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Maternally inherited mitochondrial (mt)DNA mutations can cause fatal or severely debilitating syndromes in children, with disease severity dependent on the specific gene mutation and the ratio of mutant to wild-type mtDNA (heteroplasmy) in each cell and tissue. Pathogenic mtDNA mutations are relatively common, with an estimated 778 affected children born each year in the United States. Mitochondrial replacement therapies or techniques (MRT) circumventing mother-to-child mtDNA disease transmission involve replacement of oocyte maternal mtDNA. Here we report MRT outcomes in several families with common mtDNA syndromes. The mother's oocytes were of normal quality and mutation levels correlated with those in existing children. Efficient replacement of oocyte mutant mtDNA was performed by spindle transfer, resulting in embryos containing >99% donor mtDNA. Donor mtDNA was stably maintained in embryonic stem cells (ES cells) derived from most embryos. However, some ES cell lines demonstrated gradual loss of donor mtDNA and reversal to the maternal haplotype. In evaluating donor-to-maternal mtDNA interactions, it seems that compatibility relates to mtDNA replication efficiency rather than to mismatch or oxidative phosphorylation dysfunction. We identify a polymorphism within the conserved sequence box II region of the D-loop as a plausible cause of preferential replication of specific mtDNA haplotypes. In addition, some haplotypes confer proliferative and growth advantages to cells. Hence, we propose a matching paradigm for selecting compatible donor mtDNA for MRT.

Although previous studies have documented a bottleneck in the transmission of mtDNA genomes from mothers to offspring, several aspects remain unclear, including the size and nature of the bottleneck. Here, we analyze the dynamics of mtDNA heteroplasmy transmission in the Genomes of the Netherlands (GoNL) data, which consists of complete mtDNA genome sequences from 228 trios, eight dizygotic (DZ) twin quartets, and 10 monozygotic (MZ) twin quartets. Using a minor allele frequency (MAF) threshold of 2%, we identified 189 heteroplasmies in the trio mothers, of which 59% were transmitted to offspring, and 159 heteroplasmies in the trio offspring, of which 70% were inherited from the mothers. MZ twin pairs exhibited greater similarity in MAF at heteroplasmic sites than DZ twin pairs, suggesting that the heteroplasmy MAF in the oocyte is the major determinant of the heteroplasmy MAF in the offspring. We used a likelihood method to estimate the effective number of mtDNA genomes transmitted to offspring under different bottleneck models; a variable bottleneck size model provided the best fit to the data, with an estimated mean of nine individual mtDNA genomes transmitted. We also found evidence for negative selection during transmission against novel heteroplasmies (in which the minor allele has never been observed in polymorphism data). These novel heteroplasmies are enhanced for tRNA and rRNA genes, and mutations associated with mtDNA diseases frequently occur in these genes. Our results thus suggest that the female germ line is able to recognize and select against deleterious heteroplasmies.

The maternally inherited mitochondrial genome (mtDNA) is present in multimeric form within cells and harbors sequence variants (heteroplasmy). While a single mtDNA variant at high load can cause disease, naturally occurring variants likely persist at low levels across generations of healthy populations. To determine how naturally occurring variants are segregated and transmitted, we generated a mini-pig model, which originates from the same maternal ancestor. Following next-generation sequencing, we identified a series of low-level mtDNA variants in blood samples from the female founder and her daughters. Four variants, ranging from 3% to 20%, were selected for validation by high-resolution melting analysis in 12 tissues from 31 animals across three generations. All four variants were maintained in the offspring, but variant load fluctuated significantly across the generations in several tissues, with sex-specific differences in heart and liver. Moreover, variant load was persistently reduced in high-respiratory organs (heart, brain, diaphragm, and muscle), which correlated significantly with higher mtDNA copy number. However, oocytes showed increased heterogeneity in variant load, which correlated with increased mtDNA copy number during in vitro maturation. Altogether, these outcomes show that naturally occurring mtDNA variants segregate and are maintained in a tissue-specific manner across generations. This segregation likely involves the maintenance of selective mtDNA variants during organogenesis, which can be differentially regulated in oocytes and preimplantation embryos during maturation.

The inheritance of mitochondrial DNA (mtDNA) is predominantly uniparental in most sexual eukaryotes. In this study, we examined the mitochondrial inheritance pattern of Cryptococcus gattii, a basidiomycetous yeast responsible for the recent and ongoing outbreak of cryptococcal infections in the US Pacific Northwest and British Columbia (especially Vancouver Island) in Canada. Using molecular markers, we analyzed the inheritance of mtDNA in 14 crosses between strains within and between divergent lineages in C. gattii. Consistent with results from recent studies, our analyses identified significant variations in mtDNA inheritance patterns among strains and crosses, ranging from strictly uniparental to biparental. For two of the crosses that showed uniparental mitochondrial inheritance in standard laboratory conditions, we further investigated the effects of the following environmental variables on mtDNA inheritance: UV exposure, temperature, and treatments with the methylation inhibitor 5-aza-2'-deoxycytidine and with the ubiquitination inhibitor ammonium chloride. Interestingly, one of these crosses showed no response to these environmental variables while the other exhibited diverse patterns ranging from complete uniparental inheritance of the MATa parent mtDNA, to biparental inheritance, and to a significant bias toward inheritance of the MATα parental mtDNA. Our results indicate that mtDNA inheritance in C. gattii differs from that in its closely related species Cryptococcus neoformans.

Mutation rate in the nuclear genome differs between sexes, with males contributing more mutations than females to their offspring. The male-biased mutation rates in the nuclear genome is most likely to be driven by a higher number of cell divisions in spermatogenesis than in oogenesis, generating more opportunities for DNA replication errors. However, it remains unknown whether male-biased mutation rates are present in mitochondrial DNA (mtDNA). Although mtDNA is maternally inherited and male mtDNA mutation typically does not contribute to genetic variation in offspring, male mtDNA mutations are critical for male reproductive health. In this study, we measured male mtDNA mutation rate using publicly available whole-genome sequences of single sperm of the freshwater microcrustacean Daphnia pulex . Using a stringent mutation detection pipeline, we found that the male mtDNA mutation rate is 3.32 × 10 −6 per site per generation. All the detected mutations are heteroplasmic base substitutions, with 57% of mutations converting G/C to A/T nucleotides. Consistent with the male-biased mutation in the nuclear genome, the male mtDNA mutation rate in D. pulex is approximately 20 times higher than the female rate per generation. We propose that the elevated mutation rate per generation in male mtDNA is consistent with an increased number of cell divisions during male gametogenesis.

