Vegfa signals through ERK to promote angiogenesis, but not artery differentiation

Vascular endothelial growth factor a (Vegfa) is essential for blood vessel formation and can induce activation of numerous signaling effectors in endothelial cells. However, it is unclear how and where these function in developmental contexts during vascular morphogenesis. To address this issue, we have visualized activation of presumptive Vegfa effectors at single-cell resolution in zebrafish blood vessels. From these studies, we find that phosphorylation of the serine/threonine kinase ERK (pERK) preferentially occurs in endothelial cells undergoing angiogenesis, but not in committed arterial endothelial cells. pERK in endothelial cells was ectopically induced by Vegfa and lost in Vegfa signaling mutants. Both chemical and endothelial autonomous inhibition of ERK prevented endothelial sprouting, but did not prevent initial artery differentiation. Timed chemical inhibition during angiogenesis caused a loss of genes implicated in coordinating tip/stalk cell behaviors, including flt4 and, at later stages, dll4 ERK inhibition also blocked excessive angiogenesis and ectopic flt4 expression in Notch-deficient blood vessels. Together, these studies implicate ERK as a specific effector of Vegfa signaling in the induction of angiogenic genes during sprouting.