues when we publish snapshot data corresponding to a publication. Ideally, there would be a mechanism where remote collaborators could modify branches of the data and then flexibly reintegrate results via moderated acceptance of changes. The DVID system provides a web-based connectomics API and the first steps toward such a distributed versioning approach to EM-based connectomics datasets. Through its use as the central data resource for Janelia{\textquoteright}s FlyEM team, we have integrated the concepts of distributed versioning into reconstruction workflows, allowing support for proofreader training and segmentation experiments through branched, versioned data. DVID also supports persistence to a variety of storage systems from high-speed local SSDs to cloud-based object stores, which allows its deployment on laptops as well as large servers. The tailoring of the backend storage to each type of connectomics data leads to efficient storage and fast queries. DVID is freely available as open-source software with an increasing number of supported storage options.

}, doi = {10.3389/fncir.2019.00005}, author = {Katz, William T and Plaza, Stephen M} } @article {64889, title = {Dynamic super-resolution structured illumination imaging in the living brain.}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {116}, year = {2019}, month = {2019 Apr 26}, pages = {9586-91}, abstract = {

Cells in the brain act as components of extended networks. Therefore, to understand neurobiological processes in a physiological context, it is essential to study them in vivo. Super-resolution microscopy has spatial resolution beyond the diffraction limit, thus promising to provide structural and functional insights that are not accessible with conventional microscopy. However, to apply it to in vivo brain imaging, we must address the challenges of 3D imaging in an optically heterogeneous tissue that is constantly in motion. We optimized image acquisition and reconstruction to combat sample motion and applied adaptive optics to correcting sample-induced optical aberrations in super-resolution structured illumination microscopy (SIM) in vivo. We imaged the brains of live zebrafish larvae and mice and observed the dynamics of dendrites and dendritic spines at nanoscale resolution.

}, issn = {1091-6490}, doi = {10.1073/pnas.1819965116}, author = {Turcotte, Raphael and Liang, Yajie and Tanimoto, Masashi and Zhang, Qinrong and Li, Ziwei and Koyama, Minoru and Betzig, Eric and Ji, Na} } @article {64875, title = {Enabling cell-type-specific behavioral epigenetics in Drosophila: a modified high-yield INTACT method reveals the impact of social environment on the epigenetic landscape in dopaminergic neurons.}, journal = {BMC Biology}, volume = {17}, year = {2019}, month = {2019 Apr 10}, pages = {30}, abstract = {

BACKGROUND: Epigenetic mechanisms play fundamental roles in brain function and behavior and stressors such as social isolation can alter animal behavior via epigenetic mechanisms. However, due to cellular heterogeneity, identifying cell-type-specific epigenetic changes in the brain is challenging. Here, we report the first use of a modified isolation of nuclei tagged in specific cell type (INTACT) method in behavioral epigenetics of Drosophila melanogaster, a method we call mini-INTACT.

RESULTS: Using ChIP-seq on mini-INTACT purified dopaminergic nuclei, we identified epigenetic signatures in socially isolated and socially enriched Drosophila males. Social experience altered the epigenetic landscape in clusters of genes involved in transcription and neural function. Some of these alterations could be predicted by expression changes of four transcription factors and the prevalence of their binding sites in several clusters. These transcription factors were previously identified as activity-regulated genes, and their knockdown in dopaminergic neurons reduced the effects of social experience on sleep.

CONCLUSIONS: Our work enables the use of Drosophila as a model for cell-type-specific behavioral epigenetics and establishes that social environment shifts the epigenetic landscape in dopaminergic neurons. Four activity-related transcription factors are required in dopaminergic neurons for the effects of social environment on sleep.

}, issn = {1741-7007}, doi = {10.1186/s12915-019-0646-4}, author = {Agrawal, Pavan and Chung, Phuong and Heberlein, Ulrike and Kent, Clement} } @article {64958, title = {Evolution of Mechanisms that Control Mating in Drosophila Males.}, journal = {Cell Reports}, volume = {27}, year = {2019}, month = {2019 May 28}, pages = {2527-2536.e4}, abstract = {

Genetically wired neural mechanisms inhibit mating between species because even naive animals rarely mate with other species. These mechanisms can evolve through changes in expression or function of key genes in sensory pathways or central circuits. Gr32a is a gustatory chemoreceptor that, in D.\ melanogaster, is essential to inhibit interspecies courtship and sense quinine. Similar to D.\ melanogaster, we find that D.\ simulans Gr32a is expressed in foreleg tarsi, sensorimotor appendages that inhibit interspecies courtship, and it is required to sense quinine. Nevertheless, Gr32a is not required to inhibit interspecies mating by D.\ simulans males. However, and similar to its function in D.\ melanogaster, Ppk25, a member of the Pickpocket family, promotes conspecific courtship in D.\ simulans. Together, we have identified distinct evolutionary mechanisms underlying chemosensory control of taste and courtship in closely related Drosophila species.

}, issn = {2211-1247}, doi = {10.1016/j.celrep.2019.04.104}, author = {Ahmed, Osama M and Avila-Herrera, Aram and Tun, Khin May and Serpa, Paula H and Peng, Justin and Parthasarathy, Srinivas and Knapp, Jon-Michael and Stern, David L and Davis, Graeme W and Pollard, Katherine S and Shah, Nirao M} } @article {64934, title = {Functional and anatomical specificity in a higher olfactory centre.}, journal = {Elife}, volume = {8}, year = {2019}, month = {2019 May 21}, abstract = {

Most sensory systems are organized into parallel neuronal pathways that process distinct aspects of incoming stimuli. In the insect olfactory system, second order projection neurons target both the mushroom body, required for learning, and the lateral horn (LH), proposed to mediate innate olfactory behavior. Mushroom body neurons form a sparse olfactory population code, which is not stereotyped across animals. In contrast, odor coding in the LH remains poorly understood. We combine genetic driver lines, anatomical and functional criteria to show that the LH has ~1400 neurons and \>165 cell types. Genetically labeled LHNs have stereotyped odor responses across animals and on average respond to three times more odors than single projection neurons. LHNs are better odor categorizers than projection neurons, likely due to stereotyped pooling of related inputs. Our results reveal some of the principles by which a higher processing area can extract innate behavioral significance from sensory stimuli.

}, issn = {2050-084X}, doi = {10.7554/eLife.44590}, author = {Frechter, Shahar and Bates, Alexander Shakeel and Tootoonian, Sina and Dolan, Michael-John and Manton, James D and Jamasb, Arian Rokkum and Kohl, Johannes and Bock, Davi and Jefferis, Gregory Sxe} } @article {65205, title = {Functional clustering of dendritic activity during decision-making.}, journal = {Elife}, volume = {8}, year = {2019}, month = {2019 Oct 30}, abstract = {

The active properties of dendrites can support local nonlinear operations, but previous imaging and electrophysiological measurements have produced conflicting views regarding the prevalence and selectivity of local nonlinearities in vivo. We imaged calcium signals in pyramidal cell dendrites in the motor cortex of mice performing a tactile decision task. A custom microscope allowed us to image the soma and up to 300 μm of contiguous dendrite at 15 Hz, while resolving individual spines. New analysis methods were used to estimate the frequency and spatial scales of activity in dendritic branches and spines. The majority of dendritic calcium transients were coincident with global events. However, task-associated calcium signals in dendrites and spines were compartmentalized by dendritic branching and clustered within branches over approximately 10 μm. Diverse behavior-related signals were intermingled and distributed throughout the dendritic arbor, potentially supporting a large learning capacity in individual neurons.

}, issn = {2050-084X}, doi = {10.7554/eLife.46966}, author = {Kerlin, Aaron and Boaz, Mohar and Flickinger, Daniel and MacLennan, Bryan J and Dean, Matthew B and Davis, Courtney and Spruston, Nelson and Svoboda, Karel} } @article {65224, title = {Gas cluster ion beam SEM for imaging of large tissue samples with 10 nm isotropic resolution.}, journal = {Nature Methods}, volume = {17}, year = {2019}, month = {2020 Jan 01}, pages = {68-71}, abstract = {

We demonstrate gas cluster ion beam scanning electron microscopy (SEM), in which wide-area ion milling is performed on a series of thick tissue sections. This three-dimensional electron microscopy technique acquires datasets with \<10 nm isotropic resolution of each section, and these can then be stitched together to span the sectioned volume. Incorporating gas cluster ion beam SEM into existing single-beam and multibeam SEM workflows should be straightforward, increasing reliability while improving z resolution by a factor of three or more.

}, issn = {1548-7105}, doi = {10.1038/s41592-019-0641-2}, author = {Hayworth, Kenneth J and Peale, David and Januszewski, Micha{\l} and Knott, Graham W and Lu, Zhiyuan and Xu, C Shan and Hess, Harald F} } @article {64823, title = {GCIB-SEM: A path to 10 nm isotropic imaging of cubic millimeter volumes.}, journal = {bioRxiv}, year = {2019}, month = {2019 Feb 28}, abstract = {Focused Ion Beam Scanning Electron Microscopy (FIB-SEM) generates 3D datasets optimally suited for segmentation of cell ultrastructure and automated connectome tracing but is limited to small fields of view and is therefore incompatible with the new generation of ultrafast multibeam SEMs. In contrast, section-based techniques are multibeam-compatible but are limited in z-resolution making automatic segmentation of cellular ultrastructure difficult. Here we demonstrate a novel 3D electron microscopy technique, Gas Cluster Ion Beam SEM (GCIB-SEM), in which top-down, wide-area ion milling is performed on a series of thick sections, acquiring < 10 nm isotropic datasets of each which are then stitched together to span the full sectioned volume. Based on our results, incorporating GCIB-SEM into existing single beam and multibeam SEM workflows should be straightforward and should dramatically increase reliability while simultaneously improving z-resolution by a factor of 3 or more.}, doi = {10.1101/563239}, author = {Hayworth, Kenneth J and Peale, David R and Januszewski, Michal and Knott, Graham and Lu, Zhiyuan and Xu, C. Shan and Hess, Harald F} } @article {65234, title = {Generation of stable heading representations in diverse visual scenes.}, journal = {Nature}, volume = {576}, year = {2019}, month = {2019 Nov 20}, pages = {126-31}, abstract = {

Many animals rely on an internal heading representation when navigating in varied environments. How this representation is linked to the sensory cues that define different surroundings is unclear. In the fly brain, heading is represented by {\textquoteright}compass{\textquoteright} neurons that innervate a ring-shaped structure known as the ellipsoid body. Each compass neuron receives inputs from {\textquoteright}ring{\textquoteright} neurons that are selective for particular visual features; this combination provides an ideal substrate for the extraction of directional information from a visual scene. Here we combine two-photon calcium imaging and optogenetics in tethered flying flies with circuit modelling, and show how the correlated activity of compass and visual neurons drives plasticity, which flexibly transforms two-dimensional visual cues into a stable heading representation. We also describe how this plasticity enables the fly to convert a partial heading representation, established from orienting within part of a novel setting, into a complete heading representation. Our results provide mechanistic insight into the memory-related computations that are essential for flexible navigation in varied surroundings.

}, issn = {1476-4687}, doi = {10.1038/s41586-019-1767-1}, author = {Kim, Sung Soo and Hermundstad, Ann M and Romani, Sandro and Abbott, L F and Jayaraman, Vivek} } @article {68360, title = {Genetic Identification of Vagal Sensory Neurons That Control Feeding}, journal = {Cell}, volume = {179}, year = {2019}, month = {11/2019}, pages = {1129-1143.e23}, abstract = {

Summary Energy homeostasis requires precise measurement of the quantity and quality of ingested food. The vagus nerve innervates the gut and can detect diverse interoceptive cues, but the identity of the key sensory neurons and corresponding signals that regulate food intake remains unknown. Here, we use an approach for target-specific, single-cell RNA sequencing to generate a map of the vagal cell types that innervate the gastrointestinal tract. We show that unique molecular markers identify vagal neurons with distinct innervation patterns, sensory endings, and function. Surprisingly, we find that food intake is most sensitive to stimulation of mechanoreceptors in the intestine, whereas nutrient-activated mucosal afferents have no effect. Peripheral manipulations combined with central recordings reveal that intestinal mechanoreceptors, but not other cell types, potently and durably inhibit hunger-promoting AgRP neurons in the hypothalamus. These findings identify a key role for intestinal mechanoreceptors in the regulation of feeding.

}, keywords = {AgRP Neurons, chemogenetics, fiber photometry, Hypothalamus, Optogenetics, RNA sequencing, Satiation, stretch, vagal afferents, vagus nerve}, issn = {0092-8674}, doi = {https://doi.org/10.1016/j.cell.2019.10.031}, url = {https://www.sciencedirect.com/science/article/pii/S009286741931181X}, author = {Ling Bai and Sheyda Mesgarzadeh and Karthik S. Ramesh and Erica L. Huey and Yin Liu and Lindsay A. Gray and Tara J. Aitken and Yiming Chen and Lisa R. Beutler and Jamie S. Ahn and Linda Madisen and Hongkui Zeng and Mark A. Krasnow and Zachary A. Knight} } @article {65056, title = {A genetically encoded fluorescent sensor for in vivo imaging of GABA.}, journal = {Nature Methods}, volume = {16}, year = {2019}, month = {2019 Jul 15}, pages = {763-770}, abstract = {

Current techniques for monitoring GABA (γ-aminobutyric acid), the primary inhibitory neurotransmitter in vertebrates, cannot follow transients in intact neural circuits. To develop a GABA sensor, we applied the design principles used to create the fluorescent glutamate receptor iGluSnFR. We used a protein derived from a previously unsequenced Pseudomonas fluorescens strain and performed structure-guided mutagenesis and library screening to obtain intensity-based GABA sensing fluorescence reporter (iGABASnFR) variants. iGABASnFR is genetically encoded, detects GABA release evoked by electric stimulation of afferent fibers in acute brain slices and produces readily detectable fluorescence increases in vivo in mice and zebrafish. We applied iGABASnFR to track mitochondrial GABA content and its modulation by an anticonvulsant, swimming-evoked, GABA-mediated transmission in zebrafish cerebellum, GABA release events during interictal spikes and seizures in awake mice, and found that GABA-mediated tone decreases during isoflurane anesthesia.

}, issn = {1548-7105}, doi = {10.1038/s41592-019-0471-2}, author = {Marvin, Jonathan S and Shimoda, Yoshiteru and Magloire, Vincent and Leite, Marco and Kawashima, Takashi and Jensen, Thomas P and Kolb, Ilya and Knott, Erika L and Novak, Ondrej and Podgorski, Kaspar and Leidenheimer, Nancy J and Rusakov, Dmitri A and Ahrens, Misha B and Kullmann, Dimitri M and Looger, Loren L} } @article {64796, title = {A genetically encoded single-wavelength sensor for imaging cytosolic and cell surface ATP.}, journal = {Nature Communications}, volume = {10}, year = {2019}, month = {2019 Feb 12}, pages = {711}, abstract = {

Adenosine 5{\textquoteright} triphosphate (ATP) is a universal intracellular energy source and an evolutionarily ancient, ubiquitous extracellular signal in diverse species. Here, we report the generation and characterization of single-wavelength genetically encoded fluorescent sensors (iATPSnFRs) for imaging extracellular and cytosolic ATP from insertion of circularly permuted superfolder GFP into the epsilon subunit of FF-ATPase from Bacillus PS3. On the cell surface and within the cytosol, iATPSnFR responds to relevant ATP concentrations (30 μM to 3 mM) with fast increases in fluorescence. iATPSnFRs can be genetically targeted to specific cell types and sub-cellular compartments, imaged with standard light microscopes, do not respond to other nucleotides and nucleosides, and when fused with a red fluorescent protein function as ratiometric indicators. After careful consideration of their modest pH sensitivity, iATPSnFRs represent promising reagents for imaging ATP in the extracellular space and within cells during a variety of settings, and for further application-specific refinements.

}, issn = {2041-1723}, doi = {10.1038/s41467-019-08441-5}, author = {Lobas, Mark A and Tao, Rongkun and Nagai, Jun and Kronschl{\"a}ger, Mira T and Borden, Philip M and Marvin, Jonathan S and Looger, Loren L and Khakh, Baljit S} } @article {65020, title = {Genetics of intellectual disability in consanguineous families.}, journal = {Mol Psychiatry}, volume = {24}, year = {2019}, month = {2018 Jan 04}, pages = {1027-1039}, abstract = {

Autosomal recessive (AR) gene defects are the leading genetic cause of intellectual disability (ID) in countries with frequent parental consanguinity, which account for about 1/7th of the world population. Yet, compared to autosomal dominant de novo mutations, which are the predominant cause of ID in Western countries, the identification of AR-ID genes has lagged behind. Here, we report on whole exome and whole genome sequencing in 404 consanguineous predominantly Iranian families with two or more affected offspring. In 219 of these, we found likely causative variants, involving 77 known and 77 novel AR-ID (candidate) genes, 21 X-linked genes, as well as 9 genes previously implicated in diseases other than ID. This study, the largest of its kind published to date, illustrates that high-throughput DNA sequencing in consanguineous families is a superior strategy for elucidating the thousands of hitherto unknown gene defects underlying AR-ID, and it sheds light on their prevalence.

}, issn = {1476-5578}, doi = {10.1038/s41380-017-0012-2}, author = {Hu, Hao and Kahrizi, Kimia and Musante, Luciana and Fattahi, Zohreh and Herwig, Ralf and Hosseini, Masoumeh and Oppitz, Cornelia and Abedini, Seyedeh Sedigheh and Suckow, Vanessa and Larti, Farzaneh and Beheshtian, Maryam and Lipkowitz, Bettina and Akhtarkhavari, Tara and Mehvari, Sepideh and Otto, Sabine and Mohseni, Marzieh and Arzhangi, Sanaz and Jamali, Payman and Mojahedi, Faezeh and Taghdiri, Maryam and Papari, Elaheh and Soltani Banavandi, Mohammad Javad and Akbari, Saeide and Tonekaboni, Seyed Hassan and Dehghani, Hossein and Ebrahimpour, Mohammad Reza and Bader, Ingrid and Davarnia, Behzad and Cohen, Monika and Khodaei, Hossein and Albrecht, Beate and Azimi, Sarah and Zirn, Birgit and Bastami, Milad and Wieczorek, Dagmar and Bahrami, Gholamreza and Keleman, Krystyna and Vahid, Leila Nouri and Tzschach, Andreas and G{\"a}rtner, Jutta and Gillessen-Kaesbach, Gabriele and Varaghchi, Jamileh Rezazadeh and Timmermann, Bernd and Pourfatemi, Fatemeh and Jankhah, Aria and Chen, Wei and Nikuei, Pooneh and Kalscheuer, Vera M and Oladnabi, Morteza and Wienker, Thomas F and Ropers, Hans-Hilger and Najmabadi, Hossein} } @article {68757, title = {A high-content imaging approach to profile C. elegans embryonic development}, journal = {Development}, volume = {146}, year = {2019}, month = {04-2019}, abstract = {

The Caenorhabditis elegans embryo is an important model for analyzing mechanisms of cell fate specification and tissue morphogenesis. Sophisticated lineage-tracing approaches for analyzing embryogenesis have been developed but are labor intensive and do not naturally integrate morphogenetic readouts. To enable the rapid classification of developmental phenotypes, we developed a high-content method that employs two custom strains: a Germ Layer strain that expresses nuclear markers in the ectoderm, mesoderm and endoderm/pharynx; and a Morphogenesis strain that expresses markers labeling epidermal cell junctions and the neuronal cell surface. We describe a procedure that allows simultaneous live imaging of development in 80-100 embryos and provide a custom program that generates cropped, oriented image stacks of individual embryos to facilitate analysis. We demonstrate the utility of our method by perturbing 40 previously characterized developmental genes in variants of the two strains containing RNAi-sensitizing mutations. The resulting datasets yielded distinct, reproducible signature phenotypes for a broad spectrum of genes that are involved in cell fate specification and morphogenesis. In addition, our analysis provides new in vivo evidence for MBK-2 function in mesoderm fate specification and LET-381 function in elongation.

}, issn = {0950-1991}, doi = {10.1242/dev.174029}, url = {https://doi.org/10.1242/dev.174029}, author = {Wang, Shaohe and Ochoa, Stacy D. and Khaliullin, Renat N. and Gerson-Gurwitz, Adina and Hendel, Jeffrey M. and Zhao, Zhiling and Biggs, Ronald and Chisholm, Andrew D. and Desai, Arshad and Oegema, Karen and Green, Rebecca A.} } @article {64986, title = {High-performance calcium sensors for imaging activity in neuronal populations and microcompartments.}, journal = {Nature Methods}, volume = {16}, year = {2019}, month = {2019 Jun 17}, pages = {649-57}, abstract = {

Calcium imaging with genetically encoded calcium indicators (GECIs) is routinely used to measure neural activity in intact nervous systems. GECIs are frequently used in one of two different modes: to track activity in large populations of neuronal cell bodies, or to follow dynamics in subcellular compartments such as axons, dendrites and individual synaptic compartments. Despite major advances, calcium imaging is still limited by the biophysical properties of existing GECIs, including affinity, signal-to-noise ratio, rise and decay kinetics and dynamic range. Using structure-guided mutagenesis and neuron-based screening, we optimized the green fluorescent protein-based GECI GCaMP6 for different modes of in vivo imaging. The resulting jGCaMP7 sensors provide improved detection of individual spikes (jGCaMP7s,f), imaging in neurites and neuropil (jGCaMP7b), and may allow tracking larger populations of neurons using two-photon (jGCaMP7s,f) or wide-field (jGCaMP7c) imaging.

}, issn = {1548-7105}, doi = {10.1038/s41592-019-0435-6}, author = {Dana, Hod and Sun, Yi and Mohar, Boaz and Hulse, Brad K and Kerlin, Aaron M and Hasseman, Jeremy P and Tsegaye, Getahun and Tsang, Arthur and Wong, Allan and Patel, Ronak and Macklin, John J and Chen, Yang and Konnerth, Arthur and Jayaraman, Vivek and Looger, Loren L and Schreiter, Eric R and Svoboda, Karel and Kim, Douglas S} } @article {65041, title = {High-resolution imaging reveals how the spindle midzone impacts chromosome movement.}, journal = {The Journal of Cell Biology}, volume = {218}, year = {2019}, month = {27 Jun 2019}, pages = {2529-44}, abstract = {

In the spindle midzone, microtubules from opposite half-spindles form bundles between segregating chromosomes. Microtubule bundles can either push or restrict chromosome movement during anaphase in different cellular contexts, but how these activities are achieved remains poorly understood. Here, we use high-resolution live-cell imaging to analyze individual microtubule bundles, growing filaments, and chromosome movement in dividing human cells. Within bundles, filament overlap length marked by the cross-linking protein PRC1 decreases during anaphase as chromosome segregation slows. Filament ends within microtubule bundles appear capped despite dynamic PRC1 turnover and submicrometer proximity to growing microtubules. Chromosome segregation distance and rate are increased in two human cell lines when microtubule bundle assembly is prevented via PRC1 knockdown. Upon expressing a mutant PRC1 with reduced microtubule affinity, bundles assemble but chromosome hypersegregation is still observed. We propose that microtubule overlap length reduction, typically linked to pushing forces generated within filament bundles, is needed to properly restrict spindle elongation and position chromosomes within daughter cells.

}, issn = {0021-9525}, doi = {10.1083/jcb.201904169}, url = {http://www.jcb.org/lookup/doi/10.1083/jcb.201904169https://syndication.highwire.org/content/doi/10.1083/jcb.201904169}, author = {Pamula, Melissa C. and Carlini, Lina and Forth, Scott and Verma, Priyanka and Suresh, Subbulakshmi and Legant, Wesley R. and Khodjakov, Alexey and Betzig, Eric and Kapoor, Tarun M.} } @article {65204, title = {High-yield, automated intracellular electrophysiology in retinal pigment epithelia.}, journal = {Journal of Neuroscience Methods}, volume = {328}, year = {2019}, month = {2019 Dec 01}, pages = {108442}, abstract = {

BACKGROUND: Recent advancements with induced pluripotent stem cell-derived (iPSC) retinal pigment epithelium (RPE) have made disease modeling and cell therapy for macular degeneration feasible. However, current techniques for intracellular electrophysiology - used to validate epithelial function - are painstaking and require manual skill; limiting experimental throughput.

NEW METHOD: A five-stage algorithm, leveraging advances in automated patch clamping, systematically derived and optimized, improves yield and reduces skill when compared to conventional, manual techniques.

RESULTS: The automated algorithm improves yield per attempt from 17\% (manually, n = 23) to 22\% (automated, n = 120) (chi-squared, p = 0.004). Specifically for RPE, depressing the local cell membrane by 6 μm and electroporating (buzzing) just prior to this depth (5 μm) maximized yield.

COMPARISON WITH EXISTING METHOD: Conventionally, intracellular epithelial electrophysiology is performed by manually lowering a pipette with a micromanipulator, blindly, towards a monolayer of cells and spontaneously stopping when the magnitude of the instantaneous measured membrane potential decreased below a predetermined threshold. The new method automatically measures the pipette tip resistance during the descent, detects the cell surface, indents the cell membrane, and briefly buzzes to electroporate the membrane while descending, overall achieving a higher yield than conventional methods.

CONCLUSIONS: This paper presents an algorithm for high-yield, automated intracellular electrophysiology in epithelia; optimized for human RPE. Automation reduces required user skill and training while, simultaneously, improving yield. This algorithm could enable large-scale exploration of drug toxicity and physiological function verification for numerous kinds of epithelia.

}, issn = {1872-678X}, doi = {10.1016/j.jneumeth.2019.108442}, author = {Lewallen, Colby F and Wan, Qin and Maminishkis, Arvydas and Stoy, William and Kolb, Ilya and Hotaling, Nathan and Bharti, Kapil and Forest, Craig R} } @article {65175, title = {Histone H3K27 acetylation precedes active transcription during zebrafish zygotic genome activation as revealed by live-cell analysis.}, journal = {Development}, volume = {146}, year = {2019}, month = {2019 Sep 30}, abstract = {

Histone post-translational modifications are key gene expression regulators, but their rapid dynamics during development remain difficult to capture. We applied a Fab-based live endogenous modification labeling technique to monitor the changes in histone modification levels during zygotic genome activation (ZGA) in living zebrafish embryos. Among various histone modifications, H3 Lys27 acetylation (H3K27ac) exhibited most drastic changes, accumulating in two nuclear foci in the 64- to 1k-cell-stage embryos. The elongating form of RNA polymerase II, which is phosphorylated at Ser2 in heptad repeats within the C-terminal domain (RNAP2 Ser2ph), and miR-430 transcripts were also concentrated in foci closely associated with H3K27ac. When treated with α-amanitin to inhibit transcription or JQ-1 to inhibit binding of acetyl-reader proteins, H3K27ac foci still appeared but RNAP2 Ser2ph and miR-430 morpholino were not concentrated in foci, suggesting that H3K27ac precedes active transcription during ZGA. We anticipate that the method presented here could be applied to a variety of developmental processes in any model and non-model organisms.

}, issn = {1477-9129}, doi = {10.1242/dev.179127}, author = {Sato, Yuko and Hilbert, Lennart and Oda, Haruka and Wan, Yinan and Heddleston, John M and Chew, Teng-Leong and Zaburdaev, Vasily and Keller, Philipp and Lionnet, Timoth{\'e}e and Vastenhouw, Nadine and Kimura, Hiroshi} } @article {65178, title = {ilastik: interactive machine learning for (bio)image analysis.}, journal = {Nature Methods}, volume = {16}, year = {2019}, month = {2019 Sep 30}, pages = {1226-32}, abstract = {

We present ilastik, an easy-to-use interactive tool that brings machine-learning-based (bio)image analysis to end users without substantial computational expertise. It contains pre-defined workflows for image segmentation, object classification, counting and tracking. Users adapt the workflows to the problem at hand by interactively providing sparse training annotations for a nonlinear classifier. ilastik can process data in up to five dimensions (3D, time and number of channels). Its computational back end runs operations on-demand wherever possible, allowing for interactive prediction on data larger than RAM. Once the classifiers are trained, ilastik workflows can be applied to new data from the command line without further user interaction. We describe all ilastik workflows in detail, including three case studies and a discussion on the expected performance.

}, issn = {1548-7105}, doi = {10.1038/s41592-019-0582-9}, author = {Berg, Stuart and Kutra, Dominik and Kroeger, Thorben and Straehle, Christoph N and Kausler, Bernhard X and Haubold, Carsten and Schiegg, Martin and Ales, Janez and Beier, Thorsten and Rudy, Markus and Eren, Kemal and Cervantes, Jaime I and Xu, Buote and Beuttenmueller, Fynn and Wolny, Adrian and Zhang, Chong and Koethe, Ullrich and Hamprecht, Fred A and Kreshuk, Anna} } @article {65092, title = {Imaging striatal dopamine release using a nongenetically encoded near infrared fluorescent catecholamine nanosensor.}, journal = {Science Advances}, volume = {5}, year = {2019}, month = {2019 Jul 10}, pages = {eaaw3108}, abstract = {

Neuromodulation plays a critical role in brain function in both health and disease, and new tools that capture neuromodulation with high spatial and temporal resolution are needed. Here, we introduce a synthetic catecholamine nanosensor with fluorescent emission in the near infrared range (1000{\textendash}1300 nm), near infrared catecholamine nanosensor (nIRCat). We demonstrate that nIRCats can be used to measure electrically and optogenetically evoked dopamine release in brain tissue, revealing hotspots with a median size of 2 {\textmu}m. We also demonstrated that nIRCats are compatible with dopamine pharmacology and show D2 autoreceptor modulation of evoked dopamine release, which varied as a function of initial release magnitude at different hotspots. Together, our data demonstrate that nIRCats and other nanosensors of this class can serve as versatile synthetic optical tools to monitor neuromodulatory neurotransmitter release with high spatial resolution.

}, doi = {10.1126/sciadv.aaw3108}, url = {http://advances.sciencemag.org/lookup/doi/10.1126/sciadv.aaw3108https://syndication.highwire.org/content/doi/10.1126/sciadv.aaw3108}, author = {Beyene, Abraham G. and Delevich, Kristen and Del Bonis-O{\textquoteright}Donnell, Jackson Travis and Piekarski, David J. and Lin, Wan Chen and Thomas, A. Wren and Yang, Sarah J. and Kosillo, Polina and Yang, Darwin and Prounis, George S. and Wilbrecht, Linda and Landry, Markita P.} } @article {64866, title = {In vivo glucose imaging in multiple model organisms with an engineered single-wavelength sensor.}, journal = {bioRxiv}, year = {2019}, month = {2019 Mar 8}, abstract = {Glucose is arguably the most important molecule in metabolism, and its mismanagement underlies diseases of vast societal import, most notably diabetes. Although glucose-related metabolism has been the subject of intense study for over a century, tools to track glucose in living organisms with high spatio-temporal resolution are lacking. We describe the engineering of a family of genetically encoded glucose sensors with high signal-to-noise ratio, fast kinetics and affinities varying over four orders of magnitude (1 {\textmu}M to 10 mM). The sensors allow rigorous mechanistic characterization of glucose transporters expressed in cultured cells with high spatial and temporal resolution. Imaging of neuron/glia co-cultures revealed \~{}3-fold higher glucose changes in astrocytes versus neurons. In larval Drosophila central nervous system explants, imaging of intracellular neuronal glucose suggested a novel rostro-caudal transport pathway in the ventral nerve cord neuropil, with paradoxically slower uptake into the peripheral cell bodies and brain lobes. In living zebrafish, expected glucose-related physiological sequelae of insulin and epinephrine treatments were directly visualized in real time. Additionally, spontaneous muscle twitches induced glucose uptake in muscle, and sensory- and pharmacological perturbations gave rise to large but enigmatic changes in the brain. These sensors will enable myriad experiments, most notably rapid, high-resolution imaging of glucose influx, efflux, and metabolism in behaving animals.}, doi = {10.1101/571422}, author = {Keller, Jacob P and Marvin, Jonathan S and Lacin, Haluk and Lemon, William C and Shea, Jamien and Kim, Soomin and Lee, Richard T and Koyama, Minoru and Keller, Philipp J and Looger, Loren L} } @article {65212, title = {Interactions between Dpr11 and DIP-γ control selection of amacrine neurons in color vision circuits.}, journal = {eLife}, volume = {8}, year = {2019}, month = {2019 Nov 06}, abstract = {

R7 UV photoreceptors (PRs) are divided into yellow (y) and pale (p) subtypes. yR7 PRs express the Dpr11 cell surface protein and are presynaptic to Dm8 amacrine neurons (yDm8) that express Dpr11{\textquoteright}s binding partner DIP-g, while pR7 PRs synapse onto DIP-g-negative pDm8. Dpr11 and DIP-g expression patterns define {\textquoteright}yellow{\textquoteright} and {\textquoteright}pale{\textquoteright} color vision circuits. We examined Dm8 neurons in these circuits by electron microscopic reconstruction and expansion microscopy. and mutations affect the morphologies of yDm8 distal ({\textquoteright}home column{\textquoteright}) dendrites. yDm8 neurons are generated in excess during development and compete for presynaptic yR7 PRs, and interactions between Dpr11 and DIP-g are required for yDm8 survival. These interactions also allow yDm8 neurons to select yR7 PRs as their appropriate home column partners. yDm8 and pDm8 neurons do not normally compete for survival signals or R7 partners, but can be forced to do so by manipulation of R7 subtype fate.

}, issn = {2050-084X}, doi = {10.7554/eLife.48935}, author = {Menon, Kaushiki P and Kulkarni, Vivek and Takemura, Shin-ya and Anaya, Michael and Zinn, Kai} } @article {64851, title = {Interfacial actin protrusions mechanically enhance killing by cytotoxic T cells.}, journal = {Science Immunology}, volume = {4}, year = {2019}, month = {2019 Mar 22}, abstract = {

Cytotoxic T lymphocytes (CTLs) kill by forming immunological synapses with target cells and secreting toxic proteases and the pore-forming protein perforin into the intercellular space. Immunological synapses are highly dynamic structures that boost perforin activity by applying mechanical force against the target cell. Here, we used high-resolution imaging and microfabrication to investigate how CTLs exert synaptic forces and coordinate their mechanical output with perforin secretion. Using micropatterned stimulatory substrates that enable synapse growth in three dimensions, we found that perforin release occurs at the base of actin-rich protrusions that extend from central and intermediate locations within the synapse. These protrusions, which depended on the cytoskeletal regulator WASP and the Arp2/3 actin nucleation complex, were required for synaptic force exertion and efficient killing. They also mediated physical deformation of the target cell surface during CTL-target cell interactions. Our results reveal the mechanical basis of cellular cytotoxicity and highlight the functional importance of dynamic, three-dimensional architecture in immune cell-cell interfaces.

}, issn = {2470-9468}, doi = {10.1126/sciimmunol.aav5445}, author = {Tamzalit, Fella and Wang, Mitchell S and Jin, Weiyang and Tello-Lafoz, Maria and Boyko, Vitaly and Heddleston, John M and Black, Charles T and Kam, Lance C and Huse, Morgan} } @article {49329, title = {Kilohertz frame-rate two-photon tomography.}, journal = {Nature Methods}, volume = {16}, year = {2019}, month = {2019 Jul 29}, pages = {778-86}, abstract = {

Point-scanning two-photon microscopy enables high-resolution imaging within scattering specimens such as the mammalian brain, but sequential acquisition of voxels fundamentally limits imaging speed. We developed a two-photon imaging technique that scans lines of excitation across a focal plane at multiple angles and uses prior information to recover high-resolution images at over 1.4 billion voxels per second. Using a structural image as a prior for recording neural activity, we imaged visually-evoked and spontaneous glutamate release across hundreds of dendritic spines in mice at depths over 250 microns and frame-rates over 1 kHz. Dendritic glutamate transients in anaesthetized mice are synchronized within spatially-contiguous domains spanning tens of microns at frequencies ranging from 1-100 Hz. We demonstrate high-speed recording of acetylcholine and calcium sensors, 3D single-particle tracking, and imaging in densely-labeled cortex. Our method surpasses limits on the speed of raster-scanned imaging imposed by fluorescence lifetime.

}, doi = {10.1101/357269}, author = {Kazemipour, Abbas and Novak, Ondrej and Flickinger, Daniel and Marvin, Jonathan S and Abdelfattah, Ahmed S and King, J. and Borden, PM and Kim, JJ and Al-Abdullatif, SH and Deal, PE and Miller, EW and Schreiter, ER and Druckmann, S and Svoboda, K and Looger, LL and Podgorski, K} } @article {68758, title = {The Kinetochore-Microtubule Coupling Machinery Is Repurposed in Sensory Nervous System Morphogenesis}, journal = {Developmental Cell}, volume = {48}, year = {2019}, month = {03/2019}, pages = {864-872.e7}, abstract = {

Summary Dynamic coupling of microtubule ends to kinetochores, built on the centromeres of chromosomes, directs chromosome segregation during cell division. Here, we report that the evolutionarily ancient kinetochore-microtubule\ coupling machine, the KMN (Knl1/Mis12/Ndc80-complex) network, plays a critical role in neuronal morphogenesis. We show that the KMN network concentrates in microtubule-rich dendrites of developing sensory neurons that collectively extend in a multicellular morphogenetic event that occurs during C.\ elegans embryogenesis. Post-mitotic degradation of KMN components in sensory neurons disrupts dendritic extension, leading to patterning and functional defects in the sensory nervous system. Structure-guided mutations revealed that the molecular interface that couples kinetochores to spindle microtubules also functions in neuronal development. These results identify a cell-division-independent function for the chromosome-segregation machinery and define a microtubule-coupling-dependent event in sensory nervous system morphogenesis.

}, keywords = {Chromosome Segregation, dendrite, kinetochore, KMN network, Knl1, microtubule, Mis12 complex, Mitosis, Morphogenesis, Ndc80 complex, Nervous System, sensory neuron}, issn = {1534-5807}, doi = {https://doi.org/10.1016/j.devcel.2019.02.002}, url = {https://www.sciencedirect.com/science/article/pii/S1534580719300577}, author = {Dhanya K. Cheerambathur and Bram Prevo and Tiffany-Lynn Chow and Neil Hattersley and Shaohe Wang and Zhiling Zhao and Taekyung Kim and Adina Gerson-Gurwitz and Karen Oegema and Rebecca Green and Arshad Desai} } @article {65057, title = {Light-sheet microscopy and its potential for understanding developmental processes.}, journal = {Annual Review of Cell and Developmental Biology}, volume = {35}, year = {2019}, month = {2019 Oct 6}, pages = {655-81}, abstract = {

The ability to visualize and quantitatively measure dynamic biological processes in vivo and at high spatiotemporal resolution is of fundamental importance to experimental investigations in developmental biology. Light-sheet microscopy is particularly well suited to providing such data, since it offers exceptionally high imaging speed and good spatial resolution while minimizing light-induced damage to the specimen. We review core principles and recent advances in light-sheet microscopy, with a focus on concepts and implementations relevant for applications in developmental biology. We discuss how light-sheet microcopy has helped advance our understanding of developmental processes from single-molecule to whole-organism studies, assess the potential for synergies with other state-of-the-art technologies, and introduce methods for computational image and data analysis. Finally, we explore the future trajectory of light-sheet microscopy, discuss key efforts to disseminate new light-sheet technology, and identify exciting opportunities for further advances.

}, issn = {1530-8995}, doi = {10.1146/annurev-cellbio-100818-125311}, author = {Wan, Yinan and McDole, Katie and Keller, Philipp J} } @article {64973, title = {Mechanistic characterization of RASGRP1 variants identifies an hnRNP K-regulated transcriptional enhancer contributing to SLE susceptibility.}, journal = {Frontiers in Immunology}, year = {2019}, month = {2019 May 20}, abstract = {

Systemic lupus erythematosus (SLE) is an autoimmune disease with a strong genetic component. We recently identified a novel SLE susceptibility locus near\ RASGRP1, which governs the ERK/MAPK kinase cascade and B-/T-cell differentiation and development. However, precise causal\ RASGRP1functional variant(s) and their mechanisms of action in SLE pathogenesis remain undefined. Our goal was to fine-map this locus, prioritize genetic variants likely to be functional, experimentally validate their biochemical mechanisms, and determine the contribution of these SNPs to SLE risk. We performed a meta-analysis across six Asian and European cohorts (9,529 cases; 22,462 controls), followed by\ in silico\ bioinformatic and epigenetic analyses to prioritize potentially functional SNPs. We experimentally validated the functional significance and mechanism of action of three SNPs in cultured T-cells. Meta-analysis identified 18 genome-wide significant (p\ \< 5 {\texttimes} 10-8) SNPs, mostly concentrated in two haplotype blocks, one intronic and the other intergenic. Epigenetic fine-mapping, allelic, eQTL, and imbalance analyses predicted three transcriptional regulatory regions with four SNPs (rs7170151, rs11631591-rs7173565, and rs9920715) prioritized for functional validation. Luciferase reporter assays indicated significant allele-specific enhancer activity for intronic rs7170151 and rs11631591-rs7173565 in T-lymphoid (Jurkat) cells, but not in HEK293 cells. Following up with EMSA, mass spectrometry, and ChIP-qPCR, we detected allele-dependent interactions between heterogeneous nuclear ribonucleoprotein K (hnRNP-K) and rs11631591. Furthermore, inhibition of hnRNP-K in Jurkat and primary T-cells downregulated\ RASGRP1\ and ERK/MAPK signaling. Comprehensive association, bioinformatics, and epigenetic analyses yielded putative functional variants of\ RASGRP1, which were experimentally validated. Notably, intronic variant (rs11631591) is located in a cell type-specific enhancer sequence, where its risk allele binds to the hnRNP-K protein and modulates\ RASGRP1\ expression in Jurkat and primary T-cells. As risk allele dosage of rs11631591 correlates with increased\ RASGRP1\ expression and ERK activity, we suggest that this SNP may underlie SLE risk at this locus.

}, doi = {10.3389/fimmu.2019.01066}, author = {Molineros, Julio E. and Singh, Bhupinder and Terao, Chikashi and Okada, Yukinori and Kaplan, Jakub and McDaniel, Barbara and Akizuki, Shuji and Sun, Celi and Webb, Carol F. and Looger, Loren L and Nath, Swapan K.} } @article {64853, title = {Membrane-cytoskeletal crosstalk mediated by myosin-I regulates adhesion turnover during phagocytosis.}, journal = {Nature Communications}, volume = {10}, year = {2019}, month = {2019 03 19}, pages = {1249}, abstract = {

Phagocytosis of invading pathogens or cellular debris requires a dramatic change in cell shape driven by actin polymerization. For antibody-covered targets, phagocytosis is thought to proceed through the sequential engagement of Fc-receptors on the phagocyte with antibodies on the target surface, leading to the extension and closure of the phagocytic cup around the target. We find that two actin-dependent molecular motors, class 1 myosins myosin 1e and myosin 1f, are specifically localized to Fc-receptor adhesions and required for efficient phagocytosis of antibody-opsonized targets. Using primary macrophages lacking both myosin 1e and myosin 1f, we find that without the actin-membrane linkage mediated by these myosins, the organization of individual adhesions is compromised, leading to excessive actin polymerization, slower adhesion turnover, and deficient phagocytic internalization. This work identifies a role for class 1 myosins in coordinated adhesion turnover during phagocytosis and supports a mechanism involving membrane-cytoskeletal crosstalk for phagocytic cup closure.

}, issn = {2041-1723}, doi = {10.1038/s41467-019-09104-1}, author = {Barger, Sarah R and Reilly, Nicholas S and Shutova, Maria S and Li, Qingsen and Maiuri, Paolo and Heddleston, John M and Mooseker, Mark S and Flavell, Richard A and Svitkina, Tatyana and Oakes, Patrick W and Krendel, Mira and Gauthier, Nils C} } @article {64842, title = {Metabolic regulation of developmental cell cycles and zygotic transcription.}, journal = {Current Biology}, volume = {29}, year = {2019}, month = {2019 Mar 01}, pages = {1193-8}, abstract = {

The thirteen nuclear cleavages that give rise to the Drosophila blastoderm are some of the fastest known cell cycles. Surprisingly, the fertilized egg is provided with at most one-third of the dNTPs needed to complete the thirteen rounds of DNA replication. The rest must be synthesized by the embryo, concurrent with cleavage divisions. What is the reason for the limited supply of DNA building blocks? We propose that frugal control of dNTP synthesis contributes to the well-characterized deceleration of the cleavage cycles and is needed for robust accumulation of zygotic gene products. In support of this model, we demonstrate that when the levels of dNTPs are abnormally high, nuclear cleavages fail to sufficiently decelerate, the levels of zygotic transcription are dramatically reduced, and the embryo catastrophically fails early in gastrulation. Our work reveals a direct connection between metabolism, the cell cycle, and zygotic transcription.

}, issn = {1879-0445}, doi = {10.1016/j.cub.2019.02.028}, author = {Djabrayan, Nareg J-V and Smits, Celia M and Krajnc, Matej and Stern, Tomer and Yamada, Shigehiro and Lemon, William C and Keller, Philipp J and Rushlow, Christine A and Shvartsman, Stanislav Y} } @article {69337, title = {Molecular Profiling to Infer Neuronal Cell Identity: Lessons from small ganglia of the Crab Cancer borealis}, journal = {bioRxiv}, year = {2019}, month = {2019 Jul 2}, abstract = {

Understanding circuit organization depends on identification of cell types. Recent advances in transcriptional profiling methods have enabled classification of cell types by their gene expression. While exceptionally powerful and high throughput, the ground-truth validation of these methods is difficult: if cell type is unknown, how does one assess whether a given analysis accurately captures neuronal identity? To shed light on the capabilities and limitations of solely using transcriptional profiling for cell type classification, we performed two forms of transcriptional profiling {\textendash} RNA-seq and quantitative RT-PCR, in single, unambiguously identified neurons from two small crustacean networks: the stomatogastric and cardiac ganglia. We then combined our knowledge of cell type with unbiased clustering analyses and supervised machine learning to determine how accurately functionally-defined neuron types can be classified by expression profile alone. Our results demonstrate that expression profile is able to capture neuronal identity most accurately when combined with multimodal information that allows for post-hoc grouping so analysis can proceed from a supervised perspective. Solely unsupervised clustering can lead to misidentification and an inability to distinguish between two or more cell types. Therefore, our study supports the general utility of cell identification by transcriptional profiling, but adds a caution: it is difficult or impossible to know under what conditions transcriptional profiling alone is capable of assigning cell identity. Only by combining multiple modalities of information such as physiology, morphology or innervation target can neuronal identity be unambiguously determined.SIGNIFICANCE STATEMENT Single cell transcriptional profiling has become a widespread tool in cell identification, particularly in the nervous system, based on the notion that genomic information determines cell identity. However, many cell type classification studies are unconstrained by other cellular attributes (e.g., morphology, physiology). Here, we systematically test how accurately transcriptional profiling can assign cell identity to well-studied anatomically- and functionally-identified neurons in two small neuronal networks. While these neurons clearly possess distinct patterns of gene expression across cell types, their expression profiles are not sufficient to unambiguously confirm their identity. We suggest that true cell identity can only be determined by combining gene expression data with other cellular attributes such as innervation pattern, morphology, or physiology.

}, doi = {10.1101/690388}, url = {https://www.biorxiv.org/content/early/2019/07/02/690388}, author = {Adam J. Northcutt and Daniel R. Kick and Adriane G. Otopalik and Benjamin M. Goetz and Rayna M. Harris and Joseph M. Santin and Hans A. Hofmann and Eve Marder and David J. Schulz} } @article {66012, title = {The mucin-selective protease StcE enables molecular and functional analysis of human cancer-associated mucins}, journal = {Proceedings of the National Academy of Sciences}, volume = {116}, year = {2019}, month = {Sep-04-2019}, pages = {7278 - 7287}, abstract = {

Mucin domains are densely\ O-glycosylated modular protein domains that are found in a wide variety of cell surface and secreted proteins. Mucin-domain glycoproteins are known to be key players in a host of human diseases, especially cancer, wherein mucin expression and glycosylation patterns are altered. Mucin biology has been difficult to study at the molecular level, in part, because methods to manipulate and structurally characterize mucin domains are lacking. Here, we demonstrate that secreted protease of C1 esterase inhibitor (StcE), a bacterial protease from\ Escherichia coli, cleaves mucin domains by recognizing a discrete peptide- and glycan-based motif. We exploited StcE{\textquoteright}s unique properties to improve sequence coverage, glycosite mapping, and glycoform analysis of recombinant human mucins by mass spectrometry. We also found that StcE digests cancer-associated mucins from cultured cells and from ascites fluid derived from patients with ovarian cancer. Finally, using StcE, we discovered that sialic acid-binding Ig-type lectin-7 (Siglec-7), a glycoimmune checkpoint receptor, selectively binds sialomucins as biological ligands, whereas the related receptor Siglec-9 does not. Mucin-selective proteolysis, as exemplified by StcE, is therefore a powerful tool for the study of mucin domain structure and function.

}, issn = {0027-8424}, doi = {10.1073/pnas.1813020116}, url = {http://www.pnas.org/lookup/doi/10.1073/pnas.1813020116https://syndication.highwire.org/content/doi/10.1073/pnas.1813020116http://www.pnas.org/syndication/doi/10.1073/pnas.1813020116}, author = {Malaker, Stacy A. and Pedram, Kayvon and Ferracane, Michael J. and Bensing, Barbara A. and Krishnan, Venkatesh and Pett, Christian and Yu, Jin and Woods, Elliot C. and Kramer, Jessica R. and Westerlind, Ulrika and Dorigo, Oliver and Bertozzi, Carolyn R.} } @article {64972, title = {Multi-color single molecule imaging uncovers extensive heterogeneity in mRNA decoding.}, journal = {Cell}, volume = {178}, year = {2019}, month = {2019 Jul 11}, pages = {458-72}, abstract = {

mRNA translation is a key step in decoding genetic information. Genetic decoding is surprisingly heterogeneous, as multiple distinct polypeptides can be synthesized from a single mRNA sequence. To study translational heterogeneity, we developed the MoonTag, a new fluorescence labeling system to visualize translation of single mRNAs. When combined with the orthogonal SunTag system, the MoonTag enables dual readouts of translation, greatly expanding the possibilities to interrogate complex translational heterogeneity. By placing MoonTag and SunTag sequences in different translation reading frames, each driven by distinct translation start sites, start site selection of individual ribosomes can be visualized in real-time. We find that start site selection is largely stochastic, but that the probability of using a particular start site differs among mRNA molecules, and can be dynamically regulated over time. Together, this study provides key insights into translation start site selection heterogeneity, and provides a powerful toolbox to visualize complex translation dynamics.

}, doi = {10.1016/j.cell.2019.05.001}, author = {Boersma, Sanne and Khuperkar, Deepak and Verhagen, Bram M.P. and Sonneveld, Stijn and Grimm, Jonathan B. and Lavis, Luke D. and Tanenbaum, Marvin E.} } @article {65141, title = {A neural circuit arbitrates between persistence and withdrawal in hungry drosophila.}, journal = {Neuron}, volume = {104}, year = {2019}, month = {2019 Nov 6}, pages = {544-58}, abstract = {

In pursuit of food, hungry animals mobilize significant energy resources and overcome exhaustion and fear. How need and motivation control the decision to continue or change behavior is not understood. Using a single fly treadmill, we show that hungry flies persistently track a food odor and increase their effort over repeated trials in the absence of reward suggesting that need dominates negative experience. We further show that odor tracking is regulated by two mushroom body output neurons (MBONs) connecting the MB to the lateral horn. These MBONs, together with dopaminergic neurons and Dop1R2 signaling, control behavioral persistence. Conversely, an octopaminergic neuron, VPM4, which directly innervates one of the MBONs, acts as a brake on odor tracking by connecting feeding and olfaction. Together, our data suggest a function for the\ MB in internal state-dependent expression of behavior that can be suppressed by external inputs conveying a competing behavioral drive.

}, issn = {1097-4199}, doi = {10.1016/j.neuron.2019.07.028}, author = {Sayin, Sercan and De Backer, Jean-Francois and Siju, K P and Wosniack, Marina E and Lewis, Laurence P and Frisch, Lisa-Marie and Gansen, Benedikt and Schlegel, Philipp and Edmondson-Stait, Amelia and Sharifi, Nadiya and Fisher, Corey B and Calle-Schuler, Steven A and Lauritzen, J Scott and Bock, Davi D and Costa, Marta and Jefferis, Gregory S X E and Gjorgjieva, Julijana and Grunwald Kadow, Ilona C} } @article {65207, title = {A Neural Circuit Arbitrates between Persistence and Withdrawal in Hungry Drosophila.}, journal = {Neuron}, year = {2019}, month = {2019 Aug 09}, abstract = {

In pursuit of food, hungry animals mobilize significant energy resources and overcome exhaustion and fear. How need and motivation control the decision to continue or change behavior is not understood. Using a single fly treadmill, we show that hungry flies persistently track a food odor and increase their effort over repeated trials in the absence of reward suggesting that need dominates negative experience. We further show that odor tracking is regulated by two mushroom body output neurons (MBONs) connecting the MB to the lateral horn. These MBONs, together with dopaminergic neurons and Dop1R2 signaling, control behavioral persistence. Conversely, an octopaminergic neuron, VPM4, which directly innervates one of the MBONs, acts as a brake on odor tracking by connecting feeding and olfaction. Together, our data suggest a function for the\ MB in internal state-dependent expression of behavior that can be suppressed by external inputs conveying a competing behavioral drive.

}, issn = {1097-4199}, doi = {10.1016/j.neuron.2019.07.028}, author = {Sayin, Sercan and De Backer, Jean-Francois and Siju, K P and Wosniack, Marina E and Lewis, Laurence P and Frisch, Lisa-Marie and Gansen, Benedikt and Schlegel, Philipp and Edmondson-Stait, Amelia and Sharifi, Nadiya and Fisher, Corey B and Calle-Schuler, Steven A and Lauritzen, J Scott and Bock, Davi D and Costa, Marta and Jefferis, Gregory S X E and Gjorgjieva, Julijana and Grunwald Kadow, Ilona C} } @article {64917, title = {A neural circuit encoding the experience of copulation in female Drosophila.}, journal = {Neuron}, volume = {102}, year = {2019}, month = {2019 Apr 26}, pages = {1025}, abstract = {

Female behavior changes profoundly after mating. In Drosophila, the mechanisms underlying the long-term changes led by seminal products have been extensively studied. However, the effect of the sensory component of copulation on the female{\textquoteright}s internal state and behavior remains elusive. We pursued this question by dissociating the effect of coital sensory inputs from those of male ejaculate. We found that the sensory inputs of copulation cause a reduction of post-coital receptivity in females, referred to as the "copulation effect." We identified three layers of a neural circuit underlying this phenomenon. Abdominal neurons expressing the mechanosensory channel Piezo convey the signal of copulation to female-specific ascending neurons, LSANs, in the ventral nerve cord. LSANs relay this information to neurons expressing myoinhibitory peptides in the brain. We hereby provide a\ neural mechanism by which the experience of\ copulation facilitates females encoding their mating\ status, thus adjusting behavior to optimize reproduction.

}, issn = {1097-4199}, doi = {10.1016/j.neuron.2019.04.009}, author = {Shao, Lisha and Chung, Phuong and Wong, Allan and Siwanowicz, Igor and Kent, Clement F and Long, Xi and Heberlein, Ulrike} } @conference {67294, title = {Neural Correlates of Cognition in Primary Visual versus Downstream Posterior Cortices During Evidence Accumulation}, booktitle = {APS March Meeting Abstracts}, year = {2019}, month = {01/2019}, abstract = {

The ability of animals to accumulate sensory information across time is fundamental to decision-making. Using a mouse behavioral paradigm where navigational decisions are based on accumulating pulses of visual cues, I compared neural activity in primary visual (V1) to secondary visual and retrosplenial cortices. Even in V1, only a small fraction of neurons had sensory-like responses to cues. Instead, all areas were grossly similar in how neural populations contained a large variety of task-related information from sensory to cognitive, including cue timings, accumulated counts, place/time, decision and reward outcome. Across-trial influences were prevalent, possibly relevant to how animal behavior incorporates past contexts. Intriguingly, all these variables also modulated the amplitudes of sensory responses. While previous work often modeled the accumulation process as integration, the observed scaling of sensory responses by accumulated counts instead suggests a recursive process where sensory responses are gradually amplified. I show that such a multiplicative feedback-loop algorithm better explains psychophysical data than integration, particularly in how the performance transitions into following Weber-Fechner{\textquoteright}s Law only at high counts.

}, url = {https://ui.adsabs.harvard.edu/abs/2019APS..MARP67013K/abstract}, author = {Koay, Sue Ann and Tank, David and Brody, Carlos} } @article {67287, title = {Neural Correlates of Cognition in Primary Visual versus Neighboring Posterior Cortices during Visual Evidence-Accumulation-based Navigation}, journal = {bioRxiv}, year = {2019}, month = {03/2019}, abstract = {

Studies of perceptual decision-making have often assumed that the main role of sensory cortices is to provide sensory input to downstream processes that accumulate and drive behavioral decisions. We performed a systematic comparison of neural activity in primary visual (V1) to secondary visual and retrosplenial cortices, as mice performed a task where they should accumulate pulsatile visual cues through time to inform a navigational decision. Even in V1, only a small fraction of neurons had sensory-like responses to cues. Instead, in all areas neurons were sequentially active, and contained information ranging from sensory to cognitive, including cue timings, evidence, place/time, decision and reward outcome. Per-cue sensory responses were amplitude-modulated by various cognitive quantities, notably accumulated evidence. This inspired a multiplicative feedback-loop circuit hypothesis that proposes a more intricate role of sensory areas in the accumulation process, and furthermore explains a surprising observation that perceptual discrimination deviates from Weber-Fechner Law.Highlights / eTOC BlurbMice made navigational decisions based on accumulating pulsatile visual cuesThe bulk of neural activity in visual cortices was sequential and beyond-sensoryAccumulated pulse-counts modulated sensory (cue) responses, suggesting feedbackA feedback-loop neural circuit explains behavioral deviations from Weber{\textquoteright}s LawHighlights / eTOC BlurbIn a task where navigation was informed by accumulated pulsatile visual evidence, neural activity in visual cortices predominantly coded for cognitive variables across multiple timescales, including outside of a visual processing context. Even sensory responses to visual pulses were amplitude-modulated by accumulated pulse counts and other variables, inspiring a multiplicative feedback-loop circuit hypothesis that in turn explained behavioral deviations from Weber-Fechner Law.

}, doi = {10.1101/568766}, url = {https://www.biorxiv.org/content/early/2019/03/06/568766}, author = {Koay, Sue Ann and Thiberge, Stephan Y. and Brody, Carlos D. and Tank, David W.} } @article {64815, title = {Neuronal reactivation during post-learning sleep consolidates long-term memory in .}, journal = {eLife}, volume = {8}, year = {2019}, month = {2019 Feb 25}, abstract = {

Animals consolidate some, but not all, learning experiences into long-term memory. Across the animal kingdom, sleep has been found to have a beneficial effect on the consolidation of recently formed memories into long-term storage. However, the underlying mechanisms of sleep dependent memory consolidation are poorly understood. Here, we show that consolidation of courtship long-term memory in is mediated by reactivation during sleep of dopaminergic neurons that were earlier involved in memory acquisition. We identify specific fan-shaped body neurons that induce sleep after the learning experience and activate dopaminergic neurons for memory consolidation. Thus, we provide a direct link between sleep, neuronal reactivation of dopaminergic neurons, and memory consolidation.

}, issn = {2050-084X}, doi = {10.7554/eLife.42786}, author = {Dag, Ugur and Lei, Zhengchang and Le, Jasmine Q and Wong, Allan and Bushey, Daniel and Keleman, Krystyna} } @article {65261, title = {Neuropixels data-acquisition system: A scalable platform for parallel recording of 10 000+ electrophysiological signals.}, journal = {IEEE Transactions on Biomedical Circuits and Systems}, volume = {13}, year = {2019}, month = {2019 Dec 01}, pages = {1635-1644}, abstract = {

Although CMOS fabrication has enabled a quick evolution in the design of high-density neural probes and neural-recording chips, the scaling and miniaturization of the complete data-acquisition systems has happened at a slower pace. This is mainly due to the complexity and the many requirements that change depending on the specific experimental settings. In essence, the fundamental challenge of a neural-recording system is getting the signals describing the largest possible set of neurons out of the brain and down to data storage for analysis. This requires a complete system optimization that considers the physical, electrical, thermal and signal-processing requirements, while accounting for available technology, manufacturing constraints and budget. Here we present a scalable and open-standards-based open-source data-acquisition system capable of recording from over 10,000 channels of raw neural data simultaneously. The components and their interfaces have been optimized to ensure robustness and minimum invasiveness in small-rodent electrophysiology.

}, issn = {1940-9990}, doi = {10.1109/TBCAS.2019.2943077}, author = {Putzeys, Jan and Musa, Silke and Mora Lopez, Carolina and Raducanu, Bogdan C and Carton, Alain and De Ceulaer, Jef and Karsh, Bill and Siegle, Joshua H and Van Helleputte, Nick and Harris, Timothy D and Dutta, Barundeb} } @article {59912, title = {NWB:N 2.0: An accessible data standard for neurophysiology.}, journal = {bioRxiv}, year = {2019}, month = {2019 Jan 17}, abstract = {

Neurodata Without Borders: Neurophysiology (NWB:N) is a data standard for neurophysiology, providing neuroscientists with a common standard to share, archive, use, and build common analysis tools for neurophysiology data. With NWB:N version 2.0 (NWB:N 2.0) we made significant advances towards creating a usable standard, software ecosystem, and vibrant community for standardizing neurophysiology data. In this manuscript we focus in particular on the NWB:N data standard schema and present advances towards creating an accessible data standard for neurophysiology.

}, doi = {10.1101/523035}, author = {Rubel, Oliver and Tritt, Andrew and Dichter, Benjamin and Braun, Thomas and Cain, Nicholas and Clack, Nathan G and Davidson, Thomas J and Dougherty, Max and Fillion-Rubin, Jean-Christophe and Graddis, Nile and Grauer, Michael and Kiggins, Justin T and Niu, Lawrence and Ozturk, Doruk and Schroeder, William and Soltesz, Ivan and Sommer, Friedrich T and Svoboda, Karel and Ng, Lydia and et al} } @article {66014, title = {Physical Principles of Membrane Shape Regulation by the Glycocalyx}, journal = {Cell}, volume = {177}, year = {2019}, month = {Jan-06-2019}, pages = {1757 - 1770.e21}, abstract = {

Cells bend their plasma membranes into highly curved forms to interact with the local environment, but how shape generation is regulated is not fully resolved. Here, we report a synergy between shape-generating processes in the cell interior and the external organization and composition of the cell-surface\ glycocalyx.\ Mucinbiopolymers\ and long-chain\ polysaccharides\ within the glycocalyx can generate entropic forces that favor or disfavor the projection of spherical and finger-like extensions from the cell surface. A polymer brush model of the glycocalyx successfully predicts the effects of polymer size and cell-surface density on membrane morphologies. Specific glycocalyx compositions can also induce plasma membrane instabilities to generate more exotic undulating and pearled\ membrane structures\ and drive secretion of extracellular vesicles. Together, our results suggest a fundamental role for the glycocalyx in regulating curved membrane features that serve in communication between cells and with the\ extracellular matrix.

}, issn = {00928674}, doi = {10.1016/j.cell.2019.04.017}, url = {https://www.sciencedirect.com/science/article/pii/S0092867419304039?via\%3Dihub}, author = {Shurer, Carolyn R. and Kuo, Joe Chin-Hun and Roberts, LaDeidra Mon{\'e}t and Gandhi, Jay G. and Colville, Marshall J. and Enoki, Thais A. and Pan, Hao and Su, Jin and Noble, Jade M. and Hollander, Michael J. and O{\textquoteright}Donnell, John P. and Yin, Rose and Pedram, Kayvon and M{\"o}ckl, Leonhard and Kourkoutis, Lena F. and Moerner, W.E. and Bertozzi, Carolyn R. and Feigenson, Gerald W. and Reesink, Heidi L. and Paszek, Matthew J.} } @article {64935, title = {Quantitative in vivo imaging of neuronal glucose concentrations with a genetically encoded fluorescence lifetime sensor.}, journal = {Journal of Neuroscience Research}, volume = {97}, year = {2019}, month = {2019 Aug 01}, pages = {946-60}, abstract = {

Glucose is an essential source of energy for the brain. Recently, the development of genetically encoded fluorescent biosensors has allowed real time visualization of glucose dynamics from individual neurons and astrocytes. A major difficulty for this approach, even for ratiometric sensors, is the lack of a practical method to convert such measurements into actual concentrations in ex vivo brain tissue or in vivo. Fluorescence lifetime imaging provides a strategy to overcome this. In a previous study, we reported the lifetime glucose sensor iGlucoSnFR-TS (then called SweetieTS) for monitoring changes in neuronal glucose levels in response to stimulation. This genetically encoded sensor was generated by combining the Thermus thermophilus glucose-binding protein with a circularly permuted variant of the monomeric fluorescent protein T-Sapphire. Here, we provide more details on iGlucoSnFR-TS design and characterization, as well as pH and temperature sensitivities. For accurate estimation of glucose concentrations, the sensor must be calibrated at the same temperature as the experiments. We find that when the extracellular glucose concentration is in the range 2-10\ mM, the intracellular glucose concentration in hippocampal neurons from acute brain slices is ~20\% of the nominal external glucose concentration (~0.4-2\ mM). We also measured the cytosolic neuronal glucose concentration in vivo, finding a range of ~0.7-2.5\ mM in cortical neurons from awake mice.

}, issn = {1097-4547}, doi = {10.1002/jnr.24433}, author = {D{\'\i}az-Garc{\'\i}a, Carlos Manlio and Lahmann, Carolina and Mart{\'\i}nez-Fran{\c c}ois, Juan Ram{\'o}n and Li, Binsen and Koveal, Dorothy and Nathwani, Nidhi and Rahman, Mahia and Keller, Jacob P and Marvin, Jonathan S and Looger, Loren L and Yellen, Gary} } @article {66013, title = {Quantitative Super-Resolution Microscopy of the Mammalian Glycocalyx}, journal = {Developmental Cell}, volume = {50}, year = {2019}, month = {Jan-07-2019}, pages = {57 - 72.e6}, abstract = {

The mammalian glycocalyx is a heavily glycosylated extramembrane compartment found on nearly every cell. Despite its relevance in both health and disease, studies of the glycocalyx remain hampered by a paucity of methods to spatially classify its components. We combine metabolic labeling, bioorthogonal chemistry, and super-resolution localization microscopy to image two constituents of cell-surface glycans,\ N-acetylgalactosamine (GalNAc) and sialic acid, with 10{\textendash}20\ nm precision in 2D and 3D. This approach enables two measurements: glycocalyx height and the distribution of individual sugars distal from the membrane. These measurements show that the glycocalyx exhibits nanoscale organization on both cell lines and primary human tumor cells. Additionally, we observe enhanced glycocalyx height in response to epithelial-to-mesenchymal transition and to oncogenic\ KRAS\ activation. In the latter case, we trace increased height to an effector gene,\ GALNT7. These data highlight the power of advanced imaging methods to provide molecular and functional insights into glycocalyx biology.

}, issn = {15345807}, doi = {10.1016/j.devcel.2019.04.035}, url = {https://www.sciencedirect.com/science/article/pii/S1534580719303326?via\%3Dihub}, author = {M{\"o}ckl, Leonhard and Pedram, Kayvon and Roy, Anish R. and Krishnan, Venkatesh and Gustavsson, Anna-Karin and Dorigo, Oliver and Bertozzi, Carolyn R. and Moerner, W.E.} } @article {65138, title = {Reconstruction of 1,000 projection neurons reveals new cell types and organization of long-range connectivity in the mouse brain.}, journal = {Cell}, volume = {179}, year = {2019}, month = {2019 Sep 19}, pages = {268-81}, abstract = {

Neuronal\ cell\ types\ are the nodes of neural circuits that determine the flow of information within the\ brain. Neuronal morphology, especially the shape of the axonal arbor, provides an essential descriptor of\ cell\ type and\ reveals\ how individual\ neurons\ route their output across the\ brain. Despite the importance of morphology, few\ projection\ neurons\ in the\ mouse\ brain\ have been reconstructed in their entirety. Here we present a robust and efficient platform for imaging and reconstructing complete neuronal morphologies, including axonal arbors that span substantial portions of the\ brain. We used this platform to reconstruct more than\ 1,000\ projection\ neurons\ in the motor cortex, thalamus, subiculum, and hypothalamus. Together, the reconstructed\ neurons\ constitute more than 85\ meters of axonal length and are available in a searchable online database. Axonal shapes revealed previously unknown subtypes of\ projection\ neurons\ and suggest organizational principles of\ long-range\ connectivity.

}, issn = {00928674}, doi = {10.1016/j.cell.2019.07.042}, author = {Winnubst, Johan and Bas, Erhan and Ferreira, Tiago A. and Wu, Zhuhao and Economo, Michael N. and Edson, Patrick and Arthur, Ben J. and Bruns, Christopher and Rokicki, Konrad and Schauder, David and Olbris, Donald J. and Murphy, Sean D. and Ackerman, David G. and Arshadi, Cameron and Baldwin, Perry and Blake, Regina and Elsayed, Ahmad and Hasan, Mashtura and Ramirez, Daniel and Dos Santos, Bruno and Weldon, Monet and Zafar, Amina and Dudman, Joshua T. and Gerfen, Charles R. and Hantman, Adam W. and Korff, Wyatt and Sternson, Scott M. and Spruston, Nelson and Svoboda, Karel and Chandrashekar, Jayaram} } @article {64988, title = {Regulation of forward and backward locomotion through intersegmental feedback circuits in Drosophila larvae.}, journal = {Nature Communications}, volume = {10}, year = {2019}, month = {2019 Jun 14}, pages = {2654}, abstract = {

Animal locomotion requires spatiotemporally coordinated contraction of muscles throughout the body. Here, we investigate how contractions of antagonistic groups of muscles are intersegmentally coordinated during bidirectional crawling of Drosophila larvae. We identify two pairs of higher-order premotor excitatory interneurons present in each abdominal neuromere that intersegmentally provide feedback to the adjacent neuromere during motor propagation. The two feedback neuron pairs are differentially active during either forward or backward locomotion but commonly target a group of premotor interneurons that together provide excitatory inputs to transverse muscles and inhibitory inputs to the antagonistic longitudinal muscles. Inhibition of either feedback neuron pair compromises contraction of transverse muscles in a direction-specific manner. Our results suggest that the intersegmental feedback neurons coordinate contraction of synergistic muscles by acting as delay circuits representing the phase lag between segments. The identified circuit architecture also shows how bidirectional motor networks could be economically embedded in the nervous system.

}, issn = {2041-1723}, doi = {10.1038/s41467-019-10695-y}, author = {Kohsaka, Hiroshi and Zwart, Maarten F and Fushiki, Akira and Fetter, Richard D and Truman, James W and Cardona, Albert and Nose, Akinao} } @article {65159, title = {A repeated molecular architecture across thalamic pathways.}, journal = {Nature Neuroscience}, volume = {22}, year = {2019}, month = {2019 Sep 16}, pages = {1925-35}, abstract = {

The thalamus is the central communication hub of the forebrain and provides the cerebral cortex with inputs from sensory organs, subcortical systems and the cortex itself. Multiple thalamic regions send convergent information to each cortical region, but the organizational logic of thalamic projections has remained elusive. Through comprehensive transcriptional analyses of retrogradely labeled thalamic neurons in adult mice, we identify three major profiles of thalamic pathways. These profiles exist along a continuum that is repeated across all major projection systems, such as those for vision, motor control and cognition. The largest component of gene expression variation in the mouse thalamus is topographically organized, with features conserved in humans. Transcriptional differences between these thalamic neuronal identities are tied to cellular features that are critical for function, such as axonal morphology and membrane properties. Molecular profiling therefore reveals covariation in the properties of thalamic pathways serving all major input modalities and output targets, thus establishing a molecular framework for understanding the thalamus.

}, issn = {1546-1726}, doi = {10.1038/s41593-019-0483-3}, author = {Phillips, James W and Schulmann, Anton and Hara, Erina and Winnubst, Johan and Liu, Chenghao and Valakh, Vera and Wang, Lihua and Shields, Brenda C and Korff, Wyatt and Chandrashekar, Jayaram and Lemire, Andrew L and Mensh, Brett and Dudman, Joshua T and Nelson, Sacha B and Hantman, Adam W} } @article {64544, title = {Rhomboid distorts lipids to break the viscosity-imposed speed limit of membrane diffusion.}, journal = {Science (New York, N.Y.)}, volume = {363}, year = {2019}, month = {2019 Feb 01}, abstract = {

Enzymes that cut proteins inside membranes regulate diverse cellular events, including cell signaling, homeostasis, and host-pathogen interactions. Adaptations that enable catalysis in this exceptional environment are poorly understood. We visualized single molecules of multiple rhomboid intramembrane proteases and unrelated proteins in living cells (human and ) and planar lipid bilayers. Notably, only rhomboid proteins were able to diffuse above the Saffman-Delbr{\"u}ck viscosity limit of the membrane. Hydrophobic mismatch with the irregularly shaped rhomboid fold distorted surrounding lipids and propelled rhomboid diffusion. The rate of substrate processing in living cells scaled with rhomboid diffusivity. Thus, intramembrane proteolysis is naturally diffusion-limited, but cells mitigate this constraint by using the rhomboid fold to overcome the "speed limit" of membrane diffusion.

}, issn = {1095-9203}, doi = {10.1126/science.aao0076}, author = {Kreutzberger, Alex J B and Ji, Ming and Aaron, Jesse and Mihaljevi{\'c}, Ljubica and Urban, Sini{\v s}a} } @article {65156, title = {RNA granules hitchhike on lysosomes for long-distance transport, Using annexin A11 as a molecular tether.}, journal = {Cell}, volume = {179}, year = {2019}, month = {2019 Sep 19}, pages = {147-164.e20}, abstract = {

Long-distance RNA transport enables local protein synthesis at metabolically-active sites distant from the nucleus. This process ensures an appropriate spatial organization of proteins, vital to polarized cells such as neurons. Here, we present a mechanism for RNA transport in which RNA granules "hitchhike" on moving lysosomes. In\ vitro biophysical modeling, live-cell microscopy, and unbiased proximity labeling proteomics reveal that annexin A11 (ANXA11), an RNA granule-associated phosphoinositide-binding protein, acts as a molecular tether between RNA granules and lysosomes. ANXA11 possesses an N-terminal low complexity domain, facilitating its phase separation into membraneless RNA granules, and a C-terminal membrane binding domain, enabling interactions with lysosomes. RNA granule transport requires ANXA11, and amyotrophic lateral sclerosis (ALS)-associated mutations in ANXA11 impair RNA granule transport by disrupting their interactions with lysosomes. Thus, ANXA11 mediates neuronal RNA transport by tethering RNA granules to actively-transported lysosomes, performing a critical cellular function that is disrupted in ALS.

}, issn = {1097-4172}, doi = {10.1016/j.cell.2019.08.050}, author = {Liao, Ya-Cheng and Fernandopulle, Michael S and Wang, Guozhen and Choi, Heejun and Hao, Ling and Drerup, Catherine M and Patel, Rajan and Qamar, Seema and Nixon-Abell, Jonathon and Shen, Yi and Meadows, William and Vendruscolo, Michele and Knowles, Tuomas P J and Nelson, Matthew and Czekalska, Magdalena A and Musteikyte, Greta and Gachechiladze, Mariam A and Stephens, Christina A and Pasolli, H Amalia and Forrest, Lucy R and St George-Hyslop, Peter and Lippincott-Schwartz, Jennifer and Ward, Michael E} } @article {68754, title = {A Semi-high-throughput Imaging Method and Data Visualization Toolkit to Analyze C. elegans Embryonic Development}, journal = {Journal of Visualized Experiments}, year = {2019}, month = {10/2019}, abstract = {

C. elegans\ is the premier system for the systematic analysis of cell fate specification and morphogenetic events during embryonic development. One challenge is that embryogenesis dynamically unfolds over a period of about 13 h; this half day-long timescale has constrained the scope of experiments by limiting the number of embryos that can be imaged. Here, we describe a semi-high-throughput protocol that allows for the simultaneous 3D time-lapse imaging of development in 80{\textendash}100 embryos at moderate time resolution, from up to 14 different conditions, in a single overnight run. The protocol is straightforward and can be implemented by any laboratory with access to a microscope with point visiting capacity. The utility of this protocol is demonstrated by using it to image two custom-built strains expressing fluorescent markers optimized to visualize key aspects of germ-layer specification and morphogenesis. To analyze the data, a custom program that crops individual embryos out of a broader field of view in all channels, z-steps, and timepoints and saves the sequences for each embryo into a separate tiff stack was built. The program, which includes a user-friendly graphical user interface (GUI), streamlines data processing by isolating, pre-processing, and uniformly orienting individual embryos in preparation for visualization or automated analysis. Also supplied is an ImageJ macro that compiles individual embryo data into a multi-panel file that displays maximum intensity fluorescence projection and brightfield images for each embryo at each time point. The protocols and tools described herein were validated by using them to characterize embryonic development following knock-down of 40 previously described developmental genes; this analysis visualized previously annotated developmental phenotypes and revealed new ones. In summary, this work details a semi-high-throughput imaging method coupled with a cropping program and ImageJ visualization tool that, when combined with strains expressing informative fluorescent markers, greatly accelerates experiments to analyze embryonic development.

}, doi = {10.3791/60362}, url = {https://www.jove.com/t/60362/a-semi-high-throughput-imaging-method-data-visualization-toolkit-tohttps://www.jove.com/t/60362/a-semi-high-throughput-imaging-method-data-visualization-toolkit-to}, author = {Khaliullin, Renat N. and Hendel, Jeffrey M. and Gerson-Gurwitz, Adina and Wang, Shaohe and Ochoa, Stacy D. and Zhao, Zhiling and Desai, Arshad and Oegema, Karen and Green, Rebecca A.} } @article {67290, title = {Sequential and efficient neural-population coding of complex task information}, journal = {bioRxiv}, year = {2019}, month = {10/2019}, abstract = {

Recent work has highlighted that many types of variables are represented in each neocortical area. How can these many neural representations be organized together without interference, and coherently maintained/updated through time? We recorded from large neural populations in posterior cortices as mice performed a complex, dynamic task involving multiple interrelated variables. The neural encoding implied that correlated task variables were represented by uncorrelated modes in an information-coding subspace. We show via theory that this can enable optimal decoding directions to be insensitive to neural noise levels. Across posterior cortex, principles of efficient coding thus applied to task-specific information, with neural-population modes as the encoding unit. Remarkably, this encoding function was multiplexed with rapidly changing, sequential neural dynamics, yet reliably followed slow changes in task-variable correlations through time. We can explain this as due to a mathematical property of high-dimensional spaces that the brain might exploit as a temporal scaffold.

}, doi = {10.1101/801654}, url = {https://www.biorxiv.org/content/early/2019/10/13/801654}, author = {Koay, Sue Ann and Thiberge, Stephan Y. and Brody, Carlos D. and Tank, David W.} } @article {65764, title = {Simple imaging protocol for autofluorescence elimination and optical sectioning in fluorescence endomicroscopy}, journal = {Optica}, volume = {6}, year = {2019}, month = {07/2019}, pages = {972}, abstract = {

Fiber-optic epifluorescence imaging with one-photon excitation benefits from its ease of use, cheap light sources, and full-frame acquisition, which enables it for favorable temporal resolution of image acquisition. However, it suffers from a lack of robustness against autofluorescence and light scattering. Moreover, it cannot easily eliminate the out-of-focus background, which generally results in low-contrast images. In order to overcome these limitations, we have implemented fast out-of-phase imaging after optical modulation (Speed OPIOM) for dynamic contrast in fluorescence endomicroscopy. Using a simple and cheap optical-fiber bundle-based endomicroscope integrating modulatable light sources, we first showed that Speed OPIOM provides intrinsic optical sectioning, which restricts the observation of fluorescent labels at targeted positions within a sample. We also demonstrated that this imaging protocol efficiently eliminates the interference of autofluorescence arising from both the fiber bundle and the specimen in several biological samples. Finally, we could perform multiplexed observations of two spectrally similar fluorophores differing by their photoswitching dynamics. Such attractive features of Speed OPIOM in fluorescence endomicroscopy should find applications in bioprocessing, clinical diagnostics, plant observation, and surface imaging.

}, doi = {10.1364/optica.6.000972}, author = {Zhang, Ruikang and Chouket, Raja and Tebo, Alison G and Plamont, Marie-Aude and Kelemen, Zsolt and Gissot, Lionel and Faure, Jean-Denis and Gautier, Arnaud and Croquette, Vincent and Jullien, Ludovic and Saux, Thomas Le} } @article {65162, title = {Single-cell reconstruction of emerging population activity in an entire developing circuit.}, journal = {Cell}, volume = {179}, year = {2019}, month = {2019 Sep 23}, abstract = {

Animal survival requires a functioning nervous system to develop during embryogenesis. Newborn neurons must assemble into circuits producing activity patterns capable of instructing behaviors. Elucidating how this process is coordinated requires new methods that follow maturation and activity of all cells across a developing circuit. We present an imaging method for comprehensively tracking neuron lineages, movements, molecular identities, and activity in the entire developing zebrafish spinal cord, from neurogenesis until the emergence of patterned activity instructing the earliest spontaneous motor behavior. We found that motoneurons are active first and form local patterned ensembles with neighboring neurons. These ensembles merge, synchronize globally after reaching a threshold size, and finally recruit commissural interneurons to orchestrate the left-right alternating patterns important for locomotion in vertebrates. Individual neurons undergo functional maturation stereotypically based on their birth time and anatomical origin. Our study provides a general strategy for reconstructing how functioning circuits emerge during embryogenesis.

}, issn = {1097-4172}, doi = {10.1016/j.cell.2019.08.039}, author = {Wan, Yinan and Wei, Ziqiang and Looger, Loren L and Koyama, Minoru and Druckmann, Shaul and Keller, Philipp J} } @article {65089, title = {A small number of cholinergic neurons mediate hyperaggression in female Drosophila.}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {116}, year = {2019}, month = {2019 Aug 07}, pages = {17029-38}, abstract = {

In the\ Drosophila\ model of aggression, males and females fight in same-sex pairings, but a wide disparity exists in the levels of aggression displayed by the 2 sexes. A screen of\ Drosophila Flylight Gal4\ lines by driving expression of the gene coding for the temperature sensitive dTRPA1 channel, yielded a single line (GMR26E01-Gal4) displaying greatly enhanced aggression when thermoactivated. Targeted neurons were widely distributed throughout male and female nervous systems, but the enhanced aggression was seen only in females. No effects were seen on female mating behavior, general arousal, or male aggression. We quantified the enhancement by measuring fight patterns characteristic of female and male aggression and confirmed that the effect was female-specific. To reduce the numbers of neurons involved, we used an intersectional approach with our library of enhancer trap flp-recombinase lines. Several crosses reduced the populations of labeled neurons, but only 1 cross yielded a large reduction while maintaining the phenotype. Of particular interest was a small group (2 to 4 pairs) of neurons in the approximate position of the pC1 cluster important in governing male and female social behavior. Female brains have approximately 20\ doublesex (dsx)-expressing neurons within pC1 clusters. Using\ dsxFLP\ instead of\ 357FLP\ for the intersectional studies, we found that the same 2 to 4 pairs of neurons likely were identified with both. These neurons were cholinergic and showed no immunostaining for other transmitter compounds. Blocking the activation of these neurons blocked the enhancement of aggression.

}, issn = {0027-8424}, doi = {10.1073/pnas.1907042116}, url = {http://www.pnas.org/lookup/doi/10.1073/pnas.1907042116https://syndication.highwire.org/content/doi/10.1073/pnas.1907042116http://www.pnas.org/syndication/doi/10.1073/pnas.1907042116}, author = {Palavicino-Maggio, Caroline B. and Chan, Yick-Bun and McKellar, Claire and Kravitz, Edward A.} } @article {67281, title = {Specialized coding of sensory, motor and cognitive variables in VTA dopamine neurons}, journal = {Nature}, volume = {570}, year = {2019}, month = {05/2019}, pages = {509 - 513}, abstract = {

There is increased appreciation that dopamine neurons in the midbrain respond not only to reward1\ and reward-predicting cues1,2, but also to other variables such as the distance to reward3, movements4,5,6,7,8,9\ and behavioural choices10,11. An important question is how the responses to these diverse variables are organized across the population of dopamine neurons. Whether individual dopamine neurons multiplex several variables, or whether there are subsets of neurons that are specialized in encoding specific behavioural variables remains unclear. This fundamental question has been difficult to resolve because recordings from large populations of individual dopamine neurons have not been performed in a behavioural task with sufficient complexity to examine these diverse variables simultaneously. Here, to address this gap, we used two-photon calcium imaging through an implanted lens to record the activity of more than 300 dopamine neurons from the ventral tegmental area of the mouse midbrain during a complex decision-making task. As mice navigated in a virtual-reality environment, dopamine neurons encoded an array of sensory, motor and cognitive variables. These responses were functionally clustered, such that subpopulations of neurons transmitted information about a subset of behavioural variables, in addition to encoding reward. These functional clusters were spatially organized, with neighbouring neurons more likely to be part of the same cluster. Together with the topography between dopamine neurons and their projections, this specialization and anatomical organization may aid downstream circuits in correctly interpreting the wide range of signals transmitted by dopamine neurons.

}, issn = {0028-0836}, doi = {10.1038/s41586-019-1261-9}, url = {http://www.nature.com/articles/s41586-019-1261-9}, author = {Engelhard, Ben and Finkelstein, Joel and Cox, Julia and Fleming, Weston and Jang, Hee Jae and Ornelas, Sharon and Koay, Sue Ann and Thiberge, Stephan Y. and Daw, Nathaniel D. and Tank, David W. and Witten, Ilana B.} } @article {64943, title = {Time-variant SRC kinase activation determines endothelial permeability response.}, journal = {Cell Chemical Biology}, volume = {26}, year = {2019}, month = {2019 Aug 15}, pages = {1081-94}, abstract = {

In the current model of endothelial barrier regulation, the tyrosine kinase SRC is purported to induce disassembly of endothelial adherens junctions (AJs) via\ phosphorylation of VE cadherin, and thereby increase\ junctional permeability. Here, using a chemical biology approach to temporally control SRC activation,\ we show that SRC exerts distinct time-variant effects on the endothelial barrier. We discovered that the immediate effect of SRC activation was to transiently enhance endothelial barrier function as the result of accumulation of VE cadherin at AJs and formation of morphologically distinct reticular AJs. Endothelial barrier enhancement via SRC required phosphorylation of VE cadherin at Y731. In contrast, prolonged SRC activation induced VE cadherin phosphorylation at Y685, resulting in increased endothelial permeability. Thus, time-variant SRC activation differentially phosphorylates VE cadherin and shapes AJs to fine-tune endothelial barrier function. Our work demonstrates important advantages of synthetic biology tools in dissecting complex signaling systems.

}, issn = {2451-9448}, doi = {10.1016/j.chembiol.2019.04.007}, author = {Klomp, Jennifer E and Shaaya, Mark and Matsche, Jacob and Rebiai, Rima and Aaron, Jesse S and Collins, Kerrie B and Huyot, Vincent and Gonzalez, Annette M and Muller, William A and Chew, Teng-Leong and Malik, Asrar B and Karginov, Andrei V} } @article {65078, title = {Unlimited genetic switches for cell-type-specific manipulation.}, journal = {Neuron}, volume = {104}, year = {2019}, month = {2019 Oct 23}, pages = {227-38}, abstract = {

Gaining independent genetic access to discrete cell types is critical to interrogate their biological functions as well as to deliver precise gene therapy. Transcriptomics has allowed us to profile cell populations with extraordinary precision, revealing that cell types are typically defined by a unique combination of genetic markers. Given the lack of adequate tools to target cell types based on multiple markers, most cell types remain inaccessible to genetic manipulation. Here we present CaSSA, a platform to create unlimited genetic switches based on CRISPR/Cas9 (Ca) and the DNA repair mechanism known as single-strand annealing (SSA). CaSSA allows engineering of independent genetic switches, each responding to a specific gRNA. Expressing multiple gRNAs in specific patterns enables multiplex cell-type-specific manipulations and combinatorial genetic targeting. CaSSA is a new genetic tool that conceptually works as an unlimited number of recombinases and will facilitate genetic access to cell types in diverse organisms.

}, doi = {https://doi.org/10.1016/j.neuron.2019.07.005}, author = {Garcia-Marques, Jorge and Yang, Ching-Po and Isabel Espinosa Medina and Mok, Kent and Koyama, Minoru and Lee, Tzumin} } @article {49151, title = {50 Hz volumetric functional imaging with continuously adjustable depth of focus.}, journal = {Biomedical Optics Express}, volume = {9}, year = {2018}, month = {2018 Apr}, pages = {1964-76}, abstract = {

Understanding how neural circuits control behavior requires monitoring a large population of neurons with high spatial resolution and volume rate. Here we report an axicon-based Bessel beam module with continuously adjustable depth of focus (CADoF), that turns frame rate into volume rate by extending the excitation focus in the axial direction while maintaining high lateral resolutions. Cost-effective and compact, this CADoF Bessel module can be easily integrated into existing two-photon fluorescence microscopes. Simply translating one of the relay lenses along its optical axis enabled continuous adjustment of the axial length of the Bessel focus. We used this module to simultaneously monitor activity of spinal projection neurons extending over 60 {\textmu}m depth in larval zebrafish at 50 Hz volume rate with adjustable axial extent of the imaged volume.

}, doi = {10.1364/BOE.9.001964}, author = {Lu, Rongwen and Tanimoto, Masashi and Koyama, Minoru and Na, Ji} } @article {67283, title = {An Accumulation-of-Evidence Task Using Visual Pulses for Mice Navigating in Virtual Reality}, journal = {Frontiers in Behavioral Neuroscience}, volume = {12}, year = {2018}, month = {Jun-03-2018}, abstract = {

The gradual accumulation of sensory evidence is a crucial component of perceptual decision making, but its neural mechanisms are still poorly understood. Given the wide availability of genetic and optical tools for mice, they can be useful model organisms for the study of these phenomena; however, behavioral tools are largely lacking. Here, we describe a new evidence-accumulation task for head-fixed mice navigating in a virtual reality (VR) environment. As they navigate down the stem of a virtual T-maze, they see brief pulses of visual evidence on either side, and retrieve a reward on the arm with the highest number of pulses. The pulses occur randomly with Poisson statistics, yielding a diverse yet well-controlled stimulus set, making the data conducive to a variety of computational approaches. A large number of mice of different genotypes were able to learn and consistently perform the task, at levels similar to rats in analogous tasks. They are sensitive to side differences of a single pulse, and their memory of the cues is stable over time. Moreover, using non-parametric as well as modeling approaches, we show that the mice indeed accumulate evidence: they use multiple pulses of evidence from throughout the cue region of the maze to make their decision, albeit with a small overweighting of earlier cues, and their performance is affected by the magnitude but not the duration of evidence. Additionally, analysis of the mice{\textquoteright}s running patterns revealed that trajectories are fairly stereotyped yet modulated by the amount of sensory evidence, suggesting that the navigational component of this task may provide a continuous readout correlated to the underlying cognitive variables. Our task, which can be readily integrated with state-of-the-art techniques, is thus a valuable tool to study the circuit mechanisms and dynamics underlying perceptual decision making, particularly under more complex behavioral contexts.

}, doi = {10.3389/fnbeh.2018.00036}, url = {http://journal.frontiersin.org/article/10.3389/fnbeh.2018.00036/full}, author = {Pinto, Lucas and Koay, Sue A. and Engelhard, Ben and Yoon, Alice M. and Deverett, Ben and Thiberge, Stephan Y. and Witten, Ilana B. and Tank, David W. and Brody, Carlos D.} } @article {49128, title = {Accurate and sensitive quantification of protein-DNA binding affinity.}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {115}, year = {2018}, month = {2018 Apr 02}, pages = {E3692-701}, abstract = {

Transcription factors (TFs) control gene expression by binding to genomic DNA in a sequence-specific manner. Mutations in TF binding sites are increasingly found to be associated with human disease, yet we currently lack robust methods to predict these sites. Here, we developed a versatile maximum likelihood framework named No Read Left Behind (NRLB) that infers a biophysical model of protein-DNA recognition across the full affinity range from a library of in vitro selected DNA binding sites. NRLB predicts human Max homodimer binding in near-perfect agreement with existing low-throughput measurements. It can capture the specificity of the p53 tetramer and distinguish multiple binding modes within a single sample. Additionally, we confirm that newly identified low-affinity enhancer binding sites are functional in vivo, and that their contribution to gene expression matches their predicted affinity. Our results establish a powerful paradigm for identifying protein binding sites and interpreting gene regulatory sequences in eukaryotic genomes.

}, issn = {1091-6490}, doi = {10.1073/pnas.1714376115}, author = {Rastogi, Chaitanya and Rube, H Tomas and Kribelbauer, Judith F and Crocker, Justin and Loker, Ryan E and Martini, Gabriella D and Laptenko, Oleg and Freed-Pastor, William A and Prives, Carol and Stern, David L and Mann, Richard S and Bussemaker, Harmen J} } @article {49341, title = {The actin cytoskeletal architecture of estrogen receptor positive breast cancer cells suppresses invasion.}, journal = {Nature Communications}, volume = {9}, year = {2018}, month = {2018 Jul 30}, pages = {2980}, abstract = {

Estrogen promotes growth of estrogen receptor-positive (ER+) breast tumors. However, epidemiological studies examining the prognostic characteristics of breast cancer in postmenopausal women receiving hormone replacement therapy reveal a significant decrease in tumor dissemination, suggesting that estrogen has potential protective effects against cancer cell invasion. Here, we show that estrogen suppresses invasion of ER+ breast cancer cells by increasing transcription of the Ena/VASP protein, EVL, which promotes the generation of suppressive cortical actin bundles that inhibit motility dynamics, and is crucial for the ER-mediated suppression of invasion in vitro and in vivo. Interestingly, despite its benefits in suppressing tumor growth, anti-estrogenic endocrine therapy decreases EVL expression and increases local invasion in patients. Our results highlight the dichotomous effects of estrogen on tumor progression and suggest that, in contrast to its established role in promoting growth of ER+ tumors, estrogen has a significant role in suppressing invasion through actin cytoskeletal remodeling.

}, issn = {2041-1723}, doi = {10.1038/s41467-018-05367-2}, author = {Padilla-Rodriguez, Marco and Parker, Sara S and Adams, Deanna G and Westerling, Thomas and Puleo, Julieann I and Watson, Adam W and Hill, Samantha M and Noon, Muhammad and Gaudin, Rapha{\"e}l and Aaron, Jesse and Tong, Daoqin and Roe, Denise J and Knudsen, Beatrice and Mouneimne, Ghassan} } @article {49056, title = {BAK/BAX macropores facilitate mitochondrial herniation and mtDNA efflux during apoptosis.}, journal = {Science (New York, N.Y.)}, volume = {359}, year = {2018}, month = {2018 02 23}, abstract = {

Mitochondrial apoptosis is mediated by BAK and BAX, two proteins that induce mitochondrial outer membrane permeabilization, leading to cytochrome c release and activation of apoptotic caspases. In the absence of active caspases, mitochondrial DNA (mtDNA) triggers the innate immune cGAS/STING pathway, causing dying cells to secrete type I interferon. How cGAS gains access to mtDNA remains unclear. We used live-cell lattice light-sheet microscopy to examine the mitochondrial network in mouse embryonic fibroblasts. We found that after BAK/BAX activation and cytochrome c loss, the mitochondrial network broke down and large BAK/BAX pores appeared in the outer membrane. These BAK/BAX macropores allowed the inner mitochondrial membrane to herniate into the cytosol, carrying with it mitochondrial matrix components, including the mitochondrial genome. Apoptotic caspases did not prevent herniation but dismantled the dying cell to suppress mtDNA-induced innate immune signaling.

}, issn = {1095-9203}, doi = {10.1126/science.aao6047}, author = {McArthur, Kate and Whitehead, Lachlan W and Heddleston, John M and Li, Lucy and Padman, Benjamin S and Oorschot, Viola and Geoghegan, Niall D and Chappaz, Stephane and Davidson, Sophia and San Chin, Hui and Lane, Rachael M and Dramicanin, Marija and Saunders, Tahnee L and Sugiana, Canny and Lessene, Romina and Osellame, Laura D and Chew, Teng-Leong and Dewson, Grant and Lazarou, Michael and Ramm, Georg and Lessene, Guillaume and Ryan, Michael T and Rogers, Kelly L and van Delft, Mark F and Kile, Benjamin T} } @article {49535, title = {Brain-wide circuit interrogation at the cellular level guided by online analysis of neuronal function.}, journal = {Nature Methods}, volume = {15}, year = {2018}, month = {2018 Nov 30}, pages = {1117-1125}, abstract = {

Whole-brain imaging allows for comprehensive functional mapping of distributed neural pathways, but neuronal perturbation experiments are usually limited to targeting predefined regions or genetically identifiable cell types. To complement whole-brain measures of activity with brain-wide manipulations for testing causal interactions, we introduce a system that uses measuredactivity patterns to guide optical perturbations of any subset of neurons in the same fictively behaving larval zebrafish. First, a light-sheet microscope collects whole-brain data that are rapidly analyzed by a distributed computing system to generate functional brain maps. On the basis of these maps, the experimenter can then optically ablate neurons and image activity changes across the brain. We applied this method to characterize contributions of behaviorally tuned populations to the optomotor response. We extended the system to optogenetically stimulate arbitrary subsets of neurons during whole-brain imaging. These open-source methods enable delineating the contributions of neurons to brain-wide circuit dynamics and behavior in individual animals.

}, doi = {10.1038/s41592-018-0221-x}, author = {Vladimirov, Nikita and Wang, Chen and H{\"o}ckendorf, Burkhard and Pujala, Avinash and Tanimoto, Masashi and Mu, Yu and Yang, Chao-Tsung and Wittenbach, Jason and Freeman, Jeremy and Preibisch, Stephan and Koyama, Minoru and Keller, Philipp J and Ahrens, Misha B} } @article {49520, title = {Brain-wide organization of neuronal activity and convergent sensorimotor transformations in larval zebrafish.}, journal = {Neuron}, volume = {100}, year = {2018}, month = {2018 Nov 21}, pages = {876-890.e5}, abstract = {

Simultaneous recordings of large populations of neurons in behaving animals allow detailed observation of high-dimensional, complex brain activity. However, experimental approaches often focus on singular behavioral paradigms or brain areas. Here, we recorded whole-brain neuronal activity of larval zebrafish presented with a battery of visual stimuli while recording fictive motor output. We identified neurons tuned to each stimulus type and motor output and discovered groups of neurons in the anterior hindbrain that respond to different stimuli eliciting similar behavioral responses. These convergent sensorimotor representations were only weakly correlated to instantaneous motor activity, suggesting that they critically inform, but do not directly generate, behavioral choices. To catalog brain-wide activity beyond explicit sensorimotor processing, we developed an unsupervised clustering technique that organizes neurons into functional groups. These analyses enabled a broad overview of the functional organization of the brain and revealed numerous brain nuclei whose neurons exhibit concerted activity patterns.

}, issn = {1097-4199}, doi = {10.1016/j.neuron.2018.09.042}, author = {Chen, Xiuye and Mu, Yu and Hu, Yu and Kuan, Aaron T and Nikitchenko, Maxim and Randlett, Owen and Chen, Alex B and Gavornik, Jeffery P and Sompolinsky, Haim and Engert, Florian and Ahrens, Misha B} } @article {67289, title = {CalmAn: An open source tool for scalable Calcium Imaging data Analysis}, journal = {bioRxiv}, year = {2018}, month = {06/2018}, abstract = {

Advances in fluorescence microscopy enable monitoring larger brain areas in-vivo with finer time resolution. The resulting data rates require reproducible analysis pipelines that are reliable, fully automated, and scalable to datasets generated over the course of months. Here we present CaImAn, an open-source library for calcium imaging data analysis. CaImAn provides automatic and scalable methods to address problems common to pre-processing, including motion correction, neural activity identification, and registration across different sessions of data collection. It does this while requiring minimal user intervention, with good performance on computers ranging from laptops to high-performance computing clusters. CaImAn is suitable for two-photon and one-photon imaging, and also enables real-time analysis on streaming data. To benchmark the performance of CaImAn we collected a corpus of ground truth annotations from multiple labelers on nine mouse two-photon datasets. We demonstrate that CaImAn achieves near-human performance in detecting locations of active neurons.

}, doi = {10.1101/339564}, url = {https://www.biorxiv.org/content/early/2018/06/05/339564}, author = {Giovannucci, Andrea and Friedrich, Johannes and Gunn, Pat and Kalfon, J{\'e}r{\'e}mie and Koay, Sue Ann and Taxidis, Jiannis and Najafi, Farzaneh and Gauthier, Jeffrey L. and Zhou, Pengcheng and Tank, David W. and Chklovskii, Dmitri and Pnevmatikakis, Eftychios A.} } @article {49406, title = {Cell-specific chemical delivery using a selective Nitroreductase-Nitroaryl pair.}, journal = {ACS Chemical Biology}, volume = {13}, year = {2018}, month = {2018 Aug 29}, pages = {1888-96}, abstract = {

The utility of small molecules to probe or perturb biological systems is limited by the lack of cell-specificity. "Masking" the activity of small molecules using a general chemical modification and "unmasking" it only within target cells overcomes this limitation. To this end, we have developed a selective enzyme-substrate pair consisting of engineered variants of E. coli nitroreductase (NTR) and a 2-nitro- N-methylimidazolyl (NM) masking group. To discover and optimize this NTR-NM system, we synthesized a series of fluorogenic substrates containing different nitroaromatic masking groups, confirmed their stability in cells, and identified the best substrate for NTR. We then engineered the enzyme for improved activity in mammalian cells, ultimately yielding an enzyme variant (enhanced NTR, or eNTR) that possesses up to 100-fold increased activity over wild-type NTR. These improved NTR enzymes combined with the optimal NM masking group enable rapid, selective unmasking of dyes, indicators, and drugs to genetically defined populations of cells.

}, issn = {1554-8937}, doi = {10.1021/acschembio.8b00524}, author = {Gruber, Todd D and Krishnamurthy, Chithra and Grimm, Jonathan B and Tadross, Michael R and Wysocki, Laura M and Gartner, Zev J and Lavis, Luke D} } @article {67284, title = {Cerebellar involvement in an evidence-accumulation decision-making task}, journal = {eLife}, volume = {7}, year = {2018}, month = {08/2018}, abstract = {

To make successful evidence-based decisions, the brain must rapidly and accurately transform sensory inputs into specific goal-directed behaviors. Most experimental work on this subject has focused on forebrain mechanisms. Using a novel evidence-accumulation task for mice, we performed recording and perturbation studies of crus I of the lateral posterior cerebellum, which communicates bidirectionally with numerous forebrain regions. Cerebellar inactivation led to a reduction in the fraction of correct trials. Using two-photon fluorescence imaging of calcium, we found that Purkinje cell somatic activity contained choice/evidence-related information. Decision errors were represented by dendritic calcium spikes, which in other contexts are known to drive cerebellar plasticity. We propose that cerebellar circuitry may contribute to computations that support accurate performance in this perceptual decision-making task.

}, doi = {10.7554/eLife.36781}, url = {https://elifesciences.org/articles/36781}, author = {Deverett, Ben and Koay, Sue Ann and Oostland, Marlies and Wang, Samuel S-H} } @article {67292, title = {A cerebellar role in evidence-guided decision-making}, journal = {bioRxiv}, year = {2018}, month = {06/2018}, abstract = {

To make successful evidence-based decisions, the brain must rapidly and accurately transform sensory inputs into specific goal-directed behaviors. Most experimental work on this subject has focused on forebrain mechanisms. Here we show that during perceptual decision-making over a period of seconds, decision-, sensory-, and error-related information converge on the lateral posterior cerebellum in crus I, a structure that communicates bidirectionally with numerous forebrain regions. We trained mice on a novel evidence-accumulation task and demonstrated that cerebellar inactivation reduces behavioral accuracy without impairing motor parameters of action. Using two-photon calcium imaging, we found that Purkinje cell somatic activity encoded choice- and evidence-related variables. Decision errors were represented by dendritic calcium spikes, which are known to drive plasticity. We propose that cerebellar circuitry may contribute to the set of distributed computations in the brain that support accurate perceptual decision-making.

}, doi = {10.1101/343095}, url = {https://www.biorxiv.org/content/early/2018/06/11/343095}, author = {Deverett, Ben and Koay, Sue Ann and Oostland, Marlies and Wang, Samuel S.-H.} } @article {65772, title = {Circularly Permuted Fluorogenic Proteins for the Design of Modular Biosensors.}, journal = {ACS Chemical Biology}, volume = {13}, year = {2018}, month = {09/2018}, pages = {2392 {\textendash} 2397}, abstract = {

Fluorescent reporters are essential components for the design of optical biosensors that are able to image intracellular analytes in living cells. Herein, we describe the development of circularly permuted variants of Fluorescence-Activating and absorption-Shifting Tag (FAST) and demonstrate their potential as reporting module in biosensors. Circularly permutated FAST (cpFAST) variants allow one to condition the binding and activation of a fluorogenic ligand (and thus fluorescence) to analyte recognition by coupling them with analyte-binding domains. We demonstrated their use for biosensor design by generating multicolor plug-and-play fluorogenic biosensors for imaging the intracellular levels of Ca2+ in living mammalian cells in real time.

}, doi = {10.1021/acschembio.8b00417}, author = {Tebo, Alison G and Pimenta, Frederico M and Zoumpoulaki, Martha and Kikuti, Carlos and Sirkia, Helena and Plamont, Marie-Aude and Houdusse, Anne and Gautier, Arnaud} } @article {49236, title = {Color depth MIP mask search: a new tool to expedite Split-GAL4 creation.}, journal = {bioRxiv}, year = {2018}, month = {2018 May 09}, abstract = {

The GAL4-UAS system has proven its versatility in studying the function and expression patterns of neurons the Drosophila central nervous system. Although the GAL4 system has been used for 25 years, recent genetic intersectional tools have enabled genetic targeting of very small numbers of neurons aiding in the understanding of their function. This split-GAL4 system is extremely powerful for studying neuronal morphology and the neural basis of animal behavior. However, choosing lines to intersect that have overlapping patterns restricted to one to a few neurons has been cumbersome. This challenge is now growing as the collections of GAL4 driver lines has increased. Here we present a new method and software plug-in for Fiji to dramatically improve the speed of querying large databases of potential lines to intersect and aid in the split-GAL4 creation. We also provide pre-computed datasets for the Janelia GAL4 (5,738 lines) and VT GAL4 (7,429 lines) of the Drosophila central nervous system (CNS). The tool reduced our split-GAL4 creation effort dramatically.

}, doi = {10.1101/318006}, author = {Otsuna, Hideo and Ito, Masayoshi and Kawase, Takashi} } @article {49149, title = {A community-developed Open-Source computational ecosystem for big neuro data.}, journal = {Nature Methods}, volume = {15}, year = {2018}, month = {2018 Nov}, pages = {846-7}, abstract = {

Big imaging data is becoming more prominent in brain sciences across spatiotemporal scales and phylogenies. We have developed a computational ecosystem that enables storage, visualization, and analysis of these data in the cloud, thusfar spanning 20+ publications and 100+ terabytes including nanoscale ultrastructure, microscale synaptogenetic diversity, and mesoscale whole brain connectivity, making NeuroData the largest and most diverse open repository of brain data.

}, doi = {10.1038/s41592-018-0181-1}, author = {Vogelstein, Joshua T and Perlman, Eric and Falk, Benjamin and Baden, Alex and Roncal, William Gray and Chandrashekhar, Vikram and Collman, Forrest and Seshamani, Sharmishtaa and Patsolic, Jesse and Lillaney, Kunal and Kazhdan, Michael and Hider Jr., Robert and Pryor, Derek and Matelsky, Jordan and Gion, Timothy and Manavalan, Priya and Wester, Brock and Chevillet, Mark and Trautman, Eric T and Khairy, Khaled} } @article {48620, title = {A complete electron microscopy volume of the brain of adult Drosophila melanogaster.}, journal = {Cell}, volume = {174}, year = {2018}, month = {2018 Jul 12}, pages = {730-43}, abstract = {

Drosophila melanogaster has a rich repertoire of innate and learned behaviors. Its 100,000-neuron brain is a large but tractable target for comprehensive neural circuit mapping. Only electron microscopy (EM) enables complete, unbiased mapping of synaptic connectivity; however, the fly brain is too large for conventional EM. We developed a custom high-throughput EM platform and imaged the entire brain of an adult female fly at synaptic resolution. To validate the dataset, we traced brain-spanning circuitry involving the mushroom body (MB), which has been extensively studied for its role in learning. All inputs to Kenyon cells (KCs), the intrinsic neurons of the MB, were mapped, revealing a previously unknown cell type, postsynaptic partners of KC dendrites, and unexpected clustering of olfactory projection neurons. These reconstructions show that this freely available EM volume supports mapping of brain-spanning circuits, which will significantly accelerate Drosophila neuroscience..

}, doi = {10.1016/j.cell.2018.06.019}, author = {Zheng, Zhihao and Lauritzen, J Scott and Perlman, Eric and Robinson, Camenzind G and Nichols, Matthew and Milkie, Daniel E and Torrens, Omar and Price, John and Fisher, Corey B and Sharifi, Nadiya and Calle-Schuler, Steven A and Kmecova, Lucia and Ali, Iqbal J and Karsh, Bill and Trautman, Eric T and Bogovic, John A and Hanslovsky, Philipp and Jefferis, Gregory S X E and Kazhdan, Michael and Khairy, Khaled} } @article {68759, title = {CYK-4 functions independently of its centralspindlin partner ZEN-4 to cellularize oocytes in germline syncytia}, journal = {eLife}, volume = {7}, year = {2018}, month = {07/2018}, pages = {e36919}, abstract = {

Throughout metazoans, germ cells undergo incomplete cytokinesis to form syncytia connected by intercellular bridges. Gamete formation ultimately requires bridge closure, yet how bridges are reactivated to close is not known. The most conserved bridge component is centralspindlin, a complex of the Rho family GTPase-activating protein (GAP) CYK-4/MgcRacGAP and the microtubule motor ZEN-4/kinesin-6. Here, we show that oocyte production by the syncytial \textitCaenorhabditis elegans germline requires CYK-4 but not ZEN-4, which contrasts with cytokinesis, where both are essential. Longitudinal imaging after conditional inactivation revealed that CYK-4 activity is important for oocyte cellularization, but not for the cytokinesis-like events that generate syncytial compartments. CYK-4{\textquoteright}s lipid-binding C1 domain and the GTPase-binding interface of its GAP domain were both required to target CYK-4 to intercellular bridges and to cellularize oocytes. These results suggest that the conserved C1-GAP region of CYK-4 constitutes a targeting module required for closure of intercellular bridges in germline syncytia.

}, keywords = {centralspindlin, germline syncytia, intercellular bridge, Kinesin-6, MgcRacGAP, ring canal}, issn = {2050-084X}, doi = {10.7554/eLife.36919}, url = {https://doi.org/10.7554/eLife.36919}, author = {Lee, Kian-Yong and Green, Rebecca A and Gutierrez, Edgar and Gomez-Cavazos, J Sebastian and Kolotuev, Irina and Wang, Shaohe and Desai, Arshad and Groisman, Alex and Oegema, Karen}, editor = {Balasubramanian, Mohan K and Akhmanova, Anna} } @article {49415, title = {Defective cortex glia plasma membrane structure underlies light-induced epilepsy in mutants.}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {115}, year = {2018}, month = {2018 Sep 05}, pages = {E8919-28}, abstract = {

Seizures induced by visual stimulation (photosensitive epilepsy; PSE) represent a common type of epilepsy in humans, but the molecular mechanisms and genetic drivers underlying PSE remain unknown, and no good genetic animal models have been identified as yet. Here, we show an animal model of PSE, in , owing to defective cortex glia. The cortex glial membranes are severely compromised in ceramide phosphoethanolamine synthase ()-null mutants and fail to encapsulate the neuronal cell bodies in the neuronal cortex. Expression of human sphingomyelin synthase 1, which synthesizes the closely related ceramide phosphocholine (sphingomyelin), rescues the cortex glial abnormalities and PSE, underscoring the evolutionarily conserved role of these lipids in glial membranes. Further, we show the compromise in plasma membrane structure that underlies the glial cell membrane collapse in mutants and leads to the PSE phenotype.

}, issn = {1091-6490}, doi = {10.1073/pnas.1808463115}, author = {Kunduri, Govind and Turner-Evans, Daniel and Konya, Yutaka and Izumi, Yoshihiro and Nagashima, Kunio and Lockett, Stephen and Holthuis, Joost and Bamba, Takeshi and Acharya, Usha and Acharya, Jairaj K} } @article {67291, title = {Detecting and Correcting False Transients in Calcium Imaging}, journal = {bioRxiv}, year = {2018}, month = {11/2018}, abstract = {

Population recordings of calcium activity are a major source of insight into neural function. Large dataset sizes often require automated methods, but automation can introduce errors that are difficult to detect. Here we show that automatic time course estimation can sometimes lead to significant misattribution errors, in which fluorescence is ascribed to the wrong cell. Misattribution arises when the shapes of overlapping cells are imperfectly defined, or when entire cells or processes are not identified, and misattribution can even be produced by methods specifically designed to handle overlap. To diagnose this problem, we develop a transient-by-transient metric and a visualization tool that allow users to quickly assess the degree of misattribution in large populations. To filter out misattribution, we also design a robust estimator that explicitly accounts for contaminating signals in a generative model. Our methods can be combined with essentially any cell finding technique, empowering users to diagnose and correct at large scale a problem that has the potential to significantly alter scientific conclusions.

}, doi = {10.1101/473470}, url = {https://www.biorxiv.org/content/early/2018/11/19/473470}, author = {Gauthier, Jeff L. and Koay, Sue Ann and Nieh, Edward H. and Tank, David W. and Pillow, Jonathan W. and Charles, Adam S.} } @article {49467, title = {Development of 2-colour and 3D SMLM data analysis methods for fibrous spatial point patterns.}, journal = {Journal of Physics D: Applied Physics}, volume = {52}, year = {2018}, month = {2018 Oct 11}, pages = {1}, abstract = {

Abstract ingle molecule localisation microscopy (SMLM), experimentally achieved over a decade ago, has become a routinely used analytical tool across the life sciences. Synergistic advances in probe chemistry, optical physics and data analysis has propelled SMLM into the quantitative realm, enabling unprecedented access to the cellular machinery at the nanoscale. In its early years, SMLM primarily served as a platform for impressive rendered images of sub diffraction scale structures, however more recently a shift towards interrogating SMLM point pattern data in a robust mathematical framework has occurred. A prevalent theme in the SMLM field is the need for quantitative analytical methods, to better understand the underlying processes on which SMLM reports and to extract statistically valid biological insights. Whilst some forms of post processing analytics, for example cluster analysis, have been widely studied, others such as fibre analysis remain in their infancy. Here, we review the current state of the art of cluster analysis and fibre analysis and present new methods for their implementation in both 3D SMLM data sets and multi-colour data.

}, doi = {10.1088/1361-6463/aae7ac}, author = {Peters, Ruby and Griffi{\'e}, Juliette and Williamson, David and Aaron, Jesse and Khuon, Satya and Owen, Dylan} } @article {49342, title = {The ESCRT-III protein CHMP1A mediates secretion of sonic hedgehog on a distinctive subtype of extracellular vesicles.}, journal = {Cell Reports}, volume = {24}, year = {2018}, month = {2018 Jul 24}, pages = {973-986.e8}, abstract = {

Endosomal sorting complex required for transport (ESCRT) complex proteins regulate biogenesis and release of extracellular vesicles (EVs), which enable cell-to-cell communication in the nervous system essential for development and adult function. We recently showed human loss-of-function (LOF) mutations in ESCRT-III member CHMP1A cause autosomal recessive microcephaly with pontocerebellar hypoplasia, but its mechanism was unclear. Here, we show Chmp1a is required for progenitor proliferation in mouse cortex and cerebellum and progenitor maintenance in human cerebral organoids. In Chmp1a null mice, this defect is associated with impaired sonic hedgehog (Shh) secretion and intraluminal vesicle (ILV) formation in multivesicular bodies (MVBs). Furthermore, we show CHMP1A is important for release of an EV subtype that contains AXL, RAB18, and TMED10 (ART) and SHH. Our findings show CHMP1A loss impairs secretion of SHH on ART-EVs, providing molecular mechanistic insights into the role of ESCRT proteins and EVs in the brain.

}, issn = {2211-1247}, doi = {10.1016/j.celrep.2018.06.100}, author = {Coulter, Michael E and Dorobantu, Cristina M and Lodewijk, Gerrald A and Delalande, Fran{\c c}ois and Cianferani, Sarah and Ganesh, Vijay S and Smith, Richard S and Lim, Elaine T and Xu, C Shan and Pang, Song and Wong, Eric T and Lidov, Hart G W and Calicchio, Monica L and Yang, Edward and Gonzalez, Dilenny M and Schlaeger, Thorsten M and Mochida, Ganeshwaran H and Hess, Harald and Lee, Wei-Chung Allen and Lehtinen, Maria K and Kirchhausen, Tomas and Haussler, David and Jacobs, Frank M J and Gaudin, Rapha{\"e}l and Walsh, Christopher A} } @article {49474, title = {Expanding the optogenetics toolkit by topological inversion of rhodopsins.}, journal = {Cell}, volume = {175}, year = {2018}, month = {2018 Oct 16}, pages = {1131-40}, abstract = {

Targeted manipulation of activity in specific populations of neurons is important for investigating the neural circuit basis of behavior. Optogenetic approaches using light-sensitive microbial rhodopsins have permitted manipulations to reach a level of temporal precision that is enabling functional circuit dissection. As demand for more precise perturbations to serve specific experimental goals increases, a palette of opsins with diverse selectivity, kinetics, and spectral properties will be needed. Here, we introduce a novel approach of "topological engineering"-inversion of opsins in the plasma membrane-and demonstrate that it can produce variants with unique functional properties of interest for circuit neuroscience. In one striking example, inversion of a Channelrhodopsin variant converted it from a potent activator into a fast-acting inhibitor that operates as a cation pump. Our findings argue that membrane topology provides a useful orthogonal dimension of protein engineering that immediately permits as much as a doubling of the available toolkit.

}, issn = {1097-4172}, doi = {10.1016/j.cell.2018.09.026}, author = {Brown, Jennifer and Behnam, Reza and Coddington, Luke and Tervo, D G R and Martin, Kathleen and Proskurin, Mikhail and Kuleshova, Elena and Park, Junchol and Phillips, James and Bergs, Amelie C F and Gottschalk, Alexander and Dudman, Joshua T and Karpova, Alla Y} } @article {49324, title = {Fluorogenic structure activity library pinpoints molecular variations in substrate specificity of structurally homologous esterases.}, journal = {The Journal of Biological Chemistry}, volume = {293}, year = {2018}, month = {2018 Jul 13}, pages = {13851-62}, abstract = {

Cellular esterases catalyze many essential biological functions by performing hydrolysis reactions on diverse substrates. The promiscuity of esterases complicates assignment of their substrate preferences and biological functions. To identify universal factors controlling esterase substrate recognition, we designed a 32-member structure-activity relationship (SAR) library of fluorogenic ester substrates and used this library to systematically interrogate esterase preference for chain length, branching patterns, and polarity to differentiate common classes of esterase substrates. Two structurally homologous bacterial esterases were screened against this library, refining their previously broad overlapping substrate specificity. esterase ybfF displayed a preference for γ-position thioethers and ethers, whereas Rv0045c from displayed a preference for branched substrates with and without thioethers. We determined that this substrate differentiation was partially controlled by individual substrate selectivity residues Tyr119 in ybfF and His187 in Rv0045c; reciprocal substitution of these residues shifted each esterase{\textquoteright}s substrate preference. This work demonstrates that the selectivity of esterases is tuned based on transition state stabilization, identifies thioethers as an underutilized functional group for esterase substrates, and provides a rapid method for differentiating structural isozymes. This SAR library could have multi-faceted future applications including in vivo imaging, biocatalyst screening, molecular fingerprinting, and inhibitor design.

}, issn = {1083-351X}, doi = {10.1074/jbc.RA118.003972}, author = {White, Alex and Koelper, Andrew and Russell, Arielle and Larsen, Erik M and Kim, Charles and Lavis, Luke D and Hoops, Geoffrey C and Johnson, R Jeremy} } @article {49093, title = {Functional architecture of reward learning in mushroom body extrinsic neurons of larval Drosophila.}, journal = {Nature Communications}, volume = {9}, year = {2018}, month = {2018 Mar 16}, pages = {1104}, abstract = {

The brain adaptively integrates present sensory input, past experience, and options for future action. The insect mushroom body exemplifies how a central brain structure brings about such integration. Here we use a combination of systematic single-cell labeling, connectomics, transgenic silencing, and activation experiments to study the mushroom body at single-cell resolution, focusing on the behavioral architecture of its input and output neurons (MBINs and MBONs), and of the mushroom body intrinsic APL neuron. Our results reveal the identity and morphology of almost all of these 44 neurons in stage 3 Drosophila larvae. Upon an initial screen, functional analyses focusing on the mushroom body medial lobe uncover sparse and specific functions of its dopaminergic MBINs, its MBONs, and of the GABAergic APL neuron across three behavioral tasks, namely odor preference, taste preference, and associative learning between odor and taste. Our results thus provide a cellular-resolution study case of how brains organize behavior.

}, issn = {2041-1723}, doi = {10.1038/s41467-018-03130-1}, author = {Saumweber, Timo and Rohwedder, Astrid and Schleyer, Michael and Eichler, Katharina and Chen, Yi-chun and Aso, Yoshinori and Cardona, Albert and Eschbach, Claire and Kobler, Oliver and Voigt, Anne and Durairaja, Archana and Mancini, Nino and Zlatic, Marta and Truman, James W and Thum, Andreas S and Gerber, Bertram} } @article {49573, title = {Gene flow mediates the role of sex chromosome meiotic drive during complex speciation.}, journal = {eLife}, volume = {7}, year = {2018}, month = {2018 Dec 13}, abstract = {

During speciation, sex chromosomes often accumulate interspecific genetic incompatibilities faster than the rest of the genome. The drive theory posits that sex chromosomes are susceptible to recurrent bouts of meiotic drive and suppression, causing the evolutionary build-up of divergent cryptic sex-linked drive systems and, incidentally, genetic incompatibilities. To assess the role of drive during speciation, we combine high-resolution genetic mapping of X-linked hybrid male sterility with population genomics analyses of divergence and recent gene flow between the fruitfly species, and . Our findings reveal a high density of genetic incompatibilities and a corresponding dearth of gene flow on the X chromosome. Surprisingly, we find that a known drive element recently migrated between species and, rather than contributing to interspecific divergence, caused a strong reduction in local sequence divergence, undermining the evolution of hybrid sterility. Gene flow can therefore mediate the effects of selfish genetic elements during speciation.

}, issn = {2050-084X}, doi = {10.7554/eLife.35468}, author = {Meiklejohn, Colin D and Landeen, Emily L and Gordon, Kathleen E and Rzatkiewicz, Thomas and Kingan, Sarah B and Geneva, Anthony J and Vedanayagam, Jeffrey P and Muirhead, Christina A and Garrigan, Daniel and Stern, David L and Presgraves, Daven C} } @article {49010, title = {A genetically encoded Ca2+ indicator based on circularly permutated sea anemone red fluorescent protein eqFP578.}, journal = {BMC Biology}, volume = {16}, year = {2018}, month = {2018 Jan 16}, pages = {9}, abstract = {

BACKGROUND: Genetically encoded calcium ion (Ca2+) indicators (GECIs) are indispensable tools for measuring Ca2+ dynamics and neuronal activities in vitro and in vivo. Red fluorescent protein (RFP)-based GECIs have inherent advantages relative to green fluorescent protein-based GECIs due to the longer wavelength light used for excitation. Longer wavelength light is associated with decreased phototoxicity and deeper penetration through tissue. Red GECI can also enable multicolor visualization with blue- or cyan-excitable fluorophores.

RESULTS: Here we report the development, structure, and validation of a new RFP-based GECI, K-GECO1, based on a circularly permutated RFP derived from the sea anemone Entacmaea quadricolor. We have characterized the performance of K-GECO1 in cultured HeLa cells, dissociated neurons, stem-cell-derived cardiomyocytes, organotypic brain slices, zebrafish spinal cord in vivo, and mouse brain in vivo.

CONCLUSION: K-GECO1 is the archetype of a new lineage of GECIs based on the RFP eqFP578 scaffold. It offers high sensitivity and fast kinetics, similar or better than those of current state-of-the-art indicators, with diminished lysosomal accumulation and minimal blue-light photoactivation. Further refinements of the K-GECO1 lineage could lead to further improved variants with overall performance that exceeds that of the most highly optimized red GECIs.

}, issn = {1741-7007}, doi = {10.1186/s12915-018-0480-0}, author = {Shen, Yi and Dana, Hod and Abdelfattah, Ahmed S and Patel, Ronak and Shea, Jamien and Molina, Rosana S and Rawal, Bijal and Rancic, Vladimir and Chang, Yu-Fen and Wu, Lanshi and Chen, Yingche and Qian, Yong and Wiens, Matthew D and Hambleton, Nathan and Ballanyi, Klaus and Hughes, Thomas E and Drobizhev, Mikhail and Kim, Douglas S and Koyama, Minoru and Schreiter, Eric R and Campbell, Robert E} } @article {49813, title = {High-performance GFP-based calcium indicators for imaging activity in neuronal populations and microcompartments.}, journal = {bioRxiv}, year = {2018}, month = {2018 Oct 3}, abstract = {

Calcium imaging with genetically encoded calcium indicators (GECIs) is routinely used to measure neural activity in intact nervous systems. GECIs are frequently used in one of two different modes: to track activity in large populations of neuronal cell bodies, or to follow dynamics in subcellular compartments such as axons, dendrites and individual synaptic compartments. Despite major advances, calcium imaging is still limited by the biophysical properties of existing GECIs, including affinity, signal-to-noise ratio, rise and decay kinetics, and dynamic range. Using structure-guided mutagenesis and neuron-based screening, we optimized the green fluorescent protein-based GECI GCaMP6 for different modes of in vivo imaging. The jGCaMP7 sensors provide improved detection of individual spikes (jGCaMP7s,f), imaging in neurites and neuropil (jGCaMP7b), and tracking large populations of neurons using 2-photon (jGCaMP7s,f) or wide-field (jGCaMP7c) imaging.

}, doi = {10.1101/434589}, author = {Dana, Hod and Sun, Yi and Mohar, Boaz and Hulse, Brad and Hasseman, Jeremy P and Tsegaye, Getahun and Tsang, Arthur and Wong, Allan and Patel, Ronak and Macklin, John J and Chen, Yang and Konnerth, Arthur and Jayaraman, Vivek and Looger, Loren L and Schreiter, Eric R and Svoboda, Karel and Kim, Douglas S} } @article {51794, title = {High-resolution transcriptional dissection of in vivo Atoh1-mediated hair cell conversion in mature cochleae identifies Isl1 as a co-reprogramming factor.}, journal = {PLoS Genetics}, volume = {14}, year = {2018}, month = {2018 07}, pages = {e1007552}, abstract = {

In vivo direct conversion of differentiated cells holds promise for regenerative medicine; however, improving the conversion efficiency and producing functional target cells remain challenging. Ectopic Atoh1 expression in non-sensory supporting cells (SCs) in mouse cochleae induces their partial conversion to hair cells (HCs) at low efficiency. Here, we performed single-cell RNA sequencing of whole mouse sensory epithelia harvested at multiple time points after conditional overexpression of Atoh1. Pseudotemporal ordering revealed that converted HCs (cHCs) are present along a conversion continuum that correlates with both endogenous and exogenous Atoh1 expression. Bulk sequencing of isolated cell populations and single-cell qPCR confirmed 51 transcription factors, including Isl1, are differentially expressed among cHCs, SCs and HCs. In transgenic mice, co-overexpression of Atoh1 and Isl1 enhanced the HC conversion efficiency. Together, our study shows how high-resolution transcriptional profiling of direct cell conversion can identify co-reprogramming factors required for efficient conversion.

}, keywords = {Animals, Animals, Newborn, Basic Helix-Loop-Helix Transcription Factors, Cellular Reprogramming, Gene Expression Profiling, Hair Cells, Auditory, LIM-Homeodomain Proteins, Mice, Mice, Inbred C57BL, Mice, Transgenic, Regeneration, Sequence Analysis, RNA, Single-Cell Analysis, Transcription Factors}, issn = {1553-7404}, doi = {10.1371/journal.pgen.1007552}, author = {Yamashita, Tetsuji and Zheng, Fei and Finkelstein, David and Kellard, Zoe and Carter, Robert and Rosencrance, Celeste D and Sugino, Ken and Easton, John and Gawad, Charles and Zuo, Jian} } @article {49545, title = {Imaging cortical dynamics in GCaMP transgenic rats with a head-mounted widefield macroscope.}, journal = {Neuron}, year = {2018}, month = {2018 Oct 22}, abstract = {

Widefield imaging of calcium dynamics is an emerging method for mapping regional neural activity but is currently limited to restrained animals. Here we describe cScope, a head-mounted widefield macroscope\ developed to image large-scale cortical dynamics in rats during natural behavior. cScope provides a 7.8\ {\texttimes} 4\ mm field of view and dual illumination paths for both fluorescence and hemodynamic correction and can be fabricated at low cost using readily attainable components. We also report the development of Thy-1 transgenic rat strains with widespread neuronal expression of the calcium indicator GCaMP6f. We combined these two technologies to image large-scale calcium dynamics in the dorsal neocortex during a visual evidence accumulation\ task. Quantitative analysis of task-related dynamics revealed multiple regions having neural signals that encode behavioral choice and sensory evidence. Our results provide a new transgenic resource for calcium imaging in rats and extend the domain of head-mounted microscopes to larger-scale cortical dynamics.

}, issn = {1097-4199}, doi = {10.1016/j.neuron.2018.09.050}, author = {Scott, Benjamin B and Thiberge, Stephan Y and Guo, Caiying and Tervo, D Gowanlock R and Brody, Carlos D and Karpova, Alla Y and Tank, David W} } @article {49466, title = {In toto imaging and reconstruction of post-implantation mouse development at the single-cell level.}, journal = {Cell}, volume = {175}, year = {2018}, month = {2018 Oct 10}, pages = {859-876}, abstract = {

The mouse embryo has long been central to the study of mammalian development; however, elucidating the cell behaviors governing gastrulation and the formation of tissues and organs remains a fundamental challenge. A major obstacle is the lack of live imaging and image analysis technologies capable of systematically following cellular dynamics across the developing embryo. We developed a light-sheet microscope that adapts itself to the dramatic changes in size, shape, and optical properties of the post-implantation\ mouse embryo and captures its development from gastrulation to early organogenesis at\ the cellular level. We furthermore developed a computational framework for reconstructing long-term cell tracks, cell divisions, dynamic fate maps, and maps of tissue morphogenesis across the entire embryo. By jointly analyzing cellular dynamics in multiple embryos registered in space and time, we built a dynamic atlas of post-implantation mouse development that, together with our microscopy and computational methods, is provided as a resource.

}, issn = {1097-4172}, doi = {10.1016/j.cell.2018.09.031}, author = {McDole, Katie and Guignard, Leo and Amat, Fernando and Berger, Andrew and Malandain, Gr{\'e}goire and Royer, Lo{\"\i}c A and Turaga, Srinivas C and Branson, Kristin and Keller, Philipp J} } @article {49369, title = {Inhibitory control of prefrontal cortex by the claustrum.}, journal = {Neuron}, volume = {99}, year = {2018}, month = {2018 Aug 07}, pages = {1029-39}, abstract = {

The claustrum is a small subcortical nucleus that has extensive excitatory connections with many cortical areas. While the anatomical connectivity from the claustrum to the cortex has been studied intensively, the physiological effect and underlying circuit mechanisms of claustrocortical communication remain elusive. Here we show that the claustrum provides strong, widespread, and long-lasting feedforward inhibition of the prefrontal cortex (PFC) sufficient to silence ongoing neural activity. This claustrocortical feedforward inhibition was predominantly mediated by interneurons containing neuropeptide Y, and to a lesser extent those containing parvalbumin. Therefore, in contrast to other long-range excitatory inputs to the PFC, the claustrocortical pathway is designed to provide overall inhibition of cortical activity. This unique circuit organization allows the claustrum to rapidly and powerfully suppress cortical networks and suggests a distinct role for the claustrum in regulating cognitive processes in prefrontal circuits.

}, issn = {1097-4199}, doi = {10.1016/j.neuron.2018.07.031}, author = {Jackson, Jesse and Karnani, Mahesh M and Zemelman, Boris V and Burdakov, Denis and Lee, Albert K} } @article {49094, title = {Integrative structure and functional anatomy of a nuclear pore complex.}, journal = {Nature}, volume = {555}, year = {2018}, month = {2018 Mar 14}, pages = {475-82}, abstract = {

Nuclear pore complexes play central roles as gatekeepers of RNA and protein transport between the cytoplasm and nucleoplasm. However, their large size and dynamic nature have impeded a full structural and functional elucidation. Here we determined the structure of the entire 552-protein nuclear pore complex of the yeast Saccharomyces cerevisiae at sub-nanometre precision by satisfying a wide range of data relating to the molecular arrangement of its constituents. The nuclear pore complex incorporates sturdy diagonal columns and connector cables attached to these columns, imbuing the structure with strength and flexibility. These cables also tie together all other elements of the nuclear pore complex, including membrane-interacting regions, outer rings and RNA-processing platforms. Inwardly directed anchors create a high density of transport factor-docking Phe-Gly repeats in the central channel, organized into distinct functional units. This integrative structure enables us to rationalize the architecture, transport mechanism and evolutionary origins of the nuclear pore complex.

}, issn = {1476-4687}, doi = {10.1038/nature26003}, author = {Kim, Seung Joong and Fernandez-Martinez, Javier and Nudelman, Ilona and Shi, Yi and Zhang, Wenzhu and Raveh, Barak and Herricks, Thurston and Slaughter, Brian D and Hogan, Joanna A and Upla, Paula and Chemmama, Ilan E and Pellarin, Riccardo and Echeverria, Ignacia and Shivaraju, Manjunatha and Chaudhury, Azraa S and Wang, Junjie and Williams, Rosemary and Unruh, Jay R and Greenberg, Charles H and Jacobs, Erica Y and Yu, Zhiheng and de la Cruz, M Jason and Mironska, Roxana and Stokes, David L and Aitchison, John D and Jarrold, Martin F and Gerton, Jennifer L and Ludtke, Steven J and Akey, Christopher W and Chait, Brian T and Sali, Andrej and Rout, Michael P} } @article {49177, title = {Joint deformable registration of large EM image volumes: a matrix solver approach.}, journal = {arXiv}, year = {2018}, month = {2018 Apr 26}, abstract = {

Large electron microscopy image datasets for connectomics are typically composed of thousands to millions of partially overlapping two-dimensional images (tiles), which must be registered into a coherent volume prior to further analysis. A common registration strategy is to find matching features between neighboring and overlapping image pairs, followed by a numerical estimation of optimal image deformation using a so-called solver program.\ 
Existing solvers are inadequate for large data volumes, and inefficient for small-scale image registration.\ 
In this work, an efficient and accurate matrix-based solver method is presented. A linear system is constructed that combines minimization of feature-pair square distances with explicit constraints in a regularization term. In absence of reliable priors for regularization, we show how to construct a rigid-model approximation to use as prior. The linear system is solved using available computer programs, whose performance on typical registration tasks we briefly compare, and to which future scale-up is delegated. Our method is applied to the joint alignment of 2.67 million images, with more than 200 million point-pairs and has been used for successfully aligning the first full adult fruit fly brain.

}, url = {https://arxiv.org/abs/1804.10019}, author = {Khairy, Khaled and Denisov, Gennady and Saalfeld, Stephan} } @article {64996, title = {Knock-in rats expressing Cre and Flp recombinases at the Parvalbumin locus.}, journal = {bioRxiv}, year = {2018}, month = {2018 Aug 07}, abstract = {

Rats have the ability to learn and perform sophisticated behavioral tasks, making them very useful for investigating neural circuit functions. In contrast to the extensive mouse genetic toolkit, the paucity of recombinase-expressing rat models has limited the ability to monitor and manipulate molecularly-defined neural populations in this species. Here we report the generation and validation of two knock-in rat strains expressing either\ Cre\ or\ Flp\ recombinase under the control of\ Parvalbumin (Pvalb), a gene expressed in the critical {\textquotedblleft}fast-spiking{\textquotedblright} subset of inhibitory interneurons (FSIs). These strains were generated with CRISPR-Cas9 gene editing and show highly specific and penetrant labeling of\ Pvalb-expressing neurons, as demonstrated by\ in situ\ hybridization and immunohistochemistry. We validated these models in both prefrontal cortex and striatum using both\ ex vivo\ and\ in vivo\ approaches, including whole-cell recording, optogenetics, extracellular physiology and photometry. Our results demonstrate the utility of these new transgenic models for a wide range of neuroscience experiments.

}, doi = {10.1101/386474}, author = {Jai Y. Yu and Jeffrey R. Pettibone and Caiying Guo and Shuqin Zhang and Thomas L. Saunders and Elizabeth D. Hughes and Wanda E. Filipiak and Michael G. Zeidler and Kevin J. Bender and Frederic Hopf and Clay N. Smyth and Viktor Kharazia and Anna Kiseleva and Thomas J. Davidson and Loren M. Frank and Joshua D. Berke} } @article {64995, title = {Lamellar junctions in the endolymphatic sac act as a relief valve to regulate inner ear pressure.}, journal = {eLife}, year = {2018}, month = {2018 Jun 19}, doi = {10.7554/eLife.37131}, author = {Ian A. Swinburne and Kishore R. Mosaliganti and Srigokul Upadhyayula and Tsung-Li Liu and David G. C. Hildebrand and Tony Y.-C. Tsai and Anzhi Chen and Ebaa Al-Obeidi and Anna K. Fass and Samir Malhotra and Florian Engert and Jeff W. Lichtman and Tom Kirchhausen and Sean G. Megason and Eric Betzig} } @article {49275, title = {Lamellar projections in the endolymphatic sac act as a relief valve to regulate inner ear pressure.}, journal = {eLife}, volume = {7}, year = {2018}, month = {2018 Jun 19}, abstract = {

The inner ear is a fluid-filled closed-epithelial structure whose function requires maintenance of an internal hydrostatic pressure and fluid composition. The endolymphatic sac (ES) is a dead-end epithelial tube connected to the inner ear whose function is unclear. ES defects can cause distended ear tissue, a pathology often seen in hearing and balance disorders. Using live imaging of zebrafish larvae, we reveal that the ES undergoes cycles of slow pressure-driven inflation followed by rapid deflation. Absence of these cycles in mutants leads to distended ear tissue. Using serial-section electron microscopy and adaptive optics lattice light-sheet microscopy, we find a pressure relief valve in the ES comprised of partially separated apical junctions and dynamic overlapping basal lamellae that separate under pressure to release fluid. We propose that this lmx1-dependent pressure relief valve is required to maintain fluid homeostasis in the inner ear and other fluid-filled cavities.

}, issn = {2050-084X}, doi = {10.7554/eLife.37131}, author = {Swinburne, Ian A and Mosaliganti, Kishore R and Upadhyayula, Srigokul and Liu, Tsung-Li and Hildebrand, David G C and Tsai, Tony Y-C and Chen, Anzhi and Al-Obeidi, Ebaa and Fass, Anna K and Malhotra, Samir and Engert, Florian and Lichtman, Jeff W and Kirchausen, Tomas and Betzig, Eric and Megason, Sean G} } @article {49473, title = {Long-Term Consolidation of Ensemble Neural Plasticity Patterns in Hippocampal Area CA1.}, journal = {Cell reports}, volume = {25}, year = {2018}, month = {2018 Oct 16}, pages = {640-650.e2}, abstract = {

Neural network remodeling underpins the ability to remember life experiences, but little is known about the long-term plasticity of neural populations. To study how the brain encodes episodic events, we used time-lapse two-photon microscopy and a fluorescent reporter of neural plasticity based on an enhanced form of the synaptic activity-responsive element (E-SARE) within the Arc promoter to track thousands of CA1 hippocampal pyramidal cells over weeks in mice that repeatedly encountered different environments. Each environment evokes characteristic patterns of ensemble neural plasticity, but with each encounter, the set of activated cells gradually evolves. After repeated exposures, the plasticity patterns evoked by an individual environment progressively stabilize. Compared with young adults, plasticity patterns in aged mice are less specific to individual environments and less stable across repeat experiences. Long-term consolidation of hippocampal plasticity patterns may support long-term memory formation, whereas weaker consolidation in aged subjects might reflect declining memory function.

}, issn = {2211-1247}, doi = {10.1016/j.celrep.2018.09.064}, author = {Attardo, Alessio and Lu, Ju and Kawashima, Takashi and Okuno, Hiroyuki and Fitzgerald, James E and Bito, Haruhiko and Schnitzer, Mark J} } @article {65768, title = {Macroscale fluorescence imaging against autofluorescence under ambient light}, journal = {Light: Science \& Applications}, year = {2018}, month = {11/2018}, pages = {1 {\textendash} 12}, abstract = {

Macroscale fluorescence imaging is increasingly used to observe biological samples. However, it may suffer from spectral interferences that originate from ambient light or autofluorescence of the sample or its support. In this manuscript, we built a simple and inexpensive fluorescence macroscope, which has been used to evaluate the performance of Speed OPIOM (Out of Phase Imaging after Optical Modulation), which is a reference-free dynamic contrast protocol, to selectively image reversibly photoswitchable fluorophores as labels against detrimental autofluorescence and ambient light. By tuning the intensity and radial frequency of the modulated illumination to the Speed OPIOM resonance and adopting a phase-sensitive detection scheme that ensures noise rejection, we enhanced the sensitivity and the signal-to-noise ratio for fluorescence detection in blot assays by factors of 50 and 10, respectively, over direct fluorescence observation under constant illumination. Then, we overcame the strong autofluorescence of growth media that are currently used in microbiology and realized multiplexed fluorescence observation of colonies of spectrally similar fluorescent bacteria with a unique configuration of excitation and emission wavelengths. Finally, we easily discriminated fluorescent labels from the autofluorescent and reflective background in labeled leaves, even under the interference of incident light at intensities that are comparable to sunlight. The proposed approach is expected to find multiple applications, from biological assays to outdoor observations, in fluorescence macroimaging.

}, doi = {10.1038/s41377-018-0098-6}, author = {Zhang, Ruikang and Chouket, Raja and Plamont, Marie-Aude and Kelemen, Zsolt and Espagne, Agathe and Tebo, Alison G and Gautier, Arnaud and Gissot, Lionel and Faure, Jean-Denis and Jullien, Ludovic and Croquette, Vincent and Saux, Thomas Le} } @article {49526, title = {Magnetocaloric materials as switchable high contrast ratio MRI labels.}, journal = {Magnetic Resonance in Medicine}, volume = {81}, year = {2018}, month = {2018 Nov 25}, pages = {2238-46}, abstract = {

PURPOSE: To develop switchable and tunable labels with high contrast ratio for MRI using magnetocaloric materials that have sharp first-order magnetic phase transitions at physiological temperatures and typical MRI magnetic field strengths.

METHODS: A prototypical magnetocaloric material iron-rhodium (FeRh) was prepared by melt mixing, high-temperature annealing, and ice-water quenching. Temperature- and magnetic field-dependent magnetization measurements of wire-cut FeRh samples were performed on a vibrating sample magnetometer. Temperature-dependent MRI of FeRh samples was performed on a 4.7T MRI.

RESULTS: Temperature-dependent MRI clearly demonstrated image contrast changes due to the sharp magnetic state transition of the FeRh samples in the MRI magnetic field (4.7T) and at a physiologically relevant temperature (~37{\textdegree}C).

CONCLUSION: A magnetocaloric material, FeRh, was demonstrated to act as a high contrast ratio switchable MRI contrast agent due to its sharp first-order magnetic phase transition in the DC magnetic field of MRI and at physiologically relevant temperatures. A wide range of magnetocaloric materials are available that can be tuned by materials science techniques to optimize their response under MRI-appropriate conditions and be controllably switched in situ with temperature, magnetic field, or a combination of both.

}, issn = {1522-2594}, doi = {10.1002/mrm.27615}, author = {Barbic, Mladen and Dodd, Stephen J and Morris, H Douglas and Dilley, Neil and Marcheschi, Barbara and Huston, Alan and Harris, Tim D and Koretsky, Alan P} } @article {49153, title = {Measuring the global substrate specificity of mycobacterial serine hydrolases using a library of fluorogenic ester substrates.}, journal = {ACS Infectious Diseases}, volume = {4}, year = {2018}, month = {2018 Jun 8}, pages = {904-11}, abstract = {

Among the proteins required for lipid metabolism in Mycobacterium tuberculosis are a significant number of uncharacterized serine hydrolases, especially lipases and esterases. Using a streamlined synthetic method, a library of immolative fluorogenic ester substrates was expanded to better represent the natural lipidomic diversity of Mycobacterium. This expanded fluorogenic library was then used to rapidly characterize the global structure activity relationship (SAR) of mycobacterial serine hydrolases in M.\ smegmatis under different growth conditions. Confirmation of fluorogenic substrate activation by mycobacterial serine hydrolases was performed using nonspecific serine hydrolase inhibitors and reinforced the biological significance of the SAR. The hydrolases responsible for the global SAR were then assigned using gel-resolved activity measurements, and these assignments were used to rapidly identify the relative substrate specificity of previously uncharacterized mycobacterial hydrolases. These measurements provide a global SAR of mycobacterial hydrolase activity, a picture of cycling hydrolase activity, and a detailed substrate specificity profile for previously uncharacterized hydrolases.

}, issn = {2373-8227}, doi = {10.1021/acsinfecdis.7b00263}, author = {Bassett, Braden and Waibel, Brent and White, Alex and Hansen, Heather and Stephens, Dominique and Koelper, Andrew and Larsen, Erik M and Kim, Charles and Glanzer, Adam and Lavis, Luke D and Hoops, Geoffrey C and Johnson, R Jeremy} } @article {65030, title = {Membrane bending moduli of coexisting liquid phases containing transmembrane peptide.}, journal = {Biophys J}, volume = {114}, year = {2018}, month = {2018 May 08}, pages = {2152-2164}, abstract = {

A number of highly curved membranes in\ vivo, such as epithelial cell microvilli, have the relatively high sphingolipid content associated with "raft-like" composition. Given the much lower bending energy measured for bilayers with "nonraft" low sphingomyelin and low cholesterol content, observing high curvature for presumably more rigid compositions seems counterintuitive. To understand this behavior, we measured membrane rigidity by fluctuation analysis of giant unilamellar vesicles. We found that including a transmembrane helical GWALP peptide increases the membrane bending modulus of the liquid-disordered (Ld) phase. We observed this increase at both low-cholesterol fraction and higher, more physiological cholesterol fraction. We find that simplified, commonly used Ld and liquid-ordered (Lo) phases are not representative of those that coexist. When Ld and Lo phases coexist, GWALP peptide favors the Ld phase with a partition coefficient of 3-10 depending on mixture composition. In model membranes at high cholesterol fractions, Ld phases with GWALP have greater bending moduli than the Lo phase that would coexist.

}, keywords = {Amino Acid Sequence, Biomechanical Phenomena, Cell Membrane, Cholesterol, Mechanical Phenomena, Models, Molecular, Oligopeptides, Protein Conformation, alpha-Helical}, issn = {1542-0086}, doi = {10.1016/j.bpj.2018.03.026}, author = {Usery, Rebecca D and Enoki, Thais A and Wickramasinghe, Sanjula P and Nguyen, Vanessa P and Ackerman, David G and Greathouse, Denise V and Koeppe, Roger E and Barrera, Francisco N and Feigenson, Gerald W} } @article {65766, title = {Modifying the Steric Properties in the Second Coordination Sphere of Designed Peptides Leads to Enhancement of Nitrite Reductase Activity}, journal = {Angewandte Chemie International Edition}, volume = {57}, year = {2018}, month = {01/2018}, pages = {3954 {\textendash} 3957}, abstract = {

Protein design is a useful strategy to interrogate the protein structure-function relationship. We demonstrate using a highly modular 3-stranded coiled coil (TRI-peptide system) that a functional type 2 copper center exhibiting copper nitrite reductase (NiR) activity exhibits the highest homogeneous catalytic efficiency under aqueous conditions for the reduction of nitrite to NO and H2O. Modification of the amino acids in the second coordination sphere of the copper center increases the nitrite reductase activity up to 75-fold compared to previously reported systems. We find also that steric bulk can be used to enforce a three-coordinate CuI in a site, which tends toward two-coordination with decreased steric bulk. This study demonstrates the importance of the second coordination sphere environment both for controlling metal-center ligation and enhancing the catalytic efficiency of metalloenzymes and their analogues.

}, doi = {10.1002/anie.201712757}, url = {https://onlinelibrary.wiley.com/doi/full/10.1002/anie.201712757}, author = {Koebke, Karl J and Yu, Fangting and Salerno, Elvin and Stappen, Casey Van and Tebo, Alison G and Penner-Hahn, James E and Pecoraro, Vincent L} } @article {49585, title = {Motor cortex is an input-driven dynamical system controlling dexterous movement.}, journal = {bioRxiv}, year = {2018}, month = {2018-12-14}, pages = {266320}, abstract = {

Skillful control of movement is central to our ability to sense and manipulate the world. A large body of work in nonhuman primates has demonstrated that motor cortex provides flexible, time-varying activity patterns that control the arm during reaching and grasping. Previous studies have suggested that these patterns are generated by strong local recurrent dynamics operating autonomously from inputs during movement execution. An alternative possibility is that motor cortex requires coordination with upstream brain regions throughout the entire movement in order to yield these patterns. Here, we developed an experimental preparation in the mouse to directly test these possibilities using optogenetics and electrophysiology during a skilled reach-to-grab-to-eat task. To validate this preparation, we first established that a specific, time-varying pattern of motor cortical activity was required to produce coordinated movement. Next, in order to disentangle the contribution of local recurrent motor cortical dynamics from external input, we optogenetically held the recurrent contribution constant, then observed how motor cortical activity recovered following the end of this perturbation. Both the neural responses and hand trajectory varied from trial to trial, and this variability reflected variability in external inputs. To directly probe the role of these inputs, we used optogenetics to perturb activity in the thalamus. Thalamic perturbation at the start of the trial prevented movement initiation, and perturbation at any stage of the movement prevented progression of the hand to the target; this demonstrates that input is required throughout the movement. By comparing motor cortical activity with and without thalamic perturbation, we were able to estimate the effects of external inputs on motor cortical population activity. Thus, unlike pattern-generating circuits that are local and autonomous, such as those in the spinal cord that generate left-right alternation during locomotion, the pattern generator for reaching and grasping is distributed across multiple, strongly-interacting brain regions.

}, doi = {10.1101/266320}, author = {Sauerbrei, Britton and Guo, Jian-Zhong and Mischiati, Matteo and Guo, Wendy and Kabra, Mayank and Verma, Nakul and Branson, Kristin M and Hantman, Adam W} } @article {49455, title = {moxMaple3: a photoswitchable fluorescent protein for PALM and protein highlighting in oxidizing cellular environments.}, journal = {Scientific Reports}, volume = {8}, year = {2018}, month = {2018 Oct 03}, pages = {14738}, abstract = {

The ability of fluorescent proteins (FPs) to fold robustly is fundamental to the autocatalytic formation of the chromophore. While the importance of the tertiary protein structure is well appreciated, the impact of individual amino acid mutations for FPs is often not intuitive and requires direct testing. In this study, we describe the engineering of a monomeric photoswitchable FP, moxMaple3, for use in oxidizing cellular environments, especially the eukaryotic secretory pathway. Surprisingly, a point mutation to replace a cysteine substantially improved the yield of correctly folded FP capable of chromophore formation, regardless of cellular environment. The improved folding of moxMaple3 increases the fraction of visibly tagged fusion proteins, as well as FP performance in PALM super-resolution microscopy, and thus makes moxMaple3 a robust monomeric FP choice for PALM and optical highlighting applications.

}, issn = {2045-2322}, doi = {10.1038/s41598-018-32955-5}, author = {Kaberniuk, Andrii A and Mohr, Manuel A and Verkhusha, Vladislav V and Snapp, Erik Lee} } @article {65047, title = {Multiplicative updates for optimization problems with dynamics.}, journal = {IEEE Xplore}, year = {2018}, month = {2018 Apr 16}, abstract = {

We consider the problem of optimizing general convex objective functions with nonnegativity constraints. Using the Karush-Kuhn-Tucker (KKT) conditions for the nonnegativity constraints we will derive fast multiplicative update rules for several problems of interest in signal processing, including non-negative deconvolution, point-process smoothing, ML estimation for Poisson Observations, nonnegative least squares and nonnegative matrix factorization (NMF). Our algorithm can also account for temporal and spatial structure and regularization. We will analyze the performance of our algorithm on simultaneously recorded neuronal calcium imaging and electrophysiology data.

}, doi = {10.1109/ACSSC.2017.8335723}, author = {Abbas Kazemipour and Behtash Babadi and wu, min and Podgorski, Kaspar and Shaul Druckmann} } @article {49011, title = {Mutual inhibition of lateral inhibition: a network motif for an elementary computation in the brain.}, journal = {Current Opinion in Neurobiology}, volume = {49}, year = {2018}, month = {2018 Jan 16}, pages = {69-74}, abstract = {

A series of classical studies in non-human primates has revealed the neuronal activity patterns underlying decision-making. However, the circuit mechanisms for such patterns remain largely unknown. Recent detailed circuit analyses in simpler neural systems have started to reveal the connectivity patterns underlying analogous processes. Here we review a few of these systems that share a particular connectivity pattern, namely mutual inhibition of lateral inhibition. Close examination of these systems suggests that this recurring connectivity pattern ({\textquoteright}network motif{\textquoteright}) is a building block to enforce particular dynamics, which can be used not only for simple behavioral choice but also for more complex choices and other brain functions. Thus, a network motif provides an elementary computation that is not specific to a particular brain function and serves as an elementary building block in the brain.

