A link of Ca(2+ )to cAMP oscillations in Dictyostelium: the calmodulin antagonist W-7 potentiates cAMP relay and transiently inhibits the acidic Ca(2+)-store

BACKGROUND: During early differentiation of Dictyostelium the attractant cAMP is released periodically to induce aggregation of the cells. Here we pursue the question whether pulsatile cAMP signaling is coupled to a basic Ca(2+)-oscillation. RESULTS: We found that the calmodulin antagonist W-7 transiently enhanced cAMP spikes. We show that W-7 acts on an acidic Ca(2+)-store: it abolished ATP-dependent vesicular acidification, inhibited V-type H(+)ATPase activity more potently than the weaker antagonist W-5 and caused vesicular Ca(2+)-leakage. Concanamycin A, an inhibitor of the V-type H(+)-pump, blocked the Ca(2+)-leakage elicited by W-7 as well as cAMP-oscillations in the presence of W-7. Concanamycin A caused an increase of the cytosolic Ca(2+)-concentration whereas W-7 did not. In case of the latter, Ca(2+ )was secreted by the cells. In accord with our hypothesis that the link from Ca(2+ )to cAMP synthesis is mediated by a Ca(2+)-dependent phospholipase C we found that W-7 was not active in the phospholipase C knockout mutant. CONCLUSION: We conclude that the potentiation of cAMP relay by W-7 is due to a transient inhibition of the acidic Ca(2+)-store. The inhibition of the proton pump by W-7 causes a leakage of Ca(2+ )that indirectly stimulates adenylyl cyclase activity via phospholipase C.