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Sample records for mitochondrial andnuclear dna

  1. The complete mitochondrial genome of the enigmatic bigheadedturtle (Platysternon): description of unusual genomic features and thereconciliation of phylogenetic hypotheses based on mitochondrial andnuclear DNA

    Energy Technology Data Exchange (ETDEWEB)

    Parham, James F.; Feldman, Chris R.; Boore, Jeffrey L.

    2005-12-28

    The big-headed turtle (Platysternon megacephalum) from east Asia is the sole living representative of a poorly-studied turtle lineage (Platysternidae). It has no close living relatives, and its phylogenetic position within turtles is one of the outstanding controversies in turtle systematics. Platysternon was traditionally considered to be close to snapping turtles (Chelydridae) based on some studies of its morphology and mitochondrial (mt) DNA, however, other studies of morphology and nuclear (nu) DNA do not support that hypothesis. We sequenced the complete mt genome of Platysternon and the nearly complete mt genomes of two other relevant turtles and compared them to turtle mt genomes from the literature to form the largest molecular dataset used to date to address this issue. The resulting phylogeny robustly rejects the placement of Platysternon with Chelydridae, but instead shows that it is a member of the Testudinoidea, a diverse, nearly globally-distributed group that includes pond turtles and tortoises. We also discovered that Platysternon mtDNA has large-scale gene rearrangements and possesses two, nearly identical, control regions, features that distinguish it from all other studied turtles. Our study robustly determines the phylogenetic placement of Platysternon and provides a well-resolved outline of major turtle lineages, while demonstrating the significantly greater resolving power of comparing large amounts of mt sequence over that of short fragments. Earlier phylogenies placing Platysternon with chelydrids required a temporal gap in the fossil record that is now unnecessary. The duplicated control regions and gene rearrangements of the Platysternon mt DNA probably resulted from the duplication of part of the genome and then the subsequent loss of redundant genes. Although it is possible that having two control regions may provide some advantage, explaining why the control regions would be maintained while some of the duplicated genes were eroded

  2. What Is Mitochondrial DNA?

    Science.gov (United States)

    ... DNA What is mitochondrial DNA? What is mitochondrial DNA? Although most DNA is packaged in chromosomes within ... proteins. For more information about mitochondria and mitochondrial DNA: Molecular Expressions, a web site from the Florida ...

  3. Defects of mitochondrial DNA replication.

    Science.gov (United States)

    Copeland, William C

    2014-09-01

    Mitochondrial DNA is replicated by DNA polymerase γ in concert with accessory proteins such as the mitochondrial DNA helicase, single-stranded DNA binding protein, topoisomerase, and initiating factors. Defects in mitochondrial DNA replication or nucleotide metabolism can cause mitochondrial genetic diseases due to mitochondrial DNA deletions, point mutations, or depletion, which ultimately cause loss of oxidative phosphorylation. These genetic diseases include mitochondrial DNA depletion syndromes such as Alpers or early infantile hepatocerebral syndromes, and mitochondrial DNA deletion disorders, such as progressive external ophthalmoplegia, ataxia-neuropathy, or mitochondrial neurogastrointestinal encephalomyopathy. This review focuses on our current knowledge of genetic defects of mitochondrial DNA replication (POLG, POLG2, C10orf2, and MGME1) that cause instability of mitochondrial DNA and mitochondrial disease.

  4. Replicating animal mitochondrial DNA

    Directory of Open Access Journals (Sweden)

    Emily A. McKinney

    2013-01-01

    Full Text Available The field of mitochondrial DNA (mtDNA replication has been experiencing incredible progress in recent years, and yet little is certain about the mechanism(s used by animal cells to replicate this plasmid-like genome. The long-standing strand-displacement model of mammalian mtDNA replication (for which single-stranded DNA intermediates are a hallmark has been intensively challenged by a new set of data, which suggests that replication proceeds via coupled leading-and lagging-strand synthesis (resembling bacterial genome replication and/or via long stretches of RNA intermediates laid on the mtDNA lagging-strand (the so called RITOLS. The set of proteins required for mtDNA replication is small and includes the catalytic and accessory subunits of DNA polymerase y, the mtDNA helicase Twinkle, the mitochondrial single-stranded DNA-binding protein, and the mitochondrial RNA polymerase (which most likely functions as the mtDNA primase. Mutations in the genes coding for the first three proteins are associated with human diseases and premature aging, justifying the research interest in the genetic, biochemical and structural properties of the mtDNA replication machinery. Here we summarize these properties and discuss the current models of mtDNA replication in animal cells.

  5. Mitochondrial DNA Alterations and Reduced Mitochondrial Function in Aging

    OpenAIRE

    Hebert, Sadie L.; Lanza, Ian R.; Nair, K. Sreekumaran

    2010-01-01

    Oxidative damage to mitochondrial DNA increases with aging. This damage has the potential to affect mitochondrial DNA replication and transcription which could alter the abundance or functionality of mitochondrial proteins. This review describes mitochondrial DNA alterations and changes in mitochondrial function that occur with aging. Age-related alterations in mitochondrial DNA as a possible contributor to the reduction in mitochondrial function are discussed.

  6. Mitochondrial Myopathy with DNA Deletions

    OpenAIRE

    J Gordon Millichap

    1992-01-01

    Deletions of mitochondrial DNA (mtDNA) are reported in 19 of 56 patients with mitochondrial myopathy examined in the Department of Neurology and Neuromuscular Research Laboratory, Mayo Clinic, Rochester, MN.

  7. Mitochondrial DNA maintenance: an appraisal.

    Science.gov (United States)

    Akhmedov, Alexander T; Marín-García, José

    2015-11-01

    Mitochondria play a crucial role in a variety of cellular processes ranging from energy metabolism, generation of reactive oxygen species (ROS), and Ca(2+) handling to stress responses, cell survival, and death. Malfunction of the organelle may contribute to the pathogenesis of neuromuscular disorders, cancer, premature aging, and cardiovascular diseases, including myocardial ischemia, cardiomyopathy, and heart failure. Mitochondria are unique as they contain their own genome organized into DNA-protein complexes, so-called mitochondrial nucleoids, along with multiprotein machineries, which promote mitochondrial DNA (mtDNA) replication, transcription, and repair. Although the organelle possesses almost all known nuclear DNA repair pathways, including base excision repair, mismatch repair, and recombinational repair, the proximity of mtDNA to the main sites of ROS production and the lack of protective histones may result in increased susceptibility to oxidative stress and other types of mtDNA damage. Defects in the components of these highly organized machineries, which mediate mtDNA maintenance (replication and repair), may result in accumulation of point mutations and/or deletions in mtDNA and decreased mtDNA copy number impairing mitochondrial function. This review will focus on the mechanisms of mtDNA maintenance with emphasis on the proteins implicated in these processes and their functional role in various disease conditions and aging.

  8. Implications of mitochondrial DNA mutations and mitochondrial dysfunction in tumorigenesis

    Institute of Scientific and Technical Information of China (English)

    Jianxin Lu; Lokendra Kumar Sharma; Yidong Bai

    2009-01-01

    Alterations in oxidative phosphorylation resulting from mitochondrial dysfunction have long been hypothesized to be involved in tumorigenesis. Mitochondria have recently been shown to play an important role in regulating both programmed cell death and cell proliferation. Furthermore, mitochondrial DNA (mtDNA) mutations have been found in various cancer cells. However, the role of these mtDNA mutations in tumorigenesis remains largely unknown. This review focuses on basic mitochondrial genetics, mtDNA mutations and consequential mitochondrial dysfunction associated with cancer. The potential molecular mechanisms, mediating the pathogenesis from mtDNA mutations and mitochondrial dysfunction to tumorigenesis are also discussed.

  9. Mitochondrial DNA inheritance after SCNT.

    Science.gov (United States)

    Hiendleder, Stefan

    2007-01-01

    Mitochondrial biogenesis and function is under dual genetic control and requires extensive interaction between biparentally inherited nuclear genes and maternally inherited mitochondrial genes. Standard SCNT procedures deprive an oocytes' mitochondrial DNA (mtDNA) of the corresponding maternal nuclear DNA and require it to interact with an entirely foreign nucleus that is again interacting with foreign somatic mitochondria. As a result, most SCNT embryos, -fetuses, and -offspring carry somatic cell mtDNA in addition to recipient oocyte mtDNA, a condition termed heteroplasmy. It is thus evident that somatic cell mtDNA can escape the selective mechanism that targets and eliminates intraspecific sperm mitochondria in the fertilized oocyte to maintain homoplasmy. However, the factors responsible for the large intra- and interindividual differences in heteroplasmy level remain elusive. Furthermore, heteroplasmy is probably confounded with mtDNA recombination. Considering the essential roles of mitochondria in cellular metabolism, cell signalling, and programmed cell death, future experiments will need to assess the true extent and impact of unorthodox mtDNA transmission on various aspects of SCNT success.

  10. Mitochondrial DNA and Cancer Epidemiology Workshop

    Science.gov (United States)

    A workshop to review the state-of-the science in the mitochondrial DNA field and its use in cancer epidemiology, and to develop a concept for a research initiative on mitochondrial DNA and cancer epidemiology.

  11. Minisequencing mitochondrial DNA pathogenic mutations

    Directory of Open Access Journals (Sweden)

    Carracedo Ángel

    2008-04-01

    Full Text Available Abstract Background There are a number of well-known mutations responsible of common mitochondrial DNA (mtDNA diseases. In order to overcome technical problems related to the analysis of complete mtDNA genomes, a variety of different techniques have been proposed that allow the screening of coding region pathogenic mutations. Methods We here propose a minisequencing assay for the analysis of mtDNA mutations. In a single reaction, we interrogate a total of 25 pathogenic mutations distributed all around the whole mtDNA genome in a sample of patients suspected for mtDNA disease. Results We have detected 11 causal homoplasmic mutations in patients suspected for Leber disease, which were further confirmed by standard automatic sequencing. Mutations m.11778G>A and m.14484T>C occur at higher frequency than expected by change in the Galician (northwest Spain patients carrying haplogroup J lineages (Fisher's Exact test, P-value Conclusion We here developed a minisequencing genotyping method for the screening of the most common pathogenic mtDNA mutations which is simple, fast, and low-cost. The technique is robust and reproducible and can easily be implemented in standard clinical laboratories.

  12. The inheritance of pathogenic mitochondrial DNA mutations

    OpenAIRE

    Cree, L.M.; Samuels, D.C.; Chinnery, P F

    2009-01-01

    Abstract Mitochondrial DNA mutations cause disease in >1 in 5000 of the population, and ~1 in 200 of the population are asymptomatic carriers of a pathogenic mtDNA mutation. Many patients with these pathogenic mtDNA mutations present with a progressive, disabling neurological syndrome that leads to major disability and premature death. There is currently no effective treatment for mitochondrial disorders, placing great emphasis on preventing the transmission of these diseases. An e...

  13. Mitochondrial DNA, restoring Beethovens music.

    Science.gov (United States)

    Merheb, Maxime; Vaiedelich, Stéphane; Maniguet, Thiérry; Hänni, Catherine

    2016-01-01

    Great ancient composers have endured many obstacles and constraints which are very difficult to understand unless we perform the restoration process of ancient music. Species identification in leather used during manufacturing is the key step to start such a restoration process in order to produce a facsimile of a museum piano. Our study reveals the species identification in the leather covering the hammer head in a piano created by Erard in 1802. This is the last existing piano similar to the piano that Beethoven used with its leather preserved in its original state. The leather sample was not present in a homogeneous piece, yet combined with glue. Using a DNA extraction method that avoids PCR inhibitors; we discovered that sheep and cattle are the origin of the combination. To identify the species in the leather, we focused on the amounts of mitochondrial DNA in both leather and glue and results have led us to the conclusion that the leather used to cover the hammer head in this piano was made of cattle hide.

  14. Mitochondrial DNA sequence evolution in shorebird populations.

    NARCIS (Netherlands)

    Wenink, P.W.

    1994-01-01

    This thesis describes the global molecular population structure of two shorebird species, in particular of the dunlin, Calidris alpina, by means of comparative sequence analysis of the most variable part of the mitochondrial DNA (mtDNA) genome. There are several reasons why mtDNA is the molecule of

  15. Mitochondrial DNA plasticity is an essential inducer of tumorigenesis.

    Science.gov (United States)

    Lee, W T Y; Cain, J E; Cuddihy, A; Johnson, J; Dickinson, A; Yeung, K-Y; Kumar, B; Johns, T G; Watkins, D N; Spencer, A; St John, J C

    2016-01-01

    Although mitochondrial DNA has been implicated in diseases such as cancer, its role remains to be defined. Using three models of tumorigenesis, namely glioblastoma multiforme, multiple myeloma and osteosarcoma, we show that mitochondrial DNA plays defining roles at early and late tumour progression. Specifically, tumour cells partially or completely depleted of mitochondrial DNA either restored their mitochondrial DNA content or actively recruited mitochondrial DNA, which affected the rate of tumorigenesis. Nevertheless, non-depleted tumour cells modulated mitochondrial DNA copy number at early and late progression in a mitochondrial DNA genotype-specific manner. In glioblastoma multiforme and osteosarcoma, this was coupled with loss and gain of mitochondrial DNA variants. Changes in mitochondrial DNA genotype affected tumour morphology and gene expression patterns at early and late progression. Importantly, this identified a subset of genes that are essential to early progression. Consequently, mitochondrial DNA and commonly expressed early tumour-specific genes provide novel targets against tumorigenesis.

  16. (Somatic mutations in nuclear and mitochondrial DNA)

    Energy Technology Data Exchange (ETDEWEB)

    1992-01-01

    The study is concerned the design of new assays that may detect rare somatic mutations in nuclear and mitochondrial DNA, which may increase upon exposure to mutagens, and thus become a marker of human exposure to such mutagens. Two assays for somatic mutation were presented, one for mitochondrial DNA deletions which was developed by the author, and one for deletions of the ADA gene which resides in the nucleus.

  17. Preventing Mitochondrial Fission Impairs Mitochondrial Function and Leads to Loss of Mitochondrial DNA

    OpenAIRE

    Parone, Philippe A.; Sandrine Da Cruz; Daniel Tondera; Yves Mattenberger; James, Dominic I.; Pierre Maechler; François Barja; Jean-Claude Martinou

    2008-01-01

    Mitochondria form a highly dynamic tubular network, the morphology of which is regulated by frequent fission and fusion events. However, the role of mitochondrial fission in homeostasis of the organelle is still unknown. Here we report that preventing mitochondrial fission, by down-regulating expression of Drp1 in mammalian cells leads to a loss of mitochondrial DNA and a decrease of mitochondrial respiration coupled to an increase in the levels of cellular reactive oxygen species (ROS). At t...

  18. Role and Treatment of Mitochondrial DNA-Related Mitochondrial Dysfunction in Sporadic Neurodegenerative Diseases

    OpenAIRE

    Swerdlow, Russell H.

    2011-01-01

    Several sporadic neurodegenerative diseases display phenomena that directly or indirectly relate to mitochondrial function. Data suggesting altered mitochondrial function in these diseases could arise from mitochondrial DNA (mtDNA) are reviewed. Approaches for manipulating mitochondrial function and minimizing the downstream consequences of mitochondrial dysfunction are discussed.

  19. Mitochondrial DNA sequence evolution in shorebird populations

    NARCIS (Netherlands)

    Wenink, P.W.

    1994-01-01

    This thesis describes the global molecular population structure of two shorebird species, in particular of the dunlin, Calidris alpina, by means of comparative sequence analysis of the most variable part of the mitochondrial DNA (mtDNA) genome. There are several reasons

  20. [Mitochondrial DNA diversity in Kazym Khanty].

    Science.gov (United States)

    Naumova, O Iu; Khaiat, S Sh; Rychkov, S Iu

    2009-06-01

    New data on mitochondrial DNA polymorphism in the representatives of Kazym territorial group of Northern Khanty are presented. MtDNA diversity observed in Kazym Khanty was compared with that in Khanty from Shuryshkarskii raion of Yamalo-Nenets Autonomous Okrug.

  1. Mitochondrial DNA depletion analysis by pseudogene ratioing.

    Science.gov (United States)

    Swerdlow, Russell H; Redpath, Gerard T; Binder, Daniel R; Davis, John N; VandenBerg, Scott R

    2006-01-30

    The mitochondrial DNA (mtDNA) depletion status of rho(0) cell lines is typically assessed by hybridization or polymerase chain reaction (PCR) experiments, in which the failure to hybridize mtDNA or amplify mtDNA using mtDNA-directed primers suggests thorough mitochondrial genome removal. Here, we report the use of an mtDNA pseudogene ratioing technique for the additional confirmation of rho0 status. Total genomic DNA from a U251 human glioma cell line treated with ethidium bromide was amplified using primers designed to anneal either mtDNA or a previously described nuclear DNA-embedded mtDNA pseudogene (mtDNApsi). The resultant PCR product was used to generate plasmid clones. Sixty-two plasmid clones were genotyped, and all arose from mtDNApsi template. These data allowed us to determine with 95% confidence that the resultant mtDNA-depleted cell line contains less than one copy of mtDNA per 10 cells. Unlike previous hybridization or PCR-based analyses of mtDNA depletion, this mtDNApsi ratioing technique does not rely on interpretation of a negative result, and may prove useful as an adjunct for the determination of rho0 status or mtDNA copy number.

  2. Maternal inheritance and mitochondrial DNA variants in familial Parkinson's disease

    OpenAIRE

    Pfeiffer Ronald F; Rudolph Alice; Halter Cheryl A; Pauciulo Michael W; Kissell Diane K; Pankratz Nathan; Simon David K; Nichols William C; Foroud Tatiana

    2010-01-01

    Abstract Background Mitochondrial function is impaired in Parkinson's disease (PD) and may contribute to the pathogenesis of PD, but the causes of mitochondrial impairment in PD are unknown. Mitochondrial dysfunction is recapitulated in cell lines expressing mitochondrial DNA (mtDNA) from PD patients, implicating mtDNA variants or mutations, though the role of mtDNA variants or mutations in PD risk remains unclear. We investigated the potential contribution of mtDNA variants or mutations to t...

  3. Preventing mitochondrial fission impairs mitochondrial function and leads to loss of mitochondrial DNA.

    Directory of Open Access Journals (Sweden)

    Philippe A Parone

    Full Text Available Mitochondria form a highly dynamic tubular network, the morphology of which is regulated by frequent fission and fusion events. However, the role of mitochondrial fission in homeostasis of the organelle is still unknown. Here we report that preventing mitochondrial fission, by down-regulating expression of Drp1 in mammalian cells leads to a loss of mitochondrial DNA and a decrease of mitochondrial respiration coupled to an increase in the levels of cellular reactive oxygen species (ROS. At the cellular level, mitochondrial dysfunction resulting from the lack of fission leads to a drop in the levels of cellular ATP, an inhibition of cell proliferation and an increase in autophagy. In conclusion, we propose that mitochondrial fission is required for preservation of mitochondrial function and thereby for maintenance of cellular homeostasis.

