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. 2016 May 3;44(15):7173–7188. doi: 10.1093/nar/gkw327

Figure 5.

Figure 5.

Depletion of Setd1a inhibits β-globin gene transcription as well as recruitment of the Setd1a complex and H3K4me3 levels at the β-globin promoter. (A) Comparison of H3K4me3 enrichment at stemness promoters and erythroid promoters in ES cells and differentiated EBs. (B) Western blot analysis of Setd1a protein levels in R1E ES cells harboring shRNA against Setd1a compared to control; α-tubulin served as a loading control. (C) Stable ESC clones harboring vector control or shSetd1a-expressing constructs were induced with EPO and other cytokines and total RNA was extracted at day 19. Expression of genes important for erythroid differentiation was analyzed by qRT-PCR. (D) Stable ESC clones harboring vector control or shSetd1a-expressing constructs were induced with EPO and other cytokines for erythroid differentiation and total RNA was extracted at day 0, 4 and 19. The βmaj-globin transcripts were analyzed by qRT-PCR and normalized with β-actin. (E) ChIP analysis of H3K4me1, me2 and me3 patterns over the β-globin locus at day 19 upon the Setd1a KD. (F) ChIP analysis of ASH2L (TOP) and Setd1a (Bottom) recruitment across the β-globin locus at days 19 upon Setd1a KD. Data are collected from at least three independent experiments and shown as mean ± SD. *P < 0.05; **P < 0.01 by student t-test.