Overton, M. S. | Search bioRxiv

bioRxiv2025

Improved Genotype Inference Reveals Cis- and Trans-Driven Variation in the Loss-of-Heterozygosity Rates in Yeast

Loss-of-heterozygosity (LOH) events are an important source of genetic variation in diploids and are implicated in cancer. LOH-event rates vary across the genome and across genetic backgrounds, but our understanding of this variation is in its infancy. State-of-the-art measurements of LOH rates are obtained from mutation accumulation (MA) experiments in heterozygous hybrids and hinge on the accurate inference of diploid genotypes from short sequencing reads. We analyzed a new large MA dataset in yeast Saccharomyces cerevisiae and found that the currently standard "single-reference" genotyping approach can lead to errors in LOH-rate estimates and produce spurious homolog biases. To address this problem, we develop a novel genotyping approach that is symmetric with respect to both homologs, removes dubious heterozygous markers and corrects for undetected LOH events. We report revised estimates of LOH rates across 12 yeast hybrids, which differ by factors between 0.19 and 5.3 from previously published ones. Our revised estimates do not support the previously reported positive correlation between the rate of terminal LOH events and the hybrid heterozygosity. Finally, our analysis reveals that the 60-fold variation in the rates of interstitial LOH events across yeast hybrids is driven overwhelmingly by genome-wide (trans) genetic factors. In contrast, the 6-fold variation in terminal LOH events is driven by both trans and local (cis) factors. Our results provide a foundation for reliable detection of LOH events and further investigations into the genetic underpinnings of LOH-rate variation.

evolutionary biology↗

bioRxiv2023

Upper bound on the mutational burden imposed by a CRISPR-Cas9 gene-drive element

CRISPR-Cas9 gene drives (CCGDs) are powerful tools for genetic control of wild populations, with applications from disease eradication to species conservation. However, Cas9 alone and in a complex with gRNA can cause double-stranded DNA breaks at off-target sites, which could increase the mutational load and lead to unintended loss-of-heterozygosity (LOH) events. These undesired effects raise potential concerns about the long-term evolutionary safety of CCGDs, but the magnitude of these effects is unknown. To measure how the presence of a CCGD or a Cas9 alone in the genome affects the rates of LOH events and de novo mutations, we carried out a mutation accumulation experiment in yeast Saccharomyces cerevisiae. We found no detectable effects on the genome-wide rates of mutations or LOH events. Our power calculations suggest that CCGD or Cas9 affect these rates by less than 30%, which is much less than natural variation for these traits in yeast. A more detailed examination shows that CCGD or Cas9 may alter the lengths and genomic distributions of LOH events, but the statistical support for these effects is weak. Thus, our results demonstrate that CCGDs impose at most a weak additional mutational burden in the yeast model. Although mutagenic effects of gene drives need to be further evaluated in other systems, our results add credence to the proposition that the evolutionary risks posed by well designed gene drives are likely acceptable.

evolutionary biology↗