Mitochondria contain their own DNA (mtDNA). In most sexually reproducing organisms, mtDNA is inherited maternally (uniparentally); this type of inheritance is thus referred to as 'maternal (uniparental) inheritance'. Recent studies have revealed various mechanisms to prevent the transmission of sperm-derived paternal mtDNA to the offspring, thereby ensuring maternal inheritance of mtDNA. In the nematode Caenorhabditis elegans, paternal mitochondria and their mtDNA degenerate almost immediately after fertilization and are selectively degraded by autophagy, which is referred to as 'allophagy' (allogeneic [non-self] organelle autophagy). In the fruit fly Drosophila melanogaster, paternal mtDNA is largely eliminated by an endonuclease G-mediated mechanism. Paternal mitochondria are subsequently removed by endocytic and autophagic pathways after fertilization. In many mammals, including humans, paternal mitochondria enter fertilized eggs. However, the fate of paternal mitochondria and their mtDNA in mammals is still a matter of debate. In this review, we will summarize recent knowledge on the molecular mechanisms underlying the prevention of paternal mtDNA transmission, which ensures maternal mtDNA inheritance in animals.

Mitochondrial DNA is typically inherited from only one parent [1–3]. In animals this is usually the mother. Maternal inheritance is often presented as the passive outcome of the difference in cytoplasmic content of egg and sperm, however active programs enforce uniparental inheritance at two levels, eliminating paternal mitochondrial genomes or destroying mitochondria delivered to the zygote by the sperm [4–14]. Both levels operate in Drosophila [8,12,13]. As sperm formation begins, hundreds of doomed mitochondrial genomes are visualized within the two huge mitochondria of each spermatid. These genomes abruptly disappear during spermatogenesis. Genome elimination, which is not in the interests of the restricted genomes, is directed by nuclear genes. Mutation of EndoG, which encodes a mitochondria-targeted endonuclease, retarded elimination [8]. Here, we show that knockdown of the nuclear-encoded mitochondrial DNA polymerase (Pol γ-α), Tamas, produces a more complete block of mtDNA elimination. Tamas is found in large particles that localize to mtDNA during genome elimination. We discount a simple possible mechanism by showing that the 3′-exonuclease function of the polymerase is not needed. While DNA elimination is a surprising function for DNA polymerase, it could provide a robust nexus for nuclear control of mitochondrial genome copy number since use of common interactions for elimination and replication might limit options for the mitochondrial genome to escape restriction. We suggest the DNA polymerase may play this role more widely and that inappropriate activation of its elimination ability might underlie association of DNA loss syndromes with mutations of the human mitochondrial DNA polymerase [15–17].

Slides presented at the Mitochondrial Genomics and Evolution SMBE Satellite Meeting in Ein Gedi (Israel) 3-6 September 2017.ABSTRACTMitochondria cannot be produced de novo and are inherited across generations. Their peculiar genetic features and the exposition to Reactive Oxygen Species (ROS) are predicted to cause the accumulation of deleterious mutations. How is the inheritance of a viable mitochondrial genetic information achieved?The “division of labour” (DOL) hypothesis postulates that oocyte mitochondria function as a genetic template by suppressing oxidative phosphorylation (OXPHOS) thus being protected from ROS-induced damage, while male gametes sacrifice mtDNA integrity to OXPHOS for motility. However, there is a lack of support for a causal link between high OXPHOS activity and generation of hazardous amounts of ROS, and there is no clear-cut evidence about oocyte mitochondria being quiescent. A well-suited biological system to study such issue, is the Doubly Uniparental Inheritance (DUI) of mitochondria, as yet reported in ~100 species of bivalves in which two sex-linked mitochondrial lineages are inherited uniparentally. Under DUI, sperm mitochondria are regularly transmitted to male progeny, and the long evolutionary persistence of DUI (>200 Myr) indicates that sperm mtDNA can be a viable genetic template. Accordingly, our data show that in DUI animals mitochondria of both gamete types are active, and we did not find evidence of mutation accumulation due to sperm swimming.Although DUI might look like a weird exception to a quite conserved biological feature, molecular and phylogenetic evidence suggests that it evolved from the common maternal inheritance so the two systems share the same underlying molecular mechanism. Is DUI undermining the DOL hypothesis? More work is needed to assess this point, and there are at least two possibilities by which this would not be the case: DUI species might use alternative energy-production pathways and/or produce less ROS; DUI species might have evolved specific mechanisms of ROS scavenging and/or mtDNA protection.

Freshwater mussel species with doubly uniparental inheritance (DUI) of mtDNA are unique because they are naturally heteroplasmic for two extremely divergent mtDNAs with ~50% amino acid differences for protein-coding genes. The paternally-transmitted mtDNA (or M mtDNA) clearly functions in sperm in these species, but it is still unknown whether it is transcribed when present in male or female soma. In the present study, we used PCR and RT-PCR to detect the presence and expression of the M mtDNA in male and female somatic and gonadal tissues of the freshwater mussel species Venustaconcha ellipsiformis and Utterbackia peninsularis (Unionidae). This is the first study demonstrating that the M mtDNA is transcribed not only in male gonads, but also in male and female soma in freshwater mussels with DUI. Because of the potentially deleterious nature of heteroplasmy, we suggest the existence of different mechanisms in DUI species to deal with this possibly harmful situation, such as silencing mechanisms for the M mtDNA at the transcriptional, post-transcriptional and/or post-translational levels. These hypotheses will necessitate additional studies in distantly-related DUI species that could possess different mechanisms of action to deal with heteroplasmy.

The mitochondrion is an organelle that has its own DNA (mtDNA). Mitochondria play essential roles in energy production and in various cellular processes such as metabolism and signal transduction. In most animals, including humans, although the sperm-derived paternal mitochondria enter the oocyte cytoplasm after fertilization, their mtDNA is never transmitted to the offspring. This pattern of mtDNA inheritance is well known as “maternal inheritance.” However, how the paternal mitochondria and mtDNA are eliminated from the cytoplasm of gametes or zygotes remains an enigma. Recently, a variety of mechanisms, including specific nuclease-dependent systems, ubiquitin–proteasome system, and autophagy have been shown to degrade the paternal mtDNA or the paternal mitochondria themselves in order to prevent paternal mtDNA transmission. In this review, we will address the current state of knowledge of the molecular mechanisms underlying the elimination of paternal mtDNA or mitochondrial structures for ensuring the maternal transmission of mtDNA.

Species within the marine mussel Mytilus edulis complex (M. edulis, M. galloprovin- cialis, and M. trossulus) have an unusual form of mitochondrial DNA (mtDNA) inheritance com- monly referred to as doubly uniparental inheritance (DUI). With DUI, all progeny inherit mtDNA maternally (F), while male progeny also inherit mtDNA paternally (M) through their father's sperm. Therefore, females are normally homoplasmic for the F mtDNA, and males are heteroplas- mic for the F and M mtDNA. In this study, we show that the regulation of DUI in populations of blue mussels in northern Japan is severely disrupted; consequently, the majority of individuals (89%) are heteroplasmic for M and F mtDNA. Disruption of DUI is primarily due to the failure of female embryos to exclude the father's M mtDNA from their somatic and germinal tissues. High proportions of heteroplasmic females have only been reported in Japanese mussel populations thus far. We show that this disruption occurs in both parental species (M. trossulus and M. gallo- provincialis) as well as hybrids and is independent of geographic location around Hokkaido, Japan. This finding differs from many previous studies, which have shown that disruption in DUI is confined to hybrid zones. In addition, some individuals were heteroplasmic for M or F mtDNA, indicating that DUI disruption is multigenerational. Triplasmic individuals were 3 times more often female and were mostly confined within the hybrid zones; however, a few individuals were found within M. galloprovincialis parental populations.

