Filopodia are dispensable for endothelial tip cell guidance.

Actin filaments are instrumental in driving processes such as migration, cytokinesis and endocytosis and provide cells with mechanical support. During angiogenesis, actin-rich filopodia protrusions have been proposed to drive endothelial tip cell functions by translating guidance cues into directional migration and mediating new contacts during anastomosis. To investigate the structural organisation, dynamics and functional importance of F-actin in endothelial cells (ECs) during angiogenesis in vivo, we generated a transgenic zebrafish line expressing Lifeact-EGFP in ECs. Live imaging identifies dynamic and transient F-actin-based structures, such as filopodia, contractile ring and cell cortex, and more persistent F-actin-based structures, such as cell junctions. For functional analysis, we used low concentrations of Latrunculin B that preferentially inhibited F-actin polymerisation in filopodia. In the absence of filopodia, ECs continued to migrate, albeit at reduced velocity. Detailed morphological analysis reveals that ECs generate lamellipodia that are sufficient to drive EC migration when filopodia formation is inhibited. Vessel guidance continues unperturbed during intersegmental vessel development in the absence of filopodia. Additionally, hypersprouting induced by loss of Dll4 and attraction of aberrant vessels towards ectopic sources of Vegfa165 can occur in the absence of endothelial filopodia protrusion. These results reveal that the induction of tip cells and the integration of endothelial guidance cues do not require filopodia. Anastomosis, however, shows regional variations in filopodia requirement, suggesting that ECs might rely on different protrusive structures depending on the nature of the environment or of angiogenic cues.