Resistance of mitochondrial DNA to chemical-induced germline mutations is independent of mitophagy in C. elegans

Mitochondrial DNA (mtDNA) is prone to mutation in aging and over evolutionary time, yet the processes that regulate the accumulation of de novo mtDNA mutations and modulate mtDNA heteroplasmy are not fully elucidated. Mitochondria lack certain DNA repair processes, which could contribute to polymerase error-induced mutations and increase susceptibility to chemical-induced mtDNA mutagenesis. We conducted error-corrected, ultra-sensitive Duplex Sequencing to investigate the effects of two known nuclear genome mutagens, cadmium and Aflatoxin B1, on germline mtDNA mutagenesis in Caenorhabditis elegans . After 1,750 generations, we detected 2,270 single nucleotide mutations. Heteroplasmy is pervasive in C. elegans and mtDNA mutagenesis is dominated by C:G A:T mutations generally attributed to oxidative damage, yet there was no effect of either exposure on mtDNA mutation frequency, spectrum, or trinucleotide context signature. Mitophagy may play a role in eliminating mtDNA damage or deleterious mutations, and mitophagy-deficient mutants pink-1 and dct-1 accumulated significantly higher levels of mtDNA damage compared to wild-type C. elegans after exposures. However, there were only small differences in overall mutation frequency, spectrum, or trinucleotide context signature compared to wild-type after 3,050 generations, across all treatments. These findings suggest mitochondria harbor additional previously uncharacterized mechanisms that regulate mtDNA mutational processes across generations. ### Competing Interest Statement S.R.K. is an equity holder and paid consultant for Twinstrand Biosciences, a for-profit company commercializing Duplex Sequencing. No Twinstrand products were used in the generation of the data.