USF1 regulates ESC fate and hematopoietic differentiation by modulating bivalent domains of HS/PC-specific genes.

(A) Western blotting assay of hSET1A levels in undifferentiated (day 0) and differentiated (day 13) ESCs comparing pcDNA vector transfected control and AUSF1 overexpressing cells. (B) FACS analysis of c-Kit and Tie2 expressing hemogenic endothelium cell population in pcDNA tranfected control and AUSF1 overexpressing ESCs upon hematopoietic differentiation at day 6. (C) FACS analysis of CD41 and c-Kit expressing early hematopoietic stem and progenitor population in pcDNA transfected control and AUSF1 overexpressing ESCs upon hematopoietic differentiation at day 10. (D) Real-time RT-qPCR analysis of hematopoietic markers and transcription factors, c-Myb, CD150, CD34, Tal1, Runx1, and HoxB4 mRNA transcript levels upon hematopoietic differentiation comparing pcDNA transfected control and two AUSF1 overexpressing ES clones. Data are shown as mean ± SD. * P<0.05. (E) to (H). ChIP analyses of USF1 binding (E), hSET1A recruitment (F), H3K4me3 enrichment (G), and RNAPII loading (H) at the HoxB4 promoter in AUSF1 overexpressing ES clones. Data are shown as mean ± SD. * P<0.05.