}, issn = {1873-6882}, doi = {10.1016/j.conb.2017.12.019}, author = {Koyama, Minoru and Pujala, Avinash} } @article {49116, title = {A Neural Circuit for the Suppression of Pain by a Competing Need State.}, journal = {Cell}, volume = {173}, year = {2018}, month = {2018 Mar 22}, pages = {140-52}, abstract = {

Hunger and pain are two competing signals that individuals must resolve to ensure survival. However, the neural processes that prioritize conflicting survival needs are poorly understood. We discovered that hunger attenuates behavioral responses and affective properties of inflammatory pain without altering acute\ nociceptive responses. This effect is centrally controlled, as activity in hunger-sensitive agouti-related protein (AgRP)-expressing neurons abrogates\ inflammatory pain. Systematic analysis of AgRP projection subpopulations revealed that the\ neural processing of hunger and inflammatory pain converge in the hindbrain parabrachial nucleus (PBN). Strikingly, activity in AgRP {\textrightarrow} PBN neurons blocked the behavioral response to inflammatory pain as effectively as hunger or analgesics. The anti-nociceptive effect of hunger is mediated by neuropeptide Y (NPY) signaling in the PBN. By investigating the intersection between hunger and pain, we have identified a neural circuit that mediates competing survival needs and uncovered NPY Y1 receptor signaling in the PBN as a target for pain suppression.

}, issn = {1097-4172}, doi = {10.1016/j.cell.2018.02.057}, author = {Alhadeff, Amber L and Su, Zhenwei and Hernandez, Elen and Klima, Michelle L and Phillips, Sophie Z and Holland, Ruby A and Guo, Caiying and Hantman, Adam W and De Jonghe, Bart C and Betley, J Nicholas} } @article {49529, title = {Neuron-subtype-specific expression, interaction affinities, and specificity determinants of DIP/Dpr cell recognition proteins.}, journal = {Neuron}, volume = {100}, year = {2018}, month = {2018 Nov 12}, pages = {1385-1400}, abstract = {

Binding between DIP and Dpr neuronal recognition proteins has been proposed to regulate synaptic connections between lamina and medulla neurons in the Drosophila visual system. Each lamina neuron was previously shown to express many Dprs. Here, we demonstrate, by contrast, that their synaptic partners typically express one or two DIPs, with binding specificities matched to the lamina neuron-expressed Dprs. A deeper understanding of the molecular logic of DIP/Dpr interaction requires quantitative studies on the properties of these proteins. We thus generated a quantitative affinity-based DIP/Dpr interactome for all DIP/Dpr protein family members. This revealed a broad range of affinities and identified homophilic binding for some DIPs and some Dprs. These data, along with full-length ectodomain DIP/Dpr and DIP/DIP crystal structures, led to the identification of molecular determinants of DIP/Dpr specificity. This structural knowledge, along with a comprehensive set of quantitative binding affinities, provides new tools for functional studies in\ vivo.

}, issn = {1097-4199}, doi = {10.1016/j.neuron.2018.10.046}, author = {Cosmanescu, Filip and Katsamba, Phinikoula S and Sergeeva, Alina P and Ahlsen, Goran and Patel, Saurabh D and Brewer, Joshua J and Tan, Liming and Xu, Shuwa and Xiao, Qi and Nagarkar-Jaiswal, Sonal and Nern, Aljoscha and Bellen, Hugo J and Zipursky, S Lawrence and Honig, Barry and Shapiro, Lawrence} } @article {49095, title = {NeuroStorm: accelerating brain science discovery in the cloud.}, journal = {arXiv}, year = {2018}, month = {2018 Mar 09}, abstract = {

Neuroscientists are now able to acquire data at staggering rates across spatiotemporal scales. However, our ability to capitalize on existing datasets, tools, and intellectual capacities is hampered by technical challenges. The key barriers to accelerating scientific discovery correspond to the FAIR data principles: findability, global access to data, software interoperability, and reproducibility/re-usability. We conducted a hackathon dedicated to making strides in those steps. This manuscript is a technical report summarizing these achievements, and we hope serves as an example of the effectiveness of focused, deliberate hackathons towards the advancement of our quickly-evolving field.

}, url = {https://arxiv.org/abs/1803.03367}, author = {Kiar, Gregory and Anderson, Robert J and Baden, Alex and Badea, Alexandra and Bridgeford, Eric W and Champion, Andrew and Chandrashekar, Jayaram and Collman, Forrest and Duderstadt, Brandon and Evans, Alan C and Engert, Florian and Falk, Benjamin and Glatard, Tristan and Roncal, William Gray and Kennedy, David N and Maitlin-Shepard and Marren, Ryan A and Nnaemeka, Onyeka and Perlman, Eric and Seshamani, Sharmishtaas} } @article {68994, title = {Neutralizing Gatad2a-Chd4-Mbd3/NuRD Complex Facilitates Deterministic Induction of Naive Pluripotency.}, journal = {Cell Stem Cell}, volume = {23}, year = {2018}, month = {09/2018}, pages = {412-425.e10}, abstract = {

Mbd3, a member of nucleosome remodeling and deacetylase (NuRD) co-repressor complex, was previously identified as an inhibitor for deterministic induced pluripotent stem cell (iPSC) reprogramming, where up to 100\% of donor cells successfully complete the process. NuRD can assume multiple mutually exclusive conformations, and it remains unclear whether this deterministic phenotype can be attributed to a specific Mbd3/NuRD subcomplex. Moreover, since complete ablation of Mbd3 blocks somatic cell proliferation, we aimed to explore functionally relevant alternative ways to neutralize Mbd3-dependent NuRD activity. We identify Gatad2a, a NuRD-specific subunit, whose complete deletion specifically disrupts Mbd3/NuRD repressive activity on the pluripotency circuitry during iPSC differentiation and reprogramming without ablating somatic cell proliferation. Inhibition of Gatad2a facilitates deterministic murine iPSC reprogramming within 8\ days. We validate a distinct molecular axis, Gatad2a-Chd4-Mbd3, within Mbd3/NuRD as being critical for blocking reestablishment of naive pluripotency and further highlight signaling-dependent and post-translational modifications of Mbd3/NuRD that influence its interactions and assembly.

}, keywords = {Animals, Cells, Cultured, DNA Helicases, DNA-Binding Proteins, Female, GATA Transcription Factors, Induced Pluripotent Stem Cells, Male, Mi-2 Nucleosome Remodeling and Deacetylase Complex, Mice, Mice, Inbred C57BL, Mice, Inbred CBA, Mice, Knockout, Mice, Transgenic, Transcription Factors}, issn = {1875-9777}, doi = {10.1016/j.stem.2018.07.004}, author = {Mor, Nofar and Rais, Yoach and Sheban, Daoud and Peles, Shani and Aguilera-Castrejon, Alejandro and Zviran, Asaf and Elinger, Dalia and Viukov, Sergey and Geula, Shay and Krupalnik, Vladislav and Zerbib, Mirie and Chomsky, Elad and Lasman, Lior and Shani, Tom and Bayerl, Jonathan and Gafni, Ohad and Hanna, Suhair and Buenrostro, Jason D and Hagai, Tzachi and Masika, Hagit and Vainorius, Gintautas and Bergman, Yehudit and Greenleaf, William J and Esteban, Miguel A and Elling, Ulrich and Levin, Yishai and Massarwa, Rada and Merbl, Yifat and Novershtern, Noa and Hanna, Jacob H} } @article {49244, title = {Nicotinic cholinergic receptors in VTA glutamate neurons modulate excitatory transmission.}, journal = {Cell Reports}, volume = {23}, year = {2018}, month = {2018 May 22}, pages = {2236-2244}, abstract = {

Ventral tegmental area (VTA) glutamate neurons are important components of reward circuitry, but whether they are subject to cholinergic modulation is unknown. To study this, we used molecular, physiological, and photostimulation techniques to examine nicotinic acetylcholine receptors (nAChRs) in VTA glutamate neurons. Cells in the medial VTA, where glutamate neurons are enriched, are responsive to acetylcholine (ACh) released from cholinergic axons. VTA VGLUT2 neurons express mRNA and protein subunits known to comprise heteromeric nAChRs. Electrophysiology, coupled with two-photon microscopy and laser flash photolysis of photoactivatable nicotine, was used to demonstrate nAChR functional activity in the somatodendritic subcellular compartment of VTA VGLUT2 neurons. Finally, optogenetic isolation of intrinsic VTA glutamatergic microcircuits along with gene-editing techniques demonstrated that nicotine potently modulates excitatory transmission within the VTA via heteromeric nAChRs. These results indicate that VTA glutamate neurons are modulated by cholinergic mechanisms and participate in the cascade of physiological responses to nicotine exposure.

}, issn = {2211-1247}, doi = {10.1016/j.celrep.2018.04.062}, author = {Yan, Yijin and Peng, Can and Arvin, Matthew C and Jin, Xiao-Tao and Kim, Veronica J and Ramsey, Matthew D and Wang, Yong and Banala, Sambashiva and Wokosin, David L and McIntosh, J Michael and Lavis, Luke D and Drenan, Ryan M} } @article {49176, title = {A novel sheet-like virus particle array is a hallmark of Zika virus infection.}, journal = {Emerging Microbes \& Infections}, volume = {7}, year = {2018}, month = {2018 Apr 25}, pages = {69}, abstract = {

Zika virus (ZIKV) is an emerging flavivirus that caused thousands of human infections in recent years. Compared to other human flaviviruses, ZIKV replication is not well understood. Using fluorescent, transmission electron, and focused ion beam-scanning electron microscopy, we examined ZIKV replication dynamics in Vero 76 cells and in the brains of infected laboratory mice. We observed the progressive development of a perinuclear flaviviral replication factory both in vitro and in vivo. In vitro, we illustrated the ZIKV lifecycle from particle cell entry to egress. ZIKV particles assembled and aggregated in an induced convoluted membrane structure and ZIKV strain-specific membranous vesicles. While most mature virus particles egressed via membrane budding, some particles also likely trafficked through late endosomes and egressed through membrane abscission. Interestingly, we consistently observed a novel sheet-like virus particle array consisting of a single layer of ZIKV particles. Our study further defines ZIKV replication and identifies a novel hallmark of ZIKV infection.

}, issn = {2222-1751}, doi = {10.1038/s41426-018-0071-8}, author = {Liu, Jun and Kline, Brandon A and Kenny, Tara A and Smith, Darci R and Soloveva, Veronica and Beitzel, Brett and Pang, Song and Lockett, Stephen and Hess, Harald F and Palacios, Gustavo and Kuhn, Jens H and Sun, Mei G and Zeng, Xiankun} } @article {49173, title = {Observing the cell in its native state: Imaging subcellular dynamics in multicellular organisms.}, journal = {Science (New York, N.Y.)}, volume = {360}, year = {2018}, month = {2018 Apr 20}, abstract = {

True physiological imaging of subcellular dynamics requires studying cells within their parent organisms, where all the environmental cues that drive gene expression, and hence the phenotypes that we actually observe, are present. A complete understanding also requires volumetric imaging of the cell and its surroundings at high spatiotemporal resolution, without inducing undue stress on either. We combined lattice light-sheet microscopy with adaptive optics to achieve, across large multicellular volumes, noninvasive aberration-free imaging of subcellular processes, including endocytosis, organelle remodeling during mitosis, and the migration of axons, immune cells, and metastatic cancer cells in vivo. The technology reveals the phenotypic diversity within cells across different organisms and developmental stages and may offer insights into how cells harness their intrinsic variability to adapt to different physiological environments.

}, issn = {1095-9203}, doi = {10.1126/science.aaq1392}, author = {Liu, Tsung-Li and Upadhyayula, Srigokul and Milkie, Daniel E and Singh, Ved and Wang, Kai and Swinburne, Ian A and Mosaliganti, Kishore R and Collins, Zach M and Hiscock, Tom W and Shea, Jamien and Kohrman, Abraham Q and Medwig, Taylor N and Dambournet, Daphne and Forster, Ryan and Cunniff, Brian and Ruan, Yuan and Yashiro, Hanako and Scholpp, Steffen and Meyerowitz, Elliot M and Hockemeyer, Dirk and Drubin, David G and Martin, Benjamin L and Matus, David Q and Koyama, Minoru and Megason, Sean G and Kirchhausen, Tom and Betzig, Eric} } @article {49141, title = {Odorant binding protein 69a connects social interaction to modulation of social responsiveness in Drosophila.}, journal = {PLoS Genetics}, volume = {14}, year = {2018}, month = {2018 Apr 09}, pages = {e1007328}, abstract = {

Living in a social environment requires the ability to respond to specific social stimuli and to incorporate information obtained from prior interactions into future ones. One of the mechanisms that facilitates social interaction is pheromone-based communication. In Drosophila melanogaster, the male-specific pheromone cis-vaccenyl acetate (cVA) elicits different responses in male and female flies, and functions to modulate behavior in a context and experience-dependent manner. Although it is the most studied pheromone in flies, the mechanisms that determine the complexity of the response, its intensity and final output with respect to social context, sex and prior interaction, are still not well understood. Here we explored the functional link between social interaction and pheromone-based communication and discovered an odorant binding protein that links social interaction to sex specific changes in cVA related responses. Odorant binding protein 69a (Obp69a) is expressed in auxiliary cells and secreted into the olfactory sensilla. Its expression is inversely regulated in male and female flies by social interactions: cVA exposure reduces its levels in male flies and increases its levels in female flies. Increasing or decreasing Obp69a levels by genetic means establishes a functional link between Obp69a levels and the extent of male aggression and female receptivity. We show that activation of cVA-sensing neurons is sufficeint to regulate Obp69a levels in the absence of cVA, and requires active neurotransmission between the sensory neuron to the second order olfactory neuron. The cross-talk between sensory neurons and non-neuronal auxiliary cells at the olfactory sensilla, represents an additional component in the machinery that promotes behavioral plasticity to the same sensory stimuli in male and female flies.

}, issn = {1553-7404}, doi = {10.1371/journal.pgen.1007328}, author = {Bentzur, Assa and Shmueli, Anat and Omesi, Liora and Ryvkin, Julia and Knapp, Jon-Michael and Parnas, Moshe and Davis, Fred P and Shohat-Ophir, Galit} } @article {49115, title = {Opportunities and obstacles for deep learning in biology and medicine.}, journal = {Journal of The Royal Society Interface}, year = {2018}, month = {2018 Apr 4}, abstract = {

Deep learning describes a class of machine learning algorithms that are capable of combining raw inputs into layers of intermediate features. These algorithms have recently shown impressive results across a variety of domains. Biology and medicine are data-rich disciplines, but the data are complex and often ill-understood. Hence, deep learning techniques may be particularly well suited to solve problems of these fields. We examine applications of deep learning to a variety of biomedical problems{\textemdash}patient classification, fundamental biological processes and treatment of patients{\textemdash}and discuss whether deep learning will be able to transform these tasks or if the biomedical sphere poses unique challenges. Following from an extensive literature review, we find that deep learning has yet to revolutionize biomedicine or definitively resolve any of the most pressing challenges in the field, but promising advances have been made on the prior state of the art. Even though improvements over previous baselines have been modest in general, the recent progress indicates that deep learning methods will provide valuable means for speeding up or aiding human investigation. Though progress has been made linking a specific neural network{\textquoteright}s prediction to input features, understanding how users should interpret these models to make testable hypotheses about the system under study remains an open challenge. Furthermore, the limited amount of labelled data for training presents problems in some domains, as do legal and privacy constraints on work with sensitive health records. Nonetheless, we foresee deep learning enabling changes at both bench and bedside with the potential to transform several areas of biology and medicine.

}, doi = {10.1098/rsif.2017.0387}, author = {Ching, Travers and Himmelstein, Daniel S and Beaulieu-Jones, Brett K and Kalinin, Alexandr A and Do, Brian T and Way, Gregory P and Ferrero, Enrico and Agapow, Paul-Michael and Zietz, Michael and Hoffman, Michael M and Xie, Wei and Rosen, Gail L and Lengerich, Benjamin J and Israeli, Johnny and Lanchantin, Jack and Woloszynek, Stephen and Carpenter, Anne E and Shrikumar, Avanti and Xu, Jinbo and Cofer, Evan M} } @article {48976, title = {Optogenetic dissection of descending behavioral control in Drosophila.}, journal = {eLife}, year = {2018}, month = {2018}, pages = {e34275}, abstract = {

In most animals, the brain makes behavioral decisions that are transmitted by descending neurons to the nerve cord circuitry that produces behaviors. In insects, only a few descending neurons have been associated with specific behaviors. To explore how descending neurons control an insect{\textquoteright}s movements, we developed a novel method to systematically assay the behavioral effects of activating individual neurons on freely behaving terrestrial\ D. melanogaster. We calculated a two-dimensional representation of the entire behavior space explored by these flies and we associated descending neurons with specific behaviors by identifying regions of this space that were visited with increased frequency during optogenetic activation. Applying this approach across a large collection of descending neurons, we found that (1) activation of most of the descending neurons drove stereotyped behaviors, (2) in many cases multiple descending neurons activated similar behaviors, and (3) optogenetically-activated behaviors were often dependent on the behavioral state prior to activation.

}, doi = {10.7554/eLife.34275}, author = {Cande, Jessica and Namiki, Shigehiro and Qiu, Jirui and Korff, Wyatt and Card, Gwyneth M and Shaevitz, Joshua W and Stern, David L and Berman, Gordon J} } @article {49002, title = {Persistent activity in a recurrent circuit underlies courtship memory in Drosophila.}, journal = {eLife}, volume = {7}, year = {2018}, month = {2018 Jan 11}, abstract = {

Recurrent connections are thought to be a common feature of the neural circuits that encode memories, but how memories are laid down in such circuits is not fully understood. Here we present evidence that courtship memory in Drosophila relies on the recurrent circuit between mushroom body gamma (MBg), M6 output, and aSP13 dopaminergic neurons. We demonstrate persistent neuronal activity of aSP13 neurons and show that it transiently potentiates synaptic transmission from MBγ\>M6 neurons. M6 neurons in turn provide input to aSP13 neurons, prolonging potentiation of MBγ\>M6 synapses over time periods that match short-term memory. These data support a model in which persistent aSP13 activity within a recurrent circuit lays the foundation for a short-term memory.

}, issn = {2050-084X}, doi = {10.7554/eLife.31425}, author = {Zhao, Xiaoliang and Lenek, Daniela and Dag, Ugur and Dickson, Barry and Keleman, Krystyna} } @article {49121, title = {Photoactivatable drugs for nicotinic optopharmacology.}, journal = {Nature Methods}, volume = {15}, year = {2018}, month = {2018 Mar 26}, pages = {347-50}, abstract = {

Photoactivatable pharmacological agents have revolutionized neuroscience, but the palette of available compounds is limited. We describe a general method for caging tertiary amines by using a stable quaternary ammonium linkage that elicits a red shift in the activation wavelength. We prepared a photoactivatable nicotine (PA-Nic), uncageable via one- or two-photon excitation, that is useful to study nicotinic acetylcholine receptors (nAChRs) in different experimental preparations and spatiotemporal scales.

}, issn = {1548-7105}, doi = {10.1038/nmeth.4637}, author = {Banala, Sambashiva and Arvin, Matthew C and Bannon, Nicholas M and Jin, Xiao-Tao and Macklin, John J and Wang, Yong and Peng, Can and Zhao, Guiqing and Marshall, John J and Gee, Kyle R and Wokosin, David L and Kim, Veronica J and McIntosh, J Michael and Contractor, Anis and Lester, Henry A and Kozorovitskiy, Yevgenia and Drenan, Ryan M and Lavis, Luke D} } @article {49487, title = {A practical guide to adaptive light-sheet microscopy.}, journal = {Nature Protocols}, volume = {13}, year = {2018}, month = {2018 Oct 26}, pages = {2462{\textendash}2500}, abstract = {

We describe the implementation and use of an adaptive imaging framework for optimizing spatial resolution and signal strength in a light-sheet microscope. The framework, termed AutoPilot, comprises hardware and software modules for automatically measuring and compensating for mismatches between light-sheet and detection focal planes in living specimens. Our protocol enables researchers to introduce adaptive imaging capabilities in an existing light-sheet microscope or use our SiMView microscope blueprint to set up a new adaptive multiview light-sheet microscope. The protocol describes (i) the mechano-optical implementation of the adaptive imaging hardware, including technical drawings for all custom microscope components; (ii) the algorithms and software library for automated adaptive imaging, including the pseudocode and annotated source code for all software modules; and (iii) the execution of adaptive imaging experiments, as well as the configuration and practical use of the AutoPilot framework. Setup of the adaptive imaging hardware and software takes 1-2 weeks each. Previous experience with light-sheet microscopy and some familiarity with software engineering and building of optical instruments are recommended. Successful implementation of the protocol recovers near diffraction-limited performance in many parts of typical multicellular organisms studied with light-sheet microscopy, such as fruit fly and zebrafish embryos, for which resolution and signal strength are improved two- to fivefold.

}, issn = {1750-2799}, doi = {10.1038/s41596-018-0043-4}, author = {Royer, Lo{\"\i}c A and Lemon, William C and Chhetri, Raghav K and Keller, Philipp J} } @article {49012, title = {A preferred curvature-based continuum mechanics framework for modeling embryogenesis.}, journal = {Biophysical Journal}, volume = {114}, year = {2018}, month = {2018 Jan 23}, pages = {267-77}, abstract = {

Mechanics plays a key role in the development of higher organisms. However, understanding this relationship is complicated by the difficulty of modeling the link between local forces generated at the subcellular level and deformations observed at the tissue and whole-embryo levels. Here we propose an approach first developed for lipid bilayers and cell membranes, in which force-generation by cytoskeletal elements enters a continuum mechanics formulation for the full system in the form of local changes in preferred curvature. This allows us to express and solve the system using only tissue strains. Locations of preferred curvature are simply related to products of gene expression. A solution, in that context, means relaxing the system{\textquoteright}s mechanical energy to yield global morphogenetic predictions that accommodate a tendency toward the local preferred curvature, without a need to explicitly model force-generation mechanisms at the molecular level. Our computational framework, which we call SPHARM-MECH, extends a 3D spherical harmonics parameterization known as SPHARM to combine this level of abstraction with a sparse shape representation. The integration of these two principles allows computer simulations to be performed in three dimensions on highly complex shapes, gene expression patterns, and mechanical constraints. We demonstrate our approach by modeling mesoderm invagination in the fruit-fly embryo, where local forces generated by the acto-myosin meshwork in the region of the future mesoderm lead to formation of a ventral tissue fold. The process is accompanied by substantial changes in cell shape and long-range cell movements. Applying SPHARM-MECH to whole-embryo live imaging data acquired with light-sheet microscopy reveals significant correlation between calculated and observed tissue movements. Our analysis predicts the observed cell shape anisotropy on the ventral side of the embryo and suggests an active mechanical role of mesoderm invagination in supporting the onset of germ-band extension.

}, doi = {10.1016/j.bpj.2017.11.015}, author = {Khairy, Khaled and Lemon, William C and Amat, Fernando and Keller, Philipp J} } @article {49233, title = {Reinforcement signaling of punishment versus relief in fruit flies.}, journal = {Learning \& Memory (Cold Spring Harbor, N.Y.)}, volume = {25}, year = {2018}, month = {2018 Jun}, pages = {247-257}, abstract = {

Painful events establish opponent memories: cues that precede pain are remembered negatively, whereas cues that follow pain, thus coinciding with relief are recalled positively. How do individual reinforcement-signaling neurons contribute to this "timing-dependent valence-reversal?" We addressed this question using an optogenetic approach in the fruit fly. Two types of fly dopaminergic neuron, each comprising just one paired cell, indeed established learned avoidance of odors that preceded their photostimulation during training, and learned approach to odors that followed the photostimulation. This is in striking parallel to punishment versus relief memories reinforced by a real noxious event. For only one of these neuron types, both effects were strong enough for further analyses. Notably, interfering with dopamine biosynthesis in these neurons partially impaired the punishing effect, but not the relieving after-effect of their photostimulation. We discuss how this finding constraints existing computational models of punishment versus relief memories and introduce a new model, which also incorporates findings from mammals. Furthermore, whether using dopaminergic neuron photostimulation or a real noxious event, more prolonged punishment led to stronger relief. This parametric feature of relief may also apply to other animals and may explain particular aspects of related behavioral dysfunction in humans.

}, issn = {1549-5485}, doi = {10.1101/lm.047308.118}, author = {K{\"o}nig, Christian and Khalili, Afshin and Ganesan, Mathangi and Nishu, Amrita P and Garza, Alejandra P and Niewalda, Thomas and Gerber, Bertram and Aso, Yoshinori and Yarali, Ayse} } @article {49013, title = {Repetitive aggressive encounters generate a long-lasting internal state in Drosophila melanogaster males.}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {115}, year = {2018}, month = {2018 Jan 16}, pages = {1099-104}, abstract = {

Multiple studies have investigated the mechanisms of aggressive behavior in Drosophila; however, little is known about the effects of chronic fighting experience. Here, we investigated if repeated fighting encounters would induce an internal state that could affect the expression of subsequent behavior. We trained wild-type males to become winners or losers by repeatedly pairing them with hypoaggressive or hyperaggressive opponents, respectively. As described previously, we observed that chronic losers tend to lose subsequent fights, while chronic winners tend to win them. Olfactory conditioning experiments showed that winning is perceived as rewarding, while losing is perceived as aversive. Moreover, the effect of chronic fighting experience generalized to other behaviors, such as gap-crossing and courtship. We propose that in response to repeatedly winning or losing aggressive encounters, male flies form an internal state that displays persistence and generalization; fight outcomes can also have positive or negative valence. Furthermore, we show that the activities of the PPL1-γ1pedc dopaminergic neuron and the MBON-γ1pedc\>α/β mushroom body output neuron are required for aversion to an olfactory cue associated with losing fights.

}, issn = {1091-6490}, doi = {10.1073/pnas.1716612115}, author = {Kim, Yong-Kyu and Saver, Mathias and Simon, Jasper and Kent, Clement F and Shao, Lisha and Eddison, Mark and Agrawal, Pavan and Texada, Michael and Truman, James W and Heberlein, Ulrike} } @article {49060, title = {A robotic multidimensional directed evolution approach applied to fluorescent voltage reporters.}, journal = {Nature Chemical Biology}, year = {2018}, month = {2018 Feb 26}, abstract = {

We developed a new way to engineer complex proteins toward multidimensional specifications using a simple, yet scalable, directed evolution strategy. By robotically picking mammalian cells that were identified, under a microscope, as expressing proteins that simultaneously exhibit several specific properties, we can screen hundreds of thousands of proteins in a library in just a few hours, evaluating each along multiple performance axes. To demonstrate the power of this approach, we created\ a genetically encoded fluorescent voltage indicator, simultaneously optimizing its\ brightness and membrane localization using our microscopy-guided cell-picking strategy. We produced the high-performance opsin-based fluorescent voltage reporter Archon1 and demonstrated its utility by imaging spiking and millivolt-scale subthreshold and synaptic activity in acute mouse brain slices and in larval zebrafish in vivo. We also measured postsynaptic responses downstream of optogenetically controlled neurons in C. elegans.

}, issn = {1552-4469}, doi = {10.1038/s41589-018-0004-9}, author = {Piatkevich, Kiryl D and Jung, Erica E and Straub, Christoph and Linghu, Changyang and Park, Demian and Suk, Ho-Jun and Hochbaum, Daniel R and Goodwin, Daniel and Pnevmatikakis, Eftychios and Pak, Nikita and Kawashima, Takashi and Yang, Chao-Tsung and Jeff L Rhoades and Shemesh, Or and Asano, Shoh and Yoon, Young-Gyu and Freifeld, Limor and Saulnier, Jessica L and Riegler, Clemens and Engert, Florian and Hughes, Thom and Drobizhev, Mikhail and Szabo, Balint and Ahrens, Misha B and Flavell, Steven W and Sabatini, Bernardo L and Boyden, Edward S} } @article {48702, title = {The role of the serotonergic system in motor control.}, journal = {Neuroscience Research}, volume = {129}, year = {2018}, month = {2018 Apr}, pages = {32-9}, abstract = {

The serotonergic system in the vertebrate brain is implicated in various behaviors and diseases. Its involvement in motor control has been studied for over half a century, but efforts to build a unified model of its functions have been hampered due to the complexity of serotonergic neuromodulation. This review summarizes the anatomical structure of the serotonergic system, its afferent and efferent connections to other brain regions, and recent insights into the sensorimotor computations in the serotonergic system, and considers future research directions into the roles of serotonergic system in motor control.

}, doi = {10.1016/j.neures.2017.07.005}, author = {Kawashima, Takashi} } @article {49196, title = {Schaffer collateral inputs to CA1 excitatory and inhibitory neurons follow different connectivity rules.}, journal = {The Journal of Neuroscience : the official journal of the Society for Neuroscience}, volume = {38}, year = {2018}, month = {2018 May 04}, pages = {5140-52}, abstract = {

Neural circuits, governed by a complex interplay between excitatory and inhibitory neurons, are the substrate for information processing, and the organization of synaptic connectivity in neural network is an important determinant of circuit function. Here, we analyzed the fine structure of connectivity in hippocampal CA1 excitatory and inhibitory neurons innervated by Schaffer collaterals (SCs) using mGRASP in male mice. Our previous study revealed spatially structured synaptic connectivity between CA3-CA1 pyramidal cells (PCs). Surprisingly, parvalbumin-positive interneurons (PVs) showed a significantly more random pattern spatial structure. Notably, application of Peters{\textquoteright} Rule for synapse prediction by random overlap between axons and dendrites enhanced structured connectivity in PCs, but, by contrast, made the connectivity pattern in PVs more random. In addition, PCs in a deep sublayer of striatum pyramidale appeared more highly structured than PCs in superficial layers, and little or no sublayer specificity was found in PVs. Our results show that CA1 excitatory PCs and inhibitory PVs innervated by the same SC inputs follow different connectivity rules. The different organizations of fine scale structured connectivity in hippocampal excitatory and inhibitory neurons provide important insights into the development and functions of neural networks.Understanding how neural circuits generate behavior is one of the central goals of neuroscience. An important component of this endeavor is the mapping of fine-scale connection patterns that underlie, and help us infer, signal processing in the brain. Here, using our recently developed synapse detection technology (mGRASP and neuTube), we provide detailed profiles of synaptic connectivity in excitatory (CA1 pyramidal) and inhibitory (CA1 parvalbumin-positive) neurons innervated by the same presynaptic inputs (CA3 Schaffer collaterals). Our results reveal that these two types of CA1 neurons follow different connectivity patterns. Our new evidence for differently structured connectivity at a fine scale in hippocampal excitatory and inhibitory neurons provides a better understanding of hippocampal networks and will guide theoretical and experimental studies.

}, issn = {1529-2401}, doi = {10.1523/JNEUROSCI.0155-18.2018}, author = {Kwon, Osung and Feng, Linqing and Druckmann, Shaul and Kim, Jinhyun} } @article {49523, title = {Sensorimotor pathway controlling stopping behavior during chemotaxis in the larva.}, journal = {eLife}, volume = {7}, year = {2018}, month = {2018 Nov 22}, abstract = {

Sensory navigation results from coordinated transitions between distinct behavioral programs. During chemotaxis in the larva, the detection of positive odor gradients extends runs while negative gradients promote stops and turns. This algorithm represents a foundation for the control of sensory navigation across phyla. In the present work, we identified an olfactory descending neuron, PDM-DN, which plays a pivotal role in the organization of stops and turns in response to the detection of graded changes in odor concentrations. Artificial activation of this descending neuron induces deterministic stops followed by the initiation of turning maneuvers through head casts. Using electron microscopy, we reconstructed the main pathway that connects the PDM-DN neuron to the peripheral olfactory system and to the pre-motor circuit responsible for the actuation of forward peristalsis. Our results set the stage for a detailed mechanistic analysis of the sensorimotor conversion of graded olfactory inputs into action selection to perform goal-oriented navigation.

}, issn = {2050-084X}, doi = {10.7554/eLife.38740}, author = {Tastekin, Ibrahim and Khandelwal, Avinash and Tadres, David and Fessner, Nico D and Truman, James W and Zlatic, Marta and Cardona, Albert and Louis, Matthieu} } @article {49495, title = {Shared and distinct transcriptomic cell types across neocortical areas.}, journal = {Nature}, volume = {563}, year = {2018}, month = {2018 Nov}, pages = {72-78}, abstract = {

The neocortex contains a multitude of cell types that are segregated into layers and functionally distinct areas. To investigate the diversity of cell types across the mouse neocortex, here we analysed 23,822 cells from two areas at distant poles of the mouse neocortex: the primary visual cortex and the anterior lateral motor cortex. We define 133 transcriptomic cell types by deep, single-cell RNA sequencing. Nearly all types of GABA (γ-aminobutyric acid)-containing neurons are shared across both areas, whereas most types of glutamatergic neurons were found in one of the two areas. By combining single-cell RNA sequencing and retrograde labelling, we match transcriptomic types of glutamatergic neurons to their long-range projection specificity. Our study establishes a combined transcriptomic and projectional taxonomy of cortical cell types from functionally distinct areas of the adult mouse cortex.

}, issn = {1476-4687}, doi = {10.1038/s41586-018-0654-5}, author = {Tasic, Bosiljka and Yao, Zizhen and Graybuck, Lucas T and Smith, Kimberly A and Nguyen, Thuc Nghi and Bertagnolli, Darren and Goldy, Jeff and Garren, Emma and Economo, Michael N and Viswanathan, Sarada and Penn, Osnat and Bakken, Trygve and Menon, Vilas and Miller, Jeremy and Fong, Olivia and Hirokawa, Karla E and Lathia, Kanan and Rimorin, Christine and Tieu, Michael and Larsen, Rachael and Casper, Tamara and Barkan, Eliza and Kroll, Matthew and Parry, Sheana and Shapovalova, Nadiya V and Hirschstein, Daniel and Pendergraft, Julie and Sullivan, Heather A and Kim, Tae Kyung and Szafer, Aaron and Dee, Nick and Groblewski, Peter and Wickersham, Ian and Cetin, Ali and Harris, Julie A and Levi, Boaz P and Sunkin, Susan M and Madisen, Linda and Daigle, Tanya L and Looger, Loren and Bernard, Amy and Phillips, John and Lein, Ed and Hawrylycz, Michael and Svoboda, Karel and Jones, Allan R and Koch, Christof and Zeng, Hongkui} } @article {49046, title = {Single excitatory axons form clustered synapses onto CA1 pyramidal cell dendrites.}, journal = {Nature Neuroscience}, volume = {21}, year = {2018}, month = {2018 Mar}, pages = {353-63}, abstract = {

CA1 pyramidal neurons are a major output of the hippocampus and encode features of experience that constitute episodic memories. Feature-selective firing of these neurons results from the dendritic integration of inputs from multiple brain regions. While it is known that synchronous activation of spatially clustered inputs can contribute to firing through the generation of dendritic spikes, there is no established mechanism for spatiotemporal synaptic clustering. Here we show that single presynaptic axons form multiple, spatially clustered inputs onto the distal, but not proximal, dendrites of CA1 pyramidal neurons. These compound connections exhibit ultrastructural features indicative of strong synapses and occur much more commonly in entorhinal than in thalamic afferents. Computational simulations revealed that compound connections depolarize dendrites in a biophysically efficient manner, owing to their inherent spatiotemporal clustering. Our results suggest that distinct afferent projections use different connectivity motifs that differentially contribute to dendritic integration.

}, doi = {10.1038/s41593-018-0084-6}, author = {Bloss, Erik B and Cembrowski, Mark S and Karsh, Bill and Colonell, Jennifer and Fetter, Richard D and Spruston, Nelson} } @article {49485, title = {Social isolation-induced epigenetic and transcriptional changes in Drosophila dopaminergic neurons.}, journal = {bioRxiv}, year = {2018}, month = {2018 Oct 14}, abstract = {Epigenetic mechanisms play fundamental roles in brain function and behavior and stressors such as social isolation can alter animal behavior via epigenetic mechanisms. However, due to cellular heterogeneity, identifying cell-type-specific epigenetic changes in the brain is challenging. Here we report first use of a modified INTACT method in behavioral epigenetics of Drosophila: a method we call mini-INTACT. Using ChIP-seq on mini-INTACT purified dopaminergic nuclei, we identified epigenetic signatures in socially-isolated and socially-enriched Drosophila males. Social experience altered the epigenetic landscape in clusters of genes involved in transcription and neural function. Some of these alterations were predicted by expression changes of four transcription factors and the prevalence of their binding sites in several clusters. These transcription factors were previously identified as activity-regulated genes and their knockdown in dopaminergic neurons reduced the effects of social experience on sleep. Our work enables the use of Drosophila as a model for cell-type-specific behavioral epigenetics. }, doi = {10.1101/443226}, author = {Agrawal, Pavan and Chung, Phuong and Heberlein, Ulrike and Kent, Clement F} } @article {67288, title = {Specialized and spatially organized coding of sensory, motor, and cognitive variables in midbrain dopamine neurons}, journal = {bioRxiv}, year = {2018}, month = {10/2018}, abstract = {

There is increased appreciation that dopamine (DA) neurons in the midbrain respond not only to reward 1,2 and reward-predicting cues 1,3,4, but also to other variables such as distance to reward 5, movements 6{\textendash}11 and behavioral choices 12{\textendash}15. Based on these findings, a major open question is how the responses to these diverse variables are organized across the population of DA neurons. In other words, do individual DA neurons multiplex multiple variables, or are subsets of neurons specialized in encoding specific behavioral variables? The reason that this fundamental question has been difficult to resolve is that recordings from large populations of individual DA neurons have not been performed in a behavioral task with sufficient complexity to examine these diverse variables simultaneously. To address this gap, we used 2-photon calcium imaging through an implanted lens to record activity of \>300 midbrain DA neurons in the VTA during a complex decision-making task. As mice navigated in a virtual reality (VR) environment, DA neurons encoded an array of sensory, motor, and cognitive variables. These responses were functionally clustered, such that subpopulations of neurons transmitted information about a subset of behavioral variables, in addition to encoding reward. These functional clusters were spatially organized, such that neighboring neurons were more likely to be part of the same cluster. Taken together with the topography between DA neurons and their projections, this specialization and anatomical organization may aid downstream circuits in correctly interpreting the wide range of signals transmitted by DA neurons.

}, doi = {10.1101/456194}, url = {https://www.biorxiv.org/content/early/2018/10/29/456194}, author = {Engelhard, Ben and Finkelstein, Joel and Cox, Julia and Fleming, Weston and Jang, Hee Jae and Ornelas, Sharon and Koay, Sue Ann and Thiberge, Stephan and Daw, Nathaniel and Tank, David W. and Witten, Ilana B.} } @article {48984, title = {Stability, affinity and chromatic variants of the glutamate sensor iGluSnFR.}, journal = {Nature Methods}, volume = {15}, year = {2018}, month = {2018 Nov}, pages = {9386-9}, abstract = {

Single-wavelength fluorescent reporters allow visualization of specific neurotransmitters with high spatial and temporal resolution. We report variants of intensity-based glutamate-sensing fluorescent reporter (iGluSnFR) that are functionally brighter; detect submicromolar to millimolar amounts of glutamate; and have blue, cyan, green, or yellow emission profiles. These variants could be imaged in vivo in cases where original iGluSnFR was too dim, resolved glutamate transients in dendritic spines and axonal boutons, and allowed imaging at kilohertz rates.

}, doi = {10.1038/s41592-018-0171-3}, author = {Marvin, Jonathan S and Scholl, Benjamin and Wilson, Daniel E and Podgorski, Kaspar and Kazemipour, Abbas and Mueller, Johannes Alexander and Schoch-McGovern, Susanne and Wang, Samuel S-H and Quiroz, Francisco Jose Urra and Rebola, Nelson and Bao, Huan and Little, Justin P and Tkachuk, Ariana N and Hantman, Adam W and Chapman, Edwin R and Dietrich, Dirk and DiGregorio, David A and Fitzpatrick, David and Looger, Loren L} } @article {49152, title = {Structural mechanism of functional modulation by gene splicing in NMDA receptors.}, journal = {Neuron}, volume = {98}, year = {2018}, month = {2018 Apr 10}, pages = {521-9}, abstract = {

Alternative gene splicing gives rise to N-methyl-D-aspartate (NMDA) receptor ion channels with defined functional properties and unique contributions to calcium signaling in a given chemical environment in the mammalian brain. Splice variants possessing the exon-5-encoded motif at the amino-terminal domain (ATD) of the GluN1 subunit are known to display robustly altered deactivation rates and pH sensitivity, but the underlying mechanism for this functional modification is largely unknown. Here, we show through cryoelectron microscopy (cryo-EM) that the presence of the exon 5 motif in GluN1 alters the local architecture of heterotetrameric GluN1-GluN2 NMDA receptors and creates contacts with the ligand-binding domains (LBDs) of the GluN1 and GluN2 subunits, which are absent in NMDA receptors lacking the exon 5 motif. The unique interactions established by the exon 5 motif are essential to the stability of the ATD/LBD and LBD/LBD interfaces that are critically involved in controlling proton sensitivity and deactivation.

}, issn = {1097-4199}, doi = {10.1016/j.neuron.2018.03.034}, author = {Regan, Michael C and Grant, Timothy and McDaniel, Miranda J and Karakas, Erkan and Zhang, Jing and Traynelis, Stephen F and Grigorieff, Nikolaus and Furukawa, Hiro} } @article {49574, title = {Structure of Plasmodium falciparum Rh5-CyRPA-Ripr invasion complex.}, journal = {Nature}, volume = {565}, year = {2018}, month = {2018 Dec 12}, pages = {118-21}, abstract = {

Plasmodium falciparum causes the severe form of malaria that has high levels of mortality in humans. Blood-stage merozoites of P. falciparum invade erythrocytes, and this requires interactions between multiple ligands from the parasite and receptors in hosts. These interactions include the binding of the Rh5-CyRPA-Ripr complex with the erythrocyte receptor basigin, which is an essential step for entry into human erythrocytes. Here we show that the Rh5-CyRPA-Ripr complex binds the erythrocyte cell line JK-1 significantly better than does Rh5 alone, and that this binding occurs through the insertion of Rh5 and Ripr into host membranes as a complex with high molecular weight. We report a cryo-electron microscopy structure of the Rh5-CyRPA-Ripr complex at subnanometre resolution, which reveals the organization of this essential invasion complex and the mode of interactions between members of the complex, and shows that CyRPA is a critical mediator of complex assembly. Our structure identifies blades 4-6 of the β-propeller of CyRPA as contact sites for Rh5 and Ripr. The limited contacts between Rh5-CyRPA and CyRPA-Ripr are consistent with the dissociation of Rh5 and Ripr from CyRPA for membrane insertion. A comparision of the crystal structure of Rh5-basigin with the cryo-electron microscopy structure of Rh5-CyRPA-Ripr suggests that Rh5 and Ripr are positioned parallel to the erythrocyte membrane before membrane insertion. This provides information on the function of this complex, and thereby provides insights into invasion by P. falciparum.

}, issn = {1476-4687}, doi = {10.1038/s41586-018-0779-6}, author = {Wong, Wilson and Huang, Rick and Menant, S{\'e}bastien and Hong, Chuan and Sandow, Jarrod J and Birkinshaw, Richard W and Healer, Julie and Hodder, Anthony N and Kanjee, Usheer and Tonkin, Christopher J and Heckmann, Denise and Soroka, Vladislav and S{\o}gaard, Teit Max Moscote and J{\o}rgensen, Thomas and Duraisingh, Manoj T and Czabotar, Peter E and de Jongh, Willem A and Tham, Wai-Hong and Webb, Andrew I and Yu, Zhiheng and Cowman, Alan F} } @article {49371, title = {Structure of the mammalian TRPM7, a magnesium channel required during embryonic development.}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {115}, year = {2018}, month = {2018 Aug 14}, pages = {E8201-10}, abstract = {

The transient receptor potential ion channel subfamily M, member 7 (TRPM7), is a ubiquitously expressed protein that is required for mouse embryonic development. TRPM7 contains both an ion channel and an α-kinase. The channel domain comprises a nonselective cation channel with notable permeability to Mg and Zn Here, we report the closed state structures of the mouse TRPM7 channel domain in three different ionic conditions to overall resolutions of 3.3, 3.7, and 4.1 {\r A}. The structures reveal key residues for an ion binding site in the selectivity filter, with proposed partially hydrated Mg ions occupying the center of the conduction pore. In high [Mg], a prominent external disulfide bond is found in the pore helix, which is essential for ion channel function. Our results provide a structural framework for understanding the TRPM1/3/6/7 subfamily and extend the knowledge base upon which to study the diversity and evolution of TRP channels.

}, issn = {1091-6490}, doi = {10.1073/pnas.1810719115}, author = {Duan, Jingjing and Li, Zongli and Li, Jian and Hulse, Raymond E and Santa-Cruz, Ana and Valinsky, William C and Abiria, Sunday A and Krapivinsky, Grigory and Zhang, Jin and Clapham, David E} } @article {49292, title = {Synaptic partner prediction from point annotations in insect brains.}, journal = {MICCAI 2018: Medical Image Computing and Computer Assisted Intervention}, year = {2018}, month = {2018 Sep 26}, abstract = {

High-throughput electron microscopy allows recording of lar- ge stacks of neural tissue with sufficient resolution to extract the wiring diagram of the underlying neural network. Current efforts to automate this process focus mainly on the segmentation of neurons. However, in order to recover a wiring diagram, synaptic partners need to be identi- fied as well. This is especially challenging in insect brains like Drosophila melanogaster, where one presynaptic site is associated with multiple post- synaptic elements. Here we propose a 3D U-Net architecture to directly identify pairs of voxels that are pre- and postsynaptic to each other. To that end, we formulate the problem of synaptic partner identification as a classification problem on long-range edges between voxels to encode both the presence of a synaptic pair and its direction. This formulation allows us to directly learn from synaptic point annotations instead of more ex- pensive voxel-based synaptic cleft or vesicle annotations. We evaluate our method on the MICCAI 2016 CREMI challenge and improve over the current state of the art, producing 3\% fewer errors than the next best method.

}, doi = {10.1007/978-3-030-00934-2_35}, author = {Buhmann, Julia and Krause, Renate and Lentini, Rodrigo Ceballos and Eckstein, Nils and Cook, Matthew and Turaga, Srinivas C and Funke, Jan} } @article {49470, title = {Thy1 transgenic mice expressing the red fluorescent calcium indicator jRGECO1a for neuronal population imaging in vivo.}, journal = {PloS One}, volume = {13}, year = {2018}, month = {2018}, pages = {e0205444}, abstract = {

Calcium imaging is commonly used to measure the neural activity of large groups of neurons in mice. Genetically encoded calcium indicators (GECIs) can be delivered for this purpose using non-invasive genetic methods. Compared to viral gene transfer, transgenic targeting of GECIs provides stable long-term expression and obviates the need for invasive viral injections. Transgenic mice expressing the green GECI GCaMP6 are already widely used. Here we present the generation and characterization of transgenic mice expressing the sensitive red GECI jRGECO1a, driven by the Thy1 promoter. Four transgenic lines with different expression patterns showed sufficiently high expression for cellular in vivo imaging. We used two-photon microscopy to characterize visual responses of individual neurons in the visual cortex in vivo. The signal-to-noise ratio in transgenic mice was comparable to, or better than, mice transduced with adeno-associated virus. In addition, we show that Thy1-jRGECO1a transgenic mice are useful for transcranial population imaging and functional mapping using widefield fluorescence microscopy. We also demonstrate imaging of visual responses in retinal ganglion cells in vitro. Thy1-jRGECO1a transgenic mice are therefore a useful addition to the toolbox for imaging activity in intact neural networks.

}, issn = {1932-6203}, doi = {10.1371/journal.pone.0205444}, author = {Dana, Hod and Novak, Ondrej and Guardado-Montesino, Michael and Fransen, James W and Hu, Amy and Borghuis, Bart G and Guo, Caiying and Kim, Douglas S and Svoboda, Karel} } @article {65093, title = {Ultralarge modulation of fluorescence by neuromodulators in carbon nanotubes functionalized with self-assembled oligonucleotide rings.}, journal = {Nano Letters}, volume = {18}, year = {2018}, month = {2018 Nov 14}, pages = {6995-7003}, abstract = {

Noncovalent interactions between single-stranded DNA (ssDNA) oligonucleotides and single wall carbon nanotubes (SWNTs) have provided a unique class of tunable chemistries for a variety of applications. However, mechanistic insight into both the photophysical and intermolecular phenomena underlying their utility is lacking, which results in obligate heuristic approaches for producing ssDNA-SWNT based technologies. In this work, we present an ultrasensitive "turn-on" nanosensor for neuromodulators dopamine and norepinephrine with strong relative change in fluorescence intensity (Δ F/ F) of up to 3500\%, a signal appropriate for in vivo neuroimaging, and uncover the photophysical principles and intermolecular interactions that govern the molecular recognition and fluorescence modulation of this nanosensor synthesized from the spontaneous self-assembly of (GT) ssDNA rings on SWNTs. The fluorescence modulation of the ssDNA-SWNT conjugate is shown to exhibit remarkable sensitivity to the ssDNA sequence chemistry, length, and surface density, providing a set of parameters with which to tune nanosensor dynamic range, analyte selectivity\ and strength of fluorescence turn-on. We employ classical and quantum mechanical molecular dynamics simulations to rationalize our experimental findings. Calculations show that (GT) ssDNA form ordered rings around (9,4)\ SWNTs, inducing periodic surface potentials that modulate exciton recombination lifetimes. Further evidence is presented to elucidate how dopamine analyte binding modulates SWNT fluorescence. We discuss the implications of our findings for SWNT-based molecular imaging applications.

}, keywords = {Biosensing Techniques, DNA, Single-Stranded, Dopamine, Fluorescence, Nanotubes, Carbon, Neurotransmitter Agents, Norepinephrine, Oligonucleotides}, issn = {1530-6992}, doi = {10.1021/acs.nanolett.8b02937}, author = {Beyene, Abraham G and Alizadehmojarad, Ali A and Dorlhiac, Gabriel and Goh, Natalie and Streets, Aaron M and Kr{\'a}l, Petr and Vukovi{\'c}, Lela and Landry, Markita P} } @article {48839, title = {Vision and locomotion shape the interactions between neuron types in mouse visual cortex.}, journal = {Neuron}, volume = {98}, year = {2018}, month = {2018 May 2}, pages = {602-15}, abstract = {

Cortical computation arises from the interaction of multiple neuronal\ types, including pyramidal (Pyr) cells and interneurons expressing Sst, Vip, or Pvalb. To study the circuit underlying such\ interactions, we imaged these four\ types\ of cells in\ mouse\ primary\ visual\ cortex(V1). Our recordings in darkness were consistent with a "disinhibitory" model in which\ locomotion\ activates Vip cells, thus inhibiting Sst cells and disinhibiting Pyr cells. However, the disinhibitory model failed when\ visual\ stimuli were present:\ locomotion\ increased Sst cell responses to large stimuli and Vip cell responses to small stimuli. A recurrent network model successfully predicted each cell type{\textquoteright}s activity from the measured activity of other\ types. Capturing the effects of\ locomotion, however, required allowing it to increase feedforward synaptic weights and modulate recurrent weights. This network model summarizes interneuron\ interactions\ and suggests that\ locomotion\ may alter cortical computation by changing effective synaptic connectivity.

}, doi = {https://doi.org/10.1101/058396}, author = {Dipoppa, Mario and Ranson, Adam and Krumin, Michael and Pachitariu, Marius and Carandini, Matteo and Harris, Kenneth D} } @article {49518, title = {Visual physiology of the layer 4 cortical circuit in silico.}, journal = {PLoS Computational Biology}, volume = {14}, year = {2018}, month = {2018 Nov 12}, pages = {e1006535}, abstract = {

Despite advances in experimental techniques and accumulation of large datasets concerning the composition and properties of the cortex, quantitative modeling of cortical circuits under in-vivo-like conditions remains challenging. Here we report and publicly release a biophysically detailed circuit model of layer 4 in the mouse primary visual cortex, receiving thalamo-cortical visual inputs. The 45,000-neuron model was subjected to a battery of visual stimuli, and results were compared to published work and new in vivo experiments. Simulations reproduced a variety of observations, including effects of optogenetic perturbations. Critical to the agreement between responses in silico and in vivo were the rules of functional synaptic connectivity between neurons. Interestingly, after extreme simplification the model still performed satisfactorily on many measurements, although quantitative agreement with experiments suffered. These results emphasize the importance of functional rules of cortical wiring and enable a next generation of data-driven models of in vivo neural activity and computations.

}, issn = {1553-7358}, doi = {10.1371/journal.pcbi.1006535}, author = {Arkhipov, Anton and Gouwens, Nathan W and Billeh, Yazan N and Gratiy, Sergey and Iyer, Ramakrishnan and Wei, Ziqiang and Xu, Zihao and Abbasi-Asl, Reza and Berg, Jim and Buice, Michael and Cain, Nicholas and da Costa, Nuno and de Vries, Saskia and Denman, Daniel and Durand, Severine and Feng, David and Jarsky, Tim and Lecoq, Jerome and Lee, Brian and Li, Lu and Mihalas, Stefan and Ocker, Gabriel K and Olsen, Shawn R and Reid, R Clay and Soler-Llavina, Gilberto and Sorensen, Staci A and Wang, Quanxin and Waters, Jack and Scanziani, Massimo and Koch, Christof} } @article {49512, title = {Visualizing intracellular organelle and cytoskeletal interactions at nanoscale resolution on millisecond timescales. }, journal = {Cell}, volume = {175}, year = {2018}, month = {2018 Nov 15}, pages = {1430-42}, abstract = {In eukaryotic cells, organelles and the cytoskeleton undergo highly dynamic yet organized interactions capable of orchestrating complex cellular functions. Visualizing these interactions requires noninvasive, long-duration imaging of the intracellular environment at high spatiotemporal resolution and low background. To achieve these normally opposing goals, we developed grazing incidence structured illumination microscopy (GI-SIM) that is capable of imaging dynamic events near the basal cell cortex at 97-nm resolution and 266 frames/s over thousands of time points. We employed multi-color GI-SIM to characterize the fast dynamic interactions of diverse organelles and the cytoskeleton, shedding new light on the complex behaviors of these structures. Precise measurements of microtubule growth or shrinkage events helped distinguish among models of microtubule dynamic instability. Analysis of endoplasmic reticulum (ER) interactions with other organelles or microtubules uncovered new ER remodeling mechanisms, such as hitchhiking of the ER on motile organelles. Finally, ER-mitochondria contact sites were found to promote both mitochondrial fission and fusion.}, doi = {10.1016/j.cell.2018.09.057}, author = {Guo, Yuting and Li, Di and Zhang, Siwei and Yang, Yanrui and Liu, Jia-Jia and Wang, Xinyu and Liu, Chong and Milkie, Daniel E and Moore, Regan P and Tulu, U Serdar and Kiehart, Daniel P and Hu, Junjie and Lippincott-Schwartz, Jennifer and Betzig, Eric and Li, Dong} } @article {48621, title = {3D Bayesian cluster analysis of super-resolution data reveals LAT recruitment to the T cell synapse.}, journal = {Scientific Reports}, volume = {7}, year = {2017}, month = {2017 Jun 22}, pages = {4077}, abstract = {

Single-molecule localisation microscopy (SMLM) allows the localisation of fluorophores with a precision of 10-30 nm, revealing the cell{\textquoteright}s nanoscale architecture at the molecular level. Recently, SMLM has been extended to 3D, providing a unique insight into cellular machinery. Although cluster analysis techniques have been developed for 2D SMLM data sets, few have been applied to 3D. This lack of quantification tools can be explained by the relative novelty of imaging techniques such as interferometric photo-activated localisation microscopy (iPALM). Also, existing methods that could be extended to 3D SMLM are usually subject to user defined analysis parameters, which remains a major drawback. Here, we present a new open source cluster analysis method for 3D SMLM data, free of user definable parameters, relying on a model-based Bayesian approach which takes full account of the individual localisation precisions in all three dimensions. The accuracy and reliability of the method is validated using simulated data sets. This tool is then deployed on novel experimental data as a proof of concept, illustrating the recruitment of LAT to the T-cell immunological synapse in data acquired by iPALM providing ~10 nm isotropic resolution.

}, issn = {2045-2322}, doi = {10.1038/s41598-017-04450-w}, author = {Griffi{\'e}, Juliette and Shlomovich, Leigh and Williamson, David J and Shannon, Michael and Aaron, Jesse and Khuon, Satya and L Burn, Garth and Boelen, Lies and Peters, Ruby and Cope, Andrew P and Cohen, Edward A K and Rubin-Delanchy, Patrick and Owen, Dylan M} } @article {49082, title = {Advances in neural engineering for rehabilitation.}, journal = {Behavioural Neurology}, volume = {2017}, year = {2017}, month = {2017}, pages = {9240921}, issn = {1875-8584}, doi = {10.1155/2017/9240921}, author = {Hu, Xiaoling and Zhao, Ting and Yao, Jun and Kuang, Yu and Yang, Yuan} } @article {48449, title = {An Amidase_3 domain-containing N-acetylmuramyl-L-alanine amidase is required for mycobacterial cell division.}, journal = {Scientific Reports}, volume = {7}, year = {2017}, month = {2017 Apr 25}, pages = {1140}, abstract = {

Mycobacteria possess a multi-layered cell wall that requires extensive remodelling during cell division. We investigated the role of an amidase_3 domain-containing N-acetylmuramyl-L-alanine amidase, a peptidoglycan remodelling enzyme implicated in cell division. We demonstrated that deletion of MSMEG_6281 (Ami1) in Mycobacterium smegmatis resulted in the formation of cellular chains, illustrative of cells that were unable to complete division. Suprisingly, viability in the Δami1 mutant was maintained through atypical lateral branching, the products of which proceeded to form viable daughter cells. We showed that these lateral buds resulted from mislocalization of DivIVA, a major determinant in facilitating polar elongation in mycobacterial cells. Failure of Δami1 mutant cells to separate also led to dysregulation of FtsZ ring bundling. Loss of Ami1 resulted in defects in septal peptidoglycan turnover with release of excess cell wall material from the septum or newly born cell poles. We noted signficant accumulation of 3-3 crosslinked muropeptides in the Δami1 mutant. We further demonstrated that deletion of ami1 leads to increased cell wall permeability and enhanced susceptiblity to cell wall targeting antibiotics. Collectively, these data provide novel insight on cell division in actinobacteria and highlights a new class of potential drug targets for mycobacterial diseases.

}, issn = {2045-2322}, doi = {10.1038/s41598-017-01184-7}, author = {Senzani, Sibusiso and Li, Dong and Bhaskar, Ashima and Ealand, Christopher and Chang, James and Rimal, Binayak and Liu, Chengyin and Joon Kim, Sung and Dhar, Neeraj and Kana, Bavesh} } @article {67220, title = {Anisotropic EM Segmentation by 3D Affinity Learning and Agglomeration}, journal = {CoRR}, volume = {abs/1707.08935}, year = {2017}, abstract = {

The field of connectomics has recently produced neuron wiring diagrams from relatively large brain regions from multiple animals. Most of these neural reconstructions were computed from isotropic (e.g., FIBSEM) or near isotropic (e.g., SBEM) data. In spite of the remarkable progress on algorithms in recent years, automatic dense reconstruction from anisotropic data remains a challenge for the connectomics community. One significant hurdle in the segmentation of anisotropic data is the difficulty in generating a suitable initial over-segmentation. In this study, we present a segmentation method for anisotropic EM data that agglomerates a 3D over-segmentation computed from the 3D affinity prediction. A 3D U-net is trained to predict 3D affinities by the MALIS approach. Experiments on multiple datasets demonstrates the strength and robustness of the proposed method for anisotropic EM segmentation.

}, url = {https://arxiv.org/abs/1707.08935v2}, author = {Toufiq Parag and Fabian Tschopp and William Grisaitis and Srinivas C. Turaga and Xuewen Zhang and Brian Matejek and Lee Kamentsky and Jeff W. Lichtman and Hanspeter Pfister} } @article {65893, title = {Antibodies to biotin enable large-scale detection of biotinylation sites on proteins}, journal = {Nature Methods}, volume = {14}, year = {2017}, month = {Jan-12-2017}, pages = {1167 - 1170}, abstract = {

Although purification of biotinylated molecules is highly efficient, identifying specific sites of biotinylation remains challenging. We show that anti-biotin antibodies enable unprecedented enrichment of biotinylated peptides from complex peptide mixtures. Live-cell proximity labeling using APEX peroxidase followed by anti-biotin enrichment and mass spectrometry yielded over 1,600 biotinylation sites on hundreds of proteins, an increase of more than 30-fold in the number of biotinylation sites identified compared to streptavidin-based enrichment of proteins.

}, issn = {1548-7091}, doi = {10.1038/nmeth.4465}, url = {http://www.nature.com/articles/nmeth.4465http://www.nature.com/doifinder/10.1038/nmeth.4465http://www.nature.com/articles/nmeth.4465.pdfhttp://www.nature.com/articles/nmeth.4465.pdfhttp://www.nature.com/articles/nmeth.4465}, author = {Udeshi, Namrata D and Pedram, Kayvon and Svinkina, Tanya and Fereshetian, Shaunt and Myers, Samuel A and Aygun, Ozan and Krug, Karsten and Clauser, Karl and Ryan, Dominic and Ast, Tslil and Mootha, Vamsi K and Ting, Alice Y and Carr, Steven A} } @article {48156, title = {Atomic structures of fibrillar segments of hIAPP suggest tightly mated β-sheets are important for cytotoxicity.}, journal = {eLife}, volume = {6}, year = {2017}, month = {2017 Jan 03}, abstract = {

hIAPP fibrils are associated with Type-II Diabetes, but the link of hIAPP structure to islet cell death remains elusive. Here we observe that hIAPP fibrils are cytotoxic to cultured pancreatic β-cells, leading us to determine the structure and cytotoxicity of protein segments composing the amyloid spine of hIAPP. Using the cryoEM method MicroED, we discover that one segment, 19-29 S20G, forms pairs of β-sheets mated by a dry interface that share structural features with and are similarly cytotoxic to full-length hIAPP fibrils. In contrast, a second segment, 15-25 WT, forms non-toxic labile β-sheets. These segments possess different structures and cytotoxic effects, however, both can seed full-length hIAPP, and cause hIAPP to take on the cytotoxic and structural features of that segment. These results suggest that protein segment structures represent polymorphs of their parent protein and that segment 19-29 S20G may serve as a model for the toxic spine of hIAPP.

}, issn = {2050-084X}, doi = {10.7554/eLife.19273}, author = {Krotee, Pascal and Rodriguez, Jose A and Sawaya, Michael R and Cascio, Duilio and Reyes, Francis E and Shi, Dan and Hattne, Johan and Nannenga, Brent L and Oskarsson, Marie E and Philipp, Stephan and Griner, Sarah and Jiang, Lin and Glabe, Charles G and Westermark, Gunilla T and Gonen, Tamir and Eisenberg, David S} } @article {48265, title = {Atomic-resolution structures from fragmented protein crystals with the cryoEM method MicroED.}, journal = {Nature Methods}, volume = {14}, year = {2017}, month = {2017 Feb 13}, pages = {399-402}, abstract = {

Traditionally, crystallographic analysis of macromolecules has depended on large, well-ordered crystals, which often require significant effort to obtain. Even sizable crystals sometimes suffer from pathologies that render them inappropriate for high-resolution structure determination. Here we show that fragmentation of large, imperfect crystals into microcrystals or nanocrystals can provide a simple path for high-resolution structure determination by the cryoEM method MicroED and potentially by serial femtosecond crystallography.

}, issn = {1548-7105}, doi = {10.1038/nmeth.4178}, author = {de la Cruz, M Jason and Hattne, Johan and Shi, Dan and Seidler, Paul and Rodriguez, Jose and Reyes, Francis E and Sawaya, Michael R and Cascio, Duilio and Weiss, Simon C and Kim, Sun Kyung and Hinck, Cynthia S and Hinck, Andrew P and Calero, Guillermo and Eisenberg, David and Gonen, Tamir} } @article {48266, title = {Axonal Endoplasmic Reticulum Ca(2+) Content Controls Release Probability in CNS Nerve Terminals.}, journal = {Neuron}, volume = {93}, year = {2017}, month = {2017 Jan 30}, pages = {867-81}, abstract = {

Although the endoplasmic reticulum (ER) extends throughout axons and axonal ER dysfunction is implicated in numerous neurological diseases, its role at nerve terminals is poorly understood. We developed novel genetically encoded ER-targeted low-affinity Ca(2+) indicators optimized for examining axonal ER Ca(2+). Our experiments revealed that presynaptic function is tightly controlled by ER Ca(2+) content. We found that neuronal activity drives net Ca(2+) uptake into presynaptic ER although this activity does not contribute significantly to shaping cytosolic Ca(2+) except during prolonged repetitive firing. In contrast, we found that axonal ER acts as an actuator of plasma membrane (PM) function: [Ca(2+)]ER controls STIM1 activation in presynaptic terminals, which results in the local modulation of presynaptic function, impacting activity-driven Ca(2+) entry and release probability. These experiments reveal a critical role of presynaptic ER in the control of neurotransmitter release and will help frame future investigations into the molecular basis of ER-driven neuronal disease states.

}, issn = {1097-4199}, doi = {10.1016/j.neuron.2017.01.010}, author = {de Juan-Sanz, Jaime and Holt, Graham T and Schreiter, Eric R and de Juan, Fernando and Kim, Douglas S and Ryan, Timothy A} } @article {48715, title = {Best practices for managing large CryoEM facilities.}, journal = {Journal of Structural Biology}, volume = {199}, year = {2017}, month = {2017-08-04}, pages = {225-36}, abstract = {

This paper provides an overview of the discussion and presentations from the Workshop on the Management of Large CryoEM Facilities held at the New York Structural Biology Center, New York, NY on February 6{\textendash}7, 2017. A major objective of the workshop was to discuss best practices for managing cryoEM facilities. The discussions were largely focused on supporting single-particle methods for cryoEM and topics included: user access, assessing projects, workflow, sample handling, microscopy, data management and processing, and user training.

}, doi = {10.1016/j.jsb.2017.07.011}, author = {Alewijnse, Bart and Ashton, Alun W and Chambers, Melissa G and Chen, Songye and Cheng, Anchi and Ebrahim, Mark and Eng, Edward T and Hagen, Wim JH and Koster, Abraham J and Lopez, Claudia S and Lukoyanova, Natalya and Ortega, Joaquin and Renault, Ludovic and Reyntjens, Steve and Rice, William J and Scapin, Giovanna and Schrijver, Raymond and Siebert, Alistair and Stagg, Scott M and et al} } @article {48645, title = {Building bridges between cellular and molecular structural biology.}, journal = {eLife}, volume = {6}, year = {2017}, month = {2017 Jul 06}, abstract = {

The integration of cellular and molecular structural data is key to understanding the function of macromolecular assemblies and complexes in their in vivo context. Here we report on the outcomes of a workshop that discussed how to integrate structural data from a range of public archives. The workshop identified two main priorities: the development of tools and file formats to support segmentation (that is, the decomposition of a three-dimensional volume into regions that can be associated with defined objects), and the development of tools to support the annotation of biological structures.

}, issn = {2050-084X}, doi = {10.7554/eLife.25835}, author = {Patwardhan, Ardan and Brandt, Robert and Butcher, Sarah J and Collinson, Lucy and Gault, David and Gr{\"u}newald, Kay and Hecksel, Corey and Huiskonen, Juha T and Iudin, Andrii and Jones, Martin L and Korir, Paul K and Koster, Abraham J and Lagerstedt, Ingvar and Lawson, Catherine L and Mastronarde, David and McCormick, Matthew and Parkinson, Helen and Rosenthal, Peter B and Saalfeld, Stephan and Saibil, Helen R and Sarntivijai, Sirarat and Solanes Valero, Irene and Subramaniam, Sriram and Swedlow, Jason R and Tudose, Ilinca and Winn, Martyn and Kleywegt, Gerard J} } @article {65894, title = {A bulky glycocalyx fosters metastasis formation by promoting G1 cell cycle progression}, journal = {eLife}, volume = {6}, year = {2017}, month = {Sep-12-2018}, abstract = {

Metastasis depends upon cancer cell growth and survival within the metastatic niche. Tumors which remodel their glycocalyces, by overexpressing bulky glycoproteins like mucins, exhibit a higher predisposition to metastasize, but the role of mucins in oncogenesis remains poorly understood. Here we report that a bulky glycocalyx promotes the expansion of disseminated tumor cells in vivo by fostering integrin adhesion assembly to permit G1 cell cycle progression. We engineered tumor cells to display glycocalyces of various thicknesses by coating them with synthetic mucin-mimetic glycopolymers. Cells adorned with longer glycopolymers showed increased metastatic potential, enhanced cell cycle progression, and greater levels of integrin-FAK mechanosignaling and Akt signaling in a syngeneic mouse model of metastasis. These effects were mirrored by expression of the ectodomain of cancer-associated mucin MUC1. These findings functionally link mucinous proteins with tumor aggression, and offer a new view of the cancer glycocalyx as a major driver of disease progression.

}, doi = {https://doi.org/10.7554/eLife.25752}, url = {https://elifesciences.org/articles/25752}, author = {Woods, Elliot C and Kai, FuiBoon and Barnes, J Matthew and Pedram, Kayvon and Pickup, Michael W and Hollander, Michael J and Weaver, Valerie M and Bertozzi, Carolyn R} } @article {48960, title = {Cell dynamics underlying oriented growth of the Drosophila wing imaginal disc.}, journal = {Development (Cambridge, England)}, volume = {144}, year = {2017}, month = {2017 Dec 01}, pages = {4406-4421}, abstract = {

Quantitative analysis of the dynamic cellular mechanisms shaping the Drosophila wing during its larval growth phase has been limited, impeding our ability to understand how morphogen patterns regulate tissue shape. Such analysis requires explants to be imaged under conditions that maintain both growth and patterning, as well as methods to quantify how much cellular behaviors change tissue shape. Here, we demonstrate a key requirement for the steroid hormone 20-hydroxyecdysone (20E) in the maintenance of numerous patterning systems in vivo and in explant culture. We find that low concentrations of 20E support prolonged proliferation in explanted wing discs in the absence of insulin, incidentally providing novel insight into the hormonal regulation of imaginal growth. We use 20E-containing media to observe growth directly and to apply recently developed methods for quantitatively decomposing tissue shape changes into cellular contributions. We discover that whereas cell divisions drive tissue expansion along one axis, their contribution to expansion along the orthogonal axis is cancelled by cell rearrangements and cell shape changes. This finding raises the possibility that anisotropic mechanical constraints contribute to growth orientation in the wing disc.

}, keywords = {Animals, Animals, Genetically Modified, Body Patterning, Cell Proliferation, Drosophila melanogaster, Ecdysterone, Gene Expression Profiling, Gene Expression Regulation, Developmental, Genes, Insect, Imaginal Discs, Insulin, Morphogenesis, Signal Transduction, Transcription, Genetic, Wings, Animal}, issn = {1477-9129}, doi = {10.1242/dev.155069}, author = {Dye, Natalie A and Popovi{\'c}, Marko and Spannl, Stephanie and Etournay, Rapha{\"e}l and Kainm{\"u}ller, Dagmar and Ghosh, Suhrid and Myers, Eugene W and J{\"u}licher, Frank and Eaton, Suzanne} } @article {48858, title = {Cell volume change through water efflux impacts cell stiffness and stem cell fate.}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {114}, year = {2017}, month = {2017 Sep 25}, pages = {E8618-27}, abstract = {

Cells alter their mechanical properties in response to their local microenvironment; this plays a role in determining cell function and can even influence stem cell fate. Here, we identify a robust and unified relationship between cell stiffness and cell volume. As a cell spreads on a substrate, its volume decreases, while its stiffness concomitantly increases. We find that both cortical and cytoplasmic cell stiffness scale with volume for numerous perturbations, including varying substrate stiffness, cell spread area, and external osmotic pressure. The reduction of cell volume is a result of water efflux, which leads to a corresponding increase in intracellular molecular crowding. Furthermore, we find that changes in cell volume, and hence stiffness, alter stem-cell differentiation, regardless of the method by which these are induced. These observations reveal a surprising, previously unidentified relationship between cell stiffness and cell volume that strongly influences cell biology.

}, issn = {1091-6490}, doi = {10.1073/pnas.1705179114}, author = {Guo, Ming and Pegoraro, Adrian F and Mao, Angelo and Zhou, Enhua H and Arany, Praveen R and Han, Yulong and Burnette, Dylan T and Jensen, Mikkel H and Kasza, Karen E and Moore, Jeffrey R and Mackintosh, Frederick C and Fredberg, Jeffrey J and Mooney, David J and Lippincott-Schwartz, Jennifer and Weitz, David A} } @article {48751, title = {Circuits in the rodent brainstem that control whisking in concert with other orofacial motor actions.}, journal = {Neuroscience}, volume = {368}, year = {2017}, month = {2017 Aug 23}, pages = {152-70}, abstract = {

The world view of rodents is largely determined by sensation on two length scales. One is within the animal{\textquoteright}s peri-personal space. Sensorimotor control on this scale involves active movements of the nose, tongue, head, and vibrissa, along with sniffing to determine olfactory clues. The second scale involves the detection of more distant space through vision and audition; these detection processes also impact repositioning of the head, eyes, and ears. Here we focus on orofacial motor actions, primarily vibrissa-based touch but including nose twitching, head bobbing, and licking, that control sensation at short, peri-personal distances. The orofacial nuclei for control of the motor plants, as well as primary and secondary sensory nuclei associated with these motor actions, lie within the hindbrain. The current data support three themes: First, the position of the sensors is determined by the summation of two drive signals, i.e., a fast rhythmic component and an evolving orienting component. Second, the rhythmic component is coordinated across all orofacial motor actions and is phase-locked to sniffing as the animal explores. Reverse engineering reveals that the preB{\"o}tzinger inspiratory complex provides the reset to the relevant premotor oscillators. Third, direct feedback from somatosensory trigeminal nuclei can rapidly alter motion of the sensors. This feedback is disynaptic and can be tuned by high-level inputs. The elucidation of synergistic coordination of orofacial motor actions to form behaviors, beyond that of a common rhythmic component, represents a work in progress that encompasses feedback through the midbrain and forebrain as well as hindbrain areas.

}, issn = {1873-7544}, doi = {10.1016/j.neuroscience.2017.08.034}, author = {McElvain, Lauren E and Friedman, Beth and Karten, Harvey J and Svoboda, Karel and Wang, Fan and Desch{\^e}nes, Martin and Kleinfeld, David} } @article {48991, title = {Common fibrillar spines of amyloid-β and human Islet Amyloid Polypeptide revealed by Micro Electron Diffraction and inhibitors developed using structure-based design.}, journal = {The Journal of Biological Chemistry}, volume = {293}, year = {2017}, month = {2017 Dec 27}, pages = {2888-902}, abstract = {

Amyloid-β (Aβ) and human islet amyloid polypeptide (hIAPP) aggregate to form amyloid fibrils that deposit in tissues, and are associated with Alzheimer{\textquoteright}s disease (AD) and Type-II Diabetes (T2D), respectively. Individuals with T2D have an increased risk of developing AD, and conversely, AD patients have an increased risk of developing T2D. Evidence suggests that this link between AD and T2D might originate from a structural similarity between aggregates of Aβ and hIAPP. Using the cryoEM method Micro-Electron Diffraction (MicroED) we determined the atomic structures of 11-residue segments from both Aβ and hIAPP, termed Aβ 24-34 WT and hIAPP 19-29 S20G, with 64\% sequence similarity. We observe a high degree of structural similarity between their backbone atoms (0.96 {\r A} RMSD). Moreover, fibrils of these segments induce amyloid formation through self- and cross-seeding. Furthermore, inhibitors designed for one segment show cross-efficacy for full-length Aβ and hIAPP and reduce cytotoxicity of both proteins, though by apparently blocking different cytotoxic mechanisms. The similarity of the atomic structures of Aβ 24-34 WT and hIAPP 19-29 S20G offers a molecular model for cross-seeding between Aβ and hIAPP.

}, issn = {1083-351X}, doi = {10.1074/jbc.M117.806109}, author = {Krotee, Pascal and Griner, Sarah L and Sawaya, Michael R and Cascio, Duilio and Rodriguez, Jose A and Shi, Dan and Philipp, Stephan and Murray, Kevin and Saelices, Lorena and Lee, Ji and Seidler, Paul and Glabe, Charles G and Jiang, Lin and Gonen, Tamir and Eisenberg, David S} } @article {48488, title = {Confirmation of five novel susceptibility loci for systemic lupus erythematosus (SLE) and integrated network analysis of 82 SLE susceptibility loci.}, journal = {Human Molecular Genetics}, volume = {26}, year = {2017}, month = {2017 Mar 15}, pages = {1205-1216}, abstract = {

We recently identified ten novel SLE susceptibility loci in Asians and uncovered several additional suggestive loci requiring further validation. This study aimed to replicate five of these suggestive loci in a Han Chinese cohort from Hong Kong, followed by meta-analysis (11,656 cases and 23,968 controls) on previously reported Asian and European populations, and to perform bioinformatic analyses on all 82 reported SLE loci to identify shared regulatory signatures. We performed a battery of analyses for these five loci, as well as joint analyses on all 82 SLE loci. All five loci passed genome-wide significance: MYNN (rs10936599, Pmeta = 1.92 {\texttimes} 10-13, OR = 1.14), ATG16L2 (rs11235604, Pmeta = 8.87 {\texttimes} 10 -12, OR = 0.78), CCL22 (rs223881, Pmeta = 5.87 {\texttimes} 10-16, OR = 0.87), ANKS1A (rs2762340, Pmeta = 4.93 {\texttimes} 10-15, OR = 0.87) and RNASEH2C (rs1308020, Pmeta = 2.96 {\texttimes} 10-19, OR = 0.84) and co-located with annotated gene regulatory elements. The novel loci share genetic signatures with other reported SLE loci, including effects on gene expression, transcription factor binding, and epigenetic characteristics. Most (56\%) of the correlated (r2 \> 0.8) SNPs from the 82 SLE loci were implicated in differential expression (9.81 {\texttimes} 10-198 \< P \< 5 {\texttimes} 10-3) of cis-genes. Transcription factor binding sites for p53, MEF2A and E2F1 were significantly (P \< 0.05) over-represented in SLE loci, consistent with apoptosis playing a critical role in SLE. Enrichment analysis revealed common pathways, gene ontology, protein domains, and cell type-specific expression. In summary, we provide evidence of five novel SLE susceptibility loci. Integrated bioinformatics using all 82 loci revealed that SLE susceptibility loci share many gene regulatory features, suggestive of conserved mechanisms of SLE etiopathogenesis.

}, keywords = {Adaptor Proteins, Signal Transducing, Asian Continental Ancestry Group, Autophagy-Related Proteins, Chemokine CCL22, Gene Expression Regulation, Genetic Predisposition to Disease, Genome-Wide Association Study, Genotype, Humans, Kruppel-Like Transcription Factors, Lupus Erythematosus, Systemic, Polymorphism, Single Nucleotide, Ribonuclease H}, issn = {1460-2083}, doi = {10.1093/hmg/ddx026}, author = {Molineros, Julio E and Yang, Wanling and Zhou, Xu-Jie and Sun, Celi and Okada, Yukinori and Zhang, Huoru and Heng Chua, Kek and Lau, Yu-Lung and Kochi, Yuta and Suzuki, Akari and Yamamoto, Kazuhiko and Ma, Jianyang and Bang, So-Young and Lee, Hye-Soon and Kim, Kwangwoo and Bae, Sang-Cheol and Zhang, Hong and Shen, Nan and Looger, Loren L and Nath, Swapan K} } @article {48658, title = {A connectome of a learning and memory center in the adult Drosophila brain.}, journal = {eLife}, volume = {6}, year = {2017}, month = {2017 Jul 18}, pages = {e26975}, abstract = {

Understanding memory formation, storage and retrieval requires knowledge of the underlying neuronal circuits. In\ Drosophila, the mushroom body (MB) is the major site of associative learning. We reconstructed the morphologies and synaptic connections of all 983 neurons within the three functional units, or compartments, that compose the adult MB{\textquoteright}s α lobe, using a dataset of isotropic 8-nm voxels collected by focused ion-beam milling scanning electron microscopy. We found that Kenyon cells (KCs), whose sparse activity encodes sensory information, each make multiple\ en passant\ synapses to MB output neurons (MBONs) in each compartment. Some MBONs have inputs from all KCs, while others differentially sample sensory modalities. Only six percent of KC\>MBON synapses receive a direct synapse from a dopaminergic neuron (DAN). We identified two unanticipated classes of synapses, KC\>DAN and DAN\>MBON. DAN activation produces a slow depolarization of the MBON in these DAN\>MBON synapses and can weaken memory recall.

}, doi = {10.7554/eLife.26975}, author = {Takemura, Shin-ya and Aso, Yoshinori and Hige, Toshihide and Wong, Allan M and Lu, Zhiyuan and Xu, C Shan and Rivlin, Patricia K and Hess, Harald F and Zhao, Ting and Parag, Toufiq and Berg, Stuart and Huang, Gary and Katz, William T and Olbris, Donald J and Plaza, Stephen M and Umayam, Lowell A and Aniceto, Roxanne and Chang, Lei-Ann and Lauchie, Shirley and et al} } @article {48729, title = {Cortical actin recovery at the immunological synapse leads to termination of lytic granule secretion in cytotoxic T lymphocytes.}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {114}, year = {2017}, month = {2017 Aug 08}, pages = {E6585-94}, abstract = {

CD8(+) cytotoxic T lymphocytes (CTLs) eliminate virally infected cells through directed secretion of specialized lytic granules. Because a single CTL can kill multiple targets, degranulation must be tightly regulated. However, how CTLs regulate the termination of granule secretion remains unclear. Previous work demonstrated that centralized actin reduction at the immune synapse precedes degranulation. Using a combination of live confocal, total internal reflection fluorescence, and superresolution microscopy, we now show that, after granule fusion, actin recovers at the synapse and no further secretion is observed. Depolymerization of actin led to resumed granule secretion, suggesting that recovered actin acts as a barrier preventing sustained degranulation. Furthermore, RAB27a-deficient CTLs, which do not secrete cytotoxic granules, failed to recover actin at the synapse, suggesting that RAB27a-mediated granule secretion is required for actin recovery. Finally, we show that both actin clearance and recovery correlated with synaptic phosphatidylinositol 4,5-bisphosphate (PIP2) and that alterations in PIP2 at the immunological synapse regulate cortical actin in CTLs, providing a potential mechanism through which CTLs control cortical actin density. Our work provides insight into actin-related mechanisms regulating CTL secretion that may facilitate serial killing during immune responses.

}, issn = {1091-6490}, doi = {10.1073/pnas.1710751114}, author = {Ritter, Alex T and Kapnick, Senta M and Murugesan, Sricharan and Schwartzberg, Pamela L and Griffiths, Gillian M and Lippincott-Schwartz, Jennifer} } @article {48413, title = {Deconstructing behavioral neuropharmacology with cellular specificity.}, journal = {Science (New York, N.Y.)}, volume = {356}, year = {2017}, month = {2017 Apr 07}, abstract = {

Behavior has molecular, cellular, and circuit determinants. However, because many proteins are broadly expressed, their acute manipulation within defined cells has been difficult. Here, we combined the speed and molecular specificity of pharmacology with the cell type specificity of genetic tools. DART (drugs acutely restricted by tethering) is a technique that rapidly localizes drugs to the surface of defined cells, without prior modification of the native target. We first developed an AMPAR antagonist DART, with validation in cultured neuronal assays, in slices of mouse dorsal striatum, and in behaving mice. In parkinsonian animals, motor deficits were causally attributed to AMPARs in indirect spiny projection neurons (iSPNs) and to excess phasic firing of tonically active interneurons (TANs). Together, iSPNs and TANs (i.e., D2 cells) drove akinesia, whereas movement execution deficits reflected the ratio of AMPARs in D2 versus D1 cells. Finally, we designed a muscarinic antagonist DART in one iteration, demonstrating applicability of the method to diverse targets.

}, issn = {1095-9203}, doi = {10.1126/science.aaj2161}, author = {Shields, Brenda C and Kahuno, Elizabeth and Kim, Charles and Apostolides, Pierre F and Brown, Jennifer and Lindo, Sarah and Mensh, Brett D and Dudman, Joshua T and Lavis, Luke D and Tadross, Michael R} } @article {48463, title = {Detachable glass microelectrodes for recording action potentials in active moving organs.}, journal = {American Journal of Physiology. Heart and Circulatory Physiology}, volume = {312}, year = {2017}, month = {2017 May 05}, pages = {H1248-59}, abstract = {

We describe new detachable floating glass micropipette electrode devices that provide targeted action potential recordings in active moving organs without requiring constant mechanical constraint or pharmacological inhibition of tissue motion. The technology is based on the concept of a glass micropipette electrode that is held firmly during cell targeting and intracellular insertion, after which a 100{\textmu}g glass microelectrode, a "microdevice", is gently released to remain within the moving organ. The microdevices provide long-term recordings of action potentials, even during millimeter-scale movement of tissue in which the device is embedded. We demonstrate two different glass micropipette electrode holding and detachment designs appropriate for the heart (sharp glass microdevices for cardiac myocytes in rats, guinea pigs and humans) and the brain (patch glass microdevices for neurons in rats). We explain how microdevices enable measurements of multiple cells within a moving organ that are typically difficult with other technologies. Using sharp microdevices, action potential duration (APD) was monitored continuously for 15 minutes in unconstrained perfused hearts during global ischemia-reperfusion, providing beat-to-beat measurements of changes in APD. Action potentials from neurons in the hippocampus of anaesthetized rats were measured with patch microdevices, which provided stable base potentials during long-term recordings. Our results demonstrate that detachable microdevices are an elegant and robust tool to record electrical activity with high temporal resolution and cellular level localization without disturbing the physiological working conditions of the organ.

}, issn = {1522-1539}, doi = {10.1152/ajpheart.00741.2016}, author = {Barbic, Mladen and Moreno, Angel and Harris, Timothy D and Kay, Matthew W} } @article {48786, title = {Dissection of the Drosophila neuropeptide F circuit using a high-throughput two-choice assay.}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {114}, year = {2017}, month = {2017 Sep 05}, pages = {e8091-9}, abstract = {

In their classic experiments, Olds and Milner showed that rats learn to lever press to receive an electric stimulus in specific brain regions. This led to the identification of mammalian reward centers. Our interest in defining the neuronal substrates of reward perception in the fruit fly Drosophila melanogaster prompted us to develop a simpler experimental approach wherein flies could implement behavior that induces self-stimulation of specific neurons in their brains. The high-throughput assay employs optogenetic activation of neurons when the fly occupies a specific area of a behavioral chamber, and the flies{\textquoteright} preferential occupation of this area reflects their choosing to experience optogenetic stimulation. Flies in which neuropeptide F (NPF) neurons are activated display preference for the illuminated side of the chamber. We show that optogenetic activation of NPF neuron is rewarding in olfactory conditioning experiments and that the preference for NPF neuron activation is dependent on NPF signaling. Finally, we identify a small subset of NPF-expressing neurons located in the dorsomedial posterior brain that are sufficient to elicit preference in our assay. This assay provides the means for carrying out unbiased screens to map reward neurons in flies.

}, issn = {1091-6490}, doi = {10.1073/pnas.1710552114}, author = {Shao, Lisha and Saver, Mathias and Chung, Phuong and Ren, Qingzhong and Lee, Tzumin and Kent, Clement F and Heberlein, Ulrike} } @article {48983, title = {Divergent connectivity of homologous command-like neurons mediates segment-specific touch responses in Drosophila.}, journal = {Neuron}, volume = {96}, year = {2017}, month = {2017 Dec 20}, pages = {1373-87}, abstract = {

Animals adaptively respond to a tactile stimulus by choosing an ethologically relevant behavior depending on the location of the stimuli. Here, we investigate how somatosensory inputs on different body segments are linked to distinct motor outputs in Drosophila larvae. Larvae escape by backward locomotion when touched on the head, while they crawl forward when touched on the tail. We identify a class of segmentally repeated second-order somatosensory interneurons, that we named Wave, whose activation in anterior and posterior segments elicit backward and forward locomotion, respectively. Anterior and posterior Wave neurons extend their dendrites in opposite directions to receive somatosensory inputs from the head and tail, respectively. Downstream of anterior Wave neurons, we identify premotor circuits including the neuron A03a5, which together with Wave, is necessary for the backward locomotion touch response. Thus, Wave neurons match their receptive field to appropriate motor programs by participating in different circuits in different segments.

}, issn = {1097-4199}, doi = {10.1016/j.neuron.2017.10.030}, author = {Takagi, Suguru and Cocanougher, Benjamin Thomas and Niki, Sawako and Miyamoto, Dohjin and Kohsaka, Hiroshi and Kazama, Hokto and Fetter, Richard Doty and Truman, James William and Zlatic, Marta and Cardona, Albert and Nose, Akinao} } @article {48416, title = {Diverse protocols for correlative super-resolution fluorescence imaging and electron microscopy of chemically fixed samples.}, journal = {Nature Protocols}, volume = {12}, year = {2017}, month = {2017 May}, pages = {916-946}, abstract = {

Our groups have recently developed related approaches for sample preparation for super-resolution imaging within endogenous cellular environments using correlative light and electron microscopy (CLEM). Four distinct techniques for preparing and acquiring super-resolution CLEM data sets for aldehyde-fixed specimens are provided, including Tokuyasu cryosectioning, whole-cell mount, cell unroofing and platinum replication, and resin embedding and sectioning. The choice of the best protocol for a given application depends on a number of criteria that are discussed in detail. Tokuyasu cryosectioning is relatively rapid but is limited to small, delicate specimens. Whole-cell mount has the simplest sample preparation but is restricted to surface structures. Cell unroofing and platinum replication creates high-contrast, 3D images of the cytoplasmic surface of the plasma membrane but is more challenging than whole-cell mount. Resin embedding permits serial sectioning of large samples but is limited to osmium-resistant probes, and is technically difficult. Expected results from these protocols include super-resolution localization (\~{}10-50 nm) of fluorescent targets within the context of electron microscopy ultrastructure, which can help address cell biological questions. These protocols can be completed in 2-7 d, are compatible with a number of super-resolution imaging protocols, and are broadly applicable across biology.

}, issn = {1750-2799}, doi = {10.1038/nprot.2017.017}, author = {Kopek, Benjamin G and Paez-Segala, Maria G and Shtengel, Gleb and Sochacki, Kem A and Sun, Mei G and Wang, Yalin and Xu, C Shan and Van Engelenburg, Schuyler B and Taraska, Justin W and Looger, Loren L and Hess, Harald F} } @article {48561, title = {Drosophila courtship conditioning as a measure of learning and memory.}, journal = {Journal of Visualized Experiments - Neuroscience }, year = {2017}, month = {2017-06-05}, pages = {e55808}, abstract = {

Many insights into the molecular mechanisms underlying learning and memory have been elucidated through the use of simple behavioral assays in model organisms such as the fruit fly,\ Drosophila melanogaster.\ Drosophila\ is useful for understanding the basic neurobiology underlying cognitive deficits resulting from mutations in genes associated with human cognitive disorders, such as intellectual disability (ID) and autism. This work describes a methodology for testing learning and memory using a classic paradigm in\ Drosophilaknown as courtship conditioning. Male flies\ court females using a distinct pattern of easily recognizable behaviors. Premated females are not receptive to mating and will reject the male{\textquoteright}s copulation attempts. In response to this rejection, male flies reduce their courtship behavior. This learned reduction in courtship behavior is measured over time, serving as an indicator of learning and memory. The basic numerical output of this assay is the courtship index (CI), which is defined as the percentage of time that a male spends courting during a 10 min interval. The learning index (LI) is the relative reduction of CI in flies that have been exposed to a premated female compared to na{\"\i}ve flies with no previous social encounters. For the statistical comparison of LIs between genotypes, a randomization test with bootstrapping is used. To illustrate how the assay can be used to address the role of a gene relating to learning and memory, the pan-neuronal knockdown of\ Dihydroxyacetone phosphate acyltransferase\ (Dhap-at) was characterized here. The human ortholog\ of\ Dhap-at,\ glyceronephosphate O-acyltransferase\ (GNPT), is involved in rhizomelic chondrodysplasia punctata type 2, an autosomal-recessive syndrome characterized by severe ID. Using the courtship conditioning assay, it was determined that Dhap-at is required for long-term memory, but not for short-term memory. This result serves as a basis for further investigation of the underlying molecular mechanisms.

}, doi = {10.3791/55808}, author = {Koemans, Tom S and Oppitz, Cornelia and Donders, Rogier AT and van Bokhoven, Hans and Schenck, Annette and Keleman, Krystyna and Kramer, Jamie M} } @article {65018, title = {Ensemble cryo-EM elucidates the mechanism of translation fidelity.}, journal = {Nature}, volume = {546}, year = {2017}, month = {2017 Jun 01}, pages = {113-117}, abstract = {

Gene translation depends on accurate decoding of mRNA, the structural mechanism of which remains poorly understood. Ribosomes decode mRNA codons by selecting cognate aminoacyl-tRNAs delivered by elongation factor Tu (EF-Tu). Here we present high-resolution structural ensembles of ribosomes with cognate or near-cognate aminoacyl-tRNAs delivered by EF-Tu. Both cognate and near-cognate tRNA anticodons explore the aminoacyl-tRNA-binding site (A site) of an open 30S subunit, while inactive EF-Tu is separated from the 50S subunit. A transient conformation of decoding-centre nucleotide G530 stabilizes the cognate codon-anticodon helix, initiating step-wise {\textquoteright}latching{\textquoteright} of the decoding centre. The resulting closure of the 30S subunit docks EF-Tu at the sarcin-ricin loop of the 50S subunit, activating EF-Tu for GTP hydrolysis and enabling accommodation of the aminoacyl-tRNA. By contrast, near-cognate complexes fail to induce the G530 latch, thus favouring open 30S pre-accommodation intermediates with inactive EF-Tu. This work reveals long-sought structural differences between the pre-accommodation of cognate and near-cognate tRNAs that elucidate the mechanism of accurate decoding.

}, keywords = {Anticodon, Codon, Cryoelectron Microscopy, Escherichia coli, GTP Phosphohydrolases, Guanosine Triphosphate, Hydrolysis, Models, Molecular, Peptide Elongation Factor Tu, Protein Biosynthesis, Protein Domains, Ribosome Subunits, Ribosomes, RNA, Ribosomal, 16S, RNA, Transfer, Amino Acyl}, issn = {1476-4687}, doi = {10.1038/nature22397}, author = {Loveland, Anna B and Demo, Gabriel and Grigorieff, Nikolaus and Korostelev, Andrei A} } @article {48935, title = {Fully integrated silicon probes for high-density recording of neural activity.}, journal = {Nature}, volume = {551}, year = {2017}, month = {2017 Nov 08}, pages = {232-236}, abstract = {

Sensory, motor and cognitive operations involve the coordinated action of large neuronal populations across multiple brain regions in both superficial and deep structures. Existing extracellular probes record neural activity with excellent spatial and temporal (sub-millisecond) resolution, but from only a few dozen neurons per shank. Optical Ca(2+) imaging offers more coverage but lacks the temporal resolution needed to distinguish individual spikes reliably and does not measure local field potentials. Until now, no technology compatible with use in unrestrained animals has combined high spatiotemporal resolution with large volume coverage. Here we design, fabricate and test a new silicon probe known as Neuropixels to meet this need. Each probe has 384 recording channels that can programmably address 960 complementary metal-oxide-semiconductor (CMOS) processing-compatible low-impedance TiN sites that tile a single 10-mm long, 70 {\texttimes} 20-μm cross-section shank. The 6 {\texttimes} 9-mm probe base is fabricated with the shank on a single chip. Voltage signals are filtered, amplified, multiplexed and digitized on the base, allowing the direct transmission of noise-free digital data from the probe. The combination of dense recording sites and high channel count yielded well-isolated spiking activity from hundreds of neurons per probe implanted in mice and rats. Using two probes, more than 700 well-isolated single neurons were recorded simultaneously from five brain structures in an awake mouse. The fully integrated functionality and small size of Neuropixels probes allowed large populations of neurons from several brain structures to be recorded in freely moving animals. This combination of high-performance electrode technology and scalable chip fabrication methods opens a path towards recording of brain-wide neural activity during behaviour.

}, issn = {1476-4687}, doi = {10.1038/nature24636}, author = {Jun, James J and Steinmetz, Nicholas A and Siegle, Joshua H and Denman, Daniel J and Bauza, Marius and Barbarits, Brian and Lee, Albert K and Anastassiou, Costas A and Andrei, Alexandru and Ayd{\i}n, {\c C}a{\u g}atay and Barbic, Mladen and Blanche, Timothy J and Bonin, Vincent and Couto, Jo{\~a}o and Dutta, Barundeb and Gratiy, Sergey L and Gutnisky, Diego A and H{\"a}usser, Michael and Karsh, Bill and Ledochowitsch, Peter and Lopez, Carolina Mora and Mitelut, Catalin and Musa, Silke and Okun, Michael and Pachitariu, Marius and Putzeys, Jan and Rich, P Dylan and Rossant, Cyrille and Sun, Wei-Lung and Svoboda, Karel and Carandini, Matteo and Harris, Kenneth D and Koch, Christof and O{\textquoteright}Keefe, John and Harris, Timothy D} } @article {48975, title = {The functional organization of descending sensory-motor pathways in Drosophila.}, journal = {eLife}, year = {2017}, month = {2018 Jun 26}, pages = {e34272}, abstract = {

In most animals, the brain controls the body via a set of descending neurons (DNs) that traverse the neck. DN activity activates, maintains or modulates locomotion and other behaviors. Individual DNs have been well-studied in species from insects to primates, but little is known about overall connectivity patterns across the DN population. We systematically investigated DN anatomy in\ Drosophila melanogaster\ and created over 100 transgenic lines targeting individual cell types. We identified roughly half of all\ Drosophila\ DNs and comprehensively map connectivity between sensory and motor neuropils in the brain and nerve cord, respectively. We find the nerve cord is a layered system of neuropils reflecting the fly{\textquoteright}s capability for two largely independent means of locomotion -- walking and flight -- using distinct sets of appendages. Our results reveal the basic functional map of descending pathways in flies and provide tools for systematic interrogation of neural circuits.

}, doi = {10.7554/eLife.34272}, author = {Namiki, Shigehiro and Dickinson, Michael H and Wong, Allan M and Korff, Wyatt and Card, Gwyneth M} } @article {48891, title = {A general method to fine-tune fluorophores for live-cell and in vivo imaging.}, journal = {Nature Methods}, volume = {14}, year = {2017}, month = {2017 Oct}, pages = {987-994}, abstract = {

Pushing the frontier of fluorescence microscopy requires the design of enhanced fluorophores with finely tuned properties. We recently discovered that incorporation of four-membered azetidine rings into classic fluorophore structures elicits substantial increases in brightness and photostability, resulting in the Janelia Fluor (JF) series of dyes. We refined and extended this strategy, finding that incorporation of 3-substituted azetidine groups allows rational tuning of the spectral and chemical properties of rhodamine dyes with unprecedented precision. This strategy allowed us to establish principles for fine-tuning the properties of fluorophores and to develop a palette of new fluorescent and fluorogenic labels with excitation ranging from blue to the far-red. Our results demonstrate the versatility of these new dyes in cells, tissues and animals.

}, keywords = {Animals, Brain, Cell Line, Coloring Agents, Drosophila, Image Processing, Computer-Assisted, Larva, Mice, Microscopy, Fluorescence, Photochemical Processes, Staining and Labeling}, issn = {1548-7105}, doi = {10.1038/nmeth.4403}, author = {Grimm, Jonathan B and Muthusamy, Anand K and Liang, Yajie and Brown, Timothy A and Lemon, William C and Patel, Ronak and Lu, Rongwen and Macklin, John J and Keller, Philipp J and Ji, Na and Lavis, Luke D} } @article {48319, title = {Genetic and transgenic reagents for Drosophila simulans, D. mauritiana, D. yakuba, D. santomea and D. virilis.}, journal = {G3 (Bethesda, Md.)}, volume = {7}, year = {2017}, month = {2017 Mar 09}, pages = {1339-47}, abstract = {

Species of the Drosophila melanogaster species subgroup, including the species D. simulans, D. mauritiana, D. yakuba, and D. santomea, have long served as model systems for studying evolution. Studies in these species have been limited, however, by a paucity of genetic and transgenic reagents. Here we describe a collection of transgenic and genetic strains generated to facilitate genetic studies within and between these species. We have generated many strains of each species containing mapped piggyBac transposons including an enhanced yellow fluorescent protein gene expressed in the eyes and a phiC31 attP site-specific integration site. We have tested a subset of these lines for integration efficiency and reporter gene expression levels. We have also generated a smaller collection of other lines expressing other genetically encoded fluorescent molecules in the eyes and a number of other transgenic reagents that will be useful for functional studies in these species. In addition, we have mapped the insertion locations of 58 transposable elements in D. virilis that will be useful for genetic mapping studies.

}, issn = {2160-1836}, doi = {10.1534/g3.116.038885}, author = {Stern, David L and Crocker, Justin and Ding, Yun and Frankel, Nicol{\'a}s and Kappes, Gretchen and Kim, Elizabeth and Kuzmickas, Ryan and Lemire, Andrew and Mast, Joshua D and Picard, Serge} } @article {48845, title = {Genetic reagents for making split-GAL4 lines in Drosophila.}, journal = {Genetics }, volume = {209}, year = {2017}, month = {2018 March}, pages = {31-5}, abstract = {

The ability to reproducibly target expression of transgenes to small, defined subsets of cells is a key experimental tool for understanding many biological processes. The Drosophila nervous system contains thousands of distinct cell types and it has generally not been possible to limit expression to one or a few cell types when using a single segment of genomic DNA as an enhancer to drive expression. Intersectional methods, in which expression of the transgene only occurs where two different enhancers overlap in their expression patterns, can be used to achieve the desired specificity. This report describes a set of over 2,800 transgenic lines for use with the split-GAL4 intersectional method.

}, doi = {10.1101/197509}, author = {Dionne, Heather and Hibbard, Karen L and Cavallaro, Amanda and Kao, Jui-Chun and Rubin, Gerald M} } @article {48252, title = {The glia of the adult Drosophila nervous system.}, journal = {Glia}, volume = {65}, year = {2017}, month = {2017 Jan 30}, pages = {606-38}, abstract = {

Glia play crucial roles in the development and homeostasis of the nervous system. While the GLIA in the Drosophila embryo have been well characterized, their study in the adult nervous system has been limited. Here, we present a detailed description of the glia in the adult nervous system, based on the analysis of some 500 glial drivers we identified within a collection of synthetic GAL4 lines. We find that glia make up \~{}10\% of the cells in the nervous system and envelop all compartments of neurons (soma, dendrites, axons) as well as the nervous system as a whole. Our morphological analysis suggests a set of simple rules governing the morphogenesis of glia and their interactions with other cells. All glial subtypes minimize contact with their glial neighbors but maximize their contact with neurons and adapt their macromorphology and micromorphology to the neuronal entities they envelop. Finally, glial cells show no obvious spatial organization or registration with neuronal entities. Our detailed description of all glial subtypes and their regional specializations, together with the powerful genetic toolkit we provide, will facilitate the functional analysis of glia in the mature nervous system. GLIA 2017.

}, issn = {1098-1136}, doi = {10.1002/glia.23115}, author = {Kremer, Malte C and Jung, Christophe and Batelli, Sara and Rubin, Gerald M and Gaul, Ulrike} } @article {65886, title = {Homeostatic plasticity shapes the visual system{\textquoteright}s first synapse}, journal = {Nature Communications}, volume = {8}, year = {2017}, month = {10/2017}, abstract = {

Vision in dim light depends on synapses between rods and rod bipolar cells (RBCs). Here, we find that these synapses exist in multiple configurations, in which single release sites of rods are apposed by one to three postsynaptic densities (PSDs). Single RBCs often form multiple PSDs with one rod; and neighboring RBCs share ~13\% of their inputs. Rod-RBC synapses develop while ~7\% of RBCs undergo programmed cell death (PCD). Although PCD is common throughout the nervous system, its influences on circuit development and function are not well understood. We generate mice in which ~53 and ~93\% of RBCs, respectively, are removed during development. In these mice, dendrites of the remaining RBCs expand in graded fashion independent of light-evoked input. As RBC dendrites expand, they form fewer multi-PSD contacts with rods. Electrophysiological recordings indicate that this homeostatic co-regulation of neurite and synapse development preserves retinal function in dim light.

}, doi = {10.1038/s41467-017-01332-7}, url = {http://www.nature.com/articles/s41467-017-01332-7http://www.nature.com/articles/s41467-017-01332-7.pdfhttp://www.nature.com/articles/s41467-017-01332-7.pdfhttp://www.nature.com/articles/s41467-017-01332-7}, author = {Johnson, Robert E. and Tien, Nai-Wen and Shen, Ning and Pearson, James T. and Soto, Florentina and Kerschensteiner, Daniel} } @article {48669, title = {How to make a worm twitch.}, journal = {Biophysical Journal}, volume = {112}, year = {2017}, month = {2017 May 09}, pages = {1737-1738}, issn = {1542-0086}, doi = {10.1016/j.bpj.2017.03.035}, author = {Keller, Philipp J} } @article {48643, title = {Ion Channels: History, Diversity, and Impact.}, journal = {Cold Spring Harbor Protocols}, volume = {2017}, year = {2017}, month = {2017 Jul 05}, pages = {pdb.top092288}, abstract = {

From patch-clamp techniques to recombinant DNA technologies, three-dimensional protein modeling, and optogenetics, diverse and sophisticated methods have been used to study ion channels and how they determine the electrical properties of cells.

}, issn = {1559-6095}, doi = {10.1101/pdb.top092288}, author = {Brenowitz, Stephan and Duguid, Ian and Kammermeier, Paul J} } @article {48646, title = {Mapping the neural substrates of behavior.}, journal = {Cell}, volume = {170}, year = {2017}, month = {2017-07-13}, pages = {393-406}, abstract = {

Assigning behavioral functions to neural structures has long been a central goal in neuroscience and is a necessary first step toward a circuit-level understanding of how the brain generates behavior. Here, we map the neural substrates of locomotion and social behaviors for Drosophila melanogaster using automated machine-vision and machine-learning techniques. From videos of 400,000 flies, we quantified the behavioral effects of activating 2,204 genetically targeted populations of neurons. We combined a novel quantification of anatomy with our behavioral analysis to create brain-behavior correlation maps, which are shared as browsable web pages and interactive software. Based on these maps, we generated hypotheses of regions of the brain causally related to sensory processing, locomotor control, courtship, aggression, and sleep. Our maps directly specify genetic tools to target these regions, which we used to identify a small population of neurons with a role in the control of walking.

{\textbullet}We developed machine-vision methods to broadly and precisely quantify fly behavior{\textbullet}We measured effects of activating 2,204 genetically targeted neuronal populations{\textbullet}We created whole-brain maps of neural substrates of locomotor and social behaviors{\textbullet}We created resources for exploring our results and enabling further investigation

Machine-vision analyses of large behavior and neuroanatomy data reveal whole-brain maps of regions associated with numerous complex behaviors.

}, doi = {10.1016/j.cell.2017.06.032}, url = {http://dx.doi.org/10.1016/j.cell.2017.06.032}, author = {Robie, Alice A and Hirokawa, Jonathan and Edwards, Austin W and Umayam, Lowell A and Lee, Allen and Phillips, Mary L and Card, Gwyneth M and Korff, Wyatt and Rubin, Gerald M and Simpson, Julie H and Reiser, Michael B and Branson, Kristin M} } @article {48344, title = {Mechanism of ribosome rescue by ArfA and RF2.}, journal = {eLife}, volume = {6}, year = {2017}, month = {2017 Mar 16}, pages = {e23687}, abstract = {

ArfA rescues ribosomes stalled on truncated mRNAs by recruiting release factor RF2, which normally binds stop codons to catalyze peptide release. We report two 3.2-{\r A} resolution cryo-EM structures - determined from a single sample - of the 70S ribosome with ArfA{\textbullet}RF2 in the A site. In both states, the ArfA C-terminus occupies the mRNA tunnel downstream of the A site. One state contains a compact inactive RF2 conformation. Ordering of the ArfA N-terminus in the second state rearranges RF2 into an extended conformation that docks the catalytic GGQ motif into the peptidyl-transferase center. Our work thus reveals the structural dynamics of ribosome rescue. The structures demonstrate how ArfA "senses" the vacant mRNA tunnel and activates RF2 to mediate peptide release without a stop codon, allowing stalled ribosomes to be recycled.

}, issn = {2050-084X}, doi = {10.7554/eLife.23687}, author = {Demo, Gabriel and Svidritskiy, Egor and Madireddy, Rohini and Diaz-Avalos, Ruben and Grant, Timothy and Grigorieff, Nikolaus and Sousa, Duncan and Korostelev, Andrei A} } @article {48557, title = {Memory retrieval from first principles.}, journal = {Neuron}, volume = {94}, year = {2017}, month = {2017 Jun 07}, pages = {1027-1032}, abstract = {

The dilemma that neurotheorists face is that (1) detailed biophysical models that can be constrained by direct measurements, while being of great importance, offer no immediate insights into cognitive processes in the brain, and (2) high-level abstract cognitive models, on the other hand, while relevant for understanding behavior, are largely detached from neuronal processes and typically have many free, experimentally unconstrained parameters that have to be tuned to a particular data set and, hence, cannot be readily generalized to other experimental paradigms. In this contribution, we propose a set of "first principles" for neurally inspired cognitive modeling of memory retrieval that has no biologically unconstrained parameters and can be analyzed mathematically both at neuronal and cognitive levels. We apply this framework to the classical cognitive paradigm of free recall. We show that the resulting model accounts well for puzzling behavioral data on human participants and makes predictions that could potentially be tested with neurophysiological recording techniques.

}, issn = {1097-4199}, doi = {10.1016/j.neuron.2017.03.048}, author = {Katkov, M and Romani, S and Tsodyks, M} } @article {48558, title = {Mesoscale-duration activated states gate spiking in response to fast rises in membrane voltage in the awake brain.}, journal = {Journal of Neurophysiology}, volume = {118}, year = {2017}, month = {2017 May 31}, pages = {1270-91}, abstract = {

Seconds-scale network states, affecting many neurons within a network, modulate neural activity by complementing fast integration of neuron-specific inputs that arrive in the milliseconds before spiking. Non-rhythmic subthreshold dynamics at intermediate timescales, however, are less well-characterized. We found, using automated whole cell patch clamping in vivo, that spikes recorded in CA1 and barrel cortex in awake mice are often preceded not only by monotonic voltage rises lasting milliseconds, but also by more gradual (lasting 10s-100s of ms) depolarizations. The latter exert a gating function on spiking, in a fashion that depends on the gradual rise duration: the probability of spiking was higher for longer gradual rises, even controlling for the amplitude of the gradual rises. Barrel cortex double-autopatch recordings show that gradual rises are shared across some but not all neurons. The gradual rises may represent a new kind of state, intermediate both in timescale and in proportion of neurons participating, which gates a neuron{\textquoteright}s ability to respond to subsequent inputs.

}, issn = {1522-1598}, doi = {10.1152/jn.00116.2017}, author = {Singer, Annabelle C and Talei Franzesi, Giovanni and Kodandaramaiah, Suhasa B and Flores, Francisco J and Cohen, Jeremy D and Lee, Albert K and B{\"o}rgers, Christoph and Forest, Craig R and Kopell, Nancy J and Boyden, Edward S} } @article {48223, title = {moxDendra2: an inert photoswitchable protein for oxidizing environments.}, journal = {Chemical Communications}, volume = {53}, year = {2017}, month = {2017 Jan 19}, pages = {2106-9}, abstract = {

Fluorescent proteins (FPs) that can be optically highlighted enable PALM super-resolution microscopy and pulse-chase experiments of cellular molecules. Most FPs evolved in cytoplasmic environments either in the original source organism or in the cytoplasm of bacteria during the course of optimization for research applications. Consequently, many FPs may fold incorrectly in the chemically distinct environments in subcellular organelles. Here, we describe the first monomeric photoswitchable (from green to bright red) FP adapted for oxidizing environments.

}, doi = {10.1039/C6CC09997A}, author = {Kaberniuk, Andrii A and Morano, Nicholas C and Verkhusha, Vladislav V and Snapp, Erik Lee} } @article {48254, title = {Multicut brings automated neurite segmentation closer to human performance.}, journal = {Nature Methods}, volume = {14}, year = {2017}, month = {2017 Jan 31}, pages = {101-102}, issn = {1548-7105}, doi = {10.1038/nmeth.4151}, author = {Beier, Thorsten and Pape, Constantin and Rahaman, Nasim and Prange, Timo and Berg, Stuart and Bock, Davi D and Cardona, Albert and Knott, Graham W and Plaza, Stephen M and Scheffer, Louis K and Koethe, Ullrich and Kreshuk, Anna and Hamprecht, Fred A} } @article {48278, title = {Multi-view light-sheet imaging and tracking with the MaMuT software reveals the cell lineage of a direct developing arthropod limb.}, journal = {eLife}, volume = {7}, year = {2017}, month = {2018 Mar 29}, pages = {e34410}, abstract = {

During development, coordinated cell behaviors orchestrate tissue and organ morphogenesis. Detailed descriptions of cell lineages and behaviors provide a powerful framework to elucidate the mechanisms of morphogenesis. To study the cellular basis of limb development, we imaged transgenic fluorescently-labeled embryos from the crustacean Parhyale hawaiensis with multi-view light-sheet microscopy at high spatiotemporal resolution over several days of embryogenesis. The cell lineage of outgrowing thoracic limbs was reconstructed at single-cell resolution with new software called Massive Multi-view Tracker (MaMuT). In silico clonal analyses suggested that the early limb primordium becomes subdivided into anterior-posterior and dorsal-ventral compartments whose boundaries intersect at the distal tip of the growing limb. Limb-bud formation is associated with spatial modulation of cell proliferation, while limb elongation is also driven by preferential orientation of cell divisions along the proximal-distal growth axis. Cellular reconstructions were predictive of the expression patterns of limb development genes including the BMP morphogen Decapentaplegic.

}, doi = {10.7554/eLife.34410}, author = {Wolff, Carsten and Tinevez, Jean-Yves and Pietzsch, Tobias and Stamataki, Evangelia and Harich, Benjamin and Guignard, Leo and Preibisch, Stephan and Shorte, Spencer and Keller, Philipp J and Tomancak, Pavel and Pavlopoulos, Anastasios} } @article {48731, title = {Myc Regulates Chromatin Decompaction and Nuclear Architecture during B Cell Activation.}, journal = {Molecular Cell}, volume = {67}, year = {2017}, month = {2017 Jul 28}, pages = {566-78}, abstract = {

50 years ago, Vincent Allfrey and colleagues discovered that lymphocyte activation triggers massive acetylation of chromatin. However, the molecular mechanisms driving epigenetic accessibility are still unknown. We here show that stimulated lymphocytes decondense chromatin by three differentially regulated steps. First, chromatin is repositioned away from the nuclear periphery in response to global acetylation. Second, histone nanodomain clusters decompact into mononucleosome fibers through a mechanism that requires Myc and continual energy input. Single-molecule imaging shows that this step lowers transcription factor residence time and non-specific collisions during sampling for DNA targets. Third, chromatin interactions shift from long range to predominantly short range, and CTCF-mediated loops and contact domains double in numbers. This architectural change facilitates cognate promoter-enhancer contacts and also requires Myc and continual ATP production. Our results thus define the nature and transcriptional impact of chromatin decondensation and reveal an unexpected role for Myc in the establishment of nuclear topology in mammalian cells.

}, issn = {1097-4164}, doi = {10.1016/j.molcel.2017.07.013}, author = {Kieffer-Kwon, Kyong-Rim and Nimura, Keisuke and Rao, Suhas S P and Xu, Jianliang and Jung, Seolkyoung and Pekowska, Aleksandra and Dose, Marei and Stevens, Evan and Mathe, Ewy and Dong, Peng and Huang, Su-Chen and Ricci, Maria Aurelia and Baranello, Laura and Zheng, Ying and Ardori, Francesco Tomassoni and Resch, Wolfgang and Stavreva, Diana and Nelson, Steevenson and McAndrew, Michael and Casellas, Adriel and Finn, Elizabeth and Gregory, Charles and St Hilaire, Brian Glenn and Johnson, Steven M and Dubois, Wendy and Cosma, Maria Pia and Batchelor, Eric and Levens, David and Phair, Robert D and Misteli, Tom and Tessarollo, Lino and Hager, Gordon and Lakadamyali, Melike and Liu, Zhe and Floer, Monique and Shroff, Hari and Aiden, Erez Lieberman and Casellas, Rafael} } @article {48417, title = {Near-infrared fluorescent protein iRFP713 as a reporter protein for optogenetic vectors, a transgenic Cre-reporter rat, and other neuronal studies.}, journal = {Journal of Neuroscience Methods}, volume = {284}, year = {2017}, month = {2017 Apr 02}, pages = {1-14}, abstract = {

BACKGROUND: The use of genetically-encoded fluorescent reporters is essential for the identification and observation of cells that express transgenic modulatory proteins. Near-infrared (NIR) fluorescent proteins have superior light penetration through biological tissue, but are not yet widely adopted.

NEW METHOD: Using the near-infrared fluorescent protein, iRFP713, improves the imaging resolution in thick tissue sections or the intact brain due to the reduced light-scattering at the longer, NIR wavelengths used to image the protein. Additionally, iRFP713 can be used to identify transgenic cells without photobleaching other fluorescent reporters or affecting opsin function. We have generated a set of adeno-associated vectors in which iRFP713 has been fused to optogenetic channels, and can be expressed constitutively or Cre-dependently.

RESULTS: iRFP713 is detectable when expressed in neurons both in vitro and in vivo without exogenously supplied chromophore biliverdin. Neuronally-expressed iRFP713 has similar properties to GFP-like fluorescent proteins, including the ability to be translationally fused to channelrhodopsin or halorhodopsin, however, it shows superior photostability compared to EYFP. Furthermore, electrophysiological recordings from iRFP713-labeled cells compared to cells labeled with mCherry suggest that iRFP713 cells are healthier and therefore more stable and reliable in an ex vivo preparation. Lastly, we have generated a transgenic rat that expresses iRFP713 in a Cre-dependent manner.

CONCLUSIONS: Overall, we have demonstrated that iRFP713 can be used as a reporter in neurons without the use of exogenous biliverdin, with minimal impact on viability and function thereby making it feasible to extend the capabilities for imaging genetically-tagged neurons in slices and in vivo.

}, issn = {1872-678X}, doi = {10.1016/j.jneumeth.2017.03.020}, author = {Richie, Christopher T and Whitaker, Leslie R and Whitaker, Keith W and Necarsulmer, Julie and Baldwin, Heather A and Zhang, Yajun and Fortuno, Lowella and Hinkle, Josh J and Koivula, Pyry and Henderson, Mark J and Sun, Wenzhi and Wang, Kai and Smith, Jeffrey C and Pickel, Jim and Ji, Na and Hope, Bruce T and Harvey, Brandon K} } @article {48919, title = {Network-size independent covering number bounds for deep networks.}, journal = {arXiv}, year = {2017}, month = {2017 Nov 02}, pages = {arXiv:1711.00753 }, abstract = {

We give a covering number bound for deep learning networks that is independent of the size of the network. The key for the simple analysis is that for linear classifiers, rotating the data doesn{\textquoteright}t affect the covering number. Thus, we can ignore the rotation part of each layer{\textquoteright}s linear transformation, and get the covering number bound by concentrating on the scaling part.

}, url = {https://arxiv.org/abs/1711.00753}, author = {Kabra, Mayank and Branson, Kristin M} } @article {48549, title = {Neural signatures of dynamic stimulus selection in Drosophila.}, journal = {Nature Neuroscience}, volume = {20}, year = {2017}, month = {2017 Jun 12}, pages = {1104-13}, abstract = {

Many animals orient using visual cues, but how a single cue is selected from among many is poorly understood. Here we show that Drosophila ring neurons{\textemdash}central brain neurons implicated in navigation{\textemdash}display visual stimulus selection. Using\ in vivo\ two-color two-photon imaging with genetically encoded calcium indicators, we demonstrate that individual ring neurons inherit simple-cell-like receptive fields from their upstream partners. Stimuli in the contralateral visual field suppressed responses to ipsilateral stimuli in both populations. Suppression strength depended on when and where the contralateral stimulus was presented, an effect stronger in ring neurons than in their upstream inputs. This history-dependent effect on the temporal structure of visual responses, which was well modeled by a simple biphasic filter, may determine how visual references are selected for the fly{\textquoteright}s internal compass. Our approach highlights how two-color calcium imaging can help identify and localize the origins of sensory transformations across synaptically connected neural populations.

}, doi = {10.1038/nn.4581}, author = {Sun, Yi and Nern, Aljoscha and Franconville, Romain and Dana, Hod and Schreiter, Eric R and Looger, Loren L and Svoboda, Karel and Kim, Douglas S and Hermundstad, Ann M and Jayaraman, Vivek} } @article {48642, title = {The Ol1mpiad: concordance of behavioural faculties of stage 1 and stage 3 Drosophila larvae.}, journal = {The Journal of Experimental Biology}, volume = {220}, year = {2017}, month = {2017 Jul 01}, pages = {2452-2475}, abstract = {

Mapping brain function to brain structure is a fundamental task for neuroscience. For such an endeavour, the Drosophila larva is simple enough to be tractable, yet complex enough to be interesting. It features about 10,000 neurons and is capable of various taxes, kineses and Pavlovian conditioning. All its neurons are currently being mapped into a light-microscopical atlas, and Gal4 strains are being generated to experimentally access neurons one at a time. In addition, an electron microscopic reconstruction of its nervous system seems within reach. Notably, this electron microscope-based connectome is being drafted for a stage 1 larva - because stage 1 larvae are much smaller than stage 3 larvae. However, most behaviour analyses have been performed for stage 3 larvae because their larger size makes them easier to handle and observe. It is therefore warranted to either redo the electron microscopic reconstruction for a stage 3 larva or to survey the behavioural faculties of stage 1 larvae. We provide the latter. In a community-based approach we called the Ol1mpiad, we probed stage 1 Drosophila larvae for free locomotion, feeding, responsiveness to substrate vibration, gentle and nociceptive touch, burrowing, olfactory preference and thermotaxis, light avoidance, gustatory choice of various tastants plus odour-taste associative learning, as well as light/dark-electric shock associative learning. Quantitatively, stage 1 larvae show lower scores in most tasks, arguably because of their smaller size and lower speed. Qualitatively, however, stage 1 larvae perform strikingly similar to stage 3 larvae in almost all cases. These results bolster confidence in mapping brain structure and behaviour across developmental stages.

}, issn = {1477-9145}, doi = {10.1242/jeb.156646}, author = {Almeida-Carvalho, Maria J and Berh, Dimitri and Braun, Andreas and Chen, Yi-chun and Eichler, Katharina and Eschbach, Claire and Fritsch, Pauline M J and Gerber, Bertram and Hoyer, Nina and Jiang, Xiaoyi and Kleber, J{\"o}rg and Kl{\"a}mbt, Christian and K{\"o}nig, Christian and Louis, Matthieu and Michels, Birgit and Miroschnikow, Anton and Mirth, Christen and Miura, Daisuke and Niewalda, Thomas and Otto, Nils and Paisios, Emmanouil and Pankratz, Michael J and Petersen, Meike and Ramsperger, Noel and Randel, Nadine and Risse, Benjamin and Saumweber, Timo and Schlegel, Philipp and Schleyer, Michael and Soba, Peter and Sprecher, Simon G and Tanimura, Teiichi and Thum, Andreas S and Toshima, Naoko and Truman, Jim W and Yarali, Ayse and Zlatic, Marta} } @article {48799, title = {Optogenetic control with a photocleavable protein, PhoCl.}, journal = {Nature Methods}, volume = {14}, year = {2017}, month = {2017 Apr}, pages = {391-394}, abstract = {

To expand the range of experiments that are accessible with optogenetics, we developed a photocleavable protein (PhoCl) that spontaneously dissociates into two fragments after violet-light-induced cleavage of a specific bond in the protein backbone. We demonstrated that PhoCl can be used to engineer light-activatable Cre recombinase, Gal4 transcription factor, and a viral protease that in turn was used to activate opening of the large-pore ion channel Pannexin-1.

}, keywords = {Connexins, Directed Molecular Evolution, Green Fluorescent Proteins, HEK293 Cells, HSP90 Heat-Shock Proteins, Humans, Luminescent Proteins, Nerve Tissue Proteins, Nuclear Localization Signals, Optogenetics, Patch-Clamp Techniques, Photochemistry, Protein Engineering, Recombinant Proteins, Viral Nonstructural Proteins}, issn = {1548-7105}, doi = {10.1038/nmeth.4222}, author = {Zhang, Wei and Lohman, Alexander W and Zhuravlova, Yevgeniya and Lu, Xiaocen and Wiens, Matthew D and Hoi, Hiofan and Yaganoglu, Sine and Mohr, Manuel A and Kitova, Elena N and Klassen, John S and Pantazis, Periklis and Thompson, Roger J and Campbell, Robert E} } @article {67286, title = {Predictive Coding of Novel versus Familiar Stimuli in the Primary Visual Cortex}, journal = {bioRxiv}, year = {2017}, month = {10/2017}, abstract = {

To explore theories of predictive coding, we presented mice with repeated sequences of images with novel images sparsely substituted. Under these conditions, mice could be rapidly trained to lick in response to a novel image, demonstrating a high level of performance on the first day of testing. Using 2-photon calcium imaging to record from layer 2/3 neurons in the primary visual cortex, we found that novel images evoked excess activity in the majority of neurons. When a new stimulus sequence was repeatedly presented, a majority of neurons had similarly elevated activity for the first few presentations, which then decayed to almost zero activity. The decay time of these transient responses was not fixed, but instead scaled with the length of the stimulus sequence. However, at the same time, we also found a small fraction of the neurons within the population (\~2\%) that continued to respond strongly and periodically to the repeated stimulus. Decoding analysis demonstrated that both the transient and sustained responses encoded information about stimulus identity. We conclude that the layer 2/3 population uses a two-channel predictive code: a dense transient code for novel stimuli and a sparse sustained code for familiar stimuli. These results extend and unify existing theories about the nature of predictive neural codes.

}, doi = {10.1101/197608}, url = {https://www.biorxiv.org/content/early/2017/10/03/197608}, author = {Homann, Jan and Koay, Sue Ann and Glidden, Alistair M. and Tank, David W. and Berry, Michael J.} } @article {48992, title = {Protein-templated gold nanoparticle synthesis: protein organization, controlled gold sequestration, and unexpected reaction products.}, journal = {Dalton Transactions (Cambridge, England : 2003)}, volume = {46}, year = {2017}, month = {2017 Dec 21}, pages = {16465-16473}, abstract = {

Emerging applications that exploit the properties of nanoparticles for biotechnology require that the nanoparticles be biocompatible or support biological recognition. These types of particles can be produced through syntheses that involve biologically relevant molecules (proteins or natural extracts, for example). Many of the protocols that rely on these molecules are performed without a clear understanding of the mechanism by which the materials are produced. We have investigated a previously described reaction in which gold nanoparticles are produced from the reaction of chloroauric acid and proteins in solution. We find that modifications to the starting conditions can alter the product from the expected solution-suspended colloids to a product where colloids are formed within a solid, fibrous protein structure. We have interrogated this synthesis, exploiting the change in products to better understand this reaction. We have evaluated the kinetics and products for 7 different proteins over a range of concentrations and temperatures. The key factor that controls the synthetic outcome (colloid or fiber) is the concentration of the protein relative to the gold concentration. We find that the observed fibrous structures are more likely to form at low protein concentrations and when hydrophilic proteins are used. An analysis of the reaction kinetics shows that AuNP formation occurs faster at lower protein (fiber-forming) concentrations than at higher protein (colloid-forming) concentrations. These results contradict traditional expectations for reaction kinetics and protein-fiber formation and are instructive of the manner in which proteins template gold nanoparticle production.

}, issn = {1477-9234}, doi = {10.1039/c7dt03275g}, author = {Hart, Cassidy and Abuladel, Nouf and Bee, Madeleine and Kreider, Megan C and CVitan, Alexander C and Esson, Moira M and Farag, Andrew and Ibeh, Trisha and Kalivas, Eleni N and Larco, Daniel-Mario and Walker Long, Andrew and Lymperopoulos, Loukas and Mendel, Zachary and Miles, Nancy and Zareba, Carly M and Schwabacher, James C and Slucher, Helen and Vinals, Javier and Heddleston, John M and Li, Wenyue and Fox, Douglas M and Hartings, Matthew R} } @article {48343, title = {Quantifying transcription factor binding dynamics at the single-molecule level in live cells.}, journal = {Methods (San Diego, Calif.)}, year = {2017}, month = {2017 Mar 15}, abstract = {

Progressive, technological achievements in the quantitative fluorescence microscopy field are allowing researches from many different areas to start unraveling the dynamic intricacies of biological processes inside living cells. From super-resolution microscopy techniques to tracking of individual proteins, fluorescence microscopy is changing our perspective on how the cell works. Fortunately, a growing number of research groups are exploring single-molecule studies in living cells. However, no clear consensus exists on several key aspects of the technique such as image acquisition conditions, or analysis of the obtained data. Here, we describe a detailed approach to perform single-molecule tracking (SMT) of transcription factors in living cells to obtain key binding characteristics, namely their residence time and bound fractions. We discuss different types of fluorophores, labeling density, microscope, cameras, data acquisition, and data analysis. Using the glucocorticoid receptor as a model transcription factor, we compared alternate tags (GFP, mEOS, HaloTag, SNAP-tag, CLIP-tag) for potential multicolor applications. We also examine different methods to extract the dissociation rates and compare them with simulated data. Finally, we discuss several challenges that this exciting technique still faces.

}, issn = {1095-9130}, doi = {10.1016/j.ymeth.2017.03.014}, author = {Presman, Diego M and Ball, David A and Paakinaho, Ville and Grimm, Jonathan B and Lavis, Luke D and Karpova, Tatiana S and Hager, Gordon L} } @article {48650, title = {Rational engineering of photoconvertible fluorescent proteins for dual-color fluorescence nanoscopy enabled by a triplet-state mechanism of primed conversion.}, journal = {Angewandte Chemie (International ed. in English)}, volume = {56}, year = {2017}, month = {2017 Jun 29}, pages = {11628-33}, abstract = {

Green-to-red photoconvertible fluorescent proteins (pcFPs) are powerful tools for super-resolution localization microscopy and protein tagging. Recently, they have been found to undergo efficient photoconversion not only by the traditional 400-nm illumination but also by an alternative method termed primed conversion, employing dual wavelength illumination with blue and far-red/near-infrared light. Primed conversion has been reported only for Dendra2 and its mechanism has remained elusive. Here, we uncover the molecular mechanism of primed conversion by reporting the intermediate "primed" state to be a triplet dark state formed by intersystem crossing. We show that formation of this state can be influenced by the introduction of serine or threonine at sequence position 69 (Eos notation) and use this knowledge to create "pr"- (for primed convertible) variants of most known green-to-red pcFPs.

}, issn = {1521-3773}, doi = {10.1002/anie.201706121}, author = {Mohr, Manuel Alexander and Kobitski, Andrei Yu and Sabater, Lluc Rullan and Nienhaus, Karin and Obara, Christopher John and Lippincott-Schwartz, Jennifer and Nienhaus, Gerd Ulrich and Pantazis, Periklis} } @article {48462, title = {Ring attractor dynamics in the Drosophila central brain.}, journal = {Science (New York, N.Y.)}, volume = {356}, year = {2017}, month = {2017 May 04}, pages = {849-53}, abstract = {

Ring attractors are a class of recurrent networks hypothesized to underlie the representation of heading direction. Such network structures, schematized as a ring of neurons whose connectivity depends on their heading preferences, can sustain a bump-like activity pattern whose location can be updated by continuous shifts along either turn direction. We recently reported that a population of fly neurons represents the animal{\textquoteright}s heading via bump-like activity dynamics. We combined two-photon calcium imaging in head-fixed flying flies with optogenetics to overwrite the existing population representation with an artificial one, which was then maintained by the circuit with naturalistic dynamics. A network with local excitation and global inhibition enforces this unique and persistent heading representation. Ring attractor networks have long been invoked in theoretical work; our study provides physiological evidence of their existence and functional architecture.

}, issn = {1095-9203}, doi = {10.1126/science.aal4835}, author = {Kim, Sung S and Rouault, Herv{\'e} and Druckmann, Shaul and Jayaraman, Vivek} } @article {48787, title = {The role of disulfide bond replacements in analogues of the tarantula toxin ProTx-II and their effects on inhibition of the voltage-gated sodium ion channel Nav1.7.}, journal = {Journal of the American Chemical Society}, volume = {139}, year = {2017}, month = {2017 Sep 07}, pages = {13063-75}, abstract = {

Spider venom toxins, such as Protoxin-II (ProTx-II), have recently received much attention as selective Nav1.7 channel blockers, with potential to be developed as leads for the treatment of chronic nocioceptive pain. ProTx-II is a 30-amino acid peptide with three disulfide bonds that has been reported to adopt a well-defined inhibitory cystine knot (ICK) scaffold structure. Potential drawbacks with such peptides include poor pharmacodynamics and potential scrambling of the disulfide bonds in vivo. In order to address these issues, in the present study we report the solid-phase synthesis of lanthionine-bridged analogues of ProTx-II, in which one of the three disulfide bridges is replaced with a thioether linkage, and evaluate the biological properties of these analogues. We have also investigated the folding and disulfide bridging patterns arising from different methods of oxidation of the linear peptide precursor. Finally, we report the X-ray crystal structure of ProTx-II to atomic resolution; to our knowledge this is the first crystal structure of an ICK spider venom peptide not bound to a substrate.

}, issn = {1520-5126}, doi = {10.1021/jacs.7b06506}, author = {Wright, Zo{\"e} V F and McCarthy, Stephen and Dickman, Rachael and Reyes, Francis E and Sanchez-Martinez, Silvia and Cryar, Adam and Kilford, Ian and Hall, Adrian and Takle, Andrew K and Topf, Maya and Gonen, Tamir and Thalassinos, Konstantinos and Tabor, Alethea B} } @article {48159, title = {Solution of the structure of a calmodulin-peptide complex in a novel configuration from a variably twinned data set.}, journal = {Acta Crystallographica. Section D, Structural Biology}, volume = {73}, year = {2017}, month = {2017 Jan 01}, pages = {22-31}, abstract = {

Structure determination of conformationally variable proteins can prove challenging even when many possible molecular-replacement (MR) search models of high sequence similarity are available. Calmodulin (CaM) is perhaps the best-studied archetype of these flexible proteins: while there are currently \~{}450 structures of significant sequence similarity available in the Protein Data Bank (PDB), novel conformations of CaM and complexes thereof continue to be reported. Here, the details of the solution of a novel peptide-CaM complex structure by MR are presented, in which only one MR solution of marginal quality was found despite the use of 120 different search models, an exclusivity enhanced by the presence of a high degree of hemihedral twinning (overall refined twin fraction = 0.43). Ambiguities in the initial MR electron-density maps were overcome by using MR-SAD: phases from the MR partial model were used to identify weak anomalous scatterers (calcium, sulfur and chloride), which were in turn used to improve the phases, automatically rebuild the structure and resolve sequence ambiguities. Retrospective analysis of consecutive wedges of the original data sets showed twin fractions ranging from 0.32 to 0.55, suggesting that the data sets were variably twinned. Despite these idiosyncrasies and obstacles, the data themselves and the final model were of high quality and indeed showed a novel, nearly right-angled conformation of the bound peptide.

}, issn = {2059-7983}, doi = {10.1107/S2059798316019318}, author = {Keller, Jacob Pearson} } @conference {67293, title = {Stochastic filtering of two-photon imaging using reweighted l1}, booktitle = {2017 IEEE International Conference on Acoustics, Speech and Signal Processing (ICASSP)}, year = {2017}, month = {03/2017}, publisher = {IEEE}, organization = {IEEE}, address = {New Orleans, LA}, abstract = {
Two-photon (TP) calcium imaging is an important imaging modality in neuroscience, allowing for large-scale recording of neural activity in awake, behaving animals at behavior-relevant timescales. Interpretation of TP data requires the accurate extraction of temporal neural activity traces, which can be accomplished via manual or automated methods. In this work we seek to improve the accuracy of both manual and automated TP microscopy demixing methods by introducing a denoising algorithm based on a statistical model of TP data which includes spatial contiguity, sparse activity and Poisson observations. Our method leverages recent developments in stochastic filtering of structured signals based on Laplacian-scale mixture models (LSMs) to model the neural activity in TP data as a set of spatially correlated sparse variables. We apply our method on TP images taken from the visual cortex of an awake, behaving mouse, and demonstrate improved neural activity demixing over current pre-processing techniques.
}, doi = {10.1109/ICASSP.2017.7952314}, url = {https://ieeexplore.ieee.org/abstract/document/7952314}, author = {Charles, Adam S. and Song, Alexander and Koay, Sue Ann and Tank, David W. and Pillow, Jonathan W.} } @article {48346, title = {Stochastic protein labeling enables long-term single molecule observation in vivo.}, journal = {bioRxiv}, year = {2017}, month = {2017 Mar 13}, abstract = {Our ability to unambiguously image and track individual molecules in live cells is limited by packing of multiple copies of labeled molecules within the resolution limit. Here we devise a universal genetic strategy to precisely control protein copy number in a cell. This system has a dynamic titration range of more than 10,000 fold, enabling sparse labeling of proteins expressed at widely different levels. Combined with fluorescence signal amplification tags, this system extends the duration of automated single-molecule tracking by 2 orders of magnitude. We demonstrate long-term imaging of synaptic vesicle dynamics in cultured neurons as well as in live zebrafish. We found that axon initial segment utilizes a waterfall mechanism gating synaptic vesicle transport polarity by promoting anterograde transport processivity. Long-time observation also reveals that transcription factor Sox2 samples clustered binding sites in spatially-restricted sub-nuclear regions, suggesting that topological structures in the nucleus shape local gene activities by a sequestering mechanism. This strategy thus greatly expands the spatiotemporal length scales of live-cell single-molecule measurements for a quantitative understanding of complex control of molecular dynamics in vivo.}, doi = {10.1101/116186}, author = {Liu, Hui and Dong, Peng and Ioannou, Maria S and Li, Li and Shea, Jamien and Pasolli, H Amalia and Grimm, Jonathan B and Rivlin, Patricia K and Lavis, Luke D and Koyama, Minoru and Liu, Zhe} } @article {48699, title = {Structure and topology around the cleavage site regulate post-translational cleavage of the HIV-1 gp160 signal peptide.}, journal = {eLife}, volume = {6}, year = {2017}, month = {2017 Jul 28}, abstract = {

Like all other secretory proteins, the HIV-1 envelope glycoprotein gp160, is targeted to the endoplasmic reticulum (ER) by its signal peptide during synthesis. Proper gp160 folding in the ER requires core glycosylation, disulfide-bond formation and proline isomerization. Signal-peptide cleavage occurs only late after gp160 chain termination and is dependent on folding of the soluble subunit gp120 to a near-native conformation. We here detail the mechanism by which co-translational signal-peptide cleavage is prevented. Conserved residues from the signal peptide and residues downstream of the canonical cleavage site form an extended alpha-helix in the ER membrane that covers the cleavage site, thus preventing cleavage. A point mutation in the signal peptide breaks the alpha helix allowing co-translational cleavage. We demonstrate that postponed cleavage of gp160 enhances functional folding of the molecule. The change to early cleavage results in decreased viral fitness compared to wild-type HIV.

}, issn = {2050-084X}, doi = {10.7554/eLife.26067}, author = {Snapp, Erik Lee and McCaul, Nicholas and Quandte, Matthias and Cabartova, Zuzana and Bontjer, Ilja and K{\"a}llgren, Carolina and Nilsson, IngMarie and Land, Aafke and von Heijne, Gunnar and Sanders, Rogier W and Braakman, Ineke} } @article {48865, title = {Structure of RNA polymerase bound to ribosomal 30S subunit.}, journal = {eLife}, volume = {6}, year = {2017}, month = {2017 Oct 13}, abstract = {

In bacteria, mRNA transcription and translation are coupled to coordinate optimal gene expression and maintain genome stability. Coupling is thought to involve direct interactions between RNA polymerase (RNAP) and the translational machinery. We present cryo-EM structures of E. coli RNAP core bound to the small ribosomal 30S subunit. The complex is stable under cell-like ionic conditions, consistent with functional interaction between RNAP and the 30S subunit. The RNA exit tunnel of RNAP aligns with the Shine-Dalgarno-binding site of the 30S subunit. Ribosomal protein S1 forms a wall of the tunnel between RNAP and the 30S subunit, consistent with its role in directing mRNAs onto the ribosome. The nucleic-acid-binding cleft of RNAP samples distinct conformations, suggesting different functional states during transcription-translation coupling. The architecture of the 30S{\textbullet}RNAP complex provides a structural basis for co-localization of the transcriptional and translational machineries, and inform future mechanistic studies of coupled transcription and translation.

}, issn = {2050-084X}, doi = {10.7554/eLife.28560}, author = {Demo, Gabriel and Rasouly, Aviram and Vasilyev, Nikita and Svetlov, Vladimir and Loveland, Anna B and Diaz-Avalos, Ruben and Grigorieff, Nikolaus and Nudler, Evgeny and Korostelev, Andrei A} } @article {48657, title = {A systematic nomenclature for the Drosophila ventral nervous system.}, journal = {bioRxiv}, year = {2017}, month = {2017 Apr 26}, abstract = {

Insect nervous systems are proven and powerful model systems for neuroscience research with wide relevance in biology and medicine. However, descriptions of insect brains have suffered from a lack of a complete and uniform nomenclature. Recognising this problem the Insect Brain Name Working Group produced the first agreed hierarchical nomenclature system for the adult insect brain, using Drosophila melanogaster as the reference framework, with other insect taxa considered to ensure greater consistency and expandability (Ito et al., 2014). Ito et al. (2014) purposely focused on the gnathal regions that account for approximately 50\% of the adult CNS. We extend this nomenclature system to the sub-gnathal regions of the adult Drosophila nervous system to provide a nomenclature of the so-called ventral nervous system (VNS), which includes the thoracic and abdominal neuromeres that was not included in the original work and contains the neurons that play critical roles underpinning most fly behaviours.

}, doi = {10.1101/122952}, author = {Court, Robert Christopher and Armstrong, James Douglas and Borner, Jana and Card, Gwyneth M and Costa, Marta and Dickinson, Michael H and Duch, Carsten and Korff, Wyatt and Mann, Richard S and Merritt, David and Murphey, Rodney K and Namiki, Shigehiro and Seeds, Andrew M and Shepherd, David and Shirangi, Troy R and Simpson, Julie H and Truman, James W and Tuthill, John C and Williams, Darren W} } @article {48193, title = {TED: A Tolerant Edit Distance for segmentation evaluation.}, journal = {Methods}, volume = {115}, year = {2017}, month = {2017 Jan 17}, pages = {119-27}, abstract = {

In this paper, we present a novel error measure to compare a computer-generated segmentation of images or volumes against ground truth. This measure, which we call Tolerant Edit Distance (TED), is motivated by two observations that we usually encounter in biomedical image processing: (1) Some errors, like small boundary shifts, are tolerable in practice. Which errors are tolerable is application dependent and should be explicitly expressible in the measure. (2) Non-tolerable errors have to be corrected manually. The effort needed to do so should be reflected by the error measure. Our measure is the minimal weighted sum of split and merge operations to apply to one segmentation such that it resembles another segmentation within specified tolerance bounds. This is in contrast to other commonly used measures like Rand index or variation of information, which integrate small, but tolerable, differences. Additionally, the TED provides intuitive numbers and allows the localization and classification of errors in images or volumes. We demonstrate the applicability of the TED on 3D segmentations of neurons in electron microscopy images where topological correctness is arguable more important than exact boundary locations. Furthermore, we show that the TED is not just limited to evaluation tasks. We use it as the loss function in a max-margin learning framework to find parameters of an automatic neuron segmentation algorithm. We show that training to minimize the TED, i.e., to minimize crucial errors, leads to higher segmentation accuracy compared to other learning methods.

}, doi = {10.1016/j.ymeth.2016.12.013}, author = {Funke, Jan and Klein, Jonas and Moreno-Noguer, Francesc and Cardona, Albert and Cook, Matthew} } @article {48512, title = {Time-accuracy tradeoffs in kernel prediction: controlling prediction quality.}, journal = {Journal of Machine Learning Research}, volume = {18}, year = {2017}, month = {2017 Apr 1 }, pages = {1-29}, abstract = {

Kernel regression or classification (also referred to as weighted ε-NN methods in Machine Learning) are appealing for their simplicity and therefore ubiquitous in data analysis. How- ever, practical implementations of kernel regression or classification consist of quantizing or sub-sampling data for improving time efficiency, often at the cost of prediction quality. While such tradeoffs are necessary in practice, their statistical implications are generally not well understood, hence practical implementations come with few performance guaran- tees. In particular, it is unclear whether it is possible to maintain the statistical accuracy of kernel prediction{\textemdash}crucial in some applications{\textemdash}while improving prediction time.

The present work provides guiding principles for combining kernel prediction with data- quantization so as to guarantee good tradeoffs between prediction time and accuracy, and in particular so as to approximately maintain the good accuracy of vanilla kernel prediction.

Furthermore, our tradeoff guarantees are worked out explicitly in terms of a tuning parameter which acts as a knob that favors either time or accuracy depending on practical needs. On one end of the knob, prediction time is of the same order as that of single-nearest- neighbor prediction (which is statistically inconsistent) while maintaining consistency; on the other end of the knob, the prediction risk is nearly minimax-optimal (in terms of the original data size) while still reducing time complexity. The analysis thus reveals the interaction between the data-quantization approach and the kernel prediction method, and most importantly gives explicit control of the tradeoff to the practitioner rather than fixing the tradeoff in advance or leaving it opaque.

The theoretical results are validated on data from a range of real-world application domains; in particular we demonstrate that the theoretical knob performs as expected.\ 

}, url = {http://jmlr.org/papers/volume18/16-538/16-538.pdf}, author = {Kpotufe, Samory and Verma, Nakul} } @article {48985, title = {A topographic axis of transcriptional identity in thalamus.}, journal = {bioRxiv}, year = {2017}, month = {2017 Dec 31}, pages = {241315}, abstract = {

A fundamental goal in neuroscience is to uncover common principles by which different modalities of information are processed. In the mammalian brain, thalamus acts as the essential hub for forebrain circuits handling inputs from sensory, motor, limbic, and cognitive pathways. Whether thalamus imposes common transformations on each of these modalities is unknown. Molecular characterization offers a principled approach to revealing the organization of thalamus. Using near-comprehensive and projection-specific transcriptomic sequencing, we found that almost all thalamic nuclei fit into one of three profiles. These profiles lie on a single axis of genetic variance which is aligned with the mediolateral spatial axis of thalamus. Genes defining this axis of variance include receptors and ion channels, providing a systematic diversification of input/output transformations across the topography of thalamus. Single cell transcriptional profiling revealed graded heterogeneity within individual thalamic nuclei, demonstrating that a spectrum of cell types and potentially diverse input/output transforms exist within a given thalamic nucleus. Together, our data argue for an archetypal organization of pathways serving diverse input modalities, and provides a comprehensive organizational scheme for thalamus.

}, doi = {10.1101/241315}, author = {Phillips, James W and Schulman, Anton and Hara, Erina and Liu, Chenghao and Shields, Brenda C and Korff, Wyatt and Lemire, Andrew and Dudman, Joshua T and Nelson, Sacha B and Hantman, Adam W} } @article {48367, title = {Trainable Weka Segmentation: a machine learning tool for microscopy pixel classification.}, journal = {Bioinformatics (Oxford, England)}, volume = {33}, year = {2017}, month = {2017 Mar 29}, pages = {2424-6}, abstract = {

Summary: State-of-the-art light and electron microscopes are capable of acquiring large image datasets, but quantitatively evaluating the data often involves manually annotating structures of interest. This processis time-consuming and often a major bottleneck in the evaluation pipeline. To overcome this problem, we have introduced the Trainable Weka Segmentation (TWS), a machine learning tool that leveragesa limited number of manual annotations in order to train a classifier and segment the remaining dataautomatically. In addition, TWS can provide unsupervised segmentation learning schemes (clustering) and can be customized to employ user-designed image features or classifiers.

Availability and Implementation: TWS is distributed as open-source software as part of the Fiji image processing distribution of ImageJ at http://imagej.net/Trainable_Weka_Segmentation.

Contact: ignacio.arganda@ehu.eus.

Supplementary information: Supplementary data are available at Bioinformatics online.

}, issn = {1367-4811}, doi = {10.1093/bioinformatics/btx180}, author = {Arganda-Carreras, Ignacio and Kaynig, Verena and Rueden, Curtis and Eliceiri, Kevin W and Schindelin, Johannes and Cardona, Albert and Seung, H Sebastian} } @article {48934, title = {Ultra-selective looming detection from radial motion opponency.}, journal = {Nature}, volume = {551}, year = {2017}, month = {2017 Nov 08}, pages = {237-241}, abstract = {

Nervous systems combine lower-level sensory signals to detect higher-order stimulus features critical to survival, such as the visual looming motion created by an imminent collision or approaching predator. Looming-sensitive neurons have been identified in diverse animal species. Different large-scale visual features such as looming often share local cues, which means loom-detecting neurons face the challenge of rejecting confounding stimuli. Here we report the discovery of an ultra-selective looming detecting neuron, lobula plate/lobula columnar, type II (LPLC2) in Drosophila, and show how its selectivity is established by radial motion opponency. In the fly visual system, directionally selective small-field neurons called T4 and T5 form a spatial map in the lobula plate, where they each terminate in one of four retinotopic layers, such that each layer responds to motion in a different cardinal direction. Single-cell anatomical analysis reveals that each arm of the LPLC2 cross-shaped primary dendrites ramifies in one of these layers and extends along that layer{\textquoteright}s preferred motion direction. In vivo calcium imaging demonstrates that, as their shape predicts, individual LPLC2 neurons respond strongly to outward motion emanating from the centre of the neuron{\textquoteright}s receptive field. Each dendritic arm also receives local inhibitory inputs directionally selective for inward motion opposing the excitation. This radial motion opponency generates a balance of excitation and inhibition that makes LPLC2 non-responsive to related patterns of motion such as contraction, wide-field rotation or luminance change. As a population, LPLC2 neurons densely cover visual space and terminate onto the giant fibre descending neurons, which drive the jump muscle motor neuron to trigger an escape take off. Our findings provide a mechanistic description of the selective feature detection that flies use to discern and escape looming threats.

}, issn = {1476-4687}, doi = {10.1038/nature24626}, author = {Klapoetke, Nathan C and Nern, Aljoscha and Peek, Martin Y and Rogers, Edward M and Breads, Patrick and Rubin, Gerald M and Reiser, Michael B and Card, Gwyneth M} } @article {48304, title = {Video-rate volumetric functional imaging of the brain at synaptic resolution.}, journal = {Nature Neuroscience}, volume = {20}, year = {2017}, month = {2017 Feb 27}, pages = {620-8}, abstract = {

Neurons and neural networks often extend hundreds of micrometers in three dimensions. Capturing the calcium transients associated with their activity requires volume imaging methods with subsecond temporal resolution. Such speed is a challenge for conventional two-photon laser-scanning microscopy, because it depends on serial focal scanning in 3D and indicators with limited brightness. Here we present an optical module that is easily integrated into standard two-photon laser-scanning microscopes to generate an axially elongated Bessel focus, which when scanned in 2D turns frame rate into volume rate. We demonstrated the power of this approach in enabling discoveries for neurobiology by imaging the calcium dynamics of volumes of neurons and synapses in fruit flies, zebrafish larvae, mice and ferrets in vivo. Calcium signals in objects as small as dendritic spines could be resolved at video rates, provided that the samples were sparsely labeled to limit overlap in their axially projected images.

}, issn = {1546-1726}, doi = {10.1038/nn.4516}, author = {Lu, Rongwen and Sun, Wenzhi and Liang, Yajie and Kerlin, Aaron and Bierfeld, Jens and Seelig, Johannes D and Wilson, Daniel E and Scholl, Benjamin and Mohar, Boaz and Tanimoto, Masashi and Koyama, Minoru and Fitzpatrick, David and Orger, Michael B and Ji, Na} } @article {48496, title = {Visualizing dynamic microvillar search and stabilization during ligand detection by T cells.}, journal = {Science (New York, N.Y.)}, volume = {356}, year = {2017}, month = {2017 May 12}, abstract = {

During immune surveillance, T cells survey the surface of antigen-presenting cells. In searching for peptide-loaded major histocompatibility complexes (pMHCs), they must solve a classic trade-off between speed and sensitivity. It has long been supposed that microvilli on T cells act as sensory organs to enable search, but their strategy has been unknown. We used lattice light-sheet and quantum dot-enabled synaptic contact mapping microscopy to show that anomalous diffusion and fractal organization of microvilli survey the majority of opposing surfaces within 1 minute. Individual dwell times were long enough to discriminate pMHC half-lives and T cell receptor (TCR) accumulation selectively stabilized microvilli. Stabilization was independent of tyrosine kinase signaling and the actin cytoskeleton, suggesting selection for avid TCR microclusters. This work defines the efficient cellular search process against which ligand detection takes place.

