Comprehensive analysis of retinal development at single cell resolution identifies NFI factors as essential for mitotic exit and specification of late-born cells

SUMMARYPrecise temporal control of gene expression in neuronal progenitors is necessary for correct regulation of neurogenesis and cell fate specification. However, the extensive cellular heterogeneity of the developing CNS has posed a major obstacle to identifying the gene regulatory networks that control these processes. To address this, we used single cell RNA-sequencing to profile ten developmental stages encompassing the full course of retinal neurogenesis. This allowed us to comprehensively characterize changes in gene expression that occur during initiation of neurogenesis, changes in developmental competence, and specification and differentiation of each of the major retinal cell types. These data identify transitions in gene expression between early and late-stage retinal progenitors, as well as a classification of neurogenic progenitors. We identify here the NFI family of transcription factors (Nfia, Nfib, and Nfix) as genes with enriched expression within late RPCs, and show they are regulators of bipolar interneuron and Müller glia specification and the control of proliferative quiescence.

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The single-cell epigenetic regulatory landscape in mammalian perinatal testis development

10.1101/2021.03.17.435776 ◽

2021 ◽

Author(s):

Jinyue Liao ◽

Hoi Ching Suen ◽

Shitao Rao ◽

Alfred Chun Shui Luk ◽

Ruoyu Zhang ◽

...

Keyword(s):

Gene Expression ◽

Single Cell ◽

Cell Fate ◽

Germ Cells ◽

Somatic Cells ◽

Cell Types ◽

Regulatory Elements ◽

Cellular Heterogeneity ◽

Cell Populations ◽

Cell Type

AbstractSpermatogenesis depends on an orchestrated series of developing events in germ cells and full maturation of the somatic microenvironment. To date, the majority of efforts to study cellular heterogeneity in testis has been focused on single-cell gene expression rather than the chromatin landscape shaping gene expression. To advance our understanding of the regulatory programs underlying testicular cell types, we analyzed single-cell chromatin accessibility profiles in more than 25,000 cells from mouse developing testis. We showed that scATAC-Seq allowed us to deconvolve distinct cell populations and identify cis-regulatory elements (CREs) underlying cell type specification. We identified sets of transcription factors associated with cell type-specific accessibility, revealing novel regulators of cell fate specification and maintenance. Pseudotime reconstruction revealed detailed regulatory dynamics coordinating the sequential developmental progressions of germ cells and somatic cells. This high-resolution data also revealed putative stem cells within the Sertoli and Leydig cell populations. Further, we defined candidate target cell types and genes of several GWAS signals, including those associated with testosterone levels and coronary artery disease. Collectively, our data provide a blueprint of the ‘regulon’ of the mouse male germline and supporting somatic cells.

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Transcriptional Control of Gene Expression and the Heterogeneous Cellular Identity of Erythroblastic Island Macrophages

Frontiers in Genetics ◽

10.3389/fgene.2021.756028 ◽

2021 ◽

Vol 12 ◽

Author(s):

Kaustav Mukherjee ◽

James J. Bieker

Keyword(s):