Mitochondrial homoplasmy signifies the existence of identical copies of mitochondrial DNA (mtDNA) and is essential for normal development, as heteroplasmy causes abnormal development and diseases in human. Homoplasmy in many organisms is ensured by maternal mtDNA inheritance through either absence of paternal mtDNA delivery or early elimination of paternal mtDNA. However, whether paternal mtDNA is transcribed has remained unknown. Here we report that paternal mtDNA shows late elimination and transcriptional quiescence in cyprinid fishes. Paternal mtDNA was present in zygotes but absent in larvae and adult organs of goldfish and blunt-snout bream, demonstrating paternal mtDNA delivery and elimination for maternal mtDNA inheritance. Surprisingly, paternal mtDNA remained detectable up to the heartbeat stage, suggesting its late elimination leading to embryonic heteroplasmy up to advanced embryogenesis. Most importantly, we never detected the cytb RNA of paternal mtDNA at all stages when paternal mtDNA was easily detectable, which reveals that paternal mtDNA is transcriptionally quiescent and thus excludes its effect on the development of heteroplasmic embryos. Therefore, paternal mtDNA in cyprinids shows late elimination and transcriptional quiescence. Clearly, transcriptional quiescence of paternal mtDNA represents a new mechanism for maternal mtDNA inheritance and provides implications for treating mitochondrion-associated diseases by mitochondrial transfer or replacement.

Although the uniparental (or maternal) inheritance of mitochondrial DNA (mtDNA) is widespread, the reasons for its evolution remain unclear. Two main hypotheses have been proposed: selection against individuals containing different mtDNAs (heteroplasmy) and selection against "selfish" mtDNA mutations. Recently, uniparental inheritance was shown to promote adaptive evolution in mtDNA, potentially providing a third hypothesis for its evolution. Here, we explore this hypothesis theoretically and ask if the accumulation of beneficial mutations provides a sufficient fitness advantage for uniparental inheritance to invade a population in which mtDNA is inherited biparentally. In a deterministic model, uniparental inheritance increases in frequency but cannot replace biparental inheritance if only a single beneficial mtDNA mutation sweeps through the population. When we allow successive selective sweeps of mtDNA, however, uniparental inheritance can replace biparental inheritance. Using a stochastic model, we show that a combination of selection and drift facilitates the fixation of uniparental inheritance (compared to a neutral trait) when there is only a single selective mtDNA sweep. When we consider multiple mtDNA sweeps in a stochastic model, uniparental inheritance becomes even more likely to replace biparental inheritance. Our findings thus suggest that selective sweeps of beneficial mtDNA haplotypes can drive the evolution of uniparental inheritance.

Mitochondrial homoplasmy, which is maintained by strictly maternal inheritance and a series of bottlenecks, is thought to be an adaptive condition for metazoans. Doubly uniparental inheritance (DUI) is a unique mode of mitochondrial transmission found in bivalve species, in which two distinct mitochondrial genome (mtDNA) lines are present, one inherited through eggs (F) and one through sperm (M). During development, the two lines segregate in a sex- and tissue-specific manner: females lose M during embryogenesis, whereas males actively segregate it in the germ line. These two pivotal events are still poorly characterized. Here we investigated mtDNA replication dynamics during embryogenesis and pre-adulthood of the venerid Ruditapes philippinarum using real-time quantitative PCR. We found that both mtDNAs do not detectably replicate during early embryogenesis, and that the M line might be lost from females around 24 h of age. A rise in mtDNA copy number was observed before the first reproductive season in both sexes, with the M mitochondrial genome replicating more than the F in males, and we associate these boosts to the early phase of gonad production. As evidence indicates that DUI relies on the same molecular machine of mitochondrial maternal inheritance that is common in most animals, our data are relevant not only to DUI but also to shed light on how differential segregations of mtDNA variants, in the same nuclear background, may be controlled during development.

SummaryThe timing and mechanisms of mitochondrial DNA (mtDNA) segregation and transmission in mammals are poorly understood. Genetic bottleneck in female germ cells has been proposed as the main phenomenon responsible for rapid intergenerational segregation of heteroplasmic mtDNA. We demonstrate here that mtDNA segregation occurs during primate preimplantation embryogenesis resulting in partitioning of mtDNA variants between daughter blastomeres. A substantial shift toward homoplasmy occurred in fetuses and embryonic stem cells (ESCs) derived from these heteroplasmic embryos. We also observed a wide range of heteroplasmic mtDNA variants distributed in individual oocytes recovered from these fetuses. Thus, we present here evidence for a previously unknown mtDNA segregation and bottleneck during preimplantation embryo development, suggesting that return to the homoplasmic condition can occur during development of an individual organism from the zygote to birth, without a passage through the germline.

Mitochondria are factories for energy production and genetic alterations in mtDNA will directly impact OXPHOS function. The copy number of mtDNA (i.e., the number of mtDNA per spermatozoon) is one of the major mitochondrial genetic features. Besides mtDNA copy number, the change of either mtDNA or nDNA integrity is another important factor causing asthenospermia, or poor sperm motility in infertile men. In this study, we investigated the mtDNA copy number and the integrities of mtDNA and nDNA respectively in semen samples from different donors at 5,300 m altitudes. Total DNA was extracted from semen samples from donors in two different altitudes. Quantitative PCR was performed to evaluate the mtDNA copy number. PCR amplification was used to examine the integrity of sperm mtDNA. Flow cytometry was carried out to investigate sperm nDNA integrity. All data were analyzed to show the statistical significance. Sperm mtDNA copy number for those living at high altitude (5,300 m) for one month was significantly higher (P

Doubly uniparental inheritance (DUI) of mitochondrial (mt) DNA has been reported in the blue mussel Mytilus galloprovincialis. In DUI, males inherit both paternal (M type) and maternal (F type) mtDNA. Here we investigated changes in M type mtDNA copy numbers and mitochondrial mass in testicular cells by real-time polymerase chain reaction and flow cytometry. The ratios of M type mtDNA copy numbers to nuclear DNA content were not different between haploid (1n), diploid (2n) and tetraploid (4n) spermatogenic cells. The mitochondrial mass decreased gradually during spermatogenesis. These results suggest that mtDNA and mitochondrial mass are maintained during spermatogenesis. We then traced M type mtDNA in larvae after fertilization. M type mtDNA was maintained up to 24 h after fertilization in the male-biased crosses, but decreased significantly in female-biased crosses (predicted by Mito Tracker staining pattern). These results are strikingly different from those reported for mammals and fish, where it is well known that the mitochondria and mtDNA are reduced during spermatogenesis and that sperm mitochondria and mtDNA are eliminated soon after fertilization. Thus, the M type mtDNA copy number is maintained during spermatogenesis and in the development of male larvae to sustain the DUI system in the blue mussel.

