Trans-generational maintenance of mitochondrial DNA integrity in oocytes during early folliculogenesis

Mutations in mitochondrial DNA (mtDNA) can lead to mitochondrial and cellular dysfunction. However, recent studies suggest that purifying selection acts against mutant mtDNAs during transgenerational transmission. We investigated the mtDNA dynamics during ovarian follicle development. Using base-editing, we generated mice harboring a 3177 G > A mutation corresponding to the human Leber hereditary optic neuropathy (LHON)-related mtDNA mutation and confirmed a transgenerational reduction of the mutant mtDNA. Utilizing a mouse follicle culture system in which pathogenic mtDNA mutations were introduced in vitro, followed by mtDNA sequencing and digital PCR, we found that the germline heteroplasmy shift during early folliculogenesis was driven by a decrease in mutant mtDNA along with compensatory replication of wild-type mtDNA. In contrast, synonymous mtDNA mutations did not affect mtDNA dynamics. These findings demonstrate that mice can eliminate certain pathogenic mtDNA mutations in the germline during early folliculogenesis, thus advancing our understanding of mtDNA purifying selection during oogenesis. Furthermore, our use of mtDNA editing in in vitro-cultured follicles provides a novel approach to create and monitor mitochondrial DNA mutations.