}, issn = {1095-9203}, doi = {10.1126/science.aal3118}, author = {Cai, En and Marchuk, Kyle and Beemiller, Peter and Beppler, Casey and Rubashkin, Matthew G and Weaver, Valerie M and G{\'e}rard, Audrey and Liu, Tsung-Li and Chen, Bi-Chang and Betzig, Eric and Bartumeus, Frederic and Krummel, Matthew F} } @article {48752, title = {Visualizing long-term single-molecule dynamics in vivo by stochastic protein labeling.}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {115}, year = {2017}, month = {2017 Jan 09}, pages = {343-8}, abstract = {

Our ability to unambiguously image and track individual molecules in live cells is limited by packing of multiple copies of labeled molecules within the resolution limit. Here we devise a universal genetic strategy to precisely control copy number of fluorescently labeled molecules in a cell. This system has a dynamic titration range of \>10,000 fold, enabling sparse labeling of proteins expressed at different abundance levels. Combined with photostable labels, this system extends the duration of automated single-molecule tracking by 2 orders of magnitude. We demonstrate long-term imaging of synaptic vesicle dynamics in cultured neurons as well as in intact\ zebrafish. We found axon initial segment utilizes a "waterfall" mechanism gating synaptic vesicle transport polarity by promoting anterograde transport processivity. Long-time observation also reveals that transcription factor hops between clustered binding sites in spatially-restricted sub-nuclear regions, suggesting that topological structures in the nucleus shape local gene activities by a sequestering mechanism. This strategy thus greatly expands the spatiotemporal length scales of live-cell single-molecule measurements, enabling new experiments to quantitatively understand complex control of molecular dynamics\ in vivo.

}, doi = {10.1073/pnas.1713895115}, author = {Liu, Hui and Dong, Peng and Ioannou, Maria S and Li, Li and Shea, Jamien and Pasolli, H Amalia and Grimm, Jonathan B and Rivlin, Patricia K and Lavis, Luke D and Koyama, Minoru and Liu, Zhe} } @article {48730, title = {Volume imaging.}, volume = {B2}, number = {14/452,181}, year = {2017}, month = {2017 Aug 01}, edition = {G01J 4/00 (20060101); G02B 21/00 (20060101); G02B 21/36 (20060101)}, abstract = {

A system for a laser-scanning microscope includes an optical element configured to transmit light in a first direction onto a first beam path and to reflect light in a second direction to a second beam path that is different from the first beam path; a reflector on the first beam path; and a lens including a variable focal length, the lens positioned on the first beam path. The lens and reflector are positioned relative to each other to cause light transmitted by the optical element to pass through the lens a plurality of times and in a different direction each time. In some implementations, the system also can include a feedback system that receives a signal that represents an amount of focusing of the lens, and changes the focal length of the lens based on the received signal.

}, url = {http://patft.uspto.gov/netacgi/nph-Parser?Sect1=PTO2\&Sect2=HITOFF\&u=\%2Fnetahtml\%2FPTO\%2Fsearch-adv.htm\&r=1\&p=1\&f=G\&l=50\&d=PTXT\&S1=(\%22Howard+Hughes+medical\%22.ASNM.)\&OS=AN/\%22Howard+Hughes+medical\%22\&RS=AN/\%22Howard+Hughes+medical\%22}, author = {Cui, Meng and Kong, Lingjie} } @article {67282, title = {Volumetric two-photon imaging of neurons using stereoscopy (vTwINS)}, journal = {Nature Methods}, volume = {14}, year = {2017}, month = {03/2017}, pages = {420 - 426}, abstract = {

Two-photon laser scanning microscopy of calcium dynamics using fluorescent indicators is a widely used imaging method for large-scale recording of neural activity\ in vivo. Here, we introduce volumetric two-photon imaging of neurons using stereoscopy (vTwINS), a volumetric calcium imaging method that uses an elongated, V-shaped point spread function to image a 3D brain volume. Single neurons project to spatially displaced {\textquoteright}image pairs{\textquoteright} in the resulting 2D image, and the separation distance between projections is proportional to depth in the volume. To demix the fluorescence time series of individual neurons, we introduce a modified orthogonal matching pursuit algorithm that also infers source locations within the 3D volume. We illustrated vTwINS by imaging neural population activity in the mouse primary visual cortex and hippocampus. Our results demonstrated that vTwINS provides an effective method for volumetric two-photon calcium imaging that increases the number of neurons recorded while maintaining a high frame rate.

}, issn = {1548-7091}, doi = {10.1038/nmeth.4226}, url = {http://www.nature.com/articles/nmeth.4226http://www.nature.com/articles/nmeth.4226.pdfhttp://www.nature.com/articles/nmeth.4226.pdfhttp://www.nature.com/articles/nmeth.4226}, author = {Song, Alexander and Charles, Adam S and Koay, Sue Ann and Gauthier, Jeff L and Thiberge, Stephan Y and Pillow, Jonathan W and Tank, David W} } @article {48305, title = {Zyxin regulates endothelial von Willebrand factor secretion by reorganizing actin filaments around exocytic granules.}, journal = {Nature Communications}, volume = {8}, year = {2017}, month = {2017 Mar 03}, pages = {14639}, abstract = {

Endothelial exocytosis of Weibel-Palade body (WPB) is one of the first lines of defence against vascular injury. However, the mechanisms that control WPB exocytosis in the final stages (including the docking, priming and fusion of granules) are poorly understood. Here we show that the focal adhesion protein zyxin is crucial in this process. Zyxin downregulation inhibits the secretion of von Willebrand factor (VWF), the most abundant cargo in WPBs, from human primary endothelial cells (ECs) induced by cAMP agonists. Zyxin-deficient mice exhibit impaired epinephrine-stimulated VWF release, prolonged bleeding time and thrombosis, largely due to defective endothelial secretion of VWF. Using live-cell super-resolution microscopy, we visualize previously unappreciated reorganization of pre-existing actin filaments around WPBs before fusion, dependent on zyxin and an interaction with the actin crosslinker α-actinin. Our findings identify zyxin as a physiological regulator of endothelial exocytosis through reorganizing local actin network in the final stage of exocytosis.

}, issn = {2041-1723}, doi = {10.1038/ncomms14639}, author = {Han, Xiaofan and Li, Pin and Yang, Zhenghao and Huang, Xiaoshuai and Wei, Guoqin and Sun, Yujie and Kang, Xuya and Hu, Xueting and Deng, Qiuping and Chen, Liangyi and He, Aibin and Huo, Yingqing and Li, Dong and Betzig, Eric and Luo, Jincai} } @article {47711, title = {Accurate design of megadalton-scale two-component icosahedral protein complexes.}, journal = {Science (New York, N.Y.)}, volume = {353}, year = {2016}, month = {2016 Jul 22}, pages = {389-94}, abstract = {

Nature provides many examples of self- and co-assembling protein-based molecular machines, including icosahedral protein cages that serve as scaffolds, enzymes, and compartments for essential biochemical reactions and icosahedral virus capsids, which encapsidate and protect viral genomes and mediate entry into host cells. Inspired by these natural materials, we report the computational design and experimental characterization of co-assembling, two-component, 120-subunit icosahedral protein nanostructures with molecular weights (1.8 to 2.8 megadaltons) and dimensions (24 to 40 nanometers in diameter) comparable to those of small viral capsids. Electron microscopy, small-angle x-ray scattering, and x-ray crystallography show that 10 designs spanning three distinct icosahedral architectures form materials closely matching the design models. In vitro assembly of icosahedral complexes from independently purified components occurs rapidly, at rates comparable to those of viral capsids, and enables controlled packaging of molecular cargo through charge complementarity. The ability to design megadalton-scale materials with atomic-level accuracy and controllable assembly opens the door to a new generation of genetically programmable protein-based molecular machines.

}, issn = {1095-9203}, doi = {10.1126/science.aaf8818}, author = {Bale, Jacob B and Gonen, Shane and Liu, Yuxi and Sheffler, William and Ellis, Daniel and Thomas, Chantz and Cascio, Duilio and Yeates, Todd O and Gonen, Tamir and King, Neil P and Baker, David} } @article {47509, title = {Activation of NMDA receptors and the mechanism of inhibition by ifenprodil.}, journal = {Nature}, volume = {534}, year = {2016}, month = {2016 Jun 2}, pages = {63-8}, abstract = {

The physiology of N-methyl-d-aspartate (NMDA) receptors is fundamental to brain development and function. NMDA receptors are ionotropic glutamate receptors that function as heterotetramers composed mainly of GluN1 and GluN2 subunits. Activation of NMDA receptors requires binding of neurotransmitter agonists to a ligand-binding domain (LBD) and structural rearrangement of an amino-terminal domain (ATD). Recent crystal structures of GluN1-GluN2B NMDA receptors bound to agonists and an allosteric inhibitor, ifenprodil, represent the allosterically inhibited state. However, how the ATD and LBD move to activate the NMDA receptor ion channel remains unclear. Here we applied X-ray crystallography, single-particle electron cryomicroscopy and electrophysiology to rat NMDA receptors to show that, in the absence of ifenprodil, the bi-lobed structure of GluN2 ATD adopts an open conformation accompanied by rearrangement of the GluN1-GluN2 ATD heterodimeric interface, altering subunit orientation in the ATD and LBD and forming an active receptor conformation that gates the ion channel.

}, issn = {1476-4687}, doi = {10.1038/nature17679}, author = {Tajima, Nami and Karakas, Erkan and Grant, Timothy and Simorowski, Noriko and Diaz-Avalos, Ruben and Grigorieff, Nikolaus and Furukawa, Hiro} } @article {47987, title = {Adaptive light-sheet microscopy for long-term, high-resolution imaging in living organisms.}, journal = {Nature Biotechnology}, volume = {34}, year = {2016}, month = {2016 Oct 31}, pages = {1267-78}, abstract = {

Optimal image quality in light-sheet microscopy requires a perfect overlap between the illuminating light sheet and the focal plane of the detection objective. However, mismatches between the light-sheet and detection planes are common owing to the spatiotemporally varying optical properties of living specimens. Here we present the AutoPilot framework, an automated method for spatiotemporally adaptive imaging that integrates (i) a multi-view light-sheet microscope capable of digitally translating and rotating light-sheet and detection planes in three dimensions and (ii) a computational method that continuously optimizes spatial resolution across the specimen volume in real time. We demonstrate long-term adaptive imaging of entire developing zebrafish (Danio rerio) and Drosophila melanogaster embryos and perform adaptive whole-brain functional imaging in larval zebrafish. Our method improves spatial resolution and signal strength two to five-fold, recovers cellular and sub-cellular structures in many regions that are not resolved by non-adaptive imaging, adapts to spatiotemporal dynamics of genetically encoded fluorescent markers and robustly optimizes imaging performance during large-scale morphogenetic changes in living organisms.

}, issn = {1546-1696}, doi = {10.1038/nbt.3708}, author = {Royer, Lo{\"\i}c A and Lemon, William C and Chhetri, Raghav K and Wan, Yinan and Coleman, Michael and Myers, Eugene W and Keller, Philipp J} } @article {47894, title = {Adult mouse cortical cell taxonomy revealed by single cell transcriptomics.}, journal = {Nature neuroscience}, volume = {19}, year = {2016}, month = {2016 Feb}, pages = {335-46}, abstract = {

Nervous systems are composed of various cell types, but the extent of cell type diversity is poorly understood. We constructed a cellular taxonomy of one cortical region, primary visual cortex, in adult mice on the basis of single-cell RNA sequencing. We identified 49 transcriptomic cell types, including 23 GABAergic, 19 glutamatergic and 7 non-neuronal types. We also analyzed cell type-specific mRNA processing and characterized genetic access to these transcriptomic types by many transgenic Cre lines. Finally, we found that some of our transcriptomic cell types displayed specific and differential electrophysiological and axon projection properties, thereby confirming that the single-cell transcriptomic signatures can be associated with specific cellular properties.

}, keywords = {Animals, Cell Line, Cerebral Cortex, Classification, gamma-Aminobutyric Acid, Gene Library, Genetic Markers, Glutamic Acid, Interneurons, Male, Mice, Mice, Inbred C57BL, Mice, Transgenic, Neurons, RNA, Sequence Analysis, RNA, Transcriptome, Visual Cortex}, issn = {1546-1726}, doi = {10.1038/nn.4216}, author = {Tasic, Bosiljka and Menon, Vilas and Nguyen, Thuc Nghi and Kim, Tae Kyung and Jarsky, Tim and Yao, Zizhen and Levi, Boaz and Gray, Lucas T and Sorensen, Staci A and Dolbeare, Tim and Bertagnolli, Darren and Goldy, Jeff and Shapovalova, Nadiya and Parry, Sheana and Lee, Changkyu and Smith, Kimberly and Bernard, Amy and Madisen, Linda and Sunkin, Susan M and Hawrylycz, Michael and Koch, Christof and Zeng, Hongkui} } @article {47994, title = {AMPK activation prevents and reverses drug-induced mitochondrial and hepatocyte injury by promoting mitochondrial fusion and function.}, journal = {PLoS One}, volume = {11}, year = {2016}, month = {2016 Oct 31}, pages = {e0165638}, abstract = {

Mitochondrial damage is the major factor underlying drug-induced liver disease but whether conditions that thwart mitochondrial injury can prevent or reverse drug-induced liver damage is unclear. A key molecule regulating mitochondria quality control is AMP activated kinase (AMPK). When activated, AMPK causes mitochondria to elongate/fuse and proliferate, with mitochondria now producing more ATP and less reactive oxygen species. Autophagy is also triggered, a process capable of removing damaged/defective mitochondria. To explore whether AMPK activation could potentially prevent or reverse the effects of drug-induced mitochondrial and hepatocellular damage, we added an AMPK activator to collagen sandwich cultures of rat and human hepatocytes exposed to the hepatotoxic drugs, acetaminophen or diclofenac. In the absence of AMPK activation, the drugs caused hepatocytes to lose polarized morphology and have significantly decreased ATP levels and viability. At the subcellular level, mitochondria underwent fragmentation and had decreased membrane potential due to decreased expression of the mitochondrial fusion proteins Mfn1, 2 and/or Opa1. Adding AICAR, a specific AMPK activator, at the time of drug exposure prevented and reversed these effects. The mitochondria became highly fused and ATP production increased, and hepatocytes maintained polarized morphology. In exploring the mechanism responsible for this preventive and reversal effect, we found that AMPK activation prevented drug-mediated decreases in Mfn1, 2 and Opa1. AMPK activation also stimulated autophagy/mitophagy, most significantly in acetaminophen-treated cells. These results suggest that activation of AMPK prevents/reverses drug-induced mitochondrial and hepatocellular damage through regulation of mitochondrial fusion and autophagy, making it a potentially valuable approach for treatment of drug-induced liver injury.

}, issn = {1932-6203}, doi = {10.1371/journal.pone.0165638}, author = {Kang, Sun Woo Sophie and Haydar, Ghada and Taniane, Caitlin and Farrell, Geoffrey and Arias, Irwin M and Lippincott-Schwartz, Jennifer and Fu, Dong} } @article {48163, title = {Automated detection and quantification of single RNAs at cellular resolution in zebrafish embryos.}, journal = {Development (Cambridge, England)}, volume = {143}, year = {2016}, month = {2016 Feb 01}, pages = {540-6}, abstract = {

Analysis of differential gene expression is crucial for the study of cell fate and behavior during embryonic development. However, automated methods for the sensitive detection and quantification of RNAs at cellular resolution in embryos are lacking. With the advent of single-molecule fluorescence in situ hybridization (smFISH), gene expression can be analyzed at single-molecule resolution. However, the limited availability of protocols for smFISH in embryos and the lack of efficient image analysis pipelines have hampered quantification at the (sub)cellular level in complex samples such as tissues and embryos. Here, we present a protocol for smFISH on zebrafish embryo sections in combination with an image analysis pipeline for automated transcript detection and cell segmentation. We use this strategy to quantify gene expression differences between different cell types and identify differences in subcellular transcript localization between genes. The combination of our smFISH protocol and custom-made, freely available, analysis pipeline will enable researchers to fully exploit the benefits of quantitative transcript analysis at cellular and subcellular resolution in tissues and embryos.

}, keywords = {Animals, Automation, Cell Membrane, Embryo, Nonmammalian, Gene Expression Regulation, Developmental, In Situ Hybridization, Fluorescence, RNA, RNA, Messenger, Subcellular Fractions, Transcription, Genetic, Zebrafish}, issn = {1477-9129}, doi = {10.1242/dev.128918}, author = {Stapel, L Carine and Lombardot, Benoit and Broaddus, Coleman and Kainmueller, Dagmar and Jug, Florian and Myers, Eugene W and Vastenhouw, Nadine L} } @article {47393, title = {The basal ganglia: from motor commands to the control of vigor.}, journal = {Current Opinion in Neurobiology}, volume = {37}, year = {2016}, month = {2016 Mar 21}, pages = {158-66}, abstract = {

Vertebrates are remarkable for their ability to select and execute goal-directed actions: motor skills critical for thriving in complex, competitive environments. A key aspect of a motor skill is the ability to execute its component movements over a range of speeds, amplitudes and frequencies (vigor). Recent work has indicated that a subcortical circuit, the basal ganglia, is a critical determinant of movement vigor in rodents and primates. We propose that the basal ganglia evolved from a circuit that in lower vertebrates and some mammals is sufficient to directly command simple or stereotyped movements to one that indirectly controls the vigor of goal-directed movements. The implications of a dual role of the basal ganglia in the control of vigor and response to reward are also discussed.

}, issn = {1873-6882}, doi = {10.1016/j.conb.2016.02.005}, author = {Dudman, Joshua T and Krakauer, John W} } @article {47463, title = {Brain derived neurotrophic factor differentially modulates excitability of two classes of hippocampal output neurons.}, journal = {Journal of Neurophysiology}, volume = {116}, year = {2016}, month = {2016 May 4}, pages = {466-71}, abstract = {

Brain-derived neurotrophic factor (BDNF) plays an important role in hippocampus-dependent learning and memory. Canonically, this has been ascribed to an enhancing effect on neuronal excitability and synaptic plasticity in the CA1 region. However, it is the pyramidal neurons in the subiculum that form the primary efferent pathways conveying hippocampal information to other areas of the brain, and yet the effect of BDNF on these neurons has remained unexplored. We present new data that BDNF regulates neuronal excitability and cellular plasticity in a much more complex manner than previously suggested. Subicular pyramidal neurons can be divided into two major classes, which have different electrophysiological and morphological properties, different requirements for the induction of plasticity and different extra-hippocampal projections. We found that BDNF increases excitability in one class of subicular pyramidal neurons, yet decreases excitability of the other class. Further, while endogenous BDNF was necessary for the induction of synaptic plasticity in both cell types, BDNF enhanced intrinsic plasticity in one class of pyramidal neurons, yet suppressed intrinsic plasticity in the other. Taken together, these data suggest a novel role for BDNF signaling, as it appears to dynamically and bidirectionally regulate the output of hippocampal information to different regions of the brain.

}, issn = {1522-1598}, doi = {10.1152/jn.00186.2016}, author = {Graves, Austin R and Moore, Shannon J and Spruston, Nelson and Tryba, Andrew Kieran and Kaczorowski, Catherine C} } @article {47527, title = {A bright cyan-excitable orange fluorescent protein facilitates dual-emission microscopy and enhances bioluminescence imaging in vivo.}, journal = {Nature Biotechnology}, volume = {34}, year = {2016}, month = {2016 May 30}, pages = {760-7}, abstract = {

Orange-red fluorescent proteins (FPs) are widely used in biomedical research for multiplexed epifluorescence microscopy with GFP-based probes, but their different excitation requirements make multiplexing with new advanced microscopy methods difficult. Separately, orange-red FPs are useful for deep-tissue imaging in mammals owing to the relative tissue transmissibility of orange-red light, but their dependence on illumination limits their sensitivity as reporters in deep tissues. Here we describe CyOFP1, a bright, engineered, orange-red FP that is excitable by cyan light. We show that CyOFP1 enables single-excitation multiplexed imaging with GFP-based probes in single-photon and two-photon microscopy, including time-lapse imaging in light-sheet systems. CyOFP1 also serves as an efficient acceptor for resonance energy transfer from the highly catalytic blue-emitting luciferase NanoLuc. An optimized fusion of CyOFP1 and NanoLuc, called Antares, functions as a highly sensitive bioluminescent reporter in vivo, producing substantially brighter signals from deep tissues than firefly luciferase and other bioluminescent proteins.

}, issn = {1546-1696}, doi = {10.1038/nbt.3550}, author = {Chu, Jun and Oh, Younghee and Sens, Alex and Ataie, Niloufar and Dana, Hod and Macklin, John J and Laviv, Tal and Welf, Erik S and Dean, Kevin M and Zhang, Feijie and Kim, Benjamin B and Tang, Clement Tran and Hu, Michelle and Baird, Michelle A and Davidson, Michael W and Kay, Mark A and Fiolka, Reto and Yasuda, Ryohei and Kim, Douglas S and Ng, Ho-Leung and Lin, Michael Z} } @article {47324, title = {A circuit mechanism for the propagation of waves of muscle contraction in Drosophila.}, journal = {eLife}, volume = {5}, year = {2016}, month = {2016 Feb 18}, abstract = {

Animals move by adaptively coordinating the sequential activation of muscles. The circuit mechanisms underlying coordinated locomotion are poorly understood. Here, we report on a novel circuit for propagation of waves of muscle contraction, using the peristaltic locomotion of Drosophila larvae as a model system. We found an intersegmental chain of synaptically connected neurons, alternating excitatory and inhibitory, necessary for wave propagation and active in phase with the wave. The excitatory neurons (A27h) are premotor and necessary only for forward locomotion, and are modulated by stretch receptors and descending inputs. The inhibitory neurons (GDL) are necessary for both forward and backward locomotion, suggestive of different yet coupled central pattern generators, and its inhibition is necessary for wave propagation. The circuit structure and functional imaging indicated that the commands to contract one segment promote the relaxation of the next segment, revealing a mechanism for wave propagation in peristaltic locomotion.

}, issn = {2050-084X}, doi = {10.7554/eLife.13253}, author = {Fushiki, Akira and Zwart, Maarten F and Kohsaka, Hiroshi and Fetter, Richard D and Cardona, Albert and Nose, Akinao} } @article {65008, title = {A circuit motif in the zebrafish hindbrain for a two alternative behavioral choice to turn left or right.}, journal = {Elife}, volume = {5}, year = {2016}, month = {2016 08 09}, abstract = {

Animals collect sensory information from the world and make adaptive choices about how to respond to it. Here, we reveal a network motif in the brain for one of the most fundamental behavioral choices made by bilaterally symmetric animals: whether to respond to a sensory stimulus by moving to the left or to the right. We define network connectivity in the hindbrain important for the lateralized escape behavior of zebrafish and then test the role of neurons by using laser ablations and behavioral studies. Key inhibitory neurons in the circuit lie in a column of morphologically similar cells that is one of a series of such columns that form a developmental and functional ground plan for building hindbrain networks. Repetition within the columns of the network motif we defined may therefore lie at the foundation of other lateralized behavioral choices.

}, keywords = {Action Potentials, Animals, Animals, Genetically Modified, Behavior, Animal, Choice Behavior, Laser Therapy, Locomotion, Neural Pathways, Neurons, Patch-Clamp Techniques, Perception, Rhombencephalon, Zebrafish}, issn = {2050-084X}, doi = {10.7554/eLife.16808}, author = {Koyama, Minoru and Minale, Francesca and Shum, Jennifer and Nishimura, Nozomi and Schaffer, Chris B and Fetcho, Joseph R} } @conference {48179, title = {Convexity shape constraints for image segmentation.}, booktitle = {IEEE Conference on Computer Vision and Pattern Recognition (CVPR)}, year = {2016}, month = {2016 Jun 27}, publisher = {IEEE}, organization = {IEEE}, address = {Las Vegas, NV}, abstract = {

Segmenting an image into multiple components is a central task in computer vision. In many practical scenarios, prior knowledge about plausible components is available. Incorporating such prior knowledge into models and algorithms for image segmentation is highly desirable, yet can be non-trivial. In this work, we introduce a new approach that allows, for the first time, to constrain some or all components of a segmentation to have convex shapes. Specifically, we extend the Minimum Cost Multicut Problem by a class of constraints that enforce convexity. To solve instances of this NP-hard integer linear program to optimality, we separate the proposed constraints in the branch-and-cut loop of a state-of-the-art ILP solver. Results on photographs and micrographs demonstrate the effectiveness of the approach as well as its advantages over the state-of-the-art heuristic.

}, doi = {10.1109/CVPR.2016.50}, author = {Royer, Lo{\"\i}c A and Richmond, David L and Rother, Carsten and Andres, Bjoern and Kainmueller, Dagmar} } @article {47353, title = {Data publication with the structural biology data grid supports live analysis.}, journal = {Nature Communications}, volume = {7}, year = {2016}, month = {2016 Mar 07}, pages = {10882}, abstract = {

Access to experimental X-ray diffraction image data is fundamental for validation and reproduction of macromolecular models and indispensable for development of structural biology processing methods. Here, we established a diffraction data publication and dissemination system, Structural Biology Data Grid (SBDG; data.sbgrid.org), to preserve primary experimental data sets that support scientific publications. Data sets are accessible to researchers through a community driven data grid, which facilitates global data access. Our analysis of a pilot collection of crystallographic data sets demonstrates that the information archived by SBDG is sufficient to reprocess data to statistics that meet or exceed the quality of the original published structures. SBDG has extended its services to the entire community and is used to develop support for other types of biomedical data sets. It is anticipated that access to the experimental data sets will enhance the paradigm shift in the community towards a much more dynamic body of continuously improving data analysis.

}, issn = {2041-1723}, doi = {10.1038/ncomms10882}, author = {Meyer, Peter A and Socias, Stephanie and Key, Jason and Ransey, Elizabeth and Tjon, Emily C and Buschiazzo, Alejandro and Lei, Ming and Botka, Chris and Withrow, James and Neau, David and Rajashankar, Kanagalaghatta and Anderson, Karen S and Baxter, Richard H and Blacklow, Stephen C and Boggon, Titus J and Bonvin, Alexandre M J J and Borek, Dominika and Brett, Tom J and Caflisch, Amedeo and Chang, Chung-I and Chazin, Walter J and Corbett, Kevin D and Cosgrove, Michael S and Crosson, Sean and Dhe-Paganon, Sirano and Di Cera, Enrico and Drennan, Catherine L and Eck, Michael J and Eichman, Brandt F and Fan, Qing R and Ferr{\'e}-D{\textquoteright}Amar{\'e}, Adrian R and Christopher Fromme, J and Garcia, K Christopher and Gaudet, Rachelle and Gong, Peng and Harrison, Stephen C and Heldwein, Ekaterina E and Jia, Zongchao and Keenan, Robert J and Kruse, Andrew C and Kvansakul, Marc and McLellan, Jason S and Modis, Yorgo and Nam, Yunsun and Otwinowski, Zbyszek and Pai, Emil F and Pereira, Pedro Jos{\'e} Barbosa and Petosa, Carlo and Raman, C S and Rapoport, Tom A and Roll-Mecak, Antonina and Rosen, Michael K and Rudenko, Gabby and Schlessinger, Joseph and Schwartz, Thomas U and Shamoo, Yousif and Sondermann, Holger and Tao, Yizhi J and Tolia, Niraj H and Tsodikov, Oleg V and Westover, Kenneth D and Wu, Hao and Foster, Ian and Fraser, James S and Maia, Filipe R N C and Gonen, Tamir and Kirchhausen, Tom and Diederichs, Kay and Crosas, Merc{\`e} and Sliz, Piotr} } @article {47508, title = {Design and synthesis of a calcium-sensitive photocage.}, journal = {Angewandte Chemie (International ed. in English)}, year = {2016}, month = {2016 May 24}, abstract = {

Photolabile protecting groups (or "photocages") enable precise spatiotemporal control of chemical functionality and facilitate advanced biological experiments. Extant photocages exhibit a simple input-output relationship, however, where application of light elicits a photochemical reaction irrespective of the environment. Herein, we refine and extend the concept of photolabile groups, synthesizing the first Ca(2+) -sensitive photocage. This system functions as a chemical coincidence detector, releasing small molecules only in the presence of both light and elevated [Ca(2+) ]. Caging a fluorophore with this ion-sensitive moiety yields an "ion integrator" that permanently marks cells undergoing high Ca(2+) flux during an illumination-defined time period. Our general design concept demonstrates a new class of light-sensitive material for cellular imaging, sensing, and targeted molecular delivery.

}, issn = {1521-3773}, doi = {10.1002/anie.201602941}, author = {Heckman, Laurel M and Grimm, Jonathan B and Schreiter, Eric R and Kim, Charles and Verdecia, Mark A and Shields, Brenda C and Lavis, Luke D} } @article {47562, title = {Design of a hyperstable 60-subunit protein icosahedron.}, journal = {Nature}, year = {2016}, month = {2016 Jun 15}, abstract = {

The icosahedron is the largest of the Platonic solids, and icosahedral protein structures are widely used in biological systems for packaging and transport. There has been considerable interest in repurposing such structures for applications ranging from targeted delivery to multivalent immunogen presentation. The ability to design proteins that self-assemble into precisely specified, highly ordered icosahedral structures would open the door to a new generation of protein containers with properties custom-tailored to specific applications. Here we describe the computational design of a 25-nanometre icosahedral nanocage that self-assembles from trimeric protein building blocks. The designed protein was produced in Escherichia coli, and found by electron microscopy to assemble into a homogenous population of icosahedral particles nearly identical to the design model. The particles are stable in 6.7 molar guanidine hydrochloride at up to 80 degrees Celsius, and undergo extremely abrupt, but reversible, disassembly between 2 molar and 2.25 molar guanidinium thiocyanate. The icosahedron is robust to genetic fusions: one or two copies of green fluorescent protein (GFP) can be fused to each of the 60 subunits to create highly fluorescent {\textquoteright}standard candles{\textquoteright} for use in light microscopy, and a designed protein pentamer can be placed in the centre of each of the 20 pentameric faces to modulate the size of the entrance/exit channels of the cage. Such robust and customizable nanocages should have considerable utility in targeted drug delivery, vaccine design and synthetic biology.

}, issn = {1476-4687}, doi = {10.1038/nature18010}, author = {Hsia, Yang and Bale, Jacob B and Gonen, Shane and Shi, Dan and Sheffler, William and Fong, Kimberly K and Nattermann, Una and Xu, Chunfu and Huang, Po-Ssu and Ravichandran, Rashmi and Yi, Sue and Davis, Trisha N and Gonen, Tamir and King, Neil P and Baker, David} } @article {47934, title = {A designer AAV variant permits efficient retrograde access to projection neurons.}, journal = {Neuron}, volume = {92}, year = {2016}, month = {2016 Oct 19}, pages = {372-82}, abstract = {

Efficient retrograde access to projection neurons for the delivery of sensors and effectors constitutes an important and enabling capability for neural circuit dissection. Such an approach would also be useful for gene therapy, including the treatment of neurodegenerative disorders characterized by pathological spread through functionally connected and highly distributed networks. Viral vectors, in particular, are powerful gene delivery vehicles for the nervous system, but all available tools suffer from inefficient retrograde transport or limited clinical potential. To address this need, we applied in\ vivo directed evolution to engineer potent retrograde functionality into the capsid of adeno-associated virus (AAV), a vector that has shown promise in neuroscience research and the clinic. A newly evolved variant, rAAV2-retro, permits robust retrograde access to projection neurons with efficiency comparable to classical synthetic retrograde tracers and enables sufficient sensor/effector expression for functional circuit interrogation and in\ vivo genome editing in targeted neuronal populations. VIDEO ABSTRACT.

}, issn = {1097-4199}, doi = {10.1016/j.neuron.2016.09.021}, author = {Tervo, D Gowanlock R and Hwang, Bum-Yeol and Viswanathan, Sarada and Gaj, Thomas and Lavzin, Maria and Ritola, Kimberly D and Lindo, Sarah and Michael, Susan and Kuleshova, Elena and Ojala, David and Huang, Cheng-Chiu and Gerfen, Charles R and Schiller, Jackie and Dudman, Joshua T and Hantman, Adam W and Looger, Loren L and Schaffer, David V and Karpova, Alla Y} } @article {47434, title = {Direct neural pathways convey distinct visual information to Drosophila mushroom bodies.}, journal = {eLife}, volume = {5}, year = {2016}, month = {2016 Apr 15}, pages = {e14009}, abstract = {

Previously, we identified that visual and olfactory associative memories of Drosophila share the mushroom body (MB) circuits (Vogt et al. 2014). Despite well-characterized odor representations in the Drosophila MB, the MB circuit for visual information is totally unknown. Here we show that a small subset of MB Kenyon cells (KCs) selectively responds to visual but not olfactory stimulation. The dendrites of these atypical KCs form a ventral accessory calyx (vAC), distinct from the main calyx that receives olfactory input. We identified two types of visual projection neurons (VPNs) directly connecting the optic lobes and the vAC. Strikingly, these VPNs are differentially required for visual memories of color and brightness. The segregation of visual and olfactory domains in the MB allows independent processing of distinct sensory memories and may be a conserved form of sensory representations among insects.

}, issn = {2050-084X}, doi = {10.7554/eLife.14009}, author = {Vogt, Katrin and Aso, Yoshinori and Hige, Toshihide and Knapek, Stephan and Ichinose, Toshiharu and Friedrich, Anja B and Turner, Glenn C and Rubin, Gerald M and Tanimoto, Hiromu} } @article {48300, title = {Distinct subpopulations of FOXD1 stroma-derived cells regulate renal erythropoietin.}, journal = {The Journal of Clinical Investigation}, volume = {126}, year = {2016}, month = {2016 May 02}, pages = {1926-38}, abstract = {

Renal peritubular interstitial fibroblast-like cells are critical for adult erythropoiesis, as they are the main source of erythropoietin (EPO). Hypoxia-inducible factor 2 (HIF-2) controls EPO synthesis in the kidney and liver and is regulated by prolyl-4-hydroxylase domain (PHD) dioxygenases PHD1, PHD2, and PHD3, which function as cellular oxygen sensors. Renal interstitial cells with EPO-producing capacity are poorly characterized, and the role of the PHD/HIF-2 axis in renal EPO-producing cell (REPC) plasticity is unclear. Here we targeted the PHD/HIF-2/EPO axis in FOXD1 stroma-derived renal interstitial cells and examined the role of individual PHDs in REPC pool size regulation and renal EPO output. Renal interstitial cells with EPO-producing capacity were entirely derived from FOXD1-expressing stroma, and Phd2 inactivation alone induced renal Epo in a limited number of renal interstitial cells. EPO induction was submaximal, as hypoxia or pharmacologic PHD inhibition further increased the REPC fraction among Phd2-/- renal interstitial cells. Moreover, Phd1 and Phd3 were differentially expressed in renal interstitium, and heterozygous deficiency for Phd1 and Phd3 increased REPC numbers in Phd2-/- mice. We propose that FOXD1 lineage renal interstitial cells consist of distinct subpopulations that differ in their responsiveness to Phd2 inactivation and thus regulation of HIF-2 activity and EPO production under hypoxia or conditions of pharmacologic or genetic PHD inactivation.

}, issn = {1558-8238}, doi = {10.1172/JCI83551}, author = {Kobayashi, Hanako and Liu, Qingdu and Binns, Thomas C and Urrutia, Andres A and Davidoff, Olena and Kapitsinou, Pinelopi P and Pfaff, Andrew S and Olauson, Hannes and Wernerson, Annika and Fogo, Agnes B and Fong, Guo-Hua and Gross, Kenneth W and Haase, Volker H} } @article {47394, title = {Dysfunctional calcium and glutamate signaling in striatal astrocytes from Huntington{\textquoteright}s Disease model mice.}, journal = {The Journal of Neuroscience : the official journal of the Society for Neuroscience}, volume = {36}, year = {2016}, month = {2016 Mar 23}, pages = {3453-70}, abstract = {

UNLABELLED: Astrocytes tile the entire CNS, but their functions within neural circuits in health and disease remain incompletely understood. We used genetically encoded Ca(2+)and glutamate indicators to explore the rules for astrocyte engagement in the corticostriatal circuit of adult wild-type (WT) and Huntington{\textquoteright}s disease (HD) model mice at ages not accompanied by overt astrogliosis (at approximately postnatal days 70-80). WT striatal astrocytes displayed extensive spontaneous Ca(2+)signals, but did not respond to cortical stimulation, implying that astrocytes were largely disengaged from cortical input in healthy tissue. In contrast, in HD model mice, spontaneous Ca(2+)signals were significantly reduced in frequency, duration, and amplitude, but astrocytes responded robustly to cortical stimulation with evoked Ca(2+)signals. These action-potential-dependent astrocyte Ca(2+)signals were mediated by neuronal glutamate release during cortical stimulation, accompanied by prolonged extracellular glutamate levels near astrocytes and tightly gated by Glt1 glutamate transporters. Moreover, dysfunctional Ca(2+)and glutamate signaling that was observed in HD model mice was largely, but not completely, rescued by astrocyte specific restoration of Kir4.1, emphasizing the important contributions of K(+)homeostatic mechanisms that are known to be reduced in HD model mice. Overall, our data show that astrocyte engagement in the corticostriatal circuit is markedly altered in HD. Such prodromal astrocyte dysfunctions may represent novel therapeutic targets in HD and other brain disorders.

SIGNIFICANCE STATEMENT: We report how early-onset astrocyte dysfunction without detectable astrogliosis drives disease-related processes in a mouse model of Huntington{\textquoteright}s disease (HD). The cellular mechanisms involve astrocyte homeostasis and signaling mediated by Kir4.1, Glt1, and Ca(2+) The data show that the rules for astrocyte engagement in a neuronal circuit are fundamentally altered in a brain disease caused by a known molecular defect and that fixing early homeostasis dysfunction remedies additional cellular deficits. Overall, our data suggest that key aspects of altered striatal function associated with HD may be triggered, at least in part, by dysfunctional astrocytes, thereby providing details of an emerging striatal microcircuit mechanism in HD. Such prodromal changes in astrocytes may represent novel therapeutic targets.

}, issn = {1529-2401}, doi = {10.1523/JNEUROSCI.3693-15.2016}, author = {Jiang, Ruotian and Diaz-Castro, Blanca and Looger, Loren L and Khakh, Baljit S} } @article {47423, title = {Editorial overview: Neurobiology of cognitive behavior: Complexity of neural computation and cognition.}, journal = {Current Opinion in Neurobiology}, volume = {37}, year = {2016}, month = {2016 Mar 14}, pages = {v-viii}, issn = {1873-6882}, doi = {10.1016/j.conb.2016.03.003}, author = {Karpova, Alla and Kiani, Roozbeh} } @article {47386, title = {Emerging imaging and genomic tools for developmental systems biology.}, journal = {Developmental Cell}, volume = {36}, year = {2016}, month = {2016 Mar 21}, pages = {597-610}, abstract = {

Animal development is a complex and dynamic process orchestrated by exquisitely timed cell lineage commitment, divisions, migration, and morphological changes at the single-cell level. In the past decade, extensive genetic, stem cell, and genomic studies provided crucial insights into molecular underpinnings and the functional importance of genetic pathways governing various cellular differentiation processes. However, it is still largely unknown how the precise coordination of these pathways is achieved at the whole-organism level and how the highly regulated spatiotemporal choreography of development is established in turn. Here, we discuss the latest technological advances in imaging and single-cell genomics that hold great promise for advancing our understanding of this intricate process. We propose an integrated approach that combines such methods to quantitatively decipher in\ vivo cellular dynamic behaviors and their underlying molecular mechanisms at the systems level with single-cell, single-molecule resolution.

}, issn = {1878-1551}, doi = {10.1016/j.devcel.2016.02.016}, author = {Liu, Zhe and Keller, Philipp J} } @article {47465, title = {Ensemble cryo-EM uncovers inchworm-like translocation of a viral IRES through the ribosome.}, journal = {eLife}, volume = {5}, year = {2016}, month = {2016 May 9}, abstract = {

Internal ribosome entry sites (IRESs) mediate cap-independent translation of viral mRNAs. Using electron cryo-microscopy of a single specimen, we present five ribosome structures formed with the Taura syndrome virus IRES and translocase eEF2{\textbullet}GTP bound with sordarin. The structures suggest a trajectory of IRES translocation, required for translation initiation, and provide an unprecedented view of eEF2 dynamics. The IRES rearranges from extended to bent to extended conformations. This inchworm-like movement is coupled with ribosomal inter-subunit rotation and 40S head swivel. eEF2, attached to the 60S subunit, slides along the rotating 40S subunit to enter the A site. Its diphthamide-bearing tip at domain IV separates the tRNA-mRNA-like pseudoknot I (PKI) of the IRES from the decoding center. This unlocks 40S domains, facilitating head swivel and biasing IRES translocation via hitherto-elusive intermediates with PKI captured between the A and P sites. The structures suggest missing links in our understanding of tRNA translocation.

}, issn = {2050-084X}, doi = {10.7554/eLife.14874}, author = {Abeyrathne, Priyanka D and Koh, Cha San and Grant, Timothy and Grigorieff, Nikolaus and Korostelev, Andrei A} } @conference {48832, title = {Fast and accurate spike sorting of high-channel count probes with KiloSort.}, booktitle = {Neural Information Processing Systems (NIPS 2016)}, year = {2016}, month = {2016 Dec 05}, address = {Barcelona, Spain}, abstract = {

New silicon technology is enabling large-scale electrophysiological recordings in vivo from hundreds to thousands of channels. Interpreting these recordings requires scalable and accurate automated methods for spike sorting, which should minimize the time required for manual curation of the results. Here we introduce KiloSort, a new integrated spike sorting framework that uses template matching both during spike detection and during spike clustering. KiloSort models the electrical voltage as a sum of template waveforms triggered on the spike times, which allows overlapping spikes to be identified and resolved. Unlike previous algorithms that compress the data with PCA, KiloSort operates on the raw data which allows it to construct a more accurate model of the waveforms. Processing times are faster than in previous algorithms thanks to batch-based optimization on GPUs. We compare KiloSort to an established algorithm and show favorable performance, at much reduced processing times. A novel post-clustering merging step based on the continuity of the templates further reduced substantially the number of manual operations required on this data, for the neurons with near-zero error rates, paving the way for fully automated spike sorting of multichannel electrode recordings.

}, url = {http://papers.nips.cc/paper/6326-fast-and-accurate-spike-sorting-of-high-channel-count-probes-with-kilosort}, author = {Pachitariu, Marius and Steinmetz, Nicholas A and Kadir, Shabnam N and Carandini, Matteo and Harris, Kenneth D} } @article {47726, title = {The genome of the crustacean Parhyale hawaiensis: a model for animal development, regeneration, immunity and lignocellulose digestion.}, journal = {eLife}, volume = {5}, year = {2016}, month = {2016 Nov 16}, pages = {e20062}, abstract = {

Parhyale hawaiensis is a blossoming model system for studies of developmental mechanisms and more recently adult regeneration. We have sequenced the genome allowing annotation of all key signaling pathways, small non-coding RNAs and transcription factors that will enhance ongoing functional studies. Parhayle is a member of the Malacostraca, which includes crustacean food crop species. We analysed the immunity related genes of Parhyale as an important comparative system for these species, where immunity related aquaculture problems have increased as farming has intensified. We also find that Parhyale and other species within Multicrustacea contain the enzyme sets necessary to perform lignocellulose digestion (wood eating), suggesting this ability may predate the diversification of this lineage. Our data provide an essential resource for further development of the Parhyale model. The first Malacostracan genome sequence will underpin ongoing comparative work in important food crop species and research investigating lignocellulose as energy source.

Publication first appeared in BioRxiv on August 2, 2016.\ http://dx.doi.org/10.1101/065789

}, doi = {10.7554/eLife.20062}, author = {Kao, Damian and Lai, Alvina G and Stamataki, Evangelia and Rosic, Silvana and Konstantinides, Nikolaos and Jarvis, Erin and Di Donfrancesco, Alessia and Pouchkina-Stantcheva, Natalia and Semon, Marie and Grillo, Marco and Bruce, Heather and Kumar, Suyash and Siwanowicz, Igor and Le, Andy and Lemire, Andrew and Extavour, Cassandra and Browne, William and Wolff, Carsten and Averof, Michalis and et al} } @article {47348, title = {High-density genotyping of immune-related loci identifies new SLE risk variants in individuals with Asian ancestry.}, journal = {Nature Genetics}, volume = {48}, year = {2016}, month = {2016 Mar}, pages = {323-30}, abstract = {

Systemic lupus erythematosus (SLE) has a strong but incompletely understood genetic architecture. We conducted an association study with replication in 4,478 SLE cases and 12,656 controls from six East Asian cohorts to identify new SLE susceptibility loci and better localize known loci. We identified ten new loci and confirmed 20 known loci with genome-wide significance. Among the new loci, the most significant locus was GTF2IRD1-GTF2I at 7q11.23 (rs73366469, Pmeta = 3.75 {\texttimes} 10(-117), odds ratio (OR) = 2.38), followed by DEF6, IL12B, TCF7, TERT, CD226, PCNXL3, RASGRP1, SYNGR1 and SIGLEC6. We identified the most likely functional variants at each locus by analyzing epigenetic marks and gene expression data. Ten candidate variants are known to alter gene expression in cis or in trans. Enrichment analysis highlights the importance of these loci in B cell and T cell biology. The new loci, together with previously known loci, increase the explained heritability of SLE to 24\%. The new loci share functional and ontological characteristics with previously reported loci and are possible drug targets for SLE therapeutics.

}, issn = {1546-1718}, doi = {10.1038/ng.3496}, author = {Sun, Celi and Molineros, Julio E and Looger, Loren L and Zhou, Xu-Jie and Kim, Kwangwoo and Okada, Yukinori and Ma, Jianyang and Qi, Yuan-Yuan and Kim-Howard, Xana and Motghare, Prasenjeet and Bhattarai, Krishna and Adler, Adam and Bang, So-Young and Lee, Hye-Soon and Kim, Tae-Hwan and Kang, Young Mo and Suh, Chang-Hee and Chung, Won Tae and Park, Yong-Beom and Choe, Jung-Yoon and Shim, Seung Cheol and Kochi, Yuta and Suzuki, Akari and Kubo, Michiaki and Sumida, Takayuki and Yamamoto, Kazuhiko and Lee, Shin-Seok and Kim, Young Jin and Han, Bok-Ghee and Dozmorov, Mikhail and Kaufman, Kenneth M and Wren, Jonathan D and Harley, John B and Shen, Nan and Chua, Kek Heng and Zhang, Hong and Bae, Sang-Cheol and Nath, Swapan K} } @article {47877, title = {Imaging far and wide.}, journal = {eLife}, volume = {5}, year = {2016}, month = {2016 Sep 23}, pages = {e21072}, abstract = {

A custom-built objective lens called the Mesolens allows relatively large biological specimens to be imaged with cellular resolution.

}, doi = {10.7554/eLife.18659}, author = {Chhetri, Raghav K and Keller, Philipp J} } @article {47551, title = {A large field of view two-photon mesoscope with subcellular resolution for in vivo imaging.}, journal = {eLife}, volume = {5}, year = {2016}, month = {2016 Jun 14}, pages = {e14472}, abstract = {

Imaging is used to map activity across populations of neurons. Microscopes with cellular resolution have small (\<1 millimeter) fields of view and cannot simultaneously image activity distributed across multiple brain areas. Typical large field of view microscopes do not resolve single cells, especially in the axial dimension. We developed a 2-photon random access mesoscope (2p-RAM) that allows high-resolution imaging anywhere within a volume spanning multiple brain areas (∅ 5 mm x 1 mm cylinder). 2p-RAM resolution is near diffraction limited (lateral, 0.66 μm, axial 4.09 μm at the center; excitation wavelength = 970 nm; numerical aperture = 0.6) over a large range of excitation wavelengths. A fast three-dimensional scanning system allows efficient sampling of neural activity in arbitrary regions of interest across the entire imaging volume. We illustrate the use of the 2p-RAM by imaging neural activity in multiple, non-contiguous brain areas in transgenic mice expressing protein calcium sensors.

}, issn = {2050-084X}, doi = {10.7554/eLife.14472}, author = {Sofroniew, Nicholas James and Flickinger, Daniel and King, Jonathon and Svoboda, Karel} } @article {47933, title = {Live-cell single-molecule tracking reveals co-recognition of H3K27me3 and DNA targets polycomb Cbx7-PRC1 to chromatin.}, journal = {eLife}, volume = {5}, year = {2016}, month = {2016 Oct 10}, abstract = {

The Polycomb PRC1 plays essential roles in development and disease pathogenesis. Targeting of PRC1 to chromatin is thought to be mediated by the Cbx family proteins (Cbx2/4/6/7/8) binding to histone H3 with a K27me3 modification (H3K27me3). Despite this prevailing view, the molecular mechanisms of targeting remain poorly understood. Here, by combining live-cell single-molecule tracking (SMT) and genetic engineering, we reveal that H3K27me3 contributes significantly to the targeting of Cbx7 and Cbx8 to chromatin, but less to Cbx2, Cbx4, and Cbx6. Genetic disruption of the complex formation of PRC1 facilitates the targeting of Cbx7 to chromatin. Biochemical analyses uncover that the CD and AT-hook-like (ATL) motif of Cbx7 constitute a functional DNA-binding unit. Live-cell SMT of Cbx7 mutants demonstrates that Cbx7 is targeted to chromatin by co-recognizing of H3K27me3 and DNA. Our data suggest a novel hierarchical cooperation mechanism by which histone modifications and DNA coordinate to target chromatin regulatory complexes.

}, issn = {2050-084X}, doi = {10.7554/eLife.17667}, author = {Zhen, Chao Yu and Tatavosian, Roubina and Huynh, Thao Ngoc and Duc, Huy Nguyen and Das, Raibatak and Kokotovic, Marko and Grimm, Jonathan B and Lavis, Luke D and Lee, Jun and Mejia, Frances J and Li, Yang and Yao, Tingting and Ren, Xiaojun} } @article {47565, title = {Macular telangiectasia type 1 managed with long-term aflibercept therapy.}, journal = {Ophthalmic Surgery, Lasers and Imaging Retina}, volume = {47}, year = {2016}, month = {2016 Jun}, pages = {593-5}, abstract = {

A 60-year-old man diagnosed with macular telangiectasia type 1 (MacTel 1) was treated for 3 years with monthly aflibercept (Eylea; Regeneron, Tarrytown, NY) and serially imaged with spectral-domain optical coherence tomography. When administered monthly, aflibercept appeared to have a beneficial effect on macular edema secondary to MacTel 1. Visual acuity preservation despite minimal chronic macular edema could be attributed to the lack of significant photoreceptor disruption.

}, doi = {10.3928/23258160-20160601-14}, author = {Kovach, Jaclyn L and Hess, Harald F and Rosenfeld, Philip J} } @conference {48178, title = {Mapping auto-context decision forests to deep ConvNets for semantic segmentation.}, booktitle = {British Machine Vision Conference}, year = {2016}, month = {2016 Sep 19}, address = {York}, abstract = {

In this paper, we propose a mapping from the Auto-context model to a deep Convolutional Neural Network (ConvNet), bridging the gap be- tween these two models, and helping address the challenge of training ConvNets with limited training data.\ 

}, url = {http://www.bmva.org/bmvc/2016/abstracts.pdf}, author = {Richmond, David L and Kainmueller, Dagmar and Yang, Michael Y and Myers, Eugene W and Rother, Carsten} } @article {47398, title = {Mapping translation {\textquoteright}hot-spots{\textquoteright} in live cells by tracking single molecules of mRNA and ribosomes.}, journal = {eLife}, volume = {5}, year = {2016}, month = {2016}, abstract = {

Messenger RNA localization is important for cell motility by local protein translation. However, while single mRNAs can be imaged and their movements tracked in single cells, it has not yet been possible to determine whether these mRNAs are actively translating. Therefore, we imaged single β-actin mRNAs tagged with MS2 stem loops colocalizing with labeled ribosomes to determine when polysomes formed. A dataset of tracking information consisting of thousands of trajectories per cell demonstrated that mRNAs co-moving with ribosomes have significantly different diffusion properties from non-translating mRNAs that were exposed to translation inhibitors. These data indicate that ribosome load changes mRNA movement and therefore highly translating mRNAs move slower. Importantly, β-actin mRNA near focal adhesions exhibited sub-diffusive corralled movement characteristic of increased translation. This method can identify where ribosomes become engaged for local protein production and how spatial regulation of mRNA-protein interactions mediates cell directionality.

}, issn = {2050-084X}, doi = {10.7554/eLife.10415}, author = {Katz, Zachary B and English, Brian P and Lionnet, Timoth{\'e}e and Yoon, Young J and Monnier, Nilah and Ovryn, Ben and Bathe, Mark and Singer, Robert H} } @article {47790, title = {Membrane dynamics of dividing cells imaged by lattice light-sheet microscopy.}, journal = {Molecular Biology of the Cell}, volume = {27}, year = {2016}, month = {2016 Aug 17}, pages = {3418-35}, abstract = {

Membrane remodeling is an essential part for transfer of components to and from the cell surface and membrane-bound organelles, and for changes in cell shape, particularly critical during cell division. Earlier analyses, based on classical optical live-cell imaging, mostly restricted by technical necessity to the attached bottom surface, showed persistent formation of endocytic clathrin pits and vesicles during mitosis. Taking advantage of the resolution, speed, and non-invasive illumination of the newly developed lattice light sheet fluorescence microscope, we reexamined their assembly dynamics over the entire cell surface and showed that clathrin pits form at a lower rate during late mitosis. Full-cell imaging measurements of cell surface area and volume throughout the cell cycle of single cells in culture and in zebrafish embryos showed that the total surface increased rapidly during the transition from telophase to cytokinesis, whereas cell volume increased slightly in metaphase and remained relatively constant during cytokinesis. These applications demonstrate the advantage of lattice light sheet microscopy and enable a new standard for imaging membrane dynamics in single cells and in multicellular assemblies.

}, issn = {1939-4586}, doi = {10.1091/mbc.E16-03-0164}, author = {Aguet, Fran{\c c}ois and Upadhyayula, Srigokul and Gaudin, Rapha{\"e}l and Chou, Yi-ying and Cocucci, Emanuele and He, Kangmin and Chen, Bi-Chang and Mosaliganti, Kishore and Pasham, Mithun and Skillern, Wesley and Legant, Wesley R and Liu, Tsung-Li and Findlay, Greg and Marino, Eric and Danuser, Gaudenz and Megason, Sean and Betzig, Eric and Kirchhausen, Tom} } @article {47912, title = {Molecularly Defined Subplate Neurons Project Both to Thalamocortical Recipient Layers and Thalamus.}, journal = {Cerebral Cortex (New York, N.Y. : 1991)}, volume = {27}, year = {2016}, month = {2016 Sep 20}, pages = {4759-68}, abstract = {

In mammals, subplate neurons (SPNs) are among the first generated cortical neurons. While most SPNs exist only transiently during development, a number of SPNs persist among adult Layer 6b (L6b). During development, SPNs receive thalamic and intra-cortical input, and primarily project to Layer 4 (L4). SPNs are critical for the anatomical and functional development of thalamocortical connections and also pioneer corticothalamic projections. Since SPNs are heterogeneous, SPN subpopulations might serve different roles. Here, we investigate the connectivity of one subpopulation, complexin-3 (Cplx3)-positive SPNs (Cplx3-SPNs), in mouse whisker somatosensory (barrel) cortex (S1). We find that many Cplx3-SPNs survive into adulthood and become a subpopulation of L6b. Cplx3-SPNs axons project to thalamorecipient layers, that is, L4, 5a, and 1. The L4 projections are biased towards the septal regions between barrels in the second postnatal week. Thus, S1 Cplx3-SPN targets co-localize with the eventual projections of the medial posterior thalamic nucleus (POm). In addition to their cortical targets, Cplx3-SPNs also extend long-range axons to several thalamic nuclei, including POm. Thus, Cplx3-SPN/L6b neurons are associated with paralemniscal pathways and can potentially directly link thalamocortical and corticothalamic circuits. This suggests an additional key role for SPNs in the establishment and maintenance of thalamocortical processing.

}, issn = {1460-2199}, doi = {10.1093/cercor/bhw271}, author = {Viswanathan, Sarada and Sheikh, Aminah and Looger, Loren L and Kanold, Patrick O} } @article {47973, title = {Motion quantification during multi-photon functional imaging in behaving animals.}, journal = {Biomedical Optics Express}, volume = {7}, year = {2016}, month = {2016 Sep 1}, pages = {3686-3695}, abstract = {

Functional imaging in behaving animals is essential to understanding brain function. However, artifacts resulting from animal motion, including locomotion, can severely corrupt functional measurements. To dampen tissue motion, we designed a new optical window with minimal optical aberrations. Using the newly developed high-speed continuous volumetric imaging system based on an optical phase-locked ultrasound lens, we quantified motion of the cerebral cortex and hippocampal surface during two-photon functional imaging in behaving mice. We find that the out-of-plane motion is generally greater than the axial dimension of the point-spread-function during mouse locomotion, which indicates that high-speed continuous volumetric imaging is necessary to minimize motion artifacts.

}, doi = {10.1364/BOE.7.003686}, author = {Kong, Lingjie and Little, Justin P and Cui, Meng} } @article {47424, title = {Movement and structure of mitochondria in oligodendrocytes and their myelin sheaths.}, journal = {Glia}, volume = {64}, year = {2016}, month = {2016 May}, pages = {810-25}, abstract = {

Mitochondria play several crucial roles in the life of oligodendrocytes. During development of the myelin sheath they are essential providers of carbon skeletons and energy for lipid synthesis. During normal brain function their consumption of pyruvate will be a key determinant of how much lactate is available for oligodendrocytes to export to power axonal function. Finally, during calcium-overload induced pathology, as occurs in ischemia, mitochondria may buffer calcium or induce apoptosis. Despite their important functions, very little is known of the properties of oligodendrocyte mitochondria, and mitochondria have never been observed in the myelin sheaths. We have now used targeted expression of fluorescent mitochondrial markers to characterize the location and movement of mitochondria within oligodendrocytes. We show for the first time that mitochondria are able to enter and move within the myelin sheath. Within the myelin sheath the highest number of mitochondria was in the cytoplasmic ridges along the sheath. Mitochondria moved more slowly than in neurons and, in contrast to their behavior in neurons and astrocytes, their movement was increased rather than inhibited by glutamate activating NMDA receptors. By electron microscopy we show that myelin sheath mitochondria have a low surface area of cristae, which suggests a low ATP production. These data specify fundamental properties of the oxidative phosphorylation system in oligodendrocytes, the glial cells that enhance cognition by speeding action potential propagation and provide metabolic support to axons. GLIA 2016;64:810-825.

}, issn = {1098-1136}, doi = {10.1002/glia.22965}, author = {Rinholm, Johanne E and Vervaeke, Koen and Tadross, Michael R and Tkachuk, Ariana N and Kopek, Benjamin G and Brown, Timothy A and Bergersen, Linda H and Clayton, David A} } @article {47564, title = {Neural circuits that drive startle behavior, with a focus on the Mauthner cells and spiral fiber neurons of fishes.}, journal = {Journal of Neurogenetics}, volume = {30}, year = {2016}, month = {2016 Jun}, pages = {89-100}, abstract = {

Startle behaviors are rapid, high-performance motor responses to threatening stimuli. Startle responses have been identified in a broad range of species across animal diversity. For investigations of neural circuit structure and function, these behaviors offer a number of benefits, including that they are driven by large and identifiable neurons and their neural control is simple in comparison to other behaviors. Among vertebrates, the best-known startle circuit is the Mauthner cell circuit of fishes. In recent years, genetic approaches in zebrafish have provided key tools for morphological and physiological dissection of circuits and greatly extended understanding of their architecture. Here we discuss the startle circuit of fishes, with a focus on the Mauthner cells and associated interneurons called spiral fiber neurons and we add new observations on hindbrain circuit organization. We also briefly review and compare startle circuits of several other taxa, paying particular attention to how movement direction is controlled.

}, issn = {1563-5260}, doi = {10.1080/01677063.2016.1182526}, author = {Hale, Melina E and Katz, Hilary R and Peek, Martin Y and Fremont, Rachel T} } @article {47936, title = {The Oscillating Stimulus Transporter Assay, OSTA: Quantitative functional imaging of transporter protein activity in time and frequency domains.}, journal = {Molecular Cell}, volume = {64}, year = {2016}, month = {2016 Oct 6}, pages = {199-212}, abstract = {

Transmembrane transporter proteins allow the passage of essentially all biologically important molecules across the lipid membranes of cells and organelles and are therefore of central importance to all forms of life. Current methods of transporter measurement, however, are lacking in several dimensions. Herein, a method is presented in which oscillating stimuli are presented to transporter-expressing cells, and activity is measured through imaging the corresponding oscillating responses of intracellular fluorescent sensors. This approach yields continuous temporal readouts of transporter activity and can therefore be used to measure time-dependent responses to drugs and other stimuli. Because of the periodic nature of the response, temporal Fourier transforms can be used to identify and quantify regions of interest in the xy plane and to overcome noise. This technique, called the Oscillating Stimulus Transporter Assay (OSTA), should greatly facilitate both functional characterization of transporters as well as high-throughput screening of drugs for transporters of particular pathophysiological interest.

}, issn = {1097-4164}, doi = {10.1016/j.molcel.2016.09.001}, author = {Keller, Jacob P and Looger, Loren L} } @article {48118, title = {A plasma membrane template for macropinocytic cups.}, journal = {eLife}, volume = {5}, year = {2016}, month = {2016 Dec 13}, abstract = {

Macropinocytosis is a fundamental mechanism that allows cells to take up extracellular liquid into large vesicles. It critically depends on the formation of a ring of protrusive actin beneath the plasma membrane, which develops into the macropinocytic cup. We show that macropinocytic cups in Dictyostelium are organised around coincident intense patches of PIP3, active Ras and active Rac. These signalling patches are invariably associated with a ring of active SCAR/WAVE at their periphery, as are all examined structures based on PIP3 patches, including phagocytic cups and basal waves. Patch formation does not depend on the enclosing F-actin ring, and patches become enlarged when the RasGAP NF1 is mutated, showing that Ras plays an instructive role. New macropinocytic cups predominantly form by splitting from existing ones. We propose that cup-shaped plasma membrane structures form from self-organizing patches of active Ras/PIP3, which recruit a ring of actin nucleators to their periphery.

}, issn = {2050-084X}, doi = {10.7554/eLife.20085}, author = {Veltman, Douwe M and Williams, Thomas D and Bloomfield, Gareth and Chen, Bi-Chang and Betzig, Eric and Insall, Robert H and Kay, Robert R} } @article {47382, title = {Practice makes perfect in memory recall.}, journal = {Learning \& Memory (Cold Spring Harbor, N.Y.)}, volume = {23}, year = {2016}, month = {2016 Apr}, pages = {169-73}, abstract = {

A large variability in performance is observed when participants recall briefly presented lists of words. The sources of such variability are not known. Our analysis of a large data set of free recall revealed a small fraction of participants that reached an extremely high performance, including many trials with the recall of complete lists. Moreover, some of them developed a number of consistent input-position-dependent recall strategies, in particular recalling words consecutively ("chaining") or in groups of consecutively presented words ("chunking"). The time course of acquisition and particular choice of positional grouping were variable among participants. Our results show that acquiring positional strategies plays a crucial role in improvement of recall performance.

}, issn = {1549-5485}, doi = {10.1101/lm.041178.115}, author = {Romani, Sandro and Katkov, Mikhail and Tsodyks, Misha} } @article {47062, title = {Quantitative neuroanatomy for connectomics in Drosophila.}, journal = {eLife}, year = {2016}, month = {2016 Mar 18}, pages = {e12059}, abstract = {

Neuronal circuit mapping using electron microscopy demands laborious proofreading or reconciliation of multiple independent reconstructions. Here, we describe new methods to apply quantitative arbor and network context to iteratively proofread and reconstruct circuits and create anatomically enriched wiring diagrams. We measured the morphological underpinnings of connectivity in new and existing reconstructions of\ Drosophila\ sensorimotor (larva) and visual (adult) systems. Synaptic inputs were preferentially located on numerous small, microtubule-free {\textquoteright}twigs{\textquoteright} which branch off a single microtubule-containing {\textquoteright}backbone{\textquoteright}. Omission of individual twigs accounted for 96\% of errors. However, the synapses of highly connected neurons were distributed across multiple twigs. Thus, the robustness of a strong connection to detailed twig anatomy was associated with robustness to reconstruction error. By comparing iterative reconstruction to the consensus of multiple reconstructions, we show that our method overcomes the need for redundant effort through the discovery and application of relationships between cellular neuroanatomy and synaptic connectivity.

}, doi = {10.7554/eLife.12059}, author = {Schneider-Mizell, Casey M. and Gerhard, Stephan and Longair, Mark and Kazimiers, Tom and Li, Feng and Zwart, Maarten and Champion, Andrew and Midgley, Frank and Fetter, Richard D and Saalfeld, Stephan and Cardona, Albert} } @article {47938, title = {Rapid dynamics of general transcription factor TFIIB binding during preinitiation complex assembly revealed by single-molecule analysis.}, journal = {Genes and Development}, volume = {30}, year = {2016}, month = {2016 Sep 15}, pages = {2106-18}, abstract = {

Transcription of protein-encoding genes in eukaryotic cells requires the coordinated action of multiple general transcription factors (GTFs) and RNA polymerase II (Pol II). A {\textquotedblleft}step-wise{\textquotedblright} preinitiation complex (PIC) assembly model has been suggested based on conventional ensemble biochemical measurements, in which protein factors bind stably to the promoter DNA sequentially to build a functional PIC. However, recent dynamic measurements in live cells suggest that transcription factors mostly interact with chromatin DNA rather transiently. To gain a clearer dynamic picture of PIC assembly, we established an integrated in vitro single-molecule transcription platform reconstituted from highly purified human transcription factors and complemented it by live-cell imaging. Here we performed real-time measurements of the hierarchal promoter-specific binding of TFIID, TFIIA, and TFIIB. Surprisingly, we found that while promoter binding of TFIID and TFIIA is stable, promoter binding by TFIIB is highly transient and dynamic (with an average residence time of 1.5 sec). Stable TFIIB{\textendash}promoter association and progression beyond this apparent PIC assembly checkpoint control occurs only in the presence of Pol II{\textendash}TFIIF. This transient-to-stable transition of TFIIB-binding dynamics has gone undetected previously and underscores the advantages of single-molecule assays for revealing the dynamic nature of complex biological reactions.

}, doi = {10.1101/gad.285395.116}, author = {Zhang, Zhengjian and English, Brian P and Grimm, Jonathan B and Kazane, Stephanie A and Hu, Wenxin and Tsai, Albert and Inouye, Carla and You, Changjiang and Piehler, Jacob and Schultz, Peter G and Lavis, Luke D and Revyakin, Andrey and Tjian, Robert} } @article {47267, title = {Real-time three-dimensional cell segmentation in large-scale microscopy data of developing embryos.}, journal = {Developmental Cell}, volume = {36}, year = {2016}, month = {2016 Jan 25}, pages = {225-40}, abstract = {

We present the Real-time Accurate Cell-shape Extractor (RACE), a high-throughput image analysis framework for automated three-dimensional cell segmentation in large-scale images. RACE is 55{\textendash}330 times faster and 2{\textendash}5 times more accurate than state-of-the-art methods. We demonstrate the generality of RACE by extracting cell-shape information from entire Drosophila, zebrafish, and mouse embryos imaged with confocal and light-sheet microscopes. Using RACE, we automatically reconstructed cellular-resolution tissue anisotropy maps across developing Drosophila embryos and quantified differences in cell-shape dynamics in wild-type and mutant embryos. We furthermore integrated RACE with our framework for automated cell lineaging and performed joint segmentation and cell tracking in entire Drosophila embryos. RACE processed these terabyte-sized datasets on a single computer within 1.4\ days. RACE is easy to use, as it requires adjustment of only three parameters, takes full advantage of state-of-the-art multi-core processors and graphics cards, and is available as open-source software for Windows, Linux, and Mac\ OS.

}, doi = {10.1016/j.devcel.2015.12.028}, author = {Stegmaier, Johannes and Amat, Fernando and Lemon, William C and McDole, Katie and Wan, Yinan and Teodoro, George and Mikut, Ralf and Keller, Philipp J} } @article {47440, title = {Repulsive cues combined with physical barriers and cell{\textendash}cell adhesion determine progenitor cell positioning during organogenesis}, journal = {Nature Communications}, volume = {7}, year = {2016}, month = {2016 Apr 18}, pages = {11288}, abstract = {

The precise positioning of organ progenitor cells constitutes an essential, yet poorly understood step during organogenesis. Using primordial germ cells that participate in gonad formation, we present the developmental mechanisms maintaining a motile progenitor cell population at the site where the organ develops. Employing high-resolution live-cell microscopy, we find that repulsive cues coupled with physical barriers confine the cells to the correct bilateral positions. This analysis revealed that cell polarity changes on interaction with the physical barrier and that the establishment of compact clusters involves increased cell-cell interaction time. Using particle-based simulations, we demonstrate the role of reflecting barriers, from which cells turn away on contact, and the importance of proper cell-cell adhesion level for maintaining the tight cell clusters and their correct positioning at the target region. The combination of these developmental and cellular mechanisms prevents organ fusion, controls organ positioning and is thus critical for its proper function.

}, doi = {10.1038/ncomms11288}, author = {Paksa, Azadeh and Bandemer, Jan and H{\"o}ckendorf, Burkhard and Razin, Nitzan and Tarbashevich, Katsiaryna and Minina, Sofia and Meyen, Dana and Gov, Nir S and Keller, Philipp J and Raz, Erez} } @article {65009, title = {Ribosome{\textbullet}RelA structures reveal the mechanism of stringent response activation.}, journal = {Elife}, volume = {5}, year = {2016}, month = {2016 07 19}, abstract = {

Stringent response is a conserved bacterial stress response underlying virulence and antibiotic resistance. RelA/SpoT-homolog proteins synthesize transcriptional modulators (p)ppGpp, allowing bacteria to adapt to stress. RelA is activated during amino-acid starvation, when cognate deacyl-tRNA binds to the ribosomal A (aminoacyl-tRNA) site. We report four cryo-EM structures of E. coli RelA bound to the 70S ribosome, in the absence and presence of deacyl-tRNA accommodating in the 30S A site. The boomerang-shaped RelA with a wingspan of more than 100 {\r A} wraps around the A/R (30S A-site/RelA-bound) tRNA. The CCA end of the A/R tRNA pins the central TGS domain against the 30S subunit, presenting the (p)ppGpp-synthetase domain near the 30S spur. The ribosome and A/R tRNA are captured in three conformations, revealing hitherto elusive states of tRNA engagement with the ribosomal decoding center. Decoding-center rearrangements are coupled with the step-wise 30S-subunit {\textquoteright}closure{\textquoteright}, providing insights into the dynamics of high-fidelity tRNA decoding.

}, keywords = {Cryoelectron Microscopy, Escherichia coli, Ligases, Protein Binding, Ribosomes, RNA, Transfer, Stress, Physiological}, issn = {2050-084X}, doi = {10.7554/eLife.17029}, author = {Loveland, Anna B and Bah, Eugene and Madireddy, Rohini and Zhang, Ying and Brilot, Axel F and Grigorieff, Nikolaus and Korostelev, Andrei A} } @article {48069, title = {RNA-binding profiles of Drosophila CPEB proteins Orb and Orb2.}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, year = {2016}, month = {2016 Oct 24}, abstract = {

Localized protein translation is critical in many biological contexts, particularly in highly polarized cells, such as neurons, to regulate gene expression in a spatiotemporal manner. The cytoplasmic polyadenylation element-binding (CPEB) family of RNA-binding proteins has emerged as a key regulator of mRNA transport and local translation required for early embryonic development, synaptic plasticity, and long-term memory (LTM). Drosophila Orb and Orb2 are single members of the CPEB1 and CPEB2 subfamilies of the CPEB proteins, respectively. At present, the identity of the mRNA targets they regulate is not fully known, and the binding specificity of the CPEB2 subfamily is a matter of debate. Using transcriptome-wide UV cross-linking and immunoprecipitation, we define the mRNA-binding sites and targets of Drosophila CPEBs. Both Orb and Orb2 bind linear cytoplasmic polyadenylation element-like sequences in the 3{\textquoteright} UTRs of largely overlapping target mRNAs, with Orb2 potentially having a broader specificity. Both proteins use their RNA-recognition motifs but not the Zinc-finger region for RNA binding. A subset of Orb2 targets is translationally regulated in cultured S2 cells and fly head extracts. Moreover, pan-neuronal RNAi knockdown of these targets suggests that a number of these targets are involved in LTM. Our results provide a comprehensive list of mRNA targets of the two CPEB proteins in Drosophila, thus providing insights into local protein synthesis involved in various biological processes, including LTM.

}, issn = {1091-6490}, doi = {10.1073/pnas.1603715113}, author = {Stepien, Barbara Krystyna and Oppitz, Cornelia and Gerlach, Daniel and Dag, Ugur and Novatchkova, Maria and Kr{\"u}ttner, Sebastian and Stark, Alexander and Keleman, Krystyna} } @article {47957, title = {Sampling Requirements for Stable Autoregressive Estimation}, journal = {CoRR}, volume = {abs/1605.01436}, year = {2016}, abstract = {

We consider the problem of estimating the parameters of a linear autoregressive model with sub-Gaussian innovations from a limited sequence of consecutive observations. Assuming that the parameters are compressible, we analyze the performance of the\ l1-regularized least squares as well as a greedy estimator of the parameters and characterize the sampling trade-offs required for stable recovery in the non-asymptotic regime. Our results extend those of compressed sensing for linear models where the covariates are i.i.d. and independent of the observation history to autoregressive processes with highly inter-dependent covariates. We also derive sufficient conditions on the sparsity level that guarantee the minimax optimality of the\ l1-regularized least squares estimate. Applying these techniques to simulated data as well as real-world datasets from crude oil prices and traffic speed data confirm our predicted theoretical performance gains in terms of estimation accuracy and model selection.

}, url = {http://arxiv.org/abs/1605.01436}, author = {Abbas Kazemipour and Sina Miran and Piya Pal and Behtash Babadi and Min Wu} } @article {47767, title = {Satb2 stations neurons along reflex arcs.}, journal = {Neuron}, volume = {91}, year = {2016}, month = {2016 Aug 17}, pages = {711-3}, abstract = {

The nociceptive flexor withdrawal reflex has an august place in the history of neuroscience. In this issue of\ Neuron, Hilde et\ al. (2016) advance our understanding of this reflex by characterizing the molecular identity and circuit connectivity of component interneurons. They assess how a DNA-binding factor Satb2 controls\ cell position, molecular identity, pre-and postsynaptic targeting, and function of a population of inhibitory sensory relay interneurons that serve to integrate both proprioceptive and nociceptive afferent information.

The nociceptive flexor withdrawal reflex has an august place in the history of neuroscience. In this issue of\ Neuron, Hilde et\ al. (2016) advance our understanding of this reflex by characterizing the molecular identity and circuit connectivity of component interneurons. They assess how a DNA-binding factor Satb2 controls cell position, molecular identity, pre-and postsynaptic targeting, and function of a population of inhibitory sensory relay interneurons that serve to integrate both proprioceptive and nociceptive afferent information.

}, doi = {10.1016/j.neuron.2016.08.005}, author = {Hantman, Adam W and Kaltschmidt, Julia A} } @article {47538, title = {A screen for constituents of motor control and decision making in Drosophila reveals visual distance-estimation neurons.}, journal = {Scientific Reports}, volume = {6}, year = {2016}, month = {2016}, pages = {27000}, abstract = {

Climbing over chasms larger than step size is vital to fruit flies, since foraging and mating are achieved while walking. Flies avoid futile climbing attempts by processing parallax-motion vision to estimate gap width. To identify neuronal substrates of climbing control, we screened a large collection of fly lines with temporarily inactivated neuronal populations in a novel high-throughput assay described here. The observed climbing phenotypes were classified; lines in each group are reported. Selected lines were further analysed by high-resolution video cinematography. One striking class of flies attempts to climb chasms of unsurmountable width; expression analysis guided us to C2 optic-lobe interneurons. Inactivation of C2 or the closely related C3 neurons with highly specific intersectional driver lines consistently reproduced hyperactive climbing whereas strong or weak artificial depolarization of C2/C3 neurons strongly or mildly decreased climbing frequency. Contrast-manipulation experiments support our conclusion that C2/C3 neurons are part of the distance-evaluation system.

}, issn = {2045-2322}, doi = {10.1038/srep27000}, author = {Triphan, Tilman and Nern, Aljoscha and Roberts, Sonia F and Korff, Wyatt and Naiman, Daniel Q and Strauss, Roland} } @article {47511, title = {Segmenting and Tracking Multiple Dividing Targets Using ilastik.}, journal = {Advances in anatomy, embryology, and cell biology}, volume = {219}, year = {2016}, month = {2016 May 21}, pages = {199-229}, abstract = {

Tracking crowded cells or other targets in biology is often a challenging task due to poor signal-to-noise ratio, mutual occlusion, large displacements, little discernibility, and the ability of cells to divide. We here present an open source implementation of conservation tracking (Schiegg et\ al., IEEE international conference on computer vision (ICCV). IEEE, New York, pp\ 2928-2935, 2013) in the ilastik software framework. This robust tracking-by-assignment algorithm explicitly makes allowance for false positive detections, undersegmentation, and cell division. We give an overview over the underlying algorithm and parameters, and explain the use for a light sheet microscopy sequence of a Drosophila embryo. Equipped with this knowledge, users will be able to track targets of interest in their own data.

}, issn = {0301-5556}, doi = {10.1007/978-3-319-28549-8_8}, author = {Haubold, Carsten and Schiegg, Martin and Kreshuk, Anna and Berg, Stuart and Koethe, Ullrich and Hamprecht, Fred A} } @article {47396, title = {Sensitive red protein calcium indicators for imaging neural activity.}, journal = {eLife}, volume = {5}, year = {2016}, month = {2016 Mar 24}, abstract = {

Genetically encoded calcium indicators (GECIs) allow measurement of activity in large populations of neurons and in small neuronal compartments, over times of milliseconds to months. Although GFP-based GECIs are widely used for in vivo neurophysiology, GECIs with red-shifted excitation and emission spectra have advantages for in vivo imaging because of reduced scattering and absorption in tissue, and a consequent reduction in phototoxicity. However, current red GECIs are inferior to the state-of-the-art GFP-based GCaMP6 indicators for detecting and quantifying neural activity. Here we present improved red GECIs based on mRuby (jRCaMP1a, b) and mApple (jRGECO1a), with sensitivity comparable to GCaMP6. We characterized the performance of the new red GECIs in cultured neurons and in mouse, Drosophila, zebrafish and C. elegans in vivo. Red GECIs facilitate deep-tissue imaging, dual-color imaging together with GFP-based reporters, and the use of optogenetics in combination with calcium imaging.

}, issn = {2050-084X}, doi = {10.7554/eLife.12727}, author = {Dana, Hod and Mohar, Boaz and Sun, Yi and Narayan, Sujatha and Gordus, Andrew and Hasseman, Jeremy P and Tsegaye, Getahun and Holt, Graham T and Hu, Amy and Walpita, Deepika and Patel, Ronak and Macklin, John J and Bargmann, Cornelia I and Ahrens, Misha B and Schreiter, Eric R and Jayaraman, Vivek and Looger, Loren L and Svoboda, Karel and Kim, Douglas S} } @article {47991, title = {The serotonergic system tracks the outcomes of actions to mediate short-term motor learning.}, journal = {Cell}, volume = {167}, year = {2016}, month = {2016 Oct 27}, pages = {933-46}, abstract = {

To execute accurate movements, animals must continuously adapt their behavior to changes in their\ bodies and environments. Animals can learn changes in the relationship between their locomotor commands and the resulting distance moved, then adjust command strength to achieve a desired travel distance. It is largely unknown which circuits implement this form of motor learning, or how. Using whole-brain neuronal imaging and circuit manipulations in larval zebrafish, we discovered that the serotonergic dorsal raphe nucleus (DRN) mediates short-term locomotor learning. Serotonergic DRN neurons respond phasically to swim-induced visual motion, but little to motion that is not self-generated. During prolonged exposure to a given motosensory gain, persistent DRN activity emerges that stores the learned efficacy of motor commands and adapts future locomotor drive for tens of seconds. The DRN{\textquoteright}s ability to track the effectiveness of motor intent may constitute a computational building block\ for the broader functions of the serotonergic system.

}, doi = {10.1016/j.cell.2016.09.055}, author = {Kawashima, Takashi and Zwart, Maarten F and Yang, Chao-Tsung and Mensh, Brett D and Ahrens, Misha B} } @article {47420, title = {Steroid receptors reprogram FoxA1 occupancy through dynamic chromatin transitions.}, journal = {Cell}, year = {2016}, month = {2016 Apr 6}, abstract = {

The estrogen receptor (ER), glucocorticoid receptor (GR), and forkhead box protein 1 (FoxA1) are significant factors in breast cancer progression. FoxA1 has been implicated in establishing ER-binding patterns though its unique ability to serve as a pioneer factor. However, the molecular interplay between ER, GR, and FoxA1 requires further investigation. Here we show that ER and GR both have the ability to alter the genomic distribution of the FoxA1 pioneer factor. Single-molecule tracking experiments in live cells reveal a highly dynamic interaction of FoxA1 with chromatin in\ vivo. Furthermore, the FoxA1 factor is not associated with detectable footprints at its binding sites throughout the genome. These findings support a model wherein interactions between transcription factors and pioneer factors are highly dynamic. Moreover, at a subset of genomic sites, the role of pioneer can be reversed, with the steroid receptors serving to enhance binding of FoxA1.

}, issn = {1097-4172}, doi = {10.1016/j.cell.2016.02.067}, author = {Swinstead, Erin E and Miranda, Tina B and Paakinaho, Ville and Baek, Songjoon and Goldstein, Ido and Hawkins, Mary and Karpova, Tatiana S and Ball, David and Mazza, Davide and Lavis, Luke D and Grimm, Jonathan B and Morisaki, Tatsuya and Gr{\o}ntved, Lars and Presman, Diego M and Hager, Gordon L} } @article {47322, title = {Structured dendritic inhibition supports branch-selective integration in CA1 pyramidal cells.}, journal = {Neuron}, year = {2016}, month = {2016 Feb 18}, abstract = {

Neuronal circuit function is governed by precise patterns of connectivity between specialized groups of neurons. The diversity of GABAergic interneurons is a hallmark of cortical circuits, yet little is known about their targeting to individual postsynaptic dendrites. We examined synaptic connectivity between molecularly defined inhibitory interneurons and CA1 pyramidal cell dendrites using correlative light-electron microscopy and large-volume array tomography. We show that interneurons can be highly selective in their connectivity to specific dendritic branch types and, furthermore, exhibit precisely targeted connectivity to the origin or end of individual branches. Computational simulations indicate that the observed subcellular targeting enables control over the nonlinear integration of synaptic input or the initiation and backpropagation of action potentials in a branch-selective manner. Our results demonstrate that connectivity between interneurons and pyramidal cell dendrites is more precise and spatially segregated than previously appreciated, which may be a critical determinant of how inhibition shapes dendritic computation.

}, issn = {1097-4199}, doi = {10.1016/j.neuron.2016.01.029}, author = {Bloss, Erik B and Cembrowski, Mark S and Karsh, Bill and Colonell, Jennifer and Fetter, Richard D and Spruston, Nelson} } @article {47560, title = {Structured learning of assignment models for neuron reconstruction to minimize topological errors.}, journal = {IEEE 13th International Symposium on Biomedical Imaging (ISBI)}, year = {2016}, month = {2016 Ap 13}, pages = {607-11}, abstract = {

Structured learning provides a powerful framework for empirical risk minimization on the predictions of structured models. It allows end-to-end learning of model parameters to minimize an application specific loss function. This framework is particularly well suited for discrete optimization models that are used for neuron reconstruction from anisotropic electron microscopy (EM) volumes. However, current methods are still learning unary potentials by training a classifier that is agnostic about the model it is used in. We believe the reason for that lies in the difficulties of (1) finding a representative training sample, and (2) designing an application specific loss function that captures the quality of a proposed solution. In this paper, we show how to find a representative training sample from human generated ground truth, and propose a loss function that is suitable to minimize topological errors in the reconstruction. We compare different training methods on two challenging EM-datasets. Our structured learning approach shows consistently higher reconstruction accuracy than other current learning methods.

}, doi = {10.1109/ ISBI.2016.7493341}, author = {Funke, Jan and Klein, Jonas and Moreno-Noguer, Francesc and Cardona, Albert and Cook, Matthew} } @article {47709, title = {Super-resolution spectroscopic microscopy via photon localization.}, journal = {Nature Communications}, volume = {7}, year = {2016}, month = {2016 Jul 25}, pages = {12290}, abstract = {

Traditional photon localization microscopy analyses only the spatial distributions of photons emitted by individual molecules to reconstruct super-resolution optical images. Unfortunately, however, the highly valuable spectroscopic information from these photons have been overlooked. Here we report a spectroscopic photon localization microscopy that is capable of capturing the inherent spectroscopic signatures of photons from individual stochastic radiation events. Spectroscopic photon localization microscopy achieved higher spatial resolution than traditional photon localization microscopy through spectral discrimination to identify the photons emitted from individual molecules. As a result, we resolved two fluorescent molecules, which were 15 nm apart, with the corresponding spatial resolution of 10 nm-a four-fold improvement over photon localization microscopy. Using spectroscopic photon localization microscopy, we further demonstrated simultaneous multi-colour super-resolution imaging of microtubules and mitochondria in COS-7 cells and showed that background autofluorescence can be identified through its distinct emission spectra.

}, issn = {2041-1723}, doi = {10.1038/ncomms12290}, author = {Dong, Biqin and Almassalha, Luay and Urban, Ben E and Nguyen, The-Quyen and Khuon, Satya and Chew, Teng-Leong and Backman, Vadim and Sun, Cheng and Zhang, Hao F} } @article {47972, title = {An unsupervised method for quantifying the behavior of interacting individuals.}, journal = {Physical Biology}, volume = {14}, year = {2016}, month = {2017 Feb16}, pages = {1609.09345}, abstract = {

Behaviors involving the interaction of multiple individuals are complex and frequently crucial for an animal{\textquoteright}s survival. These interactions, ranging across sensory modalities, length scales, and time scales, are often subtle and difficult to characterize. Contextual effects on the frequency of behaviors become even more difficult to quantify when physical interaction between animals interferes with conventional data analysis, e.g. due to visual occlusion. We introduce a method for quantifying behavior in fruit fly interaction that combines high-throughput video acquisition and tracking of individuals with recent unsupervised methods for capturing an animal{\textquoteright}s entire behavioral repertoire. We find behavioral differences between solitary flies and those paired with an individual of the opposite sex, identifying specific behaviors that are affected by social and spatial context. Our pipeline allows for a comprehensive description of the interaction between two individuals using unsupervised machine learning methods, and will be used to answer questions about the depth of complexity and variance in fruit fly courtship.

}, doi = {10.1088/1478-3975/aa5c50}, url = {https://arxiv.org/abs/1609.09345v1}, author = {Klibaite, Ugne and Berman, Gordon J and Cande, Jessica and Stern, David L} } @article {47814, title = {Whole-animal imaging with high spatio-temporal resolution.}, journal = {Proceedings of SPIE}, volume = {9720}, year = {2016}, month = {2016 Jul 18}, pages = {97200R}, abstract = {

We developed isotropic multiview (IsoView) light-sheet microscopy in order to image fast cellular dynamics, such as cell movements in an entire developing embryo or neuronal activity throughput an entire brain or nervous system, with high resolution in all dimensions, high imaging speeds, good physical coverage and low photo-damage. To achieve high temporal resolution and high spatial resolution at the same time, IsoView microscopy rapidly images large specimens via simultaneous light-sheet illumination and fluorescence detection along four orthogonal directions. In a post-processing step, these four views are then combined by means of high-throughput multiview deconvolution to yield images with a system resolution of <= 450 nm in all three dimensions. Using IsoView microscopy, we performed whole-animal functional imaging of Drosophila embryos and larvae at a spatial resolution of 1.1-2.5 μm and at a temporal resolution of 2 Hz for up to 9 hours. We also performed whole-brain functional imaging in larval zebrafish and multicolor imaging of fast cellular dynamics across entire, gastrulating Drosophila embryos with isotropic, sub-cellular resolution. Compared with conventional (spatially anisotropic) light-sheet microscopy, IsoView microscopy improves spatial resolution at least sevenfold and decreases resolution anisotropy at least threefold. Compared with existing high-resolution light-sheet techniques, such as lattice lightsheet microscopy or diSPIM, IsoView microscopy effectively doubles the penetration depth and provides subsecond temporal resolution for specimens 400-fold larger than could previously be imaged.

}, isbn = {978-162841954-2}, issn = { 0277786X}, doi = {10.1117/12.2212564}, author = {Chhetri, Raghav and Amat, Fernando and Wan, Yinan and H{\"o}ckendorf, Burkhard and Lemon, William C and Keller, Philipp J} } @article {47486, title = {The wiring diagram of a glomerular olfactory system.}, journal = {eLife}, volume = {5}, year = {2016}, month = {2016 May 13}, abstract = {

The sense of smell enables animals to react to long-distance cues according to learned and innate valences. Here, we have mapped with electron microscopy the complete wiring diagram of the Drosophila larval antennal lobe, an olfactory neuropil similar to the vertebrate olfactory bulb. We found a canonical circuit with uniglomerular projection neurons (uPNs) relaying gain-controlled ORN activity to the mushroom body and the lateral horn. A second, parallel circuit with multiglomerular projection neurons (mPNs) and hierarchically connected local neurons (LNs) selectively integrates multiple ORN signals already at the first synapse. LN-LN synaptic connections putatively implement a bistable gain control mechanism that either computes odor saliency through panglomerular inhibition, or allows some glomeruli to respond to faint aversive odors in the presence of strong appetitive odors. This complete wiring diagram will support experimental and theoretical studies towards bridging the gap between circuits and behavior.

}, issn = {2050-084X}, doi = {10.7554/eLife.14859}, author = {Berck, Matthew E and Khandelwal, Avinash and Claus, Lindsey and Hernandez-Nunez, Luis and Si, Guangwei and Tabone, Christopher J and Li, Feng and Truman, James W and Fetter, Richard D and Louis, Matthieu and Samuel, Aravinthan Dt and Cardona, Albert} } @article {46525, title = {Age-dependent changes in intrinsic neuronal excitability in subiculum after status epilepticus.}, journal = {PLoS One}, volume = {10}, year = {2015}, month = {2015 Mar 16}, pages = {e0119411}, abstract = {

Kainic acid-induced status epilepticus (KA-SE) in mature rats results in the development of spontaneous recurrent seizures and a pattern of cell death resembling hippocampal sclerosis in patients with temporal lobe epilepsy. In contrast, KA-SE in young animals before postnatal day (P) 18 is less likely to cause cell death or epilepsy. To investigate whether changes in neuronal excitability occur in the subiculum after KA-SE, we examined the age-dependent effects of SE on the bursting neurons of subiculum, the major output region of the hippocampus. Patch-clamp recordings were used to monitor bursting in pyramidal neurons in the subiculum of rat hippocampal slices. Neurons were studied either one or 2-3 weeks following injection of KA or saline (control) in immature (P15) or more mature (P30) rats, which differ in their sensitivity to KA as well as the long-term sequelae of the KA-SE. A significantly greater proportion of subicular pyramidal neurons from P15 rats were strong-bursting neurons and showed increased frequency-dependent bursting compared to P30 animals. Frequency-dependent burst firing was enhanced in P30, but not in P15 rats following KA-SE. The enhancement of bursting induced by KA-SE in more mature rats suggests that the frequency-dependent limitation of repetitive burst firing, which normally occurs in the subiculum, is compromised following SE. These changes could facilitate the initiation of spontaneous recurrent seizures or their spread from the hippocampus to other parts of the brain.

}, issn = {1932-6203}, doi = {10.1371/journal.pone.0119411}, author = {Chung, Sungkwon and Spruston, Nelson and Koh, Sookyong} } @article {68446, title = {Ankyrin Repeats Convey Force to Gate the NOMPC Mechanotransduction Channel.}, journal = {Cell}, volume = {162}, year = {2015}, month = {09/2015}, pages = {1391-403}, abstract = {

How metazoan mechanotransduction channels sense mechanical stimuli is not well understood. The NOMPC channel in the transient receptor potential (TRP) family, a mechanotransduction channel for Drosophila touch sensation and hearing, contains 29 Ankyrin repeats (ARs) that associate with microtubules. These ARs have been postulated to act as a tether that conveys force to the channel. Here, we report that these N-terminal ARs form a cytoplasmic domain essential for NOMPC mechanogating in vitro, mechanosensitivity of touch receptor neurons in vivo, and touch-induced behaviors of Drosophila larvae. Duplicating the ARs elongates the filaments that tether NOMPC to microtubules in mechanosensory neurons. Moreover, microtubule association is required for NOMPC mechanogating. Importantly, transferring the NOMPC ARs to mechanoinsensitive voltage-gated potassium channels confers mechanosensitivity to the chimeric channels. These experiments strongly support a tether mechanism of mechanogating for the NOMPC channel, providing insights into the basis of mechanosensitivity of mechanotransduction channels.

}, keywords = {Animals, Drosophila, Drosophila Proteins, Kv1.2 Potassium Channel, Larva, Mechanotransduction, Cellular, Microtubules, Protein Structure, Tertiary, Touch, Transient Receptor Potential Channels}, issn = {1097-4172}, doi = {10.1016/j.cell.2015.08.024}, author = {Zhang, Wei and Cheng, Li E and Kittelmann, Maike and Li, Jiefu and Petkovic, Maja and Cheng, Tong and Jin, Peng and Guo, Zhenhao and G{\"o}pfert, Martin C and Jan, Lily Yeh and Jan, Yuh Nung} } @article {46530, title = {Asymmetric formation of coated pits on dorsal and ventral surfaces at the leading edge of motile cells and on protrusions of immobile cells.}, journal = {Molecular Biology of the Cell}, volume = {26}, year = {2015}, month = {2015 Apr 7}, pages = {2044-53}, abstract = {

Clathrin/AP2-coated vesicles are the principal endocytic carriers originating at the plasma membrane. In experiments reported here, we have used spinning disk confocal and lattice light sheet microscopy to study the assembly dynamics of coated pits on the dorsal and ventral membranes of migrating U373 glioblastoma cells stably expressing AP2-EGFP and on lateral protrusions from immobile SUM159 breast carcinoma cells, gene edited to express AP2-EGFP. On U373 cells, coated pits initiated on the dorsal membrane at the front of the lamellipodium, as well as at the approximate boundary between the lamellipodium and lamella, and continued to grow as they were swept back toward the cell body; coated pits were absent from the corresponding ventral membrane. We observed a similar dorsal/ventral asymmetry on membrane protrusions from SUM159 cells. Stationary-coated pits formed and budded on the remainder of the dorsal and ventral surfaces of both types of cells. These observations support a previously proposed model that invokes net membrane deposition at the leading edge due to an imbalance between the endocytic and exocytic membrane flow at the front of a migrating cell.

}, issn = {1939-4586}, doi = {10.1091/mbc.E15-01-0055}, author = {Kural, Comert and Akatay, Ahmet Ata and Gaudin, Rapha{\"e}l and Chen, Bi-Chang and Legant, Wesley R and Betzig, Eric and Kirchhausen, Tom} } @article {47895, title = {Canonical genetic signatures of the adult human brain.}, journal = {Nature neuroscience}, volume = {18}, year = {2015}, month = {2015 Dec}, pages = {1832-44}, abstract = {

The structure and function of the human brain are highly stereotyped, implying a conserved molecular program responsible for its development, cellular structure and function. We applied a correlation-based metric called differential stability to assess reproducibility of gene expression patterning across 132 structures in six individual brains, revealing mesoscale genetic organization. The genes with the highest differential stability are highly biologically relevant, with enrichment for brain-related annotations, disease associations, drug targets and literature citations. Using genes with high differential stability, we identified 32 anatomically diverse and reproducible gene expression signatures, which represent distinct cell types, intracellular components and/or associations with neurodevelopmental and neurodegenerative disorders. Genes in neuron-associated compared to non-neuronal networks showed higher preservation between human and mouse; however, many diversely patterned genes displayed marked shifts in regulation between species. Finally, highly consistent transcriptional architecture in neocortex is correlated with resting state functional connectivity, suggesting a link between conserved gene expression and functionally relevant circuitry.

}, keywords = {Adult, Animals, Brain, Gene Regulatory Networks, Humans, Mice, Nerve Net, Transcriptome}, issn = {1546-1726}, doi = {10.1038/nn.4171}, author = {Hawrylycz, Michael and Miller, Jeremy A and Menon, Vilas and Feng, David and Dolbeare, Tim and Guillozet-Bongaarts, Angela L and Jegga, Anil G and Aronow, Bruce J and Lee, Chang-Kyu and Bernard, Amy and Glasser, Matthew F and Dierker, Donna L and Menche, J{\"o}rg and Szafer, Aaron and Collman, Forrest and Grange, Pascal and Berman, Kenneth A and Mihalas, Stefan and Yao, Zizhen and Stewart, Lance and Barab{\'a}si, Albert-L{\'a}szl{\'o} and Schulkin, Jay and Phillips, John and Ng, Lydia and Dang, Chinh and Haynor, David R and Jones, Allan and Van Essen, David C and Koch, Christof and Lein, Ed} } @article {46546, title = {Cellular levels of signaling factors are sensed by β-actin alleles to modulate transcriptional pulse intensity.}, journal = {Cell Reports}, volume = {11}, year = {2015}, month = {2015 Apr 7}, pages = {419-32}, abstract = {

The transcriptional response of β-actin to extra-cellular stimuli is a paradigm for transcription factor complex assembly and regulation. Serum induction leads to a precisely timed pulse of β-actin transcription in the cell population. Actin protein is proposed to be involved in this response, but it is not known whether cellular actin levels affect nuclear β-actin transcription. We perturbed the levels of key signaling factors and examined the effect on the induced transcriptional pulse by following endogenous β-actin alleles in single living cells. Lowering serum response factor (SRF) protein levels leads to loss of pulse integrity, whereas reducing actin protein levels reveals positive feedback regulation, resulting in elevated gene activation and a prolonged transcriptional response. Thus, transcriptional pulse fidelity requires regulated amounts of signaling proteins, and perturbations in factor levels eliminate the physiological response, resulting in either tuning down or exaggeration of the transcriptional pulse.

}, issn = {2211-1247}, doi = {10.1016/j.celrep.2015.03.039}, author = {Kalo, Alon and Kanter, Itamar and Shraga, Amit and Sheinberger, Jonathan and Tzemach, Hadar and Kinor, Noa and Singer, Robert H and Lionnet, Timoth{\'e}e and Shav-Tal, Yaron} } @article {47034, title = {Chemical perturbation of an intrinsically disordered region of TFIID distinguishes two modes of transcription initiation.}, journal = {eLife}, volume = {4}, year = {2015}, month = {2015 Aug 28}, abstract = {

Intrinsically disordered protein regions (IDRs) are peptide segments that fail to form stable 3-dimensional structures in the absence of partner proteins. They are abundant in eukaryotic proteomes and are often associated with human diseases, but their biological functions have been elusive to study. Here we report the identification of a tin(IV) oxochloride-derived cluster that binds an evolutionarily conserved IDR within the metazoan TFIID transcription complex. Binding arrests an isomerization of promoter-bound TFIID that is required for the engagement of Pol II during the first (de novo) round of transcription initiation. However, the specific chemical probe does not affect reinitiation, which requires the re-entry of Pol II, thus, mechanistically distinguishing these two modes of transcription initiation. This work also suggests a new avenue for targeting the elusive IDRs by harnessing certain features of metal-based complexes for mechanistic studies, and for the development of novel pharmaceutical interventions.

}, issn = {2050-084X}, doi = {10.7554/eLife.07777}, author = {Zhang, Zhengjian and Boskovic, Zarko and Hussain, Mahmud M and Hu, Wenxin and Inouye, Carla and Kim, Han-Je and Abole, A Katherine and Doud, Mary K and Lewis, Timothy A and Koehler, Angela N and Schreiber, Stuart L and Tjian, Robert} } @article {45854, title = {A consistent muscle activation strategy underlies crawling and swimming in Caenorhabditis elegans.}, journal = {Journal of the Royal Society Interface}, volume = {12}, year = {2015}, month = {2015 Jan 6}, pages = {20140963}, abstract = {

Although undulatory swimming is observed in many organisms, the neuromuscular basis for undulatory movement patterns is not well understood. To better understand the basis for the generation of these movement patterns, we studied muscle activity in the nematode Caenorhabditis elegans. Caenorhabditis elegans exhibits a range of locomotion patterns: in low viscosity fluids the undulation has a wavelength longer than the body and propagates rapidly, while in high viscosity fluids or on agar media the undulatory waves are shorter and slower. Theoretical treatment of observed behaviour has suggested a large change in force-posture relationships at different viscosities, but analysis of bend propagation suggests that short-range proprioceptive feedback is used to control and generate body bends. How muscles could be activated in a way consistent with both these results is unclear. We therefore combined automated worm tracking with calcium imaging to determine muscle activation strategy in a variety of external substrates. Remarkably, we observed that across locomotion patterns spanning a threefold change in wavelength, peak muscle activation occurs approximately 45{\textdegree} (1/8th of a cycle) ahead of peak midline curvature. Although the location of peak force is predicted to vary widely, the activation pattern is consistent with required force in a model incorporating putative length- and velocity-dependence of muscle strength. Furthermore, a linear combination of local curvature and velocity can match the pattern of activation. This suggests that proprioception can enable the worm to swim effectively while working within the limitations of muscle biomechanics and neural control.

}, issn = {1742-5662}, author = {Butler, Victoria J and Branicky, Robyn and Yemini, Eviatar and Liewald, Jana F and Gottschalk, Alexander and Kerr, Rex A and Chklovskii, Dmitri B and Schafer, William R} } @article {46890, title = {Continuous volumetric imaging via an optical phase-locked ultrasound lens.}, journal = {Nature Methods}, volume = {12}, year = {2015}, month = {2015-Jul 13}, pages = {759-62}, abstract = {

In vivo imaging at high spatiotemporal resolution is key to the understanding of complex biological systems. We integrated an optical phase-locked ultrasound lens into a two-photon fluorescence microscope and achieved microsecond-scale axial scanning, thus enabling volumetric imaging at tens of hertz. We applied this system to multicolor volumetric imaging of processes sensitive to motion artifacts, including calcium dynamics in behaving mouse brain and transient morphology changes and trafficking of immune cells.

}, doi = {10.1038/nmeth.3476}, author = {Kong, Lingjie and Tang, Jianyong and Little, Justin P and Yu, Yang and L{\"a}mmermann, Tim and Lin, Charles P and Germain, Ronald N and Cui, Meng} } @article {46022, title = {Dendritic function in vivo.}, journal = {Trends in Neuroscience}, volume = {38}, year = {2015}, month = {2015 Jan}, pages = {45-54}, abstract = {

Dendrites are the predominant entry site for excitatory synaptic potentials in most types of central neurons. There is increasing evidence that dendrites are not just passive transmitting devices but play active roles in synaptic integration through linear and non-linear mechanisms. Frequently, excitatory synapses are formed on dendritic spines. In addition to relaying incoming electrical signals, spines can play important roles in modifying these signals through complex biochemical processes and, thereby, determine learning and memory formation. Here, we review recent advances in our understanding of the function of spines and dendrites in central mammalian neurons in vivo by focusing particularly on insights obtained from Ca(2+) imaging studies.

}, issn = {1878-108X}, doi = {10.1016/j.tins.2014.11.002}, author = {Grienberger, Christine and Chen, Xiaowei and Konnerth, Arthur} } @article {47201, title = {Dendritic sodium spikes are required for long-term potentiation at distal synapses on hippocampal pyramidal neurons.}, journal = {eLife}, volume = {4}, year = {2015}, month = {2015 Aug 06}, abstract = {

Dendritic integration of synaptic inputs mediates rapid neural computation as well as longer-lasting plasticity. Several channel types can mediate dendritically initiated spikes (dSpikes), which may impact information processing and storage across multiple timescales; however, the roles of different channels in the rapid vs long-term effects of dSpikes are unknown. We show here that dSpikes mediated by Nav channels (blocked by a low concentration of TTX) are required for long-term potentiation (LTP) in the distal apical dendrites of hippocampal pyramidal neurons. Furthermore, imaging, simulations, and buffering experiments all support a model whereby fast Nav channel-mediated dSpikes (Na-dSpikes) contribute to LTP induction by promoting large, transient, localized increases in intracellular calcium concentration near the calcium-conducting pores of NMDAR and L-type Cav channels. Thus, in addition to contributing to rapid neural processing, Na-dSpikes are likely to contribute to memory formation via their role in long-lasting synaptic plasticity.

}, issn = {2050-084X}, doi = {10.7554/eLife.06414}, author = {Kim, Yujin and Hsu, Ching-Lung and Cembrowski, Mark S and Mensh, Brett D and Spruston, Nelson} } @article {47215, title = {Direct in~vivo manipulation and imaging of calcium transients in neutrophils identify a critical role for leading-edge calcium flux.}, journal = {Cell Reports}, volume = {13}, year = {2015}, month = {2015 Dec 15}, pages = {2107-17}, abstract = {

Calcium signaling has long been associated with key events of immunity, including chemotaxis, phagocytosis, and activation. However, imaging and manipulation of calcium flux in motile immune cells in live animals remain challenging. Using light-sheet microscopy for in\ vivo calcium imaging in zebrafish, we observe characteristic patterns of calcium flux triggered by distinct events, including phagocytosis of pathogenic bacteria and migration of neutrophils toward inflammatory stimuli. In contrast to findings from ex\ vivo studies, we observe enriched calcium influx at the leading edge of migrating neutrophils. To directly manipulate calcium dynamics in\ vivo, we have developed transgenic lines with cell-specific expression of the mammalian TRPV1 channel, enabling ligand-gated, reversible, and spatiotemporal control of calcium influx. We find that controlled calcium influx can function to help define the neutrophil{\textquoteright}s leading edge. Cell-specific TRPV1 expression may have broad utility for precise control of calcium dynamics in other immune cell types and organisms.

}, issn = {2211-1247}, doi = {10.1016/j.celrep.2015.11.010}, author = {Beerman, Rebecca W and Matty, Molly A and Au, Gina G and Looger, Loren L and Choudhury, Kingshuk Roy and Keller, Philipp J and Tobin, David M} } @article {47036, title = {Dopamine is required for the neural representation and control of movement vigor.}, journal = {Cell}, volume = {162}, year = {2015}, month = {2015 Sep 10}, pages = {1418-30}, abstract = {

Progressive depletion of midbrain dopamine neurons (PDD) is associated with deficits in the initiation, speed, and fluidity of voluntary movement. Models of\ basal ganglia function focus on initiation deficits; however, it is unclear how they account for deficits in the speed or amplitude of movement (vigor). Using an effort-based operant conditioning task for head-fixed mice, we discovered distinct functional classes of neurons in the dorsal striatum that represent movement vigor. Mice with PDD exhibited a progressive reduction in vigor, along with a selective impairment of its neural representation in striatum. Restoration of dopaminergic tone with a synthetic precursor ameliorated deficits in movement vigor and its neural representation, while suppression of striatal activity during movement was sufficient to reduce vigor. Thus, dopaminergic input to the dorsal striatum is indispensable for the emergence of striatal activity that mediates adaptive changes in movement vigor. These results suggest refined intervention strategies for Parkinson{\textquoteright}s disease.

}, doi = {10.1016/j.cell.2015.08.014}, author = {Panigrahi, Babita and Martin, Kathleen A and Li, Yi and Graves, Austin R and Vollmer, Alison and Olson, Lars and Mensh, Brett D and Karpova, Alla Y and Dudman, Joshua T} } @article {46773, title = {A dopamine-modulated neural circuit regulating aversive taste memory in Drosophila.}, journal = {Current Biology}, volume = {25}, year = {2015}, month = {2015 May 14}, pages = {1535-41}, abstract = {

Taste memories allow animals to modulate feeding behavior in accordance with past experience and avoid the consumption of potentially harmful food [1]. We have developed a single-fly taste memory assay to functionally interrogate the neural circuitry encoding taste memories [2]. Here, we screen a collection of Split-GAL4 lines that label small populations of neurons associated with the fly memory center-the mushroom bodies (MBs) [3]. Genetic silencing of PPL1 dopamine neurons disrupts conditioned, but not naive, feeding behavior, suggesting these neurons are selectively involved in the conditioned taste response. We identify two PPL1 subpopulations that innervate the MB α lobe and are essential for aversive taste memory. Thermogenetic activation of these dopamine neurons during training induces memory, indicating these neurons are sufficient for the reinforcing properties of bitter tastant to the MBs. Silencing of either the intrinsic MB neurons or the output neurons from the α lobe disrupts taste conditioning. Thermogenetic manipulation of these output neurons alters naive feeding response, suggesting that dopamine neurons modulate the threshold of response to appetitive tastants. Taken together, these findings detail a neural mechanism underlying the formation of taste memory and provide a functional model for dopamine-dependent plasticity in Drosophila.

}, issn = {1879-0445}, doi = {10.1016/j.cub.2015.04.027}, author = {Masek, Pavel and Worden, Kurtresha and Aso, Yoshinori and Rubin, Gerald M and Keene, Alex C} } @article {47160, title = {Dynamics of CRISPR-Cas9 genome interrogation in living cells.}, journal = {Science (New York, N.Y.)}, volume = {350}, year = {2015}, month = {2015 Nov 13}, pages = {823-6}, abstract = {

The RNA-guided CRISPR-associated protein Cas9 is used for genome editing, transcriptional modulation, and live-cell imaging. Cas9-guide RNA complexes recognize and cleave double-stranded DNA sequences on the basis of 20-nucleotide RNA-DNA complementarity, but the mechanism of target searching in mammalian cells is unknown. Here, we use single-particle tracking to visualize diffusion and chromatin binding of Cas9 in living cells. We show that three-dimensional diffusion dominates Cas9 searching in vivo, and off-target binding events are, on average, short-lived (\<1 second). Searching is dependent on the local chromatin environment, with less sampling and slower movement within heterochromatin. These results reveal how the bacterial Cas9 protein interrogates mammalian genomes and navigates eukaryotic chromatin structure.

}, issn = {1095-9203}, doi = {10.1126/science.aac6572}, author = {Knight, Spencer C and Xie, Liangqi and Deng, Wulan and Guglielmi, Benjamin and Witkowsky, Lea B and Bosanac, Lana and Zhang, Elisa T and El Beheiry, Mohamed and Masson, Jean-Baptiste and Dahan, Maxime and Liu, Zhe and Doudna, Jennifer A and Tjian, Robert} } @article {46283, title = {Effects of long-term representations on free recall of unrelated words.}, journal = {Learning \& Memory}, volume = {22}, year = {2015}, month = {2015 Jan 15}, pages = {101-8}, abstract = {

Human memory stores vast amounts of information. Yet recalling this information is often challenging when specific cues are lacking. Here we consider an associative model of retrieval where each recalled item triggers the recall of the next item based on the similarity between their long-term neuronal representations. The model predicts that different items stored in memory have different probability to be recalled depending on the size of their representation. Moreover, items with high recall probability tend to be recalled earlier and suppress other items. We performed an analysis of a large data set on free recall and found a highly specific pattern of statistical dependencies predicted by the model, in particular negative correlations between the number of words recalled and their average recall probability. Taken together, experimental and modeling results presented here reveal complex interactions between memory items during recall that severely constrain recall capacity.

}, issn = {1549-5485}, doi = {10.1101/lm.035238.114}, author = {Katkov, Mikhail and Romani, Sandro and Tsodyks, Misha} } @article {47082, title = {Efficient processing and analysis of large-scale light-sheet microscopy data.}, journal = {Nature Protocols}, volume = {10}, year = {2015}, month = {2015 Oct 2}, pages = {1679-96}, abstract = {

Light-sheet microscopy is a powerful method for imaging the development and function of complex biological systems at high spatiotemporal resolution and over long time scales. Such experiments typically generate terabytes of multidimensional image data, and thus they demand efficient computational solutions for data management, processing and analysis. We present protocols and software to tackle these steps, focusing on the imaging-based study of animal development. Our protocols facilitate (i) high-speed lossless data compression and content-based multiview image fusion optimized for multicore CPU architectures, reducing image data size 30{\textendash}500-fold; (ii) automated large-scale cell tracking and segmentation; and (iii) visualization, editing and annotation of multiterabyte image data and cell-lineage reconstructions with tens of millions of data points. These software modules are open source. They provide high data throughput using a single computer workstation and are readily applicable to a wide spectrum of biological model systems.

}, doi = {10.1038/nprot.2015.111}, author = {Amat, Fernando and H{\"o}ckendorf, Burkhard and Wan, Yinan and Lemon, William C and McDole, Katie and Keller, Philipp J} } @article {47302, title = {Emotor control: computations underlying bodily resource allocation, emotions, and confidence.}, journal = {Dialogues in Clinical Neuroscience}, volume = {17}, year = {2015}, month = {2015 Dec}, pages = {391-401}, abstract = {

Emotional processes are central to behavior, yet their deeply subjective nature has been a challenge for neuroscientific study as well as for psychiatric diagnosis. Here we explore the relationships between subjective feelings and their underlying brain circuits from a computational perspective. We apply recent insights from systems neuroscience-approaching subjective behavior as the result of mental computations instantiated in the brain-to the study of emotions. We develop the hypothesis that emotions are the product of neural computations whose motor role is to reallocate bodily resources mostly gated by smooth muscles. This "emotor" control system is analagous to the more familiar motor control computations that coordinate skeletal muscle movements. To illustrate this framework, we review recent research on "confidence." Although familiar as a feeling, confidence is also an objective statistical quantity: an estimate of the probability that a hypothesis is correct. This model-based approach helped reveal the neural basis of decision confidence in mammals and provides a bridge to the subjective feeling of confidence in humans. These results have important implications for psychiatry, since disorders of confidence computations appear to contribute to a number of psychopathologies. More broadly, this computational approach to emotions resonates with the emerging view that psychiatric nosology may be best parameterized in terms of disorders of the cognitive computations underlying complex behavior.

}, issn = {1958-5969}, author = {Kepecs, Adam and Mensh, Brett D} } @article {47020, title = {Extended-resolution structured illumination imaging of endocytic and cytoskeletal dynamics.}, journal = {Science (New York, N.Y.)}, volume = {349}, year = {2015}, month = {2015 Aug 28}, abstract = {

Super-resolution fluorescence microscopy is distinct among nanoscale imaging tools in its ability to image protein dynamics in living cells. Structured illumination microscopy (SIM) stands out in this regard because of its high speed and low illumination intensities, but typically offers only a twofold resolution gain. We extended the resolution of live-cell SIM through two approaches: ultrahigh numerical aperture SIM at 84-nanometer lateral resolution for more than 100 multicolor frames, and nonlinear SIM with patterned activation at 45- to 62-nanometer resolution for approximately 20 to 40 frames. We applied these approaches to image dynamics near the plasma membrane of spatially resolved assemblies of clathrin and caveolin, Rab5a in early endosomes, and α-actinin, often in relationship to cortical actin. In addition, we examined mitochondria, actin, and the Golgi apparatus dynamics in three dimensions.

}, issn = {1095-9203}, doi = {10.1126/science.aab3500}, author = {Li, Dong and Shao, Lin and Chen, Bi-Chang and Zhang, Xi and Zhang, Mingshu and Moses, Brian and Milkie, Daniel E and Beach, Jordan R and Hammer, John A and Pasham, Mithun and Kirchhausen, Tomas and Baird, Michelle A and Davidson, Michael W and Xu, Pingyong and Betzig, Eric} } @article {46862, title = {From a meso- to micro-scale connectome: array tomography and mGRASP.}, journal = {Frontiers in Neuroanatomy}, volume = {9}, year = {2015}, month = {2015 Jun 04}, pages = {78}, abstract = {

Mapping mammalian synaptic connectivity has long been an important goal of neuroscience because knowing how neurons and brain areas are connected underpins an understanding of brain function. Meeting this goal requires advanced techniques with single synapse resolution and large-scale capacity, especially at multiple scales tethering the meso- and micro-scale connectome. Among several advanced LM-based connectome technologies, Array Tomography (AT) and mammalian GFP-Reconstitution Across Synaptic Partners (mGRASP) can provide relatively high-throughput mapping synaptic connectivity at multiple scales. AT- and mGRASP-assisted circuit mapping (ATing and mGRASPing), combined with techniques such as retrograde virus, brain clearing techniques, and activity indicators will help unlock the secrets of complex neural circuits. Here, we discuss these useful new tools to enable mapping of brain circuits at multiple scales, some functional implications of spatial synaptic distribution, and future challenges and directions of these endeavors.

}, issn = {1662-5129}, doi = {10.3389/fnana.2015.00078}, author = {Rah, Jong-Cheol and Feng, Linqing and Druckmann, Shaul and Lee, Hojin and Kim, Jinhyun} } @article {46549, title = {Generating customized transgene landing sites and multi-transgene arrays in Drosophila using phiC31 integrase.}, journal = {Genetics}, volume = {199}, year = {2015}, month = {2015 Apr}, pages = {919-34}, abstract = {

Transgenesis in numerous eukaryotes has been facilitated by the use of site-specific integrases to stably insert transgenes at predefined genomic positions (landing sites). However, the utility of integrase-mediated transgenesis in any system is constrained by the limited number and variable expression properties of available landing sites. By exploiting the nonstandard recombination activity exhibited by a phiC31 integrase mutant, we developed a rapid and inexpensive method for isolating landing sites that exhibit desired expression properties. Additionally, we devised a simple technique for constructing arrays of transgenes at a single landing site, thereby extending the utility of previously characterized landing sites. Using the fruit fly Drosophila melanogaster, we demonstrate the feasibility of these approaches by isolating new landing sites optimized to express transgenes in the nervous system and by building fluorescent reporter arrays at several landing sites. Because these strategies require the activity of only a single exogenous protein, we anticipate that they will be portable to species such as nonmodel organisms, in which genetic manipulation is more challenging, expediting the development of genetic resources in these systems.

}, issn = {1943-2631}, doi = {10.1534/genetics.114.173187}, author = {Knapp, Jon-Michael and Chung, Phuong and Simpson, Julie H} } @article {47896, title = {Genome engineering of isogenic human ES cells to model autism disorders.}, journal = {Nucleic acids research}, volume = {43}, year = {2015}, month = {2015 May 26}, pages = {e65}, abstract = {

Isogenic pluripotent stem cells are critical tools for studying human neurological diseases by allowing one to study the effects of a mutation in a fixed genetic background. Of particular interest are the spectrum of autism disorders, some of which are monogenic such as Timothy syndrome (TS); others are multigenic such as the microdeletion and microduplication syndromes of the 16p11.2 chromosomal locus. Here, we report engineered human embryonic stem cell (hESC) lines for modeling these two disorders using locus-specific endonucleases to increase the efficiency of homology-directed repair (HDR). We developed a system to: (1) computationally identify unique transcription activator-like effector nuclease (TALEN) binding sites in the genome using a new software program, TALENSeek, (2) assemble the TALEN genes by combining golden gate cloning with modified constructs from the FLASH protocol, and (3) test the TALEN pairs in an amplification-based HDR assay that is more sensitive than the typical non-homologous end joining assay. We applied these methods to identify, construct, and test TALENs that were used with HDR donors in hESCs to generate an isogenic TS cell line in a scarless manner and to model the 16p11.2 copy number disorder without modifying genomic loci with high sequence similarity.

}, keywords = {Autistic Disorder, Binding Sites, Cell Engineering, Cell Line, Child Development Disorders, Pervasive, Deoxyribonucleases, Embryonic Stem Cells, Gene Targeting, Genome, Human, Humans, Long QT Syndrome, Models, Genetic, Recombinational DNA Repair, Software, Syndactyly}, issn = {1362-4962}, doi = {10.1093/nar/gkv164}, author = {Martinez, Refugio A and Stein, Jason L and Krostag, Anne-Rachel F and Nelson, Angelique M and Marken, John S and Menon, Vilas and May, Ryan C and Yao, Zizhen and Kaykas, Ajamete and Geschwind, Daniel H and Grimley, Joshua S} } @article {47014, title = {Genome-wide errant targeting by Hairy.}, journal = {eLife}, volume = {4}, year = {2015}, month = {2015 Aug 25}, abstract = {

Metazoan transcriptional repressors regulate chromatin through diverse histone modifications. Contributions of individual factors to the chromatin landscape in development is difficult to establish, as global surveys reflect multiple changes in regulators. Therefore, we studied the conserved Hairy/Enhancer of Split family repressor Hairy, analyzing histone marks and gene expression in Drosophila embryos. This long-range repressor mediates histone acetylation and methylation in large blocks, with highly context-specific effects on target genes. Most strikingly, Hairy exhibits biochemical activity on many loci that are uncoupled to changes in gene expression. Rather than representing inert binding sites, as suggested for many eukaryotic factors, many regions are targeted errantly by Hairy to modify the chromatin landscape. Our findings emphasize that identification of active cis-regulatory elements must extend beyond the survey of prototypical chromatin marks. We speculate that this errant activity may provide a path for creation of new regulatory elements, facilitating the evolution of novel transcriptional circuits.

}, issn = {2050-084X}, doi = {10.7554/eLife.06394}, author = {Kok, Kurtulus and Ay, Ahmet and Li, Li M and Arnosti, David N} } @article {46756, title = {Genomic signatures of evolutionary transitions from solitary to group living.}, journal = {Science}, year = {2015}, month = {2015 May 14}, abstract = {

The evolution of eusociality is one of the major transitions in evolution, but the underlying genomic changes are unknown. We compared the genomes of ten bee species that vary in social complexity, representing multiple independent transitions in social evolution, and report three major findings. First, many important genes show evidence of neutral evolution as a consequence of relaxed selection with increasing social complexity. Second, there is no single road map to eusociality; independent evolutionary transitions in sociality have independent genetic underpinnings. Third, though clearly independent in detail, these transitions do have similar general features, including an increase in constrained protein evolution accompanied by increases in the potential for gene regulation and decreases in diversity and abundance of transposable elements. Eusociality may arise through different mechanisms each time, but would likely always involve an increase in the complexity of gene networks.

}, issn = {1095-9203}, doi = {10.1126/science.aaa4788}, author = {Kapheim, Karen M and Pan, Hailin and Li, Cai and Salzberg, Steven L and Puiu, Daniela and Magoc, Tanja and Robertson, Hugh M and Hudson, Matthew E and Venkat, Aarti and Fischman, Brielle J and Hernandez, Alvaro and Yandell, Mark and Ence, Daniel and Holt, Carson and Yocum, George D and Kemp, William P and Bosch, Jordi and Waterhouse, Robert M and Zdobnov, Evgeny M and Stolle, Eckart and Kraus, F Bernhard and Helbing, Sophie and Moritz, Robin F A and Glastad, Karl M and Hunt, Brendan G and Goodisman, Michael A D and Hauser, Frank and Grimmelikhuijzen, Cornelis J P and Pinheiro, Daniel Guariz and Nunes, Francis Morais Franco and Soares, Michelle Prioli Miranda and Tanaka, {\'E}rica Donato and Sim{\~o}es, Zil{\'a} Luz Paulino and Hartfelder, Klaus and Evans, Jay D and Barribeau, Seth M and Johnson, Reed M and Massey, Jonathan H and Southey, Bruce R and Hasselmann, Martin and Hamacher, Daniel and Biewer, Matthias and Kent, Clement F and Zayed, Amro and Blatti, Charles and Sinha, Saurabh and Johnston, J Spencer and Hanrahan, Shawn J and Kocher, Sarah D and Wang, Jun and Robinson, Gene E and Zhang, Guojie} } @article {47084, title = {Green-to-red photoconversion of GCaMP.}, journal = {PLoS One}, volume = {10}, year = {2015}, month = {2015 Sep 18}, pages = {e0138127}, abstract = {

Genetically encoded calcium indicators (GECIs) permit imaging intracellular calcium transients. Among GECIs, the GFP-based GCaMPs are the most widely used because of their high sensitivity and rapid response to changes in intracellular calcium concentrations. Here we report that the fluorescence of GCaMPs-including GCaMP3, GCaMP5 and GCaMP6-can be converted from green to red following exposure to blue-green light (450-500 nm). This photoconversion occurs in both insect and mammalian cells and is enhanced in a low oxygen environment. The red fluorescent GCaMPs retained calcium responsiveness, albeit with reduced sensitivity. We identified several amino acid residues in GCaMP important for photoconversion and generated a GCaMP variant with increased photoconversion efficiency in cell culture. This light-induced spectral shift allows the ready labeling of specific, targeted sets of GCaMP-expressing cells for functional imaging in the red channel. Together, these findings indicate the potential for greater utility of existing GCaMP reagents, including transgenic animals.

}, issn = {1932-6203}, doi = {10.1371/journal.pone.0138127}, author = {Ai, Minrong and Mills, Holly and Kanai, Makoto and Lai, Jason and Deng, Jingjing and Schreiter, Eric and Looger, Loren and Neubert, Thomas and Suh, Greg} } @article {46895, title = {A high throughput (>90\%), large compensation range, single-prism femtosecond pulse compressor.}, journal = {arXiv}, year = {2015}, month = {2015 Jun 20}, pages = {arXiv:1306.5011 }, abstract = {

We demonstrate a high throughput, large compensation range, single-prism femtosecond pulse compressor, using a single prism and two roof mirrors. The compressor has zero angular dispersion, zero spatial dispersion, zero pulse-front tilt, and unity magnification. The high efficiency is achieved by adopting two roof mirrors as the retroreflectors. We experimentally achieved ~ -14500 fs2 group delay dispersion (GDD) with 30 cm of prism tip-roof mirror prism separation, and ~90.7\% system throughput with the current implementation. With better components, the throughput can be even higher.

}, url = {http://arxiv.org/abs/1306.5011}, author = {Kong, Lingjie and Cui, Meng} } @article {47128, title = {A higher order visual neuron tuned to the spatial amplitude spectra of natural scenes.}, journal = {Nature Communications}, volume = {6}, year = {2015}, month = {2015 Oct 06}, pages = {8522}, abstract = {

Animal sensory systems are optimally adapted to those features typically encountered in natural surrounds, thus allowing neurons with limited bandwidth to encode challengingly large input ranges. Natural scenes are not random, and peripheral visual systems in vertebrates and insects have evolved to respond efficiently to their typical spatial statistics. The mammalian visual cortex is also tuned to natural spatial statistics, but less is known about coding in higher order neurons in insects. To redress this we here record intracellularly from a higher order visual neuron in the hoverfly. We show that the cSIFE neuron, which is inhibited by stationary images, is maximally inhibited when the slope constant of the amplitude spectrum is close to the mean in natural scenes. The behavioural optomotor response is also strongest to images with naturalistic image statistics. Our results thus reveal a close coupling between the inherent statistics of natural scenes and higher order visual processing in insects.

}, issn = {2041-1723}, doi = {10.1038/ncomms9522}, author = {Dyakova, Olga and Lee, Yu-Jen and Longden, Kit D and Kiselev, Valerij G and Nordstr{\"o}m, Karin} } @article {46819, title = {High-resolution live imaging reveals axon-glia interactions during peripheral nerve injury and repair in zebrafish.}, journal = {Disease Models \& Mechanisms}, volume = {8}, year = {2015}, month = {2015 Jun 1}, pages = {553-64}, abstract = {

Neural damage is a devastating outcome of physical trauma. The glia are one of the main effectors of neuronal repair in the nervous system, but the dynamic interactions between peripheral neurons and Schwann cells during injury and regeneration remain incompletely characterized. Here, we combine laser microsurgery, genetic analysis, high-resolution intravital imaging and lattice light-sheet microscopy to study the interaction between Schwann cells and sensory neurons in a zebrafish model of neurotrauma. We found that chronic denervation by neuronal ablation leads to Schwann-cell death, whereas acute denervation by axonal severing does not affect the overall complexity and architecture of the glia. Neuronal-circuit regeneration begins when Schwann cells extend bridging processes to close the injury gap. Regenerating axons grow faster and directionally after the physiological clearing of distal debris by the Schwann cells. This might facilitate circuit repair by ensuring that axons are guided through unoccupied spaces within bands of B{\"u}ngner towards their original peripheral target. Accordingly, in the absence of Schwann cells, regenerating axons are misrouted, impairing the re-innervation of sensory organs. Our results indicate that regenerating axons use haptotaxis as a directional cue during the reconstitution of a neural circuit. These findings have implications for therapies aimed at neurorepair, which will benefit from preserving the architecture of the peripheral glia during periods of denervation.

}, issn = {1754-8411}, doi = {10.1242/dmm.018184}, author = {Xiao, Yan and Faucherre, Ad{\`e}le and Pola-Morell, Laura and Heddleston, John M and Liu, Tsung-Li and Chew, Teng-Leong and Sato, Fuminori and Sehara-Fujisawa, Atsuko and Kawakami, Koichi and L{\'o}pez-Schier, Hern{\'a}n} } @article {46522, title = {Identification of genes that promote or inhibit olfactory memory formation in Drosophila.}, journal = {Genetics}, volume = {199}, year = {2015}, month = {2015 Apr}, pages = {1173-82}, abstract = {

Genetic screens in Drosophila melanogaster and other organisms have been pursued to filter the genome for genetic functions important for memory formation. Such screens have employed primarily chemical or transposon-mediated mutagenesis and have identified numerous mutants including classical memory mutants, dunce and rutabaga. Here, we report the results of a large screen using panneuronal RNAi expression to identify additional genes critical for memory formation. We identified \>500 genes that compromise memory when inhibited (low hits), either by disrupting the development and normal function of the adult animal or by participating in the neurophysiological mechanisms underlying memory formation. We also identified \>40 genes that enhance memory when inhibited (high hits). The dunce gene was identified as one of the low hits and further experiments were performed to map the effects of the dunce RNAi to the α/β and γ mushroom body neurons. Additional behavioral experiments suggest that dunce knockdown in the mushroom body neurons impairs memory without significantly affecting acquisition. We also characterized one high hit, sickie, to show that RNAi knockdown of this gene enhances memory through effects in dopaminergic neurons without apparent effects on acquisition. These studies further our understanding of two genes involved in memory formation, provide a valuable list of genes that impair memory that may be important for understanding the neurophysiology of memory or neurodevelopmental disorders, and offer a new resource of memory suppressor genes that will aid in understanding restraint mechanisms employed by the brain to optimize resources.

}, issn = {1943-2631}, doi = {10.1534/genetics.114.173575}, author = {Walkinshaw, Erica and Gai, Yunchao and Farkas, Caitlin and Richter, Daniel and Nicholas, Eric and Keleman, Krystyna and Davis, Ronald L} } @article {47037, title = {Identification of inhibitory premotor interneurons activated at a late phase in a motor cycle during Drosophila larval locomotion.}, journal = {PLoS One}, volume = {10}, year = {2015}, month = {2015 Sep 03}, pages = {e0136660}, abstract = {

Rhythmic motor patterns underlying many types of locomotion are thought to be produced by central pattern generators (CPGs). Our knowledge of how CPG networks generate motor patterns in complex nervous systems remains incomplete, despite decades of work in a variety of model organisms. Substrate borne locomotion in Drosophila larvae is driven by waves of muscular contraction that propagate through multiple body segments. We use the motor circuitry underlying crawling in larval Drosophila as a model to try to understand how segmentally coordinated rhythmic motor patterns are generated. Whereas muscles, motoneurons and sensory neurons have been well investigated in this system, far less is known about the identities and function of interneurons. Our recent study identified a class of glutamatergic premotor interneurons, PMSIs (period-positive median segmental interneurons), that regulate the speed of locomotion. Here, we report on the identification of a distinct class of glutamatergic premotor interneurons called Glutamatergic Ventro-Lateral Interneurons (GVLIs). We used calcium imaging to search for interneurons that show rhythmic activity and identified GVLIs as interneurons showing wave-like activity during peristalsis. Paired GVLIs were present in each abdominal segment A1-A7 and locally extended an axon towards a dorsal neuropile region, where they formed GRASP-positive putative synaptic contacts with motoneurons. The interneurons expressed vesicular glutamate transporter (vGluT) and thus likely secrete glutamate, a neurotransmitter known to inhibit motoneurons. These anatomical results suggest that GVLIs are premotor interneurons that locally inhibit motoneurons in the same segment. Consistent with this, optogenetic activation of GVLIs with the red-shifted channelrhodopsin, CsChrimson ceased ongoing peristalsis in crawling larvae. Simultaneous calcium imaging of the activity of GVLIs and motoneurons showed that GVLIs{\textquoteright} wave-like activity lagged behind that of motoneurons by several segments. Thus, GVLIs are activated when the front of a forward motor wave reaches the second or third anterior segment. We propose that GVLIs are part of the feedback inhibition system that terminates motor activity once the front of the motor wave proceeds to anterior segments.

}, issn = {1932-6203}, doi = {10.1371/journal.pone.0136660}, author = {Itakura, Yuki and Kohsaka, Hiroshi and Ohyama, Tomoko and Zlatic, Marta and Pulver, Stefan R and Nose, Akinao} } @article {46534, title = {In vivo neuroimaging through the highly scattering tissue via iterative multi-photon adaptive compensation technique.}, journal = {Optics Express}, volume = {23}, year = {2015}, month = {2015 Mar 9}, pages = {6145-50}, abstract = {

For in vivo deep tissue imaging, high order wavefront measurement and correction is needed for handling the severe wavefront distortion. Towards such a goal, we have developed the iterative multi-photon adaptive compensation technique (IMPACT). In this work, we explore using IMPACT to perform calcium imaging of neocortex through the intact skull of adult mice, and to image through the highly scattering white matter on the hippocampus surface.

}, issn = {1094-4087}, doi = {10.1364/OE.23.006145}, author = {Kong, Lingjie and Cui, Meng} } @article {46929, title = {Inferring transient particle transport dynamics in live cells.}, journal = {Nature Methods}, volume = {12}, year = {2015}, month = {2015 Jul 20}, pages = {838-40}, abstract = {

Live-cell imaging and particle tracking provide rich information on mechanisms of intracellular transport. However, trajectory analysis procedures to infer complex transport dynamics involving stochastic switching between active transport and diffusive motion are lacking. We applied Bayesian model selection to hidden Markov modeling to infer transient transport states from trajectories of mRNA-protein complexes in live mouse hippocampal neurons and metaphase kinetochores in dividing human cells. The software is available at http://hmm-bayes.org/.

}, issn = {1548-7105}, doi = {10.1038/nmeth.3483}, author = {Monnier, Nilah and Barry, Zachary and Park, Hye Yoon and Su, Kuan-Chung and Katz, Zachary and English, Brian P and Dey, Arkajit and Pan, Keyao and Cheeseman, Iain M and Singer, Robert H and Bathe, Mark} } @conference {47229, title = {Interactive exemplar-based segmentation toolkit for biomedical image analysis.}, booktitle = {International Symposium on Biomedical Imaging}, year = {2015}, month = {2015 Apr}, abstract = {

In the field of biomedical imaging analysis on single-cell level, reliable and fast segmentation of the cell nuclei from the background on three-dimensional images is highly needed for the further analysis. In this work we propose an interactive cell segmentation toolkit that first establishes a set of exemplar regions from user input through an easy and intuitive interface in both 2D and 3D in real-time, then
extracts the shape and intensity features from those exemplars. Based on a local contrast-constrained region growing scheme, each connected component in the whole image would be filtered by the features from exemplars, forming an {\textquotedblleft}exemplar-matching{\textquotedblright} group which passed the filtering and would be part of the final segmentation result, and a {\textquotedblleft}non-exemplar-matching{\textquotedblright} group in which components
would be further segmented by the gradient vector field based algorithm. The results of the filtering process are visualized back to the user in near real-time, thus enhancing the experience in exemplar selecting and parameter tuning. The toolkit is distributed as a plugin within the open source Vaa3D system (http://vaa3d.org).

}, author = {Li, Xiang and Zhou, Zhi and Keller, Philipp J and Zeng, Hongkui and Liu, Tianming and Peng, Hanchuan} } @article {46368, title = {Labeling of active neural circuits in vivo with designed calcium integrators.}, journal = {Science}, volume = {347}, year = {2015}, month = {2015 Feb 13}, pages = {755-60}, abstract = {

The identification of active neurons and circuits in vivo is a fundamental challenge in understanding the neural basis of behavior. Genetically encoded calcium (Ca(2+)) indicators (GECIs) enable quantitative monitoring of cellular-resolution activity during behavior. However, such indicators require online monitoring within a limited field of view. Alternatively, post hoc staining of immediate early genes (IEGs) indicates highly active cells within the entire brain, albeit with poor temporal resolution. We designed a fluorescent sensor, CaMPARI, that combines the genetic targetability and quantitative link to neural activity of GECIs with the permanent, large-scale labeling of IEGs, allowing a temporally precise "activity snapshot" of a large tissue volume. CaMPARI undergoes efficient and irreversible green-to-red conversion only when elevated intracellular Ca(2+) and experimenter-controlled illumination coincide. We demonstrate the utility of CaMPARI in freely moving larvae of zebrafish and flies, and in head-fixed mice and adult flies.

}, issn = {1095-9203}, doi = {10.1126/science.1260922}, author = {Fosque, Benjamin F and Sun, Yi and Dana, Hod and Yang, Chao-Tsung and Ohyama, Tomoko and Tadross, Michael R and Patel, Ronak and Zlatic, Marta and Kim, Douglas S and Ahrens, Misha B and Jayaraman, Vivek and Looger, Loren L and Schreiter, Eric R} } @article {48830, title = {Learning enhances sensory and multiple non-sensory representations in primary visual cortex.}, journal = {Neuron}, volume = {86}, year = {2015}, month = {2015 Jun 17}, pages = {1478-90}, abstract = {

We determined how learning modifies neural representations in primary visual cortex (V1) during acquisition of a visually guided behavioral task. We imaged the activity of the same layer 2/3 neuronal populations as mice learned to discriminate two visual patterns while running through a virtual corridor, where one pattern was rewarded. Improvements in behavioral performance were closely associated with increasingly distinguishable population-level representations of task-relevant stimuli, as a result of stabilization of existing and recruitment of new neurons selective for these stimuli. These effects correlated with the appearance of multiple task-dependent signals during learning: those that increased neuronal selectivity across the population when expert animals engaged in the task, and those reflecting anticipation or behavioral choices specifically in neuronal subsets preferring the rewarded stimulus. Therefore, learning engages diverse mechanisms that modify sensory and non-sensory representations in V1 to adjust its processing to task requirements and the behavioral relevance of visual stimuli.

}, keywords = {Animals, Calcium, Discrimination (Psychology), Female, Learning, Luminescent Proteins, Male, Mice, Mice, Inbred C57BL, Mice, Transgenic, Models, Neurological, Neurons, Nonlinear Dynamics, Optogenetics, Photic Stimulation, Sensory Receptor Cells, User-Computer Interface, Vision, Ocular, Visual Cortex}, issn = {1097-4199}, doi = {10.1016/j.neuron.2015.05.037}, author = {Poort, Jasper and Khan, Adil G and Pachitariu, Marius and Nemri, Abdellatif and Orsolic, Ivana and Krupic, Julija and Bauza, Marius and Sahani, Maneesh and Keller, Georg B and Mrsic-Flogel, Thomas D and Hofer, Sonja B} } @article {45457, title = {Light sheet-based imaging and analysis of early embryogenesis in the fruit fly.}, journal = {Methods in Molecular Biology}, volume = {1189}, year = {2015}, month = {2015}, pages = {79-97}, abstract = {

The fruit fly is an excellent model system for investigating the sequence of epithelial tissue invaginations constituting the process of gastrulation. By combining recent advancements in light sheet fluorescence microscopy (LSFM) and image processing, the three-dimensional fly embryo morphology and relevant gene expression patterns can be accurately recorded throughout the entire process of embryogenesis. LSFM provides exceptionally high imaging speed, high signal-to-noise ratio, low level of photoinduced damage, and good optical penetration depth. This powerful combination of capabilities makes LSFM particularly suitable for live imaging of the fly embryo.The resulting high-information-content image data are subsequently processed to obtain the outlines of cells and cell nuclei, as well as the geometry of the whole embryo tissue by image segmentation. Furthermore, morphodynamics information is extracted by computationally tracking objects in the image. Towards that goal we describe the successful implementation of a fast fitting strategy of Gaussian mixture models.The data obtained by image processing is well-suited for hypothesis testing of the detailed biomechanics of the gastrulating embryo. Typically this involves constructing computational mechanics models that consist of an objective function providing an estimate of strain energy for a given morphological configuration of the tissue, and a numerical minimization mechanism of this energy, achieved by varying morphological parameters.In this chapter, we provide an overview of in vivo imaging of fruit fly embryos using LSFM, computational tools suitable for processing the resulting images, and examples of computational biomechanical simulations of fly embryo gastrulation.

}, issn = {1940-6029}, doi = {10.1007/978-1-4939-1164-6_6}, author = {Khairy, Khaled and Lemon, William C and Amat, Fernando and Keller, Philipp J} } @article {47144, title = {A Low Affinity GCaMP3 Variant (GCaMPer) for Imaging the Endoplasmic Reticulum Calcium Store.}, journal = {PloS one}, volume = {10}, year = {2015}, month = {2015 Oct 09}, pages = {e0139273}, abstract = {

Endoplasmic reticulum calcium homeostasis is critical for cellular functions and is disrupted in diverse pathologies including neurodegeneration and cardiovascular disease. Owing to the high concentration of calcium within the ER, studying this subcellular compartment requires tools that are optimized for these conditions. To develop a single-fluorophore genetically encoded calcium indicator for this organelle, we targeted a low affinity variant of GCaMP3 to the ER lumen (GCaMPer (10.19)). A set of viral vectors was constructed to express GCaMPer in human neuroblastoma cells, rat primary cortical neurons, and human induced pluripotent stem cell-derived cardiomyocytes. We observed dynamic changes in GCaMPer (10.19) fluorescence in response to pharmacologic manipulations of the ER calcium store. Additionally, periodic calcium efflux from the ER was observed during spontaneous beating of cardiomyocytes. GCaMPer (10.19) has utility in imaging ER calcium in living cells and providing insight into luminal calcium dynamics under physiologic and pathologic states.

}, issn = {1932-6203}, doi = {10.1371/journal.pone.0139273}, author = {Henderson, Mark J and Baldwin, Heather A and Werley, Christopher A and Boccardo, Stefano and Whitaker, Leslie R and Yan, Xiaokang and Holt, Graham T and Schreiter, Eric R and Looger, Loren L and Cohen, Adam E and Kim, Douglas S and Harvey, Brandon K} } @article {46326, title = {Mapping social behavior-induced brain activation at cellular resolution in the mouse.}, journal = {Cell Reports}, volume = {10}, year = {2015}, month = {2015 Jan 13}, pages = {292-305}, abstract = {

Understanding how brain activation mediates behaviors is a central goal of systems neuroscience. Here, we apply an automated method for mapping brain activation in the mouse in order to probe how sex-specific social behaviors are represented in the male brain. Our method uses the immediate-early-gene c-fos, a marker of neuronal activation, visualized by serial two-photon tomography: the c-fos-GFP+ neurons are computationally detected, their distribution is registered to a reference brain and a brain atlas, and their numbers are analyzed by statistical tests. Our results reveal distinct and shared female and male interaction-evoked patterns of male brain activation representing sex discrimination and social recognition. We also identify brain regions whose degree of activity correlates to specific features of social behaviors and estimate the total numbers and the densities of activated neurons per brain areas. Our study opens the door to automated screening of behavior-evoked brain activation in the mouse.

}, issn = {2211-1247}, doi = {10.1016/j.celrep.2014.12.014}, author = {Kim, Yongsoo and Venkataraju, Kannan Umadevi and Pradhan, Kith and Mende, Carolin and Taranda, Julian and Turaga, Srinivas C and Arganda-Carreras, Ignacio and Ng, Lydia and Hawrylycz, Michael J and Rockland, Kathleen S and Seung, H Sebastian and Osten, Pavel} } @article {47240, title = {Neural Network Model of Memory Retrieval.}, journal = {Frontiers in Computational Neuroscience}, volume = {9}, year = {2015}, month = {2015 Dec 17}, pages = {149}, abstract = {

Human memory can store large amount of information. Nevertheless, recalling is often a challenging task. In a classical free recall paradigm, where participants are asked to repeat a briefly presented list of words, people make mistakes for lists as short as 5 words. We present a model for memory retrieval based on a Hopfield neural network where transition between items are determined by similarities in their long-term memory representations. Meanfield analysis of the model reveals stable states of the network corresponding (1) to single memory representations and (2) intersection between memory representations. We show that oscillating feedback inhibition in the presence of noise induces transitions between these states triggering the retrieval of different memories. The network dynamics qualitatively predicts the distribution of time intervals required to recall new memory items observed in experiments. It shows that items having larger number of neurons in their representation are statistically easier to recall and reveals possible bottlenecks in our ability of retrieving memories. Overall, we propose a neural network model of information retrieval broadly compatible with experimental observations and is consistent with our recent graphical model (Romani et al., 2013).

}, issn = {1662-5188}, doi = {10.3389/fncom.2015.00149}, author = {Recanatesi, Stefano and Katkov, Mikhail and Romani, Sandro and Tsodyks, Misha} } @article {46972, title = {Neuronal representation of ultraviolet visual stimuli in mouse primary visual cortex.}, journal = {Scientific Reports}, volume = {5}, year = {2015}, month = {2015 Jul 29}, pages = {12597}, abstract = {

The mouse has become an important model for understanding the neural basis of visual perception. Although it has long been known that mouse lens transmits ultraviolet (UV) light and mouse opsins have absorption in the UV band, little is known about how UV visual information is processed in the mouse brain. Using a custom UV stimulation system and in vivo calcium imaging, we characterized the feature selectivity of layer 2/3 neurons in mouse primary visual cortex (V1). In adult mice, a comparable percentage of the neuronal population responds to UV and visible stimuli, with similar pattern selectivity and receptive field properties. In young mice, the orientation selectivity for UV stimuli increased steadily during development, but not direction selectivity. Our results suggest that, by expanding the spectral window through which the mouse can acquire visual information, UV sensitivity provides an important component for mouse vision.

}, issn = {2045-2322}, doi = {10.1038/srep12597}, author = {Tan, Zhongchao and Sun, Wenzhi and Chen, Tsai-Wen and Kim, Douglas and Ji, Na} } @article {46816, title = {The nutrient-responsive hormone CCHamide-2 controls growth by regulating insulin-like peptides in the brain of Drosophila melanogaster.}, journal = {PLoS Genetics}, volume = {11}, year = {2015}, month = {2015 May}, pages = {e1005209}, abstract = {

The coordination of growth with nutritional status is essential for proper development and physiology. Nutritional information is mostly perceived by peripheral organs before being relayed to the brain, which modulates physiological responses. Hormonal signaling ensures this organ-to-organ communication, and the failure of endocrine regulation in humans can cause diseases including obesity and diabetes. In Drosophila melanogaster, the fat body (adipose tissue) has been suggested to play an important role in coupling growth with nutritional status. Here, we show that the peripheral tissue-derived peptide hormone CCHamide-2 (CCHa2) acts as a nutrient-dependent regulator of Drosophila insulin-like peptides (Dilps). A BAC-based transgenic reporter revealed strong expression of CCHa2 receptor (CCHa2-R) in insulin-producing cells (IPCs) in the brain. Calcium imaging of brain explants and IPC-specific CCHa2-R knockdown demonstrated that peripheral-tissue derived CCHa2 directly activates IPCs. Interestingly, genetic disruption of either CCHa2 or CCHa2-R caused almost identical defects in larval growth and developmental timing. Consistent with these phenotypes, the expression of dilp5, and the release of both Dilp2 and Dilp5, were severely reduced. Furthermore, transcription of CCHa2 is altered in response to nutritional levels, particularly of glucose. These findings demonstrate that CCHa2 and CCHa2-R form a direct link between peripheral tissues and the brain, and that this pathway is essential for the coordination of systemic growth with nutritional availability. A mammalian homologue of CCHa2-R, Bombesin receptor subtype-3 (Brs3), is an orphan receptor that is expressed in the islet β-cells; however, the role of Brs3 in insulin regulation remains elusive. Our genetic approach in Drosophila melanogaster provides the first evidence, to our knowledge, that bombesin receptor signaling with its endogenous ligand promotes insulin production.

}, issn = {1553-7404}, doi = {10.1371/journal.pgen.1005209}, author = {Sano, Hiroko and Nakamura, Akira and Texada, Michael J and Truman, James W and Ishimoto, Hiroshi and Kamikouchi, Azusa and Nibu, Yutaka and Kume, Kazuhiko and Ida, Takanori and Kojima, Masayasu} } @article {47017, title = {Optogenetics: 10 years after ChR2 in neurons-views from the community.}, journal = {Nature Neuroscience}, volume = {18}, year = {2015}, month = {2015 Aug 26}, pages = {1202-12}, issn = {1546-1726}, doi = {10.1038/nn.4106}, author = {Adamantidis, Antoine and Arber, Silvia and Bains, Jaideep S and Bamberg, Ernst and Bonci, Antonello and Buzs{\'a}ki, Gy{\"o}rgy and Cardin, Jessica A and Costa, Rui M and Dan, Yang and Goda, Yukiko and Graybiel, Ann M and H{\"a}usser, Michael and Hegemann, Peter and Huguenard, John R and Insel, Thomas R and Janak, Patricia H and Johnston, Daniel and Josselyn, Sheena A and Koch, Christof and Kreitzer, Anatol C and L{\"u}scher, Christian and Malenka, Robert C and Miesenb{\"o}ck, Gero and Nagel, Georg and Roska, Botond and Schnitzer, Mark J and Shenoy, Krishna V and Soltesz, Ivan and Sternson, Scott M and Tsien, Richard W and Tsien, Roger Y and Turrigiano, Gina G and Tye, Kay M and Wilson, Rachel I} } @inbook {46570, title = {Population genomic and phylogenomic insights into the evolution of physiology and behaviour in social insects}, booktitle = {Advances in Insect Physiology}, number = {48}, year = {2015}, pages = {293{\textendash}324}, abstract = {

Genomics revolutionized the field of social insect research by providing powerful tools to understand the relationship between genes, physiology and behaviour of social insects. Notably, analysis of gene expression and methylation patterns in the different castes of insect colonies highlighted many genes that likely underlie caste-specific physiological and behavioural phenotypes. However, earlier studies of social insect genomes lacked an {\textquoteleft}evolutionary{\textquoteright} context.\ Out of the millions of DNA bases found in the genome of a social insect, which pieces were most important to fitness over the timescale of social evolution?\ Here, we review a burgeoning body of literature that utilizes between-species or within-species genomic comparisons to highlight the evolutionary forces that have shaped social insect genomes. These pioneering phylogenetic and population genomic studies provide a critically needed evolutionary context to social insect genomes and underscore the importance of adaptive changes in physiology and behaviour in social evolution.

}, doi = {10.1016/bs.aiip.2015.01.002}, author = {Kent, Clement F and Zayed, Amro} } @article {46497, title = {Proper migration and axon outgrowth of zebrafish cranial motoneuron subpopulations require the cell adhesion molecule MDGA2A.}, journal = {Biology Open}, volume = {4}, year = {2015}, month = {2015}, pages = {146-54}, abstract = {

The formation of functional neuronal circuits relies on accurate migration and proper axonal outgrowth of neuronal precursors. On the route to their targets migrating cells and growing axons depend on both, directional information from neurotropic cues and adhesive interactions mediated via extracellular matrix molecules or neighbouring cells. The inactivation of guidance cues or the interference with cell adhesion can cause severe defects in neuronal migration and axon guidance. In this study we have analyzed the function of the MAM domain containing glycosylphosphatidylinositol anchor 2A (MDGA2A) protein in zebrafish cranial motoneuron development. MDGA2A is prominently expressed in distinct clusters of cranial motoneurons, especially in the ones of the trigeminal and facial nerves. Analyses of MDGA2A knockdown embryos by light sheet and confocal microscopy revealed impaired migration and aberrant axonal outgrowth of these neurons; suggesting that adhesive interactions mediated by MDGA2A are required for the proper arrangement and outgrowth of cranial motoneuron subtypes.

}, issn = {2046-6390}, doi = {10.1242/bio.20148482}, author = {Ingold, Esther and Vom Berg-Maurer, Colette M and Burckhardt, Christoph J and Lehnherr, Andr{\'e} and Rieder, Philip and Keller, Philipp J and Stelzer, Ernst H and Greber, Urs F and Neuhauss, Stephan C F and Gesemann, Matthias} } @article {46499, title = {The Release 6 reference sequence of the Drosophila melanogaster genome.}, journal = {Genome Research}, volume = {25}, year = {2015}, month = {2015 Mar}, pages = {445-58}, abstract = {

Drosophila melanogaster plays an important role in molecular, genetic, and genomic studies of heredity, development, metabolism, behavior, and human disease. The initial reference genome sequence reported more than a decade ago had a profound impact on progress in Drosophila research, and improving the accuracy and completeness of this sequence continues to be important to further progress. We previously described improvement of the 117-Mb sequence in the euchromatic portion of the genome and 21 Mb in the heterochromatic portion, using a whole-genome shotgun assembly, BAC physical mapping, and clone-based finishing. Here, we report an improved reference sequence of the single-copy and middle-repetitive regions of the genome, produced using cytogenetic mapping to mitotic and polytene chromosomes, clone-based finishing and BAC fingerprint verification, ordering of scaffolds by alignment to cDNA sequences, incorporation of other map and sequence data, and validation by whole-genome optical restriction mapping. These data substantially improve the accuracy and completeness of the reference sequence and the order and orientation of sequence scaffolds into chromosome arm assemblies. Representation of the Y chromosome and other heterochromatic regions is particularly improved. The new 143.9-Mb reference sequence, designated Release 6, effectively exhausts clone-based technologies for mapping and sequencing. Highly repeat-rich regions, including large satellite blocks and functional elements such as the ribosomal RNA genes and the centromeres, are largely inaccessible to current sequencing and assembly methods and remain poorly represented. Further significant improvements will require sequencing technologies that do not depend on molecular cloning and that produce very long reads.

