Gαq potentiation of adenylate cyclase type 9 activity through a Ca2+/calmodulin-dependent pathway

Adenylate cyclase (EC 4.6.1.1) type 9 (AC9) activity has been shown to be inhibited by PMA activation of novel protein kinase C (nPKC) isoforms. In the current study the effect on AC9 activity of activating PKC in physiological relevant manner was examined. Contrary to the anticipated inhibitory effect of activating PKCs through G(q)-coupled receptors, activation of transiently expressed G,coupled serotonin 5-HT2A or muscarinic M-5 receptors resulted in the potentiation of isoproterenol-stimulated cyclic AMP accumulation in HEK293 cells stably expressing AC9 (HEK-AC9). Consistent with G(q)-mediated activation of PKC, the addition of the PKC inhibitor bisindolylmaleimide further potentiated isoproterenol-stimulated cyclic AMP accumulation. Expression of a constitutively active mutant of G alpha(q) in HEK-AC9 cells also produced an enhancement in basal and isoproterenol-stimulated cyclic AMP accumulation. We also examined the role of G alpha(q)-mediated release of intracellular Ca2+ on the observed potentiation of AC9 activity, by depleting intracellular Ca2+, stores with thapsigargin. In Ca2+-depleted HEK-AC9 cells, activation of transiently expressed M5 receptors resulted in inhibition of isoproterenol-stimulated cyclic AMP accumulation that was blocked by bisindolylmaleimide, indicating that M5 potentiation of AC9 activity requires Ca2+. This prompted us to examine the effects of the calmodulin antagonist W7 and the Ca2+/calmodulin-dependent kinase II (CaMK II) inhibitor KN-93. Pretreating cells with W7 and KN-93 significantly inhibited M-5-mediated potentiation of isoproterenol-stimulated cyclic AMP accumulation in HEK-AC9 cells, suggesting that Got, potentiation of AC9 activity involves Ca2+/calmodulin and CaMK H. This data provides evidence for Ca2+-mediated potentiation of AC9 activity. (c) 2005 Elsevier Inc. All rights reserved.