Luteinizing hormone/choriogonadotropin receptor-mediated activation of heterotrimeric guanine nucleotide binding proteins in ovarian follicular membranes

The LH/CG receptor signals to adenylyl cyclase via the stimulatory heterotrimeric GTP binding regulatory protein, Gs, and to phospholipase C and potentially to other effectors, such as ion channels, via a G protein or proteins that have not been identified in gonadal cells. To identify G proteins activated in a physiological membrane environment upon LH/CG receptor activation, we used the ability of activated G proteins to bind GTP and incubated ovarian follicular membranes with the photoaffinity GTP analog, P³-(4-azidoanilido)-P¹-5'-GTP ([³²P]AAGTP). Results showed that human CG (hCG) stimulated the binding of [³²P]AAGTP to a 45-kDa protein(s) in follicular membranes that comigrated with immunoreactive Gαs, Gαq/11, and Gα13. When Gα proteins were immunoprecipitated from Triton X-100 solubilized membrane extracts after photoaffinity labeling with [³²P]AAGTP, a time-dependent increase in hCG-dependent [³²P]AAGTP binding to Gαs, Gαq/11, and Gαi was detected. hCG-dependent [³²P]AAGTP binding to Gα13 was also detected. These results demonstrate that agonist-dependent LH/CG receptor activation promotes the activation of Gs, Gi, Gq/11, and G13 in porcine ovarian follicular membranes. These results further suggest that Gαs remains coupled to the agonist-bound LH/CG receptor during at least the initial 10 min after agonist-dependent receptor activation.