Phosphorylation and inhibition of type III adenylyl cyclase by calmodulin-dependent protein kinase II in vivo.
Inhibition of type III adenylyl cyclase (III-AC) by intracellular Ca2+ in vivo provides a mechanism for attenuation of hormone-stimulated cAMP signals in olfactory epithelium, heart, and other tissues (Wayman, G. A., Impey, S., and Storm, D. R. (1995) J. Biol. Chem. 270, 21480-21486). Although the mechanism for Ca2+ inhibition of III-AC in vivo has not been defined, inhibition is not mediated by Gi, cAMP-dependent protein kinase, or protein kinase C. However, Ca2+ inhibition of III-AC is antagonized by KN-62, a CaM-dependent kinase inhibitor. In addition, constitutively activated CaM kinase II inhibits the enzyme. These data suggest that CaM kinase II regulates the activity of III-AC by direct phosphorylation or by an indirect mechanism involving phosphorylation of a protein that inhibits III-AC. Here we report that III-AC is phosphorylated in vivo when intracellular Ca2+ is increased and that phosphorylation is prevented by CaM-dependent kinase inhibitors. Site-directed mutagenesis of a CaM kinase II consensus site (Ser-1076 to Ala-1076) in III-AC greatly reduced Ca2+-stimulated phosphorylation and inhibition of III-AC in vivo. These data support the hypothesis that Ca2+ inhibition of III-AC is due to direct phosphorylation of the enzyme by CaM kinase II in vivo.
Wei
J
J
Department of Pharmacology, University of Washington, Seattle, Washington 98195-7820, USA.
Wayman
G
G
Storm
D R
DR
D000067956:Adenylyl Cyclase Inhibitors; D000262:Adenylyl Cyclases / Q000378:metabolism*; D002118:Calcium / Q000502:physiology; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism*; D002460:Cell Line; D017868:Cyclic AMP-Dependent Protein Kinases / Q000037:antagonists & inhibitors; D018076:DNA, Complementary / Q000235:genetics; D006801:Humans; D016297:Mutagenesis, Site-Directed; D010766:Phosphorylation; D010768:Phosphoserine / Q000378:metabolism; D011493:Protein Kinase C / Q000037:antagonists & inhibitors; D011994:Recombinant Proteins; D013329:Structure-Activity Relationship
D000067956:Adenylyl Cyclase Inhibitors; D018076:DNA, Complementary; D011994:Recombinant Proteins; D010768:Phosphoserine; D017868:Cyclic AMP-Dependent Protein Kinases; D011493:Protein Kinase C; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D000262:Adenylyl Cyclases; D002118:Calcium