Yap1p Activates Gene Transcription in an Oxidant-Specific Fashion

Abstract

ABSTRACT Positive regulation of gene expression by the yeast Saccharomyces cerevisiae transcription factor Yap1p is required for normal tolerance of oxidative stress elicited by the redox-active agents diamide and H 2 O 2 . Several groups have provided evidence that a cluster of cysteine residues in the extreme C terminus of the factor are required for normal modulation of Yap1p by oxidant challenge. Deletion of this C-terminal cysteine-rich domain (c-CRD) produces a protein that is highly active under both stressed and nonstressed conditions and is constitutively located in the nucleus. We have found that a variety of different c-CRD mutant proteins are hyperactive in terms of their ability to confer diamide tolerance to cells but fail to provide even normal levels of H 2 O 2 resistance. Although the c-CRD mutant forms of Yap1p activate an artificial Yap1p-responsive gene to the same high level in the presence of either diamide or H 2 O 2 , these mutant factors confer hyperresistance to diamide but hypersensitivity to H 2 O 2 . To address this discrepancy, we have examined the ability of c-CRD mutant forms of Yap1p to activate expression of an authentic target gene required for H 2 O 2 tolerance, TRX2 . When assayed in the presence of c-CRD mutant forms of Yap1p, a TRX2-lacZ fusion gene fails to induce in response to H 2 O 2 . We have also identified a second cysteine-rich domain, in the N terminus (n-CRD), that is required for H 2 O 2 but not diamide resistance and influences the localization of the protein. These data are consistent with the idea that the function of Yap1p is different at promoters of loci involved in H 2 O 2 tolerance from promoters of genes involved in diamide resistance.