Chloroplast and mitochondrial DNA (mtDNA) is inherited exclusively from the female parent in the plant and animal kingdoms. In isogamous green algae of the genus Chlamydomonas, however, chloroplast DNA (cpDNA) is inherited maternally while mtDNA is inherited paternally. To study the paternal inheritance of mtDNA in Chlamydomonas, markers were constructed to distinguish mtDNA origins, diploid cells were constructed, and genetic, molecular biological and microscopic analyses of mt+ and mt- mtDNA in zygotes were conducted. Here, the literature on mitochondrial inheritance in Chlamydomonas is reviewed.

Mammalian mitochondrial DNA (mtDNA) is a small, maternally inherited genome that codes for 13 essential proteins in the respiratory chain. Mature oocytes contain more than 150 000 copies of mtDNA, at least an order of magnitude greater than the number in most somatic cells, but sperm contain only approximately 100 copies. Mitochondrial oxidative phosphorylation has been suggested to be an important determinant of oocyte quality and sperm motility; however, the functional significance of the high mtDNA copy number in oocytes, and of the low copy number in sperm, remains unclear. To investigate the effects of mtDNA copy number on fertility, we genetically manipulated mtDNA copy number in the mouse by deleting one copy of Tfam, an essential component of the mitochondrial nucleoid, at different stages of germline development. We show that males can tolerate at least a threefold reduction in mtDNA copy number in their sperm without impaired fertility, and in fact, they preferentially transmit a deleted Tfam allele. Surprisingly, oocytes with as few as 4000 copies of mtDNA can be fertilized and progress normally through preimplantation development to the blastocyst stage. The mature oocyte, however, has a critical postimplantation developmental threshold of 40 000-50 000 copies of mtDNA in the mature oocyte. These observations suggest that the high mtDNA copy number in the mature oocyte is a genetic device designed to distribute mitochondria and mtDNAs to the cells of the early postimplantation embryo before mitochondrial biogenesis and mtDNA replication resumes, whereas down-regulation of mtDNA copy number is important for normal sperm function.

Mutations in mitochondrial DNA (mtDNA) are a common cause of genetic disease. Pathogenic mutations in mtDNA are detected in approximately 1 in 250 live births and at least 1 in 10,000 adults in the UK are affected by mtDNA disease. Treatment options for patients with mtDNA disease are extremely limited and are predominantly supportive in nature. Mitochondrial DNA is transmitted maternally and it has been proposed that nuclear transfer techniques may be an approach for the prevention of transmission of human mtDNA disease. Here we show that transfer of pronuclei between abnormally fertilized human zygotes results in minimal carry-over of donor zygote mtDNA and is compatible with onward development to the blastocyst stage in vitro. By optimizing the procedure we found the average level of carry-over after transfer of two pronuclei is less than 2.0%, with many of the embryos containing no detectable donor mtDNA. We believe that pronuclear transfer between zygotes, as well as the recently described metaphase II spindle transfer, has the potential to prevent the transmission of mtDNA disease in humans.

Cann et al. have claimed on the basis of mitochondrial DNA (mtDNA) data that our direct ancestral Homo sapiens evolved in the African continent and spread to other continents, followed by the total replacement of the indigenous population. Their “Out-of-Africa” model is based on the assumption that mtDNA inheritance is simply maternal. Recent findings suggest the possibility that in between-population, e.g. African and Asian, mating, the African paternal mtDNA was transferred to the egg cell of an Asian together with Y-chromosomal DNA in the human past. Considering that Y-chromos- omal DNA and mtDNA sequences of African origin coexist together with Asian X-chromos- omal and autosomal DNA sequences in a current Asian, the observations by Cann et al. suggest the full/near full replacement of mtDNA in the human past, but do not necessarily imply the total replacement of indigenous populations with African migrants.

Mitochondria are found in all eukaryotic cells and contain their own genome (mitochondrial DNA or mtDNA). Unlike the nuclear genome, which is derived from both the egg and sperm at fertilization, the mtDNA in the embryo is derived almost exclusively from the egg; that is, it is of maternal origin. Mutations in mtDNA contribute to a diverse range of currently incurable human diseases and disorders. To establish preclinical models for new therapeutic approaches, we demonstrate here that the mitochondrial genome can be efficiently replaced in mature non-human primate oocytes (Macaca mulatta) by spindle-chromosomal complex transfer from one egg to an enucleated, mitochondrial-replete egg. The reconstructed oocytes with the mitochondrial replacement were capable of supporting normal fertilization, embryo development and produced healthy offspring. Genetic analysis confirmed that nuclear DNA in the three infants born so far originated from the spindle donors whereas mtDNA came from the cytoplast donors. No contribution of spindle donor mtDNA was detected in offspring. Spindle replacement is shown here as an efficient protocol replacing the full complement of mitochondria in newly generated embryonic stem cell lines. This approach may offer a reproductive option to prevent mtDNA disease transmission in affected families.

Although it is known that the hybrid male mouse is sterile just like any other animal's heterogametic sex, the reason why only the male germ cells are impaired has yet to be discovered. TdT-mediated dUTP nick end labeling assay using a confocal fluorescence microscope and DNA fragmentation assay of hybrid testis indicated destruction of the mitochondrial DNA (mtDNA) rather than the nuclear DNA. Previously we reported that maternal mtDNA inheritance is through selective sperm mtDNA elimination based on the sperm factor and two egg factors, and expression of these three factors was recognized in the hybrid testis. It was thereby assumed that mtDNA destruction caused by the expression of maternal mtDNA inheritance system in male germ cells is implicated in the hybrid male sterility of mice., Molecular biology reports, 36(6), pp.1365-1369; 2009

The mechanisms by which paternal inheritance of mitochondrial DNA (mtDNA) (paternal leakage) and, subsequently, recombination of mtDNA are prevented vary in a species-specific manner with one mechanism in common: paternally derived mtDNA is assumed to be vastly outnumbered by maternal mtDNA in the zygote. To date, this dilution effect has only been described for two mammalian species, human and mouse. Here, we estimate the mtDNA content of chinook salmon oocytes to evaluate the dilution effect operating in another vertebrate; the first such study outside a mammalian system. Employing real-time PCR, we determined the mtDNA content of chinook salmon oocytes to be 3.2 x 10(9)+/-1.0 x 10(9), and recently, we determined the mtDNA content of chinook salmon sperm to be 5.73+/-2.28 per gamete. Accordingly, the ratio of paternal-to-maternal mtDNA if paternal leakage occurs is estimated to be 1:5.5 x 10(8). This contribution of paternal mtDNA to the overall mtDNA pool in salmon zygotes is three to five orders of magnitude smaller than those revealed for the mammalian system, strongly suggesting that paternal inheritance of mtDNA per offspring will be much less likely in this system than in mammals.

