The absence of canonical respiratory complex I subunits in male-type mitogenomes of three Donax species.

Bivalves are an extraordinary class of animals in which species with a doubly uniparental inheritance (DUI) of mitochondrial DNA have been described. DUI is characterized as a mitochondrial homoplasmy of females and heteroplasmy of male individuals where F-type mitogenomes are passed to the progeny with mother egg cells and divergent M-type mitogenomes are inherited with fathers sperm cells. However, in most cases only male individuals retain divergent mitogenome inherited with spermatozoa. Additionally, in many of bivalves, unique mitochondrial features, like additional genes, gene duplication, gene extensions, mitochondrial introns, and recombination, were observed. In this study, we sequenced and assembled male-type mitogenomes of three Donax species. Comparative analysis of mitochondrial sequences revealed a lack of all seven NADH dehydrogenase subunits as well as the presence of three long additional open reading frames lacking identifiable homology to any of the existing genes.

Confirmation of the first intronic sequence in the bivalvian mitochondrial genome of Macoma balthica (Linnaeus, 1758).

In 2020, the first male-type mitochondrial genome from the clam Macoma balthica was published. Apart from the unusual doubly uniparental inheritance of mtDNA, scientists observed a unique (over 4k bp long) extension in the middle of the cox2 gene. We have attempted to replicate these data by NGS DNA sequencing and explore further the expression of the long cox2 gene. In our study, we report an even longer cox2 gene (over 5.5 kbp) with no stop codon separating conserved cox2 domains, as well as, based on the rtPCR, a lower relative gene expression pattern of the middle part of the gene (5' = 1; mid = 0.46; 3' = 0.89). Lastly, we sequenced the cox2 gene transcript proving the excision of the intronic sequence.

No evidence of DUI in the Mediterranean alien species Brachidontes pharaonis (P. Fisher, 1870) despite mitochondrial heteroplasmy.

Two genetically different mitochondrial haplogroups of Brachidontes pharaonis (p-distance 6.8%) have been identified in the Mediterranean Sea. This hinted at a possible presence of doubly uniparental inheritance in this species. To ascertain this possibility, we sequenced two complete mitogenomes of Brachidontes pharaonis mussels and performed a qPCR analysis to measure the relative mitogenome copy numbers of both mtDNAs. Despite the presence of two very similar regions composed entirely of repetitive sequences in the two haplogroups, no recombination between mitogenomes was detected. In heteroplasmic individuals, both mitogenomes were present in the generative tissues of both sexes, which argues against the presence of doubly uniparental inheritance in this species.

Semimytilus algosus: first known hermaphroditic mussel with doubly uniparental inheritance of mitochondrial DNA.

Doubly uniparental inheritance (DUI) of mitochondrial DNA is a rare phenomenon occurring in some freshwater and marine bivalves and is usually characterized by the mitochondrial heteroplasmy of male individuals. Previous research on freshwater Unionida mussels showed that hermaphroditic species do not have DUI even if their closest gonochoristic counterparts do. No records showing DUI in a hermaphrodite have ever been reported. Here we show for the first time that the hermaphroditic mussel Semimytilus algosus (Mytilida), very likely has DUI, based on the complete sequences of both mitochondrial DNAs and the distribution of mtDNA types between male and female gonads. The two mitogenomes show considerable divergence (34.7%). The presumably paternal M type mitogenome dominated the male gonads of most studied mussels, while remaining at very low or undetectable levels in the female gonads of the same individuals. If indeed DUI can function in the context of simultaneous hermaphroditism, a change of paradigm regarding its involvement in sex determination is needed. It is apparently associated with gonadal differentiation rather than with sex determination in bivalves.

The ORF in the control region of the female-transmitted Mytilus mtDNA codes for a protein.

Bivalve species with doubly uniparental inheritance of mitochondria have been shown to contain additional mtDNA-encoded proteins suspected to be involved in sex-specific transmission of the female (F) and male (M) mitochondrial genomes. This is true for freshwater mussels and marine clams but was still unclear for marine mussel Mytilus spp. Here we present evidence that a F mtDNA-specific open reading frame (ORF) identified in the control region of M. edulis codes for a protein. The protein was detected, using western blots, in both female and male mantle tissues, which contain the gonads. The protein was also localized, using immunochemistry, in sperm mitochondria.