  4. Mitochondrial DNA mutation in essential hypertension

    Institute of Scientific and Technical Information of China (English)

    Yuqi Liu; Shiwen Wang

    2008-01-01

    Essential hypertension (EH) is an escalating problem for developed and developing countries.It is currently seen as a 'complex' genetic trait caused by multiple susceptibility genes which are modulated by gene-environment and gene-gene interactions.Over the past 10 years,mitochondrial defects have been implicated in a wide variety of degenerative diseases,aging,and cancer.Recently several studies showed that human essential hypertension has excess maternal transmission which suggests a possible mitochondrial involvement.However,the exact pathophysiology of mitochondrial DNA mutation (mtDNA) in essential hypertension still remains perplexing.With the application of a variety of imaging approaches and successive mouse model of mitochonddal diseases we convince that these problems will be resolved in the near future.(J Geriatr Cardiol 2008;5(1):60-64)

  5. Mitochondrial DNA diagnosis for taeniasis and cysticercosis.

    Science.gov (United States)

    Yamasaki, Hiroshi; Nakao, Minoru; Sako, Yasuhito; Nakaya, Kazuhiro; Sato, Marcello Otake; Ito, Akira

    2006-01-01

    Molecular diagnosis for taeniasis and cysticercosis in humans on the basis of mitochondrial DNA analysis was reviewed. Development and application of three different methods, including restriction fragment length polymorphism analysis, base excision sequence scanning thymine-base analysis and multiplex PCR, were described. Moreover, molecular diagnosis of cysticerci found in specimens submitted for histopathology and the molecular detection of taeniasis using copro-DNA were discussed.

  6. [Hepatomioneuropathy secondary to mitochondrial DNA depletion].

    Science.gov (United States)

    Blanco-Barca, M O; Gómez-Lado, C; Campos-González, Y; Castro-Gago, M

    2007-04-01

    Mitochondrial DNA depletion (mtDNA) is an highly heterogeneous condition characterized by a decreased number of mtDNA copies. The patient is a 22-month-old girl with generalized hypotonia, marked weakness, respiratory failure, arterial hypertension, hyperlactacidemia, hepatosplenomegaly and mild hypertransaminasemia without hepatic failure neither hypoketotic hypoglycemia. Electromyographic findings were consistent with neuromyopathy and muscle biopsy suggested a neurogenic atrophy. Electron microscopy revealed lipid droplets, subsarcolemmal accumulation of mitochondrias and glycogen granules. Respiratory chain enzime activities were normal. Genetic study in muscle showed mtDNA depletion, and the diagnosis of spinal muscular atrophy caused by survival motoneuron gene deletion was excluded. This case might be a novel phenotype of mtDNA depletion which could be named hepatomioneuropatyc form. A normal result of respiratory chain enzimes in muscle doesn't excluded mtDNA depletion.

  7. Zen and the art of mitochondrial DNA maintenance.

    Science.gov (United States)

    Holt, Ian J

    2010-03-01

    Because mitochondrial genes encode proteins essential for aerobic ATP production, mitochondrial DNA defects can cause an energy crisis. These defects fall into two broad categories: primary mutations in mitochondrial DNA and mutations in nuclear genes, whose protein products are involved in mitochondrial DNA maintenance. Evidence is accumulating that both types of defects can cause mitochondrial DNA loss. Hence, regulatory factors, which determine whether mitochondrial DNA molecules are maintained or lost, potentially play a more important role in these disorders than hitherto recognised. Candidates include reactive oxygen species (ROS) and the tumour suppressor p53. The cell might not always be the best judge of when to dispense with the services of mitochondrial DNA, and so interventions that favour its retention could potentially limit the adverse effects of pathological mitochondrial DNAs.

  8. Restoration of normal embryogenesis by mitochondrial supplementation in pig oocytes exhibiting mitochondrial DNA deficiency.

    Science.gov (United States)

    Cagnone, Gael L M; Tsai, Te-Sha; Makanji, Yogeshwar; Matthews, Pamela; Gould, Jodee; Bonkowski, Michael S; Elgass, Kirstin D; Wong, Ashley S A; Wu, Lindsay E; McKenzie, Matthew; Sinclair, David A; St John, Justin C

    2016-03-18

    An increasing number of women fail to achieve pregnancy due to either failed fertilization or embryo arrest during preimplantation development. This often results from decreased oocyte quality. Indeed, reduced mitochondrial DNA copy number (mitochondrial DNA deficiency) may disrupt oocyte quality in some women. To overcome mitochondrial DNA deficiency, whilst maintaining genetic identity, we supplemented pig oocytes selected for mitochondrial DNA deficiency, reduced cytoplasmic maturation and lower developmental competence, with autologous populations of mitochondrial isolate at fertilization. Supplementation increased development to blastocyst, the final stage of preimplantation development, and promoted mitochondrial DNA replication prior to embryonic genome activation in mitochondrial DNA deficient oocytes but not in oocytes with normal levels of mitochondrial DNA. Blastocysts exhibited transcriptome profiles more closely resembling those of blastocysts from developmentally competent oocytes. Furthermore, mitochondrial supplementation reduced gene expression patterns associated with metabolic disorders that were identified in blastocysts from mitochondrial DNA deficient oocytes. These results demonstrate the importance of the oocyte's mitochondrial DNA investment in fertilization outcome and subsequent embryo development to mitochondrial DNA deficient oocytes.

  9. Quantification of human mitochondrial DNA using synthesized DNA standards.

    Science.gov (United States)

    Kavlick, Mark F; Lawrence, Helen S; Merritt, R Travis; Fisher, Constance; Isenberg, Alice; Robertson, James M; Budowle, Bruce

    2011-11-01

    Successful mitochondrial DNA (mtDNA) forensic analysis depends on sufficient quantity and quality of mtDNA. A real-time quantitative PCR assay was developed to assess such characteristics in a DNA sample, which utilizes a duplex, synthetic DNA to ensure optimal quality assurance and quality control. The assay's 105-base pair target sequence facilitates amplification of degraded DNA and is minimally homologous to nonhuman mtDNA. The primers and probe hybridize to a region that has relatively few sequence polymorphisms. The assay can also identify the presence of PCR inhibitors and thus indicate the need for sample repurification. The results show that the assay provides information down to 10 copies and provides a dynamic range spanning seven orders of magnitude. Additional experiments demonstrated that as few as 300 mtDNA copies resulted in successful hypervariable region amplification, information that permits sample conservation and optimized downstream PCR testing. The assay described is rapid, reliable, and robust.

  10. Mitochondrial DNA disease: new options for prevention.

    Science.gov (United States)

    Craven, Lyndsey; Elson, Joanna L; Irving, Laura; Tuppen, Helen A; Lister, Lisa M; Greggains, Gareth D; Byerley, Samantha; Murdoch, Alison P; Herbert, Mary; Turnbull, Doug

    2011-10-15

    Very recently, two papers have presented intriguing data suggesting that prevention of transmission of human mitochondrial DNA (mtDNA) disease is possible. [Craven, L., Tuppen, H.A., Greggains, G.D., Harbottle, S.J., Murphy, J.L., Cree, L.M., Murdoch, A.P., Chinnery, P.F., Taylor, R.W., Lightowlers, R.N. et al. (2010) Pronuclear transfer in human embryos to prevent transmission of mitochondrial DNA disease. Nature, 465, 82-85. Tachibana, M., Sparman, M., Sritanaudomchai, H., Ma, H., Clepper, L., Woodward, J., Li, Y., Ramsey, C., Kolotushkina, O. and Mitalipov, S. (2009) Mitochondrial gene replacement in primate offspring and embryonic stem cells. Nature, 461, 367-372.] These recent advances raise hopes for families with mtDNA disease; however, the successful translational of these techniques to clinical practice will require further research to test for safety and to maximize efficacy. Furthermore, in the UK, amendment to the current legislation will be required. Here, we discuss the clinical and scientific background, studies we believe are important to establish safety and efficacy of the techniques and some of the potential concerns about the use of these approaches.

  11. Mitochondrial DNA repair and association with aging--an update

    DEFF Research Database (Denmark)

    Diaz, Ricardo Gredilla; Bohr, Vilhelm A; Stevnsner, Tinna V.

    2010-01-01

    Mitochondrial DNA is constantly exposed to oxidative injury. Due to its location close to the main site of reactive oxygen species, the inner mitochondrial membrane, mtDNA is more susceptible than nuclear DNA to oxidative damage. The accumulation of DNA damage is thought to play a critical role...

  12. Mitochondrial transcription termination factor 2 binds to entire mitochondrial DNA and negatively regulates mitochondrial gene expression

    Institute of Scientific and Technical Information of China (English)

    Weiwei Huang; Min Yu; Yang Jiao; Jie Ma; Mingxing Ma; Zehua Wang; Hong Wu; Deyong Tan

    2011-01-01

    Mitochondrial transcription termination factor 2 (mTERF2) is a mitochondriai matrix protein that binds to the mitochondriai DNA.Previous studies have shown that overexpression of mTERF2 can inhibit cell proliferation, but the mechanism has not been well defined so far.This study aimed to present the binding pattern of mTERF2 to the mitochondrial DNA (mtDNA) in vivo, and investigated the biological function of mTERF2 on the replication of mtDNA, mRNA transcription, and protein translation.The mTERF2 binding to entire mtDNA was identified via the chromatin immunoprecipitation analysis.The mtDNA replication efficiency and expression levels of mitochondria genes were significantly inhibited when the mTERF2 was overexpressed in HeLa cells.The inhibition level of mtDNA content was the same with the decreased levels of mRNA and mitochondrial protein expression.Overall, the mTERF2 might be a cell growth inhibitor based on its negative effect on mtDNA replication, which eventually own-regulated all of the oxidative phosphorylation components in the mitochondria that were essential for the cell's energy metabolism.

  13. Mitochondrial DNA analysis in Parkinson's disease.

    Science.gov (United States)

    Schapira, A H; Holt, I J; Sweeney, M; Harding, A E; Jenner, P; Marsden, C D

    1990-01-01

    The reduced form of nicotinamide adenine dinucleotide coenzyme Q reductase (complex I) activity has recently been shown to be deficient in the substantia nigra of patients dying with Parkinson's disease. This biochemical defect is identical to that produced by the neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), which also produces parkinsonism in humans. Complex I comprises 25 polypeptides, seven of which are encoded by mitochondrial DNA. Restriction fragment analysis of substantia nigra DNA from six patients with Parkinson's disease did not show any major deletion. In two cases, there were different novel polymorphisms that were not observed in control brain (n = 6) or blood (n = 34) samples.

  14. Mitochondrial DNA sequence variation in Greeks.

    Science.gov (United States)

    Kouvatsi, A; Karaiskou, N; Apostolidis, A; Kirmizidis, G

    2001-12-01

    Mitochondrial DNA (mtDNA) control region sequences were determined in 54 unrelated Greeks, coming from different regions in Greece, for both segments HVR-I and HVR-II. Fifty-two different mtDNA haplotypes were revealed, one of which was shared by three individuals. A very low heterogeneity was found among Greek regions. No one cluster of lineages was specific to individuals coming from a certain region. The average pairwise difference distribution showed a value of 7.599. The data were compared with that for other European or neighbor populations (British, French, Germans, Tuscans, Bulgarians, and Turks). The genetic trees that were constructed revealed homogeneity between Europeans. Median networks revealed that most of the Greek mtDNA haplotypes are clustered to the five known haplogroups and that a number of haplotypes are shared among Greeks and other European and Near Eastern populations.

  15. Number matters: control of mammalian mitochondrial DNA copy number.

    Science.gov (United States)

    Clay Montier, Laura L; Deng, Janice J; Bai, Yidong

    2009-03-01

    Regulation of mitochondrial biogenesis is essential for proper cellular functioning. Mitochondrial DNA (mtDNA) depletion and the resulting mitochondrial malfunction have been implicated in cancer, neurodegeneration, diabetes, aging, and many other human diseases. Although it is known that the dynamics of the mammalian mitochondrial genome are not linked with that of the nuclear genome, very little is known about the mechanism of mtDNA propagation. Nevertheless, our understanding of the mode of mtDNA replication has advanced in recent years, though not without some controversies. This review summarizes our current knowledge of mtDNA copy number control in mammalian cells, while focusing on both mtDNA replication and turnover. Although mtDNA copy number is seemingly in excess, we reason that mtDNA copy number control is an important aspect of mitochondrial genetics and biogenesis and is essential for normal cellular function.

  16. Mitochondrial DNA: Radically free of free-radical driven mutations.

    Science.gov (United States)

    Kauppila, Johanna H K; Stewart, James B

    2015-11-01

    Mitochondrial DNA has long been posited as a likely target of oxidative damage induced mutation during the ageing process. Research over the past decades has uncovered the accumulation of mitochondrial DNA mutations in association with a mosaic pattern of cells displaying mitochondrial dysfunction in ageing individuals. Unfortunately, the underlying mechanisms are far less straightforward than originally anticipated. Recent research on mitochondria reveals that these genomes are far less helpless than originally envisioned. Additionally, new technologies have allowed us to analyze the mutational signatures of many more somatic mitochondrial DNA mutations, revealing surprising patterns that are inconsistent with a DNA-oxidative damage based hypothesis. In this review, we will discuss these recent observations and new insights into the eccentricities of mitochondrial genetics, and their impact on our understanding of mitochondrial mutations and their role in the ageing process. This article is part of a Special Issue entitled: Mitochondrial Dysfunction in Aging.

  17. Sephardic signature in haplogroup T mitochondrial DNA

    OpenAIRE

    Bedford, Felice L

    2011-01-01

    A rare combination of mutations within mitochondrial DNA subhaplogroup T2e is identified as affiliated with Sephardic Jews, a group that has received relatively little attention. Four investigations were pursued: Search of the motif in 250 000 control region records across 8 databases, comparison of frequencies of T subhaplogroups (T1, T2b, T2c, T2e, T4, T*) across 11 diverse populations, creation of a phylogenic median-joining network from public T2e control region entries, and analysis of o...

  18. Disruption of mitochondrial DNA replication in Drosophila increases mitochondrial fast axonal transport in vivo.

    Directory of Open Access Journals (Sweden)

    Rehan M Baqri

    Full Text Available Mutations in mitochondrial DNA polymerase (pol gamma cause several progressive human diseases including Parkinson's disease, Alper's syndrome, and progressive external ophthalmoplegia. At the cellular level, disruption of pol gamma leads to depletion of mtDNA, disrupts the mitochondrial respiratory chain, and increases susceptibility to oxidative stress. Although recent studies have intensified focus on the role of mtDNA in neuronal diseases, the changes that take place in mitochondrial biogenesis and mitochondrial axonal transport when mtDNA replication is disrupted are unknown. Using high-speed confocal microscopy, electron microscopy and biochemical approaches, we report that mutations in pol gamma deplete mtDNA levels and lead to an increase in mitochondrial density in Drosophila proximal nerves and muscles, without a noticeable increase in mitochondrial fragmentation. Furthermore, there is a rise in flux of bidirectional mitochondrial axonal transport, albeit with slower kinesin-based anterograde transport. In contrast, flux of synaptic vesicle precursors was modestly decreased in pol gamma-alpha mutants. Our data indicate that disruption of mtDNA replication does not hinder mitochondrial biogenesis, increases mitochondrial axonal transport, and raises the question of whether high levels of circulating mtDNA-deficient mitochondria are beneficial or deleterious in mtDNA diseases.

  19. Acceptance of Domestic Cat Mitochondrial DNA in a Criminal Proceeding

    Science.gov (United States)

    Lyons, Leslie A.; Grahn, Robert A.; Kun, Teri J.; Netzel, Linda R.; Wictum, Elizabeth E.; Halverson, Joy L.

    2014-01-01

    Shed hair from domestic animals readily adheres to clothing and other contact items, providing a source of transfer evidence for criminal investigations. Mitochondrial DNA is often the only option for DNA analysis of shed hair. Human mitochondrial DNA analysis has been accepted in the US court system since 1996. The murder trial of the State of Missouri versus Henry L. Polk, Jr. represents the first legal proceeding where cat mitochondrial DNA analysis was introduced into evidence. The mitochondrial DNA evidence was initially considered inadmissible due to concerns about the cat dataset and the scientific acceptance of the marker. Those concerns were subsequently addressed, and the evidence was deemed admissible. This report reviews the case in regards to the cat biological evidence and its ultimate admission as generally accepted and reliable. Expansion and saturation analysis of the cat mitochondrial DNA control region dataset supported the initial interpretation of the evidence. PMID:25086413

  20. Acceptance of domestic cat mitochondrial DNA in a criminal proceeding.

    Science.gov (United States)

    Lyons, Leslie A; Grahn, Robert A; Kun, Teri J; Netzel, Linda R; Wictum, Elizabeth E; Halverson, Joy L

    2014-11-01

    Shed hair from domestic animals readily adheres to clothing and other contact items, providing a source of transfer evidence for criminal investigations. Mitochondrial DNA is often the only option for DNA analysis of shed hair. Human mitochondrial DNA analysis has been accepted in the US court system since 1996. The murder trial of the State of Missouri versus Henry L. Polk, Jr. represents the first legal proceeding where cat mitochondrial DNA analysis was introduced into evidence. The mitochondrial DNA evidence was initially considered inadmissible due to concerns about the cat dataset and the scientific acceptance of the marker. Those concerns were subsequently addressed, and the evidence was deemed admissible. This report reviews the case in regards to the cat biological evidence and its ultimate admission as generally accepted and reliable. Expansion and saturation analysis of the cat mitochondrial DNA control region dataset supported the initial interpretation of the evidence.

  1. Taenia hydatigena: isolation of mitochondrial DNA, molecular cloning, and physical mitochondrial genome mapping.

    Science.gov (United States)

    Yap, K W; Thompson, R C; Rood, J I; Pawlowski, I D

    1987-06-01

    Mitochondrial DNA was isolated from Taenia hydatigena, T. crassiceps, and Echinococcus granulosus using a cetyltrimethylammonium bromide precipitation technique. The technique is simple, rapid, reproducible, and does not require extensive high speed ultracentrifugation. The advantage of using mitochondrial DNA from taeniid cestodes for comparative restriction analysis was demonstrated. Mitochondrial DNA of T. hydatigena was isolated as covalently closed circular molecules. These were linearized by single digestion with BamHI and the molecular weight was estimated from the linear form of 17.6 kb. The mitochondrial DNA of T. hydatigena is therefore similar in size and structure to that of many other animal species. The entire mitochondrial genome was cloned into pBR322 in Escherichia coli and a restriction map of the recombinant molecule was constructed. The potential of using the cloned mitochondrial genome as a probe in speciation studies as well as for providing functional information on the role of the cestode mitochondrion is discussed.

