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single cell RNA sequencing

Simone Mayer, Jiadong Chen, Dmitry Velmeshev, Andreas Mayer, Ugomma C Eze, Aparna Bhaduri, Carlos E Cunha, Diane Jung, Arpana Arjun, Emmy Li, Beatriz Alvarado, Shaohui Wang, Nils Lovegren, Michael L Gonzales, Lukasz Szpankowski, Anne Leyrat, Jay A A West, Georgia Panagiotakos, Arturo Alvarez-Buylla, Mercedes F Paredes, Tomasz J Nowakowski, Alex A Pollen, Arnold R Kriegstein
In the developing human neocortex, progenitor cells generate diverse cell types prenatally. Progenitor cells and newborn neurons respond to signaling cues, including neurotransmitters. While single-cell RNA sequencing has revealed cellular diversity, physiological heterogeneity has yet to be mapped onto these developing and diverse cell types. By combining measurements of intracellular Ca2+ elevations in response to neurotransmitter receptor agonists and RNA sequencing of the same single cells, we show that Ca2+ responses are cell-type-specific and change dynamically with lineage progression...
February 7, 2019: Neuron
Renée R C E Schreurs, Martin E Baumdick, Adrian F Sagebiel, Max Kaufmann, Michal Mokry, Paul L Klarenbeek, Nicola Schaltenberg, Fenja L Steinert, Jorik M van Rijn, Agata Drewniak, Sarah-May M L The, Roel Bakx, Joep P M Derikx, Niek de Vries, Willemijn E Corpeleijn, Steven T Pals, Nicola Gagliani, Manuel A Friese, Sabine Middendorp, Edward E S Nieuwenhuis, Konrad Reinshagen, Teunis B H Geijtenbeek, Johannes B van Goudoever, Madeleine J Bunders
Although the fetal immune system is considered tolerogenic, preterm infants can suffer from severe intestinal inflammation, including necrotizing enterocolitis (NEC). Here, we demonstrate that human fetal intestines predominantly contain tumor necrosis factor-α (TNF-α)+ CD4+ CD69+ T effector memory (Tem) cells. Single-cell RNA sequencing of fetal intestinal CD4+ T cells showed a T helper 1 phenotype and expression of genes mediating epithelial growth and cell cycling. Organoid co-cultures revealed a dose-dependent, TNF-α-mediated effect of fetal intestinal CD4+ T cells on intestinal stem cell (ISC) development, in which low T cell numbers supported epithelial development, whereas high numbers abrogated ISC proliferation...
February 5, 2019: Immunity
Lauren E Higdon, Steven Schaffert, Purvesh Khatri, Jonathan S Maltzman
Recently developed single-cell profiling technologies hold promise to provide new insights including analysis of population heterogeneity and linkage of antigen receptors with gene expression. These technologies produce complex data sets that require knowledge of bioinformatics for appropriate analysis. In this minireview, we discuss several single-cell immune profiling technologies for gene and protein expression, including cytometry by time-of-flight, RNA sequencing, and antigen receptor sequencing, as well as key considerations for analysis that apply to each...
February 15, 2019: American Journal of Transplantation
Chikako Ragan, Gregory J Goodall, Nikolay E Shirokikh, Thomas Preiss
Circular RNAs (circRNAs) exhibit unique properties due to their covalently closed nature. Models of circRNAs synthesis and function are emerging but much remains undefined about this surprisingly prevalent class of RNA. Here, we identified exonic circRNAs from human and mouse RNA-sequencing datasets, documenting multiple new examples. Addressing function, we found that many circRNAs co-sediment with ribosomes, indicative of their translation potential. By contrast, circRNAs with potential to act as microRNA sponges were scarce, with some support for a collective sponge function by groups of circRNAs...
February 14, 2019: Scientific Reports
Alba Rodriguez-Meira, Gemma Buck, Sally-Ann Clark, Benjamin J Povinelli, Veronica Alcolea, Eleni Louka, Simon McGowan, Angela Hamblin, Nikolaos Sousos, Nikolaos Barkas, Alice Giustacchini, Bethan Psaila, Sten Eirik W Jacobsen, Supat Thongjuea, Adam J Mead
Single-cell RNA sequencing (scRNA-seq) has emerged as a powerful tool for resolving transcriptional heterogeneity. However, its application to studying cancerous tissues is currently hampered by the lack of coverage across key mutation hotspots in the vast majority of cells; this lack of coverage prevents the correlation of genetic and transcriptional readouts from the same single cell. To overcome this, we developed TARGET-seq, a method for the high-sensitivity detection of multiple mutations within single cells from both genomic and coding DNA, in parallel with unbiased whole-transcriptome analysis...
