Hey2 acts upstream of Notch in hematopoietic stem cell specification in zebrafish embryos

Haematopoietic stem cells (HSCs) are essential for homeostasis and injury-induced regeneration of the vertebrate blood system. Although HSC transplantations constitute the most common type of stem cell therapy applied in the clinic we know relatively little about the molecular programming of HSCs during vertebrate embryogenesis. In vertebrate embryos, HSCs form in close association with the ventral wall of the dorsal aorta (DA). We have shown previously that in zebrafish HSC formation depends on the presence of a signalling cascade that involves Hedgehog, vascular endothelial growth factor (Vegf) and Notch signalling. Here, we reveal that Hey2, a hairy and enhancer-of-split related basic helix-loop-helix transcription factor often believed to act downstream of Notch, is also required for HSC formation. In DA progenitors, Hey2 expression is induced downstream of cloche and the transcription factor Scl/Tal1, and is maintained by Hedgehog and Vegf signalling. While knock-down of Hey2 expression results in a loss of Notch receptor expression in the DA angioblasts, activation of Notch signalling in hey2 morphants rescues HSC formation in zebrafish embryos. These results establish an essential role for Hey2 upstream of Notch in HSC formation.