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3836 Publications

Showing 121-130 of 3836 results
09/01/23 | OME-Zarr: a cloud-optimized bioimaging file format with international community support.
Josh Moore , Daniela Basurto-Lozada , Sébastien Besson , John Bogovic , Eva M. Brown , Jean-Marie Burel , Gustavo de Medeiros , Erin E. Diel , David Gault , Satrajit S. Ghosh , Ilan Gold , Yaroslav O. Halchenko , Matthew Hartley , Dave Horsfall , Mark S. Keller , Mark Kittisopikul , Gabor Kovacs , Aybüke Küpcü Yoldaş , Albane le Tournoulx de la Villegeorges , Tong Li , Prisca Liberali , Melissa Linkert , Dominik Lindner , Joel Lüthi , Jeremy Maitin-Shepard , Trevor Manz , Matthew McCormick , Khaled Mohamed , William Moore , Bugra Özdemir , Constantin Pape , Lucas Pelkmans , Martin Prete , Tobias Pietzsch , Stephan Preibisch , Norman Rzepka , David R. Stirling , Jonathan Striebel , Christian Tischer , Daniel Toloudis , Petr Walczysko , Alan M. Watson , Frances Wong , Kevin A. Yamauchi , Omer Bayraktar , Muzlifah Haniffa , Stephan Saalfeld , Jason R. Swedlow
Histochemistry and Cell Biology. 2023 Feb 25;160(3):223-251. doi: 10.1007/s00418-023-02209-1

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 – OME-Zarr – 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 — the file format that underlies so many personal, institutional, and global data management and analysis tasks.

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09/01/23 | The Neural Basis of Drosophila Courtship Song
Joshua L. Lillvis , Kaiyu Wang , Hiroshi M. Shiozaki , Min Xu , David L. Stern , Barry J. Dickson
bioRxiv. 2023 Sep 01:. doi: 10.1101/2023.08.30.555537

Animal sounds are produced by patterned vibrations of specific organs, but the neural circuits that drive these vibrations are not well defined in any animal. Here we provide a functional and synaptic map of most of the neurons in the Drosophila male ventral nerve cord (the analog of the vertebrate spinal cord) that drive complex, patterned song during courtship. Male Drosophila vibrate their wings toward females during courtship to produce two distinct song modes – pulse and sine song – with characteristic features that signal species identity and male quality. We identified song-producing neural circuits by optogenetically activating and inhibiting identified cell types in the ventral nerve cord (VNC) and by tracing their patterns of synaptic connectivity in the male VNC connectome. The core song circuit consists of at least eight cell types organized into overlapping circuits, where all neurons are required for pulse song and a subset are required for sine song. The pulse and sine circuits each include a feed-forward pathway from brain descending neurons to wing motor neurons, with extensive reciprocal and feed-back connections. We also identify specific neurons that shape the individual features of each song mode. These results reveal commonalities amongst diverse animals in the neural mechanisms that generate diverse motor patterns from a single set of muscles.

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08/31/23 | Direct observation of the conformational states of PIEZO1.
Mulhall EM, Gharpure A, Lee RM, Dubin AE, Aaron JS, Marshall KL, Spencer KR, Reiche MA, Henderson SC, Chew T, Patapoutian A
Nature. 2023 Aug 31;620(7976):1117-1125. doi: 10.1038/s41586-023-06427-4

PIEZOs are mechanosensitive ion channels that convert force into chemoelectric signals and have essential roles in diverse physiological settings. In vitro studies have proposed that PIEZO channels transduce mechanical force through the deformation of extensive blades of transmembrane domains emanating from a central ion-conducting pore. However, little is known about how these channels interact with their native environment and which molecular movements underlie activation. Here we directly observe the conformational dynamics of the blades of individual PIEZO1 molecules in a cell using nanoscopic fluorescence imaging. Compared with previous structural models of PIEZO1, we show that the blades are significantly expanded at rest by the bending stress exerted by the plasma membrane. The degree of expansion varies dramatically along the length of the blade, where decreased binding strength between subdomains can explain increased flexibility of the distal blade. Using chemical and mechanical modulators of PIEZO1, we show that blade expansion and channel activation are correlated. Our findings begin to uncover how PIEZO1 is activated in a native environment. More generally, as we reliably detect conformational shifts of single nanometres from populations of channels, we expect that this approach will serve as a framework for the structural analysis of membrane proteins through nanoscopic imaging.