}, issn = {1549-5469}, doi = {10.1101/gr.185579.114}, author = {Hoskins, Roger A and Carlson, Joseph W and Wan, Kenneth H and Park, Soo and Mendez, Ivonne and Galle, Samuel E and Booth, Benjamin W and Pfeiffer, Barret D and George, Reed A and Svirskas, Robert and Krzywinski, Martin and Schein, Jacqueline and Accardo, Maria Carmela and Damia, Elisabetta and Messina, Giovanni and Mendez-Lago, Maria and de Pablos, Beatriz and Demakova, Olga V and Andreyeva, Evgeniya N and Boldyreva, Lidiya V and Marra, Marco and Carvalho, A Bernardo and Dimitri, Patrizio and Villasante, Alfredo and Zhimulev, Igor F and Rubin, Gerald M and Karpen, Gary H and Celniker, Susan E} } @article {47956, title = {Robust estimation of self-exciting point process models with application to neuronal modeling.}, journal = {CoRR}, volume = {abs/1507.03955}, year = {2015}, abstract = {

We consider the problem of estimating discrete selfexciting point process models from limited binary observations, where the history of the process serves as the covariate. We analyze the performance of two classes of estimators, namely the {\textquoteleft}1-regularized maximum likelihood and greedy estimators, for a canonical self-exciting point process and characterize the sampling tradeoffs required for stable recovery in the non-asymptotic regime. Our results extend those of compressed sensing for linear and generalized linear models with i.i.d. covariates to point processes with highly inter-dependent covariates. We further provide simulation studies as well as application to real spiking data from mouse{\textquoteright}s lateral geniculate nucleus and ferret{\textquoteright}s retinal ganglion cells which agree with our theoretical predictions.

}, url = {http://arxiv.org/abs/1507.03955}, author = {Abbas Kazemipour and Min Wu and Behtash Babadi} } @article {46454, title = {A sensitive and robust enzyme kinetic experiment using microplates and fluorogenic ester substrates}, journal = {Journal of Chemical Education}, volume = {92}, year = {2015}, month = {2015 Feb}, pages = {385-8}, abstract = {

Enzyme kinetics measurements are a standard component of undergraduate biochemistry laboratories. The combination of serine hydrolases and fluorogenic enzyme substrates provides a rapid, sensitive, and general method for measuring enzyme kinetics in an undergraduate biochemistry laboratory. In this method, the kinetic activity of multiple protein variants is determined in parallel using a microplate reader, multichannel pipets, serial dilutions, and fluorogenic ester substrates. The utility of this methodology is illustrated by the measurement of differential enzyme activity in microplate volumes in triplicate with small protein samples and low activity enzyme variants. Enzyme kinetic measurements using fluorogenic substrates are, thus, adaptable for use with student-purified enzyme variants and for comparative enzyme kinetics studies. The rapid setup and analysis of these kinetic experiments not only provides advanced undergraduates with experience in a fundamental biochemical technique, but also provides the adaptability for use in inquiry-based laboratories.

}, issn = {0021-9584}, doi = {10.1021/ed500452f}, author = {Johnson, R. Jeremy and Hoops, Geoffrey C. and Savas, Christopher J. and Kartje, Zachary and Lavis, Luke D} } @article {46187, title = {Sensory determinants of behavioral dynamics in Drosophila thermotaxis.}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {112}, year = {2015}, month = {2015 Jan 13}, pages = {E220-9}, abstract = {

Complex animal behaviors are built from dynamical relationships between sensory inputs, neuronal activity, and motor outputs in patterns with strategic value. Connecting these patterns illuminates how nervous systems compute behavior. Here, we study Drosophila larva navigation up temperature gradients toward preferred temperatures (positive thermotaxis). By tracking the movements of animals responding to fixed spatial temperature gradients or random temperature fluctuations, we calculate the sensitivity and dynamics of the conversion of thermosensory inputs into motor responses. We discover three thermosensory neurons in each dorsal organ ganglion (DOG) that are required for positive thermotaxis. Random optogenetic stimulation of the DOG thermosensory neurons evokes behavioral patterns that mimic the response to temperature variations. In vivo calcium and voltage imaging reveals that the DOG thermosensory neurons exhibit activity patterns with sensitivity and dynamics matched to the behavioral response. Temporal processing of temperature variations carried out by the DOG thermosensory neurons emerges in distinct motor responses during thermotaxis.

}, issn = {1091-6490}, doi = {10.1073/pnas.1416212112}, author = {Klein, Mason and Afonso, Bruno and Vonner, Ashley J and Hernandez-Nunez, Luis and Berck, Matthew and Tabone, Christopher J and Kane, Elizabeth A and Pieribone, Vincent A and Nitabach, Michael N and Cardona, Albert and Zlatic, Marta and Sprecher, Simon G and Gershow, Marc and Garrity, Paul A and Samuel, Aravinthan D T} } @article {46700, title = {Specific interaction of KIF11 with ZBP1 regulates the transport of β-actin mRNA and cell motility.}, journal = {Journal of Cell Science}, volume = {128}, year = {2015}, month = {2015 Mar 1}, pages = {1001-10}, abstract = {

ZBP1-modulated localization of β-actin mRNA enables a cell to establish polarity and structural asymmetry. Although the mechanism of β-actin mRNA localization has been well established, the underlying mechanism of how a specific molecular motor contributes to the transport of the ZBP1 (also known as IGF2BP1) complex in non-neuronal cells remains elusive. In this study, we report the isolation and identification of KIF11, a microtubule motor, which physically interacts with ZBP1 and is a component of β-actin messenger ribonucleoprotein particles (mRNPs). We show that KIF11 colocalizes with the β-actin mRNA, and the ability of KIF11 to transport β-actin mRNA is dependent on ZBP1. We characterize the corresponding regions of ZBP1 and KIF11 that mediate the interaction of the two proteins in vitro and in vivo. Disruption of the in vivo interaction of KIF11 with ZBP1 delocalizes β-actin mRNA and affects cell migration. Our study reveals a molecular mechanism by which a particular microtubule motor mediates the transport of an mRNP through direct interaction with an mRNA-binding protein.

}, issn = {1477-9137}, doi = {10.1242/jcs.161679}, author = {Song, Tingting and Zheng, Yi and Wang, Yarong and Katz, Zachary and Liu, Xin and Chen, Shaoying and Singer, Robert H and Gu, Wei} } @article {47133, title = {Stochastic electrotransport selectively enhances the transport of highly electromobile molecules.}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {112}, year = {2015}, month = {2015 Nov 2}, pages = {E6274-83}, abstract = {

Nondestructive chemical processing of porous samples such as fixed biological tissues typically relies on molecular diffusion. Diffusion into a porous structure is a slow process that significantly delays completion of chemical processing. Here, we present a novel electrokinetic method termed stochastic electrotransport for rapid nondestructive processing of porous samples. This method uses a rotational electric field to selectively disperse highly electromobile molecules throughout a porous sample without displacing the low-electromobility molecules that constitute the sample. Using computational models, we show that stochastic electrotransport can rapidly disperse electromobile molecules in a porous medium. We apply this method to completely clear mouse organs within 1{\textendash}3 days and to stain them with nuclear dyes, proteins, and antibodies within 1 day. Our results demonstrate the potential of stochastic electrotransport to process large and dense tissue samples that were previously infeasible in time when relying on diffusion.

}, doi = {10.1073/pnas.1510133112}, author = {Kim, Sung-Yon and Cho, Jae Hun and Murray, Evan and Bakh, Naveed and Choi, Heejin and Ohn, Kimberly and Ruelas, Luzdary and Hubbert, Austin and McCue, Meg and Vassallo, Sara L and Keller, Philipp J and Chung, Kwanghun} } @article {46886, title = {Structural basis for the antipolymer activity of Hb ζ22βsζ2βs2 trapped in a tense conformation.}, journal = {Journal of Molecular Structure}, volume = {1099}, year = {2015}, month = {2015 Nov}, pages = {99-107}, abstract = {

The phenotypical severity of sickle cell disease (SCD) can be mitigated by modifying mutant hemoglobin S (Hb S, Hb α2β2s) to contain embryonic ζ globin in place of adult α-globin subunits (Hb ζ2β2s). Crystallographical analyses of liganded Hb ζζ2β2s, though, demonstrate a tense (T-state) quaternary structure that paradoxically predicts its participation in--rather than its exclusion from--pathological deoxyHb S polymers. We resolved this structure-function conundrum by examining the effects of α\ {\textrightarrow}\ ζ exchange on the characteristics of specific amino acids that mediate sickle polymer assembly. Superposition analyses of the βs subunits of T-state deoxyHb α2β2s and T-state CO-liganded Hb ζ2β2s reveal significant displacements of both mutant βsVal6 and conserved β-chain contact residues, predicting weakening of corresponding polymer-stabilizing interactions. Similar comparisons of the α- and ζ-globin subunits implicate four amino acids that are either repositioned or undergo non-conservative substitution, abrogating critical polymer contacts. CO-Hb ζ2βs2 additionally exhibits a unique trimer-of-heterotetramers crystal packing that is sustained by novel intermolecular interactions involving the pathological βsVal6, contrasting sharply with the classical double-stranded packing of deoxyHb S. Finally, the unusually large buried solvent-accessible surface area for CO-Hb ζ2β2s suggests that it does not co-assemble with deoxyHb S in\ vivo \ . In sum, the antipolymer activities of Hb ζ2β2s appear to arise from both repositioning and replacement of specific α- and βs-chain residues, favoring an alternate T-state solution structure that is excluded from pathological deoxyHb S polymers. These data account for the antipolymer activity of Hb ζ2β2s, and recommend the utility of SCD therapeutics that capitalize on α-globin exchange strategies.

}, issn = {0022-2860}, doi = {10.1016/j.molstruc.2015.06.047}, author = {Safo, Martin K and Ko, Tzu-Ping and Schreiter, Eric R and Russell, J. Eric} } @article {46923, title = {Structure of a designed tetrahedral protein assembly variant engineered to have improved soluble expression.}, journal = {Protein Science}, volume = {24}, year = {2015}, month = {2015 Jul 15}, pages = {1695-701}, abstract = {

We recently reported the development of a computational method for the design of co-assembling, multi-component protein nanomaterials. While four such materials were validated at high-resolution by X-ray crystallography, low yield of soluble protein prevented X-ray structure determination of a fifth designed material, T33-09. Here we report the design and crystal structure of T33-31, a variant of T33-09 with improved soluble yield resulting from redesign efforts focused on mutating solvent-exposed side chains to charged amino acids. The structure is found to match the computational design model with atomic-level accuracy, providing further validation of the design approach and demonstrating a simple and potentially general means of improving the yield of designed protein nanomaterials. This article is protected by copyright. All rights reserved.

}, issn = {1469-896X}, doi = {10.1002/pro.2748}, author = {Bale, Jacob B and Park, Rachel U and Liu, Yuxi and Gonen, Shane and Gonen, Tamir and Cascio, Duilio and King, Neil P and Yeates, Todd O and Baker, David} } @article {47039, title = {A subset of serotonergic neurons evokes hunger in adult Drosophila.}, journal = {Current Biology : CB}, volume = {25}, year = {2015}, month = {2015 Sep 2}, pages = {2435-40}, abstract = {

Hunger is a complex motivational state that drives multiple behaviors. The sensation of hunger is caused by an imbalance between energy intake and expenditure. One immediate response to hunger is increased food consumption. Hunger also modulates behaviors related to food seeking such as increased locomotion and enhanced sensory sensitivity in both insects [1-5] and vertebrates [6, 7]. In addition, hunger can promote the expression of food-associated memory [8, 9]. Although progress is being made [10], how hunger is represented in the brain and how it coordinates these behavioral responses is not fully understood in any system. Here, we use Drosophila melanogaster to identify neurons encoding hunger. We found a small group of neurons that, when activated, induced a fed fly to eat as though it were starved, suggesting that these neurons are downstream of the metabolic regulation of hunger. Artificially activating these neurons also promotes appetitive memory performance in sated flies, indicating that these neurons are not simply feeding command neurons but likely play a more general role in encoding hunger. We determined that the neurons relevant for the feeding effect are serotonergic and project broadly within the brain, suggesting a possible mechanism for how various responses to hunger are coordinated. These findings extend our\ understanding of the neural circuitry that drives feeding and enable future exploration of how state influences neural activity within this circuit.

}, issn = {1879-0445}, doi = {10.1016/j.cub.2015.08.005}, author = {Albin, Stephanie D and Kaun, Karla R and Knapp, Jon-Michael and Chung, Phuong and Heberlein, Ulrike and Simpson, Julie H} } @article {47152, title = {Synaptic circuits and their variations within different columns in the visual system of Drosophila.}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {112}, year = {2015}, month = {2015 Nov 3}, pages = {13711-6}, abstract = {

We reconstructed the synaptic circuits of seven columns in the second neuropil or medulla behind the fly{\textquoteright}s compound eye. These neurons embody some of the most stereotyped circuits in one of the most miniaturized of animal brains. The reconstructions allow us, for the first time to our knowledge, to study variations between circuits in the medulla{\textquoteright}s neighboring columns. This variation in the number of synapses and the types of their synaptic partners has previously been little addressed because methods that visualize multiple circuits have not resolved detailed connections, and existing connectomic studies, which can see such connections, have not so far examined multiple reconstructions of the same circuit. Here, we address the omission by comparing the circuits common to all seven columns to assess variation in their connection strengths and the resultant rates of several different and distinct types of connection error. Error rates reveal that, overall, \<1\% of contacts are not part of a consensus circuit, and we classify those contacts that supplement (E+) or are missing from it (E-). Autapses, in which the same cell is both presynaptic and postsynaptic at the same synapse, are occasionally seen; two cells in particular, Dm9 and Mi1, form >=20-fold more autapses than do other neurons. These results delimit the accuracy of developmental events that establish and normally maintain synaptic circuits with such precision, and thereby address the operation of such circuits. They also establish a precedent for error rates that will be required in the new science of connectomics.

}, issn = {1091-6490}, doi = {10.1073/pnas.1509820112}, author = {Takemura, Shin-ya and Xu, C Shan and Lu, Zhiyuan and Rivlin, Patricia K and Parag, Toufiq and Olbris, Donald J and Plaza, Stephen and Zhao, Ting and Katz, William T and Umayam, Lowell and Weaver, Charlotte and Hess, Harald F and Horne, Jane Anne and Nunez-Iglesias, Juan and Aniceto, Roxanne and Chang, Lei-Ann and Lauchie, Shirley and Nasca, Ashley and Ogundeyi, Omotara and Sigmund, Christopher and Takemura, Satoko and Tran, Julie and Langille, Carlie and Le Lacheur, Kelsey and McLin, Sari and Shinomiya, Aya and Chklovskii, Dmitri B and Meinertzhagen, Ian A and Scheffer, Louis K} } @article {46887, title = {Synaptic Orb2A bridges memory acquisition and late memory consolidation in Drosophila.}, journal = {Cell Reports}, volume = {11}, year = {2015}, month = {2015 Jun 30}, pages = {1953-65}, abstract = {

To adapt to an ever-changing environment, animals consolidate some, but not all, learning experiences to long-term memory. In mammals, long-term memory consolidation often involves neural pathway reactivation hours after memory acquisition. It is not known whether this delayed-reactivation schema is\ common across the animal kingdom or how information is stored during the delay period. Here, we show\ that, during courtship suppression learning, Drosophila exhibits delayed long-term memory consolidation. We also show that the same class of dopaminergic neurons engaged earlier in memory acquisition is also both necessary and sufficient for delayed long-term memory consolidation. Furthermore, we present evidence that, during learning, the translational regulator Orb2A tags specific synapses of mushroom body neurons for later consolidation. Consolidation involves the subsequent recruitment of Orb2B and the activity-dependent synthesis of CaMKII. Thus, our results provide evidence for the role of a neuromodulated, synapse-restricted molecule bridging memory acquisition and long-term memory consolidation in a learning animal.

}, issn = {2211-1247}, doi = {10.1016/j.celrep.2015.05.037}, author = {Kr{\"u}ttner, Sebastian and Traunm{\"u}ller, Lisa and Dag, Ugur and Jandrasits, Katharina and Stepien, Barbara and Iyer, Nirmala and Fradkin, Lee G and Noordermeer, Jasprina N and Mensh, Brett D and Keleman, Krystyna} } @article {46697, title = {Synonymous modification results in high-fidelity gene expression of repetitive protein and nucleotide sequences.}, journal = {Genes \& Development}, volume = {29}, year = {2015}, month = {2015 Apr 15}, pages = {876-86}, abstract = {

Repetitive nucleotide or amino acid sequences are often engineered into probes and biosensors to achieve functional readouts and robust signal amplification. However, these repeated sequences are notoriously prone to aberrant deletion and degradation, impacting the ability to correctly detect and interpret biological functions. Here, we introduce a facile and generalizable approach to solve this often unappreciated problem by modifying the nucleotide sequences of the target mRNA to make them nonrepetitive but still functional ("synonymous"). We first demonstrated the procedure by designing a cassette of synonymous MS2 RNA motifs and tandem coat proteins for RNA imaging and showed a dramatic improvement in signal and reproducibility in single-RNA detection in live cells. The same approach was extended to enhancing the stability of engineered fluorescent biosensors containing a fluorescent resonance energy transfer (FRET) pair of fluorescent proteins on which a great majority of systems thus far in the field are based. Using the synonymous modification to FRET biosensors, we achieved correct expression of full-length sensors, eliminating the aberrant truncation products that often were assumed to be due to nonspecific proteolytic cleavages. Importantly, the biological interpretations of the sensor are significantly different when a correct, full-length biosensor is expressed. Thus, we show here a useful and generally applicable method to maintain the integrity of expressed genes, critical for the correct interpretation of probe readouts.

}, issn = {1549-5477}, doi = {10.1101/gad.259358.115}, author = {Wu, Bin and Miskolci, Veronika and Sato, Hanae and Tutucci, Evelina and Kenworthy, Charles A and Donnelly, Sara K and Yoon, Young J and Cox, Dianne and Singer, Robert H and Hodgson, Louis} } @article {47242, title = {Synthesis of a far-red photoactivatable silicon-containing rhodamine for super-resolution microscopy.}, journal = {Angewandte Chemie (International ed. in English)}, volume = {55}, year = {2015}, month = {2015 Dec 11}, pages = {1723-7}, abstract = {

The rhodamine system is a flexible framework for building small-molecule fluorescent probes. Changing N-substitution patterns and replacing the xanthene oxygen with a dimethylsilicon moiety can shift the absorption and fluorescence emission maxima of rhodamine dyes to longer wavelengths. Acylation of the rhodamine nitrogen atoms forces the molecule to adopt a nonfluorescent lactone form, providing a convenient method to make fluorogenic compounds. Herein, we take advantage of all of these structural manipulations and describe a novel photoactivatable fluorophore based on a Si-containing analogue of Q-rhodamine. This probe is the first example of a "caged" Si-rhodamine, exhibits higher photon counts compared to established localization microscopy dyes, and is sufficiently red-shifted to allow multicolor imaging. The dye is a useful label for super-resolution imaging and constitutes a new scaffold for far-red fluorogenic molecules.

}, issn = {1521-3773}, doi = {10.1002/anie.201509649}, author = {Grimm, Jonathan B and Klein, Teresa and Kopek, Benjamin G and Shtengel, Gleb and Hess, Harald F and Sauer, Markus and Lavis, Luke D} } @article {46787, title = {Systematic imaging reveals features and changing localization of mRNAs in Drosophila development.}, journal = {Elife}, volume = {4}, year = {2015}, month = {2015}, abstract = {

mRNA localization is critical for eukaryotic cells and affects numerous transcripts, yet how cells regulate distribution of many mRNAs to their subcellular destinations is still unknown. We combined transcriptomics and systematic imaging to determine the tissue-specific expression and subcellular distribution of 5862 mRNAs during Drosophila oogenesis. mRNA localization is widespread in the ovary and detectable in all of its cell types-the somatic epithelial, the nurse cells, and the oocyte. Genes defined by a common RNA localization share distinct gene features and differ in expression level, 3{\textquoteright}UTR length and sequence conservation from unlocalized mRNAs. Comparison of mRNA localizations in different contexts revealed that localization of individual mRNAs changes over time in the oocyte and between ovarian and embryonic cell types. This genome scale image-based resource (Dresden Ovary Table, DOT, http://tomancak-srv1.mpi-cbg.de/DOT/main.html) enables the transition from mechanistic dissection of singular mRNA localization events towards global understanding of how mRNAs transcribed in the nucleus distribute in cells.

}, issn = {2050-084X}, doi = {10.7554/eLife.05003}, author = {Jambor, Helena and Surendranath, Vineeth and Kalinka, Alex T and Mejstrik, Pavel and Saalfeld, Stephan and Tomancak, Pavel} } @article {47025, title = {The transgenic RNAi project at Harvard Medical School: resources and validation.}, journal = {Genetics}, volume = {201}, year = {2015}, month = {2015 Aug 28}, pages = {843-52}, abstract = {

To facilitate large scale functional studies in Drosophila, the Drosophila Transgenic RNAi Project (TRiP) at Harvard Medical School (HMS) was established along with several goals: developing efficient vectors for RNAi that work in all tissues, generating a genome scale collection of RNAi stocks with input from the community, distributing the lines as they are generated through existing stock centers, validating as many lines as possible using RT-qPCR and phenotypic analyses, and developing tools and web resources for identifying RNAi lines and retrieving existing information on their quality. With these goals in mind, here we describe in detail the various tools we developed and the status of the collection, which is currently comprised of 11,491 lines and covering 71\% of Drosophila genes. Data on the characterization of the lines either by RT-qPCR or phenotype is available on a dedicated web site, the RNAi Stock Validation and Phenotypes Project (RSVP; www.flyrnai.org/RSVP.html), and stocks are available from three stock centers, the Bloomington Drosophila Stock Center (USA), National Institute of Genetics (Japan), and TsingHua Fly Center (China).

}, issn = {1943-2631}, doi = {10.1534/genetics.115.180208}, author = {Perkins, L A and Holderbaum, L and Tao, R and Hu, Y and Sopko, R and McCall, K and Yang-Zhou, D and Flockhart, I and Binari, R and Shim, H-S and Miller, A and Housden, A and Foos, M and Randkelv, S and Kelley, C and Namgyal, P and Villalta, C and Liu, L-P and Jiang, X and Huan-Huan, Q and Xia, W and Fujiyama, A and Toyoda, A and Ayers, K and Blum, A and Czech, B and Neumuller, R and Yan, D and Cavallaro, A and Hibbard, K and Hall, D and Cooley, L and Hannon, G J and Lehmann, R and Parks, A and Mohr, S E and Ueda, R and Kondo, S and Ni, J-Q and Perrimon, Norbert} } @article {46325, title = {Ultrastructurally smooth thick partitioning and volume stitching for large-scale connectomics.}, journal = {Nature Methods}, volume = {12}, year = {2015}, month = {2015 Feb 16}, pages = {319-22}, abstract = {

Focused-ion-beam scanning electron microscopy (FIB-SEM) has become an essential tool for studying neural tissue at resolutions below 10 nm {\texttimes} 10 nm {\texttimes} 10 nm, producing data sets optimized for automatic connectome tracing. We present a technical advance, ultrathick sectioning, which reliably subdivides embedded tissue samples into chunks (20 μm thick) optimally sized and mounted for efficient, parallel FIB-SEM imaging. These chunks are imaged separately and then {\textquoteright}volume stitched{\textquoteright} back together, producing a final three-dimensional data set suitable for connectome tracing.

}, issn = {1548-7105}, doi = {10.1038/nmeth.3292}, author = {Hayworth, Kenneth J and Xu, C Shan and Lu, Zhiyuan and Knott, Graham W and Fetter, Richard D and Tapia, Juan Carlos and Lichtman, Jeff W and Hess, Harald F} } @proceedings {46519, title = {Understanding classifier errors by examining influential neighbors.}, year = {2015}, month = {06/2015}, publisher = {IEEE}, address = {Boston, MA}, abstract = {

Modern supervised learning algorithms can learn very accurate and complex discriminating functions. But when these classifiers fail, this complexity can also be a drawback because there is no easy, intuitive way to diagnose why they are failing and remedy the problem. This important question has received little attention. To address this problem, we propose a novel method to analyze and understand a classifier{\textquoteright}s errors. Our method centers around a measure of how much influence a training example has on the classifier{\textquoteright}s prediction for a test example. To understand why a classifier is\ mispredicting\ the label of a given test example, the user can find and review the most influential training examples that caused this\ misprediction, allowing them to focus their attention on relevant areas of the data space. This will aid the user in determining if and how the training data is inconsistently labeled or lacking in diversity, or if the feature representation is insufficient. As computing the influence of each training example is computationally impractical, we propose a novel distance metric to approximate influence for boosting classifiers that is fast enough to be used interactively. We also show several novel use paradigms of our distance metric. Through experiments, we show that it can be used to find incorrectly or inconsistently labeled training examples, to find specific areas of the data space that need more training data, and to gain insight into which features are missing from the current representation.\ 

Code is available at\ https://github.com/kristinbranson/InfluentialNeighbors.

}, author = {Mayank Kabra and Alice A. Robie and Kristin Branson} } @article {46922, title = {Vinculin is required for cell polarization, migration, and extracellular matrix remodeling in 3D collagen.}, journal = {FASEB Journal}, volume = {29}, year = {2015}, month = {2015 Nov}, pages = {4555-67}, abstract = {

Vinculin is filamentous (F)-actin-binding protein enriched in integrin-based adhesions to the extracellular matrix (ECM). Whereas studies in 2-dimensional (2D) tissue culture models have suggested that vinculin negatively regulates cell migration by promoting cytoskeleton-ECM coupling to strengthen and stabilize adhesions, its role in regulating cell migration in more physiologic, 3-dimensional (3D) environments is unclear. To address the role of vinculin in 3D cell migration, we analyzed the morphodynamics, migration, and ECM remodeling of primary murine embryonic fibroblasts (MEFs) with cre/loxP-mediated vinculin gene disruption in 3D collagen I cultures. We found that vinculin promoted 3D cell migration by increasing directional persistence. Vinculin was necessary for persistent cell protrusion, cell elongation, and stable cell orientation in 3D collagen, but was dispensable for lamellipodia formation, suggesting that vinculin-mediated cell adhesion to the ECM is needed to convert actin-based cell protrusion into persistent cell shape change and migration. Consistent with this finding, vinculin was necessary for efficient traction force generation in 3D collagen without affecting myosin II activity and promoted 3D collagen fiber alignment and macroscopical gel contraction. Our results suggest that vinculin promotes directionally persistent cell migration and tension-dependent ECM remodeling in complex 3D environments by increasing cell-ECM adhesion and traction force generation.-Thievessen, I., Fakhri, N., Steinwachs, J., Kraus, V., McIsaac, R. S., Gao, L., Chen, B.-C., Baird, M. A., Davidson, M. W., Betzig, E., Oldenbourg, R., Waterman, C., M., Fabry, B. Vinculin is required for cell polarization, migration, and extracellular matrix remodeling in 3D collagen.

}, issn = {1530-6860}, doi = {10.1096/fj.14-268235}, author = {Thievessen, Ingo and Fakhri, Nikta and Steinwachs, Julian and Kraus, Viola and McIsaac, R Scott and Gao, Liang and Chen, Bi-Chang and Baird, Michelle A and Davidson, Michael W and Betzig, Eric and Oldenbourg, Rudolf and Waterman, Clare M and Fabry, Ben} } @article {46020, title = {Visualizing whole-brain activity and development at the single-cell level using light-sheet microscopy.}, journal = {Neuron}, volume = {85}, year = {2015}, month = {2015 Feb 4}, pages = {462-83}, abstract = {

The nature of nervous system function and development is inherently global, since all components eventually influence one another. Networks communicate through dense synaptic, electric, and modulatory connections and develop through concurrent growth and interlinking of their neurons, processes, glia, and blood vessels. These factors drive the development of techniques capable of imaging neural signaling, anatomy, and developmental processes at ever-larger scales. Here, we discuss the nature of questions benefitting from large-scale imaging techniques and introduce recent applications. We focus on emerging light-sheet microscopy approaches, which are well suited for live imaging of large systems with high spatiotemporal resolution and over long periods of time. We also discuss computational methods suitable for extracting biological information from the resulting system-level image data sets. Together with new tools for reporting and manipulating neuronal activity and gene expression, these techniques promise new insights into the large-scale function and development of neural systems.

}, issn = {1097-4199}, doi = {10.1016/j.neuron.2014.12.039}, author = {Keller, Philipp J and Ahrens, Misha B} } @inbook {47184, title = {Who is talking to whom: Synaptic partner detection in anisotropic volumes of insect brain.}, booktitle = {Medical Image Computing and Computer-Assisted Intervention -- MICCAI 2015}, number = {9349}, year = {2015}, pages = {661-8}, abstract = {

Automated reconstruction of neural connectivity graphs from electron microscopy image stacks is an essential step towards large-scale neural circuit mapping. While significant progress has recently been made in automated segmentation of neurons and detection of synapses, the problem of synaptic partner assignment for polyadic (one-to-many) synapses, prevalent in the\ Drosophila\ brain, remains unsolved. In this contribution, we propose a method which automatically assigns pre- and postsynaptic roles to neurites adjacent to a synaptic site. The method constructs a probabilistic graphical model over potential synaptic partner pairs which includes factors to account for a high rate of one-to-many connections, as well as the possibility of the same neuron to be pre-synaptic in one synapse and post-synaptic in another. The algorithm has been validated on a publicly available stack of ssTEM images of\ Drosophila\ neural tissue and has been shown to reconstruct most of the synaptic relations correctly.

}, doi = {10.1007/978-3-319-24553-9_81}, author = {Kreshuk, Anna and Funke, Jan and Cardona, Albert and Hamprecht, Fred A} } @article {47116, title = {Whole-animal functional and developmental imaging with isotropic spatial resolution}, journal = {Nature Methods}, volume = {12}, year = {2015}, month = {2015 Oct 26}, pages = {1171-8}, abstract = {

Imaging fast cellular dynamics across large specimens requires high resolution in all dimensions, high imaging speeds, good physical coverage and low photo-damage. To meet these requirements, we developed isotropic multiview (IsoView) light-sheet microscopy, which rapidly images large specimens via simultaneous light-sheet illumination and fluorescence detection along four orthogonal directions. Combining these four views by means of high-throughput multiview deconvolution yields images with high resolution in all three dimensions. We demonstrate whole-animal functional imaging of Drosophila larvae at a spatial resolution of 1.1-2.5 μm and temporal resolution of 2 Hz for several hours. We also present spatially isotropic whole-brain functional imaging in Danio rerio larvae and spatially isotropic multicolor imaging of fast cellular dynamics across gastrulating Drosophila embryos. Compared with conventional light-sheet microscopy, IsoView microscopy improves spatial resolution at least sevenfold and decreases resolution anisotropy at least threefold. Compared with existing high-resolution light-sheet techniques, IsoView microscopy effectively doubles the penetration depth and provides subsecond temporal resolution for specimens 400-fold larger than could previously be imaged.

}, doi = {10.1038/nmeth.3632}, author = {Chhetri, Raghav K and Amat, Fernando and Wan, Yinan and H{\"o}ckendorf, Burkhard and Lemon, William C and Keller, Philipp J} } @article {46984, title = {Whole-central nervous system functional imaging in larval Drosophila.}, journal = {Nature Communications}, volume = {6}, year = {2015}, month = {2015 Aug 11}, pages = {7924}, abstract = {

Understanding how the brain works in tight concert with the rest of the central nervous system (CNS) hinges upon knowledge of coordinated activity patterns across the whole CNS. We present a method for measuring activity in an entire, non-transparent CNS with high spatiotemporal resolution. We combine a light-sheet microscope capable of simultaneous multi-view imaging at volumetric speeds 25-fold faster than the state-of-the-art, a whole-CNS imaging assay for the isolated Drosophila larval CNS and a computational framework for analysing multi-view, whole-CNS calcium imaging data. We image both brain and ventral nerve cord, covering the entire CNS at 2 or 5 Hz with two- or one-photon excitation, respectively. By mapping network activity during fictive behaviours and quantitatively comparing high-resolution whole-CNS activity maps across individuals, we predict functional connections between CNS regions and reveal neurons in the brain that identify type and temporal state of motor programs executed in the ventral nerve cord.

}, doi = {10.1038/ncomms8924}, author = {Lemon, William C and Pulver, Stefan R and H{\"o}ckendorf, Burkhard and McDole, Katie and Branson, Kristin M and Freeman, Jeremy and Keller, Philipp J} } @article {45180, title = {Accurate design of co-assembling multi-component protein nanomaterials.}, journal = {Nature}, volume = {510}, year = {2014}, month = {2014 Jun 5}, pages = {103-8}, abstract = {

The self-assembly of proteins into highly ordered nanoscale architectures is a hallmark of biological systems. The sophisticated functions of these molecular machines have inspired the development of methods to engineer self-assembling protein nanostructures; however, the design of multi-component protein nanomaterials with high accuracy remains an outstanding challenge. Here we report a computational method for designing protein nanomaterials in which multiple copies of two distinct subunits co-assemble into a specific architecture. We use the method to design five 24-subunit cage-like protein nanomaterials in two distinct symmetric architectures and experimentally demonstrate that their structures are in close agreement with the computational design models. The accuracy of the method and the number and variety of two-component materials that it makes accessible suggest a route to the construction of functional protein nanomaterials tailored to specific applications.

}, keywords = {Computer Simulation, Crystallography, X-Ray, Drug Design, Models, Molecular, Nanostructures, Protein Subunits, Proteins}, issn = {1476-4687}, doi = {10.1038/nature13404}, author = {King, Neil P and Bale, Jacob B and Sheffler, William and McNamara, Dan E and Gonen, Shane and Gonen, Tamir and Yeates, Todd O and Baker, David} } @article {48164, title = {Active graph matching for automatic joint segmentation and annotation of C. elegans.}, journal = {Medical image computing and computer-assisted intervention : MICCAI ... International Conference on Medical Image Computing and Computer-Assisted Intervention}, volume = {17}, year = {2014}, month = {2014}, pages = {81-8}, abstract = {

In this work we present a novel technique we term active graph matching, which integrates the popular active shape model into a sparse graph matching problem. This way we are able to combine the benefits of a global, statistical deformation model with the benefits of a local deformation model in form of a second-order random field. We present a new iterative energy minimization technique which achieves empirically good results. This enables us to exceed state-of-the art results for the task of annotating nuclei in 3D microscopic images of C. elegans. Furthermore with the help of the generalized Hough transform we are able to jointly segment and annotate a large set of nuclei in a fully automatic fashion for the first time.

}, keywords = {Algorithms, Animals, Caenorhabditis elegans, Cell Nucleus, Image Enhancement, Image Interpretation, Computer-Assisted, Microscopy, Pattern Recognition, Automated, Reproducibility of Results, Sensitivity and Specificity, Subtraction Technique}, author = {Kainmueller, Dagmar and Jug, Florian and Rother, Carsten and Myers, Gene} } @article {46049, title = {Adaptation to background light enables contrast coding at rod bipolar cell synapses.}, journal = {Neuron}, volume = {81}, year = {2014}, month = {2014 Jan 22}, pages = {388-401}, abstract = {

Rod photoreceptors contribute to vision over an \~{} 6-log-unit range of light intensities. The wide dynamic range of rod vision is thought to depend upon light intensity-dependent switching between two parallel pathways linking rods to ganglion cells: a rod {\textrightarrow} rod bipolar (RB) cell pathway that operates at dim backgrounds and a rod {\textrightarrow} cone {\textrightarrow} cone bipolar cell pathway that operates at brighter backgrounds. We evaluated this conventional model of rod vision by recording rod-mediated light responses from ganglion and AII amacrine cells and by recording RB-mediated synaptic currents from AII amacrine cells in mouse retina. Contrary to the conventional model, we found that the RB pathway functioned at backgrounds sufficient to activate the rod {\textrightarrow} cone pathway. As background light intensity increased, the RB{\textquoteright}s role changed from encoding the absorption of single photons to encoding contrast modulations around mean luminance. This transition is explained by the intrinsic dynamics of transmission from RB synapses.

}, keywords = {2-Amino-5-phosphonovalerate, Adaptation, Ocular, Animals, Biophysics, Computer Simulation, Excitatory Amino Acid Antagonists, Glucosamine 6-Phosphate N-Acetyltransferase, Green Fluorescent Proteins, In Vitro Techniques, Light, Mice, Mice, Inbred C57BL, Mice, Knockout, Mice, Transgenic, Models, Neurological, Patch-Clamp Techniques, Quinoxalines, Retinal Bipolar Cells, Retinal Rod Photoreceptor Cells, Synapses, Visual Pathways}, issn = {1097-4199}, doi = {10.1016/j.neuron.2013.10.054}, author = {Ke, Jiang-Bin and Wang, Yanbin V and Borghuis, Bart G and Cembrowski, Mark S and Riecke, Hermann and Kath, William L and Demb, Jonathan B and Singer, Joshua H} } @article {6935, title = {Allelic heterogeneity in NCF2 associated with systemic lupus erythematosus (SLE) susceptibility across four ethnic populations.}, journal = {Human Molecular Genetics}, volume = {23}, year = {2014}, month = {2014 Mar 15}, pages = {1656-68}, abstract = {

Recent reports have associated NCF2, encoding a core component of the multi-protein NADPH oxidase (NADPHO), with systemic lupus erythematosus (SLE) susceptibility in individuals of European ancestry. To identify ethnicity-specific and -robust variants within NCF2, we assessed 145 SNPs in and around the NCF2 gene in 5325 cases and 21 866 controls of European-American (EA), African-American (AA), Hispanic (HS) and Korean (KR) ancestry. Subsequent imputation, conditional, haplotype and bioinformatic analyses identified seven potentially functional SLE-predisposing variants. Association with non-synonymous rs17849502, previously reported in EA, was detected in EA, HS and AA (P(EA) = 1.01 {\texttimes} 10(-54), PHS = 3.68 {\texttimes} 10(-10), P(AA) = 0.03); synonymous rs17849501 was similarly significant. These SNPs were monomorphic in KR. Novel associations were detected with coding variants at rs35937854 in AA (PAA = 1.49 {\texttimes} 10(-9)), and rs13306575 in HS and KR (P(HS) = 7.04 {\texttimes} 10(-7), P(KR) = 3.30 {\texttimes} 10(-3)). In KR, a 3-SNP haplotype was significantly associated (P = 4.20 {\texttimes} 10(-7)), implying that SLE predisposing variants were tagged. Significant SNP-SNP interaction (P = 0.02) was detected between rs13306575 and rs17849502 in HS, and a dramatically increased risk (OR = 6.55) with a risk allele at each locus. Molecular modeling predicts that these non-synonymous mutations could disrupt NADPHO complex assembly. The risk allele of rs17849501, located in a conserved transcriptional regulatory region, increased reporter gene activity, suggesting in vivo enhancer function. Our results not only establish allelic heterogeneity within NCF2 associated with SLE, but also emphasize the utility of multi-ethnic cohorts to identify predisposing variants explaining additional phenotypic variance ({\textquoteright}missing heritability{\textquoteright}) of complex diseases like SLE.

}, issn = {1460-2083}, doi = {10.1093/hmg/ddt532}, author = {Kim-Howard, Xana and Sun, Celi and Molineros, Julio E and Maiti, Amit K and Chandru, Hema and Adler, Adam and Wiley, Graham B and Kaufman, Kenneth M and Kottyan, Leah and Guthridge, Joel M and Rasmussen, Astrid and Kelly, Jennifer and S{\'a}nchez, Elena and Raj, Prithvi and Li, Quan-Zhen and Bang, So-Young and Lee, Hye-Soon and Kim, Tae-Hwan and Kang, Young Mo and Suh, Chang-Hee and Chung, Won Tae and Park, Yong-Beom and Choe, Jung-Yoon and Shim, Seung Cheol and Lee, Shin-Seok and Han, Bok-Ghee and Olsen, Nancy J and Karp, David R and Moser, Kathy and Pons-Estel, Bernardo A and Wakeland, Edward K and James, Judith A and Harley, John B and Bae, Sang-Cheol and Gaffney, Patrick M and Alarc{\'o}n-Riquelme, Marta and Looger, Loren L and Nath, Swapan K} } @article {6880, title = {Automated detection of synapses in serial section transmission electron microscopy image stacks.}, journal = {PloS one}, volume = {9}, year = {2014}, month = {2014}, pages = {e87351}, abstract = {We describe a method for fully automated detection of chemical synapses in serial electron microscopy images with highly anisotropic axial and lateral resolution, such as images taken on transmission electron microscopes. Our pipeline starts from classification of the pixels based on 3D pixel features, which is followed by segmentation with an Ising model MRF and another classification step, based on object-level features. Classifiers are learned on sparse user labels; a fully annotated data subvolume is not required for training. The algorithm was validated on a set of 238 synapses in 20 serial 7197{\texttimes}7351 pixel images (4.5{\texttimes}4.5{\texttimes}45 nm resolution) of mouse visual cortex, manually labeled by three independent human annotators and additionally re-verified by an expert neuroscientist. The error rate of the algorithm (12\% false negative, 7\% false positive detections) is better than state-of-the-art, even though, unlike the state-of-the-art method, our algorithm does not require a prior segmentation of the image volume into cells. The software is based on the ilastik learning and segmentation toolkit and the vigra image processing library and is freely available on our website, along with the test data and gold standard annotations (http://www.ilastik.org/synapse-detection/sstem).}, issn = {1932-6203}, doi = {10.1371/journal.pone.0087351}, author = {Kreshuk, Anna and Koethe, Ullrich and Pax, Elizabeth and Bock, Davi D and Hamprecht, Fred A} } @article {45458, title = {Behavioral variability through stochastic choice and its gating by anterior cingulate cortex.}, journal = {Cell}, volume = {159}, year = {2014}, month = {2014 Sep 25}, pages = {21-32}, abstract = {

Behavioral choices that ignore prior experience promote exploration and unpredictability but are seemingly at odds with the brain{\textquoteright}s tendency to use experience to optimize behavioral choice. Indeed, when faced with virtual competitors, primates resort to strategic counterprediction rather than to stochastic choice. Here, we show that rats also use history- and model-based strategies when faced with similar competitors but can switch to a "stochastic" mode when challenged with a competitor that they cannot defeat by counterprediction. In this mode, outcomes associated with an animal{\textquoteright}s actions are ignored, and normal engagement of anterior cingulate cortex (ACC) is suppressed. Using circuit perturbations in\ transgenic rats, we demonstrate that switching between strategic and stochastic behavioral modes is controlled by locus coeruleus input into ACC. Our\ findings suggest that, under conditions of uncertainty about environmental rules, changes in noradrenergic input alter ACC output and prevent erroneous beliefs from guiding decisions, thus enabling behavioral variation.

}, issn = {1097-4172}, doi = {10.1016/j.cell.2014.08.037}, author = {Tervo, Dougal G R and Proskurin, Mikhail and Manakov, Maxim and Kabra, Mayank and Vollmer, Alison and Branson, Kristin and Karpova, Alla Y} } @article {45638, title = {Cell-type-specific repression by methyl-CpG-binding protein 2 is biased toward long genes.}, journal = {Journal of Neuroscience}, volume = {34}, year = {2014}, month = {2014 Sep 17}, pages = {12877-83}, abstract = {

Mutations in methyl-CpG-binding protein 2 (MeCP2) cause Rett syndrome and related autism spectrum disorders (Amir et al., 1999). MeCP2 is believed to be required for proper regulation of brain gene expression, but prior microarray studies in Mecp2 knock-out mice using brain tissue homogenates have revealed only subtle changes in gene expression (Tudor et al., 2002; Nuber et al., 2005; Jordan et al., 2007; Chahrour et al., 2008). Here, by profiling discrete subtypes of neurons we uncovered more dramatic effects of MeCP2 on gene expression, overcoming the "dilution problem" associated with assaying homogenates of complex tissues. The results reveal misregulation of genes involved in neuronal connectivity and communication. Importantly, genes upregulated following loss of MeCP2 are biased toward longer genes but this is not true for downregulated genes, suggesting MeCP2 may selectively repress long genes. Because genes involved in neuronal connectivity and communication, such as cell adhesion and cell-cell signaling genes, are enriched among longer genes, their misregulation following loss of MeCP2 suggests a possible etiology for altered circuit function in Rett syndrome.

}, issn = {1529-2401}, doi = {10.1523/JNEUROSCI.2674-14.2014}, author = {Sugino, Ken and Hempel, Chris M and Okaty, Benjamin W and Arnson, Hannah A and Kato, Saori and Dani, Vardhan S and Nelson, Sacha B} } @article {45181, title = {Combined protein- and nucleic acid-level effects of rs1143679 (R77H), a lupus-predisposing variant within ITGAM.}, journal = {Human Molecular Genetics}, volume = {23}, year = {2014}, month = {2014 Aug 1}, pages = {4161-76}, abstract = {

Integrin alpha M (ITGAM; CD11b) is a component of the macrophage-1 antigen complex, which mediates leukocyte adhesion, migration and phagocytosis as part of the immune system. We previously identified a missense polymorphism, rs1143679 (R77H), strongly associated with systemic lupus erythematosus (SLE). However, the molecular mechanisms of this variant are incompletely understood. A meta-analysis of published and novel data on 28 439 individuals with European, African, Hispanic and Asian ancestries reinforces genetic association between rs1143679 and SLE [Pmeta = 3.60 {\texttimes} 10(-90), odds ratio (OR) = 1.76]. Since rs1143679 is in the most active region of chromatin regulation and transcription factor binding in ITGAM, we quantitated ITGAM RNA and surface protein levels in monocytes from patients with each rs1143679 genotype. We observed that transcript levels significantly decreased for the risk allele ({\textquoteright}A{\textquoteright}) relative to the non-risk allele ({\textquoteright}G{\textquoteright}), in a dose-dependent fashion: ({\textquoteright}AA{\textquoteright} \< {\textquoteright}AG{\textquoteright} \< {\textquoteright}GG{\textquoteright}). CD11b protein levels in patients{\textquoteright} monocytes were directly correlated with RNA levels. Strikingly, heterozygous individuals express much lower (average 10- to 15-fold reduction) amounts of the {\textquoteright}A{\textquoteright} transcript than {\textquoteright}G{\textquoteright} transcript. We found that the non-risk sequence surrounding rs1143679 exhibits transcriptional enhancer activity in vivo and binds to Ku70/80, NFKB1 and EBF1 in vitro, functions that are significantly reduced with the risk allele. Mutant CD11b protein shows significantly reduced binding to fibrinogen and vitronectin, relative to non-risk, both in purified protein and in cellular models. This two-pronged contribution (nucleic acid- and protein-level) of the rs1143679 risk allele to decreasing ITGAM activity provides insight into the molecular mechanisms of its potent association with SLE.

}, issn = {1460-2083}, doi = {10.1093/hmg/ddu106}, author = {Maiti, Amit K and Kim-Howard, Xana and Motghare, Prasenjeet and Pradhan, Vandana and Chua, Kek Heng and Sun, Celi and Arango-Guerrero, Mar{\'\i}a Teresa and Ghosh, Kanjaksha and Niewold, Timothy B and Harley, John B and Anaya, Juan-Manual and Looger, Loren L and Nath, Swapan K} } @article {6960, title = {Conditions and constraints for astrocyte calcium signaling in the hippocampal mossy fiber pathway.}, journal = {Neuron}, volume = {82}, year = {2014}, month = {2014 Apr 16}, pages = {413-29}, abstract = {

The spatiotemporal activities of astrocyte Ca(2+) signaling in mature neuronal circuits remain unclear. We used genetically encoded Ca(2+) and glutamate indicators as well as pharmacogenetic and electrical control of neurotransmitter release to explore astrocyte activity in the hippocampal mossy fiber pathway. Our data revealed numerous localized, spontaneous Ca(2+) signals in astrocyte branches and territories, but these were not driven by neuronal activity or glutamate. Moreover, evoked astrocyte Ca(2+) signaling changed linearly with the number of mossy fiber action potentials. Under these settings, astrocyte responses were global, suppressed by neurotransmitter clearance, and mediated by glutamate and GABA. Thus, astrocyte engagement in the fully developed mossy fiber pathway was slow and territorial, contrary to that frequently proposed for astrocytes within microcircuits. We show that astrocyte Ca(2+) signaling functionally segregates large volumes of neuropil and that these transients are not suited for responding to, or regulating, single synapses in the mossy fiber pathway.

}, issn = {1097-4199}, doi = {10.1016/j.neuron.2014.02.041}, author = {Haustein, Martin D and Kracun, Sebastian and Lu, Xiao-Hong and Shih, Tiffany and Jackson-Weaver, Olan and Tong, Xiaoping and Xu, Ji and Yang, X William and O{\textquoteright}Dell, Thomas J and Marvin, Jonathan S and Ellisman, Mark H and Bushong, Eric A and Looger, Loren L and Khakh, Baljit S.} } @article {45419, title = {Development of the annelid axochord: insights into notochord evolution.}, journal = {Science}, volume = {345}, year = {2014}, month = {2014 Sep 12}, pages = {1365-8}, abstract = {

The origin of chordates has been debated for more than a century, with one key issue being the emergence of the notochord. In vertebrates, the notochord develops by convergence and extension of the chordamesoderm, a population of midline cells of unique molecular identity. We identify a population of mesodermal cells in a developing invertebrate, the marine annelid Platynereis dumerilii, that converges and extends toward the midline and expresses a notochord-specific combination of genes. These cells differentiate into a longitudinal muscle, the axochord, that is positioned between central nervous system and axial blood vessel and secretes a strong collagenous extracellular matrix. Ancestral state reconstruction suggests that contractile mesodermal midline cells existed in bilaterian ancestors. We propose that these cells, via vacuolization and stiffening, gave rise to the chordate notochord.

}, issn = {1095-9203}, doi = {10.1126/science.1253396}, author = {Lauri, Antonella and Brunet, Thibaut and Handberg-Thorsager, Mette and Fischer, Antje H L and Simakov, Oleg and Steinmetz, Patrick R H and Tomer, Raju and Keller, Philipp J and Arendt, Detlev} } @article {6891, title = {Direct phase measurement in zonal wavefront reconstruction using multidither coherent optical adaptive technique.}, journal = {Optics Express}, volume = {22}, year = {2014}, month = {2014 Jan 27}, pages = {1619-28}, abstract = {

In traditional zonal wavefront sensing for adaptive optics, after local wavefront gradients are obtained, the entire wavefront can be calculated by assuming that the wavefront is a continuous surface. Such an approach will lead to sub-optimal performance in reconstructing wavefronts which are either discontinuous or undersampled by the zonal wavefront sensor. Here, we report a new method to reconstruct the wavefront by directly measuring local wavefront phases in parallel using multidither coherent optical adaptive technique. This method determines the relative phases of each pupil segment independently, and thus produces an accurate wavefront for even discontinuous wavefronts. We implemented this method in an adaptive optical two-photon fluorescence microscopy and demonstrated its superior performance in correcting large or discontinuous aberrations.

}, issn = {1094-4087}, doi = {10.1364/OE.22.001619}, author = {Liu, Rui and Milkie, Daniel E and Kerlin, Aaron and Maclennan, Bryan and Ji, Na} } @article {6911, title = {Discovery of brainwide neural-behavioral maps via multiscale unsupervised structure learning}, journal = {Science}, volume = {344}, year = {2014}, month = {2014 Mar 27}, pages = {386-92}, chapter = {386}, abstract = {

A single nervous system can generate many distinct motor patterns. Identifying which neurons and circuits control which behaviors has been a laborious piecemeal process, usually for one observer-defined behavior at a time. We present a fundamentally different approach to neuron-behavior mapping. We optogenetically activated 1,054 identified neuron lines in Drosophila larva and tracked the behavioral responses from 37,780 animals. Applying multiscale unsupervised structure learning methods to the behavioral data identified 29 discrete statistically distinguishable and observer-unbiased behavioral phenotypes. Mapping the neural lines to the behavior(s) they evoke provides a behavioral reference atlas for neuron subsets covering a large fraction of larval neurons. This atlas is a starting point for connectivity- and activity-mapping studies to further investigate the mechanisms by which neurons mediate diverse behaviors.

}, issn = {1095-9203}, doi = {10.1126/science.1250298}, author = {Vogelstein, Joshua T and Park, Youngser and Ohyama, Tomoko and Kerr, Rex and Truman, James W and Priebe, Carey E. and Zlatic, Marta} } @article {46595, title = {EGFR and FGFR pathways have distinct roles in Drosophila mushroom body development and ethanol-induced behavior.}, journal = {PLoS One}, volume = {9}, year = {2014}, month = {2014 Jan 31}, pages = {e87714}, abstract = {

Epidermal Growth Factor Receptor (EGFR) signaling has a conserved role in ethanol-induced behavior in flies and mice, affecting ethanol-induced sedation in both species. However it is not known what other effects EGFR signaling may have on ethanol-induced behavior, or what roles other Receptor Tyrosine Kinase (RTK) pathways may play in ethanol induced behaviors. We examined the effects of both the EGFR and Fibroblast Growth Factor Receptor (FGFR) RTK signaling pathways on ethanol-induced enhancement of locomotion, a behavior distinct from sedation that may be associated with the rewarding effects of ethanol. We find that both EGFR and FGFR genes influence ethanol-induced locomotion, though their effects are opposite - EGFR signaling suppresses this behavior, while FGFR signaling promotes it. EGFR signaling affects development of the Drosophila mushroom bodies in conjunction with the JNK MAP kinase basket (bsk), and with the Ste20 kinase tao, and we hypothesize that the EGFR pathway affects ethanol-induced locomotion through its effects on neuronal development. We find, however, that FGFR signaling most likely affects ethanol-induced behavior through a different mechanism, possibly through acute action in adult neurons.

}, keywords = {Animals, Behavior, Animal, Central Nervous System Depressants, Drosophila melanogaster, Drosophila Proteins, Ethanol, Locomotion, Mice, Mushroom Bodies, Receptor, Epidermal Growth Factor, Receptors, Invertebrate Peptide, Signal Transduction}, issn = {1932-6203}, doi = {10.1371/journal.pone.0087714}, author = {King, Ian F G and Eddison, Mark and Kaun, Karla R and Heberlein, Ulrike} } @article {6895, title = {Excitatory synaptic inputs to mouse on-off direction-selective retinal ganglion cells lack direction tuning.}, journal = {The Journal of Neuroscience : the Official Journal of the Society for Neuroscience}, volume = {34}, year = {2014}, month = {2014 Mar 12}, pages = {3976-81}, abstract = {

Direction selectivity represents a fundamental visual computation. In mammalian retina, On-Off direction-selective ganglion cells (DSGCs) respond strongly to motion in a preferred direction and weakly to motion in the opposite, null direction. Electrical recordings suggested three direction-selective (DS) synaptic mechanisms: DS GABA release during null-direction motion from starburst amacrine cells (SACs) and DS acetylcholine and glutamate release during preferred direction motion from SACs and bipolar cells. However, evidence for DS acetylcholine and glutamate release has been inconsistent and at least one bipolar cell type that contacts another DSGC (On-type) lacks DS release. Here, whole-cell recordings in mouse retina showed that cholinergic input to On-Off DSGCs lacked DS, whereas the remaining (glutamatergic) input showed apparent DS. Fluorescence measurements with the glutamate biosensor intensity-based glutamate-sensing fluorescent reporter (iGluSnFR) conditionally expressed in On-Off DSGCs showed that glutamate release in both On- and Off-layer dendrites lacked DS, whereas simultaneously recorded excitatory currents showed apparent DS. With GABA-A receptors blocked, both iGluSnFR signals and excitatory currents lacked DS. Our measurements rule out DS release from bipolar cells onto On-Off DSGCs and support a theoretical model suggesting that apparent DS excitation in voltage-clamp recordings results from inadequate voltage control of DSGC dendrites during null-direction inhibition. SAC GABA release is the apparent sole source of DS input onto On-Off DSGCs.

}, issn = {1529-2401}, doi = {10.1523/JNEUROSCI.5017-13.2014}, author = {Park, Silvia J H and Kim, In-Jung and Looger, Loren L and Demb, Jonathan B and Borghuis, Bart G} } @article {45058, title = {Fast, accurate reconstruction of cell lineages from large-scale fluorescence microscopy data.}, journal = {Nature Methods}, volume = {11}, year = {2014}, month = {2014 Jul 20}, pages = {951-8}, abstract = {

The comprehensive reconstruction of cell lineages in complex multicellular organisms is a central goal of developmental biology. We present an open-source computational framework for the segmentation and tracking of cell nuclei with high accuracy and speed. We demonstrate its (i) generality by reconstructing cell lineages in four-dimensional, terabyte-sized image data sets of fruit fly, zebrafish and mouse embryos acquired with three types of fluorescence microscopes, (ii) scalability by analyzing advanced stages of development with up to 20,000 cells per time point at 26,000 cells min(-1) on a single computer workstation and (iii) ease of use by adjusting only two parameters across all data sets and providing visualization and editing tools for efficient data curation. Our approach achieves on average 97.0\% linkage accuracy across all species and imaging modalities. Using our system, we performed the first cell lineage reconstruction of early Drosophila melanogaster nervous system development, revealing neuroblast dynamics throughout an entire embryo.

}, issn = {1548-7105}, doi = {10.1038/nmeth.3036}, author = {Amat, Fernando and Lemon, William and Mossing, Daniel P and McDole, Katie and Wan, Yinan and Branson, Kristin and Myers, Eugene W and Keller, Philipp J} } @article {47624, title = {Flat clathrin lattices: stable features of the plasma membrane.}, journal = {Molecular biology of the cell}, volume = {25}, year = {2014}, month = {2014 Nov 5}, pages = {3581-94}, abstract = {

Clathrin-mediated endocytosis (CME) is a fundamental property of eukaryotic cells. Classical CME proceeds via the formation of clathrin-coated pits (CCPs) at the plasma membrane, which invaginate to form clathrin-coated vesicles, a process that is well understood. However, clathrin also assembles into flat clathrin lattices (FCLs); these structures remain poorly described, and their contribution to cell biology is unclear. We used quantitative imaging to provide the first comprehensive description of FCLs and explore their influence on plasma membrane organization. Ultrastructural analysis by electron and superresolution microscopy revealed two discrete populations of clathrin structures. CCPs were typified by their sphericity, small size, and homogeneity. FCLs were planar, large, and heterogeneous and present on both the dorsal and ventral surfaces of cells. Live microscopy demonstrated that CCPs are short lived and culminate in a peak of dynamin recruitment, consistent with classical CME. In contrast, FCLs were long lived, with sustained association with dynamin. We investigated the biological relevance of FCLs using the chemokine receptor CCR5 as a model system. Agonist activation leads to sustained recruitment of CCR5 to FCLs. Quantitative molecular imaging indicated that FCLs partitioned receptors at the cell surface. Our observations suggest that FCLs provide stable platforms for the recruitment of endocytic cargo.

}, keywords = {Animals, Chemokine CCL5, CHO Cells, Clathrin, Clathrin-Coated Vesicles, Coated Pits, Cell-Membrane, Cricetulus, Dynamins, Endocytosis, Genes, Reporter, Green Fluorescent Proteins, HEK293 Cells, Hela Cells, Humans, Microscopy, Electron, Microscopy, Fluorescence, Molecular Imaging, Protein Transport, Receptors, CCR5}, issn = {1939-4586}, doi = {10.1091/mbc.E14-06-1154}, author = {Grove, Joe and Metcalf, Daniel J and Knight, Alex E and Wavre-Shapton, Sil{\`e}ne T and Sun, Tony and Protonotarios, Emmanouil D and Griffin, Lewis D and Lippincott-Schwartz, Jennifer and Marsh, Mark} } @article {45334, title = {A GAL4 driver resource for developmental and behavioral studies on the larval CNS of Drosophila.}, journal = {Cell Reports}, volume = {8}, year = {2014}, month = {2014 Aug 7}, pages = {897-908}, abstract = {

We report the larval CNS expression patterns for 6,650 GAL4 lines based on cis-regulatory regions (CRMs) from the Drosophila genome. Adult CNS expression patterns were previously reported for this collection, thereby providing a unique resource for determining the origins of adult cells. An illustrative example reveals the origin of the astrocyte-like glia of the ventral\ CNS. Besides larval neurons and glia, the larval CNS\ contains scattered lineages of immature, adult-specific\ neurons. Comparison of lineage expression within this large collection of CRMs provides insight into the codes used for designating neuronal types. The CRMs encode both dense and sparse patterns of lineage expression. There is little correlation between brain and thoracic lineages in patterns of sparse expression, but expression in the two regions is highly correlated in the dense mode. The optic lobes, by comparison, appear to use a different set of genetic instructions in their development.

}, issn = {2211-1247}, doi = {10.1016/j.celrep.2014.06.065}, author = {Li, Hsing-Hsi and Kroll, Jason R and Lennox, Sara M and Ogundeyi, Omotara and Jeter, Jennifer and Depasquale, Gina and Truman, James W} } @article {6934, title = {Graph-based active learning of agglomeration (GALA): a Python library to segment 2D and 3D neuroimages}, journal = {Frontiers in Neuroinformatics}, volume = {8}, year = {2014}, month = {2014 Apr 4}, pages = {34}, abstract = {

The aim in high-resolution connectomics is to reconstruct complete neuronal connectivity in a tissue. Currently, the only technology capable of resolving the smallest neuronal processes is electron microscopy (EM). Thus, a common approach to network reconstruction is to perform (error-prone) automatic segmentation of EM images, followed by manual proofreading by experts to fix errors. We have developed an algorithm and software library to not only improve the accuracy of the initial automatic segmentation, but also point out the image coordinates where it is likely to have made errors. Our software, called gala (graph-based active learning of agglomeration), improves the state of the art in agglomerative image segmentation. It is implemented in Python and makes extensive use of the scientific Python stack (numpy, scipy, networkx, scikit-learn, scikit-image, and others). We present here the software architecture of the gala library, and discuss several designs that we consider would be generally useful for other segmentation packages. We also discuss the current limitations of the gala library and how we intend to address them.

}, issn = {1662-5196}, doi = {10.3389/fninf.2014.00034}, author = {Nunez-Iglesias, Juan and Kennedy, Ryan and Plaza, Stephen M and Chakraborty, Anirban and William T. Katz} } @article {46704, title = {Heterogeneity in periodontitis prevalence in the Hispanic Community Health Study/Study of Latinos.}, journal = {Annals of Epidemiology}, volume = {24}, year = {2014}, month = {2014 Jun}, pages = {455-62}, abstract = {

PURPOSE: The aim of the study was to examine acculturation and established risk factors in explaining variation in periodontitis prevalence among Hispanic/Latino subgroups.

METHODS: Participants were 12,730 dentate adults aged 18-74 years recruited into the Hispanic Community Health Study/Study of Latinos (HCHS/SOL) from four U.S. field centers between 2008 and 2011. A standardized periodontal assessment measured probing pocket depth and gingival recession at six sites per tooth for up to 28 teeth. Periodontitis was defined according to the Centers for Disease Control and Prevention and American Academy of Periodontology case classifications developed for population surveillance. Covariates included acculturation indicators and established periodontitis risk factors. Survey estimation procedures took account of the complex sampling design. Adjusted multivariate binomial regression estimated prevalence ratios and 95\% confidence limits (CLs).

RESULTS: Unadjusted prevalence of moderate and severe periodontitis was 38.5\% and ranged from 24.7\% among Dominicans to 52.1\% among Cubans. Adjusted prevalence ratios for subgroups relative to Dominicans were as follows: (1) 1.34 (95\% CL, 1.13-1.58) among South Americans; (2) 1.37 (95\% CL, 1.17-1.61) among Puerto Ricans; (3) 1.43 (95\% CL, 1.25-1.64) among Mexicans; (4) 1.53 (95\% CL, 1.32-1.76) among Cubans; and (5) 1.55 (95\% CL, 1.35-1.78) among Central Americans.

CONCLUSIONS: Heterogeneity in prevalence of moderate/severe periodontitis among Hispanic/Latino subpopulations was not explained by acculturation or periodontitis risk factors.

}, keywords = {Adult, Aged, Attitude to Health, Dental Health Surveys, Female, Hispanic Americans, Humans, Male, Middle Aged, Oral Hygiene, Periodontitis, Population Surveillance, Prevalence, Risk Assessment, Risk Factors, United States, Young Adult}, issn = {1873-2585}, doi = {10.1016/j.annepidem.2014.02.018}, author = {Sanders, Anne E and Campbell, Steven M and Mauriello, Sally M and Beck, James D and Jimenez, Monik C and Kaste, Linda M and Singer, Richard H and Beaver, Shirley M and Finlayson, Tracy L and Badner, Victor M} } @article {47897, title = {A high-resolution spatiotemporal atlas of gene expression of the developing mouse brain.}, journal = {Neuron}, volume = {83}, year = {2014}, month = {2014 Jul 16}, pages = {309-23}, abstract = {

To provide a temporal framework for the genoarchitecture of brain development, we generated in situ hybridization data for embryonic and postnatal mouse brain at seven developmental stages for \~{}2,100 genes, which were processed with an automated informatics pipeline and manually annotated. This resource comprises 434,946 images, seven reference atlases, an ontogenetic ontology, and tools to explore coexpression of genes across neurodevelopment. Gene sets coinciding with developmental phenomena were identified. A temporal shift in the principles governing the molecular organization of the brain was detected, with transient neuromeric, plate-based organization of the brain present at E11.5 and E13.5. Finally, these data provided a transcription factor code that discriminates brain structures and identifies the developmental age of a tissue, providing a foundation for eventual genetic manipulation or tracking of specific brain structures over development. The resource is available as the Allen Developing Mouse Brain Atlas (http://developingmouse.brain-map.org).

}, keywords = {Animals, Brain, Brain Mapping, Gene Expression, Gene Expression Profiling, Gene Expression Regulation, Developmental, Mice}, issn = {1097-4199}, doi = {10.1016/j.neuron.2014.05.033}, author = {Thompson, Carol L and Ng, Lydia and Menon, Vilas and Martinez, Salvador and Lee, Chang-Kyu and Glattfelder, Katie and Sunkin, Susan M and Henry, Alex and Lau, Christopher and Dang, Chinh and Garcia-Lopez, Raquel and Martinez-Ferre, Almudena and Pombero, Ana and Rubenstein, John L R and Wakeman, Wayne B and Hohmann, John and Dee, Nick and Sodt, Andrew J and Young, Rob and Smith, Kimberly and Nguyen, Thuc-Nghi and Kidney, Jolene and Kuan, Leonard and Jeromin, Andreas and Kaykas, Ajamete and Miller, Jeremy and Page, Damon and Orta, Geri and Bernard, Amy and Riley, Zackery and Smith, Simon and Wohnoutka, Paul and Hawrylycz, Michael J and Puelles, Luis and Jones, Allan R} } @article {45060, title = {Imaging cellular ultrastructure by PALM, iPALM, and correlative iPALM-EM.}, journal = {Methods in Cell Biology}, volume = {123}, year = {2014}, month = {2014}, pages = {273-94}, abstract = {

Many biomolecules in cells can be visualized with high sensitivity and specificity by fluorescence microscopy. However, the resolution of conventional light microscopy is limited by diffraction to ~200-250nm laterally and \>500nm axially. Here, we describe superresolution methods based on single-molecule localization analysis of photoswitchable fluorophores (PALM: photoactivated localization microscopy) as well as our recent three-dimensional (3D) method (iPALM: interferometric PALM) that allows imaging with a resolution better than 20nm in all three dimensions. Considerations for their implementations, applications to multicolor imaging, and a recent development that extend the imaging depth of iPALM to ~750nm are discussed. As the spatial resolution of superresolution fluorescence microscopy converges with that of electron microscopy (EM), direct imaging of the same specimen using both approaches becomes feasible. This could be particularly useful for cross validation of experiments, and thus, we also describe recent methods that were developed for correlative superresolution fluorescence and EM.

}, issn = {0091-679X}, doi = {10.1016/B978-0-12-420138-5.00015-X}, author = {Shtengel, Gleb and Wang, Yilin and Zhang, Zhen and Goh, Wah Ing and Hess, Harald F and Kanchanawong, Pakorn} } @article {47898, title = {Improving reliability and absolute quantification of human brain microarray data by filtering and scaling probes using RNA-Seq.}, journal = {BMC genomics}, volume = {15}, year = {2014}, month = {2014}, pages = {154}, abstract = {

BACKGROUND: High-throughput sequencing is gradually replacing microarrays as the preferred method for studying mRNA expression levels, providing nucleotide resolution and accurately measuring absolute expression levels of almost any transcript, known or novel. However, existing microarray data from clinical, pharmaceutical, and academic settings represent valuable and often underappreciated resources, and methods for assessing and improving the quality of these data are lacking.

RESULTS: To quantitatively assess the quality of microarray probes, we directly compare RNA-Seq to Agilent microarrays by processing 231 unique samples from the Allen Human Brain Atlas using RNA-Seq. Both techniques provide highly consistent, highly reproducible gene expression measurements in adult human brain, with RNA-Seq slightly outperforming microarray results overall. We show that RNA-Seq can be used as ground truth to assess the reliability of most microarray probes, remove probes with off-target effects, and scale probe intensities to match the expression levels identified by RNA-Seq. These sequencing scaled microarray intensities (SSMIs) provide more reliable, quantitative estimates of absolute expression levels for many genes when compared with unscaled intensities. Finally, we validate this result in two human cell lines, showing that linear scaling factors can be applied across experiments using the same microarray platform.

CONCLUSIONS: Microarrays provide consistent, reproducible gene expression measurements, which are improved using RNA-Seq as ground truth. We expect that our strategy could be used to improve probe quality for many data sets from major existing repositories.

}, keywords = {Brain, Cluster Analysis, Computational Biology, Gene Expression, Gene Expression Profiling, High-Throughput Nucleotide Sequencing, Humans, Neocortex, Oligonucleotide Array Sequence Analysis, Reproducibility of Results, Sequence Analysis, RNA, Transcriptome}, issn = {1471-2164}, doi = {10.1186/1471-2164-15-154}, author = {Miller, Jeremy A and Menon, Vilas and Goldy, Jeff and Kaykas, Ajamete and Lee, Chang-Kyu and Smith, Kimberly A and Shen, Elaine H and Phillips, John W and Lein, Ed S and Hawrylycz, Mike J} } @article {45786, title = {In vivo fluorescence microscopy via iterative multi-photon adaptive compensation technique.}, journal = {Optical Express}, volume = {22}, year = {2014}, month = {2014 Oct 6}, pages = {23786-94}, abstract = {

Iterative multi-photon adaptive compensation technique (IMPACT) has been developed for wavefront measurement and compensation in highly scattering tissues. Our previous report was largely based on the measurements of fixed tissue. Here we demonstrate the advantages of IMPACT for in vivo imaging and report the latest results. In particular, we show that IMPACT can be used for functional imaging of awake mice, and greatly improve the in vivo neuron imaging in mouse cortex at large depth (~660 microns). Moreover, IMPACT enables neuron imaging through the intact skull of adult mice, which promises noninvasive optical measurements in mouse brain.

}, issn = {1094-4087}, doi = {10.1364/OE.22.023786}, author = {Kong, Lingjie and Cui, Meng} } @article {6719, title = {Independent optical excitation of distinct neural populations.}, journal = {Nature Methods}, volume = {11}, year = {2014}, month = {2014 Feb 9}, pages = {338-46}, abstract = {

Optogenetic tools enable examination of how specific cell types contribute to brain circuit functions. A long-standing question is whether it is possible to independently activate two distinct neural populations in mammalian brain tissue. Such a capability would enable the study of how different synapses or pathways interact to encode information in the brain. Here we describe two channelrhodopsins, Chronos and Chrimson, discovered through sequencing and physiological characterization of opsins from over 100 species of alga. Chrimson{\textquoteright}s excitation spectrum is red shifted by 45 nm relative to previous channelrhodopsins and can enable experiments in which red light is preferred. We show minimal visual system-mediated behavioral interference when using Chrimson in neurobehavioral studies in Drosophila melanogaster. Chronos has faster kinetics than previous channelrhodopsins yet is effectively more light sensitive. Together these two reagents enable two-color activation of neural spiking and downstream synaptic transmission in independent neural populations without detectable cross-talk in mouse brain slice.

}, issn = {1548-7105}, doi = {10.1038/nmeth.2836}, author = {Klapoetke, Nathan C and Murata, Yasunobu and Kim, Sung Soo and Pulver, Stefan R and Birdsey-Benson, Amanda and Cho, Yong Ku and Morimoto, Tania K and Chuong, Amy S and Carpenter, Eric J and Tian, Zhijian and Wang, Jun and Xie, Yinlong and Yan, Zhixiang and Zhang, Yong and Chow, Brian Y and Surek, Barbara and Melkonian, Michael and Jayaraman, Vivek and Constantine-Paton, Martha and Wong, Gane Ka-Shu and Boyden, Edward S} } @article {45459, title = {Intrinsic bursting of aII amacrine cells underlies oscillations in the rd1 mouse retina.}, journal = {Journal of Neurophysiology}, volume = {112}, year = {2014}, month = {2014 Sep 15}, pages = {1491-504}, abstract = {

In many forms of retinal degeneration, photoreceptors die but inner retinal circuits remain intact. In the rd1 mouse, an established model for blinding retinal diseases, spontaneous activity in the coupled network of AII amacrine and ON cone bipolar cells leads to rhythmic bursting of ganglion cells. Since such activity could impair retinal and/or cortical responses to restored photoreceptor function, understanding its nature is important for developing treatments of retinal pathologies. Here we analyzed a compartmental model of the wild-type mouse AII amacrine cell to predict that the cell{\textquoteright}s intrinsic membrane properties, specifically, interacting fast Na and slow, M-type K conductances, would allow its membrane potential to oscillate when light-evoked excitatory synaptic inputs were withdrawn following photoreceptor degeneration. We tested and confirmed this hypothesis experimentally by recording from AIIs in a slice preparation of rd1 retina. Additionally, recordings from ganglion cells in a whole mount preparation of rd1 retina demonstrated that activity in AIIs was propagated unchanged to elicit bursts of action potentials in ganglion cells. We conclude that oscillations are not an emergent property of a degenerated retinal network. Rather, they arise largely from the intrinsic properties of a single retinal interneuron, the AII amacrine cell.

}, issn = {1522-1598}, doi = {10.1152/jn.00437.2014}, author = {Choi, Hannah and Zhang, Lei and Cembrowski, Mark S and Sabottke, Carl F and Markowitz, Alexander L and Butts, Daniel A and Kath, William L and Singer, Joshua H and Riecke, Hermann} } @article {45190, title = {Juvenile hormone regulates body size and perturbs insulin signaling in Drosophila.}, journal = {Proceedings of the National Academy of Science of the United States of America}, volume = {111}, year = {2014}, month = {2014 May 13}, pages = {7018-23}, abstract = {

The role of juvenile hormone (JH) in regulating the timing and nature of insect molts is well-established. Increasing evidence suggests that JH is also involved in regulating final insect size. Here we elucidate the developmental mechanism through which JH regulates body size in developing Drosophila larvae by genetically ablating the JH-producing organ, the corpora allata (CA). We found that larvae that lack CA pupariated at smaller sizes than control larvae due to a reduced larval growth rate. Neither the timing of the metamorphic molt nor the duration of larval growth was affected by the loss of JH. Further, we show that the effects of JH on growth rate are dependent on the forkhead box O transcription factor (FOXO), which is negatively regulated by the insulin-signaling pathway. Larvae that lacked the CA had elevated levels of FOXO activity, whereas a loss-of-function mutation of FOXO rescued the effects of CA ablation on final body size. Finally, the effect of JH on growth appears to be mediated, at least in part, via ecdysone synthesis in the prothoracic gland. These results indicate a role of JH in regulating growth rate via the ecdysone- and insulin-signaling pathways.

}, keywords = {Animals, Basic Helix-Loop-Helix Transcription Factors, Body Size, Corpora Allata, Denervation, Drosophila melanogaster, Drosophila Proteins, Ecdysone, Forkhead Transcription Factors, Insulin, Juvenile Hormones, Larva, Signal Transduction}, issn = {1091-6490}, doi = {10.1073/pnas.1313058111}, author = {Mirth, C K and Tang, H Y and Makohon-Moore, S C and Salhadar, S and Gokhale, R H and Warner, R D and Koyama, T and Riddiford, L M and Shingleton, A W} } @article {45617, title = {Lattice light-sheet microscopy: imaging molecules to embryos at high spatiotemporal resolution.}, journal = {Science}, volume = {346}, year = {2014}, month = {2014 Oct 24}, pages = {1257998}, abstract = {

Although fluorescence microscopy provides a crucial window into the physiology of living specimens, many biological processes are too fragile, are too small, or occur too rapidly to see clearly with existing tools. We crafted ultrathin light sheets from two-dimensional optical lattices that allowed us to image three-dimensional (3D) dynamics for hundreds of volumes, often at subsecond intervals, at the diffraction limit and beyond. We applied this to systems spanning four orders of magnitude in space and time, including the diffusion of single transcription factor molecules in stem cell spheroids, the dynamic instability of mitotic microtubules, the immunological synapse, neutrophil motility in a 3D matrix, and embryogenesis in Caenorhabditis elegans and Drosophila melanogaster. The results provide a visceral reminder of the beauty and the complexity of living systems.

}, issn = {1095-9203}, doi = {10.1126/science.1257998}, author = {Chen, Bi-Chang and Legant, Wesley R and Wang, Kai and Shao, Lin and Milkie, Daniel E and Davidson, Michael W and Janetopoulos, Chris and Wu, Xufeng S and Hammer, John A and Liu, Zhe and English, Brian P and Mimori-Kiyosue, Yuko and Romero, Daniel P and Ritter, Alex T and Lippincott-Schwartz, Jennifer and Fritz-Laylin, Lillian and Mullins, R Dyche and Mitchell, Diana M and Bembenek, Joshua N and Reymann, Anne-Cecile and B{\"o}hme, Ralph and Grill, Stephan W and Wang, Jennifer T and Seydoux, Geraldine and Tulu, U Serdar and Kiehart, Daniel P and Betzig, Eric} } @article {45841, title = {Leptin mediates the increase in blood pressure associated with obesity.}, journal = {Cell}, volume = {159}, year = {2014}, month = {2014 Dec 4}, pages = {1404-16}, abstract = {

Obesity is associated with increased blood pressure (BP), which in turn increases the risk of cardiovascular diseases. We found that the increase in leptin levels seen in diet-induced obesity (DIO) drives an increase in BP in rodents, an effect that was not seen in animals deficient in leptin or leptin receptors (LepR). Furthermore, humans with loss-of-function mutations in leptin and the LepR have low BP despite severe obesity. Leptin{\textquoteright}s effects on BP are mediated by neuronal circuits in the dorsomedial hypothalamus (DMH), as blocking leptin with a specific antibody, antagonist, or inhibition of the activity of LepR-expressing neurons in the DMH caused a rapid reduction of BP in DIO mice, independent of changes in weight. Re-expression of LepRs in the DMH of DIO LepR-deficient mice caused an increase in BP. These studies demonstrate that leptin couples changes in weight to changes in BP in mammalian species.

}, issn = {1097-4172}, doi = {10.1016/j.cell.2014.10.058}, author = {Simonds, Stephanie E and Pryor, Jack T and Ravussin, Eric and Greenway, Frank L and Dileone, Ralph and Allen, Andrew M and Bassi, Jaspreet and Elmquist, Joel K and Keogh, Julia M and Henning, Elana and Myers, Martin G and Licinio, Julio and Brown, Russell D and Enriori, Pablo J and O{\textquoteright}Rahilly, Stephen and Sternson, Scott M and Grove, Kevin L and Spanswick, David C and Farooqi, I Sadaf and Cowley, Michael A} } @article {45177, title = {Light-sheet functional imaging in fictively behaving zebrafish.}, journal = {Nature Methods}, volume = {11}, year = {2014}, month = {2014 Jul 27}, pages = {883-4}, abstract = {

The processing of sensory input and the generation of behavior involves large networks of neurons, which necessitates new technology for recording from many neurons in behaving animals. In the larval zebrafish, light-sheet microscopy can be used to record the activity of almost all neurons in the brain simultaneously at single-cell resolution. Existing implementations, however, cannot be combined with visually driven behavior because the light sheet scans over the eye, interfering with presentation of controlled visual stimuli. Here we describe a system that overcomes the confounding eye stimulation through the use of two light sheets and combines whole-brain light-sheet imaging with virtual reality for fictively behaving larval zebrafish.

}, issn = {1548-7105}, doi = {10.1038/nmeth.3040}, author = {Vladimirov, Nikita and Mu, Yu and Kawashima, Takashi and Bennett, Davis V and Yang, Chao-Tsung and Looger, Loren L and Keller, Philipp J and Freeman, Jeremy and Ahrens, Misha B} } @article {45868, title = {Light-sheet imaging for systems neuroscience.}, journal = {Nature Methods}, volume = {12}, year = {2014}, month = {2014 Dec 30}, pages = {27-9}, abstract = {

Developments in electrical and optical recording technology are scaling up the size of neuronal populations that can be monitored simultaneously. Light-sheet imaging is rapidly gaining traction as a method for optically interrogating activity in large networks and presents both opportunities and challenges for understanding circuit function.

}, issn = {1548-7105}, doi = {10.1038/nmeth.3214}, author = {Keller, Philipp J and Ahrens, Misha B and Freeman, Jeremy} } @article {6896, title = {Live imaging and quantitative analysis of gastrulation in mouse embryos using light-sheet microscopy and 3D tracking tools.}, journal = {Nature Protocols}, volume = {9}, year = {2014}, month = {2014 Mar}, pages = {575-85}, abstract = {

This protocol describes how to observe gastrulation in living mouse embryos by using light-sheet microscopy and computational tools to analyze the resulting image data at the single-cell level. We describe a series of techniques needed to image the embryos under physiological conditions, including how to hold mouse embryos without agarose embedding, how to transfer embryos without air exposure and how to construct environmental chambers for live imaging by digital scanned light-sheet microscopy (DSLM). Computational tools include manual and semiautomatic tracking programs that are developed for analyzing the large 4D data sets acquired with this system. Note that this protocol does not include details of how to build the light-sheet microscope itself. Time-lapse imaging ends within 12 h, with subsequent tracking analysis requiring 3-6 d. Other than some mouse-handling skills, this protocol requires no advanced skills or knowledge. Light-sheet microscopes are becoming more widely available, and thus the techniques outlined in this paper should be helpful for investigating mouse embryogenesis.

}, issn = {1750-2799}, doi = {10.1038/nprot.2014.035}, author = {Ichikawa, Takehiko and Nakazato, Kenichi and Keller, Philipp J and Kajiura-Kobayashi, Hiroko and Stelzer, Ernst H K and Mochizuki, Atsushi and Nonaka, Shigenori} } @article {45718, title = {Making biology transparent.}, journal = {Nature Biotechnology}, volume = {32}, year = {2014}, month = {2014 Nov 7}, pages = {1104-5}, abstract = {

The molecular and cellular architecture of the organs in a whole mouse is revealed through optical clearing.

}, issn = {1546-1696}, doi = {10.1038/nbt.3061}, author = {H{\"o}ckendorf, Burkhard and Lavis, Luke D and Keller, Philipp J} } @article {6898, title = {Making Drosophila lineage-restricted drivers via patterned recombination in neuroblasts.}, journal = {Nature Neuroscience}, volume = {17}, year = {2014}, month = {2014 Apr}, pages = {631-7}, abstract = {

The Drosophila cerebrum originates from about 100 neuroblasts per hemisphere, with each neuroblast producing a characteristic set of neurons. Neurons from a neuroblast are often so diverse that many neuron types remain unexplored. We developed new genetic tools that target neuroblasts and their diverse descendants, increasing our ability to study fly brain structure and development. Common enhancer-based drivers label neurons on the basis of terminal identities rather than origins, which provides limited labeling in the heterogeneous neuronal lineages. We successfully converted conventional drivers that are temporarily expressed in neuroblasts, into drivers expressed in all subsequent neuroblast progeny. One technique involves immortalizing GAL4 expression in neuroblasts and their descendants. Another depends on loss of the GAL4 repressor, GAL80, from neuroblasts during early neurogenesis. Furthermore, we expanded the diversity of MARCM-based reagents and established another site-specific mitotic recombination system. Our transgenic tools can be combined to map individual neurons in specific lineages of various genotypes.

}, issn = {1546-1726}, doi = {10.1038/nn.3654}, author = {Awasaki, Takeshi and Kao, Chih-Fei and Lee, Ying-Jou and Yang, Ching-Po and Huang, Yaling and Pfeiffer, Barret D and Luan, Haojiang and Jing, Xiaotang and Huang, Yu-Fen and He, Yisheng and Schroeder, Mark David and Kuzin, Alexander and Brody, Thomas and Zugates, Christopher T and Odenwald, Ward F and Lee, Tzumin} } @article {45065, title = {Mapping brain activity at scale with cluster computing.}, journal = {Nature Methods}, volume = {11}, year = {2014}, month = {2014 Jul 27}, pages = {941-950}, abstract = {

Understanding brain function requires monitoring and interpreting the activity of large networks of neurons during behavior. Advances in recording technology are greatly increasing the size and complexity of neural data. Analyzing such data will pose a fundamental bottleneck for neuroscience. We present a library of analytical tools called Thunder built on the open-source Apache Spark platform for large-scale distributed computing. The library implements a variety of univariate and multivariate analyses with a modular, extendable structure well-suited to interactive exploration and analysis development. We demonstrate how these analyses find structure in large-scale neural data, including whole-brain light-sheet imaging data from fictively behaving larval zebrafish, and two-photon imaging data from behaving mouse. The analyses relate neuronal responses to sensory input and behavior, run in minutes or less and can be used on a private cluster or in the cloud. Our open-source framework thus holds promise for turning brain activity mapping efforts into biological insights.

}, issn = {1548-7091}, doi = {10.1038/nmeth.3041}, author = {Freeman, Jeremy and Vladimirov, Nikita and Kawashima, Takashi and Mu, Yu and Sofroniew, Nicholas J. and Bennett, Davis V. and Rosen, Joshua and Yang, Chao-Tsung and Looger, Loren L. and Ahrens, Misha B.} } @article {45184, title = {Molecular basis for age-dependent microtubule acetylation by tubulin acetyltransferase.}, journal = {Cell}, volume = {157}, year = {2014}, month = {2014 Jun 5}, pages = {1405-15}, abstract = {

Acetylation of α-tubulin Lys40 by tubulin acetyltransferase (TAT) is the only known posttranslational modification in the microtubule lumen. It marks stable microtubules and is required for polarity establishment and directional migration. Here, we elucidate the mechanistic underpinnings for TAT activity and its preference for microtubules with slow turnover. 1.35\ {\r A} TAT cocrystal structures with bisubstrate analogs constrain TAT action to the microtubule lumen and reveal Lys40 engaged in a suboptimal active site. Assays with diverse tubulin polymers show that TAT is stimulated by microtubule interprotofilament contacts. Unexpectedly, despite the confined intraluminal location of Lys40, TAT efficiently scans the microtubule bidirectionally and acetylates stochastically without preference for ends. First-principles modeling and single-molecule measurements demonstrate that TAT catalytic activity, not constrained luminal diffusion, is rate limiting for acetylation. Thus, because of its preference for microtubules over free tubulin and its modest catalytic rate, TAT can function as a slow clock for microtubule lifetimes.

}, issn = {1097-4172}, doi = {10.1016/j.cell.2014.03.061}, author = {Szyk, Agnieszka and Deaconescu, Alexandra M and Spector, Jeffrey and Goodman, Benjamin and Valenstein, Max L and Ziolkowska, Natasza E and Kormendi, Vasilisa and Grigorieff, Nikolaus and Roll-Mecak, Antonina} } @article {45373, title = {Multiplexed aberration measurement for deep tissue imaging in vivo.}, journal = {Nature Methods}, year = {2014}, month = {2014 Aug 17}, abstract = {

We describe an adaptive optics method that modulates the intensity or phase of light rays at multiple pupil segments in parallel to determine the sample-induced aberration. Applicable to fluorescent protein-labeled structures of arbitrary complexity, it allowed us to obtain diffraction-limited resolution in various samples in vivo. For the strongly scattering mouse brain, a single aberration correction improved structural and functional imaging of fine neuronal processes over a large imaging volume.

}, issn = {1548-7105}, doi = {10.1038/nmeth.3068}, author = {Wang, Chen and Liu, Rui and Milkie, Daniel E and Sun, Wenzhi and Tan, Zhongchao and Kerlin, Aaron and Chen, Tsai-Wen and Kim, Douglas S and Ji, Na} } @article {45811, title = {Mushroom body output neurons encode valence and guide memory-based action selection in Drosophila.}, journal = {eLife}, volume = {4}, year = {2014}, month = {12/2014}, abstract = {

Animals discriminate stimuli, learn their predictive value and use this knowledge to modify their behavior. In Drosophila, the mushroom body (MB) plays a key role in these processes. Sensory stimuli are sparsely represented by \~{}2000 Kenyon cells, which converge onto 34 output neurons (MBONs) of 21 types. We studied the role of MBONs in several associative learning tasks and in sleep regulation, revealing the extent to which information flow is segregated into distinct channels and suggesting possible roles for the multi-layered MBON network. We also show that optogenetic activation of MBONs can, depending on cell type, induce repulsion or attraction in flies. The behavioral effects of MBON perturbation are combinatorial, suggesting that the MBON ensemble collectively represents valence. We propose that local, stimulus-specific dopaminergic modulation selectively alters the balance within the MBON network for those stimuli. Our results suggest that valence encoded by the MBON ensemble biases memory-based action selection.

}, issn = {2050-084X}, doi = {10.7554/eLife.04580}, author = {Aso, Yoshinori and Sitaraman, Divya and Ichinose, Toshiharu and Kaun, Karla R and Vogt, Katrin and Belliart-Gu{\'e}rin, Ghislain and Placais, Pierre-Yves and Robie, Alice A and Yamagata, Nobuhiro and Schnaitmann, Christopher and Rowell, William J and Johnston, Rebecca M and Ngo, Teri-T B and Chen, Nan and Korff, Wyatt and Nitabach, Michael N and Heberlein, Ulrike and Preat, Thomas and Branson, Kristin M and Tanimoto, Hiromu and Rubin, Gerald M} } @article {46297, title = {Novel cell-based odorant sensor elements based on insect odorant receptors.}, journal = {Biosensors \& Bioelectronics}, volume = {65C}, year = {2014}, month = {2014 Oct 17}, pages = {287-294}, abstract = {

Development of cell-based odorant sensor elements combined not only high degree of sensitivity and selectivity but also long-term stability is crucial for their practical applications. Here we report the development of a novel cell-based odorant sensor element that sensitively and selectively detects odorants and displays increased fluorescent intensities over a long period of time. Our odorant sensor elements, based on Sf21 cell lines expressing insect odorant receptors, are sensitive to the level of several tens of parts per billion in solution, can selectively distinguish between different types of odorants based on the odorant selectivity intrinsic to the expressed receptors, and have response times of approximately 13s. Specifically, with the use of Sf21 cells and insect odorant receptors, we demonstrated that the established cell lines stably expressing insect odorant receptors are able to detect odorants with consistent responsiveness for at least 2 months, thus exceeding the short life-span normally associated with cell-based sensors. We also demonstrated the development of a compact odorant sensor chip by integrating the established insect cell lines into a microfluidic chip. The methodology we established in this study, in conjunction with the large repertoire of insect odorant receptors, will aid in the development of practical cell-based odorant sensors for various applications, including food administration and health management.

}, issn = {1873-4235}, doi = {10.1016/j.bios.2014.10.026}, author = {Mitsuno, Hidefumi and Sakurai, Takeshi and Namiki, Shigehiro and Mitsuhashi, Hiroyuki and Kanzaki, Ryohei} } @article {6892, title = {Optimized ratiometric calcium sensors for functional in vivo imaging of neurons and T lymphocytes.}, journal = {Nature Methods}, volume = {11}, year = {2014}, month = {2014 Feb}, pages = {175-82}, abstract = {

The quality of genetically encoded calcium indicators (GECIs) has improved dramatically in recent years, but high-performing ratiometric indicators are still rare. Here we describe a series of fluorescence resonance energy transfer (FRET)-based calcium biosensors with a reduced number of calcium binding sites per sensor. These {\textquoteright}Twitch{\textquoteright} sensors are based on the C-terminal domain of Opsanus troponin C. Their FRET responses were optimized by a large-scale functional screen in bacterial colonies, refined by a secondary screen in rat hippocampal neuron cultures. We tested the in vivo performance of the most sensitive variants in the brain and lymph nodes of mice. The sensitivity of the Twitch sensors matched that of synthetic calcium dyes and allowed visualization of tonic action potential firing in neurons and high resolution functional tracking of T lymphocytes. Given their ratiometric readout, their brightness, large dynamic range and linear response properties, Twitch sensors represent versatile tools for neuroscience and immunology.

}, issn = {1548-7105}, doi = {10.1038/nmeth.2773}, author = {Thestrup, Thomas and Litzlbauer, Julia and Bartholom{\"a}us, Ingo and Mues, Marsilius and Russo, Luigi and Dana, Hod and Kovalchuk, Yuri and Liang, Yajie and Kalamakis, Georgios and Laukat, Yvonne and Becker, Stefan and Witte, Gregor and Geiger, Anselm and Allen, Taylor and Rome, Lawrence C and Chen, Tsai-Wen and Kim, Douglas S. and Garaschuk, Olga and Griesinger, Christian and Griesbeck, Oliver} } @article {45719, title = {The PAR complex controls the spatiotemporal dynamics of F-actin and the MTOC in directionally migrating leukocytes.}, journal = {Journal of Cell Science}, volume = {127}, year = {2014}, month = {2014 Oct 15}, pages = {4381-95}, abstract = {

Inflammatory cells acquire a polarized phenotype to migrate towards sites of infection or injury. A conserved polarity complex comprising PAR-3, PAR-6 and atypical protein kinase C (aPKC) relays extracellular polarizing cues to control cytoskeletal and signaling networks affecting morphological and functional polarization. However, there is no evidence that myeloid cells use PAR signaling to migrate vectorially in three-dimensional (3D) environments in vivo. Using genetically encoded bioprobes and high-resolution live imaging, we reveal the existence of F-actin oscillations in the trailing edge and constant repositioning of the microtubule organizing center (MTOC) to direct leukocyte migration in wounded medaka fish larvae (Oryzias latipes). Genetic manipulation in live myeloid cells demonstrates that the catalytic activity of aPKC and the regulated interaction with PAR-3 and PAR-6 are required for consistent F-actin oscillations, MTOC perinuclear mobility, aPKC repositioning and wound-directed migration upstream of Rho kinase (also known as ROCK or ROK) activation. We propose that the PAR complex coordinately controls cytoskeletal changes affecting both the generation of traction force and the directionality of leukocyte migration to sites of injury.

}, issn = {1477-9137}, doi = {10.1242/jcs.146217}, author = {Crespo, Carolina and Vernieri, Claudio and Keller, Philipp J and Garr{\`e}, Massimiliano and Bender, Jeffrey R and Wittbrodt, Joachim and Pardi, Ruggero} } @article {6717, title = {Population genomics of the honey bee reveals strong signatures of positive selection on worker traits.}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {111}, year = {2014}, month = {2014 Jan 31}, pages = {2614-19}, abstract = {

Most theories used to explain the evolution of eusociality rest upon two key assumptions: mutations affecting the phenotype of sterile workers evolve by positive selection if the resulting traits benefit fertile kin, and that worker traits provide the primary mechanism allowing social insects to adapt to their environment. Despite the common view that positive selection drives phenotypic evolution of workers, we know very little about the prevalence of positive selection acting on the genomes of eusocial insects. We mapped the footprints of positive selection in Apis mellifera through analysis of 40 individual genomes, allowing us to identify thousands of genes and regulatory sequences with signatures of adaptive evolution over multiple timescales. We found Apoidea- and Apis-specific genes to be enriched for signatures of positive selection, indicating that novel genes play a disproportionately large role in adaptive evolution of eusocial insects. Worker-biased proteins have higher signatures of adaptive evolution relative to queen-biased proteins, supporting the view that worker traits are key to adaptation. We also found genes regulating worker division of labor to be enriched for signs of positive selection. Finally, genes associated with worker behavior based on analysis of brain gene expression were highly enriched for adaptive protein and cis-regulatory evolution. Our study highlights the significant contribution of worker phenotypes to adaptive evolution in social insects, and provides a wealth of knowledge on the loci that influence fitness in honey bees.

}, issn = {1091-6490}, doi = {10.1073/pnas.1315506111}, author = {Harpur, Brock A and Kent, Clement F and Molodtsova, Daria and Lebon, Jonathan M D and Alqarni, Abdulaziz S and Owayss, Ayman A and Zayed, Amro} } @inbook {65784, title = {Porphyrins and Metalloporphyrins at Components in Artificial Photosynthesis Research}, booktitle = {Handbook of Porphyrin Science}, volume = {34}, year = {2014}, pages = {196 {\textendash} 233}, publisher = {World Scientific Pulishing}, organization = {World Scientific Pulishing}, isbn = {978-981-4425-10-0}, doi = {10.1142/9789814417297_0016}, author = {Tebo, Alison G. and Herrero, C and Aukauloo, A}, editor = {Kadish, Karl M and Smith, Kevin M and undefined, and Guilard, Roger} } @article {45067, title = {Precise spatial coding is preserved along the longitudinal hippocampal axis.}, journal = {Hippocampus}, volume = {24}, year = {2014}, month = {2014 Jul 18}, pages = {1-16}, abstract = {

Compared to the dorsal hippocampus, relatively few studies have characterized neuronal responses in the ventral hippocampus. In particular, it is unclear whether and how cells in the ventral region represent space and/or respond to contextual changes. We recorded from dorsal and ventral CA1 neurons in freely moving mice exposed to manipulations of visuospatial and olfactory contexts. We found that ventral cells respond to alterations of the visuospatial environment such as exposure to novel local cues, cue rotations, and contextual expansion in similar ways to dorsal cells, with the exception of cue rotations. Furthermore, we found that ventral cells responded to odors much more strongly than dorsal cells, particularly to odors of high valence. Similar to earlier studies recording from the ventral hippocampus in CA3, we also found increased scaling of place cell field size along the longitudinal hippocampal axis. Although the increase in place field size observed toward the ventral pole has previously been taken to suggest a decrease in spatial information coded by ventral place cells, we hypothesized that a change in spatial scaling could instead signal a shift in representational coding that preserves the resolution of spatial information. To explore this possibility, we examined population activity using principal component analysis (PCA) and neural location reconstruction techniques. Our results suggest that ventral populations encode a distributed representation of space, and that the resolution of spatial information at the population level is comparable to that of dorsal populations of similar size. Finally, through the use of neural network modeling, we suggest that the redundancy in spatial representation along the longitudinal hippocampal axis may allow the hippocampus to overcome the conflict between memory interference and generalization inherent in neural network memory. Our results suggest that ventral population activity is well suited for generalization across locations and contexts. {\textcopyright} 2014 Wiley Periodicals, Inc.

}, issn = {1098-1063}, doi = {10.1002/hipo.22333}, author = {Keinath, Alexander T and Wang, Melissa E and Wann, Ellen G and Yuan, Robin K and Dudman, Joshua T and Muzzio, Isabel A} } @article {6897, title = {Procedures for behavioral experiments in head-fixed mice.}, journal = {PLoS One}, volume = {9}, year = {2014}, month = {2014}, pages = {e88678}, abstract = {

The mouse is an increasingly prominent model for the analysis of mammalian neuronal circuits. Neural circuits ultimately have to be probed during behaviors that engage the circuits. Linking circuit dynamics to behavior requires precise control of sensory stimuli and measurement of body movements. Head-fixation has been used for behavioral research, particularly in non-human primates, to facilitate precise stimulus control, behavioral monitoring and neural recording. However, choice-based, perceptual decision tasks by head-fixed mice have only recently been introduced. Training mice relies on motivating mice using water restriction. Here we describe procedures for head-fixation, water restriction and behavioral training for head-fixed mice, with a focus on active, whisker-based tactile behaviors. In these experiments mice had restricted access to water (typically 1 ml/day). After ten days of water restriction, body weight stabilized at approximately 80\% of initial weight. At that point mice were trained to discriminate sensory stimuli using operant conditioning. Head-fixed mice reported stimuli by licking in go/no-go tasks and also using a forced choice paradigm using a dual lickport. In some cases mice learned to discriminate sensory stimuli in a few trials within the first behavioral session. Delay epochs lasting a second or more were used to separate sensation (e.g. tactile exploration) and action (i.e. licking). Mice performed a variety of perceptual decision tasks with high performance for hundreds of trials per behavioral session. Up to four months of continuous water restriction showed no adverse health effects. Behavioral performance correlated with the degree of water restriction, supporting the importance of controlling access to water. These behavioral paradigms can be combined with cellular resolution imaging, random access photostimulation, and whole cell recordings.

}, issn = {1932-6203}, doi = {10.1371/journal.pone.0088678}, author = {Guo, Zengcai V and Hires, S Andrew and Li, Nuo and O{\textquoteright}Connor, Daniel H and Komiyama, Takaki and Ophir, Eran and Huber, Daniel and Bonardi, Claudia and Morandell, Karin and Gutnisky, Diego and Peron, Simon and Xu, Ning-long and Cox, James and Svoboda, Karel} } @article {46062, title = {Quantifying histone and RNA polymerase II post-translational modification dynamics in mother and daughter cells.}, journal = {Methods}, volume = {70}, year = {2014}, month = {2014 Dec}, pages = {77-88}, abstract = {

Post-translational histone modifications are highly correlated with transcriptional activity, but the relative timing of these marks and their dynamic interplay during gene regulation remains controversial. To shed light on this problem and clarify the connections between histone modifications and transcription, we demonstrate how FabLEM (Fab-based Live Endogenous Modification labeling) can be used to simultaneously track histone H3 Lysine 9 acetylation (H3K9ac) together with RNA polymerase II Serine 2 and Serine 5 phosphorylation (RNAP2 Ser2ph/Ser5ph) in single living cells and their progeny. We provide a detailed description of the FabLEM methodology, including helpful tips for preparing and loading fluorescently conjugated antigen binding fragments (Fab) into cells for optimal results. We also introduce simple procedures for analyzing and visualizing FabLEM data, including color-coded scatterplots to track correlations between modifications through the cell cycle and temporal cross-correlation analysis to dissect modification dynamics. Using these methods, we find significant correlations that span cell generations, with a relatively strong correlation between H3K9ac and Ser5ph that appears to peak a few hours before mitosis and may reflect the bookmarking of genes for efficient re-initiation following mitosis. The techniques we have developed are broadly applicable and should help clarify how histone modifications dynamically contribute to gene regulation.

}, issn = {1095-9130}, doi = {10.1016/j.ymeth.2014.08.002}, author = {Stasevich, Timothy J and Sato, Yuko and Nozaki, Naohito and Kimura, Hiroshi} } @article {45412, title = {Reactivation of developmentally silenced globin genes by forced chromatin looping.}, journal = {Cell}, volume = {158}, year = {2014}, month = {2014 Aug 14}, pages = {849-60}, abstract = {

Distal enhancers commonly contact target promoters via chromatin looping. In erythroid cells, the locus control region (LCR) contacts β-type globin genes in a developmental stage-specific manner to\ stimulate transcription. Previously, we induced LCR-promoter looping by tethering the self-association domain (SA) of Ldb1 to the β-globin promoter via artificial zinc fingers. Here, we show that targeting the SA to a developmentally silenced embryonic globin gene in adult murine erythroblasts triggers its transcriptional reactivation. This activity depends on the LCR, consistent with an LCR-promoter looping mechanism. Strikingly, targeting the SA to the fetal γ-globin promoter in primary adult human erythroblasts increases γ-globin promoter-LCR contacts, stimulating transcription to approximately 85\% of total β-globin synthesis, with a reciprocal reduction in adult β-globin expression. Our findings demonstrate that forced chromatin looping can override a stringent developmental gene expression program and suggest a novel approach to control the balance\ of globin gene transcription for therapeutic applications.

}, issn = {1097-4172}, doi = {10.1016/j.cell.2014.05.050}, author = {Deng, Wulan and Rupon, Jeremy W and Krivega, Ivan and Breda, Laura and Motta, Irene and Jahn, Kristen S and Reik, Andreas and Gregory, Philip D and Rivella, Stefano and Dean, Ann and Blobel, Gerd A} } @article {45038, title = {Reconciling the deep homology of neuromodulation with the evolution of behavior.}, journal = {Current Opinion in Neurobiology}, volume = {29C}, year = {2014}, month = {2014 May 27}, pages = {39-47}, abstract = {

The evolution of behavior seems inconsistent with the deep homology of neuromodulatory signaling. G protein coupled receptors (GPCRs) evolved slowly from a common ancestor through a process involving gene duplication, neofunctionalization, and loss. Neuropeptides co-evolved with their receptors and exhibit many conserved functions. Furthermore, brain areas are highly conserved with suggestions of deep anatomical homology between arthropods and vertebrates. Yet, behavior evolved more rapidly; even members of the same genus or species can differ in heritable behavior. The solution to the paradox involves changes in the compartmentalization, or subfunctionalization, of neuromodulation; neurons shift their expression of GPCRs and the content of monoamines and neuropeptides. Furthermore, parallel evolution of neuromodulatory signaling systems suggests a route for repeated evolution of similar behaviors.

}, issn = {1873-6882}, doi = {10.1016/j.conb.2014.05.002}, author = {Katz, Paul S and Lillvis, Joshua L} } @article {45843, title = {Regulation of RNA polymerase II activation by histone acetylation in single living cells.}, journal = {Nature}, volume = {516}, year = {2014}, month = {2014 Dec 11}, pages = {272-5}, abstract = {

In eukaryotic cells, post-translational histone modifications have an important role in gene regulation. Starting with early work on histone acetylation, a variety of residue-specific modifications have now been linked to RNA polymerase II (RNAP2) activity, but it remains unclear if these markers are active regulators of transcription or just passive byproducts. This is because studies have traditionally relied on fixed cell populations, meaning temporal resolution is limited to minutes at best, and correlated factors may not actually be present in the same cell at the same time. Complementary approaches are therefore needed to probe the dynamic interplay of histone modifications and RNAP2 with higher temporal resolution in single living cells. Here we address this problem by developing a system to track residue-specific histone modifications and RNAP2 phosphorylation in living cells by fluorescence microscopy. This increases temporal resolution to the tens-of-seconds range. Our single-cell analysis reveals histone H3 lysine-27 acetylation at a gene locus can alter downstream transcription kinetics by as much as 50\%, affecting two temporally separate events. First acetylation enhances the search kinetics of transcriptional activators, and later the acetylation accelerates the transition of RNAP2 from initiation to elongation. Signatures of the latter can be found genome-wide using chromatin immunoprecipitation followed by sequencing. We argue that this regulation leads to a robust and potentially tunable transcriptional response.

}, issn = {1476-4687}, doi = {10.1038/nature13714}, author = {Stasevich, Timothy J and Hayashi-Takanaka, Yoko and Sato, Yuko and Maehara, Kazumitsu and Ohkawa, Yasuyuki and Sakata-Sogawa, Kumiko and Tokunaga, Makio and Nagase, Takahiro and Nozaki, Naohito and McNally, James G and Kimura, Hiroshi} } @article {6961, title = {Reproductive status, endocrine physiology and chemical signaling in the Neotropical, swarm-founding eusocial wasp, Polybia micans Ducke (Vespidae: Epiponini).}, journal = {The Journal of Experimental Biology}, volume = {217}, year = {2014}, month = {2014 Apr 17}, pages = {2399-410}, abstract = {

In the evolution of caste-based societies in Hymenoptera, the classical insect hormones, juvenile hormone (JH) and ecdysteroids, were co-opted into new functions. Social wasps, which show all levels of sociality and lifestyles, are an ideal group to study such functional changes. Virtually all studies on the physiological mechanisms underlying reproductive division of labor and caste functions in wasps have been done on independent-founding paper wasps, and the majority of these studies have focused on species specially adapted for overwintering. The relatively little studied tropical swarming-founding wasps of the Epiponini (Vespidae) are a diverse group of permanently social wasps, with some species maintaining caste flexibility well into the adult phase. We investigated the behavior, reproductive status, JH and ecdysteroid titers in hemolymph, ecdysteroid content of the ovary and cuticular hydrocarbon (CHC) profiles in the caste-monomorphic, epiponine wasp Polybia micans Ducke. We found that the JH titer was not elevated in competing queens from established multiple-queen nests, but increased in lone queens that lack direct competition. In queenless colonies, JH titers rose transiently in young potential reproductives upon challenge by nestmates, suggesting that JH may prime the ovaries for further development. Ovarian ecdysteroids were very low in workers but higher and correlated with the number of vitellogenic oocytes in the queens. Hemolymph ecdysteroid levels were low and variable in both. Profiles of P. micans CHCs reflected caste, age and reproductive status, but were not tightly linked to either hormone. These findings show a significant divergence in hormone function in swarm-founding wasps compared to independent-founding ones.

}, issn = {1477-9145}, doi = {10.1242/jeb.096750}, author = {Kelstrup, Hans C and Hartfelder, Klaus and Nascimento, Fabio S and Riddiford, Lynn M} } @article {65887, title = {Retrograde Plasticity and Differential Competition of Bipolar Cell Dendrites and Axons in the Developing Retina}, journal = {Current Biology}, volume = {24}, year = {2014}, month = {Jan/2014}, pages = {2301 - 2306}, abstract = {

Most neurons function in the context of pathways that process and propagate information through a series of stages, e.g., from the sensory periphery to cerebral cortex. Because activity at each stage of a neural pathway depends on connectivity at the preceding one, we hypothesized that during development, axonal output of a neuron may regulate synaptic development of its dendrites (i.e., retrograde plasticity). Within pathways, neurons often receive input from multiple partners and provide output to targets shared with other neurons (i.e., convergence). Converging axons can intermingle or occupy separate territories on target dendrites. Activity-dependent competition has been shown to bias target innervation by overlapping axons in several systems. By contrast, whether territorial axons or dendrites compete for targets and inputs, respectively, has not been tested. Here, we generate transgenic mice in which glutamate release from specific sets of retinal bipolar cells (BCs) is suppressed. We find that dendrites of silenced BCs recruit fewer inputs when their neighbors are active and that dendrites of active BCs recruit more inputs when their neighbors are silenced than either active or silenced BCs with equal neighbors. By contrast, axons of silenced BCs form fewer synapses with their targets, irrespective of the activity of their neighbors. These findings reveal that retrograde plasticity guides BC dendritic development in vivo and demonstrate that dendrites, but not territorial axons, in a convergent neural pathway engage in activity-dependent competition. We propose that at a population level, retrograde plasticity serves to maximize functional representation of inputs.

}, issn = {09609822}, doi = {10.1016/j.cub.2014.08.018}, url = {https://linkinghub.elsevier.com/retrieve/pii/S0960982214010318https://api.elsevier.com/content/article/PII:S0960982214010318?httpAccept=text/xmlhttps://api.elsevier.com/content/article/PII:S0960982214010318?httpAccept=text/plain}, author = {Johnson, Robert~E. and Kerschensteiner, Daniel} } @article {45785, title = {The role of juvenile hormone in dominance behavior, reproduction and cuticular pheromone signaling in the caste-flexible epiponine wasp, Synoeca surinama.}, journal = {Frontiers in Zoology}, volume = {11}, year = {2014}, month = {2014}, pages = {78}, abstract = {

BACKGROUND: The popular view on insect sociality is that of a harmonious division of labor among two morphologically distinct and functionally non-overlapping castes. But this is a highly derived state and not a prerequisite for a functional society. Rather, caste-flexibility is a central feature in many eusocial wasps, where adult females have the potential to become queens or workers, depending on the social environment. In non-swarming paper wasps (e.g., Polistes), prospective queens fight one another to assert their dominance, with losers becoming workers if they remain on the nest. This aggression is fueled by juvenile hormone (JH) and ecdysteroids, major factors involved in caste differentiation in most eusocial insects. We tested whether these hormones have conserved aggression-promoting functions in Synoeca surinama, a caste-flexible swarm-founding wasp (Epiponini) where reproductive competition is high and aggressive displays are common.

RESULTS: We observed the behavioral interactions of S. surinama females in field nests before and after we had removed the egg-laying queen(s). We measured the ovarian reproductive status, hemolymph JH and ecdysteroid titers, ovarian ecdysteroid content, and analyzed the cuticular hydrocarbon (CHC) composition of females engaged in competitive interactions in both queenright and queenless contexts. These data, in combination with hormone manipulation experiments, revealed that neither JH nor ecdysteroids are necessary for the expression of dominance behaviors in S. surinama. Instead, we show that JH likely functions as a gonadotropin and directly modifies the cuticular hydrocarbon blend of young workers to match that of a reproductive. Hemolymph ecdysteroids, in contrast, are not different between queens and workers despite great differences in ovarian ecdysteroid content.

CONCLUSIONS: The endocrine profile of S. surinama shows surprising differences from those of other caste-flexible wasps, although a rise in JH titers in replacement queens is a common theme. Extensive remodeling of hormone functions is also evident in the highly eusocial bees, which has been attributed to the evolution of morphologically defined castes. Our results show that hormones which regulate caste-plasticity can lose these roles even while caste-plasticity is preserved.

}, issn = {1742-9994}, doi = {10.1186/s12983-014-0078-5}, author = {Kelstrup, Hans C and Hartfelder, Klaus and Nascimento, Fabio S and Riddiford, Lynn M} } @article {45416, title = {Shared mushroom body circuits underlie visual and olfactory memories in Drosophila.}, journal = {eLife}, volume = {3}, year = {2014}, month = {2014}, pages = {e02395}, abstract = {

In nature, animals form memories associating reward or punishment with stimuli from different sensory modalities, such as smells and colors. It is unclear, however, how distinct sensory memories are processed in the brain. We established appetitive and aversive visual learning assays for Drosophila that are comparable to the widely used olfactory learning assays. These assays share critical features, such as reinforcing stimuli (sugar reward and electric shock punishment), and allow direct comparison of the cellular requirements for visual and olfactory memories. We found that the same subsets of dopamine neurons drive formation of both sensory memories. Furthermore, distinct yet partially overlapping subsets of mushroom body intrinsic neurons are required for visual and olfactory memories. Thus, our results suggest that distinct sensory memories are processed in a common brain center. Such centralization of related brain functions is an economical design that avoids the repetition of similar circuit motifs.

}, issn = {2050-084X}, doi = {10.7554/eLife.02395}, author = {Vogt, Katrin and Schnaitmann, Christopher and Dylla, Kristina V and Knapek, Stephan and Aso, Yoshinori and Rubin, Gerald M and Tanimoto, Hiromu} } @article {46327, title = {Space-time wiring specificity supports direction selectivity in the retina.}, journal = {Nature}, volume = {509}, year = {2014}, month = {2014 May 15}, pages = {331-6}, abstract = {

How does the mammalian retina detect motion? This classic problem in visual neuroscience has remained unsolved for 50 years. In search of clues, here we reconstruct Off-type starburst amacrine cells (SACs) and bipolar cells (BCs) in serial electron microscopic images with help from EyeWire, an online community of {\textquoteright}citizen neuroscientists{\textquoteright}. On the basis of quantitative analyses of contact area and branch depth in the retina, we find evidence that one BC type prefers to wire with a SAC dendrite near the SAC soma, whereas another BC type prefers to wire far from the soma. The near type is known to lag the far type in time of visual response. A mathematical model shows how such {\textquoteright}space-time wiring specificity{\textquoteright} could endow SAC dendrites with receptive fields that are oriented in space-time and therefore respond selectively to stimuli that move in the outward direction from the soma.

}, keywords = {Amacrine Cells, Animals, Artificial Intelligence, Brain Mapping, Crowdsourcing, Dendrites, Mice, Models, Neurological, Motion, Neural Pathways, Presynaptic Terminals, Retina, Retinal Bipolar Cells, Spatio-Temporal Analysis}, issn = {1476-4687}, doi = {10.1038/nature13240}, author = {Kim, Jinseop S and Greene, Matthew J and Zlateski, Aleksandar and Lee, Kisuk and Richardson, Mark and Turaga, Srinivas C and Purcaro, Michael and Balkam, Matthew and Robinson, Amy and Behabadi, Bardia F and Campos, Michael and Denk, Winfried and Seung, H Sebastian} } @article {6963, title = {Structural plasticity within the barrel cortex during initial phases of whisker-dependent learning.}, journal = {The Journal of Neuroscience}, volume = {34}, year = {2014}, month = {2014 Apr 23}, pages = {6078-83}, abstract = {

We report learning-related structural plasticity in layer 1 branches of pyramidal neurons in the barrel cortex, a known site of sensorimotor integration. In mice learning an active, whisker-dependent object localization task, layer 2/3 neurons showed enhanced spine growth during initial skill acquisition that both preceded and predicted expert performance. Preexisting spines were stabilized and new persistent spines were formed. These findings suggest rapid changes in connectivity between motor centers and sensory cortex guide subsequent sensorimotor learning.

}, issn = {1529-2401}, doi = {10.1523/JNEUROSCI.4919-12.2014}, author = {Kuhlman, Sandra J and O{\textquoteright}Connor, Daniel H and Fox, Kevin and Svoboda, Karel} } @article {6713, title = {Structured synaptic connectivity between hippocampal regions.}, journal = {Neuron}, volume = {81}, year = {2014}, month = {2014 Feb 5}, pages = {629-40}, abstract = {

The organization of synaptic connectivity within a neuronal circuit is a prime determinant of circuit function. We performed a comprehensive fine-scale circuit mapping of hippocampal regions (CA3-CA1) using the newly developed synapse labeling method, mGRASP. This mapping revealed spatially nonuniform and clustered synaptic connectivity patterns. Furthermore, synaptic clustering was enhanced between groups of neurons that shared a similar developmental/migration time window, suggesting a mechanism for establishing the spatial structure of synaptic connectivity. Such connectivity patterns are thought to effectively engage active dendritic processing and storage mechanisms, thereby potentially enhancing neuronal feature selectivity.

}, issn = {1097-4199}, doi = {10.1016/j.neuron.2013.11.026}, author = {Shaul Druckmann and Feng, Linqing and Lee, Bokyoung and Yook, Chaehyun and Zhao, Ting and Magee, Jeffrey C and Kim, Jinhyun} } @article {6884, title = {A systematic nomenclature for the insect brain.}, journal = {Neuron}, volume = {81}, year = {2014}, month = {2014 Feb 19}, pages = {755-65}, abstract = {

Despite the importance of the insect nervous system for functional and developmental neuroscience, descriptions of insect brains have suffered from a lack of uniform nomenclature. Ambiguous definitions of brain regions and fiber bundles have contributed to the variation of names used to describe the same structure. The lack of clearly determined neuropil boundaries has made it difficult to document precise locations of neuronal projections for connectomics study. To address such issues, a consortium of\ neurobiologists studying arthropod brains, the Insect Brain Name Working Group, has established the present hierarchical nomenclature system, using the brain of Drosophila melanogaster as the reference framework, while taking the brains of other taxa into careful consideration for maximum consistency and expandability. The following summarizes the consortium{\textquoteright}s nomenclature system and highlights examples of existing ambiguities and remedies for them. This nomenclature is intended to serve as a\ standard of reference for the study of the brain of Drosophila and other insects.

}, issn = {1097-4199}, doi = {10.1016/j.neuron.2013.12.017}, author = {Ito, Kei and Shinomiya, Kazunori and Ito, Masayoshi and Armstrong, J Douglas and Boyan, George and Hartenstein, Volker and Harzsch, Steffen and Heisenberg, Martin and Homberg, Uwe and Jenett, Arnim and Keshishian, Haig and Restifo, Linda L and R{\"o}ssler, Wolfgang and Simpson, Julie H and Strausfeld, Nicholas J and Strauss, Roland and Vosshall, Leslie B} } @article {65021, title = {Taura syndrome virus IRES initiates translation by binding its tRNA-mRNA-like structural element in the ribosomal decoding center.}, journal = {Proc Natl Acad Sci U S A}, volume = {111}, year = {2014}, month = {2014 Jun 24}, pages = {9139-44}, abstract = {

In cap-dependent translation initiation, the open reading frame (ORF) of mRNA is established by the placement of the AUG start codon and initiator tRNA in the ribosomal peptidyl (P) site. Internal ribosome entry sites (IRESs) promote translation of mRNAs in a cap-independent manner. We report two structures of the ribosome-bound Taura syndrome virus (TSV) IRES belonging to the family of Dicistroviridae intergenic IRESs. Intersubunit rotational states differ in these structures, suggesting that ribosome dynamics play a role in IRES translocation. Pseudoknot I of the IRES occupies the ribosomal decoding center at the aminoacyl (A) site in a manner resembling that of the tRNA anticodon-mRNA codon. The structures reveal that the TSV IRES initiates translation by a previously unseen mechanism, which is conceptually distinct from initiator tRNA-dependent mechanisms. Specifically, the ORF of the IRES-driven mRNA is established by the placement of the preceding tRNA-mRNA-like structure in the A site, whereas the 40S P site remains unoccupied during this initial step.

}, keywords = {Nucleic Acid Conformation, Open Reading Frames, Peptide Chain Initiation, Translational, Picornaviridae, Ribosomes, RNA, Messenger, RNA, Transfer, RNA, Viral}, issn = {1091-6490}, doi = {10.1073/pnas.1406335111}, author = {Koh, Cha San and Brilot, Axel F and Grigorieff, Nikolaus and Korostelev, Andrei A} } @article {45353, title = {Thy1 - GCaMP6 transgenic mice for neuronal population imaging in vivo.}, journal = {PLoS One}, volume = {9}, year = {2014}, month = {2014}, pages = {e108697}, chapter = {e108697}, abstract = {

Genetically-encoded calcium indicators (GECIs) facilitate imaging activity of genetically defined neuronal populations in vivo. The high intracellular GECI concentrations required for in vivo imaging are usually achieved by viral gene transfer using adeno-associated viruses. Transgenic expression of GECIs promises important advantages, including homogeneous, repeatable, and stable expression without the need for invasive virus injections. Here we present the generation and characterization of transgenic mice expressing the GECIs GCaMP6s or GCaMP6f under the Thy1 promoter. We quantified GCaMP6 expression across brain regions and neurons and compared to other transgenic mice and AAV-mediated expression. We tested three mouse lines for imaging in the visual cortex in vivo and compared their performance to mice injected with AAV expressing GCaMP6. Furthermore, we show that GCaMP6 Thy1 transgenic mice are useful for long-term, high-sensitivity imaging in behaving mice.

}, issn = {1932-6203}, doi = {10.1371/journal.pone.0108697}, author = {Dana, Hod and Chen, Tsai-Wen and Hu, Amy and Shields, Brenda C and Cui, Guohong and Looger, Loren and Kim, Douglas S. and Svoboda, Karel} } @conference {48180, title = {Tracking by assignment facilitates data curation.}, booktitle = {Medical Image Computing and Computer-Assisted Intervention {\textendash} MICCAI Workshop 2014}, year = {2014}, month = {2014 Sep 14}, address = {Boston, MA}, abstract = {

Object tracking is essential for a multitude of biomedical re- search projects. Automated methods are desired in order to avoid im- possible amounts of manual tracking efforts. However, automatically found solutions are not free of errors, and these errors again have to be identified and resolved manually. We propose six innovative ways for semi-automatic curation of automatically found tracking solutions. Respective user interactions are six simple operations: Inclusion and ex- clusion of objects and tracking decisions, specification of the number of objects, and one-click altering of object segmentations. We show how all proposed interactions can be elegantly incorporated into {\textquotedblleft}assignment models{\textquotedblright} [1,2,3,4,5,6], an innovative and increasingly popular tracking paradigm. Given some user interaction, the tracking engine is capable of computing the respective globally optimal tracking solution efficiently, even benefitting from {\textquotedblleft}warm start{\textquotedblright}-capabilities. We show that after in- teractively pointing at a single mistake, multiple segmentation and track- ing errors can be fixed automatically in one single re-evaluation, provably leading to the new, feedback-conscious global optimum.\ 

}, url = {http://interactivemedical.org/imic2014/CameraReadyPapers/Paper\%203/Jug_Interactive_IMIC2014.pdf}, author = {Jug, Florian and Tobias Pietzsch and Kainmueller, Dagmar and Myers, Eugene W} } @article {46377, title = {Transgenesis in non-model organisms: the case of Parhyale.}, journal = {Methods Mol Biol}, volume = {1196}, year = {2014}, month = {2014}, pages = {145-81}, abstract = {

One of the most striking manifestations of Hox gene activity is the morphological and functional diversity of arthropod body plans, segments, and associated appendages. Among arthropod models, the amphipod crustacean Parhyale hawaiensis satisfies a number of appealing biological and technical requirements to study the Hox control of tissue and organ morphogenesis. Parhyale embryos undergo direct development from fertilized eggs into miniature adults within 10 days and are amenable to all sorts of embryological and functional genetic manipulations. Furthermore, each embryo develops a series of specialized appendages along the anterior-posterior body axis, offering exceptional material to probe the genetic basis of appendage patterning, growth, and differentiation. Here, we describe the methodologies and techniques required for transgenesis-based gain-of-function studies of Hox genes in Parhyale embryos. First, we introduce a protocol for efficient microinjection of early-stage Parhyale embryos. Second, we describe the application of fast and reliable assays to test the activity of the Minos DNA transposon in embryos. Third, we present the use of Minos-based transgenesis vectors to generate stable and transient transgenic Parhyale. Finally, we describe the development and application of a conditional heat-inducible misexpression system to study the role of the Hox gene Ultrabithorax in Parhyale appendage specialization. Beyond providing a useful resource for Parhyalists, this chapter also aims to provide a road map for researchers working on other emerging model organisms. Acknowledging the time and effort that need to be invested in developing transgenic approaches in new species, it is all worth it considering the wide scope of experimentation that opens up once transgenesis is established.

}, issn = {1940-6029}, doi = {10.1007/978-1-4939-1242-1_10}, author = {Kontarakis, Zacharias and Pavlopoulos, Anastasios} } @article {46284, title = {Word length effect in free recall of randomly assembled word lists.}, journal = {Frontiers in Computational Neuroscience}, volume = {8}, year = {2014}, month = {2014 Oct 14}, pages = {129}, abstract = {

In serial recall experiments, human subjects are requested to retrieve a list of words in the same order as they were presented. In a classical study, participants were reported to recall more words from study lists composed of short words compared to lists of long words, the word length effect. The world length effect was also observed in free recall experiments, where subjects can retrieve the words in any order. Here we analyzed a large dataset from free recall experiments of unrelated words, where short and long words were randomly mixed, and found a seemingly opposite effect: long words are recalled better than the short ones. We show that our recently proposed mechanism of associative retrieval can explain both these observations. Moreover, the direction of the effect depends solely on the way study lists are composed.

}, issn = {1662-5188}, doi = {10.3389/fncom.2014.00129}, author = {Katkov, Mikhail and Romani, Sandro and Tsodyks, Misha} } @conference {5905, title = {3D Haar-like elliptical features for object classification in microscopy.}, booktitle = {International Symposium on Biomedical Imaging}, year = {2013}, month = {2013 Apr}, abstract = {

Object detection and classification are key tasks in computer vision that can facilitate high-throughput image analysis of microscopy data. We present a set of local image descriptors for three-dimensional (3D) microscopy datasets inspired by the well-known Haar wavelet framework. We add orientation, illumination and scale information by assuming that the neighborhood surrounding points of interests in the image can be described with ellipsoids, and we increase discriminative power by incorporating edge and shape information into the features. The calculation of the local image descriptors is implemented in a Graphics Processing Unit (GPU) in order to reduce computation time to 1 millisecond per object of interest. We present results for cell division detection in 3D time-lapse fluorescence microscopy with 97.6\% accuracy.

}, author = {Amat, Fernando and Keller, Philipp J} } @article {6127, title = {Admixture mapping in lupus identifies multiple functional variants within IFIH1 associated with apoptosis, inflammation, and autoantibody production.}, journal = {PLoS Genetics}, volume = {9}, year = {2013}, month = {2013 Feb 18}, pages = {e1003222}, abstract = {

Systemic lupus erythematosus (SLE) is an inflammatory autoimmune disease with a strong genetic component. African-Americans (AA) are at increased risk of SLE, but the genetic basis of this risk is largely unknown. To identify causal variants in SLE loci in AA, we performed admixture mapping followed by fine mapping in AA and European-Americans (EA). Through genome-wide admixture mapping in AA, we identified a strong SLE susceptibility locus at 2q22-24 (LOD=6.28), and the admixture signal is associated with the European ancestry (ancestry risk ratio \ 1.5). Large-scale genotypic analysis on 19,726 individuals of African and European ancestry revealed three independently associated variants in the IFIH1 gene: an intronic variant, rs13023380 [P(meta) = 5.20{\texttimes}10(-14); odds ratio, 95\% confidence interval = 0.82 (0.78-0.87)], and two missense variants, rs1990760 (Ala946Thr) [P(meta) = 3.08{\texttimes}10(-7); 0.88 (0.84-0.93)] and rs10930046 (Arg460His) [P(dom) = 1.16{\texttimes}10(-8); 0.70 (0.62-0.79)]. Both missense variants produced dramatic phenotypic changes in apoptosis and inflammation-related gene expression. We experimentally validated function of the intronic SNP by DNA electrophoresis, protein identification, and in vitro protein binding assays. DNA carrying the intronic risk allele rs13023380 showed reduced binding efficiency to a cellular protein complex including nucleolin and lupus autoantigen Ku70/80, and showed reduced transcriptional activity in vivo. Thus, in SLE patients, genetic susceptibility could create a biochemical imbalance that dysregulates nucleolin, Ku70/80, or other nucleic acid regulatory proteins. This could promote antibody hypermutation and auto-antibody generation, further destabilizing the cellular network. Together with molecular modeling, our results establish a distinct role for IFIH1 in apoptosis, inflammation, and autoantibody production, and explain the molecular basis of these three risk alleles for SLE pathogenesis.

}, keywords = {African Americans, Alleles, Antigens, Apoptosis, Autoantibodies, Chromosome Mapping, DEAD-box RNA Helicases, DNA-Binding Proteins, European Continental Ancestry Group, Genetic Predisposition to Disease, Genome, Haplotypes, Human, Humans, Inflammation, Lupus Erythematosus, Nuclear, Polymorphism, Protein Binding, Single Nucleotide, Systemic}, issn = {1553-7404}, doi = {10.1371/journal.pgen.1003222}, author = {Molineros, Julio E and Maiti, Amit K and Sun, Celi and Looger, Loren L and Han, Shizhong and Kim-Howard, Xana and Glenn, Stuart and Adler, Adam and Kelly, Jennifer A and Niewold, Timothy B and Gilkeson, Gary S and Brown, Elizabeth E and Alarc{\'o}n, Graciela S and Edberg, Jeffrey C and Petri, Michelle and Ramsey-Goldman, Rosalind and Reveille, John D and Vil{\'a}, Luis M and Freedman, Barry I and Tsao, Betty P and Criswell, Lindsey A and Jacob, Chaim O and Moore, Jason H and Vyse, Timothy J and Langefeld, Carl L and Guthridge, Joel M and Gaffney, Patrick M and Moser, Kathy L and Scofield, R Hal and Alarc{\'o}n-Riquelme, Marta E and Williams, Scott M and Merrill, Joan T and James, Judith A and Kaufman, Kenneth M and Kimberly, Robert P and Harley, John B and Nath, Swapan K} } @article {6452, title = {ASI regulates satiety quiescence in C. elegans.}, journal = {The Journal of Neuroscience : The Official Journal of the Society for Neuroscience}, volume = {33}, year = {2013}, month = {2013 Jun 5}, pages = {9716-24}, abstract = {

In Caenorhabditis elegans, satiety quiescence mimics behavioral aspects of satiety and postprandial sleep in mammals. On the basis of calcium-imaging, genetics, and behavioral studies, here we report that a pair of amphid neurons, ASI, is activated by nutrition and regulates worms{\textquoteright} behavioral states specifically promoting satiety quiescence; ASI inhibits the switch from quiescence to dwelling (a browsing state) and accelerates the switch from dwelling to quiescence. The canonical TGFβ pathway, whose ligand is released from ASI, regulates satiety quiescence. The mutants of a ligand, a receptor and SMADs in the TGFβ pathway all eat more and show less quiescence than wild-type. The TGFβ receptor in downstream neurons RIM and RIC is sufficient for worms to exhibit satiety quiescence, suggesting neuronal connection from ASI to RIM and RIC is essential for feeding regulation through the TGFβ pathway. ASI also regulates satiety quiescence partly through cGMP signaling; restoring cGMP signaling in ASI rescues the satiety quiescence defect of cGMP signaling mutants. From these results, we propose that TGFβ and cGMP pathways in ASI connect nutritional status to promotion of satiety quiescence, a sleep-like behavioral state.

}, keywords = {Animals, Caenorhabditis elegans, Caenorhabditis elegans Proteins, Cyclic GMP, Eating, Genetically Modified, Motor Activity, Protein Kinases, Satiation, Sensory Receptor Cells, Transforming Growth Factor beta}, issn = {1529-2401}, doi = {10.1523/JNEUROSCI.4493-12.2013}, author = {Gallagher, Thomas and Kim, Jeongho and Oldenbroek, Marieke and Kerr, Rex and You, Young-Jai} } @article {6174, title = {Automated alignment of imperfect EM images for neural reconstruction.}, journal = {arXiv}, year = {2013}, month = {2013 Apr-22}, pages = {arXiv:1304.6034 [q-bio.QM]}, abstract = {

The most established method of reconstructing neural circuits from animals involves slicing tissue very thin, then taking mosaics of electron microscope (EM) images. To trace neurons across different images and through different sections, these images must be accurately aligned, both with the others in the same section and to the sections above and below. Unfortunately, sectioning and imaging are not ideal processes - some of the problems that make alignment difficult include lens distortion, tissue shrinkage during imaging, tears and folds in the sectioned tissue, and dust and other artifacts. In addition the data sets are large (hundreds of thousands of images) and each image must be aligned with many neighbors, so the process must be automated and reliable. This paper discusses methods of dealing with these problems, with numeric results describing the accuracy of the resulting alignments.

}, url = {http://arxiv.org/abs/1304.6034}, author = {Scheffer, L. K. and Karsh, Bill and Vitaladevun, S.} } @article {45850, title = {Automated long-term tracking and social behavioural phenotyping of animal colonies within a semi-natural environment.}, journal = {Nature Communications}, volume = {4}, year = {2013}, month = {2013}, pages = {2018}, abstract = {

Social behaviour has a key role in animal survival across species, ranging from insects to primates and humans. However, the biological mechanisms driving natural interactions between multiple animals, over long-term periods, are poorly studied and remain elusive. Rigorous and objective quantification of behavioural parameters within a group poses a major challenge as it requires simultaneous monitoring of the positions of several individuals and comprehensive consideration of many complex factors. Automatic tracking and phenotyping of interacting animals could thus overcome the limitations of manual tracking methods. Here we report a broadly applicable system that automatically tracks the locations of multiple, uniquely identified animals, such as mice, within a semi-natural setting. The system combines video and radio frequency identified tracking data to obtain detailed behavioural profiles of both individuals and groups. We demonstrate the usefulness of these data in characterizing individual phenotypes, interactions between pairs and the collective social organization of groups.

}, keywords = {Animals, Automation, Behavior, Animal, Crosses, Genetic, Data Collection, Environment, Female, Hierarchy, Social, Locomotion, Male, Mice, Mice, Inbred C57BL, Phenotype, Radio Frequency Identification Device, Social Behavior, Time Factors, Video Recording}, issn = {2041-1723}, doi = {10.1038/ncomms3018}, author = {Weissbrod, Aharon and Shapiro, Alexander and Vasserman, Genadiy and Edry, Liat and Dayan, Molly and Yitzhaky, Assif and Hertzberg, Libi and Feinerman, Ofer and Kimchi, Tali} } @article {6268, title = {Aβ induces astrocytic glutamate release, extrasynaptic NMDA receptor activation, and synaptic loss.}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {110}, year = {2013}, month = {2013 Jul 2}, pages = {E2518-27}, abstract = {

Synaptic loss is the cardinal feature linking neuropathology to cognitive decline in Alzheimer{\textquoteright}s disease (AD). However, the mechanism of synaptic damage remains incompletely understood. Here, using FRET-based glutamate sensor imaging, we show that amyloid-β peptide (Aβ) engages α7 nicotinic acetylcholine receptors to induce release of astrocytic glutamate, which in turn activates extrasynaptic NMDA receptors (eNMDARs) on neurons. In hippocampal autapses, this eNMDAR activity is followed by reduction in evoked and miniature excitatory postsynaptic currents (mEPSCs). Decreased mEPSC frequency may reflect early synaptic injury because of concurrent eNMDAR-mediated NO production, tau phosphorylation, and caspase-3 activation, each of which is implicated in spine loss. In hippocampal slices, oligomeric Aβ induces eNMDAR-mediated synaptic depression. In AD-transgenic mice compared with wild type, whole-cell recordings revealed excessive tonic eNMDAR activity accompanied by eNMDAR-sensitive loss of mEPSCs. Importantly, the improved NMDAR antagonist NitroMemantine, which selectively inhibits extrasynaptic over physiological synaptic NMDAR activity, protects synapses from Aβ-induced damage both in vitro and in vivo.

}, issn = {1091-6490}, doi = {10.1073/pnas.1306832110}, author = {Talantova, Maria and Sanz-Blasco, Sara and Zhang, Xiaofei and Xia, Peng and Akhtar, Mohd Waseem and Okamoto, Shu-ichi and Dziewczapolski, Gustavo and Nakamura, Tomohiro and Cao, Gang and Pratt, Alexander E and Kang, Yeon-Joo and Tu, Shichun and Molokanova, Elena and McKercher, Scott R and Hires, Samuel Andrew and Sason, Hagit and Stouffer, David G and Buczynski, Matthew W and Solomon, James P and Michael, Sarah and Powers, Evan T and Kelly, Jeffery W and Roberts, Amanda and Tong, Gary and Fang-Newmeyer, Traci and Parker, James and Holland, Emily A and Zhang, Dongxian and Nakanishi, Nobuki and Chen, H-S Vincent and Wolosker, Herman and Wang, Yuqiang and Parsons, Loren H and Ambasudhan, Rajesh and Masliah, Eliezer and Heinemann, Stephen F and Pi{\~n}a-Crespo, Juan C and Lipton, Stuart A} } @article {6715, title = {Balanced synaptic impact via distance-dependent synapse distribution and complementary expression of AMPARs and NMDARs in hippocampal dendrites.}, journal = {Neuron}, volume = {80}, year = {2013}, month = {2013 Dec 18}, pages = {1451-63}, abstract = {

Neuronal computation involves the integration of synaptic inputs that are often distributed over expansive dendritic trees, suggesting the need for compensatory mechanisms that enable spatially disparate synapses to influence neuronal output. In hippocampal CA1 pyramidal neurons, such mechanisms have indeed been reported, which normalize either the ability of distributed synapses to drive action potential initiation in the axon or their ability to drive dendritic spiking locally. Here we report that these mechanisms can coexist, through an elegant combination of distance-dependent regulation of synapse number and synaptic expression of AMPA and NMDA receptors. Together, these complementary gradients allow individual dendrites in both the apical and basal dendritic trees of hippocampal neurons to operate as facile computational subunits capable of supporting both global integration in the soma/axon and local integration in the dendrite.

}, issn = {1097-4199}, doi = {10.1016/j.neuron.2013.09.027}, author = {Menon, Vilas and Musial, Timothy F and Liu, Annie and Katz, Yael and Kath, William L and Spruston, Nelson and Nicholson, Daniel A} } @article {6282, title = {Cbl-associated protein regulates assembly and function of two tension-sensing structures in Drosophila.}, journal = {Development}, volume = {140}, year = {2013}, month = {2013 Feb 1}, pages = {627-38}, abstract = {

Cbl-associated protein (CAP) localizes to focal adhesions and associates with numerous cytoskeletal proteins; however, its physiological roles remain unknown. Here, we demonstrate that Drosophila CAP regulates the organization of two actin-rich structures in Drosophila: muscle attachment sites (MASs), which connect somatic muscles to the body wall; and scolopale cells, which form an integral component of the fly chordotonal organs and mediate mechanosensation. Drosophila CAP mutants exhibit aberrant junctional invaginations and perturbation of the cytoskeletal organization at the MAS. CAP depletion also results in collapse of scolopale cells within chordotonal organs, leading to deficits in larval vibration sensation and adult hearing. We investigate the roles of different CAP protein domains in its recruitment to, and function at, various muscle subcellular compartments. Depletion of the CAP-interacting protein Vinculin results in a marked reduction in CAP levels at MASs, and vinculin mutants partially phenocopy Drosophila CAP mutants. These results show that CAP regulates junctional membrane and cytoskeletal organization at the membrane-cytoskeletal interface of stretch-sensitive structures, and they implicate integrin signaling through a CAP/Vinculin protein complex in stretch-sensitive organ assembly and function.

}, keywords = {Actin Cytoskeleton, Amino Acid, Amino Acid Sequence, Animal Structures, Animals, Binding Sites, Cell Membrane, Cell-Matrix Junctions, Cellular, Cytoskeletal Proteins, Developmental, Drosophila, Electron, Electrophysiological Phenomena, Gene Expression Regulation, Genome, Hearing Disorders, Insect, Integrins, Larva, Mechanotransduction, Microscopy, Multiprotein Complexes, Muscles, Protein Interaction Mapping, Sequence Homology, Signal Transduction, src Homology Domains, Talin, Transmission, Vibration, Vinculin}, issn = {1477-9129}, doi = {10.1242/dev.085100}, author = {Bharadwaj, Rajnish and Roy, Madhuparna and Ohyama, Tomoko and Sivan-Loukianova, Elena and Delannoy, Michael and Lloyd, Thomas E and Zlatic, Marta and Eberl, Daniel F and Kolodkin, Alex L} } @article {6284, title = {Clonal development and organization of the adult Drosophila central brain.}, journal = {Current biology : CB}, volume = {23}, year = {2013}, month = {2013 Apr 22}, pages = {633-43}, abstract = {BACKGROUND: The insect brain can be divided into neuropils that are formed by neurites of both local and remote origin. The complexity of the interconnections obscures how these neuropils are established and interconnected through development. The Drosophila central brain develops from a fixed number of neuroblasts (NBs) that deposit neurons in regional clusters. RESULTS: By determining individual NB clones and pursuing their projections into specific neuropils, we unravel the regional development of the brain neural network. Exhaustive clonal analysis revealed 95 stereotyped neuronal lineages with characteristic cell-body locations and neurite trajectories. Most clones show complex projection patterns, but despite the complexity, neighboring clones often coinnervate the same local neuropil or neuropils and further target a restricted set of distant neuropils. CONCLUSIONS: These observations argue for regional clonal development of both neuropils and neuropil connectivity throughout the Drosophila central brain.}, issn = {1879-0445}, doi = {10.1016/j.cub.2013.02.057}, author = {Yu, Hung-Hsiang and Awasaki, Takeshi and Schroeder, Mark David and Long, Fuhui and Yang, Jacob S and He, Yisheng and Ding, Peng and Kao, Jui-Chun and Wu, Gloria Yueh-Yi and Peng, Hanchuan and Myers, Gene and Lee, Tzumin} } @article {46596, title = {Competing dopamine neurons drive oviposition choice for ethanol in Drosophila.}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {110}, year = {2013}, month = {2013 Dec 24}, pages = {21153-8}, abstract = {

The neural circuits that mediate behavioral choice evaluate and integrate information from the environment with internal demands and then initiate a behavioral response. Even circuits that support simple decisions remain poorly understood. In Drosophila melanogaster, oviposition on a substrate containing ethanol enhances fitness; however, little is known about the neural mechanisms mediating this important choice behavior. Here, we characterize the neural modulation of this simple choice and show that distinct subsets of dopaminergic neurons compete to either enhance or inhibit egg-laying preference for ethanol-containing food. Moreover, activity in α{\textquoteright}β{\textquoteright} neurons of the mushroom body and a subset of ellipsoid body ring neurons (R2) is required for this choice. We propose a model where competing dopaminergic systems modulate oviposition preference to adjust to changes in natural oviposition substrates.

}, keywords = {Analysis of Variance, Animals, Choice Behavior, Dopaminergic Neurons, Drosophila melanogaster, Ethanol, Female, Fermentation, Fruit, Mushroom Bodies, Oviposition}, issn = {1091-6490}, doi = {10.1073/pnas.1320208110}, author = {Azanchi, Reza and Kaun, Karla R and Heberlein, Ulrike} } @article {6712, title = {Correlative photoactivated localization and scanning electron microscopy.}, journal = {PLoS One}, volume = {8}, year = {2013}, month = {2013 Oct 25}, pages = {e77209}, abstract = {

The ability to localize proteins precisely within subcellular space is crucial to understanding the functioning of biological systems. Recently, we described a protocol that correlates a precise map of fluorescent fusion proteins localized using three-dimensional super-resolution optical microscopy with the fine ultrastructural context of three-dimensional electron micrographs. While it achieved the difficult simultaneous objectives of high photoactivated fluorophore preservation and ultrastructure preservation, it required a super-resolution optical and specialized electron microscope that is not available to many researchers. We present here a faster and more practical protocol with the advantage of a simpler two-dimensional optical (Photoactivated Localization Microscopy (PALM)) and scanning electron microscope (SEM) system that retains the often mutually exclusive attributes of fluorophore preservation and ultrastructure preservation. As before, cryosections were prepared using the Tokuyasu protocol, but the staining protocol was modified to be amenable for use in a standard SEM without the need for focused ion beam ablation. We show the versatility of this technique by labeling different cellular compartments and structures including mitochondrial nucleoids, peroxisomes, and the nuclear lamina. We also demonstrate simultaneous two-color PALM imaging with correlated electron micrographs. Lastly, this technique can be used with small-molecule dyes as demonstrated with actin labeling using phalloidin conjugated to a caged dye. By retaining the dense protein labeling expected for super-resolution microscopy combined with ultrastructural preservation, simplifying the tools required for correlative microscopy, and expanding the number of useful labels we expect this method to be accessible and valuable to a wide variety of researchers.

}, issn = {1932-6203}, doi = {10.1371/journal.pone.0077209}, author = {Kopek, Benjamin G and Shtengel, Gleb and Grimm, Jonathan B and Clayton, David A and Hess, Harald F} } @article {6277, title = {Diverse neuronal lineages make stereotyped contributions to the Drosophila locomotor control center, the central complex.}, journal = {The Journal of Comparative Neurology}, volume = {521}, year = {2013}, month = {2013 Aug 15}, pages = {2645-62, Spc1}, abstract = {

The Drosophila central brain develops from a fixed number of neuroblasts. Each neuroblast makes a clone of neurons that exhibit common trajectories. Here we identified 15 distinct clones that carry larval-born neurons innervating the Drosophila central complex (CX), which consists of four midline structures including the protocerebral bridge (PB), fan-shaped body (FB), ellipsoid body (EB), and noduli (NO). Clonal analysis revealed that the small-field CX neurons, which establish intricate projections across different CX substructures, exist in four isomorphic groups that respectively derive from four complex posterior asense-negative lineages. In terms of the region-characteristic large-field CX neurons, we found that two lineages make PB neurons, 10 lineages produce FB neurons, three lineages generate EB neurons, and two lineages yield NO neurons. The diverse FB developmental origins reflect the discrete input pathways for different FB subcompartments. Clonal analysis enlightens both development and anatomy of the insect locomotor control center.

}, issn = {1096-9861}, doi = {10.1002/cne.23339}, author = {Yang, Jacob S and Awasaki, Takeshi and Yu, Hung-Hsiang and He, Yisheng and Ding, Peng and Kao, Jui-Chun and Lee, Tzumin} } @article {6846, title = {Dual functions of TAF7L in adipocyte differentiation.}, journal = {eLife}, volume = {2}, year = {2013}, month = {2013 Jan 8}, pages = {e00170}, abstract = {

The diverse transcriptional mechanisms governing cellular differentiation and development of mammalian tissue remains poorly understood. Here we report that TAF7L, a paralogue of TFIID subunit TAF7, is enriched in adipocytes and white fat tissue (WAT) in mouse. Depletion of TAF7L reduced adipocyte-specific gene expression, compromised adipocyte differentiation, and WAT development as well. Ectopic expression of TAF7L in myoblasts reprograms these muscle precursors into adipocytes upon induction. Genome-wide mRNA-seq expression profiling and ChIP-seq binding studies confirmed that TAF7L is required for activating adipocyte-specific genes via a dual mechanism wherein it interacts with PPARγ at enhancers and TBP/Pol II at core promoters. In vitro binding studies confirmed that TAF7L forms complexes with both TBP and PPARγ. These findings suggest that TAF7L plays an integral role in adipocyte gene expression by targeting enhancers as a cofactor for PPARγ and promoters as a component of the core transcriptional machinery.DOI:http://dx.doi.org/10.7554/eLife.00170.001.

}, issn = {2050-084X}, doi = {10.7554/eLife.00170}, author = {Zhou, Haiying and Kaplan, Tommy and Li, Yan and Grubisic, Ivan and Zhang, Zhengjian and Wang, P Jeremy and Eisen, Michael B and Tjian, Robert} } @article {6270, title = {Essential role of the mushroom body in context-dependent CO2 avoidance in Drosophila.}, journal = {Current Biology}, volume = {23}, year = {2013}, month = {2013 Jul 8}, pages = {1228-34}, abstract = {

Internal state as well as environmental conditions influence choice behavior. The neural circuits underpinning state-dependent behavior remain largely unknown. Carbon dioxide (CO2) is an important olfactory cue for many insects, including mosquitoes, flies, moths, and honeybees [1]. Concentrations of CO2 higher than 0.02\% above atmospheric level trigger a strong innate avoidance in the fly Drosophila melanogaster [2, 3]. Here, we show that the mushroom body (MB), a brain center essential for olfactory associative memories [4-6] but thought to be dispensable for innate odor processing [7], is essential for CO2 avoidance behavior only in the context of starvation or in the context of a food-related odor. Consistent with this, CO2 stimulation elicits Ca(2+) influx into the MB intrinsic cells (Kenyon cells: KCs) in\ vivo. We identify an atypical projection neuron (bilateral ventral projection neuron, biVPN) that connects CO2 sensory input bilaterally to the MB calyx. Blocking synaptic output of the biVPN completely abolishes CO2 avoidance in food-deprived flies, but not in fed flies. These findings show that two alternative neural pathways control innate choice behavior, and they are dependent on the animal{\textquoteright}s internal state. In addition, they suggest that, during innate choice behavior, the MB serves as an integration site for internal state and olfactory input.

}, issn = {1879-0445}, doi = {10.1016/j.cub.2013.05.029}, author = {Br{\"a}cker, Lasse B and Siju, K P and Varela, N{\'e}lia and Aso, Yoshinori and Zhang, Mo and Hein, Irina and Vasconcelos, Maria Lu{\'\i}sa and Grunwald Kadow, Ilona C} } @article {6454, title = {Extremes of lineage plasticity in the Drosophila brain.}, journal = {Current Biology}, volume = {23}, year = {2013}, month = {2013 Sep 19}, pages = {1908-13}, abstract = {

An often-overlooked aspect of neural plasticity is the plasticity of neuronal composition, in which the numbers of neurons of particular classes are altered in response to environment and experience. The Drosophila brain features several well-characterized lineages in which a single neuroblast gives rise to multiple neuronal classes in a stereotyped sequence during development [1]. We find that in the\ intrinsic mushroom body neuron lineage, the numbers for each class are highly plastic, depending on the timing of temporal fate transitions and the rate of neuroblast proliferation. For example, mushroom body neuroblast cycling can continue under starvation conditions, uncoupled from\ temporal fate transitions that depend on extrinsic cues reflecting organismal growth and development. In contrast, the proliferation rates of antennal lobe lineages are closely associated with organismal development, and\ their temporal fate changes appear to be cell cycle-dependent, such that the same numbers and types of uniglomerular projection neurons innervate the antennal lobe following various perturbations. We propose that this surprising difference in plasticity for these brain lineages is adaptive, given their respective roles as parallel\ processors versus discrete carriers of olfactory information.

}, issn = {1879-0445}, doi = {10.1016/j.cub.2013.07.074}, author = {Lin, Suewei and Marin, Elizabeth C and Yang, Ching-Po and Kao, Chih-Fei and Apenteng, Bettye A and Huang, Yaling and O{\textquoteright}Connor, Michael B and Truman, James W and Lee, Tzumin} } @article {5536, title = {Fast and robust optical flow for time-lapse microscopy using super-voxels.}, journal = {Bioinformatics}, volume = {29}, year = {2013}, month = {2013 Feb}, pages = {373-80}, abstract = {

Optical flow is a key method used for quantitative motion estimation of biological structures in light microscopy. It has also been used as a key module in segmentation and tracking systems and is considered a mature technology in the field of computer vision. However, most of the research focused on 2D natural images, which are small in size and rich in edges and texture information. In contrast, 3D time-lapse recordings of biological specimens comprise up to several terabytes of image data and often exhibit complex object dynamics as well as blurring due to the point-spread-function of the microscope. Thus, new approaches to optical flow are required to improve performance for such data. We solve optical flow in large 3D time-lapse microscopy datasets by defining a Markov random field (MRF) over super-voxels in the foreground and applying motion smoothness constraints between super-voxels instead of voxel-wise. This model is tailored to the specific characteristics of light microscopy datasets: super-voxels help registration in textureless areas, the MRF over super-voxels efficiently propagates motion information between neighboring cells and the background subtraction and super-voxels reduce the dimensionality of the problem by an order of magnitude. We validate our approach on large 3D time-lapse datasets of Drosophila and zebrafish development by analyzing cell motion patterns. We show that our approach is, on average, 10x faster than commonly used optical flow implementations in the Insight Tool-Kit (ITK) and reduces the average flow end point error by 50\% in regions with complex dynamic processes, such as cell divisions.

}, issn = {1367-4811}, doi = {10.1093/bioinformatics/bts706}, author = {Amat, Fernando and Myers, Eugene W and Keller, Philipp J} } @article {5522, title = {Fast multicolor 3D imaging using aberration-corrected multifocus microscopy.}, journal = {Nature Methods}, volume = {10}, year = {2013}, month = {2013}, pages = {60-3}, abstract = {

Conventional acquisition of three-dimensional (3D) microscopy data requires sequential z scanning and is often too slow to capture biological events. We report an aberration-corrected multifocus microscopy method capable of producing an instant focal stack of nine 2D images. Appended to an epifluorescence microscope, the multifocus system enables high-resolution 3D imaging in multiple colors with single-molecule sensitivity, at speeds limited by the camera readout time of a single image.

