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 (G*s) 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 G*s concentration. At the highest concentration of G*s 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. G*s was not able to stimulate or potentiate a calmodulin-stimulated adenylate cyclase purified from membranes pretreated with the nonhydrolyzable GTP analog, guanyl-5'-yl beta,gamma-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 G*s. The results demonstrate that the guanyl nucleotide-dependent enhancement of calmodulin-stimulated adenylate cyclase activity is mediated by G*s and suggest that G*s modulates the enzymatic turnover of the calmodulin-stimulated activity.
Harrison
J K
JK
Department of Pharmacology, University of Michigan, Ann Arbor 48109.
Hewlett
G H
GH
Gnegy
M E
ME
D000262:Adenylyl Cyclases / Q000302:isolation & purification / Q000378:metabolism*; D000818:Animals; D002147:Calmodulin / Q000494:pharmacology*; D002417:Cattle; D003342:Corpus Striatum / Q000201:enzymology*; D019204:GTP-Binding Proteins / Q000302:isolation & purification / Q000502:physiology*; D006165:Guanylyl Imidodiphosphate / Q000494:pharmacology; D007700:Kinetics; D008099:Liver / Q000378:metabolism; D008970:Molecular Weight; D011817:Rabbits
D002147:Calmodulin; D006165:Guanylyl Imidodiphosphate; D019204:GTP-Binding Proteins; D000262:Adenylyl Cyclases