Inhibition of cloned adenylyl cyclases by mutant-activated Gi-alpha and specific suppression of type 2 adenylyl cyclase inhibition by phorbol ester treatment.
Inhibition of the various cloned Gs-sensitive adenylyl cyclases was studied. The effect of mutant-activated Gi2-alpha on types 2, 3, and 6 adenylyl cyclases was analyzed after co-transfection into COS-7 cells. These adenylyl cyclases were chosen as representative members of the distinct families of mammalian adenylyl cyclases. COS-7 cells were transfected with cDNAs encoding (a) the required type of adenylyl cyclase, (b) luteinizing hormone receptor, and (c) Q205LGi2-alpha or Q205LGo1-alpha. Human chorionic gonadotropin (hCG)-stimulated cAMP accumulation was then measured. Transfection with Q205LGi2-alpha but not Q205LGo1-alpha resulted in 50-60% inhibition of hCG-stimulated cAMP accumulation of the type 2 adenylyl cyclase. hCG stimulation of types 3 and 6 adenylyl cyclases was also extensively inhibited by co-transfection with the Q205LGi2-alpha cDNA. Treatment with 4 beta-phorbol 12-myristate 13-acetate of cells transfected with the type 2 enzyme cDNA resulted in a 2-fold increase of the hCG-stimulated cAMP accumulation and a complete suppression of the Q205LGi2-alpha inhibition of the type 2 adenylyl cyclase. However, similar treatment did not affect Q205LGi2-alpha inhibition of the type 3 and type 6 adenylyl cyclases. These data indicate that many if not all types of adenylyl cyclases may be susceptible to inhibition by Gi2-alpha, but this inhibition can be regulated by protein kinase C in a type-specific manner.
Department of Pharmacology, Mount Sinai School of Medicine, City University of New York, New York 10029.
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J
Chen
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Iyengar
D000067956:Adenylyl Cyclase Inhibitors; D000262:Adenylyl Cyclases / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D002460:Cell Line; D006063:Chorionic Gonadotropin / Q000494:pharmacology*; D003001:Cloning, Molecular; D000242:Cyclic AMP / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000378:metabolism*; D006801:Humans; D007527:Isoenzymes / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D007700:Kinetics; D046911:Macromolecular Substances; D016296:Mutagenesis; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*; D014162:Transfection
D000067956:Adenylyl Cyclase Inhibitors; D006063:Chorionic Gonadotropin; D007527:Isoenzymes; D046911:Macromolecular Substances; D011994:Recombinant Proteins; D000242:Cyclic AMP; D019204:GTP-Binding Proteins; D000262:Adenylyl Cyclases; D013755:Tetradecanoylphorbol Acetate