Distinct patterns of bidirectional regulation of mammalian adenylyl cyclases

The capacities of the α subunits of pertussis toxin-sensitive guanine nucleotide-binding regulatory proteins (G proteins) to inhibit different isoforms of mammalian adenylyl cyclases were assessed. Membranes from Sf9 cells infected with recombinant baculoviruses encoding either type I, II, V, or VI adenylyl cyclase were reconstituted with purified G protein subunits. Types V and VI adenylyl cyclase are most sensitive to inhibition by G(iα1), G(iα2), and G(iα3); type I adenylyl cyclase can be inhibited by these three G(iα) proteins and by G(oα) as well. Type II adenylyl cyclase appears to be immune to inhibition by these proteins. Examination of the effects of native and mutant G(iα) proteins, as well as analysis of competition for binding of G(sα) to adenylyl cyclases, indicate that at least certain adenylyl cyclases have independent sites for interaction with G(sα) (site 1, stimulatory) and G(iα) (site 2, inhibitory). High concentrations of G(iα) can interact with site 1 on types I and II adenylyl cyclase and activate the enzymes. Types I and II adenylyl cyclase also appear to have independent sites for interaction with G protein βγ subunits. The type I enzyme is strongly inhibited, while type II adenylyl cyclase is activated if G(sα) is also present.