ABSTRACT
The growth of new blood vessels by angiogenesis and their stabilization by the recruitment of perivascular mural cells are thought to be two sequential, yet independent events. Here we identify molecular links between both processes through the ßPix and integrin α(v)ß(8) proteins. Bubblehead (bbh) mutants with a genetic mutation in ßPix show defective vascular stabilization. ßPix is a guanine nucleotide exchange factor and scaffold protein that binds many proteins including Git1, which bridges ßPix to integrins at focal adhesions. Here we show that the ability of ßPix to stabilize vessels requires Git1 binding residues. Knockdown of Git1 leads to a hemorrhage phenotype similar to loss of integrin α(v), integrin ß(8) or ßPix, suggesting that vascular stabilization through ßPix involves interactions with integrins. Furthermore, double loss of function of ßPix and integrin α(v) shows enhanced hemorrhage rates. Not only is vascular stability impaired in these embryos, but we also uncover a novel role of both ßPix and integrin α(v)ß(8) in cerebral angiogenesis. Downregulation of either ßPix or integrin α(v)ß(8) results in fewer and morphologically abnormal cerebral arteries penetrating the hindbrain. We show that this is coupled with a significant reduction in endothelial cell proliferation in bbh mutants or integrin α(v)ß(8) morphants. These data suggest that a complex involving ßPix, GIT1 and integrin α(v)ß(8) may regulate vascular stability, cerebral angiogenesis and endothelial cell proliferation in the developing embryo.
