Cross-talk between Transcriptional Regulators of Multidrug Resistance in Saccharomyces cerevisiae

Xiaoting Zhang,Zhifeng Cui,Tokichi Miyakawa,W Scott Moye-Rowley

doi:10.1074/jbc.m010686200

Abstract

Multiple or pleiotropic drug resistance often arises in the yeast Saccharomyces cerevisiae due to genetic alterations of the functional state of the Cys(6)-Zn(II)(2) transcription factors Pdr1p and Pdr3p. Single amino acid substitutions give rise to hyperactive forms of these regulatory proteins, which in turn cause overproduction of downstream target genes that directly mediate multidrug resistance. Previous work has identified a novel Cys(6)-Zn(II)(2) transcription factor designated Yrr1p as mutant forms of this protein confer high level resistance to the cell cycle inhibitor reveromycin A and DNA damaging agent 4-nitroquinoline-N-oxide. In the present study, we demonstrate that Yrr1p also mediates oligomycin resistance through activation of the ATP-binding cassette transporter-encoding gene YOR1. Additionally, insertion of triplicated copies of the hemagglutinin epitope in the C-terminal region of Yrr1p causes the protein to behave as a hyperactive regulator of transcription. We have found that YRR1 expression is both controlled in a Pdr1p/Pdr3p-dependent manner and autoregulated. Chromatin immunoprecipitation experiments also show that Yrr1p associates with target promoters in vivo. Together these data argue that the signal generated by activation of Pdr1p and/or Pdr3p can be amplified through the action of these transcriptional regulatory proteins on downstream target genes, like YRR1, that also encode transcription factors.

Highlights

Multiple or pleiotropic drug resistance often arises in the yeast Saccharomyces cerevisiae due to genetic alterations of the functional state of the Cys6-Zn(II)2 transcription factors Pdr1p and Pdr3p
These data indicate that, YRR1 does contribute to oligomycin resistance in S. cerevisiae, its contribution is masked in the presence of Pdr1p and Pdr3p
YRR1 Activates YOR1 Gene Expression Independent of the PDRE and Its Binding Proteins—We have previously demonstrated that Pdr1p and Pdr3p regulate YOR1 gene expression via directly binding to the single PDRE (Pdr1p/Pdr3p response element) located in the YOR1 promoter [28]

Summary

Introduction

Multiple or pleiotropic drug resistance often arises in the yeast Saccharomyces cerevisiae due to genetic alterations of the functional state of the Cys6-Zn(II) transcription factors Pdr1p and Pdr3p. Gene amplification is typically not seen associated with pleiotropic drug resistance (Pdr) of the yeast Saccharomyces cerevisiae [2], yet these highly drug-resistant strains commonly possess high level transcription of ABC transporter genes like PDR5 [3,4,5] or YOR1 [6, 7]. Overexpression of these ABC transporters is required for the Pdr phenotype and most commonly results from genetic lesions that activate function of the Cys6-Zn(II) transcription factors Pdr1p and Pdr3p [8, 9]. Chromatin immunoprecipitation experiments using these 3X-HA-tagged forms of Yrr1p demonstrate that this protein associates with both the YRR1 and YOR1 promoters in vivo

Methods

Results

Conclusion