Mitochondrial Encephalomyopathies: Defects of Nuclear DNA

The term "mitochondrial diseases" encompasses a heterogeneous group of disorders in which a primary mitochondrial dysfunction is suspected or proven by morphologic, genetic, or biochemical criteria. Clinically, these progressive disorders usually affect muscle, either alone (mitochondrial myopathies) or in combination with other systems, most often brain (encephalomyopathies). Mitochondria are unique among intracellular organelles in that mitochondrial proteins are encoded by two genomes, nuclear DNA (nDNA) and mitochondrial DNA (mtDNA). The vast majority of mitochondrial proteins are encoded by the nuclear genome, whereas mtDNA (a circular, double stranded 16.5 kb molecule) encodes only 13 polypeptides, all of them subunits of respiratory chain complexes. In addition to structural genes, mtDNA also codes for 22 transfer RNAs and two ribosomal RNAs. Our understanding of mitochondrial diseases has grown at an impressive rate in the past few years, and most of the progress has been in the area of mtDNA genetics, where several mtDNA mutations have been associated with specific diseases (reviewed in this issue by Zeviani et al.). In comparison, our understanding of mitochondrial disorders due to nDNA lesions has lagged behind and, to date, molecular defects of nuclear genes have been documented in only a few patients. We will review which alterations in the nuclear genome can cause mitochondrial disorders and which criteria are useful in identifying such mutations. While several examples will be provided, this is not intended as a complete review of the subject.     

Mitochondrial DNA and disease

Mitochondrial DNA (mtDNA) defects are a relatively common cause of inherited disease and have been implicated in both ageing and cancer. MtDNA encodes essential subunits of the mitochondrial respiratory chain and defects result in impaired oxidative phosphorylation (OXPHOS). Similar OXPHOS defects have been shown to be present in a number of neurodegenerative conditions, including Parkinson's disease, as well as in normal ageing human tissues. Additionally, a number of tumours have been shown to contain mtDNA mutations and an altered metabolic phenotype. In this review we outline the unique characteristics of mitochondrial genetics before detailing important pathological features of mtDNA diseases, focusing on adult neurological disease as well as the role of mtDNA mutations in neurodegenerative diseases, ageing and cancer.     

Mitochondrial DNA Mutations and Mitochondrial Abnormalities in Dilated Cardiomyopathy

Mitochondrial (mt)DNA defects, both deletions and tRNA point mutations, have been associated with cardiomyopathies. The aim of the study was to determine the prevalence of pathological mtDNA mutations and to assess associated defects of mitochondrial enzyme activity in dilated cardiomyopathy (DCM) patients with ultrastructural abnormalities of cardiac mitochondria. In a large cohort of 601 DCM patients we performed conventional light and electron microscopy on endomyocardial biopsy samples. Cases with giant organelles, angulated, tubular, and concentric cristae, and crystalloid or osmiophilic inclusion bodies were selected for mtDNA analysis. Mutation screening techniques, automated DNA sequencing, restriction enzyme digestion, and densitometric assays were performed to identify mtDNA mutations, assess heteroplasmy, and quantify the amount of mutant in myocardial and blood DNA. Of 601 patients (16 to 63 years; mean, 43.5 ± 12.7 years), 85 had ultrastructural evidence of giant organelles, with abnormal cristae and inclusion bodies; 19 of 85 (22.35%) had heteroplasmic mtDNA mutations (9 tRNA, 5 rRNA, and 4 missense, one in two patients) that were not found in 111 normal controls and in 32 DCM patients without the above ultrastructural mitochondrial abnormalities. In all cases, the amount of mutant was higher in heart than in blood. In hearts of patients that later underwent transplantation, cytochrome c oxidase (Cox) activity was significantly lower in cases with mutations than in those without or controls (P = 0.0008). NADH dehydrogenase activity was only slightly reduced in cases with mutations (P = 0.0388), whereas succinic dehydrogenase activity did not significantly differ between DCM patients with mtDNA mutations and those without or controls. The present study represents the first attempt to detect a morphological, easily identifiable marker to guide mtDNA mutation screening. Pathological mtDNA mutations are associated with ultrastructurally abnormal mitochondria, and reduced Cox activity in a small subgroup of non-otherwise-defined, idiopathic DCMs, in which mtDNA defects may constitute the basis for, or contribute to, the development of congestive heart failure.     

