VEGFR2 induces c-Src signaling and vascular permeability in vivo via the adaptor protein TSAd

Abstract

Regulation of vascular endothelial (VE) growth factor (VEGF)–induced permeability is critical in physiological and pathological processes. We show that tyrosine phosphorylation of VEGF receptor 2 (VEGFR2) at Y951 facilitates binding of VEGFR2 to the Rous sarcoma (Src) homology 2-domain of T cell–specific adaptor (TSAd), which in turn regulates VEGF-induced activation of the c-Src tyrosine kinase and vascular permeability. c-Src was activated in vivo and in vitro in a VEGF/TSAd-dependent manner, and was regulated via increased phosphorylation at pY418 and reduced phosphorylation at pY527. Tsad silencing blocked VEGF-induced c-Src activation, but did not affect pathways involving phospholipase Cγ, extracellular regulated kinase, and endothelial nitric oxide. VEGF-induced rearrangement of VE–cadherin–positive junctions in endothelial cells isolated from mouse lungs, or in mouse cremaster vessels, was dependent on TSAd expression, and TSAd formed a complex with VE-cadherin, VEGFR2, and c-Src at endothelial junctions. Vessels in tsad−/− mice showed undisturbed flow and pressure, but impaired VEGF-induced permeability, as measured by extravasation of Evans blue, dextran, and microspheres in the skin and the trachea. Histamine-induced extravasation was not affected by TSAd deficiency. We conclude that TSAd is required for VEGF-induced, c-Src-mediated regulation of endothelial cell junctions and for vascular permeability.