Robust manipulation of the behavior of Drosophila melanogaster by a fungal pathogen in the laboratory, bioRxiv, 2017-12-11
AbstractMany microbes induce striking behavioral changes in their animal hosts, but how they achieve this is poorly understood, especially at the molecular level. Mechanistic understanding has been largely constrained by the lack of a model system with advanced tools for molecular manipulation. We recently discovered a strain of the behavior-manipulating fungal pathogen Entomophthora muscae infecting wild Drosophila, and established methods to infect D. melanogaster in the lab. Lab-infected flies manifest the moribund behaviors characteristic of E. muscae infection hours before death, they climb upward, extend their proboscides and affix in place, then raise their wings, clearing a path for infectious spores to launch from their abdomens. We found that E. muscae invades the fly nervous system, suggesting a direct means by which the fungus could induce behavioral changes. Given the vast molecular toolkit available for D. melanogaster, we believe this new system will enable rapid progress in understanding the mechanistic basis of E. muscae’s behavioral manipulation in the fly.
biorxiv animal-behavior-and-cognition 200-500-users 2017Shared and distinct transcriptomic cell types across neocortical areas, bioRxiv, 2017-12-07
ABSTRACTNeocortex contains a multitude of cell types segregated into layers and functionally distinct regions. To investigate the diversity of cell types across the mouse neocortex, we analyzed 12,714 cells from the primary visual cortex (VISp), and 9,035 cells from the anterior lateral motor cortex (ALM) by deep single-cell RNA-sequencing (scRNA-seq), identifying 116 transcriptomic cell types. These two regions represent distant poles of the neocortex and perform distinct functions. We define 50 inhibitory transcriptomic cell types, all of which are shared across both cortical regions. In contrast, 49 of 52 excitatory transcriptomic types were found in either VISp or ALM, with only three present in both. By combining single cell RNA-seq and retrograde labeling, we demonstrate correspondence between excitatory transcriptomic types and their region-specific long-range target specificity. This study establishes a combined transcriptomic and projectional taxonomy of cortical cell types from functionally distinct regions of the mouse cortex.
biorxiv neuroscience 200-500-users 2017Exponential fluorescent amplification of individual RNAs using clampFISH probes, bioRxiv, 2017-12-06
AbstractNon-enzymatic, high-gain signal amplification methods with single-cell, single-molecule resolution are in great need. We present click-amplifying FISH (clampFISH) for the fluorescent detection of RNA that combines the specificity of oligonucleotides with bioorthogonal click chemistry in order to achieve high specificity and extremely high-gain (>400x) signal amplification. We show that clampFISH signal enables detection with low magnification microscopy and separation of cells by RNA levels via flow cytometry. Additionally, we show that the modular design of clampFISH probes enables multiplexing, that the locking mechanism prevents probe detachment in expansion microscopy, and that clampFISH works in tissue samples.
biorxiv bioengineering 200-500-users 2017High-throughput ANI Analysis of 90K Prokaryotic Genomes Reveals Clear Species Boundaries, bioRxiv, 2017-11-28
A fundamental question in microbiology is whether there is a continuum of genetic diversity among genomes or clear species boundaries prevail instead. Answering this question requires robust measurement of whole-genome relatedness among thousands of genomes and from diverge phylogenetic lineages. Whole-genome similarity metrics such as Average Nucleotide Identity (ANI) can provide the resolution needed for this task, overcoming several limitations of traditional techniques used for the same purposes. Although the number of genomes currently available may be adequate, the associated bioinformatics tools for analysis are lagging behind these developments and cannot scale to large datasets. Here, we present a new method, FastANI, to compute ANI using alignment-free approximate sequence mapping. Our analyses demonstrate that FastANI produces an accurate ANI estimate and is up to three orders of magnitude faster when compared to an alignment (e.g., BLAST)-based approach. We leverage FastANI to compute pairwise ANI values among all prokaryotic genomes available in the NCBI database. Our results reveal a clear genetic discontinuity among the database genomes, with 99.8% of the total 8 billion genome pairs analyzed showing either >95% intra-species ANI or <83% inter-species ANI values. We further show that this discontinuity is recovered with or without the most frequently represented species in the database and is robust to historic additions in the public genome databases. Therefore, 95% ANI represents an accurate threshold for demarcating almost all currently named prokaryotic species, and wide species boundaries may exist for prokaryotes.
biorxiv bioinformatics 200-500-users 2017A suite of transgenic driver and reporter mouse lines with enhanced brain cell type targeting and functionality, bioRxiv, 2017-11-26
SUMMARYModern genetic approaches are powerful in providing access to diverse types of neurons within the mammalian brain and greatly facilitating the study of their function. We here report a large set of driver and reporter transgenic mouse lines, including 23 new driver lines targeting a variety of cortical and subcortical cell populations and 26 new reporter lines expressing an array of molecular tools. In particular, we describe the TIGRE2.0 transgenic platform and introduce Cre-dependent reporter lines that enable optical physiology, optogenetics, and sparse labeling of genetically-defined cell populations. TIGRE2.0 reporters broke the barrier in transgene expression level of single-copy targeted-insertion transgenesis in a wide range of neuronal types, along with additional advantage of a simplified breeding strategy compared to our first-generation TIGRE lines. These novel transgenic lines greatly expand the repertoire of high-precision genetic tools available to effectively identify, monitor, and manipulate distinct cell types in the mouse brain.
biorxiv neuroscience 200-500-users 2017Common risk variants identified in autism spectrum disorder, bioRxiv, 2017-11-26
AbstractAutism spectrum disorder (ASD) is a highly heritable and heterogeneous group of neurodevelopmental phenotypes diagnosed in more than 1% of children. Common genetic variants contribute substantially to ASD susceptibility, but to date no individual variants have been robustly associated with ASD. With a marked sample size increase from a unique Danish population resource, we report a genome-wide association meta-analysis of 18,381 ASD cases and 27,969 controls that identifies five genome-wide significant loci. Leveraging GWAS results from three phenotypes with significantly overlapping genetic architectures (schizophrenia, major depression, and educational attainment), seven additional loci shared with other traits are identified at equally strict significance levels. Dissecting the polygenic architecture we find both quantitative and qualitative polygenic heterogeneity across ASD subtypes, in contrast to what is typically seen in other complex disorders. These results highlight biological insights, particularly relating to neuronal function and corticogenesis and establish that GWAS performed at scale will be much more productive in the near term in ASD, just as it has been in a broad range of important psychiatric and diverse medical phenotypes.
biorxiv genetics 200-500-users 2017