Gene Expression ◽

Single Cell ◽

Transcriptional Control ◽

Cell Types ◽

Cellular Heterogeneity ◽

Erythroid Progenitors ◽

Distinct Cell Type ◽

Control Of Gene Expression ◽

Erythroblastic Island ◽

Isolation And Characterization

During definitive erythropoiesis, maturation of erythroid progenitors into enucleated reticulocytes requires the erythroblastic island (EBI) niche comprising a central macrophage attached to differentiating erythroid progenitors. Normally, the macrophage provides a nurturing environment for maturation of erythroid cells. Its critical physiologic importance entails aiding in recovery from anemic insults, such as systemic stress or acquired disease. Considerable interest in characterizing the central macrophage of the island niche led to the identification of putative cell surface markers enriched in island macrophages, enabling isolation and characterization. Recent studies focus on bulk and single cell transcriptomics of the island macrophage during adult steady-state erythropoiesis and embryonic erythropoiesis. They reveal that the island macrophage is a distinct cell type but with widespread cellular heterogeneity, likely suggesting distinct developmental origins and biological function. These studies have also uncovered transcriptional programs that drive gene expression in the island macrophage. Strikingly, the master erythroid regulator EKLF/Klf1 seems to also play a major role in specifying gene expression in island macrophages, including a putative EKLF/Klf1-dependent transcription circuit. Our present review and analysis of mouse single cell genetic patterns suggest novel expression characteristics that will enable a clear enrichment of EBI subtypes and resolution of island macrophage heterogeneity. Specifically, the discovery of markers such as Epor, and specific features for EKLF/Klf1-expressing island macrophages such as Sptb and Add2, or for SpiC-expressing island macrophage such as Timd4, or for Maf/Nr1h3-expressing island macrophage such as Vcam1, opens exciting possibilities for further characterization of these unique macrophage cell types in the context of their critical developmental function.

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Single-cell Transcriptomic Landscape of Nucleated Cells in Umbilical Cord Blood

10.1101/346106 ◽

2018 ◽

Cited By ~ 1

Author(s):

Yi Zhao ◽

Xiao Li ◽

Jingwan Wang ◽

Ziyun Wan ◽

Kai Gao ◽

...

Keyword(s):

Gene Expression ◽

Cord Blood ◽

Single Cell ◽

Cell Fate ◽

Umbilical Cord ◽

Umbilical Cord Blood ◽

Therapeutic Option ◽

Cell Types ◽

Cellular Heterogeneity ◽

Specific Cell

ABSTRACTUmbilical cord blood (UCB) transplant is a therapeutic option for both pediatric and adult patients with a variety of hematologic diseases such as several types of blood cancers, myeloproliferative disorders, genetic diseases, and metabolic disorders. However, the level of cellular heterogeneity and diversity of nucleated cells in the UCB has not yet been assessed in an unbiased and systemic fashion. In the current study, nucleated cells from UCB were subjected to single-cell RNA sequencing, a technology enabled simultaneous profiling of the gene expression signatures of thousands of cells, generating rich resources for further functional studies. Here, we report the transcriptomic maps of 19,052 UCB cells, covering 11 major cell types. Many of these cell types are comprised of distinct subpopulations, including distinct signatures in NK and NKT cell types in the UCB. Pseudotime ordering of nucleated red blood cells (NRBC) identifies wave-like activation and suppression of transcription regulators, leading to a polarized cellular state, which may reflect the NRBC maturation. Progenitor cells in the UBC also consist two subpopulations with divergent transcription programs activated, leading to specific cell-fate commitment. Collectively, we provide this comprehensive single-cell transcriptomic landscape and show that it can uncover previously unrecognized cell types, pathways and gene expression regulations that may contribute to the efficacy and outcome of UCB transplant, broadening the scope of research and clinical innovations.

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Single-cell analysis of human retina identifies evolutionarily conserved and species-specific mechanisms controlling development

10.1101/779694 ◽

2019 ◽

Cited By ~ 1

Author(s):

Yufeng Lu ◽

Fion Shiau ◽

Wenyang Yi ◽

Suying Lu ◽

Qian Wu ◽

...

Keyword(s):

Gene Expression ◽

Single Cell ◽

Cell Fate ◽

Single Cell Analysis ◽

Retinal Development ◽

Cell Types ◽

Retinal Cell ◽

Transcriptional Networks ◽

Species Specific

SummaryThe development of single-cell RNA-Sequencing (scRNA-Seq) has allowed high resolution analysis of cell type diversity and transcriptional networks controlling cell fate specification. To identify the transcriptional networks governing human retinal development, we performed scRNA-Seq over retinal organoid and in vivo retinal development, across 20 timepoints. Using both pseudotemporal and cross-species analyses, we examined the conservation of gene expression across retinal progenitor maturation and specification of all seven major retinal cell types. Furthermore, we examined gene expression differences between developing macula and periphery and between two distinct populations of horizontal cells. We also identify both shared and species-specific patterns of gene expression during human and mouse retinal development. Finally, we identify an unexpected role for ATOH7 expression in regulation of photoreceptor specification during late retinogenesis. These results provide a roadmap to future studies of human retinal development, and may help guide the design of cell-based therapies for treating retinal dystrophies.

