Project description:In order to identify genes that are activated in differentiating WT ESCs, but are missing in Tal-1-/- and Runx1-/- ESCs, and which might be involved in the generation of definitive hematopoietic progenitors and their specification thereafter, we performed microarray analyses on purified Flk-1+ cells, differentiated from these ESCs for 4, 5, and 6 days “in vitro”.
Project description:Cellular binary fate decisions require the progeny to silence genes associated with the alternative fate. The major subsets of alpha:beta T cells have been extensively studied as a model system for fate decisions. While the transcription factor RUNX3 is required for the initiation of Cd4 silencing in CD8 T cell progenitors, it is not required to maintain the silencing of Cd4 and other helper T lineage genes. The other runt domain containing protein, RUNX1, silences Cd4 in an earlier T cell progenitor, but this silencing is reversed whereas the gene silencing after RUNX3 expression is not reverse. Therefore, we hypothesized that RUNX3 and not RUNX1 recruits other factors that maintains the silencing of helper T lineage genes in CD8 T cells. To this end, we performed a proteomics screen of RUNX1 and RUNX3 to determine candidate silencing factors.
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:ES cells differentiated in the presence of the Wnt inhibitor DKK1 fail to express the transcription factor Snail and undergo EMT or mesoderm differentiation. We generated an ES cell line, A2.snail, that induced Snail expression upon addition of doxycycline addition. Microarrays were used to gain a global picture of Flk1- and Flk1+ cells generated one day after Snail was expressed during Wnt inhibition. A2.snail ES cells, which express Snail upon addition of doxycycline, were differentiated as embryoid bodies in differentiation media and DKK1. Snail-induced cultures uniquely develop a select population of Flk1+ cells. Total RNA was harvested from sorted control (no doxycycline) Flk1- cells and sorted Snail-induced (doxycycline at day 2) Flk1- and Flk1+ cells at day 3 of differentiation.
Project description:ES cells differentiated in the presence of the Wnt inhibitor DKK1 fail to express the transcription factor Snail and undergo EMT or mesoderm differentiation. We generated an ES cell line, A2.snail, that induced Snail expression upon addition of doxycycline addition. Microarrays were used to gain a global picture of Flk1- and Flk1+ cells generated one day after Snail was expressed during Wnt inhibition.
