Live-cell assays reveal selectivity and sensitivity of the multidrug response in budding yeast

Pleiotropic drug resistance arises by the enhanced extrusion of bioactive molecules and is present in a wide range of organisms, ranging from fungi to human cells. A key feature of this adaptation is the sensitive detection of intracellular xenobiotics by transcriptional activators, activating expression of multiple drug exporters. Here, we investigated the selectivity and sensitivity of the budding yeast (Saccharomyces cerevisiae) multidrug response to better understand how differential drug recognition leads to specific activation of drug exporter genes and to drug resistance. Applying live-cell luciferase reporters, we demonstrate that the SNQ2, PDR5, PDR15, and YOR1 transporter genes respond to different mycotoxins, menadione, and hydrogen peroxide in a distinguishable manner and with characteristic amplitudes, dynamics, and sensitivities. These responses correlated with differential sensitivities of the respective transporter mutants to the specific xenobiotics. We further establish a binary vector system, enabling quantitative determination of xenobiotic-transcription factor (TF) interactions in real time. Applying this system we found that the TFs Pdr1, Pdr3, Yrr1, Stb5, and Pdr8 have largely different drug recognition patterns. We noted that Pdr1 is the most promiscuous activator, whereas Yrr1 and Stb5 are selective for ochratoxin A and hydrogen peroxide, respectively. We also show that Pdr1 is rapidly degraded after xenobiotic exposure, which leads to a desensitization of the Pdr1-specific response upon repeated activation. The findings of our work indicate that in the yeast multidrug system, several transcriptional activators with distinguishable selectivities trigger differential activation of the transporter genes.