Figure 8.

GPR30 signaling promotes the formation of Src-dependent, Shc-integrin α5β1 protein complexes. Triton X-100 extracts prepared from MDA-MB-231 cells stably expressing vector or GPR30 were left untreated (UN) or stimulated with 17β-E2, EGF, or pretreated with the Src family kinase inhibitor, PP1 (50 nm; 15 min) before stimulation with 17β-E2. Integrin α5β1 or α2β1 was immunoprecipitated with rabbit antibodies specific for the integrin α5 or α2 subunit cytoplasmic tails, respectively. Associated Shc proteins were detected with Shc mAB, PG797. Recovery of integrin α5β1 or α2β1 was determined by blotting with integrin α5 or α2 subunit protein antibodies. Relative positions of endogenous p66, p52, and p46 Shc are designated.