Preactivation permits subsequent stimulation of phospholipase C by G(i)-coupled receptors

In the complex signal transduction networks involving G protein-coupled rec eptors there are numerous examples where G(i)-linked receptors augment G(q) -dependent signals. The mechanistic basis of such occurrences is thought to entail signal convergence at phospholipase C beta (PLC beta) via the G pro tein beta gamma-dimers. Herein, we explored the possibility that augmentati on by beta gamma-dimers requires preactivation of PLC beta. COS-7 cells wer e transiently cotransfected with cDNAs encoding various combinations of rec eptors and G protein subunits. The G(i) coupled delta- and kappa-opioid rec eptors could not stimulate PLC beta unless they were coexpressed with G alp ha(16). The opioid-induced response was dose-dependent and partially inhibi ted by pertussis toxin or coexpression with transducin, indicating the invo lvement of beta gamma-subunits released from the G(i) proteins. When PLC be ta was preactivated by constitutively active mutants of G alpha(16),G alpha (q), or G alpha(14), opioids enhanced the activity by 80 to 300% and such r esponses were mostly pertussis toxin-sensitive. The opioid-induced enhancem ent was dose-dependent and could not be blocked by staurosporin, a protein kinase C inhibitor. Other G(i)-coupled receptors that were ineffective on t heir own also acquired the ability to stimulate PLCb in the presence of a c onstitutively active mutant of G alpha(q). Coactivation of endogenous or ex ogenous G(q)-coupled receptors with the delta-opioid receptor produced stro ng stimulations of PLC beta and such responses could be partially blocked b y pertussis toxin. These results show that enhancement of G(q)-dependent si gnals by G(i)-coupled receptors requires activated PLC beta and is mediated via the beta gamma-dimer.