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Joint profiling of gene expression and chromatin accessibility of amphioxus development at single cell resolution

10.21203/rs.3.rs-504113/v1 ◽

2021 ◽

Author(s):

Pengcheng Ma ◽

Xingyan Liu ◽

Huimin Liu ◽

Zaoxu Xu ◽

Xiangning Ding ◽

...

Keyword(s):

Gene Expression ◽

Single Cell ◽

Cell Fate ◽

Regulatory Networks ◽

Functional Divergence ◽

Expression Patterns ◽

Genetic Regulatory Networks ◽

Public Access ◽

Origin And Evolution ◽

Key Species

Abstract Vertebrate evolution was accompanied with two rounds of whole genome duplication followed by functional divergence in terms of regulatory circuits and gene expression patterns. As a basal and slow-evolving chordate species, amphioxus is an ideal paradigm for exploring the origin and evolution of vertebrates. Single cell sequencing has been widely employed to construct the developmental cell atlas of several key species of vertebrates (human, mouse, zebrafish and frog) and tunicate (sea squirts). Here, we performed single-nucleus RNA sequencing (snRNA-seq) and single-cell assay for transposase accessible chromatin sequencing (scATAC-seq) for different stages of amphioxus (covering embryogenesis and adult tissues). With the datasets generated we constructed the developmental tree for amphioxus cell fate commitment and lineage specification, and revealed the underlying key regulators and genetic regulatory networks. The generated data were integrated into an online platform, AmphioxusAtlas, for public access at http://120.79.46.200:81/AmphioxusAtlas.

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Engineering Gene Control Circuits with Allosteric Ribozymes in Human Cells as a Medicine of the Future

Bioinformatics ◽

10.4018/978-1-4666-3604-0.ch047 ◽

2013 ◽

pp. 860-883

Author(s):

Robert Penchovsky

Keyword(s):

Gene Expression ◽

Cell Fate ◽

Regulatory Networks ◽

Control Of Gene Expression ◽

Practical Applications ◽

Gene Therapies ◽

Sequencing Technologies ◽

The Future ◽

Control Circuits ◽

Exogenous Control

Systems and synthetic biology promise to develop new approaches for analysis and design of complex gene expression regulatory networks in living cells with many practical applications to the pharmaceutical and biotech industries. In this chapter the development of novel universal strategies for exogenous control of gene expression is discussed. They are based on designer allosteric ribozymes that can function in the cell. The synthetic riboswitches are obtained by a patented computational procedure that provides fast and accurate modular designs with various Boolean logic functions. The riboswitches can be designed to sense in the cell either the presence or the absence of disease indicative RNA(s) or small molecules, and to switch on or off the gene expression of any exogenous protein. In addition, the riboswitches can be engineered to induce RNA interference or microRNA pathways that can conditionally down regulate the expression of key proteins in the cell. That can prevent a disease’s development. Therefore, the presented synthetic riboswitches can be used as truly universal cellular biosensors. Nowadays, disease indicative RNA(s) can be precisely identified by employing next-generation sequencing technologies with high accuracy . The methods can be employed not only for exogenous control of gene expression but also for re-programming the cell fate, anticancer, and antiviral gene therapies. Such approaches may be employed as potent molecular medicines of the future.

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A Single-Cell Transcriptome Atlas of the Mouse Glomerulus

Journal of the American Society of Nephrology ◽

10.1681/asn.2018030238 ◽

2018 ◽

Vol 29 (8) ◽

pp. 2060-2068 ◽

Cited By ~ 53

Author(s):

Nikos Karaiskos ◽

Mahdieh Rahmatollahi ◽

Anastasiya Boltengagen ◽

Haiyue Liu ◽

Martin Hoehne ◽

...

