Heterologous expression of a pleiotropic drug resistance transporter from Phytophthora sojae in yeast transporter mutants.
A system for the expression of an ATP binding cassette (ABC) transporter from the soybean pathogen Phytophthora sojae is described. Pdr1, an ABC transporter with homology to the pleiotropic drug resistance (PDR) family of transporters, was cloned by primer walking from a P. sojae genomic library. Reverse transcriptase PCR assays showed that the transcript disappeared after encystment of zoospores and was not detected in hyphal germlings in dilute salts, in hyphae growing in liquid V8 media, or in tissue extracts from infected hypocotyls. BLAST analysis of Pdr1 against the P. sojae EST database also revealed that this gene was present only in zoospore libraries. Comparison of the number of hits to Pdr1 with that of a set of housekeeping genes revealed that Pdr1 was expressed at rates two- to threefold higher than other transcripts. To test the hypothesis that Pdr1p functions as a broad substrate membrane transporter, Pdr1 was transformed into yeast mutants deficient in several drug resistance transporters. Yeast mutants transformed with Pdr1 possessed partial drug resistance against only 5 of 17 chemically distinct compounds. Thus, when expressed in yeast, this transporter has a significantly narrower substrate specificity in comparison to the yeast transporters, Pdr5p, Yorlp, and Snq2p.
Connolly
Mary S
MS
Biology Sciences, Bowling Green State University, OH, USA.
Sakihama
Yasuko
Y
Phuntumart
Vipaporn
V
Jiang
Yinjun
Y
Warren
Franklin
F
Mourant
Lindsay
L
Morris
Paul F
PF
D000595:Amino Acid Sequence; D004268:DNA-Binding Proteins / Q000235:genetics*; D026141:Drug Resistance, Multiple, Fungal / Q000235:genetics*; D015966:Gene Expression Regulation, Fungal; D026901:Membrane Transport Proteins / Q000235:genetics*; D008969:Molecular Sequence Data; D009154:Mutation; D009868:Oomycetes / Q000235:genetics; D010838:Phytophthora / Q000166:cytology / Q000235:genetics*; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics*; D016415:Sequence Alignment; D015534:Trans-Activators / Q000235:genetics*; D014157:Transcription Factors
D004268:DNA-Binding Proteins; D026901:Membrane Transport Proteins; C085480:PDR1 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D015534:Trans-Activators; D014157:Transcription Factors