}, issn = {1548-7105}, doi = {10.1038/nmeth.2277}, author = {Abrahamsson, Sara and Chen, Jiji and Hajj, Bassam and Stallinga, Sjoerd and Katsov, Alexander Y and Wisniewski, Jan and Mizuguchi, Gaku and Soule, Pierre and Mueller, Florian and Darzacq, Claire Dugast and Darzacq, Xavier and Wu, Carl and Bargmann, Cornelia I and Agard, David A and Dahan, Maxime and Gustafsson, Mats G L} } @article {6669, title = {Fast structural responses of gap junction membrane domains to AB5 toxins.}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {110}, year = {2013}, month = {2013 Oct 29}, pages = {E4125-33}, abstract = {

Gap junctions (GJs) represent connexin-rich membrane domains that connect interiors of adjoining cells in mammalian tissues. How fast GJs can respond to bacterial pathogens has not been known previously. Using Bessel beam plane illumination and confocal spinning disk microscopy, we found fast (\ 500 ms) formation of connexin-depleted regions (CDRs) inside GJ plaques between cells exposed to AB5 toxins. CDR formation appears as a fast redistribution of connexin channels within GJ plaques with minor changes in outline or geometry. CDR formation does not depend on membrane trafficking or submembrane cytoskeleton and has no effect on GJ conductance. However, CDR responses depend on membrane lipids, can be modified by cholesterol-clustering agents and extracellular K(+) ion concentration, and influence cAMP signaling. The CDR response of GJ plaques to bacterial toxins is a phenomenon observed for all tested connexin isoforms. Through signaling, the CDR response may enable cells to sense exposure to AB5 toxins. CDR formation may reflect lipid-phase separation events in the biological membrane of the GJ plaque, leading to increased connexin packing and lipid reorganization. Our data demonstrate very fast dynamics (in the millisecond-to-second range) within GJ plaques, which previously were considered to be relatively stable, long-lived structures.

}, issn = {1091-6490}, doi = {10.1073/pnas.1315850110}, author = {Majoul, Irina V and Gao, Liang and Betzig, Eric and Onichtchouk, Daria and Butkevich, Eugenia and Kozlov, Yuri and Bukauskas, Feliksas and Bennett, Michael V L and Lippincott-Schwartz, Jennifer and Duden, Rainer} } @article {47637, title = {Fast structural responses of gap junction membrane domains to AB5 toxins.}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {110}, year = {2013}, month = {2013 Oct 29}, pages = {E4125-33}, abstract = {

Gap junctions (GJs) represent connexin-rich membrane domains that connect interiors of adjoining cells in mammalian tissues. How fast GJs can respond to bacterial pathogens has not been known previously. Using Bessel beam plane illumination and confocal spinning disk microscopy, we found fast (~500 ms) formation of connexin-depleted regions (CDRs) inside GJ plaques between cells exposed to AB5 toxins. CDR formation appears as a fast redistribution of connexin channels within GJ plaques with minor changes in outline or geometry. CDR formation does not depend on membrane trafficking or submembrane cytoskeleton and has no effect on GJ conductance. However, CDR responses depend on membrane lipids, can be modified by cholesterol-clustering agents and extracellular K(+) ion concentration, and influence cAMP signaling. The CDR response of GJ plaques to bacterial toxins is a phenomenon observed for all tested connexin isoforms. Through signaling, the CDR response may enable cells to sense exposure to AB5 toxins. CDR formation may reflect lipid-phase separation events in the biological membrane of the GJ plaque, leading to increased connexin packing and lipid reorganization. Our data demonstrate very fast dynamics (in the millisecond-to-second range) within GJ plaques, which previously were considered to be relatively stable, long-lived structures.

}, keywords = {Analysis of Variance, Animals, Bacterial Toxins, Bicyclo Compounds, Heterocyclic, Cercopithecus aethiops, Connexins, Cyclic AMP, DNA Primers, Filipin, Fluorescence, Gap Junctions, Image Processing, Computer-Assisted, Membrane Lipids, Microscopy, Confocal, Patch-Clamp Techniques, Potassium, Thiazolidines, Vero Cells}, issn = {1091-6490}, doi = {10.1073/pnas.1315850110}, author = {Majoul, Irina V and Gao, Liang and Betzig, Eric and Onichtchouk, Daria and Butkevich, Eugenia and Kozlov, Yuri and Bukauskas, Feliksas and Bennett, Michael V L and Lippincott-Schwartz, Jennifer and Duden, Rainer} } @article {6133, title = {Functional labeling of neurons and their projections using the synthetic activity-dependent promoter E-SARE.}, journal = {Nature Methods}, volume = {10}, year = {2013}, month = {2013 Jul 14}, pages = {889-95}, abstract = {

Identifying the neuronal ensembles that respond to specific stimuli and mapping their projection patterns in living animals are fundamental challenges in neuroscience. To this end, we engineered a synthetic promoter, the enhanced synaptic activity-responsive element (E-SARE), that drives neuronal activity-dependent gene expression more potently than other existing immediate-early gene promoters. Expression of a drug-inducible Cre recombinase downstream of E-SARE enabled imaging of neuronal populations that respond to monocular visual stimulation and tracking of their long-distance thalamocortical projections in living mice. Targeted cell-attached recordings and calcium imaging of neurons in sensory cortices revealed that E-SARE reporter expression correlates with sensory-evoked neuronal activity at the single-cell level and is highly specific to the type of stimuli presented to the animals. This activity-dependent promoter can expand the repertoire of genetic approaches for high-resolution anatomical and functional analysis of neural circuits.

}, issn = {1548-7105}, doi = {10.1038/nmeth.2559}, author = {Kawashima, Takashi and Kitamura, Kazuo and Suzuki, Kanzo and Nonaka, Mio and Kamijo, Satoshi and Takemoto-Kimura, Sayaka and Kano, Masanobu and Okuno, Hiroyuki and Ohki, Kenichi and Bito, Haruhiko} } @article {6923, title = {Genetic and neural mechanisms that inhibit Drosophila from mating with other species.}, journal = {Cell}, volume = {154}, year = {2013}, month = {2013 Jul 3}, pages = {89-102}, abstract = {

Genetically hard-wired neural mechanisms must enforce behavioral reproductive isolation because interspecies courtship is rare even in sexually na{\"{\i}ve animals of most species. We find that the chemoreceptor Gr32a inhibits male D. melanogaster from courting diverse fruit fly species. Gr32a recognizes nonvolatile aversive cues present on these reproductively dead-end targets, and activity of Gr32a neurons is necessary and sufficient to inhibit interspecies courtship. Male-specific Fruitless (Fru(M)), a master regulator of courtship, also inhibits interspecies courtship. Gr32a and Fru(M) are not coexpressed, but Fru(M) neurons contact Gr32a neurons, suggesting that these genes influence a shared neural circuit that inhibits interspecies courtship. Gr32a and Fru(M) also suppress within-species intermale courtship, but we show that distinct mechanisms preclude sexual displays toward conspecific males and other species. Although this chemosensory pathway does not inhibit interspecies mating in D. melanogaster females, similar mechanisms appear to inhibit this behavior in many other male drosophilids.

}, keywords = {Animal, Animals, Cell Surface, Courtship, Drosophila, Drosophila melanogaster, Drosophila Proteins, Female, Genetic Speciation, Male, Mating Preference, Nerve Tissue Proteins, Neurons, Receptors, Transcription Factors}, issn = {1097-4172}, doi = {10.1016/j.cell.2013.06.008}, author = {Fan, Pu and Manoli, Devanand S and Ahmed, Osama M and Chen, Yi and Agarwal, Neha and Kwong, Sara and Cai, Allen G and Neitz, Jeffrey and Renslo, Adam and Baker, Bruce S and Shah, Nirao M} } @article {6126, title = {Genetically encoded calcium indicators and astrocyte calcium microdomains.}, journal = {The Neuroscientist : A Review Journal Bringing Neurobiology, Neurology and Psychiatry}, volume = {19}, year = {2013}, month = {2013 Jun}, pages = {274-91}, abstract = {

The discovery of intracellular Ca(2+) signals within astrocytes has changed our view of how these ubiquitous cells contribute to brain function. Classically thought merely to serve supportive functions, astrocytes are increasingly thought to respond to, and regulate, neurons. The use of organic Ca(2+) indicator dyes such as Fluo-4 and Fura-2 has proved instrumental in the study of astrocyte physiology. However, progress has recently been accelerated by the use of cytosolic and membrane targeted genetically encoded calcium indicators (GECIs). Herein, we review these recent findings, discuss why studying astrocyte Ca(2+) signals is important to understand brain function, and summarize work that led to the discovery of TRPA1 channel-mediated near-membrane Ca(2+) signals in astrocytes and their indirect neuromodulatory roles at inhibitory synapses in the CA1 stratum radiatum region of the hippocampus. We suggest that the use of membrane-targeted and cytosolic GECIs holds great promise to explore the diversity of Ca(2+) signals within single astrocytes and also to study diversity of function for astrocytes in different parts of the brain.

}, issn = {1089-4098}, doi = {10.1177/1073858412468794}, author = {Tong, Xiaoping and Shigetomi, Eiji and Looger, Loren L and Khakh, Baljit S.} } @article {6541, title = {Genetically encoded calcium indicators for multi-color neural activity imaging and combination with optogenetics.}, journal = {Frontiers in Molecular Neuroscience}, volume = {6}, year = {2013}, month = {2013 Mar 4}, pages = {2}, abstract = {

Genetically encoded calcium indicators (GECIs) are powerful tools for systems neuroscience. Here we describe red, single-wavelength GECIs, "RCaMPs," engineered from circular permutation of the thermostable red fluorescent protein mRuby. High-resolution crystal structures of mRuby, the red sensor RCaMP, and the recently published red GECI R-GECO1 give insight into the chromophore environments of the Ca(2+)-bound state of the sensors and the engineered protein domain interfaces of the different indicators. We characterized the biophysical properties and performance of RCaMP sensors in vitro and in vivo in Caenorhabditis elegans, Drosophila larvae, and larval zebrafish. Further, we demonstrate 2-color calcium imaging both within the same cell (registering mitochondrial and somatic [Ca(2+)]) and between two populations of cells: neurons and astrocytes. Finally, we perform integrated optogenetics experiments, wherein neural activation via channelrhodopsin-2 (ChR2) or a red-shifted variant, and activity imaging via RCaMP or GCaMP, are conducted simultaneously, with the ChR2/RCaMP pair providing independently addressable spectral channels. Using this paradigm, we measure calcium responses of naturalistic and ChR2-evoked muscle contractions in vivo in crawling C. elegans. We systematically compare the RCaMP sensors to R-GECO1, in terms of action potential-evoked fluorescence increases in neurons, photobleaching, and photoswitching. R-GECO1 displays higher Ca(2+) affinity and larger dynamic range than RCaMP, but exhibits significant photoactivation with blue and green light, suggesting that integrated channelrhodopsin-based optogenetics using R-GECO1 may be subject to artifact. Finally, we create and test blue, cyan, and yellow variants engineered from GCaMP by rational design. This engineered set of chromatic variants facilitates new experiments in functional imaging and optogenetics.

}, issn = {1662-5099}, doi = {10.3389/fnmol.2013.00002}, author = {Akerboom, Jasper and Carreras Calder{\'o}n, Nicole and Tian, Lin and Wabnig, Sebastian and Prigge, Matthias and Tol{\"o}, Johan and Gordus, Andrew and Orger, Michael B. and Severi, Kristen E and Macklin, John J and Patel, Ronak and Pulver, Stefan R and Wardill, Trevor J. and Fischer, Elisabeth and Sch{\"u}ler, Christina and Chen, Tsai-Wen and Sarkisyan, Karen S and Marvin, Jonathan S and Bargmann, Cornelia I and Kim, Douglas S. and K{\"u}gler, Sebastian and Lagnado, Leon and Hegemann, Peter and Gottschalk, Alexander and Schreiter, Eric R and Looger, Loren L} } @article {5803, title = {HCV IRES manipulates the ribosome to promote the switch from translation initiation to elongation.}, journal = {Nature Structural \& Molecular Biology}, volume = {20}, year = {2013}, month = {2013 Feb}, pages = {150-8}, abstract = {

The internal ribosome entry site (IRES) of the hepatitis C virus (HCV) drives noncanonical initiation of protein synthesis necessary for viral replication. Functional studies of the HCV IRES have focused on 80S ribosome formation but have not explored its role after the 80S ribosome is poised at the start codon. Here, we report that mutations of an IRES domain that docks in the 40S subunit{\textquoteright}s decoding groove cause only a local perturbation in IRES structure and result in conformational changes in the IRES-rabbit 40S subunit complex. Functionally, the mutations decrease IRES activity by inhibiting the first ribosomal translocation event, and modeling results suggest that this effect occurs through an interaction with a single ribosomal protein. The ability of the HCV IRES to manipulate the ribosome provides insight into how the ribosome{\textquoteright}s structure and function can be altered by bound RNAs, including those derived from cellular invaders.

}, issn = {1545-9985}, doi = {10.1038/nsmb.2465}, author = {Filbin, Megan E and Vollmar, Breanna S and Shi, Dan and Gonen, Tamir and Kieft, Jeffrey S} } @article {6262, title = {High-throughput analysis of stimulus-evoked behaviors in Drosophila larva reveals multiple modality-specific escape strategies.}, journal = {PLoS One}, volume = {8}, year = {2013}, month = {2013 Aug 20}, pages = {e71706}, abstract = {

All organisms react to noxious and mechanical stimuli but we still lack a complete understanding of cellular and molecular mechanisms by which somatosensory information is transformed into appropriate motor outputs. The small number of neurons and excellent genetic tools make Drosophila larva an especially tractable model system in which to address this problem. We developed high throughput assays with which we can simultaneously expose more than 1,000 larvae per man-hour to precisely timed noxious heat, vibration, air current, or optogenetic stimuli. Using this hardware in combination with custom software we characterized larval reactions to somatosensory stimuli in far greater detail than possible previously. Each stimulus evoked a distinctive escape strategy that consisted of multiple actions. The escape strategy was context-dependent. Using our system we confirmed that the nociceptive class IV multidendritic neurons were involved in the reactions to noxious heat. Chordotonal (ch) neurons were necessary for normal modulation of head casting, crawling and hunching, in response to mechanical stimuli. Consistent with this we observed increases in calcium transients in response to vibration in ch neurons. Optogenetic activation of ch neurons was sufficient to evoke head casting and crawling. These studies significantly increase our understanding of the functional roles of larval ch neurons. More generally, our system and the detailed description of wild type reactions to somatosensory stimuli provide a basis for systematic identification of neurons and genes underlying these behaviors.

}, issn = {1932-6203}, doi = {10.1371/journal.pone.0071706}, author = {Ohyama, Tomoko and Jovanic, Tihana and Denisov, Gennady and Dang, Tam C and Hoffmann, Dominik and Kerr, Rex A and Zlatic, Marta} } @article {68819, title = {Hindbrain V2a Neurons in the Excitation of Spinal Locomotor Circuits during Zebrafish Swimming}, journal = {Current Biology}, volume = {23}, year = {2013}, month = {05/2013}, pages = {843-849}, abstract = {

During locomotion in vertebrates, reticulospinal neurons in the hindbrain play critical roles in providing descending excitation to the spinal cord locomotor systems. However, despite the fact that many genes that are used to classify the neuronal identities of neurons in the hindbrain have been identified, the molecular identity of the reticulospinal neurons that are critically involved in locomotor drive is not well understood. Chx10-expressing neurons (V2a neurons) are ipsilaterally projecting glutamatergic neurons in the spinal cord and the hindbrain. Many of the V2a neurons in the hindbrain are known to project to the spinal cord in zebrafish, making hindbrain V2a neurons a prime candidate in descending locomotor drive. Results We investigated the roles of hindbrain V2a neurons using optogenetic and electrophysiological approaches. The forced activation of hindbrain V2a neurons using channelrhodopsin efficiently evoked swimming, whereas the forced inactivation of them using Archearhodopsin3 or Halorhodpsin reliably stopped ongoing swimming. Electrophysiological recordings of two populations of hindbrain reticulospinal V2a neurons showed that they were active during swimming. One population of neurons, small V2a neurons in the caudal hindbrain, fired with low rhythmicity, whereas the other population of neurons, large reticulospinal V2a neurons, called MiV1 neurons, fired more rhythmically. Conclusions These results indicated that hindbrain reticulospinal V2a neurons play critical roles in providing excitation to the spinal locomotor circuits during swimming by providing both tonic and phasic inputs to the circuits.

}, issn = {0960-9822}, doi = {https://doi.org/10.1016/j.cub.2013.03.066}, url = {https://www.sciencedirect.com/science/article/pii/S0960982213003680}, author = {Yukiko Kimura and Chie Satou and Shunji Fujioka and Wataru Shoji and Keiko Umeda and Toru Ishizuka and Hiromu Yawo and Shin-ichi Higashijima} } @article {5840, title = {Identification of nonvisual photomotor response cells in the vertebrate hindbrain.}, journal = {The Journal of Neuroscience}, volume = {33}, year = {2013}, month = {2013 Feb 27}, pages = {3834-43}, abstract = {

Nonvisual photosensation enables animals to sense light without sight. However, the cellular and molecular mechanisms of nonvisual photobehaviors are poorly understood, especially in vertebrate animals. Here, we describe the photomotor response (PMR), a robust and reproducible series of motor behaviors in zebrafish that is elicited by visual wavelengths of light but does not require the eyes, pineal gland, or other canonical deep-brain photoreceptive organs. Unlike the relatively slow effects of canonical nonvisual pathways, motor circuits are strongly and quickly (seconds) recruited during the PMR behavior. We find that the hindbrain is both necessary and sufficient to drive these behaviors. Using in vivo calcium imaging, we identify a discrete set of neurons within the hindbrain whose responses to light mirror the PMR behavior. Pharmacological inhibition of the visual cycle blocks PMR behaviors, suggesting that opsin-based photoreceptors control this behavior. These data represent the first known light-sensing circuit in the vertebrate hindbrain.

}, issn = {1529-2401}, doi = {10.1523/JNEUROSCI.3689-12.2013}, author = {Kokel, David and Dunn, Timothy W. and Ahrens, Misha B and Alshut, R{\"u}diger and Cheung, Chung Yan J. and Saint-Amant, Louis and Bruni, Giancarlo and Mateus, Rita and van Ham, Tjakko J. and Shiraki, Tomoya and Fukada, Yoshitaka and Kojima, Daisuke and Yeh, Jing-Ruey J. and Mikut, Ralf and von Lintig, Johannes and Engert, Florian and Peters, Randall T.} } @article {5982, title = {Imaging morphogenesis: technological advances and biological insights.}, journal = {Science}, volume = {340}, year = {2013}, month = {2013 Jun 7}, pages = {1234168}, abstract = {

Morphogenesis, the development of the shape of an organism, is a dynamic process on a multitude of scales, from fast subcellular rearrangements and cell movements to slow structural changes at the whole-organism level. Live-imaging approaches based on light microscopy reveal the intricate dynamics of this process and are thus indispensable for investigating the underlying mechanisms. This Review discusses emerging imaging techniques that can record morphogenesis at temporal scales from seconds to days and at spatial scales from hundreds of nanometers to several millimeters. To unlock their full potential, these methods need to be matched with new computational approaches and physical models that help convert highly complex image data sets into biological insights.

Science Profile: A Research Career in Focus

}, issn = {1095-9203}, doi = {10.1126/science.1234168}, author = {Keller, Philipp J} } @article {6670, title = {Imaging neuronal populations in behaving rodents: paradigms for studying neural circuits underlying behavior in the mammalian cortex.}, journal = {The Journal of Neuroscience : the Official Journal of the Society for Neuroscience}, volume = {33}, year = {2013}, month = {2013 Nov 6}, pages = {17631-40}, abstract = {

Understanding the neural correlates of behavior in the mammalian cortex requires measurements of activity in awake, behaving animals. Rodents have emerged as a powerful model for dissecting the cortical circuits underlying behavior attributable to the convergence of several methods. Genetically encoded calcium indicators combined with viral-mediated or transgenic tools enable chronic monitoring of calcium signals in neuronal populations and subcellular structures of identified cell types. Stable one- and two-photon imaging of neuronal activity in awake, behaving animals is now possible using new behavioral paradigms in head-fixed animals, or using novel miniature head-mounted microscopes in freely moving animals. This mini-symposium will highlight recent applications of these methods for studying sensorimotor integration, decision making, learning, and memory in cortical and subcortical brain areas. We will outline future prospects and challenges for identifying the neural underpinnings of task-dependent behavior using cellular imaging in rodents.

}, issn = {1529-2401}, doi = {10.1523/JNEUROSCI.3255-13.2013}, author = {Chen, Jerry L and Andermann, Mark L and Keck, Tara and Xu, Ning-long and Ziv, Yaniv} } @article {5884, title = {In vivo imaging of zebrafish embryogenesis.}, journal = {Methods}, volume = {62}, year = {2013}, month = {2013 Aug 15}, pages = {268-78}, abstract = {

The zebrafish Danio rerio has emerged as a powerful vertebrate model system that lends itself particularly well to quantitative investigations with live imaging approaches, owing to its exceptionally high optical clarity in embryonic and larval stages. Recent advances in light microscopy technology enable comprehensive analyses of cellular dynamics during zebrafish embryonic development, systematic mapping of gene expression dynamics, quantitative reconstruction of mutant phenotypes and the system-level biophysical study of morphogenesis. Despite these technical breakthroughs, it remains challenging to design and implement experiments for in vivo long-term imaging at high spatio-temporal resolution. This article discusses the fundamental challenges in zebrafish long-term live imaging, provides experimental protocols and highlights key properties and capabilities of advanced fluorescence microscopes. The article focuses in particular on experimental assays based on light sheet-based fluorescence microscopy, an emerging imaging technology that achieves exceptionally high imaging speeds and excellent signal-to-noise ratios, while minimizing light-induced damage to the specimen. This unique combination of capabilities makes light sheet microscopy an indispensable tool for the in vivo long-term imaging of large developing organisms.

}, issn = {1095-9130}, doi = {10.1016/j.ymeth.2013.03.015}, author = {Keller, Philipp J} } @article {47899, title = {The influence of synaptic weight distribution on neuronal population dynamics.}, journal = {PLoS computational biology}, volume = {9}, year = {2013}, month = {2013 Oct}, pages = {e1003248}, abstract = {

The manner in which different distributions of synaptic weights onto cortical neurons shape their spiking activity remains open. To characterize a homogeneous neuronal population, we use the master equation for generalized leaky integrate-and-fire neurons with shot-noise synapses. We develop fast semi-analytic numerical methods to solve this equation for either current or conductance synapses, with and without synaptic depression. We show that its solutions match simulations of equivalent neuronal networks better than those of the Fokker-Planck equation and we compute bounds on the network response to non-instantaneous synapses. We apply these methods to study different synaptic weight distributions in feed-forward networks. We characterize the synaptic amplitude distributions using a set of measures, called tail weight numbers, designed to quantify the preponderance of very strong synapses. Even if synaptic amplitude distributions are equated for both the total current and average synaptic weight, distributions with sparse but strong synapses produce higher responses for small inputs, leading to a larger operating range. Furthermore, despite their small number, such synapses enable the network to respond faster and with more stability in the face of external fluctuations.

}, keywords = {Action Potentials, Computer Simulation, Models, Neurological, Neurons, Synapses}, issn = {1553-7358}, doi = {10.1371/journal.pcbi.1003248}, author = {Iyer, Ramakrishnan and Menon, Vilas and Buice, Michael and Koch, Christof and Mihalas, Stefan} } @article {6528, title = {Intensity discrimination deficits cause habituation changes in middle-aged Caenorhabditis elegans.}, journal = {Neurobiology of Aging}, volume = {34}, year = {2013}, month = {2013 Feb}, pages = {621-31}, abstract = {

The ability to learn and remember is critical for all animals to survive in the ever-changing environment. As we age, many of our biological faculties decay and of these, decline in learning and memory can be the most distressing. To carefully define age-dependent changes in learning during reproductive age in the nematode Caenorhabditis elegans, we performed a parametric behavioral study of habituation to nonlocalized mechanical stimuli (petri plate taps) over a range of intensities in middle-aged worms. We found that as worms age (from the onset of reproduction to the end of egg laying), response probability habituation increases (at both 10- and 60-second interstimulus intervals) and that these age-related changes were associated with a decrease in the discrimination between stimuli of different intensities. We also used optogenetics to investigate where these age-dependent changes occur. Our data suggest that the changes occur upstream of mechanosensory neuron depolarization. These data support the idea that declines in stimulus intensity discrimination abilities during aging may be one variable underlying age-related cognitive deficits.

}, keywords = {Age Factors, Aging, Animal, Animals, Behavior, Caenorhabditis elegans, Discrimination (Psychology), Habituation, Memory, Physical Stimulation, Psychophysiologic, Short-Term}, issn = {1558-1497}, doi = {10.1016/j.neurobiolaging.2012.03.016}, author = {Timbers, Tiffany A and Giles, Andrew C and Ardiel, Evan L and Kerr, Rex A and Rankin, Catharine H} } @article {5763, title = {JAABA: interactive machine learning for automatic annotation of animal behavior.}, journal = {Nature Methods}, volume = {10}, year = {2013}, month = {2012 Dec}, pages = {64-7}, abstract = {

We present a machine learning{\textendash}based system for automatically computing interpretable, quantitative measures of animal behavior. Through our interactive system, users encode their intuition about behavior by annotating a small set of video frames. These manual labels are converted into classifiers that can automatically annotate behaviors in screen-scale data sets. Our general-purpose system can create a variety of accurate individual and social behavior classifiers for different organisms, including mice and adult and larval Drosophila.

}, issn = {1548-7091}, author = {Kabra, Mayank and Robie, Alice A and Rivera-Alba, Marta and Branson, Steven and Branson, Kristin} } @article {65022, title = {Large scale structural rearrangement of a serine hydrolase from Francisella tularensis facilitates catalysis.}, journal = {J Biol Chem}, volume = {288}, year = {2013}, month = {2013 Apr 12}, pages = {10522-35}, abstract = {

Tularemia is a deadly, febrile disease caused by infection by the gram-negative bacterium, Francisella tularensis. Members of the ubiquitous serine hydrolase protein family are among current targets to treat diverse bacterial infections. Herein we present a structural and functional study of a novel bacterial carboxylesterase (FTT258) from F. tularensis, a homologue of human acyl protein thioesterase (hAPT1). The structure of FTT258 has been determined in multiple forms, and unexpectedly large conformational changes of a peripheral flexible loop occur in the presence of a mechanistic cyclobutanone ligand. The concomitant changes in this hydrophobic loop and the newly exposed hydrophobic substrate binding pocket suggest that the observed structural changes are essential to the biological function and catalytic activity of FTT258. Using diverse substrate libraries, site-directed mutagenesis, and liposome binding assays, we determined the importance of these structural changes to the catalytic activity and membrane binding activity of FTT258. Residues within the newly exposed hydrophobic binding pocket and within the peripheral flexible loop proved essential to the hydrolytic activity of FTT258, indicating that structural rearrangement is required for catalytic activity. Both FTT258 and hAPT1 also showed significant association with liposomes designed to mimic bacterial or human membranes, respectively, even though similar structural rearrangements for hAPT1 have not been reported. The necessity for acyl protein thioesterases to have maximal catalytic activity near the membrane surface suggests that these conformational changes in the protein may dually regulate catalytic activity and membrane association in bacterial and human homologues.

}, keywords = {Bacterial Proteins, Cell Membrane, Francisella tularensis, Humans, Mutagenesis, Site-Directed, Serine Endopeptidases, Structural Homology, Protein, Structure-Activity Relationship, Thiolester Hydrolases}, issn = {1083-351X}, doi = {10.1074/jbc.M112.446625}, author = {Filippova, Ekaterina V and Weston, Leigh A and Kuhn, Misty L and Geissler, Brett and Gehring, Alexandra M and Armoush, Nicola and Adkins, Chinessa T and Minasov, George and Dubrovska, Ievgeniia and Shuvalova, Ludmilla and Winsor, James R and Lavis, Luke D and Satchell, Karla J F and Becker, Daniel P and Anderson, Wayne F and Johnson, R Jeremy} } @article {5346, title = {Light sheet microscopy in cell biology.}, journal = {Methods in Molecular Biology}, volume = {931}, year = {2013}, month = {2013}, pages = {123-37}, abstract = {

Light sheet-based fluorescence microscopy (LSFM) is emerging as a powerful imaging technique for the life sciences. LSFM provides an exceptionally high imaging speed, high signal-to-noise ratio, low level of photo-bleaching, and good optical penetration depth. This unique combination of capabilities makes light sheet-based microscopes highly suitable for live imaging applications. Here, we provide an overview of light sheet-based microscopy assays for in vitro and in vivo imaging of biological samples, including cell extracts, soft gels, and large multicellular organisms. We furthermore describe computational tools for basic image processing and data inspection.

}, issn = {1940-6029}, doi = {10.1007/978-1-62703-056-4_7}, author = {Tomer, Raju and Khairy, Khaled and Keller, Philipp J} } @article {6918, title = {Live imaging of nervous system development and function using light-sheet microscopy.}, journal = {Molecular Reproduction and Development}, volume = {82}, year = {2013}, month = {2015 Jul}, pages = {605-18}, abstract = {

In vivo imaging applications typically require carefully balancing conflicting parameters. Often it is necessary to achieve high imaging speed, low photo-bleaching, and photo-toxicity, good three-dimensional resolution, high signal-to-noise ratio, and excellent physical coverage at the same time. Light-sheet microscopy provides good performance in all of these categories, and is thus emerging as a particularly powerful live imaging method for the life sciences. We see an outstanding potential for applying light-sheet microscopy to the study of development and function of the early nervous system in vertebrates and higher invertebrates. Here, we review state-of-the-art approaches to live imaging of early development, and show how the unique capabilities of light-sheet microscopy can further advance our understanding of the development and function of the nervous system. We discuss key considerations in the design of light-sheet microscopy experiments, including sample preparation and fluorescent marker strategies, and provide an outlook for future directions in the field.

}, issn = {1098-2795}, doi = {10.1002/mrd.22258}, author = {Lemon, William C and Keller, Philipp J} } @article {6073, title = {Live imaging of whole mouse embryos during gastrulation: migration analyses of epiblast and mesodermal cells.}, journal = {PLoS One}, volume = {8}, year = {2013}, month = {2013 Jul 8}, pages = {e64506}, abstract = {

During gastrulation in the mouse embryo, dynamic cell movements including epiblast invagination and mesodermal layer expansion lead to the establishment of the three-layered body plan. The precise details of these movements, however, are sometimes elusive, because of the limitations in live imaging. To overcome this problem, we developed techniques to enable observation of living mouse embryos with digital scanned light sheet microscope (DSLM). The achieved deep and high time-resolution images of GFP-expressing nuclei and following 3D tracking analysis revealed the following findings: (i) Interkinetic nuclear migration (INM) occurs in the epiblast at embryonic day (E)6 and 6.5. (ii) INM-like migration occurs in the E5.5 embryo, when the epiblast is a monolayer and not yet pseudostratified. (iii) Primary driving force for INM at E6.5 is not pressure from neighboring nuclei. (iv) Mesodermal cells migrate not as a sheet but as individual cells without coordination.

}, issn = {1932-6203}, doi = {10.1371/journal.pone.0064506}, author = {Ichikawa, Takehiko and Nakazato, Kenichi and Keller, Philipp J and Kajiura-Kobayashi, Hiroko and Stelzer, Ernst H K and Mochizuki, Atsushi and Nonaka, Shigenori} } @article {6263, title = {Machine learning of hierarchical clustering to segment 2D and 3D images.}, journal = {PLoS One}, volume = {8}, year = {2013}, month = {2013}, pages = {e71715}, abstract = {

We aim to improve segmentation through the use of machine learning tools during region agglomeration. We propose an active learning approach for performing hierarchical agglomerative segmentation from superpixels. Our method combines multiple features at all scales of the agglomerative process, works for data with an arbitrary number of dimensions, and scales to very large datasets. We advocate the use of variation of information to measure segmentation accuracy, particularly in 3D electron microscopy (EM) images of neural tissue, and using this metric demonstrate an improvement over competing algorithms in EM and natural images.

}, issn = {1932-6203}, doi = {10.1371/journal.pone.0071715}, author = {Nunez-Iglesias, Juan and Kennedy, Ryan and Toufiq Parag and Shi, Jianbo and Chklovskii, Dmitri B} } @article {6925, title = {Male-specific fruitless isoforms have different regulatory roles conferred by distinct zinc finger DNA binding domains.}, journal = {BMC Genomics}, volume = {14}, year = {2013}, month = {2013 Sep 27}, pages = {659}, abstract = {

BACKGROUND: Drosophila melanogaster adult males perform an elaborate courtship ritual to entice females to mate. fruitless (fru), a gene that is one of the key regulators of male courtship behavior, encodes multiple male-specific isoforms (Fru(M)). These isoforms vary in their carboxy-terminal zinc finger domains, which are predicted to facilitate DNA binding. RESULTS: By over-expressing individual Fru(M) isoforms in fru-expressing neurons in either males or females and assaying the global transcriptional response by RNA-sequencing, we show that three Fru(M) isoforms have different regulatory activities that depend on the sex of the fly. We identified several sets of genes regulated downstream of Fru(M) isoforms, including many annotated with neuronal functions. By determining the binding sites of individual Fru(M) isoforms using SELEX we demonstrate that the distinct zinc finger domain of each Fru(M) isoforms confers different DNA binding specificities. A genome-wide search for these binding site sequences finds that the gene sets identified as induced by over-expression of Fru(M) isoforms in males are enriched for genes that contain the binding sites. An analysis of the chromosomal distribution of genes downstream of Fru(M) shows that those that are induced and repressed in males are highly enriched and depleted on the X chromosome, respectively. CONCLUSIONS: This study elucidates the different regulatory and DNA binding activities of three Fru(M) isoforms on a genome-wide scale and identifies genes regulated by these isoforms. These results add to our understanding of sex chromosome biology and further support the hypothesis that in some cell-types genes with male-biased expression are enriched on the X chromosome.

}, issn = {1471-2164}, doi = {10.1186/1471-2164-14-659}, author = {Dalton, Justin E and Fear, Justin M and Knott, Simon and Baker, Bruce S and McIntyre, Lauren M and Arbeitman, Michelle N} } @article {6714, title = {Mapping mammalian synaptic connectivity.}, journal = {Cellular and Molecular Life Sciences: CMLS}, volume = {70}, year = {2013}, month = {2013 Dec}, pages = {4747-57}, abstract = {

Mapping mammalian synaptic connectivity has long been an important goal of neuroscientists since it is considered crucial for explaining human perception and behavior. Yet, despite enormous efforts, the overwhelming complexity of the neural circuitry and the lack of appropriate techniques to unravel it have limited the success of efforts to map connectivity. However, recent technological advances designed to overcome the limitations of conventional methods for connectivity mapping may bring about a turning point. Here, we address the promises and pitfalls of these new mapping technologies.

}, keywords = {Animals, Brain, Computational Biology, Connectome, Humans, Mammals, Nervous System Diseases, Synapses}, issn = {1420-9071}, doi = {10.1007/s00018-013-1417-y}, author = {Yook, Chaehyun and Druckmann, S and Kim, Jinhyun} } @article {6383, title = {A molt timer is involved in the metamorphic molt in Manduca sexta larvae.}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {110}, year = {2013}, month = {2013 Jul 30}, pages = {12518-25}, abstract = {

Manduca sexta larvae are a model for growth control in insects, particularly for the demonstration of critical weight, a threshold weight that the larva must surpass before it can enter metamorphosis on a normal schedule, and the inhibitory action of juvenile hormone on this checkpoint. We examined the effects of nutrition on allatectomized (CAX) larvae that lack juvenile hormone to impose the critical weight checkpoint. Normal larvae respond to prolonged starvation at the start of the last larval stage, by extending their subsequent feeding period to ensure that they begin metamorphosis above critical weight. CAX larvae, by contrast, show no homeostatic adjustment to starvation but start metamorphosis 4 d after feeding onset, regardless of larval size or the state of development of their imaginal discs. By feeding starved CAX larvae for various durations, we found that feeding for only 12-24 h was sufficient to result in metamorphosis on day 4, regardless of further feeding or body size. Manipulation of diet composition showed that protein was the critical macronutrient to initiate this timing. This constant period between the start of feeding and the onset of metamorphosis suggests that larvae possess a molt timer that establishes a minimal time to metamorphosis. Ligation experiments indicate that a portion of the timing may occur in the prothoracic glands. This positive system that promotes molting and the negative control via the critical weight checkpoint provide antagonistic pathways that evolution can modify to adapt growth to the ecological needs of different insects.

}, issn = {1091-6490}, doi = {10.1073/pnas.1311405110}, author = {Suzuki, Yuichiro and Koyama, Takashi and Hiruma, Kiyoshi and Riddiford, Lynn M and Truman, James W} } @article {46710, title = {mRNA on the move: the road to its biological destiny.}, journal = {The Journal of Biological Chemistry}, volume = {288}, year = {2013}, month = {2013 Jul 12}, pages = {20361-8}, abstract = {

Cells have evolved to regulate the asymmetric distribution of specific mRNA targets to institute spatial and temporal control over gene expression. Over the last few decades, evidence has mounted as to the importance of localization elements in the mRNA sequence and their respective RNA-binding proteins. Live imaging methodologies have shown mechanistic details of this phenomenon. In this minireview, we focus on the advanced biochemical and cell imaging techniques used to tweeze out the finer aspects of mechanisms of mRNA movement.

}, keywords = {Animals, Cytoplasm, Humans, In Situ Hybridization, Microscopy, Fluorescence, Models, Genetic, RNA Precursors, RNA Transport, RNA, Messenger, RNA-Binding Proteins}, issn = {1083-351X}, doi = {10.1074/jbc.R113.452094}, author = {Eliscovich, Carolina and Buxbaum, Adina R and Katz, Zachary B and Singer, Robert H} } @article {47644, title = {NBR1 acts as an autophagy receptor for peroxisomes.}, journal = {Journal of cell science}, volume = {126}, year = {2013}, month = {2013 Feb 15}, pages = {939-52}, abstract = {

Selective macro-autophagy is an intracellular process by which large cytoplasmic materials are selectively sequestered and degraded in the lysosomes. Substrate selection is mediated by ubiquitylation and recruitment of ubiquitin-binding autophagic receptors such as p62, NBR1, NDP52 and Optineurin. Although it has been shown that these receptors act cooperatively to target some types of substrates to nascent autophagosomes, their precise roles are not well understood. We examined selective autophagic degradation of peroxisomes (pexophagy), and found that NBR1 is necessary and sufficient for pexophagy. Mutagenesis studies of NBR1 showed that the amphipathic α-helical J domain, the ubiquitin-associated (UBA) domain, the LC3-interacting region and the coiled-coil domain are necessary to mediate pexophagy. Strikingly, substrate selectivity is partly achieved by NBR1 itself by coincident binding of the J and UBA domains to peroxisomes. Although p62 is not required when NBR1 is in excess, its binding to NBR1 increases the efficiency of NBR1-mediated pexophagy. Together, these results suggest that NBR1 is the specific autophagy receptor for pexophagy.

}, keywords = {Autophagy, Blotting, Western, Cell Line, Hela Cells, Humans, Microscopy, Confocal, Microscopy, Electron, Peroxisomes, Proteins}, issn = {1477-9137}, doi = {10.1242/jcs.114819}, author = {Deosaran, Elizabeth and Larsen, Kenneth B and Hua, Rong and Sargent, Graeme and Wang, Yuqing and Kim, Sarah and Lamark, Trond and Jauregui, Miluska and Law, Kelsey and Lippincott-Schwartz, Jennifer and Brech, Andreas and Johansen, Terje and Kim, Peter K} } @article {6579, title = {A neuron-based screening platform for optimizing genetically-encoded calcium indicators.}, journal = {PLoS One}, volume = {8}, year = {2013}, month = {2013}, pages = {e77728}, abstract = {

Fluorescent protein-based sensors for detecting neuronal activity have been developed largely based on non-neuronal screening systems. However, the dynamics of neuronal state variables (e.g., voltage, calcium, etc.) are typically very rapid compared to those of non-excitable cells. We developed an electrical stimulation and fluorescence imaging platform based on dissociated rat primary neuronal cultures. We describe its use in testing genetically-encoded calcium indicators (GECIs). Efficient neuronal GECI expression was achieved using lentiviruses containing a neuronal-selective gene promoter. Action potentials (APs) and thus neuronal calcium levels were quantitatively controlled by electrical field stimulation, and fluorescence images were recorded. Images were segmented to extract fluorescence signals corresponding to individual GECI-expressing neurons, which improved sensitivity over full-field measurements. We demonstrate the superiority of screening GECIs in neurons compared with solution measurements. Neuronal screening was useful for efficient identification of variants with both improved response kinetics and high signal amplitudes. This platform can be used to screen many types of sensors with cellular resolution under realistic conditions where neuronal state variables are in relevant ranges with respect to timing and amplitude.

}, issn = {1932-6203}, doi = {10.1371/journal.pone.0077728}, author = {Wardill, Trevor J. and Chen, Tsai-Wen and Schreiter, Eric R and Hasseman, Jeremy P and Tsegaye, Getahun and Fosque, Benjamin F and Behnam, Reza and Shields, Brenda C and Ramirez, Melissa and Kimmel, Bruce E and Kerr, Rex A and Jayaraman, Vivek and Looger, Loren L and Svoboda, Karel and Kim, Douglas S.} } @article {5883, title = {New technologies in imaging.}, journal = {Molecular Biology of the Cell}, volume = {24}, year = {2013}, month = {2013 Mar}, pages = {669}, abstract = {

Visualization of cellular and molecular processes is an indispensable tool for cell biologists, and innovations in microscopy methods unfailingly lead to new biological discoveries. Today, light microscopy (LM) provides ever-higher spatial and temporal resolution and visualization of biological process over enormous ranges. Electron microscopy (EM) is moving into the atomic resolution regime and allowing cellular analyses that are more physiological and sophisticated in scope. Importantly, much is being gained by combining multiple approaches, (e.g., LM and EM) to take advantage of their complementary strengths. The advent of high-throughput microscopies has led to a common need for sophisticated computational methods to quantitatively analyze huge amounts of data and translate images into new biological insights.

}, issn = {1939-4586}, doi = {10.1091/mbc.E12-12-0867}, author = {Galbraith, Catherine G and Keller, Philipp J and Nogales, Eva} } @article {65888, title = {NGL-2 Regulates Pathway-Specific Neurite Growth and Lamination, Synapse Formation, and Signal Transmission in the Retina}, journal = {Journal of Neuroscience}, volume = {33}, year = {2013}, month = {May-07-2014}, pages = {11949 - 11959}, abstract = {

Parallel processing is an organizing principle of many neural circuits. In the retina, parallel neuronal pathways process signals from rod and cone photoreceptors and support vision over a wide range of light levels. Toward this end, rods and cones form triad synapses with dendrites of distinct bipolar cell types, and the axons or dendrites, respectively, of horizontal cells (HCs). The molecular cues that promote the formation of specific neuronal pathways remain largely unknown. Here, we discover that developing and mature HCs express the leucine-rich repeat (LRR)-containing protein netrin-G ligand 2 (NGL-2). NGL-2 localizes selectively to the tips of HC axons, which form reciprocal connections with rods. In mice with null mutations in Ngl-2 (Ngl-2$^{-}$/$^{-}$), many branches of HC axons fail to stratify in the outer plexiform layer (OPL) and invade the outer nuclear layer. In addition, HC axons expand lateral territories and increase coverage of the OPL, but establish fewer synapses with rods. NGL-2 can form transsynaptic adhesion complexes with netrin-G2, which we show to be expressed by photoreceptors. In Ngl-2$^{-}$/$^{-}$ mice, we find specific defects in the assembly of presynaptic ribbons in rods, indicating that reverse signaling of complexes involving NGL-2 regulates presynaptic maturation. The development of HC dendrites and triad synapses of cone photoreceptors proceeds normally in the absence of NGL-2 and in vivo electrophysiology reveals selective defects in rod-mediated signal transmission in Ngl-2$^{-}$/$^{-}$ mice. Thus, our results identify NGL-2 as a central component of pathway-specific development in the outer retina.

}, issn = {0270-6474}, doi = {10.1523/JNEUROSCI.1521-13.2013}, url = {http://www.jneurosci.org/cgi/doi/10.1523/JNEUROSCI.1521-13.2013}, author = {Soto, F. and Watkins, K. L. and Johnson, R. E. and Schottler, F. and Kerschensteiner, D.} } @article {48165, title = {Omnidirectional displacements for deformable surfaces.}, journal = {Medical image analysis}, volume = {17}, year = {2013}, month = {2013 May}, pages = {429-41}, abstract = {

Deformable surface models are often represented as triangular meshes in image segmentation applications. For a fast and easily regularized deformation onto the target object boundary, the vertices of the mesh are commonly moved along line segments (typically surface normals). However, in case of high mesh curvature, these lines may not intersect with the target boundary at all. Consequently, certain deformations cannot be achieved. We propose omnidirectional displacements for deformable surfaces (ODDS) to overcome this limitation. ODDS allow each vertex to move not only along a line segment but within the volumetric inside of a surrounding sphere, and achieve globally optimal deformations subject to local regularization constraints. However, allowing a ball-shaped instead of a linear range of motion per vertex significantly increases runtime and memory. To alleviate this drawback, we propose a hybrid approach, fastODDS, with improved runtime and reduced memory requirements. Furthermore, fastODDS can also cope with simultaneous segmentation of multiple objects. We show the theoretical benefits of ODDS with experiments on synthetic data, and evaluate ODDS and fastODDS quantitatively on clinical image data of the mandible and the hip bones. There, we assess both the global segmentation accuracy as well as local accuracy in high curvature regions, such as the tip-shaped mandibular coronoid processes and the ridge-shaped acetabular rims of the hip bones.

}, keywords = {Algorithms, Computer Simulation, Humans, Image Enhancement, Image Interpretation, Computer-Assisted, Imaging, Three-Dimensional, Models, Biological, Pattern Recognition, Automated, Reproducibility of Results, Sensitivity and Specificity}, issn = {1361-8423}, doi = {10.1016/j.media.2012.11.006}, author = {Kainmueller, Dagmar and Lamecker, Hans and Heller, Markus O and Weber, Britta and Hege, Hans-Christian and Zachow, Stefan} } @article {6904, title = {The Open Connectome Project Data Cluster: Scalable analysis and vision for high-throughput neuroscience.}, journal = {Scientific and Statistical Database Management: International Conference, SSDBM ... : Proceedings. International Conference on Scientific and Statistical Database Management}, year = {2013}, month = {2013 Jun 18}, abstract = {

We describe a scalable database cluster for the spatial analysis and annotation of high-throughput brain imaging data, initially for 3-d electron microscopy image stacks, but for time-series and multi-channel data as well. The system was designed primarily for workloads that build connectomes- neural connectivity maps of the brain-using the parallel execution of computer vision algorithms on high-performance compute clusters. These services and open-science data sets are publicly available at openconnecto.me. The system design inherits much from NoSQL scale-out and data-intensive computing architectures. We distribute data to cluster nodes by partitioning a spatial index. We direct I/O to different systems-reads to parallel disk arrays and writes to solid-state storage-to avoid I/O interference and maximize throughput. All programming interfaces are RESTful Web services, which are simple and stateless, improving scalability and usability. We include a performance evaluation of the production system, highlighting the effec-tiveness of spatial data organization.

}, doi = {10.1145/2484838.2484870}, author = {Burns, Randal and Roncal, William Gray and Kleissas, Dean and Lillaney, Kunal and Manavalan, Priya and Perlman, Eric and Berger, Daniel R. and Bock, Davi D and Chung, Kwanghun and Grosenick, Logan and Kasthuri, Narayanan and Weiler, Nicholas C and Deisseroth, Karl and Kazhdan, Michael and Lichtman, Jeff and Reid, R Clay and Smith, Stephen J and Szalay, Alexander S and Vogelstein, Joshua T and Vogelstein, R Jacob} } @article {6526, title = {The Oxytricha trifallax macronuclear genome: a complex eukaryotic genome with 16,000 tiny chromosomes.}, journal = {PLoS Biology}, volume = {11}, year = {2013}, month = {2013 Jan}, pages = {e1001473}, abstract = {

The macronuclear genome of the ciliate Oxytricha trifallax displays an extreme and unique eukaryotic genome architecture with extensive genomic variation. During sexual genome development, the expressed, somatic macronuclear genome is whittled down to the genic portion of a small fraction (\~{}5\%) of its precursor "silent" germline micronuclear genome by a process of "unscrambling" and fragmentation. The tiny macronuclear "nanochromosomes" typically encode single, protein-coding genes (a small portion, 10\%, encode 2-8 genes), have minimal noncoding regions, and are differentially amplified to an average of \~{}2,000 copies. We report the high-quality genome assembly of \~{}16,000 complete nanochromosomes (\~{}50 Mb haploid genome size) that vary from 469 bp to 66 kb long (mean \~{}3.2 kb) and encode \~{}18,500 genes. Alternative DNA fragmentation processes \~{}10\% of the nanochromosomes into multiple isoforms that usually encode complete genes. Nucleotide diversity in the macronucleus is very high (SNP heterozygosity is \~{}4.0\%), suggesting that Oxytricha trifallax may have one of the largest known effective population sizes of eukaryotes. Comparison to other ciliates with nonscrambled genomes and long macronuclear chromosomes (on the order of 100 kb) suggests several candidate proteins that could be involved in genome rearrangement, including domesticated MULE and IS1595-like DDE transposases. The assembly of the highly fragmented Oxytricha macronuclear genome is the first completed genome with such an unusual architecture. This genome sequence provides tantalizing glimpses into novel molecular biology and evolution. For example, Oxytricha maintains tens of millions of telomeres per cell and has also evolved an intriguing expansion of telomere end-binding proteins. In conjunction with the micronuclear genome in progress, the O. trifallax macronuclear genome will provide an invaluable resource for investigating programmed genome rearrangements, complementing studies of rearrangements arising during evolution and disease.

}, keywords = {Base Sequence, DNA, DNA Copy Number Variations, DNA Fragmentation, Gene Amplification, Gene Rearrangement, Genes, Genetic Variation, Genome, Macronucleus, Messenger, Molecular Sequence Data, Oxytricha, Protein Binding, Protozoan, RNA, Sequence Analysis, Telomere}, issn = {1545-7885}, doi = {10.1371/journal.pbio.1001473}, author = {Swart, Estienne C and Bracht, John R and Magrini, Vincent and Minx, Patrick and Chen, Xiao and Zhou, Yi and Khurana, Jaspreet S and Goldman, Aaron D and Nowacki, Mariusz and Schotanus, Klaas and Jung, Seolkyoung and Fulton, Robert S and Ly, Amy and McGrath, Sean and Haub, Kevin and Wiggins, Jessica L and Storton, Donna and Matese, John C and Parsons, Lance and Chang, Wei-Jen and Bowen, Michael S and Stover, Nicholas A and Jones, Thomas A and Eddy, Sean R. and Herrick, Glenn A and Doak, Thomas G and Wilson, Richard K and Mardis, Elaine R and Landweber, Laura F} } @article {6436, title = {Sensorimotor structure of Drosophila larva phototaxis.}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {110}, year = {2013}, month = {2013 Sep 16}, pages = {E3868-77}, abstract = {

The avoidance of light by fly larvae is a classic paradigm for sensorimotor behavior. Here, we use behavioral assays and video microscopy to quantify the sensorimotor structure of phototaxis using the Drosophila larva. Larval locomotion is composed of sequences of runs (periods of forward movement) that are interrupted by abrupt turns, during which the larva pauses and sweeps its head back and forth, probing local light information to determine the direction of the successive run. All phototactic responses are mediated by the same set of sensorimotor transformations that require temporal processing of sensory inputs. Through functional imaging and genetic inactivation of specific neurons downstream of the sensory periphery, we have begun to map these sensorimotor circuits into the larval central brain. We find that specific sensorimotor pathways that govern distinct light-evoked responses begin to segregate at the first relay after the photosensory neurons.

}, issn = {1091-6490}, doi = {10.1073/pnas.1215295110}, author = {Kane, Elizabeth A and Gershow, Marc and Afonso, Bruno and Larderet, Ivan and Klein, Mason and Carter, Ashley R and de Bivort, Benjamin L and Sprecher, Simon G and Samuel, Aravinthan D. T.} } @article {46711, title = {Single cell analysis of RNA-mediated histone H3.3 recruitment to a cytomegalovirus promoter-regulated transcription site.}, journal = {The Journal of Biological Chemistry}, volume = {288}, year = {2013}, month = {2013 Jul 5}, pages = {19882-99}, abstract = {

Unlike the core histones, which are incorporated into nucleosomes concomitant with DNA replication, histone H3.3 is synthesized throughout the cell cycle and utilized for replication-independent (RI) chromatin assembly. The RI incorporation of H3.3 into nucleosomes is highly conserved and occurs at both euchromatin and heterochromatin. However, neither the mechanism of H3.3 recruitment nor its essential function is well understood. Several different chaperones regulate H3.3 assembly at distinct sites. The H3.3 chaperone, Daxx, and the chromatin-remodeling factor, ATRX, are required for H3.3 incorporation and heterochromatic silencing at telomeres, pericentromeres, and the cytomegalovirus (CMV) promoter. By evaluating H3.3 dynamics at a CMV promoter-regulated transcription site in a genetic background in which RI chromatin assembly is blocked, we have been able to decipher the regulatory events upstream of RI nucleosomal deposition. We find that at the activated transcription site, H3.3 accumulates with sense and antisense RNA, suggesting that it is recruited through an RNA-mediated mechanism. Sense and antisense transcription also increases after H3.3 knockdown, suggesting that the RNA signal is amplified when chromatin assembly is blocked and attenuated by nucleosomal deposition. Additionally, we find that H3.3 is still recruited after Daxx knockdown, supporting a chaperone-independent recruitment mechanism. Sequences in the H3.3 N-terminal tail and αN helix mediate both its recruitment to RNA at the activated transcription site and its interaction with double-stranded RNA in vitro. Interestingly, the H3.3 gain-of-function pediatric glioblastoma mutations, G34R and K27M, differentially affect H3.3 affinity in these assays, suggesting that disruption of an RNA-mediated regulatory event could drive malignant transformation.

}, keywords = {Cell Line, Chromatin Assembly and Disassembly, Cytomegalovirus, Histones, Humans, Nucleosomes, Promoter Regions, Genetic, Protein Structure, Secondary, RNA, Viral, Transcription, Genetic}, issn = {1083-351X}, doi = {10.1074/jbc.M113.473181}, author = {Newhart, Alyshia and Rafalska-Metcalf, Ilona U and Yang, Tian and Joo, Lucy M and Powers, Sara Lawrence and Kossenkov, Andrew V and Lopez-Jones, Melissa and Singer, Robert H and Showe, Louise C and Skordalakes, Emmanuel and Janicki, Susan M} } @article {6125, title = {{\ss}-synuclein aggregates and induces neurodegeneration in dopaminergic neurons.}, journal = {Annals of Neurology}, volume = {74}, year = {2013}, month = {2013 Mar 28}, pages = {109-18}, abstract = {

Objective: While the contribution of α-Synuclein to neurodegeneration in Parkinson{\textquoteright}s disease is well accepted, the putative impact of its close homologue, β-Synuclein, is enigmatic. β-Synuclein is widely expressed throughout the central nervous system as is α-Synuclein, but the physiological functions of both proteins remain unknown. Recent findings supported the view that β-Synuclein can act as an ameliorating regulator of α-Synuclein-induced neurotoxicity, having neuroprotective rather than neurodegenerative capabilities, and being non-aggregating due to absence of most part of the aggregation-promoting NAC domain. However, a mutation of β-Synuclein linked to dementia with Lewy bodies rendered the protein neurotoxic in transgenic mice and fibrillation of β-Synuclein has been demonstrated in vitro. Methods / Results: Supporting the hypothesis that β-Synuclein can act as a neurodegeneration-inducing factor we now demonstrate that wild-type β-Synuclein is neurotoxic for cultured primary neurons. Furthermore, β-Synuclein formed proteinase K resistant aggregates in dopaminergic neurons in vivo, leading to pronounced and progressive neurodegeneration in rats. Expression of β-Synuclein caused mitochondrial fragmentation, but this fragmentation did not render mitochondria non-functional in terms of ion handling and respiration even in late stages of neurodegeneration. A comparison of the neurodegenerative effects induced by α-, β-, and γ-Synuclein revealed that β-Synuclein was eventually as neurotoxic as α-Synuclein for nigral dopaminergic neurons, while γ-Synuclein proved to be non-toxic and had very low aggregation propensity. Interpretation: Our results suggest that the role of β-Synuclein as a putative modulator of neuropathology in aggregopathies like Parkinson{\textquoteright}s disease and dementia with Lewy bodies needs to be revisited. ANN NEUROL 2013. {\textcopyright} 2013 American Neurological Association.

}, issn = {1531-8249}, doi = {10.1002/ana.23905}, author = {Taschenberger, G and Toloe, J and Tereshchenko, J and Akerboom, J and Wales, P and Benz, R and Becker, S and Outeiro, Tf and Looger, L and B{\"a}hr, M and Zweckstetter, M and K{\"u}gler, S} } @article {6582, title = {Structure of fully liganded Hb zeta(2)beta(s)(2) trapped in a tense conformation.}, journal = {Acta Crystallographica Section D:Biological Crystallography}, volume = {69}, year = {2013}, month = {2013 Oct}, pages = {2061-71}, abstract = {

A variant Hb zeta(2)beta(s)(2) that is formed from sickle hemoglobin (Hb S; alpha(2)beta(s)(2)) by exchanging adult alpha-globin with embryonic zeta-globin subunits shows promise as a therapeutic agent for sickle-cell disease (SCD). Hb zeta(2)beta(s)(2) inhibits the polymerization of deoxygenated Hb S in vitro and reverses characteristic features of SCD in vivo in mouse models of the disorder. When compared with either Hb S or with normal human adult Hb A (alpha(2)beta(2)), Hb zeta(2)beta(s)(2) exhibits atypical properties that include a high oxygen affinity, reduced cooperativity, a weak Bohr effect and blunted 2,3-diphosphoglycerate allostery. Here, the 1.95 angstrom resolution crystal structure of human Hb zeta(2)beta(s)(2) that was expressed in complex transgenic knockout mice and purified from their erythrocytes is presented. When fully liganded with carbon monoxide, Hb zeta(2)beta(s)(2) displays a central water cavity, a zeta 1-beta(s)2 (or zeta 2-beta(s)1) interface, intersubunit salt-bridge/hydrogen-bond interactions, C-terminal beta His146 salt-bridge interactions, and a beta-cleft, that are highly unusual for a relaxed hemoglobin structure and are more typical of a tense conformation. These quaternary tense-like features contrast with the tertiary relaxed-like conformations of the zeta 1-beta(s1) dimer and the CD and FG corners, as well as the overall structures of the heme cavities. This crystallographic study provides insights into the altered oxygen-transport properties of Hb zeta(2)beta(s)(2) and, moreover, decouples tertiary- and quaternary-structural events that are critical to Hb ligand binding and allosteric function.

}, issn = {0907-444910.1107/s0907444913019197}, doi = {10.1107/S0907444913019197}, author = {Safo, M. K. and Ko, T. P. and Abdulmalik, O. and He, Z. N. and Wang, A. H. J. and Schreiter, E. R. and Russell, J. E.} } @article {47633, title = {Superresolution imaging with standard fluorescent probes.}, journal = {Current protocols in cell biology / editorial board, Juan S. Bonifacino ... [et al.]}, volume = {60}, year = {2013}, month = {2013}, pages = {Unit 21.8.}, abstract = {

For more than 100 years, the ultimate resolution of a light microscope (\~{} 200 nm) has been constrained by the fundamental physical phenomenon of diffraction, as described by Ernst Abbe in 1873. While this limitation is just as applicable to today{\textquoteright}s light microscopes, it is the combination of high-end optics, clever methods of sample illumination, and computational techniques that has enabled researchers to access information at an order of magnitude greater resolution than once thought possible. This combination, broadly termed superresolution microscopy, has been increasingly practical for many labs to implement from both a hardware and software standpoint, but, as with many cutting-edge techniques, it also comes with limitations. One of the current drawbacks to superresolution microscopy is the limited number of probes and conditions that have been suitable for imaging. Here, a technique termed bleaching/blinking-assisted localization microscopy (BaLM) makes use of the inherent blinking and bleaching properties of almost all fluorophores as a means to generate superresolution images.

}, keywords = {Animals, Cercopithecus aethiops, COS Cells, Microscopy, Fluorescence, Signal-To-Noise Ratio}, issn = {1934-2616}, doi = {10.1002/0471143030.cb2108s60}, author = {Millis, Bryan A and Burnette, Dylan T and Lippincott-Schwartz, Jennifer and Kachar, Bechara} } @article {46606, title = {Taok2 controls behavioral response to ethanol in mice.}, journal = {Genes, Brain, and Behavior}, volume = {12}, year = {2013}, month = {2013 Feb}, pages = {87-97}, abstract = {

Despite recent advances in the understanding of ethanol{\textquoteright}s biological action, many of the molecular targets of ethanol and mechanisms behind ethanol{\textquoteright}s effect on behavior remain poorly understood. In an effort to identify novel genes, the products of which regulate behavioral responses to ethanol, we recently identified a mutation in the dtao gene that confers resistance to the locomotor stimulating effect of ethanol in Drosophila. dtao encodes a member of the Ste20 family of serine/threonine kinases implicated in MAP kinase signaling pathways. In this study, we report that conditional ablation of the mouse dtao homolog, Taok2, constitutively and specifically in the nervous system, results in strain-specific and overlapping alterations in ethanol-dependent behaviors. These data suggest a functional conservation of dtao and Taok2 in mediating ethanol{\textquoteright}s biological action and identify Taok2 as a putative candidate gene for ethanol use disorders in humans.

}, keywords = {Alcohol Drinking, Alcoholism, Animals, Behavior, Animal, Conditioning, Classical, Ethanol, Gait Ataxia, Locomotion, MAP Kinase Kinase Kinases, Mice, Mice, Inbred C57BL, Mice, Transgenic, Protein-Serine-Threonine Kinases}, issn = {1601-183X}, doi = {10.1111/j.1601-183X.2012.00834.x}, author = {Kapfhamer, D and Taylor, S and Zou, M E and Lim, J P and Kharazia, V and Heberlein, U} } @article {6674, title = {Thalamocortical input onto layer 5 pyramidal neurons measured using quantitative large-scale array tomography.}, journal = {Frontiers in Neural Circuits}, volume = {7}, year = {2013}, month = {2013}, pages = {177}, abstract = {

The subcellular locations of synapses on pyramidal neurons strongly influences dendritic integration and synaptic plasticity. Despite this, there is little quantitative data on spatial distributions of specific types of synaptic input. Here we use array tomography (AT), a high-resolution optical microscopy method, to examine thalamocortical (TC) input onto layer 5 pyramidal neurons. We first verified the ability of AT to identify synapses using parallel electron microscopic analysis of TC synapses in layer 4. We then use large-scale array tomography (LSAT) to measure TC synapse distribution on L5 pyramidal neurons in a 1.00 {\texttimes} 0.83 {\texttimes} 0.21 mm(3) volume of mouse somatosensory cortex. We found that TC synapses primarily target basal dendrites in layer 5, but also make a considerable input to proximal apical dendrites in L4, consistent with previous work. Our analysis further suggests that TC inputs are biased toward certain branches and, within branches, synapses show significant clustering with an excess of TC synapse nearest neighbors within 5-15 μm compared to a random distribution. Thus, we show that AT is a sensitive and quantitative method to map specific types of synaptic input on the dendrites of entire neurons. We anticipate that this technique will be of wide utility for mapping functionally-relevant anatomical connectivity in neural circuits.

}, issn = {1662-5110}, doi = {10.3389/fncir.2013.00177}, author = {Rah, Jong-Cheol and Bas, Erhan and Colonell, Jennifer and Mishchenko, Yuriy and Karsh, Bill and Fetter, Richard D and Myers, Eugene W and Chklovskii, Dmitri B and Svoboda, Karel and Harris, Timothy D and Isaac, John T R} } @article {5935, title = {Towards comprehensive cell lineage reconstructions in complex organisms using light-sheet microscopy.}, journal = {Development, Growth and Differentiation}, volume = {55}, year = {2013}, month = {2013 Apr 29}, pages = {563-78}, abstract = {

Understanding the development of complex multicellular organisms as a function of the underlying cell behavior is one of the most fundamental goals of developmental biology. The ability to quantitatively follow cell dynamics in entire developing embryos is an indispensable step towards such a system-level understanding. In recent years, light-sheet fluorescence microscopy has emerged as a particularly promising strategy for recording the in vivo data required to realize this goal. Using light-sheet fluorescence microscopy, entire complex organisms can be rapidly imaged in three dimensions at sub-cellular resolution, achieving high temporal sampling and excellent signal-to-noise ratio without damaging the living specimen or bleaching fluorescent markers. The resulting datasets allow following individual cells in vertebrate and higher invertebrate embryos over up to several days of development. However, the complexity and size of these multi-terabyte recordings typically preclude comprehensive manual analyses. Thus, new computational approaches are required to automatically segment cell morphologies, accurately track cell identities and systematically analyze cell behavior throughout embryonic development. We review current efforts in light-sheet microscopy and bioimage informatics towards this goal, and argue that comprehensive cell lineage reconstructions are finally within reach for many key model organisms, including fruit fly, zebrafish and mouse.

}, issn = {1440-169X}, doi = {10.1111/dgd.12063}, author = {Amat, Fernando and Keller, Philipp J} } @article {68814, title = {Transgenic tools to characterize neuronal properties of discrete populations of zebrafish neurons}, journal = {Development}, volume = {140}, year = {2013}, month = {09/2013}, pages = {3927-3931}, abstract = {

The developing nervous system consists of a variety of cell types. Transgenic animals expressing reporter genes in specific classes of neuronal cells are powerful tools for the study of neuronal network formation. We generated a wide variety of transgenic zebrafish that expressed reporter genes in specific classes of neurons or neuronal progenitors. These include lines in which neurons of specific neurotransmitter phenotypes expressed fluorescent proteins or Gal4, and lines in which specific subsets of the dorsal progenitor domain in the spinal cord expressed fluorescent proteins. Using these, we examined domain organization in the developing dorsal spinal cord, and found that there are six progenitor domains in zebrafish, which is similar to the domain organization in mice. We also systematically characterized neurotransmitter properties of the neurons that are produced from each domain. Given that reporter gene expressions occurs in a wide area of the nervous system in the lines generated, these transgenic fish should serve as powerful tools for the investigation of not only the neurons in the dorsal spinal cord but also neuronal structures and functions in many other regions of the nervous system.

}, issn = {0950-1991}, doi = {10.1242/dev.099531}, url = {https://doi.org/10.1242/dev.099531}, author = {Satou, Chie and Kimura, Yukiko and Hirata, Hiromi and Suster, Maximiliano L. and Kawakami, Koichi and Higashijima, Shin-Ichi} } @article {6124, title = {Ultrasensitive fluorescent proteins for imaging neuronal activity.}, journal = {Nature}, volume = {499}, year = {2013}, month = {2013 Jul 18}, pages = {295-300}, abstract = {

Fluorescent calcium sensors are widely used to image neural activity. Using structure-based mutagenesis and neuron-based screening, we developed a family of ultrasensitive protein calcium sensors (GCaMP6) that outperformed other sensors in cultured neurons and in zebrafish, flies and mice in vivo. In layer 2/3 pyramidal neurons of the mouse visual cortex, GCaMP6 reliably detected single action potentials in neuronal somata and orientation-tuned synaptic calcium transients in individual dendritic spines. The orientation tuning of structurally persistent spines was largely stable over timescales of weeks. Orientation tuning averaged across spine populations predicted the tuning of their parent cell. Although the somata of GABAergic neurons showed little orientation tuning, their dendrites included highly tuned dendritic segments (5{\textendash}40-{\textmu}m long). GCaMP6 sensors thus provide new windows into the organization and dynamics of neural circuits over multiple spatial and temporal scales.

}, doi = {10.1038/nature12354}, author = {Chen, Tsai-Wen and Wardill, Trevor J. and Sun, Yi and Pulvar, Stefan R. and Renninger, Sabine L. and Baohan, Amy and Schreiter, Eric R and Kerr, Rex A and Orger, Michael B. and Jayaraman, Vivek and Looger, Loren L and Svoboda, Karel and Kim, Douglas S.} } @article {6508, title = {Vertebrate versus invertebrate neural circuits.}, journal = {Current Biology}, volume = {23}, year = {2013}, month = {2013 Jun 17}, pages = {R504-6}, issn = {1879-0445}, author = {Katz, Paul and Grillner, Sten and Wilson, Rachel and Borst, Alexander and Greenspan, Ralph and Buzs{\'a}ki, Gy{\"o}rgy and Martin, Kevan and Marder, Eve and Kristan, William and Friedrich, Rainer and Chklovskii, Dmitri Mitya} } @article {6178, title = {A visual motion detection circuit suggested by Drosophila connectomics.}, journal = {Nature}, volume = {500}, year = {2013}, month = {2013 Aug 7}, pages = {175{\textendash}81}, abstract = {

Animal behaviour arises from computations in neuronal circuits, but our understanding of these computations has been frustrated by the lack of detailed synaptic connection maps, or connectomes. For example, despite intensive investigations over half a century, the neuronal implementation of local motion detection in the insect visual system remains elusive. Here we develop a semi-automated pipeline using electron microscopy to reconstruct a connectome, containing 379 neurons and 8,637 chemical synaptic contacts, within the Drosophila optic medulla. By matching reconstructed neurons to examples from light microscopy, we assigned neurons to cell types and assembled a connectome of the repeating module of the medulla. Within this module, we identified cell types constituting a motion detection circuit, and showed that the connections onto individual motion-sensitive neurons in this circuit were consistent with their direction selectivity. Our results identify cellular targets for future functional investigations, and demonstrate that connectomes can provide key insights into neuronal computations.

}, issn = {1476-4687}, doi = {doi:10.1038/nature12450}, url = {http://www.nature.com/nature/journal/v500/n7461/full/nature12450.html}, author = {Takemura, Shin-ya and Bharioke, Arjun and Lu, Zhiyuan and Nern, Aljoscha and Vitaladevuni, Shiv and Rivlin, Patricia K. and Katz, William T. and Olbris, Donald J. and Plaza, Stephen M and Winston, Philip and Zhao, Ting and Horne, Jane Anne and Fetter, Richard D and Takemura, Satoko and Blazek, Katerina and Chang, Lei-Ann and Ogundeyi, Omotara and Saunders, Mathew A. and Shapiro, Victor and Sigmund, Christopher and Rubin, Gerald M and Scheffer, Louis K and Meinertzhagen, Ian A. and Chklovskii, Dmitri B} } @article {5937, title = {Whole-brain functional imaging at cellular resolution using light-sheet microscopy.}, journal = {Nature Methods}, volume = {10}, year = {2013}, month = {2013 May}, pages = {413-20}, abstract = {

Brain function relies on communication between large populations of neurons across multiple brain areas, a full understanding of which would require knowledge of the time-varying activity of all neurons in the central nervous system. Here we use light-sheet microscopy to record activity, reported through the genetically encoded calcium indicator GCaMP5G, from the entire volume of the brain of the larval zebrafish in vivo at 0.8 Hz, capturing more than 80\% of all neurons at single-cell resolution. Demonstrating how this technique can be used to reveal functionally defined circuits across the brain, we identify two populations of neurons with correlated activity patterns. One circuit consists of hindbrain neurons functionally coupled to spinal cord neuropil. The other consists of an anatomically symmetric population in the anterior hindbrain, with activity in the left and right halves oscillating in antiphase, on a timescale of 20 s, and coupled to equally slow oscillations in the inferior olive.

}, issn = {1548-7105}, doi = {10.1038/nmeth.2434}, author = {Ahrens, Misha B and Orger, Michael B. and Robson, Drew N and Li, Jennifer M and Keller, Philipp J} } @article {48166, title = {Automatic detection and classification of teeth in CT data.}, journal = {Medical image computing and computer-assisted intervention : MICCAI ... International Conference on Medical Image Computing and Computer-Assisted Intervention}, volume = {15}, year = {2012}, month = {2012}, pages = {609-16}, abstract = {

We propose a fully automatic method for tooth detection and classification in CT or cone-beam CT image data. First we compute an accurate segmentation of the maxilla bone. Based on this segmentation, our method computes a complete and optimal separation of the row of teeth into 16 subregions and classifies the resulting regions as existing or missing teeth. This serves as a prerequisite for further individual tooth segmentation. We show the robustness of our approach by providing extensive validation on 43 clinical head CT scans.

}, keywords = {Algorithms, Artifacts, Bone and Bones, Humans, Image Processing, Computer-Assisted, Imaging, Three-Dimensional, Maxilla, Models, Statistical, Pattern Recognition, Automated, Radiographic Image Interpretation, Computer-Assisted, Reproducibility of Results, Tomography, X-Ray Computed, Tooth}, author = {Duy, Nguyen The and Lamecker, Hans and Kainmueller, Dagmar and Zachow, Stefan} } @article {4920, title = {Computational design of self-assembling protein nanomaterials with atomic level accuracy.}, journal = {Science}, volume = {336}, year = {2012}, month = {2012 Jun 1}, pages = {1171-4}, abstract = {

We describe a general computational method for designing proteins that self-assemble to a desired symmetric architecture. Protein building blocks are docked together symmetrically to identify complementary packing arrangements, and low-energy protein-protein interfaces are then designed between the building blocks in order to drive self-assembly. We used trimeric protein building blocks to design a 24-subunit, 13-nm diameter complex with octahedral symmetry and a 12-subunit, 11-nm diameter complex with tetrahedral symmetry. The designed proteins assembled to the desired oligomeric states in solution, and the crystal structures of the complexes revealed that the resulting materials closely match the design models. The method can be used to design a wide variety of self-assembling protein nanomaterials.

}, issn = {1095-9203}, doi = {10.1126/science.1219364}, author = {King, Neil P and Sheffler, William and Sawaya, Michael R and Vollmar, Breanna S and Sumida, John P and Andr{\'e}, Ingemar and Gonen, Tamir and Yeates, Todd O and Baker, David} } @article {47649, title = {Computational model of cytokinetic abscission driven by ESCRT-III polymerization and remodeling.}, journal = {Biophysical journal}, volume = {102}, year = {2012}, month = {2012 May 16}, pages = {2309-20}, abstract = {

The endosomal sorting complex required for transport (ESCRT)-III complex, capable of polymerization and remodeling, participates in abscission of the intercellular membrane bridge connecting two daughter cells at the end of cytokinesis. Here, we integrate quantitative imaging of ESCRT-III during cytokinetic abscission with biophysical properties of ESCRT-III complexes to formulate and test a computational model for ESCRT-mediated cytokinetic abscission. We propose that cytokinetic abscission is driven by an ESCRT-III fission complex, which arises from ESCRT-III polymerization at the edge of the cytokinetic midbody structure, located at the center of the intercellular bridge. Formation of the fission complex is completed by remodeling and breakage of the ESCRT-III polymer assisted by VPS4. Subsequent spontaneous constriction of the fission complex generates bending deformation of the intercellular bridge membrane. The related membrane elastic force propels\ the fission complex along the intercellular bridge away from the midbody until it reaches an equilibrium position, determining the scission site. Membrane attachment to the dome-like end-cap of the fission complex drives membrane fission, completing the abscission process. We substantiate the model by theoretical analysis of the membrane elastic energy and by experimental verification of the major model assumptions.

}, keywords = {Adenosine Triphosphate, Animals, Cell Line, Cell Membrane, Computer Simulation, Cytokinesis, Dogs, Endosomal Sorting Complexes Required for Transport, Extracellular Space, Humans, Models, Biological, Polymerization, Reproducibility of Results, Vacuolar Proton-Translocating ATPases}, issn = {1542-0086}, doi = {10.1016/j.bpj.2012.04.007}, author = {Elia, Natalie and Fabrikant, Gur and Kozlov, Michael M and Lippincott-Schwartz, Jennifer} } @article {5924, title = {Control of timing, rate and bursts of hippocampal place cells by dendritic and somatic inhibition.}, journal = {Nature neuroscience}, volume = {15}, year = {2012}, month = {2012 May}, pages = {769-75}, abstract = {A consortium of inhibitory neurons control the firing patterns of pyramidal cells, but their specific roles in the behaving animal are largely unknown. We performed simultaneous physiological recordings and optogenetic silencing of either perisomatic (parvalbumin (PV) expressing) or dendrite-targeting (somatostatin (SOM) expressing) interneurons in hippocampal area CA1 of head-fixed mice actively moving a treadmill belt rich with visual-tactile stimuli. Silencing of either PV or SOM interneurons increased the firing rates of pyramidal cells selectively in their place fields, with PV and SOM interneurons having their largest effect during the rising and decaying parts of the place field, respectively. SOM interneuron silencing powerfully increased burst firing without altering the theta phase of spikes. In contrast, PV interneuron silencing had no effect on burst firing, but instead shifted the spikes{\textquoteright} theta phase toward the trough of theta. These findings indicate that perisomatic and dendritic inhibition have distinct roles in controlling the rate, burst and timing of hippocampal pyramidal cells.}, keywords = {Action Potentials, Animal, Animals, Behavior, Biophysics, Dendrites, Electric Stimulation, Exercise Test, Green Fluorescent Proteins, Halorhodopsins, Hippocampus, Humans, Inbred C57BL, Light, Mice, Movement, Neural Inhibition, Neurons, Optics and Photonics, Parvalbumins, Patch-Clamp Techniques, Photic Stimulation, Probability, Somatostatin, Spectrum Analysis, Time Factors, Time Perception, Touch, Transgenic}, issn = {1546-1726}, doi = {10.1038/nn.3077}, author = {Royer, S{\'e}bastien and Zemelman, Boris V and Losonczy, Attila and Kim, Jinhyun and Chance, Frances and Magee, Jeffrey C and Buzs{\'a}ki, Gy{\"o}rgy} } @article {45187, title = {Correlative 3D superresolution fluorescence and electron microscopy reveal the relationship of mitochondrial nucleoids to membranes.}, journal = {Proceedings of the National Academy of Science of the United States of America}, volume = {109}, year = {2012}, month = {2012 Apr 17}, pages = {6136-41}, abstract = {

Microscopic images of specific proteins in their cellular context yield important insights into biological processes and cellular architecture. The advent of superresolution optical microscopy techniques provides the possibility to augment EM with nanometer-resolution fluorescence microscopy to access the precise location of proteins in the context of cellular ultrastructure. Unfortunately, efforts to combine superresolution fluorescence and EM have been stymied by the divergent and incompatible sample preparation protocols of the two methods. Here, we describe a protocol that preserves both the delicate photoactivatable fluorescent protein labels essential for superresolution microscopy and the fine ultrastructural context of EM. This preparation enables direct 3D imaging in 500- to 750-nm sections with interferometric photoactivatable localization microscopy followed by scanning EM images generated by focused ion beam ablation. We use this process to "colorize" detailed EM images of the mitochondrion with the position of labeled proteins. The approach presented here has provided a new level of definition of the in vivo nature of organization of mitochondrial nucleoids, and we expect this straightforward method to be applicable to many other biological questions that can be answered by direct imaging.

}, keywords = {3T3 Cells, Animals, DNA, Mitochondrial, DNA-Binding Proteins, Fibroblasts, High Mobility Group Proteins, Imaging, Three-Dimensional, Luminescent Proteins, Mice, Microscopy, Electron, Microscopy, Fluorescence, Microscopy, Interference, Mitochondria, Mitochondrial Membranes, Mitochondrial Proteins, Reproducibility of Results}, issn = {1091-6490}, doi = {10.1073/pnas.1121558109}, author = {Kopek, Benjamin G and Shtengel, Gleb and Xu, C Shan and Clayton, David A and Hess, Harald F} } @article {69345, title = {Differential uptake of MRI contrast agents indicates charge-selective blood-brain interface in the crayfish}, journal = {Cell and Tissue Research}, volume = {349}, year = {2012}, month = {2012 Apr 19}, pages = {493 - 503}, abstract = {

This study provides a new perspective on the long-standing problem of the nature of the decapod crustacean blood-brain interface. Previous studies of crustacean blood-brain interface permeability have relied on invasive histological, immunohistochemical and electrophysiological techniques, indicating a leaky non-selective blood-brain barrier. The present investigation involves the use of magnetic resonance imaging (MRI), a method for non-invasive longitudinal tracking of tracers in real-time. Differential uptake rates of two molecularly distinct MRI contrast agents, namely manganese (Mn(II)) and Magnevist{\textregistered} (Gd-DTPA), were observed and quantified in the crayfish,\ Cherax destructor. Contrast agents were injected into the pericardium and uptake was observed with longitudinal MRI for approximately 14.5\ h. Mn(II) was taken up quickly into neural tissue (within 6.5\ min), whereas Gd-DTPA was not taken up into neural tissue and was instead restricted to the intracerebral vasculature or excreted into nearby sinuses. Our results provide evidence for a charge-selective intracerebral blood-brain interface in the crustacean nervous system, a structural characteristic once considered too complex for a lower-order arthropod.

}, issn = {0302-766X}, doi = {10.1007/s00441-012-1413-9}, url = {http://link.springer.com/10.1007/s00441-012-1413-9http://link.springer.com/content/pdf/10.1007/s00441-012-1413-9http://link.springer.com/content/pdf/10.1007/s00441-012-1413-9.pdfhttp://link.springer.com/article/10.1007/s00441-012-1413-9/fulltext.html}, author = {Otopalik, Adriane G. and Shin, Jane and Beltz, Barbara S. and Sandeman, David C. and Kolodny, Nancy H.} } @article {46605, title = {DlgS97/SAP97, a neuronal isoform of discs large, regulates ethanol tolerance.}, journal = {PLoS One}, volume = {7}, year = {2012}, month = {2012 Nov 7}, pages = {e48967}, abstract = {

From a genetic screen for Drosophila melanogaster mutants with altered ethanol tolerance, we identified intolerant (intol), a novel allele of discs large 1 (dlg1). Dlg1 encodes Discs Large 1, a MAGUK (Membrane Associated Guanylate Kinase) family member that is the highly conserved homolog of mammalian PSD-95 and SAP97. The intol mutation disrupted specifically the expression of DlgS97, a SAP97 homolog, and one of two major protein isoforms encoded by dlg1 via alternative splicing. Expression of the major isoform, DlgA, a PSD-95 homolog, appeared unaffected. Ethanol tolerance in the intol mutant could be partially restored by transgenic expression of DlgS97, but not DlgA, in specific neurons of the fly{\textquoteright}s brain. Based on co-immunoprecipitation, DlgS97 forms a complex with N-methyl-D-aspartate (NMDA) receptors, a known target of ethanol. Consistent with these observations, flies expressing reduced levels of the essential NMDA receptor subunit dNR1 also showed reduced ethanol tolerance, as did mutants in the gene calcium/calmodulin-dependent protein kinase (caki), encoding the fly homolog of mammalian CASK, a known binding partner of DlgS97. Lastly, mice in which SAP97, the mammalian homolog of DlgS97, was conditionally deleted in adults failed to develop rapid tolerance to ethanol{\textquoteright}s sedative/hypnotic effects. We propose that DlgS97/SAP97 plays an important and conserved role in the development of tolerance to ethanol via NMDA receptor-mediated synaptic plasticity.

}, keywords = {Alleles, Alternative Splicing, Animals, Drosophila melanogaster, Drosophila Proteins, Ethanol, Female, Guanylate Kinase, Male, Membrane Proteins, Mice, Mice, Inbred C57BL, Mutation, Neurons, Protein Isoforms, Receptors, N-Methyl-D-Aspartate, Repressor Proteins, Tumor Suppressor Proteins}, issn = {1932-6203}, doi = {10.1371/journal.pone.0048967}, author = {Maiya, Rajani and Lee, Seonok and Berger, Karen H and Kong, Eric C and Slawson, Justin B and Griffith, Leslie C and Takamiya, Kogo and Huganir, Richard L and Margolis, Ben and Heberlein, Ulrike} } @article {6854, title = {Dopamine neurons modulate pheromone responses in Drosophila courtship learning.}, journal = {Nature}, volume = {489}, year = {2012}, month = {2012 Sep 6}, pages = {145-9}, abstract = {

Learning through trial-and-error interactions allows animals to adapt innate behavioural {\textquoteleft}rules of thumb{\textquoteright} to the local environment, improving their prospects for survival and reproduction. Naive Drosophila melanogaster males, for example, court both virgin and mated females, but learn through experience to selectively suppress futile courtship towards females that have already mated. Here we show that courtship learning reflects an enhanced response to the male pheromone cis-vaccenyl acetate (cVA), which is deposited on females during mating and thus distinguishes mated females from virgins. Dissociation experiments suggest a simple learning rule in which unsuccessful courtship enhances sensitivity to cVA. The learning experience can be mimicked by artificial activation of dopaminergic neurons, and we identify a specific class of dopaminergic neuron that is critical for courtship learning. These neurons provide input to the mushroom body (MB) γ lobe, and the DopR1 dopamine receptor is required in MBγ neurons for both natural and artificial courtship learning. Our work thus reveals critical behavioural, cellular and molecular components of the learning rule by which Drosophila adjusts its innate mating strategy according to experience.

}, keywords = {Acetates, Animal, Animals, Brain, Courtship, Dopamine, Dopaminergic Neurons, Drosophila melanogaster, Female, Learning, Male, Mushroom Bodies, Oleic Acids, Pheromones, Presynaptic Terminals, Receptors, Sex Attractants, Sexual Behavior, Synaptic Transmission}, issn = {1476-4687}, doi = {10.1038/nature11345}, author = {Keleman, Krystyna and Vrontou, Eleftheria and Kr{\"u}ttner, Sebastian and Yu, Jai Y and Kurtovic-Kozaric, Amina and Dickson, Barry J} } @article {5319, title = {Dorsorostral snout muscles in the rat subserve coordinated movement for whisking and sniffing.}, journal = {Anatomical Record}, volume = {295}, year = {2012}, month = {2012 Jul}, pages = {1181-91}, abstract = {

Histochemical examination of the dorsorostral quadrant of the rat snout revealed superficial and deep muscles that are involved in whisking, sniffing, and airflow control. The part of M. nasolabialis profundus that acts as an intrinsic (follicular) muscle to facilitate protraction and translation of the vibrissae is described. An intraturbinate and selected rostral-most nasal muscles that can influence major routs of inspiratory airflow and rhinarial touch through their control of nostril configuration, atrioturbinate and rhinarium position, were revealed.

}, issn = {1932-8494}, doi = {10.1002/ar.22501}, author = {Haidarliu, Sebastian and Golomb, David and Kleinfeld, David and Ahissar, Ehud} } @article {6856, title = {Drosophila CPEB Orb2A mediates memory independent of Its RNA-binding domain.}, journal = {Neuron}, volume = {76}, year = {2012}, month = {2012 Oct 18}, pages = {383-95}, abstract = {

Long-term memory and synaptic plasticity are thought to require the synthesis of new proteins at activated synapses. The CPEB family of RNA binding proteins, including Drosophila Orb2, has been implicated in this process. The precise mechanism by which these molecules regulate memory formation is however poorly understood. We used gene targeting and site-specific transgenesis to specifically modify the endogenous orb2 gene in order to investigate its role in long-term memory formation. We show that the Orb2A and Orb2B isoforms, while both essential, have distinct functions in memory formation. These two isoforms have common glutamine-rich and RNA-binding domains, yet Orb2A uniquely requires the former and Orb2B the latter. We further show that Orb2A induces Orb2 complexes in a manner dependent upon both its glutamine-rich region and neuronal activity. We propose that Orb2B acts as a conventional CPEB to regulate transport and/or translation of specific mRNAs, whereas Orb2A acts in an unconventional manner to form stable Orb2 complexes that are essential for memory to persist.

}, keywords = {Animals, Biogenic Amines, Brain, Cell Line, Chromatography, Courtship, Developmental, Drosophila, Drosophila Proteins, Embryo, Gene Expression Regulation, Genetically Modified, Genotype, Green Fluorescent Proteins, High Pressure Liquid, Immunoelectron, Immunoprecipitation, Larva, Learning, Male, Mass Spectrometry, Memory, Messenger, Microscopy, Mitogen-Activated Protein Kinases, mRNA Cleavage and Polyadenylation Factors, Mushroom Bodies, Mutation, Nonmammalian, Protein Isoforms, Protein Structure, RNA, RNA-Binding Proteins, Tertiary, Transcription Factors}, issn = {1097-4199}, doi = {10.1016/j.neuron.2012.08.028}, author = {Kr{\"u}ttner, Sebastian and Stepien, Barbara and Noordermeer, Jasprina N and Mommaas, Mieke A and Mechtler, Karl and Dickson, Barry J and Keleman, Krystyna} } @article {4522, title = {Drosophila melanogaster as a model to study drug addiction.}, journal = {Human Genetics}, volume = {131}, year = {2012}, month = {2012 Feb 17}, pages = {959-75}, abstract = {

Animal studies have been instrumental in providing knowledge about the molecular and neural mechanisms underlying drug addiction. Recently, the fruit fly Drosophila melanogaster has become a valuable system to model not only the acute stimulating and sedating effects of drugs but also their more complex rewarding properties. In this review, we describe the advantages of using the fly to study drug-related behavior, provide a brief overview of the behavioral assays used, and review the molecular mechanisms and neural circuits underlying drug-induced behavior in flies. Many of these mechanisms have been validated in mammals, suggesting that the fly is a useful model to understand the mechanisms underlying addiction.

}, issn = {1432-1203}, doi = {10.1007/s00439-012-1146-6}, author = {Kaun, Karla R and Devineni, Anita V and Heberlein, Ulrike} } @article {4596, title = {Excitation spectra and brightness optimization of two-photon excited probes.}, journal = {Biophysical Journal}, volume = {102}, year = {2012}, month = {2012 Feb 22}, pages = {934-44}, abstract = {

Two-photon probe excitation data are commonly presented as absorption cross section or molecular brightness (the detected fluorescence rate per molecule). We report two-photon molecular brightness spectra for a diverse set of organic and genetically encoded probes with an automated spectroscopic system based on fluorescence correlation spectroscopy. The two-photon action cross section can be extracted from molecular brightness measurements at low excitation intensities, while peak molecular brightness (the maximum molecular brightness with increasing excitation intensity) is measured at higher intensities at which probe photophysical effects become significant. The spectral shape of these two parameters was similar across all dye families tested. Peak molecular brightness spectra, which can be obtained rapidly and with reduced experimental complexity, can thus serve as a first-order approximation to cross-section spectra in determining optimal wavelengths for two-photon excitation, while providing additional information pertaining to probe photostability. The data shown should assist in probe choice and experimental design for multiphoton microscopy studies. Further, we show that, by the addition of a passive pulse splitter, nonlinear bleaching can be reduced-resulting in an enhancement of the fluorescence signal in fluorescence correlation spectroscopy by a factor of two. This increase in fluorescence signal, together with the observed resemblance of action cross section and peak brightness spectra, suggests higher-order photobleaching pathways for two-photon excitation.

}, issn = {1542-0086}, doi = {10.1016/j.bpj.2011.12.056}, author = {M{\"u}tze, J{\"o}rg and Iyer, Vijay and Macklin, John J and Colonell, Jennifer and Karsh, Bill and Petr{\'a}\v sek, Zden\v ek and Schwille, Petra and Looger, Loren L and Lavis, Luke D and Harris, Timothy D} } @article {5634, title = {Fiji: an open-source platform for biological-image analysis.}, journal = {Nature Methods}, volume = {9}, year = {2012}, month = {2012 Jul}, pages = {676-82}, abstract = {

Fiji is a distribution of the popular open-source software ImageJ focused on biological-image analysis. Fiji uses modern software engineering practices to combine powerful software libraries with a broad range of scripting languages to enable rapid prototyping of image-processing algorithms. Fiji facilitates the transformation of new algorithms into ImageJ plugins that can be shared with end users through an integrated update system. We propose Fiji as a platform for productive collaboration between computer science and biology research communities.

}, keywords = {Algorithms, Animals, Brain, Computational Biology, Computer-Assisted, Drosophila melanogaster, Image Enhancement, Image Processing, Imaging, Information Dissemination, Software, Software Design, Three-Dimensional}, issn = {1548-7105}, doi = {10.1038/nmeth.2019}, author = {Schindelin, Johannes and Arganda-Carreras, Ignacio and Frise, Erwin and Kaynig, Verena and Longair, Mark and Pietzsch, Tobias and Preibisch, Stephan and Rueden, Curtis and Saalfeld, Stephan and Schmid, Benjamin and Tinevez, Jean-Yves and White, Daniel James and Hartenstein, Volker and Eliceiri, Kevin and Tomancak, Pavel and Cardona, Albert} } @article {68815, title = {Generation of Multiple Classes of V0 Neurons in Zebrafish Spinal Cord: Progenitor Heterogeneity and Temporal Control of Neuronal Diversity}, journal = {Journal of Neuroscience}, volume = {32}, year = {2012}, month = {02/2012}, pages = {1771{\textendash}1783}, abstract = {

The developing spinal cord is subdivided into distinct progenitor domains, each of which gives rise to different types of neurons. However, the developmental mechanisms responsible for generating neuronal diversity within a domain are not well understood. Here, we have studied zebrafish V0 neurons, those that derive from the p0 progenitor domain, to address this question. We find that all V0 neurons have commissural axons, but they can be divided into excitatory and inhibitory classes. V0 excitatory neurons (V0-e) can be further categorized into three groups based on their axonal trajectories; V0-eA (ascending), V0-eB (bifurcating), and V0-eD (descending) neurons. By using time-lapse imaging of p0 progenitors and their progeny, we show that inhibitory and excitatory neurons are produced from different progenitors. We also demonstrate that V0-eA neurons are produced from distinct progenitors, while V0-eB and V0-eD neurons are produced from common progenitors. We then use birth-date analysis to reveal that V0-eA, V0-eB, and V0-eD neurons arise in this order. By perturbing Notch signaling and accelerating neuronal differentiation, we predictably alter the generation of early born V0-e neurons at the expense of later born ones. These results suggest that multiple types of V0 neurons are produced by two distinct mechanisms; from heterogeneous p0 progenitors and from the same p0 progenitor, but in a time-dependent manner.

}, issn = {0270-6474}, doi = {10.1523/JNEUROSCI.5500-11.2012}, url = {https://www.jneurosci.org/content/32/5/1771}, author = {Chie Satou and Yukiko Kimura and Shin-ichi Higashijima} } @article {4590, title = {Hierarchical deployment of factors regulating temporal fate in a diverse neuronal lineage of the Drosophila central brain.}, journal = {Neuron}, volume = {73}, year = {2012}, month = {2012 Feb 23}, pages = {677-84}, abstract = {

The anterodorsal projection neuron lineage of Drosophila melanogaster produces 40 neuronal types in a stereotypic order. Here we take advantage of this complete lineage sequence to examine the role of known temporal fating factors, including Chinmo and the Hb/Kr/Pdm/Cas transcriptional cascade, within this diverse central brain lineage. Kr mutation affects the temporal fate of the neuroblast (NB) itself, causing a single fate to be skipped, whereas Chinmo null only elicits fate transformation of NB progeny without altering cell counts. Notably, Chinmo operates in two separate windows to prevent fate transformation (into the subsequent Chinmo-indenpendent fate) within each window. By contrast, Hb/Pdm/Cas play no detectable role, indicating that Kr either acts outside of the cascade identified in the ventral nerve cord or that redundancy exists at the level of fating factors. Therefore, hierarchical fating mechanisms operate within the lineage to generate neuronal diversity in an unprecedented fashion.

}, issn = {1097-4199}, doi = {10.1016/j.neuron.2011.12.018}, author = {Kao, Chih-Fei and Yu, Hung-Hsiang and He, Yisheng and Kao, Jui-Chun and Lee, Tzumin} } @article {6720, title = {Hippocampal pyramidal neurons comprise two distinct cell types that are countermodulated by metabotropic receptors.}, journal = {Neuron}, volume = {76}, year = {2012}, month = {2012 Nov 21}, pages = {776-89}, abstract = {

Relating the function of neuronal cell types to information processing and behavior is a central goal of neuroscience. In the hippocampus, pyramidal cells in CA1 and the subiculum process sensory and motor cues to form a cognitive map encoding spatial, contextual, and emotional information, which they transmit throughout the brain. Do these cells constitute a single class or are there multiple cell types with specialized functions? Using unbiased cluster analysis, we show that there are two morphologically and electrophysiologically distinct principal cell types that carry hippocampal output. We show further that these two cell types are inversely modulated by the synergistic action of glutamate and acetylcholine acting on metabotropic receptors that are central to hippocampal function. Combined with prior connectivity studies, our results support a model of hippocampal processing in which the two pyramidal cell types are predominantly segregated into two parallel pathways that process distinct modalities of information.

}, keywords = {3-dione, 6-Cyano-7-nitroquinoxaline-2, Analysis of Variance, Animals, Benzylamines, Biophysics, CA1 Region, Electric Stimulation, Excitatory Amino Acid Antagonists, Excitatory Amino Acid Transporter 3, GABA Antagonists, Glutamate Decarboxylase, Hippocampal, Lysine, Male, Membrane Potentials, Metabotropic Glutamate, Neurons, Newborn, Patch-Clamp Techniques, Phosphinic Acids, Pyridazines, Rats, Receptors, Time Factors}, issn = {1097-4199}, doi = {10.1016/j.neuron.2012.09.036}, author = {Graves, Austin R and Moore, Shannon J and Bloss, Erik B and Mensh, Brett D and Kath, William L and Spruston, Nelson} } @article {46726, title = {IGF2BP1 promotes cell migration by regulating MK5 and PTEN signaling.}, journal = {Genes \& Development}, volume = {26}, year = {2012}, month = {2012 Jan 15}, pages = {176-89}, abstract = {

In primary neurons, the oncofetal RNA-binding protein IGF2BP1 (IGF2 mRNA-binding protein 1) controls spatially restricted β-actin (ACTB) mRNA translation and modulates growth cone guidance. In cultured tumor-derived cells, IGF2BP1 was shown to regulate the formation of lamellipodia and invadopodia. However, how and via which target mRNAs IGF2BP1 controls the motility of tumor-derived cells has remained elusive. In this study, we reveal that IGF2BP1 promotes the velocity and directionality of tumor-derived cell migration by determining the cytoplasmic fate of two novel target mRNAs: MAPK4 and PTEN. Inhibition of MAPK4 mRNA translation by IGF2BP1 antagonizes MK5 activation and prevents phosphorylation of HSP27, which sequesters actin monomers available for F-actin polymerization. Consequently, HSP27-ACTB association is reduced, mobilizing cellular G-actin for polymerization in order to promote the velocity of cell migration. At the same time, stabilization of the PTEN mRNA by IGF2BP1 enhances PTEN expression and antagonizes PIP(3)-directed signaling. This enforces the directionality of cell migration in a RAC1-dependent manner by preventing additional lamellipodia from forming and sustaining cell polarization intrinsically. IGF2BP1 thus promotes the velocity and persistence of tumor cell migration by controlling the expression of signaling proteins. This fine-tunes and connects intracellular signaling networks in order to enhance actin dynamics and cell polarization.

}, keywords = {Actins, Cell Line, Tumor, Cell Movement, Cell Polarity, Enzyme Activation, Extracellular Signal-Regulated MAP Kinases, HSP27 Heat-Shock Proteins, Humans, Intracellular Signaling Peptides and Proteins, Phosphorylation, Protein-Serine-Threonine Kinases, PTEN Phosphohydrolase, RNA, Messenger, Signal Transduction}, issn = {1549-5477}, doi = {10.1101/gad.177642.111}, author = {St{\"o}hr, Nadine and K{\"o}hn, Marcel and Lederer, Marcell and Glass, Markus and Reinke, Claudia and Singer, Robert H and H{\"u}ttelmaier, Stefan} } @article {4595, title = {An improved Greengenes taxonomy with explicit ranks for ecological and evolutionary analyses of bacteria and archaea.}, journal = {The ISME Journal}, volume = {6}, year = {2012}, month = {2012 Mar}, pages = {610-8}, abstract = {

Reference phylogenies are crucial for providing a taxonomic framework for interpretation of marker gene and metagenomic surveys, which continue to reveal novel species at a remarkable rate. Greengenes is a dedicated full-length 16S rRNA gene database that provides users with a curated taxonomy based on de novo tree inference. We developed a {\textquoteright}taxonomy to tree{\textquoteright} approach for transferring group names from an existing taxonomy to a tree topology, and used it to apply the Greengenes, National Center for Biotechnology Information (NCBI) and cyanoDB (Cyanobacteria only) taxonomies to a de novo tree comprising 408 315 sequences. We also incorporated explicit rank information provided by the NCBI taxonomy to group names (by prefixing rank designations) for better user orientation and classification consistency. The resulting merged taxonomy improved the classification of 75\% of the sequences by one or more ranks relative to the original NCBI taxonomy with the most pronounced improvements occurring in under-classified environmental sequences. We also assessed candidate phyla (divisions) currently defined by NCBI and present recommendations for consolidation of 34 redundantly named groups. All intermediate results from the pipeline, which includes tree inference, jackknifing and transfer of a donor taxonomy to a recipient tree (tax2tree) are available for download. The improved Greengenes taxonomy should provide important infrastructure for a wide range of megasequencing projects studying ecosystems on scales ranging from our own bodies (the Human Microbiome Project) to the entire planet (the Earth Microbiome Project). The implementation of the software can be obtained from http://sourceforge.net/projects/tax2tree/.

}, issn = {1751-7370}, doi = {10.1038/ismej.2011.139}, author = {McDonald, Daniel and Price, Morgan N and Goodrich, Julia and Nawrocki, Eric P and Desantis, Todd Z and Probst, Alexander and Andersen, Gary L and Knight, Rob and Hugenholtz, Philip} } @proceedings {45795, title = {Investigation of the role of Aubergine RNA-binding proteins in the reproductive plasticity of the pea aphid, Acyrthosiphon pisum}, volume = {52}, year = {2012}, month = {01/2012}, pages = {E202-E202}, publisher = {OXFORD UNIV PRESS INC}, abstract = {

Environmental changes can elicit alterations in the form, behavior and/or physiology of all species, and this developmental response to environment is known as phenotypic plasticity. Despite its ubiquity, the molecular basis for phenotypic plasticity is not fully understood. The pea aphid, Acyrthosiphon pisum, serves as a model for an extreme form of phenotypic plasticity, known as polyphenism. Changes in photoperiod stimulate a switch in female aphid reproductive mode from asexual to sexual reproduction over the course of one generation without changes in genotype. This reproductive polyphenism results in female aphids with ovaries of one of two types: sexual ovaries (producing haploid oocytes via meiosis), or asexual ovaries (producing identical diploid aphid clones via parthenogenesis). To better understand how aphid ovaries could produce different outputs, we surveyed the transcriptomes of sexual and asexual ovaries using RNA-seq. Among genes that exhibited greater than two-fold differences in gene expression between sexual and asexual ovaries, we identified several aubergine paralogs, which encode for germline-specific members of the Argonaute small RNA-binding protein family. The A. pisum genome contains eight aubergine paralogs and at least two piwi paralogs. We are currently comparing the expression patterns of these aphid aubergine paralogs between asexual and sexual aphid ovaries. Aubergine proteins in other species are thought to help suppress the activity of transposable elements, which are found in high quantities throughout the A. pisum genome. Together, these experiments will help elucidate a potential relationship between aubergine paralogs and aphid reproductive plasticity.

}, url = {http://sicb.org/meetings/2012/schedule/abstractdetails.php?id=959}, author = {A Abdelhady and R Cortes and S Musumeci and D Srinivasan and S Shigenobu and D Stern and S Kobayashi} } @article {46603, title = {JNK pathway activation is controlled by Tao/TAOK3 to modulate ethanol sensitivity.}, journal = {PLoS One}, volume = {7}, year = {2012}, month = {2012 Dec 5}, pages = {e50594}, abstract = {

Neuronal signal transduction by the JNK MAP kinase pathway is altered by a broad array of stimuli including exposure to the widely abused drug ethanol, but the behavioral relevance and the regulation of JNK signaling is unclear. Here we demonstrate that JNK signaling functions downstream of the Sterile20 kinase family gene tao/Taok3 to regulate the behavioral effects of acute ethanol exposure in both the fruit fly Drosophila and mice. In flies tao is required in neurons to promote sensitivity to the locomotor stimulant effects of acute ethanol exposure and to establish specific brain structures. Reduced expression of key JNK pathway genes substantially rescued the structural and behavioral phenotypes of tao mutants. Decreasing and increasing JNK pathway activity resulted in increased and decreased sensitivity to the locomotor stimulant properties of acute ethanol exposure, respectively. Further, JNK expression in a limited pattern of neurons that included brain regions implicated in ethanol responses was sufficient to restore normal behavior. Mice heterozygous for a disrupted allele of the homologous Taok3 gene (Taok3Gt) were resistant to the acute sedative effects of ethanol. JNK activity was constitutively increased in brains of Taok3Gt/+ mice, and acute induction of phospho-JNK in brain tissue by ethanol was occluded in Taok3Gt/+ mice. Finally, acute administration of a JNK inhibitor conferred resistance to the sedative effects of ethanol in wild-type but not Taok3Gt/+ mice. Taken together, these data support a role of a TAO/TAOK3-JNK neuronal signaling pathway in regulating sensitivity to acute ethanol exposure in flies and in mice.

}, keywords = {Animals, Base Sequence, Behavior, Animal, DNA Primers, Drosophila, Drosophila Proteins, Ethanol, Immunohistochemistry, MAP Kinase Kinase 4, Mice, Mice, Inbred C57BL, Mutation, Protein-Serine-Threonine Kinases, Reverse Transcriptase Polymerase Chain Reaction}, issn = {1932-6203}, doi = {10.1371/journal.pone.0050594}, author = {Kapfhamer, David and King, Ian and Zou, Mimi E and Lim, Jana P and Heberlein, Ulrike and Wolf, Fred W} } @article {3836, title = {Light sheet microscopy of living or cleared specimens.}, journal = {Current Opinion in Neurobiology}, volume = {22}, year = {2012}, month = {2012 Feb}, pages = {138-43}, abstract = {

Light sheet microscopy is a versatile imaging technique with a unique combination of capabilities. It provides high imaging speed, high signal-to-noise ratio and low levels of photobleaching and phototoxic effects. These properties are crucial in a wide range of applications in the life sciences, from live imaging of fast dynamic processes in single cells to long-term observation of developmental dynamics in entire large organisms. When combined with tissue clearing methods, light sheet microscopy furthermore allows rapid imaging of large specimens with excellent coverage and high spatial resolution. Even samples up to the size of entire mammalian brains can be efficiently recorded and quantitatively analyzed. Here, we provide an overview of the history of light sheet microscopy, review the development of tissue clearing methods, and discuss recent technical breakthroughs that have the potential to influence the future direction of the field.

}, issn = {1873-6882}, doi = {10.1016/j.conb.2011.08.003}, author = {Keller, Philipp J and Dodt, Hans-Ulrich} } @article {6290, title = {Lineage analysis of Drosophila lateral antennal lobe neurons reveals notch-dependent binary temporal fate decisions.}, journal = {PLoS Biology}, volume = {10}, year = {2012}, month = {2012 Nov 20}, pages = {e1001425}, abstract = {

Binary cell fate decisions allow the production of distinct sister neurons from an intermediate precursor. Neurons are further diversified based on the birth order of intermediate precursors. Here we examined the interplay between binary cell fate and birth-order-dependent temporal fate in the Drosophila lateral antennal lobe (lAL) neuronal lineage. Single-cell mapping of the lAL lineage by twin-spot mosaic analysis with repressible cell markers (ts-MARCM) revealed that projection neurons (PNs) and local interneurons (LNs) are made in pairs through binary fate decisions. Forty-five types of PNs innervating distinct brain regions arise in a stereotyped sequence; however, the PNs with similar morphologies are not necessarily born in a contiguous window. The LNs are morphologically less diverse than the PNs, and the sequential morphogenetic changes in the two pairs occur independently. Sanpodo-dependent Notch activity promotes and patterns the LN fates. By contrast, Notch diversifies PN temporal fates in a Sanpodo-dispensable manner. These pleiotropic Notch actions underlie the differential temporal fate specification of twin neurons produced by common precursors within a lineage, possibly by modulating postmitotic neurons{\textquoteright} responses to Notch-independent transcriptional cascades.

}, keywords = {Animals, Biological Markers, Brain, Cell Division, Cell Lineage, Cellular, Developmental, Drosophila, Drosophila Proteins, Embryo, Gene Expression Regulation, Genes, Immunohistochemistry, Insect, Larva, Mechanotransduction, Nerve Tissue Proteins, Neurogenesis, Neurons, Nonmammalian, Notch, Receptors, Single-Cell Analysis}, issn = {1545-7885}, doi = {10.1371/journal.pbio.1001425}, author = {Lin, Suewei and Kao, Chih-Fei and Yu, Hung-Hsiang and Huang, Yaling and Lee, Tzumin} } @article {46607, title = {Lmo4 in the basolateral complex of the amygdala modulates fear learning.}, journal = {PLoS One}, volume = {7}, year = {2012}, month = {2012 Apr 3}, pages = {e34559}, abstract = {

Pavlovian fear conditioning is an associative learning paradigm in which mice learn to associate a neutral conditioned stimulus with an aversive unconditioned stimulus. In this study, we demonstrate a novel role for the transcriptional regulator Lmo4 in fear learning. LMO4 is predominantly expressed in pyramidal projection neurons of the basolateral complex of the amygdala (BLC). Mice heterozygous for a genetrap insertion in the Lmo4 locus (Lmo4gt/+), which express 50\% less Lmo4 than their wild type (WT) counterparts display enhanced freezing to both the context and the cue in which they received the aversive stimulus. Small-hairpin RNA-mediated knockdown of Lmo4 in the BLC, but not the dentate gyrus region of the hippocampus recapitulated this enhanced conditioning phenotype, suggesting an adult- and brain region-specific role for Lmo4 in fear learning. Immunohistochemical analyses revealed an increase in the number of c-Fos positive puncta in the BLC of Lmo4gt/+ mice in comparison to their WT counterparts after fear conditioning. Lastly, we measured anxiety-like behavior in Lmo4gt/+ mice and in mice with BLC-specific downregulation of Lmo4 using the elevated plus maze, open field, and light/dark box tests. Global or BLC-specific knockdown of Lmo4 did not significantly affect anxiety-like behavior. These results suggest a selective role for LMO4 in the BLC in modulating learned but not unlearned fear.

}, keywords = {Adaptor Proteins, Signal Transducing, Amygdala, Animals, Behavior, Animal, Conditioning (Psychology), Cues, Fear, Gene Knockdown Techniques, LIM Domain Proteins, Male, Mice, Mice, Inbred C57BL, Neurons, Protein Transport}, issn = {1932-6203}, doi = {10.1371/journal.pone.0034559}, author = {Maiya, Rajani and Kharazia, Viktor and Lasek, Amy W and Heberlein, Ulrike} } @article {46048, title = {The mechanisms of repetitive spike generation in an axonless retinal interneuron.}, journal = {Cell Reports}, volume = {1}, year = {2012}, month = {2012 Feb 23}, pages = {155-66}, abstract = {

Several types of retinal interneurons exhibit spikes but lack axons. One such neuron is the AII amacrine cell, in which spikes recorded at the soma exhibit small amplitudes (\<10 mV) and broad time courses (\>5 ms). Here, we used electrophysiological recordings and computational analysis to examine the mechanisms underlying this atypical spiking. We found that somatic spikes likely represent large, brief action potential-like events initiated in a single, electrotonically distal dendritic compartment. In this same compartment, spiking undergoes slow modulation, likely by an M-type K conductance. The structural correlate of this compartment is a thin neurite that extends from the primary dendritic tree: local application of TTX to this neurite, or excision of it, eliminates spiking. Thus, the physiology of the axonless AII is much more complex than would be anticipated from morphological descriptions and somatic recordings; in particular, the AII possesses a single dendritic structure that controls its firing pattern.

}, keywords = {Action Potentials, Amacrine Cells, Animals, Axons, Female, Interneurons, Ion Channel Gating, Mice, Mice, Inbred C57BL, Models, Neurological, Potassium Channels, Retina, Time Factors}, issn = {2211-1247}, doi = {10.1016/j.celrep.2011.12.006}, author = {Cembrowski, Mark S and Logan, Stephen M and Tian, Miao and Jia, Li and Li, Wei and Kath, William L and Riecke, Hermann and Singer, Joshua H} } @article {4487, title = {mGRASP enables mapping mammalian synaptic connectivity with light microscopy.}, journal = {Nature Methods}, volume = {9}, year = {2012}, month = {2012 Jan}, pages = {96-102}, abstract = {

The GFP reconstitution across synaptic partners (GRASP) technique, based on functional complementation between two nonfluorescent GFP fragments, can be used to detect the location of synapses quickly, accurately and with high spatial resolution. The method has been previously applied in the nematode and the fruit fly but requires substantial modification for use in the mammalian brain. We developed mammalian GRASP (mGRASP) by optimizing transmembrane split-GFP carriers for mammalian synapses. Using in silico protein design, we engineered chimeric synaptic mGRASP fragments that were efficiently delivered to synaptic locations and reconstituted GFP fluorescence in vivo. Furthermore, by integrating molecular and cellular approaches with a computational strategy for the three-dimensional reconstruction of neurons, we applied mGRASP to both long-range circuits and local microcircuits in the mouse hippocampus and thalamocortical regions, analyzing synaptic distribution in single neurons and in dendritic compartments.

}, issn = {1548-7105}, doi = {10.1038/nmeth.1784}, author = {Kim, Jinhyun and Zhao, Ting and Petralia, Ronald S and Yu, Yang and Peng, Hanchuan and Myers, Eugene and Magee, Jeffrey C} } @article {5710, title = {Network resets in medial prefrontal cortex mark the onset of behavioral uncertainty.}, journal = {Science}, volume = {338}, year = {2012}, month = {2012 Oct 5}, pages = {135-9}, abstract = {

Regions within the prefrontal cortex are thought to process beliefs about the world, but little is known about the circuit dynamics underlying the formation and modification of these beliefs. Using a task that permits dissociation between the activity encoding an animal{\textquoteright}s internal state and that encoding aspects of behavior, we found that transient increases in the volatility of activity in the rat medial prefrontal cortex accompany periods when an animal{\textquoteright}s belief is modified after an environmental change. Activity across the majority of sampled neurons underwent marked, abrupt, and coordinated changes when prior belief was abandoned in favor of exploration of alternative strategies. These dynamics reflect network switches to a state of instability, which diminishes over the period of exploration as new stable representations are formed.

}, keywords = {Animal, Animals, Behavior, Long-Evans, Male, Nerve Net, Neurons, Prefrontal Cortex, Rats, Rejection (Psychology), Reward, Uncertainty}, issn = {1095-9203}, doi = {10.1126/science.1226518}, author = {Karlsson, Mattias P and Tervo, Dougal G R and Karpova, Alla Y} } @article {4489, title = {Nonlinear structured-illumination microscopy with a photoswitchable protein reveals cellular structures at 50-nm resolution.}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {109}, year = {2012}, month = {2012 Jan 17}, pages = {E135-43}, abstract = {

Using ultralow light intensities that are well suited for investigating biological samples, we demonstrate whole-cell superresolution imaging by nonlinear structured-illumination microscopy. Structured-illumination microscopy can increase the spatial resolution of a wide-field light microscope by a factor of two, with greater resolution extension possible if the emission rate of the sample responds nonlinearly to the illumination intensity. Saturating the fluorophore excited state is one such nonlinear response, and a realization of this idea, saturated structured-illumination microscopy, has achieved approximately 50-nm resolution on dye-filled polystyrene beads. Unfortunately, because saturation requires extremely high light intensities that are likely to accelerate photobleaching and damage even fixed tissue, this implementation is of limited use for studying biological samples. Here, reversible photoswitching of a fluorescent protein provides the required nonlinearity at light intensities six orders of magnitude lower than those needed for saturation. We experimentally demonstrate approximately 40-nm resolution on purified microtubules labeled with the fluorescent photoswitchable protein Dronpa, and we visualize cellular structures by imaging the mammalian nuclear pore and actin cytoskeleton. As a result, nonlinear structured-illumination microscopy is now a biologically compatible superresolution imaging method.

}, issn = {1091-6490}, doi = {10.1073/pnas.1107547108}, author = {Rego, E Hesper and Shao, Lin and Macklin, John J and Winoto, Lukman and Johansson, G{\"o}ran A and Kamps-Hughes, Nicholas and Davidson, Michael W and Gustafsson, Mats G L} } @article {5164, title = {Optimization of a GCaMP calcium indicator for neural activity imaging.}, journal = {The Journal of Neuroscience}, volume = {32}, year = {2012}, month = {2012 Oct 3}, pages = {13819-40}, chapter = {13819}, abstract = {

Genetically encoded calcium indicators (GECIs) are powerful tools for systems neuroscience. Recent efforts in protein engineering have significantly increased the performance of GECIs. The state-of-the art single-wavelength GECI, GCaMP3, has been deployed in a number of model organisms and can reliably detect three or more action potentials in short bursts in several systems in vivo . Through protein structure determination, targeted mutagenesis, high-throughput screening, and a battery of in vitro assays, we have increased the dynamic range of GCaMP3 by severalfold, creating a family of {\textquotedblleft}GCaMP5{\textquotedblright} sensors. We tested GCaMP5s in several systems: cultured neurons and astrocytes, mouse retina, and in vivo in Caenorhabditis chemosensory neurons, Drosophila larval neuromuscular junction and adult antennal lobe, zebrafish retina and tectum, and mouse visual cortex. Signal-to-noise ratio was improved by at least 2- to 3-fold. In the visual cortex, two GCaMP5 variants detected twice as many visual stimulus-responsive cells as GCaMP3. By combining in vivo imaging with electrophysiology we show that GCaMP5 fluorescence provides a more reliable measure of neuronal activity than its predecessor GCaMP3.GCaMP5allows more sensitive detection of neural activity in vivo andmayfind widespread applications for cellular imaging in general.

}, doi = {10.1523/​JNEUROSCI.2601-12.2012}, author = {Akerboom, Jasper and Chen, Tsai-Wen and Wardill, Trevor J. and Marvin, Jonathan S and Mutlu, Sevinc and Carreras Caldero, Nicole and Esposti, Federico and Borghuis, Bart G and Sun, Xiaonan Richard and Gordus, Andrew and Orger, Michael B. and Portugues, Ruben and Engert, Florian and Macklin, John J and Filosa, Alessandro and Aggarwal, Aman and Kerr, Rex and Takagi, Ryousuke and Kracun, Sebastian and Shigetomi, Eiji and Khakh, Baljit S. and Baier, Herwig and Lagnado, Leon and Wang, Samuel S-H and Bargmann, Cornelia and Kimmel, Bruce and Jayaraman, Vivek and Svoboda, Karel and Kim, Douglas S. and Schreiter, Eric R and Looger, Loren L} } @article {6330, title = {Outcome of the first electron microscopy validation task force meeting.}, journal = {Structure}, volume = {20}, year = {2012}, month = {2012 Feb 8}, pages = {205-14}, abstract = {

This Meeting Review describes the proceedings and conclusions from the inaugural meeting of the Electron Microscopy Validation Task Force organized by the Unified Data Resource for 3DEM (http://www.emdatabank.org) and held at Rutgers University in New Brunswick, NJ on September 28 and 29, 2010. At the workshop, a group of scientists involved in collecting electron microscopy data, using the data to determine three-dimensional electron microscopy (3DEM) density maps, and building molecular models into the maps explored how to assess maps, models, and other data that are deposited into the Electron Microscopy Data Bank and Protein Data Bank public data archives. The specific recommendations resulting from the workshop aim to increase the impact of 3DEM in biology and medicine.

}, keywords = {Animals, Databases, Electron, Factual, Guidelines as Topic, Humans, Information Storage and Retrieval, Macromolecular Substances, Microscopy, Models, Molecular, Molecular Conformation, Molecular Sequence Annotation}, issn = {1878-4186}, doi = {10.1016/j.str.2011.12.014}, author = {Henderson, Richard and Sali, Andrej and Baker, Matthew L and Carragher, Bridget and Devkota, Batsal and Downing, Kenneth H and Egelman, Edward H and Feng, Zukang and Frank, Joachim and Grigorieff, Nikolaus and Jiang, Wen and Ludtke, Steven J and Medalia, Ohad and Penczek, Pawel A and Rosenthal, Peter B and Rossmann, Michael G and Schmid, Michael F and Schr{\"o}der, Gunnar F and Steven, Alasdair C and Stokes, David L and Westbrook, John D and Wriggers, Willy and Yang, Huanwang and Young, Jasmine and Berman, Helen M and Chiu, Wah and Kleywegt, Gerard J and Lawson, Catherine L} } @article {5460, title = {Proprioceptive coupling within motor neurons drives C. elegans forward locomotion.}, journal = {Neuron}, volume = {76}, year = {2012}, month = {2012 Nov 21}, pages = {750-61}, abstract = {

Locomotion requires coordinated motor activity throughout an animal{\textquoteright}s body. In both vertebrates and invertebrates, chains of coupled central pattern generators (CPGs) are commonly evoked to explain local rhythmic behaviors. In C.\ elegans, we report that proprioception within the motor circuit is responsible for propagating and coordinating rhythmic undulatory waves from head to tail during forward movement. Proprioceptive coupling between adjacent body regions transduces rhythmic movement initiated near the head into bending waves driven along the body by a chain of reflexes. Using optogenetics and calcium imaging to manipulate and monitor motor circuit activity of moving C.\ elegans held in microfluidic devices, we found that the B-type cholinergic motor neurons transduce the proprioceptive signal. In C.\ elegans, a sensorimotor feedback loop operating within a specific type of motor neuron both drives and organizes body movement.

}, issn = {1097-4199}, doi = {10.1016/j.neuron.2012.08.039}, author = {Wen, Quan and Po, Michelle D. and Hulme, Elizabeth and Chen, Sway and Liu, Xinyu and Kwok, Sen Wai and Gershow, Marc and Leifer, Andrew M. and Butler, Victoria and Fang-Yen, Christopher and Kawano, Taizo and Schafer, William R and Whitesides, George and Wyart, Matthieu and Chklovskii, Dmitri B and Samuel, Aravinthan D. T.} } @article {4898, title = {Quantitative high-speed imaging of entire developing embryos with simultaneous multiview light-sheet microscopy.}, journal = {Nature Methods}, volume = {9}, year = {2012}, month = {2012 Jun 3}, pages = {755-63}, abstract = {

Live imaging of large biological specimens is fundamentally limited by the short optical penetration depth of light microscopes. To maximize physical coverage, we developed the SiMView technology framework for high-speed in vivo imaging, which records multiple views of the specimen simultaneously. SiMView consists of a light-sheet microscope with four synchronized optical arms, real-time electronics for long-term sCMOS-based image acquisition at 175 million voxels per second, and computational modules for high-throughput image registration, segmentation, tracking and real-time management of the terabytes of multiview data recorded per specimen. We developed one-photon and multiphoton SiMView implementations and recorded cellular dynamics in entire Drosophila melanogaster embryos with 30-s temporal resolution throughout development. We furthermore performed high-resolution long-term imaging of the developing nervous system and followed neuroblast cell lineages in vivo. SiMView data sets provide quantitative morphological information even for fast global processes and enable accurate automated cell tracking in the entire early embryo.

High-resolution movies in the Digital Embryo repository
Nature News: "Fruitfly development, cell by cell" by Lauren Gravitz
Nature Methods Technology Feature: "Faster frames, clearer pictures" by Monya Baker
Andor Insight Awards: Life Sciences Winner

}, issn = {1548-7105}, doi = {10.1038/nmeth.2062}, author = {Tomer, Raju and Khairy, Khaled and Amat, Fernando and Keller, Philipp J} } @article {6297, title = {Quantitative semi-automated analysis of morphogenesis with single-cell resolution in complex embryos.}, journal = {Development}, volume = {139}, year = {2012}, month = {2012 Nov}, pages = {4271-9}, abstract = {

A quantitative understanding of tissue morphogenesis requires description of the movements of individual cells in space and over time. In transparent embryos, such as C. elegans, fluorescently labeled nuclei can be imaged in three-dimensional time-lapse (4D) movies and automatically tracked through early cleavage divisions up to \ 350 nuclei. A similar analysis of later stages of C. elegans development has been challenging owing to the increased error rates of automated tracking of large numbers of densely packed nuclei. We present Nucleitracker4D, a freely available software solution for tracking nuclei in complex embryos that integrates automated tracking of nuclei in local searches with manual curation. Using these methods, we have been able to track \>99\% of all nuclei generated in the C. elegans embryo. Our analysis reveals that ventral enclosure of the epidermis is accompanied by complex coordinated migration of the neuronal substrate. We can efficiently track large numbers of migrating nuclei in 4D movies of zebrafish cardiac morphogenesis, suggesting that this approach is generally useful in situations in which the number, packing or dynamics of nuclei present challenges for automated tracking.

}, keywords = {Animals, Caenorhabditis elegans, Cell Differentiation, Cell Division, Cell Movement, Cell Nucleus, Computer-Assisted, Computers, Embryo, Epidermis, Image Processing, Morphogenesis, Nonmammalian, Single-Cell Analysis, Software, Statistics as Topic, Zebrafish}, issn = {1477-9129}, doi = {10.1242/dev.086256}, author = {Giurumescu, Claudiu A and Kang, Sukryool and Planchon, Thomas A and Betzig, Eric and Bloomekatz, Joshua and Yelon, Deborah and Cosman, Pamela and Chisholm, Andrew D} } @article {46730, title = {Regulation of local expression of cell adhesion and motility-related mRNAs in breast cancer cells by IMP1/ZBP1.}, journal = {The Journal of Cell Science}, volume = {125}, year = {2012}, month = {2012 Jan 1}, pages = {81-91}, abstract = {

Metastasis involves tumor cell detachment from the primary tumor, and acquisition of migratory and invasive capabilities. These capabilities are mediated by multiple events, including loss of cell-cell contact, an increase in focal adhesion turnover and failure to maintain a normal cell polarity. We have previously reported that silencing of the expression of the zipcode-binding protein IMP1/ZBP1 in breast tumor patients is associated with metastasis. IMP1/ZBP1 selectively binds to a group of mRNAs that encode important mediators for cell adhesion and motility. Here, we show that in both T47D and MDA231 human breast carcinoma cells IMP1/ZBP1 functions to suppress cell invasion. Binding of ZBP1 to the mRNAs encoding E-cadherin, β-actin, α-actinin and the Arp2/3 complex facilitates localization of the mRNAs, which stabilizes cell-cell connections and focal adhesions. Our studies suggest a novel mechanism through which IMP1/ZBP1 simultaneously regulates the local expression of many cell-motility-related mRNAs to maintain cell adherence and polarity, decrease focal adhesion turnover and maintain a persistent and directional motility.

}, keywords = {Actin-Related Protein 2-3 Complex, Actinin, Actins, Breast Neoplasms, Cadherins, Cell Adhesion, Cell Line, Tumor, Cell Movement, Focal Adhesions, Gene Expression Regulation, Neoplastic, Humans, Neoplasm Invasiveness, Neoplasm Metastasis, RNA, Messenger, RNA, Neoplasm, RNA-Binding Proteins}, issn = {1477-9137}, doi = {10.1242/jcs.086132}, author = {Gu, Wei and Katz, Zachary and Wu, Bin and Park, Hye Yoon and Li, Deling and Lin, Stanley and Wells, Amber L and Singer, Robert H} } @article {4594, title = {Regulation of neuronal input transformations by tunable dendritic inhibition.}, journal = {Nature Neuroscience}, volume = {15}, year = {2012}, month = {2012 Jan 15}, pages = {423-30}, abstract = {

Transforming synaptic input into action potential output is a fundamental function of neurons. The pattern of action potential output from principal cells of the mammalian hippocampus encodes spatial and nonspatial information, but the cellular and circuit mechanisms by which neurons transform their synaptic input into a given output are unknown. Using a combination of optical activation and cell type-specific pharmacogenetic silencing in vitro, we found that dendritic inhibition is the primary regulator of input-output transformations in mouse hippocampal CA1 pyramidal cells, and acts by gating the dendritic electrogenesis driving burst spiking. Dendrite-targeting interneurons are themselves modulated by interneurons targeting pyramidal cell somata, providing a synaptic substrate for tuning pyramidal cell output through interactions in the local inhibitory network. These results provide evidence for a division of labor in cortical circuits, where distinct computational functions are implemented by subtypes of local inhibitory neurons.

}, issn = {1546-1726}, doi = {10.1038/nn.3024}, author = {Lovett-Barron, Matthew and Turi, Gergely F and Kaifosh, Patrick and Lee, Peter H and Bolze, Fr{\'e}d{\'e}ric and Sun, Xiao-Hua and Nicoud, Jean-Fran\c cois and Zemelman, Boris V and Sternson, Scott M and Losonczy, Attila} } @article {6515, title = {A resource for manipulating gene expression and analyzing cis-regulatory modules in the Drosophila CNS.}, journal = {Cell Reports}, volume = {2}, year = {2012}, month = {2012 Oct 25}, pages = {1002-13}, abstract = {

Here, we describe the embryonic central nervous system expression of 5,000 GAL4 lines made using molecularly defined cis-regulatory DNA inserted into a single attP genomic location. We document and annotate the patterns in early embryos when neurogenesis is at its peak, and in older embryos where there is maximal neuronal diversity and the first neural circuits are established. We note expression in other tissues, such as the lateral body wall (muscle, sensory neurons, and trachea) and viscera. Companion papers report on the adult brain and larval imaginal discs, and the integrated data sets\ are available online (http://www.janelia.org/gal4-gen1). This collection of embryonically expressed GAL4 lines will be valuable for determining neuronal morphology and function. The 1,862 lines expressed in small subsets of neurons (\<20/segment) will be especially valuable for characterizing\ interneuronal diversity and function, because although interneurons comprise the majority of all central nervous system neurons, their gene expression profile and function remain virtually unexplored.

}, keywords = {Animals, Central Nervous System, Databases, Drosophila, Drosophila Proteins, Embryo, Factual, Female, Gene Expression, Genes, Genetically Modified, Internet, Male, Nonmammalian, Regulatory Elements, Reporter, Transcription Factors, Transcriptional}, issn = {2211-1247}, doi = {10.1016/j.celrep.2012.09.009}, author = {Manning, Laurina and Heckscher, Ellie S and Purice, Maria D and Roberts, Jourdain and Bennett, Alysha L and Kroll, Jason R and Pollard, Jill L and Strader, Marie E and Lupton, Josh R and Dyukareva, Anna V and Doan, Phuong Nam and Bauer, David M and Wilbur, Allison N and Tanner, Stephanie and Kelly, Jimmy J and Lai, Sen-Lin and Tran, Khoa D and Kohwi, Minoree and Laverty, Todd R and Pearson, Joseph C and Crews, Stephen T and Rubin, Gerald M and Doe, Chris Q} } @article {5480, title = {Sexual deprivation increases ethanol intake in Drosophila.}, journal = {Science}, volume = {335}, year = {2012}, month = {2012 Mar 16}, pages = {1351-5}, abstract = {

The brain{\textquoteright}s reward systems reinforce behaviors required for species survival, including sex, food consumption, and social interaction. Drugs of abuse co-opt these neural pathways, which can lead to addiction. Here, we used Drosophila melanogaster to investigate the relationship between natural and drug rewards. In males, mating increased, whereas sexual deprivation reduced, neuropeptide F (NPF) levels. Activation or inhibition of the NPF system in turn reduced or enhanced ethanol preference. These results thus link sexual experience, NPF system activity, and ethanol consumption. Artificial activation of NPF neurons was in itself rewarding and precluded the ability of ethanol to act as a reward. We propose that activity of the NPF-NPF receptor axis represents the state of the fly reward system and modifies behavior accordingly.

}, keywords = {Acetates, Addictive, Alcohol Drinking, Animal, Animals, Behavior, Cohort Studies, Conditioning (Psychology), Cues, Drosophila melanogaster, Drosophila Proteins, Ethanol, Female, Male, Neurons, Neuropeptide, Neuropeptides, Oleic Acids, Pheromones, Receptors, Reward, Sexual Behavior, TRPC Cation Channels}, issn = {1095-9203}, doi = {10.1126/science.1215932}, author = {Shohat-Ophir, G. and Kaun, K.R. and Azanchi, R. and Mohammed, H and Heberlein, U} } @article {4521, title = {Sexual experience affects ethanol intake in Drosophila through Neuropeptide F.}, journal = {Science}, volume = {335}, year = {2012}, month = {03/2012}, pages = {1351-5}, abstract = {

The brain{\textquoteright}s reward systems reinforce behaviors required for species survival, including sex, food consumption, and social interaction. Drugs of abuse co-opt these neural pathways, which can lead to addiction. Here, we used Drosophila melanogaster to investigate the relationship between natural and drug rewards. In males, mating increased, whereas sexual deprivation reduced, neuropeptide F (NPF) levels. Activation or inhibition of the NPF system in turn reduced or enhanced ethanol preference. These results thus link sexual experience, NPF system activity, and ethanol consumption. Artificial activation of NPF neurons was in itself rewarding and precluded the ability of ethanol to act as a reward. We propose that activity of the NPF{\textendash}NPF receptor axis represents the state of the fly reward system and modifies behavior accordingly.

}, doi = {10.1126/science.1215932 }, author = {Shohat-Ophir, G. and Kaun, K.R. and Azanchi, R. and Heberlein, U} } @article {67279, title = {Simplified models for LHC new physics searches}, journal = {Journal of Physics G: Nuclear and Particle Physics}, volume = {39}, year = {2012}, month = {Jan-10-2012}, pages = {105005}, abstract = {

This document proposes a collection of simplified models relevant to the design of new-physics searches at the Large Hadron Collider (LHC) and the characterization of their results. Both ATLAS and CMS have already presented some results in terms of simplified models, and we encourage them to continue and expand this effort, which supplements both signature-based results and benchmark model interpretations. A simplified model is defined by an effective Lagrangian describing the interactions of a small number of new particles. Simplified models can equally well be described by a small number of masses and cross-sections. These parameters are directly related to collider physics observables, making simplified models a particularly effective framework for evaluating searches and a useful starting point for characterizing positive signals of new physics. This document serves as an official summary of the results from the {\textquoteright}Topologies for Early LHC Searches{\textquoteright} workshop, held at SLAC in September of 2010, the purpose of which was to develop a set of representative models that can be used to cover all relevant phase space in experimental searches. Particular emphasis is placed on searches relevant for the first ~50{\textendash}500 pb-1\ of data and those motivated by supersymmetric models. This note largely summarizes material posted at\ http://lhcnewphysics.org/, which includes simplified model definitions, Monte Carlo material, and supporting contacts within the theory community. We also comment on future developments that may be useful as more data is gathered and analyzed by the experiments.

}, issn = {0954-3899}, doi = {10.1088/0954-3899/39/10/105005}, url = {https://iopscience.iop.org/article/10.1088/0954-3899/39/10/105005}, author = {Alves, Daniele and Arkani-Hamed, Nima and Arora, Sanjay and Bai, Yang and Baumgart, Matthew and Berger, Joshua and Buckley, Matthew and Butler, Bart and Chang, Spencer and Cheng, Hsin-Chia and Cheung, Clifford and Chivukula, R Sekhar and Cho, Won Sang and Cotta, Randy and D{\textquoteright}Alfonso, Mariarosaria and Hedri, Sonia El and Essig, Rouven and Evans, Jared A and Fitzpatrick, Liam and Fox, Patrick and Franceschini, Roberto and Freitas, Ayres and Gainer, James S and Gershtein, Yuri and Gray, Richard and Gregoire, Thomas and Gripaios, Ben and Gunion, Jack and Han, Tao and Haas, Andy and Hansson, Per and Hewett, JoAnne and Hits, Dmitry and Hubisz, Jay and Izaguirre, Eder and Kaplan, Jared and Katz, Emanuel and Kilic, Can and Kim, Hyung-Do and Kitano, Ryuichiro and Koay, Sue Ann and Ko, Pyungwon and Krohn, David and Kuflik, Eric and Lewis, Ian and Lisanti, Mariangela and Liu, Tao and Liu, Zhen and Lu, Ran and Luty, Markus and Meade, Patrick and Morrissey, David and Mrenna, Stephen and Nojiri, Mihoko and Okui, Takemichi and Padhi, Sanjay and Papucci, Michele and Park, Michael and Park, Myeonghun and Perelstein, Maxim and Peskin, Michael and Phalen, Daniel and Rehermann, Keith and Rentala, Vikram and Roy, Tuhin and Ruderman, Joshua T and Sanz, Veronica and Schmaltz, Martin and Schnetzer, Stephen and Schuster, Philip and Schwaller, Pedro and Schwartz, Matthew D and Schwartzman, Ariel and Shao, Jing and Shelton, Jessie and Shih, David and Shu, Jing and Silverstein, Daniel and Simmons, Elizabeth and Somalwar, Sunil and Spannowsky, Michael and Spethmann, Christian and Strassler, Matthew and Su, Shufang and Tait, Tim and Thomas, Brooks and Thomas, Scott and Toro, Natalia and Volansky, Tomer and Wacker, Jay and Waltenberger, Wolfgang and Yavin, Itay and Yu, Felix and Zhao, Yue and Zurek, Kathryn} } @article {5922, title = {Synaptic amplification by dendritic spines enhances input cooperativity.}, journal = {Nature}, volume = {491}, year = {2012}, month = {2012 Nov 22}, pages = {599-602}, abstract = {

Dendritic spines are the nearly ubiquitous site of excitatory synaptic input onto neurons and as such are critically positioned to influence diverse aspects of neuronal signalling. Decades of theoretical studies have proposed that spines may function as highly effective and modifiable chemical and electrical compartments that regulate synaptic efficacy, integration and plasticity. Experimental studies have confirmed activity-dependent structural dynamics and biochemical compartmentalization by spines. However, there is a longstanding debate over the influence of spines on the electrical aspects of synaptic transmission and dendritic operation. Here we measure the amplitude ratio of spine head to parent dendrite voltage across a range of dendritic compartments and calculate the associated spine neck resistance (R(neck)) for spines at apical trunk dendrites in rat hippocampal CA1 pyramidal neurons. We find that R(neck) is large enough (\ 500 MΩ) to amplify substantially the spine head depolarization associated with a unitary synaptic input by \ 1.5- to \ 45-fold, depending on parent dendritic impedance. A morphologically realistic compartmental model capable of reproducing the observed spatial profile of the amplitude ratio indicates that spines provide a consistently high-impedance input structure throughout the dendritic arborization. Finally, we demonstrate that the amplification produced by spines encourages electrical interaction among coactive inputs through an R(neck)-dependent increase in spine head voltage-gated conductance activation. We conclude that the electrical properties of spines promote nonlinear dendritic processing and associated forms of plasticity and storage, thus fundamentally enhancing the computational capabilities of neurons.

}, keywords = {Animals, Dendritic Spines, Electric Impedance, Excitatory Postsynaptic Potentials, Male, Models, Neurological, Pyramidal Cells, Rats, Sprague-Dawley, Synapses, Wistar}, issn = {1476-4687}, doi = {10.1038/nature11554}, author = {Harnett, Mark T and Makara, Judit K and Spruston, Nelson and Kath, William L and Magee, Jeffrey C} } @article {43051, title = {Tandem fluorescent protein timers for in vivo analysis of protein dynamics.}, journal = {Nature Biotechnology}, volume = {30}, year = {2012}, month = {2012 Jun 24}, pages = {708-14}, abstract = {

The functional state of a cell is largely determined by the spatiotemporal organization of its proteome. Technologies exist for measuring particular aspects of protein turnover and localization, but comprehensive analysis of protein dynamics across different scales is possible only by combining several methods. Here we describe tandem fluorescent protein timers (tFTs), fusions of two single-color fluorescent proteins that mature with different kinetics, which we use to analyze protein turnover and mobility in living cells. We fuse tFTs to proteins in yeast to study the longevity, segregation and inheritance of cellular components and the mobility of proteins between subcellular compartments; to measure protein degradation kinetics without the need for time-course measurements; and to conduct high-throughput screens for regulators of protein turnover. Our experiments reveal the stable nature and asymmetric inheritance of nuclear pore complexes and identify regulators of N-end rule{\textendash}mediated protein degradation.

}, keywords = {Green Fluorescent Proteins, High-Throughput Screening Assays, Kinetics, Nuclear Pore, Protein Stability, Proteins, Proteolysis, Saccharomyces cerevisiae, Subcellular Fractions}, issn = {1546-1696}, doi = {10.1038/nbt.2281}, author = {Khmelinskii, Anton and Keller, Philipp J and Bartosik, Anna and Meurer, Matthias and Barry, Joseph D and Mardin, Balca R and Kaufmann, Andreas and Trautmann, Susanne and Wachsmuth, Malte and Pereira, Gislene and Huber, Wolfgang and Schiebel, Elmar and Knop, Michael} } @article {6778, title = {Target-specific output patterns are predicted by the distribution of regular-spiking and bursting pyramidal neurons in the subiculum.}, journal = {Hippocampus}, volume = {22}, year = {2012}, month = {2012 Apr}, pages = {693-706}, abstract = {

Pyramidal neurons in the subiculum project to a variety of cortical and subcortical areas in the brain to convey information processed in the hippocampus. Previous studies have shown that two groups of subicular pyramidal neurons{\textendash}regular-spiking and bursting neurons{\textendash}are distributed in an organized fashion along the proximal-distal axis, with more regular-spiking neurons close to CA1 (proximal) and more bursting neurons close to presubiculum (distal). Anatomically, neurons projecting to some targets are located more proximally along this axis, while others are located more distally. However, the relationship between the firing properties and the targets of subicular pyramidal neurons is not known. To study this relationship, we used in vivo injections of retrogradely transported fluorescent beads into each of nine different regions and conducted whole-cell current-clamp recordings from the bead-containing subicular neurons in acute brain slices. We found that subicular projections to each area were composed of a mixture of regular-spiking and bursting neurons. Neurons projecting to amygdala, lateral entorhinal cortex, nucleus accumbens, and medial/ventral orbitofrontal cortex were located primarily in the proximal subiculum and consisted mostly of regular-spiking neurons (\~{}80\%). By contrast, neurons projecting to medial EC, presubiculum, retrosplenial cortex, and ventromedial hypothalamus were located primarily in the distal subiculum and consisted mostly of bursting neurons (\~{}80\%). Neurons projecting to a thalamic nucleus were located in the middle portion of subiculum, and their probability of bursting was close to 50\%. Thus, the fraction of bursting neurons projecting to each target region was consistent with the known distribution of regular-spiking and bursting neurons along the proximal-distal axis of the subiculum. Variation in the distribution of regular-spiking and bursting neurons suggests that different types of information are conveyed from the subiculum to its various targets.

}, keywords = {Action Potentials, Animals, Biophysical Phenomena, CA1 Region, CA3 Region, Efferent Pathways, Electrophysiological Phenomena, Hippocampal, Hippocampus, Male, Patch-Clamp Techniques, Pyramidal Cells, Rats, Wistar}, issn = {1098-1063}, doi = {10.1002/hipo.20931}, author = {Kim, Yujin and Spruston, Nelson} } @article {6301, title = {A toolbox of Cre-dependent optogenetic transgenic mice for light-induced activation and silencing.}, journal = {Nature Neuroscience}, volume = {15}, year = {2012}, month = {2012 May}, pages = {793-802}, abstract = {

Cell type-specific expression of optogenetic molecules allows temporally precise manipulation of targeted neuronal activity. Here we present a toolbox of four knock-in mouse lines engineered for strong, Cre-dependent expression of channelrhodopsins ChR2-tdTomato and ChR2-EYFP, halorhodopsin eNpHR3.0 and archaerhodopsin Arch-ER2. All four transgenes mediated Cre-dependent, robust activation or silencing of cortical pyramidal neurons in vitro and in vivo upon light stimulation, with ChR2-EYFP and Arch-ER2 demonstrating light sensitivity approaching that of in utero or virally transduced neurons. We further show specific photoactivation of parvalbumin-positive interneurons in behaving ChR2-EYFP reporter mice. The robust, consistent and inducible nature of our ChR2 mice represents a significant advance over previous lines, and the Arch-ER2 and eNpHR3.0 mice are to our knowledge the first demonstration of successful conditional transgenic optogenetic silencing. When combined with the hundreds of available Cre driver lines, this optimized toolbox of reporter mice will enable widespread investigations of neural circuit function with unprecedented reliability and accuracy.

}, keywords = {Action Potentials, Animals, Archaeal Proteins, Brain, Electroporation, Halorhodopsins, Integrases, Light, Luminescent Proteins, Mice, Neurogenesis, Neurons, Optics and Photonics, Proteins, Rhodopsin, Transgenic, Wakefulness}, issn = {1546-1726}, doi = {10.1038/nn.3078}, author = {Madisen, Linda and Mao, Tianyi and Koch, Henner and Zhuo, Jia-min and Berenyi, Antal and Fujisawa, Shigeyoshi and Hsu, Yun-Wei A and Garcia, Alfredo J and Gu, Xuan and Zanella, Sebastien and Kidney, Jolene and Gu, Hong and Mao, Yimei and Hooks, Bryan M and Boyden, Edward S and Buzs{\'a}ki, Gy{\"o}rgy and Ramirez, Jan Marino and Jones, Allan R and Svoboda, Karel and Han, Xue and Turner, Eric E and Zeng, Hongkui} } @article {4598, title = {Triggering a cell shape change by exploiting preexisting actomyosin contractions.}, journal = {Science}, volume = {335}, year = {2012}, month = {2012 Mar 9}, pages = {1232-5}, abstract = {

Apical constriction changes cell shapes, driving critical morphogenetic events, including gastrulation in diverse organisms and neural tube closure in vertebrates. Apical constriction is thought to be triggered by contraction of apical actomyosin networks. We found that apical actomyosin contractions began before cell shape changes in both Caenorhabitis elegans and Drosophila. In C. elegans, actomyosin networks were initially dynamic, contracting and generating cortical tension without substantial shrinking of apical surfaces. Apical cell-cell contact zones and actomyosin only later moved increasingly in concert, with no detectable change in actomyosin dynamics or cortical tension. Thus, apical constriction appears to be triggered not by a change in cortical tension, but by dynamic linking of apical cell-cell contact zones to an already contractile apical cortex.

}, issn = {1095-9203}, doi = {10.1126/science.1217869}, author = {Roh-Johnson, Minna and Shemer, Gidi and Higgins, Christopher D and McClellan, Joseph H and Werts, Adam D and Tulu, U Serdar and Gao, Liang and Betzig, Eric and Kiehart, Daniel P and Goldstein, Bob} } @article {4486, title = {Use of a Drosophila genome-wide conserved sequence database to identify functionally related cis-regulatory enhancers.}, journal = {Developmental Dynamics: An Official Publication of the American Association of Anatomists}, volume = {241}, year = {2012}, month = {2012 Jan}, pages = {169-89}, abstract = {

Phylogenetic footprinting has revealed that cis-regulatory enhancers consist of conserved DNA sequence clusters (CSCs). Currently, there is no systematic approach for enhancer discovery and analysis that takes full-advantage of the sequence information within enhancer CSCs.

}, issn = {1097-0177}, doi = {10.1002/dvdy.22728}, author = {Brody, Thomas and Yavatkar, Amarendra S and Kuzin, Alexander and Kundu, Mukta and Tyson, Leonard J and Ross, Jermaine and Lin, Tzu-Yang and Lee, Chi-Hon and Awasaki, Takeshi and Lee, Tzumin and Odenwald, Ward F} } @article {46721, title = {β-Actin mRNA compartmentalization enhances focal adhesion stability and directs cell migration.}, journal = {Genes \& Development}, volume = {26}, year = {2012}, month = {2012 Sep 1}, pages = {1885-90}, abstract = {

Directed cell motility is at the basis of biological phenomena such as development, wound healing, and metastasis. It has been shown that substrate attachments mediate motility by coupling the cell{\textquoteright}s cytoskeleton with force generation. However, it has been unclear how the persistence of cell directionality is facilitated. We show that mRNA localization plays an important role in this process, but the mechanism of action is still unknown. In this study, we show that the zipcode-binding protein 1 transports β-actin mRNA to the focal adhesion compartment, where it dwells for minutes, suggesting a means for associating its localization with motility through the formation of stable connections between adhesions and newly synthesized actin filaments. In order to demonstrate this, we developed an approach for assessing the functional consequences of β-actin mRNA and protein localization by tethering the mRNA to a specific location-in this case, the focal adhesion complex. This approach will have a significant impact on cell biology because it is now possible to forcibly direct any mRNA and its cognate protein to specific locations in the cell. This will reveal the importance of localized protein translation on various cellular processes.

}, keywords = {Actins, Animals, Cell Compartmentation, Cell Line, Cell Movement, Cells, Cultured, Fibroblasts, Focal Adhesions, Glycoproteins, Mice, Mice, Knockout, RNA, Messenger}, issn = {1549-5477}, doi = {10.1101/gad.190413.112}, author = {Katz, Zachary B and Wells, Amber L and Park, Hye Yoon and Wu, Bin and Shenoy, Shailesh M and Singer, Robert H} } @article {46346, title = {β-secretase cleavage of the fly amyloid precursor protein is required for glial survival.}, journal = {Journal of Neuroscience}, volume = {32}, year = {2012}, month = {2012 Nov 14}, pages = {16181-92}, abstract = {

β-secretase (or BACE1) is the key enzyme in the production of β-amyloid (Aβ), which accumulates in the senile plaques characteristic for Alzheimer{\textquoteright}s disease. Consequently, the lack of BACE1 prevents β-processing of the amyloid precursor protein and Aβ production, which made it a promising target for drug development. However, the loss of BACE1 is also detrimental, leading to myelination defects and altered neuronal activity, functions that have been associated with the cleavage of Neuregulin and a voltage-gated sodium channel subunit. Here we show that the Drosophila ortholog of BACE, dBACE, is required for glial survival. Cell-specific knockdown experiments reveal that this is a non-cell autonomous function, as a knockdown of dBACE in photoreceptor neurons leads to progressive degeneration of glia in their target zone, the lamina. Interestingly, this phenotype is suppressed by the loss of the fly amyloid precursor protein (APPL), whereas a secretion-deficient form of APPL enhances the degeneration. This shows that full-length APPL in neurons promotes the death of neighboring glial cells and that β-processing of APPL is needed to prevent glial death. These results therefore not only demonstrate a novel function for an APP protein in glia, but they also show this function specifically requires regulation by β-cleavage.

}, keywords = {Amyloid beta-Protein Precursor, Amyloid Precursor Protein Secretases, Animals, Axons, Blotting, Western, Cell Death, Cell Movement, Cell Survival, Drosophila, Immunohistochemistry, Microscopy, Electron, Neuregulins, Neuroglia, Polymerase Chain Reaction, Retina, RNA, RNA Interference, Vacuoles}, issn = {1529-2401}, doi = {10.1523/JNEUROSCI.0228-12.2012}, author = {Bolkan, Bonnie J and Triphan, Tilman and Kretzschmar, Doris} } @article {46614, title = {Abnormal behavior in mice mutant for the Disc1 binding partner, Dixdc1.}, journal = {Translational Psychiatry}, volume = {1}, year = {2011}, month = {2011}, pages = {e43}, abstract = {

Disrupted-in-Schizophrenia-1 (DISC1) is a genetic susceptibility locus for major mental illness, including schizophrenia and depression. The Disc1 protein was recently shown to interact with the Wnt signaling protein, DIX domain containing 1 (Dixdc1). Both proteins participate in neural progenitor proliferation dependent on Wnt signaling, and in neural migration independently of Wnt signaling. Interestingly, their effect on neural progenitor proliferation is additive. By analogy to Disc1, mutations in Dixdc1 may lead to abnormal behavior in mice, and to schizophrenia or depression in humans. To explore this hypothesis further, we generated mice mutant at the Dixdc1 locus and analyzed their behavior. Dixdc1(-/-) mice had normal prepulse inhibition, but displayed decreased spontaneous locomotor activity, abnormal behavior in the elevated plus maze and deficits in startle reactivity. Our results suggest that Dixdc1(-/-) mice will be a useful tool to elucidate molecular pathophysiology involving Disc1 in major mental illnesses.

}, keywords = {Animals, Behavior, Animal, Epistasis, Genetic, Humans, Intracellular Signaling Peptides and Proteins, Male, Mice, Mice, 129 Strain, Mice, Mutant Strains, Mice, Transgenic, Microfilament Proteins, Nerve Tissue Proteins}, issn = {2158-3188}, doi = {10.1038/tp.2011.41}, author = {Kivim{\"a}e, S and Martin, P-M and Kapfhamer, D and Ruan, Y and Heberlein, U and Rubenstein, J L R and Cheyette, B N R} } @article {5483, title = {Alk is a transcriptional target of LMO4 and ERα that promotes cocaine sensitization and reward.}, journal = {The Journal of Neuroscience: The Official Journal of the Society for Neuroscience}, volume = {31}, year = {2011}, month = {2011 Oct 5}, pages = {14134-41}, abstract = {

Previously, we showed that the mouse LIM-domain only 4 (Lmo4) gene, which encodes a protein containing two zinc-finger LIM domains that interact with various DNA-binding transcription factors, attenuates behavioral sensitivity to repeated cocaine administration. Here we show that transcription of anaplastic lymphoma kinase (Alk) is repressed by LMO4 in the striatum and that Alk promotes the development of cocaine sensitization and conditioned place preference, a measure of cocaine reward. Since LMO4 is known to interact with estrogen receptor α (ERα) at the promoters of target genes, we investigated whether Alk expression might be controlled by a similar mechanism. We found that LMO4 and ERα are associated with the Alk promoter by chromatin immunoprecipitation and that Alk is an estrogen-responsive gene in the striatum. Moreover, we show that ERα knock-out mice exhibit enhanced cocaine sensitization and conditioned place preference and an increase in Alk expression in the nucleus accumbens. These data define a novel regulatory network involved in behavioral responses to cocaine. Interestingly, sex differences in several behavioral responses to cocaine in humans and rodents have been described, and estrogen is thought to mediate some of these differences. Our data suggest that estrogen regulation of Alk may be one mechanism responsible for sexually dimorphic responses to cocaine.