Keyword(s):

Gene Expression ◽

Endothelial Cells ◽

Single Cell ◽

Mesangial Cells ◽

Transcriptional Profiling ◽

Wild Type Mouse ◽

Cell Types ◽

Cellular Heterogeneity ◽

Marker Genes ◽

Glomerular Cell

Background Three different cell types constitute the glomerular filter: mesangial cells, endothelial cells, and podocytes. However, to what extent cellular heterogeneity exists within healthy glomerular cell populations remains unknown.Methods We used nanodroplet-based highly parallel transcriptional profiling to characterize the cellular content of purified wild-type mouse glomeruli.Results Unsupervised clustering of nearly 13,000 single-cell transcriptomes identified the three known glomerular cell types. We provide a comprehensive online atlas of gene expression in glomerular cells that can be queried and visualized using an interactive and freely available database. Novel marker genes for all glomerular cell types were identified and supported by immunohistochemistry images obtained from the Human Protein Atlas. Subclustering of endothelial cells revealed a subset of endothelium that expressed marker genes related to endothelial proliferation. By comparison, the podocyte population appeared more homogeneous but contained three smaller, previously unknown subpopulations.Conclusions Our study comprehensively characterized gene expression in individual glomerular cells and sets the stage for the dissection of glomerular function at the single-cell level in health and disease.

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A single cell brain atlas in human Alzheimer’s disease

10.1101/628347 ◽

2019 ◽

Cited By ~ 4

Author(s):

Alexandra Grubman ◽

Gabriel Chew ◽

John F. Ouyang ◽

Guizhi Sun ◽

Xin Yi Choo ◽

...

Keyword(s):

Gene Expression ◽

Transcription Factor ◽

Single Cell ◽

Cell Fate ◽

Expression Patterns ◽

Cell Types ◽

Gene Expression Patterns ◽

Cell Type ◽

Web Resource ◽

Cell Type Specific

AbstractAlzheimer’s disease (AD) is a heterogeneous disease that is largely dependent on the complex cellular microenvironment in the brain. This complexity impedes our understanding of how individual cell types contribute to disease progression and outcome. To characterize the molecular and functional cell diversity in the human AD brain we utilized single nuclei RNA- seq in AD and control patient brains in order to map the landscape of cellular heterogeneity in AD. We detail gene expression changes at the level of cells and cell subclusters, highlighting specific cellular contributions to global gene expression patterns between control and Alzheimer’s patient brains. We observed distinct cellular regulation of APOE which was repressed in oligodendrocyte progenitor cells (OPCs) and astrocyte AD subclusters, and highly enriched in a microglial AD subcluster. In addition, oligodendrocyte and microglia AD subclusters show discordant expression of APOE. Integration of transcription factor regulatory modules with downstream GWAS gene targets revealed subcluster-specific control of AD cell fate transitions. For example, this analysis uncovered that astrocyte diversity in AD was under the control of transcription factor EB (TFEB), a master regulator of lysosomal function and which initiated a regulatory cascade containing multiple AD GWAS genes. These results establish functional links between specific cellular sub-populations in AD, and provide new insights into the coordinated control of AD GWAS genes and their cell-type specific contribution to disease susceptibility. Finally, we created an interactive reference web resource which will facilitate brain and AD researchers to explore the molecular architecture of subtype and AD-specific cell identity, molecular and functional diversity at the single cell level.HighlightsWe generated the first human single cell transcriptome in AD patient brainsOur study unveiled 9 clusters of cell-type specific and common gene expression patterns between control and AD brains, including clusters of genes that present properties of different cell types (i.e. astrocytes and oligodendrocytes)Our analyses also uncovered functionally specialized sub-cellular clusters: 5 microglial clusters, 8 astrocyte clusters, 6 neuronal clusters, 6 oligodendrocyte clusters, 4 OPC and 2 endothelial clusters, each enriched for specific ontological gene categoriesOur analyses found manifold AD GWAS genes specifically associated with one cell-type, and sets of AD GWAS genes co-ordinately and differentially regulated between different brain cell-types in AD sub-cellular clustersWe mapped the regulatory landscape driving transcriptional changes in AD brain, and identified transcription factor networks which we predict to control cell fate transitions between control and AD sub-cellular clustersFinally, we provide an interactive web-resource that allows the user to further visualise and interrogate our dataset.Data resource web interface:http://adsn.ddnetbio.com

