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Single-cell transcriptomic analysis of embryonic vasculogenesis identifies the conversion of Etv2-deficient vascular progenitors into skeletal muscle

ABSTRACT: An ETS transcription factor Etv2 functions as an evolutionarily conserved master regulator of vasculogenesis. Here we performed single-cell transcriptomic analysis of hematovascular development in wild-type and etv2 mutant zebrafish embryos. Distinct transcriptional signatures of different types of hematopoietic and vascular progenitors were identified using an etv2ci32Gt gene trap line, in which the Gal4 transcriptional activator is integrated into the etv2 gene locus. Unexpectedly, a cell population with a skeletal muscle signature was observed in etv2-deficient embryos. We demonstrate that multiple etv2ci32Gt; UAS:GFP cells migrate into the somites, elongate and differentiate as skeletal muscle cells instead of contributing to vasculature in etv2-deficient embryos.

ORGANISM(S): Danio rerio

PROVIDER: GSE143750 | GEO | 2020/01/16

REPOSITORIES: GEO

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Single-cell transcriptomic analysis of embryonic vasculogenesis identifies the conversion of Etv2-deficient vascular progenitors into skeletal muscle

Project description:During vertebrate embryogenesis, vascular endothelial cells originate in the lateral plate mesoderm (LPM) next to the progenitors of skeletal muscle. It is currently not clear what prevents vascular progenitors from responding to the adjacent signals that promote muscle development. An ETS transcription factor Etv2 functions as an evolutionarily conserved master regulator of vasculogenesis. Here we performed single-cell transcriptomic analysis of hematovascular development in wild-type and etv2 mutant zebrafish embryos. Distinct transcriptional signatures of different types of hematopoietic and vascular progenitors were identified using an etv2ci32Gt gene trap line, in which Gal4 transcriptional activator has integrated into the etv2 gene locus. Unexpectedly, a cell population with the skeletal muscle signature was observed in etv2-deficient embryos. We demonstrate that multiple etv2ci32Gt; UAS:GFP cells migrate into the somites, elongate and differentiate as skeletal muscle cells instead of contributing to vasculature in etv2-deficient embryos. Wnt and FGF signaling promoted the differentiation of these putative multipotent etv2 progenitor cells into skeletal muscle cells. We conclude that etv2 actively represses muscle differentiation in vascular progenitors, thus locking these cells into vascular endothelial fate. We also identified the transcriptional signature of putative multipotent progenitors within the LPM that may give rise to vascular progenitor and skeletal muscle cells. Finally, we demonstrate that arterial progenitors co-express multiple arterial and venous markers during early stages of vasculogenesis, suggesting multi-potency of early vascular progenitors.These findings will be important in understanding the ontogeny of different mesodermal lineages and will help in designing in vitro differentiation strategies to generate vascular, muscle and other types of progenitors for therapeutic purposes.

2020-03-10 | GSE142484 | GEO

Single-cell transcriptomic analysis of embryonic vasculogenesis identifies the conversion of Etv2-deficient vascular progenitors into skeletal muscle

Project description:Single-cell transcriptomic analysis of embryonic vasculogenesis identifies the conversion of Etv2-deficient vascular progenitors into skeletal muscle

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Single-cell transcriptomic analysis of embryonic vasculogenesis identifies the conversion of Etv2-deficient vascular progenitors into skeletal muscle

Project description:Single-cell transcriptomic analysis of embryonic vasculogenesis identifies the conversion of Etv2-deficient vascular progenitors into skeletal muscle

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Comparison of Flk-1+/Etv2- vs Flk-1+/Etv2+ populations

Project description:Screening for genes up in Etv2+ cells within Flk-1+ ES derived mesoderm Microarray analysis performed to screen for the candidate genes regulated by Etv2. Differentiated Flk-1+ mesoderm can be devided into Etv2+ or-. Etv2+ cells are assumed to be committed to hemato/endothelial cells. Comparison of two populations can reveal genes relevant in this commitment. Extract RNA from sorted Flk-1+/Etv2- vs Flk-1+/Etv2+ populations.Etv2-Venus KI ES cells were differentiated on OP9 for 4-5 days and Flk-1+ population was separated into Etv2-Venus+ or- cells. Total RNA was purified from each population for analysis.

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RNA-seq of etv2 mutant knock-in heterozygous and homozygous zebrafish embryos

Project description:Zebrafish etv2^Gt(2A-Gal4)ci32 line was generated using CRISPR-Cas9 mediated NHEJ approach which has 2A-Gal4 sequence inserted with the exon 5 of etv2 gene thus interrupting etv2 coding sequence. Heterozygous and homozogous embryos were subjected to bulk RNA-seq

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Comparison of Flk-1+/Etv2- vs Flk-1+/Etv2+ populations

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Gene expression profiling in Tie-2-Etv2 endothelial and/or hematopoietic precursors

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Endocardial identity is established during early somitogenesis by Bmp signalling acting upstream of npas4l and etv2

Project description:The endocardium plays important roles in the development and function of the vertebrate heart; however, few molecular markers of this tissue have been identified and little is known about what regulates its differentiation. Here, we describe the Gt(SAGFF27C); Tg(4xUAS:egfp) line as a marker of endocardial development in zebrafish. Transcriptomic comparison between endocardium and pan-endothelium confirms molecular distinction between these populations and time-course analysis suggests differentiation as early as eight somites. To investigate what regulates endocardial identity, we employed npas4l, etv2 and scl loss-of-function models. Endocardial expression is lost in npas4l mutants, significantly reduced in etv2 mutants and only modestly affected upon scl loss-of-function. Bmp signalling was also examined: overactivation of Bmp signalling increased endocardial expression, whereas Bmp inhibition decreased expression. Finally, epistasis experiments showed that overactivation of Bmp signalling was incapable of restoring endocardial expression in etv2 mutants. By contrast, overexpression of either npas4l or etv2 was sufficient to rescue endocardial expression upon Bmp inhibition. Together, these results describe the differentiation of the endocardium, distinct from vasculature, and place npas4l and etv2 downstream of Bmp signalling in regulating its differentiation.

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Gene expression profiling in Tie-2-Etv2 endothelial and/or hematopoietic precursors

Project description:Etv2 transgene was expressed from ROSA26 locus by removing floxed STOP cassette by Tie-2 Cre transgene. VE-Cad+/CD45= or VE-Cad+/CD45+ cells were sorted from control or tie-2-Etv2 E11.5 embryos (YS;yolk sac and Emb;embryo proper)and compared for gene expression in duplicate. Sample ID #1 VECAD+CD45-P3;Emb;Control #2 VECAD+CD45-P3;Emb;Tg #3 VECAD+CD45+P4;Emb;Control #4 VECAD+CD45+P4;Emb;Tg #5 VECAD+CD45-P3;YS;Control #6 VECAD+CD45-P3;YS;Tg #7 VECAD+CD45+P4;YS;Control #8 VECAD+CD45+P4;YS;Tg

2012-05-09 | E-GEOD-36516 | biostudies-arrayexpress

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