Ethanol is a ubiquitous environmental stressor that is toxic to all lifeforms. Here, we use the model eukaryote Saccharomyces cerevisiae to show that exposure to sublethal ethanol concentrations causes DNA replication stress and an increased mutation rate. Specifically, we find that ethanol slows down replication and affects localization of Mrc1, a conserved protein that helps stabilize the replisome. In addition, ethanol exposure also results in the recruitment of error-prone DNA polymerases to the replication fork. Interestingly, preventing this recruitment through mutagenesis of the PCNA/Pol30 polymerase clamp or deleting specific error-prone polymerases abolishes the mutagenic effect of ethanol. Taken together, this suggests that the mutagenic effect depends on a complex mechanism, where dysfunctional replication forks lead to recruitment of error-prone polymerases. Apart from providing a general mechanistic framework for the mutagenic effect of ethanol, our findings may also provide a route to better understand and prevent ethanol-associated carcinogenesis in higher eukaryotes.

Alcohol consumption is an underestimated risk factor for the development of precancerous lesions in the oral cavity. Although alcohol is a well-accepted recreational drug, 26.4% of all lip and oral cavity cancers worldwide are related to heavy drinking. Molecular mechanisms underlying this carcinogenic effect of ethanol are still under investigation. An important damaging effect comes from the first metabolite of ethanol, being acetaldehyde. Concentrations of acetaldehyde detected in the oral cavity are relatively high due to the metabolization of ethanol by oral microbes. Acetaldehyde can directly damage the DNA by the formation of mutagenic DNA adducts and interstrand crosslinks. Additionally, ethanol is known to affect epigenetic methylation and acetylation patterns, which are important regulators of gene expression. Ethanol-induced hypomethylation can activate the expression of oncogenes which subsequently can result in malignant transformation. The recent identification of ethanol-related mutational signatures emphasizes the role of acetaldehyde in alcohol-associated carcinogenesis. However, not all signatures associated with alcohol intake also relate to acetaldehyde. This finding highlights that there might be other effects of ethanol yet to be discovered.