In 2014, the coal cleaning chemical 4-methylcyclohexane methanol (MCHM) spilled into the water supply for 300,000 West Virginians. Initial toxicology tests showed relatively mild results, but the underlying effects on cellular biology were underexplored. Treated wildtype yeast cells grew poorly, but there was only a small decrease in cell viability. Cell cycle analysis revealed an absence of cells in S phase within thirty minutes of treatment. Cells accumulated in G1 over a six-hour time course, indicating arrest instead of death. A genetic screen of the haploid knockout collection revealed 329 high confidence genes required for optimal growth in MCHM. These genes encode three major cell processes: mitochondrial gene expression/translation, the vacuolar ATPase, and aromatic amino acid biosynthesis. The transcriptome showed an upregulation of pleiotropic drug response genes and amino acid biosynthetic genes and downregulation in ribosome biosynthesis. Analysis of these datasets pointed to environmental stress response activation upon treatment. Overlap in datasets included the aromatic amino acid genes ARO1, ARO3, and four of the five TRP genes. This implicated nutrient deprivation as the signal for stress response. Excess supplementation of nutrients and amino acids did not improve growth on MCHM, so the source of nutrient deprivation signal is still unclear. Reactive oxygen species and DNA damage were directly detected with MCHM treatment, but timepoints showed these accumulated slower than cells arrested. We propose that wildtype cells arrest from nutrient deprivation and survive, accumulating oxidative damage through the implementation of robust environmental stress responses.
Keywords: MCHM; Saccharomyces cerevisiae; aromatic amino acids; environmental stress response; genetic screen; nutrient starvation; reactive oxygen species; tryptophan.
Copyright © 2020 Ayers et al.