Animal mitochondrial DNA (mtDNA) is predominantly inherited maternally. Various mechanisms to avoid the transmission of paternal mtDNA to offspring have been proposed, including the dilution of paternal mtDNA by maternal mtDNA in the zygote. The effectiveness of dilution as a barrier will be determined by the number of mtDNA molecules contributed by each parental gamete, and is expected to be highly variable among different taxa due to interspecific differences in mating systems and gamete investment. Estimates of this ratio are currently limited to few mammalian species, and data from other taxa are therefore needed to better understand the mechanisms of mitochondrial inheritance. The present study estimates mtDNA content in salmon sperm, the first nonmammalian vertebrate to be examined. Although highly divergent, it appears that the mtDNA content may be conserved within vertebrate taxa, indicating that the reduction of mtDNA is a key factor of spermatogenesis to ensure mitochondrial functionality on the one hand, and to avoid paternal leakage at a significant or detectable level on the other hand. We employ quantitative real-time PCR (Q-PCR) and demonstrate the accuracy and high reproducibility of our experiments. Furthermore, we compare and evaluate two standard approaches used for the quantification of genes, Q-PCR and blotting methods, in regard to their utility in the accurate quantification of mitochondrial genes.

The majority of mitochondrial DNA (mtDNA) mutations that cause human disease are mild to moderately deleterious, yet many random mtDNA mutations would be expected to be severe. To determine the fate of the more severe mtDNA mutations, we introduced mtDNAs containing two mutations that affect oxidative phosphorylation into the female mouse germ line. The severe ND6 mutation was selectively eliminated during oogenesis within four generations, whereas the milder COI mutation was retained throughout multiple generations even though the offspring consistently developed mitochondrial myopathy and cardiomyopathy. Thus, severe mtDNA mutations appear to be selectively eliminated from the female germ line, thereby minimizing their impact on population fitness.

The first complete mitochondrial DNA (mtDNA) sequences (approximately 16,569 bp) in 20 patients with asthenozoospermia and a comparison with 23 new complete mtDNA sequences in teratoasthenozoospermic individuals, confirmed no sharing of specific polymorphisms or specific mitochondrial lineages between these individuals. This is strong evidence against the accepted claim of a major role played by mtDNA in male fertility, once supported by haplogroup association studies based on the screening of hypervariable region I. The hypothesis of maternally driven selection acting in male reproductive success must thus be treated with caution. This work was partially supported by Fundação para a Ciência e a Tecnologia—Programa de Financiamento Plirianual do Centro de Investigação de Genética Molecular Humana and by Research Project POCTI/SAU/97/2001 from Fundação para a Ciência e a Tecnologia.

Mitochondrial DNA (mtDNA) is strictly maternally inherited in metazoans. The major exception to this rule has been found in many bivalve species which allow the presence of different sex-linked mtDNA molecules. This mechanism, named Doubly Uniparental Inheritance (DUI), is characterized by the presence of 2 mtDNAs: the female mtDNA is found in somatic tissue and female gonads whereas the male mtDNA is usually found in male gonads and sperm. In this study we highlight the existence of two divergent mitochondrial haplotypes with a low genetic difference around 6-8% in Arctica islandica, a long-lived clam belonging to the Arcticidae, a sister group to the Veneridae in which DUI has been found. Phylogenetic analysis on cytochrome b and 16S sequences from somatic and gonadic tissues of clams belonging to different populations reveal the presence of the “divergent” type in male gonads only and the “normal” type in somatic tissues and female gonads. This peculiar segregation of divergent mtDNA types speaks for the occurrence of the DUI mechanism in Arctica islandica. This example also highlights the difficulties to assess the presence of such particular mitochondrial inheritance system and underlines the possible misinterpretations in phylogeographic and phylogenetic studies of bivalve species linked to the presence of two poorly differentiated mitochondrial genomes.

Some regions of the genome exhibit sexual asymmetries in inheritance and are thus subjected to sex-biased evolutionary forces. Maternal inheritance of mitochondrial DNA (mtDNA) enables mtDNA mutations harmful to males, but not females, to accumulate. In the face of male-harmful mtDNA mutation accumulation, selection will favour the evolution of compensatory modifiers in the nuclear genome that offset fitness losses to males. The Y chromosome is a candidate to host these modifiers, because it is paternally inherited, known to harbour an abundance of genetic variation for male fertility, and therefore likely to be under strong selection to uphold male viability. Here, we test for intergenomic interactions involving mtDNA and Y chromosomes in male Drosophila melanogaster. Specifically, we examine effects of each of these genomic regions, and their interaction, on locomotive activity, across different environmental contexts – both dietary and social. We found that both the mtDNA haplotype and Y chromosome haplotype affected activity in males assayed in an environment perceived as social. These effects, however, were not evident in males assayed in perceived solitary environments, and neither social nor solitary treatments revealed evidence for intergenomic interactions. Finally, the magnitude and direction of these genetic effects was further contingent on the diet treatment of the males. Thus, genes within the mtDNA and Y chromosome are involved in genotype-by-environment interactions. These interactions might contribute to the maintenance of genetic variation within these asymmetrically inherited gene regions and complicate the dynamics of genetic interactions between the mtDNA and the Y chromosome.

Recent reports have questioned the accepted dogma that mammalian mitochondrial DNA (mtDNA) is strictly maternally inherited. In humans, the argument hinges on detecting a signature of inter-molecular recombination in mtDNA sequences sampled at the population level, inferring a paternal source for the mixed haplotypes. However, interpreting these data is fraught with difficulty, and direct experimental evidence is lacking. Using extreme-high depth mtDNA re-sequencing up to ~1.2 million-fold coverage, we find no evidence that paternal mtDNA haplotypes are transmitted to offspring in humans, thus excluding a simple dilution mechanism for uniparental transmission of mtDNA present in all healthy individuals. Our findings indicate that an active mechanism eliminates paternal mtDNA which likely acts at the molecular level., Author Summary Emerging evidence raises the possibility that human mitochondrial DNA (mtDNA) is not strictly maternally inherited, but it has not been technically possible to test this hypothesis directly. We identified trios with discordant mtDNA haplotypes, parent-offspring trios were validated using polymorphic microsatellites, and then used extreme-high depth mtDNA re-sequencing to look for paternally transmitted mtDNA. Despite having up to ~1.2 million-fold coverage of mtDNA, we find no evidence that paternal mtDNA haplotypes are transmitted to offspring in humans. Our findings exclude a simple dilution mechanism for uniparental transmission of mtDNA present in all healthy individuals.

