Role of intracellular glutathione in the response of Saccharomyces cerevisiae to H2O2 was investigated. Depletion of cellular glutathione or inhibition of gamma-glutamylcysteine synthetase (GSH-I) enhanced the sensitivity to H2O2 and suppressed the adaptation to H2O2. A mutant deficient in GSH-I also showed the hypersensitivity and could not adapt to H2O2. Incubation of the cell with amino acids constituting glutathione (L-Glu, L-Cys, Gly) increased the intracellular glutathione content, and subsequently the cell acquired resistance against H2O2. These results strongly suggest that intracellular glutathione plays an important role in the adaptive response in S. cerevisiae to oxidative damage.