Mitochondrial DNA deletion and sarcopenia

PurposeAccumulation of mitochondrial DNA deletions and the resultant impaired oxidative phosphorylation may play a pathogenic role in the mediation of age-related sarcopenia.MethodsTwenty four participants of the New Mexico Aging Process Study were classified as normal lean (n = 15) or sarcopenic (n = 9) based on body composition determined by Dual Energy x-ray Absorptiometry. Complex I and Complex IV activities were measured in the skeletal muscle samples obtained from gastrocnemius muscle. A two-stage nested polymerase chain reaction strategy was used to identify the mitochondrial DNA deletions in the entire mitochondrial genome in the skeletal muscle samples.ResultsAlthough Complex I activity was not significantly different (5.5 ± 0.9 vs. 4.6 ± 0.7 mU/mg protein, P > 0.05), Complex IV activity was higher in sarcopenic subjects (1.4 ± 0.3 vs. 1.0 ± 0.1 mU/mg protein, P < 0.05). Mitochondrial DNA deletions were mostly located in the region of Complex I and spanned from nicotinamide adenine dinucleotide dehydrogenase 1 to nicotinamide adenine dinucleotide dehydrogenase 6. Deletions in the 8,577–10,407 bp and 10,233–11,249 bp regions were associated with a significant decrease in Complex I activity (P < 0.05 and P = 0.02, respectively). Total cumulative deletion, defined as the sum of individual length of deletions in a subject, was comparable in subjects with and without sarcopenia (1760 ± 726 vs. 1782 ± 888 bp, P > 0.05). The magnitude of mitochondrial DNA deletion, however, correlated positively with lean body mass (r = 0.43, P < 0.05).ConclusionThus, mitochondrial DNA deletions are common in elderly subjects and are negatively related to Complex I activity. The positive association between mitochondrial DNA deletions and lean body mass needs to be confirmed by studies in a larger study population.     

线粒体DNA损伤与细胞凋亡

细胞核DNA（nuclear DNA,nDNA）损伤后诱导细胞凋亡的信号转导途径已经逐渐清晰,而线粒体DNA（mitochondrial DNA,mtDNA）损伤与细胞凋亡之间的关系尚有待进一步明确.目前已有研究结果表明：mtDNA断裂、缺失或功能缺陷能够显著影响细胞凋亡发生率;线粒体缺失细胞（p0细胞）同其亲代细胞相比,对多种凋亡诱导因素的反应存在显著差异;ROS的产生并非mtDNA损伤诱导凋亡的必要条件,mtDNA损伤断裂本身可能启动了凋亡的级联反应.但mtDNA的损伤,在细胞凋亡的信号转导途径具体扮演何种角色,还有待深入研究.     

Mitochondrial DNA polymorphisms in Thailand

Nucleotide sequences of the D-loop region of human mitochondrial DNA from six small ethnic groups of Thailand i.e., Hill tribes (Lisu and Mussur), Phuthai, Lao Song, Chong, and aboriginal Sakai, were analyzed. The sequences were compared with those of native Thai populations from two provinces, Chiang Mai and Khon Kaen. Based on a comparison of the 563-bp sequences in 215 Thai individuals, 137 different sequence types were observed. Of these, 124 were unique to their respective populations, whereas 13 were shared between two to five populations. The intergenic COII/tRNA^Lys 9-bp deletion was observed in every Thai population examined, except for the Sakai, with varying frequencies ranging from 18% to 40%. The D-loop sequences variation, and phylogenetic analysis, suggested that the 9-bp deletion had occurred in a very ancient ancestry of Southeast Asians, although multiple origins of the deletion cannot be ruled out. Genetic distances, based on net nucleotide diversities, between populations revealed that the Sakai were distantly related to the other Thai populations, while the Lao Song and Chong were closely related to each other. Close genetic affinities were also observed among the Hill tribes, Phuthai, and native northeast Thai (Khon Kaen), indicating that they may share some degree of the common ancestral maternal lineages. Received: October 23, 2000 / Accepted: December 4, 2000     

