Protein engineering of Saccharomyces cerevisiae transporter Pdr5p identifies key residues that impact Fusarium mycotoxin export and resistance to inhibition.
Cereal infection by the broad host range fungal pathogen Fusarium graminearum is a significant global agricultural and food safety issue due to the deposition of mycotoxins within infected grains. Methods to study the intracellular effects of mycotoxins often use the baker's yeast model system (Saccharomyces cerevisiae); however, this organism has an efficient drug export network known as the pleiotropic drug resistance (PDR) network, which consists of a family of multidrug exporters. This study describes the first study that has evaluated the potential involvement of all known or putative ATP-binding cassette (ABC) transporters from the PDR network in exporting the F. graminearum trichothecene mycotoxins deoxynivalenol (DON) and 15-acetyl-deoxynivalenol (15A-DON) from living yeast cells. We found that Pdr5p appears to be the only transporter from the PDR network capable of exporting these mycotoxins. We engineered mutants of Pdr5p at two sites previously identified as important in determining substrate specificity and inhibitor susceptibility. These results indicate that it is possible to alter inhibitor insensitivity while maintaining the ability of Pdr5p to export the mycotoxins DON and 15A-DON, which may enable the development of resistance strategies to generate more Fusarium-tolerant crop plants.
Ottawa Research and Development Centre, Agriculture and Agri-Food Canada, Ottawa, ON, K1A 0C6, Canada.|Ottawa-Carleton Institute of Biology, University of Ottawa, Ottawa, ON, K1N 6N5, Canada.
Amanda B
AB
Gunter
Ottawa Research and Development Centre, Agriculture and Agri-Food Canada, Ottawa, ON, K1A 0C6, Canada.
Anne
A
Hermans
Ottawa Research and Development Centre, Agriculture and Agri-Food Canada, Ottawa, ON, K1A 0C6, Canada.
Whynn
W
Bosnich
Ottawa-Carleton Institute of Biology, University of Ottawa, Ottawa, ON, K1N 6N5, Canada.
Douglas A
DA
Johnson
Ottawa Research and Development Centre, Agriculture and Agri-Food Canada, Ottawa, ON, K1A 0C6, Canada.
Linda J
LJ
Harris
Ottawa Research and Development Centre, Agriculture and Agri-Food Canada, Ottawa, ON, K1A 0C6, Canada.
Steve
S
Gleddie
D018528:ATP-Binding Cassette Transporters / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D019943:Amino Acid Substitution / Q000235:genetics; D002523:Edible Grain / Q000382:microbiology*; D059022:Food Safety; D005670:Fusarium / Q000472:pathogenicity*; D010935:Plant Diseases / Q000382:microbiology; D015202:Protein Engineering; D021381:Protein Transport / Q000235:genetics*; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D029701:Saccharomyces cerevisiae Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D014255:Trichothecenes / Q000378:metabolism*
D018528:ATP-Binding Cassette Transporters; C092438:PDR5 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D014255:Trichothecenes; C046760:15-acetyldeoxynivalenol; C007262:deoxynivalenol

Fusarium graminearum
; ABC transporters; deoxynivalenol; drug resistance; enniatin; yeast