Understanding the molecular mechanisms of killer toxin defence in the budding yeast Saccharomyces cerevisiae

A/B toxins are a well-characterized family of toxins—including cholera, Shiga, and ricin toxins—that cause severe disease and significant mortality worldwide. Due to their pathogenic impact, there is growing interest in understanding mechanisms related to A/B toxin biology and identifying potential therapeutic targets. A recent discovery in yeast has revealed a defence factor, Ktd1 (Killer toxin defence factor), that provides resistance against K28, a yeast-secreted A/B toxin. As Saccharomyces cerevisiae is a eukaryotic model organism and naturally harbours a defence factor against its own A/B toxin, it presents a valuable system to study toxin resistance and regulatory mechanisms relevant to higher organisms. As Ktd1 was only recently identified protein, very little is known about how it is regulated. This study investigates whether Ktd1 function is modulated by post-translational phosphorylation. To test this, we optimized two complementary assays and screened S. cerevisiae phosphatase and kinase mutant strains for sensitivity to K28. Of the strains screened, 7 phosphatases and 28 kinases showed potential sensitivity in the primary assay. After validation using a more sensitive assay, the candidates were narrowed to 5 phosphatases (glc7-DAmP, sit4Δ, yvh1Δ, ssu72-DAmP, and ptc4Δ) and 22 kinases, including (ctk3Δ, ctk2Δ, bud23Δ, hog1Δ, pbs2Δ, skm1Δ, bck1Δ, and ste11Δ). Complementation of mutants by Ktd1 overexpression was used to prioritise enzymes that potentially co-function with Ktd1. Initial fluorescence microscopy work on the kinase Hog1 and the phosphatase Glc7 suggest they may be involved in Ktd1-mediated defence. Understanding how Ktd1 is modulated could offer insights into similar defence systems in higher eukaryotes and support the development of strategies to combat A/B toxin-related diseases in humans.