Unstable mitochondrial heteroplasmy in Mytilus edulis primary cell cultures

Doubly uniparental inheritance is a phenomenon unique to bivalve mollusks characterized by the inheritance of mitochondria from both parents by male embryos and the maintenance of paternal mitochondria in male gonads. This lineage-specific mechanism has been described in several bivalve species, where male mitochondrial DNA is also often found in somatic tissues. We exploited the development of a primary cell culture model for the blue mussel Mytilus edulis to investigate mitochondrial DNA dynamics at the cellular level and over time, using cells dissociated from male and female mantle (gonadal) and gill (somatic) tissues. We employed quantitative polymerase chain reactions to monitor fluctuations in the relative abundance of male and female mitochondrial DNAs. In male mantle cells, the amount of male mtDNA increased over time in culture across all tested samples. In male gill cells, male mtDNA was detectable in approximately 50% of the samples at the initial time point and generally decreased over time. No heteroplasmic female could be detected in this study. Illumina sequencing, comparing the proportion of the sex-linked mtDNAs between male mantle tissue and derived cultured cells, revealed a sharp increase in the proportion of male mtDNA in cultured mantle cells. Finally, immunofluorescence experiments using antibodies against open reading frames (ORFs) encoded by male or female mtDNA genomes showed that these peptides are present in the mitochondria of all male mantle cells after 10 days in culture. Overall, these findings support the idea that heteroplasmy is maintained in male germ cells in culture, whereas heteroplasmic cells in somatic tissues tend to be scarce and are lost over time.