Objective Mitochondrial DNA (mtDNA) encodes the proteins of the electron transfer chain to produce adenosine triphosphate through oxidative phosphorylation, and is essential to sustain life. mtDNA is unique from the nuclear genome in so much as it is solely maternally inherited (non-mendelian patterning), and shows a relatively high rate of mutation due to the absence of error checking capacity. While it is generally assumed that most new mutations accumulate through the process of heteroplasmy, it is unknown whether mutations initiated in the mother are inherited, occur in utero, or occur and accumulate early in life. The purpose of this study is to examine the maternally heritable and de novo mutation rate in the fetal mtDNA through high-fidelity sequencing from a large population-based cohort. Study Design Samples were obtained from 90 matched maternal (blood) and fetal (placental) pairs. In addition, a smaller cohort (n = 5) of maternal (blood), fetal (placental), and neonatal (cord blood) trios were subjected to DNA extraction and shotgun sequencing. The whole genome was sequenced on the Illumina HiSeq platform (Illumina Inc., San Diego, CA), and haplogroups and mtDNA variants were identified through mapping to reference mitochondrial genomes (NC_012920). Results We observed 665 single nucleotide polymorphisms and 82 insertions-deletions variants identified in the cohort at large. We achieved high sequencing depth of the mtDNA to an average depth of 65X (range, 20–171X) coverage. The proportions of haplogroups identified in the cohort are consistent with the patient’s self-identified ethnicity (>90% Hispanic), and all maternal-fetal pairs mapped to the identical haplogroup. Only variants from samples with average depth >20X and allele frequency >1% were included for further analysis. While the majority of the maternal-fetal pairs (>90%) demonstrated identical variants at the single nucleotide level, we observed rare mitochondrial single nucleotide polymorphism discordance between maternal and fetal mitochondrial genomes. Conclusion In this first in-depth sequencing analysis of mtDNA from maternal-fetal pairs at the time of birth, a low rate of de novo mutations appears in the fetal mitochondrial genome. This implies that these mutations likely arise from the maternal heteroplasmic pool (eg, in the oocyte), and accumulate later in the offspring’s life. These findings have key implications for both the occurrence and screening for mitochondrial disorders.

Background The retention of a genome in mitochondria (mtDNA) has several consequences, among which the problem of ensuring a faithful transmission of its genetic information through generations despite the accumulation of oxidative damage by reactive oxygen species (ROS) predicted by the free radical theory of ageing. A division of labour between male and female germ line mitochondria was proposed: since mtDNA is maternally inherited, female gametes would prevent damages by repressing oxidative phosphorylation, thus being quiescent genetic templates. We assessed mitochondrial activity in gametes of an unusual biological system (doubly uniparental inheritance of mitochondria, DUI), in which also sperm mtDNA is transmitted to the progeny, thus having to overcome the problem of maintaining genetic information viability while producing ATP for swimming. Results Ultrastructural analysis shows no difference in the conformation of mitochondrial cristae in male and female mature gametes, while mitochondria in immature oocytes exhibit a simpler internal structure. Our data on transcriptional activity in germ line mitochondria show variability between sexes and different developmental stages, but we do not find evidence for transcriptional quiescence of mitochondria. Our observations on mitochondrial membrane potential are consistent with mitochondria being active in both male and female gametes. Conclusions Our findings and the literature we discussed may be consistent with the hypothesis that template mitochondria are not functionally silenced, on the contrary their activity might be fundamental for the inheritance mechanism. We think that during gametogenesis, fertilization and embryo development, mitochondria undergo selection for different traits (e.g. replication, membrane potential), increasing the probability of the transmission of functional organelles. In these phases of life cycle, the great reduction in mtDNA copy number per organelle/cell and the stochastic segregation of mtDNA variants would greatly improve the efficiency of selection. When a higher mtDNA copy number per organelle/cell is present, selection on mtDNA deleterious mutants is less effective, due to the buffering effect of wild-type variants. In our opinion, a combination of drift and selection on germ line mtDNA population, might be responsible for the maintenance of viable mitochondrial genetic information through generations, and a mitochondrial activity would be necessary for the selective process. Reviewers This article was reviewed by Nick Lane, Fedor S Severin and Fyodor Kondrashov.

Several species from a number of bivalve molluscan families are known to have a paternally transmitted mitochondrial genome, along with the standard maternally transmitted one. The main characteristic of the phenomenon, known as doubly uniparental inheritance (DUI), is the coupling of sex and mtDNA inheritance: males receive both genomes but transmit only the paternal to their progeny; females either do not have the paternal genome or, if they do, they do not transmit it to their progeny. In the families Mytilidae and Veneridae, both of which have DUI, a female individual is either female-biased (it produces only, or nearly so, female progeny), male-biased (it produces mainly male progeny) or non-biased (it produces both genders in intermediate frequencies). Here we present evidence for a same pattern in the freshwater mussel, Unio delphinus (Unionidae). These results suggest that the maternal control of whether a fertilized egg will develop into a male or a female individual (and the associated feature of whether it will inherited or not inherit the paternal mtDNA) is a general characteristic of species with DUI.

Women who have had a child with mitochondrial DNA (mtDNA) disease need to know the risk of recurrence, but this risk is difficult to estimate because mutant and wild-type (normal) mtDNA coexist in the same person (heteroplasmy). The possibility that a single sample may not reflect the whole organism both impedes prenatal diagnosis of most mtDNA diseases, and suggests radical alternative strategies such as nuclear transfer. We used naturally occurring mtDNA variants to investigate mtDNA segregation in placenta. Using large samples of control placenta, we demonstrated that the level of polymorphic heteroplasmic mtDNA variants is very similar in mother, cord blood and placenta. However, where placental samples were very small (

It has previously been shown that paternal mitochondrial DNA (mtDNA) can be detected in later generations in Drosophila. To further analyze the paternal transmission of mtDNA, the progeny of two intraspecific and three interspecific crosses were examined in the frequency of the paternal transmission of mtDNA, using closely related species of the melanogaster species subgroup. Types of mtDNA in the progeny of the individual backcrosses of F(1) females were analyzed by selective amplification of paternal mtDNA. More than 100 F(1) females were examined for each backcross. The same type of mtDNA as that of the paternal mtDNA was detected in approximately 20-60% of the backcrosses. The present results indicate that paternal leakage occurs in the intraspecific crosses as well as in the interspecific crosses in Drosophila.

