ABSTRACT
Mitochondrial (mt) function depends critically on optimal interactions between components encoded by mt and nuclear DNAs. mitochondrial DNA (mtDNA) inheritance (SMI) is thought to have evolved in animal species to maintain mito-nuclear complementarity by preventing the spread of selfish mt elements thus typically rendering mtDNA heteroplasmy evolutionarily ephemeral. Here, we show that mtDNA intraorganismal heteroplasmy can have deterministic underpinnings and persist for hundreds of millions of years. We demonstrate that the only exception to SMI in the animal kingdom, that is, the doubly uniparental mtDNA inheritance system in bivalves, with its three-way interactions among egg mt-, sperm mt- and nucleus-encoded gene products, is tightly associated with the maintenance of separate male and female sexes (dioecy) in freshwater mussels. Specifically, this mother-through-daughter and father-through-son mtDNA inheritance system, containing highly differentiated mt genomes, is found in all dioecious freshwater mussel species. Conversely, all hermaphroditic species lack the paternally transmitted mtDNA (=possess SMI) and have heterogeneous macromutations in the recently discovered, novel protein-coding gene (F-orf) in their maternally transmitted mt genomes. Using immunoelectron microscopy, we have localized the F-open reading frame (ORF) protein, likely involved in specifying separate sexes, in mitochondria and in the nucleus. Our results support the hypothesis that proteins coded by the highly divergent maternally and paternally transmitted mt genomes could be directly involved in sex determination in freshwater mussels. Concomitantly, our study demonstrates novel features for animal mt genomes: the existence of additional, lineage-specific, mtDNA-encoded proteins with functional significance and the involvement of mtDNA-encoded proteins in extra-mt functions. Our results open new avenues for the identification, characterization, and functional analyses of ORFs in the intergenic regions, previously defined as "noncoding," found in a large proportion of animal mt genomes.
Subject(s)
DNA, Mitochondrial/genetics , Mitochondrial Proteins/genetics , Sex Determination Analysis/methods , Unionidae/genetics , Amino Acid Sequence , Animals , Chromosome Mapping , Electron Transport Complex IV/genetics , Electron Transport Complex IV/metabolism , Female , Fresh Water , Likelihood Functions , Male , Mitochondrial Proteins/metabolism , Molecular Sequence Data , Open Reading Frames , Ovum/metabolism , Phylogeny , Protein Structure, Secondary , Sequence Analysis, DNA , Sex Characteristics , Transcription, Genetic , Unionidae/classification , Unionidae/metabolismABSTRACT
Our previous study documented a reproductive function for the male-transmitted mitochondrial DNA (mtDNA)-encoded cytochrome c oxidase subunit II (MCOX2) protein in a unionoid bivalve. Here, immunoblotting, immunohistochemistry and immunoelectron microscopy analyses demonstrate that the female-transmitted protein (FCOX2) is: (i) expressed in both male and female gonads; (ii) maximally expressed in ovaries just prior to the time of the annual fertilization event; (iii) displayed in the cytoplasm and more strongly in the plasma membrane (microvilli), vitelline matrix and vitelline envelope of mature ovarian eggs; and (iv) strongly localized to the vitelline matrix of some eggs just prior to fertilization. These findings represent evidence for the extra-mitochondrial localization of an mtDNA-encoded gene product and are consistent with multifunctionality for FCOX2 in eggs.