  2. Uniparental Inheritance and Replacement of Mitochondrial DNA in Neurospora Tetrasperma

    OpenAIRE

    Lee, S B; Taylor, J. W.

    1993-01-01

    This study tested mechanisms proposed for maternal uniparental mitochondrial inheritance in Neurospora: (1) exclusion of conidial mitochondria by the specialized female reproductive structure, trichogyne, due to mating locus heterokaryon incompatibility and (2) mitochondrial input bias favoring the larger trichogyne over the smaller conidium. These mechanisms were tested by determining the modes of mitochondrial DNA (mtDNA) inheritance and transmission in the absence of mating locus heterokar...

  3. Mitochondrial DNA Mutations Associated with Aminoglycoside Ototoxicity

    Institute of Scientific and Technical Information of China (English)

    GUAN Min-Xin

    2006-01-01

    The mitochondrial 12S rRNA has been shown to be the hot spot for mutations associated with both aminoglycoside-induced and non-syndromic hearing loss. Of all the mutations, the homoplasmic A1555G and C1494T mutations at a highly conserved decoding region in the 12S rRNA have been associated with aminoglycoside-induced and non-syndromic hearing loss in many families worldwide. The A1555G or C1494T mutation is expected to form novel 1494C-G1555 or 1494U-A1555 base-pair at the highly conserved A-site of 12S rRNA. These transitions make the secondary structure of this RNA more closely resemble the corresponding region of bacterial 16S rRNA. Thus, the new U - A or G-C pair in 12S rRNA created by the C1494T or A1555G transition facilitates the binding of aminoglycosides, thereby accounting for the fact that the exposure to aminoglycosides can induce or worsen hearing loss in individuals carrying these mutations. Furthermore, the growth defect and impairment of mitochondrial translation were observed in cell lines carrying the A1555G or C1494T mutation in the presence of high concentration of aminoglycosides. In addition, nuclear modifier genes and mitochondrial haplotypes modulate the phenotypic manifestation of the A1555G and C1494T mutations. These observations provide the direct genetic and biochemical evidences that the A1555G or C1494T mutation is a pathogenic mtDNA mutation associated with aminoglycoside-induced and nonsyndromic hearing loss. Therefore, these data have been providing valuable information and technology to predict which individuals are at risk for ototoxicity, to improve the safety of aminoglycoside antibiotic therapy, and eventually to decrease the incidence of deafness.

  4. Nuclear responses to depletion of mitochondrial DNA in human cells.

    Science.gov (United States)

    Li, K; Neufer, P D; Williams, R S

    1995-11-01

    The derivation of human cell lines devoid of mitochondrial (mt) DNA (rho 0) provides an opportunity to study nuclear responses to a chronic impairment of mitochondrial oxidative phosphorylation. Expression of several nuclear genes is induced in human rho 0 cells, including those encoding integral proteins of the mitochondrial inner membrane, intermediate filaments, and ribosomes. In contrast to conditions in which mitochondrial respiration is altered acutely, expression of heat shock proteins and immediate early genes is not induced. Mitochondria from rho 0 cells maintain a transmembrane electrochemical potential and are distributed within the cytoplasm of these cells in a manner indistinguishable from that of wild-type cells. We conclude that a chronic deficiency of mitochondrial oxidative phosphorylation produced by elimination of mtDNA is associated with a different pattern of gene induction than that provoked by other acute or subacute conditions that impair mitochondrial respiration or create energy demands in excess of mitochondrial respiratory capacity.

  5. Staphylococcus aureus sepsis induces early renal mitochondrial DNA repair and mitochondrial biogenesis in mice.

    Directory of Open Access Journals (Sweden)

    Raquel R Bartz

    Full Text Available Acute kidney injury (AKI contributes to the high morbidity and mortality of multi-system organ failure in sepsis. However, recovery of renal function after sepsis-induced AKI suggests active repair of energy-producing pathways. Here, we tested the hypothesis in mice that Staphyloccocus aureus sepsis damages mitochondrial DNA (mtDNA in the kidney and activates mtDNA repair and mitochondrial biogenesis. Sepsis was induced in wild-type C57Bl/6J and Cox-8 Gfp-tagged mitochondrial-reporter mice via intraperitoneal fibrin clots embedded with S. aureus. Kidneys from surviving mice were harvested at time zero (control, 24, or 48 hours after infection and evaluated for renal inflammation, oxidative stress markers, mtDNA content, and mitochondrial biogenesis markers, and OGG1 and UDG mitochondrial DNA repair enzymes. We examined the kidneys of the mitochondrial reporter mice for changes in staining density and distribution. S. aureus sepsis induced sharp amplification of renal Tnf, Il-10, and Ngal mRNAs with decreased renal mtDNA content and increased tubular and glomerular cell death and accumulation of protein carbonyls and 8-OHdG. Subsequently, mtDNA repair and mitochondrial biogenesis was evidenced by elevated OGG1 levels and significant increases in NRF-1, NRF-2, and mtTFA expression. Overall, renal mitochondrial mass, tracked by citrate synthase mRNA and protein, increased in parallel with changes in mitochondrial GFP-fluorescence especially in proximal tubules in the renal cortex and medulla. Sub-lethal S. aureus sepsis thus induces widespread renal mitochondrial damage that triggers the induction of the renal mtDNA repair protein, OGG1, and mitochondrial biogenesis as a conspicuous resolution mechanism after systemic bacterial infection.

  6. Maternal inheritance of mitochondrial DNA by diverse mechanisms to eliminate paternal mitochondrial DNA.

    Science.gov (United States)

    Sato, Miyuki; Sato, Ken

    2013-08-01

    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.

  7. Mitochondrial DNA recombination in a free-ranging Australian lizard.

    Science.gov (United States)

    Ujvari, Beata; Dowton, Mark; Madsen, Thomas

    2007-04-22

    Mitochondrial DNA (mtDNA) is the traditional workhorse for reconstructing evolutionary events. The frequent use of mtDNA in such analyses derives from the apparent simplicity of its inheritance: maternal and lacking bi-parental recombination. However, in hybrid zones, the reproductive barriers are often not completely developed, resulting in the breakdown of male mitochondrial elimination mechanisms, leading to leakage of paternal mitochondria and transient heteroplasmy, resulting in an increased possibility of recombination. Despite the widespread occurrence of heteroplasmy and the presence of the molecular machinery necessary for recombination, we know of no documented example of recombination of mtDNA in any terrestrial wild vertebrate population. By sequencing the entire mitochondrial genome (16761bp), we present evidence for mitochondrial recombination in the hybrid zone of two mitochondrial haplotypes in the Australian frillneck lizard (Chlamydosaurus kingii).

  8. A Spatio-Temporal Analysis of Mitochondrial DNA Haplogroup I

    Directory of Open Access Journals (Sweden)

    Revesz Peter Z.

    2016-01-01

    Full Text Available The recent recovery of ancient DNA from a growing number of human samples shows that mitochondrial DNA haplogroup I was introduced to Europe after the end of the Last Glacial Maximum. This paper provides a spatio-temporal analysis of the various subhaplogroups of mitochondrial DNA I. The study suggests that haplogroup I diversified into haplogroups I1, I2’3, I4 and I5 at specific regions in Eurasia and then spread southward to Crete and Egypt.

  9. Mitochondrial DNA recombination in a free-ranging Australian lizard

    OpenAIRE

    Ujvari, Beata; Dowton, Mark; Madsen, Thomas

    2007-01-01

    Mitochondrial DNA (mtDNA) is the traditional workhorse for reconstructing evolutionary events. The frequent use of mtDNA in such analyses derives from the apparent simplicity of its inheritance: maternal and lacking bi-parental recombination. However, in hybrid zones, the reproductive barriers are often not completely developed, resulting in the breakdown of male mitochondrial elimination mechanisms, leading to leakage of paternal mitochondria and transient heteroplasmy, resulting in an incre...

  10. Proteomic Dissection of the Mitochondrial DNA Metabolism Apparatus in Arabidopsis

    Energy Technology Data Exchange (ETDEWEB)

    SAlly A. Mackenzie

    2004-01-06

    This study involves the investigation of nuclear genetic components that regulate mitochondrial genome behavior in higher plants. The approach utilizes the advanced plant model system of Arabidopsis thaliana to identify and functionally characterize multiple components of the mitochondrial DNA replication, recombination and mismatch repair system and their interaction partners. The rationale for the research stems from the central importance of mitochondria to overall cellular metabolism and the essential nature of the mitochondrial genome to mitochondrial function. Relatively little is understood about mitochondrial DNA maintenance and transmission in higher eukaryotes, and the higher plant mitochondrial genome displays unique properties and behavior. This investigation has revealed at least three important properties of plant mitochondrial DNA metabolism components. (1) Many are dual targeted to mitochondrial and chloroplasts by novel mechanisms, suggesting that the mitochondria a nd chloroplast share their genome maintenance apparatus. (2)The MSH1 gene, originating as a component of mismatch repair, has evolved uniquely in plants to participate in differential replication of the mitochondrial genome. (3) This mitochondrial differential replication process, termed substoichiometric shifting and also involving a RecA-related gene, appears to represent an adaptive mechanism to expand plant reproductive capacity and is likely present throughout the plant kingdom.

  11. Altofrequency SNPs of mitochondrial DNA in 26 Han Chinese

    Institute of Scientific and Technical Information of China (English)

    LUO Yong-jun; GAO Wen-xiang; GAO Yu-qi; CHEN Jian; TAN Xiao-ling; LIU Xin; CHEN Hai-hua

    2007-01-01

    Objective:To explore the possible mitochondrial DNA (mtDNA) polymorphism in Han Chinese.Methods:The complete mitochondrial genome of 26 unrelated healthy Han Chinese were extracted and sequenced.Results:The mtDNA nucleotide sites (2 706,7 028,8 860,11 719,and 15 326)were found totally different from the Revised Cambridge Reference Sequence (rCRS).These single nucleotide polymorphisms (SNPs) were 2 706 A→G,7 028 C→T,8 860 A→G,11 719 G→A,15 326 A→G.Conclusion:These findings provide new insights into the characteristics of Han Chinese mitochondrial genetic diversity.

  12. Mitochondrial DNA structure in the Arabian Peninsula.

    Science.gov (United States)

    Abu-Amero, Khaled K; Larruga, José M; Cabrera, Vicente M; González, Ana M

    2008-02-12

    Two potential migratory routes followed by modern humans to colonize Eurasia from Africa have been proposed. These are the two natural passageways that connect both continents: the northern route through the Sinai Peninsula and the southern route across the Bab al Mandab strait. Recent archaeological and genetic evidence have favored a unique southern coastal route. Under this scenario, the study of the population genetic structure of the Arabian Peninsula, the first step out of Africa, to search for primary genetic links between Africa and Eurasia, is crucial. The haploid and maternally inherited mitochondrial DNA (mtDNA) molecule has been the most used genetic marker to identify and to relate lineages with clear geographic origins, as the African Ls and the Eurasian M and N that have a common root with the Africans L3. To assess the role of the Arabian Peninsula in the southern route, we genetically analyzed 553 Saudi Arabs using partial (546) and complete mtDNA (7) sequencing, and compared the lineages obtained with those present in Africa, the Near East, central, east and southeast Asia and Australasia. The results showed that the Arabian Peninsula has received substantial gene flow from Africa (20%), detected by the presence of L, M1 and U6 lineages; that an 18% of the Arabian Peninsula lineages have a clear eastern provenance, mainly represented by U lineages; but also by Indian M lineages and rare M links with Central Asia, Indonesia and even Australia. However, the bulk (62%) of the Arabian lineages has a Northern source. Although there is evidence of Neolithic and more recent expansions in the Arabian Peninsula, mainly detected by (preHV)1 and J1b lineages, the lack of primitive autochthonous M and N sequences, suggests that this area has been more a receptor of human migrations, including historic ones, from Africa, India, Indonesia and even Australia, than a demographic expansion center along the proposed southern coastal route.

  13. Mitochondrial DNA structure in the Arabian Peninsula

    Directory of Open Access Journals (Sweden)

    Cabrera Vicente M

    2008-02-01

    Full Text Available Abstract Background Two potential migratory routes followed by modern humans to colonize Eurasia from Africa have been proposed. These are the two natural passageways that connect both continents: the northern route through the Sinai Peninsula and the southern route across the Bab al Mandab strait. Recent archaeological and genetic evidence have favored a unique southern coastal route. Under this scenario, the study of the population genetic structure of the Arabian Peninsula, the first step out of Africa, to search for primary genetic links between Africa and Eurasia, is crucial. The haploid and maternally inherited mitochondrial DNA (mtDNA molecule has been the most used genetic marker to identify and to relate lineages with clear geographic origins, as the African Ls and the Eurasian M and N that have a common root with the Africans L3. Results To assess the role of the Arabian Peninsula in the southern route, we genetically analyzed 553 Saudi Arabs using partial (546 and complete mtDNA (7 sequencing, and compared the lineages obtained with those present in Africa, the Near East, central, east and southeast Asia and Australasia. The results showed that the Arabian Peninsula has received substantial gene flow from Africa (20%, detected by the presence of L, M1 and U6 lineages; that an 18% of the Arabian Peninsula lineages have a clear eastern provenance, mainly represented by U lineages; but also by Indian M lineages and rare M links with Central Asia, Indonesia and even Australia. However, the bulk (62% of the Arabian lineages has a Northern source. Conclusion Although there is evidence of Neolithic and more recent expansions in the Arabian Peninsula, mainly detected by (preHV1 and J1b lineages, the lack of primitive autochthonous M and N sequences, suggests that this area has been more a receptor of human migrations, including historic ones, from Africa, India, Indonesia and even Australia, than a demographic expansion center along the

  14. Sephardic signature in haplogroup T mitochondrial DNA.

    Science.gov (United States)

    Bedford, Felice L

    2012-04-01

    A rare combination of mutations within mitochondrial DNA subhaplogroup T2e is identified as affiliated with Sephardic Jews, a group that has received relatively little attention. Four investigations were pursued: Search of the motif in 250 000 control region records across 8 databases, comparison of frequencies of T subhaplogroups (T1, T2b, T2c, T2e, T4, T(*)) across 11 diverse populations, creation of a phylogenic median-joining network from public T2e control region entries, and analysis of one Sephardic mitochondrial full genomic sequence with the motif. It was found that the rare motif belonged only to Sephardic descendents (Turkey, Bulgaria), to inhabitants of North American regions known for secret Spanish-Jewish colonization, or were consistent with Sephardic ancestry. The incidence of subhaplogroup T2e decreased from the Western Arabian Peninsula to Italy to Spain and into Western Europe. The ratio of sister subhaplogroups T2e to T2b was found to vary 40-fold across populations from a low in the British Isles to a high in Saudi Arabia with the ratio in Sephardim more similar to Saudi Arabia, Egypt, and Italy than to hosts Spain and Portugal. Coding region mutations of 2308G and 14499T may locate the Sephardic signature within T2e, but additional samples and reworking of current T2e phylogenetic branch structure is needed. The Sephardic Turkish community has a less pronounced founder effect than some Ashkenazi groups considered singly (eg, Polish), but other comparisons of interest await comparable averaging. Registries of signatures will benefit the study of populations with a large number of smaller-size founders.

  15. Phenotype of Mitochondrial DNA 3243A > G Mutation

    Directory of Open Access Journals (Sweden)

    J Gordon Millichap

    2007-10-01

    Full Text Available The prevalence and common clinical manifestations of the mitochondrial DNA 3243A > G mutation in children in a defined population in Finland were studied at the Universities of Oulu and Turku and other centers.

  16. [Somatic mutations in nuclear and mitochondrial DNA]. Progress report

    Energy Technology Data Exchange (ETDEWEB)

    1992-09-01

    The study is concerned the design of new assays that may detect rare somatic mutations in nuclear and mitochondrial DNA, which may increase upon exposure to mutagens, and thus become a marker of human exposure to such mutagens. Two assays for somatic mutation were presented, one for mitochondrial DNA deletions which was developed by the author, and one for deletions of the ADA gene which resides in the nucleus.

  17. Mitochondrial DNA Fails to Pinpoint African Ancestry

    Institute of Scientific and Technical Information of China (English)

    葛群燕

    2006-01-01

    最新的线粒体基因(Mitochondrial DNA)研究结果恐怕要让一些黑人失望了:他们想要确定自己究竟属于哪一个具体的非洲部族,然而在抽样研究中,只有不到10%的样本可以确定与之完全吻合的部落族群,有一半的基因样本与多种部族相吻合,另有40%的样本基因则根本找不到与其吻合的非洲部族。这项新的基因研究技术最终能否给黑人们带来满意的答案呢?有限的数据限制了目前的研究,我们只能拭目以待。

  18. Mitochondrial DNA mutations provoke dominant inhibition of mitochondrial inner membrane fusion.

    Directory of Open Access Journals (Sweden)

    Cécile Sauvanet

    Full Text Available Mitochondria are highly dynamic organelles that continuously move, fuse and divide. Mitochondrial dynamics modulate overall mitochondrial morphology and are essential for the proper function, maintenance and transmission of mitochondria and mitochondrial DNA (mtDNA. We have investigated mitochondrial fusion in yeast cells with severe defects in oxidative phosphorylation (OXPHOS due to removal or various specific mutations of mtDNA. We find that, under fermentative conditions, OXPHOS deficient cells maintain normal levels of cellular ATP and ADP but display a reduced mitochondrial inner membrane potential. We demonstrate that, despite metabolic compensation by glycolysis, OXPHOS defects are associated to a selective inhibition of inner but not outer membrane fusion. Fusion inhibition was dominant and hampered the fusion of mutant mitochondria with wild-type mitochondria. Inhibition of inner membrane fusion was not systematically associated to changes of mitochondrial distribution and morphology, nor to changes in the isoform pattern of Mgm1, the major fusion factor of the inner membrane. However, inhibition of inner membrane fusion correlated with specific alterations of mitochondrial ultrastructure, notably with the presence of aligned and unfused inner membranes that are connected to two mitochondrial boundaries. The fusion inhibition observed upon deletion of OXPHOS related genes or upon removal of the entire mtDNA was similar to that observed upon introduction of point mutations in the mitochondrial ATP6 gene that are associated to neurogenic ataxia and retinitis pigmentosa (NARP or to maternally inherited Leigh Syndrome (MILS in humans. Our findings indicate that the consequences of mtDNA mutations may not be limited to OXPHOS defects but may also include alterations in mitochondrial fusion. Our results further imply that, in healthy cells, the dominant inhibition of fusion could mediate the exclusion of OXPHOS-deficient mitochondria from

  19. RECQL4 localizes to mitochondria and preserves mitochondrial DNA integrity

    DEFF Research Database (Denmark)

    Croteau, Deborah L; Rossi, Marie L; Canugovi, Chandrika

    2012-01-01

    RECQL4 is associated with Rothmund-Thomson Syndrome (RTS), a rare autosomal recessive disorder characterized by premature aging, genomic instability, and cancer predisposition. RECQL4 is a member of the RecQ helicase family, and has many similarities to WRN protein, which is also implicated...... in premature aging. There is no information about whether any of the RecQ helicases play roles in mitochondrial biogenesis, which is strongly implicated in the aging process. Here, we used microscopy to visualize RECQL4 in mitochondria. Fractionation of human and mouse cells also showed that RECQL4 was present...... in mitochondria. Q-PCR amplification of mitochondrial DNA demonstrated that mtDNA damage accumulated in RECQL4-deficient cells. Microarray analysis suggested that mitochondrial bioenergetic pathways might be affected in RTS. Measurements of mitochondrial bioenergetics showed a reduction in the mitochondrial...

