Gata2is Required For The Runx1-Independent Hematopoiesis In Zebrafish

The current notion about how hematopoietic stem cells (HSCs) are generated identifies the transcription factor RUNX1 as an essential factor for the emergence of definitive hematopoietic stem cells (HSCs) from the hemogenic endothelium. Consequently, Runx1knockout mice fail to develop definitive hematopoiesis and lack all definitive blood lineages and cannot survive past embryonic day 12. However, even though zebrafish with arunx1stop codon mutation (runx1W84X/W84X) presented defects in definitive hematopoiesis during embryogenesis, runx1W84X/W84Xembryos could develop to fertile adults with blood cells of multi-lineages, raising the possibility that HSCs can emerge without RUNX1. In order to determine if a RUNX1-independent mechanism can support the generation of HSCs we have generated three new zebrafish runx1-/- with engineered deletions of the runx1gene using TALEN and CRISPR-Cas9. Our analysis shows that all three mutants have identical phenotypei.e., failure to develop definitive hematopoiesis during early embryogenesis, with later reemergence of hematopoietic cells and survivalof therunx1 mutants to adulthood, further confirming the existence of a RUNX1-independent mechanism for the emergence of HSCs. In the absence of a functional runx1, a cd41-GFP+population of hematopoietic precursors can still be detected in the aorta-gonad-mesonephros (AGM) region and in the hematopoietic tissues of the mutant embryos.