}, keywords = {Adaptor Proteins, Animals, Cells, Cocaine, Cultured, Estrogen Receptor alpha, Gene Targeting, Genetic, HEK293 Cells, Humans, Inbred C57BL, Knockout, LIM Domain Proteins, Male, Mice, Motor Activity, Newborn, Receptor Protein-Tyrosine Kinases, Reward, Signal Transducing, Transcription}, issn = {1529-2401}, doi = {10.1523/JNEUROSCI.3415-11.2011}, author = {Lasek, Amy W and Gesch, Julie and Giorgetti, Francesco and Kharazia, Viktor and Heberlein, Ulrike} } @article {4928, title = {The binding of cholera toxin to the periplasmic vestibule of the type II secretion channel.}, journal = {Channels}, volume = {5}, year = {2011}, month = {2011 May-Jun}, pages = {215-8}, abstract = {

The type II secretion system (T2SS) is a large macromolecular complex spanning the inner and outer membranes of many gram-negative bacteria. The T2SS is responsible for the secretion of virulence factors such as cholera toxin (CT) and heat-labile enterotoxin (LT) from Vibrio cholerae and enterotoxigenic Escherichia coli, respectively. CT and LT are closely related AB5 heterohexamers, composed of one A subunit and a B-pentamer. Both CT and LT are translocated, as folded protein complexes, from the periplasm across the outer membrane through the type II secretion channel, the secretin GspD. We recently published the 19 {\r A} structure of the V. cholerae secretin (VcGspD) in its closed state and showed by SPR measurements that the periplasmic domain of GspD interacts with the B-pentamer complex. Here we extend these studies by characterizing the binding of the cholera toxin B-pentamer to VcGspD using electron microscopy of negatively stained preparations. Our studies indicate that the pentamer is captured within the large periplasmic vestibule of VcGspD. These new results agree well with our previously published studies and are in accord with a piston-driven type II secretion mechanism.

}, keywords = {Bacterial Proteins, Bacterial Secretion Systems, Cholera Toxin, Periplasm, Porins, Protein Folding, Protein Structure, Protein Subunits, Protein Transport, Quaternary, Tertiary, Vibrio cholerae}, issn = {1933-6969}, author = {Reichow, Steve L and Korotkov, Konstantin V and Gonen, Melissa and Sun, Ji and Delarosa, Jaclyn R and Hol, Wim G J and Gonen, Tamir} } @article {47660, title = {Bleaching/blinking assisted localization microscopy for superresolution imaging using standard fluorescent molecules.}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {108}, year = {2011}, month = {2011 Dec 27}, pages = {21081-6}, abstract = {

Superresolution imaging techniques based on the precise localization of single molecules, such as photoactivated localization microscopy (PALM) and stochastic optical reconstruction microscopy (STORM), achieve high resolution by fitting images of single fluorescent molecules with a theoretical Gaussian to localize them with a precision on the order of tens of nanometers. PALM/STORM rely on photoactivated proteins or photoswitching dyes, respectively, which makes them technically challenging. We present a simple and practical way of producing point localization-based superresolution images that does not require photoactivatable or photoswitching probes. Called bleaching/blinking assisted localization microscopy (BaLM), the technique relies on the intrinsic bleaching and blinking behaviors characteristic of all commonly used fluorescent probes. To detect single fluorophores, we simply acquire a stream of fluorescence images. Fluorophore bleach or blink-off events are detected by subtracting from each image of the series the subsequent image. Similarly, blink-on events are detected by subtracting from each frame the previous one. After image subtractions, fluorescence emission signals from single fluorophores are identified and the localizations are determined by fitting the fluorescence intensity distribution with a theoretical Gaussian. We also show that BaLM works with a spectrum of fluorescent molecules in the same sample. Thus, BaLM extends single molecule-based superresolution localization to samples labeled with multiple conventional fluorescent probes.

}, keywords = {Animals, Cercopithecus aethiops, COS Cells, Fluorescent Dyes, Green Fluorescent Proteins, Image Interpretation, Computer-Assisted, Microscopy, Fluorescence, Photobleaching, Software}, issn = {1091-6490}, doi = {10.1073/pnas.1117430109}, author = {Burnette, Dylan T and Sengupta, Prabuddha and Dai, Yuhai and Lippincott-Schwartz, Jennifer and Kachar, Bechara} } @article {2755, title = {A combinatorial semaphorin code instructs the initial steps of sensory circuit assembly in the Drosophila CNS.}, journal = {Neuron}, volume = {70}, year = {2011}, month = {2011 Apr 28}, pages = {281-98}, abstract = {

Longitudinal axon fascicles within the Drosophila embryonic CNS provide connections between body segments and are required for coordinated neural signaling along the anterior-posterior axis. We show here that establishment of select CNS longitudinal tracts and formation of precise mechanosensory afferent innervation to the same CNS region are coordinately regulated by the secreted semaphorins Sema-2a and Sema-2b. Both Sema-2a and Sema-2b utilize the same neuronal receptor, plexin B (PlexB), but serve distinct guidance functions. Localized Sema-2b attraction promotes the initial assembly of a subset of CNS longitudinal projections and subsequent targeting of chordotonal sensory afferent axons to these same longitudinal connectives, whereas broader Sema-2a repulsion serves to prevent aberrant innervation. In the absence of Sema-2b or PlexB, chordotonal afferent connectivity within the CNS is severely disrupted, resulting in specific larval behavioral deficits. These results reveal that distinct semaphorin-mediated guidance functions converge at PlexB and are critical for functional neural circuit assembly.

}, issn = {1097-4199}, doi = {10.1016/j.neuron.2011.02.050}, author = {Wu, Zhuhao and Sweeney, Lora B and Ayoob, Joseph C and Chak, Kayam and Andreone, Benjamin J and Ohyama, Tomoko and Kerr, Rex and Luo, Liqun and Zlatic, Marta and Kolodkin, Alex L} } @article {4070, title = {A computational statistics approach for estimating the spatial range of morphogen gradients.}, journal = {Development}, volume = {138}, year = {2011}, month = {2011 Nov}, pages = {4867-74}, abstract = {

A crucial issue in studies of morphogen gradients relates to their range: the distance over which they can act as direct regulators of cell signaling, gene expression and cell differentiation. To address this, we present a straightforward statistical framework that can be used in multiple developmental systems. We illustrate the developed approach by providing a point estimate and confidence interval for the spatial range of the graded distribution of nuclear Dorsal, a transcription factor that controls the dorsoventral pattern of the Drosophila embryo.

}, issn = {1477-9129}, doi = {10.1242/dev.071571}, author = {Kanodia, Jitendra S and Kim, Yoosik and Tomer, Raju and Khan, Zia and Chung, Kwanghun and Storey, John D and Lu, Hang and Keller, Philipp J and Shvartsman, Stansilas Y} } @article {4071, title = {Digital scanned laser light-sheet fluorescence microscopy (DSLM) of zebrafish and Drosophila embryonic development.}, journal = {Cold Spring Harbor Protocols}, volume = {2011}, year = {2011}, month = {2011 Oct}, pages = {1235-43}, abstract = {

Embryonic development is one of the most complex processes encountered in biology. In vertebrates and higher invertebrates, a single cell transforms into a fully functional organism comprising several tens of thousands of cells, arranged in tissues and organs that perform impressive tasks. In vivo observation of this biological process at high spatiotemporal resolution and over long periods of time is crucial for quantitative developmental biology. Importantly, such recordings must be realized without compromising the physiological development of the specimen. In digital scanned laser light-sheet fluorescence microscopy (DSLM), a specimen is rapidly scanned with a thin sheet of light while fluorescence is recorded perpendicular to the axis of illumination with a camera. Combining light-sheet technology and fast laser scanning, DSLM delivers quantitative data for entire embryos at high spatiotemporal resolution. Compared with confocal and two-photon fluorescence microscopy, DSLM exposes the embryo to at least three orders of magnitude less light energy, but still provides up to 50 times faster imaging speeds and a 10{\textendash}100-fold higher signal-to-noise ratio. By using automated image processing algorithms, DSLM images of embryogenesis can be converted into a digital representation. These digital embryos permit following cells as a function of time, revealing cell fate as well as cell origin. By means of such analyses, developmental building plans of tissues and organs can be determined in a whole-embryo context. This article presents a sample preparation and imaging protocol for studying the development of whole zebrafish and Drosophila embryos using DSLM.

}, issn = {1559-6095}, doi = {10.1101/pdb.prot065839}, author = {Keller, Philipp J and Schmidt, Annette D and Wittbrodt, Jochen and Stelzer, Ernst H K} } @article {46042, title = {Disruption of circadian clocks has ramifications for metabolism, brain, and behavior.}, journal = {Proceedings of the National Academy of Sciences USA}, volume = {108}, year = {2011}, month = {2011 Jan 25}, pages = {1657-62}, abstract = {

Circadian (daily) rhythms are present in almost all plants and animals. In mammals, a brain clock located in the hypothalamic suprachiasmatic nucleus maintains synchrony between environmental light/dark cycles and physiology and behavior. Over the past 100 y, especially with the advent of electric lighting, modern society has resulted in a round-the-clock lifestyle, in which natural connections between rest/activity cycles and environmental light/dark cycles have been degraded or even broken. Instances in which rapid changes to sleep patterns are necessary, such as transmeridian air travel, demonstrate negative effects of acute circadian disruption on physiology and behavior. However, the ramifications of chronic disruption of the circadian clock for mental and physical health are not yet fully understood. By housing mice in 20-h light/dark cycles, incongruous with their endogenous \~{}24-h circadian period, we were able to model the effects of chronic circadian disruption noninvasively. Housing in these conditions results in accelerated weight gain and obesity, as well as changes in metabolic hormones. In the brain, circadian-disrupted mice exhibit a loss of dendritic length and decreased complexity of neurons in the prelimbic prefrontal cortex, a brain region important in executive function and emotional control. Disrupted animals show decreases in cognitive flexibility and changes in emotionality consistent with the changes seen in neural architecture. How our findings translate to humans living and working in chronic circadian disruption is unknown, but we believe that this model can provide a foundation to understand how environmental disruption of circadian rhythms impacts the brain, behavior, and physiology.

}, keywords = {Animals, Behavior, Animal, Biological Clocks, Blood Glucose, Body Temperature, Brain, Circadian Clocks, Circadian Rhythm, Energy Metabolism, Humans, Insulin, Leptin, Male, Maze Learning, Mice, Mice, Inbred C57BL, Motor Activity, Neurons, Photoperiod, Prefrontal Cortex, Weight Gain}, issn = {1091-6490}, doi = {10.1073/pnas.1018375108}, author = {Karatsoreos, Ilia N and Bhagat, Sarah and Bloss, Erik B and Morrison, John H and McEwen, Bruce S} } @article {46050, title = {Distinct functions of kainate receptors in the brain are determined by the auxiliary subunit Neto1.}, journal = {Nature Neuroscience}, volume = {14}, year = {2011}, month = {2011 Jul}, pages = {866-73}, abstract = {

Ionotropic glutamate receptors principally mediate fast excitatory transmission in the brain. Among the three classes of ionotropic glutamate receptors, kainate receptors (KARs) have a unique brain distribution, which has been historically defined by (3)H-radiolabeled kainate binding. Compared with recombinant KARs expressed in heterologous cells, synaptic KARs exhibit characteristically slow rise-time and decay kinetics. However, the mechanisms responsible for these distinct KAR properties remain unclear. We found that both the high-affinity binding pattern in the mouse brain and the channel properties of native KARs are determined by the KAR auxiliary subunit Neto1. Through modulation of agonist binding affinity and off-kinetics of KARs, but not trafficking of KARs, Neto1 determined both the KAR high-affinity binding pattern and the distinctively slow kinetics of postsynaptic KARs. By regulating KAR excitatory postsynaptic current kinetics, Neto1 can control synaptic temporal summation, spike generation and fidelity.

}, keywords = {Animals, Animals, Newborn, Biophysical Phenomena, Biophysics, CA1 Region, Hippocampal, Cell Line, Transformed, Cerebellum, Dizocilpine Maleate, Dose-Response Relationship, Drug, Drug Interactions, Electric Stimulation, Excitatory Amino Acid Agonists, Excitatory Amino Acid Antagonists, Excitatory Postsynaptic Potentials, Gene Expression Regulation, Green Fluorescent Proteins, Guanylate Kinase, Humans, Immunoprecipitation, In Vitro Techniques, Intracellular Signaling Peptides and Proteins, Kainic Acid, Lipoproteins, LDL, Membrane Potentials, Membrane Proteins, Mice, Mice, Knockout, Neurons, Patch-Clamp Techniques, Presynaptic Terminals, Protein Binding, Protein Subunits, Receptors, Kainic Acid, Synaptophysin, Transfection, Tritium}, issn = {1546-1726}, doi = {10.1038/nn.2837}, author = {Straub, Christoph and Hunt, David L and Yamasaki, Miwako and Kim, Kwang S and Watanabe, Masahiko and Castillo, Pablo E and Tomita, Susumu} } @article {4524, title = {A Drosophila model for alcohol reward.}, journal = {Nature Neuroscience}, volume = {14}, year = {2011}, month = {2011 May}, pages = {612-9}, abstract = {

The rewarding properties of drugs contribute to the development of abuse and addiction. We developed a new assay for investigating the motivational properties of ethanol in the genetically tractable model Drosophila melanogaster. Flies learned to associate cues with ethanol intoxication and, although transiently aversive, the experience led to a long-lasting attraction for the ethanol-paired cue, implying that intoxication is rewarding. Temporally blocking transmission in dopaminergic neurons revealed that flies require activation of these neurons to express, but not develop, conditioned preference for ethanol-associated cues. Moreover, flies acquired, consolidated and retrieved these rewarding memories using distinct sets of neurons in the mushroom body. Finally, mutations in scabrous, encoding a fibrinogen-related peptide that regulates Notch signaling, disrupted the formation of memories for ethanol reward. Our results thus establish that Drosophila can be useful for understanding the molecular, genetic and neural mechanisms underling the rewarding properties of ethanol.

}, keywords = {Alcohol-Related Disorders, Alcohols, Analysis of Variance, Animal, Animals, Behavior, Chromatography, Conditioning (Psychology), Disease Models, Dopamine, Drosophila, Drosophila Proteins, Electric Stimulation, Ethanol, Food Preferences, Genetically Modified, Green Fluorescent Proteins, High Pressure Liquid, Male, Maze Learning, Mushroom Bodies, Neurons, Odors, Reward, Synaptic Transmission, Time Factors, Tyrosine 3-Monooxygenase}, issn = {1546-1726}, doi = {10.1038/nn.2805}, author = {Kaun, Karla R and Azanchi, Reza and Maung, Zaw and Hirsh, Jay and Heberlein, Ulrike} } @article {4525, title = {Drosophila tao controls mushroom body development and ethanol-stimulated behavior through par-1.}, journal = {The Journal of Neuroscience: The Official Journal of the Society for Neuroscience}, volume = {31}, year = {2011}, month = {2011 Jan 19}, pages = {1139-48}, abstract = {

In both mammalian and insect models of ethanol-induced behavior, low doses of ethanol stimulate locomotion. However, the mechanisms of the stimulant effects of ethanol on the CNS are mostly unknown. We have identified tao, encoding a serine-threonine kinase of the Ste20 family, as a gene necessary for ethanol-induced locomotor hyperactivity in Drosophila. Mutations in tao also affect behavioral responses to cocaine and nicotine, making flies resistant to the effects of both drugs. We show that tao function is required during the development of the adult nervous system and that tao mutations cause defects in the development of central brain structures, including the mushroom body. Silencing of a subset of mushroom body neurons is sufficient to reduce ethanol-induced hyperactivity, revealing the mushroom body as an important locus mediating the stimulant effects of ethanol. We also show that mutations in par-1 suppress both the mushroom body morphology and behavioral phenotypes of tao mutations and that the phosphorylation state of the microtubule-binding protein Tau can be altered by RNA interference knockdown of tao, suggesting that tao and par-1 act in a pathway to control microtubule dynamics during neural development.

}, keywords = {Analysis of Variance, Animal, Animals, Behavior, Biological, Blotting, Drosophila, Drosophila Proteins, Ethanol, Hyperkinesis, Immunohistochemistry, Metamorphosis, Motor Activity, Mushroom Bodies, Neurons, Protein-Serine-Threonine Kinases, Western}, issn = {1529-2401}, doi = {10.1523/JNEUROSCI.4416-10.2011}, author = {King, Ian and Tsai, Linus T-Y and Pflanz, Ralf and Voigt, Aaron and Lee, Seongsoo and J{\"a}ckle, Herbert and Lu, Bingwei and Heberlein, Ulrike} } @article {46610, title = {An evolutionary conserved role for anaplastic lymphoma kinase in behavioral responses to ethanol.}, journal = {PLoS One}, volume = {6}, year = {2011}, month = {2011 Jul 22}, pages = {e22636}, abstract = {

Anaplastic lymphoma kinase (Alk) is a gene expressed in the nervous system that encodes a receptor tyrosine kinase commonly known for its oncogenic function in various human cancers. We have determined that Alk is associated with altered behavioral responses to ethanol in the fruit fly Drosophila melanogaster, in mice, and in humans. Mutant flies containing transposon insertions in dAlk demonstrate increased resistance to the sedating effect of ethanol. Database analyses revealed that Alk expression levels in the brains of recombinant inbred mice are negatively correlated with ethanol-induced ataxia and ethanol consumption. We therefore tested Alk gene knockout mice and found that they sedate longer in response to high doses of ethanol and consume more ethanol than wild-type mice. Finally, sequencing of human ALK led to the discovery of four polymorphisms associated with a low level of response to ethanol, an intermediate phenotype that is predictive of future alcohol use disorders (AUDs). These results suggest that Alk plays an evolutionary conserved role in ethanol-related behaviors. Moreover, ALK may be a novel candidate gene conferring risk for AUDs as well as a potential target for pharmacological intervention.

}, keywords = {Alcohol Drinking, Alcoholics, Animals, Behavior, Animal, Conscious Sedation, Drosophila melanogaster, Drosophila Proteins, Ethanol, Evolution, Molecular, Female, Gene Expression Regulation, Enzymologic, Homeodomain Proteins, Humans, Male, Mice, Polymorphism, Genetic, Receptor Protein-Tyrosine Kinases}, issn = {1932-6203}, doi = {10.1371/journal.pone.0022636}, author = {Lasek, Amy W and Lim, Jana and Kliethermes, Christopher L and Berger, Karen H and Joslyn, Geoff and Brush, Gerry and Xue, Liquan and Robertson, Margaret and Moore, Monica S and Vranizan, Karen and Morris, Stephan W and Schuckit, Marc A and White, Raymond L and Heberlein, Ulrike} } @article {5280, title = {Fast filtering for RNA homology search.}, journal = {Bioinformatics}, volume = {27}, year = {2011}, month = {2011 Nov 15}, pages = {3102-9}, abstract = {

MOTIVATION: Homology search for RNAs can use secondary structure information to increase power by modeling base pairs, as in covariance models, but the resulting computational costs are high. Typical acceleration strategies rely on at least one filtering stage using sequence-only search. RESULTS: Here we present the multi-segment CYK (MSCYK) filter, which implements a heuristic of ungapped structural alignment for RNA homology search. Compared to gapped alignment, this approximation has lower computation time requirements (O(N$^{4}$) reduced to O(N{\textthreesuperior}), and space requirements (O(N{\textthreesuperior}) reduced to O(N{\texttwosuperior}). A vector-parallel implementation of this method gives up to 100-fold speed-up; vector-parallel implementations of standard gapped alignment at two levels of precision give 3- and 6-fold speed-ups. These approaches are combined to create a filtering pipeline that scores RNA secondary structure at all stages, with results that are synergistic with existing methods.

}, keywords = {Algorithms, Base Sequence, Consensus Sequence, Humans, Nucleic Acid, Nucleic Acid Conformation, RNA, Sequence Alignment, Sequence Analysis, Sequence Homology, Software}, issn = {1367-4811}, doi = {10.1093/bioinformatics/btr545}, author = {Kolbe, Diana L and Eddy, Sean R.} } @article {3359, title = {Functional dissection of the neural substrates for sexual behaviors in Drosophila melanogaster.}, journal = {Genetics}, volume = {189}, year = {2011}, month = {2011 Jun 24}, pages = {195-211}, abstract = {

The male-specific Fruitless proteins (Fru(M)) act to establish the potential for male courtship behavior in Drosophila melanogaster and are expressed in small groups of neurons throughout the nervous system. We screened \ 1000 GAL4 lines, using assays for general courtship, male-male interactions, and male fertility to determine the phenotypes resulting from the GAL4 driven inhibition of Fru(M) expression in subsets of these neurons. A battery of secondary assays showed that the phenotypic classes of GAL4 lines could be divided into subgroups based on additional neurobiological and behavioral criteria. For example, in some lines restoration of Fru(M) expression in cholinergic neurons restores fertility or reduces male-male courtship. Persistent chains of males courting each other in some lines results from males courting both sexes indiscriminately whereas in other lines this phenotype result from apparent habituation deficits. Inhibition of ectopic Fru(M) expression in females, in populations of neurons where Fru(M) is necessary for male fertility, can rescue female infertility. To identify the neurons responsible for some of the observed behavioral alterations, we determined the overlap between the identified GAL4 lines and endogenous Fru(M) expression in lines with fertility defects. The GAL4 lines causing fertility defects generally had widespread overlap with Fru(M) expression in many regions of the nervous system suggesting likely redundant Fru(M)-expressing neuronal pathways capable of conferring male fertility. From associations between the screened behaviors, we propose a functional model for courtship initiation.

}, issn = {1943-2631}, doi = {10.1534/genetics.111.129940}, author = {Meissner, Geoffrey W and Manoli, Devanand S and Chavez, Jose F and Knapp, Jon-Michael and Lin, Tasha L and Stevens, Robin J and Mellert, David J and Tran, David H and Baker, Bruce S} } @article {47658, title = {Fuse or die: Shaping mitochondrial fate during starvation.}, journal = {Communicative \& integrative biology}, volume = {4}, year = {2011}, month = {2011 Nov 1}, pages = {752-4}, abstract = {

Mitochondria continuously change their shape and thereby influence different cellular processes like cell death or development. Recently, we showed that during starvation mitochondria fuse into a highly connected network. The change in mitochondrial shape was dependent on inactivation of the fission protein Drp1, through targeting of two different phosphorylation sites. This rapid inhibition of mitochondrial fission led to unopposed fusion, protecting mitochondria from starvation-induced degradation and enabling the cell to survive nutrient scarce conditions.

}, issn = {1942-0889}, author = {Rambold, Angelika S and Kostelecky, Brenda and Lippincott-Schwartz, Jennifer} } @article {46613, title = {Genome-wide association and genetic functional studies identify autism susceptibility candidate 2 gene (AUTS2) in the regulation of alcohol consumption.}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {108}, year = {2011}, month = {2011 Apr 26}, pages = {7119-24}, abstract = {

Alcohol consumption is a moderately heritable trait, but the genetic basis in humans is largely unknown, despite its clinical and societal importance. We report a genome-wide association study meta-analysis of \~{}2.5 million directly genotyped or imputed SNPs with alcohol consumption (gram per day per kilogram body weight) among 12 population-based samples of European ancestry, comprising 26,316 individuals, with replication genotyping in an additional 21,185 individuals. SNP rs6943555 in autism susceptibility candidate 2 gene (AUTS2) was associated with alcohol consumption at genome-wide significance (P = 4 {\texttimes} 10(-8) to P = 4 {\texttimes} 10(-9)). We found a genotype-specific expression of AUTS2 in 96 human prefrontal cortex samples (P = 0.026) and significant (P \< 0.017) differences in expression of AUTS2 in whole-brain extracts of mice selected for differences in voluntary alcohol consumption. Down-regulation of an AUTS2 homolog caused reduced alcohol sensitivity in Drosophila (P \< 0.001). Our finding of a regulator of alcohol consumption adds knowledge to our understanding of genetic mechanisms influencing alcohol drinking behavior.

}, keywords = {Alcohol Drinking, Animals, Drosophila melanogaster, European Continental Ancestry Group, Female, Gene Expression Regulation, Genome-Wide Association Study, Genotype, Humans, Male, Mice, Nuclear Proteins, Polymorphism, Single Nucleotide, Proteins, Quantitative Trait, Heritable}, issn = {1091-6490}, doi = {10.1073/pnas.1017288108}, author = {Schumann, Gunter and Coin, Lachlan J and Lourdusamy, Anbarasu and Charoen, Pimphen and Berger, Karen H and Stacey, David and Desrivi{\`e}res, Sylvane and Aliev, Fazil A and Khan, Anokhi A and Amin, Najaf and Aulchenko, Yurii S and Bakalkin, Georgy and Bakker, Stephan J and Balkau, Beverley and Beulens, Joline W and Bilbao, Ainhoa and de Boer, Rudolf A and Beury, Delphine and Bots, Michiel L and Breetvelt, Elemi J and Cauchi, St{\'e}phane and Cavalcanti-Proen{\c c}a, Christine and Chambers, John C and Clarke, Toni-Kim and Dahmen, Norbert and de Geus, Eco J and Dick, Danielle and Ducci, Francesca and Easton, Alanna and Edenberg, Howard J and Esko, T{\~o}nu and Esk, T{\~o}nu and Fern{\'a}ndez-Medarde, Alberto and Foroud, Tatiana and Freimer, Nelson B and Girault, Jean-Antoine and Grobbee, Diederick E and Guarrera, Simonetta and Gudbjartsson, Daniel F and Hartikainen, Anna-Liisa and Heath, Andrew C and Hesselbrock, Victor and Hofman, Albert and Hottenga, Jouke-Jan and Isohanni, Matti K and Kaprio, Jaakko and Khaw, Kay-Tee and Kuehnel, Brigitte and Laitinen, Jaana and Lobbens, St{\'e}phane and Luan, Jian{\textquoteright}an and Mangino, Massimo and Maroteaux, Matthieu and Matullo, Giuseppe and McCarthy, Mark I and Mueller, Christian and Navis, Gerjan and Numans, Mattijs E and N{\'u}{\~n}ez, Alejandro and Nyholt, Dale R and Onland-Moret, Charlotte N and Oostra, Ben A and O{\textquoteright}Reilly, Paul F and Palkovits, Miklos and Penninx, Brenda W and Polidoro, Silvia and Pouta, Anneli and Prokopenko, Inga and Ricceri, Fulvio and Santos, Eugenio and Smit, Johannes H and Soranzo, Nicole and Song, Kijoung and Sovio, Ulla and Stumvoll, Michael and Surakk, Ida and Thorgeirsson, Thorgeir E and Thorsteinsdottir, Unnur and Troakes, Claire and Tyrfingsson, Thorarinn and T{\"o}njes, Anke and Uiterwaal, Cuno S and Uitterlinden, Andre G and van der Harst, Pim and van der Schouw, Yvonne T and Staehlin, Oliver and Vogelzangs, Nicole and Vollenweider, Peter and Waeber, Gerard and Wareham, Nicholas J and Waterworth, Dawn M and Whitfield, John B and Wichmann, Erich H and Willemsen, Gonneke and Witteman, Jacqueline C and Yuan, Xin and Zhai, Guangju and Zhao, Jing H and Zhang, Weihua and Martin, Nicholas G and Metspalu, Andres and Doering, Angela and Scott, James and Spector, Tim D and Loos, Ruth J and Boomsma, Dorret I and Mooser, Vincent and Peltonen, Leena and Stefansson, Kari and van Duijn, Cornelia M and Vineis, Paolo and Sommer, Wolfgang H and Kooner, Jaspal S and Spanagel, Rainer and Heberlein, Ulrike A and Jarvelin, Marjo-Riitta and Elliott, Paul} } @article {46425, title = {Glial-derived prodegenerative signaling in the Drosophila neuromuscular system.}, journal = {Neuron}, volume = {72}, year = {2011}, month = {2011 Dec 8}, pages = {760-75}, abstract = {

We provide evidence for a prodegenerative, glial-derived signaling framework in the Drosophila neuromuscular system that includes caspase and mitochondria-dependent signaling. We demonstrate that Drosophila TNF-α (eiger) is expressed in a subset of peripheral glia, and the TNF-α receptor (TNFR), Wengen, is expressed in motoneurons. NMJ degeneration caused by disruption of the spectrin/ankyrin skeleton is suppressed by an eiger mutation or by eiger knockdown within a subset of peripheral glia. Loss of wengen in motoneurons causes a similar suppression providing evidence for glial-derived prodegenerative TNF-α signaling. Neither JNK nor NFκβ is required for prodegenerative signaling. However, we provide evidence for the involvement of both an initiator and effector caspase, Dronc and Dcp-1, and mitochondrial-dependent signaling. Mutations that deplete the axon and nerve terminal of mitochondria suppress degeneration as do mutations in Drosophila Bcl-2 (debcl), a mitochondria-associated protein, and Apaf-1 (dark), which links mitochondrial signaling with caspase activity in other systems.

}, keywords = {Animals, Animals, Genetically Modified, Apoptotic Protease-Activating Factor 1, Caspases, Disease Models, Animal, Drosophila, Drosophila Proteins, Excitatory Postsynaptic Potentials, Fluorescence Recovery After Photobleaching, Green Fluorescent Proteins, Mitochondria, Motor Neuron Disease, Mutation, Nerve Degeneration, Nerve Tissue Proteins, Neuroglia, Neuromuscular Junction, Proto-Oncogene Proteins, Receptors, Tumor Necrosis Factor, RNA Interference, Signal Transduction}, issn = {1097-4199}, doi = {10.1016/j.neuron.2011.09.031}, author = {Keller, Lani C and Cheng, Ling and Locke, Cody J and M{\"u}ller, Martin and Fetter, Richard D and Davis, Graeme W} } @article {47905, title = {Hallmarks of molecular action of microtubule stabilizing agents: effects of epothilone B, ixabepilone, peloruside A, and laulimalide on microtubule conformation.}, journal = {The Journal of biological chemistry}, volume = {286}, year = {2011}, month = {2011 Apr 1}, pages = {11765-78}, abstract = {

Microtubule stabilizing agents (MSAs) comprise a class of drugs that bind to microtubule (MT) polymers and stabilize them against disassembly. Several of these agents are currently in clinical use as anticancer drugs, whereas others are in various stages of development. Nonetheless, there is insufficient knowledge about the molecular modes of their action. Recent studies from our laboratory utilizing hydrogen-deuterium exchange in combination with mass spectrometry (MS) provide new information on the conformational effects of Taxol and discodermolide on microtubules isolated from chicken erythrocytes (CET). We report here a comprehensive analysis of the effects of epothilone B, ixabepilone (IXEMPRA(TM)), laulimalide, and peloruside A on CET conformation. The results of our comparative hydrogen-deuterium exchange MS studies indicate that all MSAs have significant conformational effects on the C-terminal H12 helix of α-tubulin, which is a likely molecular mechanism for the previously observed modulations of MT interactions with microtubule-associated and motor proteins. More importantly, the major mode of MT stabilization by MSAs is the tightening of the longitudinal interactions between two adjacent αβ-tubulin heterodimers at the interdimer interface. In contrast to previous observations reported with bovine brain tubulin, the lateral interactions between the adjacent protofilaments in CET are particularly strongly stabilized by peloruside A and laulimalide, drugs that bind outside the taxane site. This not only highlights the significance of tubulin isotype composition in modulating drug effects on MT conformation and stability but also provides a potential explanation for the synergy observed when combinations of taxane and alternative site binding drugs are used.

}, keywords = {Animals, Bicyclo Compounds, Heterocyclic, Binding Sites, Brain, Brain Chemistry, Cattle, Epothilones, Lactones, Macrolides, Mass Spectrometry, Microtubules, Nerve Tissue Proteins, Protein Structure, Secondary, Tubulin, Tubulin Modulators}, issn = {1083-351X}, doi = {10.1074/jbc.M110.162214}, author = {Khrapunovich-Baine, Marina and Menon, Vilas and Yang, Chia-Ping Huang and Northcote, Peter T and Miller, John H and Angeletti, Ruth Hogue and Fiser, Andras and Horwitz, Susan Band and Xiao, Hui} } @article {6135, title = {High-throughput behavioral analysis in C. elegans.}, journal = {Nature Methods}, volume = {8}, year = {2011}, month = {2011 Jun 5}, pages = {592-8}, abstract = {

We designed a real-time computer vision system, the Multi-Worm Tracker (MWT), which can simultaneously quantify the behavior of dozens of Caenorhabditis elegans on a Petri plate at video rates. We examined three traditional behavioral paradigms using this system: spontaneous movement on food, where the behavior changes over tens of minutes; chemotaxis, where turning events must be detected accurately to determine strategy; and habituation of response to tap, where the response is stochastic and changes over time. In each case, manual analysis or automated single-worm tracking would be tedious and time-consuming, but the MWT system allowed rapid quantification of behavior with minimal human effort. Thus, this system will enable large-scale forward and reverse genetic screens for complex behaviors.

}, keywords = {Animal, Animals, Behavior, Caenorhabditis elegans, Chemotaxis, Computer-Assisted, Image Processing, Movement, Software, Stochastic Processes, Time Factors}, issn = {1548-7105}, doi = {10.1038/nmeth.1625}, author = {Swierczek, Nicholas A and Giles, Andrew C and Rankin, Catharine H and Kerr, Rex A} } @article {3522, title = {In search of the structure of human olfactory space.}, journal = {Frontiers in Systems Neuroscience}, volume = {5}, year = {2011}, month = {2011 Jun 2}, pages = {65}, abstract = {

We analyze the responses of human observers to an ensemble of monomolecular odorants. Each odorant is characterized by a set of 146 perceptual descriptors obtained from a database of odor character profiles. Each odorant is therefore represented by a point in a highly multidimensional sensory space. In this work we study the arrangement of odorants in this perceptual space. We argue that odorants densely sample a two-dimensional curved surface embedded in the multidimensional sensory space. This surface can account for more than half of the variance of the perceptual data. We also show that only 12\% of experimental variance cannot be explained by curved surfaces of substantially small dimensionality (\<10). We suggest that these curved manifolds represent the relevant spaces sampled by the human olfactory system, thereby providing surrogates for olfactory sensory space. For the case of 2D approximation, we relate the two parameters on the curved surface to the physico-chemical parameters of odorant molecules. We show that one of the dimensions is related to eigenvalues of molecules{\textquoteright} connectivity matrix, while the other is correlated with measures of molecules{\textquoteright} polarity. We discuss the behavioral significance of these findings.

}, author = {Koulakov, Alexei and Kolterman, Brian E. and Enikolopov, Armen and Rinberg, Dmitry} } @article {46608, title = {Insulin attenuates the acquisition and expression of ethanol-induced locomotor sensitization in DBA/2J mice.}, journal = {Life Sciences}, volume = {89}, year = {2011}, month = {2011 Dec 19}, pages = {968-74}, abstract = {

AIM: Ethanol-induced locomotor sensitization is a behavioral manifestation of physiological responses to repeated ethanol exposures. While ethanol exerts direct effects on multiple neurotransmitter systems in the brain, ethanol-induced changes in metabolic state, including acute hyperglycemia and inhibition of insulin signaling, also have plausible roles in the expression of ethanol-related behaviors through direct and indirect effects on brain function. The current experiments examined whether insulin administration or the resultant hypoglycemia might attenuate the development of sensitization to the locomotor stimulant effect of ethanol.

MAIN METHODS: Male and female DBA/2J mice received daily injections of 5 or 10 IU/kg insulin before or after a stimulating dose of ethanol and subsequent testing in an automated activity monitor. Blood glucose levels were determined upon the completion of the experiments.

KEY FINDINGS: Insulin injected prior to ethanol blunted the acute stimulant response as well as the acquisition and expression of locomotor sensitization, while insulin given after ethanol did not affect the development of the sensitized response. In a separate experiment, mice given glucose concurrently with insulin developed ethanol-induced locomotor sensitization normally.

SIGNIFICANCE: These experiments suggest that insulin attenuates the development of ethanol-induced locomotor sensitization, and that blood glucose levels can largely account for this effect. Further studies of the role of ethanol-induced metabolic states should provide novel information on the expression of ethanol-related behaviors.

}, keywords = {Animals, Behavior, Animal, Blood Glucose, Central Nervous System Depressants, Dose-Response Relationship, Drug, Ethanol, Female, Glucose, Hypoglycemia, Hypoglycemic Agents, Insulin, Male, Mice, Mice, Inbred DBA, Motor Activity}, issn = {1879-0631}, doi = {10.1016/j.lfs.2011.10.011}, author = {Kliethermes, Christopher L and Heberlein, Ulrike} } @article {4326, title = {Large-scale automated histology in the pursuit of connectomes.}, journal = {The Journal of Neuroscience: The Official Journal of the Society for Neuroscience}, volume = {31}, year = {2011}, month = {2011 Nov 9}, pages = {16125-38}, abstract = {

How does the brain compute? Answering this question necessitates neuronal connectomes, annotated graphs of all synaptic connections within defined brain areas. Further, understanding the energetics of the brain{\textquoteright}s computations requires vascular graphs. The assembly of a connectome requires sensitive hardware tools to measure neuronal and neurovascular features in all three dimensions, as well as software and machine learning for data analysis and visualization. We present the state of the art on the reconstruction of circuits and vasculature that link brain anatomy and function. Analysis at the scale of tens of nanometers yields connections between identified neurons, while analysis at the micrometer scale yields probabilistic rules of connection between neurons and exact vascular connectivity.

}, issn = {1529-2401}, doi = {10.1523/JNEUROSCI.4077-11.2011}, author = {Kleinfeld, David and Bharioke, Arjun and Blinder, Pablo and Bock, Davi D and Briggman, Kevin L and Chklovskii, Dmitri B and Denk, Winfried and Helmstaedter, Moritz and Kaufhold, John P and Lee, Wei-Chung Allen and Meyer, Hanno S and Micheva, Kristina D and Oberlaender, Marcel and Prohaska, Steffen and Reid, R Clay and Smith, Stephen J and Takemura, Shinya and Tsai, Philbert S and Sakmann, Bert} } @article {5283, title = {Long-range neuronal circuits underlying the interaction between sensory and motor cortex.}, journal = {Neuron}, volume = {72}, year = {2011}, month = {2011 Oct 6}, pages = {111-23}, abstract = {

In the rodent vibrissal system, active sensation and sensorimotor integration are mediated in part by connections between barrel cortex and vibrissal motor cortex. Little is known about how these structures interact at the level of neurons. We used Channelrhodopsin-2 (ChR2) expression, combined with anterograde and retrograde labeling, to map connections between barrel cortex and pyramidal neurons in mouse motor cortex. Barrel cortex axons preferentially targeted upper layer (L2/3, L5A) neurons in motor cortex; input to neurons projecting back to barrel cortex was particularly strong. Barrel cortex input to deeper layers (L5B, L6) of motor cortex, including neurons projecting to the brainstem, was weak, despite pronounced geometric overlap of dendrites with axons from barrel cortex. Neurons in different layers received barrel cortex input within stereotyped dendritic domains. The cortico-cortical neurons in superficial layers of motor cortex thus couple motor and sensory signals and might mediate sensorimotor integration and motor learning.

}, keywords = {Animals, Inbred C57BL, Membrane Potentials, Mice, Molecular Imaging, Motor Cortex, Neural Pathways, Neuroanatomical Tract-Tracing Techniques, Neurons, Somatosensory Cortex, Vibrissae}, issn = {1097-4199}, doi = {10.1016/j.neuron.2011.07.029}, author = {Mao, Tianyi and Kusefoglu, Deniz and Hooks, Bryan M and Huber, Daniel and Petreanu, Leopoldo and Svoboda, Karel} } @article {6445, title = {Mapping a sensory-motor network onto a structural and functional ground plan in the hindbrain.}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {108}, year = {2011}, month = {2011 Jan 18}, pages = {1170-5}, abstract = {

The hindbrain of larval zebrafish contains a relatively simple ground plan in which the neurons throughout it are arranged into stripes that represent broad neuronal classes that differ in transmitter identity, morphology, and transcription factor expression. Within the stripes, neurons are stacked continuously according to age as well as structural and functional properties, such as axonal extent, input resistance, and the speed at which they are recruited during movements. Here we address the question of how particular networks among the many different sensory-motor networks in hindbrain arise from such an orderly plan. We use a combination of transgenic lines and pairwise patch recording to identify excitatory and inhibitory interneurons in the hindbrain network for escape behaviors initiated by the Mauthner cell. We map this network onto the ground plan to show that an individual hindbrain network is built by drawing components in predictable ways from the underlying broad patterning of cell types stacked within stripes according to their age and structural and functional properties. Many different specialized hindbrain networks may arise similarly from a simple early patterning.

}, keywords = {Animals, Body Patterning, Brain Mapping, Computer-Assisted, Electrophysiology, Escape Reaction, Genetically Modified, Image Processing, Immunohistochemistry, Interneurons, Larva, Nerve Net, Patch-Clamp Techniques, Psychomotor Performance, Rhombencephalon, Zebrafish}, issn = {1091-6490}, doi = {10.1073/pnas.1012189108}, author = {Koyama, Minoru and Kinkhabwala, Amina and Satou, Chie and Higashijima, Shin-Ichi and Fetcho, Joseph} } @article {5381, title = {Minimum-energy vesicle and cell shapes calculated using spherical harmonics parameterization.}, journal = {Soft Matter}, volume = {7}, year = {2011}, month = {2011 Jan 21}, pages = {2138-43}, abstract = {

An important open question in biophysics is to understand how mechanical forces shape membrane-bounded cells and their organelles. A general solution to this problem is to calculate the bending energy of an arbitrarily shaped membrane surface, which can include both lipids and cytoskeletal proteins, and minimize the energy subject to all mechanical constraints. However, the calculations are difficult to perform, especially for shapes that do not possess axial symmetry. We show that the spherical harmonics parameterization (SHP) provides an analytic description of shape that can be used to quickly and reliably calculate minimum energy shapes of both symmetric and asymmetric surfaces. Using this method, we probe the entire set of shapes predicted by the bilayer couple model, unifying work based on different computational approaches, and providing additional details of the transitions between different shape classes. In addition, we present new minimum-energy morphologies based on non-linear models of membrane skeletal elasticity that closely mimic extreme shapes of red blood cells. The SHP thus provides a versatile shape description that can be used to investigate forces that shape cells.

}, doi = {10.1039/c0sm01193b}, author = {Khairy, Khaled and Howard, Jonathon} } @article {2164, title = {Network anatomy and in vivo physiology of visual cortical neurons.}, journal = {Nature}, volume = {471}, year = {2011}, month = {2011 Mar 10}, pages = {177-82}, abstract = {

In the cerebral cortex, local circuits consist of tens of thousands of neurons, each of which makes thousands of synaptic connections. Perhaps the biggest impediment to understanding these networks is that we have no wiring diagrams of their interconnections. Even if we had a partial or complete wiring diagram, however, understanding the network would also require information about each neuron{\textquoteright}s function. Here we show that the relationship between structure and function can be studied in the cortex with a combination of in vivo physiology and network anatomy. We used two-photon calcium imaging to characterize a functional property{\textendash}the preferred stimulus orientation{\textendash}of a group of neurons in the mouse primary visual cortex. Large-scale electron microscopy of serial thin sections was then used to trace a portion of these neurons{\textquoteright} local network. Consistent with a prediction from recent physiological experiments, inhibitory interneurons received convergent anatomical input from nearby excitatory neurons with a broad range of preferred orientations, although weak biases could not be rejected.

}, keywords = {Animals, Calcium Signaling, Electron, Fluorescence, Interneurons, Male, Mice, Microscopy, Microtomy, Nerve Net, Neural Inhibition, Neurons, Pyramidal Cells, Synapses, Transmission, Visual Cortex}, issn = {1476-4687}, doi = {10.1038/nature09802}, author = {Bock, Davi D and Lee, Wei-Chung Allen and Kerlin, Aaron M and Andermann, Mark L and Hood, Greg and Wetzel, Arthur W and Yurgenson, Sergey and Soucy, Edward R and Kim, Hyon Suk and Reid, R Clay} } @article {3629, title = {N-terminal domain of alphaB-crystallin provides a conformational switch for multimerization and structural heterogeneity.}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {108}, year = {2011}, month = {2011 Apr 19}, pages = {6409-14}, abstract = {

The small heat shock protein (sHSP) αB-crystallin (αB) plays a key role in the cellular protection system against stress. For decades, high-resolution structural studies on heterogeneous sHSPs have been confounded by the polydisperse nature of αB oligomers. We present an atomic-level model of full-length αB as a symmetric 24-subunit multimer based on solid-state NMR, small-angle X-ray scattering (SAXS), and EM data. The model builds on our recently reported structure of the homodimeric α-crystallin domain (ACD) and C-terminal IXI motif in the context of the multimer. A hierarchy of interactions contributes to build multimers of varying sizes: Interactions between two ACDs define a dimer, three dimers connected by their C-terminal regions define a hexameric unit, and variable interactions involving the N-terminal region define higher-order multimers. Within a multimer, N-terminal regions exist in multiple environments, contributing to the heterogeneity observed by NMR. Analysis of SAXS data allows determination of a heterogeneity parameter for this type of system. A mechanism of multimerization into higher-order asymmetric oligomers via the addition of up to six dimeric units to a 24-mer is proposed. The proposed asymmetric multimers explain the homogeneous appearance of αB in negative-stain EM images and the known dynamic exchange of αB subunits. The model of αB provides a structural basis for understanding known disease-associated missense mutations and makes predictions concerning substrate binding and the reported fibrilogenesis of αB.

}, keywords = {alpha-Crystallin B Chain, Animals, Biomolecular, Humans, Missense, Models, Molecular, Mutation, Nuclear Magnetic Resonance, Protein Multimerization, Protein Structure, Quaternary, Tertiary}, issn = {1091-6490}, doi = {10.1073/pnas.1014656108}, author = {Jehle, Stefan and Vollmar, Breanna S and Bardiaux, Benjamin and Dove, Katja K and Rajagopal, Ponni and Gonen, Tamir and Oschkinat, Hartmut and Klevit, Rachel E} } @article {3421, title = {Precise olfactory responses tile the sniff cycle.}, journal = {Nature Neuroscience}, volume = {14}, year = {2011}, month = {2011 Jul 17}, pages = {1039-44}, abstract = {

In terrestrial vertebrates, sniffing controls odorant access to receptors, and therefore sets the timescale of olfactory stimuli. We found that odorants evoked precisely sniff-locked activity in mitral/tufted cells in the olfactory bulb of awake mouse. The trial-to-trial response jitter averaged 12 ms, a precision comparable to other sensory systems. Individual cells expressed odor-specific temporal patterns of activity and, across the population, onset times tiled the duration of the sniff cycle. Responses were more tightly time-locked to the sniff phase than to the time after inhalation onset. The spikes of single neurons carried sufficient information to discriminate odors. In addition, precise locking to sniff phase may facilitate ensemble coding by making synchrony relationships across neurons robust to variation in sniff rate. The temporal specificity of mitral/tufted cell output provides a potentially rich source of information for downstream olfactory areas.

}, issn = {1546-1726}, doi = {10.1038/nn.2877}, author = {Shusterman, Roman and Smear, Matthew C and Koulakov, Alexei A and Rinberg, Dmitry} } @article {5383, title = {Probing tension and dynamics in actomyosin mediated cell shape change.}, journal = {Molecular Biology of the Cell}, volume = {22}, year = {2011}, month = {2011}, author = {Higgins, C. D. and Tulu, U. S. and Gao, L. and Betzig, E and Kiehart, D. P. and Goldstein, B.} } @article {46739, title = {Qualification of a new and precise automatic tool for the assessment of hair diameters in phototrichograms.}, journal = {Skin Research \& Technology}, volume = {17}, year = {2011}, month = {2011 May}, pages = {186-95}, abstract = {

BACKGROUND/PURPOSE: To automatically assess hair growth during cosmetic trials, incorporating parameters such as anagen-to-telogen rate, growth rate, and especially hair diameter.

METHODS: We designed and qualified a new and automatic phototrichogram system based on a high-resolution DSLR camera system (theoretical resolution of 2.557 μm/pixel) and modular macrolens system with fixed focus, combined with a trainable pattern recognition software for automated analysis.

RESULTS: We improved the standard routine for dermatological phototrichogram technique to overcome inaccuracy in thickness measurements due to hair swelling by using an alternative immersion fluid, and increased the effective resolution for hair size and thickness measurement to \<4 μm. After having qualified manual measurements as gold standard for the determination of hair diameters, we established a new trainable automatic picture analysis software able to locate and measure individual hairs in length and thickness even in picture series taken from the same skin area at different time points. Comparisons between manual and automatic measurements of the same hairs showed a \>90\% correlation, and by comparing the automatic results with manual measurements of the same images without individual hair annotation, we could find a correlation of at least 80\%.

CONCLUSION: According to the results and findings generated in this qualification study, we have a reliable tool now that enables us to test cosmetic products for hair treatment in a highly automated way with a sufficient degree of precision and accuracy to detect even small changes in hair diameter during cosmetic trials.

}, keywords = {Alopecia, Dermatology, Equipment Design, Female, Hair, Hair Preparations, Humans, Image Processing, Computer-Assisted, Male, Photography, Software}, issn = {1600-0846}, doi = {10.1111/j.1600-0846.2010.00482.x}, author = {Scheede, Stefan and Herpens, Andreas and Burmeister, Frank and Oltrogge, Bernhard and Saenger, K and Schmidt-Rose, Thomas and Schreiner, Volker and Wenck, Horst and Knieps, Torsten and Berlage, Thomas} } @article {1680, title = {Reconstructing embryonic development.}, journal = {Genesis}, volume = {49}, year = {2011}, month = {2010 Dec 7}, pages = {488-513}, abstract = {

Novel approaches to bio-imaging and automated computational image processing allow the design of truly quantitative studies in developmental biology. Cell behavior, cell fate decisions, cell interactions during tissue morphogenesis, and gene expression dynamics can be analyzed in vivo for entire complex organisms and throughout embryonic development. We review state-of-the-art technology for live imaging, focusing on fluorescence light microscopy techniques for system-level investigations of animal development and discuss computational approaches to image segmentation, cell tracking, automated data annotation, and biophysical modeling. We argue that the substantial increase in data complexity and size requires sophisticated new strategies to data analysis to exploit the enormous potential of these new resources.

}, issn = {1526-968X}, doi = {10.1002/dvg.20698}, author = {Khairy, Khaled and Keller, Philipp J} } @article {1927, title = {Rfam: Wikipedia, clans and the "decimal" release.}, journal = {Nucleic Acids Research}, volume = {39}, year = {2011}, month = {2011 Jan}, pages = {D141-5}, abstract = {

The Rfam database aims to catalogue non-coding RNAs through the use of sequence alignments and statistical profile models known as covariance models. In this contribution, we discuss the pros and cons of using the online encyclopedia, Wikipedia, as a source of community-derived annotation. We discuss the addition of groupings of related RNA families into clans and new developments to the website. Rfam is available on the Web at http://rfam.sanger.ac.uk.

}, issn = {1362-4962}, doi = {10.1093/nar/gkq1129}, author = {Gardner, Paul P and Daub, Jennifer and Tate, John and Moore, Benjamin L and Osuch, Isabelle H and Griffiths-Jones, Sam and Finn, Robert D and Nawrocki, Eric P and Kolbe, Diana L and Eddy, Sean R. and Bateman, Alex} } @article {47671, title = {A role for actin arcs in the leading-edge advance of migrating cells.}, journal = {Nature cell biology}, volume = {13}, year = {2011}, month = {2011 Apr}, pages = {371-81}, abstract = {

Epithelial cell migration requires coordination of two actin modules at the leading edge: one in the lamellipodium and one in the lamella. How the two modules connect mechanistically to regulate directed edge motion is not understood. Using live-cell imaging and photoactivation approaches, we demonstrate that the actin network of the lamellipodium evolves spatio-temporally into the lamella. This occurs during the retraction phase of edge motion, when myosin II redistributes to the lamellipodial actin and condenses it into an actin arc parallel to the edge. The new actin arc moves rearward, slowing down at focal adhesions in the lamella. We propose that net edge extension occurs by nascent focal adhesions advancing the site at which new actin arcs slow down and form the base of the next protrusion event. The actin arc thereby serves as a structural element underlying the temporal and spatial connection between the lamellipodium and the lamella during directed cell motion.

}, keywords = {Actins, Cell Movement, Cytoskeleton, Epithelial Cells, Focal Adhesions, Myosin Type II, Pseudopodia, Recombinant Fusion Proteins}, issn = {1476-4679}, doi = {10.1038/ncb2205}, author = {Burnette, Dylan T and Manley, Suliana and Sengupta, Prabuddha and Sougrat, Rachid and Davidson, Michael W and Kachar, Bechara and Lippincott-Schwartz, Jennifer} } @article {4926, title = {Secretins: dynamic channels for protein transport across membranes.}, journal = {Trends in Biochemical Sciences}, volume = {36}, year = {2011}, month = {2011 Aug}, pages = {433-43}, abstract = {

Secretins form megadalton bacterial-membrane channels in at least four sophisticated multiprotein systems that are crucial for translocation of proteins and assembled fibers across the outer membrane of many species of bacteria. Secretin subunits contain multiple domains, which interact with numerous other proteins, including pilotins, secretion-system partner proteins, and exoproteins. Our understanding of the structure of secretins is rapidly progressing, and it is now recognized that features common to all secretins include a cylindrical arrangement of 12-15 subunits, a large periplasmic vestibule with a wide opening at one end and a periplasmic gate at the other. Secretins might also play a key role in the biogenesis of their cognate secretion systems.

}, keywords = {Bacterial Outer Membrane Proteins, Bacterial Proteins, Carrier Proteins, Gram-Negative Bacteria, Membrane Transport Proteins, Models, Molecular, Protein Binding, Protein Conformation, Protein Structure, Protein Transport, Secondary, Secretin, Tertiary}, issn = {0968-0004}, doi = {10.1016/j.tibs.2011.04.002}, author = {Korotkov, Konstantin V and Gonen, Tamir and Hol, Wim G J} } @article {3684, title = {Shedding light on the system: studying embryonic development with light sheet microscopy.}, journal = {Current Opinion in Genetics and Development}, volume = {21}, year = {2011}, month = {2011 Aug}, pages = {558-65}, abstract = {

Light sheet-based fluorescence microscopy (LSFM) is emerging as a powerful imaging technique for the life sciences. LSFM provides an exceptionally high imaging speed, high signal-to-noise ratio, low level of photo-bleaching and good optical penetration depth. This unique combination of capabilities makes light sheet-based microscopes highly suitable for live imaging applications. There is an outstanding potential in applying this technology to the quantitative study of embryonic development. Here, we provide an overview of the different basic implementations of LSFM, review recent technical advances in the field and highlight applications in the context of embryonic development. We conclude with a discussion of promising future directions.

}, issn = {1879-0380}, doi = {10.1016/j.gde.2011.07.003}, author = {Tomer, Raju and Khairy, Khaled and Keller, Philipp J} } @article {3839, title = {Simultaneous recognition and segmentation of cells: application in C. elegans.}, journal = {Bioinformatics}, volume = {27}, year = {2011}, month = {2011 Aug 17}, pages = {2895-902}, abstract = {

MOTIVATION: Automatic recognition of cell identities is critical for quantitative measurement, targeting, and manipulation of cells of model animals at single-cell resolution. It has been shown to be a powerful tool for studying gene expression and regulation, cell lineages, and cell fates. Existing methods first segment cells, before applying a recognition algorithm in the second step. As a result, the segmentation errors in the first step directly affect and complicate the subsequent cell recognition step. Moreover, in new experimental settings, some of the image features that have been previously relied upon to recognize cells may not be easy to reproduce, due to limitations on the number of color channels available for fluorescent imaging or to the cost of building transgenic animals. An approach that is more accurate and relies on only a single signal channel is clearly desirable. RESULTS: We have developed a new method, called SRS (for Simultaneous Recognition and Segmentation of cells), and applied it to 3D image stacks of the model organism C. elegans. Given a 3D image stack of the animal and a 3D atlas of target cells, SRS is effectively an atlas-guided voxel classification process: cell recognition is realized by smoothly deforming the atlas to best fit the image, where the segmentation is obtained naturally via classification of all image voxels. The method achieved a 97.7\% overall recognition accuracy in recognizing a key class of marker cells, the body wall muscle (BWM) cells, on a data set of 175 C. elegans image stacks containing 14,118 manually curated BWM cells providing the "ground-truth" for accuracy. This result was achieved without any additional fiducial image features. SRS also automatically identified 14 of the image stacks as involving {\textpm}90-degree rotations. With these stacks excluded from the data set, the recognition accuracy rose to 99.1\%. We also show SRS is generally applicable to other cell-types, e.g. intestinal cells. AVAILABILITY: The supplementary movies can be downloaded from our website http://penglab.janelia.org/proj/celegans_seganno. The method has been implemented as a plug-in program within the V3D system (http://penglab.janelia.org/proj/v3d) and will be released in the V3D plugin source code repository.

}, issn = {1367-4811}, doi = {10.1093/bioinformatics/btr480}, author = {Qu, Lei and Long, Fuhui and Liu, Xiao and Kim, Stuart and Myers, Eugene and Peng, Hanchuan} } @article {6779, title = {Slow integration leads to persistent action potential firing in distal axons of coupled interneurons.}, journal = {Nature Neuroscience}, volume = {14}, year = {2011}, month = {2011 Feb}, pages = {200-7}, abstract = {

The conventional view of neurons is that synaptic inputs are integrated on a timescale of milliseconds to seconds in the dendrites, with action potential initiation occurring in the axon initial segment. We found a much slower form of integration that leads to action potential initiation in the distal axon, well beyond the initial segment. In a subset of rodent hippocampal and neocortical interneurons, hundreds of spikes, evoked over minutes, resulted in persistent firing that lasted for a similar duration. Although axonal action potential firing was required to trigger persistent firing, somatic depolarization was not. In paired recordings, persistent firing was not restricted to the stimulated neuron; it could also be produced in the unstimulated cell. Thus, these interneurons can slowly integrate spiking, share the output across a coupled network of axons and respond with persistent firing even in the absence of input to the soma or dendrites.

}, keywords = {Action Potentials, Animals, Axons, Cerebral Cortex, Dendrites, Hippocampus, Interneurons, Mice, Nerve Net, Patch-Clamp Techniques, Synapses, Transgenic}, issn = {1546-1726}, doi = {10.1038/nn.2728}, author = {Sheffield, Mark E J and Best, Tyler K and Mensh, Brett D and Kath, William L and Spruston, Nelson} } @article {4147, title = {Sparse incomplete representations: a potential role of olfactory granule cells.}, journal = {Neuron}, volume = {72}, year = {2011}, month = {2011 Oct 6}, pages = {124-36}, abstract = {

Mitral/tufted cells of the olfactory bulb receive odorant information from receptor neurons and transmit this information to the cortex. Studies in awake behaving animals have found that sustained responses of mitral cells to odorants are rare, suggesting sparse combinatorial representation of the odorants. Careful alignment of mitral cell firing with the phase of the respiration cycle revealed brief transient activity in the larger population of mitral cells, which respond to odorants during a small fraction of the respiration cycle. Responses of these cells are therefore temporally sparse. Here, we propose a mathematical model for the olfactory bulb network that can reproduce both combinatorially and temporally sparse mitral cell codes. We argue that sparse codes emerge as a result of the balance between mitral cells{\textquoteright} excitatory inputs and inhibition provided by the granule cells. Our model suggests functional significance for the dendrodendritic synapses mediating interactions between mitral and granule cells.

}, issn = {1097-4199}, doi = {10.1016/j.neuron.2011.07.031}, author = {Koulakov, Alexei A and Rinberg, Dmitry} } @article {6444, title = {A structural and functional ground plan for neurons in the hindbrain of zebrafish.}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {108}, year = {2011}, month = {2011 Jan 18}, pages = {1164-9}, abstract = {

The vertebrate hindbrain contains various sensory-motor networks controlling movements of the eyes, jaw, head, and body. Here we show that stripes of neurons with shared neurotransmitter phenotype that extend throughout the hindbrain of young zebrafish reflect a broad underlying structural and functional patterning. The neurotransmitter stripes contain cell types with shared gross morphologies and transcription factor markers. Neurons within a stripe are stacked systematically by extent and location of axonal projections, input resistance, and age, and are recruited along the axis of the stripe during behavior. The implication of this pattern is that the many networks in hindbrain are constructed from a series of neuronal components organized into stripes that are ordered from top to bottom according to a neuron{\textquoteright}s age, structural and functional properties, and behavioral roles. This simple organization probably forms a foundation for the construction of the networks underlying the many behaviors produced by the hindbrain.

}, keywords = {Age Factors, Animals, Body Patterning, Calcium, Confocal, Electrophysiology, Genetically Modified, Interneurons, Microscopy, Models, Nerve Net, Neurological, Neurotransmitter Agents, Rhombencephalon, Zebrafish}, issn = {1091-6490}, doi = {10.1073/pnas.1012185108}, author = {Kinkhabwala, Amina and Riley, Michael and Koyama, Minoru and Monen, Joost and Satou, Chie and Kimura, Yukiko and Higashijima, Shin-Ichi and Fetcho, Joseph} } @article {68822, title = {A structural and functional ground plan for neurons in the hindbrain of zebrafish}, journal = {Proceedings of the National Academy of Sciences}, volume = {108}, year = {2011}, month = {01/2011}, pages = {1164-1169}, abstract = {

The vertebrate hindbrain contains various sensory-motor networks controlling movements of the eyes, jaw, head, and body. Here we show that stripes of neurons with shared neurotransmitter phenotype that extend throughout the hindbrain of young zebrafish reflect a broad underlying structural and functional patterning. The neurotransmitter stripes contain cell types with shared gross morphologies and transcription factor markers. Neurons within a stripe are stacked systematically by extent and location of axonal projections, input resistance, and age, and are recruited along the axis of the stripe during behavior. The implication of this pattern is that the many networks in hindbrain are constructed from a series of neuronal components organized into stripes that are ordered from top to bottom according to a neuron{\textquoteright}s age, structural and functional properties, and behavioral roles. This simple organization probably forms a foundation for the construction of the networks underlying the many behaviors produced by the hindbrain.

}, doi = {10.1073/pnas.1012185108}, url = {https://www.pnas.org/doi/abs/10.1073/pnas.1012185108}, author = {Amina Kinkhabwala and Michael Riley and Minoru Koyama and Joost Monen and Chie Satou and Yukiko Kimura and Shin-ichi Higashijima and Joseph Fetcho} } @article {5309, title = {Super-resolution 3D microscopy of live whole cells using structured illumination.}, journal = {Nature Methods}, volume = {8}, year = {2011}, month = {2011 Dec}, pages = {1044-6}, abstract = {

Three-dimensional (3D) structured-illumination microscopy (SIM) can double the lateral and axial resolution of a wide-field fluorescence microscope but has been too slow for live imaging. Here we apply 3D SIM to living samples and record whole cells at up to 5 s per volume for \>50 time points with 120-nm lateral and 360-nm axial resolution. We demonstrate the technique by imaging microtubules in S2 cells and mitochondria in HeLa cells.

}, keywords = {Animals, Cell Line, Cell Survival, Drosophila melanogaster, Fluorescence, Hela Cells, Humans, Imaging, Microscopy, Microtubules, Mitochondria, Three-Dimensional}, issn = {1548-7105}, doi = {10.1038/nmeth.1734}, author = {Shao, Lin and Kner, Peter and Rego, E Hesper and Gustafsson, Mats G L} } @article {4469, title = {Superresolution fluorescence imaging of mitochondrial nucleoids reveals their spatial range, limits, and membrane interaction.}, journal = {Molecular and Cellular Biology}, volume = {31}, year = {2011}, month = {2011 Dec}, pages = {4994-5010}, abstract = {

A fundamental objective in molecular biology is to understand how DNA is organized in concert with various proteins, RNA, and biological membranes. Mitochondria maintain and express their own DNA (mtDNA), which is arranged within structures called nucleoids. Their functions, dimensions, composition, and precise locations relative to other mitochondrial structures are poorly defined. Superresolution fluorescence microscopy techniques that exceed the previous limits of imaging within the small and highly compartmentalized mitochondria have been recently developed. We have improved and employed both two- and three-dimensional applications of photoactivated localization microscopy (PALM and iPALM, respectively) to visualize the core dimensions and relative locations of mitochondrial nucleoids at an unprecedented resolution. PALM reveals that nucleoids differ greatly in size and shape. Three-dimensional volumetric analysis indicates that, on average, the mtDNA within ellipsoidal nucleoids is extraordinarily condensed. Two-color PALM shows that the freely diffusible mitochondrial matrix protein is largely excluded from the nucleoid. In contrast, nucleoids are closely associated with the inner membrane and often appear to be wrapped around cristae or crista-like inner membrane invaginations. Determinations revealing high packing density, separation from the matrix, and tight association with the inner membrane underscore the role of mechanisms that regulate access to mtDNA and that remain largely unknown.

}, keywords = {3T3 Cells, Animals, Confocal, DNA, Fluorescence, Mice, Microscopy, Mitochondria, Mitochondrial, Mitochondrial Proteins, Plasmids, Sequence Alignment, Sequence Analysis}, issn = {1098-5549}, doi = {10.1128/MCB.05694-11}, url = {http://mcb.asm.org/content/31/24/4994.short}, author = {Brown, Timothy A and Tkachuk, Ariana N and Shtengel, Gleb and Kopek, Benjamin G and Bogenhagen, Daniel F and Hess, Harald F and Clayton, David A} } @article {48393, title = {Symmetries in stimulus statistics shape the form of visual motion estimators.}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {108}, year = {2011}, month = {2011 Aug 02}, pages = {12909-14}, abstract = {

The estimation of visual motion has long been studied as a paradigmatic neural computation, and multiple models have been advanced to explain behavioral and neural responses to motion signals. A broad class of models, originating with the Reichardt correlator model, proposes that animals estimate motion by computing a temporal cross-correlation of light intensities from two neighboring points in visual space. These models provide a good description of experimental data in specific contexts but cannot explain motion percepts in stimuli lacking pairwise correlations. Here, we develop a theoretical formalism that can accommodate diverse stimuli and behavioral goals. To achieve this, we treat motion estimation as a problem of Bayesian inference. Pairwise models emerge as one component of the generalized strategy for motion estimation. However, correlation functions beyond second order enable more accurate motion estimation. Prior expectations that are asymmetric with respect to bright and dark contrast use correlations of both even and odd orders, and we show that psychophysical experiments using visual stimuli with symmetric probability distributions for contrast cannot reveal whether the subject uses odd-order correlators for motion estimation. This result highlights a gap in previous experiments, which have largely relied on symmetric contrast distributions. Our theoretical treatment provides a natural interpretation of many visual motion percepts, indicates that motion estimation should be revisited using a broader class of stimuli, demonstrates how correlation-based motion estimation is related to stimulus statistics, and provides multiple experimentally testable predictions.

}, keywords = {Algorithms, Animals, Computer Simulation, Models, Neurological, Motion, Motion Perception, Photic Stimulation, Time Factors}, issn = {1091-6490}, doi = {10.1073/pnas.1015680108}, author = {Fitzgerald, James E and Katsov, Alexander Y and Clandinin, Thomas R and Schnitzer, Mark J} } @article {46047, title = {A synaptic mechanism for retinal adaptation to luminance and contrast.}, journal = {Journal of Neuroscience}, volume = {31}, year = {2011}, month = {2011 Jul 27}, pages = {11003-15}, abstract = {

The gain of signaling in primary sensory circuits is matched to the stimulus intensity by the process of adaptation. Retinal neural circuits adapt to visual scene statistics, including the mean (background adaptation) and the temporal variance (contrast adaptation) of the light stimulus. The intrinsic properties of retinal bipolar cells and synapses contribute to background and contrast adaptation, but it is unclear whether both forms of adaptation depend on the same cellular mechanisms. Studies of bipolar cell synapses identified synaptic mechanisms of gain control, but the relevance of these mechanisms to visual processing is uncertain because of the historical focus on fast, phasic transmission rather than the tonic transmission evoked by ambient light. Here, we studied use-dependent regulation of bipolar cell synaptic transmission evoked by small, ongoing modulations of membrane potential (V(M)) in the physiological range. We made paired whole-cell recordings from rod bipolar (RB) and AII amacrine cells in a mouse retinal slice preparation. Quasi-white noise voltage commands modulated RB V(M) and evoked EPSCs in the AII. We mimicked changes in background luminance or contrast, respectively, by depolarizing the V(M) or increasing its variance. A linear systems analysis of synaptic transmission showed that increasing either the mean or the variance of the presynaptic V(M) reduced gain. Further electrophysiological and computational analyses demonstrated that adaptation to mean potential resulted from both Ca channel inactivation and vesicle depletion, whereas adaptation to variance resulted from vesicle depletion alone. Thus, background and contrast adaptation apparently depend in part on a common synaptic mechanism.

}, keywords = {Adaptation, Physiological, Amacrine Cells, Animals, Biophysical Phenomena, Biophysics, Calcium, Contrast Sensitivity, Electric Stimulation, Excitatory Postsynaptic Potentials, Female, In Vitro Techniques, Lighting, Male, Mice, Mice, Inbred C57BL, Models, Neurological, Numerical Analysis, Computer-Assisted, Patch-Clamp Techniques, Photic Stimulation, Presynaptic Terminals, Retina, Retinal Bipolar Cells, Synaptic Transmission}, issn = {1529-2401}, doi = {10.1523/JNEUROSCI.2631-11.2011}, author = {Jarsky, Tim and Cembrowski, Mark and Logan, Stephen M and Kath, William L and Riecke, Hermann and Demb, Jonathan B and Singer, Joshua H} } @article {46609, title = {Tao kinases as coordinators of actin and microtubule dynamics in developing neurons.}, journal = {Communicative \& Integrative Biology}, volume = {4}, year = {2011}, month = {2011 Sep}, pages = {554-6}, abstract = {

Drosophila tao, encoding a Ste20 family kinase, was identified as a gene involved in ethanol, cocaine and nicotine sensitivity. The behavioral phenotypes appear to be caused by defects in the development of the adult brain. Specifically, Drosophila tao functions to promote axon guidance of mushroom body (MB) neurons. The MB is a large structure in the central brain of the fly whose development and function have been well characterized. tao interacts genetically with mutations in the par-1 gene, also encoding a serine-threonine kinase. Since Par-1 has been implicated in the regulation of microtubule dynamics, this suggests that tao regulates the microtubule cytoskeleton in developing MB neurons. Here we discuss these results in light of previous studies that have proposed that Drosophila tao and its mammalian homologs function as a link between the actin and microtubule cytoskeletons, regulating microtubule stability in response to actin signals.

}, issn = {1942-0889}, doi = {10.4161/cib.4.5.16051}, author = {King, Ian and Heberlein, Ulrike} } @article {47665, title = {Together we are stronger: fusion protects mitochondria from autophagosomal degradation.}, journal = {Autophagy}, volume = {7}, year = {2011}, month = {2011 Dec}, pages = {1568-9}, abstract = {

Starvation induces a protective process of self-cannibalization called autophagy that is thought to mediate nonselective degradation of cytoplasmic material. We recently reported that mitochondria escape autophagosomal degradation through extensive fusion into mitochondrial networks upon certain starvation conditions. The extent of mitochondrial elongation is dependent on the type of nutrient deprivation, with amino acid depletion having a particularly strong effect. Downregulation of the mitochondrial fission protein Drp1 was determined to be important in bringing about starvation-induced mitochondrial fusion. The formation of mitochondrial networks during nutrient depletion selectively blocked their autophagic degradation, presumably allowing cells to sustain efficient ATP production and thereby survive starvation.

}, keywords = {Animals, Autophagy, Cyclic AMP-Dependent Protein Kinases, Membrane Fusion, Mice, Mitochondria, Phagosomes, Signal Transduction, TOR Serine-Threonine Kinases}, issn = {1554-8635}, author = {Rambold, Angelika S and Kostelecky, Brenda and Lippincott-Schwartz, Jennifer} } @article {47669, title = {Tubular network formation protects mitochondria from autophagosomal degradation during nutrient starvation.}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {108}, year = {2011}, month = {2011 Jun 21}, pages = {10190-5}, abstract = {

Mitochondria are highly dynamic organelles that mediate essential cell functions such as apoptosis and cell-cycle control in addition to their role as efficient ATP generators. Mitochondrial morphology changes are tightly regulated, and their shape can shift between small, fragmented units and larger networks of elongated mitochondria. We demonstrate that mitochondrial elements become significantly elongated and interconnected shortly after nutrient depletion. This mitochondrial morphological shift depends on the type of starvation, with an additive effect observed when multiple nutrients are depleted simultaneously. We further show that starvation-induced mitochondrial elongation is mediated by down-regulation of dynamin-related protein 1 (Drp1) through modulation of two Drp1 phosphorylation sites, leading to unopposed mitochondrial fusion. Finally, we establish that mitochondrial tubulation upon nutrient deprivation protects mitochondria from autophagosomal degradation, which could permit mitochondria to maximize energy production and supply autophagosomal membranes during starvation.

}, keywords = {Animals, Autophagy, Cells, Cultured, Dynamins, Fibroblasts, GTP Phosphohydrolases, Mice, Microtubule-Associated Proteins, Mitochondria, Mitochondrial Proteins, Phagosomes, Protein Processing, Post-Translational, Starvation}, issn = {1091-6490}, doi = {10.1073/pnas.1107402108}, author = {Rambold, Angelika S and Kostelecky, Brenda and Elia, Natalie and Lippincott-Schwartz, Jennifer} } @article {4305, title = {Two-photon calcium imaging of evoked activity from L5 somatosensory neurons in vivo.}, journal = {Nature Neuroscience}, volume = {14}, year = {2011}, month = {2011 Aug}, pages = {1089-93}, abstract = {

Multiphoton imaging (MPI) is widely used for recording activity simultaneously from many neurons in superficial cortical layers in vivo. We combined regenerative amplification multiphoton microscopy (RAMM) with genetically encoded calcium indicators to extend MPI of neuronal population activity into layer 5 (L5) of adult mouse somatosensory cortex. We found that this approach could be used to record and quantify spontaneous and sensory-evoked activity in populations of L5 neuronal somata located as much as 800 μm below the pia. In addition, we found that RAMM could be used to simultaneously image activity from large (80) populations of apical dendrites and follow these dendrites down to their somata of origin.

}, keywords = {Animals, Axons, Calcium, Dendrites, Evoked Potentials, Green Fluorescent Proteins, Inbred C57BL, Mice, Nerve Net, Neurons, Newborn, Photons, Physical Stimulation, Probability, Reaction Time, Somatosensory, Somatosensory Cortex, Statistics as Topic, Transgenic}, issn = {1546-1726}, doi = {10.1038/nn.2879}, author = {Mittmann, Wolfgang and Wallace, Damian J and Czubayko, Uwe and Herb, Jan T and Schaefer, Andreas T and Looger, Loren L and Denk, Winfried and Kerr, Jason N D} } @article {46737, title = {Variegated gene expression caused by cell-specific long-range DNA interactions.}, journal = {Nature Cell Biology}, volume = {13}, year = {2011}, month = {2011 Aug}, pages = {944-51}, abstract = {

Mammalian genomes contain numerous regulatory DNA sites with unknown target genes. We used mice with an extra β-globin locus control region (LCR) to investigate how a regulator searches the genome for target genes. We find that the LCR samples a restricted nuclear subvolume, wherein it preferentially contacts genes controlled by shared transcription factors. No contacted gene is detectably upregulated except for endogenous β-globin genes located on another chromosome. This demonstrates genetically that mammalian trans activation is possible, but suggests that it will be rare. Trans activation occurs not pan-cellularly, but in {\textquoteright}jackpot{\textquoteright} cells enriched for the interchromosomal interaction. Therefore, cell-specific long-range DNA contacts can cause variegated expression.

}, keywords = {Animals, beta-Globins, DNA, GATA1 Transcription Factor, Gene Expression, In Situ Hybridization, Fluorescence, Kruppel-Like Transcription Factors, Locus Control Region, Mice, Mice, Transgenic, RNA, Messenger, Transcriptional Activation}, issn = {1476-4679}, doi = {10.1038/ncb2278}, author = {Noordermeer, Daan and de Wit, Elzo and Klous, Petra and van de Werken, Harmen and Simonis, Marieke and Lopez-Jones, Melissa and Eussen, Bert and de Klein, Annelies and Singer, Robert H and de Laat, Wouter} } @article {6357, title = {A versatile strategy for gene trapping and trap conversion in emerging model organisms.}, journal = {Development }, volume = {138}, year = {2011}, month = {2011 Jun}, pages = {2625-30}, abstract = {

Genetic model organisms such as Drosophila, C. elegans and the mouse provide formidable tools for studying mechanisms of development, physiology and behaviour. Established models alone, however, allow us to survey only a tiny fraction of the morphological and functional diversity present in the animal kingdom. Here, we present iTRAC, a versatile gene-trapping approach that combines the implementation of unbiased genetic screens with the generation of sophisticated genetic tools both in established and emerging model organisms. The approach utilises an exon-trapping transposon vector that carries an integrase docking site, allowing the targeted integration of new constructs into trapped loci. We provide proof of principle for iTRAC in the emerging model crustacean Parhyale hawaiensis: we generate traps that allow specific developmental and physiological processes to be visualised in unparalleled detail, we show that trapped genes can be easily cloned from an unsequenced genome, and we demonstrate targeting of new constructs into a trapped locus. Using this approach, gene traps can serve as platforms for generating diverse reporters, drivers for tissue-specific expression, gene knockdown and other genetic tools not yet imagined.

}, keywords = {Animal, Animals, Cloning, Crustacea, DNA Transposable Elements, Genetic Techniques, Genetic Vectors, Integrases, Models, Molecular}, issn = {1477-9129}, doi = {10.1242/dev.066324}, author = {Kontarakis, Zacharias and Pavlopoulos, Anastasios and Kiupakis, Alexandros and Konstantinides, Nikolaos and Douris, Vassilis and Averof, Michalis} } @article {5385, title = {Wireless neural/EMG telemetry systems for small freely moving animals.}, journal = {IEEE Transactions on Biomedical Circuits and Systems}, volume = {5}, year = {2011}, month = {2011 Apr}, pages = {103-11}, abstract = {

We have developed miniature telemetry systems that capture neural, EMG, and acceleration signals from a freely moving insect or other small animal and transmit the data wirelessly to a remote digital receiver. The systems are based on custom low-power integrated circuits (ICs) that amplify, filter, and digitize four biopotential signals using low-noise circuits. One of the chips also digitizes three acceleration signals from an off-chip microelectromechanical-system accelerometer. All information is transmitted over a wireless ~ 900-MHz telemetry link. The first unit, using a custom chip fabricated in a 0.6- μm BiCMOS process, weighs 0.79 g and runs for two hours on two small batteries. We have used this system to monitor neural and EMG signals in jumping and flying locusts as well as transdermal potentials in weakly swimming electric fish. The second unit, using a custom chip fabricated in a 0.35-μ m complementary metal-oxide semiconductor CMOS process, weighs 0.17 g and runs for five hours on a single 1.5-V battery. This system has been used to monitor neural potentials in untethered perching dragonflies.

}, issn = {1932-4545}, doi = {10.1109/TBCAS.2011.2131140}, author = {Harrison, Reid R. and Fotowat, Haleh and Chan, Raymond and Kier, Ryan J and Olberg, Robert and Leonardo, Anthony and Gabbiani, Fabrizio} } @article {48172, title = {3D reconstruction of the human rib cage from 2D projection images using a statistical shape model.}, journal = {International journal of computer assisted radiology and surgery}, volume = {5}, year = {2010}, month = {2010 Mar}, pages = {111-24}, abstract = {

PURPOSE: This paper describes an approach for the three-dimensional (3D) shape and pose reconstruction of the human rib cage from few segmented two-dimensional (2D) projection images. Our work is aimed at supporting temporal subtraction techniques of subsequently acquired radiographs by establishing a method for the assessment of pose differences in sequences of chest radiographs of the same patient.

METHODS: The reconstruction method is based on a 3D statistical shape model (SSM) of the rib cage, which is adapted to binary 2D projection images of an individual rib cage. To drive the adaptation we minimize a distance measure that quantifies the dissimilarities between 2D projections of the 3D SSM and the projection images of the individual rib cage. We propose different silhouette-based distance measures and evaluate their suitability for the adaptation of the SSM to the projection images.

RESULTS: An evaluation was performed on 29 sets of biplanar binary images (posterior-anterior and lateral). Depending on the chosen distance measure, our experiments on the combined reconstruction of shape and pose of the rib cages yield reconstruction errors from 2.2 to 4.7 mm average mean 3D surface distance. Given a geometry of an individual rib cage, the rotational errors for the pose reconstruction range from 0.1 degrees to 0.9 degrees.

CONCLUSIONS: The results show that our method is suitable for the estimation of pose differences of the human rib cage in binary projection images. Thus, it is able to provide crucial 3D information for registration during the generation of 2D subtraction images.

}, keywords = {Humans, Image Interpretation, Computer-Assisted, Image Processing, Computer-Assisted, Imaging, Three-Dimensional, Models, Anatomic, Radiographic Image Enhancement, Ribs, Subtraction Technique}, issn = {1861-6429}, doi = {10.1007/s11548-009-0390-2}, author = {Dworzak, Jalda and Lamecker, Hans and von Berg, Jens and Klinder, Tobias and Lorenz, Cristian and Kainm{\"u}ller, Dagmar and Seim, Heiko and Hege, Hans-Christian and Zachow, Stefan} } @article {1454, title = {3D segmentation of cell boundaries from whole cell cryogenic electron tomography volumes.}, journal = {Journal of Structural Biology}, volume = {170}, year = {2010}, month = {2010 Apr}, pages = {134-45}, abstract = {

Cryogenic electron tomography (cryo-ET) has gained increasing interest in recent years due to its ability to image whole cells and subcellular structures in 3D at nanometer resolution in their native environment. However, due to dose restrictions and the inability to acquire high tilt angle images, the reconstructed volumes are noisy and have missing information. Thus, features are unreliable, and precision extraction of the cell boundary is difficult, manual and time intensive. This paper presents an efficient recursive algorithm called BLASTED (Boundary Localization using Adaptive Shape and Texture Discovery) to automatically extract the cell boundary using a conditional random field (CRF) framework in which boundary points and shape are jointly inferred. The algorithm learns the texture of the boundary region progressively, and uses a global shape model and shape-dependent features to propose candidate boundary points on a slice of the membrane. It then updates the shape of that slice by accepting the appropriate candidate points using local spatial clustering, the global shape model, and trained boosted texture classifiers. The BLASTED algorithm segmented the cell membrane over an average of 93\% of the length of the cell in 19 difficult cryo-ET datasets.

}, keywords = {Algorithms, Cell Membrane, Cryoelectron Microscopy, Electron Microscope Tomography}, issn = {1095-8657}, doi = {10.1016/j.jsb.2009.12.015}, author = {Moussavi, Farshid and Heitz, Geremy and Amat, Fernando and Comolli, Luis R and Koller, Daphne and Horowitz, Mark} } @article {4287, title = {Birth time/order-dependent neuron type specification.}, journal = {Current Opinion in Neurobiology}, volume = {20}, year = {2010}, month = {2010 Feb}, pages = {14-21}, abstract = {

Neurons derived from the same progenitor may acquire different fates according to their birth timing/order. To reveal temporally guided cell fates, we must determine neuron types as well as their lineage relationships and times of birth. Recent advances in genetic lineage analysis and fate mapping are facilitating such studies. For example, high-resolution lineage analysis can identify each sequentially derived neuron of a lineage and has revealed abrupt temporal identity changes in diverse Drosophila neuronal lineages. In addition, fate mapping of mouse neurons made from the same pool of precursors shows production of specific neuron types in specific temporal patterns. The tools used in these analyses are helping to further our understanding of the genetics of neuronal temporal identity.

}, keywords = {Animals, Cell Lineage, Drosophila, Mice, Neurogenesis, Neurons, Time Factors}, issn = {1873-6882}, doi = {10.1016/j.conb.2009.10.017}, author = {Kao, Chih-Fei and Lee, Tzumin} } @conference {3858, title = {Boundary learning by optimization with topological constraints.}, booktitle = {IEEE Conference on Computer Vision and Pattern Recognition}, year = {2010}, month = {2010}, abstract = {

Recent studies have shown that machine learning can improve the accuracy of detecting object boundaries in images. In the standard approach, a boundary detector is trained by minimizing its pixel-level disagreement with human boundary tracings. This naive metric is problematic because it is overly sensitive to boundary locations. This problem is solved by metrics provided with the Berkeley Segmentation Dataset, but these can be insensitive to topological differences, such as gaps in boundaries. Furthermore, the Berkeley metrics have not been useful as cost functions for supervised learning. Using concepts from digital topology, we propose a new metric called the warping error that tolerates disagreements over boundary location, penalizes topological disagreements, and can be used directly as a cost function for learning boundary detection, in a method that we call Boundary Learning by Optimization with Topological Constraints (BLOTC). We trained boundary detectors on electron microscopic images of neurons, using both BLOTC and standard training. BLOTC produced substantially better performance on a 1.2 million pixel test set, as measured by both the warping error and the Rand index evaluated on segmentations generated from the boundary labelings. We also find our approach yields significantly better segmentation performance than either gPb-OWT-UCM or multiscale normalized cut, as well as Boosted Edge Learning trained directly on our data.

}, author = {Jain, Viren and Bollmann, Benjamin and Richardson, Mark and Berger, Daniel R. and Helmstaedter, Moritz N. and Briggman, Kevin L and Denk, Winfried and Bowden, Jared B. and Mendenhall, John M. and Abraham, Wickliffe C. and Harris, Kristen M and Kasthuri, Narayanan and Hayworth, Ken J. and Schalek, Richard and Tapia, Juan Carlos and Lichtman, Jeff W. and Seung, Hyunjune Sebastian} } @article {2550, title = {Capture of particles by direct interception by cilia during feeding of a gastropod veliger.}, journal = {The Biological Bulletin}, volume = {218}, year = {2010}, month = {2010 Apr}, pages = {145-59}, abstract = {

Ciliary feeders vary in the arrangement of ciliary bands and mechanisms of capture of food. Some larvae use opposed parallel bands of preoral (prototroch) and postoral (metatroch) cilia. Hypotheses for the mechanism of particle capture include filtration by adhesion to a cilium that overtakes a particle (direct interception), but until now unequivocal evidence for this mechanism has been lacking. Here, high-speed video recordings of veliger larvae of the gastropod Lacuna vincta demonstrated direct interception of particles by prototrochal cilia. Adhesion between cilium and particle was seen when a prototrochal cilium tugged a diatom chain into the food groove while in contact with one part of the chain. In several recorded events, a prototochal cilium overtook a particle during its effective stroke and subsequently pulled the particle inward with its recovery stroke; thereupon, the particle was deposited onto the food groove and transported to the mouth. Captures varied, however. In some cases the particle was intercepted multiple times in one capture event; in others, several cilia passed a particle without interception. Particles occasionally remained in the area of recovery strokes, indicating retention without continuing adhesion to a cilium. In three events, a particle lost from prototrochal cilia was intercepted and moved into the food groove by metatrochal cilia. Particles as wide as or wider than the food groove were also captured and transported but were not ingested.

}, keywords = {Animals, Biological, Cilia, Feeding Behavior, Gastropoda, Microscopy, Models, Particulate Matter, Video}, issn = {1939-8697}, author = {Romero, Melissa R and Kelstrup, Hans C P and Strathmann, Richard R} } @article {1767, title = {The cells and peripheral representation of sodium taste in mice.}, journal = {Nature}, volume = {464}, year = {2010}, month = {2010 Mar 11}, pages = {297-301}, abstract = {

Salt taste in mammals can trigger two divergent behavioural responses. In general, concentrated saline solutions elicit robust behavioural aversion, whereas low concentrations of NaCl are typically attractive, particularly after sodium depletion. Notably, the attractive salt pathway is selectively responsive to sodium and inhibited by amiloride, whereas the aversive one functions as a non-selective detector for a wide range of salts. Because amiloride is a potent inhibitor of the epithelial sodium channel (ENaC), ENaC has been proposed to function as a component of the salt-taste-receptor system. Previously, we showed that four of the five basic taste qualities-sweet, sour, bitter and umami-are mediated by separate taste-receptor cells (TRCs) each tuned to a single taste modality, and wired to elicit stereotypical behavioural responses. Here we show that sodium sensing is also mediated by a dedicated population of TRCs. These taste cells express the epithelial sodium channel ENaC, and mediate behavioural attraction to NaCl. We genetically engineered mice lacking ENaCalpha in TRCs, and produced animals exhibiting a complete loss of salt attraction and sodium taste responses. Together, these studies substantiate independent cellular substrates for all five basic taste qualities, and validate the essential role of ENaC for sodium taste in mice.

}, keywords = {Animals, Behavior, Epithelial Sodium Channel, Mice, Sodium, Taste, Taste Buds, Transgenic}, issn = {1476-4687}, doi = {10.1038/nature08783}, author = {Chandrashekar, Jayaram and Kuhn, Christina and Oka, Yuki and Yarmolinsky, David A and Hummler, Edith and Ryba, Nicholas J P and Zuker, Charles S} } @article {6862, title = {Cellular organization of the neural circuit that drives Drosophila courtship behavior.}, journal = {Current Biology}, volume = {20}, year = {2010}, month = {2010 Sep 28}, pages = {1602-14}, abstract = {

BACKGROUND: Courtship behavior in Drosophila has been causally linked to the activity of the heterogeneous set of \~{}1500 neurons that express the sex-specific transcripts of the fruitless (fru) gene, but we currently lack an appreciation of the cellular diversity within this population, the extent to which these cells are sexually dimorphic, and how they might be organized into functional circuits. RESULTS: We used genetic methods to define 100 distinct classes of fru neuron, which we compiled into a digital 3D atlas at cellular resolution. We determined the polarity of many of these neurons and computed their likely patterns of connectivity, thereby assembling them into a neural circuit that extends from sensory input to motor output. The cellular organization of this circuit reveals neuronal pathways in the brain that are likely to integrate multiple sensory cues from other flies and to issue descending control signals to motor circuits in the thoracic ganglia. We identified 11 anatomical dimorphisms within this circuit: neurons that are male specific, are more numerous in males than females, or have distinct arborization patterns in males and females. CONCLUSIONS: The cellular organization of the fru circuit suggests how multiple distinct sensory cues are integrated in the fly{\textquoteright}s brain to drive sex-specific courtship behavior. We propose that sensory processing and motor control are mediated through circuits that are largely similar in males and females. Sex-specific behavior may instead arise through dimorphic circuits in the brain and nerve cord that differentially couple sensory input to motor output.

}, keywords = {Animal, Animals, Brain, Drosophila melanogaster, Female, Male, Nerve Net, Neurons, Sexual Behavior}, issn = {1879-0445}, doi = {10.1016/j.cub.2010.08.025}, author = {Yu, Jai Y and Kanai, Makoto I and Demir, Ebru and Jefferis, Gregory S X E and Dickson, Barry J} } @article {783, title = {A complete developmental sequence of a Drosophila neuronal lineage as revealed by twin-spot MARCM.}, journal = {PLoS Biology}, volume = {8}, year = {2010}, month = {2010 Aug 24}, abstract = {

Drosophila brains contain numerous neurons that form complex circuits. These neurons are derived in stereotyped patterns from a fixed number of progenitors, called neuroblasts, and identifying individual neurons made by a neuroblast facilitates the reconstruction of neural circuits. An improved MARCM (mosaic analysis with a repressible cell marker) technique, called twin-spot MARCM, allows one to label the sister clones derived from a common progenitor simultaneously in different colors. It enables identification of every single neuron in an extended neuronal lineage based on the order of neuron birth. Here we report the first example, to our knowledge, of complete lineage analysis among neurons derived from a common neuroblast that relay olfactory information from the antennal lobe (AL) to higher brain centers. By identifying the sequentially derived neurons, we found that the neuroblast serially makes 40 types of AL projection neurons (PNs). During embryogenesis, one PN with multi-glomerular innervation and 18 uniglomerular PNs targeting 17 glomeruli of the adult AL are born. Many more PNs of 22 additional types, including four types of polyglomerular PNs, derive after the neuroblast resumes dividing in early larvae. Although different offspring are generated in a rather arbitrary sequence, the birth order strictly dictates the fate of each post-mitotic neuron, including the fate of programmed cell death. Notably, the embryonic progenitor has an altered temporal identity following each self-renewing asymmetric cell division. After larval hatching, the same progenitor produces multiple neurons for each cell type, but the number of neurons for each type is tightly regulated. These observations substantiate the origin-dependent specification of neuron types. Sequencing neuronal lineages will not only unravel how a complex brain develops but also permit systematic identification of neuron types for detailed structure and function analysis of the brain.

}, keywords = {Animals, Brain, Cell Lineage, Clone Cells, Drosophila, Genetic Techniques, Mosaicism, Neurogenesis, Neurons, Olfactory Pathways}, issn = {1545-7885}, doi = {10.1371/journal.pbio.1000461}, author = {Yu, Hung-Hsiang and Kao, Chih-Fei and He, Yisheng and Ding, Peng and Kao, Jui-Chun and Lee, Tzumin} } @article {68825, title = {Dermal morphogenesis controls lateral line patterning during postembryonic development of teleost fish}, journal = {Developmental Biology}, volume = {340}, year = {2010}, month = {04/2010}, pages = {583-594}, abstract = {

The lateral line system displays highly divergent patterns in adult teleost fish. The mechanisms underlying this variability are poorly understood. Here, we demonstrate that the lateral line mechanoreceptor, the neuromast, gives rise to a series of accessory neuromasts by a serial budding process during postembryonic development in zebrafish. We also show that accessory neuromast formation is highly correlated to the development of underlying dermal structures such as bones and scales. Abnormalities in opercular bone morphogenesis, in endothelin 1-knockdown embryos, are accompanied by stereotypic errors in neuromast budding and positioning, further demonstrating the tight correlation between the patterning of neuromasts and of the underlying dermal bones. In medaka, where scales form between peridermis and opercular bones, the lateral line displays a scale-specific pattern which is never observed in zebrafish. These results strongly suggest a control of postembryonic neuromast patterns by underlying dermal structures. This dermal control may explain some aspects of the evolution of lateral line patterns.

}, keywords = {Dermal bone, Lateral line, Medaka, Neuromast, Opercle, Postembryonic development, Scale, Zebrafish}, issn = {0012-1606}, doi = {https://doi.org/10.1016/j.ydbio.2010.02.017}, url = {https://www.sciencedirect.com/science/article/pii/S0012160610001028}, author = {Hironori Wada and Alain Ghysen and Chie Satou and Shin-ichi Higashijima and Koichi Kawakami and Satoshi Hamaguchi and Mitsuru Sakaizumi} } @article {65010, title = {Development-based compartmentalization of the Drosophila central brain.}, journal = {J Comp Neurol}, volume = {518}, year = {2010}, month = {2010 Aug 01}, pages = {2996-3023}, abstract = {

The neuropile of the Drosophila brain is subdivided into anatomically discrete compartments. Compartments are rich in terminal neurite branching and synapses; they are the neuropile domains in which signal processing takes place. Compartment boundaries are defined by more or less dense layers of glial cells as well as long neurite fascicles. These fascicles are formed during the larval period, when the approximately 100 neuronal lineages that constitute the Drosophila central brain differentiate. Each lineage forms an axon tract with a characteristic trajectory in the neuropile; groups of spatially related tracts congregate into the brain fascicles that can be followed from the larva throughout metamorphosis into the adult stage. Here we provide a map of the adult brain compartments and the relevant fascicles defining compartmental boundaries. We have identified the neuronal lineages contributing to each fascicle, which allowed us to compare compartments of the larval and adult brain directly. Most adult compartments can be recognized already in the early larval brain, where they form a "protomap" of the later adult compartments. Our analysis highlights the morphogenetic changes shaping the Drosophila brain; the data will be important for studies that link early-acting genetic mechanisms to the adult neuronal structures and circuits controlled by these mechanisms.

}, keywords = {Animals, Antibodies, Monoclonal, Axons, Brain, Brain Mapping, Cadherins, Chemokine CX3CL1, Drosophila, Image Processing, Computer-Assisted, Immunohistochemistry, Larva, Neural Pathways, Neuroglia, Synapses}, issn = {1096-9861}, doi = {10.1002/cne.22376}, author = {Pereanu, Wayne and Kumar, Abilasha and Jennett, Arnim and Reichert, Heinrich and Hartenstein, Volker} } @article {1683, title = {Digital scanned laser light sheet fluorescence microscopy.}, journal = {Cold Spring Harbor Protocols}, volume = {2010}, year = {2010}, month = {2011 Oct}, pages = {pdb.top78}, abstract = {

Modern applications in the life sciences are frequently based on in vivo imaging of biological specimens, a domain for which light microscopy approaches are typically best suited. Often, quantitative information must be obtained from large multicellular organisms at the cellular or even subcellular level and with a good temporal resolution. However, this usually requires a combination of conflicting features: high imaging speed, low photobleaching and low phototoxicity in the specimen, good three-dimensional (3D) resolution, an excellent signal-to-noise ratio, and multiple-view imaging capability. The latter feature refers to the capability of recording a specimen along multiple directions, which is crucial for the imaging of large specimens with strong light-scattering or light-absorbing tissue properties. An imaging technique that fulfills these requirements is essential for many key applications: For example, studying fast cellular processes over long periods of time, imaging entire embryos throughout development, or reconstructing the formation of morphological defects in mutants. Here, we discuss digital scanned laser light sheet fluorescence microscopy (DSLM) as a novel tool for quantitative in vivo imaging in the post-genomic era and show how this emerging technique relates to the currently most widely applied 3D microscopy techniques in biology: confocal fluorescence microscopy and two-photon microscopy.

}, keywords = {Computer-Assisted, Confocal, Fluorescence, Image Processing, Microscopy}, issn = {1559-6095}, doi = {10.1101/pdb.top78}, author = {Keller, Philipp J and Stelzer, Ernst H K} } @article {65007, title = {Ephus: multipurpose data acquisition software for neuroscience experiments.}, journal = {Front Neural Circuits}, volume = {4}, year = {2010}, month = {2010}, pages = {100}, abstract = {

Physiological measurements in neuroscience experiments often involve complex stimulus paradigms and multiple data channels. Ephus (http://www.ephus.org) is an open-source software package designed for general-purpose data acquisition and instrument control. Ephus operates as a collection of modular programs, including an ephys program for standard whole-cell recording with single or multiple electrodes in typical electrophysiological experiments, and a mapper program for synaptic circuit mapping experiments involving laser scanning photostimulation based on glutamate uncaging or channelrhodopsin-2 excitation. Custom user functions allow user-extensibility at multiple levels, including on-line analysis and closed-loop experiments, where experimental parameters can be changed based on recently acquired data, such as during in vivo behavioral experiments. Ephus is compatible with a variety of data acquisition and imaging hardware. This paper describes the main features and modules of Ephus and their use in representative experimental applications.

}, issn = {1662-5110}, doi = {10.3389/fncir.2010.00100}, author = {Suter, Benjamin A and O{\textquoteright}Connor, Timothy and Iyer, Vijay and Petreanu, Leopoldo T and Hooks, Bryan M and Kiritani, Taro and Svoboda, Karel and Shepherd, Gordon M G} } @article {2326, title = {Error tolerant indexing and alignment of short reads with covering template families.}, journal = {Journal of Computational Biology: A Journal of Computational Molecular Cell Biology}, volume = {17}, year = {2010}, month = {2010 Oct}, pages = {1397-1411}, abstract = {

The rapid adoption of high-throughput next generation sequence data in biological research is presenting a major challenge for sequence alignment tools{\textemdash}specifically, the efficient alignment of vast amounts of short reads to large references in the presence of differences arising from sequencing errors and biological sequence variations. To address this challenge, we developed a short read aligner for high-throughput sequencer data that is tolerant of errors or mutations of all types{\textemdash}namely, substitutions, deletions, and insertions. The aligner utilizes a multi-stage approach in which template-based indexing is used to identify candidate regions for alignment with dynamic programming. A template is a pair of gapped seeds, with one used with the read and one used with the reference. In this article, we focus on the development of template families that yield error-tolerant indexing up to a given error-budget. A general algorithm for finding those families is presented, and a recursive construction that creates families with higher error tolerance from ones with a lower error tolerance is developed.

}, keywords = {Algorithms, Base Sequence, DNA, Genetic, Molecular Sequence Data, Sequence Alignment, Sequence Analysis, Software, Templates}, issn = {1557-8666}, doi = {10.1089/cmb.2010.0005}, author = {Giladi, Eldar and Healy, John and Myers, Gene and Hart, Chris and Kapranov, Philipp and Lipson, Doron and Roels, Steve and Thayer, Edward and Letovsky, Stan} } @article {46619, title = {Ethanol-regulated genes that contribute to ethanol sensitivity and rapid tolerance in Drosophila.}, journal = {Alcoholism, Clinical and Experimental Research}, volume = {34}, year = {2010}, month = {2010 Feb}, pages = {302-16}, abstract = {

BACKGROUND: Increased ethanol intake, a major predictor for the development of alcohol use disorders, is facilitated by the development of tolerance to both the aversive and pleasurable effects of the drug. The molecular mechanisms underlying ethanol tolerance development are complex and are not yet well understood.

METHODS: To identify genetic mechanisms that contribute to ethanol tolerance, we examined the time course of gene expression changes elicited by a single sedating dose of ethanol in Drosophila, and completed a behavioral survey of strains harboring mutations in ethanol-regulated genes.

RESULTS: Enrichment for genes in metabolism, nucleic acid binding, olfaction, regulation of signal transduction, and stress suggests that these biological processes are coordinately affected by ethanol exposure. We also detected a coordinate up-regulation of genes in the Toll and Imd innate immunity signal transduction pathways. A multi-study comparison revealed a small set of genes showing similar regulation, including increased expression of 3 genes for serine biosynthesis. A survey of Drosophila strains harboring mutations in ethanol-regulated genes for ethanol sensitivity and tolerance phenotypes revealed roles for serine biosynthesis, olfaction, transcriptional regulation, immunity, and metabolism. Flies harboring deletions of the genes encoding the olfactory co-receptor Or83b or the sirtuin Sir2 showed marked changes in the development of ethanol tolerance.

CONCLUSIONS: Our findings implicate novel roles for these genes in regulating ethanol behavioral responses.

}, keywords = {Animals, Behavior, Animal, Central Nervous System Depressants, Coenzyme A Ligases, Drosophila, Drosophila Proteins, Drug Tolerance, Ethanol, Female, Gene Expression Regulation, Histone Deacetylases, Male, Motor Activity, Mutation, Oligonucleotide Array Sequence Analysis, Receptors, Odorant, Reverse Transcriptase Polymerase Chain Reaction, Serpins, Sirtuins, Species Specificity}, issn = {1530-0277}, doi = {10.1111/j.1530-0277.2009.01093.x}, author = {Kong, Eric C and Allouche, Lorien and Chapot, Paul A and Vranizan, Karen and Moore, Monica S and Heberlein, Ulrike and Wolf, Fred W} } @article {2752, title = {Examining the combinatorial model of motor neuron survival by expression profiling of trophic factors and their receptors in the embryonic Gallus gallus.}, journal = {Developmental Dynamics: An Official Publication of the American Association of Anatomists}, volume = {239}, year = {2010}, month = {2010 Mar}, pages = {965-79}, abstract = {

During embryogenesis, limb-innervating lateral motor column (LMC) spinal motor neurons (MN) are generated in excess and subsequently nearly half of them die. Many motor neuron survival factors (MnSFs) have been shown to suppress this default programmed cell death (PCD) program through their receptors (MnSFRs), raising the possibility that they are involved in matching specific MNs with their target muscles. Published observations suggest a combinatorial model of MnSF/Rs function, which assumes that during the PCD phase, MNs are expressing combinations of MnSFRs, whereas the limb muscles innervated by these MNs express cognate combinations of MnSFs. We tested this model by expression profiling of MnSFs and their receptors in the avian lumbosacral spinal cord and limb muscles during the peak PCD period. Our findings highlight the complexity of MnSF/Rs function in the control of LMC motor neuron survival.

}, keywords = {Animals, Biological, Cell Death, Chick Embryo, Chickens, Computer-Assisted, Developmental, Developmental Biology, DNA Primers, Gene Expression Profiling, Gene Expression Regulation, Image Processing, Ligands, Lumbosacral Region, Models, Motor Neurons, Reverse Transcriptase Polymerase Chain Reaction, Spinal Cord}, issn = {1097-0177}, doi = {10.1002/dvdy.22215}, author = {Gu, Wendy X W and Kania, Artur} } @article {2027, title = {Fast, high-contrast imaging of animal development with scanned light sheet-based structured-illumination microscopy.}, journal = {Nature Methods}, volume = {7}, year = {2010}, month = {08/2010}, pages = {637-42}, abstract = {

Recording light-microscopy images of large, nontransparent specimens, such as developing multicellular organisms, is complicated by decreased contrast resulting from light scattering. Early zebrafish development can be captured by standard light-sheet microscopy, but new imaging strategies are required to obtain high-quality data of late development or of less transparent organisms. We combined digital scanned laser light-sheet fluorescence microscopy with incoherent structured-illumination microscopy (DSLM-SI) and created structured-illumination patterns with continuously adjustable frequencies. Our method discriminates the specimen-related scattered background from signal fluorescence, thereby removing out-of-focus light and optimizing the contrast of in-focus structures. DSLM-SI provides rapid control of the illumination pattern, exceptional imaging quality, and high imaging speeds. We performed long-term imaging of zebrafish development for 58 h and fast multiple-view imaging of early Drosophila melanogaster development. We reconstructed cell positions over time from the Drosophila DSLM-SI data and created a fly digital embryo.

}, keywords = {Animals, Drosophila melanogaster, Embryo, Microscopy, Nonmammalian, Zebrafish}, issn = {1548-7105}, doi = {10.1038/nmeth.1476}, author = {Keller, Philipp J and Schmidt, Annette D and Santella, Anthony and Khairy, Khaled and Zhirong Bao and Wittbrodt, Joachim and Stelzer, Ernst H K} } @article {2955, title = {Fast live simultaneous multiwavelength four-dimensional optical microscopy.}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {107}, year = {2010}, month = {2010 Sep 14}, pages = {16016-22}, abstract = {

Live fluorescence microscopy has the unique capability to probe dynamic processes, linking molecular components and their localization with function. A key goal of microscopy is to increase spatial and temporal resolution while simultaneously permitting identification of multiple specific components. We demonstrate a new microscope platform, OMX, that enables subsecond, multicolor four-dimensional data acquisition and also provides access to subdiffraction structured illumination imaging. Using this platform to image chromosome movement during a complete yeast cell cycle at one 3D image stack per second reveals an unexpected degree of photosensitivity of fluorophore-containing cells. To avoid perturbation of cell division, excitation levels had to be attenuated between 100 and 10,000{\texttimes} below the level normally used for imaging. We show that an image denoising algorithm that exploits redundancy in the image sequence over space and time allows recovery of biological information from the low light level noisy images while maintaining full cell viability with no fading.

}, keywords = {Algorithms, Animals, Cell Survival, Drosophila melanogaster, Fluorescence, Microscopy, Saccharomyces cerevisiae, Software}, issn = {1091-6490}, doi = {10.1073/pnas.1004037107}, author = {Carlton, Peter M and Boulanger, J{\'e}r{\^o}me and Kervrann, Charles and Sibarita, Jean-Baptiste and Salamero, Jean and Gordon-Messer, Susannah and Bressan, Debra and Haber, James E and Haase, Sebastian and Shao, Lin and Winoto, Lukman and Matsuda, Atsushi and Kner, Peter and Uzawa, Satoru and Gustafsson, Mats and Kam, Zvi and Agard, David A and Sedat, John W} } @article {4403, title = {A full-length cDNA resource for the pea aphid, acyrthosiphon pisum.}, journal = {Insect Molecular Biology}, volume = {19 Suppl 2}, year = {2010}, month = {2010 Mar}, pages = {23-31}, abstract = {

Large collections of full-length cDNAs are important resources for genome annotation and functional genomics. We report the creation of a collection of 50 599 full-length cDNA clones from the pea aphid, Acyrthosiphon pisum. Sequencing from 5{\textquoteright} and 3{\textquoteright} ends of the clones generated 97 828 high-quality expressed sequence tags, representing approximately 9000 genes. These sequences were imported to AphidBase and are shown to play crucial roles in both automatic gene prediction and manual annotation. Our detailed analyses demonstrated that the full-length cDNAs can further improve gene models and can even identify novel genes that are not included in the current version of the official gene set. This full-length cDNA collection can be utilized for a wide variety of functional studies, serving as a community resource for the study of the functional genomics of the pea aphid.

}, keywords = {3{\textquoteright} Untranslated Regions, 5{\textquoteright} Untranslated Regions, Animals, Aphids, Base Sequence, Complementary, DNA, DNA Primers, Expressed Sequence Tags, Gene Library, Genetic, Genome, Host-Pathogen Interactions, Insect, Models, Peas}, issn = {1365-2583}, doi = {10.1111/j.1365-2583.2009.00946.x}, author = {Shigenobu, S and Richards, S and Cree, A G and Morioka, M and Fukatsu, T and Kudo, T and Miyagishima, S and Gibbs, R A and Stern, D L and Nakabachi, A} } @article {2751, title = {Identification of genes controlled by LMX1B in E13.5 mouse limbs.}, journal = {Developmental Dynamics: An Official Publication of the American Association of Anatomists}, volume = {239}, year = {2010}, month = {2010 Aug}, pages = {2246-55}, abstract = {

During limb development, the dorsal limb mesenchyme expression of the transcription factor LMX1B is required for dorsoventral limb patterning. In mice, Lmx1b mutations result in the mirror-image duplication of ventral limb structures and loss of dorsal limb structures. Heterozygous LMX1B mutations in humans cause the Nail-Patella Syndrome characterized by limb, kidney, and eye developmental defects. We used DNA microarrays to compare the mRNAs in E13.5 mouse Lmx1b mutant and wild-type limbs. We report 14 genes that require Lmx1b for their normal expression in the dorsal limb or the restriction of their expression to the ventral limb.

}, keywords = {Animals, Body Patterning, Extremities, Gene Expression, Genes, Homeodomain Proteins, Humans, Messenger, Mice, Oligonucleotide Array Sequence Analysis, RNA, Transcription Factors}, issn = {1097-0177}, doi = {10.1002/dvdy.22357}, author = {Gu, Wendy X W and Kania, Artur} } @article {48167, title = {Improving deformable surface meshes through omni-directional displacements and MRFs.}, journal = {Medical image computing and computer-assisted intervention : MICCAI ... International Conference on Medical Image Computing and Computer-Assisted Intervention}, volume = {13}, year = {2010}, month = {2010}, pages = {227-34}, abstract = {

Deformable surface models are often represented as triangular meshes in image segmentation applications. For a fast and easily regularized deformation onto the target object boundary, the vertices of the mesh are commonly moved along line segments (typically surface normals). However, in case of high mesh curvature, these lines may intersect with the target boundary at "non-corresponding" positions, or even not at all. Consequently, certain deformations cannot be achieved. We propose an approach that allows each vertex to move not only along a line segment, but within a surrounding sphere. We achieve globally regularized deformations via Markov Random Field optimization. We demonstrate the potential of our approach with experiments on synthetic data, as well as an evaluation on 2 x 106 coronoid processes of the mandible in Cone-Beam CTs, and 56 coccyxes (tailbones) in low-resolution CTs.

}, keywords = {Algorithms, Computer Simulation, Cone-Beam Computed Tomography, Finite Element Analysis, Humans, Imaging, Three-Dimensional, Models, Biological, Radiographic Image Enhancement, Radiographic Image Interpretation, Computer-Assisted, Reproducibility of Results, Sensitivity and Specificity}, author = {Kainmueller, D and Lamecker, H and Seim, H and Zachow, S and Hege, H C} } @article {1245, title = {Increasing depth resolution of electron microscopy of neural circuits using sparse tomographic reconstruction.}, journal = {Computer Vision and Pattern Recognition (CVPR)}, year = {2010}, month = {2010}, pages = {1767-74}, issn = {1063-6919}, doi = {10.1109/CVPR.2010.5539846}, author = {Veeraraghavan, Ashok and Genkin, Alex V and Vitaladevuni, Shiv and Scheffer, Lou and Xu, CShan and Hess, Harald and Fetter, Richard and Cantoni, Marco and Knott, Graham and Chklovskii, Dmitri B} } @article {4288, title = {Learning-related fine-scale specificity imaged in motor cortex circuits of behaving mice.}, journal = {Nature}, volume = {464}, year = {2010}, month = {2010 Apr 22}, pages = {1182-6}, abstract = {

Cortical neurons form specific circuits, but the functional structure of this microarchitecture and its relation to behaviour are poorly understood. Two-photon calcium imaging can monitor activity of spatially defined neuronal ensembles in the mammalian cortex. Here we applied this technique to the motor cortex of mice performing a choice behaviour. Head-fixed mice were trained to lick in response to one of two odours, and to withhold licking for the other odour. Mice routinely showed significant learning within the first behavioural session and across sessions. Microstimulation and trans-synaptic tracing identified two non-overlapping candidate tongue motor cortical areas. Inactivating either area impaired voluntary licking. Imaging in layer 2/3 showed neurons with diverse response types in both areas. Activity in approximately half of the imaged neurons distinguished trial types associated with different actions. Many neurons showed modulation coinciding with or preceding the action, consistent with their involvement in motor control. Neurons with different response types were spatially intermingled. Nearby neurons (within approximately 150 mum) showed pronounced coincident activity. These temporal correlations increased with learning within and across behavioural sessions, specifically for neuron pairs with similar response types. We propose that correlated activity in specific ensembles of functionally related neurons is a signature of learning-related circuit plasticity. Our findings reveal a fine-scale and dynamic organization of the frontal cortex that probably underlies flexible behaviour.

}, keywords = {Animal, Animals, Axonal Transport, Behavior, Chemical, Choice Behavior, Inbred C57BL, Learning, Male, Mice, Motor Cortex, Motor Neurons, Neural Pathways, Odors, Pyramidal Cells, Reward, Stimulation, Time Factors, Tongue}, issn = {1476-4687}, doi = {10.1038/nature08897}, author = {Komiyama, Takaki and Sato, Takashi R and O{\textquoteright}Connor, Daniel H and Zhang, Ying-Xin and Huber, Daniel and Hooks, Bryan M and Gabitto, Mariano and Svoboda, Karel} } @article {46616, title = {Lmo4 in the nucleus accumbens regulates cocaine sensitivity.}, journal = {Genes, Brain, and Behavior}, volume = {9}, year = {2010}, month = {2010 Oct}, pages = {817-24}, abstract = {

An estimated 2 million Americans use cocaine, resulting in large personal and societal costs. Discovery of the genetic factors that contribute to cocaine abuse is important for understanding this complex disease. Previously, mutations in the Drosophila LIM-only (dLmo) gene were identified because of their increased behavioral sensitivity to cocaine. Here we show that the mammalian homolog Lmo4, which is highly expressed in brain regions implicated in drug addiction, plays a similar role in cocaine-induced behaviors. Mice with a global reduction in Lmo4 levels show increased sensitivity to the locomotor stimulatory effects of cocaine upon chronic cocaine administration. This effect is reproduced with downregulation of Lmo4 in the nucleus accumbens by RNA interference. Thus, Lmo genes play conserved roles in regulating the behavioral effects of cocaine in invertebrate and mammalian models of drug addiction.

}, keywords = {Adaptor Proteins, Signal Transducing, Animals, Behavior, Animal, beta-Galactosidase, Cells, Cultured, Cloning, Molecular, Cocaine, Dendritic Spines, Dopamine Uptake Inhibitors, Gene Expression, Homeodomain Proteins, Immunohistochemistry, Lentivirus, LIM Domain Proteins, Male, Mice, Mice, Inbred C57BL, Microdissection, Motor Activity, Mutagenesis, Insertional, Nucleus Accumbens, Reverse Transcriptase Polymerase Chain Reaction, RNA, RNA Interference, Stereotaxic Techniques, Transcription Factors}, issn = {1601-183X}, doi = {10.1111/j.1601-183X.2010.00620.x}, author = {Lasek, A W and Kapfhamer, D and Kharazia, V and Gesch, J and Giorgetti, F and Heberlein, U} } @article {2540, title = {Midline crossing by gustatory receptor neuron axons is regulated by fruitless, doublesex and the roundabout receptors.}, journal = {Development}, volume = {137}, year = {2010}, month = {2010 Jan}, pages = {323-32}, abstract = {

Although nervous system sexual dimorphisms are known in many species, relatively little is understood about the molecular mechanisms generating these dimorphisms. Recent findings in Drosophila provide the tools for dissecting how neurogenesis and neuronal differentiation are modulated by the Drosophila sex-determination regulatory genes to produce nervous system sexual dimorphisms. Here we report studies aimed at illuminating the basis of the sexual dimorphic axonal projection patterns of foreleg gustatory receptor neurons (GRNs): only in males do GRN axons project across the midline of the ventral nerve cord. We show that the sex determination genes fruitless (fru) and doublesex (dsx) both contribute to establishing this sexual dimorphism. Male-specific Fru (Fru(M)) acts in foreleg GRNs to promote midline crossing by their axons, whereas midline crossing is repressed in females by female-specific Dsx (Dsx(F)). In addition, midline crossing by these neurons might be promoted in males by male-specific Dsx (Dsx(M)). Finally, we (1) demonstrate that the roundabout (robo) paralogs also regulate midline crossing by these neurons, and (2) provide evidence that Fru(M) exerts its effect on midline crossing by directly or indirectly regulating Robo signaling.

}, keywords = {Animals, Axons, Cell Surface, Confocal, DNA-Binding Proteins, Drosophila melanogaster, Drosophila Proteins, Female, Immunologic, Male, Microscopy, Nerve Tissue Proteins, Neurons, Receptors, Sex Characteristics, Transcription Factors}, issn = {1477-9129}, doi = {10.1242/dev.045047}, author = {Mellert, David J and Knapp, Jon-Michael and Manoli, Devanand S and Meissner, Geoffrey W and Baker, Bruce S} } @article {47681, title = {Mitochondria supply membranes for autophagosome biogenesis during starvation.}, journal = {Cell}, volume = {141}, year = {2010}, month = {2010 May 14}, pages = {656-67}, abstract = {

Starvation-induced autophagosomes engulf cytosol and/or organelles and deliver them to lysosomes for degradation, thereby resupplying depleted nutrients. Despite advances in understanding the molecular basis of this process, the membrane origin of autophagosomes remains unclear. Here, we demonstrate that, in starved cells, the outer membrane of mitochondria participates in autophagosome biogenesis. The early autophagosomal marker, Atg5, transiently localizes to punctae on mitochondria, followed by the late autophagosomal marker, LC3. The tail-anchor of an outer mitochondrial membrane protein also labels autophagosomes and is sufficient to deliver another outer mitochondrial membrane protein, Fis1, to autophagosomes. The fluorescent lipid NBD-PS (converted to NBD-phosphotidylethanolamine in mitochondria) transfers from mitochondria to autophagosomes. Photobleaching reveals membranes of mitochondria and autophagosomes are transiently shared. Disruption of mitochondria/ER connections by mitofusin2 depletion dramatically impairs starvation-induced autophagy. Mitochondria thus play a central role in starvation-induced autophagy, contributing membrane to autophagosomes.

}, keywords = {Animals, Cell Line, Cell Physiological Phenomena, Culture Media, Membrane Proteins, Mitochondria, Mitochondrial Membranes, Mitochondrial Proteins, Phagosomes, Rats}, issn = {1097-4172}, doi = {10.1016/j.cell.2010.04.009}, author = {Hailey, Dale W and Rambold, Angelika S and Satpute-Krishnan, Prasanna and Mitra, Kasturi and Sougrat, Rachid and Kim, Peter K and Lippincott-Schwartz, Jennifer} } @conference {48181, title = {Model-based auto-segmentation of knee bones and cartilage in MRI data.}, booktitle = {Medical Image Analysis for the Clinic - A Grand Challenge, MICCAI 2010, the 13th International Conference on Medical Image Computing and Computer Assisted Intervention}, year = {2010}, month = {2010 Sep 24}, address = {Beijing, China}, abstract = {

We present a method for fully automatic segmentation of the bones and cartilages of the human knee from MRI data. Based on statistical shape models and graph-based optimization, first the femoral and tibial bone surfaces are reconstructed. Starting from the bone sur- faces the cartilages are segmented simultaneously with a multi object technique using prior knowledge on the variation of cartilage thickness. We validate our method on 40 clinical MRI datasets acquired before knee replacement.\ 

}, url = {http://www.ski10.org/data/2010-11-09-1131.pdf}, author = {Seim, Heiko and Kainmueller, Dagmar and Lamecker, Hans and Bindernagel, Matthias and Malinowski, Jana and Zachow, Stefan} } @article {4158, title = {Mutation of the Drosophila vesicular GABA transporter disrupts visual figure detection.}, journal = {The Journal of Experimental Biology}, volume = {213}, year = {2010}, month = {2010 May}, pages = {1717-30}, abstract = {

The role of gamma amino butyric acid (GABA) release and inhibitory neurotransmission in regulating most behaviors remains unclear. The vesicular GABA transporter (VGAT) is required for the storage of GABA in synaptic vesicles and provides a potentially useful probe for inhibitory circuits. However, specific pharmacologic agents for VGAT are not available, and VGAT knockout mice are embryonically lethal, thus precluding behavioral studies. We have identified the Drosophila ortholog of the vesicular GABA transporter gene (which we refer to as dVGAT), immunocytologically mapped dVGAT protein expression in the larva and adult and characterized a dVGAT(minos) mutant allele. dVGAT is embryonically lethal and we do not detect residual dVGAT expression, suggesting that it is either a strong hypomorph or a null. To investigate the function of VGAT and GABA signaling in adult visual flight behavior, we have selectively rescued the dVGAT mutant during development. We show that reduced GABA release does not compromise the active optomotor control of wide-field pattern motion. Conversely, reduced dVGAT expression disrupts normal object tracking and figure-ground discrimination. These results demonstrate that visual behaviors are segregated by the level of GABA signaling in flies, and more generally establish dVGAT as a model to study the contribution of GABA release to other complex behaviors.

}, keywords = {Alleles, Amino Acid Sequence, Animal, Animals, Behavior, Brain, Drosophila melanogaster, Drosophila Proteins, Fixation, Ganglia, Gene Knockdown Techniques, Larva, Mice, Molecular Sequence Data, Movement, Mutation, Ocular, Protein Transport, Synaptic Vesicles, Transgenes, Vesicular Inhibitory Amino Acid Transport Proteins, Visual Perception}, issn = {1477-9145}, doi = {10.1242/jeb.036053}, author = {Fei, Hao and Chow, Dawnis M and Chen, Audrey and Romero-Calder{\'o}n, Rafael and Ong, Wei S and Ackerson, Larry C and Maidment, Nigel T and Simpson, Julie H and Frye, Mark A and Krantz, David E} } @article {926, title = {Nanoscale architecture of integrin-based cell adhesions.}, journal = {Nature}, volume = {468}, year = {2010}, month = {2010 Nov 25}, pages = {580-4}, abstract = {

Cell adhesions to the extracellular matrix (ECM) are necessary for morphogenesis, immunity, and wound healing. Focal adhesions are multifunctional organelles that mediate cell-ECM adhesion, force transmission, cytoskeletal regulation and signaling. Focal adhesions consist of a complex network of trans-plasma-membrane integrins and cytoplasmic proteins that form a \<200-nm plaque linking the ECM to the actin cytoskeleton. The complexity of focal adhesion composition and dynamics implicate an intricate molecular machine. However, focal adhesion molecular architecture remains unknown. Here we used three-dimensional super-resolution fluorescence microscopy (interferometric photoactivated localization microscopy) to map nanoscale protein organization in focal adhesions. Our results reveal that integrins and actin are vertically separated by a \~{}40-nm focal adhesion core region consisting of multiple protein-specific strata: a membrane-apposed integrin signaling layer containing integrin cytoplasmic tails, focal adhesion kinase, and paxillin; an intermediate force-transduction layer containing talin and vinculin; and an uppermost actin-regulatory layer containing zyxin, vasodilator-stimulated phosphoprotein and α-actinin. By localizing amino- and carboxy-terminally tagged talins, we reveal talin{\textquoteright}s polarized orientation, indicative of a role in organizing the focal adhesion strata. The composite multilaminar protein architecture provides a molecular blueprint for understanding focal adhesion functions.

}, keywords = {Actins, Animals, Biological, Cell Adhesion, Cell Line, Cell Membrane, Extracellular Matrix, Humans, Integrins, Mice, Models, Tumor}, issn = {1476-4687}, doi = {10.1038/nature09621}, author = {Kanchanawong, Pakorn and Shtengel, Gleb and Pasapera, Ana M and Ramko, Ericka B and Davidson, Michael W and Hess, Harald F and Waterman, Clare M} } @article {1681, title = {Nlcam modulates midline convergence during anterior neural plate morphogenesis.}, journal = {Developmental Biology}, volume = {339}, year = {2010}, month = {2010 Mar 1}, pages = {14-25}, abstract = {

During development, different cell types must undergo distinct morphogenetic programs so that tissues develop the right dimensions in the appropriate place. In early eye morphogenesis, retinal progenitor cells (RPCs) move first towards the midline, before turning around to migrate out into the evaginating optic vesicles. Neighbouring forebrain cells, however, converge rapidly and then remain at the midline. These differential behaviours are regulated by the transcription factor Rx3. Here, we identify a downstream target of Rx3, the Ig-domain protein Nlcam, and characterise its role in regulating cell migration during the initial phase of optic vesicle morphogenesis. Through sophisticated live imaging and comprehensive cell tracking experiments in zebrafish, we show that ectopic expression of Nlcam in RPCs, as is observed in Rx3 mutants, causes enhanced convergence of these cells. Expression levels of Nlcam therefore regulate the migratory properties of RPCs. Our results provide evidence that the two phases of optic vesicle morphogenesis: slowed convergence and outward-directed migration, are under different genetic control. We propose that Nlcam forms part of the guidance machinery directing rapid midline migration of forebrain precursors, where it is normally expressed, and that its ectopic expression upon loss of Rx3 imparts these migratory characteristics upon RPCs.

}, keywords = {Animals, Base Sequence, Body Patterning, Cell Adhesion Molecules, Chick Embryo, DNA Primers, Electrophoretic Mobility Shift Assay, Homeodomain Proteins, Morphogenesis, Neural Plate, Protein Binding, Zebrafish}, issn = {1095-564X}, doi = {10.1016/j.ydbio.2009.12.003}, author = {Brown, Katherine E and Keller, Philipp J and Ramialison, Mirana and Rembold, Martina and Stelzer, Ernst H K and Loosli, Felix and Wittbrodt, Joachim} } @article {1541, title = {Nodavirus-induced membrane rearrangement in replication complex assembly requires replicase protein a, RNA templates, and polymerase activity.}, journal = {Journal of Virology}, volume = {84}, year = {2010}, month = {2010 Dec}, pages = {12492-503}, abstract = {

Positive-strand RNA [(+)RNA] viruses invariably replicate their RNA genomes on modified intracellular membranes. In infected Drosophila cells, Flock House nodavirus (FHV) RNA replication complexes form on outer mitochondrial membranes inside \~{}50-nm, virus-induced spherular invaginations similar to RNA replication-linked spherules induced by many (+)RNA viruses at various membranes. To better understand replication complex assembly, we studied the mechanisms of FHV spherule formation. FHV has two genomic RNAs; RNA1 encodes multifunctional RNA replication protein A and RNA interference suppressor protein B2, while RNA2 encodes the capsid proteins. Expressing genomic RNA1 without RNA2 induced mitochondrial spherules indistinguishable from those in FHV infection. RNA1 mutation showed that protein B2 was dispensable and that protein A was the only FHV protein required for spherule formation. However, expressing protein A alone only "zippered" together the surfaces of adjacent mitochondria, without inducing spherules. Thus, protein A is necessary but not sufficient for spherule formation. Coexpressing protein A plus a replication-competent FHV RNA template induced RNA replication in trans and membrane spherules. Moreover, spherules were not formed when replicatable FHV RNA templates were expressed with protein A bearing a single, polymerase-inactivating amino acid change or when wild-type protein A was expressed with a nonreplicatable FHV RNA template. Thus, unlike many (+)RNA viruses, the membrane-bounded compartments in which FHV RNA replication occurs are not induced solely by viral protein(s) but require viral RNA synthesis. In addition to replication complex assembly, the results have implications for nodavirus interaction with cell RNA silencing pathways and other aspects of virus control.

}, keywords = {Animals, Blotting, Capsid Proteins, DNA-Directed RNA Polymerases, Drosophila melanogaster, Genome, Intracellular Membranes, Mitochondria, Nodaviridae, Northern, Plasmids, RNA, RNA Replicase, RNA Virus Infections, Small Interfering, Viral, Virus Replication, Western}, issn = {1098-5514}, doi = {10.1128/JVI.01495-10}, url = {http://jvi.asm.org/content/84/24/12492.short}, author = {Kopek, Benjamin G and Settles, Erik W and Friesen, Paul D and Ahlquist, Paul} } @article {46618, title = {A pair of dopamine neurons target the D1-like dopamine receptor DopR in the central complex to promote ethanol-stimulated locomotion in Drosophila.}, journal = {PLoS One}, volume = {5}, year = {2010}, month = {2010 Apr 1}, pages = {e9954}, abstract = {

Dopamine is a mediator of the stimulant properties of drugs of abuse, including ethanol, in mammals and in the fruit fly Drosophila. The neural substrates for the stimulant actions of ethanol in flies are not known. We show that a subset of dopamine neurons and their targets, through the action of the D1-like dopamine receptor DopR, promote locomotor activation in response to acute ethanol exposure. A bilateral pair of dopaminergic neurons in the fly brain mediates the enhanced locomotor activity induced by ethanol exposure, and promotes locomotion when directly activated. These neurons project to the central complex ellipsoid body, a structure implicated in regulating motor behaviors. Ellipsoid body neurons are required for ethanol-induced locomotor activity and they express DopR. Elimination of DopR blunts the locomotor activating effects of ethanol, and this behavior can be restored by selective expression of DopR in the ellipsoid body. These data tie the activity of defined dopamine neurons to D1-like DopR-expressing neurons to form a neural circuit that governs acute responding to ethanol.

}, keywords = {Animals, Behavior, Animal, Central Nervous System Depressants, Dopamine, Drosophila, Drosophila Proteins, Ethanol, Locomotion, Motor Activity, Neurons, Receptors, Dopamine}, issn = {1932-6203}, doi = {10.1371/journal.pone.0009954}, author = {Kong, Eric C and Woo, Katherine and Li, Haiyan and Lebestky, Tim and Mayer, Nasima and Sniffen, Melissa R and Heberlein, Ulrike and Bainton, Roland J and Hirsh, Jay and Wolf, Fred W} } @article {6780, title = {A post-burst after depolarization is mediated by group i metabotropic glutamate receptor-dependent upregulation of Ca(v)2.3 R-type calcium channels in CA1 pyramidal neurons.}, journal = {PLoS Biology}, volume = {8}, year = {2010}, month = {2010}, pages = {e1000534}, abstract = {

Activation of group I metabotropic glutamate receptors (subtypes mGluR1 and mGluR5) regulates neural activity in a variety of ways. In CA1 pyramidal neurons, activation of group I mGluRs eliminates the post-burst afterhyperpolarization (AHP) and produces an afterdepolarization (ADP) in its place. Here we show that upregulation of Ca(v)2.3 R-type calcium channels is responsible for a component of the ADP lasting several hundred milliseconds. This medium-duration ADP is rapidly and reversibly induced by activation of mGluR5 and requires activation of phospholipase C (PLC) and release of calcium from internal stores. Effects of mGluR activation on subthreshold membrane potential changes are negligible but are large following action potential firing. Furthermore, the medium ADP exhibits a biphasic activity dependence consisting of short-term facilitation and longer-term inhibition. These findings suggest that mGluRs may dramatically alter the firing of CA1 pyramidal neurons via a complex, activity-dependent modulation of Ca(v)2.3 R-type channels that are activated during spiking at physiologically relevant rates and patterns.

}, keywords = {Action Potentials, Animals, Calcium Channels, Cation Transport Proteins, Female, Inbred C57BL, Ion Channel Gating, Male, Metabotropic Glutamate, Methoxyhydroxyphenylglycol, Mice, Neurons, Patch-Clamp Techniques, Pyramidal Cells, R-Type, Rats, Receptors, Up-Regulation, Wistar, Xenopus}, issn = {1545-7885}, doi = {10.1371/journal.pbio.1000534}, author = {Park, Jin-Yong and Remy, Stefan and Varela, Juan and Cooper, Donald C and Chung, Sungkwon and Kang, Ho-Won and Lee, Jung-Ha and Spruston, Nelson} } @article {68824, title = {Proneural gene-linked neurogenesis in zebrafish cerebellum}, journal = {Developmental Biology}, volume = {343}, year = {2010}, month = {07/2010}, pages = {1-17}, abstract = {

In mammals, cerebellar neurons are categorized as glutamatergic or GABAergic, and are derived from progenitors that express the proneural genes atoh1 or ptf1a, respectively. In zebrafish, three atoh1 genes, atoh1a, atoh1b, and atoh1c, are expressed in overlapping but distinct expression domains in the upper rhombic lip (URL): ptf1a is expressed exclusively in the ventricular zone (VZ). Using transgenic lines expressing fluorescent proteins under the control of the regulatory elements of atoh1a and ptf1a, we traced the lineages of the cerebellar neurons. The atoh1+ progenitors gave rise not only to granule cells but also to neurons of the anteroventral rhombencephalon. The ptf1a+ progenitors generated Purkinje cells. The olig2+ eurydendroid cells, which are glutamatergic, were derived mostly from ptf1a+ progenitors in the VZ but some originated from the atoh1+ progenitors in the URL. In the adult cerebellum, atoh1a, atoh1b, and atoh1c are expressed in the molecular layer of the valvula cerebelli and of the medial corpus cerebelli, and ptf1a was detected in the VZ. The proneural gene expression patterns coincided with the sites of proliferating neuronal progenitors in the adult cerebellum. Our data indicate that proneural gene-linked neurogenesis is evolutionarily conserved in the cerebellum among vertebrates, and that the continuously generated neurons help remodel neural circuits in the adult zebrafish cerebellum.

}, keywords = {Adult neurogenesis, Cerebellum, GABAergic neuron, Glutamatergic neuron, Hindbrain, Proneural gene, Zebrafish}, issn = {0012-1606}, doi = {https://doi.org/10.1016/j.ydbio.2010.03.024}, url = {https://www.sciencedirect.com/science/article/pii/S0012160610002071}, author = {Shuichi Kani and Young-Ki Bae and Takashi Shimizu and Koji Tanabe and Chie Satou and Michael J. Parsons and Ethan Scott and Shin-ichi Higashijima and Masahiko Hibi} } @article {46617, title = {Protein Phosphatase 2a and glycogen synthase kinase 3 signaling modulate prepulse inhibition of the acoustic startle response by altering cortical M-Type potassium channel activity.}, journal = {The Journal of Neuroscience}, volume = {30}, year = {2010}, month = {2010 Jun 30}, pages = {8830-40}, abstract = {

There is considerable interest in the regulation of sensorimotor gating, since deficits in this process could play a critical role in the symptoms of schizophrenia and other psychiatric disorders. Sensorimotor gating is often studied in humans and rodents using the prepulse inhibition of the acoustic startle response (PPI) model, in which an acoustic prepulse suppresses behavioral output to a startle-inducing stimulus. However, the molecular and neural mechanisms underlying PPI are poorly understood. Here, we show that a regulatory pathway involving protein phosphatase 2A (PP2A), glycogen synthase kinase 3 beta (GSK3beta), and their downstream target, the M-type potassium channel, regulates PPI. Mice (Mus musculus) carrying a hypomorphic allele of Ppp2r5delta, encoding a regulatory subunit of PP2A, show attenuated PPI. This PPP2R5delta reduction increases the phosphorylation of GSK3beta at serine 9, which inactivates GSK3beta, indicating that PPP2R5delta positively regulates GSK3beta activity in the brain. Consistently, genetic and pharmacological manipulations that reduce GSK3beta function attenuate PPI. The M-type potassium channel subunit, KCNQ2, is a putative GSK3beta substrate. Genetic reduction of Kcnq2 also reduces PPI, as does systemic inhibition of M-channels with linopirdine. Importantly, both the GSK3 inhibitor 3-(2,4-dichlorophenyl)-4-(1-methyl-1H-indol-3-yl)1H-pyrrole-2,5-dione (SB216763) and linopirdine reduce PPI when directly infused into the medial prefrontal cortex (mPFC). Whole-cell electrophysiological recordings of mPFC neurons show that SB216763 and linopirdine have similar effects on firing, and GSK3 inhibition occludes the effects of M-channel inhibition. These data support a previously uncharacterized mechanism by which PP2A/GSK3beta signaling regulates M-type potassium channel activity in the mPFC to modulate sensorimotor gating.

}, keywords = {Amino Acid Sequence, Animals, Auditory Perception, Brain, Glycogen Synthase Kinase 3, Inhibition (Psychology), KCNQ2 Potassium Channel, Mice, Mice, Inbred C57BL, Mice, Transgenic, Models, Neurological, Neurons, Phosphorylation, Potassium Channels, Prefrontal Cortex, Protein Phosphatase 2, Reflex, Startle, Signal Transduction}, issn = {1529-2401}, doi = {10.1523/JNEUROSCI.1292-10.2010}, author = {Kapfhamer, David and Berger, Karen H and Hopf, F Woodward and Seif, Taban and Kharazia, Viktor and Bonci, Antonello and Heberlein, Ulrike} } @article {2549, title = {Reproductive constraints, direct fitness and indirect fitness benefits explain helping behaviour in the primitively eusocial wasp, Polistes canadensis.}, journal = {Proceedings. Biological Sciences / The Royal Society}, volume = {277}, year = {2010}, month = {2010 Jun 7}, pages = {1721-8}, abstract = {

A key step in the evolution of sociality is the abandonment of independent breeding in favour of helping. In cooperatively breeding vertebrates and primitively eusocial insects, helpers are capable of leaving the group and reproducing independently, and yet many do not. A fundamental question therefore is why do helpers help? Helping behaviour may be explained by constraints on independent reproduction and/or benefits to individuals from helping. Here, we examine simultaneously the reproductive constraints and fitness benefits underlying helping behaviour in a primitively eusocial paper wasp. We gave 31 helpers the opportunity to become egg-layers on their natal nests by removing nestmates. This allowed us to determine whether helpers are reproductively constrained in any way. We found that age strongly influenced whether an ex-helper could become an egg-layer, such that young ex-helpers could become egg-layers while old ex-helpers were less able. These differential reproductive constraints enabled us to make predictions about the behaviours of ex-helpers, depending on the relative importance of direct and indirect fitness benefits. We found little evidence that indirect fitness benefits explain helping behaviour, as 71 per cent of ex-helpers left their nests before the end of the experiment. In the absence of reproductive constraints, however, young helpers value direct fitness opportunities over indirect fitness. We conclude that a combination of reproductive constraints and potential for future direct reproduction explain helping behaviour in this species. Testing several competing explanations for helping behaviour simultaneously promises to advance our understanding of social behaviour in animal groups.

}, keywords = {Animal, Animals, Behavior, Female, Helping Behavior, Population Density, Reproduction, Social Behavior, Wasps}, issn = {1471-2954}, doi = {10.1098/rspb.2009.2289}, author = {Sumner, Seirian and Kelstrup, Hans and Fanelli, Daniele} } @article {6388, title = {Scanning light-sheet microscopy in the whole mouse brain with HiLo background rejection.}, journal = {Journal of Biomedical Optics}, volume = {15}, year = {2010}, month = {2010 Jan-Feb}, pages = {016027}, abstract = {

It is well known that light-sheet illumination can enable optically sectioned wide-field imaging of macroscopic samples. However, the optical sectioning capacity of a light-sheet macroscope is undermined by sample-induced scattering or aberrations that broaden the thickness of the sheet illumination. We present a technique to enhance the optical sectioning capacity of a scanning light-sheet microscope by out-of-focus background rejection. The technique, called HiLo microscopy, makes use of two images sequentially acquired with uniform and structured sheet illumination. An optically sectioned image is then synthesized by fusing high and low spatial frequency information from both images. The benefits of combining light-sheet macroscopy and HiLo background rejection are demonstrated in optically cleared whole mouse brain samples, using both green fluorescent protein (GFP)-fluorescence and dark-field scattered light contrast.

}, keywords = {Animals, Background Radiation, Brain, Computer-Assisted, Fluorescence, Image Processing, Mice, Microscopy}, issn = {1560-2281}, doi = {10.1117/1.3324890}, author = {Mertz, Jerome and Kim, Jinhyun} } @article {1682, title = {Segregation of yeast nuclear pores.}, journal = {Nature}, volume = {466}, year = {2010}, month = {2010 Jul 22}, pages = {E1}, abstract = {

During mitosis in Saccharomyces cerevisiae, senescence factors such as extrachromosomal ribosomal DNA circles (ERCs) are retained in the mother cell and excluded from the bud/daughter cell. Shcheprova et al. proposed a model suggesting segregation of ERCs through their association with nuclear pore complexes (NPCs) and retention of preexisting NPCs in the mother cell during mitosis. However, this model is inconsistent with previous data and we demonstrate here that NPCs do efficiently migrate from the mother into the bud. Therefore, binding to NPCs does not seem to explain the retention of ERCs in the mother cell.

}, keywords = {Chromosome Segregation, DNA, Fungal, Mitosis, Nuclear Pore, Nuclear Pore Complex Proteins, Ribosomal, Saccharomyces cerevisiae, Saccharomyces cerevisiae Proteins}, issn = {1476-4687}, doi = {10.1038/nature09255}, author = {Khmelinskii, Anton and Keller, Philipp J and Lorenz, Holger and Schiebel, Elmar and Knop, Michael} } @article {1032, title = {Sex and the single cell. II. There is a time and place for sex.}, journal = {PLoS Biology}, volume = {8}, year = {2010}, month = {2010 May}, pages = {e1000365}, abstract = {

The Drosophila melanogaster sex hierarchy controls sexual differentiation of somatic cells via the activities of the terminal genes in the hierarchy, doublesex (dsx) and fruitless (fru). We have targeted an insertion of GAL4 into the dsx gene, allowing us to visualize dsx-expressing cells in both sexes. Developmentally and as adults, we find that both XX and XY individuals are fine mosaics of cells and tissues that express dsx and/or fruitless (fru(M)), and hence have the potential to sexually differentiate, and those that don{\textquoteright}t. Evolutionary considerations suggest such a mosaic expression of sexuality is likely to be a property of other animal species having two sexes. These results have also led to a major revision of our view of how sex-specific functions are regulated by the sex hierarchy in flies. Rather than there being a single regulatory event that governs the activities of all downstream sex determination regulatory genes-turning on Sex lethal (Sxl) RNA splicing activity in females while leaving it turned off in males-there are, in addition, elaborate temporal and spatial transcriptional controls on the expression of the terminal regulatory genes, dsx and fru. Thus tissue-specific aspects of sexual development are jointly specified by post-transcriptional control by Sxl and by the transcriptional controls of dsx and fru expression.

}, keywords = {Animals, Developmental, DNA-Binding Proteins, Drosophila melanogaster, Drosophila Proteins, Female, Gene Expression Regulation, Male, Nerve Tissue Proteins, Organ Specificity, RNA-Binding Proteins, Sex Characteristics, Sex Differentiation, Transcription Factors}, issn = {1545-7885}, doi = {10.1371/journal.pbio.1000365}, author = {Robinett, Carmen C and Vaughan, Alexander G and Knapp, Jon-Michael and Baker, Bruce S} } @article {2046, title = {Single-molecule discrimination of discrete perisynaptic and distributed sites of actin filament assembly within dendritic spines. (With commentary)}, journal = {Neuron}, volume = {67}, year = {2010}, month = {2010 Jul 15}, pages = {86-99}, abstract = {

Within dendritic spines, actin is presumed to anchor receptors in the postsynaptic density and play numerous roles regulating synaptic transmission. However, the submicron dimensions of spines have hindered examination of actin dynamics within them and prevented live-cell discrimination of perisynaptic actin filaments. Using photoactivated localization microscopy, we measured movement of individual actin molecules within living spines. Velocity of single actin molecules along filaments, an index of filament polymerization rate, was highly heterogeneous within individual spines. Most strikingly, molecular velocity was elevated in discrete, well-separated foci occurring not principally at the spine tip, but in subdomains throughout the spine, including the neck. Whereas actin velocity on filaments at the synapse was substantially elevated, at the endocytic zone there was no enhanced polymerization activity. We conclude that actin subserves spatially diverse, independently regulated processes throughout spines. Perisynaptic actin forms a uniquely dynamic structure well suited for direct, active regulation of the synapse.

Commentary: A nice application of single particle tracking PALM (sptPALM), showing the flow of actin in the spines of live cultured neurons. Since 2008, the PALM in our lab has largely become a user facility, available to outside users as well as Janelians. Grad student Nick Frost in Tom Blanpied{\textquoteright}s group at the U. of Maryland Med School visited on a number of occasions to use the PALM, with training and assistance from Hari.

}, keywords = {Animals, Biological, Cells, Cultured, Dendritic Spines, Embryo, Green Fluorescent Proteins, Hippocampus, Mammalian, Microfilaments, Models, Nerve Tissue Proteins, Neuronal Plasticity, Neurons, Presynaptic Terminals, Protein Transport, Rats, Sprague-Dawley, Synaptic Transmission, Tissue Distribution, Transfection}, issn = {1097-4199}, doi = {10.1016/j.neuron.2010.05.026}, url = {http://www.ncbi.nlm.nih.gov/pubmed/20624594}, author = {Frost, Nicholas A and Shroff, Hari and Kong, Huihui and Betzig, Eric and Blanpied, Thomas A} } @article {2768, title = {Snapshot Image Mapping Spectrometer (IMS) with high sampling density for hyperspectral microscopy.}, journal = {Optics Express}, volume = {18}, year = {2010}, month = {2010 Jul 5}, pages = {14330-44}, abstract = {

A snapshot Image Mapping Spectrometer (IMS) with high sampling density is developed for hyperspectral microscopy, measuring a datacube of dimensions 285 x 285 x 60 (x, y, lambda). The spatial resolution is approximately 0.45 microm with a FOV of 100 x 100 microm(2). The measured spectrum is from 450 nm to 650 nm and is sampled by 60 spectral channels with average sampling interval approximately 3.3 nm. The channel{\textquoteright}s spectral resolution is approximately 8nm. The spectral imaging results demonstrate the potential of the IMS for real-time cellular fluorescence imaging.

}, keywords = {Animals, Cattle, Cell Nucleus, Endothelial Cells, Fluorescence, Imaging, Microscopy, Pulmonary Artery, Spectrum Analysis, Staining and Labeling, Three-Dimensional}, issn = {1094-4087}, doi = {10.1364/AO.50.001792}, author = {Gao, Liang and Kester, Robert T and Hagen, Nathan and Tkaczyk, Tomasz S} } @article {6340, title = {Structure of clathrin coat with bound Hsc70 and auxilin: mechanism of Hsc70-facilitated disassembly.}, journal = {The EMBO Journal}, volume = {29}, year = {2010}, month = {2010 Feb 3}, pages = {655-65}, abstract = {

The chaperone Hsc70 drives the clathrin assembly-disassembly cycle forward by stimulating dissociation of a clathrin lattice. A J-domain containing co-chaperone, auxilin, associates with a freshly budded clathrin-coated vesicle, or with an in vitro assembled clathrin coat, and recruits Hsc70 to its specific heavy-chain-binding site. We have determined by electron cryomicroscopy (cryoEM), at about 11 A resolution, the structure of a clathrin coat (in the D6-barrel form) with specifically bound Hsc70 and auxilin. The Hsc70 binds a previously analysed site near the C-terminus of the heavy chain, with a stoichiometry of about one per three-fold vertex. Its binding is accompanied by a distortion of the clathrin lattice, detected by a change in the axial ratio of the D6 barrel. We propose that when Hsc70, recruited to a position close to its target by the auxilin J-domain, splits ATP, it clamps firmly onto its heavy-chain site and locks in place a transient fluctuation. Accumulation of the local strain thus imposed at multiple vertices can then lead to disassembly.

}, keywords = {Animals, Auxilins, Biological, Cattle, Clathrin, Clathrin-Coated Vesicles, Cryoelectron Microscopy, HSC70 Heat-Shock Proteins, Models, Molecular, Multiprotein Complexes, Protein Binding, Protein Multimerization, Protein Structure, Quaternary, Secondary}, issn = {1460-2075}, doi = {10.1038/emboj.2009.383}, author = {Xing, Yi and B{\"o}cking, Till and Wolf, Matthias and Grigorieff, Nikolaus and Kirchhausen, Tomas and Harrison, Stephen C} } @article {3632, title = {Structure of the cholera toxin secretion channel in its closed state.}, journal = {Nature Structural \& Molecular Biology}, volume = {17}, year = {2010}, month = {2010 Oct}, pages = {1226-32}, abstract = {

The type II secretion system (T2SS) is a macromolecular complex spanning the inner and outer membranes of Gram-negative bacteria. Remarkably, the T2SS secretes folded proteins, including multimeric assemblies such as cholera toxin and heat-labile enterotoxin from Vibrio cholerae and enterotoxigenic Escherichia coli, respectively. The major outer membrane T2SS protein is the {\textquoteright}secretin{\textquoteright} GspD. Cryo-EM reconstruction of the V. cholerae secretin at 19-{\r A} resolution reveals a dodecameric structure reminiscent of a barrel, with a large channel at its center that contains a closed periplasmic gate. The GspD periplasmic domain forms a vestibule with a conserved constriction, and it binds to a pentameric exoprotein and to the trimeric tip of the T2SS pseudopilus. By combining our results with structures of the cholera toxin and T2SS pseudopilus tip, we provide a structural basis for a possible secretion mechanism of the T2SS.

}, keywords = {Amino Acid, Amino Acid Sequence, Bacterial, Bacterial Outer Membrane Proteins, Cholera Toxin, Computer-Assisted, Conserved Sequence, Cryoelectron Microscopy, Fimbriae, Image Processing, Models, Molecular, Molecular Sequence Data, Periplasm, Protein Conformation, Protein Structure, Recombinant Fusion Proteins, Sequence Alignment, Sequence Homology, Surface Plasmon Resonance, Tertiary, Vibrio cholerae}, issn = {1545-9985}, doi = {10.1038/nsmb.1910}, author = {Reichow, Steve L and Korotkov, Konstantin V and Hol, Wim G J and Gonen, Tamir} } @article {6487, title = {Tetracycline-controlled transgene activation using the ROSA26-iM2-GFP knock-in mouse strain permits GFP monitoring of DOX-regulated transgene-expression.}, journal = {BMC Developmental Biology}, volume = {10}, year = {2010}, month = {2010}, pages = {95}, abstract = {

BACKGROUND: Conditional gene activation is an efficient strategy for studying gene function in genetically modified animals. Among the presently available gene switches, the tetracycline-regulated system has attracted considerable interest because of its unique potential for reversible and adjustable gene regulation. RESULTS: To investigate whether the ubiquitously expressed Gt(ROSA)26Sor locus enables uniform DOX-controlled gene expression, we inserted the improved tetracycline-regulated transcription activator iM2 together with an iM2 dependent GFP gene into the Gt(ROSA)26Sor locus, using gene targeting to generate ROSA26-iM2-GFP (R26t1Δ) mice. Despite the presence of ROSA26 promoter driven iM2, R26t1Δ mice showed very sparse DOX-activated expression of different iM2-responsive reporter genes in the brain, mosaic expression in peripheral tissues and more prominent expression in erythroid, myeloid and lymphoid lineages, in hematopoietic stem and progenitor cells and in olfactory neurons. CONCLUSIONS: The finding that gene regulation by the DOX-activated transcriptional factor iM2 in the Gt(ROSA)26Sor locus has its limitations is of importance for future experimental strategies involving transgene activation from the endogenous ROSA26 promoter. Furthermore, our ROSA26-iM2 knock-in mouse model (R26t1Δ) represents a useful tool for implementing gene function in vivo especially under circumstances requiring the side-by-side comparison of gene manipulated and wild type cells. Since the ROSA26-iM2 mouse allows mosaic gene activation in peripheral tissues and haematopoietic cells, this model will be very useful for uncovering previously unknown or unsuspected phenotypes.

}, keywords = {Animals, Doxycycline, Gene Knock-In Techniques, Genetic Techniques, Green Fluorescent Proteins, Mice, Tetracycline, Transgenes, Transgenic}, issn = {1471-213X}, doi = {10.1186/1471-213X-10-95}, author = {W{\"o}rtge, Simone and Eshkind, Leonid and Cabezas-Wallscheid, Nina and Lakaye, Bernard and Kim, Jinhyun and Heck, Rosario and Abassi, Yasmin and Diken, Mustafa and Sprengel, Rolf and Bockamp, Ernesto} } @article {6509, title = {Toward the second generation of optogenetic tools.}, journal = {The Journal of Neuroscience: The Official Journal of the Society for Neuroscience}, volume = {30}, year = {2010}, month = {2010 Nov 10}, pages = {14998-5004}, abstract = {

This mini-symposium aims to provide an integrated perspective on recent developments in optogenetics. Research in this emerging field combines optical methods with targeted expression of genetically encoded, protein-based probes to achieve experimental manipulation and measurement of neural systems with superior temporal and spatial resolution. The essential components of the optogenetic toolbox consist of two kinds of molecular devices: actuators and reporters, which respectively enable light-mediated control or monitoring of molecular processes. The first generation of genetically encoded calcium reporters, fluorescent proteins, and neural activators has already had a great impact on neuroscience. Now, a second generation of voltage reporters, neural silencers, and functionally extended fluorescent proteins hold great promise for continuing this revolution. In this review, we will evaluate and highlight the limitations of presently available optogenic tools and discuss where these technologies and their applications are headed in the future.

