Involvement of G protein βγ-subunits in diverse signaling induced by Gi/o-coupled receptors: study using theXenopusoocyte expression system

Abstract

We studied the functions of βγ-subunits of Gi/oprotein using the Xenopus oocyte expression system. Isoproterenol (ISO) elicited cAMP production and slowly activating Cl−currents in oocytes expressing β2-adrenoceptor and the protein kinase A-dependent Cl−channel encoded by the cystic fibrosis transmembrane conductance regulator (CFTR) gene. 5-Hydroxytryptamine (5-HT), [d-Ala2, d-Leu5]-enkephalin (DADLE), and baclofen enhanced ISO-induced cAMP levels and CFTR currents in oocytes expressing β2-adrenoceptor-CFTR and 5-HT1Areceptor (5-HT1AR), δ-opioid receptor, or GABABreceptor, respectively. 5-HT also enhanced pituitary adenylate cyclase activating peptide (PACAP) 38-induced cAMP levels and CFTR currents in oocytes expressing PACAP receptor, CFTR and 5-HT1AR. The 5-HT-induced enhancement of Gs-coupled receptor-mediated currents was abrogated by pretreatment with pertussis toxin (PTX) and coexpression of G transducin α (Gtα). The 5-HT-induced enhancement was further augmented by coexpression of the Gβγ-activated form of adenylate cyclase (AC) type II but not AC type III. Thus βγ-subunits of Gi/oprotein contribute to the enhancement of Gs-coupled receptor-mediated responses. 5-HT and DADLE did not elicit any currents in oocytes expressing 5-HT1AR or δ-opioid receptor alone. They elicited Ca2+-activated Cl−currents in oocytes coexpressing these receptors with the Gβγ-activated form of phospholipase C (PLC)-β2 but not with PLC-β1. These currents were inhibited by pretreatment with PTX and coexpression of Gtα, suggesting that βγ-subunits of Gi/oprotein activate PLC-β2 and then cause intracellular Ca2+mobilization. Our results indicate that βγ-subunits of Gi/oprotein participate in diverse intracellular signals, enhancement of Gs-coupled receptor-mediated responses, and intracellular Ca2+mobilization.