Abstract The 3243A>G mutation in mitochondrial DNA (mtDNA), the most common cause of the syndrome of mitochondrial encephalomyopathy, lactic acidosis and stroke-like episodes, is also associated with many other phenotypes such as hearing loss, diabetes mellitus, epilepsy, cognitive decline, myopathy and cardiomyopathy. The prevalence of the mutation has been shown to be 16.3/100 000 adults in Northern Finland. The present study was performed to estimate the frequency and progression of cardiac abnormalities and to examine causes of death in patients with 3243A>G. Left ventricular hypertrophy (LVH) was found in echocardiography in 56% of patients with 3243A>G and in 15% of age and sex-matched controls. The median thickness of the diastolic interventricular septum or posterior wall was 14 mm in the patients with LVH. The prevalence of LVH determined by echocardiography increased from 40% to 56% in 25 patients with 3243A>G during three years of follow-up, this trend being especially marked among the diabetic patients. The ultra-low-frequency (ULF) and very-low-frequency (VLF) components of the spectral analysis of heart rate variability (HRV) were lower among the patients with 3243A>G than in matched controls (p = 0.02 in ULF and p = 0.04 in VLF), and the short-term fractal scaling exponent in detrended fluctuation analysis of HRV was lower in the patients with 3243A>G (1.16 ± 0.18 vs. 1.28 ± 0.13) (p < 0.01). Survival analysis of a birth cohort from pedigrees with 3243A>G revealed excess mortality before the age of 50 years. Neurological and cardiovascular diseases accounted for 32% of all the underlying causes of death in families with 3243A>G. Death was sudden and unexpected in 31% of cases in which 3243A>G was considered to be involved in the cause of death. The results show that cardiac abnormalities are frequent and progressive in patients with the 3243A>G mtDNA mutation and that cardiac autonomic regulation is disturbed. Patients with the 3243A>G mutation and their first degree maternal relatives died younger than was presupposed by their life expectancy at birth or at 15 years. The most common causes of death were neuropsychiatric and cardiovascular diseases.

This mini-review investigates and compiles the latest knowledge regarding epigenetic changes on the mammalian mitochondrial DNA and its proteins. Methylation of the DNA, acetylation of the proteins and silencing of genes by short non-coding RNAs are the main epigenetic changes known today to affect mitochondrial DNA, mostly leading to repression. Methylation mainly occurs at non-CpG sites in the main non-coding region called the D-loop, with methylation patterns being cell type specific. Acetylation of proteins are mainly controlled by the deacetylase SIRT3, with its function being correlated to longevity. On the other hand, mitochondrial dysfunction is directly associated with a plethora of diseases, such as neurodegenerative disorders and heart disorders. The mitochondrion and nucleus are immensely dependent on each other and exchange vital proteins and RNAs, with epigenetic changes on one potentially affecting the other. Recent research shows that heteroplasmy is a proven cause of mitochondrial malfunction and that paternal inheritance is possible. The mitochondrial haplotype also shows different vulnerability to certain diets and diseases, leading to the conclusion that the mitochondrial haplotype can be used to more than just tracing human origins, such as to predicting and preventing diseases.

No description available.

MtDNA mutations in mammalian cells are implicated in cellular ageing and encephalomyopathies, although mechanisms involved are not completely understood. The mitochondrial genetic bottleneck has puzzled biologists for a long time. Approximate models of genetic bottleneck proposed in the literature do not accurately model underlying biology. Recent studies indicate mitochondrial morphology changes during cellular aging in culture. In particular, the rates of mitochondrial fission and fusion are shown to be in tight balance, though this rate decreases with age. Some proteins involved in mitochondrial morphology maintenance are implicated in apoptosis. Hence, mitochondrial genetic and morphologic dynamics are critical to the life and death of cells. By working closely with experimental collaborators and by utilizing data derived from literature, we have developed stochastic simulation models of mitochondrial genetic and morphologic dynamics. Hypotheses from the mitochondrial genetic dynamics model include: (1) the decay of mtDNA heteroplasmy in blood is exponential and not linear as reported in literature. (2) Blood heteroplasmy measurements are a good proxy for the blood stem cell heteroplasmy. (3) By analyzing our simulation results in tandem with published longitudinal clinical data, we propose for the first time, a way to correct for the patient's age in the analysis of heteroplasmy data. (4) We develop a direct model of the genetic bottleneck process during mouse embryogenesis. (5) Partitioning of mtDNA into daughter cells during blastocyst formation and relaxed replication of mtDNA during the exponential growth phase of primordial germ cells leads to the variation in heteroplasmy inherited by offspring from the same mother. (6) We develop a “simulation control” for experimental studies on mtDNA heteroplasmy variation in cell cultures. Hypothesis from the mitochondrial morphologic dynamics model: (7) A cell adjusts the mitochondrial fusion rate to compensate for the fluctuations in the fission rate, but not vice versa. A deterministic model for this control is proposed. Contributions: extensible simulation models of mitochondrial genetic and morphologic dynamics to aide in the powerful analysis of published and new experimental data. Our results have direct relevance to cell biology and clinical diagnosis. The work also illustrates scientific success by tight integration of theory with practice. / Ph. D.

