A quick and reliable method for genome-wide host factor screening of Saccharomyces cerevisiae killer toxins

Abstract

Numerous yeasts produce toxic compounds to fight the competitors. Such compounds include small molecules (like antibiotics), antibiotic peptides, and also larger proteins, including killer toxins. Their ability to affect the cell depends on the host factors modulating the killing activity. Here we describe a robotics-based method to advance the genome-wide screening for the host factors affecting sensitivity of budding yeast to the killer toxins using the K2 system as the model. We demonstrate that arraying the mutant library on the agar plates containing the K2 killer toxinproducing strain and/or purified toxin ("survival" assay) increases the sensitivity and speed of the screen and decreases the costs compared to the traditional "killer" assay. We show the applicability of a new screening method of searching for the host factors using a killer strain isolated from agricultural plant environment. In addition, the "survival" assay allows identification of previously undetected factors that could be the "missing links" in the pathways of toxininduced cellular responses.

The antifungal capacity of a 681-membered collection of environmental yeast isolates

“…Most toxins have been reported to be secreted in rather low amounts. [131][132][133][134] Thus the reported target range is only the range as it appears from our assay. Although we show examples where concentrating the spent media and hence the active proteins helps in defining the target range, we argue that our assay is still suitable to report a certain specificity in targeting (Supplementary Figures 5B and 6).…”