Activation of H(+)-ATPase by glucose in Saccharomyces cerevisiae involves a membrane serine protease.
BACKGROUND: Glucose induces H(+)-ATPase activation in Saccharomyces cerevisiae. Our previous study showed that (i) S. cerevisiae plasma membrane H(+)-ATPase forms a complex with acetylated tubulin (AcTub), resulting in inhibition of the enzyme activity; (ii) exogenous glucose addition results in the dissociation of the complex and recovery of the enzyme activity. METHODS: We used classic biochemical and molecular biology tools in order to identify the key components in the mechanism that leads to H(+)-ATPase activation after glucose treatment. RESULTS: We demonstrate that glucose-induced dissociation of the complex is due to pH-dependent activation of a protease that hydrolyzes membrane tubulin. Biochemical analysis identified a serine protease with a kDa of 35-40 and an isoelectric point between 8 and 9. Analysis of several knockout yeast strains led to the detection of Lpx1p as the serine protease responsible of tubulin proteolysis. When lpx1Δ cells were treated with glucose, tubulin was not degraded, the AcTub/H(+)-ATPase complex did not undergo dissociation, and H(+)-ATPase activation was significantly delayed. CONCLUSION: Our findings indicate that the mechanism of H(+)-ATPase activation by glucose involves a decrease in the cytosolic pH and consequent activation of a serine protease that hydrolyzes AcTub, accelerating the process of the AcTub/H(+)-ATPase complex dissociation and the activation of the enzyme. GENERAL SIGNIFICANCE: Our data sheds light into the mechanism by which acetylated tubulin dissociates from the yeast H(+)-ATPase, identifying a degradative step that remained unknown. This finding also proposes an indirect way to pharmacologically regulate yeast H(+)-ATPase activity and open the question about mechanistic similarities with other higher eukaryotes.
Pubmed ID: 23518202 RIS Download
Acetylation | Adenosine Triphosphatases | Cell Membrane | Enzyme Activation | Glucose | Membrane Proteins | Multiprotein Complexes | Phospholipases A | Saccharomyces cerevisiae | Saccharomyces cerevisiae Proteins | Serine Proteases | Tubulin