Cooperative Transcription Factor Induction Mediates Hemogenic Reprogramming

During development, hematopoietic stem/progenitor cells (HSPCs) arise from specialized endothelial cells by a process termed endothelial-to-hematopoietic transition (EHT). The genetic program driving human HSPC emergence remains largely unknown. We previously reported that the generation of hemogenic precursor cells from mouse fibroblasts recapitulates developmental hematopoiesis. Here, we demonstrate that human fibroblasts can be reprogrammed into hemogenic cells by the same transcription factors. Induced cells display dynamic EHT transcriptional programs, generate hematopoietic progeny, possess HSPC cell surface phenotype and repopulate immunodeficient mice for three months. Mechanistically, GATA2 and GFI1B interact and co-occupy a cohort of targets. This cooperative binding is reflected by engagement of open enhancers and promoters, initiating silencing of fibroblast genes and activating the hemogenic program. However, GATA2 displays dominant and independent targeting activity during the early phases of reprogramming. These findings shed light on the processes controlling human HSC specification and support generation of reprogrammed HSCs for clinical applications. mRNAseq profiling on adult human dermal fibroblasts (HDFa) at the population and single cell level and on neonatal foreskin fibroblasts (BJ) generated after transdution with GATA2, GFI1b, cFOS at day 2, day 15 and day 25. mRNAseq profiling of HDFa, BJ and Umbilical Cord Blood (UCB) Lin-Cd34+ single cells were used as controls. Examination of global occupancy for GATA2, GFI1b and FOS together or individually at day 2 of reprogramming.