February 11, 2019: Molecular Cell
Verena Haage, Marcus Semtner, Ramon Oliveira Vidal, Daniel Perez Hernandez, Winnie W Pong, Zhihong Chen, Dolores Hambardzumyan, Vincent Magrini, Amy Ly, Jason Walker, Elaine Mardis, Philipp Mertins, Sascha Sauer, Helmut Kettenmann, David H Gutmann
Monocytes/macrophages have begun to emerge as key cellular modulators of brain homeostasis and central nervous system (CNS) disease. In the healthy brain, resident microglia are the predominant macrophage cell population; however, under conditions of blood-brain barrier leakage, peripheral monocytes/macrophages can infiltrate the brain and participate in CNS disease pathogenesis. Distinguishing these two populations is often challenging, owing to a paucity of universally accepted and reliable markers. To identify discriminatory marker sets for microglia and peripheral monocytes/macrophages, we employed a large meta-analytic approach using five published murine transcriptional datasets...
February 14, 2019: Acta Neuropathologica Communications
Yoon Ha Choi, Jong Kyoung Kim
Cell-to-cell variability in gene expression exists even in a homogeneous population of cells. Dissecting such cellular heterogeneity within a biological system is a prerequisite for understanding how a biological system is developed, homeostatically regulated, and responds to external perturbations. Single-cell RNA sequencing (scRNA-seq) allows the quantitative and unbiased characterization of cellular heterogeneity by providing genome-wide molecular profiles from tens of thousands of individual cells. A major question in analyzing scRNA-seq data is how to account for the observed cell-to-cell variability...
February 12, 2019: Molecules and Cells
Szu-Hsien Wu, Ji-Hyun Lee, Bon-Kyoung Koo
Tracking the fate of individual cells and their progeny through lineage tracing has been widely used to investigate various biological processes including embryonic development, homeostatic tissue turnover, and stem cell function in regeneration and disease. Conventional lineage tracing involves the marking of cells either with dyes or nucleoside analogues or genetic marking with fluorescent and/or colorimetric protein reporters. Both are imaging-based approaches that have played a crucial role in the field of developmental biology as well as adult stem cell biology...
February 13, 2019: Molecules and Cells
Brandon Monier, Adam McDermaid, Cankun Wang, Jing Zhao, Allison Miller, Anne Fennell, Qin Ma
Next-Generation Sequencing has made available substantial amounts of large-scale Omics data, providing unprecedented opportunities to understand complex biological systems. Specifically, the value of RNA-Sequencing (RNA-Seq) data has been confirmed in inferring how gene regulatory systems will respond under various conditions (bulk data) or cell types (single-cell data). RNA-Seq can generate genome-scale gene expression profiles that can be further analyzed using correlation analysis, co-expression analysis, clustering, differential gene expression (DGE), among many other studies...
February 14, 2019: PLoS Computational Biology
Ludan Zhao, Yinhua Jin, Katie Donahue, Margaret Tsui, Matt Fish, Catriona Y Logan, Bruce Wang, Roel Nusse
In the liver, Wnt/β-catenin signaling is involved in regulating zonation and hepatocyte proliferation during homeostasis. We have examined Wnt gene expression and signaling after injury and we show by in situ hybridization that Wnts are activated by acute carbon tetrachloride (CCl4 ) toxicity. Following injury, peri-injury hepatocytes become Wnt-responsive, expressing the Wnt target gene Axin2. Lineage tracing of peri-injury Axin2+ hepatocytes shows that during recovery, the injured parenchyma becomes repopulated and repaired by Axin2+ descendants...
February 14, 2019: Hepatology: Official Journal of the American Association for the Study of Liver Diseases
Aanchal Mongia, Debarka Sengupta, Angshul Majumdar
Motivation: Single-cell RNA sequencing has been proved to be revolutionary for its potential of zooming into complex biological systems. Genome-wide expression analysis at single-cell resolution provides a window into dynamics of cellular phenotypes. This facilitates the characterization of transcriptional heterogeneity in normal and diseased tissues under various conditions. It also sheds light on the development or emergence of specific cell populations and phenotypes. However, owing to the paucity of input RNA, a typical single cell RNA sequencing data features a high number of dropout events where transcripts fail to get amplified...
2019: Frontiers in Genetics
Anil Dangi, Shuangjin Yu, Xunrong Luo
Newly emerging technologies are rapidly changing conventional approaches to organ transplantation. In the modern era, the key challenges to transplantation include (1) how to best individualize and possibly eliminate the need for life-long immunosuppression and (2) how to expand the donor pool suitable for human transplantation. This article aims to provide readers with an updated review of three new technologies that address these challenges. First, single-cell RNA sequencing technology is rapidly evolving and has recently been employed in settings related to transplantation...
