Project description:We found that mouse ES cell-derived Flk1+ cells could be subdivided into three population by the expression of PDGFRa and CAR (Flk1+PDGFRa-CAR-, Flk1+PDGFRa-CAR+, and Flk1+PDGFRa+CAR+). Therefore, global gene expression analysis was perfomed by microarray to characterize these mesodermal subsets. RNA isolated from five separate experiments was pooled and used for comparison
Project description:We found that mouse ES cell-derived Flk1+ cells could be subdivided into three population by the expression of PDGFRa and CAR (Flk1+PDGFRa-CAR-, Flk1+PDGFRa-CAR+, and Flk1+PDGFRa+CAR+). Therefore, global gene expression analysis was perfomed by microarray to characterize these mesodermal subsets.
Project description:Scl/Tal1 confers hemogenic competence and prevents cardiomyogenesis in embryonic endothelium. Here we show that Scl both directly activates a broad gene regulatory network required for hematopoietic stem/progenitor cell (HS/PC) development, and represses transcriptional regulators required for cardiogenesis. Cardiac repression occurs during a short developmental window through Scl binding to distant cardiac enhancers that harbor H3K4me1 at this stage. Scl binding to hematopoietic regulators extends throughout HS/PC and erythroid development and spreads from distant enhancers to promoters. Surprisingly, Scl complex partners Gata 1 and 2 are dispensable for hematopoietic versus cardiac specification and Scl binding to the majority of its target genes. Nevertheless, Gata factors co-operate with Scl to activate selected transcription factors to facilitate HS/PC emergence from hemogenic endothelium. These results uncover a dual function for Scl in dictating hematopoietic versus cardiac fate choice and suggest a mechanism by which lineage-specific bHLH factors direct the divergence of competing fates. ChIP-seq with Scl, Hand1, Lsd1, Ezh2, H3K4me1 and H3K27ac in different cell types with mesodermal origin. Scl ChIP-seq in WT, SclKO, SclKO-iScl and Gata12KO mES cell derived day4 EB (embryoid body) Flk1+ mesodermal cells, SclKO-iScl ES cells and MEL cells; Hand1 ChIP-seq in WT mES cell derived day4 EB Flk1+ mesodermal cells; Lsd1 and Ezh2 ChIP-seq in WT and SclKO mES cell derived day4 EB Flk1+ mesodermal cells. ChIP-seq of histone modifications H3K4me1 and H3K27ac in WT, SclKO and Gata12KO mES cell derived day4 EB Flk1+ mesodermal cells, HPC7 hematopoietic progenitor cells and HL1 cardiomyogenic cells
Project description:We identified distict mesodermal sub-populations based on Endoglin (Eng) and Flk1 expression in Brachyury (Bry) positive cells. By using whole-transcriptome analysis, we further characterized these populations and how they changed when Wnt pathway is inhibited Reaggregates mRNA profiles of unsorted, Flk1+ Eng+, and Flk1- Eng+ samples were generated by deep sequencing, in triplicate , using Ilumina.
Project description:We find a new non-coding transcript associated with active HoxB locus, which we named HoxBlinc lncRNA. We report that HoxBlinc RNA specifies Flk1+ mesoderm and then promotes the development of HS/PCs and cardiomyocytes. To understand the molecular pathways reguated by HoxBlinc, we generate two doxycycline inducible KD ES R1E cell lines, then differentiate these cell lines to day 6 via embryonic bodies stage. RNA-seq analysis were performed for day 6 EBs with or without doxycycline treatment starting at Flk1+ mesodermal stage.
Project description:Blood and endothelial cells arise from hemangiogenic progenitors that are specified from FLK1-expressing mesoderm by the transcription factor ETV2. FLK1 mesoderm also contributes to other tissues, including vascular smooth muscle (VSM) and cardiomyocytes. However, the developmental process of FLK1 mesoderm generation and its derivatives and the lineage relationship among FLK1 mesoderm derivatives these tissues remain obscure. Recent single cell RNA-sequencing (scRNA-seq) studies of early stages of embryogenesis embryos, or in vitro differentiated human embryonic stem (ES) cells have differentiation provided unprecedented information on the spatiotemporal resolution of cells in embryogenesis. Nonetheless, these snapshots still nonetheless offer insufficient information on dynamic developmental processes due to inadvertently missing intermediate states and unavoidable batch effects. Here we performed scRNA-seq of mouse ES cells in asynchronous embryoid bodies (EBs), in vitro differentiated embryonic stem (ES) cells containing undifferentiated ES cells and its differentiated hemangiogenic progeny, as well as yolk sacs, the first hematopoietic extraembryonic tissue in developing embryo that contains hemangiogenic and VSM lineages. We captured a continuous developmental process from undifferentiated pluripotent cells to FLK1 mesoderm-derived tissues involved in hemangiogenesis. This continuous transcriptome map will benefit both basic and applied studies of mesoderm and its derivatives.
Project description:We identified distict mesodermal sub-populations based on Endoglin (Eng) and Flk1 expression in Brachyury (Bry) positive cells. By using whole-transcriptome analysis, we further characterized these populations and how they changed when Wnt pathway is inhibited
Project description:We find a new non-coding transcript associated with active HoxB locus, which we named HoxBlinc lncRNA. We report that HoxBlinc RNA specifies Flk1+ mesoderm and then promotes the development of HS/PCs and cardiomyocytes. To understand the molecular pathways reguated by HoxBlinc, we generate two doxycycline inducible KD ES R1E cell lines, then differentiate these cell lines to day 6 via embryonic bodies stage. RNA-seq analysis were performed for day 6 EBs with or without doxycycline treatment starting at Flk1+ mesodermal stage. RNA from day 6 EBs targeting with 2 invidiual shRNAs with or without doxycycline treatment were combined together for RNA-seq analysis.
Project description:Mouse Embryonic Stem Cells (mESCs) were differentiated with EB formation and characterized with Flk1 and PDGFαR specific antibodies. miRNA profile of lateral mesodermal cells, paraxial mesodermal cells, DN (Double negative) and DP (Double positive) populations were firstly determined and effects of differentially expressed miRNAs transfected transiently on EB formation, and myogenic and hematopoietic differentiation potential were assessed.