Regulation of P-ATPases by acetylated tubulin: S.Cerevisiae H+-ATPase regulation mechanism

CAMPETELLI, A.N.; MONESTEROLO, N; SANTANDER, V.S; PREVITALI, G; ARCE, C.A.; CASALE, C.H.

Villa Carlos Paz, Córdoba

Congreso; XLIV Reunión Anual de la Sociedad Argentina de de Investigación en Bioquímica y Biología Molecular.; 2008

Sociedad Argentina de de Investigación en Bioquímica y Biología Molecular.

Acetylated tubulin interacts with Na/K-ATPase and as results of this interaction the ATPase is inhibited. We have been studying the regulation of several P-ATPases by tubulin. When a Ca-ATPase enriched microsomal preparation is stimulated with calmodulin or ethanol, the Ca-ATPase-tubulin complex is dissociated and the ATPase is activated. Similarly, when S.cerevisiae are incubated with glucose, the tubulin-H-ATPase complex is dissociated. In all cases, when te stimuli are eliminated, the ATPase-tubulin complexes are restored and the enzymes are inhibited again. In the H-ATPase activation, tubulin degradation has been observed. The aim of this work is to demonstrate a protease involvement in this activation mechanism. In in vivo exeriments using several protease inhibitors, we found that tubulin proteolysis is mediated by a metalloprotease. Affinity chromatography showed that tubulin is able to retain a protease from a yeast cytosolic extract and in a tubulin degradation assay using this tubulin retained protease, we obtained significative results. These yeast cytosolic proteins retained by tubulin were identified by MALDI TOF, but any protease like protein was identified. Probably, thiks enzyme is not a cytosolic protein, so next experiments are focalized in the screeening of proteins from other subcellular compartment.