A yeast cell-based system for screening Candida glabrata multidrug resistance reversal agents and selection of loss-of-function pdr1 mutants.
In the pathogenic yeast Candida glabrata, multidrug resistance is associated with the overexpression of drug efflux pumps caused by gain-of-function mutations in the CgPDR1 gene. CgPdr1p transcription factor, which activates the expression of several drug efflux transporter genes, is considered to be a promising target for compounds sensitizing the multidrug-resistant yeast cells. Here, we describe a cell-based screening system for detecting the inhibitory activity of compounds interfering with the CgPdr1p function in a heterologous genetic background of the hypersensitive Saccharomyces cerevisiae mutant strain. The screening is based on the ability to abrogate the growth defect of cells suffering from the galactose-induced and CgPdr1p-driven overexpression of a dominant lethal pma1(D378N) allele placed under the control of the ScPDR5 promoter. The system allows rapid identification of multidrug resistance reversal agents inhibiting the CgPdr1p activity or loss-of-function Cgpdr1 mutations, and is amenable to high-throughput screening on solid or liquid media.
Department of Microbiology and Virology, Faculty of Natural Sciences, Comenius University in Bratislava, Bratislava, Slovak Republic.
Eduard
E
Goffa
Alexandra
A
Bialkova
Monika
M
Batova
Vladimira
V
Dzugasova
Julius
J
Subik
D041221:Candida glabrata / Q000187:drug effects* / Q000235:genetics; D004353:Drug Evaluation, Preclinical / Q000379:methods*; D026141:Drug Resistance, Multiple, Fungal / Q000187:drug effects*; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism*; D017353:Gene Deletion; D057166:High-Throughput Screening Assays; D026901:Membrane Transport Proteins / Q000235:genetics / Q000378:metabolism*; D050296:Microbial Viability / Q000187:drug effects; D006180:Proton-Translocating ATPases / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000187:drug effects / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism
D005656:Fungal Proteins; D026901:Membrane Transport Proteins; D029701:Saccharomyces cerevisiae Proteins; C092477:PMA1 protein, S cerevisiae; D006180:Proton-Translocating ATPases