Negative transcriptional regulation of multidrug resistance gene expression by an Hsp70 protein.
One of the most common origins of multidrug resistance occurs via the overproduction of ATP-binding cassette (ABC) transporter proteins. These ABC transporters then act as broad specificity drug pumps and efflux a wide range of toxic agents out of the cell. The yeast Saccharomyces cerevisiae exhibits multiple or pleiotropic drug resistance (Pdr) often through the over-production of a plasma membrane-localized ABC transporter protein called Pdr5p. Expression of the PDR5 gene is controlled by two zinc cluster-containing transcription factors called Pdr1p and Pdr3p. Cells that lack their mitochondrial genome (rho(0) cells) strongly induce PDR5 transcription in a Pdr3p-dependent fashion. To identify proteins associated with Pdr3p that might act to regulate this factor, a tandem affinity purification (TAP) moiety was fused to Pdr3p, and this recombinant protein was purified from yeast cells. The cytosolic Hsp70 chaperone Ssa1p co-purified with TAP-Pdr3p. Overexpression of Ssa1p repressed expression of PDR5 but had no effect on expression of other genes involved in the Pdr phenotype. This Ssa1p-mediated repression required the presence of Pdr3p and did not influence Pdr1p-dependent gene expression. Loss of the nucleotide exchange factor Fes1p mimicked Ssa1p-mediated repression of PDR5. Co-immunoprecipitation experiments indicated that Ssa1p was associated with Pdr3p but not Pdr1p in yeast cells. Finally, rho(0) cells had less Ssa1p bound to Pdr3p than rho(+) cells, consistent with Ssa1p-mediated repression of Pdr3p activity serving as a key regulatory step in control of multidrug resistance in yeast.
Shahi
Puja
P
Department of Molecular Physiology and Biophysics, University of Iowa, Iowa City, Iowa 52242.
Gulshan
Kailash
K
Department of Molecular Physiology and Biophysics, University of Iowa, Iowa City, Iowa 52242.
Moye-Rowley
W Scott
WS
Department of Molecular Physiology and Biophysics, University of Iowa, Iowa City, Iowa 52242. Electronic address: scott-moyerowley@uiowa.edu.
D018528:ATP-Binding Cassette Transporters / Q000235:genetics*; D000251:Adenosine Triphosphatases / Q000502:physiology*; D004268:DNA-Binding Proteins / Q000235:genetics*; D026141:Drug Resistance, Multiple, Fungal / Q000235:genetics*; D015966:Gene Expression Regulation, Fungal; D018840:HSP70 Heat-Shock Proteins / Q000502:physiology*; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics* / Q000502:physiology*; D014157:Transcription Factors / Q000235:genetics*; D014158:Transcription, Genetic
D018528:ATP-Binding Cassette Transporters; D004268:DNA-Binding Proteins; D018840:HSP70 Heat-Shock Proteins; C089261:PDR3 protein, S cerevisiae; C092438:PDR5 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; D000251:Adenosine Triphosphatases; C095973:SSA1 protein, S cerevisiae