Control of dynamic cell behaviors during angiogenesis and anastomosis by Rasip 1

Organ morphogenesis is driven by a wealth of tightly orchestrated cellular behaviors, which ensure proper organ assembly and function. Many of these cell activities involve cell-cell interactions and remodeling of the F-actin cytoskeleton. Here, we analyze the requirement for Rasip1 (Ras-interacting protein 1), an endothelial-specific regulator of junctional dynamics, during blood vessel formation. Phenotype analysis of rasip1 mutants in zebrafish embryos reveal distinct requirements for Rasip1 during sprouting angiogenesis, vascular anastomosis and lumen formation. During angiogenic sprouting, Rasip1 is required for efficient cell pairing, which is essential for multicellular tube formation. High-resolution time-lapse analyses show that these cell pairing defects are caused by a destabilization of tricellular junctions suggesting that tri-cellular junctions may serve as a counterfort to tether sprouting endothelial cells during morphogenetic cell rearrangements. During anastomosis, Rasip1 is required to establish a stable apical membrane compartment; rasip1 mutants display ectopic, reticulated junctions and the apical compartment is frequently collapsed. Loss of Ccm1 and Heg1 function leads to junctional defects similar to those seen in rasip1 mutants. Analysis of radil-b single and rasip1/radil-b double mutants reveal distinct and overlapping functions of both proteins. While Rasip1 and Radil-b have similar functions during angiogenic sprouting, the junction formation during anastomosis may primarily depend on Rasip1.