Mitochondrial DNA mutations in human disease

The small, maternally inherited mitochondrial DNA (mtDNA) has turned out to be a Pandora's box of pathogenic mutations: 13 years into the era of "molecular mitochondrial medicine," more than 100 pathogenic point mutations and innumerable rearrangements have been associated with a striking variety of multisystemic as well as tissue-specific human diseases. After reviewing the principles of mitochondrial genetics, we consider disorders due to mutations in genes affecting mitochondrial protein synthesis and disorders due to mutations in protein-coding genes. In contrast to the remarkable progress in our understanding of etiology, pathogenesis is only partially explained by the rules of mitochondrial genetics and remains largely unclear. We review recent progress in prenatal diagnosis, epidemiology, and in the development of animal models harboring mtDNA mutations.     

Mitochondrial DNA polymorphisms in bipolar disorder

BACKGROUND: Previous studies suggested mitochondrial abnormality in bipolar disorder: (1) possible contribution of parent-of-origin effect in transmission of bipolar disorder; (2) abnormal brain phosphorus metabolism detected by phosphorus-31 magnetic resonance spectroscopy; (3) comorbidity of affective disorders in patients with mitochondrial encephalopathy; (4) increased levels of the 4977bp deletion of mitochondrial DNA (mtDNA) in the postmortem brains. We investigated mtDNA polymorphisms in association with bipolar disorder. METHODS: Twelve PCR fragments including all tRNA genes were examined by the single-strand conformation polymorphism method in 43 bipolar patients. All observed polymorphisms were sequenced. Association of these polymorphisms with bipolar disorder was examined by restriction fragment length polymorphism method in 135 bipolar patients and 187 controls. RESULTS: In total, we found 28 polymorphisms including 14 polymorphisms that have not been reported previously. The A10398G polymorphism was significantly associated with bipolar disorder (10398A genotype: 33.1% in bipolar, 22.2% in the control, P<0.05). Although this difference was not significant after Bonferroni correction, the CA haplotype of the 5178 and 10398 polymorphisms was still significantly associated with bipolar disorder (CA haplotype: 33.6% in bipolar, 16.8% in control, P<0.001). Three rare mutations substituting evolutionary conserved bases; A5539G in tRNA(Trp) gene, A5747G in the origin of L-strand replication, and A8537G in ATPase subunit-6 and -8 genes, were found in patients with family history in which maternal transmission was suspected. DISCUSSION: The 5178C/10398A haplotype in mtDNA may be a risk factor of bipolar disorder (odds ratio, 2.4). Pathophysiological significance of rare mtDNA mutations needs to be verified in the future. This finding may imply the pathophysiological significance of mtDNA in bipolar disorder.     

Mitochondrial DNA: Epigenetics and environment

Maintenance of the mitochondrial genome is essential for proper cellular function. For this purpose, mitochondrial DNA (mtDNA) needs to be faithfully replicated, transcribed, translated, and repaired in the face of constant onslaught from endogenous and environmental agents. Although only 13 polypeptides are encoded within mtDNA, the mitochondrial proteome comprises over 1500 proteins that are encoded by nuclear genes and translocated to the mitochondria for the purpose of maintaining mitochondrial function. Regulation of mtDNA and mitochondrial proteins by epigenetic changes and post-translational modifications facilitate crosstalk between the nucleus and the mitochondria and ultimately lead to the maintenance of cellular health and homeostasis. DNA methyl transferases have been identified in the mitochondria implicating that methylation occurs within this organelle; however, the extent to which mtDNA is methylated has been debated for many years. Mechanisms of demethylation within this organelle have also been postulated, but the exact mechanisms and their outcomes is still an active area of research. Mitochondrial dysfunction in the form of altered gene expression and ATP production, resulting from epigenetic changes, can lead to various conditions including aging-related neurodegenerative disorders, altered metabolism, changes in circadian rhythm, and cancer. Here, we provide an overview of the epigenetic regulation of mtDNA via methylation, long and short noncoding RNAs, and post-translational modifications of nucleoid proteins (as mitochondria lack histones). We also highlight the influence of xenobiotics such as airborne environmental pollutants, contamination from heavy metals, and therapeutic drugs on mtDNA methylation. Environ. Mol. Mutagen., 60:668–682, 2019. © 2019 Wiley Periodicals, Inc.     

