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.