The human mitochondrial genome is extremely small compared with the nuclear genome, and mitochondrial genetics presents unique clinical and experimental challenges. Despite the diminutive size of the mitochondrial genome, mitochondrial DNA (mtDNA) mutations are an important cause of inherited disease. Recent years have witnessed considerable progress in understanding basic mitochondrial genetics and the relationship between inherited mutations and disease phenotypes, and in identifying acquired mtDNA mutations in both ageing and cancer. However, many challenges remain, including the prevention and treatment of these diseases. This review explores the advances that have been made and the areas in which future progress is likely.

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Mitochondrial dna mutations in human disease

Since their first association with human disease in 1988, more than 250 pathogenic point mutations and rearrangements of the 16.6 kb mitochondrial genome (mtDNA) have been reported in a spectrum of clinical disorders which exhibit prominent muscle and central nervous system involvement. With novel mutations and disease phenotypes still being described, mtDNA disorders are recognized collectively as common, inherited genetic diseases although relatively little is still known concerning the precise pathophysiological mechanisms that lead to cell dysfunction and pathology. This review considers the basic principles of mitochondrial genetics which govern both the behaviour and investigation of pathogenic mtDNA mutations summarizing recent advances in this area, and an assessment of the ongoing debate into the role of somatic mtDNA mutations in neurodegenerative disease, ageing and cancer.

https://iubmb.onlinelibrary.wiley.com/doi/pdf/10.1080/15216540600686888

Mitochondrial DNA mutations in human disease.

Mitochondrial dysfunction has been linked to a wide range of degenerative and metabolic diseases, cancer, and aging. All these clinical manifestations arise from the central role of bioenergetics in cell biology. Although genetic therapies are maturing as the rules of bioenergetic genetics are clarified, metabolic therapies have been ineffectual. This failure results from our limited appreciation of the role of bioenergetics as the interface between the environment and the cell. A systems approach, which, ironically, was first successfully applied over 80 years ago with the introduction of the ketogenic diet, is required. Analysis of the many ways that a shift from carbohydrate glycolytic metabolism to fatty acid and ketone oxidative metabolism may modulate metabolism, signal transduction pathways, and the epigenome gives us an appreciation of the ketogenic diet and the potential for bioenergetic therapeutics.

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Mitochondrial Energetics and Therapeutics

Mitochondrial DNA mutations and human disease

Mitochondrial function in the human oocyte and embryo and their role in developmental competence.

Cystic fibrosis (CF) is the first monogenic disorder for which single cell preimplantation genetic diagnosis (PGD) has been successfully applied. The spectrum of mutations in CF is extremely heterogeneous, and hence, the development of mutation-specific PGD protocols is impracticable. The current study reports the development and evaluation of a general multiplex marker polymerase chain reaction (PCR) protocol for PGD of CF. Four closely linked highly polymorphic (CA)(n) repeat markers D7S523, D7S486, D7S480 and D7S490, flanking the cystic fibrosis transmembrane regulator (CFTR) gene, were used. In 99% of the single cells tested (100 leukocytes and 50 blastomeres), multiplex PCR results were obtained and the overall allelic drop out (ADO) rate varied from 2 to 5%. After validation for the presence of ADO and additional alleles, 95% of the multiplex PCR results were accepted to construct the marker genotypes. Depending on the genotype of the couple, and taking into account the embryos lost for transfer due to validation criteria (5%), ADO (0-2%) and single recombination (1.1-3%), in general >90% of the embryos could be reliably genotyped by PGD using a single blastomere. The risk of misdiagnosis equals the chance of a double recombination between informative flanking markers and is <0.05%. Therefore, this polymorphic and multi-allelic marker system is a reliable and generally applicable alternative for mutation-directed PGD protocols. Furthermore, it provides a test for the origin of the detected genotype and also gives an indication of the chromosomal ploidy status of the blastomere tested.

Multiplex PCR of polymorphic markers flanking the CFTR gene; a general approach for preimplantation genetic diagnosis of cystic fibrosis.

