ATAC-seq analysis in 5F cells and AGM cells with EZH1 depletion

Blood develops in distinct stages. Haematopoietic progenitors in the embryo manifest restricted differentiation potential relative to definitive haematopoietic stem cells in adult bone marrow, which support lifelong multilineage haematopoiesis. To identify regulators of embryonic haematopoiesis, we screened chromatin modifiers and identified the Polycomb group protein EZH1 as a barrier to multilineage potential from pluripotent stem cells (PSCs). EZH1 was directly bound to bivalently poised, yet restricted, HSC and lymphoid genes in primitive progenitors; knockdown enabled robust generation of multilineage progenitors. Moreover, EZH1 haploinsufficiency promoted the generation of HSCs with long-term, multilineage and self-renewal potential from sites of embryonic haematopoiesis in vivo. Together, this work identifies EZH1 as a key epigenetic barrier to definitive haematopoiesis during embryonic development, and highlights the utility of chromatin modifiers as cell engineering targets to enhance blood differentiation from PSCs. ATAC-seq was performed to determine the chromatin accessibility in 5F cells transduced with shRNAs for luciferase (control) or EZH1, or in Ezh1 WT, HET or KO embryonic day (E)10.5 AGM cells.