}, keywords = {Fluorescent Dyes, Genetic Engineering, Neurosciences}, issn = {1529-2401}, doi = {10.1523/JNEUROSCI.4190-10.2010}, author = {Kn{\"o}pfel, Thomas and Lin, Michael Z and Levskaya, Anselm and Tian, Lin and Lin, John Y and Boyden, Edward S} } @article {4064, title = {Towards automated high-throughput screening of C. elegans on agar.}, journal = {Arxiv}, year = {2010}, abstract = {

High-throughput screening (HTS) using model organisms is a promising method to identify a small number of genes or drugs potentially relevant to human biology or disease. In HTS experiments, robots and computers do a significant portion of the experimental work. However, one remaining major bottleneck is the manual analysis of experimental results, which is commonly in the form of microscopy images. This manual inspection is labor intensive, slow and subjective. Here we report our progress towards applying computer vision and machine learning methods to analyze HTS experiments that use Caenorhabditis elegans (C. elegans) worms grown on agar. Our main contribution is a robust segmentation algorithm for separating the worms from the background using brightfield images. We also show that by combining the output of this segmentation algorithm with an algorithm to detect the fluorescent dye, Nile Red, we can reliably distinguish different fluorescence-based phenotypes even though the visual differences are subtle. The accuracy of our method is similar to that of expert human analysts. This new capability is a significant step towards fully automated HTS experiments using C. elegans.

}, author = {Kabra, Mayank and Conery, Annie L. and O{\textquoteright}Rourke, Eyleen J. and Xie, Xin and Ljosa. Vebjorn and Jones, Thouis R. and Ausubel, Frederick M. and Ruvkun, Gary and Carpenter, Anne E. and Freund, Yoav} } @inbook {2558, title = {Using Drosophila to reveal insight into protein misfolding disease.}, booktitle = {Protein Misfolding Diseases: Current and Emerging Principles and Therapies}, year = {2010}, author = {Bilen, Julide and Bonini, Nancy M}, editor = {Ramirez- Alvarado, M and Kelly, J and Dobson, C} } @article {2329, title = {Vibrissa-based object localization in head-fixed mice.}, journal = {The Journal of Neuroscience}, volume = {30}, year = {2010}, month = {2010 Feb 3}, pages = {1947-67}, abstract = {

Linking activity in specific cell types with perception, cognition, and action, requires quantitative behavioral experiments in genetic model systems such as the mouse. In head-fixed primates, the combination of precise stimulus control, monitoring of motor output, and physiological recordings over large numbers of trials are the foundation on which many conceptually rich and quantitative studies have been built. Choice-based, quantitative behavioral paradigms for head-fixed mice have not been described previously. Here, we report a somatosensory absolute object localization task for head-fixed mice. Mice actively used their mystacial vibrissae (whiskers) to sense the location of a vertical pole presented to one side of the head and reported with licking whether the pole was in a target (go) or a distracter (no-go) location. Mice performed hundreds of trials with high performance (\>90\% correct) and localized to \<0.95 mm (\<6 degrees of azimuthal angle). Learning occurred over 1-2 weeks and was observed both within and across sessions. Mice could perform object localization with single whiskers. Silencing barrel cortex abolished performance to chance levels. We measured whisker movement and shape for thousands of trials. Mice moved their whiskers in a highly directed, asymmetric manner, focusing on the target location. Translation of the base of the whiskers along the face contributed substantially to whisker movements. Mice tended to maximize contact with the go (rewarded) stimulus while minimizing contact with the no-go stimulus. We conjecture that this may amplify differences in evoked neural activity between trial types.

}, keywords = {Animals, Exploratory Behavior, Head Movements, Inbred C57BL, Mice, Movement, Physical, Restraint, Somatosensory Cortex, Space Perception, Touch, Vibrissae}, issn = {1529-2401}, doi = {10.1523/JNEUROSCI.3762-09.2010}, author = {O{\textquoteright}Connor, Daniel H and Clack, Nathan G and Huber, Daniel and Komiyama, Takaki and Myers, Eugene W and Svoboda, Karel} } @proceedings {65006, title = {A wireless neural/EMG telemetry system for freely moving insects.}, year = {2010}, month = {2010 May 30}, abstract = {

We have developed a miniature telemetry system that captures neural, EMG, and acceleration signals from a freely moving insect and transmits the data wirelessly to a remote digital receiver. The system is based on a custom low-power integrated circuit that amplifies and digitizes four biopotential signals as well as three acceleration signals from an off-chip MEMS accelerometer, and transmits this information over a wireless 920-MHz telemetry link. The unit weighs 0.79 g and runs for two hours on two small batteries. We have used this system to monitor neural and EMG signals in jumping and flying locusts.

}, doi = {10.1109/ISCAS.2010.5538034}, author = {Reid R. Harrison and Ryan J. Kier and Anthony Leonardo and Haleh Fotowat and Raymond Chan and Fabrizio Gabbiani} } @article {2149, title = {A 3D digital atlas of C. elegans and its application to single-cell analyses.}, journal = {Nature Methods}, volume = {6}, year = {2009}, month = {2009 Sep}, pages = {667-72}, abstract = {

We built a digital nuclear atlas of the newly hatched, first larval stage (L1) of the wild-type hermaphrodite of Caenorhabditis elegans at single-cell resolution from confocal image stacks of 15 individual worms. The atlas quantifies the stereotypy of nuclear locations and provides other statistics on the spatial patterns of the 357 nuclei that could be faithfully segmented and annotated out of the 558 present at this developmental stage. We then developed an automated approach to assign cell names to each nucleus in a three-dimensional image of an L1 worm. We achieved 86\% accuracy in identifying the 357 nuclei automatically. This computational method will allow high-throughput single-cell analyses of the post-embryonic worm, such as gene expression analysis, or ablation or stimulation of cells under computer control in a high-throughput functional screen.

}, keywords = {Animals, Automatic Data Processing, Biological, Caenorhabditis elegans, Cell Lineage, Cell Nucleus, Models}, issn = {1548-7105}, doi = {10.1007/s12021-010-9090-x}, author = {Long, Fuhui and Peng, Hanchuan and Liu, Xiao and Kim, Stuart K and Myers, Eugene} } @article {2697, title = {Analysis of cell fate from single-cell gene expression profiles in C. elegans.}, journal = {Cell}, volume = {139}, year = {2009}, month = {2009 Oct 30}, pages = {623-33}, abstract = {

The C. elegans cell lineage provides a unique opportunity to look at how cell lineage affects patterns of gene expression. We developed an automatic cell lineage analyzer that converts high-resolution images of worms into a data table showing fluorescence expression with single-cell resolution. We generated expression profiles of 93 genes in 363 specific cells from L1 stage larvae and found that cells with identical fates can be formed by different gene regulatory pathways. Molecular signatures identified repeating cell fate modules within the cell lineage and enabled the generation of a molecular differentiation map that reveals points in the cell lineage when developmental fates of daughter cells begin to diverge. These results demonstrate insights that become possible using computational approaches to analyze quantitative expression from many genes in parallel using a digital gene expression atlas.

}, keywords = {Animals, Caenorhabditis elegans, Caenorhabditis elegans Proteins, Cell Differentiation, Cell Lineage, Gene Expression Profiling}, issn = {1097-4172}, doi = {10.1016/j.cell.2009.08.044}, author = {Liu, Xiao and Long, Fuhui and Peng, Hanchuan and Aerni, Sarah J and Jiang, Min and S{\'a}nchez-Blanco, Adolfo and Murray, John I and Preston, Elicia and Mericle, Barbara and Batzoglou, Serafim and Myers, Eugene W and Kim, Stuart K} } @article {48169, title = {An articulated statistical shape model for accurate hip joint segmentation.}, journal = {Conference proceedings : ... Annual International Conference of the IEEE Engineering in Medicine and Biology Society. IEEE Engineering in Medicine and Biology Society. Annual Conference}, volume = {2009}, year = {2009}, month = {2009}, pages = {6345-51}, abstract = {

In this paper we propose a framework for fully automatic, robust and accurate segmentation of the human pelvis and proximal femur in CT data. We propose a composite statistical shape model of femur and pelvis with a flexible hip joint, for which we extend the common definition of statistical shape models as well as the common strategy for their adaptation. We do not analyze the joint flexibility statistically, but model it explicitly by rotational parameters describing the bent in a ball-and-socket joint. A leave-one-out evaluation on 50 CT volumes shows that image driven adaptation of our composite shape model robustly produces accurate segmentations of both proximal femur and pelvis. As a second contribution, we evaluate a fine grain multi-object segmentation method based on graph optimization. It relies on accurate initializations of femur and pelvis, which our composite shape model can generate. Simultaneous optimization of both femur and pelvis yields more accurate results than separate optimizations of each structure. Shape model adaptation and graph based optimization are embedded in a fully automatic framework.

}, keywords = {Algorithms, Artificial Intelligence, Computer Simulation, Hip Joint, Imaging, Three-Dimensional, Models, Anatomic, Models, Statistical, Pattern Recognition, Automated, Radiographic Image Enhancement, Radiographic Image Interpretation, Computer-Assisted, Reproducibility of Results, Sensitivity and Specificity, Tomography, X-Ray Computed}, issn = {1557-170X}, doi = {10.1109/IEMBS.2009.5333269}, author = {Kainmueller, Dagmar and Lamecker, Hans and Zachow, Stefan and Hege, Hans-Christian} } @article {48176, title = {Automatic Extraction of Anatomical Landmarks From Medical Image Data: An Evaluation of Different Methods}, year = {2009}, abstract = {

This work presents three different methods for automatic detection of anatomical landmarks in CT data, namely for the left and right anterior superior iliac spines and the pubic symphysis. The methods exhibit different degrees of generality in terms of portability to other anatomical landmarks and require a different amount of training data. The srst method is problem-specisc and is based on the convex hull of the pelvis. Method two is a more generic approach based on a statistical shape model including the landmarks of interest for every training shape. With our third method we present the most generic approach, where only a small set of training landmarks is required. Those landmarks are transferred to the patient specisc geometry based on Mean Value Coordinates (MVCs). The methods work on surfaces of the pelvis that need to be extracted beforehand. We perform this geometry reconstruction with our previously introduced fully automatic segmentation framework for the pelvic bones. With a focus on the accuracy of our novel MVC-based approach, we evaluate and compare our methods on 100 clinical CT datasets, for which gold standard landmarks were desned manually by multiple observers.

}, author = {Kainmueller, Dagmar and Hans-Christian Hege and Heiko Seim and Markus Heller and Stefan Zachow} } @article {48174, title = {Automatic extraction of mandibular nerve and bone from cone-beam CT data.}, journal = {Medical image computing and computer-assisted intervention : MICCAI ... International Conference on Medical Image Computing and Computer-Assisted Intervention}, volume = {12}, year = {2009}, month = {2009}, pages = {76-83}, abstract = {

The exact localization of the mandibular nerve with respect to the bone is important for applications in dental implantology and maxillofacial surgery. Cone beam computed tomography (CBCT), often also called digital volume tomography (DVT), is increasingly utilized in maxillofacial or dental imaging. Compared to conventional CT, however, soft tissue discrimination is worse due to a reduced dose. Thus, small structures like the alveolar nerves are even harder recognizable within the image data. We show that it is nonetheless possible to accurately reconstruct the 3D bone surface and the course of the nerve in a fully automatic fashion, with a method that is based on a combined statistical shape model of the nerve and the bone and a Dijkstra-based optimization procedure. Our method has been validated on 106 clinical datasets: the average reconstruction error for the bone is 0.5 +/- 0.1 mm, and the nerve can be detected with an average error of 1.0 +/- 0.6 mm.

}, keywords = {Algorithms, Artificial Intelligence, Cone-Beam Computed Tomography, Humans, Mandible, Mandibular Nerve, Pattern Recognition, Automated, Radiographic Image Enhancement, Radiographic Image Interpretation, Computer-Assisted, Reproducibility of Results, Sensitivity and Specificity, Subtraction Technique}, author = {Kainmueller, Dagmar and Lamecker, Hans and Seim, Heiko and Zinser, Max and Zachow, Stefan} } @article {1495, title = {Automatic tracking of Escherichia coli in phase-contrast microscopy video.}, journal = {IEEE Transactions on Bio-Medical Engineering}, volume = {56}, year = {2009}, month = {2009 Feb}, pages = {390-9}, abstract = {

In this paper, we present an automatic method for estimating the trajectories of Escherichia coli bacteria from in vivo phase-contrast microscopy. To address the low-contrast boundaries in cellular images, an adaptive kernel-based technique is applied to detect cells in each frame. In addition to intensity features, region homogeneity measure and class uncertainty are also applied in this detection technique. To track cells with complex motion, a novel matching gain measure is introduced to cope with the challenges, particularly the presence of low-contrast boundary, the variations of appearance, and the frequent overlapping and occlusion. For multicell tracking over time, an optimal matching strategy is introduced to improve the handling of cell collision and broken trajectories. The results of successful tracking of Escherichia coli from various phase-contrast sequences are reported and compared with manually determined trajectories, as well as those obtained from existing tracking schemes. The stability of the algorithm with different parameter values is also analyzed and discussed.

}, keywords = {Algorithms, Artificial Intelligence, Automated, Chemotaxis, Computer-Assisted, Entropy, Escherichia coli, Image Processing, Microscopy, Pattern Recognition, Phase-Contrast, Video}, issn = {1558-2531}, doi = {10.1109/TBME.2008.2005956}, author = {Xie, Jun and Khan, Shahid and Shah, Mubarak} } @article {46421, title = {Caenorhabditis elegans innexins regulate active zone differentiation.}, journal = {The Journal of Neuroscience}, volume = {29}, year = {2009}, month = {2009 Apr 22}, pages = {5207-17}, abstract = {

In a genetic screen for active zone defective mutants in Caenorhabditis elegans, we isolated a loss-of-function allele of unc-7, a gene encoding an innexin/pannexin family gap junction protein. Innexin UNC-7 regulates the size and distribution of active zones at C. elegans neuromuscular junctions. Loss-of-function mutations in another innexin, UNC-9, cause similar active zone defects as unc-7 mutants. In addition to presumptive gap junction localizations, both UNC-7 and UNC-9 are also localized perisynaptically throughout development and required in presynaptic neurons to regulate active zone differentiation. Our mosaic analyses, electron microscopy, as well as expression studies suggest a novel and likely nonjunctional role of specific innexins in active zone differentiation in addition to gap junction formations.

}, keywords = {Animals, Caenorhabditis elegans, Caenorhabditis elegans Proteins, Cell Differentiation, Connexins, Gap Junctions, Gene Expression Regulation, Developmental, Male, Membrane Proteins, Organisms, Genetically Modified, Presynaptic Terminals}, issn = {1529-2401}, doi = {10.1523/JNEUROSCI.0637-09.2009}, author = {Yeh, Edward and Kawano, Taizo and Ng, Sharon and Fetter, Richard and Hung, Wesley and Wang, Ying and Zhen, Mei} } @article {48171, title = {Comparison and evaluation of methods for liver segmentation from CT datasets.}, journal = {IEEE transactions on medical imaging}, volume = {28}, year = {2009}, month = {2009 Aug}, pages = {1251-65}, abstract = {

This paper presents a comparison study between 10 automatic and six interactive methods for liver segmentation from contrast-enhanced CT images. It is based on results from the "MICCAI 2007 Grand Challenge" workshop, where 16 teams evaluated their algorithms on a common database. A collection of 20 clinical images with reference segmentations was provided to train and tune algorithms in advance. Participants were also allowed to use additional proprietary training data for that purpose. All teams then had to apply their methods to 10 test datasets and submit the obtained results. Employed algorithms include statistical shape models, atlas registration, level-sets, graph-cuts and rule-based systems. All results were compared to reference segmentations five error measures that highlight different aspects of segmentation accuracy. All measures were combined according to a specific scoring system relating the obtained values to human expert variability. In general, interactive methods reached higher average scores than automatic approaches and featured a better consistency of segmentation quality. However, the best automatic methods (mainly based on statistical shape models with some additional free deformation) could compete well on the majority of test images. The study provides an insight in performance of different segmentation approaches under real-world conditions and highlights achievements and limitations of current image analysis techniques.

}, keywords = {Algorithms, Bayes Theorem, Databases, Factual, Humans, Image Processing, Computer-Assisted, Liver, Tomography, X-Ray Computed}, issn = {1558-254X}, doi = {10.1109/TMI.2009.2013851}, author = {Heimann, Tobias and van Ginneken, Bram and Styner, Martin A and Arzhaeva, Yulia and Aurich, Volker and Bauer, Christian and Beck, Andreas and Becker, Christoph and Beichel, Reinhard and Bekes, Gy{\"o}rgy and Bello, Fernando and Binnig, Gerd and Bischof, Horst and Bornik, Alexander and Cashman, Peter M M and Chi, Ying and Cordova, Andr{\'e}s and Dawant, Benoit M and Fidrich, M{\'a}rta and Furst, Jacob D and Furukawa, Daisuke and Grenacher, Lars and Hornegger, Joachim and Kainm{\"u}ller, Dagmar and Kitney, Richard I and Kobatake, Hidefumi and Lamecker, Hans and Lange, Thomas and Lee, Jeongjin and Lennon, Brian and Li, Rui and Li, Senhu and Meinzer, Hans-Peter and Nemeth, G{\'a}bor and Raicu, Daniela S and Rau, Anne-Mareike and van Rikxoort, Eva M and Rousson, Mika{\"e}l and Rusko, L{\'a}szl{\'o} and Saddi, Kinda A and Schmidt, G{\"u}nter and Seghers, Dieter and Shimizu, Akinobu and Slagmolen, Pieter and Sorantin, Erich and Soza, Grzegorz and Susomboon, Ruchaneewan and Waite, Jonathan M and Wimmer, Andreas and Wolf, Ivo} } @article {1577, title = {Correlated mutation analyses on super-family alignments reveal functionally important residues.}, journal = {Proteins}, volume = {76}, year = {2009}, month = {2009 Aug 15}, pages = {608-16}, abstract = {

Correlated mutation analyses (CMA) on multiple sequence alignments are widely used for the prediction of the function of amino acids. The accuracy of CMA-based predictions is mainly determined by the number of sequences, by their evolutionary distances, and by the quality of the alignments. These criteria are best met in structure-based sequence alignments of large super-families. So far, CMA-techniques have mainly been employed to study the receptor interactions. The present work shows how a novel CMA tool, called Comulator, can be used to determine networks of functionally related residues in enzymes. These analyses provide leads for protein engineering studies that are directed towards modification of enzyme specificity or activity. As proof of concept, Comulator has been applied to four enzyme super-families: the isocitrate lyase/phoshoenol-pyruvate mutase super-family, the hexokinase super-family, the RmlC-like cupin super-family, and the FAD-linked oxidases super-family. In each of those cases networks of functionally related residue positions were discovered that upon mutation influenced enzyme specificity and/or activity as predicted. We conclude that CMA is a powerful tool for redesigning enzyme activity and selectivity.

}, keywords = {Algorithms, Computational Biology, Glucose-6-Phosphate Isomerase, Hexokinase, Isocitrate Lyase, Models, Molecular, Mutagenesis, Oxidoreductases, Oxidoreductases Acting on CH-CH Group Donors, Protein Structure, Secondary, Software}, issn = {1097-0134}, doi = {10.1002/prot.22374}, author = {Kuipers, Remko K P and Joosten, Henk-Jan and Verwiel, Eugene and Paans, Sjoerd and Akerboom, Jasper and van der Oost, John and Leferink, Nicole G H and van Berkel, Willem J H and Vriend, Gert and Schaap, Peter J} } @article {2910, title = {A critical role for N-WASp in cell migration during central nervous system development.}, journal = {International Journal of Developmental Neuroscience}, volume = {26}, year = {2009}, month = {2008}, pages = {413}, author = {Jin, FJ and Bharioke, A and Zhang, JY and Kuhlmann, TK and Georgiou, JG and Lommel, SL and Siminovitch, KS} } @article {6499, title = {Development of an implanted intramuscular EMG-triggered FES system for ambulation after incomplete spinal cord injury.}, journal = {Conference Proceedings: Annual International Conference of the IEEE Engineering in Medicine and Biology Society. IEEE Engineering in Medicine and Biology Society.}, volume = {2009}, year = {2009}, month = {2009}, pages = {6793-7}, abstract = {

Ambulation after spinal cord injury is possible with the aid of neuroprosthesis employing functional electrical stimulation (FES). Individuals with incomplete spinal cord injury (iSCI) retain partial volitional control of muscles below the level of injury, necessitating careful integration of FES with intact voluntary motor function for efficient walking. In this study, the intramuscular electromyogram (iEMG) was used to detect the intent to step and trigger FES-assisted walking in a volunteer with iSCI via an implanted neuroprosthesis consisting of two channels of bipolar iEMG signal acquisition and 12 independent channels of stimulation. The detection was performed with two types of classifiers- a threshold-based classifier that compared the running mean of the iEMG with a discrimination threshold to generate the trigger and a pattern recognition classifier that compared the time-history of the iEMG with a specified template of activity to generate the trigger whenever the cross-correlation coefficient exceeded a discrimination threshold. The pattern recognition classifier generally outperformed the threshold-based classifier, particularly with respect to minimizing False Positive triggers. The overall True Positive rates for the threshold-based classifier were 61.6\% and 87.2\% for the right and left steps with overall False Positive rates of 38.4\% and 33.3\%. The overall True Positive rates for the left and right step with the pattern recognition classifier were 57.2\% and 93.3\% and the overall False Positive rates were 11.9\% and 24.4\%. The subject showed no preference for either the threshold or pattern recognition-based classifier as determined by the Usability Rating Scale (URS) score collected after each trial and both the classifiers were perceived as moderately easy to use.

}, keywords = {Automated, Electric Stimulation, Electrodes, Electromyography, Humans, Implanted, Male, Muscles, Pattern Recognition, Recovery of Function, Spinal Cord Injuries, Walking}, issn = {1557-170X}, doi = {10.1109/IEMBS.2009.5333980}, author = {Dutta, Anirban and Kobetic, Rudi and Triolo, Ronald} } @article {47906, title = {Distinct pose of discodermolide in taxol binding pocket drives a complementary mode of microtubule stabilization.}, journal = {Biochemistry}, volume = {48}, year = {2009}, month = {2009 Dec 15}, pages = {11664-77}, abstract = {

The microtubule cytoskeleton has proven to be an effective target for cancer therapeutics. One class of drugs, known as microtubule stabilizing agents (MSAs), binds to microtubule polymers and stabilizes them against depolymerization. The prototype of this group of drugs, Taxol, is an effective chemotherapeutic agent used extensively in the treatment of human ovarian, breast, and lung carcinomas. Although electron crystallography and photoaffinity labeling experiments determined that the binding site for Taxol is in a hydrophobic pocket in beta-tubulin, little was known about the effects of this drug on the conformation of the entire microtubule. A recent study from our laboratory utilizing hydrogen-deuterium exchange (HDX) in concert with various mass spectrometry (MS) techniques has provided new information on the structure of microtubules upon Taxol binding. In the current study we apply this technique to determine the binding mode and the conformational effects on chicken erythrocyte tubulin (CET) of another MSA, discodermolide, whose synthetic analogues may have potential use in the clinic. We confirmed that, like Taxol, discodermolide binds to the taxane binding pocket in beta-tubulin. However, as opposed to Taxol, which has major interactions with the M-loop, discodermolide orients itself away from this loop and toward the N-terminal H1-S2 loop. Additionally, discodermolide stabilizes microtubules mainly via its effects on interdimer contacts, specifically on the alpha-tubulin side, and to a lesser extent on interprotofilament contacts between adjacent beta-tubulin subunits. Also, our results indicate complementary stabilizing effects of Taxol and discodermolide on the microtubules, which may explain the synergy observed between the two drugs in vivo.

}, keywords = {Alkanes, Animals, Binding Sites, Carbamates, Cattle, Chickens, Deuterium Exchange Measurement, Dimerization, Drug Synergism, Lactones, Microtubules, Paclitaxel, Peptide Fragments, Protein Binding, Protein Isoforms, Protein Stability, Pyrones, Tubulin, Tubulin Modulators}, issn = {1520-4995}, doi = {10.1021/bi901351q}, author = {Khrapunovich-Baine, Marina and Menon, Vilas and Verdier-Pinard, Pascal and Smith, Amos B and Angeletti, Ruth Hogue and Fiser, Andras and Horwitz, Susan Band and Xiao, Hui} } @article {46623, title = {Drosophila, a genetic model system to study cocaine-related behaviors: a review with focus on LIM-only proteins.}, journal = {Neuropharmacology}, volume = {56 Suppl 1}, year = {2009}, month = {2009}, pages = {97-106}, abstract = {

In the last decade, the fruit fly Drosophila melanogaster, highly accessible to genetic, behavioral and molecular analyses, has been introduced as a novel model organism to help decipher the complex genetic, neurochemical, and neuroanatomical underpinnings of behaviors induced by drugs of abuse. Here we review these data, focusing specifically on cocaine-related behaviors. Several of cocaine{\textquoteright}s most characteristic properties have been recapitulated in Drosophila. First, cocaine induces motor behaviors in flies that are remarkably similar to those observed in mammals. Second, repeated cocaine administration induces behavioral sensitization a form of behavioral plasticity believed to underlie certain aspects of addiction. Third, a key role for dopaminergic systems in mediating cocaine{\textquoteright}s effects has been demonstrated through both pharmacological and genetic methods. Finally, and most importantly, unbiased genetic screens, feasible because of the simplicity and scale with which flies can be manipulated in the laboratory, have identified several novel genes and pathways whose role in cocaine behaviors had not been anticipated. Many of these genes and pathways have been validated in mammalian models of drug addiction. We focus in this review on the role of LIM-only proteins in cocaine-induced behaviors.

}, keywords = {Animals, Animals, Genetically Modified, Behavior, Animal, Cocaine, Cocaine-Related Disorders, Disease Models, Animal, Dopamine Uptake Inhibitors, Drosophila melanogaster, Drosophila Proteins, Homeodomain Proteins, Models, Genetic}, issn = {0028-3908}, doi = {10.1016/j.neuropharm.2008.07.023}, author = {Heberlein, Ulrike and Tsai, Linus T-Y and Kapfhamer, David and Lasek, Amy W} } @article {5239, title = {Enzyme stabilization by domain insertion into a thermophilic protein.}, journal = {Protein Engineering, Design \& Selection : PEDS}, volume = {22}, year = {2009}, month = {2009 Oct}, pages = {615-23}, abstract = {

Insufficient kinetic stability of exoinulinase (EI) restricts its application in many areas including enzymatic transformation of inulin for production of ultra-high fructose syrup and oligofructan, as well as fermentation of inulin into bioethanol. The conventional method for enzyme stabilization involves mutagenesis and therefore risks alteration of an enzyme{\textquoteright}s desired properties, such as activity. Here, we report a novel method for stabilization of EI without any modification of its primary sequence. Our method employs domain insertion of an entire EI domain into a thermophilic scaffold protein. Insertion of EI into a loop of a thermophilic maltodextrin-binding protein from Pyrococcus furiosus (PfMBP) resulted in improvement of kinetic stability (the duration over which an enzyme remains active) at 37 degrees C without any compromise in EI activity. Our analysis suggests that the improved kinetic stability at 37 degrees C might originate from a raised kinetic barrier for irreversible conversion of unfolded intermediates to completely inactivated species, rather than an increased energy difference between the folded and unfolded forms.

}, keywords = {Aspergillus, Bacterial Proteins, Carrier Proteins, Enzyme Stability, Fungal Proteins, Glycoside Hydrolases, Kinetics, Models, Molecular, Protein Engineering, Protein Folding, Protein Structure, Pyrococcus, Recombinant Fusion Proteins, Temperature, Tertiary, Urea}, issn = {1741-0134}, doi = {10.1093/protein/gzp044}, author = {Kim, Chung-Sei and Pierre, Brennal and Ostermeier, Marc and Looger, Loren L and Kim, Jin Ryoun} } @article {1684, title = {Evolution of mutational robustness in the yeast genome: a link to essential genes and meiotic recombination hotspots.}, journal = {PLoS Genetics}, volume = {5}, year = {2009}, month = {2009 Jun}, pages = {e1000533}, abstract = {

Deleterious mutations inevitably emerge in any evolutionary process and are speculated to decisively influence the structure of the genome. Meiosis, which is thought to play a major role in handling mutations on the population level, recombines chromosomes via non-randomly distributed hot spots for meiotic recombination. In many genomes, various types of genetic elements are distributed in patterns that are currently not well understood. In particular, important (essential) genes are arranged in clusters, which often cannot be explained by a functional relationship of the involved genes. Here we show by computer simulation that essential gene (EG) clustering provides a fitness benefit in handling deleterious mutations in sexual populations with variable levels of inbreeding and outbreeding. We find that recessive lethal mutations enforce a selective pressure towards clustered genome architectures. Our simulations correctly predict (i) the evolution of non-random distributions of meiotic crossovers, (ii) the genome-wide anti-correlation of meiotic crossovers and EG clustering, (iii) the evolution of EG enrichment in pericentromeric regions and (iv) the associated absence of meiotic crossovers (cold centromeres). Our results furthermore predict optimal crossover rates for yeast chromosomes, which match the experimentally determined rates. Using a Saccharomyces cerevisiae conditional mutator strain, we show that haploid lethal phenotypes result predominantly from mutation of single loci and generally do not impair mating, which leads to an accumulation of mutational load following meiosis and mating. We hypothesize that purging of deleterious mutations in essential genes constitutes an important factor driving meiotic crossover. Therefore, the increased robustness of populations to deleterious mutations, which arises from clustered genome architectures, may provide a significant selective force shaping crossover distribution. Our analysis reveals a new aspect of the evolution of genome architectures that complements insights about molecular constraints, such as the interference of pericentromeric crossovers with chromosome segregation.

}, keywords = {Chromosomes, Essential, Evolution, Fungal, Genes, Genetic, Genome, Meiosis, Molecular, Mutation, Recombination, Saccharomyces cerevisiae}, issn = {1553-7404}, doi = {10.1371/journal.pgen.1000533}, author = {Keller, Philipp J and Knop, Michael} } @article {1116, title = {Exploring the pharmacokinetic properties of phosphorus-containing selective HDAC 1 and 2 inhibitors (SHI-1:2).}, journal = {Bioorganic \& Medicinal Chemistry Letters}, volume = {19}, year = {2009}, month = {2009 Apr 1}, pages = {2053-8}, abstract = {

We report the preparation and structure-activity relationships of phosphorus-containing histone deacetylase inhibitors. A strong trend between decreasing phosphorus functional group size and superior mouse pharmacokinetic properties was identified. In addition, optimized candidates showed tumor growth inhibition in xenograft studies.

}, keywords = {Administration, Animals, Antineoplastic Agents, Enzyme Inhibitors, Heterologous, Histone Deacetylase 1, Histone Deacetylase 2, Histone Deacetylase Inhibitors, Histone Deacetylases, Mice, Nude, Oral, Phosphonic Acids, Repressor Proteins, Transplantation}, issn = {1464-3405}, doi = {10.1016/j.bmcl.2009.02.009}, author = {Heidebrecht, Richard W and Chenard, Melissa and Close, Joshua and Dahlberg, William K and Fleming, Judith and Grimm, Jonathan B and Hamill, Julie E and Harsch, Andreas and Haines, Brian B and Hughes, Bethany and Kral, Astrid M and Middleton, Richard E and Mushti, Chandrasekhar and Ozerova, Nicole and Szewczak, Alexander A and Wang, Hongmei and Wilson, Kevin and Witter, David J and Secrist, J Paul and Miller, Thomas A} } @article {1091, title = {Fluorogenic affinity label for the facile, rapid imaging of proteins in live cells.}, journal = {Organic \& Biomolecular Chemistry}, volume = {7}, year = {2009}, month = {2009 Oct 7}, pages = {3969-75}, abstract = {

Haloalkane dehalogenase (HD) catalyzes the hydrolysis of haloalkanes via a covalent enzyme-substrate intermediate. Fusing a target protein to an HD variant that cannot hydrolyze the intermediate enables labeling of the target protein with a haloalkane in cellulo. The utility of extant probes is hampered, however, by background fluorescence as well as limited membrane permeability. Here, we report on the synthesis and use of a fluorogenic affinity label that, after unmasking by an intracellular esterase, labels an HD variant in cellulo. Labeling is rapid and specific, as expected from the reliance upon enzymic catalysts and the high membrane permeance of the probe both before and after unmasking. Most notably, even high concentrations of the fluorogenic affinity label cause minimal background fluorescence without a need to wash the cells. We envision that such fluorogenic affinity labels, which enlist catalysis by two cellular enzymes, will find utility in pulse-chase experiments, high-content screening, and numerous other protocols.

}, keywords = {Affinity Labels, Animals, Cell Line, Cell Survival, Fluorescent Dyes, Hydrolases, Hydrophobic and Hydrophilic Interactions, Time Factors}, issn = {1477-0539}, doi = {10.1039/b907664f}, author = {Watkins, Rex W and Lavis, Luke D and Kung, Vanessa M and Los, Georgyi V and Raines, Ronald T} } @article {68816, title = {Functional Role of a Specialized Class of Spinal Commissural Inhibitory Neurons during Fast Escapes in Zebrafish}, journal = {Journal of Neuroscience}, volume = {29}, year = {2009}, month = {05/2009}, pages = {6780{\textendash}6793}, abstract = {

In teleost fish, the Mauthner (M) cell, a large reticulospinal neuron in the brainstem, triggers escape behavior. Spinal commissural inhibitory interneurons that are electrotonically excited by the M-axon have been identified, but the behavioral roles of these neurons have not yet been addressed. Here, we studied these neurons, named CoLo (commissural local), in larval zebrafish using an enhancer-trap line in which the entire population of CoLos was visualized by green fluorescent protein. CoLos were present at one cell per hemi-segment. Electrophysiological recordings showed that an M-spike evoked a spike in CoLos via electrotonic transmission and that CoLos made monosynaptic inhibitory connections onto contralateral primary motoneurons, consistent with the results in adult goldfish. We further showed that CoLos were active only during escapes. We examined the behavioral roles of CoLos by investigating escape behaviors in CoLo-ablated larvae. The results showed that the escape behaviors evoked by sound/vibration stimuli were often impaired with a reduced initial bend of the body, indicating that CoLos play important roles in initiating escapes. We obtained several lines of evidence that strongly suggested that the impaired escapes occurred during bilateral activation of the M-cells: in normal larvae, CoLo-mediated inhibitory circuits enable animals to perform escapes even in these occasions by silencing the output of the slightly delayed firing of the second M-cell. This study illustrates (1) a clear example of the behavioral role of a specialized class of interneurons and (2) the capacity of the spinal circuits to filter descending commands and thereby produce the appropriate behavior.

}, issn = {0270-6474}, doi = {10.1523/JNEUROSCI.0801-09.2009}, url = {https://www.jneurosci.org/content/29/21/6780}, author = {Chie Satou and Yukiko Kimura and Tsunehiko Kohashi and Kazuki Horikawa and Hiroyuki Takeda and Yoichi Oda and Shin-ichi Higashijima} } @article {46621, title = {Genetic aspects of behavioral neurotoxicology.}, journal = {Neurotoxicology}, volume = {30}, year = {2009}, month = {2009 Sep}, pages = {741-53}, abstract = {

Considerable progress has been made over the past couple of decades concerning the molecular bases of neurobehavioral function and dysfunction. The field of neurobehavioral genetics is becoming mature. Genetic factors contributing to neurologic diseases such as Alzheimer{\textquoteright}s disease have been found and evidence for genetic factors contributing to other diseases such as schizophrenia and autism are likely. This genetic approach can also benefit the field of behavioral neurotoxicology. It is clear that there is substantial heterogeneity of response with behavioral impairments resulting from neurotoxicants. Many factors contribute to differential sensitivity, but it is likely that genetic variability plays a prominent role. Important discoveries concerning genetics and behavioral neurotoxicity are being made on a broad front from work with invertebrate and piscine mutant models to classic mouse knockout models and human epidemiologic studies of polymorphisms. Discovering genetic factors of susceptibility to neurobehavioral toxicity not only helps identify those at special risk, it also advances our understanding of the mechanisms by which toxicants impair neurobehavioral function in the larger population. This symposium organized by Edward Levin and Annette Kirshner, brought together researchers from the laboratories of Michael Aschner, Douglas Ruden, Ulrike Heberlein, Edward Levin and Kathleen Welsh-Bohmer conducting studies with Caenorhabditis elegans, Drosophila, fish, rodents and humans studies to determine the role of genetic factors in susceptibility to behavioral impairment from neurotoxic exposure.

}, keywords = {Animals, Behavior, Disease Models, Animal, Genetics, Humans, Neurotoxicity Syndromes, Neurotoxins, Phylogeny, Toxicology}, issn = {1872-9711}, doi = {10.1016/j.neuro.2009.07.014}, author = {Levin, Edward D and Aschner, Michael and Heberlein, Ulrike and Ruden, Douglas and Welsh-Bohmer, Kathleen A and Bartlett, Selena and Berger, Karen and Chen, Lang and Corl, Ammon B and Eddins, Donnie and French, Rachael and Hayden, Kathleen M and Helmcke, Kirsten and Hirsch, Helmut V B and Linney, Elwood and Lnenicka, Greg and Page, Grier P and Possidente, Debra and Possidente, Bernard and Kirshner, Annette} } @article {4277, title = {Infernal 1.0: inference of RNA alignments.}, journal = {Bioinformatics}, volume = {25}, year = {2009}, month = {2009 May 15}, pages = {1335-7}, abstract = {

SUMMARY: INFERNAL builds consensus RNA secondary structure profiles called covariance models (CMs), and uses them to search nucleic acid sequence databases for homologous RNAs, or to create new sequence- and structure-based multiple sequence alignments. AVAILABILITY: Source code, documentation and benchmark downloadable from http://infernal.janelia.org. INFERNAL is freely licensed under the GNU GPLv3 and should be portable to any POSIX-compliant operating system, including Linux and Mac OS/X.

}, keywords = {Databases, Nucleic Acid, Nucleic Acid Conformation, RNA, Sequence Alignment, Sequence Analysis, Software}, issn = {1367-4811}, doi = {10.1093/bioinformatics/btp157}, author = {Nawrocki, Eric P and Kolbe, Diana L and Eddy, Sean R.} } @article {927, title = {Interferometric fluorescent super-resolution microscopy resolves 3D cellular ultrastructure.}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {106}, year = {2009}, month = {2009 Mar 3}, pages = {3125-30}, abstract = {

Understanding molecular-scale architecture of cells requires determination of 3D locations of specific proteins with accuracy matching their nanometer-length scale. Existing electron and light microscopy techniques are limited either in molecular specificity or resolution. Here, we introduce interferometric photoactivated localization microscopy (iPALM), the combination of photoactivated localization microscopy with single-photon, simultaneous multiphase interferometry that provides sub-20-nm 3D protein localization with optimal molecular specificity. We demonstrate measurement of the 25-nm microtubule diameter, resolve the dorsal and ventral plasma membranes, and visualize the arrangement of integrin receptors within endoplasmic reticulum and adhesion complexes, 3D protein organization previously resolved only by electron microscopy. iPALM thus closes the gap between electron tomography and light microscopy, enabling both molecular specification and resolution of cellular nanoarchitecture.

}, keywords = {Animals, Cell Line, Cercopithecus aethiops, Fluorescence, Humans, Imaging, Interference, Microscopy, Microtubules, Three-Dimensional}, issn = {1091-6490}, doi = {10.1073/pnas.0813131106}, author = {Shtengel, Gleb and Galbraith, James A and Galbraith, Catherine G and Lippincott-Schwartz, Jennifer and Gillette, Jennifer M and Manley, Suliana and Sougrat, Rachid and Waterman, Clare M and Kanchanawong, Pakorn and Davidson, Michael W and Fetter, Richard D and Hess, Harald F} } @article {6360, title = {Knockdown of parhyale ultrabithorax recapitulates evolutionary changes in crustacean appendage morphology.}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {106}, year = {2009}, month = {2009 Aug 18}, pages = {13892-6}, abstract = {

Crustaceans possess remarkably diverse appendages, both between segments of a single individual as well as between species. Previous studies in a wide range of crustaceans have demonstrated a correlation between the anterior expression boundary of the homeotic (Hox) gene Ultrabithorax (Ubx) and the location and number of specialized thoracic feeding appendages, called maxillipeds. Given that Hox genes regulate regional identity in organisms as diverse as mice and flies, these observations in crustaceans led to the hypothesis that Ubx expression regulates the number of maxillipeds and that evolutionary changes in Ubx expression have generated various aspects of crustacean appendage diversity. Specifically, evolutionary changes in the expression boundary of Ubx have resulted in crustacean species with either 0, 1, 2, or 3 pairs of thoracic maxillipeds. Here we test this hypothesis by altering the expression of Ubx in Parhyale hawaiensis, a crustacean that normally possesses a single pair of maxillipeds. By reducing Ubx expression, we can generate Parhyale with additional maxillipeds in a pattern reminiscent of that seen in other crustacean species, and these morphological alterations are maintained as the animals molt and mature. These results provide critical evidence supporting the proposition that changes in Ubx expression have played a role in generating crustacean appendage diversity and lend general insights into the mechanisms of morphological evolution.

}, keywords = {Animals, Base Sequence, Biological, Biological Evolution, Cloning, Crustacea, Electron, Extremities, Gene Expression Regulation, Genes, Genetic Techniques, Homeobox, Homeodomain Proteins, In Situ Hybridization, Microscopy, Models, Molecular, Molecular Sequence Data, RNA, Scanning, Small Interfering}, issn = {1091-6490}, doi = {10.1073/pnas.0903105106}, author = {Liubicich, Danielle M and Serano, Julia M and Pavlopoulos, Anastasios and Kontarakis, Zacharias and Protas, Meredith E and Kwan, Elaine and Chatterjee, Sandip and Tran, Khoa D and Averof, Michalis and Patel, Nipam H} } @article {5243, title = {Local RNA structure alignment with incomplete sequence.}, journal = {Bioinformatics}, volume = {25}, year = {2009}, month = {2009 May 15}, pages = {1236-43}, abstract = {

Accuracy of automated structural RNA alignment is improved by using models that consider not only primary sequence but also secondary structure information. However, current RNA structural alignment approaches tend to perform poorly on incomplete sequence fragments, such as single reads from metagenomic environmental surveys, because nucleotides that are expected to be base paired are missing.

}, keywords = {Algorithms, Databases, Genetic, Models, Molecular, Nucleic Acid Conformation, RNA, Sequence Alignment, Sequence Analysis}, issn = {1367-4811}, doi = {10.1093/bioinformatics/btp154}, author = {Kolbe, Diana L and Eddy, Sean R.} } @article {48183, title = {Multi-object segmentation of head bones.}, journal = {MIDAS Journal}, year = {2009}, month = {2009 Jul 20}, abstract = {

We present a fully automatic method for 3D segmentation of the mandibular bone from CT data. The method includes an adaptation of statistical shape models of the mandible, the skull base and the midfacial bones, followed by a simultaneous graph-based optimization of adjacent deformable models. The adaptation of the models to the image data is performed according to a heuristic model of the typical intensity distribution in the vincinity of the bone boundary, with special focus on an accurate discrimination of adjacent bones in joint regions. An evaluation of our method based on 18 CT scans shows that a manual correction of the automatic segmentations is not necessary in approx. 60\% of the axial slices that contain the mandible.

}, url = {http://hdl.handle.net/10380/3099}, author = {Kainmueller, Dagmar and Lamecker, Hans and Seim, Heiko and Zachow, Stefan} } @article {48182, title = {Multi-object segmentation with coupled deformable models.}, journal = {Annals of the British Machine Vision Association}, volume = {2009}, year = {2009}, month = {2009}, pages = {1-10}, abstract = {

For biomechanical simulations, the segmentation of multiple adjacent anatomical struc- tures from medical image data is often required. If adjacent structures are barely dis- tinguishable in image data, in general automatic segmentation methods for single struc- tures do not yield sufficiently accurate results. To improve segmentation accuracy in these cases, knowledge about adjacent structures must be exploited. Optimal graph searching (graph cuts) based on deformable surface models allows for a simultaneous segmentation of multiple adjacent objects. However, this method requires a correspon- dence relation between vertices of adjacent surface meshes. Line segments, each con- taining two corresponding vertices, may then serve as shared displacement directions in the segmentation process. In this paper we propose a scheme for constructing a corre- spondence relation in adjacent regions of two arbitrary surfaces. This correspondence relation implies shared displacement directions that we apply for segmentation with de- formable surfaces. Here, overlap of the surfaces is guaranteed not to occur. We show correspondence relations for regions on a femoral head and acetabulum and other adja- cent structures, as well as an evaluation of segmentation results on 50 ct images of the hip joint.\ 

}, url = {http://www.bmva.org/annals/2009/2009-0005.pdf}, author = {Kainmueller, Dagmar and Lamecker, Hans and Zachow, Stefan} } @article {2687, title = {Neurotoxic effects induced by the Drosophila amyloid-beta peptide suggest a conserved toxic function.}, journal = {Neurobiology of Disease}, volume = {33}, year = {2009}, month = {2009 Feb}, pages = {274-81}, abstract = {

The accumulation of amyloid-beta (Abeta) into plaques is a hallmark feature of Alzheimer{\textquoteright}s disease (AD). While amyloid precursor protein (APP)-related proteins are found in most organisms, only Abeta fragments from human APP have been shown to induce amyloid deposits and progressive neurodegeneration. Therefore, it was suggested that neurotoxic effects are a specific property of human Abeta. Here we show that Abeta fragments derived from the Drosophila orthologue APPL aggregate into intracellular fibrils, amyloid deposits, and cause age-dependent behavioral deficits and neurodegeneration. We also show that APPL can be cleaved by a novel fly beta-secretase-like enzyme. This suggests that Abeta-induced neurotoxicity is a conserved function of APP proteins whereby the lack of conservation in the primary sequence indicates that secondary structural aspects determine their pathogenesis. In addition, we found that the behavioral phenotypes precede extracellular amyloid deposit formation, supporting results that intracellular Abeta plays a key role in AD.

}, keywords = {Aging, Amino Acid Sequence, Amyloid, Amyloid beta-Protein Precursor, Amyloid Precursor Protein Secretases, Animal, Animals, Apoptosis, Behavior, Blotting, Brain, Cell Surface, Drosophila melanogaster, Drosophila Proteins, Electron, Gene Expression, Immunohistochemistry, Light, Membrane Proteins, Microscopy, Molecular Sequence Data, Nerve Degeneration, Nerve Tissue Proteins, Neurons, Peptide Fragments, Protease Nexins, Receptors, Western}, issn = {1095-953X}, doi = {10.1016/j.nbd.2008.10.014}, author = {Carmine-Simmen, Katia and Proctor, Thomas and Tsch{\"a}pe, Jakob and Poeck, Burkhard and Triphan, Tilman and Strauss, Roland and Kretzschmar, Doris} } @article {4430, title = {Nonlinear integration of visual and haltere inputs in fly neck motor neurons.}, journal = {The Journal of Neuroscience: The Official Journal of the Society for Neuroscience}, volume = {29}, year = {2009}, month = {2009 Oct 21}, pages = {13097-105}, abstract = {

Animals use information from multiple sensory organs to generate appropriate behavior. Exactly how these different sensory inputs are fused at the motor system is not well understood. Here we study how fly neck motor neurons integrate information from two well characterized sensory systems: visual information from the compound eye and gyroscopic information from the mechanosensory halteres. Extracellular recordings reveal that a subpopulation of neck motor neurons display "gating-like" behavior: they do not fire action potentials in response to visual stimuli alone but will do so if the halteres are coactivated. Intracellular recordings show that these motor neurons receive small, sustained subthreshold visual inputs in addition to larger inputs that are phase locked to haltere movements. Our results suggest that the nonlinear gating-like effect results from summation of these two inputs with the action potential threshold providing the nonlinearity. As a result of this summation, the sustained visual depolarization is transformed into a temporally structured train of action potentials synchronized to the haltere beating movements. This simple mechanism efficiently fuses two different sensory signals and may also explain the context-dependent effects of visual inputs on fly behavior.

}, keywords = {Action Potentials, Age Factors, Animals, Diptera, Female, Functional Laterality, Motor Neurons, Neck, Nonlinear Dynamics, Patch-Clamp Techniques, Photic Stimulation, Sense Organs, Sensory Gating, Touch, Visual Pathways}, issn = {1529-2401}, doi = {10.1523/JNEUROSCI.2915-09.2009}, author = {Huston, Stephen J and Krapp, Holger G} } @article {2501, title = {The nuclear receptor E75A has a novel pair-rule-like function in patterning the milkweed bug, Oncopeltus fasciatus.}, journal = {Developmental Biology}, volume = {334}, year = {2009}, month = {2009 Oct 1}, pages = {300-10}, abstract = {

Genetic studies of the fruit fly Drosophila have revealed a hierarchy of segmentation genes (maternal, gap, pair-rule and HOX) that subdivide the syncytial blastoderm into sequentially finer-scale coordinates. Within this hierarchy, the pair-rule genes translate gradients of information into periodic stripes of expression. How pair-rule genes function during the progressive mode of segmentation seen in short and intermediate-germ insects is an ongoing question. Here we report that the nuclear receptor Of{\textquoteright}E75A is expressed with double segment periodicity in the head and thorax. In the abdomen, Of{\textquoteright}E75A is expressed in a unique pattern during posterior elongation, and briefly resembles a sequence that is typical of pair-rule genes. Depletion of Of{\textquoteright}E75A mRNA caused loss of a subset of odd-numbered parasegments, as well as parasegment 6. Because these parasegments straddle segment boundaries, we observe fusions between adjacent segments. Finally, expression of Of{\textquoteright}E75A in the blastoderm requires even-skipped, which is a gap gene in Oncopeltus. These data show that the function of Of{\textquoteright}E75A during embryogenesis shares many properties with canonical pair-rule genes in other insects. They further suggest that parasegment specification may occur through irregular and episodic pair-rule-like activity.

}, keywords = {Animals, Blastoderm, Body Patterning, Cytoplasmic and Nuclear, Embryo, Hemiptera, Insect Proteins, Nonmammalian, Receptors, RNA Interference}, issn = {1095-564X}, doi = {10.1016/j.ydbio.2009.06.038}, author = {Erezyilmaz, Deniz F and Kelstrup, Hans C and Riddiford, Lynn M} } @article {5486, title = {Oviposition preference for and positional avoidance of acetic acid provide a model for competing behavioral drives in Drosophila.}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {106}, year = {2009}, month = {2009 Jul 7}, pages = {11352-7}, abstract = {

Selection of appropriate oviposition sites is essential for progeny survival and fitness in generalist insect species, such as Drosophila melanogaster, yet little is known about the mechanisms regulating how environmental conditions and innate adult preferences are evaluated and balanced to yield the final substrate choice for egg-deposition. Female D. melanogaster are attracted to food containing acetic acid (AA) as an oviposition substrate. However, our observations reveal that this egg-laying preference is a complex process, as it directly opposes an otherwise strong, default behavior of positional avoidance for the same food. We show that 2 distinct sensory modalities detect AA. Attraction to AA-containing food for the purpose of egg-laying relies on the gustatory system, while positional repulsion depends primarily on the olfactory system. Similarly, distinct central brain regions are involved in AA attraction and repulsion. Given this unique situation, in which a single environmental stimulus yields 2 opposing behavioral outputs, we propose that the interaction of egg-laying attraction and positional aversion for AA provides a powerful model for studying how organisms balance competing behavioral drives and integrate signals involved in choice-like processes.

}, keywords = {Acetic Acid, Animal, Animal Feed, Animals, Avoidance Learning, Biological, Brain, Drosophila melanogaster, Female, Hydrogen-Ion Concentration, Models, Neurons, Olfactory Pathways, Oviposition, Sexual Behavior, Taste}, issn = {1091-6490}, doi = {10.1073/pnas.0901419106}, author = {Joseph, Ryan M and Devineni, Anita V and King, Ian F G and Heberlein, Ulrike} } @article {6359, title = {Probing the evolution of appendage specialization by Hox gene misexpression in an emerging model crustacean.}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {106}, year = {2009}, month = {2009 Aug 18}, pages = {13897-902}, abstract = {

Changes in the expression of Hox genes have been widely linked to the evolution of animal body plans, but functional demonstrations of this relationship have been impeded by the lack of suitable model organisms. A classic case study involves the repeated evolution of specialized feeding appendages, called maxillipeds, from anterior thoracic legs, in many crustacean lineages. These leg-to-maxilliped transformations correlate with the loss of Ultrabithorax (Ubx) expression from corresponding segments, which is proposed to be the underlying genetic cause. To functionally test this hypothesis, we establish tools for conditional misexpression and use these to misexpress Ubx in the crustacean Parhyale hawaiensis. Ectopic Ubx leads to homeotic transformations of anterior appendages toward more posterior thoracic fates, including maxilliped-to-leg transformations, confirming the capacity of Ubx to control thoracic (leg) versus gnathal (feeding) segmental identities. We find that maxillipeds not only are specified in the absence of Ubx, but also can develop in the presence of low/transient Ubx expression. Our findings suggest a path for the gradual evolutionary transition from thoracic legs to maxillipeds, in which stepwise changes in Hox gene expression have brought about this striking morphological and functional transformation.

}, keywords = {Animals, Cloning, Crustacea, Down-Regulation, Electron, Extremities, Gene Expression Regulation, Genes, Genetic, Genetically Modified, Heat-Shock Proteins, Homeobox, Homeodomain Proteins, Immunohistochemistry, Microscopy, Models, Molecular, Phenotype, Scanning, Transgenes}, issn = {1091-6490}, doi = {10.1073/pnas.0902804106}, author = {Pavlopoulos, Anastasios and Kontarakis, Zacharias and Liubicich, Danielle M and Serano, Julia M and Akam, Michael and Patel, Nipam H and Averof, Michalis} } @article {6491, title = {A proposal for a coordinated effort for the determination of brainwide neuroanatomical connectivity in model organisms at a mesoscopic scale.}, journal = {PLoS Computational Biology}, volume = {5}, year = {2009}, month = {2009 Mar}, pages = {e1000334}, abstract = {

In this era of complete genomes, our knowledge of neuroanatomical circuitry remains surprisingly sparse. Such knowledge is critical, however, for both basic and clinical research into brain function. Here we advocate for a concerted effort to fill this gap, through systematic, experimental mapping of neural circuits at a mesoscopic scale of resolution suitable for comprehensive, brainwide coverage, using injections of tracers or viral vectors. We detail the scientific and medical rationale and briefly review existing knowledge and experimental techniques. We define a set of desiderata, including brainwide coverage; validated and extensible experimental techniques suitable for standardization and automation; centralized, open-access data repository; compatibility with existing resources; and tractability with current informatics technology. We discuss a hypothetical but tractable plan for mouse, additional efforts for the macaque, and technique development for human. We estimate that the mouse connectivity project could be completed within five years with a comparatively modest budget.

}, keywords = {Animals, Brain, Databases, Factual, Humans, Macaca, Mice, Models, Nerve Net, Neuroanatomy, Neurological, Research Design}, issn = {1553-7358}, doi = {10.1371/journal.pcbi.1000334}, author = {Bohland, Jason W and Wu, Caizhi and Barbas, Helen and Bokil, Hemant and Bota, Mihail and Breiter, Hans C and Cline, Hollis T and Doyle, John C and Freed, Peter J and Greenspan, Ralph J and Haber, Suzanne N and Hawrylycz, Michael and Herrera, Daniel G and Hilgetag, Claus C and Huang, Z Josh and Jones, Allan and Jones, Edward G and Karten, Harvey J and Kleinfeld, David and K{\"o}tter, Rolf and Lester, Henry A and Lin, John M and Mensh, Brett D and Mikula, Shawn and Panksepp, Jaak and Price, Joseph L and Safdieh, Joseph and Saper, Clifford B and Schiff, Nicholas D and Schmahmann, Jeremy D and Stillman, Bruce W and Svoboda, Karel and Swanson, Larry W and Toga, Arthur W and Van Essen, David C and Watson, James D and Mitra, Partha P} } @article {5253, title = {Rapid functional maturation of nascent dendritic spines.}, journal = {Neuron}, volume = {61}, year = {2009}, month = {2009 Jan 29}, pages = {247-58}, abstract = {

Spine growth and retraction with synapse formation and elimination plays an important role in shaping brain circuits during development and in the adult brain, yet the temporal relationship between spine morphogenesis and the formation of functional synapses remains poorly defined. We imaged hippocampal pyramidal neurons to identify spines of different ages. We then used two-photon glutamate uncaging, whole-cell recording, and Ca(2+) imaging to analyze the properties of nascent spines and their older neighbors. New spines expressed glutamate-sensitive currents that were indistinguishable from mature spines of comparable volumes. Some spines exhibited negligible AMPA receptor-mediated responses, but the occurrence of these "silent" spines was uncorrelated with spine age. In contrast, NMDA receptor-mediated Ca(2+) accumulations were significantly lower in new spines. New spines reconstructed using electron microscopy made synapses. Our data support a model in which outgrowth and enlargement of nascent spines is tightly coupled to formation and maturation of glutamatergic synapses.

}, keywords = {AMPA, Animals, Calcium, Calcium Signaling, Cell Differentiation, Confocal, Dendritic Spines, Electron, Glutamic Acid, Hippocampus, Microscopy, Neuronal Plasticity, Organ Culture Techniques, Patch-Clamp Techniques, Pyramidal Cells, Rats, Receptors, Synapses, Synaptic Membranes, Synaptic Transmission, Time Factors, Transmission}, issn = {1097-4199}, doi = {10.1016/j.neuron.2008.10.054}, author = {Zito, Karen and Scheuss, Volker and Knott, Graham and Hill, Travis and Svoboda, Karel} } @article {4266, title = {Rfam: updates to the RNA families database.}, journal = {Nucleic Acids Research}, volume = {37}, year = {2009}, month = {2009 Jan}, pages = {D136-40}, abstract = {

Rfam is a collection of RNA sequence families, represented by multiple sequence alignments and covariance models (CMs). The primary aim of Rfam is to annotate new members of known RNA families on nucleotide sequences, particularly complete genomes, using sensitive BLAST filters in combination with CMs. A minority of families with a very broad taxonomic range (e.g. tRNA and rRNA) provide the majority of the sequence annotations, whilst the majority of Rfam families (e.g. snoRNAs and miRNAs) have a limited taxonomic range and provide a limited number of annotations. Recent improvements to the website, methodologies and data used by Rfam are discussed. Rfam is freely available on the Web at http://rfam.sanger.ac.uk/and http://rfam.janelia.org/.

}, keywords = {Computer Graphics, Databases, Internet, Nucleic Acid, RNA, Sequence Alignment, Sequence Analysis}, issn = {1362-4962}, doi = {10.1093/nar/gkn766}, author = {Gardner, Paul P and Daub, Jennifer and Tate, John G and Nawrocki, Eric P and Kolbe, Diana L and Lindgreen, Stinus and Wilkinson, Adam C and Finn, Robert D and Griffiths-Jones, Sam and Eddy, Sean R. and Bateman, Alex} } @article {6783, title = {A state-mutating genetic algorithm to design ion-channel models.}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {106}, year = {2009}, month = {2009 Sep 29}, pages = {16829-34}, abstract = {

Realistic computational models of single neurons require component ion channels that reproduce experimental findings. Here, a topology-mutating genetic algorithm that searches for the best state diagram and transition-rate parameters to model macroscopic ion-channel behavior is described. Important features of the algorithm include a topology-altering strategy, automatic satisfaction of equilibrium constraints (microscopic reversibility), and multiple-protocol fitting using sequential goal programming rather than explicit weighting. Application of this genetic algorithm to design a sodium-channel model exhibiting both fast and prolonged inactivation yields a six-state model that produces realistic activity-dependent attenuation of action-potential backpropagation in current-clamp simulations of a CA1 pyramidal neuron.

}, keywords = {Algorithms, Cell Membrane, Ion Channels, Models, Mutation, Neurons, Sodium Channels, Theoretical}, issn = {1091-6490}, doi = {10.1073/pnas.0903766106}, author = {Menon, Vilas and Spruston, Nelson and Kath, William L} } @article {5251, title = {Subcellular dynamics of type II PKA in neurons.}, journal = {Neuron}, volume = {62}, year = {2009}, month = {2009 May 14}, pages = {363-74}, abstract = {

Protein kinase A (PKA) plays multiple roles in neurons. The localization and specificity of PKA are largely controlled by A-kinase anchoring proteins (AKAPs). However, the dynamics of PKA in neurons and the roles of specific AKAPs are poorly understood. We imaged the distribution of type II PKA in hippocampal and cortical layer 2/3 pyramidal neurons in vitro and in vivo. PKA was concentrated in dendritic shafts compared to the soma, axons, and dendritic spines. This spatial distribution was imposed by the microtubule-binding protein MAP2, indicating that MAP2 is the dominant AKAP in neurons. Following cAMP elevation, catalytic subunits dissociated from the MAP2-tethered regulatory subunits and rapidly became enriched in nearby spines. The spatial gradient of type II PKA between dendritic shafts and spines was critical for the regulation of synaptic strength and long-term potentiation. Therefore, the localization and activity-dependent translocation of type II PKA are important determinants of PKA function.

}, keywords = {A Kinase Anchor Proteins, Animals, Cerebral Cortex, Cyclic AMP-Dependent Protein Kinases, Dendritic Spines, Hippocampus, Mice, Microtubule-Associated Proteins, Pyramidal Cells, Rats, Subcellular Fractions, Tissue Distribution, Transgenic}, issn = {1097-4199}, doi = {10.1016/j.neuron.2009.03.013}, author = {Zhong, Haining and Sia, Gek-Ming and Sato, Takashi R and Gray, Noah W and Mao, Tianyi and Khuchua, Zaza and Huganir, Richard L and Svoboda, Karel} } @article {2956, title = {Super-resolution video microscopy of live cells by structured illumination.}, journal = {Nature Methods}, volume = {6}, year = {2009}, month = {2009 May}, pages = {339-42}, abstract = {

Structured-illumination microscopy can double the resolution of the widefield fluorescence microscope but has previously been too slow for dynamic live imaging. Here we demonstrate a high-speed structured-illumination microscope that is capable of 100-nm resolution at frame rates up to 11 Hz for several hundred time points. We demonstrate the microscope by video imaging of tubulin and kinesin dynamics in living Drosophila melanogaster S2 cells in the total internal reflection mode.

}, keywords = {Algorithms, Animals, Automatic Data Processing, Cell Line, Computer-Assisted, Cytophotometry, Drosophila melanogaster, Fluorescence, Fourier Analysis, Image Processing, Kinesin, Lighting, Microscopy, Microtubules, Mitotic Spindle Apparatus, Tubulin, Video}, issn = {1548-7105}, doi = {10.1038/nmeth.1324}, author = {Kner, Peter and Chhun, Bryant B and Griffis, Eric R and Winoto, Lukman and Gustafsson, Mats G L} } @article {6784, title = {Synapse distribution suggests a two-stage model of dendritic integration in CA1 pyramidal neurons.}, journal = {Neuron}, volume = {63}, year = {2009}, month = {2009 Jul 30}, pages = {171-7}, abstract = {

Competing models have been proposed to explain how neurons integrate the thousands of inputs distributed throughout their dendritic trees. In a simple global integration model, inputs from all locations sum in the axon. In a two-stage integration model, inputs contribute directly to dendritic spikes, and outputs from multiple branches sum in the axon. These two models yield opposite predictions of how synapses at different dendritic locations should be scaled if they are to contribute equally to neuronal output. We used serial-section electron microscopy to reconstruct individual apical oblique dendritic branches of CA1 pyramidal neurons and observe a synapse distribution consistent with the two-stage integration model. Computational modeling suggests that the observed synapse distribution enhances the contribution of each dendritic branch to neuronal output.

}, keywords = {Animals, Dendrites, Electron, Hippocampus, Humans, Microscopy, Models, Neural Networks (Computer), Neural Pathways, Neurological, Neurons, Synapses}, issn = {1097-4199}, doi = {10.1016/j.neuron.2009.06.023}, author = {Katz, Yael and Menon, Vilas and Nicholson, Daniel A and Geinisman, Yuri and Kath, William L and Spruston, Nelson} } @article {6132, title = {Synaptic activity-responsive element in the Arc/Arg3.1 promoter essential for synapse-to-nucleus signaling in activated neurons.}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {106}, year = {2009}, month = {2009 Jan 6}, pages = {316-21}, abstract = {

The neuronal immediate early gene Arc/Arg-3.1 is widely used as one of the most reliable molecular markers for intense synaptic activity in vivo. However, the cis-acting elements responsible for such stringent activity dependence have not been firmly identified. Here we combined luciferase reporter assays in cultured cortical neurons and comparative genome mapping to identify the critical synaptic activity-responsive elements (SARE) of the Arc/Arg-3.1 gene. A major SARE was found as a unique approximately 100-bp element located at \>5 kb upstream of the Arc/Arg-3.1 transcription initiation site in the mouse genome. This single element, when positioned immediately upstream of a minimal promoter, was necessary and sufficient to replicate crucial properties of endogenous Arc/Arg-3.1{\textquoteright}s transcriptional regulation, including rapid onset of transcription triggered by synaptic activity and low basal expression during synaptic inactivity. We identified the major determinants of SARE as a unique cluster of neuronal activity-dependent cis-regulatory elements consisting of closely localized binding sites for CREB, MEF2, and SRF. Consistently, a SARE reporter could readily trace and mark an ensemble of cells that have experienced intense activity in the recent past in vivo. Taken together, our work uncovers a novel transcriptional mechanism by which a critical 100-bp element, SARE, mediates a predominant component of the synapse-to-nucleus signaling in ensembles of Arc/Arg-3.1-positive activated neurons.

}, keywords = {Animals, Binding Sites, Cell Nucleus, Cyclic AMP Response Element-Binding Protein, Cytoskeletal Proteins, Gene Expression Regulation, Genetic, Genomics, Mice, Myogenic Regulatory Factors, Nerve Tissue Proteins, Neurons, Nucleic Acid, Promoter Regions, Regulatory Sequences, Serum Response Factor, Signal Transduction, Synapses}, issn = {1091-6490}, doi = {10.1073/pnas.0806518106}, author = {Kawashima, Takashi and Okuno, Hiroyuki and Nonaka, Mio and Adachi-Morishima, Aki and Kyo, Nan and Okamura, Michiko and Takemoto-Kimura, Sayaka and Worley, Paul F and Bito, Haruhiko} } @article {46620, title = {Too fat to fly? New brain circuits regulate obesity in Drosophila.}, journal = {Neuron}, volume = {63}, year = {2009}, month = {2009 Aug 13}, pages = {279-81}, abstract = {

In mammals, fat store levels are regulated by brain centers that control food intake and metabolism. A new study by Al-Anzi and colleagues in this issue of Neuron identifies neurons with similar functions in Drosophila, further establishing the fly as a legitimate model to study obesity.

}, keywords = {Animals, Brain, Disease Models, Animal, Drosophila, Drosophila Proteins, Nerve Net, Neurons, Obesity}, issn = {1097-4199}, doi = {10.1016/j.neuron.2009.07.023}, author = {Kaun, Karla R and Heberlein, Ulrike} } @conference {6450, title = {Which spatial partition trees are adaptive to intrinsic dimension?}, booktitle = {Conference on Uncertainty in Artificial Intelligence}, year = {2009}, month = {2009 Jun 18}, author = {Verma, N and Kpotufe, S and Dasgupta, S} } @article {6536, title = {Wnt-Ror signaling to SIA and SIB neurons directs anterior axon guidance and nerve ring placement in C. elegans.}, journal = {Development}, volume = {136}, year = {2009}, month = {2009 Nov}, pages = {3801-10}, abstract = {

Wnt signaling through Frizzled proteins guides posterior cells and axons in C. elegans into different spatial domains. Here we demonstrate an essential role for Wnt signaling through Ror tyrosine kinase homologs in the most prominent anterior neuropil, the nerve ring. A genetic screen uncovered cwn-2, the C. elegans homolog of Wnt5, as a regulator of nerve ring placement. In cwn-2 mutants, all neuronal structures in and around the nerve ring are shifted to an abnormal anterior position. cwn-2 is required at the time of nerve ring formation; it is expressed by cells posterior of the nerve ring, but its precise site of expression is not critical for its function. In nerve ring development, cwn-2 acts primarily through the Wnt receptor CAM-1 (Ror), together with the Frizzled protein MIG-1, with parallel roles for the Frizzled protein CFZ-2. The identification of CAM-1 as a CWN-2 receptor contrasts with CAM-1 action as a non-receptor in other C. elegans Wnt pathways. Cell-specific rescue of cam-1 and cell ablation experiments reveal a crucial role for the SIA and SIB neurons in positioning the nerve ring, linking Wnt signaling to specific cells that organize the anterior nervous system.

}, keywords = {Animals, Axons, Caenorhabditis elegans, Caenorhabditis elegans Proteins, Neurogenesis, Neurons, Receptor Protein-Tyrosine Kinases, Receptor Tyrosine Kinase-like Orphan Receptors, Signal Transduction, Wnt Proteins}, issn = {1477-9129}, doi = {10.1242/dev.038109}, author = {Kennerdell, Jason R and Fetter, Richard D and Bargmann, Cornelia I} } @conference {6530, title = {Action recognition using ballistic dynamics.}, booktitle = {2008 IEEE Conference on Computer Vision and Pattern Recognition}, volume = {2008}, year = {2008}, month = {2008 Jun}, pages = {1-8}, publisher = {IEEE}, organization = {IEEE}, address = {Anchorage, AK}, abstract = {

We present a Bayesian framework for action recognition through ballistic dynamics. Psycho-kinesiological studies indicate that ballistic movements form the natural units for human movement planning. The framework leads to an efficient and robust algorithm for temporally segmenting videos into atomic movements. Individual movements are annotated with person-centric morphological labels called ballistic verbs. This is tested on a dataset of interactive movements, achieving high recognition rates. The approach is also applied on a gesture recognition task, improving a previously reported recognition rate from 84\% to 92\%. Consideration of ballistic dynamics enhances the performance of the popular Motion History Image feature. We also illustrate the approach{\textquoteright}s general utility on real-world videos. Experiments indicate that the method is robust to view, style and appearance variations.

}, isbn = {978-1-4244-2242-5}, issn = {1063-6919}, doi = {10.1109/CVPR.2008.4587806}, author = {Vitadalevuni, Shiv N. and Kellokumpu, Vili and Davis, Larry S.} } @conference {2719, title = {Automatic recognition of cells (ARC) for 3D images of C. elegans.}, booktitle = {Proceedings of the 2008 Conference on Computational Molecular Biology (RECOMB) (Singapore, 2008)}, year = {2008}, month = {2008 Apr 2}, abstract = {

The development of high-resolution microscopy makes possible the high-throughput screening of cellular information, such as gene expression at single cell resolution. One of the critical enabling techniques yet to be developed is the automatic recognition or annotation of specific cells in a 3D image stack. In this paper, we present a novel graph-based algorithm, ARC, that determines cell identities in a 3D confocal image of C. elegans based on their highly stereotyped arrangement. This is an essential step in our work on gene expression analysis of C. elegans at the resolution of single cells. Our ARC method integrates both the absolute and relative spatial locations of cells in a C. elegans body. It uses a marker-guided, spatially-constrained, two-stage bipartite matching to find the optimal match between cells in a subject image and cells in 15 template images that have been manually annotated and vetted. We applied ARC to the recognition of cells in 3D confocal images of the first larval stage (L1) of C. elegans hermaphrodites, and achieved an average accuracy of 94.91\%.

}, author = {Long, Fuhui and Peng, Hanchuan and Liu, X and Kim, Stuart K and Myers, Eugene} } @article {48177, title = {Automatic Segmentation of the Pelvic Bones from CT Data Based on a Statistical Shape Model}, year = {2008}, abstract = {

We present an algorithm for automatic segmentation of the human pelvic bones from CT datasets that is based on the application of a statistical shape model. The proposed method is divided into three steps: 1) The averaged shape of the pelvis model is initially placed within the CT data using the Generalized Hough Transform, 2) the statistical shape model is then adapted to the image data by a transformation and variation of its shape modes, and 3) a final free-form deformation step based on optimal graph searching is applied to overcome the restrictive character of the statistical shape representation. We thoroughly evaluated the method on 50 manually segmented CT datasets by performing a leave-one-out study. The Generalized Hough Transform proved to be a reliable method for an automatic initial placement of the shape model within the CT data. Compared to the manual gold standard segmentations, our automatic segmentation approach produced an average surface distance of 1.2 {\textpm} 0.3mm after the adaptation of the statistical shape model, which could be reduced to 0.7{\textpm}0.3mm using a final free-form deformation step. Together with an average segmentation time of less than 5 minutes, the results of our study indicate that our method meets the requirements of clinical routine.

}, doi = {10.2312/VCBM/VCBM08/093-100}, author = {Kainmueller, Dagmar and Seim, Heiko and Heller, Markus and Lamecker, Hans and Zachow, Stefan and Hege, Hans-Christian} } @article {1496, title = {Automatic tracking of Escherichia coli bacteria.}, journal = {Medical Image Computing and Computer-Assisted Intervention: MICCAI ... International Conference on Medical Image Computing and Computer-Assisted Intervention}, volume = {11}, year = {2008}, month = {2008 Sep 6}, pages = {824-32}, abstract = {

In this paper, we present an automatic method for estimating the trajectories of Escherichia coli bacteria from in vivo phase-contrast microscopy videos. To address the low-contrast boundaries in cellular images, an adaptive kernel-based technique is applied to detect cells in sequence of frames. Then a novel matching gain measure is introduced to cope with the challenges such as dramatic changes of cells{\textquoteright} appearance and serious overlapping and occlusion. For multiple cell tracking, an optimal matching strategy is proposed to improve the handling of cell collision and broken trajectories. The results of successful tracking of Escherichia coli from various phase-contrast sequences are reported and compared with manually-determined trajectories, as well as those obtained from existing tracking methods. The stability of the algorithm with different parameter values is also analyzed and discussed.

}, keywords = {Algorithms, Artificial Intelligence, Automated, Cell Movement, Cluster Analysis, Computer-Assisted, Escherichia coli, Image Enhancement, Image Interpretation, Microscopy, Pattern Recognition, Phase-Contrast, Reproducibility of Results, Sensitivity and Specificity, Subtraction Technique, Video}, author = {Xie, Jun and Khan, Shahid and Shah, Mubarak} } @article {48395, title = {Benchmarking implicit solvent folding simulations of the amyloid beta(10-35) fragment.}, journal = {The journal of physical chemistry. B}, volume = {112}, year = {2008}, month = {2008 May 15}, pages = {6175-86}, abstract = {

A pathogenetic feature of Alzhemier disease is the aggregation of monomeric beta-amyloid proteins (Abeta) to form oligomers. Usually these oligomers of long peptides aggregate on time scales of microseconds or longer, making computational studies using atomistic molecular dynamics models prohibitively expensive and making it essential to develop computational models that are cheaper and at the same time faithful to physical features of the process. We benchmark the ability of our implicit solvent model to describe equilibrium and dynamic properties of monomeric Abeta(10-35) using all-atom Langevin dynamics (LD) simulations, since Alphabeta(10-35) is the only fragment whose monomeric properties have been measured. The accuracy of the implicit solvent model is tested by comparing its predictions with experiment and with those from a new explicit water MD simulation, (performed using CHARMM and the TIP3P water model) which is approximately 200 times slower than the implicit water simulations. The dependence on force field is investigated by running multiple trajectories for Alphabeta(10-35) using the CHARMM, OPLS-aal, and GS-AMBER94 force fields, whereas the convergence to equilibrium is tested for each force field by beginning separate trajectories from the native NMR structure, a completely stretched structure, and from unfolded initial structures. The NMR order parameter, S2, is computed for each trajectory and is compared with experimental data to assess the best choice for treating aggregates of Alphabeta. The computed order parameters vary significantly with force field. Explicit and implicit solvent simulations using the CHARMM force fields display excellent agreement with each other and once again support the accuracy of the implicit solvent model. Alphabeta(10-35) exhibits great flexibility, consistent with experiment data for the monomer in solution, while maintaining a general strand-loop-strand motif with a solvent-exposed hydrophobic patch that is believed to be important for aggregation. Finally, equilibration of the peptide structure requires an implicit solvent LD simulation as long as 30 ns.

}, keywords = {Amyloid beta-Peptides, Computer Simulation, Diffusion, Magnetic Resonance Spectroscopy, Models, Molecular, Peptide Fragments, Protein Folding, Protein Structure, Tertiary, Solvents, Surface Properties, Water}, issn = {1520-6106}, doi = {10.1021/jp077099h}, author = {Kent, Andrew and Jha, Abhishek K and Fitzgerald, James E and Freed, Karl F} } @article {45930, title = {Compartmental neural simulations with spatial adaptivity.}, journal = {Journal of Computational Neuroscience}, volume = {25}, year = {2008}, month = {2008 Dec}, pages = {465-80}, abstract = {

Since their inception, computational models have become increasingly complex and useful counterparts to laboratory experiments within the field of neuroscience. Today several software programs exist to solve the underlying mathematical system of equations, but such programs typically solve these equations in all parts of a cell (or network of cells) simultaneously, regardless of whether or not all of the cell is active. This approach can be inefficient if only part of the cell is active and many simulations must be performed. We have previously developed a numerical method that provides a framework for spatial adaptivity by making the computations local to individual branches rather than entire cells (Rempe and Chopp, SIAM Journal on Scientific Computing, 28: 2139-2161, 2006). Once the computation is reduced to the level of branches instead of cells, spatial adaptivity is straightforward: the active regions of the cell are detected and computational effort is focused there, while saving computations in other regions of the cell that are at or near rest. Here we apply the adaptive method to four realistic neuronal simulation scenarios and demonstrate its improved efficiency over non-adaptive methods. We find that the computational cost of the method scales with the amount of activity present in the simulation, rather than the physical size of the system being simulated. For certain problems spatial adaptivity reduces the computation time by up to 80\%.

}, keywords = {Action Potentials, Adaptation, Biological, Algorithms, Animals, Computer Simulation, Dendrites, Gap Junctions, Hippocampus, Ion Channel Gating, Ion Channels, Models, Neurological, Nerve Net, Neural Networks (Computer), Pyramidal Cells, Signal Processing, Computer-Assisted}, issn = {1573-6873}, doi = {10.1007/s10827-008-0089-3}, author = {Rempe, Michael J and Spruston, Nelson and Kath, William L and Chopp, David L} } @article {45932, title = {Distribution of bursting neurons in the CA1 region and the subiculum of the rat hippocampus.}, journal = {Journal of Comparative Neurology}, volume = {506}, year = {2008}, month = {2008 Feb 1}, pages = {535-47}, abstract = {

We performed patch-clamp recordings from morphologically identified and anatomically mapped pyramidal neurons of the ventral hippocampus to test the hypothesis that bursting neurons are distributed on a gradient from the CA2/CA1 border (proximal) through the subiculum (distal), with more bursting observed at distal locations. We find that the well-defined morphological boundaries between the hippocampal subregions CA1 and subiculum do not correspond to abrupt changes in electrophysiological properties. Rather, we observed that the percentage of bursting neurons is linearly correlated with position in the proximal-distal axis across the CA1 and the subiculum, the percentages of bursting neurons being 10\% near the CA1-CA2 border, 24\% at the CA1-subiculum border, and higher than 50\% in the distal subiculum. The distribution of bursting neurons was paralleled by a gradient in afterdepolarization (ADP) amplitude. We also tested the hypothesis that there was an association between bursting and two previously described morphologically distinct groups of pyramidal neurons (twin and single apical dendrites) in the CA1 region. We found no difference in output mode between single and twin apical dendrite morphologies, which was consistent with the observation that the two morphologies were equally distributed across the transverse axis of the CA1 region. Taken together with the known organization of connections from CA3 to CA1 and CA1 to subiculum, our results indicate that bursting neurons are most likely to be connected to regular spiking neurons and vice versa.

}, keywords = {Action Potentials, Animals, Axons, Cell Shape, Dendrites, Hippocampus, Lysine, Neural Pathways, Organ Culture Techniques, Patch-Clamp Techniques, Pyramidal Cells, Rats, Rats, Wistar, Staining and Labeling}, issn = {0021-9967}, doi = {10.1002/cne.21564}, author = {Jarsky, Tim and Mady, Rina and Kennedy, Benjamin and Spruston, Nelson} } @article {46624, title = {Ethanol sensitivity and tolerance in long-term memory mutants of Drosophila melanogaster.}, journal = {Alcoholism, Clinical and Experimental Research}, volume = {32}, year = {2008}, month = {2008 May}, pages = {895-908}, abstract = {

BACKGROUND: It has become increasingly clear that molecular and neural mechanisms underlying learning and memory and drug addiction are largely shared. To confirm and extend these findings, we analyzed ethanol-responsive behaviors of a collection of Drosophila long-term memory mutants.

METHODS: For each mutant, sensitivity to the acute uncoordinating effects of ethanol was quantified using the inebriometer. Additionally, 2 distinct forms of ethanol tolerance were measured: rapid tolerance, which develops in response to a single brief exposure to a high concentration of ethanol vapor; and chronic tolerance, which develops following a sustained low-level exposure.

RESULTS: Several mutants were identified with altered sensitivity, rapid or chronic tolerance, while a number of mutants exhibited multiple defects.

CONCLUSIONS: The corresponding genes in these mutants represent areas of potential overlap between learning and memory and behavioral responses to alcohol. These genes also define components shared between different ethanol behavioral responses.

}, keywords = {Animals, Behavior, Animal, Drosophila melanogaster, Drug Tolerance, Ethanol, Learning, Memory}, issn = {1530-0277}, doi = {10.1111/j.1530-0277.2008.00659.x}, author = {Berger, Karen H and Kong, Eric C and Dubnau, Josh and Tully, Tim and Moore, Monica S and Heberlein, Ulrike} } @article {2012, title = {Fast monte carlo simulation methods for biological reaction-diffusion systems in solution and on surfaces.}, journal = {SIAM Journal on Scientific Computing: A Publication of the Society for Industrial and Applied Mathematics}, volume = {30}, year = {2008}, month = {2008 Oct 13}, pages = {3126}, abstract = {

Many important physiological processes operate at time and space scales far beyond those accessible to atom-realistic simulations, and yet discrete stochastic rather than continuum methods may best represent finite numbers of molecules interacting in complex cellular spaces. We describe and validate new tools and algorithms developed for a new version of the MCell simulation program (MCell3), which supports generalized Monte Carlo modeling of diffusion and chemical reaction in solution, on surfaces representing membranes, and combinations thereof. A new syntax for describing the spatial directionality of surface reactions is introduced, along with optimizations and algorithms that can substantially reduce computational costs (e.g., event scheduling, variable time and space steps). Examples for simple reactions in simple spaces are validated by comparison to analytic solutions. Thus we show how spatially realistic Monte Carlo simulations of biological systems can be far more cost-effective than often is assumed, and provide a level of accuracy and insight beyond that of continuum methods.

}, issn = {1095-7197}, doi = {10.1137/070692017}, author = {Kerr, Rex A and Bartol, Thomas M and Kaminsky, Boris and Dittrich, Markus and Chang, Jen-Chien Jack and Baden, Scott B and Sejnowski, Terrence J and Stiles, Joel R} } @article {2347, title = {Induction of muscle thermogenesis by high-fat diet in mice: association with obesity-resistance.}, journal = {American Journal of Physiology. Endocrinology and Metabolism}, volume = {295}, year = {2008}, month = {2008 Aug}, pages = {E356-67}, abstract = {

The obesogenic effect of a high-fat (HF) diet is counterbalanced by stimulation of energy expenditure and lipid oxidation in response to a meal. The aim of this study was to reveal whether muscle nonshivering thermogenesis could be stimulated by a HF diet, especially in obesity-resistant A/J compared with obesity-prone C57BL/6J (B/6J) mice. Experiments were performed on male mice born and maintained at 30 degrees C. Four-week-old mice were randomly weaned onto a low-fat (LF) or HF diet for 2 wk. In the A/J LF mice, cold exposure (4 degrees C) resulted in hypothermia, whereas the A/J HF, B/6J LF, and B/6J HF mice were cold tolerant. Cold sensitivity of the A/J LF mice was associated with a relatively low whole body energy expenditure under resting conditions, which was normalized by the HF diet. In both strains, the HF diet induced uncoupling protein-1-mediated thermogenesis, with a stronger induction in A/J mice. Only in A/J mice: 1) the HF diet augmented activation of whole body lipid oxidation by cold; and 2) at 30 degrees C, oxygen consumption, total content, and phosphorylation of AMP-activated protein kinase (AMPK), and AICAR-stimulated palmitate oxidation in soleus muscle was increased by the HF diet in parallel with significantly increased leptinemia. Gene expression data in soleus muscle of the A/J HF mice indicated a shift from carbohydrate to fatty acid oxidation. Our results suggest a role for muscle nonshivering thermogenesis and lipid oxidation in the obesity-resistant phenotype of A/J mice and indicate that a HF diet could induce thermogenesis in oxidative muscle, possibly via the leptin-AMPK axis.

}, keywords = {Aminoimidazole Carboxamide, AMP-Activated Protein Kinases, Animals, Basal Metabolism, Body Temperature, Body Weight, Calorimetry, Dietary Fats, Fatty Acids, Inbred C57BL, Indirect, Male, Mice, Multienzyme Complexes, Muscle, Newborn, Nonesterified, Obesity, Oxygen Consumption, Protein-Serine-Threonine Kinases, Random Allocation, Ribonucleotides, Skeletal, Thermogenesis, Triglycerides}, issn = {0193-1849}, doi = {10.1523/JNEUROSCI.3773-10.2011}, author = {Kus, Vladimir and Prazak, Tomas and Brauner, Petr and Hensler, Michal and Kuda, Ondrej and Flachs, Pavel and Janovska, Petra and Medrikova, Dasa and Rossmeisl, Martin and Jilkova, Zuzana and Stefl, Bohumir and Pastalkova, Eva and Drahota, Zdenek and Houstek, Josef and Kopecky, Jan} } @article {2503, title = {Insulin/IGF signaling regulates the change in commitment in imaginal discs and primordia by overriding the effect of juvenile hormone.}, journal = {Developmental Biology}, volume = {324}, year = {2008}, month = {2008 Dec 15}, pages = {258-65}, abstract = {

At the beginning of the final larval (fifth) instar of Manduca sexta, imaginal precursors including wing discs and eye primordia initiate metamorphic changes, such as pupal commitment, patterning and cell proliferation. Juvenile hormone (JH) prevents these changes in earlier instars and in starved final instar larvae, but nutrient intake overcomes this effect of JH in the latter. In this study, we show that a molecular marker of pupal commitment, broad, is up-regulated in the wing discs by feeding on sucrose or by bovine insulin or Manduca bombyxin in starved final instar larvae. This effect of insulin could not be prevented by JH. In vitro insulin had no effect on broad expression but relieved the suppression of broad expression by JH. This effect of insulin was directly on the disc as shown by its reduction in the presence of insulin receptor dsRNA. In starved penultimate fourth instar larvae, broad expression in the wing disc was not up-regulated by insulin. The discs became responsive to this action of insulin during the molt to the fifth instar together with the ability to become pupally committed in response to 20-hydroxyecdysone. Thus, the Manduca bombyxin acts as a metamorphosis-initiating factor in the imaginal precursors.

}, keywords = {Animals, Cells, Cloning, Cultured, Developmental, Ecdysterone, Gene Expression Regulation, Insect Proteins, Insulin, Juvenile Hormones, Larva, Manduca, Messenger, Molecular, Receptor, RNA, RNA Interference, Signal Transduction, Somatomedins, Sucrose, Wing}, issn = {1095-564X}, doi = {10.1016/j.ydbio.2008.09.017}, author = {Koyama, Takashi and Syropyatova, Maria O and Riddiford, Lynn M} } @article {1248, title = {Local potential connectivity in cat primary visual cortex.}, journal = {Cerebral Cortex}, volume = {18}, year = {2008}, month = {2008 Jan}, pages = {13-28}, abstract = {

Time invariant description of synaptic connectivity in cortical circuits may be precluded by the ongoing growth and retraction of dendritic spines accompanied by the formation and elimination of synapses. On the other hand, the spatial arrangement of axonal and dendritic branches appears stable. This suggests that an invariant description of connectivity can be cast in terms of potential synapses, which are locations in the neuropil where an axon branch of one neuron is proximal to a dendritic branch of another neuron. In this paper, we attempt to reconstruct the potential connectivity in local cortical circuits of the cat primary visual cortex (V1). Based on multiple single-neuron reconstructions of axonal and dendritic arbors in 3 dimensions, we evaluate the expected number of potential synapses and the probability of potential connectivity among excitatory (pyramidal and spiny stellate) neurons and inhibitory basket cells. The results provide a quantitative description of structural organization of local cortical circuits. For excitatory neurons from different cortical layers, we compute local domains, which contain their potentially pre- and postsynaptic excitatory partners. These domains have columnar shapes with laminar specific radii and are roughly of the size of the ocular dominance column. Therefore, connections between most excitatory neurons in the ocular dominance column can be implemented by local synaptogenesis. Structural connectivity involving inhibitory basket cells is generally weaker than excitatory connectivity. Here, only nearby neurons are capable of establishing more than one potential synapse, implying that within the ocular dominance column these connections have more limited potential for circuit remodeling.

}, keywords = {Animals, Cats, Cells, Cultured, Nerve Net, Neural Pathways, Synapses, Visual Cortex}, issn = {1460-2199}, doi = {10.1093/cercor/bhm027}, author = {Stepanyants, Armen and Hirsch, Judith A and Martinez, Luis M and Kisv{\'a}rday, Zolt{\'a}n F and Ferecsk{\'o}, Alex S and Chklovskii, Dmitri B} } @article {46626, title = {Loss of RAB-3/A in Caenorhabditis elegans and the mouse affects behavioral response to ethanol.}, journal = {Genes, Brain, and Behavior}, volume = {7}, year = {2008}, month = {2008 Aug}, pages = {669-76}, abstract = {

The mechanisms by which ethanol induces changes in behavior are not well understood. Here, we show that Caenorhabditis elegans loss-of-function mutations in the synaptic vesicle-associated RAB-3 protein and its guanosine triphosphate exchange factor AEX-3 confer resistance to the acute locomotor effects of ethanol. Similarly, mice lacking one or both copies of Rab3A are resistant to the ataxic and sedative effects of ethanol, and Rab3A haploinsufficiency increases voluntary ethanol consumption. These data suggest a conserved role of RAB-3-/RAB3A-regulated neurotransmitter release in ethanol-related behaviors.

}, keywords = {Alcohol Drinking, Alcohol-Induced Disorders, Nervous System, Animals, Ataxia, Behavior, Animal, Brain, Brain Chemistry, Caenorhabditis elegans, Caenorhabditis elegans Proteins, Central Nervous System Depressants, Consciousness Disorders, Drug Resistance, Ethanol, Haplotypes, Male, Mice, Mice, Inbred C57BL, Mice, Knockout, rab3 GTP-Binding Proteins, rab3A GTP-Binding Protein}, issn = {1601-183X}, doi = {10.1111/j.1601-183X.2008.00404.x}, author = {Kapfhamer, D and Bettinger, J C and Davies, A G and Eastman, C L and Smail, E A and Heberlein, U and McIntire, S L} } @conference {48184, title = {Model-based autosegmentation of the central brain of the honeybee, Apis mellifera, using active statistical shape models.}, booktitle = {Neuroinformatics 2008}, year = {2008}, month = {2008 Sep 07}, address = {Stockholm, Sweden}, abstract = {

The Honeybee Brain Atlas serves as 3D database and communicative platform to accumulate structural data, i.e. reconstructed neurons, derived from confocal scans (Brandt et al., 2005) (www.neurobiologie.fu-berlin.de/beebrain/) (1). Transforming neurons into the atlas requires manual segmentation of neuropils within confocal images, a time-consuming task requiring expertise in identifying biological structures which can result in different outcomes from various segmenters.

}, doi = {10.3389/conf.neuro.11.2008.01.064}, author = {Singer, Jochen and Lienhard, Matthias and Seim, Heiko and Kainmueller, Dagmar and Kuss, Anja and Lamecker, Hans and Zachow, Stefan and Menzel, Randolf and Rybak, J{\"u}rgen} } @article {2806, title = {Monitoring of toxic compounds in air using a handheld rectilinear ion trap mass spectrometer.}, journal = {Analytical Chemistry}, volume = {80}, year = {2008}, month = {2008 Feb 1}, pages = {734-41}, abstract = {

A miniature, handheld mass spectrometer, based on the rectilinear ion trap mass analyzer, has been applied to air monitoring for traces of toxic compounds. The instrument is battery-operated, hand-portable, and rugged. We anticipate its use in public safety, industrial hygiene, and environmental monitoring. Gaseous samples of nine toxic industrial compounds, phosgene, ethylene oxide, sulfur dioxide, acrylonitrile, cyanogen chloride, hydrogen cyanide, acrolein, formaldehyde, and ethyl parathion, were tested. A sorption trap inlet was constructed to serve as the interface between atmosphere and the vacuum chamber of the mass spectrometer. After selective collection of analytes on the sorbent bed, the sorbent tube was evacuated and then heated to desorb analyte into the instrument. Sampling, detection, identification, and quantitation of all compounds were readily achieved in times of less than 2 min, with detection limits ranging from 800 parts per trillion to 3 parts per million depending on the analyte. For all but one analyte, detection limits were well below (3.5-130 times below) permissible exposure limits. A linear dynamic range of 1-2 orders of magnitude was obtained over the concentration ranges studied (sub-ppbv to ppmv) for all analytes.

}, issn = {0003-2700}, doi = {10.1364/AO.50.001792}, author = {Keil, Adam and Hernandez-Soto, Heriberto and Noll, Robert J and Fico, Miriam and Gao, Liang and Ouyang, Zheng and Cooks, R Graham} } @proceedings {3300, title = {Multi-modal target tracking using heterogeneous sensor networks.}, year = {2008}, month = {2008 Aug 7}, abstract = {

The paper describes a target tracking system running on a Heterogeneous Sensor Network (HSN) and presents results gathered from a realistic deployment. The system fuses audio direction of arrival data from mote class devices and object detection measurements from embedded PCs equipped with cameras. The acoustic sensor nodes perform beamforming and measure the energy as a function of the angle. The camera nodes detect moving objects and estimate their angle. The sensor detections are sent to a centralized sensor fusion node via a combination of two wireless networks. The novelty of our system is the unique combination of target tracking methods customized for the application at hand and their implementation on an actual HSN platform.

}, author = {Kushwaha, M and Amundson, I and Volgyesi, P and Ahammad, Parvez and Simon, G and Koutsoukos, X and Ledeczi, A and Sastry, S} } @article {2397, title = {The nucleosome remodeling factor (NURF) regulates genes involved in Drosophila innate immunity.}, journal = {Developmental Biology}, volume = {316}, year = {2008}, month = {2008 Apr 15}, pages = {538-47}, abstract = {

The Drosophila nucleosome remodeling factor (NURF) is an ISWI-containing chromatin remodeling complex that catalyzes ATP-dependent nucleosome sliding. By sliding nucleosomes, NURF has the ability to alter chromatin structure and regulate transcription. Previous studies have shown that mutation of Drosophila NURF induces melanotic tumors, implicating NURF in innate immune function. Here, we show that NURF mutants exhibit identical innate immune responses to gain-of-function mutants in the Drosophila JAK/STAT pathway. Using microarrays, we identify a common set of target genes that are activated in both mutants. In silico analysis of promoter sequences of these defines a consensus regulatory element comprising a STAT-binding sequence overlapped by a binding-site for the transcriptional repressor Ken. NURF interacts physically and genetically with Ken. Chromatin immunoprecipitation (ChIP) localizes NURF to Ken-binding sites in hemocytes, suggesting that Ken recruits NURF to repress STAT responders. Loss of NURF leads to precocious activation of STAT target genes.

}, keywords = {Animals, Chromosomal Proteins, DNA-Binding Proteins, Drosophila, Drosophila Proteins, Gene Expression Profiling, Gene Expression Regulation, Genetic, Genome, Immunity, Innate, Non-Histone, Oligonucleotide Array Sequence Analysis, Phenotype, Recombinant Proteins, Reverse Transcriptase Polymerase Chain Reaction, RNA, Signal Transduction, Transcription}, issn = {1095-564X}, doi = {10.1073/pnas.1100640108}, author = {Kwon, So Yeon and Xiao, Hua and Glover, Bradley P and Tjian, Robert and Wu, Carl and Badenhorst, Paul} } @article {1115, title = {Phenylglycine and phenylalanine derivatives as potent and selective HDAC1 inhibitors (SHI-1).}, journal = {Bioorganic \& Medicinal Chemistry Letters}, volume = {18}, year = {2008}, month = {2008 Mar 15}, pages = {1859-63}, abstract = {

An HTS screening campaign identified a series of low molecular weight phenols that showed excellent selectivity (\>100-fold) for HDAC1/HDAC2 over other Class I and Class II HDACs. Evolution and optimization of this HTS hit series provided HDAC1-selective (SHI-1) compounds with excellent anti-proliferative activity and improved physical properties. Dose-dependent efficacy in a mouse HCT116 xenograft model was demonstrated with a phenylglycine SHI-1 analog.

}, keywords = {Acetylation, Amides, Animals, Cell Proliferation, Colonic Neoplasms, Cultured, Dogs, Enzyme Inhibitors, Ether-A-Go-Go Potassium Channels, Glycine, Histone Deacetylase 1, Histone Deacetylase Inhibitors, Humans, Macaca mulatta, Mice, Molecular Structure, Phenylalanine, Structure-Activity Relationship, Tumor Cells, Xenograft Model Antitumor Assays}, issn = {1464-3405}, doi = {10.1016/j.bmcl.2008.02.012}, author = {Wilson, Kevin J and Witter, David J and Grimm, Jonathan B and Siliphaivanh, Phieng and Otte, Karin M and Kral, Astrid M and Fleming, Judith C and Harsch, Andreas and Hamill, Julie E and Cruz, Jonathan C and Chenard, Melissa and Szewczak, Alexander A and Middleton, Richard E and Hughes, Bethany L and Dahlberg, William K and Secrist, J Paul and Miller, Thomas A} } @article {45962, title = {Polymorphic assemblies and crystalline arrays of lens tetraspanin MP20.}, journal = {Journal of Molecular Biology}, volume = {376}, year = {2008}, month = {2008 Feb 15}, pages = {380-92}, abstract = {

Members of the tetraspanin superfamily function as transmembrane scaffold proteins that mediate the assembly of membrane proteins into specific signaling complexes. Tetraspanins also interact with each other and concentrate membrane proteins into tetraspanin-enriched microdomains (TEMs). Here we report that lens-specific tetraspanin MP20 can form multiple types of higher-order assemblies and we present crystalline arrays of MP20. When isolated in the absence of divalent cations, MP20 is solubilized predominantly in tetrameric form, whereas the presence of divalent cations during solubilization promotes the association of MP20 tetramers into higher-order species. This effect only occurs when divalent cations are present during solubilization but not when divalent cations are added to solubilized tetrameric MP20, suggesting that other factors may also be involved. When purified MP20 tetramers are reconstituted with native lens lipids in the presence of magnesium, MP20 forms two-dimensional (2D) crystals. A projection map at 18 A resolution calculated from negatively stained 2D crystals showed that the building block of the crystal is an octamer consisting of two tetramers related to each other by 2-fold symmetry. In addition to 2D crystals, reconstitution of MP20 with native lipids also produced a variety of large protein-lipid complexes, and we present three-dimensional (3D) reconstructions of the four most abundant of these complexes in negative stain. The various complexes formed by MP20 most likely reflect the many ways in which tetraspanins can interact with each other to allow formation of TEMs.

}, keywords = {Animals, Buffers, Cations, Divalent, Cell Membrane, Chelating Agents, Crystallization, Detergents, Edetic Acid, Egtazic Acid, Eye Proteins, Hydrogen-Ion Concentration, Imaging, Three-Dimensional, Lens, Crystalline, Lipid Metabolism, Magnesium, Membrane Microdomains, Membrane Proteins, Protein Structure, Secondary, Protein Structure, Tertiary, Sheep, Silver Staining, Solubility, Tromethamine}, issn = {1089-8638}, doi = {10.1016/j.jmb.2007.09.001}, author = {Gonen, Tamir and Hite, Richard K and Cheng, Yifan and Petre, Benjamin M and Kistler, Joerg and Walz, Thomas} } @article {1685, title = {Quantitative in vivo imaging of entire embryos with digital scanned laser light sheet fluorescence microscopy.}, journal = {Current Opinion in Neurobiology}, volume = {18}, year = {2008}, month = {2008 Dec}, pages = {624-32}, abstract = {

The observation of biological processes in their natural in vivo context is a key requirement for quantitative experimental studies in the life sciences. In many instances, it will be crucial to achieve high temporal and spatial resolution over long periods of time without compromising the physiological development of the specimen. Here, we discuss the principles underlying light sheet-based fluorescence microscopes. The most recent implementation DSLM is a tool optimized to deliver quantitative data for entire embryos at high spatio-temporal resolution. We compare DSLM to the two established light microscopy techniques: confocal and two-photon fluorescence microscopy. DSLM provides up to 50 times higher imaging speeds and a 10-100 times higher signal-to-noise ratio, while exposing the specimens to at least three orders of magnitude less light energy than confocal and two-photon fluorescence microscopes. We conclude with a perspective for future development.

}, keywords = {Animals, Computer-Assisted, Confocal, Fetus, Fluorescence, Humans, Image Processing, Lasers, Microscopy, Multiphoton, Photobleaching}, issn = {1873-6882}, doi = {10.1016/j.conb.2009.03.008}, author = {Keller, Philipp J and Stelzer, Ernst H K} } @article {2363, title = {Receptors, circuits, and behaviors: new directions in chemical senses.}, journal = {The Journal of Neuroscience: The Official Journal of the Society for Neuroscience}, volume = {28}, year = {2008}, month = {2008 Nov 12}, pages = {11802-5}, abstract = {

The chemical senses, smell and taste, are the most poorly understood sensory modalities. In recent years, however, the field of chemosensation has benefited from new methods and technical innovations that have accelerated the rate of scientific progress. For example, enormous advances have been made in identifying olfactory and gustatory receptor genes and mapping their expression patterns. Genetic tools now permit us to monitor and control neural activity in vivo with unprecedented precision. New imaging techniques allow us to watch neural activity patterns unfold in real time. Finally, improved hardware and software enable multineuron electrophysiological recordings on an expanded scale. These innovations have enabled some fresh approaches to classic problems in chemosensation.

}, keywords = {Afferent Pathways, Animals, Brain, Chemoreceptor Cells, Humans, Odorant, Olfactory Pathways, Olfactory Perception, Receptors, Sensory Receptor Cells, Smell, Taste}, issn = {1529-2401}, doi = {10.1523/JNEUROSCI.3613-08.2008}, author = {Katz, Donald B and Matsunami, Hiroaki and Rinberg, Dmitry and Scott, Kristin and Wachowiak, Matt and Wilson, Rachel I} } @article {1687, title = {Reconstruction of zebrafish early embryonic development by scanned light sheet microscopy.}, journal = {Science}, volume = {322}, year = {2008}, month = {2008 Nov 14}, pages = {1065-9}, abstract = {

A long-standing goal of biology is to map the behavior of all cells during vertebrate embryogenesis. We developed digital scanned laser light sheet fluorescence microscopy and recorded nuclei localization and movement in entire wild-type and mutant zebrafish embryos over the first 24 hours of development. Multiview in vivo imaging at 1.5 billion voxels per minute provides "digital embryos," that is, comprehensive databases of cell positions, divisions, and migratory tracks. Our analysis of global cell division patterns reveals a maternally defined initial morphodynamic symmetry break, which identifies the embryonic body axis. We further derive a model of germ layer formation and show that the mesendoderm forms from one-third of the embryo{\textquoteright}s cells in a single event. Our digital embryos, with 55 million nucleus entries, are provided as a resource.

}, keywords = {Algorithms, Animals, beta Catenin, Biological, Body Patterning, Cell Division, Cell Nucleus, Computer-Assisted, Databases, Embryo, Embryonic Development, Endoderm, Factual, Fluorescence, Germ Layers, Image Processing, Mesoderm, Microscopy, Models, Motion Pictures as Topic, Mutation, Nonmammalian, Software, Zebrafish}, issn = {1095-9203}, doi = {10.1126/science.1162493}, author = {Keller, Philipp J and Schmidt, Annette D and Wittbrodt, Joachim and Stelzer, Ernst H K} } @article {6510, title = {Serial section scanning electron microscopy of adult brain tissue using focused ion beam milling.}, journal = {The Journal of Neuroscience: The Official Journal of the Society for Neuroscience}, volume = {28}, year = {2008}, month = {2008 Mar 19}, pages = {2959-64}, keywords = {Animals, Brain, Computer-Assisted, Electron, Female, Image Processing, Inbred C57BL, Ions, Mice, Microscopy, Scanning, Specimen Handling, Synapses}, issn = {1529-2401}, doi = {10.1523/JNEUROSCI.3189-07.2008}, author = {Knott, Graham and Marchman, Herschel and Wall, David and Lich, Ben} } @article {4980, title = {Standardized atlas of the brain of the desert locust, Schistocerca gregaria.}, journal = {Cell and Tissue Research}, volume = {333}, year = {2008}, month = {2008 Jul 27}, pages = {125-45}, abstract = {

In order to understand the connectivity of neuronal networks, their constituent neurons should ideally be studied in a common framework. Since morphological data from physiologically characterized and stained neurons usually arise from different individual brains, this can only be performed in a virtual standardized brain that compensates for interindividual variability. The desert locust, Schistocerca gregaria, is an insect species used widely for the analysis of olfactory and visual signal processing, endocrine functions, and neural networks controlling motor output. To provide a common multi-user platform for neural circuit analysis in the brain of this species, we have generated a standardized three-dimensional brain of this locust. Serial confocal images from whole-mount locust brains were used to reconstruct 34 neuropil areas in ten brains. For standardization, we compared two different methods: an iterative shape-averaging (ISA) procedure by using affine transformations followed by iterative nonrigid registrations, and the Virtual Insect Brain (VIB) protocol by using global and local rigid transformations followed by local nonrigid transformations. Both methods generated a standard brain, but for different applications. Whereas the VIB technique was designed to visualize anatomical variability between the input brains, the purpose of the ISA method was the opposite, i.e., to remove this variability. A novel individually labeled neuron, connecting the lobula to the midbrain and deutocerebrum, has been registered into the ISA atlas and demonstrates its usefulness and accuracy for future analysis of neural networks. The locust standard brain is accessible at http://www.3d-insectbrain.com .

}, keywords = {Anatomic, Anatomy, Animals, Artistic, Atlases as Topic, Brain, Brain Mapping, Grasshoppers, Imaging, Models, Neuroanatomy, Neurological, Neurons, Three-Dimensional}, issn = {0302-766X}, doi = {10.1007/s00441-008-0620-x}, author = {Kurylas, Angela E and Rohlfing, Torsten and Krofczik, Sabine and Jenett, Arnim and Homberg, Uwe} } @article {2695, title = {Straightening caenorhabditis elegans images.}, journal = {Bioinformatics}, volume = {24}, year = {2008}, month = {2008 Jan 15}, pages = {234-42}, abstract = {

MOTIVATION: Caenorhabditis elegans, a roundworm found in soil, is a widely studied model organism with about 1000 cells in the adult. Producing high-resolution fluorescence images of C.elegans to reveal biological insights is becoming routine, motivating the development of advanced computational tools for analyzing the resulting image stacks. For example, worm bodies usually curve significantly in images. Thus one must {\textquoteright}straighten{\textquoteright} the worms if they are to be compared under a canonical coordinate system. RESULTS: We develop a worm straightening algorithm (WSA) that restacks cutting planes orthogonal to a {\textquoteright}backbone{\textquoteright} that models the anterior-posterior axis of the worm. We formulate the backbone as a parametric cubic spline defined by a series of control points. We develop two methods for automatically determining the locations of the control points. Our experimental methods show that our approaches effectively straighten both 2D and 3D worm images.

}, keywords = {Algorithms, Animals, Caenorhabditis elegans, Computer-Assisted, Fluorescence, Image Enhancement, Image Interpretation, Imaging, Microscopy, Three-Dimensional}, issn = {1367-4811}, doi = {10.1093/bioinformatics/btm569}, author = {Peng, Hanchuan and Long, Fuhui and Liu, Xiao and Kim, Stuart K and Myers, Eugene W} } @article {2399, title = {Structural changes in TAF4b-TFIID correlate with promoter selectivity.}, journal = {Molecular Cell}, volume = {29}, year = {2008}, month = {2008 Jan 18}, pages = {81-91}, abstract = {

Proper ovarian development requires the cell type-specific transcription factor TAF4b, a subunit of the core promoter recognition complex TFIID. We present the 35 A structure of a cell type-specific core promoter recognition complex containing TAF4b and TAF4 (4b/4-IID), which is responsible for directing transcriptional synergy between c-Jun and Sp1 at a TAF4b target promoter. As a first step toward correlating potential structure/function relationships of the prototypic TFIID versus 4b/4-IID, we have compared their 3D structures by electron microscopy and single-particle reconstruction. These studies reveal that TAF4b incorporation into TFIID induces an open conformation at the lobe involved in TFIIA and putative activator interactions. Importantly, this open conformation correlates with differential activator-dependent transcription and promoter recognition by 4b/4-IID. By combining functional and structural analysis, we find that distinct localized structural changes in a megadalton macromolecular assembly can significantly alter its activity and lead to a TAF4b-induced reprogramming of promoter specificity.

}, keywords = {B-Lymphocytes, Cell Line, Computer-Assisted, Electron, Genetic, Hela Cells, Humans, Image Processing, Imaging, Microscopy, Models, Molecular, Organ Specificity, Promoter Regions, Protein Conformation, Protein Interaction Mapping, Proto-Oncogene Proteins c-jun, Sp1 Transcription Factor, Structure-Activity Relationship, TATA-Binding Protein Associated Factors, Three-Dimensional, Transcription Factor TFIIA, Transcription Factor TFIID, Transcriptional Activation, Tumor}, issn = {1097-2765}, doi = {10.1073/pnas.1100640108}, author = {Liu, Wei-Li and Coleman, Robert A and Grob, Patricia and King, David S and Florens, Laurence and Washburn, Michael P and Geles, Kenneth G and Yang, Joyce L and Ramey, Vincent and Nogales, Eva and Tjian, Robert} } @article {2957, title = {Subdiffraction multicolor imaging of the nuclear periphery with 3D structured illumination microscopy.}, journal = {Science}, volume = {320}, year = {2008}, month = {2008 Jun 6}, pages = {1332-6}, abstract = {

Fluorescence light microscopy allows multicolor visualization of cellular components with high specificity, but its utility has until recently been constrained by the intrinsic limit of spatial resolution. We applied three-dimensional structured illumination microscopy (3D-SIM) to circumvent this limit and to study the mammalian nucleus. By simultaneously imaging chromatin, nuclear lamina, and the nuclear pore complex (NPC), we observed several features that escape detection by conventional microscopy. We could resolve single NPCs that colocalized with channels in the lamin network and peripheral heterochromatin. We could differentially localize distinct NPC components and detect double-layered invaginations of the nuclear envelope in prophase as previously seen only by electron microscopy. Multicolor 3D-SIM opens new and facile possibilities to analyze subcellular structures beyond the diffraction limit of the emitted light.

}, keywords = {Animals, Cell Line, Cell Nucleus, Chromatin, Confocal, Fluorescence, Fluorescent Dyes, Heterochromatin, Imaging, Indoles, Interphase, Lamins, Mice, Microscopy, Myoblasts, Nuclear Envelope, Nuclear Lamina, Nuclear Pore, Optics and Photonics, Three-Dimensional}, issn = {1095-9203}, doi = {10.1126/science.1156947}, author = {Schermelleh, Lothar and Carlton, Peter M and Haase, Sebastian and Shao, Lin and Winoto, Lukman and Kner, Peter and Burke, Brian and Cardoso, M Cristina and Agard, David A and Gustafsson, Mats G L and Leonhardt, Heinrich and Sedat, John W} } @article {2395, title = {TBP, Mot1, and NC2 establish a regulatory circuit that controls DPE-dependent versus TATA-dependent transcription.}, journal = {Genes \& Development}, volume = {22}, year = {2008}, month = {2008 Sep 1}, pages = {2353-8}, abstract = {

The RNA polymerase II core promoter is a structurally and functionally diverse transcriptional module. RNAi depletion and overexpression experiments revealed a genetic circuit that controls the balance of transcription from two core promoter motifs, the TATA box and the downstream core promoter element (DPE). In this circuit, TBP activates TATA-dependent transcription and represses DPE-dependent transcription, whereas Mot1 and NC2 block TBP function and thus repress TATA-dependent transcription and activate DPE-dependent transcription. This regulatory circuit is likely to be one means by which biological networks can transmit transcriptional signals, such as those from DPE-specific and TATA-specific enhancers, via distinct pathways.

}, keywords = {Animals, Cells, Cultured, Drosophila, Drosophila Proteins, Enhancer Elements, Gene Expression Regulation, Genetic, Promoter Regions, RNA Polymerase II, TATA Box, TATA-Box Binding Protein, Transcription, Transcription Factors}, issn = {0890-9369}, doi = {10.1073/pnas.1100640108}, author = {Hsu, Jer-Yuan and Juven-Gershon, Tamar and Marr, Michael T and Wright, Kevin J and Tjian, Robert and Kadonaga, James T} } @article {1688, title = {Three-dimensional microtubule behavior in Xenopus egg extracts reveals four dynamic states and state-dependent elastic properties.}, journal = {Biophysical Journal}, volume = {95}, year = {2008}, month = {2008 Aug}, pages = {1474-86}, abstract = {

Although microtubules are key players in many cellular processes, very little is known about their dynamic and mechanical properties in physiological three-dimensional environments. The conventional model of microtubule dynamic instability postulates two dynamic microtubule states, growth and shrinkage. However, several studies have indicated that such a model does not provide a comprehensive quantitative and qualitative description of microtubule behavior. Using three-dimensional laser light-sheet fluorescence microscopy and a three-dimensional sample preparation in spacious Teflon cylinders, we measured microtubule dynamic instability and elasticity in interphase Xenopus laevis egg extracts. Our data are inconsistent with a two-state model of microtubule dynamic instability and favor an extended four-state model with two independent metastable pause states over a three-state model with a single pause state. Moreover, our data on kinetic state transitions rule out a simple GTP cap model as the driving force of microtubule stabilization in egg extracts on timescales of a few seconds or longer. We determined the three-dimensional elastic properties of microtubules as a function of both the contour length and the dynamic state. Our results indicate that pausing microtubules are less flexible than growing microtubules and suggest a growth-speed-dependent persistence length. These data might hint toward mechanisms that enable microtubules to efficiently perform multiple different tasks in the cell and suggest the development of a unified model of microtubule dynamics and microtubule mechanics.

}, keywords = {Animals, Elasticity, Imaging, Mechanical, Microtubules, Ovum, Stress, Three-Dimensional, Xenopus laevis}, issn = {1542-0086}, doi = {10.1529/biophysj.107.128223}, author = {Keller, Philipp J and Pampaloni, Francesco and Lattanzi, Gianluca and Stelzer, Ernst H K} } @article {988, title = {Tools for neuroanatomy and neurogenetics in Drosophila.}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {105}, year = {2008}, month = {2008 Jul 15}, pages = {9715-20}, abstract = {

We demonstrate the feasibility of generating thousands of transgenic Drosophila melanogaster lines in which the expression of an exogenous gene is reproducibly directed to distinct small subsets of cells in the adult brain. We expect the expression patterns produced by the collection of 5,000 lines that we are currently generating to encompass all neurons in the brain in a variety of intersecting patterns. Overlapping 3-kb DNA fragments from the flanking noncoding and intronic regions of genes thought to have patterned expression in the adult brain were inserted into a defined genomic location by site-specific recombination. These fragments were then assayed for their ability to function as transcriptional enhancers in conjunction with a synthetic core promoter designed to work with a wide variety of enhancer types. An analysis of 44 fragments from four genes found that \>80\% drive expression patterns in the brain; the observed patterns were, on average, comprised of \<100 cells. Our results suggest that the D. melanogaster genome contains \>50,000 enhancers and that multiple enhancers drive distinct subsets of expression of a gene in each tissue and developmental stage. We expect that these lines will be valuable tools for neuroanatomy as well as for the elucidation of neuronal circuits and information flow in the fly brain.

}, keywords = {Animals, Biomedical Research, Brain, Developmental, Drosophila melanogaster, Enhancer Elements, Gene Expression Regulation, Genes, Genetic, Genetically Modified, Insect, Neurons, Neurosciences, Recombination}, issn = {1091-6490}, doi = {https://doi.org/10.1073/pnas.0803697105}, author = {Pfeiffer, Barret D and Jenett, Arnim and Hammonds, Ann S and Ngo, Teri-T B and Misra, Sima and Murphy, Christine and Scully, Audra and Carlson, Joseph W and Wan, Kenneth H and Laverty, Todd R and Mungall, Chris and Svirskas, Rob and Kadonaga, James T and Doe, Chris Q and Eisen, Michael B and Celniker, Susan E and Rubin, Gerald M} } @article {68826, title = {V2a and V2b neurons are generated by the final divisions of pair-producing progenitors in the zebrafish spinal cord}, journal = {Development}, volume = {135}, year = {2008}, month = {09/2008}, pages = {3001-3005}, abstract = {

The p2 progenitor domain in the ventral spinal cord gives rise to two interneuron subtypes: V2a and V2b. Delta-Notch-mediated cell-cell interactions between postmitotic immature neurons have been implicated in the segregation of neuron subtypes. However, lineage relationships between V2a and V2b neurons have not been reported. We address this issue using Tg[vsx1:GFP]zebrafish, a model system in which high GFP expression is initiated near the final stage of p2 progenitors. Cell fates were followed in progeny using time-lapse microscopy. Results indicate that the vast majority, if not all, of GFP-labeled p2 progenitors divide once to produce V2a/V2b neuron pairs,indicating that V2a and V2b neurons are generated by the asymmetric division of pair-producing progenitor cells. Together with evidence that Notch signaling is involved in the cell fate specification process, our results strongly suggest that Delta-Notch interactions between sister cells play a crucial role in the final outcome of these asymmetric divisions. This mechanism for determining cell fate is similar to asymmetric divisions that occur during Drosophila neurogenesis, where ganglion mother cells divide once to produce distinct neurons. However, unlike in Drosophila, the divisional axes of p2 progenitors in zebrafish were not fixed. We report that the terminal division of pair-producing progenitor cells in vertebrate neurogenesis can reproducibly produce two distinct neurons through a mechanism that may not depend on the orientation of the division axis.

}, issn = {0950-1991}, doi = {10.1242/dev.024802}, url = {https://doi.org/10.1242/dev.024802}, author = {Kimura, Yukiko and Satou, Chie and Higashijima, Shin-Ichi} } @article {4431, title = {Visuomotor transformation in the fly gaze stabilization system.}, journal = {PLoS Biology}, volume = {6}, year = {2008}, month = {2008 Jul 22}, pages = {e173}, abstract = {

For sensory signals to control an animal{\textquoteright}s behavior, they must first be transformed into a format appropriate for use by its motor systems. This fundamental problem is faced by all animals, including humans. Beyond simple reflexes, little is known about how such sensorimotor transformations take place. Here we describe how the outputs of a well-characterized population of fly visual interneurons, lobula plate tangential cells (LPTCs), are used by the animal{\textquoteright}s gaze-stabilizing neck motor system. The LPTCs respond to visual input arising from both self-rotations and translations of the fly. The neck motor system however is involved in gaze stabilization and thus mainly controls compensatory head rotations. We investigated how the neck motor system is able to selectively extract rotation information from the mixed responses of the LPTCs. We recorded extracellularly from fly neck motor neurons (NMNs) and mapped the directional preferences across their extended visual receptive fields. Our results suggest that-like the tangential cells-NMNs are tuned to panoramic retinal image shifts, or optic flow fields, which occur when the fly rotates about particular body axes. In many cases, tangential cells and motor neurons appear to be tuned to similar axes of rotation, resulting in a correlation between the coordinate systems the two neural populations employ. However, in contrast to the primarily monocular receptive fields of the tangential cells, most NMNs are sensitive to visual motion presented to either eye. This results in the NMNs being more selective for rotation than the LPTCs. Thus, the neck motor system increases its rotation selectivity by a comparatively simple mechanism: the integration of binocular visual motion information.

}, keywords = {Animals, Diptera, Fixation, Motion Perception, Motor Neurons, Neck, Neurons, Ocular, Photic Stimulation, Visual Fields, Visual Pathways}, issn = {1545-7885}, doi = {10.1371/journal.pbio.0060173}, author = {Huston, Stephen J and Krapp, Holger G} } @article {1114, title = {Absence of the Thorpe-Ingold Eet by gem-diphenyl groups in ring-closing enyne metathesis.}, journal = {Tetrahedron Letters}, volume = {48}, year = {2007}, month = {2007 Nov 5}, pages = {7961-4}, abstract = {

In tandem ring-closing metathesis of alkynyl silaketals containing two different tethered olefins, the gem-dimethyl group showed the expected Thorpe-Ingold effect, thereby giving good level of group selectivity. Unexpectedly, however, the corresponding gem-diphenyl group did not show any Thorpe-Ingold effect for the ring closure reaction.

}, issn = {0040-4039}, doi = {10.1016/j.tetlet.2007.09.063}, author = {Kim, Yi Jin and Grimm, Jonathan B and Lee, Daesung} } @article {2810, title = {Ambient mass spectrometry with a handheld mass spectrometer at high pressure.}, journal = {Analytical Chemistry}, volume = {79}, year = {2007}, month = {2007 Oct 15}, pages = {7734-9}, abstract = {

The first coupling of atmospheric pressure ionization methods, electrospray ionization (ESI) and desorption electrospray ionization (DESI), to a miniature hand-held mass spectrometer is reported. The instrument employs a rectilinear ion trap (RIT) mass analyzer and is battery-operated, hand-portable, and rugged (total system: 10 kg, 0.014 m(3), 75 W power consumption). The mass spectrometer was fitted with an atmospheric inlet, consisting of a 10 cm x 127 microm inner diameter stainless steel capillary tube which was used to introduce gas into the vacuum chamber at 13 mL/min. The operating pressure was 15 mTorr. Ions, generated by the atmospheric pressure ion source, were directed by the inlet along the axis of the ion trap, entering through an aperture in the dc-biased end plate, which was also operated as an ion gate. ESI and DESI sources were used to generate ions; ESI-MS analysis of an aqueous mixture of drugs yielded detection limits in the low parts-per-billion range. Signal response was linear over more than 3 orders of magnitude. Tandem mass spectrometry experiments were used to identify components of this mixture. ESI was also applied to the analysis of peptides and in this case multiply charged species were observed for compounds of molecular weight up to 1200 Da. Cocaine samples deposited or already present on different surfaces, including currency, were rapidly analyzed in situ by DESI. A geometry-independent version of the DESI ion source was also coupled to the miniature mass spectrometer. These results demonstrate that atmospheric pressure ionization can be implemented on simple portable mass spectrometry systems.

}, keywords = {Alkylation, Calibration, Mass Spectrometry, Molecular Structure, Molecular Weight, Pressure, Proteins, Temperature}, issn = {0003-2700}, doi = {10.1364/AO.50.001792}, author = {Keil, Adam and Talaty, Nari and Janfelt, Christian and Noll, Robert J and Gao, Liang and Ouyang, Zheng and Cooks, R Graham} } @article {45889, title = {Coincidence detection of place and temporal context in a network model of spiking hippocampal neurons.}, journal = {PLoS Computational Biology}, volume = {3}, year = {2007}, month = {2007 Dec}, pages = {e234}, abstract = {

Recent advances in single-neuron biophysics have enhanced our understanding of information processing on the cellular level, but how the detailed properties of individual neurons give rise to large-scale behavior remains unclear. Here, we present a model of the hippocampal network based on observed biophysical properties of hippocampal and entorhinal cortical neurons. We assembled our model to simulate spatial alternation, a task that requires memory of the previous path through the environment for correct selection of the current path to a reward site. The convergence of inputs from entorhinal cortex and hippocampal region CA3 onto CA1 pyramidal cells make them potentially important for integrating information about place and temporal context on the network level. Our model shows how place and temporal context information might be combined in CA1 pyramidal neurons to give rise to splitter cells, which fire selectively based on a combination of place and temporal context. The model leads to a number of experimentally testable predictions that may lead to a better understanding of the biophysical basis of information processing in the hippocampus.

}, keywords = {Action Potentials, Algorithms, Animals, Computer Simulation, Hippocampus, Models, Neurological, Nerve Net, Neurons, Rats, Synaptic Transmission}, issn = {1553-7358}, doi = {10.1371/journal.pcbi.0030234}, author = {Katz, Yael and Kath, William L and Spruston, Nelson and Hasselmo, Michael E} } @article {6361, title = {The DNA transposon Minos as a tool for transgenesis and functional genomic analysis in vertebrates and invertebrates.}, journal = {Genome Biology}, volume = {8 Suppl 1}, year = {2007}, month = {2007 Oct 31}, pages = {S2}, abstract = {

Transposons are powerful tools for conducting genetic manipulation and functional studies in organisms that are of scientific, economic, or medical interest. Minos, a member of the Tc1/mariner family of DNA transposons, exhibits a low insertional bias and transposes with high frequency in vertebrates and invertebrates. Its use as a tool for transgenesis and genome analysis of rather different animal species is described.

}, keywords = {Animals, DNA Transposable Elements, Gene Transfer Techniques, Genomics, Invertebrates, Vertebrates}, issn = {1465-6914}, doi = {10.1186/gb-2007-8-s1-s2}, author = {Pavlopoulos, Anastasios and Oehler, Stefan and Kapetanaki, Maria G and Savakis, Charalambos} } @article {45741, title = {Evolution. An embarrassment of switches.}, journal = {Science}, volume = {317}, year = {2007}, month = {2007 Aug 10}, pages = {758-9}, keywords = {Animals, Binding Sites, Chromatin Immunoprecipitation, Evolution, Molecular, Gene Expression Regulation, Humans, Mutation, Oligonucleotide Array Sequence Analysis, Phenotype, Regulatory Sequences, Nucleic Acid, Species Specificity, Transcription Factors, Yeasts}, issn = {1095-9203}, doi = {10.1126/science.1146921}, author = {Kruglyak, Leonid and Stern, David L} } @article {6494, title = {Evolution of genes and genomes on the Drosophila phylogeny.}, journal = {Nature}, volume = {450}, year = {2007}, month = {2007 Nov 8}, pages = {203-18}, abstract = {

Comparative analysis of multiple genomes in a phylogenetic framework dramatically improves the precision and sensitivity of evolutionary inference, producing more robust results than single-genome analyses can provide. The genomes of 12 Drosophila species, ten of which are presented here for the first time (sechellia, simulans, yakuba, erecta, ananassae, persimilis, willistoni, mojavensis, virilis and grimshawi), illustrate how rates and patterns of sequence divergence across taxa can illuminate evolutionary processes on a genomic scale. These genome sequences augment the formidable genetic tools that have made Drosophila melanogaster a pre-eminent model for animal genetics, and will further catalyse fundamental research on mechanisms of development, cell biology, genetics, disease, neurobiology, behaviour, physiology and evolution. Despite remarkable similarities among these Drosophila species, we identified many putatively non-neutral changes in protein-coding genes, non-coding RNA genes, and cis-regulatory regions. These may prove to underlie differences in the ecology and behaviour of these diverse species.

}, keywords = {Animals, Codon, DNA, DNA Transposable Elements, Drosophila, Drosophila Proteins, Evolution, Gene Order, Genes, Genome, Genomics, Immunity, Insect, Mitochondrial, Molecular, Multigene Family, Phylogeny, Reproduction, RNA, Sequence Alignment, Sequence Analysis, Synteny, Untranslated}, issn = {1476-4687}, doi = {10.1038/nature06341}, author = {Clark, Andrew G and Eisen, Michael B and Smith, Douglas R and Bergman, Casey M and Oliver, Brian and Markow, Therese A and Kaufman, Thomas C and Kellis, Manolis and Gelbart, William and Iyer, Venky N and Pollard, Daniel A and Sackton, Timothy B and Larracuente, Amanda M and Singh, Nadia D and Abad, Jose P and Abt, Dawn N and Adryan, Boris and Aguade, Montserrat and Akashi, Hiroshi and Anderson, Wyatt W and Aquadro, Charles F and Ardell, David H and Arguello, Roman and Artieri, Carlo G and Barbash, Daniel A and Barker, Daniel and Barsanti, Paolo and Batterham, Phil and Batzoglou, Serafim and Begun, Dave and Bhutkar, Arjun and Blanco, Enrico and Bosak, Stephanie A and Bradley, Robert K and Brand, Adrianne D and Brent, Michael R and Brooks, Angela N and Brown, Randall H and Butlin, Roger K and Caggese, Corrado and Calvi, Brian R and Bernardo de Carvalho, A and Caspi, Anat and Castrezana, Sergio and Celniker, Susan E and Chang, Jean L and Chapple, Charles and Chatterji, Sourav and Chinwalla, Asif and Civetta, Alberto and Clifton, Sandra W and Comeron, Josep M and Costello, James C and Coyne, Jerry A and Daub, Jennifer and David, Robert G and Delcher, Arthur L and Delehaunty, Kim and Do, Chuong B and Ebling, Heather and Edwards, Kevin and Eickbush, Thomas and Evans, Jay D and Filipski, Alan and Findeiss, Sven and Freyhult, Eva and Fulton, Lucinda and Fulton, Robert and Garcia, Ana C L and Gardiner, Anastasia and Garfield, David A and Garvin, Barry E and Gibson, Greg and Gilbert, Don and Gnerre, Sante and Godfrey, Jennifer and Good, Robert and Gotea, Valer and Gravely, Brenton and Greenberg, Anthony J and Griffiths-Jones, Sam and Gross, Samuel and Guigo, Roderic and Gustafson, Erik A and Haerty, Wilfried and Hahn, Matthew W and Halligan, Daniel L and Halpern, Aaron L and Halter, Gillian M and Han, Mira V and Heger, Andreas and Hillier, LaDeana and Hinrichs, Angie S and Holmes, Ian and Hoskins, Roger A and Hubisz, Melissa J and Hultmark, Dan and Huntley, Melanie A and Jaffe, David B and Jagadeeshan, Santosh and Jeck, William R and Johnson, Justin and Jones, Corbin D and Jordan, William C and Karpen, Gary H and Kataoka, Eiko and Keightley, Peter D and Kheradpour, Pouya and Kirkness, Ewen F and Koerich, Leonardo B and Kristiansen, Karsten and Kudrna, Dave and Kulathinal, Rob J and Kumar, Sudhir and Kwok, Roberta and Lander, Eric and Langley, Charles H and Lapoint, Richard and Lazzaro, Brian P and Lee, So-Jeong and Levesque, Lisa and Li, Ruiqiang and Lin, Chiao-Feng and Lin, Michael F and Lindblad-Toh, Kerstin and Llopart, Ana and Long, Manyuan and Low, Lloyd and Lozovsky, Elena and Lu, Jian and Luo, Meizhong and Machado, Carlos A and Makalowski, Wojciech and Marzo, Mar and Matsuda, Muneo and Matzkin, Luciano and McAllister, Bryant and McBride, Carolyn S and McKernan, Brendan and McKernan, Kevin and Mendez-Lago, Maria and Minx, Patrick and Mollenhauer, Michael U and Montooth, Kristi and Mount, Stephen M and Mu, Xu and Myers, Eugene and Negre, Barbara and Newfeld, Stuart and Nielsen, Rasmus and Noor, Mohamed A F and O{\textquoteright}Grady, Patrick and Pachter, Lior and Papaceit, Montserrat and Parisi, Matthew J and Parisi, Michael and Parts, Leopold and Pedersen, Jakob S and Pesole, Graziano and Phillippy, Adam M and Ponting, Chris P and Pop, Mihai and Porcelli, Damiano and Powell, Jeffrey R and Prohaska, Sonja and Pruitt, Kim and Puig, Marta and Quesneville, Hadi and Ram, Kristipati Ravi and Rand, David and Rasmussen, Matthew D and Reed, Laura K and Reenan, Robert and Reily, Amy and Remington, Karin A and Rieger, Tania T and Ritchie, Michael G and Robin, Charles and Rogers, Yu-Hui and Rohde, Claudia and Rozas, Julio and Rubenfield, Marc J and Ruiz, Alfredo and Russo, Susan and Salzberg, Steven L and Sanchez-Gracia, Alejandro and Saranga, David J and Sato, Hajime and Schaeffer, Stephen W and Schatz, Michael C and Schlenke, Todd and Schwartz, Russell and Segarra, Carmen and Singh, Rama S and Sirot, Laura and Sirota, Marina and Sisneros, Nicholas B and Smith, Chris D and Smith, Temple F and Spieth, John and Stage, Deborah E and Stark, Alexander and Stephan, Wolfgang and Strausberg, Robert L and Strempel, Sebastian and Sturgill, David and Sutton, Granger and Sutton, Granger G and Tao, Wei and Teichmann, Sarah and Tobari, Yoshiko N and Tomimura, Yoshihiko and Tsolas, Jason M and Valente, Vera L S and Venter, Eli and Venter, J Craig and Vicario, Saverio and Vieira, Filipe G and Vilella, Albert J and Villasante, Alfredo and Walenz, Brian and Wang, Jun and Wasserman, Marvin and Watts, Thomas and Wilson, Derek and Wilson, Richard K and Wing, Rod A and Wolfner, Mariana F and Wong, Alex and Wong, Gane Ka-Shu and Wu, Chung-I and Wu, Gabriel and Yamamoto, Daisuke and Yang, Hsiao-Pei and Yang, Shiaw-Pyng and Yorke, James A and Yoshida, Kiyohito and Zdobnov, Evgeny and Zhang, Peili and Zhang, Yu and Zimin, Aleksey V and Baldwin, Jennifer and Abdouelleil, Amr and Abdulkadir, Jamal and Abebe, Adal and Abera, Brikti and Abreu, Justin and Acer, St Christophe and Aftuck, Lynne and Alexander, Allen and An, Peter and Anderson, Erica and Anderson, Scott and Arachi, Harindra and Azer, Marc and Bachantsang, Pasang and Barry, Andrew and Bayul, Tashi and Berlin, Aaron and Bessette, Daniel and Bloom, Toby and Blye, Jason and Boguslavskiy, Leonid and Bonnet, Claude and Boukhgalter, Boris and Bourzgui, Imane and Brown, Adam and Cahill, Patrick and Channer, Sheridon and Cheshatsang, Yama and Chuda, Lisa and Citroen, Mieke and Collymore, Alville and Cooke, Patrick and Costello, Maura and D{\textquoteright}Aco, Katie and Daza, Riza and De Haan, Georgius and DeGray, Stuart and DeMaso, Christina and Dhargay, Norbu and Dooley, Kimberly and Dooley, Erin and Doricent, Missole and Dorje, Passang and Dorjee, Kunsang and Dupes, Alan and Elong, Richard and Falk, Jill and Farina, Abderrahim and Faro, Susan and Ferguson, Diallo and Fisher, Sheila and Foley, Chelsea D and Franke, Alicia and Friedrich, Dennis and Gadbois, Loryn and Gearin, Gary and Gearin, Christina R and Giannoukos, Georgia and Goode, Tina and Graham, Joseph and Grandbois, Edward and Grewal, Sharleen and Gyaltsen, Kunsang and Hafez, Nabil and Hagos, Birhane and Hall, Jennifer and Henson, Charlotte and Hollinger, Andrew and Honan, Tracey and Huard, Monika D and Hughes, Leanne and Hurhula, Brian and Husby, M Erii and Kamat, Asha and Kanga, Ben and Kashin, Seva and Khazanovich, Dmitry and Kisner, Peter and Lance, Krista and Lara, Marcia and Lee, William and Lennon, Niall and Letendre, Frances and LeVine, Rosie and Lipovsky, Alex and Liu, Xiaohong and Liu, Jinlei and Liu, Shangtao and Lokyitsang, Tashi and Lokyitsang, Yeshi and Lubonja, Rakela and Lui, Annie and MacDonald, Pen and Magnisalis, Vasilia and Maru, Kebede and Matthews, Charles and McCusker, William and McDonough, Susan and Mehta, Teena and Meldrim, James and Meneus, Louis and Mihai, Oana and Mihalev, Atanas and Mihova, Tanya and Mittelman, Rachel and Mlenga, Valentine and Montmayeur, Anna and Mulrain, Leonidas and Navidi, Adam and Naylor, Jerome and Negash, Tamrat and Nguyen, Thu and Nguyen, Nga and Nicol, Robert and Norbu, Choe and Norbu, Nyima and Novod, Nathaniel and O{\textquoteright}Neill, Barry and Osman, Sahal and Markiewicz, Eva and Oyono, Otero L and Patti, Christopher and Phunkhang, Pema and Pierre, Fritz and Priest, Margaret and Raghuraman, Sujaa and Rege, Filip and Reyes, Rebecca and Rise, Cecil and Rogov, Peter and Ross, Keenan and Ryan, Elizabeth and Settipalli, Sampath and Shea, Terry and Sherpa, Ngawang and Shi, Lu and Shih, Diana and Sparrow, Todd and Spaulding, Jessica and Stalker, John and Stange-Thomann, Nicole and Stavropoulos, Sharon and Stone, Catherine and Strader, Christopher and Tesfaye, Senait and Thomson, Talene and Thoulutsang, Yama and Thoulutsang, Dawa and Topham, Kerri and Topping, Ira and Tsamla, Tsamla and Vassiliev, Helen and Vo, Andy and Wangchuk, Tsering and Wangdi, Tsering and Weiand, Michael and Wilkinson, Jane and Wilson, Adam and Yadav, Shailendra and Young, Geneva and Yu, Qing and Zembek, Lisa and Zhong, Danni and Zimmer, Andrew and Zwirko, Zac and Jaffe, David B and Alvarez, Pablo and Brockman, Will and Butler, Jonathan and Chin, CheeWhye and Gnerre, Sante and Grabherr, Manfred and Kleber, Michael and Mauceli, Evan and MacCallum, Iain} } @article {6858, title = {Function of the Drosophila CPEB protein Orb2 in long-term courtship memory.}, journal = {Nature Neuroscience}, volume = {10}, year = {2007}, month = {2007 Dec 28}, pages = {1587-93}, abstract = {

Both long-term behavioral memory and synaptic plasticity require protein synthesis, some of which may occur locally at specific synapses. Cytoplasmic polyadenylation element-binding (CPEB) proteins are thought to contribute to the local protein synthesis that underlies long-term changes in synaptic efficacy, but a role has not been established for them in the formation of long-term behavioral memory. We found that the Drosophila melanogaster CPEB protein Orb2 is acutely required for long-term conditioning of male courtship behavior. Deletion of the N-terminal glutamine-rich region of Orb2 resulted in flies that were impaired in their ability to form long-term, but not short-term, memory. Memory was restored by expressing Orb2 selectively in fruitless (fru)-positive gamma neurons of the mushroom bodies and by providing Orb2 function in mushroom bodies only during and shortly after training. Our data thus demonstrate that a CPEB protein is important in long-term memory and map the molecular, spatial and temporal requirements for its function in memory formation.

}, keywords = {Animal, Animals, Behavior, Computational Biology, Courtship, Drosophila, Drosophila Proteins, Female, Gene Expression, Genetically Modified, Green Fluorescent Proteins, Male, Memory, Memory Disorders, Mushroom Bodies, Neurons, Phylogeny, RNA-Binding Proteins, Sequence Deletion, Time Factors}, issn = {1097-6256}, doi = {10.1038/nn1996}, author = {Keleman, Krystyna and Kr{\"u}ttner, Sebastian and Alenius, Mattias and Dickson, Barry J} } @article {6857, title = {A genome-wide transgenic RNAi library for conditional gene inactivation in Drosophila.}, journal = {Nature}, volume = {448}, year = {2007}, month = {2007 Jul 12}, pages = {151-6}, abstract = {

Forward genetic screens in model organisms have provided important insights into numerous aspects of development, physiology and pathology. With the availability of complete genome sequences and the introduction of RNA-mediated gene interference (RNAi), systematic reverse genetic screens are now also possible. Until now, such genome-wide RNAi screens have mostly been restricted to cultured cells and ubiquitous gene inactivation in Caenorhabditis elegans. This powerful approach has not yet been applied in a tissue-specific manner. Here we report the generation and validation of a genome-wide library of Drosophila melanogaster RNAi transgenes, enabling the conditional inactivation of gene function in specific tissues of the intact organism. Our RNAi transgenes consist of short gene fragments cloned as inverted repeats and expressed using the binary GAL4/UAS system. We generated 22,270 transgenic lines, covering 88\% of the predicted protein-coding genes in the Drosophila genome. Molecular and phenotypic assays indicate that the majority of these transgenes are functional. Our transgenic RNAi library thus opens up the prospect of systematically analysing gene functions in any tissue and at any stage of the Drosophila lifespan.

}, keywords = {Animals, Drosophila melanogaster, Exons, Female, Genetically Modified, Genomic Library, Male, Messenger, Muscles, Neurons, Organ Specificity, Ribonuclease III, RNA, RNA Interference}, issn = {1476-4687}, doi = {10.1038/nature05954}, author = {Dietzl, Georg and Chen, Doris and Schnorrer, Frank and Su, Kuan-Chung and Barinova, Yulia and Fellner, Michaela and Gasser, Beate and Kinsey, Kaolin and Oppel, Silvia and Scheiblauer, Susanne and Couto, Africa and Marra, Vincent and Keleman, Krystyna and Dickson, Barry J} } @article {955, title = {Global analysis of patterns of gene expression during Drosophila embryogenesis.}, journal = {Genome Biology}, volume = {8}, year = {2007}, month = {2007 July 23}, pages = {R145}, abstract = {

Cell and tissue specific gene expression is a defining feature of embryonic development in multi-cellular organisms. However, the range of gene expression patterns, the extent of the correlation of expression with function, and the classes of genes whose spatial expression are tightly regulated have been unclear due to the lack of an unbiased, genome-wide survey of gene expression patterns.

}, keywords = {Animals, Drosophila melanogaster, Embryonic Development, Gene Expression Profiling}, issn = {1465-6914}, doi = {10.1186/gb-2007-8-7-r145}, author = {Tomancak, Pavel and Berman, Benjamin P and Beaton, Amy and Weiszmann, Richard and Kwan, Elaine and Hartenstein, Volker and Celniker, Susan E and Rubin, Gerald M} } @article {1582, title = {A global transcriptional regulator in Thermococcus kodakaraensis controls the expression levels of both glycolytic and gluconeogenic enzyme-encoding genes.}, journal = {The Journal of Biological Chemistry}, volume = {282}, year = {2007}, month = {2007 Nov 16}, pages = {33659-70}, abstract = {

We identified a novel regulator, Thermococcales glycolytic regulator (Tgr), functioning as both an activator and a repressor of transcription in the hyperthermophilic archaeon Thermococcus kodakaraensis KOD1. Tgr (TK1769) displays similarity (28\% identical) to Pyrococcus furiosus TrmB (PF1743), a transcriptional repressor regulating the trehalose/maltose ATP-binding cassette transporter genes, but is more closely related (67\%) to a TrmB paralog in P. furiosus (PF0124). Growth of a tgr disruption strain (Deltatgr) displayed a significant decrease in growth rate under gluconeogenic conditions compared with the wild-type strain, whereas comparable growth rates were observed under glycolytic conditions. A whole genome microarray analysis revealed that transcript levels of almost all genes related to glycolysis and maltodextrin metabolism were at relatively high levels in the Deltatgr mutant even under gluconeogenic conditions. The Deltatgr mutant also displayed defects in the transcriptional activation of gluconeogenic genes under these conditions, indicating that Tgr functions as both an activator and a repressor. Genes regulated by Tgr contain a previously identified sequence motif, the Thermococcales glycolytic motif (TGM). The TGM was positioned upstream of the Transcription factor B-responsive element (BRE)/TATA sequence in gluconeogenic promoters and downstream of it in glycolytic promoters. Electrophoretic mobility shift assay indicated that recombinant Tgr protein specifically binds to promoter regions containing a TGM. Tgr was released from the DNA when maltotriose was added, suggesting that this sugar is most likely the physiological effector. Our results strongly suggest that Tgr is a global transcriptional regulator that simultaneously controls, in response to sugar availability, both glycolytic and gluconeogenic metabolism in T. kodakaraensis via its direct binding to the TGM.

}, keywords = {Amino Acid, Amino Acid Motifs, Amino Acid Sequence, Escherichia coli, Genetic, Genome, Genotype, Gluconeogenesis, Glycolysis, Molecular Sequence Data, Multienzyme Complexes, NADH, NADPH Oxidoreductases, Oligonucleotide Array Sequence Analysis, Promoter Regions, Pyrococcus furiosus, Recombinant Proteins, Sequence Homology, Thermococcus, Transcription}, issn = {0021-9258}, doi = {10.1074/jbc.M703424200}, author = {Kanai, Tamotsu and Akerboom, Jasper and Takedomi, Shogo and van de Werken, Harmen J G and Blombach, Fabian and van der Oost, John and Murakami, Taira and Atomi, Haruyuki and Imanaka, Tadayuki} } @article {6250, title = {Lola regulates Drosophila olfactory projection neuron identity and targeting specificity.}, journal = {Neural Development}, volume = {2}, year = {2007}, month = {2007 Jul 16}, pages = {14}, abstract = {

BACKGROUND: Precise connections of neural circuits can be specified by genetic programming. In the Drosophila olfactory system, projection neurons (PNs) send dendrites to single glomeruli in the antenna lobe (AL) based upon lineage and birth order and send axons with stereotyped terminations to higher olfactory centers. These decisions are likely specified by a PN-intrinsic transcriptional code that regulates the expression of cell-surface molecules to instruct wiring specificity. RESULTS: We find that the loss of longitudinals lacking (lola), which encodes a BTB-Zn-finger transcription factor with 20 predicted splice isoforms, results in wiring defects in both axons and dendrites of all lineages of PNs. RNA in situ hybridization and quantitative RT-PCR suggest that most if not all lola isoforms are expressed in all PNs, but different isoforms are expressed at widely varying levels. Overexpression of individual lola isoforms fails to rescue the lola null phenotypes and causes additional phenotypes. Loss of lola also results in ectopic expression of Gal4 drivers in multiple cell types and in the loss of transcription factor gene lim1 expression in ventral PNs. CONCLUSION: Our results indicate that lola is required for wiring of axons and dendrites of most PN classes, and suggest a need for its molecular diversity. Expression pattern changes of Gal4 drivers in lola-/- clones imply that lola normally represses the expression of these regulatory elements in a subset of the cells surrounding the AL. We propose that Lola functions as a general transcription factor that regulates the expression of multiple genes ultimately controlling PN identity and wiring specificity.

}, keywords = {Animals, Body Patterning, Brain, Cell Differentiation, Dendrites, Developmental, Drosophila melanogaster, Drosophila Proteins, Gene Expression Regulation, Growth Cones, Larva, Olfactory Pathways, Olfactory Receptor Neurons, Regulatory Elements, Transcription Factors, Transcriptional, Transcriptional Activation}, issn = {1749-8104}, doi = {10.1186/1749-8104-2-14}, author = {Spletter, Maria Lynn and Liu, Jian and Liu, Justin and Su, Helen and Giniger, Edward and Komiyama, Takaki and Quake, Stephen and Luo, Liqun} } @article {6443, title = {MRI-based localization of electrophysiological recording sites within the cerebral cortex at single-voxel accuracy.}, journal = {Nature Methods}, volume = {4}, year = {2007}, month = {2007 Feb}, pages = {161-8}, abstract = {

The localization of microelectrode recording sites in the layers of primate cerebral cortex permits the analysis of relationships between recorded neuronal activities and underlying anatomical connections. We present a magnetic resonance imaging method for precise in vivo localization of cortical recording sites. In this method, the susceptibility-induced effect thickens the appearance of the microelectrode and enhances the detectability of the microelectrode tip, which usually occupies less than a few percent of the volume of an image voxel. In a phantom study, the optimized susceptibility-induced effect allowed tip detection with single-voxel accuracy (in-plane resolution, 50 mum). We applied this method to recording microelectrodes inserted into the brains of macaque monkeys, and localized the microelectrode tip at an in-plane resolution of 150 mum within the cortex of 2-3 mm in thickness. Subsequent histological analyses validated the single-voxel accuracy of the in vivo tip localization. This method opens up a way to investigate information flow during cognitive processes in the brain.

}, keywords = {Animal, Animals, Behavior, Cerebral Cortex, Electrophysiology, Imaging, Macaca, Magnetic Resonance Imaging, Microelectrodes, Phantoms}, issn = {1548-7091}, doi = {10.1038/nmeth987}, author = {Matsui, Teppei and Koyano, Kenji W and Koyama, Minoru and Nakahara, Kiyoshi and Takeda, Masaki and Ohashi, Yohei and Naya, Yuji and Miyashita, Yasushi} } @article {1112, title = {A new strategy for the synthesis of benzylic sulfonamides: palladium-catalyzed arylation and sulfonamide metathesis.}, journal = {The Journal of Organic Chemistry}, volume = {72}, year = {2007}, month = {2007 Oct 12}, pages = {8135-8}, abstract = {

An efficient two-step strategy has been developed to access diversely functionalized benzylic sulfonamides. Execution of this strategy required the development of two reaction methods: the palladium-catalyzed cross-coupling of aryl halides with CH-acidic methanesulfonamides and a metathesis reaction between the resulting alpha-arylated sulfonamides and diverse amines. The broad scope of the cross-coupling process combined with a versatile sulfonamide metathesis constitutes an efficient strategy for the synthesis of various benzylic sulfonamides.

}, keywords = {Benzyl Compounds, Catalysis, Molecular Structure, Palladium, Sulfonamides}, issn = {0022-3263}, doi = {10.1021/jo701431j}, author = {Grimm, Jonathan B and Katcher, Matthew H and Witter, David J and Northrup, Alan B} } @article {67285, title = {Observability of Higgs produced with top quarks and decaying to bottom quarks}, journal = {Journal of Physics G: Nuclear and Particle Physics}, volume = {34}, year = {2007}, month = {04/20027}, pages = {N221 - N250}, abstract = {

The decay,\ , is dominant for a Standard Model Higgs boson in the mass range just above the exclusion limit of 114.4 GeV/c2\ reported by the LEP experiments. Unfortunately, an overwhelming abundance of\ \ events arising from more mundane sources, together with the lack of precision inherent in the reconstruction of the Higgs mass, renders this decay mode\ a priori\ undetectable in the case of direct Higgs production at the LHC. It is therefore of no small interest to investigate whether\ \ can be observed in those cases where the Higgs is produced in association with other massive particles. In this note, the results of a study of Higgs bosons produced in association with top quarks and decaying via\ \ are presented. The study was performed as realistically as possible by employing a full and detailed Monte Carlo simulation of the CMS detector followed by the application of trigger and reconstruction algorithms that were developed for use with real data. Important systematic effects resulting from such sources as the uncertainties in the jet energy scale and the estimated rates for correctly tagging b jets or mistagging non-b jets have been taken into account. The impact of large theoretical uncertainties in the cross sections for\ \ plus\ N\ jets processes due to an absence of next-to-leading order calculations is also considered.

}, issn = {0954-3899}, doi = {10.1088/0954-3899/34/5/N03}, url = {https://iopscience.iop.org/article/10.1088/0954-3899/34/5/N03}, author = {Benedetti, D and Cucciarelli, S and Hill, C and Incandela, J and Koay, S A and Riccardi, C and Santocchia, A and Schmidt, A and Torre, P and Weiser, C} } @article {2364, title = {Olfactory coding with all-or-nothing glomeruli.}, journal = {Journal of Neurophysiology}, volume = {98}, year = {2007}, month = {2007 Dec}, pages = {3134-42}, abstract = {

We present a model for olfactory coding based on spatial representation of glomerular responses. In this model distinct odorants activate specific subsets of glomeruli, dependent on the odorant{\textquoteright}s chemical identity and concentration. The glomerular response specificities are understood statistically, based on experimentally measured distributions of activation thresholds. A simple version of the model, in which glomerular responses are binary (the all-or-nothing model), allows us to account quantitatively for the following results of human/rodent olfactory psychophysics: 1) just noticeable differences in the perceived concentration of a single odor (Weber ratios) are as low as dC/C approximately 0.04; 2) the number of simultaneously perceived odors can be as high as 12; and 3) extensive lesions of the olfactory bulb do not lead to significant changes in detection or discrimination thresholds. We conclude that a combinatorial code based on a binary glomerular response is sufficient to account for several important features of the discrimination capacity of the mammalian olfactory system.

}, keywords = {Algorithms, Animals, Discrimination (Psychology), Mice, Models, Neurological, Odors, Olfactory Bulb, Olfactory Pathways, Olfactory Receptor Neurons, Psychomotor Performance, Psychophysics, Rats, Smell}, issn = {0022-3077}, doi = {10.1523/JNEUROSCI.3613-08.2008}, author = {Koulakov, Alexei and Gelperin, Alan and Rinberg, Dmitry} } @article {2155, title = {Phenotype clustering of breast epithelial cells in confocal images based on nuclear protein distribution analysis.}, journal = {BMC Cell Biology}, volume = {8 }, year = {2007}, month = {2007 Jul 10}, pages = {S3}, abstract = {

The distribution of chromatin-associated proteins plays a key role in directing nuclear function. Previously, we developed an image-based method to quantify the nuclear distributions of proteins and showed that these distributions depended on the phenotype of human mammary epithelial cells. Here we describe a method that creates a hierarchical tree of the given cell phenotypes and calculates the statistical significance between them, based on the clustering analysis of nuclear protein distributions.