Diseases owing to defects of oxidative phosphorylation (OXPHOS) affect approximately 1 in 8000 individuals. Clinical manifestations can be extremely variable and range from single-affected tissues to multisystemic syndromes. In general, tissues with a high energy demand, like brain, heart and muscle, are affected. The OXPHOS system is under dual genetic control, and mutations in both nuclear and mitochondrial genes can cause OXPHOS diseases. The expression and segregation of mitochondrial DNA (mtDNA) mutations is different from nuclear gene defects. The mtDNA mutations can be either homoplasmic or heteroplasmic and in the latter case disease becomes manifest when the mutation exceeds a tissue-specific threshold. This mutation load can vary between tissues and often an exact correlation between mutation load and phenotypic expression is lacking. The transmission of mtDNA mutations is exclusively maternal, but the mutation load between embryos can vary tremendously because of a segregational bottleneck. Diseases by nuclear gene mutations show a normal Mendelian inheritance pattern and often have a more constant clinical manifestation. Given the prevalence and severity of OXPHOS disorders and the lack of adequate therapy, existing and new methods for the prevention of transmission of OXPHOS disorders, like prenatal diagnosis (PND), preimplantation genetic diagnosis (PGD), cytoplasmic transfer (CT) and nuclear transfer (NT), are technically and ethically evaluated.

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The transmission of OXPHOS disease and methods to prevent this

A family presented with three affected children with Leigh syndrome, a progressive neurodegenerative disorder. Analysis of the OXPHOS complexes in muscle of two affected patients showed an increase in activity of pyruvate dehydrogenase and a decrease of complex V activity. Mutation analysis revealed the T9176C mutation in the mtATPase 6 gene (OMIM 516060) and the mutation load was above 90% in the patients. Unaffected maternal relatives were tested for carrier-ship and one of them, with a mutation load of 55% in blood, was pregnant with her first child. The possibility of prenatal diagnosis was evaluated. The main problem was the lack of data on genotype-phenotype associations for the T9176C mutation and on variation of the mutation percentage in tissues and in time. Therefore, multiple tissues of affected and unaffected carriers were analysed. Eventually, prenatal diagnosis was offered with understanding by the couple that there could be considerable uncertainty in the interpretation of the results. Prenatal diagnosis was carried out twice on cultured and uncultured chorion villi and amniotic fluid cells. The result was a mutation percentage just below the assumed threshold of expression (90%). The couple decided to continue the pregnancy and an apparently healthy child was born with an as yet unclear prognosis. This is the first prenatal diagnosis for a carrier of the T9176C mutation. Prenatal diagnosis for this mutation is technically reliable, but the prognostic predictions are not straightforward.

Transmission and prenatal diagnosis of the T9176C mitochondrial DNA mutation

Pearson syndrome is an often fatal multisystem disease associated with mitochondrial DNA rearrangements. Here we report a patient with a novel mtDNA deletion of 3.4 kb ranging from nucleotides 6097 to 9541 in combination with deletion dimers. The mutation percentage in different tissues (blood, muscle and liver) varied between 64% and 95%. After a remission period of about a year, the patient suddenly died at the age of 3 years owing to a severe lactic acidosis. A second patient with a previously reported deletion of 8 kb and a milder phenotype was found to have mitochondrial duplications and died at the age of 10 years. From these data and data from previous reports, we hypothesize that duplications might be beneficial in the clinical course of the disease and in life expectancy.

Pearson syndrome and the role of deletion dimers and duplications in the mtDNA.

DNA microarrays as a method to monitor changes in mitochondria-related gene expression in patients with OXPHOS defects and/or mitochondrial cardiomyopathy

L.J.A.M. Jacobs

Papers

Multiplex PCR of polymorphic markers flanking the CFTR gene; a general approach for preimplantation genetic diagnosis of cystic fibrosis.

The transmission of OXPHOS disease and methods to prevent this

Transmission and prenatal diagnosis of the T9176C mitochondrial DNA mutation

Pearson syndrome and the role of deletion dimers and duplications in the mtDNA.

DNA microarrays as a method to monitor changes in mitochondria-related gene expression in patients with OXPHOS defects and/or mitochondrial cardiomyopathy