Integrin αvβ3 requirement for VEGFR2-mediated activation of SAPK2/p38 and for Hsp90-dependent phosphorylation of focal adhesion kinase in endothelial cells activated by VEGF

Abstract

Endothelial cell migration, a key process in angiogenesis, requires the coordinated integration of motogenic signals elicited by the adhesion of endothelial cells to extracellular matrices and by angiogenic cytokines such as the vascular endothelial growth factor (VEGF). In this study, we found that addition of VEGF to human umbilical vein endothelial cells cultivated on vitronectin triggers a synergistic interaction between the VEGF receptor VEGFR2 and the clustered integrin receptor alphavbeta3. The interaction between VEGFR2 and alphavbeta3 is required for full phosphorylation of VEGFR2 and to drive the activation of motogenic pathways involving focal adhesion kinase (FAK) and stress-activated protein kinase-2/p38 (SAPK2/p38). The signal emanating from the VEGFR2 and alphavbeta3 interaction and leading to SAPK2/p38 activation proceeds directly from VEGFR2. The chaperone Hsp90 is found in a complex that coprecipitates with inactivated VEGFR2, and the association is increased by VEGF and decreased by geldanamycin, a specific inhibitor of Hsp90-mediated events. Geldanamycin also impairs the phosphorylation of FAK that results from the interaction between VEGFR2 and alphavbeta3, and this is accompanied by an inhibition of the recruitment of vinculin to VEGFR2. We conclude that a necessary cross talk should occur between VEGFR2 and the integrin alphavbeta3, to transduce the VEGF signals to SAPK2/p38 and FAK and that Hsp90 is instrumental in the building up of focal adhesions by allowing the phosphorylation of FAK and the recruitment of vinculin to VEGFR2.

Molecular blockade of VEGFR2 in human epithelial ovarian carcinoma cells

“…Interaction between VEGFR2 and a v b 3 integrin is necessary for VEGF to signal stress-activated protein kinase-2/p38 in normal endothelial cells undergoing angiogenesis. 39 Expression of VEGFR2 and a v b 3 was detected in endothelial cells in SKOV-3 EOC xenografts, 40 but to our knowledge, this is the first report of a relationship between VEGFR2 and integrin expression in EOC cells themselves. Our shRNAKDR OVCAR-3 cells showed altered survival in suspension, which was reflected in their ability to form ascites and subcutaneous cystic tumors in vivo.…”

Src-mediated Phosphorylation of Hsp90 in Response to Vascular Endothelial Growth Factor (VEGF) Is Required for VEGF Receptor-2 Signaling to Endothelial NO Synthase

“…We have previously shown that Hsp90 interacts with the VEGFR-2 and this VEGF-promoted effect contributes to signaling to FAK and actin remodeling (Rousseau et al, 2000;Masson-Gadais et al, 2003). Using MS/MS analysis, we have now identified Hsp90␤ as a tyrosine-phosphorylated protein associated to the VEGFR-2.…”

VEGF121 and VEGF165 Regulate Blood Vessel Diameter Through Vascular Endothelial Growth Factor Receptor 2 in an in vitro Angiogenesis Model

“…We currently have no explanation for this finding; however, we do note that most previous studies on p38 signaling in ECs have been performed in monolayer cultures, whereas ECs in this study are embedded in 3-D fibrin gels, which may result in previously unidentified cross-talk between pathways that could confound more simple interpretations. Supporting this hypothesis is a report that VEGFR-2 interacts with the integrin ␣ v␤3 to transduce signals to FAK and p38 that regulate cell movement (Masson-Gadais et al, 2003) and a second report suggesting that ␣2␤1 integrins may signal through p38 to modulate focal adhesions (Sweeney et al, 2003). ECs in 3-D gels will likely be triggering many integrins through matrix interactions, and loss of integrinmediated focal adhesions in the absence of active p38 may trigger cell apoptosis (anoikis).…”