Phosphorylation of p130^Cas by Angiotensin II Is Dependent on c-Src, Intracellular Ca²⁺, and Protein Kinase C

From the Departments of Pathology (P.P.S., M.S.A., M.B.M., K.E.B.) and Medicine (J.B.H.), Emory University School of Medicine, Atlanta, Ga.

Correspondence to Kenneth E. Bernstein, MD, Department of Pathology, Emory University School of Medicine, 1639 Pierce Dr, 7107 WMB, Atlanta, GA 30322. E-mail kbernst{at}emory.edu

Abstract—p130^Cas is a signaling molecule that was initially found to be tyrosine-phosphorylated in v-Crk and v-Src transformed cells. We characterized the regulation of p130^Cas tyrosine phosphorylation in vascular smooth muscle cells by angiotensin II (Ang II). This ligand induced a transient increase in p130^Cas tyrosine phosphorylation, which was sensitive to the actin polymerization inhibitor cytochalasin D and to the intracellular Ca²⁺ chelator BAPTA-AM but not the Ca²⁺ channel blocker verapamil. The Ang II–induced tyrosine phosphorylation of p130^Cas was also dependent on an active Src family tyrosine kinase, since it could be blocked by the Src kinase inhibitors geldanamycin and PP1. Ang II treatment resulted in the ability of p130^Cas to bind at least 11 different phosphate-containing proteins. Analysis of these proteins revealed that protein kinase C and the cell adhesion signaling molecule pp120 formed temporal associations with p130^Cas in response to Ang II. c-Src was found to associate with p130^Cas in a manner that was independent of Ang II treatment. Inhibition of protein kinase C by either calphostin C or phorbol 12-myristate 13-acetate downregulation inhibited the Ang II–induced tyrosine phosphorylation of p130^Cas. These results are the first to demonstrate that the tyrosine phosphorylation of p130^Cas by Ang II is transduced by the Src, intracellular Ca²⁺, protein kinase C signaling pathway.

Key Words: p130^Cas • tyrosine phosphorylation • angiotensin II • vascular smooth muscle cell

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