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2432 Janelia Publications

Showing 1501-1510 of 2432 results
02/06/16 | Dynamin regulates metaphase furrow formation and plasma membrane compartmentalization in the syncytial Drosophila embryo.
Rikhy R, Mavrakis M, Lippincott-Schwartz J
Biology open. 2015;4(3):301-11. doi: 10.1242/bio.20149936

The successive nuclear division cycles in the syncytial Drosophila embryo are accompanied by ingression and regression of plasma membrane furrows, which surround individual nuclei at the embryo periphery, playing a central role in embryo compartmentalization prior to cellularization. Here, we demonstrate that cell cycle changes in dynamin localization and activity at the plasma membrane (PM) regulate metaphase furrow formation and PM organization in the syncytial embryo. Dynamin was localized on short PM furrows during interphase, mediating endocytosis of PM components. Dynamin redistributed off ingressed PM furrows in metaphase, correlating with stabilized PM components and the associated actin regulatory machinery on long furrows. Acute inhibition of dynamin in the temperature sensitive shibire mutant embryo resulted in morphogenetic consequences in the syncytial division cycle. These included inhibition of metaphase furrow ingression, randomization of proteins normally polarized to intercap PM and disruption of the diffusion barrier separating PM domains above nuclei. Based on these findings, we propose that cell cycle changes in dynamin orchestrate recruitment of actin regulatory machinery for PM furrow dynamics during the early mitotic cycles in the Drosophila embryo.

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02/04/16 | Discovering Neuronal Cell Types and Their Gene Expression Profiles Using a Spatial Point Process Mixture Model
Furong Huang , Animashree Anandkumar , Christian Borgs , Jennifer Chayes , Ernest Fraenkel , Michael Hawrylycz , Ed Lein , Alessandro Ingrosso , Srinivas Turaga

Cataloging the neuronal cell types that comprise circuitry of individual brain regions is a major goal of modern neuroscience and the BRAIN initiative. Single-cell RNA sequencing can now be used to measure the gene expression profiles of individual neurons and to categorize neurons based on their gene expression profiles. While the single-cell techniques are extremely powerful and hold great promise, they are currently still labor intensive, have a high cost per cell, and, most importantly, do not provide information on spatial distribution of cell types in specific regions of the brain. We propose a complementary approach that uses computational methods to infer the cell types and their gene expression profiles through analysis of brain-wide single-cell resolution in situ hybridization (ISH) imagery contained in the Allen Brain Atlas (ABA). We measure the spatial distribution of neurons labeled in the ISH image for each gene and model it as a spatial point process mixture, whose mixture weights are given by the cell types which express that gene. By fitting a point process mixture model jointly to the ISH images, we infer both the spatial point process distribution for each cell type and their gene expression profile. We validate our predictions of cell type-specific gene expression profiles using single cell RNA sequencing data, recently published for the mouse somatosensory cortex. Jointly with the gene expression profiles, cell features such as cell size, orientation, intensity and local density level are inferred per cell type.

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02/03/16 | Intracellular and extracellular forces drive primary cilia movement.
Battle C, Ott CM, Burnette DT, Lippincott-Schwartz J, Schmidt CF
Proceedings of the National Academy of Sciences of the United States of America. 2015 Feb 3;112(5):1410-5. doi: 10.1073/pnas.1421845112

Primary cilia are ubiquitous, microtubule-based organelles that play diverse roles in sensory transduction in many eukaryotic cells. They interrogate the cellular environment through chemosensing, osmosensing, and mechanosensing using receptors and ion channels in the ciliary membrane. Little is known about the mechanical and structural properties of the cilium and how these properties contribute to ciliary perception. We probed the mechanical responses of primary cilia from kidney epithelial cells [Madin-Darby canine kidney-II (MDCK-II)], which sense fluid flow in renal ducts. We found that, on manipulation with an optical trap, cilia deflect by bending along their length and pivoting around an effective hinge located below the basal body. The calculated bending rigidity indicates weak microtubule doublet coupling. Primary cilia of MDCK cells lack interdoublet dynein motors. Nevertheless, we found that the organelles display active motility. 3D tracking showed correlated fluctuations of the cilium and basal body. These angular movements seemed random but were dependent on ATP and cytoplasmic myosin-II in the cell cortex. We conclude that force generation by the actin cytoskeleton surrounding the basal body results in active ciliary movement. We speculate that actin-driven ciliary movement might tune and calibrate ciliary sensory functions.

