Isoform-specific sensitization of adenylyl cyclase activity by prior activation of inhibitory receptors: role of beta gamma subunits in transducing enhanced activity of the type VI isoform.

Many different types of cells develop increased adenylyl cyclase activity (sensitization) on prior treatment with drugs such as opiates that acutely inhibit the enzyme. We found that human embryonic kidney (HEK) 293 m2 cells, which express the inhibitory m2 muscarinic cholinergic receptor, exhibit a large increase in forskolin-stimulated cAMP synthesis when the cells are preincubated with the muscarinic agonist carbachol for > or = 5 min and forskolin stimulation is performed in the presence of the muscarinic antagonist atropine. To determine whether a specific isoform of adenylyl cyclase is susceptible to the adaptation induced by prior activation of inhibitory receptors, cells were transfected with expression vectors encoding adenylyl cyclase types I, II, and VI, representing three major groups of the adenylyl cyclase family. Preincubation of the cells with carbachol for 30 min resulted in a significant increase in prostaglandin E1-stimulated cAMP accumulation in cells expressing type VI, but not type I or type II, adenylyl cyclase. A similar selective increase in activity from type VI adenylyl cyclase was observed for prior treatment with the D2 dopamine agonist quinpirole and stimulation of cAMP synthesis with human chorionic gonadotropin in cells transfected with expression vectors coding for the cognate receptors. We next investigated whether beta gamma subunits play a role in the sensitization of type VI adenylyl cyclase activity; using expression of alpha tau to inhibit beta gamma-mediated effects, we found that the quinpirole-induced sensitization of type VI adenylyl cyclase was abolished. However, beta gamma subunits do not seem to directly activate type VI adenylyl cyclase, in contrast with their ability to directly activate the type II enzyme. Therefore, beta gamma subunits liberated after activation of inhibitory receptors seem to indirectly cause an increase in activity of type VI adenylyl cyclase. Indirect activation of the type VI enzyme by beta gamma subunits is a novel mechanism contributing to the sensitization of adenylyl cyclase.