  20. Mitochondrial DNA Copy Number in Sleep Duration Discordant Monozygotic Twins

    DEFF Research Database (Denmark)

    Wrede, Joanna E; Mengel-From, Jonas; Buchwald, Dedra

    2015-01-01

    STUDY OBJECTIVES: Mitochondrial DNA (mtDNA) copy number is an important component of mitochondrial function and varies with age, disease, and environmental factors. We aimed to determine whether mtDNA copy number varies with habitual differences in sleep duration within pairs of monozygotic twins....... SETTING: Academic clinical research center. PARTICIPANTS: 15 sleep duration discordant monozygotic twin pairs (30 twins, 80% female; mean age 42.1 years [SD 15.0]). DESIGN: Sleep duration was phenotyped with wrist actigraphy. Each twin pair included a "normal" (7-9 h/24) and "short" (sleeping...... twin. Fasting peripheral blood leukocyte DNA was assessed for mtDNA copy number via the n-fold difference between qPCR measured mtDNA and nuclear DNA creating an mtDNA measure without absolute units. We used generalized estimating equation linear regression models accounting for the correlated data...

  1. Private mitochondrial DNA variants in danish patients with hypertrophic cardiomyopathy

    DEFF Research Database (Denmark)

    Hagen, Christian M; Aidt, Frederik H; Havndrup, Ole;

    2015-01-01

    Hypertrophic cardiomyopathy (HCM) is a genetic cardiac disease primarily caused by mutations in genes coding for sarcomeric proteins. A molecular-genetic etiology can be established in ~60% of cases. Evolutionarily conserved mitochondrial DNA (mtDNA) haplogroups are susceptibility factors for HCM....... Several polymorphic mtDNA variants are associated with a variety of late-onset degenerative diseases and affect mitochondrial function. We examined the role of private, non-haplogroup associated, mitochondrial variants in the etiology of HCM. In 87 Danish HCM patients, full mtDNA sequencing revealed 446......>G, and MT-CYB: m.15024G>A, p.C93Y remained. A detailed analysis of these variants indicated that none of them are likely to cause HCM. In conclusion, private mtDNA mutations are frequent, but they are rarely, if ever, associated with HCM....

  2. Loss of mitochondrial DNA with aging in Drosophila melanogaster.

    Science.gov (United States)

    Massie, H R; Baird, M B; McMahon, M M

    1975-01-01

    The buoyant densities of nuclear and mitochondrial DNA from Drosophila melanogaster lysates has been found to show no change with increasing age in both CsCl and Cs2SO4 equilibrium density gradients. Whole fly homogenates were used to demonstrate no change in nuclear DNA content during adult life. Mitochondrial DNA increased from 1.2 to 4.3% of the total DNA during the first week of adult life and then decreased during senescence to a minimum of 1.5% at 10 weeks of age which represented a 65% loss in mitochondrial DNA content with age. These data are interpreted to support the proposal that mitochondria destruction occurs during senescence.

  3. Mitochondrial DNA deletions in patients with chronic suppurative otitis media.

    Science.gov (United States)

    Tatar, Arzu; Tasdemir, Sener; Sahin, Ibrahim; Bozoglu, Ceyda; Erdem, Haktan Bagis; Yoruk, Ozgur; Tatar, Abdulgani

    2016-09-01

    The aim of this study was to investigate the 4977 and 7400 bp deletions of mitochondrial DNA in patients with chronic suppurative otitis media and to indicate the possible association of mitochondrial DNA deletions with chronic suppurative otitis media. Thirty-six patients with chronic suppurative otitis media were randomly selected to assess the mitochondrial DNA deletions. Tympanomastoidectomy was applied for the treatment of chronic suppurative otitis media, and the curettage materials including middle ear tissues were collected. The 4977 and 7400 bp deletion regions and two control regions of mitochondrial DNA were assessed by using the four pair primers. DNA was extracted from middle ear tissues and peripheral blood samples of the patients, and then polymerase chain reactions (PCRs) were performed. PCR products were separated in 2 % agarose gel. Seventeen of 36 patients had the heterozygote 4977 bp deletion in the middle ear tissue but not in peripheral blood. There wasn't any patient who had the 7400 bp deletion in mtDNA of their middle ear tissue or peripheral blood tissue. The patients with the 4977 bp deletion had a longer duration of chronic suppurative otitis media and a higher level of hearing loss than the others (p media and the reactive oxygen species can cause the mitochondrial DNA deletions and this may be a predisposing factor to sensorineural hearing loss in chronic suppurative otitis media. An antioxidant drug as a scavenger agent may be used in long-term chronic suppurative otitis media.

  4. Oxidized mitochondrial DNA activates the NLRP3 inflammasome during apoptosis.

    Science.gov (United States)

    Shimada, Kenichi; Crother, Timothy R; Karlin, Justin; Dagvadorj, Jargalsaikhan; Chiba, Norika; Chen, Shuang; Ramanujan, V Krishnan; Wolf, Andrea J; Vergnes, Laurent; Ojcius, David M; Rentsendorj, Altan; Vargas, Mario; Guerrero, Candace; Wang, Yinsheng; Fitzgerald, Katherine A; Underhill, David M; Town, Terrence; Arditi, Moshe

    2012-03-23

    We report that in the presence of signal 1 (NF-κB), the NLRP3 inflammasome was activated by mitochondrial apoptotic signaling that licensed production of interleukin-1β (IL-1β). NLRP3 secondary signal activators such as ATP induced mitochondrial dysfunction and apoptosis, resulting in release of oxidized mitochondrial DNA (mtDNA) into the cytosol, where it bound to and activated the NLRP3 inflammasome. The antiapoptotic protein Bcl-2 inversely regulated mitochondrial dysfunction and NLRP3 inflammasome activation. Mitochondrial DNA directly induced NLRP3 inflammasome activation, because macrophages lacking mtDNA had severely attenuated IL-1β production, yet still underwent apoptosis. Both binding of oxidized mtDNA to the NLRP3 inflammasome and IL-1β secretion could be competitively inhibited by the oxidized nucleoside 8-OH-dG. Thus, our data reveal that oxidized mtDNA released during programmed cell death causes activation of the NLRP3 inflammasome. These results provide a missing link between apoptosis and inflammasome activation, via binding of cytosolic oxidized mtDNA to the NLRP3 inflammasome.

  5. Mutations in circulating mitochondrial DNA: Cassandra of oral cancer?

    Science.gov (United States)

    Kandel, Eugene S

    2012-07-01

    Cell-free circulating nucleic acids in human blood are increasing being researched as a source of diagnostic and prognostic biomarkers for clinical oncology. High copy number per cell and frequent mutations in various malignancies make mitochondrial genome an attractive target for such an investigation, but practical development and validation of biomarkers based on cell-free mitochondrial DNA has been lagging. Uzawa and colleagues report in the July issue of Oncotarget that in a retrospective study of patients with oral cancer the load of mutant mitochondrial DNA in patient's serum was a strong indicator of postoperative recurrence. Based on these observations, the predictive value of circulating mutant mitochondrial DNA merits further evaluation in patients with oral and other malignancies.

  6. DNA Sequences Proximal to Human Mitochondrial DNA Deletion Breakpoints Prevalent in Human Disease Form G-quadruplexes, a Class of DNA Structures Inefficiently Unwound by the Mitochondrial Replicative Twinkle Helicase

    NARCIS (Netherlands)

    Bharti, S.K.; Sommers, J.A.; Zhou, J.; Kaplan, D.L.; Spelbrink, J.N.; Mergny, J.L.; Brosh, R.M., Jr.

    2014-01-01

    Mitochondrial DNA deletions are prominent in human genetic disorders, cancer, and aging. It is thought that stalling of the mitochondrial replication machinery during DNA synthesis is a prominent source of mitochondrial genome instability; however, the precise molecular determinants of defective

  7. Melatonin protects against common deletion of mitochondrial DNA-augmented mitochondrial oxidative stress and apoptosis.

    Science.gov (United States)

    Jou, Mei-Jie; Peng, Tsung-I; Yu, Pai-Zu; Jou, Shuo-Bin; Reiter, Russel J; Chen, Jin-Yi; Wu, Hong-Yueh; Chen, Chih-Chun; Hsu, Lee-Fen

    2007-11-01

    Defected mitochondrial respiratory chain (RC), in addition to causing a severe ATP deficiency, often augments reactive oxygen species (ROS) generation in mitochondria (mROS) which enhances pathological conditions and diseases. Previously, we demonstrated a potent endogenously RC defect-augmented mROS associated dose-dependently with a commonly seen large-scale deletion of 4977 base pairs of mitochondrial DNA (mtDNA), i.e. the common deletion (CD). As current treatments for CD-associated diseases are rather supplementary and ineffective, we investigated whether melatonin, a potential mitochondrial protector, provides beneficial protection for CD-augmented mitochondrial oxidative stress and apoptosis particularly upon the induction of a secondary oxidative stress. Detailed mechanistic investigations were performed by using laser scanning dual fluorescence imaging microscopy to provide precise spatial and temporal resolution of mitochondrial events at single cell level. We demonstrate, for the first time, that melatonin significantly prevents CD-augmented mROS formation under basal conditions as well as at early time-points upon secondary oxidative stress induced by H2O2 exposure. Thus, melatonin prevents mROS-mediated depolarization of mitochondrial membrane potential (DeltaPsim) and subsequent opening of the mitochondrial permeability transition pore (MPTP) and cytochrome c release. Moreover, melatonin prevents depletion of cardiolipin which appears to be crucial for postponing later MPTP opening, disruption of the mitochondrial membrane and apoptosis. Finally, the protection provided by melatonin is superior to those caused by the suppression of mitochondrial Ca2+ regulators including the mitochondrial Na+-Ca2) exchanger, the MPTP, and the mitochondrial Ca2+ uniporter and by antioxidants including vitamin E and mitochondria-targeted coenzyme Q, MitoQ. As RC defect-augmented endogenous mitochondrial oxidative stress is centrally involved in a variety of pathological

  8. Strong Purifying Selection in Transmission of Mammalian Mitochondrial DNA

    Science.gov (United States)

    Stewart, James Bruce; Freyer, Christoph; Elson, Joanna L; Wredenberg, Anna; Cansu, Zekiye; Trifunovic, Aleksandra; Larsson, Nils-Göran

    2008-01-01

    There is an intense debate concerning whether selection or demographics has been most important in shaping the sequence variation observed in modern human mitochondrial DNA (mtDNA). Purifying selection is thought to be important in shaping mtDNA sequence evolution, but the strength of this selection has been debated, mainly due to the threshold effect of pathogenic mtDNA mutations and an observed excess of new mtDNA mutations in human population data. We experimentally addressed this issue by studying the maternal transmission of random mtDNA mutations in mtDNA mutator mice expressing a proofreading-deficient mitochondrial DNA polymerase. We report a rapid and strong elimination of nonsynonymous changes in protein-coding genes; the hallmark of purifying selection. There are striking similarities between the mutational patterns in our experimental mouse system and human mtDNA polymorphisms. These data show strong purifying selection against mutations within mtDNA protein-coding genes. To our knowledge, our study presents the first direct experimental observations of the fate of random mtDNA mutations in the mammalian germ line and demonstrates the importance of purifying selection in shaping mitochondrial sequence diversity. PMID:18232733

  9. Markov chain for estimating human mitochondrial DNA mutation pattern

    Science.gov (United States)

    Vantika, Sandy; Pasaribu, Udjianna S.

    2015-12-01

    The Markov chain was proposed to estimate the human mitochondrial DNA mutation pattern. One DNA sequence was taken randomly from 100 sequences in Genbank. The nucleotide transition matrix and mutation transition matrix were estimated from this sequence. We determined whether the states (mutation/normal) are recurrent or transient. The results showed that both of them are recurrent.

  10. Historically low mitochondrial DNA diversity in koalas (Phascolarctos cinereus)

    DEFF Research Database (Denmark)

    Tsangaras, Kyriakos; Avila Arcos, Maria del Carmen; Ishida, Yasuko;

    2012-01-01

    in koala population size during the late 1800s and early 1900s. To examine the effects of these reductions in population size on koala genetic diversity, we sequenced part of the hypervariable region of mitochondrial DNA (mtDNA) in koala museum specimens collected in the 19th and 20th centuries...

  11. Mitochondrial DNA sequence analysis of two mouse hepatocarcinoma cell lines

    Institute of Scientific and Technical Information of China (English)

    Ji-Gang Dai; Xia Lei; Jia-Xin Min; Guo-Qiang Zhang; Hong Wei

    2005-01-01

    AIM: To study genetic difference of mitochondrial DNA (mtDNA)between two hepatocarcinoma cell lines (Hca-F and Hca-P)with diverse metastatic characteristics and the relationship between mtDNA changes in cancer cells and their oncogenic phenotype.METHODS: Mitochondrial DNA D-loop, tRNAMet+Glu+Ile and ND3gene fragments from the hepatocarcinoma cell lines with 1100, 1126 and 534 bp in length respectively were analysed by PCR amplification and restriction fragment length polymorphism techniques. The D-loop 3' end sequence of the hepatocarcinoma cell lines was determined by sequencing.RESULTS: No amplification fragment length polymorphism and restriction fragment length polymorphism were observed in tRNAMet+Glu+Ile,ND3 and D-loop of mitochondrial DNA of the hepatocarcinoma cells. Sequence differences between Hca-F and Hca-P were found in mtDNA D-loop.CONCLUSION: Deletion mutations of mitochondrial DNA restriction fragment may not play a significant role in carcinogenesis. Genetic difference of mtDNA D-loop between Hca-F and Hca-P, which may reflect the environmental and genetic influences during tumor progression, could be linked to their tumorigenic phenotypes.

  12. Introgression of mitochondrial DNA among lineages in a hybridogenetic ant.

    Science.gov (United States)

    Darras, Hugo; Aron, Serge

    2015-02-01

    We report a remarkable pattern of incongruence between nuclear and mitochondrial variations in a social insect, the desert ant Cataglyphis hispanica. This species reproduces by social hybridogenesis. In all populations, two distinct genetic lineages coexist; non-reproductive workers develop from hybrid crosses between the lineages, whereas reproductive offspring (males and new queens) are typically produced asexually by parthenogenesis. Genetic analyses based on nuclear markers revealed that the two lineages remain highly differentiated despite constant hybridization for worker production. Here, we show that, in contrast with nuclear DNA, mitochondrial DNA (mtDNA) does not recover the two lineages as monophyletic. Rather, mitochondrial haplotypes cluster according to their geographical origin. We argue that this cytonuclear incongruence stems from introgression of mtDNA among lineages, and review the mechanisms likely to explain this pattern under social hybridogenesis.

  13. PCR primers for metazoan mitochondrial 12S ribosomal DNA sequences.

    Directory of Open Access Journals (Sweden)

    Ryuji J Machida

    Full Text Available BACKGROUND: Assessment of the biodiversity of communities of small organisms is most readily done using PCR-based analysis of environmental samples consisting of mixtures of individuals. Known as metagenetics, this approach has transformed understanding of microbial communities and is beginning to be applied to metazoans as well. Unlike microbial studies, where analysis of the 16S ribosomal DNA sequence is standard, the best gene for metazoan metagenetics is less clear. In this study we designed a set of PCR primers for the mitochondrial 12S ribosomal DNA sequence based on 64 complete mitochondrial genomes and then tested their efficacy. METHODOLOGY/PRINCIPAL FINDINGS: A total of the 64 complete mitochondrial genome sequences representing all metazoan classes available in GenBank were downloaded using the NCBI Taxonomy Browser. Alignment of sequences was performed for the excised mitochondrial 12S ribosomal DNA sequences, and conserved regions were identified for all 64 mitochondrial genomes. These regions were used to design a primer pair that flanks a more variable region in the gene. Then all of the complete metazoan mitochondrial genomes available in NCBI's Organelle Genome Resources database were used to determine the percentage of taxa that would likely be amplified using these primers. Results suggest that these primers will amplify target sequences for many metazoans. CONCLUSIONS/SIGNIFICANCE: Newly designed 12S ribosomal DNA primers have considerable potential for metazoan metagenetic analysis because of their ability to amplify sequences from many metazoans.

