MetaCell analysis of single cell RNA-seq data using k-NN graph partitions, bioRxiv, 2018-10-09
ABSTRACTSingle cell RNA-seq (scRNA-seq) has become the method of choice for analyzing mRNA distributions in heterogeneous cell populations. scRNA-seq only partially samples the cells in a tissue and the RNA in each cell, resulting in sparse data that challenge analysis. We develop a methodology that addresses scRNA-seq’s sparsity through partitioning the data into metacells disjoint, homogenous and highly compact groups of cells, each exhibiting only sampling variance. Metacells constitute local building blocks for clustering and quantitative analysis of gene expression, while not enforcing any global structure on the data, thereby maintaining statistical control and minimizing biases. We illustrate the MetaCell framework by re-analyzing cell type and transcriptional gradients in peripheral blood and whole organism scRNA-seq maps. Our algorithms are implemented in the new MetaCell RC++ software package.
biorxiv bioinformatics 0-100-users 2018Cytoskeletal tension actively sustains the migratory T cell synaptic contact, bioRxiv, 2018-10-08
SummaryWhen migratory T cells encounter antigen presenting cells (APCs), they arrest and form radially symmetric, stable intercellular junctions termed immunological synapses which facilitate exchange of crucial biochemical information and are critical for T cell immunity. While the cellular processes underlying synapse formation have been well-characterized, those that maintain the symmetry, and thereby the stability of the synapse remain unknown. Here we identify an antigen-triggered mechanism that actively promotes T cell synapse symmetry by generating cytoskeletal tension in the plane of the synapse through focal nucleation of actin via Wiskott -Aldrich syndrome Protein (WASP), and contraction of the resultant actin filaments by myosin II. Following T cell activation, WASP is degraded, leading to cytoskeletal rearrangement and tension decay, which result in synapse breaking. Thus, our study identifies and characterizes a mechanical program within otherwise highly motile T cells that sustains the symmetry and stability of the T cell-APC synaptic contact.
biorxiv cell-biology 0-100-users 2018Combining Gene Ontology with Deep Neural Networks to Enhance the Clustering of Single Cell RNA-Seq Data, bioRxiv, 2018-10-07
AbstractBackgroundSingle cell RNA sequencing (scRNA-seq) is applied to assay the individual transcriptomes of large numbers of cells. The gene expression at single-cell level provides an opportunity for better understanding of cell function and new discoveries in biomedical areas. To ensure that the single-cell based gene expression data are interpreted appropriately, it is crucial to develop new computational methods.ResultsIn this article, we try to construct the structure of neural networks based on the prior knowledge of Gene Ontology (GO). By integrating GO with both unsupervised and supervised models, two novel methods are proposed, named GOAE (Gene Ontology AutoEncoder) and GONN (Gene Ontology Neural Network) respectively, for clustering of scRNA-seq data.ConclusionsThe evaluation results show that the proposed models outperform some state-of-the-art approaches. Furthermore, incorporating with GO, we provide an opportunity to interpret the underlying biological mechanism behind the neural network-based model.
biorxiv bioinformatics 0-100-users 2018Structural variants identified by Oxford Nanopore PromethION sequencing of the human genome, bioRxiv, 2018-10-04
AbstractWe sequenced the Yoruban NA19240 genome on the long read sequencing platform Oxford Nanopore PromethION for benchmarking and evaluation of recently published aligners and structural variant calling tools. In this work, we determined the precision and recall, present high confidence and high sensitivity call sets of variants and discuss optimal parameters. The aligner Minimap2 and structural variant caller Sniffles are both the most accurate and the most computationally efficient tools in our study. We describe our scalable workflow for identification, annotation, and characterization of tens of thousands of structural variants from long read genome sequencing of an individual or population. By discussing the results of this genome we provide an approximation of what can be expected in future long read sequencing studies aiming for structural variant identification.
biorxiv bioinformatics 0-100-users 2018GeneWeld a method for efficient targeted integration directed by short homology, bioRxiv, 2018-10-03
AbstractChoices for genome engineering and integration involve high efficiency with little or no target specificity or high specificity with low activity. Here, we describe a targeted integration strategy, called GeneWeld, and a vector series for gene tagging, pGTag (plasmids for Gene Tagging), which promote highly efficient and precise targeted integration in zebrafish embryos, pig fibroblasts, and human cells utilizing the CRISPRCas9 system. Our work demonstrates that in vivo targeting of a genomic locus of interest with CRISPRCas9 and a donor vector containing as little as 24 to 48 base pairs of homology directs precise and efficient knock-in when the homology arms are exposed with a double strand break in vivo. Our results suggest that the length of homology is not important in the design of knock-in vectors but rather how the homology is presented to a double strand break in the genome. Given our results targeting multiple loci in different species, we expect the accompanying protocols, vectors, and web interface for homology arm design to help streamline gene targeting and applications in CRISPR and TALEN compatible systems.
biorxiv genetics 0-100-users 2018Single-cell in situ transcriptomic map of astrocyte cortical layer diversity, bioRxiv, 2018-10-03
AbstractDuring organogenesis, patterns and gradients of gene expression underlie organization and diversified cell specification to generate complex tissue architecture. While the cerebral cortex is organized into six excitatory neuronal layers, it is unclear whether glial cells are diversified to mimic neuronal laminae or show distinct layering. To determine the molecular architecture of the mammalian cortex, we developed a high content pipeline that can quantify single-cell gene expression in situ. The Large-area Spatial Transcriptomic (LaST) map confirmed expected cortical neuron layer organization and also revealed a novel neuronal identity signature. Screening 46 candidate genes for astrocyte diversity across the cortex, we identified grey matter superficial, mid and deep astrocyte identities in gradient layer patterns that were distinct from neurons. Astrocyte layers formed in early postnatal cortex and mostly persisted in adult mouse and human cortex. Mutations that shifted neuronal post-mitotic identity or organization were sufficient to alter glial layering, indicating an instructive role for neuronal cues. In normal mouse cortex, astrocyte layer patterns showed area diversity between functionally distinct cortical regions. These findings indicate that excitatory neurons and astrocytes cells are organized into distinct lineage-associated laminae, which give rise to higher order neuroglial complexity of cortical architecture.
biorxiv neuroscience 0-100-users 2018