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08/29/23 | Functionalization and higher-order organization of liposomes with DNA nanostructures.
Zhang Z, Feng Z, Zhao X, Jean D, Yu Z, Chapman ER
Nature Communications. 2023 Aug 29;14(1):5256. doi: 10.1038/s41467-023-41013-2

Small unilamellar vesicles (SUVs) are indispensable model membranes, organelle mimics, and drug and vaccine carriers. However, the lack of robust techniques to functionalize or organize preformed SUVs limits their applications. Here we use DNA nanostructures to coat, cluster, and pattern sub-100-nm liposomes, generating distance-controlled vesicle networks, strings and dimers, among other configurations. The DNA coating also enables attachment of proteins to liposomes, and temporal control of membrane fusion driven by SNARE protein complexes. Such a convenient and versatile method of engineering premade vesicles both structurally and functionally is highly relevant to bottom-up biology and targeted delivery.

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08/25/23 | Rational Engineering of an Improved Genetically Encoded pH Sensor Based on Superecliptic pHluorin.
Shen Y, Wen Y, Sposini S, Vishwanath AA, Abdelfattah AS, Schreiter ER, Lemieux MJ, de Juan-Sanz J, Perrais D, Campbell RE
ACS Sensors. 2023 Aug 25;8(8):3014-3022. doi: 10.1021/acssensors.3c00484

Genetically encoded pH sensors based on fluorescent proteins are valuable tools for the imaging of cellular events that are associated with pH changes, such as exocytosis and endocytosis. Superecliptic pHluorin (SEP) is a pH-sensitive green fluorescent protein (GFP) variant widely used for such applications. Here, we report the rational design, development, structure, and applications of Lime, an improved SEP variant with higher fluorescence brightness and greater pH sensitivity. The X-ray crystal structure of Lime supports the mechanistic rationale that guided the introduction of beneficial mutations. Lime provides substantial improvements relative to SEP for imaging of endocytosis and exocytosis. Furthermore, Lime and its variants are advantageous for a broader range of applications including the detection of synaptic release and neuronal voltage changes.

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08/25/23 | Ultra-high density electrodes improve detection, yield, and cell type specificity of brain recordings.
Ye Z, Shelton AM, Shaker JR, Boussard J, Colonell J, Manavi S, Chen S, Windolf C, Hurwitz C, Namima T, Pedraja F, Weiss S, Raducanu B, Ness TV, Einevoll GT, Laurent G, Sawtell NB, Bair W, Pasupathy A, Mora Lopez C, Dutta B, Paninski L, Siegle JH, Koch C, Olsen SR, Harris TD, Steinmetz NA
bioRxiv. 2023 Aug 25:. doi: 10.1101/2023.08.23.554527

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 µm center-to-center spacing and low noise. This device effectively comprises an implantable voltage-sensing camera that captures a planar image of a neuron'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.

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08/24/23 | iATPSnFR2: a high dynamic range fluorescent sensor for monitoring intracellular ATP
Jonathan S. Marvin , Alexandros C. Kokotos , Mukesh Kumar , Camila Pulido , Ariana N. Tkachuk , Jocelyn Shuxin Yao , Timothy A. Brown , Timothy A. Ryan
bioRxiv. 2023 Aug 24:. doi: 10.1101/2023.08.24.554624

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.

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08/23/23 | Brain wiring determinants uncovered by integrating connectomes and transcriptomes.
Yoo J, Dombrovski M, Mirshahidi P, Nern A, LoCascio SA, Zipursky SL, Kurmangaliyev YZ
Current Biology. 2023 Aug 23;33(18):3998-3998. doi: 10.1016/j.cub.2023.08.020

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.

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08/24/23 | Theta oscillations as a substrate for medial prefrontal-hippocampal assembly interactions.
Nardin M, Kaefer K, Stella F, Csicsvari J
Cell Reports. 2023 Aug 24;42(9):113015. doi: 10.1016/j.celrep.2023.113015

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.

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08/21/23 | Hydrophobic interactions dominate the recognition of a KRAS G12V neoantigen.
Wright KM, DiNapoli SR, Miller MS, Aitana Azurmendi P, Zhao X, Yu Z, Chakrabarti M, Shi W, Douglass J, Hwang MS, Hsiue EH, Mog BJ, Pearlman AH, Paul S, Konig MF, Pardoll DM, Bettegowda C, Papadopoulos N, Kinzler KW, Vogelstein B, Zhou S, Gabelli SB
Nature Communications. 2023 Aug 21;14(1):5063. doi: 10.1038/s41467-023-40821-w

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.

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