Mitochondrial DNA sequence analysis in Sicily

This study reports data on the sequences of the first hypervariable segment of a sample of the Sicilian population from Alia (Palermo, Italy). The results show the presence of 32 different haplotypes in the 49 individuals examined. The average number of pairwise nucleotide differences was 4.04, i.e., 1.17% per nucleotide. The distribution of the nucleotide differences matches the theoretical distribution and indicates only one major episode of expansion that occurred between 20,732 and 59,691 years ago, between the Middle Paleolithic and Upper Paleolithic. Compared with the other populations, parameters of the Sicilian sample lie in an intermediate position between the eastern and western Mediterranean populations. This is due to numerous contacts that Sicily has had with the Mediterranean area since prehistoric times. At the same time, the singularity of some of the haplotypes present in the sample studied indicates the persistence of some characteristics caused by genetic drift and isolation that the population has endured in the course of its history. Am. J. Hum. Biol. 13:576–589, 2001. © 2001 Wiley‐Liss, Inc.     

Mitochondrial DNA polymorphisms and extraversion.

Mitochondria is the major site of energy production in cells, therefore, mitochondrial abnormality may affect functions of organs including the brain, which constantly requires high levels of energy consumption. Previous studies have suggested a role of mitochondria and their DNA polymorphisms in neuro-psychiatric disorders, including Alzheimer's disease, Parkinson's disease, schizophrenia and bipolar mood disorder. Thus, we hypothesized that mitochondrial DNA polymorphisms might be related with the development of personality. The present study investigated a role of two mitochondrial DNA polymorphisms, the C5178A and A10398G, in personality traits evaluated using the NEO PI-R scores in 238 healthy Japanese volunteers. Subjects with the 5178A genotype showed significantly higher extraversion score than those with the 5178C genotype (P = 0.027), while no significant association was observed between the C5178A polymorphism and other scores. No significant association was found between the A10398G polymorphism and any scores. Regarding the 5178-10398 haplotype, the score of extraversion, not other scores, was significantly associated with the A-G haplotype (P = 0.042). Although further studies are recommended for the confirmation, the result may suggest a role of the mitochondrial DNA polymorphism in the personality trait.     

Mitochondrial DNA Variation in Karkar Islanders

We analyzed 375 base pairs (bp) of the first hypervariable region (HVS‐I) of the mitochondrial DNA (mtDNA) control region and intergenic COII/tRNA^Lys 9‐bp deletion from 47 Karkar Islanders (north coast of Papua New Guinea) belonging to the Waskia Papuan language group. To address questions concerning the origin and evolution of this population we compared the Karkar mtDNA haplotypes and haplogroups to those of neighbouring East Asians and Oceanic populations. The results of the phylogeographic analysis show grouping in three different clusters of the Karkar Islander mtDNA lineages: one group of lineages derives from the first Pleistocene settlers of New Guinea‐Island Melanesia, a second set derives from more recent arrivals of Austronesian speaking populations, and the third contains lineages specific to the Karkar Islanders, but still rooted to Austronesian and New Guinea‐Island Melanesia populations. Our results suggest (i) the absence of a strong association between language and mtDNA variation and, (ii) reveal that the mtDNA haplogroups F1a1, M7b1 and E1a, which probably originated in Island Southeast Asia and may be considered signatures of Austronesian population movements, are preserved in the Karkar Islanders but absent in other New Guinea‐Island Melanesian populations. These findings indicate that the Karkar Papuan speakers retained a certain degree of their own genetic uniqueness and a high genetic diversity. We present a hypothesis based on archaeological, linguistic and environmental datasets to argue for a succession of (partial) depopulation and repopulation and expansion events, under conditions of structured interaction, which may explain the variability expressed in the Karkar mtDNA.     

