Loss of vacuolar proton-translocating ATPase activity in yeast results in chronic oxidative stress

Yeast mutants lacking vacuolar proton-translocating ATPas (V-ATPase) subunits (vma mutants) were sensitive to several different oxidants in a recent genomic screen (Thorpe, G. W. Fong, C. S., Alic, N., Higgins, V. J., and Dawes, 1. W. (2004) Proc Natl. Acad. Sci. U. S.A. 10 1, 6564 - 6569). We confirmed that mutants lacking a V, subunit (vma2 Delta), V. subunit, or either of the two V. a subunit isoforms are acutely sensitive to H2O2 and more sensitive to menadione and diamide than wild-type cells The vma2 Delta mutant contains elevated levels of reactive oxygen species and high levels of oxidative protein damage even in the absence of an applied oxidant, suggesting an endogenous source of oxidative stress. vma2 Delta mutants lacking mitochondrial DNA showed neither improved growth nor decreased sensitivity to peroxide, excluding respiration as the major source of the endogenous reactive oxygen species in the mutant. Double mutants lacking both VAM2 and components of the major cytosolic defense systems exhibited synthetic sensitivity to H2O2. Microarray analysis comparing wild-type and vma2 Delta mutant cells grown at pH 5, permissive conditions for the vma2 Delta mutant, indicated high level up-regulation of several iron uptake and metabolism genes that are part of the Aft1/Aft2 regulon. TSA2, which encodes an isoform of the cytosolic thioredoxin peroxidase, was strongly induced, but other oxidative stress defense systems were not induced. The results indicate that V-ATPase activity helps to protect cells from endogenous oxidative stress.