Les mitochondries sont impliquées dans de nombreuses voies métaboliques, et des mutations au sein de leur génome (ADNmt) conduisent à l’apparition de nombreuses pathologies. A l’heure actuelle, il n’existe aucun traitement contre ces affections mais différentes pistes thérapeutiques sont envisagées. L’objectif de ce travail a consisté en la mise au point d’une telle stratégie, dite anti-réplicative via l’exploitation de la voie d’import naturelle des ARN dans les mitochondries. De petits ARN artificiels importables dans les mitochondries humaines ont ainsi été utilisés comme vecteurs pour y importer une séquence capable de s’hybrider spécifiquement à l’ADNmt mutant et d’en stopper sa réplication. Les résultats obtenus ont permis de prouver la validité de cette stratégie vis-à-vis d’une large délétion et de mutations ponctuelles liées à divers cas pathologiques et de caractériser l’effet de modifications chimiques sur la stabilité, l’import et l’efficacité de ces ARN recombinants. / Mitochondria are involved in many metabolic pathways, and mutations in their genome (mtDNA) can cause a wide range of human disorders. No efficient treatment against these pathologies is currently available. The objective of this work consisted in the development of a therapeutic approach, called anti-replicative, based on the use of the natural pathway of RNA import into mitochondria. Small artificial RNA molecules able to be imported into human mitochondria have been used as vectors to address oligoribonucleotides capable to hybridize specifically to mutant mtDNA and to stop its replication. The effect of various chemical modifications on the stability, import and efficiency of these recombinant RNA has been characterized. All the data obtained prove the validity of the anti-replicative strategy for mtDNA containing a large deletion or pathogenic point mutations and can be considered as an important step to further develop an efficient therapy of mitochondrial diseases.

No description available.

No description available.

