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Heterologous expression of a pleiotropic drug resistance transporter from Phytophthora sojae in yeast transporter mutants

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Abstract

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.

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Acknowledgements

We thank Annabelle Decottignies and André Goffeau for their generous gift of plasmid and yeast strains. The technical assistance of Kena Allison, Kevin Fuller, Orlando Vega and Lauren Zimmerman on some of these experiments is greatly appreciated. Financial support was provided by the Ohio Plant Biotechnology Consortium, a Faculty Research Incentives grant and additional support was provided by the Center for Biomolecular Sciences at Bowling Green State University (BGSU) to PFM. MC was supported by a dissertation fellowship from the Graduate College at BGSU. YS was supported by a visiting fellowship from the Ministry of Education, Culture, Sports, Science and Technology of Japan.

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  1. Mary S. Connolly

    Present address: Department of Biology, University of Dayton, Dayton, OH, 45469, USA

  2. Yasuko Sakihama

    Present address: Laboratory of Ecological Chemistry, Faculty of Agriculture, Hokkaido University, Sapporo, 060-8589, Japan

  3. Yinjun Jiang

    Present address: Department of Experimental Radiation Oncology, The University of Texas M.D. Anderson Cancer Center, Houston, TX, 77030, USA

Authors and Affiliations

  1. Biology Sciences, Bowling Green State University, Bowling Green, OH, 4340, USA

    Mary S. Connolly, Yasuko Sakihama, Vipaporn Phuntumart, Yinjun Jiang, Franklin Warren, Lindsay Mourant & Paul F. Morris

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  1. Mary S. Connolly
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Correspondence to Paul F. Morris.

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Communicated by S. Hohmann

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Connolly, M.S., Sakihama, Y., Phuntumart, V. et al. Heterologous expression of a pleiotropic drug resistance transporter from Phytophthora sojae in yeast transporter mutants. Curr Genet 48, 356–365 (2005). https://doi.org/10.1007/s00294-005-0015-4

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