The introduction of nuclear transfer (NT) and other technologies that involve embryo reconstruction require us to reinvestigate patterns of mitochondrial DNA (mtDNA) transmission, transcription and replication. MtDNA is a 16.6 kb genome located within each mitochondrion. The number of mitochondria and mtDNA copies per organelle is specific to each cell type. MtDNA is normally transmitted through the oocyte to the offspring. However, reconstructed oocytes often transmit both recipient oocyte mtDNA and mtDNA associated with the donor nucleus. We argue that the transmission of two populations of mtDNA may have implications for offspring survival as only one allele might be actively transcribed. This could result in the offspring phenotypically exhibiting mtDNA depletion-type syndromes. A similar occurrence could arise when nucleo–cytoplasmic interactions fail to regulate mtDNA transcription and replication, especially as the initiation of mtDNA replication post-implantation is a key developmental event. Furthermore, failure of the donor somatic nucleus to be reprogrammed could result in the early initiation of replication and the loss of cellular mtDNA specificity. We suggest investigations should be conducted to enhance our understanding of nucleo–cytoplasmic interactions in order to improve NT efficiency.

Animal mitochondrial DNA is normally inherited clonally from a mother to all her offspring. Mitochondrial heteroplasmy, the occurrence of more than one mitochondrial haplotype within an individual, can be generated by relatively common somatic mutations within an individual, by heteroplasmy of the oocytes, or by paternal leakage of mitochondria during fertilization of an egg. This biparental inheritance has so far been reported only in mice, mussels, Drosophila, and humans. Here we present evidence that paternal leakage occurs in a bird, the great tit Parus major. The major and minor subspecies groups of the great tit mix in the middle Amur Valley in far-eastern Siberia, where we found a bird that possessed the very distinct haplotypes of the two groups. To our knowledge this is the first report of paternal leakage in birds.

Previous studies have shown that in most pair matings of Mytilus edulis, M. trossulus, and M. galloprovincialis there is a large sex-ratio bias in favor of either males or females. The degree of bias is a characteristic property of the female parent, as matings of the same female with different males produce the same sex ratio, but matings of the same male with different females produce different sex ratios. All three species possess the unusual feature of doubly uniparental inheritance of mitochondrial DNA (mtDNA); i.e., they contain two distinct types of mtDNA, one that is transmitted matrilinearly and one that is transmitted patrilinearly. This coupling of sex and mtDNA transmission raises the possibility that the mechanism of sex-ratio determination in mussels might be under the control of the mtDNA of the female parent. Here we present data from pedigreed crosses that confirm the previous observations that in mussel matings there is a strong sex-ratio bias and that the bias is under the control of the female parent. In addition, these data strongly suggest that this control is exercised by the mother's nuclear rather than mitochondrial genotype. Making use of these findings we develop a model of mother-dependent sex determination and use data from crosses involving wild females to test the model's predictions at the population level.

Mitochondrial DNA (mtDNA) is almost entirely maternally inherited. Thousands of copies of mtDNA are present in every nucleated cell and in most normal individuals these are virtually identical (homoplasmy). mtDNA diseases may be caused by mutations in either mitochondrial or nuclear genes and, hence, give rise to maternal or autosomal patterns of inheritance. Antenatal diagnosis of mitochondrial diseases based on chorionic villous sampling is available for Mendelian disorders and the syndromes caused by mutations at bp 8993 (associated with Leigh's syndrome and neurogenic weakness, ataxia and retinitis pigmentosa (NARP)). However, prenatal diagnosis of many other maternally inherited mtDNA diseases is less reliable because it is not possible to predict with confidence the way in which heteroplasmic mtDNA mutations segregate within tissues and find clinical expression. This review focuses on the substantial progress in genetics that has been made recently, and on the management options that clinicians can offer to families.

Mytilus and other bivalves exhibit an unusual system of mitochondrial DNA (mtDNA) transmission termed doubly uniparental inheritance (DUI). Specifically, males transmit the mtDNA they have received from their fathers to their sons. Females transmit their mother's mtDNA to both sons and daughters. Males are normally heteroplasmic and females are normally homoplasmic, but not exclusively. This system is associated with an unusual pattern of molecular evolution. Male-transmitted mtDNA (M type) evolves faster than female-transmitted (F type) mtDNA. Relatively relaxed selection on the M type has been proposed as an explanation for this phenomenon. To further evaluate the selective forces acting upon the M-type genome, we used RT-PCR to determine where it is expressed. M-type mtDNA expression was detected in all gonad samples and in 50% of somatic tissues of males, and in a single female tissue. F-type mtDNA expression was detected in all female tissues, all male somatic tissues, and all but one male gonad sample. We argue that the expression of M-type mtDNA in male somatic and male gonad tissues has implications for the strength of selection acting upon it.Key words: gender-associated mitochondrial DNA, doubly uniparental inheritance of mtDNA, Mytilus edulis, molecular evolution.

University Department of Paediatics, Level 4, John Radcliffe Hospital, Headington, Oxford, OX3 9DU, UK.There is a risk that ICSI may increase the transmission of mtDNA diseases to children born after this technique. Knowledge of the fate and transmission of paternal mitochondrial DNA is important since mutations in mitochondrial DNA have been described in oligozoospermic males. We have used an adaptation of solid phase mini-sequencing to exclude the presence of levels of paternal mtDNA >0.001% in ICSI families. This method is more sensitive than those used in previous studies and is sufficient to detect the likely paternal contribution (approximately 0.1-0.5% from simple calculations of expected dilution during fertilization). Using this method, we were able to detect concentrations as low as 0.001% paternal mtDNA in a maternal mtDNA background. No paternal mtDNA was detected in the embryonic (blood or buccal swabs) tissue of children born after ICSI nor in extra-embryonic tissue (placenta or umbilical cord). In conclusion, we did not detect paternal mtDNA in blood, buccal swabs, placenta or umbilical cord of children born after ICSI. We have found no evidence that ICSI increases the risk of paternal transmission of mtDNA and hence of mtDNA disorders.

The manifestation of mitochondrial DNA (mtDNA) diseases depends on the frequency of heteroplasmy (the presence of several alleles in an individual), yet its transmission across generations cannot be readily predicted owing to a lack of data on the size of the mtDNA bottleneck during oogenesis. For deleterious heteroplasmies, a severe bottleneck may abruptly transform a benign (low) frequency in a mother into a disease-causing (high) frequency in her child. Here we present a high-resolution study of heteroplasmy transmission conducted on blood and buccal mtDNA of 39 healthy mother-child pairs of European ancestry (a total of 156 samples, each sequenced at ∼20,000× per site). On average, each individual carried one heteroplasmy, and one in eight individuals carried a disease-associated heteroplasmy, with minor allele frequency ≥1%. We observed frequent drastic heteroplasmy frequency shifts between generations and estimated the effective size of the germ-line mtDNA bottleneck at only ∼30-35 (interquartile range from 9 to 141). Accounting for heteroplasmies, we estimated the mtDNA germ-line mutation rate at 1.3 × 10(-8) (interquartile range from 4.2 × 10(-9) to 4.1 × 10(-8)) mutations per site per year, an order of magnitude higher than for nuclear DNA. Notably, we found a positive association between the number of heteroplasmies in a child and maternal age at fertilization, likely attributable to oocyte aging. This study also took advantage of droplet digital PCR (ddPCR) to validate heteroplasmies and confirm a de novo mutation. Our results can be used to predict the transmission of disease-causing mtDNA variants and illuminate evolutionary dynamics of the mitochondrial genome.