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02/03/16 | Neural circuits underlying visually evoked escapes in larval zebrafish.
Dunn TW, Gebhardt C, Naumann EA, Riegler C, Ahrens MB, Engert F, Del Bene F
Neuron. 2016 Feb 3;89(3):613-628. doi: 10.1016/j.neuron.2015.12.021

Escape behaviors deliver organisms away from imminent catastrophe. Here, we characterize behavioral responses of freely swimming larval zebrafish to looming visual stimuli simulating predators. We report that the visual system alone can recruit lateralized, rapid escape motor programs, similar to those elicited by mechanosensory modalities. Two-photon calcium imaging of retino-recipient midbrain regions isolated the optic tectum as an important center processing looming stimuli, with ensemble activity encoding the critical image size determining escape latency. Furthermore, we describe activity in retinal ganglion cell terminals and superficial inhibitory interneurons in the tectum during looming and propose a model for how temporal dynamics in tectal periventricular neurons might arise from computations between these two fundamental constituents. Finally, laser ablations of hindbrain circuitry confirmed that visual and mechanosensory modalities share the same premotor output network. We establish a circuit for the processing of aversive stimuli in the context of an innate visual behavior.

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02/02/16 | Tagmentation-based mapping (tagmap) of mobile DNA genomic insertion sites.
bioRxiv. 2016 Feb 2:. doi: 10.1101/037762

Multiple methods have been introduced over the past 30 years to identify the genomic insertion sites of transposable elements and other DNA elements that integrate into genomes. However, each of these methods suffer from limitations that can frustrate attempts to map multiple insertions in a single genome and to map insertions in genomes of high complexity that contain extensive repetitive DNA. I introduce a new method for transposon mapping that is simple to perform, can accurately map multiple insertions per genome, and generates long sequence reads that facilitate mapping to complex genomes. The method, called TagMap, for Tagmentation-based Mapping, relies on a modified Tn5 tagmentation protocol with a single tagmentation adaptor followed by PCR using primers specific to the tranposable element and the adaptor sequence. Several minor modifications to normal tagmentation reagents and protocols allow easy and rapid preparation of TagMap libraries. Short read sequencing starting from the adaptor sequence generates oriented reads that flank and are oriented toward the transposable element insertion site. The convergent orientation of adjacent reads at the insertion site allows straightforward prediction of the precise insertion site(s). A Linux shell script is provided to identify insertion sites from fastq files.

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Ji Lab
02/01/16 | Thalamus provides layer 4 of primary visual cortex with orientation- and direction-tuned inputs.
Sun W, Tan Z, Mensh BD, Ji N
Nature Neuroscience. 2016 Feb;19(2):308-15. doi: 10.1038/nn.4196

Understanding the functions of a brain region requires knowing the neural representations of its myriad inputs, local neurons and outputs. Primary visual cortex (V1) has long been thought to compute visual orientation from untuned thalamic inputs, but very few thalamic inputs have been measured in any mammal. We determined the response properties of ~28,000 thalamic boutons and ~4,000 cortical neurons in layers 1–5 of awake mouse V1. Using adaptive optics that allows accurate measurement of bouton activity deep in cortex, we found that around half of the boutons in the main thalamorecipient L4 carried orientation-tuned information and that their orientation and direction biases were also dominant in the L4 neuron population, suggesting that these neurons may inherit their selectivity from tuned thalamic inputs. Cortical neurons in all layers exhibited sharper tuning than thalamic boutons and a greater diversity of preferred orientations. Our results provide data-rich constraints for refining mechanistic models of cortical computation.

