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1693 Janelia Publications

Showing 1-10 of 1693 results
01/14/20 | Improved HaloTag Ligand Enables BRET Imaging With NanoLuc
Thirukkumaran OM, Wang C, Asouzu NJ, Fron E, Rocha S, Hofkens J, Lavis LD, Mizuno H
Frontiers in Chemistry. 2020 Jan 14;7:. doi: 10.3389/fchem.2019.0093810.3389/fchem.2019.00938.s001
01/13/20 | When does midbrain dopamine activity exert its effects on behavior?
Coddington LT
Nature Neuroscience. 2020 Jan 13:. doi: 10.1038/s41593-019-0577-y
01/11/20 | Tiled reconstruction improves structured illumination microscopy.
Hoffman DP, Betzig E
bioRxiv. 2020 Jan 11:. doi: 10.1101/2020.01.06.895318

Structured illumination microscopy (SIM) is widely used for fast, long-term, live-cell super-resolution imaging. However, SIM images can contain substantial artifacts if the sample does not conform to the underlying assumptions of the reconstruction algorithm. Here we describe a simple, easy to implement, process that can be combined with any reconstruction algorithm to alleviate many common SIM reconstruction artifacts and briefly discuss possible extensions.

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01/09/20 | Bright and tunable far-red chemigenetic indicators.
Deo C, Abdelfattah AS, Bhargava HK, Berro AJ, Falco N, Moeyaert B, Chupanova M, Lavis LD, Schreiter ER
bioRxiv. 2020 Jan 9:
01/08/20 | Comprehensive transcriptome analysis of cochlear spiral ganglion neurons at multiple ages.
Li C, Li X, Bi Z, Sugino K, Wang G, Zhu T, Liu Z
eLife. 2020 Jan 08;9:. doi: 10.7554/eLife.50491

Inner ear cochlear spiral ganglion neurons (SGNs) transmit auditory information to the brainstem. Recent single cell RNA-Seq studies have revealed heterogeneities within SGNs. Nonetheless, much remains unknown about the transcriptome of SGNs, especially which genes are specifically expressed in SGNs. To address these questions we needed a deeper and broader gene coverage than that in previous studies. We performed bulk RNA-Seq on mouse SGNs at five ages, and on two reference cell types (hair cells and glia). Their transcriptome comparison identified genes previously unknown to be specifically expressed in SGNs. To validate our dataset and provide useful genetic tools for this research field, we generated two knockin mouse strains: and . Our comprehensive analysis confirmed the SGN-selective expression of the candidate genes, testifying to the quality of our transcriptome data. These two mouse strains can be used to temporally label SGNs or to sort them.

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02/01/20 | Tissue clearing and its applications in neuroscience
Ueda HR, Ertürk A, Chung K, Gradinaru V, Chédotal A, Tomancak P, Keller PJ
Nature Reviews Neuroscience. 2020 Feb 1:. doi: 10.1038/s41583-019-0250-1

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.

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01/07/20 | Structure of the vesicular stomatitis virus L protein in complex with Its phosphoprotein cofactor.
Jenni S, Bloyet L, Diaz-Avalos R, Liang B, Whelan SP, Grigorieff N, Harrison SC
Cell Reports. 2020 Jan 07;30(1):53-60.e5. doi: 10.1016/j.celrep.2019.12.024

The large (L) proteins of non-segmented, negative-strand RNA viruses are multifunctional enzymes that produce capped, methylated, and polyadenylated mRNA and replicate the viral genome. A phosphoprotein (P), required for efficient RNA-dependent RNA polymerization from the viral ribonucleoprotein (RNP) template, regulates the function and conformation of the L protein. We report the structure of vesicular stomatitis virus L in complex with its P cofactor determined by electron cryomicroscopy at 3.0 Å resolution, enabling us to visualize bound segments of P. The contacts of three P segments with multiple L domains show how P induces a closed, compact, initiation-competent conformation. Binding of P to L positions its N-terminal domain adjacent to a putative RNA exit channel for efficient encapsidation of newly synthesized genomes with the nucleoprotein and orients its C-terminal domain to interact with an RNP template. The model shows that a conserved tryptophan in the priming loop can support the initiating 5' nucleotide.

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01/07/20 | Trans-endocytosis of intact IL-15Rα-IL-15 complex from presenting cells into NK cells favors signaling for proliferation.
Anton OM, Peterson ME, Hollander MJ, Dorward DW, Arora G, Traba J, Rajagopalan S, Snapp EL, Garcia KC, Waldmann TA, Long EO
Proceedings of the National Academy of Sciences of the United States of America. 2020 Jan 07;117(1):522-531. doi: 10.1073/pnas.1911678117

Interleukin 15 (IL-15) is an essential cytokine for the survival and proliferation of natural killer (NK) cells. IL-15 activates signaling by the β and common γ (γ) chain heterodimer of the IL-2 receptor through -presentation by cells expressing IL-15 bound to the α chain of the IL-15 receptor (IL-15Rα). We show here that membrane-associated IL-15Rα-IL-15 complexes are transferred from presenting cells to NK cells through -endocytosis and contribute to the phosphorylation of ribosomal protein S6 and NK cell proliferation. NK cell interaction with soluble or surface-bound IL-15Rα-IL-15 complex resulted in Stat5 phosphorylation and NK cell survival at a concentration or density of the complex much lower than required to stimulate S6 phosphorylation. Despite this efficient response, Stat5 phosphorylation was reduced after inhibition of metalloprotease-induced IL-15Rα-IL-15 shedding from -presenting cells, whereas S6 phosphorylation was unaffected. Conversely, inhibition of -endocytosis by silencing of the small GTPase TC21 or expression of a dominant-negative TC21 reduced S6 phosphorylation but not Stat5 phosphorylation. Thus, -endocytosis of membrane-associated IL-15Rα-IL-15 provides a mode of regulating NK cells that is not afforded to IL-2 and is distinct from activation by soluble IL-15. These results may explain the strict IL-15 dependence of NK cells and illustrate how the cellular compartment in which receptor-ligand interaction occurs can influence functional outcome.

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01/06/20 | Nanoscale subcellular architecture revealed by multicolor three-dimensional salvaged fluorescence imaging.
Zhang Y, Schroeder LK, Lessard MD, Kidd P, Chung J, Song Y, Benedetti L, Li Y, Ries J, Grimm JB, Lavis LD, De Camilli P, Rothman JE, Baddeley D, Bewersdorf J
Nature Methods. 2020 Jan 06:. doi: 10.1038/s41592-019-0676-4

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 'salvaged fluorescence'. 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.

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01/03/20 | The neuropeptide Drosulfakinin regulates social isolation-induced aggression in Drosophila.
Agrawal P, Kao D, Chung P, Looger LL
Journal of Experimental Biology. 2020 Jan 03:. doi: 10.1242/jeb.207407

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.

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