Introduction: Maintenance of genome integrity is critical to the cell, however exogenous and endogenous stresses menace the integrity of the genome. Overwhelming a cell’s DNA repair capacity can expose the cell to the accumulation of mutations and, in humans, predispose to cancer development. Understanding the consequences of defective DNA repair pathways and their association with specific mutational processes in tumor genome sequences is difficult to achieve. A powerful method to study the effect of defects in DNA repair mechanisms is the extraction of mutational signatures from whole-genome sequencing of model organisms carrying defined genetic alterations or exposed to mutagens. We hypothesized that mutation accumulation in S. cerevisiae caused by deletions of RAD1, MUS81, or SGS1 genes, coupled with exposure to cisplatin will result in specific mutational profiles that can be used to inform repair mechanism and potentially mutation signatures in cancer.
Methods: We sequenced whole genomes of 104 populations of diploid S. cerevisiae that were WT or carried homozygous deletions of RAD1, MUS81, or SGS1. For each genetic background 12 parallel lines were propagated for 1000 generations through single-cell bottlenecks or exposed for 3 hours to 100 uM cisplatin. Genomic DNA was isolated from single clones for whole-genome sequencing.
Results and conclusions: We identified 985 SNVs, 123 indels, 107 CNVs, and 36 aneuploidies. Rad1-deficient genomes exposed to cisplatin were characterized by an increase in single nucleotide substitutions at C/G base pairs. C to N mutations were defined by a GpCpN sequence context, which indicates that the surrounding sequence influences the base to be inserted on the opposite strand. Moreover, these genomes featured dinucleotide substitutions, particularly CpT to TpA. This data reflects the chemistry of cisplatin and indicates Rad1/XPF critically protects the cell against platinum hypermutation. Sgs1-deficient genomes propagated for 1000 generations revealed CNV events with the majority of breakpoints mapped to transposable element loci and other genetic features with high transcriptional frequency. Overall the data indicates the role of Sgs1/BLM in maintaining genomic regions difficult to replicate like retroelements. Together these data define new mutation signatures for DNA repair deficient cells whose mechanism are now under investigation.