  14. Mitochondrial DNA triplication and punctual mutations in patients with mitochondrial neuromuscular disorders

    Energy Technology Data Exchange (ETDEWEB)

    Mkaouar-Rebai, Emna, E-mail: emna.mkaouar@gmail.com [Département des Sciences de la Vie, Faculté des Sciences de Sfax, Université de Sfax (Tunisia); Felhi, Rahma; Tabebi, Mouna [Laboratoire de Génétique Moléculaire Humaine, Faculté de Médecine de Sfax, Université de Sfax (Tunisia); Alila-Fersi, Olfa; Chamkha, Imen [Département des Sciences de la Vie, Faculté des Sciences de Sfax, Université de Sfax (Tunisia); Maalej, Marwa; Ammar, Marwa [Laboratoire de Génétique Moléculaire Humaine, Faculté de Médecine de Sfax, Université de Sfax (Tunisia); Kammoun, Fatma [Service de pédiatrie, C.H.U. Hedi Chaker de Sfax (Tunisia); Keskes, Leila [Laboratoire de Génétique Moléculaire Humaine, Faculté de Médecine de Sfax, Université de Sfax (Tunisia); Hachicha, Mongia [Service de pédiatrie, C.H.U. Hedi Chaker de Sfax (Tunisia); Fakhfakh, Faiza, E-mail: faiza.fakhfakh02@gmail.com [Département des Sciences de la Vie, Faculté des Sciences de Sfax, Université de Sfax (Tunisia)

    2016-04-29

    Mitochondrial diseases are a heterogeneous group of disorders caused by the impairment of the mitochondrial oxidative phosphorylation system which have been associated with various mutations of the mitochondrial DNA (mtDNA) and nuclear gene mutations. The clinical phenotypes are very diverse and the spectrum is still expanding. As brain and muscle are highly dependent on OXPHOS, consequently, neurological disorders and myopathy are common features of mtDNA mutations. Mutations in mtDNA can be classified into three categories: large-scale rearrangements, point mutations in tRNA or rRNA genes and point mutations in protein coding genes. In the present report, we screened mitochondrial genes of complex I, III, IV and V in 2 patients with mitochondrial neuromuscular disorders. The results showed the presence the pathogenic heteroplasmic m.9157G>A variation (A211T) in the MT-ATP6 gene in the first patient. We also reported the first case of triplication of 9 bp in the mitochondrial NC7 region in Africa and Tunisia, in association with the novel m.14924T>C in the MT-CYB gene in the second patient with mitochondrial neuromuscular disorder. - Highlights: • We reported 2 patients with mitochondrial neuromuscular disorders. • The heteroplasmic MT-ATP6 9157G>A variation was reported. • A triplication of 9 bp in the mitochondrial NC7 region was detected. • The m.14924T>C transition (S60P) in the MT-CYB gene was found.

  15. Detection of heteroplasmic mitochondrial DNA in single mitochondria.

    Directory of Open Access Journals (Sweden)

    Joseph E Reiner

    Full Text Available BACKGROUND: Mitochondrial DNA (mtDNA genome mutations can lead to energy and respiratory-related disorders like myoclonic epilepsy with ragged red fiber disease (MERRF, mitochondrial myopathy, encephalopathy, lactic acidosis and stroke (MELAS syndrome, and Leber's hereditary optic neuropathy (LHON. It is not well understood what effect the distribution of mutated mtDNA throughout the mitochondrial matrix has on the development of mitochondrial-based disorders. Insight into this complex sub-cellular heterogeneity may further our understanding of the development of mitochondria-related diseases. METHODOLOGY: This work describes a method for isolating individual mitochondria from single cells and performing molecular analysis on that single mitochondrion's DNA. An optical tweezer extracts a single mitochondrion from a lysed human HL-60 cell. Then a micron-sized femtopipette tip captures the mitochondrion for subsequent analysis. Multiple rounds of conventional DNA amplification and standard sequencing methods enable the detection of a heteroplasmic mixture in the mtDNA from a single mitochondrion. SIGNIFICANCE: Molecular analysis of mtDNA from the individually extracted mitochondrion demonstrates that a heteroplasmy is present in single mitochondria at various ratios consistent with the 50/50 heteroplasmy ratio found in single cells that contain multiple mitochondria.

  16. Cockayne syndrome group B protein promotes mitochondrial DNA stability by supporting the DNA repair association with the mitochondrial membrane

    DEFF Research Database (Denmark)

    Aamann, Maria Diget; Sorensen, Martin M; Hvitby, Christina Poulsen;

    2010-01-01

    Cockayne syndrome (CS) is a human premature aging disorder associated with severe developmental deficiencies and neurodegeneration, and phenotypically it resembles some mitochondrial DNA (mtDNA) diseases. Most patients belong to complementation group B, and the CS group B (CSB) protein plays a role...

  17. Mechanisms of Uniparental Mitochondrial DNA Inheritance in Cryptococcus neoformans

    OpenAIRE

    Gyawali, Rachana; Lin, Xiaorong

    2011-01-01

    In contrast to the nuclear genome, the mitochondrial genome does not follow Mendelian laws of inheritance. The nuclear genome of meiotic progeny comes from the recombination of both parental genomes, whereas the meiotic progeny could inherit mitochondria from one, the other, or both parents. In fact, one fascinating phenomenon is that mitochondrial DNA in the majority of eukaryotes is inherited from only one particular parent. Typically, such unidirectional and uniparental inheritance of mito...

  18. "Stiff neonate" with mitochondrial DNA depletion and secondary neurotransmitter defects.

    LENUS (Irish Health Repository)

    Moran, Margaret M

    2011-12-01

    Mitochondrial disorders comprise a heterogenous group. A neonate who presented with episodes of severe truncal hypertonia and apnea progressed to a hypokinetic rigid syndrome characterized by hypokinesia, tremulousness, profound head lag, absent suck and gag reflexes, brisk deep tendon reflexes, ankle and jaw clonus, and evidence of autonomic dysfunction. Analysis of cerebrospinal fluid neurotransmitters from age 7 weeks demonstrated low levels of amine metabolites (homovanillic acid and 5-hydroxyindoleacetic acid), tetrahydrobiopterin, and pyridoxal phosphate. Mitochondrial DNA quantitative studies on muscle homogenate demonstrated a mitochondrial DNA depletion disorder. Respiratory chain enzymology demonstrated decreased complex IV activity. Screening for mitochondrial DNA rearrangement disorders and sequencing relevant mitochondrial genes produced negative results. No clinical or biochemical response to treatment with pyridoxal phosphate, tetrahydrobiopterin, or l-dopa occurred. The clinical course was progressive, and the patient died at age 19 months. Mitochondrial disorders causing secondary neurotransmitter diseases are usually severe, but are rarely reported. This diagnosis should be considered in neonates or infants who present with hypertonia, hypokinesia rigidity, and progressive neurodegeneration.

  19. Mitochondrial swinger replication: DNA replication systematically exchanging nucleotides and short 16S ribosomal DNA swinger inserts.

    Science.gov (United States)

    Seligmann, Hervé

    2014-11-01

    Assuming systematic exchanges between nucleotides (swinger RNAs) resolves genomic 'parenthood' of some orphan mitochondrial transcripts. Twenty-three different systematic nucleotide exchanges (bijective transformations) exist. Similarities between transcription and replication suggest occurrence of swinger DNA. GenBank searches for swinger DNA matching the 23 swinger versions of human and mouse mitogenomes detect only vertebrate mitochondrial swinger DNA for swinger type AT+CG (from five different studies, 149 sequences) matching three human and mouse mitochondrial genes: 12S and 16S ribosomal RNAs, and cytochrome oxidase subunit I. Exchange AT+CG conserves self-hybridization properties, putatively explaining swinger biases for rDNA, against protein coding genes. Twenty percent of the regular human mitochondrial 16S rDNA consists of short swinger repeats (from 13 exchanges). Swinger repeats could originate from recombinations between regular and swinger DNA: duplicated mitochondrial genes of the parthenogenetic gecko Heteronotia binoei include fewer short AT+CG swinger repeats than non-duplicated mitochondrial genomes of that species. Presumably, rare recombinations between female and male mitochondrial genes (and in parthenogenetic situations between duplicated genes), favors reverse-mutations of swinger repeat insertions, probably because most inserts affect negatively ribosomal function. Results show that swinger DNA exists, and indicate that swinger polymerization contributes to the genesis of genetic material and polymorphism.

  20. Inherited Mendelian defects of nuclear-mitochondrial communication affecting the stability of mitochondrial DNA.

    Science.gov (United States)

    Limongelli, Anna; Tiranti, Valeria

    2002-11-01

    The presence of mtDNA abnormalities inherited as Mendelian traits indicates the existence of mutations in nuclear genes affecting the integrity of the mitochondrial genome. Two groups of nucleus-driven abnormalities have been described: qualitative alterations of mtDNA, i.e. multiple large-scale deletions of mtDNA, and quantitative decrease of the mtDNA copy number, i.e. tissue-specific depletion of mtDNA. Autosomal dominant or recessive (adPEO), progressive ophthalmoplegia and autosomal-recessive mitochondrial neurogastrointestinal encephalomyopathy (MNGIE), are three neurodegenerative disorders associated with the coexistence of wild-type mtDNA with several deletion-containing mtDNA species. Heterozygous mutations of the genes encoding the muscle-heart isoform of the adenosine diphosphate/adenosine triphosphate mitochondrial translocator (ANT1), the main subunit of polymerase gamma (POLG1), and of the putative mtDNA helicase (Twinkle) have been found in adPEO families linked to three different loci, on chromosomes 4q34-35, 10q24, and 15q25, respectively. Mutations in the gene encoding thymidine phosphorylase have been identified in several MNGIE patients. Severe, tissue-specific depletion of mtDNA is the molecular hallmark of rapidly progressive hepatopathies or myopathies of infancy and childhood. Two genes, deoxyguanosine kinase and thymidine kinase type 2, both involved in the mitochondrion-specific salvage pathways of deoxynucleotide pools, have been associated with depletion syndromes in selected families.

  1. Maternal inheritance and mitochondrial DNA variants in familial Parkinson's disease

    Directory of Open Access Journals (Sweden)

    Pfeiffer Ronald F

    2010-04-01

    Full Text Available Abstract Background Mitochondrial function is impaired in Parkinson's disease (PD and may contribute to the pathogenesis of PD, but the causes of mitochondrial impairment in PD are unknown. Mitochondrial dysfunction is recapitulated in cell lines expressing mitochondrial DNA (mtDNA from PD patients, implicating mtDNA variants or mutations, though the role of mtDNA variants or mutations in PD risk remains unclear. We investigated the potential contribution of mtDNA variants or mutations to the risk of PD. Methods We examined the possibility of a maternal inheritance bias as well as the association between mitochondrial haplogroups and maternal inheritance and disease risk in a case-control study of 168 multiplex PD families in which the proband and one parent were diagnosed with PD. 2-tailed Fisher Exact Tests and McNemar's tests were used to compare allele frequencies, and a t-test to compare ages of onset. Results The frequency of affected mothers of the proband with PD (83/167, 49.4% was not significantly different from the frequency of affected females of the proband generation (115/259, 44.4% (Odds Ratio 1.22; 95%CI 0.83 - 1.81. After correcting for multiple tests, there were no significant differences in the frequencies of mitochondrial haplogroups or of the 10398G complex I gene polymorphism in PD patients compared to controls, and no significant associations with age of onset of PD. Mitochondrial haplogroup and 10398G polymorphism frequencies were similar in probands having an affected father as compared to probands having an affected mother. Conclusions These data fail to demonstrate a bias towards maternal inheritance in familial PD. Consistent with this, we find no association of common haplogroup-defining mtDNA variants or for the 10398G variant with the risk of PD. However, these data do not exclude a role for mtDNA variants in other populations, and it remains possible that other inherited mitochondrial DNA variants, or somatic mDNA

  2. Nonneutral mitochondrial DNA variation in humans and chimpanzees

    Energy Technology Data Exchange (ETDEWEB)

    Nachman, M.W.; Aquadro, C.F. [Cornell Univ., Ithaca, NY (United States); Brown, W.M. [Univ. of Michigan, Ann Arbor, MI (United States)] [and others

    1996-03-01

    We sequenced the NADH dehydrogenase subunit 3 (ND3) gene from a sample of 61 humans, five common chimpanzees, and one gorilla to test whether patterns of mitochondrial DNA (mtDNA) variation are consistent with a neutral model of molecular evolution. Within humans and within chimpanzees, the ratio of replacement to silent nucleotide substitutions was higher than observed in comparisons between species, contrary to neutral expectations. To test the generality of this result, we reanalyzed published human RFLP data from the entire mitochondrial genome. Gains of restriction sites relative to a known human mtDNA sequence were used to infer unambiguous nucleotide substitutions. We also compared the complete mtDNA sequences of three humans. Both the RFLP data and the sequence data reveal a higher ratio of replacement to silent nucleotide substitutions within humans than is seen between species. This pattern is observed at most or all human mitochondrial genes and is inconsistent with a strictly neutral model. These data suggest that many mitochondrial protein polymorphisms are slightly deleterious, consistent with studies of human mitochondrial diseases. 59 refs., 2 figs., 8 tabs.

  3. Blood cell mitochondrial DNA content and premature ovarian aging.

    Directory of Open Access Journals (Sweden)

    Marco Bonomi

    Full Text Available Primary ovarian insufficiency (POI is a critical fertility defect characterized by an anticipated and silent impairment of the follicular reserve, but its pathogenesis is largely unexplained. The frequent maternal inheritance of POI together with a remarkable dependence of ovarian folliculogenesis upon mitochondrial biogenesis and bioenergetics suggested the possible involvement of a generalized mitochondrial defect. Here, we verified the existence of a significant correlation between blood and ovarian mitochondrial DNA (mtDNA content in a group of women undergoing ovarian hyperstimulation (OH, and then aimed to verify whether mtDNA content was significantly altered in the blood cells of POI women. We recruited 101 women with an impaired ovarian reserve: 59 women with premature ovarian failure (POF and 42 poor responders (PR to OH. A Taqman copy number assay revealed a significant mtDNA depletion (P<0.001 in both POF and PR women in comparison with 43 women of similar age and intact ovarian reserve, or 53 very old women with a previous physiological menopause. No pathogenic variations in the mitochondrial DNA polymerase γ (POLG gene were detected in 57 POF or PR women with low blood mtDNA content. In conclusion, blood cell mtDNA depletion is a frequent finding among women with premature ovarian aging, suggesting that a still undetermined but generalized mitochondrial defect may frequently predispose to POI which could then be considered a form of anticipated aging in which the ovarian defect may represent the first manifestation. The determination of mtDNA content in blood may become an useful tool for the POI risk prediction.

  4. Blood Cell Mitochondrial DNA Content and Premature Ovarian Aging

    Science.gov (United States)

    Cacciatore, Chiara; Busnelli, Marta; Rossetti, Raffaella; Bonetti, Silvia; Paffoni, Alessio; Mari, Daniela; Ragni, Guido; Persani, Luca; Arosio, M.; Beck-Peccoz, P.; Biondi, M.; Bione, S.; Bruni, V.; Brigante, C.; Cannavo`, S.; Cavallo, L.; Cisternino, M.; Colombo, I.; Corbetta, S.; Crosignani, P.G.; D'Avanzo, M.G.; Dalpra, L.; Danesino, C.; Di Battista, E.; Di Prospero, F.; Donti, E.; Einaudi, S.; Falorni, A.; Foresta, C.; Fusi, F.; Garofalo, N.; Giotti, I.; Lanzi, R.; Larizza, D.; Locatelli, N.; Loli, P.; Madaschi, S.; Maghnie, M.; Maiore, S.; Mantero, F.; Marozzi, A.; Marzotti, S.; Migone, N.; Nappi, R.; Palli, D.; Patricelli, M.G.; Pisani, C.; Prontera, P.; Petraglia, F.; Radetti, G.; Renieri, A.; Ricca, I.; Ripamonti, A.; Rossetti, R.; Russo, G.; Russo, S.; Tonacchera, M.; Toniolo, D.; Torricelli, F.; Vegetti, W.; Villa, N.; Vineis, P.; Wasniewsk, M.; Zuffardi, O.

    2012-01-01

    Primary ovarian insufficiency (POI) is a critical fertility defect characterized by an anticipated and silent impairment of the follicular reserve, but its pathogenesis is largely unexplained. The frequent maternal inheritance of POI together with a remarkable dependence of ovarian folliculogenesis upon mitochondrial biogenesis and bioenergetics suggested the possible involvement of a generalized mitochondrial defect. Here, we verified the existence of a significant correlation between blood and ovarian mitochondrial DNA (mtDNA) content in a group of women undergoing ovarian hyperstimulation (OH), and then aimed to verify whether mtDNA content was significantly altered in the blood cells of POI women. We recruited 101 women with an impaired ovarian reserve: 59 women with premature ovarian failure (POF) and 42 poor responders (PR) to OH. A Taqman copy number assay revealed a significant mtDNA depletion (P<0.001) in both POF and PR women in comparison with 43 women of similar age and intact ovarian reserve, or 53 very old women with a previous physiological menopause. No pathogenic variations in the mitochondrial DNA polymerase γ (POLG) gene were detected in 57 POF or PR women with low blood mtDNA content. In conclusion, blood cell mtDNA depletion is a frequent finding among women with premature ovarian aging, suggesting that a still undetermined but generalized mitochondrial defect may frequently predispose to POI which could then be considered a form of anticipated aging in which the ovarian defect may represent the first manifestation. The determination of mtDNA content in blood may become an useful tool for the POI risk prediction. PMID:22879975

  5. Mitochondrial DNA mutations and male infertility

    Directory of Open Access Journals (Sweden)

    Kumar D

    2009-01-01

    Full Text Available Infertility can be defined as difficulty in conceiving a child after 1 year of unprotected intercourse. Infertility can arise either because of the male factor or female factor or both. According to the current estimates, 15% of couples attempting their first pregnancy could not succeed. Infertility is either primary or secondary. Mitochondria have profound effect on all biochemical pathways, including the one that drivessperm motility. Sperm motility is heavily dependent on the ATP generated by oxidative phosphorylation in the mitochondrial sheath. In this review, the very positive role of mitochondrial genome′s association with infertility is discussed

  6. Alterations of the mitochondrial proteome caused by the absence of mitochondrial DNA: A proteomic view

    Science.gov (United States)

    Chevallet, Mireille; Lescuyer, Pierre; Diemer, Hélène; van Dorsselaer, Alain; Leize-Wagner, Emmanuelle; Rabilloud, Thierry

    2006-01-01

    The proper functioning of mitochondria requires that both the mitochondrial and the nuclear genome are functional. To investigate the importance of the mitochondrial genome, which encodes only 13 subunits of the respiratory complexes, the mitochondrial rRNAs and a few tRNAs, we performed a comparative study on the 143B cell line and on its Rho-0 counterpart, i.e. devoid of mitochondrial DNA. Quantitative differences were found, of course in the respiratory complexes subunits, but also in the mitochondrial translation apparatus, mainly mitochondrial ribosomal proteins, and in the ion and protein import system, i.e. including membrane proteins. Various mitochondrial metabolic processes were also altered, especially electron transfer proteins and some dehydrogenases, but quite often on a few proteins for each pathway. This study also showed variations in some hypothetical or poorly characterized proteins, suggesting a mitochondrial localization for these proteins. Examples include a stomatin-like protein and a protein sharing homologies with bacterial proteins implicated in tyrosine catabolism. Proteins involved in apoptosis control are also found modulated in Rho-0 mitochondria. PMID:16548050

  7. Mitochondrial DNA repair: a novel therapeutic target for heart failure.

    Science.gov (United States)

    Marín-García, José

    2016-09-01

    Mitochondria play a crucial role in a variety of cellular processes ranging from energy metabolism, generation of reactive oxygen species (ROS) and Ca(2+) handling to stress responses, cell survival and death. Malfunction of the organelle may contribute to the pathogenesis of neuromuscular, cancer, premature aging and cardiovascular diseases (CVD), including myocardial ischemia, cardiomyopathy and heart failure (HF). Mitochondria contain their own genome organized into DNA-protein complexes, called "mitochondrial nucleoids," along with multiprotein machineries, which promote mitochondrial DNA (mtDNA) replication, transcription and repair. Although the mammalian organelle possesses almost all known nuclear DNA repair pathways, including base excision repair, mismatch repair and recombinational repair, the proximity of mtDNA to the main sites of ROS production and the lack of protective histones may result in increased susceptibility to various types of mtDNA damage. These include accumulation of mtDNA point mutations and/or deletions and decreased mtDNA copy number, which will impair mitochondrial function and finally, may lead to CVD including HF.

