Project description:Transcriptional profiling of mouse ES cell-derived hemaopoitic cells comparing common primitive-definitive hematopoietic precursors (CD41SP) with definitve hematopoietic progenitor cells (KA45) RNA isolated from two separate experiments was pooled and used for comparison
Project description:Definitive hematopoiesis generates hematopoietic stem/progenitor cells (HSPCs) that give rise to all mature blood and immune cells, but remains poorly defined in human. Here, we resolve human hematopoietic populations at the earliest hematopoiesis stage by single-cell RNA-seq. We characterize the distinct molecular profiling between early primitive and definitive hematopoiesis in both human embryonic stem cell (hESC) differentiation and early embryonic development. We generate definitive HSPCs from hESCs that hold the multipotency to differentiate various blood and immune cells, as validated by single-cell clonal assay. Strikingly, the generated HSPCs from hESCs give rise to various blood and lymphoid lineages in vivo. Lastly, we characterize gene-expression dynamics in definitive and primitive hematopoiesis and reveal an unreported role of ROCK-inhibition in enhancing human definitive hematopoiesis. Our study provides a prospect for understanding human early hematopoiesis and a firm basis to generate blood and immune cells for clinical purposes.
Project description:A transcriptome study in mouse hematopoietic stem cells was performed using a sensitive SAGE method, in an attempt to detect medium and low abundant transcripts expressed in these cells. Among a total of 31,380 unique transcript, 17,326 (55%) known genes were detected, 14,054 (45%) low-copy transcripts that have no matches to currently known genes. 3,899 (23%) were alternatively spliced transcripts of the known genes and 3,754 (22%) represent anti-sense transcripts from known genes.
Project description:Lineage specification during development involves reprogramming of chromatin states, but little is known about how this is regulated in vivo. We previously showed that the chromatin remodeler Chd1 regulates transcriptional output and self-renewal of mouse embryonic stem cells, and is essential for epiblast development. These results raise the question of whether Chd1 regulates the development of other progenitor populations. Here we report that endothelial-specific deletion of Chd1 using Tie2-Cre leads to embryonic lethality by E15.5. Development of the vasculature and of primitive hematopoiesis appears to occur normally in the mutants. However, mutant embryos show signs of anemia as early as E11.5, are depleted of definitive hematopoietic stem /progenitor cells, and display a complete failure of fetal liver erythropoiesis. While mutants at E10.5 appear morphologically normal and can develop hemogenic clusters in the dorsal aorta, the E10.5 mutant endothelium fails to activate a transcriptional program associated with hematopoiesis. This transcriptional program may serve as a resource for the identification of novel markers or regulators of definitive hematopoiesis. Finally, hematopoietic-specific Chd1 deletion using Vav-Cre yields no apparent defects during development or adulthood. These results suggest that Chd1 regulates chromatin-remodeling events critical for a specific developmental window during the transition of endothelial cells to definitive blood progenitors. Analysis of CD31+ tdTomato+ cells sorted from E10.5 whole embryos with an endothelial-specific deletion of Chd1, using 4 biological replicates of 2 genotypes (CreHet controls vs mutants).