Resolving hematopoietic stem versus progenitor cell potential in the mouse dorsal aorta by differential Runx1 +110 enhancer activity

Hematopoietic stem cells (HSCs) are important in cell-based therapies for blood-related disorders. While progress has been made in the directed differentiation of pluripotent PSCs, such cultures promote hematopoietic progenitor cells (HPCs) over HSCs. Thus, elucidating signals, factors, and markers associated with HSC versus HPC lineage development in the embryo is imperative. During mouse embryonic development, HSCs and HPCs originate from hemogenic endothelium (HE) through a process critically dependent on the transcription factor Runx1. Here, we identified a Runx1 enhancer that distinguishes emerging dorsal aorta HSCs from HPCs. Phenotypic, functional, and transcriptomic analyses of Runx1 +110 enhancer-GFP reporter (110GFP) transgenic embryos showed that 110GFP expression marks HPCs, but not the emerging HSC lineage. Comparative transcriptomics revealed a 17-gene signature associated with in vivo long-term HSC potential. Furthermore, 110GFP- preHSCs showed increased expression of Jarid2 and other PRC2 components, suggesting a role for epigenetic regulation in establishing the HSC fate during EHT. Finally, single-cell multiome analysis of dorsal aorta EHT identified the Runx1 +3 enhancer as preferentially accessible in preHSC and underlined the specific activity of the +110 enhancer in HPCs. Our study demonstrates the power of cell-type specific enhancer-reporter models to dissect cell fate decisions in development and provides new inroads to label and/or perturb HSC versus HPC fate decisions in vivo and in vitro.