  8. Automated DNA extraction of single dog hairs without roots for mitochondrial DNA analysis.

    Science.gov (United States)

    Bekaert, Bram; Larmuseau, Maarten H D; Vanhove, Maarten P M; Opdekamp, Anouschka; Decorte, Ronny

    2012-03-01

    Dogs are intensely integrated in human social life and their shed hairs can play a major role in forensic investigations. The overall aim of this study was to validate a semi-automated extraction method for mitochondrial DNA analysis of telogenic dog hairs. Extracted DNA was amplified with a 95% success rate from 43 samples using two new experimental designs in which the mitochondrial control region was amplified as a single large (± 1260 bp) amplicon or as two individual amplicons (HV1 and HV2; ± 650 and 350 bp) with tailed-primers. The results prove that the extraction of dog hair mitochondrial DNA can easily be automated to provide sufficient DNA yield for the amplification of a forensically useful long mitochondrial DNA fragment or alternatively two short fragments with minimal loss of sequence in case of degraded samples.

  9. Intrinsic mitochondrial DNA repair defects in Ataxia Telangiectasia.

    Science.gov (United States)

    Sharma, Nilesh K; Lebedeva, Maria; Thomas, Terace; Kovalenko, Olga A; Stumpf, Jeffrey D; Shadel, Gerald S; Santos, Janine H

    2014-01-01

    Ataxia Telangiectasia (A-T) is a progressive childhood disorder characterized most notably by cerebellar degeneration and predisposition to cancer. A-T is caused by mutations in the kinase ATM, a master regulator of the DNA double-strand break response. In addition to DNA-damage signaling defects, A-T cells display mitochondrial dysfunction that is thought to contribute to A-T pathogenesis. However, the molecular mechanism leading to mitochondrial dysfunction in A-T remains unclear. Here, we show that lack of ATM leads to reduced mitochondrial DNA (mtDNA) integrity and mitochondrial dysfunction, which are associated to defective mtDNA repair. While protein levels of mtDNA repair proteins are essentially normal, in the absence of ATM levels specifically of DNA ligase III (Lig3), the only DNA ligase working in mitochondria is reduced. The reduction of Lig3 is observed in different A-T patient cells, in brain and pre-B cells derived from ATM knockout mice as well as upon transient or stable knockdown of ATM. Furthermore, pharmacological inhibition of Lig3 in wild type cells phenocopies the mtDNA repair defects observed in A-T patient cells. As targeted deletion of LIG3 in the central nervous system causes debilitating ataxia in mice, reduced Lig3 protein levels and the consequent mtDNA repair defect may contribute to A-T neurodegeneration. A-T is thus the first disease characterized by diminished Lig3. Published by Elsevier B.V.

  10. Mitochondrial DNA Mutation Associated with Leber's Hereditary Optic Neuropathy

    Science.gov (United States)

    Wallace, Douglas C.; Singh, Gurparkash; Lott, Marie T.; Hodge, Judy A.; Schurr, Theodore G.; Lezza, Angela M. S.; Elsas, Louis J.; Nikoskelainen, Eeva K.

    1988-12-01

    Leber's hereditary optic neuropathy is a maternally inherited disease resulting in optic nerve degeneration and cardiac dysrhythmia. A mitochondrial DNA replacement mutation was identified that correlated with this disease in multiple families. This mutation converted a highly conserved arginine to a histidine at codon 340 in the NADH dehydrogenase subunit 4 gene and eliminated an Sfa NI site, thus providing a simple diagnostic test. This finding demonstrated that a nucleotide change in a mitochondrial DNA energy production gene can result in a neurological disease.

  11. Distribution patterns of postmortem damage in human mitochondrial DNA

    DEFF Research Database (Denmark)

    Gilbert, M Thomas P; Willerslev, Eske; Hansen, Anders J

    2002-01-01

    The distribution of postmortem damage in mitochondrial DNA retrieved from 37 ancient human DNA samples was analyzed by cloning and was compared with a selection of published animal data. A relative rate of damage (rho(v)) was calculated for nucleotide positions within the human hypervariable region......, such as MT5, have lower in vivo mutation rates and lower postmortem-damage rates. The postmortem data also identify a possible functional subregion of the HVR1, termed "low-diversity 1," through the lack of sequence damage. The amount of postmortem damage observed in mitochondrial coding regions...

  12. A novel interaction between DNA ligase III and DNA polymerase gamma plays an essential role in mitochondrial DNA stability.

    Science.gov (United States)

    De, Ananya; Campbell, Colin

    2007-02-15

    The data in the present study show that DNA polymerase gamma and DNA ligase III interact in mitochondrial protein extracts from cultured HT1080 cells. An interaction was also observed between the two recombinant proteins in vitro. Expression of catalytically inert versions of DNA ligase III that bind DNA polymerase gamma was associated with reduced mitochondrial DNA copy number and integrity. In contrast, overexpression of wild-type DNA ligase III had no effect on mitochondrial DNA copy number or integrity. Experiments revealed that wild-type DNA ligase III facilitates the interaction of DNA polymerase gamma with a nicked DNA substrate in vitro, and that the zinc finger domain of DNA ligase III is required for this activity. Mitochondrial protein extracts prepared from cells overexpressing a DNA ligase III protein that lacked the zinc finger domain had reduced base excision repair activity compared with extracts from cells overexpressing the wild-type protein. These data support the interpretation that the interaction of DNA ligase III and DNA polymerase gamma is required for proper maintenance of the mammalian mitochondrial genome.

  13. DNA methyltransferase 1 mutations and mitochondrial pathology: is mtDNA methylated?

    Directory of Open Access Journals (Sweden)

    Alessandra eMaresca

    2015-03-01

    Full Text Available Autosomal dominant cerebellar ataxia, deafness and narcolepsy (ADCA-DN and Hereditary sensory neuropathy with dementia and hearing loss (HSN1E are two rare, overlapping neurodegenerative syndromes that have been recently linked to allelic dominant pathogenic mutations in the DNMT1 gene, coding for DNA (cytosine-5-methyltransferase 1. DNMT1 is the enzyme responsible for maintaining the nuclear genome methylation patterns during the DNA replication and repair, thus regulating gene expression. The mutations responsible for ADCA-DN and HSN1E affect the replication foci targeting sequence domain, which regulates DNMT1 binding to chromatin. DNMT1 dysfunction is anticipated to lead to a global alteration of the DNA methylation pattern with predictable downstream consequences on gene expression. Interestingly, ADCA-DN and HSN1E phenotypes share some clinical features typical of mitochondrial diseases, such as optic atrophy, peripheral neuropathy and deafness, and some biochemical evidence of mitochondrial dysfunction. The recent discovery of a mitochondrial isoform of DNMT1 and its proposed role in methylating mitochondrial DNA (mtDNA suggests that DNMT1 mutations may directly affect mtDNA and mitochondrial physiology. On the basis of this latter finding the link between DNMT1 abnormal activity and mitochondrial dysfunction in ADCA-DN and HSN1E appears intuitive, however mtDNA methylation remains highly debated. In the last years several groups demonstrated the presence of 5-methylcytosine in mtDNA by different approaches, but, on the other end, the opposite evidence that mtDNA is not methylated has also been published. Since over 1500 mitochondrial proteins are encoded by the nuclear genome, the altered methylation of these genes may well have a critical role in leading to the mitochondrial impairment observed in ADCA-DN and HSN1E. Thus, many open questions still remain unanswered, such as why mtDNA should be methylated, and how this process is

  14. Effects of reduced mitochondrial DNA content on secondary mitochondrial toxicant exposure in Caenorhabditis elegans.

    Science.gov (United States)

    Luz, Anthony L; Meyer, Joel N

    2016-09-01

    The mitochondrial genome (mtDNA) is intimately linked to cellular and organismal health, as demonstrated by the fact that mutations in and depletion of mtDNA result in severe mitochondrial disease in humans. However, cells contain hundreds to thousands of copies of mtDNA, which provides genetic redundancy, and creates a threshold effect in which a large percentage of mtDNA must be lost prior to clinical pathogenesis. As certain pharmaceuticals and genetic mutations can result in depletion of mtDNA, and as many environmental toxicants target mitochondria, it is important to understand whether reduced mtDNA will sensitize an individual to toxicant exposure. Here, using ethidium bromide (EtBr), which preferentially inhibits mtDNA replication, we reduced mtDNA 35-55% in the in vivo model organism Caenorhabditis elegans. Chronic, lifelong, low-dose EtBr exposure did not disrupt nematode development or lifespan, and induced only mild alterations in mitochondrial respiration, while having no effect on steady-state ATP levels. Next, we exposed nematodes with reduced mtDNA to the known and suspected mitochondrial toxicants aflatoxin B1, arsenite, paraquat, rotenone or ultraviolet C radiation (UVC). EtBr pre-exposure resulted in mild sensitization of nematodes to UVC and arsenite, had no effect on AfB1 and paraquat, and provided some protection from rotenone toxicity. These mixed results provide a first line of evidence suggesting that reduced mtDNA content may sensitize an individual to certain environmental exposures.

  15. Fecal source tracking in water using a mitochondrial DNA microarray.

    Science.gov (United States)

    Vuong, Nguyet-Minh; Villemur, Richard; Payment, Pierre; Brousseau, Roland; Topp, Edward; Masson, Luke

    2013-01-01

    A mitochondrial-based microarray (mitoArray) was developed for rapid identification of the presence of 28 animals and one family (cervidae) potentially implicated in fecal pollution in mixed activity watersheds. Oligonucleotide probes for genus or subfamily-level identification were targeted within the 12S rRNA - Val tRNA - 16S rRNA region in the mitochondrial genome. This region, called MI-50, was selected based on three criteria: 1) the ability to be amplified by universal primers 2) these universal primer sequences are present in most commercial and domestic animals of interest in source tracking, and 3) that sufficient sequence variation exists within this region to meet the minimal requirements for microarray probe discrimination. To quantify the overall level of mitochondrial DNA (mtDNA) in samples, a quantitative-PCR (Q-PCR) universal primer pair was also developed. Probe validation was performed using DNA extracted from animal tissues and, for many cases, animal-specific fecal samples. To reduce the amplification of potentially interfering fish mtDNA sequences during the MI-50 enrichment step, a clamping PCR method was designed using a fish-specific peptide nucleic acid. DNA extracted from 19 water samples were subjected to both array and independent PCR analyses. Our results confirm that the mitochondrial microarray approach method could accurately detect the dominant animals present in water samples emphasizing the potential for this methodology in the parallel scanning of a large variety of animals normally monitored in fecal source tracking.

  16. [An efficient method for isolation of mitochondrial DNA in wheat].

    Science.gov (United States)

    Li, Wen-Qiang; Zhang, Gai-Sheng; Wang, Kui; Niu, Na; Pan, Dong-Liang

    2007-06-01

    An efficient method for isolation of mitochondrial DNA (mtDNA) from etiolated tissues of wheat was developed. The protocol consists of mitochondria isolation with differential centrifugation, Dnase I treatment, lysis with SDS and proteinase K, removing protein by TE-saturated phenol/chloroform extraction and a final RNase A treatment for obtaining mtDNA. The mtDNA samples were tested using spectrophotometry and agarose gel electrophoresis. It was proved that the mtDNA isolated by this method not only have the high yield but also structural complete, and contains no impurities, such as nuclear DNA, RNA and protein. The result showed that this high quality mtDNA can be successfully used in PCR and other genetic studies. In addition, it was found that adjusting the lysis temperature has a noticeable effect on the mtDNA yield.

  17. Hybrid male sterility is caused by mitochondrial DNA deletion.

    Science.gov (United States)

    Hayashida, Kenji; Kohno, Shigeru

    2009-07-01

    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.

  18. Statistical analysis of post mortem DNA damage-derived miscoding lesions in Neandertal mitochondrial DNA

    NARCIS (Netherlands)

    S. Vives (Sergi); M.T. Gilbert (Thomas); C. Arenas (Conchita); E. Gigli (Elena); O. Lao Grueso (Oscar); C. Lalueza-Fox (Carles)

    2008-01-01

    textabstractBackground. We have analysed the distribution of post mortem DNA damage derived miscoding lesions from the datasets of seven published Neandertal specimens that have extensive cloned sequence coverage over the mitochondrial DNA (mtDNA) hypervariable region 1 (HVS1). The analysis was rest

  19. Absence of pathogenic mitochondrial DNA mutations in mouse brain tumors

    Directory of Open Access Journals (Sweden)

    Seyfried Thomas N

    2005-08-01

    Full Text Available Abstract Background Somatic mutations in the mitochondrial genome occur in numerous tumor types including brain tumors. These mutations are generally found in the hypervariable regions I and II of the displacement loop and unlikely alter mitochondrial function. Two hypervariable regions of mononucleotide repeats occur in the mouse mitochondrial genome, i.e., the origin of replication of the light strand (OL and the Arg tRNA. Methods In this study we examined the entire mitochondrial genome in a series of chemically induced brain tumors in the C57BL/6J strain and spontaneous brain tumors in the VM mouse strain. The tumor mtDNA was compared to that of mtDNA in brain mitochondrial populations from the corresponding syngeneic mouse host strain. Results Direct sequencing revealed a few homoplasmic base pair insertions, deletions, and substitutions in the tumor cells mainly in regions of mononucleotide repeats. A heteroplasmic mutation in the 16srRNA gene was detected in a spontaneous metastatic VM brain tumor. Conclusion None of the mutations were considered pathogenic, indicating that mtDNA somatic mutations do not likely contribute to the initiation or progression of these diverse mouse brain tumors.

  20. Atomistic Molecular Dynamics Simulations of Mitochondrial DNA Polymerase γ

    DEFF Research Database (Denmark)

    Euro, Liliya; Haapanen, Outi; Róg, Tomasz

    2017-01-01

    DNA polymerase γ (Pol γ) is a key component of the mitochondrial DNA replisome and an important cause of neurological diseases. Despite the availability of its crystal structures, the molecular mechanism of DNA replication, the switch between polymerase and exonuclease activities, the site...... of replisomal interactions, and functional effects of patient mutations that do not affect direct catalysis have remained elusive. Here we report the first atomistic classical molecular dynamics simulations of the human Pol γ replicative complex. Our simulation data show that DNA binding triggers remarkable...

  1. Large sequence divergence of mitochondrial DNA genotypes of the control region within populations of the African antelope, kob (Kobus kob)

    DEFF Research Database (Denmark)

    Birungi, J.; Arctander, Peter

    2000-01-01

    conservation genetics, control region, Kobus kob, mitochondrial DNA, population expansion, population structure......conservation genetics, control region, Kobus kob, mitochondrial DNA, population expansion, population structure...

  2. Reduction of nuclear encoded enzymes of mitochondrial energy metabolism in cells devoid of mitochondrial DNA

    Energy Technology Data Exchange (ETDEWEB)

    Mueller, Edith E., E-mail: ed.mueller@salk.at [Research Program for Receptor Biochemistry and Tumor Metabolism, Department of Pediatrics, Muellner Hauptstrasse 48, 5020 Salzburg (Austria); Mayr, Johannes A., E-mail: h.mayr@salk.at [Research Program for Receptor Biochemistry and Tumor Metabolism, Department of Pediatrics, Muellner Hauptstrasse 48, 5020 Salzburg (Austria); Zimmermann, Franz A., E-mail: f.zimmermann@salk.at [Research Program for Receptor Biochemistry and Tumor Metabolism, Department of Pediatrics, Muellner Hauptstrasse 48, 5020 Salzburg (Austria); Feichtinger, Rene G., E-mail: r.feichtinger@salk.at [Research Program for Receptor Biochemistry and Tumor Metabolism, Department of Pediatrics, Muellner Hauptstrasse 48, 5020 Salzburg (Austria); Stanger, Olaf, E-mail: o.stanger@rbht.nhs.uk [Department of Cardiac Surgery, Paracelsus Medical University, Muellner Hauptstrasse 48, 5020 Salzburg (Austria); Sperl, Wolfgang, E-mail: w.sperl@salk.at [Research Program for Receptor Biochemistry and Tumor Metabolism, Department of Pediatrics, Muellner Hauptstrasse 48, 5020 Salzburg (Austria); Kofler, Barbara, E-mail: b.kofler@salk.at [Research Program for Receptor Biochemistry and Tumor Metabolism, Department of Pediatrics, Muellner Hauptstrasse 48, 5020 Salzburg (Austria)

    2012-01-20

    Highlights: Black-Right-Pointing-Pointer We examined OXPHOS and citrate synthase enzyme activities in HEK293 cells devoid of mtDNA. Black-Right-Pointing-Pointer Enzymes partially encoded by mtDNA show reduced activities. Black-Right-Pointing-Pointer Also the entirely nuclear encoded complex II and citrate synthase exhibit reduced activities. Black-Right-Pointing-Pointer Loss of mtDNA induces a feedback mechanism that downregulates complex II and citrate synthase. -- Abstract: Mitochondrial DNA (mtDNA) depletion syndromes are generally associated with reduced activities of oxidative phosphorylation (OXPHOS) enzymes that contain subunits encoded by mtDNA. Conversely, entirely nuclear encoded mitochondrial enzymes in these syndromes, such as the tricarboxylic acid cycle enzyme citrate synthase (CS) and OXPHOS complex II, usually exhibit normal or compensatory enhanced activities. Here we report that a human cell line devoid of mtDNA (HEK293 {rho}{sup 0} cells) has diminished activities of both complex II and CS. This finding indicates the existence of a feedback mechanism in {rho}{sup 0} cells that downregulates the expression of entirely nuclear encoded components of mitochondrial energy metabolism.

