Direct binding of activated c-Src to the β3-adrenergic receptor is required for MAP kinase activation

Both beta (2)- and beta (3)-adrenergic receptors (ARs) are able to activate the extracellular signal-regulated kinase (ERK) pathway. We previously showed that c-Src is required for ERK activation by beta (2)AR and that it is recruited to activated beta (2)AR through binding of the Src homology 3 (SH3) domain to proline-rich regions of the adapter protein beta -arrestin1, Despite the absence of sites for phosphorylation and beta -arrestin binding, ERK activation by P(3)AR still requires c-Src. Agonist activation of beta (2)AR, but not beta (3)AR, led to redistribution of green fluorescent protein-tagged p-arrestin to the plasma membrane. In P-arrestin-deficient COS-7 cells, beta -affonist-dependent coprecipitation of c-Src with the beta (2)AR required exogenous p-arrestin, but activated beta (3)AR co-precipitated c-Src in the absence or presence of p-arrestin, ERK activation and Src co-precipitation with beta (3)AR also occurred in adipocytes in an agonist-dependent and pertussis toxin-sensitive manner. Protein interaction studies show that the beta (3)AR interacts directly with the SH3 domain of Src through proline-rich motifs (PXXP) in the third intracellular loop and the carboxyl terminus. ERK activation and Src co-precipitation were abolished in cells expressing point mutations in these PXXP motifs. Together, these data describe a novel mechanism of ERK activation by a G protein-coupled receptor in which the intracellular domains directly recruit c-Src.