We have examined the role of the Ca(2+)-dependent protein phosphatase 2B (calcineurin) in the regulation of the vacuolar H(+)-ATPase (V-ATPase) in blowfly salivary glands. In response to the neurohormone serotonin [5-hydroxytryptamine (5-HT)] and under the mediation of the cAMP/PKA signaling pathway, the secretory cells assemble and activate V-ATPase molecules at the apical membrane. We demonstrate that the inhibition of calcineurin activity by cyclosporin A, by FK-506, or by prevention of the elevation of Ca(2+) diminishes the 5-HT-induced assembly and activation of V-ATPase. The effect of calcineurin on V-ATPase is mediated by the cAMP/PKA signaling pathway, with calcineurin acting upstream of PKA, because 1) cyclosporin A does not influence the 8-(4-chlorophenylthio)adenosine-3',5'-cyclic monophosphate (8-CPT-cAMP)-induced activation of V-ATPase, and 2) the 5-HT-induced rise in cAMP is highly reduced in the presence of cyclosporin A. Moreover, a Ca(2+) rise evoked by the sarco(endo)plasmic reticulum Ca(2+)-ATPase (SERCA) inhibitor cyclopiazonic acid leads to an increase in intracellular cAMP concentration and a calcineurin-mediated PKA-dependent activation of V-ATPase. We propose that calcineurin activity mediates cross talk between the inositol 1,4,5-trisphosphate/Ca(2+) and the cAMP/PKA signaling pathways, thereby augmenting the 5-HT-induced rise in cAMP and thus the cAMP/PKA-mediated activation of V-ATPase.

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