Mitogenomics of Perumytilus purpuratus (Bivalvia: Mytilidae) and its implications for doubly uniparental inheritance of mitochondria.

Animal mitochondria are usually inherited through the maternal lineage. The exceptional system allowing fathers to transmit their mitochondria to the offspring exists in some bivalves. Its taxonomic spread is poorly understood and new mitogenomic data are needed to fill the gap. Here, we present for the first time the two divergent mitogenomes from Chilean mussel Perumytilus purpuratus. The existence of these sex-specific mitogenomes confirms that this species has the doubly uniparental inheritance (DUI) of mitochondria. The genetic distance between the two mitochondrial lineages in P. purpuratus is not only much bigger than in the Mytilus edulis species complex but also greater than the distance observed in Musculista senhousia, the only other DUI-positive member of the Mytilidae family for which both complete mitochondrial genomes were published to date. One additional, long ORF (open reading frame) is present exclusively in the maternal mitogenome of P. purpuratus. This ORF evolves under purifying selection, and will likely be a target for future DUI research.

Actively transcribed and expressed atp8 gene in Mytilus edulis mussels.

BACKGROUND: Animal mitochondrial genomes typically encode 37 genes: 13 proteins, 22 tRNAs and two rRNAs. However, many species represent exceptions to that rule. Bivalvia along with Nematoda and Platyhelminthes are often suspected to fully or partially lack the ATP synthase subunit 8 (atp8) gene. This raises the question as to whether they are really lacking this gene or is this maybe an annotation problem? Among bivalves, Mytilus edulis has been inferred to lack an ATP8 gene since the characterization of its mitochondrial genome in 1992. Even though recent bioinformatic analyses suggested that atp8 is present in Mytilus spp., due to high divergence in predicted amino acid sequences, the existence of a functional atp8 gene in this group remains controversial. RESULTS: Here we demonstrate that M. edulis mitochondrial open reading frames suggested to be atp8 (in male and female mtDNAs) are actively translated proteins. We also provide evidence that both proteins are an integral part of the ATP synthase complex based on in-gel detection of ATP synthase activity and two-dimensional Blue-Native and SDS polyacrylamide electrophoresis. CONCLUSION: Many organisms (e.g., Bivalvia along with Nematoda and Platyhelminthes) are considered to be lacking certain mitochondrial genes often only based on poor similarity between protein coding gene sequences in genetically closed species. In some situations, this may lead to the inference that the ATP8 gene is absent, when it is in fact present, but highly divergent. This shows how important complementary role protein-based approaches, such as those in the present study, can provide to bioinformatic, genomic studies (i.e., ability to confirm the presence of a gene).

Mitochondrial genomes of the key zooplankton copepods Arctic Calanus glacialis and North Atlantic Calanus finmarchicus with the longest crustacean non-coding regions.

We determined the nearly complete mitochondrial genomes of the Arctic Calanus glacialis and its North Atlantic sibling Calanus finmarchicus, which are key zooplankton components in marine ecosystems. The sequenced part of C. glacialis mitogenome is 27,342 bp long and consists of two contigs, while for C. finmarchicus it is 29,462 bp and six contigs, what makes them the longest reported copepod mitogenomes. The typical set of metazoan mitochondrial genes is present in these mitogenomes, although the non-coding regions (NCRs) are unusually long and complex. The mitogenomes of the closest species C. glacialis and C. finmarchicus, followed by the North Pacific C. sinicus, are structurally similar and differ from the much more typical of deep-water, Arctic C. hyperboreus. This evolutionary trend for the expansion of NCRs within the Calanus mitogenomes increases mitochondrial DNA density, what resulted in its similar density to the nuclear genome. Given large differences in the length and structure of C. glacialis and C. finmarchicus mitogenomes, we conclude that the species are genetically distinct and thus cannot hybridize. The molecular resources presented here: the mitogenomic and rDNA sequences, and the database of repetitive elements should facilitate the development of genetic markers suitable in pursuing evolutionary research in copepods.