Abstract 1221: Novel Role of VE-cadherin as a Scaffold Protein to Modulate VEGF-induced Src, Akt and eNOS Activation in Endothelial Cells
Vascular endothelial growth factor (VEGF) is a potent mediator for angiogenesis and vascular permeability. Src family kinase c-Src plays an essential role in those functions of VEGF. However, the molecular mechanisms by which VEGF stimulated c-Src activation are not well defined. Since vascular endothelial cadherin (VE-cadherin) is also implicated in VEGF regulation of vascular permeability and angiogenesis, we hypothesized that VE-cadherin modulated VEGF-stimulated c-Src activation in endothelial cells. In this study, we found that VE-cadherin was associated with c-Src and Csk (C-terminal Src Kinase, a negative regulator of Src activation) in human umbilical vein endothelial cells at basal condition, and VEGF stimulated Csk (but not c-Src) release from VE-cadherin resulting in c-Src phosphorylation at tyrosine 416 (pY416) and activation. Knockdown VE-cadherin by siRNA significantly inhibited c-Src pY416 by VEGF. Moreover, VEGF stimulated VE-cadherin to recruit protein tyrosine phosphatase SHP2. Knockdown SHP2 by siRNA inhibited c-Src pY416. Infection of adenovirus SHP2 mutant with phosphatase domain deletion (SHP2dPTP) also attenuated c-Src pY416. Interestingly, SHP2dPTP inhibited the release of Csk from VE-cadherin, which resulted in an increase of c-Src phosphorylation at tyrosine 527, a site targeted by Csk for inhibiting Src activation. Furthermore, we found that VE-cadherin and SHP2 regulation of c-Src activation is critical for VEGF signaling. Src kinase inhibitor PP2 blocked VEGF-stimulated activation of Akt and eNOS (endothelial nitric oxide synthase). VE-cadherin siRNA and SHP2 siRNA as well as SHP2dPTP inhibited Akt and eNOS activation, indicating that VE-cadherin/SHP2/Csk/Src module positively regulates Akt and eNOS activation by VEGF. In contrast, this module has a negatively role for VEGF-induced ERK1/2 activation since inhibiting VE-cadherin, SHP2 or Src enhanced ERK1/2 activation. In conclusion, our results demonstrate a novel role for VE-cadherin as a scaffold protein for Csk, c-Src and SHP2 to modulate c-Src, Akt and eNOS activation in endothelial cells by VEGF, and provide mechanistic insight into the importance of VE-cadherin in VEGF regulation of vascular permeability and angiogenesis.