RUNX1 is a key inducer of human hematopoiesis controlling non-hematopoietic mesodermal development.

Abstract

The RUNX1/AML1 transcription factor is one of the key regulators of definitive hematopoietic development in mice. However, its role in early human hematopoiesis remains poorly investigated. In this study, we integrated a tdTomato reporter cassette into the RUNX1 locus of human pluripotent stem cells (hPSCs) to monitor and block the expression of the gene during hPSC differentiation. This approach demonstrated that expression of RUNX1 starts early in mesodermal specification focusing later on hemogenic endothelium (HE) and nascent hematopoietic cells. Lack of RUNX1 halted the development of CD43+ and CD235-CD45+ hematopoietic cells, preventing the production of clonogenic hematopoietic progenitors including the multilineage ones. The abrogation of RUNX1 resulted in the failure of definitive lineages, specifically T and NK cells. Remarkably, we instead observed the accumulation of RUNX1-null HE cells at the stage of blood cell generation. Moreover, the loss of the gene biased the development toward the lineage of CD43-CD146+CD90+CD73+ mesenchymal cells. RNA-seq analysis of RUNX1-null cells revealed the downregulation of top-level hematopoietic transcription factor genes and the reciprocal upregulation of genes associated with non-hematopoietic cells of mesodermal origin. Forced expression of RUNX1c in differentiating RUNX1-null hPSCs effectively rescued the development of CD45+ myeloid cells and megakaryocytes. Our data demonstrate that RUNX1 is a top hematopoietic inducer that simultaneously controls the expansion of non-hematopoietic lineages.