February 13, 2019: Cellular & Molecular Immunology
Timea Marton, Adeline Feri, Pierre-Henri Commere, Corinne Maufrais, Christophe d'Enfert, Melanie Legrand
The heterozygous diploid genome of Candida albicans is highly plastic, with frequent loss of heterozygosity (LOH) events. In the SC5314 laboratory strain, while LOH events are ubiquitous, a chromosome homozygosis bias is observed for certain chromosomes, whereby only one of the two homologs can occur in the homozygous state. This suggests the occurrence of recessive lethal allele(s) (RLA) preventing large-scale LOH events on these chromosomes from being stably maintained. To verify the presence of an RLA on chromosome 7 (Chr7), we utilized a system that allows (i) DNA double-strand break (DSB) induction on Chr7 by the I- Sce I endonuclease and (ii) detection of the resulting long-range homozygosis...
February 13, 2019: MSphere
Yueli Cui, Yuxuan Zheng, Xixi Liu, Liying Yan, Xiaoying Fan, Jun Yong, Yuqiong Hu, Ji Dong, Qingqing Li, Xinglong Wu, Shuai Gao, Jingyun Li, Lu Wen, Jie Qiao, Fuchou Tang
The heart is the central organ of the circulatory system, and its proper development is vital for maintaining human life. Here, we used single-cell RNA sequencing to profile the gene expression landscapes of ∼4,000 cardiac cells from human embryos and identified four major types of cells: cardiomyocytes (CMs), cardiac fibroblasts, endothelial cells (ECs), and valvar interstitial cells (VICs). Atrial and ventricular CMs acquired distinct features early in heart development. Furthermore, both CMs and fibroblasts show stepwise changes in gene expression...
February 12, 2019: Cell Reports
Ida Lindeman, Michael J T Stubbington
In this chapter, we describe TraCeR and BraCeR, our computational tools for reconstruction of paired full-length antigen receptor sequences and clonality inference from single-cell RNA-seq (scRNA-seq) data. In brief, TraCeR reconstructs T-cell receptor (TCR) sequences from scRNA-seq data by extracting sequencing reads derived from TCRs by aligning the reads from each cell against synthetic TCR sequences. TCR-derived reads are then assembled into full-length recombined TCR sequences. BraCeR builds on the TraCeR pipeline and accounts for somatic hypermutations (SHM) and isotype switching...
2019: Methods in Molecular Biology
Yuanhua Huang, Guido Sanguinetti
Single-cell RNA-seq (scRNA-seq) provides a comprehensive measurement of stochasticity in transcription, but the limitations of the technology have prevented its application to dissect variability in RNA processing events such as splicing. In this chapter, we review the challenges in splicing isoform quantification in scRNA-seq data and discuss BRIE (Bayesian regression for isoform estimation), a recently proposed Bayesian hierarchical model which resolves these problems by learning an informative prior distribution from sequence features...
2019: Methods in Molecular Biology
Meichen Dong, Yuchao Jiang
Allele-specific expression is traditionally studied by bulk RNA sequencing, which measures average gene expression across cells. Single-cell RNA sequencing (scRNA-seq) allows the comparison of expression distribution between the two alleles of a diploid organism, and characterization of allele-specific bursting. Here we describe SCALE, a bioinformatic and statistical framework for allele-specific gene expression analysis by scRNA-seq. SCALE estimates genome-wide bursting kinetics at the allelic level while accounting for technical bias and other complicating factors such as cell size...
2019: Methods in Molecular Biology
Alicia T Lamere, Jun Li
Single-cell RNA-Sequencing is a pioneering extension of bulk-based RNA-Sequencing technology. The "guilt-by-association" heuristic has led to the use of gene co-expression networks to identify genes that are believed to be associated with a common cellular function. Many methods that were developed for bulk-based RNA-Sequencing data can continue to be applied to single-cell data, and several of the most widely used methods are explored. Several methods for leveraging the novel time information contained in single-cell data when constructing gene co-expression networks, which allows for the incorporation of directed associations, are also discussed...
2019: Methods in Molecular Biology
Jean Fan
Integrating prior knowledge of pathway-level information can enhance power and facilitate interpretation of gene expression data analyses. Here, we provide a practical demonstration of the value of gene set or pathway enrichment testing and extend such techniques to identify and characterize transcriptional subpopulations from single-cell RNA-sequencing data using pathway and gene set overdispersion analysis (PAGODA).
2019: Methods in Molecular Biology
Karthik Shekhar, Vilas Menon
Unprecedented technological advances in single-cell RNA-sequencing (scRNA-seq) technology have now made it possible to profile genome-wide expression in single cells at low cost and high throughput. There is substantial ongoing effort to use scRNA-seq measurements to identify the "cell types" that form components of a complex tissue, akin to taxonomizing species in ecology. Cell type classification from scRNA-seq data involves the application of computational tools rooted in dimensionality reduction and clustering, and statistical analysis to identify molecular signatures that are unique to each type...
2019: Methods in Molecular Biology
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