Abundant Recurrent Mitochondrial Mutations and Widespread Mitonuclear Epistasis in Caenorhabditis elegans

We have established Caenorhabditis elegans as a model organism for the investigation of mitonuclear interactions. Leveraging the GPR-1 overexpression construct, we engineered a comprehensive collection of mitonuclear cytoplasmic hybrid (cybrid) strains, amounting to an 18x18 grid consisting of 323 distinct viable genotypes, each a unique mitonuclear genetic combination. We sequenced representative strains and confirmed that they carried the expected mitochondrial and nuclear genotypes. Subsequently, we systematically assessed phenotypes of a subset of these cybrids under varying mitochondrially taxing conditions, including exposure to paraquat, heavy metals, and high temperature. Our findings underscore the prevalence of mitonuclear epistasis across the examined conditions, dwarfing the main effects of mtDNA. Our results support the idea that the mitochondrial genome affects phenotypic variation at a far greater level than expected given its small size, while the pattern of effects provides surprisingly limited support for models of mitonuclear coadaptation. This work will help us disentangle genotype-phenotype links and provide a framework for better understanding the evolutionary processes that shape population structures and local adaptation.