Src and Pyk2 mediate G-protein-coupled receptor activation of epidermal growth factor receptor (EGFR) but are not required for coupling to the mitogen-activated protein (MAP) kinase signaling cascade

The epidermal growth factor receptor (EGFR) and the non-receptor protein ty rosine kinases Src and Pyk2 have been implicated in linking a variety of G- protein-coupled receptors (GPCR) to the mitogen-activated protein (MAP) kin ase signaling cascade. In this report we apply a genetic strategy using cel ls isolated from Src-, Pyk2-, or EGFR-deficient mice to explore the roles p layed by these protein tyrosine kinases in GPCR-induced activation of EGFR, Pyk2, and MAP kinase. We show that Src kinases are critical for activation of Pyk2 in response to GPCR-stimulation and that Pyk2 and Src are essentia l for GPCR-induced tyrosine phosphorylation of EGFR, By contrast, Pyk2, Src , and EGFR are dispensable for GPCR-induced activation of MAP kinase. Moreo ver, GPCR-induced MAP kinase activation is normal in fibroblasts deficient in both Src and Pyk2 (Src-/-Pyk2-/- cells) as well as in fibroblasts defici ent in all three Src kinases expressed in these cells (Src-/-Yes-/-Fyn-/- c ells). Finally, experiments are presented demonstrating that, upon stimulat ion of GPCR, activated Pyk2 forms a complex with Src, which in turn phospho rylates EGFR directly. These experiments reveal a role for Src kinases in P yk2 activation and a role for Pyk2 and Src in tyrosine phosphorylation of E GFR following GPCR stimulation, In addition, EGFR, Src family kinases, and Pyk2 are not required for linking GPCRs with the MAP kinase signaling casca de.