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Characterizing and inferring quantitative cell cycle phase in single-cell RNA-seq data analysis

10.1101/526848 ◽

2019 ◽

Cited By ~ 3

Author(s):

Chiaowen Joyce Hsiao ◽

PoYuan Tung ◽

John D. Blischak ◽

Jonathan E. Burnett ◽

Kenneth A. Barr ◽

...

Keyword(s):

Gene Expression ◽

Cell Cycle ◽

Single Cell ◽

Cell Cycle Progression ◽

Cell Types ◽

Cell Cycle Phase ◽

Cellular Heterogeneity ◽

Cycle Phase ◽

Cycle Progression ◽

Cellular Environment

AbstractCellular heterogeneity in gene expression is driven by cellular processes such as cell cycle and cell-type identity, and cellular environment such as spatial location. The cell cycle, in particular, is thought to be a key driver of cell-to-cell heterogeneity in gene expression, even in otherwise homogeneous cell populations. Recent advances in single-cell RNA-sequencing (scRNA-seq) facilitate detailed characterization of gene expression heterogeneity, and can thus shed new light on the processes driving heterogeneity. Here, we combined fluorescence imaging with scRNA-seq to measure cell cycle phase and gene expression levels in human induced pluripotent stem cells (iPSCs). Using these data, we developed a novel approach to characterize cell cycle progression. While standard methods assign cells to discrete cell cycle stages, our method goes beyond this, and quantifies cell cycle progression on a continuum. We found that, on average, scRNA-seq data from only five genes predicted a cell’s position on the cell cycle continuum to within 14% of the entire cycle, and that using more genes did not improve this accuracy. Our data and predictor of cell cycle phase can directly help future studies to account for cell-cycle-related heterogeneity in iPSCs. Our results and methods also provide a foundation for future work to characterize the effects of the cell cycle on expression heterogeneity in other cell types.

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Single-Cell Genomics: Catalyst for Cell Fate Engineering

Frontiers in Bioengineering and Biotechnology ◽

10.3389/fbioe.2021.748942 ◽

2021 ◽

Vol 9 ◽

Author(s):

Boxun Li ◽

Gary C. Hon

Keyword(s):

Single Cell ◽

Cell Fate ◽

Molecular Mechanisms ◽

Cell Types ◽

Cellular Heterogeneity ◽

State Transitions ◽

Small Scale ◽

Specific Cell ◽

Direct Reprogramming ◽

Cell State

As we near a complete catalog of mammalian cell types, the capability to engineer specific cell types on demand would transform biomedical research and regenerative medicine. However, the current pace of discovering new cell types far outstrips our ability to engineer them. One attractive strategy for cellular engineering is direct reprogramming, where induction of specific transcription factor (TF) cocktails orchestrates cell state transitions. Here, we review the foundational studies of TF-mediated reprogramming in the context of a general framework for cell fate engineering, which consists of: discovering new reprogramming cocktails, assessing engineered cells, and revealing molecular mechanisms. Traditional bulk reprogramming methods established a strong foundation for TF-mediated reprogramming, but were limited by their small scale and difficulty resolving cellular heterogeneity. Recently, single-cell technologies have overcome these challenges to rapidly accelerate progress in cell fate engineering. In the next decade, we anticipate that these tools will enable unprecedented control of cell state.

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