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 2018Single-cell virus sequencing of influenza infections that trigger innate immunity, bioRxiv, 2018-10-07
SUMMARYThe outcome of viral infection is extremely heterogeneous, with infected cells only sometimes activating innate immunity. Here we develop a new approach to assess how the genetic variation inherent in viral populations contributes to this heterogeneity. We do this by determining both the transcriptome and full-length sequences of all viral genes in single influenza-infected cells. Most cells are infected by virions with defects such as amino-acid mutations, internal deletions, or failure to express a gene. We identify instances of each type of defect that increase the likelihood that a cell activates an innate-immune response. However, immune activation remains stochastic in cells infected by virions with these defects, and sometimes occurs even when a cell is infected by a virion that expresses unmutated copies of all genes. Our work shows that viral genetic variation substantially contributes to but does not fully explain the heterogeneity in single influenza-infected cells.
biorxiv microbiology 100-200-users 2018Structure of transcribing RNA polymerase II-nucleosome complex, bioRxiv, 2018-10-07
Transcription of eukaryotic protein-coding genes requires passage of RNA polymerase II (Pol II) through chromatin. Pol II passage is impaired by nucleosomes and requires elongation factors that help Pol II to efficiently overcome the nucleosomal barrier1-4. How the Pol II machinery transcribes through a nucleosome remains unclear because structural studies have been limited to Pol II elongation complexes formed on DNA templates lacking nucleosomes5. Here we report the cryo-electron microscopy (cryo-EM) structure of transcribing Pol II from the yeast Saccharomyces cerevisiae engaged with a downstream nucleosome core particle (NCP) at an overall resolution of 4.4 Å with resolutions ranging from 4-6 Å in Pol II and 6-8 Å in the NCP. Pol II and the NCP adopt a defined orientation that could not be predicted from modelling. Pol II contacts DNA of the incoming NCP on both sides of the nucleosomal dyad with its domains ‘clamp head’ and ‘lobe’. Comparison of the Pol II-NCP structure to known structures of Pol II complexes reveals that the elongation factors TFIIS, DSIF, NELF, PAF1 complex, and SPT6 can be accommodated on the Pol II surface in the presence of the oriented nucleosome. Further structural comparisons show that the chromatin remodelling enzyme Chd1, which is also required for efficient Pol II passage6,7, could bind the oriented nucleosome with its motor domain. The DNA-binding region of Chd1 must however be released from DNA when Pol II approaches the nucleosome, and based on published data8,9 this is predicted to stimulate Chd1 activity and to facilitate Pol II passage. Our results provide a starting point for a mechanistic analysis of chromatin transcription.
biorxiv biochemistry 100-200-users 2018Bright and photostable chemigenetic indicators for extended in vivo voltage imaging, bioRxiv, 2018-10-06
Imaging changes in membrane potential using genetically encoded fluorescent voltage indicators (GEVIs) has great potential for monitoring neuronal activity with high spatial and temporal resolution. Brightness and photostability of fluorescent proteins and rhodopsins have limited the utility of existing GEVIs. We engineered a novel GEVI, Voltron, that utilizes bright and photostable synthetic dyes instead of protein-based fluorophores, extending the combined duration of imaging and number of neurons imaged simultaneously by more than tenfold relative to existing GEVIs. We used Voltron for in vivo voltage imaging in mice, zebrafish, and fruit flies. In mouse cortex, Voltron allowed single-trial recording of spikes and subthreshold voltage signals from dozens of neurons simultaneously, over 15 minutes of continuous imaging. In larval zebrafish, Voltron enabled the precise correlation of spike timing with behavior.
biorxiv neuroscience 200-500-users 2018