Les mitochondries et leur génome, l'ADN mitochondrial (ADNmt), sont généralement transmis uniquement par la mère aux fils et aux filles chez les métazoaires (transmission strictement maternelle, SMI). Une exception à la règle générale de la SMI se trouve dans environ 100 espèces de bivalves, qui se caractérise par une double transmission uniparentale (DUI) des mitochondries. Chez les espèces DUI, deux lignées d'ADNmt très divergentes et liées au sexe coexistent. Une lignée mitochondriale maternelle (type F), présente dans les ovocytes et les tissus somatiques des individus femelles et males, et une lignée paternelle (type M), présente dans les spermatozoïdes. Dans les tissus somatiques mâles, les deux lignées coexistent parfois, une condition appelée hétéroplasmie. En sachant que les variations génétiques dans l’ADNmt peuvent avoir un impact sur les fonctions mitochondriales, et en donnant l'association stricte des ADNmt de type M et F avec différents gamètes, il est imaginable que la forte divergence entre les deux lignées DUI puisse entraîner des adaptations bioénergétiques avec répercussion sur la reproduction. Le système DUI apporte également la nécessité pour les mitochondries paternelles de préserver leur propre intégrité génétique, ainsi que pour les cellules somatiques de faire face à l'hétéroplasmie. L'objectif de ma thèse était de lier le génotype mitochondrial des espèces bivalves DUI et SMI au phénotype. Plus précisément, j'ai exploré l'impact des variations de l'ADN mitochondrial spécifiques au sexe sur un large éventail de traits phénotypiques, allant de la bioénergétique mitochondriale et cellulaire à la performance des spermatozoïdes, en étudiant la valeur adaptative du système DUI à la lumière du fitness reproductif, de la sélection et de la transmission mitochondriales. Les résultats issus de ce projet de thèse ont révélé une nette divergence phénotypique entre les espèces DUI et SMI, reflétant peut-être les différentes pressions sélectives agissant sur les deux lignées mitochondriales. Contrairement aux espèces SMI, l'évolution sexo-spécifique des variants d'ADNmt DUI entraîne l'expression de différents phénotypes bioénergétiques mâles et femelles. Au niveau de la fonctionnalité mitochondriale, les mitochondries DUI de type M présentent une phosphorylation oxydative (OXPHOS) remodelée, caractérisée par un contrôle respiratoire inhabituel à l'extrémité de la chaîne respiratoire. La réorganisation générale de la bioénergétique des spermes DUI entraîne également une variation de l'équilibre entre les principales voies de production d'énergie, incluant la glycolyse, la glycolyse anaérobique, le métabolisme des acides gras, le cycle de l'acide tricarboxylique, l'OXPHOS, ainsi que la capacité antioxydante. Enfin, les spermatozoïdes DUI comptent entièrement sur l'énergie produite par OXPHOS pour maintenir une motilité inhabituelle caractérisée par une vitesse lente et une trajectoire plus curviligne, traits potentiellement associés à un plus grand succès de reproduction chez les organismes marins sessiles. Aussi, ils conservent la capacité de passer à une stratégie de production d'énergie mixte (aérobique et anaérobie) après la détection des ovocytes. Dans l'ensemble, ces résultats suggèrent que la variation de l'ADNmt dans les espèces DUI pourrait être adaptative, incluant adaptation bioénergétique sexo-spécifiques avec un effet en aval sur la performance des spermatozoïdes, la capacité de reproduction, la sélection et transmission des mitochondries paternelles. / Mitochondria and their genome, the mitochondrial DNA (mtDNA), are usually transmitted only by the mother to both sons and daughters in metazoan (i.e. strict maternal inheritance, SMI). An exception to the general rule of SMI is found in around 100 species of bivalves, which are characterized by a doubly uniparental inheritance (DUI) of mitochondria. In DUI species, two highly divergent and sex-linked mtDNA lineages coexist. One mitochondrial lineage is maternally inherited (F-type) and is present in oocytes and somatic tissues of both female and male individuals. The other lineage is paternally inherited (M-type) and is present in sperm. In male somatic tissues both lineages sometimes coexist, a condition named heteroplasmy. Knowing that variations in mitochondrially-encoded genes might impact mitochondrial functions, and giving the strict association of M and F-type mtDNAs with different gametes, it is conceivable that the variation between the two DUI lineages might result in sex-specific bioenergetic adaptations with repercussion on reproduction. Despite providing an unprecedented opportunity for the mtDNA to evolve for male functions, the DUI system also brings the need for sperm mitochondria to preserve their genetic integrity, as well as for somatic cells to deal with heteroplasmy. The objective of my PhD was to link the mitochondrial genotype of DUI and SMI bivalve species to the phenotype. I explored the impact of sex-specific mtDNA variations upon a wide set of phenotypic traits, ranging from mitochondrial and cellular bioenergetics to sperm performance, investigating the adaptive value of DUI system in the light of reproductive fitness, mitochondrial selection, preservation and transmission. The results stemming from this PhD project revealed a clear phenotypic divergence between DUI and SMI species, possibly reflecting the different selective pressures acting on their mitochondria as a result of their different mode of mitochondria transmission. Conversely to SMI species, the sex-specific evolution of DUI mtDNA variants results in the expression of different male and female bioenergetic phenotypes. At the level of mitochondrial functionality, M-type mitochondria exhibit a remodelled OXPHOS characterized by unusual respiratory control at the terminus of the respiratory chain. The general reorganization of DUI sperm bioenergetics also entails variation in the balance between the main energy producing pathways, including glycolysis, anaerobic glycolysis, fatty acid metabolism, tricarboxylic acid cycle, OXPHOS, as well as the antioxidant capacity. Finally, DUI sperm exhibit an unusual motility phenotype characterized by slow speed and high curvilinear trajectory, traits potentially associated with a higher reproductive success in sessile broadcast spawning marine organisms. They also completely rely on the energy produced by OXPHOS to sustain their performance, although maintaining the ability to switch to a more combined aerobic/anaerobic strategy of energy production after oocyte detection. Altogether, these results suggest that the mtDNA variation in DUI species might be adaptive, resulting in the expression of sex-specific bioenergetic adaptation with downstream effect on sperm performance, reproductive fitness, paternal mitochondria selection, preservation and transmission. The results also suggest that heteroplasmy has an impact onto the bioenergetics of male soma, and that a functional compensation between genomes might minimize any potential deleterious outcome.

