Inhibition of the Ca2+-ATPase Pmc1p by the v-SNARE protein Nyv1p

Pmc1p, the Ca2+-ATPase of budding yeast related to plasma membrane Ca2+-ATP ases of animals, is transcriptionally up-regulated in response to signaling by the calmodulin calcineurin-Tcn1p/Crz1p signaling pathway. Little is kno wn about post-translational regulation of Pmc1p. In a genetic screen for po tential negative regulators of Pmc1p, a vacuolar v-SNARE protein, Nyv1p, wa s recovered. Cells overproducing Nyv1p show decreased Ca2+ tolerance and de creased accumulation of Ca2+ in the vacuole, similar to pmc1 null mutants. Overexpression of Nyv1p had no such effects on pmc1 mutants, suggesting tha t Nyv1p may inhibit Pmc1p function. Overexpression of Nyv1p did not decreas e Pmc1p levels but decreased the specific ATP-dependent Ca2+ transport acti vity of Pmc1p in purified vacuoles by at least 2-fold. The effect of Nyv1p on Pmc1p function is likely to be direct because native immunoprecipitation experiments showed that Pmc1p coprecipitated with Nyv1p. Complexes between Nyv1p and its t-SNARE partner Vam3p were also isolated, but these complexe s lacked Pmc1p. We conclude that Nyv1p can interact physically with Pmc1p a nd inhibit its Ca2+ transport activity in the vacuole membrane. This is the first example of a Ca2+-ATPase regulation by a v-SNARE protein involved in membrane fusion reactions.