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02/01/16 | Transcriptomes of lineage-specific Drosophila neuroblasts profiled by genetic targeting and robotic sorting.
Yang C, Fu C, Sugino K, Liu Z, Ren Q, Liu L, Yao X, Lee LP, Lee T
Development (Cambridge, England). 2016 Feb 1;143(3):411-21. doi: 10.1242/dev.129163

A brain consists of numerous distinct neurons arising from a limited number of progenitors, called neuroblasts in Drosophila. Each neuroblast produces a specific neuronal lineage. To unravel the transcriptional networks that underlie the development of distinct neuroblast lineages, we marked and isolated lineage-specific neuroblasts for RNA sequencing. We labeled particular neuroblasts throughout neurogenesis by activating a conditional neuroblast driver in specific lineages using various intersection strategies. The targeted neuroblasts were efficiently recovered using a custom-built device for robotic single-cell picking. Transcriptome analysis of mushroom body, antennal lobe and type II neuroblasts compared with non-selective neuroblasts, neurons and glia revealed a rich repertoire of transcription factors expressed among neuroblasts in diverse patterns. Besides transcription factors that are likely to be pan-neuroblast, many transcription factors exist that are selectively enriched or repressed in certain neuroblasts. The unique combinations of transcription factors present in different neuroblasts may govern the diverse lineage-specific neuron fates.

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01/28/16 | Studying small brains to understand the building blocks of cognition.
Haberkern H, Jayaraman V
Current Opinion in Neurobiology. 2016 Jan 28;37:59-65. doi: 10.1016/j.conb.2016.01.007

Cognition encompasses a range of higher-order mental processes, such as attention, working memory, and model-based decision-making. These processes are thought to involve the dynamic interaction of multiple central brain regions. A mechanistic understanding of such computations requires not only monitoring and manipulating specific neural populations during behavior, but also knowing the connectivity of the underlying circuitry. These goals are experimentally challenging in mammals, but are feasible in numerically simpler insect brains. In Drosophila melanogaster in particular, genetic tools enable precisely targeted physiology and optogenetics in actively behaving animals. In this article we discuss how these advantages are increasingly being leveraged to study abstract neural representations and sensorimotor computations that may be relevant for cognition in both insects and mammals.

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01/28/16 | Visualization and quantification for interactive analysis of neural connectivity in Drosophila.
Swoboda N, Moosburner J, Bruckner S, Yu J, Dickson BJ, Bühler K
Computer Graphics Forum. 2016 Jan 28:. doi: 10.1111/cgf.12792

Neurobiologists investigate the brain of the common fruit fly Drosophila melanogaster to discover neural circuits and link them to complex behaviour. Formulating new hypotheses about connectivity requires potential connectivity information between individual neurons, indicated by overlaps of arborizations of two or more neurons. As the number of higher order overlaps (i.e. overlaps of three or more arborizations) increases exponentially with the number of neurons under investigation, visualization is impeded by clutter and quantification becomes a burden. Existing solutions are restricted to visual or quantitative analysis of pairwise overlaps, as they rely on precomputed overlap data. We present a novel tool that complements existing methods for potential connectivity exploration by providing for the first time the possibility to compute and visualize higher order arborization overlaps on the fly and to interactively explore this information in both its spatial anatomical context and on a quantitative level. Qualitative evaluation by neuroscientists and non-experts demonstrated the utility and usability of the tool.

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01/25/16 | Real-time three-dimensional cell segmentation in large-scale microscopy data of developing embryos.
Stegmaier J, Amat F, Lemon WC, McDole K, Wan Y, Teodoro G, Mikut R, Keller PJ
Developmental Cell. 2016 Jan 25;36(2):225-40. doi: 10.1016/j.devcel.2015.12.028

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 55–330 times faster and 2–5 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.

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