A genome-wide RNAi screen identifies genes and networks governing hematopoietic stem cell development

ABSTRACT: We report a functional genome-wide RNAi screen to identify genes required for embryonic stem cell (ESC) differentiation to hematopoietic stem/progenitor cells (HSPC) in vitro, and report the discovery of novel genes important for differentiation to the endothelial–hematopoietic transition and subsequently to HSPC specification. iHoxB4 ESCs were transduced with a library of shRNAs targeting >15,000 genes, and were differentiated towards hematopoietic stem and progenitor cells by forcing HoxB4 expression in the cells. Five cell populations were isolated on differentiation days 6 and 20. Differentiated mesodermal/endothelium cell population D6F (Ssea1–Flkl+Cxcr4–), endodermal cell population D6C (Ssea1–Flkl–Cxcr4+), D20LS (Lin–Sca1+c-Kit–), D20LK (Lin–Sca1–c-Kit+), and D20LSK (Lin–Sca1+c-Kit+) cells were purified by FACS. Transduced iHoxB4 cells were analyzed as undifferentiated control cells. The experiment was performed three times independently. High-throughput sequencing and bioinformatics analysis identified twelve groups of shRNA target genes implicated in ESC to HSPC development, including a group of 351 novel genes that appear to be required for HSPC specification. As proof of concept in vivo, four genes in this group, Ap2a1, Mettl22, Lrsam1, and Hal, were selected for validation and were confirmed to be essential for HSPC development in zebrafish and for maintenance of human HSCs. Taken together, our results not only identify a number of novel regulatory genes and pathways throughout HSPC development but also serve as a potentially valuable resource for directed differentiation of therapy-grade HSPCs from human pluripotent stem cells.

ORGANISM(S): Mus musculus

PROVIDER: GSE86898 | GEO | 2017/01/27

SECONDARY ACCESSION(S): PRJNA342855

REPOSITORIES: GEO