  3. MITOCHONDRIAL DNA POLYMORPHISM IN CONTROL REGION FROM CHINESE YUGU POPULATION

    Institute of Scientific and Technical Information of China (English)

    刘新社; 李生斌

    2004-01-01

    Objective To investigate the mitochondrial DNA sequence polymorphism sites in Chinese YUGU ethnic group and to provide basic data used in forensic purpose. Methods Genomic DNA was extracted from the hole blood of 100 unrelated individuals of Chinese YUGU ethnic group by standard chelex-100 method. The sequence polymorphism sites was determined by PCR amplification and direct sequencing. Results 54 polymorphic sites were noted in mtDNA np16091-16418 region, and 46 haplotypes were identified. The genetic diversity was calculated to be 0.9691, and the genetic identity was calculated to be 0.0406. Conclusion There are some particular polymorphism sites in Chinese YUGU ethnic group. The results suggest that sequence polymorphism from np16091-16418 in human mitochondrial DNA can be used as a biological marker for forensic identity.

  4. Implementation of dna mitochondrial analysis in rhinoclemmys nasuta (testudines: geoemydidae)

    OpenAIRE

    Yherson Franchesco Molina Henao; Guillermo Barreto; Alan Giraldo

    2014-01-01

    Rhinoclemmys nasuta (Testudines:Geoemydidae) is considered an almost endemic specie to Colombia and the most primitive species of Rhynoclemmys. However, it is classified data deficient by IUCN because the available information is not enough to make a direct or indirect assessment of its extinction risk. Here, we describe the implementation of the method to analyze the mitochondrial DNA control sequence (mtDNA) of R. nasuta in order to generate tools for future studies in systematics and popul...

  5. Mitochondrial DNA sequence analysis of patients with 'atypical psychosis'.

    Science.gov (United States)

    Kazuno, An-A; Munakata, Kae; Mori, Kanako; Tanaka, Masashi; Nanko, Shinichiro; Kunugi, Hiroshi; Umekage, Tadashi; Tochigi, Mamoru; Kohda, Kazuhisa; Sasaki, Tsukasa; Akiyama, Tsuyoshi; Washizuka, Shinsuke; Kato, Nobumasa; Kato, Tadafumi

    2005-08-01

    Although classical psychopathological studies have shown the presence of an independent diagnostic category, 'atypical psychosis', most psychotic patients are currently classified into two major diagnostic categories, schizophrenia and bipolar disorder, by the Diagnostic and Statistical Manual of Mental Disorders (4th edn; DSM-IV) criteria. 'Atypical psychosis' is characterized by acute confusion without systematic delusion, emotional instability, and psychomotor excitement or stupor. Such clinical features resemble those seen in organic mental syndrome, and differential diagnosis is often difficult. Because patients with mitochondrial myopathy, encephalopathy, lactic acidosis, and stroke-like episodes (MELAS) sometimes show organic mental disorder, 'atypical psychosis' may be caused by mutations of mitochondrial DNA (mtDNA) in some patients. In the present study whole mtDNA was sequenced for seven patients with various psychotic disorders, who could be categorized as 'atypical psychosis'. None of them had known mtDNA mutations pathogenic for mitochondrial encephalopathy. Two of seven patients belonged to a subhaplogroup F1b1a with low frequency. These results did not support the hypothesis that clinical presentation of some patients with 'atypical psychosis' is a reflection of subclinical mitochondrial encephalopathy. However, the subhaplogroup F1b1a may be a good target for association study of 'atypical psychosis'.

  6. A Polymorphism in Mitochondrial DNA Associated with IQ?

    Science.gov (United States)

    Skuder, Patricia; And Others

    1995-01-01

    Of 100 DNA markers examined in an allelic association study, only 1 showed a replicated association with IQ in samples totaling 107 children. How the gene marked by the particular restriction fragment length polymorphism was tracked and its mitochondrial origin identified is described. (SLD)

  7. Mitochondrial DNA insertions in the nuclear Capra hircus genome.

    Science.gov (United States)

    Ning, F Y; Fu, J; Du, Z H

    2017-01-23

    Nuclear mitochondrial pseudogenes (numts), originating from mtDNA insertions into the nuclear genome, have been detected in many species. However, the distribution of numts in the newly published nuclear genome of domestic goat (Capra hircus) has not yet been explored. We used the entire goat mtDNA sequence and nuclear genome, to identify 118 numts using BLAST. Of these, 79 were able to map sequences to the genome. Further analysis showed that the size of the numts ranged from 318 to 9608 bp, and the homologous identity between numts and their respective corresponding mtDNA fragments varied between 65 and 99%. The identified Yunnan black goat numts covered nearly all the mitochondrial genes including mtDNA control region, and were distributed over all chromosomes with the exception of chromosomes 18, 21, and 25. The Y chromosome was excluded from our analysis, as sequence data are currently not available. Among the discovered 79 numts that we were able to map to the genome, 26 relatively complete mitochondrial genes were detected. Our results constitute valuable information for subsequent studies related to mitochondrial genes and goat evolution.

  8. Mitochondrial DNA HV1 and HV2 variation in Danes

    DEFF Research Database (Denmark)

    Mikkelsen, Martin; Sørensen, Erik; Rasmussen, Erik Michael;

    2010-01-01

    Sequences from the two hypervariable regions (HV1 and HV2) of the control region of the mitochondrial DNA were obtained from a total of 201 Danes and five individuals who later were recognized to be of non-West European origin. Two fractions of each region were amplified separately and sequenced ...

  9. Fly Diversity Revealed by PCR-RFLP of Mitochondrial DNA

    Science.gov (United States)

    Asraoui, Jimmy F.; Sayar, Nancy P.; Knio, Khouzama M.; Smith, Colin A.

    2008-01-01

    In this article, we describe an inexpensive, two-session undergraduate laboratory activity that introduces important molecular biology methods in the context of biodiversity. In the first session, students bring tentatively identified flies (order Diptera, true flies) to the laboratory, extract DNA, and amplify a region of the mitochondrial gene…

  10. Paternal inheritance of mitochondrial DNA in the sheep (Ovine aries)

    Institute of Scientific and Technical Information of China (English)

    ZHAO; Xingbo

    2001-01-01

    [1]Albuquerque, L. G., Keown, J. F., Van Vleck, L. D., Variances of direct genetic effects, maternal genetic effects, and cyto-plasmic inheritance effects for milk yield, fat yield, and fat percentage, J. Dairy Sci., 1998, 81(2): 544-549.[2]Wallace, D. C., Mitochondrial diseases in man and mouse, Science, 1999, 283: 1482-1488.[3]Gray, M. W., Burger, G., Lang, B. F., Mitochondrial evolution, Science, 1999, 283: 1476-1481.[4]Sutovsky, P., Schatten, G., Paternal contributions to the mammalian zygote: fertilization after sperm-egg fusion. Int. Rev. Cytol., 2000, 195: 1-65.[5]Hutchison, C. A., Newbold, J. E., Potter, S. S. et al., Maternal inheritance of mammalian mitochondrial DNA, Nature, 1974, 251: 536-538.[6]Kondo, R., Matsuura, E. T., Chigusa, S. I., Further observation of paternal transmission of Drosophila mitochondrial DNA by PCR selective amplification method, Genet. Res., 1992, 59(2):81-84.[7]Danan, C., Sternberg, D., Van Steirteghem, A. et al., Evaluation of parental mitochondrial inheritance in neonates born af-ter intracytoplasmic sperm injection, Am. J. Hum. Genet., 1999, 65(2): 463-473.[8]Gyllensten, U., Wharton, D., Josefsson, A. et al., Paternal inheritance of mitochondrial DNA in mice, Nature, 1991, 352(6332): 255-257.[9]Shitara, H., Hayashi, J. I., Takahama, S. et al., Maternal inheritance of mouse mtDNA in interspecific hybrids: segregation of the leaked paternal mtDNA followed by the prevention of subsequent paternal leakage, Genetics, 1998, 148(2): 851-857.[10]Steinborn, R., Zakhartchenko, V., Jelyazkkov, J. et al., Composition of parental mitochondrial DNA in cloned bovine em-bryos, FEBS Lett., 1998, 426(3): 352-356.[11]Lansman, R. A., Avise, J. C., Huettel, M. D. et al., Critical experimental test of the possibility of "paternal leakage" of mitochondrial DNA, Proc. Natl. Acad. Sci. USA, 1983, 80(7): 1969-1971.[12]Meusel, M. S., Moritz, R. F., Transfer of paternal mitochondrial DNA during

  11. Mitochondrial and chloroplast DNA based phylogeny of Pelargonium (Geraniaceae)

    NARCIS (Netherlands)

    Bakker, F.T.; Culham, A.; Pankhurst, C.E.; Gibby, M.

    2000-01-01

    Overall phylogenetic relationships within the genus Pelargonium (Geraniaceae) were inferred based on DNA sequences from mitochondrial(mt)-encoded nad1 b/c exons and from chloroplast(cp)-encoded trnL (UAA) 5' exon-trnF (GAA) exon regions using two species of Geranium and Sarcocaulon vanderetiae as ou

  12. Mitochondrial DNA insertions in the nuclear horse genome.

    Science.gov (United States)

    Nergadze, S G; Lupotto, M; Pellanda, P; Santagostino, M; Vitelli, V; Giulotto, E

    2010-12-01

    The insertion of mitochondrial DNA in the nuclear genome generates numts, nuclear sequences of mitochondrial origin. In the horse reference genome, we identified 82 numts and showed that the entire horse mitochondrial DNA is represented as numts without gross bias. Numts were inserted in the horse nuclear genome at random sites and were probably generated during the repair of DNA double-strand breaks. We then analysed 12 numt loci in 20 unrelated horses and found that null alleles, lacking the mitochondrial DNA insertion, were present at six of these loci. At some loci, the null allele is prevalent in the sample analysed, suggesting that, in the horse population, the number of numt loci may be higher than 82 present in the reference genome. Contrary to humans, the insertion polymorphism of numts is extremely frequent in the horse population, supporting the hypothesis that the genome of this species is in a rapidly evolving state. © 2010 The Authors, Journal compilation © 2010 Stichting International Foundation for Animal Genetics.

  13. Replication stalling by catalytically impaired Twinkle induces mitochondrial DNA rearrangements in cultured cells

    NARCIS (Netherlands)

    Pohjoismaki, J.L.; Goffart, S.; Spelbrink, J.N.

    2011-01-01

    Pathological mitochondrial DNA (mtDNA) rearrangements have been proposed to result from repair of double-strand breaks caused by blockage of mitochondrial DNA (mtDNA) replication. As mtDNA deletions are seen only in post-mitotic tissues, it has been suggested that they are selected out in actively d

  14. Mitochondrial DNA sequence of Onychostoma rara.

    Science.gov (United States)

    Zeng, Chun-Fang; Li, Xiao-Ling; Li, Chuan-Wu; Huang, Xiang-Rong; Wan, Yi-Wen

    2015-01-01

    The complete mitochondrial genome sequence of Onychostoma rara was determined to be 16,590 bp in length and contains 13 protein-coding genes (PCGs), 22 tRNA genes, large (rrnL) and small (rrnS) rRNA and the non-coding control region. Its total A + T content is 55.65%. We also analyzed the structure of control region, 6 CSBs (CSB-1, CSB-2, CSB-3, CSB-D, CSB-E and CSB-F) and 2 bp tandem repeat were detected.

  15. Mitochondrial DNA as a potential tool for early cancer detection

    Directory of Open Access Journals (Sweden)

    Parr Ryan L

    2006-01-01

    Full Text Available Abstract The recent surge in mitochondrial research has been driven by the identification of mitochondria-associated diseases and the role of mitochondria in apoptosis. Both of these aspects have identified mitochondrial analysis as a vital component of medical research. Moreover, mitochondria have been implicated in the process of carcinogenesis because of their vital role in energy production, nuclear-cytoplasmic signal integration and control of metabolic pathways. Interestingly, at some point during neoplastic transformation, there is an increase in reactive oxygen species, which damage the mitochondrial genome. This accelerates the somatic mutation rate of mitochondrial DNA. It has been proposed that these mutations may serve as an early indication of potential cancer development and may represent a means for tracking tumour progression. The purpose of this review is to explore the potential utility that these mutations may afford for the identification and monitoring of neoplasia and malignant transformation where appropriate body fluids or non-invasive tissue access is available for mitochondrial DNA recovery. Specifically, prostate, breast, colorectal, skin and lung cancers are discussed.

  16. Amphetamines promote mitochondrial dysfunction and DNA damage in pulmonary hypertension

    Science.gov (United States)

    Chen, Pin-I; Cao, Aiqin; Miyagawa, Kazuya; Tojais, Nancy F.; Hennigs, Jan K.; Li, Caiyun G.; Sweeney, Nathaly M.; Inglis, Audrey S.; Wang, Lingli; Li, Dan; Ye, Matthew; Feldman, Brian J.

    2017-01-01

    Amphetamine (AMPH) or methamphetamine (METH) abuse can cause oxidative damage and is a risk factor for diseases including pulmonary arterial hypertension (PAH). Pulmonary artery endothelial cells (PAECs) from AMPH-associated-PAH patients show DNA damage as judged by γH2AX foci and DNA comet tails. We therefore hypothesized that AMPH induces DNA damage and vascular pathology by interfering with normal adaptation to an environmental perturbation causing oxidative stress. Consistent with this, we found that AMPH alone does not cause DNA damage in normoxic PAECs, but greatly amplifies DNA damage in hypoxic PAECs. The mechanism involves AMPH activation of protein phosphatase 2A, which potentiates inhibition of Akt. This increases sirtuin 1, causing deacetylation and degradation of HIF1α, thereby impairing its transcriptional activity, resulting in a reduction in pyruvate dehydrogenase kinase 1 and impaired cytochrome c oxidase 4 isoform switch. Mitochondrial oxidative phosphorylation is inappropriately enhanced and, as a result of impaired electron transport and mitochondrial ROS increase, caspase-3 is activated and DNA damage is induced. In mice given binge doses of METH followed by hypoxia, HIF1α is suppressed and pulmonary artery DNA damage foci are associated with worse pulmonary vascular remodeling. Thus, chronic AMPH/METH can induce DNA damage associated with vascular disease by subverting the adaptive responses to oxidative stress. PMID:28138562

  17. The use of mitochondrial DNA (mtDNA-investigations in Forensic Sciences

    Directory of Open Access Journals (Sweden)

    S. Dawson

    1996-07-01

    Full Text Available A variety of methods was developed to characterize mtDNA. The initial aim of these techniques was to try and link diseases with specific mitochondrial defects. As a result of the maternal inheritance trait of mtDNA these techniques facilitate studies of the phylogenetic history and population structure of the human population. It has been shown that mitochondrial DNA typing can be of great value for human identification in forensic cases. The identification of victims of mass-disasters or mass-murders, where human remains can be recovered only after many years have passed, is one of the most challenging fields of forensic identification. By using automated DNA sequencing with fluorescent labels, mitochondrial DNA sequences can be generated rapidly and accurately. Computer software facilitates the rapid comparison of individual and reference sequences.

  18. Mitochondrial DNA: A Blind Spot in Neuroepigenetics

    OpenAIRE

    Manev, Hari; Dzitoyeva, Svetlana; Chen, Hu

    2012-01-01

    Neuroepigenetics, which includes nuclear DNA modifications such as 5-methylcytosine and 5-hydoxymethylcytosine and modifications of nuclear proteins such as histones, is emerging as the leading field in molecular neuroscience. Historically, a functional role for epigenetic mechanisms, including in neuroepigenetics, has been sought in the area of the regulation of nuclear transcription. However, one important compartment of mammalian cell DNA, different from nuclear but equally important for p...

  19. Private mitochondrial DNA variants in danish patients with hypertrophic cardiomyopathy.

    Directory of Open Access Journals (Sweden)

    Christian M Hagen

    Full Text Available Hypertrophic cardiomyopathy (HCM is a genetic cardiac disease primarily caused by mutations in genes coding for sarcomeric proteins. A molecular-genetic etiology can be established in ~60% of cases. Evolutionarily conserved mitochondrial DNA (mtDNA haplogroups are susceptibility factors for HCM. Several polymorphic mtDNA variants are associated with a variety of late-onset degenerative diseases and affect mitochondrial function. We examined the role of private, non-haplogroup associated, mitochondrial variants in the etiology of HCM. In 87 Danish HCM patients, full mtDNA sequencing revealed 446 variants. After elimination of 312 (69.9% non-coding and synonymous variants, a further 109 (24.4% with a global prevalence > 0.1%, three (0.7% haplogroup associated and 19 (2.0% variants with a low predicted in silico likelihood of pathogenicity, three variants: MT-TC: m.5772G>A, MT-TF: m.644A>G, and MT-CYB: m.15024G>A, p.C93Y remained. A detailed analysis of these variants indicated that none of them are likely to cause HCM. In conclusion, private mtDNA mutations are frequent, but they are rarely, if ever, associated with HCM.

  20. A comprehensive characterization of mitochondrial DNA mutations in glioblastoma multiforme.

    Science.gov (United States)

    Vidone, Michele; Clima, Rosanna; Santorsola, Mariangela; Calabrese, Claudia; Girolimetti, Giulia; Kurelac, Ivana; Amato, Laura Benedetta; Iommarini, Luisa; Trevisan, Elisa; Leone, Marco; Soffietti, Riccardo; Morra, Isabella; Faccani, Giuliano; Attimonelli, Marcella; Porcelli, Anna Maria; Gasparre, Giuseppe

    2015-06-01

    Glioblastoma multiforme (GBM) is the most malignant brain cancer in adults, with a poor prognosis, whose molecular stratification still represents a challenge in pathology and clinics. On the other hand, mitochondrial DNA (mtDNA) mutations have been found in most tumors as modifiers of the bioenergetics state, albeit in GBM a characterization of the mtDNA status is lacking to date. Here, a characterization of the burden of mtDNA mutations in GBM samples was performed. First, investigation of tumor-specific vs. non tumor-specific mutations was carried out with the MToolBox bioinformatics pipeline by analyzing 45 matched tumor/blood samples, from whole genome or whole exome sequencing datasets obtained from The Cancer Genome Atlas (TCGA) consortium. Additionally, the entire mtDNA sequence was obtained in a dataset of 104 fresh-frozen GBM samples. Mitochondrial mutations with potential pathogenic interest were prioritized based on heteroplasmic fraction, nucleotide variability, and in silico prediction of pathogenicity. A preliminary biochemical analysis of the activity of mitochondrial respiratory complexes was also performed on fresh-frozen GBM samples. Although a high number of mutations was detected, we report that the large majority of them does not pass the prioritization filters. Therefore, a relatively limited burden of pathogenic mutations is indeed carried by GBM, which did not appear to determine a general impairment of the respiratory chain. This article is part of a Directed Issue entitled: Energy Metabolism Disorders and Therapies.

