Fig. 4

Dnmt1-mediated methylation controls HSPC generation through blocking arterial endothelial identity. (A) WISH analysis showing the expression of arterial endothelial genes dll4, dltC, ephrin-B2a and hey2 in siblings and dnmt1 mutants at 36 hpf. The arrowheads indicate the expression of corresponding arterial endothelial genes. n≥3 replicates. (B) Statistical analysis of the WISH in A. (C) qPCR analysis of arterial endothelial genes dltC, ephrin-B2a and hey2 in kdrl⁺ ECs in siblings and dnmt1 mutants. n=3 replicates. (D) WISH results showing expression of runx1 and cmyb (black arrowheads) at 36 hpf in sibling, dnmt1 mutant and dnmt1 mutant embryos injected with fli1a: mismatch-dnmt1-EGFP constructs (upper panels) and FISH analysis of cmyb and egfp (white arrowheads) at 36 hpf in control, dnmt1 morphants and embryos co-injected with dnmt1 atgMO and fli1a:mismatch-dnmt1-EGFP constructs showing that endothelial-derived Dnmt1-EGFP overexpression rescued the population of cmyb⁺ cells (bottom panel). n=3 replicates. (E) Statistical analysis of the WISH data in D. Data are mean±s.d. *P<0.05, **P<0.01 ***P<0.001 (unpaired two-tailed Student's t-test). Scale bars: 100 µm (A,D upper panels); 50 µm (D bottom panels).