An important goal of biology is to understand the key mechanisms of evolution underlying the diversity of living organisms on Earth. In that respect, the recent innovations in the field of new generation sequencing technologies (NGS) are bringing new and exciting opportunities. This thesis presents results obtained with these tools in the specific context of the study of two sister species of cold-adapted leaf beetles, Gonioctena intermedia and G. quinquepunctata. More specifically, this work is focused around four research directions: the two first explore methods of statistical inference using a spatially explicit model of coalescence, by (1) evaluating the potential of various summary statistics to discriminate phylogeographic hypotheses, and (2) investigating the dispersal abilities of a montane leaf beetle, G. quinquepunctata, using an original method that avoids using summary statistics. The third research direction focuses on the adaptation to cold conditions in this montane leaf beetle, by testing the association between genetic polymorphism across tens of thousands of genetic markers and altitude in samples collected at various elevation levels in the Vosges (France). Finally, the fourth, and last, research axis presents the discovery of mitochondrial heteroplasmy, i.e. the presence in an individual of multiple copies of the mitochondrial genome, in natural populations of G. intermedia. We illustrate, here, how NGS technologies could help identify this phenomenon, probably underestimated in animals, on a large scale. / Doctorat en Sciences / info:eu-repo/semantics/nonPublished

Chaque mitochondrie contient son propre génome en de multiples copies d’ADN. Les mutations de l'ADN mitochondriales (ADNmt) sont responsables de sévères dysfonctions de la chaîne respiratoire. Le ratio entre la proportion de copies sauvages et mutantes, qualifiée d'hétéroplasmie, détermine la sévérité de la pathologie. Une des mutations les plus répandues de l'ADNmt est la mutation m.3243A>G, affectant l'ARN de transfert de la leucine. Ce variant est à l'origine du syndrome mitochondrial MELAS. Il n’existe à l’heure actuelle aucun traitement curatif pour ce syndrome. Nous avons développé une série de cybrides neuronaux porteurs de la mutation m.3243A>G a différents taux d’hétéroplasmie. Nous avons mis en évidence que de fort taux de mutations sont responsables de sévères dysfonctions de la chaîne respiratoire, d’un défaut d’assemblage précoce du complexe I ainsi qu’une réduction du renouvellement mitochondrial. Différentes stratégies métaboliques ont été employées pour compenser ces déficits. L’exposition des cellules a une restriction glucidique ou à la diète cétogène associant réduction glucidique et ajout de corps cétoniques, améliore significativement les fonctions mitochondriales après 4 semaines. Ces effets passent notamment par une restauration de l’assemblage et de l’activité du complexe I médiée ces interventions métaboliques. Par ailleurs, l’administration de la diète cétogène à un patient atteint du syndrome MELAS a déjà montré des résultats encourageants. De telles approches pourraient alors, constituées des stratégies thérapeutiques futures dans le traitement du syndrome MELAS et des maladies mitochondriales. / Each mitochondrion contains its own genome in multiple copies. Mitochondrial DNA (mtDNA) mutations are responsible for respiratory chain defects. The ratio of mutant to normal mtDNA, a condition known as heteroplasmy, may determine the disease severity. The m.3243A>G mutation, which affects the leucine tRNA, is one of the most common mtDNA mutation. This variant is responsible for the MELAS syndrome, a neurodegenerative disease, characterized by pseudostrokes. Unfortunately there are no curative treatments for MELAS syndrome. We have developed series of cybrid neuronal cells lines carrying the m.3243A>G mutation with different mutant loads, within the same nuclear background. High mutation load is associated to severe respiratory chain dysfunction, an early complex I assembly defect and a mitochondrial turn-over deficit. Different strategies were used to compensate the defects in the mutant cells. Cell exposure to low glucose or ketogenic diet, combining glucose reduction and the addition of ketone bodies, greatly improves mitochondrial functions after 4 weeks. Those effects are linked to a significant increase of complex I assembly and activity mediated by those metabolic interventions. In addition, a MELAS patient treated with ketogenic diet showed significant clinical improvement. Thus, metabolic approaches may constitute promising therapeutic strategies against MELAS syndrome and mitochondrial disorders.