Mitochondrial DNA Heterogeneity in Tunisian Berbers

Berbers live in groups scattered across North Africa whose origins and genetic relationships with their neighbours are not well established. The first hypervariable segment of the mitochondrial DNA (mtDNA) control region was sequenced in a total of 155 individuals from three Tunisian Berber groups and compared to other North Africans. The mtDNA lineages found belong to a common set of mtDNA haplogroups already described in North Africa. Besides the autochthonous North African U6 haplogroup, a group of L3 lineages characterized by the transition at position 16041 seems to be restricted to North Africans, suggesting that an expansion of this group of lineages took place around 10500 years ago in North Africa, and spread to neighbouring populations. Principal components and the coordinate analyses show that some Berber groups (the Tuareg, the Mozabite, and the Chenini‐Douiret) are outliers within the North African genetic landscape. This outlier position is consistent with an isolation process followed by genetic drift in haplotype frequencies, and with the high heterogeneity displayed by Berbers compared to Arab samples as shown in the AMOVA. Despite this Berber heterogeneity, no significant differences were found between Berber and Arab samples, suggesting that the Arabization was mainly a cultural process rather than a demographic replacement.     

Disorders Associated with Depletion of Mitochondrial DNA

Quantitative defects of mtDNA have been recently described in patients with fatal mitochondrial disease of early infancy or mitochondrial myopathy of childhood. There was variable tissue expression and depletion of up to 98% of mtDNA in affected tissues. Pedigree analysis was compatible with mendelian inheritance, suggesting faulty communication between nuclear and mitochondrial genomes, but the primary molecular lesion is unknown. In muscle, morphological studies allowed to correlate mtDNA depletion, absence of mtDNA-encoded peptides, mitochondrial proliferation, and loss of cytochrome c oxidase (COX) activity in individual fibers.     

Mitochondrial DNA analysis of Sporothrix schenckii]

Restriction fragment length polymorphism (RFLP) in mitochondrial DNA (mtDNA) of clinical and environmental isolates of Sporothrix schenckii was investigated. Isolates of S. schenckii were classified into 24 mtDNA types (Types 1-24) based on mtDNA RFLP patterns with HaeIII and clustered into two major groups by phylogeny, Group A and Group B. Group A isolates are predominant in South Africa, North America, Central America and South America, while Group B isolates are predominant in Australia, Japan and China. Based on the mtDNA-RFLP patterns with HaeIII, most environmental isolates morphologically identified as S. schenckii were confirmed to be species distinct from S. schenckii and S. schenckii isolates were few, while all of more than 500 clinical isolates were confirmed as S. schenckii. Therefore, RFLP analysis of mtDNA is essential for the identification of environmental, but not clinical isolates of S. schenckii.     

Mitochondrial DNA analysis of acellular laboratory samples

The source of acellular specimens is not infrequently challenged, especially for toxicology specimens, but such specimens may not be amenable to conventional genetic testing to confirm the source. Our evaluation of genomic and mitochondrial DNA (mtDNA) amplicons using polymerase chain reaction (PCR) from centrifuged, filtered, or whole urine specimens demonstrated higher sensitivity of detection of mtDNA than genomic DNA and a higher detection rate for the mtDNA markers than genomic markers in all sample sets. The mitochondrial amplicons were sequenced to identify specific single nucleotide polymorphisms (SNPs). Subsequently, a real-time PCR technique using fluorescence resonance energy transfer (FRET) probes designed to hybridize to the published mtDNA sequence over known SNP locations present within the mitochondrial control region was developed. Our results demonstrate the feasibility of using a FRET-based assay of mitochondrial genotypes with acellular laboratory specimens to screen for specimen mix-ups or to confirm sources of controversial toxicology specimens.