Regulation by Calmodulin of Adenylate Cyclase Activity in Anterior Pituitary*

Abstract

The control of adenylate cyclase activity is exerted through separate components: receptors, guanyl nucleotide-binding protein, catalytic subunit, calmodulin (CaM), and divalent cations. The roles of CaM, Mg2+ and Ca2+ in the regulation of adenylate cyclase activity in plasma membranes from anterior pituitary [rat] were examined. Adenylate cyclase activity was reduced with 2.5 mM EGTA [ethylene glycol bis(.beta.-aminoethylether) N,N,N'',N''-tetraacetic acid], 125 .mu.g/ml compounds 48/80, and 200 .mu.M trifluoperazine, which are known inhibitors of CaM in a variety of tissues. Mg2+, in excess of ATP, stimulated adenylate cyclase activity. Ca2+ produced a biphasic effect on adenylate cyclase activity over the concentration range of 0.1-10 mM, exhibiting inhibition up to 2.0 mM and stimulation above that, GTP, 5''-guanylylimidodiphosphate and F- each enhanced adenylate cyclase activity, but activity stimulated after each of these agents was reduced or returned towards control values by administration of compound 48/80 or trifluoperazine. In the absence of free Mg2+ (i.e., Mg2+ in excess of ATP concentrations), 10 mM Ca+2 produced marked stimulation of adenylate activity which was not reduced by trifluoperazine. Apparently, the plasma membranes from anterior pituitary possess a CaM-dependent adenylate cyclase and that activation of adenylate cyclase by guanyl nucleotide-binding protein requires CaM. Ca2+ may have allosteric binding sites on the catalytic subunit; Ca2+ and Mg2+ appear to have antagonistic effects at different binding sites.