Regulation of Calmodulin-sensitive Adenylate Cyclase by the Stimulatory G-protein, Gs

Studies in bovine and rat brain membranes have suggested that calmodulin can potentiate neurotransmitter- and GTP-stimulated adenylate cyclase activities. To examine whether calmodulin and the stimulatory G-protein, Gs, are potentiative at a calmodulin-sensitive adenylate cyclase, Gs, was purified from rabbit liver and reconstituted with a partially purified calmodulin-sensitive adenylate cyclase from bovine brain. Activated Gs (Gs*) stimulated basal adenylate cyclase activity and enhanced the stimulation by calmodulin. The potentiation of the calmodulin-stimulated adenylate cyclase activity was dose-dependent with respect to Gs* concentration. At the highest concentration of Gs* tested (3 nM), a 2-fold enhancement of the calmodulin-stimulated adenylate cyclase activity was observed at all concentrations of calmodulin. The synergistic activation of adenylate cyclase by calmodulin and Gs was dependent on the presence of Ca2+ and occurred at physiologically relevant Ca2+ concentrations. The potentiation was not observed when either a nonactivated Gs or a mixture of activated Gi/Go was used. Gs* was not able to stimulate or potentiate a calmodulin-stimulated adenylate cyclase purified from membranes pretreated with the nonhydrolyzable GTP analog, guanyl-5′-yl β,γ-imidodiphosphate. Photochemical cross-linking of 125I-calmodulin-diazopyruvamide to proteins having an Mr corresponding to the known Mr of adenylate cyclase was not enhanced by Gs*. The results demonstrate that the guanyl nucleotide-dependent enhancement of calmodulin-stimulated adenylate cyclase activity is mediated by Gs* and suggest that Gs* modulates the enzymatic turnover of the calmodulin-stimulated activity.