}, keywords = {Breast, Breast Neoplasms, Cell Line, Cluster Analysis, Computer-Assisted, Confocal, Epithelial Cells, Female, Humans, Image Processing, Imaging, Microscopy, Models, Nuclear Proteins, Phenotype, Statistical, Three-Dimensional}, issn = {1471-2121}, doi = {10.1007/s12021-010-9090-x}, author = {Long, Fuhui and Peng, Hanchuan and Sudar, Damir and Leli{\`e}vre, Sophie A and Knowles, David W} } @article {4251, title = {Rapidly inducible, genetically targeted inactivation of neural and synaptic activity in vivo.}, journal = {Current Opinion in Neurobiology}, volume = {17}, year = {2007}, month = {2007 Oct}, pages = {581-6}, abstract = {

Inducible and reversible perturbation of the activity of selected neurons in vivo is critical to understanding the dynamics of brain circuits. Several genetically encoded systems for rapid inducible neuronal silencing have been developed in the past few years offering an arsenal of tools for in vivo experiments. Some systems are based on ion-channels or pumps, others on G protein coupled receptors, and yet others on modified presynaptic proteins. Inducers range from light to small molecules to peptides. This diversity results in differences in the various parameters that may determine the applicability of each tool to a particular biological question. Although further development would be beneficial, the current silencing tool kit already provides the ability to make specific perturbations of circuit function in behaving animals.

}, keywords = {Animals, Brain, G-Protein-Coupled, Gene Targeting, Neurons, Receptors, Synapses}, issn = {0959-4388}, doi = {10.1016/j.conb.2007.10.002}, author = {Tervo, Dougal and Karpova, Alla Y} } @conference {48185, title = {Shape constrained automatic segmentation of the liver based on a heuristic intensity model.}, booktitle = {MICCAI Workshop 3D Segmentation in the Clinic}, year = {2007}, month = {2007 Oct 29}, address = {Brisbane, Australia}, abstract = {

We present a fully automatic 3D segmentation method for the liver from contrast-enhanced CT data. It is based on a combination of a constrained free-form and statistical deformable model. The adap- tation of the model to the image data is performed according to a simple model of the typical intensity distribution around the liver boundary and neighboring anatomical structures, considering the potential presence of tumors in the liver. All parameters of the deformation as well as the initial positioning of the model in the data are estimated automatically.\ 

}, url = {http://mbi.dkfz-heidelberg.de/grand-challenge2007/web/p109.pdf}, author = {Kainmueller, Dagmar and Lange, Thomas and Lamecker, Hans} } @article {45895, title = {Stability and plasticity of intrinsic membrane properties in hippocampal CA1 pyramidal neurons: effects of internal anions.}, journal = {The Journal of Physiology}, volume = {578}, year = {2007}, month = {2007 Feb 1}, pages = {799-818}, abstract = {

CA1 pyramidal neurons from animals that have acquired hippocampal tasks show increased neuronal excitability, as evidenced by a reduction in the postburst afterhyperpolarization (AHP). Studies of AHP plasticity require stable long-term recordings, which are affected by the intracellular solutions potassium methylsulphate (KMeth) or potassium gluconate (KGluc). Here we show immediate and gradual effects of these intracellular solutions on measurement of the AHP and basic membrane properties, and on the induction of AHP plasticity in CA1 pyramidal neurons from rat hippocampal slices. The AHP measured immediately after establishing whole-cell recordings was larger with KMeth than with KGluc. In general, the AHP in KMeth was comparable to the AHP measured in the perforated-patch configuration. However, KMeth induced time-dependent changes in the intrinsic membrane properties of CA1 pyramidal neurons. Specifically, input resistance progressively increased by 70\% after 50 min; correspondingly, the current required to trigger an action potential and the fast afterdepolarization following action potentials gradually decreased by about 50\%. Conversely, these measures were stable in KGluc. We also demonstrate that activity-dependent plasticity of the AHP occurs with physiologically relevant stimuli in KGluc. AHPs triggered with theta-burst firing every 30 s were progressively reduced, whereas AHPs elicited every 150 s were stable. Blockade of the apamin-sensitive AHP current (I(AHP)) was insufficient to block AHP plasticity, suggesting that plasticity is manifested through changes in the apamin-insensitive slow AHP current (sI(AHP)). These changes were observed in the presence of synaptic blockers, and therefore reflect changes in the intrinsic properties of the neurons. However, no AHP plasticity was observed using KMeth. In summary, these data show that KMeth produces time-dependent changes in basic membrane properties and prevents or obscures activity-dependent reduction of the AHP. In whole-cell recordings using KGluc, repetitive theta-burst firing induced AHP plasticity that mimics learning-related reduction in the AHP.

}, keywords = {Animals, Anions, Apamin, Gluconates, Male, Membrane Potentials, Neuronal Plasticity, Norepinephrine, Patch-Clamp Techniques, Pyramidal Cells, Rats, Rats, Wistar, Sulfuric Acid Esters, Time Factors}, issn = {0022-3751}, doi = {10.1113/jphysiol.2006.124586}, author = {Kaczorowski, Catherine Cook and Disterhoft, John and Spruston, Nelson} } @article {1542, title = {Three-dimensional analysis of a viral RNA replication complex reveals a virus-induced mini-organelle.}, journal = {PLoS Biology}, volume = {5}, year = {2007}, month = {2007 Sep}, pages = {e220}, abstract = {

Positive-strand RNA viruses are the largest genetic class of viruses and include many serious human pathogens. All positive-strand RNA viruses replicate their genomes in association with intracellular membrane rearrangements such as single- or double-membrane vesicles. However, the exact sites of RNA synthesis and crucial topological relationships between relevant membranes, vesicle interiors, surrounding lumens, and cytoplasm generally are poorly defined. We applied electron microscope tomography and complementary approaches to flock house virus (FHV)-infected Drosophila cells to provide the first 3-D analysis of such replication complexes. The sole FHV RNA replication factor, protein A, and FHV-specific 5-bromouridine 5{\textquoteright}-triphosphate incorporation localized between inner and outer mitochondrial membranes inside approximately 50-nm vesicles (spherules), which thus are FHV-induced compartments for viral RNA synthesis. All such FHV spherules were outer mitochondrial membrane invaginations with interiors connected to the cytoplasm by a necked channel of approximately 10-nm diameter, which is sufficient for ribonucleotide import and product RNA export. Tomographic, biochemical, and other results imply that FHV spherules contain, on average, three RNA replication intermediates and an interior shell of approximately 100 membrane-spanning, self-interacting protein As. The results identify spherules as the site of protein A and nascent RNA accumulation and define spherule topology, dimensions, and stoichiometry to reveal the nature and many details of the organization and function of the FHV RNA replication complex. The resulting insights appear relevant to many other positive-strand RNA viruses and support recently proposed structural and likely evolutionary parallels with retrovirus and double-stranded RNA virus virions.

}, keywords = {Animals, Drosophila, Electron, Imaging, Microscopy, Mitochondrial Membranes, Nodaviridae, Organelles, RNA, Three-Dimensional, Viral, Virus Replication}, issn = {1545-7885}, doi = {10.1371/journal.pbio.0050220}, author = {Kopek, Benjamin G and Perkins, Guy and Miller, David J and Ellisman, Mark H and Ahlquist, Paul} } @article {4479, title = {Three-dimensional analysis of a viral RNA replication complex reveals a virus-induced mini-organelle.}, journal = {PLoS biology}, volume = {5}, year = {2007}, month = {2007 Sep}, pages = {e220}, abstract = {

Positive-strand RNA viruses are the largest genetic class of viruses and include many serious human pathogens. All positive-strand RNA viruses replicate their genomes in association with intracellular membrane rearrangements such as single- or double-membrane vesicles. However, the exact sites of RNA synthesis and crucial topological relationships between relevant membranes, vesicle interiors, surrounding lumens, and cytoplasm generally are poorly defined. We applied electron microscope tomography and complementary approaches to flock house virus (FHV)-infected Drosophila cells to provide the first 3-D analysis of such replication complexes. The sole FHV RNA replication factor, protein A, and FHV-specific 5-bromouridine 5{\textquoteright}-triphosphate incorporation localized between inner and outer mitochondrial membranes inside approximately 50-nm vesicles (spherules), which thus are FHV-induced compartments for viral RNA synthesis. All such FHV spherules were outer mitochondrial membrane invaginations with interiors connected to the cytoplasm by a necked channel of approximately 10-nm diameter, which is sufficient for ribonucleotide import and product RNA export. Tomographic, biochemical, and other results imply that FHV spherules contain, on average, three RNA replication intermediates and an interior shell of approximately 100 membrane-spanning, self-interacting protein As. The results identify spherules as the site of protein A and nascent RNA accumulation and define spherule topology, dimensions, and stoichiometry to reveal the nature and many details of the organization and function of the FHV RNA replication complex. The resulting insights appear relevant to many other positive-strand RNA viruses and support recently proposed structural and likely evolutionary parallels with retrovirus and double-stranded RNA virus virions.

}, keywords = {Animals, Drosophila, Electron, Imaging, Microscopy, Mitochondrial Membranes, Nodaviridae, Organelles, RNA, Three-Dimensional, Viral, Virus Replication}, issn = {1545-7885}, doi = {10.1371/journal.pbio.0050220}, url = {http://www.plosbiology.org/article/info\%3Adoi\%2F10.1371\%2Fjournal.pbio.0050220}, author = {Kopek, Benjamin G and Perkins, Guy and Miller, David J and Ellisman, Mark H and Ahlquist, Paul} } @article {1689, title = {Three-dimensional preparation and imaging reveal intrinsic microtubule properties.}, journal = {Nature Methods}, volume = {4}, year = {2007}, month = {2007 Oct}, pages = {843-6}, abstract = {

We present an experimental investigation of microtubule dynamic instability in three dimensions, based on laser light-sheet fluorescence microscopy. We introduce three-dimensional (3D) preparation of Xenopus laevis egg extracts in Teflon-based cylinders and provide algorithms for 3D image processing. Our approach gives experimental access to the intrinsic dynamic properties of microtubules and to microtubule population statistics in single asters. We obtain evidence for a stochastic nature of microtubule pausing.

}, keywords = {Animals, Computer-Assisted, Fluorescence, Image Processing, Imaging, Microscopy, Microtubules, Ovum, Three-Dimensional, Xenopus laevis}, issn = {1548-7091}, doi = {10.1038/nmeth1087}, author = {Keller, Philipp J and Pampaloni, Francesco and Stelzer, Ernst H K} } @article {2400, title = {Transcription of histone gene cluster by differential core-promoter factors.}, journal = {Genes \& Development}, volume = {21}, year = {2007}, month = {2007 Nov 15}, pages = {2936-49}, abstract = {

The 100 copies of tandemly arrayed Drosophila linker (H1) and core (H2A/B and H3/H4) histone gene cluster are coordinately regulated during the cell cycle. However, the molecular mechanisms that must allow differential transcription of linker versus core histones prevalent during development remain elusive. Here, we used fluorescence imaging, biochemistry, and genetics to show that TBP (TATA-box-binding protein)-related factor 2 (TRF2) selectively regulates the TATA-less Histone H1 gene promoter, while TBP/TFIID targets core histone transcription. Importantly, TRF2-depleted polytene chromosomes display severe chromosomal structural defects. This selective usage of TRF2 and TBP provides a novel mechanism to differentially direct transcription within the histone cluster. Moreover, genome-wide chromatin immunoprecipitation (ChIP)-on-chip analyses coupled with RNA interference (RNAi)-mediated functional studies revealed that TRF2 targets several classes of TATA-less promoters of \>1000 genes including those driving transcription of essential chromatin organization and protein synthesis genes. Our studies establish that TRF2 promoter recognition complexes play a significantly more central role in governing metazoan transcription than previously appreciated.

}, keywords = {Animals, Base Sequence, Biological, Cells, Chromatin Immunoprecipitation, Chromosomes, Cluster Analysis, Cultured, Drosophila, Firefly, Fluorescence, Genes, Genetic, Histones, Immunohistochemistry, In Situ Hybridization, Insect, Luciferases, Models, Molecular Sequence Data, Multigene Family, Nucleic Acid, Promoter Regions, Reporter, RNA Interference, Sequence Homology, TATA Box Binding Protein-Like Proteins, TATA-Box Binding Protein, Transcription}, issn = {0890-9369}, doi = {10.1073/pnas.1100640108}, author = {Isogai, Yoh and Keles, S{\"u}nd{\"u}z and Prestel, Matthias and Hochheimer, Andreas and Tjian, Robert} } @article {1410, title = {Cre recombinase-mediated restoration of nigrostriatal dopamine in dopamine-deficient mice reverses hypophagia and bradykinesia.}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {103}, year = {2006}, month = {2006 Jun 6}, pages = {8858-63}, abstract = {

A line of dopamine-deficient (DD) mice was generated to allow selective restoration of normal dopamine signaling to specific brain regions. These DD floxed stop (DDfs) mice have a nonfunctional Tyrosine hydroxylase (Th) gene because of insertion of a NeoR gene flanked by lox P sites targeted to the first intron of the Th gene. DDfs mice have trace brain dopamine content, severe hypoactivity, and aphagia, and they die without intervention. However, they can be maintained by daily treatment with l-3,4-dihydroxyphenylalanine (L-dopa). Injection of a canine adenovirus (CAV-2) engineered to express Cre recombinase into the central caudate putamen restores normal Th gene expression to the midbrain dopamine neurons that project there because CAV-2 efficiently transduces axon terminals and is retrogradely transported to neuronal cell bodies. Bilateral injection of Cre recombinase into the central caudate putamen restores feeding and normalizes locomotion in DDfs mice. Analysis of feeding behavior by using lickometer cages revealed that virally rescued DDfs mice are hyperphagic and have modified meal structures compared with control mice. The virally rescued DDfs mice are also hyperactive at night, have reduced motor coordination, and are thigmotactic compared with controls. These results highlight the critical role for dopamine signaling in the dorsal striatum for most dopamine-dependent behaviors but suggest that dopamine signaling in other brain regions is important to fine-tune these behaviors. This approach offers numerous advantages compared with previous models aimed at examining dopamine signaling in discrete dopaminergic circuits.

}, keywords = {Adenoviridae, Animals, Brain, Dogs, Dopamine, Eating Disorders, Feeding Behavior, Gene Therapy, Hypokinesia, Inbred C57BL, Integrases, Mice, Motor Activity, Tyrosine 3-Monooxygenase, Viral Proteins}, issn = {0027-8424}, doi = {10.1073/pnas.0603081103}, author = {Hnasko, Thomas S and Perez, Francisco A and Scouras, Alex D and Stoll, Elizabeth A and Gale, Samuel D and Luquet, Serge and Phillips, Paul E M and Kremer, Eric J and Palmiter, Richard D} } @article {45891, title = {Dendritic patch-clamp recording.}, journal = {Nature Protocols}, volume = {1}, year = {2006}, month = {2006}, pages = {1235-47}, abstract = {

The patch-clamp technique allows investigation of the electrical excitability of neurons and the functional properties and densities of ion channels. Most patch-clamp recordings from neurons have been made from the soma, the largest structure of individual neurons, while their dendrites, which form the majority of the surface area and receive most of the synaptic input, have been relatively neglected. This protocol describes techniques for recording from the dendrites of neurons in brain slices under direct visual control. Although the basic technique is similar to that used for somatic patching, we describe refinements and optimizations of slice quality, microscope optics, setup stability and electrode approach that are required for maximizing the success rate for dendritic recordings. Using this approach, all configurations of the patch-clamp technique (cell-attached, inside-out, whole-cell, outside-out and perforated patch) can be achieved, even for relatively distal dendrites, and simultaneous multiple-electrode dendritic recordings are also possible. The protocol--from the beginning of slice preparation to the end of the first successful recording--can be completed in 3 h.

}, keywords = {Dendrites, Patch-Clamp Techniques}, issn = {1750-2799}, doi = {10.1038/nprot.2006.164}, author = {Davie, Jenny T and Kole, Maarten H P and Letzkus, Johannes J and Rancz, Ede A and Spruston, Nelson and Stuart, Greg J and H{\"a}usser, Michael} } @article {45909, title = {Distance-dependent differences in synapse number and AMPA receptor expression in hippocampal CA1 pyramidal neurons.}, journal = {Neuron}, volume = {50}, year = {2006}, month = {2006 May 4}, pages = {431-42}, abstract = {

The ability of synapses throughout the dendritic tree to influence neuronal output is crucial for information processing in the brain. Synaptic potentials attenuate dramatically, however, as they propagate along dendrites toward the soma. To examine whether excitatory axospinous synapses on CA1 pyramidal neurons compensate for their distance from the soma to counteract such dendritic filtering, we evaluated axospinous synapse number and receptor expression in three progressively distal regions: proximal and distal stratum radiatum (SR), and stratum lacunosum-moleculare (SLM). We found that the proportion of perforated synapses increases as a function of distance from the soma and that their AMPAR, but not NMDAR, expression is highest in distal SR and lowest in SLM. Computational models of pyramidal neurons derived from these results suggest that they arise from the compartment-specific use of conductance scaling in SR and dendritic spikes in SLM to minimize the influence of distance on synaptic efficacy.

}, keywords = {Animals, Cell Polarity, Cell Shape, Crosses, Genetic, Dendritic Spines, Excitatory Postsynaptic Potentials, Glutamic Acid, Hippocampus, Image Cytometry, Immunohistochemistry, Male, Microscopy, Electron, Transmission, Perforant Pathway, Presynaptic Terminals, Pyramidal Cells, Rats, Rats, Inbred BN, Rats, Inbred F344, Receptors, AMPA, Synapses, Synaptic Transmission}, issn = {0896-6273}, doi = {10.1016/j.neuron.2006.03.022}, author = {Nicholson, Daniel A and Trana, Rachel and Katz, Yael and Kath, William L and Spruston, Nelson and Geinisman, Yuri} } @article {2013, title = {Division accuracy in a stochastic model of Min oscillations in Escherichia coli.}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {103}, year = {2006}, month = {2006 Jan 10}, pages = {347-52}, abstract = {

Accurate cell division in Escherichia coli requires the Min proteins MinC, MinD, and MinE as well as the presence of nucleoids. MinD and MinE exhibit spatial oscillations, moving from pole to pole of the bacterium, resulting in an average MinD concentration that is low at the center of the cell and high at the poles. This concentration minimum is thought to signal the site of cell division. Deterministic models of the Min oscillations reproduce many observed features of the system, including the concentration minimum of MinD. However, there are only a few thousand Min proteins in a bacterium, so stochastic effects are likely to play an important role. Here, we show that Monte Carlo simulations with a large number of proteins agree well with the results from a deterministic treatment of the equations. The location of minimum local MinD concentration is too variable to account for cell division accuracy in wild type but is consistent with the accuracy of cell division in cells without nucleoids. This finding confirms the need to include additional mechanisms, such as reciprocal interactions with the cell division ring or positioning of the nucleoids, to explain wild-type accuracy.

}, keywords = {Adenosine Triphosphatases, Biological, Biophysical Phenomena, Biophysics, Cell Cycle Proteins, Cell Division, Escherichia coli, Escherichia coli Proteins, Models, Monte Carlo Method, Stochastic Processes}, issn = {0027-8424}, doi = {10.1073/pnas.0505825102}, author = {Kerr, Rex A and Levine, Herbert and Sejnowski, Terrence J and Rappel, Wouter-Jan} } @article {792, title = {Gradients of the Drosophila Chinmo BTB-zinc finger protein govern neuronal temporal identity.}, journal = {Cell}, volume = {127}, year = {2006}, month = {2006 Oct 20}, pages = {409-22}, abstract = {

Many neural progenitors, including Drosophila mushroom body (MB) and projection neuron (PN) neuroblasts, sequentially give rise to different subtypes of neurons throughout development. We identified a novel BTB-zinc finger protein, named Chinmo (Chronologically inappropriate morphogenesis), that governs neuronal temporal identity during postembryonic development of the Drosophila brain. In both MB and PN lineages, loss of Chinmo autonomously causes early-born neurons to adopt the fates of late-born neurons from the same lineages. Interestingly, primarily due to a posttranscriptional control, MB neurons born at early developmental stages contain more abundant Chinmo than their later-born siblings. Further, the temporal identity of MB progeny can be transformed toward earlier or later fates by reducing or increasing Chinmo levels, respectively. Taken together, we suggest that a temporal gradient of Chinmo (Chinmo(high) {\textendash}\> Chinmo(low)) helps specify distinct birth order-dependent cell fates in an extended neuronal lineage.

}, keywords = {5{\textquoteright} Untranslated Regions, Amino Acid Sequence, Animals, Brain, Cell Differentiation, Cell Lineage, Developmental, Drosophila, Drosophila Proteins, Gene Expression Regulation, Larva, Molecular Sequence Data, Morphogenesis, Mushroom Bodies, Mutation, Nerve Tissue Proteins, Neurons, Post-Translational, Protein Processing, Time Factors, Transgenes, Zinc Fingers}, issn = {0092-8674}, doi = {10.1016/j.cell.2006.08.045}, author = {Zhu, Sijun and Lin, Suewei and Kao, Chih-Fei and Awasaki, Takeshi and Chiang, Ann-Shyn and Lee, Tzumin} } @article {1543, title = {An in vitro fluorescence screen to identify antivirals that disrupt hepatitis B virus capsid assembly.}, journal = {Nature Biotechnology}, volume = {24}, year = {2006}, month = {2006 Mar}, pages = {358-62}, abstract = {

Virus assembly has not been routinely targeted in the development of antiviral drugs, in part because of the lack of tractable methods for screening in vitro. We have developed an in vitro assay of hepatitis B virus (HBV) capsid assembly, based on fluorescence quenching of dye-labeled capsid protein, for testing potential inhibitors. This assay is adaptable to high-throughput screening and can identify small-molecule inhibitors of virus assembly that prevent, inappropriately accelerate and/or misdirect capsid formation to yield aberrant particles. An in vitro primary screen has the advantage of identifying promising lead compounds affecting assembly without the requirement that they be taken up by cells in culture and be nontoxic. Our approach may facilitate the identification of antivirals targeting viruses other than HBV, such as avian influenza and HIV.

}, keywords = {Antiviral Agents, Biological Assay, Boron Compounds, Capsid, Capsid Proteins, Dimerization, Fluorescence, Fluorescent Dyes, Hepatitis B virus, Kinetics, Light, Models, Molecular, Radiation, Scattering, Virus Assembly}, issn = {1087-0156}, doi = {10.1038/nbt1187}, author = {Stray, Stephen J and Johnson, Jennifer M and Kopek, Benjamin G and Zlotnick, Adam} } @article {45751, title = {Large-scale gene discovery in the pea aphid Acyrthosiphon pisum (Hemiptera).}, journal = {Genome Biol}, volume = {7}, year = {2006}, month = {2006}, pages = {R21}, abstract = {

Aphids are the leading pests in agricultural crops. A large-scale sequencing of 40,904 ESTs from the pea aphid Acyrthosiphon pisum was carried out to define a catalog of 12,082 unique transcripts. A strong AT bias was found, indicating a compositional shift between Drosophila melanogaster and A. pisum. An in silico profiling analysis characterized 135 transcripts specific to pea-aphid tissues (relating to bacteriocytes and parthenogenetic embryos). This project is the first to address the genetics of the Hemiptera and of a hemimetabolous insect.

}, keywords = {Animals, Aphids, Base Composition, Base Sequence, DNA, DNA, Complementary, Expressed Sequence Tags, Gene Library, Microsatellite Repeats, Peas, Phylogeny, Plant Diseases, Population Density, Transcription, Genetic}, issn = {1465-6914}, doi = {10.1186/gb-2006-7-3-r21}, author = {Sabater-Mu{\~n}oz, Beatriz and Legeai, Fabrice and Rispe, Claude and Bonhomme, Jo{\"e}l and Dearden, Peter and Dossat, Carole and Duclert, Aymeric and Gauthier, Jean-Pierre and Ducray, Dani{\`e}le Giblot and Hunter, Wayne and Dang, Phat and Kambhampati, Srini and Martinez-Torres, David and Cortes, Teresa and Moya, Andr{\`e}s and Nakabachi, Atsushi and Philippe, Cathy and Prunier-Leterme, Nathalie and Rahb{\'e}, Yvan and Simon, Jean-Christophe and Stern, David L and Wincker, Patrick and Tagu, Denis} } @article {1690, title = {Life sciences require the third dimension.}, journal = {Current Opinion in Cell Biology}, volume = {18}, year = {2006}, month = {2006 Feb}, pages = {117-24}, abstract = {

Novel technologies are required for three-dimensional cell biology and biophysics. By three-dimensional we refer to experimental conditions that essentially try to avoid hard and flat surfaces and favour unconstrained sample dynamics. We believe that light-sheet-based microscopes are particularly well suited to studies of sensitive three-dimensional biological systems. The application of such instruments can be illustrated with examples from the biophysics of microtubule dynamics and three-dimensional cell cultures. Our experience leads us to suggest that three-dimensional approaches reveal new aspects of a system and enable experiments to be performed in a more physiological and hence clinically more relevant context.

}, keywords = {Animals, Biological Science Disciplines, Cell Line, Computer-Assisted, Dogs, Fluorescence, Image Processing, Imaging, Microscopy, Microtubules, Paclitaxel, Three-Dimensional}, issn = {0955-0674}, doi = {10.1016/j.ceb.2005.12.012}, author = {Keller, Philipp J and Pampaloni, Francesco and Stelzer, Ernst H K} } @article {2444, title = {Local caspase activity directs engulfment of dendrites during pruning.}, journal = {Nature Neuroscience}, volume = {9}, year = {2006}, month = {2006 Oct}, pages = {1234-6}, abstract = {

Pruning is important for sculpting neural circuits, as it removes excessive or inaccurate projections. Here we show that the removal of sensory neuron dendrites during pruning in Drosophila melanogaster is directed by local caspase activity. Suppressing caspase activity prevented dendrite removal, whereas a global activation of caspases within a neuron caused cell death. A new genetically encoded caspase probe revealed that caspase activity is confined to the degenerating dendrites of pruning neurons.

}, keywords = {Afferent, Analysis of Variance, Animals, Apoptosis, Caspases, Dendrites, Drosophila melanogaster, Drosophila Proteins, Enzyme Activation, Enzyme Inhibitors, Fluorescent Antibody Technique, Ganglia, Genetically Modified, Neurons, Spinal, Time Factors}, issn = {1097-6256}, doi = {10.1038/nn1774}, author = {Williams, Darren W and Kondo, Shu and Krzyzanowska, Agnieszka and Hiromi, Yasushi and Truman, James W} } @article {1691, title = {Nud1p, the yeast homolog of Centriolin, regulates spindle pole body inheritance in meiosis.}, journal = {The EMBO Journal}, volume = {25}, year = {2006}, month = {2006 Aug 23}, pages = {3856-68}, abstract = {

Nud1p, a protein homologous to the mammalian centrosome and midbody component Centriolin, is a component of the budding yeast spindle pole body (SPB), with roles in anchorage of microtubules and regulation of the mitotic exit network during vegetative growth. Here we analyze the function of Nud1p during yeast meiosis. We find that a nud1-2 temperature-sensitive mutant has two meiosis-related defects that reflect genetically distinct functions of Nud1p. First, the mutation affects spore formation due to its late function during spore maturation. Second, and most important, the mutant loses its ability to distinguish between the ages of the four spindle pole bodies, which normally determine which SPB would be preferentially included in the mature spores. This affects the regulation of genome inheritance in starved meiotic cells and leads to the formation of random dyads instead of non-sister dyads under these conditions. Both functions of Nud1p are connected to the ability of Spc72p to bind to the outer plaque and half-bridge (via Kar1p) of the SPB.

}, keywords = {Cell Cycle Proteins, Deoxyribonucleases, Fungal, Genome, Humans, Meiosis, Microtubule-Associated Proteins, Mitotic Spindle Apparatus, Mutation, Nuclear Proteins, Protein Binding, Saccharomyces cerevisiae, Saccharomyces cerevisiae Proteins, Spores, tRNA Methyltransferases}, issn = {0261-4189}, doi = {10.1038/sj.emboj.7601254}, author = {Gordon, Oren and Taxis, Christof and Keller, Philipp J and Benjak, Aleksander and Stelzer, Ernst H K and Simchen, Giora and Knop, Michael} } @article {2365, title = {Sparse odor coding in awake behaving mice.}, journal = {The Journal of Neuroscience: The Official Journal of the Society for Neuroscience}, volume = {26}, year = {2006}, month = {2006 Aug 23}, pages = {8857-65}, abstract = {

Responses of mitral cells represent the results of the first stage of odor processing in the olfactory bulb. Most of our knowledge about mitral cell activity has been obtained from recordings in anesthetized animals. We compared odor-elicited changes in firing rate of mitral cells in awake behaving mice and in anesthetized mice. We show that odor-elicited changes in mitral cell firing rate were larger and more frequently observed in the anesthetized than in the awake condition. Only 27\% of mitral cells that showed a response to odors in the anesthetized state were also odor responsive in the awake state. The amplitude of their response in the awake state was smaller, and some of the responses changed sign compared with their responses in the anesthetized state. The odor representation in the olfactory bulb is therefore sparser in awake behaving mice than in anesthetized preparations. A qualitative explanation of the mechanism responsible for this phenomenon is proposed.

}, keywords = {Afferent, Anesthesia, Animals, Electrophysiology, Inbred C57BL, Male, Mice, Neurons, Odors, Olfactory Bulb, Smell, Wakefulness}, issn = {1529-2401}, doi = {10.1523/JNEUROSCI.3613-08.2008}, author = {Rinberg, Dmitry and Koulakov, Alex and Gelperin, Alan} } @article {2366, title = {Speed-accuracy tradeoff in olfaction.}, journal = {Neuron}, volume = {51}, year = {2006}, month = {2006 Aug 3}, pages = {351-8}, abstract = {

The basic psychophysical principle of speed-accuracy tradeoff (SAT) has been used to understand key aspects of neuronal information processing in vision and audition, but the principle of SAT is still debated in olfaction. In this study we present the direct observation of SAT in olfaction. We developed a behavioral paradigm for mice in which both the duration of odorant sampling and the difficulty of the odor discrimination task were controlled by the experimenter. We observed that the accuracy of odor discrimination increases with the duration of imposed odorant sampling, and that the rate of this increase is slower for harder tasks. We also present a unifying picture of two previous, seemingly disparate experiments on timing of odorant sampling in odor discrimination tasks. The presence of SAT in olfaction provides strong evidence for temporal integration in olfaction and puts a constraint on models of olfactory processing.

}, keywords = {Animals, Chemical, Discrimination Learning, Inbred C57BL, Male, Mice, Psychomotor Performance, Reaction Time, Smell, Stimulation}, issn = {0896-6273}, doi = {10.1523/JNEUROSCI.3613-08.2008}, author = {Rinberg, Dmitry and Koulakov, Alexei and Gelperin, Alan} } @article {2367, title = {When good enough is best.}, journal = {Neuron}, volume = {51}, year = {2006}, month = {2006 Aug 3}, pages = {277-8}, abstract = {

In this issue of Neuron, Rinberg et al. show that mice use a speed-accuracy tradeoff in odor discrimination. Shorter sampling results in high performance for easy problems, and enforced longer sampling results in higher accuracy for difficult problems, but mice freely choose intermediate sampling durations and accuracy varies with difficulty. Reward value and task requirements may determine sampling time choice and performance levels.

}, keywords = {Animals, Discrimination Learning, Mice, Reaction Time, Smell}, issn = {0896-6273}, doi = {10.1523/JNEUROSCI.3613-08.2008}, author = {Kay, Leslie M and Beshel, Jennifer and Martin, Claire} } @article {2318, title = {Antipsychotic drugs elevate mRNA levels of presynaptic proteins in the frontal cortex of the rat.}, journal = {Biological Psychiatry}, volume = {57}, year = {2005}, month = {2005 May 1}, pages = {1041-51}, abstract = {

Molecular adaptations are believed to contribute to the mechanism of action of antipsychotic drugs (APDs). We attempted to establish common gene regulation patterns induced by chronic treatment with APDs.

}, keywords = {Animals, Antipsychotic Agents, Cluster Analysis, Corpus Striatum, Frontal Lobe, Gene Expression, Male, Messenger, Microtubules, Oligonucleotide Array Sequence Analysis, Presynaptic Terminals, Rats, Reverse Transcriptase Polymerase Chain Reaction, RNA, Sprague-Dawley, Synaptic Vesicles}, issn = {0006-3223}, doi = {10.3389/fnana.2010.00147}, author = {MacDonald, Matthew L and Eaton, Molly E and Dudman, Joshua T and Konradi, Christine} } @article {45918, title = {Conditional dendritic spike propagation following distal synaptic activation of hippocampal CA1 pyramidal neurons.}, journal = {Nat Neurosci}, volume = {8}, year = {2005}, month = {2005 Dec}, pages = {1667-76}, abstract = {

The perforant-path projection to the hippocampus forms synapses in the apical tuft of CA1 pyramidal neurons. We used computer modeling to examine the function of these distal synaptic inputs, which led to three predictions that we confirmed in experiments using rat hippocampal slices. First, activation of CA1 neurons by the perforant path is limited, a result of the long distance between these inputs and the soma. Second, activation of CA1 neurons by the perforant path depends on the generation of dendritic spikes. Third, the forward propagation of these spikes is unreliable, but can be facilitated by modest activation of Schaffer-collateral synapses in the upper apical dendrites. This {\textquoteright}gating{\textquoteright} of dendritic spike propagation may be an important activation mode of CA1 pyramidal neurons, and its modulation by neurotransmitters or long-term, activity-dependent plasticity may be an important feature of dendritic integration during mnemonic processing in the hippocampus.

}, keywords = {Action Potentials, Animals, Dendrites, Hippocampus, Long-Term Potentiation, Long-Term Synaptic Depression, Male, Memory, Neural Inhibition, Neural Networks (Computer), Neural Pathways, Neurotransmitter Agents, Organ Culture Techniques, Patch-Clamp Techniques, Perforant Pathway, Pyramidal Cells, Rats, Rats, Wistar, Synaptic Transmission}, issn = {1097-6256}, doi = {10.1038/nn1599}, author = {Jarsky, Tim and Roxin, Alex and Kath, William L and Spruston, Nelson} } @article {45924, title = {Factors mediating powerful voltage attenuation along CA1 pyramidal neuron dendrites.}, journal = {J Physiol}, volume = {568}, year = {2005}, month = {2005 Oct 1}, pages = {69-82}, abstract = {

We performed simultaneous patch-electrode recordings from the soma and apical dendrite of CA1 pyramidal neurons in hippocampal slices, in order to determine the degree of voltage attenuation along CA1 dendrites. Fifty per cent attenuation of steady-state somatic voltage changes occurred at a distance of 238 microm from the soma in control and 409 microm after blocking the hyperpolarization-activated (H) conductance. The morphology of three neurons was reconstructed and used to generate computer models, which were adjusted to fit the somatic and dendritic voltage responses. These models identify several factors contributing to the voltage attenuation along CA1 dendrites, including high axial cytoplasmic resistivity, low membrane resistivity, and large H conductance. In most cells the resting membrane conductances, including the H conductances, were larger in the dendrites than the soma. Simulations suggest that synaptic potentials attenuate enormously as they propagate from the dendrite to the soma, with greater than 100-fold attenuation for synapses on many small, distal dendrites. A prediction of this powerful EPSP attenuation is that distal synaptic inputs are likely only to be effective in the presence of conductance scaling, dendritic excitability, or both.

}, keywords = {Animals, Dendrites, Excitatory Postsynaptic Potentials, Hippocampus, In Vitro Techniques, Male, Models, Neurological, Neural Conduction, Patch-Clamp Techniques, Pyramidal Cells, Rats, Rats, Wistar, Synaptic Transmission}, issn = {0022-3751}, doi = {10.1113/jphysiol.2005.086793}, author = {Golding, Nace L and Mickus, Timothy J and Katz, Yael and Kath, William L and Spruston, Nelson} } @article {2369, title = {How to find decision makers in neural networks.}, journal = {Biological Cybernetics}, volume = {93}, year = {2005}, month = {2005 Dec}, pages = {447-62}, abstract = {

Nervous systems often face the problem of classifying stimuli and making decisions based on these classifications. The neurons involved in these tasks can be characterized as sensory or motor, according to their correlation with sensory stimulus or motor response. In this study we define a third class of neurons responsible for making perceptual decisions. Our mathematical formalism enables the weighting of neuronal units according to their contribution to decision making, thus narrowing the field for more detailed studies of underlying mechanisms. We develop two definitions of a contribution to decision making. The first definition states that decision making activity can be found at the points of emergence for behavioral correlations in the system. The second definition involves the study of propagation of noise in the network. The latter definition is shown to be equivalent to the first one in the cases when they can be compared. Our results suggest a new approach to analyzing decision making networks.

}, keywords = {Animals, Computer Simulation, Decision Making, Electric Stimulation, Humans, Models, Nerve Net, Neural Networks (Computer), Neurological, Neurons, Nonlinear Dynamics, Stochastic Processes}, issn = {0340-1200}, doi = {10.1523/JNEUROSCI.3613-08.2008}, author = {Koulakov, Alexei A and Rinberg, Dmitry A and Tsigankov, Dmitry N} } @article {2412, title = {In vitro analysis of huntingtin-mediated transcriptional repression reveals multiple transcription factor targets.}, journal = {Cell}, volume = {123}, year = {2005}, month = {2005 Dec 29}, pages = {1241-53}, abstract = {

Transcriptional dysregulation has emerged as a potentially important pathogenic mechanism in Huntington{\textquoteright}s disease, a neurodegenerative disorder associated with polyglutamine expansion in the huntingtin (htt) protein. Here, we report the development of a biochemically defined in vitro transcription assay that is responsive to mutant htt. We demonstrate that both gene-specific activator protein Sp1 and selective components of the core transcription apparatus, including TFIID and TFIIF, are direct targets inhibited by mutant htt in a polyglutamine-dependent manner. The RAP30 subunit of TFIIF specifically interacts with mutant htt both in vitro and in vivo to interfere with formation of the RAP30-RAP74 native complex. Importantly, overexpression of RAP30 in cultured primary striatal cells protects neurons from mutant htt-induced cellular toxicity and alleviates the transcriptional inhibition of the dopamine D2 receptor gene by mutant htt. Our results suggest a mutant htt-directed repression mechanism involving multiple specific components of the basal transcription apparatus.

}, keywords = {Animals, Biological Assay, Cell-Free System, Cells, Cercopithecus aethiops, COS Cells, Cultured, Dopamine D2, Down-Regulation, Genetic, Mice, Mutation, Nerve Tissue Proteins, Neurons, Nuclear Proteins, Protein Binding, Protein Subunits, Rats, Receptors, Sp1 Transcription Factor, TFII, Transcription, Transcription Factor TFIID, Transcription Factors}, issn = {0092-8674}, doi = {10.1073/pnas.1100640108}, author = {Zhai, Weiguo and Jeong, Hyunkyung and Cui, Libin and Krainc, Dimitri and Tjian, Robert} } @article {46399, title = {Rapid and reversible chemical inactivation of synaptic transmission in genetically targeted neurons.}, journal = {Neuron}, volume = {48}, year = {2005}, month = {2005 Dec 8}, pages = {727-35}, abstract = {

Inducible and reversible silencing of selected neurons in vivo is critical to understanding the structure and dynamics of brain circuits. We have developed Molecules for Inactivation of Synaptic Transmission (MISTs) that can be genetically targeted to allow the reversible inactivation of neurotransmitter release. MISTs consist of modified presynaptic proteins that interfere with the synaptic vesicle cycle when crosslinked by small molecule "dimerizers." MISTs based on the vesicle proteins VAMP2/Synaptobrevin and Synaptophysin induced rapid ( approximately 10 min) and reversible block of synaptic transmission in cultured neurons and brain slices. In transgenic mice expressing MISTs selectively in Purkinje neurons, administration of dimerizer reduced learning and performance of the rotarod behavior. MISTs allow for specific, inducible, and reversible lesions in neuronal circuits and may provide treatment of disorders associated with neuronal hyperactivity.

}, keywords = {Animals, Cells, Cultured, Cross-Linking Reagents, Dimerization, Gene Targeting, In Vitro Techniques, Learning, Mice, Mice, Transgenic, Motor Activity, Neural Inhibition, Neurons, Neurotransmitter Agents, Purkinje Cells, Synaptic Transmission, Synaptic Vesicles, Synaptophysin, Time Factors, Vesicle-Associated Membrane Protein 2}, issn = {0896-6273}, doi = {10.1016/j.neuron.2005.11.015}, author = {Karpova, Alla Y and Tervo, Dougal G R and Gray, Noah W and Svoboda, Karel} } @article {1692, title = {Spore number control and breeding in Saccharomyces cerevisiae: a key role for a self-organizing system.}, journal = {The Journal of Cell Biology}, volume = {171}, year = {2005}, month = {2005 Nov 21}, pages = {627-40}, abstract = {

Spindle pole bodies (SPBs) provide a structural basis for genome inheritance and spore formation during meiosis in yeast. Upon carbon source limitation during sporulation, the number of haploid spores formed per cell is reduced. We show that precise spore number control (SNC) fulfills two functions. SNC maximizes the production of spores (1-4) that are formed by a single cell. This is regulated by the concentration of three structural meiotic SPB components, which is dependent on available amounts of carbon source. Using experiments and computer simulation, we show that the molecular mechanism relies on a self-organizing system, which is able to generate particular patterns (different numbers of spores) in dependency on one single stimulus (gradually increasing amounts of SPB constituents). We also show that SNC enhances intratetrad mating, whereby maximal amounts of germinated spores are able to return to a diploid lifestyle without intermediary mitotic division. This is beneficial for the immediate fitness of the population of postmeiotic cells.

}, keywords = {Biological, Carbon, Centromere, Computer Simulation, Cytoplasm, Dominant, Electron, Fungal, Gene Expression Regulation, Genes, Genetic, Genetic Linkage, Genotype, Green Fluorescent Proteins, Haploidy, Meiosis, Microscopy, Mitotic Spindle Apparatus, Models, Mutation, Open Reading Frames, Plasmids, Recombination, Saccharomyces cerevisiae, Saccharomyces cerevisiae Proteins, Software, Spores, Time Factors, Video}, issn = {0021-9525}, doi = {10.1083/jcb.200507168}, author = {Taxis, Christof and Keller, Philipp and Kavagiou, Zaharoula and Jensen, Lars Juhl and Colombelli, Julien and Bork, Peer and Stelzer, Ernst H K and Knop, Michael} } @article {3086, title = {Tetrodotoxin infusions into the dorsal hippocampus block non-locomotor place recognition.}, journal = {Hippocampus}, volume = {15}, year = {2005}, month = {2005 Mar 2}, pages = {460-71}, abstract = {

The hippocampus is critical for navigation in an open field. One component of this navigation requires the subject to recognize the target place using distal cues. The experiments presented in this report tested whether blocking hippocampal function would impair open field place recognition. Hungry rats were trained to press a lever on a feeder for food. In Experiment 1, they were passively transported with the feeder along a circular trajectory. Lever pressing was reinforced only if the feeder was passing through a 60 degrees -wide sector. Thus, rats preferentially lever pressed in the vicinity of the reward sector indicating that they recognized its location. Tetrodotoxin (TTX) infusions aimed at the dorsal hippocampi caused rats to substantially increase lever pressing with no preference for any region. The aim of Experiment 2 was to determine whether the TTX injections caused a loss of place recognition or a general increase of lever pressing. A separate group of rats was conditioned in a stationary apparatus to press the lever in response to a light. The TTX injections did not abolish preferential lever pressing in response to light. Lever pressing increased less than half as much as the TTX-induced increase in Experiment 1. When these animals with functional hippocampi could not determine the rewarded period because the light was always off, lever pressing increased much more and was similar to the TTX-induced increase in Experiment 1. We conclude that the TTX inactivation of the hippocampi impaired the ability to recognize the reward place.

}, keywords = {Anesthetics, Animal, Animals, Behavior, Disease Models, Feeding Behavior, Hippocampus, Local, Long-Evans, Male, Memory, Memory Disorders, Orientation, Rats, Space Perception, Tetrodotoxin}, issn = {1050-9631}, doi = {10.1002/hipo.20072}, author = {Klement, Daniel and Pastalkova, Eva and Fenton, Andr{\'e} A} } @article {45956, title = {Aquaporin-0 membrane junctions form upon proteolytic cleavage.}, journal = {Journal of Molecular Biology}, volume = {342}, year = {2004}, month = {2004 Sep 24}, pages = {1337-45}, abstract = {

Aquaporin-0 (AQP0), previously known as major intrinsic protein (MIP), is the only water pore protein expressed in lens fiber cells. AQP0 is highly specific to lens fiber cells and constitutes the most abundant intrinsic membrane protein in these cells. The protein is initially expressed as a full-length protein in young fiber cells in the lens cortex, but becomes increasingly cleaved in the lens core region. Reconstitution of AQP0 isolated from the core of sheep lenses containing a proportion of truncated protein, produced double-layered two-dimensional (2D) crystals, which displayed the same dimensions as the thin 11 nm lens fiber cell junctions, which are prominent in the lens core. In contrast reconstitution of full-length AQP0 isolated from the lens cortex reproducibly yielded single-layered 2D crystals. We present electron diffraction patterns and projection maps of both crystal types. We show that cleavage of the intracellular C terminus enhances the adhesive properties of the extracellular surface of AQP0, indicating a conformational change in the molecule. This change of function of AQP0 from a water pore in the cortex to an adhesion molecule in the lens core constitutes another manifestation of the gene sharing concept originally proposed on the basis of the dual function of crystallins.

}, keywords = {Amino Acid Sequence, Animals, Aquaporins, Biopolymers, Eye Proteins, Hydrolysis, Membrane Glycoproteins, Microscopy, Electron, Molecular Sequence Data, Sheep}, issn = {0022-2836}, doi = {10.1016/j.jmb.2004.07.076}, author = {Gonen, Tamir and Cheng, Yifan and Kistler, Joerg and Walz, Thomas} } @article {45955, title = {Aquaporin-0 membrane junctions reveal the structure of a closed water pore.}, journal = {Nature}, volume = {429}, year = {2004}, month = {2004 May 13}, pages = {193-7}, abstract = {

The lens-specific water pore aquaporin-0 (AQP0) is the only aquaporin known to form membrane junctions in vivo. We show here that AQP0 from the lens core, containing some carboxy-terminally cleaved AQP0, forms double-layered crystals that recapitulate in vivo junctions. We present the structure of the AQP0 membrane junction as determined by electron crystallography. The junction is formed by three localized interactions between AQP0 molecules in adjoining membranes, mainly mediated by proline residues conserved in AQP0s from different species but not present in most other aquaporins. Whereas all previously determined aquaporin structures show the pore in an open conformation, the water pore is closed in AQP0 junctions. The water pathway in AQP0 also contains an additional pore constriction, not seen in other known aquaporin structures, which may be responsible for pore gating.

}, keywords = {Animals, Aquaporins, Biological Transport, Cell Membrane, Cell Membrane Permeability, Cryoelectron Microscopy, Crystallization, Electric Conductivity, Eye Proteins, Hydrogen-Ion Concentration, Lens, Crystalline, Membrane Glycoproteins, Models, Molecular, Molecular Sequence Data, Protein Conformation, Sheep, Water}, issn = {1476-4687}, doi = {10.1038/nature02503}, author = {Gonen, Tamir and Sliz, Piotr and Kistler, Joerg and Cheng, Yifan and Walz, Thomas} } @article {1775, title = {Decoding cilia function: defining specialized genes required for compartmentalized cilia biogenesis.}, journal = {Cell}, volume = {117}, year = {2004}, month = {2004 May 14}, pages = {527-39}, abstract = {

The evolution of the ancestral eukaryotic flagellum is an example of a cellular organelle that became dispensable in some modern eukaryotes while remaining an essential motile and sensory apparatus in others. To help define the repertoire of specialized proteins needed for the formation and function of cilia, we used comparative genomics to analyze the genomes of organisms with prototypical cilia, modified cilia, or no cilia and identified approximately 200 genes that are absent in the genomes of nonciliated eukaryotes but are conserved in ciliated organisms. Importantly, over 80\% of the known ancestral proteins involved in cilia function are included in this small collection. Using Drosophila as a model system, we then characterized a novel family of proteins (OSEGs: outer segment) essential for ciliogenesis. We show that osegs encode components of a specialized transport pathway unique to the cilia compartment and are related to prototypical intracellular transport proteins.

}, keywords = {Afferent, Animals, Carrier Proteins, Cell Compartmentation, Cell Differentiation, Cilia, Developmental, Drosophila melanogaster, Electron, Eukaryotic Cells, Evolution, Gene Expression Regulation, Genome, Humans, Microscopy, Molecular, Neurons, Oligonucleotide Array Sequence Analysis, Phylogeny}, issn = {0092-8674}, author = {Avidor-Reiss, Tomer and Maer, Andreia M and Koundakjian, Edmund and Polyanovsky, Andrey and Keil, Thomas and Subramaniam, Shankar and Zuker, Charles S} } @article {6364, title = {Efficient transformation of the beetle Tribolium castaneum using the Minos transposable element: quantitative and qualitative analysis of genomic integration events.}, journal = {Genetics}, volume = {167}, year = {2004}, month = {2004 Jun}, pages = {737-46}, abstract = {

Genetic transformation in insects holds great promise as a tool for genetic manipulation in species of particular scientific, economic, or medical interest. A number of transposable elements have been tested recently as potential vectors for transformation in a range of insects. Minos is one of the most promising elements because it appears to be active in diverse species and has the capacity to carry large inserts. We report here the use of the Minos element as a transformation vector in the red flour beetle Tribolium castaneum (Coleoptera), an important species for comparative developmental and pest management studies. Transgenic G(1) beetles were recovered from 32.4\% of fertile G(0){\textquoteright}s injected with a plasmid carrying a 3xP3-EGFP-marked transposon and in vitro synthesized mRNA encoding the Minos transposase. This transformation efficiency is 2.8-fold higher than that observed when using a plasmid helper. Molecular and genetic analyses show that several independent insertions can be recovered from a single injected parent, but that the majority of transformed individuals carry single Minos insertions. These results establish Minos as one of the most efficient vectors for genetic transformation in insects. In combination with piggyBac-based transgenesis, our work allows the introduction of sophisticated multicomponent genetic tools in Tribolium.

}, keywords = {Animals, Base Sequence, Crosses, DNA Transposable Elements, Female, Genetic, Genome, Insertional, Male, Mutagenesis, Plasmids, Transformation, Transposases, Tribolium}, issn = {0016-6731}, doi = {10.1534/genetics.103.023085}, author = {Pavlopoulos, Anastasios and Berghammer, Andreas J and Averof, Michalis and Klingler, Martin} } @article {6442, title = {Functional magnetic resonance imaging of macaque monkeys performing visually guided saccade tasks: comparison of cortical eye fields with humans.}, journal = {Neuron}, volume = {41}, year = {2004}, month = {2004 Mar 4}, pages = {795-807}, abstract = {

The frontal and parietal eye fields serve as functional landmarks of the primate brain, although their correspondences between humans and macaque monkeys remain unclear. We conducted fMRI at 4.7 T in monkeys performing visually-guided saccade tasks and compared brain activations with those in humans using identical paradigms. Among multiple parietal activations, the dorsal lateral intraparietal area in monkeys and an area in the posterior superior parietal lobule in humans exhibited the highest selectivity to saccade directions. In the frontal cortex, the selectivity was highest at the junction of the precentral and superior frontal sulci in humans and in the frontal eye field (FEF) in monkeys. BOLD activation peaks were also found in premotor areas (BA6) in monkeys, which suggests that the apparent discrepancy in location between putative human FEF (BA6, suggested by imaging studies) and monkey FEF (BA8, identified by microstimulation studies) partly arose from methodological differences.

}, keywords = {Analysis of Variance, Animals, Cerebral Cortex, Female, Humans, Macaca, Magnetic Resonance Imaging, Male, Photic Stimulation, Psychomotor Performance, Rats, Saccades, Species Specificity, Visual Fields}, issn = {0896-6273}, author = {Koyama, Minoru and Hasegawa, Isao and Osada, Takahiro and Adachi, Yusuke and Nakahara, Kiyoshi and Miyashita, Yasushi} } @article {2319, title = {Individual differences in trait anxiety predict the response of the basolateral amygdala to unconsciously processed fearful faces.}, journal = {Neuron}, volume = {44}, year = {2004}, month = {2004 Dec 16}, pages = {1043-55}, abstract = {

Responses to threat-related stimuli are influenced by conscious and unconscious processes, but the neural systems underlying these processes and their relationship to anxiety have not been clearly delineated. Using fMRI, we investigated the neural responses associated with the conscious and unconscious (backwardly masked) perception of fearful faces in healthy volunteers who varied in threat sensitivity (Spielberger trait anxiety scale). Unconscious processing modulated activity only in the basolateral subregion of the amygdala, while conscious processing modulated activity only in the dorsal amygdala (containing the central nucleus). Whereas activation of the dorsal amygdala by conscious stimuli was consistent across subjects and independent of trait anxiety, activity in the basolateral amygdala to unconscious stimuli, and subjects{\textquoteright} reaction times, were predicted by individual differences in trait anxiety. These findings provide a biological basis for the unconscious emotional vigilance characteristic of anxiety and a means for investigating the mechanisms and efficacy of treatments for anxiety.

}, keywords = {Adult, Amygdala, Anxiety, Brain Mapping, Emotions, Facial Expression, Fear, Female, Humans, Individuality, Male, Photic Stimulation, Predictive Value of Tests, Reaction Time, Unconscious (Psychology)}, issn = {0896-6273}, doi = {10.3389/fnana.2010.00147}, author = {Etkin, Amit and Klemenhagen, Kristen C and Dudman, Joshua T and Rogan, Michael T and Hen, Ren{\'e} and Kandel, Eric R and Hirsch, Joy} } @article {2320, title = {L-type Ca2+ channel blockers promote Ca2+ accumulation when dopamine receptors are activated in striatal neurons.}, journal = {Brain Research. Molecular Brain Research}, volume = {131}, year = {2004}, month = {2004 Nov 24}, pages = {65-72}, abstract = {

Dopamine (DA) receptor-mediated signal transduction and gene expression play a central role in many brain disorders from schizophrenia to Parkinson{\textquoteright}s disease to addiction. While trying to evaluate the role of L-type Ca2+ channels in dopamine D1 receptor-mediated phosphorylation of the transcription factor cyclic AMP response element-binding protein (CREB), we found that activation of dopamine D1 receptors alters the properties of L-type Ca2+ channel inhibitors and turns them into facilitators of Ca2+ influx. In D1 receptor-stimulated neurons, L-type Ca2+ channel blockers promote cytosolic Ca2+ accumulation. This leads to the activation of a molecular signal transduction pathway and CREB phosphorylation. In the absence of dopamine receptor stimulation, L-type Ca2+ channel blockers inhibit CREB phosphorylation. The effect of dopamine on L-type Ca2+ channel blockers is dependent on protein kinase A (PKA), suggesting that protein phosphorylation plays a role in this phenomenon. Because of the adverse effect of activated dopamine receptors on L-type Ca2+ channel blocker action, the role of L-type Ca2+ channels in the dopamine D1 receptor signal transduction pathway cannot be assessed with pharmacological tools. However, with antisense technology, we demonstrate that L-type Ca2+ channels contribute to D1 receptor-mediated CREB phosphorylation. We conclude that the D1 receptor signal transduction pathway depends on L-type Ca2+ channels to mediate CREB phosphorylation.

}, keywords = {2, 3, 4, 5-Tetrahydro-7, 8-dihydroxy-1-phenyl-1H-3-benzazepine, Animals, Antisense, Calcium, Calcium Channel Blockers, Calcium Channels, Cells, Corpus Striatum, Cultured, Cyclic AMP Response Element-Binding Protein, Dizocilpine Maleate, Dopamine Antagonists, Dopamine D1, Drug Interactions, Excitatory Amino Acid Antagonists, Female, L-Type, N-Methyl-D-Aspartate, Neurons, Nifedipine, Oligonucleotides, Phosphorylation, Pregnancy, Rats, Receptors, Sprague-Dawley}, issn = {0169-328X}, doi = {10.3389/fnana.2010.00147}, author = {Eaton, Molly E and Mac{\'\i}as, Wendy and Youngs, Rachael M and Rajadhyaksha, Anjali and Dudman, Joshua T and Konradi, Christine} } @article {1256, title = {Maps in the brain: what can we learn from them?}, journal = {Annual Review of Neuroscience}, volume = {27}, year = {2004}, month = {2004}, pages = {369-92}, abstract = {

In mammalian visual cortex, neurons are organized according to their functional properties into multiple maps such as retinotopic, ocular dominance, orientation preference, direction of motion, and others. What determines the organization of cortical maps? We argue that cortical maps reflect neuronal connectivity in intracortical circuits. Because connecting distant neurons requires costly wiring (i.e., axons and dendrites), there is an evolutionary pressure to place connected neurons as close to each other as possible. Then, cortical maps may be viewed as solutions that minimize wiring cost for given intracortical connectivity. These solutions can help us in inferring intracortical connectivity and, ultimately, in understanding the function of the visual system.

}, keywords = {Animals, Axons, Brain Mapping, Dendrites, Humans, Nerve Net, Neural Pathways, Visual Cortex, Visual Fields, Visual Pathways}, issn = {0147-006X}, doi = {10.1016/j.tins.2005.05.006}, author = {Chklovskii, Dmitri B and Koulakov, Alexei A} } @article {2349, title = {Object-location memory impairment in patients with thermal lesions to the right or left hippocampus.}, journal = {Neuropsychologia}, volume = {42}, year = {2004}, month = {2004 Feb 25}, pages = {1017-28}, abstract = {

Memory for object-location was investigated by testing subjects with small unilateral thermolesions to the medial temporal lobe using small-scale 2D (Abstract) or large-scale 3D (Real) recall conditions. Four patients with lesions of the left hippocampus (LH), 10 patients with damage to the right hippocampus (RH) and 9 matched normal controls (NC) were tested. Six task levels were presented in a pseudorandom order. During each level, subjects viewed one to six different objects on the floor of a circular curtained arena 2.90 m in diameter for 10 s. Recall was tested by marking the locations of objects on a map of the arena (Abstract recall) and then by replacing the objects in the arena (Real recall). Two component errors were studied by calculating the Location Error (LE), independent of the object identity and the configuration error by finding the best match to the presented configuration. The RH group was impaired relative to the NC for nearly all combinations of recall and error types. An impairment was observed in this group even for one object and it deepened sharply with an increasing object number. Damage to the right perirhinal or parahippocampal cortices did not add to the impairment. Deficits in the LH group were also observed, but less consistently. The data indicate that spatial memory is strongly but not exclusively lateralised to the right medial temporal lobe.

}, keywords = {Adult, Anterior Temporal Lobectomy, Brain Damage, Brain Mapping, Cerebral, Chronic, Dominance, Electrocoagulation, Epilepsy, Female, Hippocampus, Humans, Male, Memory, Mental Recall, Middle Aged, Orientation, Pattern Recognition, Postoperative Complications, Reference Values, Short-Term, Temporal Lobe, Visual}, issn = {0028-3932}, doi = {10.1016/j.neuropsychologia.2004.01.002}, author = {Stepankova, Katerina and Fenton, Andr{\'e} A and Pastalkova, Eva and Kalina, Miroslav and Bohbot, V{\'e}ronique D} } @article {2416, title = {Polybromo protein BAF180 functions in mammalian cardiac chamber maturation.}, journal = {Genes \& Development}, volume = {18}, year = {2004}, month = {2004 Dec 15}, pages = {3106-16}, abstract = {

BAF and PBAF are two related mammalian chromatin remodeling complexes essential for gene expression and development. PBAF, but not BAF, is able to potentiate transcriptional activation in vitro mediated by nuclear receptors, such as RXRalpha, VDR, and PPARgamma. Here we show that the ablation of PBAF-specific subunit BAF180 in mouse embryos results in severe hypoplastic ventricle development and trophoblast placental defects, similar to those found in mice lacking RXRalpha and PPARgamma. Embryonic aggregation analyses reveal that in contrast to PPARgamma-deficient mice, the heart defects are likely a direct result of BAF180 ablation, rather than an indirect consequence of trophoblast placental defects. We identified potential target genes for BAF180 in heart development, such as S100A13 as well as retinoic acid (RA)-induced targets RARbeta2 and CRABPII. Importantly, BAF180 is recruited to the promoter of these target genes and BAF180 deficiency affects the RA response for CRABPII and RARbeta2. These studies reveal unique functions of PBAF in cardiac chamber maturation.

}, keywords = {Animals, Cell Aggregation, Chromatin Assembly and Disassembly, Congenital, Developmental, DNA Primers, Gene Expression Regulation, Heart Defects, Immunoprecipitation, In Situ Hybridization, In Situ Nick-End Labeling, Knockout, Mice, Nuclear Proteins, Oligonucleotide Array Sequence Analysis, Placenta, Receptors, Retinoic Acid, Reverse Transcriptase Polymerase Chain Reaction, S100 Proteins, Transcription Factors}, issn = {0890-9369}, doi = {10.1073/pnas.1100640108}, author = {Wang, Zhong and Zhai, Weiguo and Richardson, James A and Olson, Eric N and Meneses, Juanito J and Firpo, Meri T and Kang, Chulho and Skarnes, William C and Tjian, Robert} } @article {1189, title = {Recombination of human mitochondrial DNA.}, journal = {Science}, volume = {304}, year = {2004}, month = {2004 May 14}, pages = {981}, keywords = {DNA, DNA Replication, Fathers, Female, Genetic, Humans, Male, Mitochondria, Mitochondrial, Mothers, Muscle, Polymerase Chain Reaction, Recombination}, issn = {1095-9203}, doi = {10.1126/science.1096342}, author = {Kraytsberg, Yevgenya and Schwartz, Marianne and Brown, Timothy A and Ebralidse, Konstantin and Kunz, Wolfram S and Clayton, David A and Vissing, John and Khrapko, Konstantin} } @proceedings {4534, title = {A smorgasbord of features for statistical machine translation.}, year = {2004}, month = {2004 May}, abstract = {

We describe a methodology for rapid experimentation in statistical machine translation which we use to add a large number of features to a baseline system exploiting features from a wide range of levels of syntactic representation. Feature values were combined in a log-linear model to select the highest scoring candidate translation from an n-best list. Feature weights were optimized directly against the BLEU evaluation metric on held-out data. We present results for a small selection of features at each level of syntactic representation.

}, author = {Och, Franz J. and Gildea, Daniel and Khudanpur, Sanjeev and Sarkar, Anoop and Yamada, Kenji and Fraser, Alex and Kumar, Shankar and Shen, Libin and Smith, David and Eng, Katherine and Jain, Viren and Jin, Zhen and Radev, Dragomir} } @article {2350, title = {A battery of tests for quantitative examination of idiothetic and allothetic place navigation modes in humans.}, journal = {Behavioural Brain Research}, volume = {147}, year = {2003}, month = {2003 Dec 17}, pages = {95-105}, abstract = {

Research into the neural mechanisms of place navigation in laboratory animals has led to the definition of allothetic and idiothetic navigation modes that can be examined by quantitative analysis of the generated tracks. In an attempt to use this approach in the study of human navigation behavior, 10 young subjects were examined in an enclosed arena (2.9 m in diameter, 3 m high) equipped with a computerized tracking system. Idiothetic navigation was studied in blindfolded subjects performing the following tasks-Simple Homing, Complex Homing and Idiothesis Supported by Floor-Related Signals. Allothetic navigation was examined in sighted subjects instructed to find in an empty arena the acoustically signaled unmarked goal region and later to retrieve its position using tasks (Natural Navigation, Cue-Controlled Navigation, Snapshot Memory, Map Reading) that evaluated different aspects of allothesis. The results indicate that allothetic navigation is more accurate than idiothetic, that the poor accuracy of idiothesis is due to angular rather than to distance errors, and that navigation performance is best when both allothetic and idiothetic modes contribute to the solution of the task. The proposed test battery may contribute to better understanding of the navigation disturbances accompanying various neurological disorders and to objective evaluation of the results of drug therapy and of rehabilitation procedures.

}, keywords = {Adult, Cues, Exploratory Behavior, Female, Humans, Locomotion, Male, Memory, Orientation, Space Perception, Spatial Behavior}, issn = {0166-4328}, doi = {10.1016/S0166-4328(03)00141-4}, author = {Stepankova, Katerina and Pastalkova, Eva and Kalova, Eva and Kalina, Miroslav and Bures, Jan} } @article {45766, title = {A comparison of parthenogenetic and sexual embryogenesis of the pea aphid Acyrthosiphon pisum (Hemiptera: Aphidoidea).}, journal = {J Exp Zool B Mol Dev Evol}, volume = {295}, year = {2003}, month = {2003 Feb 15}, pages = {59-81}, abstract = {

Aphids exhibit divergent modes of embryogenesis during the sexual and asexual phases of the life cycle. To explore how a single genome can give rise to these alternative developmental modes, we have initiated embryological studies of the pea aphid, Acyrthosiphon pisum. Here we present a detailed description of parthenogenetic, viviparous embryonic development in the pea aphid. We compare and contrast development of the parthenogenetic embryo with that of the embryo resulting from sexual reproduction. The primary difference between the embryos is the scale on which development occurs: early parthenogenetic development occurs in a volume approximately three orders of magnitude smaller than the sexual egg, largely because of the apparent absence of yolk in the parthenogenetic egg. This results in a drastically different duration of syncytial energid cleavage and, presumably, patterning processes in the two embryos must act at scales that differ by orders of magnitude. The eggs also develop on time scales that differ approximately by an order of magnitude and the timing of the embryonic movements, collectively called blastokinesis, have temporally shifted relative to growth of the embryo. In addition, the endosymbiotic bacteria are transferred from mother to embryo in different ways in the two embryos. Finally, the function of the serosa has diverged greatly in the two embryos: in the sexual egg the serosa deposits a thick cuticle that protects the egg, whereas the serosa of the parthenogenetic embryo is greatly reduced and its function is unclear. The pea aphid is a useful model system for examining how a single genome has evolved to allow divergent modes of development.

}, keywords = {Animals, Aphids, Embryo, Nonmammalian, Female, Germ Cells, Male, Morphogenesis, Ovum, Parthenogenesis, Reproduction}, issn = {1552-5007}, doi = {10.1002/jez.b.3}, author = {Miura, Toru and Braendle, Christian and Shingleton, Alexander and Sisk, Geoffroy and Kambhampati, Srinivas and Stern, David L} } @article {45761, title = {Developmental origin and evolution of bacteriocytes in the aphid-Buchnera symbiosis.}, journal = {PLoS Biol}, volume = {1}, year = {2003}, month = {2003 Oct}, pages = {E21}, abstract = {

Symbiotic relationships between bacteria and insect hosts are common. Although the bacterial endosymbionts have been subjected to intense investigation, little is known of the host cells in which they reside, the bacteriocytes. We have studied the development and evolution of aphid bacteriocytes, the host cells that contain the endosymbiotic bacteria Buchnera aphidicola. We show that bacteriocytes of Acyrthosiphon pisum express several gene products (or their paralogues): Distal-less, Ultrabithorax/Abdominal-A, and Engrailed. Using these markers, we find that a subpopulation of the bacteriocytes is specified prior to the transmission of maternal bacteria to the embryo. In addition, we discovered that a second population of cells is recruited to the bacteriocyte fate later in development. We experimentally demonstrate that bacteriocyte induction and proliferation occur independently of B. aphidicola. Major features of bacteriocyte development, including the two-step recruitment of bacteriocytes, have been conserved in aphids for 80-150 million years. Furthermore, we have investigated two cases of evolutionary loss of bacterial symbionts: in one case, where novel extracellular, eukaryotic symbionts replaced the bacteria, the bacteriocyte is maintained; in another case, where symbionts are absent, the bacteriocytes are initiated but not maintained. The bacteriocyte represents an evolutionarily novel cell fate, which is developmentally determined independently of the bacteria. Three of five transcription factors we examined show novel expression patterns in bacteriocytes, suggesting that bacteriocytes may have evolved to express many additional transcription factors. The evolutionary transition to a symbiosis in which bacteria and an aphid cell form a functional unit, similar to the origin of plastids, has apparently involved extensive molecular adaptations on the part of the host cell.

}, keywords = {Animals, Aphids, Buchnera, Cell Lineage, Drosophila Proteins, Evolution, Molecular, Female, Gene Expression Regulation, Bacterial, Homeodomain Proteins, Male, Microscopy, Fluorescence, Models, Biological, Parthenogenesis, Species Specificity, Symbiosis, Time Factors, Transcription Factors}, issn = {1545-7885}, doi = {10.1371/journal.pbio.0000021}, author = {Braendle, Christian and Miura, Toru and Bickel, Ryan and Shingleton, Alexander W and Kambhampati, Srinivas and Stern, David L} } @article {2322, title = {Dopamine D1 receptors mediate CREB phosphorylation via phosphorylation of the NMDA receptor at Ser897-NR1.}, journal = {Journal of Neurochemistry}, volume = {87}, year = {2003}, month = {2003 Nov}, pages = {922-34}, abstract = {

Addictive drugs such as amphetamine and cocaine stimulate the dopaminergic system, activate dopamine receptors and induce gene expression throughout the striatum. The signal transduction pathway leading from dopamine receptor stimulation at the synapse to gene expression in the nucleus has not been fully elucidated. Here, we present evidence that D1 receptor stimulation leads to phosphorylation of the transcription factor Ca2+ and cyclic AMP response element binding protein (CREB) in the nucleus by means of NMDA receptor-mediated Ca2+ signaling. Stimulation of D1 receptors induces the phosphorylation of Ser897 on the NR1 subunit by protein kinase A (PKA). This phosphorylation event is crucial for D1 receptor-mediated CREB phosphorylation. Dopamine cannot induce CRE-mediated gene expression in neurons transfected with a phosphorylation-deficient NR1 construct. Moreover, stimulation of D1 receptors or increase in cyclic AMP levels leads to an increase in cytosolic Ca2+ in the presence of glutamate, but not in the absence of glutamate, indicating the ability of dopamine and cyclic AMP to facilitate NMDA channel activity. The recruitment of the NMDA receptor signal transduction pathway by D1 receptors may provide a general mechanism for gene regulation that is fundamental for mechanisms of drug addiction and long-term memory.

}, keywords = {Animals, Calcium, Calcium-Calmodulin-Dependent Protein Kinases, Cells, Corpus Striatum, Cultured, Cyclic AMP Response Element-Binding Protein, Cyclic AMP-Dependent Protein Kinases, Dopamine D1, Glutamic Acid, Mutagenesis, N-Methyl-D-Aspartate, Neurons, Phosphorylation, Rats, Receptors, Signal Transduction, Site-Directed, Sprague-Dawley}, issn = {0022-3042}, doi = {10.3389/fnana.2010.00147}, author = {Dudman, Joshua T and Eaton, Molly E and Rajadhyaksha, Anjali and Mac{\'\i}as, Wendy and Taher, Muffadal and Barczak, Amy and Kameyama, Kimihiko and Huganir, Richard and Konradi, Christine} } @article {2321, title = {The hyperpolarization-activated HCN1 channel is important for motor learning and neuronal integration by cerebellar Purkinje cells.}, journal = {Cell}, volume = {115}, year = {2003}, month = {2003 Nov 26}, pages = {551-64}, abstract = {

In contrast to our increasingly detailed understanding of how synaptic plasticity provides a cellular substrate for learning and memory, it is less clear how a neuron{\textquoteright}s voltage-gated ion channels interact with plastic changes in synaptic strength to influence behavior. We find, using generalized and regional knockout mice, that deletion of the HCN1 channel causes profound motor learning and memory deficits in swimming and rotarod tasks. In cerebellar Purkinje cells, which are a key component of the cerebellar circuit for learning of correctly timed movements, HCN1 mediates an inward current that stabilizes the integrative properties of Purkinje cells and ensures that their input-output function is independent of the previous history of their activity. We suggest that this nonsynaptic integrative function of HCN1 is required for accurate decoding of input patterns and thereby enables synaptic plasticity to appropriately influence the performance of motor activity.

}, keywords = {Action Potentials, Afferent Pathways, Animals, Cells, Conditioning (Psychology), Cultured, Cyclic Nucleotide-Gated Cation Channels, Ion Channels, Knockout, Learning, Maze Learning, Mice, Motor Skills, Movement, Nerve Tissue Proteins, Neuronal Plasticity, Potassium Channels, Purkinje Cells, Synapses, Synaptic Transmission}, issn = {0092-8674}, doi = {10.3389/fnana.2010.00147}, author = {Nolan, Matthew F and Malleret, Ga{\"e}l and Lee, Ka Hung and Gibbs, Emma and Dudman, Joshua T and Santoro, Bina and Yin, Deqi and Thompson, Richard F and Siegelbaum, Steven A and Kandel, Eric R and Morozov, Alexei} } @article {2014, title = {In vivo imaging of C. elegans mechanosensory neurons demonstrates a specific role for the MEC-4 channel in the process of gentle touch sensation.}, journal = {Neuron}, volume = {39}, year = {2003}, month = {2003 Sep 11}, pages = {1005-17}, abstract = {

In the nematode C. elegans, genes encoding components of a putative mechanotransducing channel complex have been identified in screens for light-touch-insensitive mutants. A long-standing question, however, is whether identified MEC proteins act directly in touch transduction or contribute indirectly by maintaining basic mechanoreceptor neuron physiology. In this study, we used the genetically encoded calcium indicator cameleon to record cellular responses of mechanosensory neurons to touch stimuli in intact, behaving nematodes. We defined a gentle touch sensory modality that adapts with a time course of approximately 500 ms and primarily senses motion rather than pressure. The DEG/ENaC channel subunit MEC-4 and channel-associated stomatin MEC-2 are specifically required for neural responses to gentle mechanical stimulation but do not affect the basic physiology of touch neurons or their in vivo responses to harsh mechanical stimulation. These results distinguish a specific role for the MEC channel proteins in the process of gentle touch mechanosensation.

}, keywords = {Animals, Caenorhabditis elegans, Caenorhabditis elegans Proteins, Calcium Channels, Cells, Cellular, Cultured, L-Type, Mechanoreceptors, Mechanotransduction, Membrane Potentials, Membrane Proteins, Neurons, Touch}, issn = {0896-6273}, author = {Suzuki, Hiroshi and Kerr, Rex and Bianchi, Laura and Fr{\o}kjaer-Jensen, Christian and Slone, Dan and Xue, Jian and Gerstbrein, Beate and Driscoll, Monica and Schafer, William R} } @article {45954, title = {Insertion of MP20 into lens fibre cell plasma membranes correlates with the formation of an extracellular diffusion barrier.}, journal = {Experimental Eye Research}, volume = {77}, year = {2003}, month = {2003 Nov}, pages = {567-74}, abstract = {

It is known that during lens differentiation a number of fibre cell specific membrane proteins change their expression profiles. In this study we have investigated how the profiles of the two most abundant fibre cell membrane proteins AQP0 (formerly known as Major Intrinsic Protein, MIP) and MP20 change as a function of fibre cell differentiation. While AQP0 was always found associated with fibre cell membranes, MP20 was initially found in the cytoplasm of peripheral fibre cells before becoming inserted into the membranes of deeper fibre cells. To determine at what stage in fibre cell differentiation MP20 becomes inserted into the membrane, sections were double-labelled with an antibody against MP20, and propidium iodide, a marker of cell nuclei. This showed that membrane insertion of MP20 occurs in a discrete transition zone that coincided with the degradation of cell nuclei. To test the significance of the membrane insertion of MP20 to overall lens function, whole lenses were incubated for varying times in a solution containing either Texas Red-dextran or Lucifer yellow as markers of extracellular space. Lenses were fixed and then processed for immunocytochemistry. Analysis of these sections showed that both tracer dyes were excluded from the extracellular space in an area that coincided with insertion of MP20 into the plasma membrane. Our results suggest that the insertion of MP20 into fibre cell membranes coincides with the creation of a barrier that restricts the diffusion of molecules into the lens core via the extracellular space.

}, keywords = {Animals, Aquaporins, Cell Differentiation, Cell Membrane, Cytoplasm, Extracellular Space, Eye Proteins, Lens, Crystalline, Membrane Glycoproteins, Rats, Sheep}, issn = {0014-4835}, author = {Grey, Angus C and Jacobs, Marc D and Gonen, Tamir and Kistler, Joerg and Donaldson, Paul J} } @article {2681, title = {Involvement of the ubiquitin-proteasome system in the early stages of wallerian degeneration.}, journal = {Neuron}, volume = {39}, year = {2003}, month = {2003 Jul 17}, pages = {217-25}, abstract = {

Local axon degeneration is a common pathological feature of many neurodegenerative diseases and peripheral neuropathies. While it is believed to operate with an apoptosis-independent molecular program, the underlying molecular mechanisms are largely unknown. In this study, we used the degeneration of transected axons, termed "Wallerian degeneration," as a model to examine the possible involvement of the ubiquitin proteasome system (UPS). Inhibiting UPS activity by both pharmacological and genetic means profoundly delays axon degeneration both in vitro and in vivo. In addition, we found that the fragmentation of microtubules is the earliest detectable change in axons undergoing Wallerian degeneration, which among other degenerative events, can be delayed by proteasome inhibitors. Interestingly, similar to transected axons, degeneration of axons from nerve growth factor (NGF)-deprived sympathetic neurons could also be suppressed by proteasome inhibitors. Our findings suggest a possibility that inhibiting UPS activity may serve to retard axon degeneration in pathological conditions.

}, keywords = {Amino Acids, Animal, Animals, Axons, Benzimidazoles, Blotting, Calpain, Cells, Chelating Agents, Cultured, Cysteine Endopeptidases, Cysteine Proteinase Inhibitors, Cytoskeleton, Disease Models, Drug Interactions, Egtazic Acid, Endopeptidases, Ganglia, Immunohistochemistry, Leupeptins, Microtubules, Multienzyme Complexes, Nerve Growth Factor, Newborn, Optic Nerve, Optic Nerve Injuries, Peptide Fragments, Proteasome Endopeptidase Complex, Rats, Sympathetic, Time Factors, Tubulin, Ubiquitin, Wallerian Degeneration, Western}, issn = {0896-6273}, author = {Zhai, Qiwei and Wang, Jing and Kim, Anna and Liu, Qing and Watts, Ryan and Hoopfer, Eric and Mitchison, Timothy and Luo, Liqun and He, Zhigang} } @article {4481, title = {Microwave oven synthesis of esters promoted by imidazole.}, journal = {Tetrahedron Letters}, volume = {44}, year = {2003}, month = {2003 Feb 24}, pages = {1831-3}, abstract = {

Using imidazole as promotion agent, primary, secondary and phenolic alcohol compounds were esterified with aliphatic and aromatic carboxylic acid anhydrides. Heating a ternary mixture of alcohol, anhydride and imidazole in an unmodified microwave oven produced esters in low to high yields, depending on the steric bulk of the alcohol.

}, url = {http://www.sciencedirect.com/science/article/pii/S0040403903001047}, author = {Hirose, Takuji and Kopek, Benjamin G and Wang, Zhao-Hui and Yusa, Ritsuko and Baldwin, Bruce W.} } @article {2138, title = {Normalization of Ca2+ signals by small oblique dendrites of CA1 pyramidal neurons.}, journal = {The Journal of Neuroscience: The Official Journal of the Society for Neuroscience}, volume = {23}, year = {2003}, month = {2003 Apr 15}, pages = {3243-50}, abstract = {

Oblique dendrites of CA1 pyramidal neurons predominate in stratum radiatum and receive approximately 80\% of the synaptic input from Schaffer collaterals. Despite this fact, most of our understanding of dendritic signal processing in these neurons comes from studies of the main apical dendrite. Using a combination of Ca2+ imaging and whole-cell recording techniques in rat hippocampal slices, we found that the properties of the oblique dendrites differ markedly from those of the main dendrites. These different properties tend to equalize the Ca2+ rise from single action potentials as they backpropagate into the oblique dendrites from the main trunk. Evidence suggests that this normalization of Ca2+ signals results from a higher density of a transient, A-type K+ current [I(K(A))] in the oblique versus the main dendrites. The higher density of I(K(A)) may have important implications for our understanding of synaptic integration and plasticity in these structures.

}, keywords = {4-Aminopyridine, Action Potentials, Animals, Calcium, Calcium Channel Blockers, Calcium Signaling, Dendrites, Electric Stimulation, Fluorescent Dyes, Hippocampus, Patch-Clamp Techniques, Potassium Channel Blockers, Potassium Channels, Pyramidal Cells, Rats, Sodium Channels, Sprague-Dawley, Tetrodotoxin}, issn = {1529-2401}, doi = {10.1002/cbic.201000254}, author = {Frick, Andreas and Magee, Jeffrey and Koester, Helmut J and Migliore, Michele and Johnston, Daniel} } @article {3087, title = {Operant behavior can be triggered by the position of the rat relative to objects rotating on an inaccessible platform.}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, volume = {100}, year = {2003}, month = {2003 Feb 18}, pages = {2094-9}, abstract = {

The present study describes a task testing the ability of rats to trigger operant behavior by their relative spatial position to inaccessible rotating objects. Rats were placed in a Skinner box with a transparent front wall through which they could observe one or two adjacent objects fixed on a slowly rotating arena (d = 1 m) surrounded by an immobile black cylinder. The direction of arena rotation was alternated at a sequence of different time intervals. Rats were reinforced for the first bar-press that was emitted when a radius separating the two adjacent objects or dividing a single object into two halves (pointing radius) entered a 60 degrees sector of its circular trajectory defined with respect to the stationary Skinner box (reward sector). Well trained rats emitted 62.1 +/- 3.6\% of responses in a 60 degrees sector preceding the reward sector and in the first 30 degrees of the reward sector. Response rate increased only when the pointing radius was approaching the reward sector, regardless of the time elapsed from the last reward. In the extinction session, when no reward was delivered, rats responded during the whole passage of the pointing radius through the former reward sector and spontaneously decreased responding after the pointing radius left this area. This finding suggests that rats perceived the reward sector as a continuous single region. The same results were obtained when the Skinner box with the rat was orbiting around the immobile scene. It is concluded that rats can recognize and anticipate their position relative to movable objects.

}, keywords = {Animal, Animals, Behavior, Conditioning, Long-Evans, Male, Operant, Rats, Reward}, issn = {0027-8424}, doi = {10.1073/pnas.0438002100}, author = {Pastalkova, E and Kelemen, E and Bures, J} } @article {2323, title = {Striatal proenkephalin gene induction: coordinated regulation by cyclic AMP and calcium pathways.}, journal = {Brain Research. Molecular Brain Research}, volume = {115}, year = {2003}, month = {2003 Jul 23}, pages = {157-61}, abstract = {

Enkephalin modulates striatal function, thereby affecting motor performance and addictive behaviors. The proenkephalin gene is also used as a model to study cyclic AMP-mediated gene expression in striatal neurons. The second messenger pathway leading to proenkephalin expression demonstrates how cyclic AMP pathways are synchronized with depolarization. We show that cyclic AMP-mediated regulation of the proenkephalin gene is dependent on the activity of L-type Ca2+ channels. Inhibition of L-type Ca2+ channels blocks forskolin-mediated induction of proenkephalin. The Ca2+-activated kinase, Ca2+/calmodulin kinase, as well as the cyclic AMP-activated kinase, protein kinase A (PKA), are both necessary for the induction of the proenkephalin promoter. Similarly, both kinases are needed for the L-type Ca2+ channel-mediated induction of proenkephalin. This synchronization of second messenger pathways provides a coincidence mechanism that gates proenkephalin synthesis in striatal neurons, ensuring that levels are increased only in the presence of activated PKA and depolarization.

}, keywords = {1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine, Animals, Antihypertensive Agents, Calcium, Calcium Channel Agonists, Calcium Channel Blockers, Carcinogens, Cells, Corpus Striatum, Cultured, Cyclic AMP, Developmental, Drug Interactions, Embryo, Enkephalins, Enzyme Inhibitors, Forskolin, Gene Expression Regulation, Humans, Isoquinolines, Mammalian, Neurons, Nifedipine, Protein Precursors, Pyrroles, Rats, Sprague-Dawley, Sulfonamides, Tetradecanoylphorbol Acetate, Transcriptional Activation, Transfection}, issn = {0169-328X}, doi = {10.3389/fnana.2010.00147}, author = {Konradi, Christine and Mac{\'\i}as, Wendy and Dudman, Joshua T and Carlson, Richard R} } @article {46635, title = {Tre1, a G protein-coupled receptor, directs transepithelial migration of Drosophila germ cells.}, journal = {PLoS Biology}, volume = {1}, year = {2003}, month = {2003 Dec}, pages = {E80}, abstract = {

In most organisms, germ cells are formed distant from the somatic part of the gonad and thus have to migrate along and through a variety of tissues to reach the gonad. Transepithelial migration through the posterior midgut (PMG) is the first active step during Drosophila germ cell migration. Here we report the identification of a novel G protein-coupled receptor (GPCR), Tre1, that is essential for this migration step. Maternal tre1 RNA is localized to germ cells, and tre1 is required cell autonomously in germ cells. In tre1 mutant embryos, most germ cells do not exit the PMG. The few germ cells that do leave the midgut early migrate normally to the gonad, suggesting that this gene is specifically required for transepithelial migration and that mutant germ cells are still able to recognize other guidance cues. Additionally, inhibiting small Rho GTPases in germ cells affects transepithelial migration, suggesting that Tre1 signals through Rho1. We propose that Tre1 acts in a manner similar to chemokine receptors required during transepithelial migration of leukocytes, implying an evolutionarily conserved mechanism of transepithelial migration. Recently, the chemokine receptor CXCR4 was shown to direct migration in vertebrate germ cells. Thus, germ cells may more generally use GPCR signaling to navigate the embryo toward their target.

}, keywords = {Animals, Cell Movement, Cell Transplantation, Cloning, Molecular, Crosses, Genetic, Drosophila melanogaster, Drosophila Proteins, Embryo, Nonmammalian, Epithelium, Evolution, Molecular, Female, Gene Expression Regulation, Germ Cells, Humans, Immunohistochemistry, In Situ Hybridization, Inflammation, Leukocytes, Male, Models, Biological, Molecular Sequence Data, Mutation, Phenotype, Phylogeny, Receptors, Chemokine, Receptors, G-Protein-Coupled, rho GTP-Binding Proteins, RNA, Signal Transduction}, issn = {1545-7885}, doi = {10.1371/journal.pbio.0000080}, author = {Kunwar, Prabhat S and Starz-Gaiano, Michelle and Bainton, Roland J and Heberlein, Ulrike and Lehmann, Ruth} } @article {2143, title = {Conditional loss of Nkx3.1 in adult mice induces prostatic intraepithelial neoplasia.}, journal = {Molecular and Cellular Biology}, volume = {22}, year = {2002}, month = {2002 Mar}, pages = {1495-503}, abstract = {

The homeodomain-containing transcription factor NKX3.1 is a putative prostate tumor suppressor that is expressed in a largely prostate-specific and androgen-regulated manner. Loss of NKX3.1 protein expression is common in human prostate carcinomas and prostatic intraepithelial neoplasia (PIN) lesions and correlates with tumor progression. Disruption of the murine Nkx3.1 gene results in defects in prostate branching morphogenesis, secretions, and growth. To more closely mimic the pattern of NKX3.1 loss that occurs in human prostate tumors, we have used Cre- and loxP-mediated recombination to delete the Nkx3.1 gene in the prostates of adult transgenic mice. Conditional deletion of one or both alleles of Nkx3.1 leads to the development of preinvasive lesions that resemble PIN. The pattern of expression of several biomarkers (Ki-67, E-cadherin, and high-molecular-weight cytokeratins) in these PIN lesions resembled that observed in human cases of PIN. Furthermore, PIN foci in mice with conditional deletion of a single Nkx3.1 allele lose expression of the wild-type allele. Our results support the role of NKX3.1 as a prostate tumor suppressor and indicate a role for this gene in tumor initiation.

}, keywords = {Animals, Gene Deletion, Genes, Genetic Predisposition to Disease, Homeodomain Proteins, Homozygote, Hyperplasia, Integrases, Male, Mice, Prostate, Prostatic Intraepithelial Neoplasia, Prostatic Neoplasms, Transcription Factors, Transgenic, Tumor Suppressor, Viral Proteins}, issn = {0270-7306}, doi = {10.1002/cbic.201000254}, author = {Abdulkadir, Sarki A and Magee, Jeffrey A and Peters, Thomas J and Kaleem, Zahid and Naughton, Cathy K and Humphrey, Peter A and Milbrandt, Jeffrey} } @article {2178, title = {Detection of large interaural delays and its implication for models of binaural interaction.}, journal = {Journal of the Association for Research in Otolaryngology: JARO}, volume = {3}, year = {2002}, month = {2002 Mar}, pages = {80-8}, abstract = {

The interaural time difference (ITD) is a major cue to sound localization along the horizontal plane. The maximum natural ITD occurs when a sound source is positioned opposite to one ear. We examined the ability of owls and humans to detect large ITDs in sounds presented through headphones. Stimuli consisted of either broad or narrow bands of Gaussian noise, 100 ms in duration. Using headphones allowed presentation of ITDs that are greater than the maximum natural ITD. Owls were able to discriminate a sound leading to the left ear from one leading to the right ear, for ITDs that are 5 times the maximum natural delay. Neural recordings from optic-tectum neurons, however, show that best ITDs are usually well within the natural range and are never as large as ITDs that are behaviorally discriminable. A model of binaural crosscorrelation with short delay lines is shown to explain behavioral detection of large ITDs. The model uses curved trajectories of a cross-correlation pattern as the basis for detection. These trajectories represent side peaks of neural ITD-tuning curves and successfully predict localization reversals by both owls and human subjects.

}, keywords = {Animals, Biological, Ear, Hearing, Humans, Models, Neurons, Noise, Strigiformes, Superior Colliculi, Time Factors}, issn = {1525-3961}, author = {Saberi, Kourosh and Takahashi, Yoshifumi and Egnor, Roian and Farahbod, Haleh and Mazer, James and Konishi, Masakazu} } @article {2189, title = {Dissecting glucose signalling with diversity-oriented synthesis and small-molecule microarrays.}, journal = {Nature}, volume = {416}, year = {2002}, month = {2002 Apr 11}, pages = {653-7}, abstract = {

Small molecules that alter protein function provide a means to modulate biological networks with temporal resolution. Here we demonstrate a potentially general and scalable method of identifying such molecules by application to a particular protein, Ure2p, which represses the transcription factors Gln3p and Nil1p. By probing a high-density microarray of small molecules generated by diversity-oriented synthesis with fluorescently labelled Ure2p, we performed 3,780 protein-binding assays in parallel and identified several compounds that bind Ure2p. One compound, which we call uretupamine, specifically activates a glucose-sensitive transcriptional pathway downstream of Ure2p. Whole-genome transcription profiling and chemical epistasis demonstrate the remarkable Ure2p specificity of uretupamine and its ability to modulate the glucose-sensitive subset of genes downstream of Ure2p. These results demonstrate that diversity-oriented synthesis and small-molecule microarrays can be used to identify small molecules that bind to a protein of interest, and that these small molecules can regulate specific functions of the protein.

}, keywords = {Biological, Dioxanes, Dose-Response Relationship, Drug, Fungal, Gene Expression Profiling, Gene Expression Regulation, Genetic, Glucose, Glutathione Peroxidase, Ligands, Messenger, Models, Oligonucleotide Array Sequence Analysis, Oxazoles, Prions, Protein Binding, Repressor Proteins, RNA, Saccharomyces cerevisiae, Saccharomyces cerevisiae Proteins, Signal Transduction, Structure-Activity Relationship, Substrate Specificity, Transcription}, issn = {0028-0836}, doi = {10.1038/416653a}, author = {Kuruvilla, Finny G and Shamji, Alykhan F and Sternson, Scott M and Hergenrother, Paul J and Schreiber, Stuart L} } @article {5493, title = {Functional dissection of neuroanatomical loci regulating ethanol sensitivity in Drosophila.}, journal = {The Journal of Neuroscience: The Official Journal of the Society for Neuroscience}, volume = {22}, year = {2002}, month = {2002 Nov 1}, pages = {9490-501}, abstract = {

Ethanol has complex but similar effects on behavior in mammals and the fruit fly Drosophila melanogaster. In addition, genetic and pharmacological approaches have implicated the cAMP pathway in the regulation of ethanol-induced behaviors in both flies and rodents. Here we examine the neuroanatomical loci that modulate ethanol sensitivity in Drosophila by targeting the expression of an inhibitor of cAMP-dependent protein kinase (PKA) to specific regions in the fly{\textquoteright}s brain. Expression of the inhibitor in most brain regions or in muscle has no effect on behavior. In contrast, inhibition of PKA in a relatively small number of cells, possibly neurosecretory cells, in the fly{\textquoteright}s brain is sufficient to decrease sensitivity to the incoordinating effects of ethanol. Additional brain areas are, however, also involved. The mushroom bodies, brain structures where cAMP signaling is required for olfactory classical conditioning, are dispensable for the regulation of ethanol sensitivity. Finally, different behavioral effects of ethanol, motor incoordination and sedation, appear to be regulated by PKA function in distinct brain regions. We conclude that the regulation of ethanol-induced behaviors by PKA involves complex interactions among groups of cells that mediate either increased or reduced sensitivity to the acute intoxicating effects of ethanol.

}, keywords = {Administration, Animal, Animals, Behavior, Catalytic Domain, Cyclic AMP-Dependent Protein Kinases, DNA-Binding Proteins, Drosophila melanogaster, Drug Resistance, Ethanol, Gene Expression, Genes, Genetically Modified, Hydroxyurea, Inhalation, Larva, Locomotion, Male, Motor Activity, Mushroom Bodies, Nervous System, Organ Specificity, Protein Subunits, Reporter, Saccharomyces cerevisiae Proteins, Transcription Factors, Transgenes}, issn = {1529-2401}, author = {Rodan, Aylin R and Kiger, John A and Heberlein, Ulrike} } @article {1265, title = {Network motifs: simple building blocks of complex networks.}, journal = {Science}, volume = {298}, year = {2002}, month = {2002 Oct 25}, pages = {824-7}, abstract = {

Complex networks are studied across many fields of science. To uncover their structural design principles, we defined {\textquotedblleft}network motifs,{\textquotedblright} patterns of interconnections occurring in complex networks at numbers that are significantly higher than those in randomized networks. We found such motifs in networks from biochemistry, neurobiology, ecology, and engineering. The motifs shared by ecological food webs were distinct from the motifs shared by the genetic networks of Escherichia coli and Saccharomyces cerevisiae or from those found in the World Wide Web. Similar motifs were found in networks that perform information processing, even though they describe elements as different as biomolecules within a cell and synaptic connections between neurons in Caenorhabditis elegans. Motifs may thus define universal classes of networks. This approach may uncover the basic building blocks of most networks.

}, issn = {1095-9203}, author = {Milo, R. and Shen-Orr, S. and Itzkovitz, S. and Kashtan, N. and Chklovskii, D.B. and Alon, U.} } @article {2295, title = {A new NOS2 promoter polymorphism associated with increased nitric oxide production and protection from severe malaria in Tanzanian and Kenyan children.}, journal = {Lancet}, volume = {360}, year = {2002}, month = {2002 Nov 9}, pages = {1468-75}, abstract = {

Nitric oxide (NO) is a mediator of immunity to malaria, and genetic polymorphisms in the promoter of the inducible NO synthase gene (NOS2) could modulate production of NO. We postulated that NOS2 promoter polymorphisms would affect resistance to severe malaria.

}, keywords = {Anemia, Cerebral, Child, DNA, Falciparum, Female, Genetic, Humans, Immunity, Infant, Innate, Kenya, Malaria, Male, Nitric Oxide, Nitric Oxide Synthase, Nitric Oxide Synthase Type II, Parasitemia, Polymorphism, Preschool, Sequence Analysis, Single-Stranded Conformational, Tanzania}, issn = {0140-6736}, doi = {10.1016/S0140-6736(02)11474-7}, author = {Hobbs, Maurine R and Udhayakumar, Venkatachalam and Levesque, Marc C and Booth, Jennifer and Roberts, Jacquelin M and Tkachuk, Ariana N and Pole, Ann and Coon, Hilary and Kariuki, Simon and Nahlen, Bernard L and Mwaikambo, Esther D and Lal, Altaf L and Granger, Donald L and Anstey, Nicholas M and Weinberg, J Brice} } @article {45890, title = {Dichotomy of action-potential backpropagation in CA1 pyramidal neuron dendrites.}, journal = {J Neurophysiol}, volume = {86}, year = {2001}, month = {2001 Dec}, pages = {2998-3010}, abstract = {

In hippocampal CA1 pyramidal neurons, action potentials are typically initiated in the axon and backpropagate into the dendrites, shaping the integration of synaptic activity and influencing the induction of synaptic plasticity. Despite previous reports describing action-potential propagation in the proximal apical dendrites, the extent to which action potentials invade the distal dendrites of CA1 pyramidal neurons remains controversial. Using paired somatic and dendritic whole cell recordings, we find that in the dendrites proximal to 280 microm from the soma, single backpropagating action potentials exhibit \<50\% attenuation from their amplitude in the soma. However, in dendritic recordings distal to 300 microm from the soma, action potentials in most cells backpropagated either strongly (26-42\% attenuation; n = 9/20) or weakly (71-87\% attenuation; n = 10/20) with only one cell exhibiting an intermediate value (45\% attenuation). In experiments combining dual somatic and dendritic whole cell recordings with calcium imaging, the amount of calcium influx triggered by backpropagating action potentials was correlated with the extent of action-potential invasion of the distal dendrites. Quantitative morphometric analyses revealed that the dichotomy in action-potential backpropagation occurred in the presence of only subtle differences in either the diameter of the primary apical dendrite or branching pattern. In addition, action-potential backpropagation was not dependent on a number of electrophysiological parameters (input resistance, resting potential, voltage sensitivity of dendritic spike amplitude). There was, however, a striking correlation of the shape of the action potential at the soma with its amplitude in the dendrite; larger, faster-rising, and narrower somatic action potentials exhibited more attenuation in the distal dendrites (300-410 microm from the soma). Simple compartmental models of CA1 pyramidal neurons revealed that a dichotomy in action-potential backpropagation could be generated in response to subtle manipulations of the distribution of either sodium or potassium channels in the dendrites. Backpropagation efficacy could also be influenced by local alterations in dendritic side branches, but these effects were highly sensitive to model parameters. Based on these findings, we hypothesize that the observed dichotomy in dendritic action-potential amplitude is conferred primarily by differences in the distribution, density, or modulatory state of voltage-gated channels along the somatodendritic axis.

}, keywords = {Action Potentials, Animals, Calcium, Dendrites, Diagnostic Imaging, Electrophysiology, Fluorescent Dyes, Fura-2, Hippocampus, In Vitro Techniques, Male, Models, Neurological, Patch-Clamp Techniques, Potassium Channels, Pyramidal Cells, Rats, Rats, Wistar, Sodium Channels}, issn = {0022-3077}, author = {Golding, N L and Kath, W L and Spruston, N} } @article {45953, title = {MP20, the second most abundant lens membrane protein and member of the tetraspanin superfamily, joins the list of ligands of galectin-3.}, journal = {BMC Cell Biology}, volume = {2}, year = {2001}, month = {2001 - Aug}, pages = {17}, abstract = {

BACKGROUND: Although MP20 is the second most highly expressed membrane protein in the lens its function remains an enigma. Putative functions for MP20 have recently been inferred from its assignment to the tetraspanin superfamily of integral membrane proteins. Members of this family have been shown to be involved in cellular proliferation, differentiation, migration, and adhesion. In this study, we show that MP20 associates with galectin-3, a known adhesion modulator.

RESULTS: MP20 and galectin-3 co-localized in selected areas of the lens fiber cell plasma membrane. Individually, these proteins purified with apparent molecular masses of 60 kDa and 22 kDa, respectively. A 104 kDa complex was formed in vitro upon mixing the purified proteins. A 102 kDa complex of MP20 and galectin-3 could also be isolated from detergent-solubilized native fiber cell membranes. Binding between MP20 and galectin-3 was disrupted by lactose suggesting the lectin site was involved in the interaction.

CONCLUSIONS: MP20 adds to a growing list of ligands of galectin-3 and appears to be the first representative of the tetraspanin superfamily identified to possess this specificity.

}, keywords = {Animals, Binding Sites, Cell Membrane, Eye Proteins, Galectin 3, Lectins, Lens, Crystalline, Ligands, Macromolecular Substances, Membrane Proteins, Rats}, issn = {1471-2121}, doi = {10.1186/1471-2121-2-17}, author = {Gonen, T and Grey, A C and Jacobs, M D and Donaldson, P J and Kistler, J} } @article {1267, title = {Orientation preference maps in mammalian visual cortex: A wire length minimization approach.}, journal = {Neuron}, volume = {29}, year = {2001}, month = {2001 Feb}, pages = {519-27}, abstract = {

In the visual cortex of many mammals, orientation preference changes smoothly along the cortical surface, with the exception of singularities such as pinwheels and fractures. The reason for the existence of these singularities has remained elusive, suggesting that they are developmental artifacts. We show that singularities reduce the length of intracortical neuronal connections for some connection rules. Therefore, pinwheels and fractures could be evolutionary adaptations keeping cortical volume to a minimum. Wire length minimization approach suggests that interspecies differences in orientation preference maps reflect differences in intracortical neuronal circuits, thus leading to experimentally testable predictions. We discuss application of our model to direction preference maps.

}, issn = {0896-6273}, author = {Koulakov, Alexei A and Chklovskii, Dmitri B} } @article {2015, title = {Rapid atomic density methods for molecular shape characterization.}, journal = {Journal of Molecular Graphics \& Modelling}, volume = {19}, year = {2001}, month = {2001 Jun}, pages = {325-30, 388-90}, abstract = {

Two methods for rapid characterization of molecular shape are presented. Both techniques are based on the density of atoms near the molecular surface. The Fast Atomic Density Evaluation (FADE) algorithm uses fast Fourier transforms to quickly estimate densities. The Pairwise Atomic Density Reverse Engineering (PADRE) method derives modified density measures from the relationship between atomic density and total potentials. While many shape-characterization techniques define shape relative to a surface, the descriptors returned by FADE and PADRE can measure local geometry from points within the three-dimensional space surrounding a molecule. The methods can be used to find crevices and protrusions near the surface of a molecule and to test shape complementarity at the interface between docking molecules.

}, keywords = {Algorithms, Binding Sites, Computer Simulation, Fourier Analysis, Models, Molecular, Protein Binding, Protein Conformation, Proteins}, issn = {1093-3263}, author = {Mitchell, J C and Kerr, R and Ten Eyck, L F} } @article {2707, title = {The sequence of the human genome.}, journal = {Science}, volume = {291}, year = {2001}, month = {2001 Feb 16}, pages = {1304-51}, abstract = {

A 2.91-billion base pair (bp) consensus sequence of the euchromatic portion of the human genome was generated by the whole-genome shotgun sequencing method. The 14.8-billion bp DNA sequence was generated over 9 months from 27,271,853 high-quality sequence reads (5.11-fold coverage of the genome) from both ends of plasmid clones made from the DNA of five individuals. Two assembly strategies-a whole-genome assembly and a regional chromosome assembly-were used, each combining sequence data from Celera and the publicly funded genome effort. The public data were shredded into 550-bp segments to create a 2.9-fold coverage of those genome regions that had been sequenced, without including biases inherent in the cloning and assembly procedure used by the publicly funded group. This brought the effective coverage in the assemblies to eightfold, reducing the number and size of gaps in the final assembly over what would be obtained with 5.11-fold coverage. The two assembly strategies yielded very similar results that largely agree with independent mapping data. The assemblies effectively cover the euchromatic regions of the human chromosomes. More than 90\% of the genome is in scaffold assemblies of 100,000 bp or more, and 25\% of the genome is in scaffolds of 10 million bp or larger. Analysis of the genome sequence revealed 26,588 protein-encoding transcripts for which there was strong corroborating evidence and an additional approximately 12,000 computationally derived genes with mouse matches or other weak supporting evidence. Although gene-dense clusters are obvious, almost half the genes are dispersed in low G+C sequence separated by large tracts of apparently noncoding sequence. Only 1.1\% of the genome is spanned by exons, whereas 24\% is in introns, with 75\% of the genome being intergenic DNA. Duplications of segmental blocks, ranging in size up to chromosomal lengths, are abundant throughout the genome and reveal a complex evolutionary history. Comparative genomic analysis indicates vertebrate expansions of genes associated with neuronal function, with tissue-specific developmental regulation, and with the hemostasis and immune systems. DNA sequence comparisons between the consensus sequence and publicly funded genome data provided locations of 2.1 million single-nucleotide polymorphisms (SNPs). A random pair of human haploid genomes differed at a rate of 1 bp per 1250 on average, but there was marked heterogeneity in the level of polymorphism across the genome. Less than 1\% of all SNPs resulted in variation in proteins, but the task of determining which SNPs have functional consequences remains an open challenge.

}, keywords = {Algorithms, Animals, Artificial, Bacterial, Chromosome Banding, Chromosome Mapping, Chromosomes, Computational Biology, Consensus Sequence, CpG Islands, Databases, DNA, Evolution, Exons, Factual, Female, Gene Duplication, Genes, Genetic Variation, Genome, Human, Human Genome Project, Humans, Intergenic, Introns, Male, Molecular, Nucleic Acid, Phenotype, Physical Chromosome Mapping, Polymorphism, Proteins, Pseudogenes, Repetitive Sequences, Retroelements, Sequence Analysis, Single Nucleotide, Species Specificity}, issn = {0036-8075}, doi = {10.1126/science.1058040}, author = {Venter, J C and Adams, M D and Myers, E W and Li, P W and Mural, R J and Sutton, G G and Smith, H O and Yandell, M and Evans, C A and Holt, R A and Gocayne, J D and Amanatides, P and Ballew, R M and Huson, D H and Wortman, J R and Zhang, Q and Kodira, C D and Zheng, X H and Chen, L and Skupski, M and Subramanian, G and Thomas, P D and Zhang, J and Gabor Miklos, G L and Nelson, C and Broder, S and Clark, A G and Nadeau, J and McKusick, V A and Zinder, N and Levine, A J and Roberts, R J and Simon, M and Slayman, C and Hunkapiller, M and Bolanos, R and Delcher, A and Dew, I and Fasulo, D and Flanigan, M and Florea, L and Halpern, A and Hannenhalli, S and Kravitz, S and Levy, S and Mobarry, C and Reinert, K and Remington, K and Abu-Threideh, J and Beasley, E and Biddick, K and Bonazzi, V and Brandon, R and Cargill, M and Chandramouliswaran, I and Charlab, R and Chaturvedi, K and Deng, Z and Di Francesco, V and Dunn, P and Eilbeck, K and Evangelista, C and Gabrielian, A E and Gan, W and Ge, W and Gong, F and Gu, Z and Guan, P and Heiman, T J and Higgins, M E and Ji, R R and Ke, Z and Ketchum, K A and Lai, Z and Lei, Y and Li, Z and Li, J and Liang, Y and Lin, X and Lu, F and Merkulov, G V and Milshina, N and Moore, H M and Naik, A K and Narayan, V A and Neelam, B and Nusskern, D and Rusch, D B and Salzberg, S and Shao, W and Shue, B and Sun, J and Wang, Z and Wang, A and Wang, X and Wang, J and Wei, M and Wides, R and Xiao, C and Yan, C and Yao, A and Ye, J and Zhan, M and Zhang, W and Zhang, H and Zhao, Q and Zheng, L and Zhong, F and Zhong, W and Zhu, S and Zhao, S and Gilbert, D and Baumhueter, S and Spier, G and Carter, C and Cravchik, A and Woodage, T and Ali, F and An, H and Awe, A and Baldwin, D and Baden, H and Barnstead, M and Barrow, I and Beeson, K and Busam, D and Carver, A and Center, A and Cheng, M L and Curry, L and Danaher, S and Davenport, L and Desilets, R and Dietz, S and Dodson, K and Doup, L and Ferriera, S and Garg, N and Gluecksmann, A and Hart, B and Haynes, J and Haynes, C and Heiner, C and Hladun, S and Hostin, D and Houck, J and Howland, T and Ibegwam, C and Johnson, J and Kalush, F and Kline, L and Koduru, S and Love, A and Mann, F and May, D and McCawley, S and McIntosh, T and McMullen, I and Moy, M and Moy, L and Murphy, B and Nelson, K and Pfannkoch, C and Pratts, E and Puri, V and Qureshi, H and Reardon, M and Rodriguez, R and Rogers, Y H and Romblad, D and Ruhfel, B and Scott, R and Sitter, C and Smallwood, M and Stewart, E and Strong, R and Suh, E and Thomas, R and Tint, N N and Tse, S and Vech, C and Wang, G and Wetter, J and Williams, S and Williams, M and Windsor, S and Winn-Deen, E and Wolfe, K and Zaveri, J and Zaveri, K and Abril, J F and Guig{\'o}, R and Campbell, M J and Sjolander, K V and Karlak, B and Kejariwal, A and Mi, H and Lazareva, B and Hatton, T and Narechania, A and Diemer, K and Muruganujan, A and Guo, N and Sato, S and Bafna, V and Istrail, S and Lippert, R and Schwartz, R and Walenz, B and Yooseph, S and Allen, D and Basu, A and Baxendale, J and Blick, L and Caminha, M and Carnes-Stine, J and Caulk, P and Chiang, Y H and Coyne, M and Dahlke, C and Mays, A and Dombroski, M and Donnelly, M and Ely, D and Esparham, S and Fosler, C and Gire, H and Glanowski, S and Glasser, K and Glodek, A and Gorokhov, M and Graham, K and Gropman, B and Harris, M and Heil, J and Henderson, S and Hoover, J and Jennings, D and Jordan, C and Jordan, J and Kasha, J and Kagan, L and Kraft, C and Levitsky, A and Lewis, M and Liu, X and Lopez, J and Ma, D and Majoros, W and McDaniel, J and Murphy, S and Newman, M and Nguyen, T and Nguyen, N and Nodell, M and Pan, S and Peck, J and Peterson, M and Rowe, W and Sanders, R and Scott, J and Simpson, M and Smith, T and Sprague, A and Stockwell, T and Turner, R and Venter, E and Wang, M and Wen, M and Wu, D and Wu, M and Xia, A and Zandieh, A and Zhu, X} } @article {1518, title = {Switching repulsion to attraction: changing responses to slit during transition in mesoderm migration.}, journal = {Science}, volume = {292}, year = {2001}, month = {2001 Apr 27}, pages = {737-40}, abstract = {

Slit is secreted by cells at the midline of the central nervous system, where it binds to Roundabout (Robo) receptors and functions as a potent repellent. We found that migrating mesodermal cells in vivo respond to Slit as both an attractant and a repellent and that Robo receptors are required for both functions. Mesoderm cells expressing Robo receptors initially migrate away from Slit at the midline. A few hours after migration, these same cells change their behavior and require Robo to extend toward Slit-expressing muscle attachment sites. Thus, Slit functions as a chemoattractant to provide specificity for muscle patterning.

}, keywords = {Animals, Body Patterning, Cell Fusion, Cell Movement, Confocal, Drosophila, Drosophila Proteins, Embryo, Epidermis, Immunologic, Mesoderm, Microscopy, Muscles, Mutation, Nerve Tissue Proteins, Nonmammalian, Receptors, Signal Transduction, Stem Cells}, issn = {0036-8075}, doi = {10.1126/science.1058766}, author = {Kramer, S G and Kidd, T and Simpson, J H and Goodman, C S} } @article {958, title = {Comparative genomics of the eukaryotes.}, journal = {Science}, volume = {287}, year = {2000}, month = {2000 Mar 24}, pages = {2204-15}, abstract = {

A comparative analysis of the genomes of Drosophila melanogaster, Caenorhabditis elegans, and Saccharomyces cerevisiae-and the proteins they are predicted to encode-was undertaken in the context of cellular, developmental, and evolutionary processes. The nonredundant protein sets of flies and worms are similar in size and are only twice that of yeast, but different gene families are expanded in each genome, and the multidomain proteins and signaling pathways of the fly and worm are far more complex than those of yeast. The fly has orthologs to 177 of the 289 human disease genes examined and provides the foundation for rapid analysis of some of the basic processes involved in human disease.

}, keywords = {Animals, Apoptosis, Biological Evolution, Caenorhabditis elegans, Cell Adhesion, Cell Cycle, Drosophila melanogaster, Duplicate, Fungal Proteins, Genes, Genetic Diseases, Genetics, Genome, Helminth Proteins, Humans, Immunity, Inborn, Insect Proteins, Medical, Multigene Family, Neoplasms, Protein Structure, Proteome, Saccharomyces cerevisiae, Signal Transduction, Tertiary}, issn = {0036-8075}, doi = {10.1186/gb-2007-8-7-r145}, author = {Rubin, G M and Yandell, M D and Wortman, J R and Gabor Miklos, G L and Nelson, C R and Hariharan, I K and Fortini, M E and Li, P W and Apweiler, R and Fleischmann, W and Cherry, J M and Henikoff, S and Skupski, M P and Misra, S and Ashburner, M and Birney, E and Boguski, M S and Brody, T and Brokstein, P and Celniker, S E and Chervitz, S A and Coates, D and Cravchik, A and Gabrielian, A and Galle, R F and Gelbart, W M and George, R A and Goldstein, L S and Gong, F and Guan, P and Harris, N L and Hay, B A and Hoskins, R A and Li, J and Li, Z and Hynes, R O and Jones, S J and Kuehl, P M and Lemaitre, B and Littleton, J T and Morrison, D K and Mungall, C and O{\textquoteright}Farrell, P H and Pickeral, O K and Shue, C and Vosshall, L B and Zhang, J and Zhao, Q and Zheng, X H and Lewis, S} } @article {45952, title = {Galectin-3 is associated with the plasma membrane of lens fiber cells.}, journal = {Investigative Ophthalmology \& Visual Science}, volume = {41}, year = {2000}, month = {2000 Jan}, pages = {199-203}, abstract = {

PURPOSE: To discover proteins that have the potential to contribute to the tight packing of fiber cells in the lens.

METHODS: Crude fiber cell membranes were isolated from ovine lens cortex. Proteins were separated by two-dimensional gel electrophoresis, and selected protein spots identified by micro-sequencing. The identification of galectin-3 was confirmed by immunoblotting with a specific antibody. The association of galectin-3 with the fiber cell plasma membrane was investigated using immunofluorescence microscopy, solubilization trials with selected reagents, and immunoprecipitation to identify candidate ligands.

RESULTS: A cluster of three protein spots with an apparent molecular weight of 31,000 and isoelectric points ranging between 7 and 8.5 were resolved and identified as galectin-3. This protein was associated peripherally with the fiber cell plasma membrane and interacted with MP20, an abundant intrinsic membrane protein that had been identified previously as a component of membrane junctions between fiber cells.

CONCLUSIONS: The detection of galectin-3 in the lens is a novel result and adds to the growing list of lens proteins with adhesive properties. Its location at the fiber cell membrane and its association with the junction-forming MP20 is consistent with a potential role in the development or maintenance of the tightly packed lens tissue architecture.

}, keywords = {Amino Acid Sequence, Animals, Antigens, Differentiation, Cell Membrane, Chromatography, High Pressure Liquid, Electrophoresis, Gel, Two-Dimensional, Eye Proteins, Galectin 3, Immunoblotting, Lens Cortex, Crystalline, Ligands, Membrane Glycoproteins, Microscopy, Fluorescence, Molecular Sequence Data, Rabbits, Sequence Homology, Amino Acid, Sheep}, issn = {0146-0404}, author = {Gonen, T and Donaldson, P and Kistler, J} } @article {957, title = {The genome sequence of Drosophila melanogaster.}, journal = {Science}, volume = {287}, year = {2000}, month = {2000 Mar 24}, pages = {2185-95}, abstract = {

The fly Drosophila melanogaster is one of the most intensively studied organisms in biology and serves as a model system for the investigation of many developmental and cellular processes common to higher eukaryotes, including humans. We have determined the nucleotide sequence of nearly all of the approximately 120-megabase euchromatic portion of the Drosophila genome using a whole-genome shotgun sequencing strategy supported by extensive clone-based sequence and a high-quality bacterial artificial chromosome physical map. Efforts are under way to close the remaining gaps; however, the sequence is of sufficient accuracy and contiguity to be declared substantially complete and to support an initial analysis of genome structure and preliminary gene annotation and interpretation. The genome encodes approximately 13,600 genes, somewhat fewer than the smaller Caenorhabditis elegans genome, but with comparable functional diversity.

}, keywords = {Animals, Biological Transport, Chromatin, Cloning, Computational Biology, Contig Mapping, Cytochrome P-450 Enzyme System, DNA, DNA Repair, DNA Replication, Drosophila melanogaster, Euchromatin, Gene Library, Genes, Genetic, Genome, Heterochromatin, Insect, Insect Proteins, Molecular, Nuclear Proteins, Protein Biosynthesis, Sequence Analysis, Transcription}, issn = {0036-8075}, doi = {10.1186/gb-2007-8-7-r145}, author = {Adams, M D and Celniker, S E and Holt, R A and Evans, C A and Gocayne, J D and Amanatides, P G and Scherer, S E and Li, P W and Hoskins, R A and Galle, R F and George, R A and Lewis, S E and Richards, S and Ashburner, M and Henderson, S N and Sutton, G G and Wortman, J R and Yandell, M D and Zhang, Q and Chen, L X and Brandon, R C and Rogers, Y H and Blazej, R G and Champe, M and Pfeiffer, B D and Wan, K H and Doyle, C and Baxter, E G and Helt, G and Nelson, C R and Gabor, G L and Abril, J F and Agbayani, A and An, H J and Andrews-Pfannkoch, C and Baldwin, D and Ballew, R M and Basu, A and Baxendale, J and Bayraktaroglu, L and Beasley, E M and Beeson, K Y and Benos, P V and Berman, B P and Bhandari, D and Bolshakov, S and Borkova, D and Botchan, M R and Bouck, J and Brokstein, P and Brottier, P and Burtis, K C and Busam, D A and Butler, H and Cadieu, E and Center, A and Chandra, I and Cherry, J M and Cawley, S and Dahlke, C and Davenport, L B and Davies, P and de Pablos, B and Delcher, A and Deng, Z and Mays, A D and Dew, I and Dietz, S M and Dodson, K and Doup, L E and Downes, M and Dugan-Rocha, S and Dunkov, B C and Dunn, P and Durbin, K J and Evangelista, C C and Ferraz, C and Ferriera, S and Fleischmann, W and Fosler, C and Gabrielian, A E and Garg, N S and Gelbart, W M and Glasser, K and Glodek, A and Gong, F and Gorrell, J H and Gu, Z and Guan, P and Harris, M and Harris, N L and Harvey, D and Heiman, T J and Hernandez, J R and Houck, J and Hostin, D and Houston, K A and Howland, T J and Wei, M H and Ibegwam, C and Jalali, M and Kalush, F and Karpen, G H and Ke, Z and Kennison, J A and Ketchum, K A and Kimmel, B E and Kodira, C D and Kraft, C and Kravitz, S and Kulp, D and Lai, Z and Lasko, P and Lei, Y and Levitsky, A A and Li, J and Li, Z and Liang, Y and Lin, X and Liu, X and Mattei, B and McIntosh, T C and McLeod, M P and McPherson, D and Merkulov, G and Milshina, N V and Mobarry, C and Morris, J and Moshrefi, A and Mount, S M and Moy, M and Murphy, B and Murphy, L and Muzny, D M and Nelson, D L and Nelson, D R and Nelson, K A and Nixon, K and Nusskern, D R and Pacleb, J M and Palazzolo, M and Pittman, G S and Pan, S and Pollard, J and Puri, V and Reese, M G and Reinert, K and Remington, K and Saunders, R D and Scheeler, F and Shen, H and Shue, B C and Sid{\'e}n-Kiamos, I and Simpson, M and Skupski, M P and Smith, T and Spier, E and Spradling, A C and Stapleton, M and Strong, R and Sun, E and Svirskas, R and Tector, C and Turner, R and Venter, E and Wang, A H and Wang, X and Wang, Z Y and Wassarman, D A and Weinstock, G M and Weissenbach, J and Williams, S M and Williams, S M and Worley, K C and Wu, D and Yang, S and Yao, Q A and Ye, J and Yeh, R F and Zaveri, J S and Zhan, M and Zhang, G and Zhao, Q and Zheng, L and Zheng, X H and Zhong, F N and Zhong, W and Zhou, X and Zhu, S and Zhu, X and Smith, H O and Gibbs, R A and Myers, E W and Rubin, G M and Venter, J C} } @article {2854, title = {Identification of a yeast transcription factor IID subunit, TSG2/TAF48.}, journal = {The Journal of Biological Chemistry}, volume = {275}, year = {2000}, month = {2000 Jun 9}, pages = {17391-8}, abstract = {

The RNA polymerase II general transcription factor TFIID is a complex containing the TATA-binding protein (TBP) and associated factors (TAFs). We have used a mutant allele of the gene encoding yeast TAF(II)68/61p to analyze its function in vivo. We provide biochemical and genetic evidence that the C-terminal alpha-helix of TAF(II)68/61p is required for its direct interaction with TBP, the stable incorporation of TBP into the TFIID complex, the integrity of the TFIID complex, and the transcription of most genes in vivo. This is the first evidence that a yeast TAF(II) other than TAF(II)145/130 interacts with TBP, and the implications of this on the interpretation of data obtained studying TAF(II) mutants in vivo are discussed. We have identified a high copy suppressor of the TAF68/61 mutation, TSG2, that has sequence similarity to a region of the SAGA subunit Ada1. We demonstrate that it directly interacts with TAF(II)68/61p in vitro, is a component of TFIID, is required for the stability of the complex in vivo, and is necessary for the transcription of many yeast genes. On the basis of these functions, we propose that Tsg2/TAF(II)48p is the histone 2A-like dimerization partner for the histone 2B-like TAF(II)68/61p in the yeast TFIID complex.

}, keywords = {Mutagenesis, Phenotype, Protein Structure, Recombinant Fusion Proteins, Saccharomyces cerevisiae, Saccharomyces cerevisiae Proteins, Secondary, TATA-Binding Protein Associated Factors, Temperature, TFII, Transcription Factor TFIID, Transcription Factors}, issn = {0021-9258}, doi = {10.1074/jbc.M001635200}, author = {Reese, J C and Zhang, Z and Kurpad, H} } @article {1783, title = {A molecular pathway for light-dependent photoreceptor apoptosis in Drosophila.}, journal = {Neuron}, volume = {28}, year = {2000}, month = {2000 Oct 5}, pages = {139-52}, abstract = {

Light-induced photoreceptor apoptosis occurs in many forms of inherited retinal degeneration resulting in blindness in both vertebrates and invertebrates. Though mutations in several photoreceptor signaling proteins have been implicated in triggering this process, the molecular events relating light activation of rhodopsin to photoreceptor death are yet unclear. Here, we uncover a pathway by which activation of rhodopsin in Drosophila mediates apoptosis through a G protein-independent mechanism. This process involves the formation of membrane complexes of phosphorylated, activated rhodopsin and its inhibitory protein arrestin, and subsequent clathrin-dependent endocytosis of these complexes into a cytoplasmic compartment. Together, these data define the proapoptotic molecules in Drosophila photoreceptors and indicate a novel signaling pathway for light-activated rhodopsin molecules in control of photoreceptor viability.

}, keywords = {Animals, Apoptosis, Arrestins, Binding, Calcium-Binding Proteins, Clathrin, Competitive, Drosophila, Drosophila Proteins, Endocytosis, Gene Expression Regulation, Invertebrate, Light, Mutation, Phosphoprotein Phosphatases, Phosphoproteins, Phosphorylation, Photoreceptor Cells, Retinal Degeneration, Rhodopsin}, issn = {0896-6273}, author = {Kiselev, A and Socolich, M and Vin{\'o}s, J and Hardy, R W and Zuker, C S and Ranganathan, R} } @proceedings {45804, title = {Morphogenesis and gene expressions in the parthenogenetic embryogenesis of the pea aphid Acyrthosiphon pisum}, volume = {17}, year = {2000}, pages = {66}, publisher = {社団法人日本動物学会}, issn = {02890003}, url = {http://ci.nii.ac.jp/naid/110003371583/}, author = {T Miura and S Kambhampati and DL Stern} } @article {2016, title = {Optical imaging of calcium transients in neurons and pharyngeal muscle of C. elegans.}, journal = {Neuron}, volume = {26}, year = {2000}, month = {2000 Jun}, pages = {583-94}, abstract = {

Electrophysiology and optical indicators have been used in vertebrate systems to investigate excitable cell firing and calcium transients, but both techniques have been difficult to apply in organisms with powerful reverse genetics. To overcome this limitation, we expressed cameleon proteins, genetically encoded calcium indicators, in the pharyngeal muscle of the nematode worm Caenorhabditis elegans. In intact transgenic animals expressing cameleons, fluorescence ratio changes accompanied muscular contraction, verifying detection of calcium transients. By comparing the magnitude and duration of calcium influx in wild-type and mutant animals, we were able to determine the effects of calcium channel proteins on pharyngeal calcium transients. We also successfully used cameleons to detect electrically evoked calcium transients in individual C. elegans neurons. This technique therefore should have broad applications in analyzing the regulation of excitable cell activity in genetically tractable organisms.

}, keywords = {Animals, Caenorhabditis elegans, Caenorhabditis elegans Proteins, Calcium, Calcium Channels, Down-Regulation, Electrophysiology, Helminth Proteins, Muscle Proteins, Mutation, Neurons, Optics and Photonics, Pharyngeal Muscles, Protein Isoforms}, issn = {0896-6273}, author = {Kerr, R and Lev-Ram, V and Baird, G and Vincent, P and Tsien, R Y and Schafer, W R} } @article {1520, title = {Short-range and long-range guidance by slit and its Robo receptors. Robo and Robo2 play distinct roles in midline guidance.}, journal = {Neuron}, volume = {28}, year = {2000}, month = {2000 Dec}, pages = {753-66}, abstract = {

Previous studies showed that Roundabout (Robo) in Drosophila is a repulsive axon guidance receptor that binds to Slit, a repellent secreted by midline glia. In robo mutants, growth cones cross and recross the midline, while, in slit mutants, growth cones enter the midline but fail to leave it. This difference suggests that Slit must have more than one receptor controlling midline guidance. In the absence of Robo, some other Slit receptor ensures that growth cones do not stay at the midline, even though they cross and recross it. Here we show that the Drosophila genome encodes three Robo receptors and that Robo and Robo2 have distinct functions, which together control repulsive axon guidance at the midline. The robo,robo2 double mutant is largely identical to slit.

}, keywords = {Animals, Axons, Cell Line, Cell Surface, Central Nervous System, Cloning, COS Cells, Dimerization, Down-Regulation, Drosophila, Drosophila Proteins, Gene Expression, Growth Cones, Humans, Immunologic, Insect Proteins, Insertional, Ligands, Membrane Proteins, Messenger, Molecular, Molecular Sequence Data, Mutagenesis, Nerve Tissue Proteins, Phenotype, Receptors, RNA}, issn = {0896-6273}, author = {Simpson, J H and Kidd, T and Bland, K S and Goodman, C S} } @article {2705, title = {A whole-genome assembly of Drosophila.}, journal = {Science}, volume = {287}, year = {2000}, month = {2000 Mar 24}, pages = {2196-204}, abstract = {

We report on the quality of a whole-genome assembly of Drosophila melanogaster and the nature of the computer algorithms that accomplished it. Three independent external data sources essentially agree with and support the assembly{\textquoteright}s sequence and ordering of contigs across the euchromatic portion of the genome. In addition, there are isolated contigs that we believe represent nonrepetitive pockets within the heterochromatin of the centromeres. Comparison with a previously sequenced 2.9- megabase region indicates that sequencing accuracy within nonrepetitive segments is greater than 99. 99\% without manual curation. As such, this initial reconstruction of the Drosophila sequence should be of substantial value to the scientific community.

}, keywords = {Algorithms, Animals, Chromatin, Computational Biology, Contig Mapping, DNA, Drosophila melanogaster, Euchromatin, Genes, Genome, Heterochromatin, Insect, Molecular Sequence Data, Nucleic Acid, Physical Chromosome Mapping, Repetitive Sequences, Sequence Analysis, Sequence Tagged Sites}, issn = {0036-8075}, author = {Myers, E W and Sutton, G G and Delcher, A L and Dew, I M and Fasulo, D P and Flanigan, M J and Kravitz, S A and Mobarry, C M and Reinert, K H and Remington, K A and Anson, E L and Bolanos, R A and Chou, H H and Jordan, C M and Halpern, A L and Lonardi, S and Beasley, E M and Brandon, R C and Chen, L and Dunn, P J and Lai, Z and Liang, Y and Nusskern, D R and Zhan, M and Zhang, Q and Zheng, X and Rubin, G M and Adams, M D and Venter, J C} } @article {1270, title = {A wire length minimization approach to ocular dominance patterns in mammalian visual cortex.}, journal = {Physica A}, volume = {284}, year = {2000}, month = {2000}, pages = {318-34}, author = {Chklovskii, Dmitri B and Koulakov, Alexei A} } @article {1745, title = {Decrystallization of adult birdsong by perturbation of auditory feedback.}, journal = {Nature}, volume = {399}, year = {1999}, month = {1999 Jun 3}, pages = {466-70}, abstract = {

Young birds learn to sing by using auditory feedback to compare their own vocalizations to a memorized or innate song pattern; if they are deafened as juveniles, they will not develop normal songs. The completion of song development is called crystallization. After this stage, song shows little variation in its temporal or spectral properties. However, the mechanisms underlying this stability are largely unknown. Here we present evidence that auditory feedback is actively used in adulthood to maintain the stability of song structure. We found that perturbing auditory feedback during singing in adult zebra finches caused their song to deteriorate slowly. This {\textquoteright}decrystallization{\textquoteright} consisted of a marked loss of the spectral and temporal stereotypy seen in crystallized song, including stuttering, creation, deletion and distortion of song syllables. After normal feedback was restored, these deviations gradually disappeared and the original song was recovered. Thus, adult birds that do not learn new songs nevertheless retain a significant amount of plasticity in the brain.

}, keywords = {Aging, Animals, Auditory Perception, Feedback, Learning, Male, Songbirds}, issn = {0028-0836}, doi = {10.1038/20933}, author = {Leonardo, A and Konishi, M} } @article {3088, title = {Hippocampal damage induced by carbon monoxide poisoning and spreading depression is alleviated by chronic treatment with brain derived polypeptides.}, journal = {Brain Research}, volume = {816}, year = {1999}, month = {1999 Jan 23}, pages = {618-27}, abstract = {

A model of acute carbon monoxide poisoning combined with spreading depression (SD) induced metabolic stress was used to examine the protective effects of cerebrolysin (CL) on the development of electrophysiological, behavioral and morphological signs of hypoxic damage. Capillary electrodes were implanted into the neocortex and hippocampus of anesthetized rats which were then exposed for 90 min to breathing of 0.8\% to 0.5\% CO, while 3 to 4 waves of cortical and hippocampal SD were elicited by microinjections of 5\% KCl. Duration of SD-provoked depolarization of cerebral cortex and hippocampus was noted. Nine and 18 to 19 days later propagation of SD waves was recorded with the same electrodes and decrease of their amplitude was used as an index of brain damage which was significant in the hippocampus but not in the cortex. CL-treatment (2.5 ml/kg per day) started after CO administration and continued for 14 days significantly improved hippocampal recovery manifested by increased amplitude of SD waves. Behavioral tests performed 10 and 20 days after CO poisoning in the Morris water maze revealed better performance (escape latency 7 s) in the CL-treated than in untreated animals (14 s). Morphological analysis showed marked damage in the hippocampus consonant with electrophysiological and behavioral findings in the same animals. No apparent histological damage was found in rats exposed to CO inhalation alone without the additional SD-provoked depolarization. It is concluded that chronic CL-treatment enhances recovery of hippocampal tissue after hypoxic damage of intermediate severity.

}, keywords = {Acute Disease, Analysis of Variance, Animals, Brain, Carbon Monoxide Poisoning, Cortical Spreading Depression, Electrodes, Hippocampus, Hypoxia, Implanted, Long-Evans, Male, Maze Learning, Membrane Potentials, Neuropeptides, Rats, Reaction Time}, issn = {0006-8993}, author = {Koroleva, V I and Korolev, O S and Mares, V and Pastalkova, E and Bures, J} } @article {45813, title = {Determining aphid taxonomic affinities and life cycles with molecular data: a case study of the tribe Cerataphidini (Hormaphididae: Aphidoidea: Hemiptera)}, journal = {Systematic Entomology}, volume = {22}, year = {1997}, month = {01/1997}, pages = {81-96}, abstract = {

Aphid taxonomy is often frustrated by the host alternation and extensive polyphenism displayed by many species. Here we examine the utility of using molecular data to assist in life cycle and taxonomic determination. We found that a relatively small amount of DNA sequence data can greatly assist in these tasks. Molecular data have identified the synonymy of five species: Tuberaphis plicator (Noordam) is a junior synonym of T.takenouchii (Takahashi), T.taiwana (Takahashi) is a junior synonym of T.coreana Takahashi, Hamiltonaphis styraci (Matsumura) is transferred to Tuberaphis Takahashi, Astegopteryx roepkei Hille Ris Lambers is transferred to Ceratoglyphina van der Goot, and A.vandermeermohri Hille Ris Lambers is transferred to Cerataphis Lichtenstein. We have elucidated the complete life cycles of five species: A.basalis (van der Goot) alternates between Styrax benzoin and bamboos, Ceratoglyphina bambusae van der Goot alternates between S.benzoin and bamboos, Pseudoregma sundanica (van der Goot) alternates between S.paralleloneura and Zingiberaceae, T.coreana alternates between S.formosana and Loranthaceae, and T.takenouchii alternates between S.japonica and Loranthaceae. In all cases the molecular data agreed with available morphological data. This analysis demonstrates the utility of DNA sequence comparisons for elucidating complex life cycles and the taxonomy of difficult insect groups.

}, doi = {10.1046/j.1365-3113.1997.d01-20.x}, author = {David Stern and Shigeyuki Aoki and Shigeyuki Kurosu} } @article {45817, title = {The life cycle and natural history of the tropical aphid Cerataphis fransseni (Homoptera: Aphididae: Hormaphidinae), with reference to the evolution of host alternation in aphids}, journal = {Journal of natural history}, volume = {29}, year = {1995}, pages = {231-242}, abstract = {

We describe the life cycle and general biology of the tropical cerataphidine aphid Cerataphis fransseni. We demonstrate that this aphid migrates between trees of Styrax benzoin and various species of palms; palm-feeding populations have previously been known as C. variabilis and C. palmae, which now become synonyms of C. fransseni. On S. benzoin the fundatrix induces a relatively simple gall which can contain \>6000 aphids at maturity with a large number of reproductively sterile soldiers that protect the gall from predators. These galls are apparently produced throughout the year. Colonies on the secondary host plants, palms, are apparently obligately tended by ants whereas colonies within galls on Styrax are never tended by ants. We discuss the life cycle of this tropical aphid with respect to hypotheses for the evolution and maintenance of host alternation.

}, doi = {10.1080/00222939500770101}, url = {http://www.tandfonline.com/doi/abs/10.1080/00222939500770101$\#$.VKrUcHt79iZ}, author = {DL Stern and S Aoki and D U Kurosu} } @article {45818, title = {A test of geometric hypotheses for soldier investment patterns in the gall producing tropical aphidCerataphis fransseni (Homoptera, Hormaphididae)}, journal = {Insectes sociaux}, volume = {41}, year = {1994}, pages = {457-460}, abstract = {

The gall-forming aphidCerataphis fransseni produces soldiers that defend against predators. Soldiers are produced soon after colony foundation and the number of soldiers increases nonlinearly during colony growth. The number of soldiers scales to the square-root of the number of non-soldiers and linearly to the surface area of the gall. This suggests that soldiers are produced to defend an area, for example the perimeter of the colony or the surface of the gall, rather than individual aphids.

}, issn = {0020-1812}, doi = {10.1007/BF01240648}, url = {http://link.springer.com/article/10.1007/BF01240648}, author = {D. L. Stern and S. Aoki and U. Kurosu} } @article {1157, title = {Sex-lethal, master and slave: the hierarchy of germline sex determination in Drosophila.}, journal = {Development}, volume = {119}, year = {1993}, month = {1993 Nov}, pages = {897-908}, abstract = {

Female sex determination in the germ line of Drosophila melanogaster is regulated by genes functioning in the soma as well as genes that function within the germ line. Genes known or suspected to be involved in germ-line sex determination in Drosophila melanogaster have been examined to determine if they are required upstream or downstream of Sex-lethal+, a known germ-line sex determination gene. Seven genes required for female-specific splicing of germ-line Sex-lethal+ pre-mRNA are identified. These results together with information about the tissues in which these genes function and whether they control sex determination and viability or just sex determination in the germ line have been used to deduce the genetic hierarchy regulating female germ-line sex determination. This hierarchy includes the somatic sex determination genes transformer+, transformer-2+ and doublesex+ (and by inference Sex-lethal+), which control a somatic signal required for female germ-line sex determination, and the germ-line ovarian tumor genes fused+, ovarian tumor+, ovo+, sans fille+, and Sex-lethal+, which are involved in either the reception or interpretation of this somatic sex determination signal. The fused+, ovarian tumor+, ovo+ and sans fille+ genes function upstream of Sex-lethal+ in the germ line.

}, issn = {0950-1991}, author = {Baker, B.S. and Oliver, B. and Kim, Yi Jin} } @article {966, title = {The argos gene encodes a diffusible factor that regulates cell fate decisions in the Drosophila eye.}, journal = {Cell}, volume = {69}, year = {1992}, month = {1992 Jun 12}, pages = {963-75}, abstract = {

The argos gene encodes a protein that is required for viability and that regulates the determination of cells in the Drosophila eye. A developmental analysis of argos mutant eyes indicates that the mystery cells, which are usually nonneuronal, are transformed into extra photoreceptors, and that supernumerary cone cells and pigment cells are also recruited. Clonal analysis indicates that argos acts nonautonomously and can diffuse over the range of several cell diameters. Conceptual translation of the argos gene suggests that it encodes a secreted protein.

}, keywords = {Amino Acid Sequence, Animals, Base Sequence, Cell Communication, Clone Cells, Cloning, DNA, Drosophila melanogaster, Embryonic Induction, Eye, Gene Expression, Genes, Messenger, Molecular, Molecular Sequence Data, Phenotype, Restriction Mapping, RNA, Signal Transduction}, issn = {0092-8674}, doi = {10.1186/gb-2007-8-7-r145}, author = {Freeman, M and Kl{\"a}mbt, C and Goodman, C S and Rubin, G M} } @article {2097, title = {Near-field magneto-optics and high density data storage. (With commentary)}, journal = {Applied Physics Letters.}, volume = {61}, year = {1992}, month = {1992}, abstract = {

Near-field scanning optical microscopy (NSOM) has been used to image and record domains in thin-film magneto-optic (MO) materials. In the imaging mode, resolution of 30-50 nm has been consistently obtained, whereas in the recording mode, domains down to -60 nm have been written reproducibly. Data densities of -45 Gbits/in.{\textquoteright} have been achieved, well in excess\ of current magnetic or MO technologies. A brief analysis of speed and other issues indicates that the technique may represent a viable alternative to density data storage needs.

Commentary: The first demonstration of optical recording and playback beyond the diffraction limit, using magneto-optic multilayer films and polarization contrast near-field microscopy. Bits as small as 60 nm were recorded {\textendash} beyond estimates at the time of the superparamagnetic limit to bit stability. Bit densities of 45 Gbits/in2 were also achieved, well in excess of optical or magnetic recording technologies of the era. In the years following this work, massive resources were spent on the commercialization of near-field data storage, largely for naught.

}, author = {Betzig, E and Trautman, J K and Wolfe, R and Gyorgy, E M and Finn, P L and Kryder, M H and Chang, C H} } @article {45819, title = {Aphid soldiers discriminate between soldiers and non-soldiers, rather than between kin and non-kin, in Ceratoglyphina bambusae}, journal = {Animal behaviour}, volume = {42}, year = {1991}, pages = {865-866}, abstract = {

Examined kin discrimination and colony defense in soldier-producing aphids from the surface of 3 galls. Soldiers always attacked conspecific nonsoldiers, regardless of origin, and never attacked conspecific soldiers. Soldier attacks of nonsoldiers may exclude unrelated nonsoldier aphids from the gall where they would propagate and compete with resident aphids. (PsycINFO Database Record (c) 2012 APA, all rights reserved)

}, doi = {10.1016/S0003-3472(05)80135-6}, url = {http://psycnet.apa.org/psycinfo/1992-15421-001}, author = {Shigeyuki Aoki and Utako Kurosu and David L Stern} } @article {2564, title = {Breaking the diffraction barrier: optical microscopy on a nanometric scale.}, journal = {Science}, volume = {251}, year = {1991}, month = {1991 Mar 22}, pages = {1468-70}, abstract = {

In near-field scanning optical microscopy, a light source or detector with dimensions less than the wavelength (lambda) is placed in close proximity (lambda/50) to a sample to generate images with resolution better than the diffraction limit. A near-field probe has been developed that yields a resolution of approximately 12 nm ( approximately lambda/43) and signals approximately 10(4)- to 10(6)-fold larger than those reported previously. In addition, image contrast is demonstrated to be highly polarization dependent. With these probes, near-field microscopy appears poised to fulfill its promise by combining the power of optical characterization methods with nanometric spatial resolution.

}, issn = {0036-8075}, doi = {10.1126/science.251.5000.1468}, author = {Betzig, E and Trautman, J K and Harris, T D and Weiner, J S and Kostelak, R L} } @article {2095, title = {Breaking the diffraction barrier: optical microscopy on a nanometric scale. (With commentary)}, journal = {Science}, volume = {251}, year = {1991}, month = {1991 Mar 22}, pages = {1468-70}, abstract = {

In near-field scanning optical microscopy, a light source or detector with dimensions less than the wavelength (lambda) is placed in close proximity (lambda/50) to a sample to generate images with resolution better than the diffraction limit. A near-field probe has been developed that yields a resolution of approximately 12 nm ( approximately lambda/43) and signals approximately 10(4)- to 10(6)-fold larger than those reported previously. In addition, image contrast is demonstrated to be highly polarization dependent. With these probes, near-field microscopy appears poised to fulfill its promise by combining the power of optical characterization methods with nanometric spatial resolution.

Commentary: Introduced the adiabatically tapered single mode fiber probe to near-field scanning optical microscopy which, together with shear force feedback, made the technique a practical reality. Although earlier claims of superresolution via near-field microscopy existed for nearly a decade, this paper was the first to convincingly break Abbe{\textquoteright}s limit with visible light, as demonstrated by reproducibly resolving known, complex nanoscale patterns having features separated by much less than the wavelength. Whereas our fiber probe and shear force technologies were soon widely adopted and key to many novel applications (see above), the earlier methods proved to be technological dead ends, never achieving the results of their original claims. This experience taught me the most valuable lesson of my career: while it{\textquoteright}s bad to bullshit others, it{\textquoteright}s even worse to bullshit yourself. It{\textquoteright}s a lesson sadly unheeded by many current practitioners of superresolution microscopy.

}, issn = {0036-8075}, doi = {10.1126/science.251.5000.1468}, url = {http://www.ncbi.nlm.nih.gov/pubmed/17779440}, author = {Betzig, E and Trautman, J K and Harris, T D and Weiner, J S and Kostelak, R L} } @article {1156, title = {The maleless protein associates with the X chromosome to regulate dosage compensation in Drosophila.}, journal = {Cell}, volume = {66}, year = {1991}, month = {1991 Sep 6}, pages = {935-48}, abstract = {

The maleless (mle) gene is one of four known regulatory loci required for increased transcription (dosage compensation) of X-linked genes in D. melanogaster males. A predicted mle protein (MLE) contains seven short segments that define a superfamily of known and putative RNA and DNA helicases. MLE, while present in the nuclei of both male and female cells, differs in its association with polytene X chromosomes in the two sexes. MLE is associated with hundreds of discrete sites along the length of the X chromosome in males and not in females. The predominant localization of MLE to the X chromosome in males makes it a strong candidate to be a direct regulator of dosage compensation.

}, issn = {0092-8674}, author = {Baker, B.S. and Kuroda, M.I. and Kreber, M.J. and Ganetzky, B.} } @article {45823, title = {Agonistic interactions between ants and gall-living soldier aphids}, journal = {Journal of Ethology}, volume = {8}, year = {1990}, pages = {139-141}, issn = {0289-0771}, doi = {10.1007/BF02350284}, author = {Utako Kurosu and David L Stern and Shigeyuki Aoki} } @article {2375, title = {Evidence for the influence of electron-electron interaction on the chemical potential of the two-dimensional electron gas.}, journal = {Physical Review. B, Condensed Matter}, volume = {42}, year = {1990}, month = {1990 Aug 15}, pages = {3741-44}, abstract = {

It is shown experimentally that the interaction between electrons strongly influences the chemical potential of the two-dimensional (2D) electron gas. At sufficiently low temperatures and in high magnetic fields, regions of filling factor appear where (i) the chemical potential μ diminishes with increasing carrier density, i.e., the thermodynamic density of states is negative; (ii) the derivative ∂μ/∂H (H is the magnetic field) is considerably higher than the maximum value for a noninteracting 2D electron gas. Using these results, we have estimated that the energy of the e-e interaction in Si inversion layers in a magnetic field is about 1 order of magnitude less than the classical Coulomb interaction calculated for Si metal-oxide-semiconductor field-effect transistors.

}, issn = {0163-1829}, doi = {10.1523/JNEUROSCI.3613-08.2008}, author = {Kravchenko, SV and Rinberg, DA and Semenchinsky, SG and Pudalov, VM} } @article {969, title = {The homeo domain protein rough is expressed in a subset of cells in the developing Drosophila eye where it can specify photoreceptor cell subtype.}, journal = {Genes \& Development}, volume = {4}, year = {1990}, month = {1990 May}, pages = {712-27}, abstract = {

The Drosophila homeo box gene rough is required in photoreceptor cells R2 and R5 for normal eye development. We show here that rough protein expression is limited to a subset of cells in the developing retina where it is transiently expressed for 30-60 hr. The rough protein is first expressed broadly in the morphogenetic furrow but is rapidly restricted to the R2, R3, R4, and R5 precursor cells. Ubiquitous expression of rough under the control of the hsp70 promoter in third-instar larvae suppresses the initial steps of ommatidial assembly. Structures derived from other imaginal discs are not affected. Ectopic expression of rough in the R7 precursor, through the use of the sevenless promoter, causes this cell to develop into an R1-6 photoreceptor subtype; however, this cell still requires sevenless function for its neural differentiation. Taken together with previous analyses of the rough mutant phenotype, these results suggest that the normal role of rough is to establish the unique cell identity of photoreceptors R2 and R5.

}, keywords = {Animals, Base Sequence, Cell Cycle Proteins, DNA-Binding Proteins, Drosophila melanogaster, Drosophila Proteins, Eye, Gene Expression, Genes, Genetic, Heat-Shock Proteins, Homeobox, Hot Temperature, Microtubule-Associated Proteins, Molecular Sequence Data, Morphogenesis, Phenotype, Photoreceptor Cells, Promoter Regions, Transcription Factors}, issn = {0890-9369}, doi = {10.1186/gb-2007-8-7-r145}, author = {Kimmel, B E and Heberlein, U and Rubin, G M} } @article {2463, title = {Postmetamorphic cell death in the nervous and muscular systems of Drosophila melanogaster.}, journal = {The Journal of Neuroscience: The Official Journal of the Society for Neuroscience}, volume = {10}, year = {1990}, month = {1990 Feb}, pages = {403-1}, abstract = {

Programmed cell death occurs in the nervous and muscular system of newly emerged adult Drosophila melanogaster. Many of the abdominal muscles that were used for eclosion and wing-spreading behavior degenerate by 12 hr after eclosion. Related neurons in the ventral ganglion also die within the first 24 hr. Ligation experiments showed that the muscle breakdown is triggered by a signal from the anterior region, presumably the head, that occurs about 1 hr before adult emergence. The timing of this signal suggests that eclosion hormone may be involved. Although muscle death is triggered prior to ecdysis, it can be delayed, at least temporarily, by forcing the emerging flies to show a prolonged ecdysis behavior. In contrast to the muscles, the death of the neurons is triggered after emergence. The signal for neuronal degeneration is closely correlated with the initiation of wing inflation behavior. Ligation and digging experiments and behavioral manipulations that either blocked or delayed wing expansion behavior had a parallel effect in suppressing or delaying neuronal death.

}, keywords = {Animal, Animals, Behavior, Biological, Cell Survival, Drosophila melanogaster, Female, Ligation, Male, Metamorphosis, Muscles, Nervous System, Neurons}, issn = {0270-6474}, author = {Kimura, K I and Truman, J W} } @article {924, title = {Evaporative cooling of spin-polarized atomic hydrogen.}, journal = {Physical Review Letters}, volume = {61}, year = {1988}, month = {1988 Aug 22}, pages = {935-8}, abstract = {

A gas of hydrogen atoms, confined in a static magnetic trap, has been evaporatively cooled to temperatures of a few millikelvin. The initial trap configuration held the gas at 38 mK for as long as 5 h. Evaporative cooling reduced the temperature to 3.0 mK while maintaining the central density at 7.6{\texttimes}10\ 12\ \  cm\ -3\ \  . These values were determined by measurement of the rate of electronic spin relaxation and are in agreement with model calculations. Further cooling to 1 mK (inferred from the model) has been achieved. Measurements were made of the efficiency of the evaporative cooling process.

}, issn = {1079-7114}, author = {Masuhara, N and Doyle, JM and Sandberg, JC and Kleppner, D and Greytak, TJ and Hess, H F and Kochanski, GP} } @article {978, title = {Analysis of P transposable element functions in Drosophila.}, journal = {Cell}, volume = {38}, year = {1984}, month = {1984 Aug}, pages = {135-46}, abstract = {

We have made a P-element derivative called Pc[ry], which carries the selectable marker gene rosy, but which acts like a nondefective, intact P element. It transposes autonomously into the germline chromosomes of an M-strain Drosophila embryo and it mobilizes in trans the defective P elements of the singed-weak allele. Frameshift mutations introduced into any of the four major open reading frames of the P sequence were each sufficient to eliminate the transposase activity, but none affected signals required in cis for transposition of the element. Complementation tests between pairs of mutant elements suggest that a single polypeptide comprises the transposase. We have examined transcripts of P elements both from natural P strains and from lines containing only nondefective Pc[ry] elements, and have identified two RNA species that appear to be specific for autonomous elements.

}, keywords = {Alleles, Animals, Base Sequence, Crosses, DNA Restriction Enzymes, DNA Transposable Elements, Drosophila, Female, Genes, Genetic, Genetic Complementation Test, Male, Mutation, Nucleic Acid Hybridization}, issn = {0092-8674}, doi = {10.1186/gb-2007-8-7-r145}, author = {Karess, R E and Rubin, G M} } @article {981, title = {The molecular basis of P-M hybrid dysgenesis: the role of the P element, a P-strain-specific transposon family.}, journal = {Cell}, volume = {29}, year = {1982}, month = {1982 Jul}, pages = {995-1004}, abstract = {

We have shown previously that four of five white mutant alleles arising in P-M dysgenic hybrids result from the insertion of strongly homologous DNA sequence elements. We have named these P elements. We report that P elements are present in 30-50 copies per haploid genome in all P strains examined and apparently are missing entirely from all M strains examined, with one exception. Furthermore, members of the P family apparently transpose frequently in P-M dysgenic hybrids; chromosomes descendant from P-M dysgenic hybrids frequently show newly acquired P elements. Finally, the strain-specific breakpoint hotspots for the rearrangement of the pi 2 P X chromosome occurring in P-M dysgenic hybrids are apparently sites of residence of P elements. These observations strongly support the P factor hypothesis for the mechanistic basis of P-M hybrid dysgenesis.

}, keywords = {Base Sequence, Biological Evolution, DNA Transposable Elements, Drosophila, Female, Gene Expression Regulation, Genetic, Hybridization, Infertility, Male, Nucleic Acid, Recombination, Repetitive Sequences}, issn = {0092-8674}, doi = {10.1186/gb-2007-8-7-r145}, author = {Bingham, P M and Kidwell, M G and Rubin, G M} } @article {68294, title = {Visual projection neuron convergence and compensation in developing sensorimotor circuits in the Drosophila optic glomeruli}, month = {2022 Aug 22}, abstract = {

Visual features detected by the early visual system must be combined into higher order representations to guide behavioral decision. Although key developmental mechanisms that enable the separation of visual feature channels in early visual circuits have been discovered, relatively little is known about the mechanisms that underlie their convergence in later stages of visual processing. Here we explore the development of a functionally well-characterized sensorimotor circuit in\ Drosophila melanogaster, the convergence of visual projection neurons (VPNs) onto the dendrites of a large descending neuron called the giant fiber (GF). We find two VPNs encoding different visual features that target the same giant fiber dendrite establish their territories on the dendrite, in part, through sequential axon arrival during development prior to synaptogenesis. Physical occupancy is important to maintain territories, as we find the ablation of one VPN results in expanded dendrite territory of the remaining VPN, and that this compensation enables the GF to remain responsive to ethologically relevant visual stimuli. Our data highlight temporal mechanisms for visual feature convergence and promote the GF circuit, and the\ Drosophila\ optic glomeruli where VPN to GF connectivity resides, as an ideal developmental model for investigating complex wiring programs and plasticity in visual feature convergence.

}, doi = {10.1101/2022.08.20.504653}, url = {https://doi.org/10.1101/2022.08.20.504653}, author = {Brennan W. McFarland and HyoJong Jang and Natalie Smolin and Tanja A. Godenschwege and Aljoscha Nern and Yerbol Z. Kurmangaliyev and Catherine R. von Reyn} }