Sema3d controls collective endothelial cell migration by distinct mechanisms via Nrp1 and PlxnD1.

During cardiovascular development, tight spatiotemporal regulation of molecular cues is essential for controlling endothelial cell (EC) migration. Secreted class III Semaphorins play an important role in guidance of neuronal cell migration and were lately linked to regulating cardiovascular development. Recently, SEMA3D gene disruptions were associated with cardiovascular defects in patients; however, the mechanisms of action were not revealed. Here we show for the first time that Sema3d regulates collective EC migration in zebrafish through two separate mechanisms. Mesenchymal Sema3d guides outgrowth of the common cardinal vein via repulsion and signals through PlexinD1. Additionally, within the same ECs, we identified a novel function of autocrine Sema3d signaling in regulating Actin network organization and EC morphology. We show that this new function requires Sema3d signaling through Neuropilin1, which then regulates Actin network organization through RhoA upstream of Rock, stabilizing the EC sheet. Our findings are highly relevant for understanding EC migration and the mechanisms of collective migration in other contexts.