βPix plays a dual role in cerebral vascular stability and angiogenesis, and interacts with integrin αvβ8.
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 βPix and integrin α(v)β(8) proteins. Bubblehead (bbh) mutants with a genetic mutation in βPix show defective vascular stabilization. βPix is a guanine nucleotide exchange factor and scaffold protein that binds many proteins including Git1, which bridges βPix to integrins at focal adhesions. Here we show that the ability of βPix to stabilize vessels requires Git1 binding residues. Knockdown of Git1 leads to a hemorrhage phenotype similar to loss of integrin α(v), integrin β(8) or βPix, suggesting that vascular stabilization through βPix involves interactions with integrins. Furthermore, double loss of function of βPix 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 βPix and integrin α(v)β(8) in cerebral angiogenesis. Downregulation of either βPix or integrin α(v)β(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)β(8) morphants. These data suggest that a complex involving βPix, GIT1 and integrin α(v)β(8) may regulate vascular stability, cerebral angiogenesis and endothelial cell proliferation in the developing embryo.
Biochemistry and Molecular Biology, and Smooth Muscle Research Group, University of Calgary, AB, Canada.
Jing
J
Liu
Lei
L
Zeng
Regan M
RM
Kennedy
Nicole M
NM
Gruenig
Sarah J
SJ
Childs
D000818:Animals; D030801:Animals, Genetically Modified; D001808:Blood Vessels / Q000196:embryology / Q000378:metabolism*; D001921:Brain / Q000098:blood supply / Q000196:embryology / Q000378:metabolism; D049109:Cell Proliferation; D002543:Cerebral Hemorrhage / Q000196:embryology / Q000235:genetics; D002560:Cerebrovascular Circulation / Q000235:genetics; D042783:Endothelial Cells / Q000378:metabolism; D020690:GTPase-Activating Proteins / Q000235:genetics / Q000378:metabolism; D018507:Gene Expression Regulation, Developmental; D055785:Gene Knockdown Techniques; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D020662:Guanine Nucleotide Exchange Factors / Q000235:genetics* / Q000378:metabolism / Q000502:physiology; D007150:Immunohistochemistry; D017403:In Situ Hybridization; D016023:Integrins / Q000235:genetics* / Q000378:metabolism; D009154:Mutation; D018919:Neovascularization, Physiologic / Q000235:genetics* / Q000502:physiology; D011485:Protein Binding; D020133:Reverse Transcriptase Polymerase Chain Reaction; D064067:Rho Guanine Nucleotide Exchange Factors; D015027:Zebrafish / Q000196:embryology / Q000235:genetics / Q000378:metabolism; D029961:Zebrafish Proteins / Q000235:genetics* / Q000378:metabolism
C572193:G protein-coupled receptor kinase-interacting target, zebrafish; D020690:GTPase-Activating Proteins; D020662:Guanine Nucleotide Exchange Factors; D016023:Integrins; D064067:Rho Guanine Nucleotide Exchange Factors; D029961:Zebrafish Proteins; C403414:integrin alphavbeta8; D049452:Green Fluorescent Proteins