Eukaryotic cells typically contain numerous mitochondria, each with multiple copies of their own genome, the mtDNA. Uniparental transmission of mitochondria, usually via the mother, prevents the mixing of mtDNA from different individuals. While on the one hand, this should resolve the potential for selection for fast-replicating mtDNA variants that reduce organismal fitness, maternal inheritance will, in theory, come with another set of problems that are specifically relevant to males. Maternal inheritance implies that the mitochondrial genome is never transmitted through males, and thus selection can target only the mtDNA sequence when carried by females. A consequence is that mtDNA mutations that confer male-biased phenotypic expression will be prone to evade selection, and accumulate. Here, we review the evidence from the ecological, evolutionary and medical literature for male specificity of mtDNA mutations affecting fertility, health and ageing. While such effects have been discovered experimentally in the laboratory, their relevance to natural populations—including the human population—remains unclear. We suggest that the existence of male expression-biased mtDNA mutations is likely to be a broad phenomenon, but that these mutations remain cryptic owing to the presence of counter-adapted nuclear compensatory modifier mutations, which offset their deleterious effects.

▪ Abstract Mitochondrial DNA (mtDNA) exists in a nested hierarchy of populations. There are multiple mtDNAs within each mitochondrion, a population of mitochondria in each cell, multiple oocytes within each reproductive female, multiple females in each population, and so on up through species and higher clades. The metabolic properties of mitochondria make them highly mutagenic environments for the naked, circular mtDNAs that lie within them. This mutational pressure introduces mtDNA variation (i.e., heteroplasmy) into the cytoplasmic population of cell lineages that are particularly prone to mutational decay and Muller's ratchet owing to the asexual, maternal inheritance of mtDNA. Neutrality tests show that deleterious mutations are common in mtDNA evolution. Population cage experiments further show that mtDNA fitnesses are influenced by nuclear-mitochondrial interactions. These selective processes are pervasive despite the long-standing use of mtDNA as a neutral marker in population and evolutionary biology. These evolutionary dynamics are also unique in the nested hierarchy of mtDNA populations because mutation, selection, and drift can act—and interact—at multiple levels. Multi-level selection can facilitate the escape from Muller's ratchet and help resolve intragenomic conflicts. This review addresses recent advances in the transmission genetics, population genetics, and evolution of mtDNA. A primary goal of the review is to motivate additional empirical studies that might clarify the many units of selection acting on mtDNA.

Paternal inheritance of mitochondria DNA in sheep was discovered by examination of 152 sheep from 38 hybrid families for mtDNA D-loop polymorphisms using PCR-RFLP, amplification of repeated sequence somain, and PCR-SSCP of the D-loop 5' end region of a 253 bp fragment. Our findings have provided the first evidence of paternal inheritance of mtDNA in sheep and possible mechanisms of paternal inheritance were discussed.

The maternal inheritance of mitochondrial DNA (mtDNA) in eukaryotic organisms occurs because of the selective destruction of paternal mtDNA molecules that may be present in the zygote. The elimination of sperm mtDNA is less efficient in interspecific crosses, and biparental inheritance of mtDNA has been observed in a variety of species. Because interspecific crosses are likely to be extremely rare in nature, parental inheritance of mtDNA has been deemed of little relevance to population genetics. The mtDNA of the parasitic trematode Schistosoma mansoni was examined for its utility in addressing epidemiological questions related to the transmission and spread of schistosomiasis. Prior to embarking on such experiments, we sought to confirm the mode of inheritance of this molecule using the highly polymorphic mtDNA minisatellite as a marker. In 3 separate crosses, mtDNA apparently identical to paternal DNA was observed in some individuals of the F2 and F3 generations. These observations thus suggest the intraspecific paternal inheritance of mtDNA across multiple generations in Schistosoma mansoni.

The strictly maternal inheritance of mitochondria and mitochondrial DNA (mtDNA) in mammals is a developmental paradox promoted by an unknown mechanism responsible for the destruction of the sperm mitochondria shortly after fertilization. We have recently reported that the sperm mitochondria are ubiquitinated inside the oocyte cytoplasm and later subjected to proteolysis during preimplantation development (P. Sutovsky et al., Nature 1999; 402:371-372). Here, we provide further evidence for this process by showing that the proteolytic destruction of bull sperm mitochondria inside cow egg cytoplasm depends upon the activity of the universal proteolytic marker, ubiquitin, and the lysosomal apparatus of the egg. Binding of ubiquitin to sperm mitochondria was visualized by monospecific antibodies throughout pronuclear development and during the first embryonic divisions. The recognition and disposal of the ubiquitinated sperm mitochondria was prevented by the microinjection of anti-ubiquitin antibodies and by the treatment of the fertilized zygotes with lysosomotropic agent ammonium chloride. The postfecundal ubiquitination of sperm mitochondria and their destruction was not seen in the hybrid embryos created using cow eggs and sperm of wild cattle, gaur, thus supporting the hypothesis that sperm mitochondrion destruction is species specific. The initial ligation of ubiquitin molecules to sperm mitochondrial membrane proteins, one of which could be prohibitin, occurs during spermatogenesis. Even though the ubiquitin cross-reactivity was transiently lost from the sperm mitochondria during epididymal passage, likely as a result of disulfide bond cross-linking, it was restored and amplified after fertilization. Ubiquitination therefore may represent a mechanism for the elimination of paternal mitochondria during fertilization. Our data have important implications for anthropology, treatment of mitochondrial disorders, and for the new methods of assisted procreation, such as cloning, oocyte cytoplasm donation, and intracytoplasmic sperm injection.

Species of the families Mytilidae (sea mussels) and Unionidae (fresh water mussels) contain two types of mitochondrial DNA (mtDNA), the F that behaves as the standard animal mtDNA and the M that is transmitted through the sperm and establishes itself only in the male gonad. The two molecules have, therefore, separate transmission routes, one through the female and the other through the male lineage. The system has been named doubly uniparental inheritance (DUI). Another important feature of sea mussels is that the sex ratio among offspring of a pair mating is determined by the female parent only. The mechanism of DUI remains unknown. One hypothesis that is consistent with all observations is that the standard maternal inheritance was modified in mussels via the evolution of a suppressor gene that is expressed during oogenesis and has two alleles, the inactive and the active allele. In the presence of the active allele in the mother's genotype the egg is supplied with a substance that interferes and the normal mechanism of elimination of sperm mitochondria. This will explain why half of mussels have the father's mtDNA and half do not, but would not explain why presence/absence of paternal mtDNA is linked with the male and female gender, respectively. To provide an explanation for this linkage, one would have to assume that there is a causal relationship between retention of paternal mtDNA and sex determination.