  1. Mitochondrial DNA damage and animal longevity: insights from comparative studies.

    Science.gov (United States)

    Pamplona, Reinald

    2011-03-02

    Chemical reactions in living cells are under strict enzyme control and conform to a tightly regulated metabolic program. However, uncontrolled and potentially deleterious endogenous reactions occur, even under physiological conditions. Aging, in this chemical context, could be viewed as an entropic process, the result of chemical side reactions that chronically and cumulatively degrade the function of biological systems. Mitochondria are a main source of reactive oxygen species (ROS) and chemical sidereactions in healthy aerobic tissues and are the only known extranuclear cellular organelles in animal cells that contain their own DNA (mtDNA). ROS can modify mtDNA directly at the sugar-phosphate backbone or at the bases, producing many different oxidatively modified purines and pyrimidines, as well as single and double strand breaks and DNA mutations. In this scenario, natural selection tends to decrease the mitochondrial ROS generation, the oxidative damage to mtDNA, and the mitochondrial mutation rate in long-lived species, in agreement with the mitochondrial oxidative stress theory of aging.

  2. Homologous DNA strand exchange activity of the human mitochondrial DNA helicase TWINKLE

    OpenAIRE

    Sen, Doyel; Patel, Gayatri; Smita S Patel

    2016-01-01

    A crucial component of the human mitochondrial DNA replisome is the ring-shaped helicase TWINKLE—a phage T7-gene 4-like protein expressed in the nucleus and localized in the human mitochondria. Our previous studies showed that despite being a helicase, TWINKLE has unique DNA annealing activity. At the time, the implications of DNA annealing by TWINKLE were unclear. Herein, we report that TWINKLE uses DNA annealing function to actively catalyze strand-exchange reaction between the unwinding su...

  3. Mitochondrial DNA analysis of ancient Sampula population in Xinjiang

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    The archaeological site of Sampula cemetery was located about 14 km to the southwest of the Luo County in Xinjiang Khotan, China, belonging to the ancient Yutian kingdom. 14C analysis showed that this cemetery was used from 217 B.C. to 283 A.D.Ancient DNA was analyzed by 364 bp of the mitochondrial DNA hypervariable region Ⅰ (mtDNA HVR-Ⅰ), and by six restriction fragment length polymorphism (RFLP) sites of mtDNA coding region. We successfully extracted and sequenced intact stretches of maternally inherited mtDNA from 13 out of 16 ancient Sampula samples. The analysis of mtDNA haplogroup distribution showed that the ancient Sampula was a complex population with both European and Asian characteristics. Median joining network of U3 sub-haplogroup and multi-dimensional scaling analysis all showed that the ancient Sampula had maternal relationship with Ossetian and Iranian.

  4. Genetic typing of mitochondrial DNA in sheep (Ovis aries )

    Institute of Scientific and Technical Information of China (English)

    赵兴波; 冯继东; 李宁; 王爱华; 吴常信

    2001-01-01

    Using PCR-RFLP and DNA sequencing, this study confirmed that HinfI polymorphism in the ovine mitochondrial COI gene resulted from the T-C substitution at the nucleotide 234 but this mutation did not encode another amino acid, which was actually a synonymous mutation. This single nucleotide polymorphism can be used as gene typing marker for mitochondrial genome in the research of the interactions between mitochondria and nucleus, extranuclear gene effects, and as molecular discriminating marker for embryo or individual in the research area of gene transfer and animal cloning.

  5. Mitochondrial DNA background modulates the assembly kinetics of OXPHOS complexes in a cellular model of mitochondrial disease.

    NARCIS (Netherlands)

    Pello, R.; Martin, M.A.; Carelli, V.; Nijtmans, L.G.J.; Achilli, A.; Pala, M.; Torroni, A.; Gomez-Duran, A.; Ruiz-Pesini, E.; Martinuzzi, A.; Smeitink, J.A.M.; Arenas, J.; Ugalde, C.

    2008-01-01

    Leber's hereditary optic neuropathy (LHON), the most frequent mitochondrial disorder, is mostly due to three mitochondrial DNA (mtDNA) mutations in respiratory chain complex I subunit genes: 3460/ND1, 11778/ND4 and 14484/ND6. Despite considerable clinical evidences, a genetic modifying role of the m

  6. Clonal expansion of early to mid-life mitochondrial DNA point mutations drives mitochondrial dysfunction during human ageing.

    NARCIS (Netherlands)

    Greaves, L.C.; Nooteboom, M.; Elson, J.L.; Tuppen, H.A.; Taylor, G.A.; Commane, D.M.; Arasaradnam, R.P.; Khrapko, K.; Taylor, R.W.; Kirkwood, T.B.; Mathers, J.C.; Turnbull, D.M.

    2014-01-01

    Age-related decline in the integrity of mitochondria is an important contributor to the human ageing process. In a number of ageing stem cell populations, this decline in mitochondrial function is due to clonal expansion of individual mitochondrial DNA (mtDNA) point mutations within single cells. Ho

  7. [The application of mitochondrial genomics to forensic investigations based on human mitochondrial DNA testing].

    Science.gov (United States)

    Skonieczna, Katarzyna; Bednarek, Jarosław; Rogalla, Urszula; Woźniak, Marcin; Gorzkiewicz, Marta; Linkowska, Katarzyna; Duleba, Anna; Sliwka, Karol; Grzybowski, Tomasz

    2012-01-01

    In this study we present two forensic cases where mitochondrial DNA HVS I and HVS II haplotypes of evidentiary hairs match reference samples. Based on the information retrieved from mtDNA coding region of reference material, we selected single nucleotide polymorphisms (SNPs) located outside the HVS I and HVS II regions that could increase the informativeness of mtDNA analysis. The SNPs were typed via SNaPshot or dideoxy sequencing technology. In both cases the SNP results allowed for unambiguous exlusion of the evidence and for determining that reference samples originated from the same person.

  8. Mitochondrial DNA diversity in the African American population.

    Science.gov (United States)

    Johnson, Derek C; Shrestha, Sadeep; Wiener, Howard W; Makowsky, Robert; Kurundkar, Ashish; Wilson, Craig M; Aissani, Brahim

    2015-06-01

    Genetic polymorphism along mitochondrial DNA (mtDNA) defines population-specific signatures called mtDNA haplogroups. Estimation of mtDNA haplogroup distribution may be prone to errors, notably if the study sample is not drawn from a multicenter cohort. Here, we report on mtDNA diversity in a sample of African American individuals (n = 343) enrolled in a multicenter cohort. Sequencing of the hypervariable regions I and II of the D-loop control region showed that the most common mitochondrial variants are 73G, 146C, 150T, 152C, 189G, 16278T, and 16311C. In agreement with the published data, we observed 17 common mtDNA haplogroups: L0, L1, L1b, L1c, L2, L2a, L2b, L2c, L2e, L3, L3b, L3d, L3e, L3f, L3h, L3x, and L4. The most commonly observed haplogroup is L2a (19.8%), followed by L1b (10.2%). Overall, the observed mtDNA haplogroup distribution in our study is similar to those published for the African American and the African populations.

  9. Mitochondrial DNA in the regulation of innate immune responses

    Directory of Open Access Journals (Sweden)

    Chunju Fang

    2015-10-01

    Full Text Available Abstract Mitochondrion is known as the energy factory of the cell, which is also a unique mammalian organelle and considered to be evolved from aerobic prokaryotes more than a billion years ago. Mitochondrial DNA, similar to that of its bacterial ancestor’s, consists of a circular loop and contains significant number of unmethylated DNA as CpG islands. The innate immune system plays an important role in the mammalian immune response. Recent research has demonstrated that mitochondrial DNA (mtDNA activates several innate immune pathways involving TLR9, NLRP3 and STING signaling, which contributes to the signaling platforms and results in effector responses. In addition to facilitating antibacterial immunity and regulating antiviral signaling, mounting evidence suggests that mtDNA contributes to inflammatory diseases following cellular damage and stress. Therefore, in addition to its well-appreciated roles in cellular metabolism and energy production, mtDNA appears to function as a key member in the innate immune system. Here, we highlight the emerging roles of mtDNA in innate immunity.

  10. Heterogeneity of mitochondrial DNA from Saccharomyces carlsbergensis. Denaturation mapping by electron microscopy.

    DEFF Research Database (Denmark)

    Christiansen, Gunna; Christiansen, C; Bak, AL

    1975-01-01

    Electronmicroscopic observation of the denaturation pattern of 130 partially denaturated linear mitochondrial DNA molecules from Saccharomyces carlsbergensis was used to investigate the distribution of AT-rich sequences within the mitochondrial genome. The molecules were observed after heating...... denaturated sequences in the mitochondrial DNA. These sequences which presumably correspond to the very AT-rich regions, known to exist in the yeast mitochondrial DNA, were found at intervals of about 0.5 - 3 mum on the map....

  11. Accumulation of mitochondrial DNA deletions within dopaminergic neurons triggers neuroprotective mechanisms.

    Science.gov (United States)

    Perier, Celine; Bender, Andreas; García-Arumí, Elena; Melià, Ma Jesus; Bové, Jordi; Laub, Christoph; Klopstock, Thomas; Elstner, Matthias; Mounsey, Ross B; Teismann, Peter; Prolla, Tomas; Andreu, Antoni L; Vila, Miquel

    2013-08-01

    Acquired alterations in mitochondrial DNA are believed to play a pathogenic role in Parkinson's disease. In particular, accumulation of mitochondrial DNA deletions has been observed in substantia nigra pars compacta dopaminergic neurons from patients with Parkinson's disease and aged individuals. Also, mutations in mitochondrial DNA polymerase gamma result in multiple mitochondrial DNA deletions that can be associated with levodopa-responsive parkinsonism and severe substantia nigra pars compacta dopaminergic neurodegeneration. However, whether mitochondrial DNA deletions play a causative role in the demise of dopaminergic neurons remains unknown. Here we assessed the potential pathogenic effects of mitochondrial DNA deletions on the dopaminergic nigrostriatal system by using mutant mice possessing a proofreading-deficient form of mitochondrial DNA polymerase gamma (POLGD257A), which results in a time-dependent accumulation of mitochondrial DNA deletions in several tissues, including the brain. In these animals, we assessed the occurrence of mitochondrial DNA deletions within individual substantia nigra pars compacta dopaminergic neurons, by laser capture microdissection and quantitative real-time polymerase chain reaction, and determined the potential deleterious effects of such mitochondrial DNA alterations on mitochondrial function and dopaminergic neuronal integrity, by cytochrome c oxidase histochemistry and quantitative morphology. Nigral dopaminergic neurons from POLGD257A mice accumulate mitochondrial DNA deletions to a similar extent (∼40-60%) as patients with Parkinson's disease and aged individuals. Despite such high levels of mitochondrial DNA deletions, the majority of substantia nigra pars compacta dopaminergic neurons from these animals did not exhibit mitochondrial dysfunction or degeneration. Only a few individual substantia nigra pars compacta neurons appeared as cytochrome c oxidase-negative, which exhibited higher levels of mitochondrial DNA

  12. Mitochondrial DNA copy number - but not a mitochondrial tandem CC to TT transition - is increased in sun-exposed skin.

    Science.gov (United States)

    Gebhard, Daniel; Mahler, Bettina; Matt, Katja; Burger, Katharina; Bergemann, Jörg

    2014-03-01

    Mitochondrial DNA (mtDNA) mutations are causatively associated with photo-ageing and are used as biomarkers of UV exposure. The most prominent mitochondrial mutation is the common deletion (CD), which is induced in many tissues by oxidative stress. More photo-specific mutations might be CC to TT tandem transitions which arise from UV-induced cyclobutane pyrimidine dimers. As nucleotide excision repair is absent in mitochondria, this DNA damage can presumably not be repaired resulting in high mitochondrial mutation levels. Here, we analysed levels of the CD, a mitochondrial and a chromosomal tandem transition in epidermis and dermis from exposed and less UV-exposed skin. We also analysed mtDNA copy number, for which changes as a result of oxidative stress have been described in different experimental settings. Whereas mitochondrial tandem transition levels were surprisingly low with no discernible correlation with UV exposure, mtDNA copy number and CD were significantly increased in UV-exposed samples.

  13. Research Progress on Mitochondrial DNA%线粒体DNA(mtDNA)的研究进展

    Institute of Scientific and Technical Information of China (English)

    王存芳; 曾勇庆; 杜立新; 高秀华

    2001-01-01

    本文概述了mtDNA的基本特征和研究方法;介绍了人及各种畜禽mtDNA的研究现状,并对mtDNA的研究作了展望。%This contribution briefly represented fundamental properties and methods of studies on mitochondrial DNA. The pre sent condition of studies among humans and animals were presented respectively. The studies on mitochondrial DNA were also forcasted.

  14. Functional Recovery of Human Cells Harbouring the Mitochondrial DNA Mutation MERRF A8344G via Peptide-Mediated Mitochondrial Delivery

    Directory of Open Access Journals (Sweden)

    Jui-Chih Chang

    2012-09-01

    Full Text Available We explored the feasibility of mitochondrial therapy using the cell-penetrating peptide Pep-1 to transfer mitochondrial DNA (mtDNA between cells and rescue a cybrid cell model of the mitochondrial disease myoclonic epilepsy with ragged-red fibres (MERRF syndrome. Pep-1-conjugated wild-type mitochondria isolated from parent cybrid cells incorporating a mitochondria-specific tag were used as donors for mitochondrial delivery into MERRF cybrid cells (MitoB2 and mtDNA-depleted Rho-zero cells (Mitoρ°. Forty-eight hours later, translocation of Pep-1-labelled mitochondria into the mitochondrial regions of MitoB2 and Mitoρ° host cells was observed (delivery efficiencies of 77.48 and 82.96%, respectively. These internalized mitochondria were maintained for at least 15 days in both cell types and were accompanied by mitochondrial function recovery and cell survival by preventing mitochondria-dependent cell death. Mitochondrial homeostasis analyses showed that peptide-mediated mitochondrial delivery (PMD also increased mitochondrial biogenesis in both cell types, but through distinct regulatory pathways involving mitochondrial dynamics. Dramatic decreases in mitofusin-2 (MFN2 and dynamin-related protein 1/fission 1 were observed in MitoB2 cells, while Mitoρ° cells showed a significant increase in optic atrophy 1 and MFN2. These findings suggest that PMD can be used as a potential therapeutic intervention for mitochondrial disorders.

  15. Patterns of mitochondrial DNA instability in Brassica campestris cultured cells.

    Science.gov (United States)

    Shirzadegan, M; Palmer, J D; Christey, M; Earle, E D

    1991-01-01

    We previously showed that the mitochondrial DNA (mtDNA) of a Brassica campestris callus culture had undergone extensive rearrangements (i.e. large inversions and a duplication) relative to DNA of the control plant [54]. In this study we observed that after continued growth, the mtDNA of this culture continues to change, with rearranged forms amplifying and diminishing to varying proportions. Strikingly similar changes were detected in the mtDNA profiles of a variety of other long- and short-term callus and cell suspension lines. However, the proportions of parental ('unrearranged') and novel ('rearranged') forms varied in different cultured cell mtDNAs. To address the source of this heterogeneity, we compared the mtDNA organization of 28 individual plants from the parental seed stock. With the exception of one plant containing high levels of a novel plasmid-like mtDNA molecule, no significant variation was detected among individual plants and therefore source plant variation is unlikely to have contributed to the diversity of mitochondrial genomes observed in cultured cells. The source of this culture-induced heterogeneity was also investigated in 16 clones derived from single protoplasts. A mixed population of unrearranged and rearranged mtDNA molecules was apparent in each protoclone, suggesting that the observed heterogeneity in various cultures might reflect the genomic composition of each individual cell; however, the induction of an intercellular heterogeneity subsequent to the protoplast isolation was not tested and therefore cannot be ruled out. The results of this study support our earlier model that the rapid structural alteration of B. campestris mtDNA in vitro results from preferential amplification and reassortment of minor pre-existing forms of the genome rather than de novo rearrangement. Infrequent recombination between short dispersed repeated elements is proposed as the underlying mechanism for the formation of these minor mtDNA molecules.

  16. Sequencing of mitochondrial HV1 and HV2 DNA with length heteroplasmy

    DEFF Research Database (Denmark)

    Rasmussen, E. Michael; Eriksen, Birthe; Larsen, Hans Jakob

    2003-01-01

    This study presents a fast method for sequencing the poly C/G regions in HV1 and HV2 in the mitochondrial DNA (mtDNA)......This study presents a fast method for sequencing the poly C/G regions in HV1 and HV2 in the mitochondrial DNA (mtDNA)...

  17. qPCR-based mitochondrial DNA quantification: Influence of template DNA fragmentation on accuracy

    Energy Technology Data Exchange (ETDEWEB)

    Jackson, Christopher B., E-mail: Christopher.jackson@insel.ch [Division of Human Genetics, Departements of Pediatrics and Clinical Research, Inselspital, University of Berne, Freiburgstrasse, CH-3010 Berne (Switzerland); Gallati, Sabina, E-mail: sabina.gallati@insel.ch [Division of Human Genetics, Departements of Pediatrics and Clinical Research, Inselspital, University of Berne, Freiburgstrasse, CH-3010 Berne (Switzerland); Schaller, Andre, E-mail: andre.schaller@insel.ch [Division of Human Genetics, Departements of Pediatrics and Clinical Research, Inselspital, University of Berne, Freiburgstrasse, CH-3010 Berne (Switzerland)

    2012-07-06

    Highlights: Black-Right-Pointing-Pointer Serial qPCR accurately determines fragmentation state of any given DNA sample. Black-Right-Pointing-Pointer Serial qPCR demonstrates different preservation of the nuclear and mitochondrial genome. Black-Right-Pointing-Pointer Serial qPCR provides a diagnostic tool to validate the integrity of bioptic material. Black-Right-Pointing-Pointer Serial qPCR excludes degradation-induced erroneous quantification. -- Abstract: Real-time PCR (qPCR) is the method of choice for quantification of mitochondrial DNA (mtDNA) by relative comparison of a nuclear to a mitochondrial locus. Quantitative abnormal mtDNA content is indicative of mitochondrial disorders and mostly confines in a tissue-specific manner. Thus handling of degradation-prone bioptic material is inevitable. We established a serial qPCR assay based on increasing amplicon size to measure degradation status of any DNA sample. Using this approach we can exclude erroneous mtDNA quantification due to degraded samples (e.g. long post-exicision time, autolytic processus, freeze-thaw cycles) and ensure abnormal DNA content measurements (e.g. depletion) in non-degraded patient material. By preparation of degraded DNA under controlled conditions using sonification and DNaseI digestion we show that erroneous quantification is due to the different preservation qualities of the nuclear and the mitochondrial genome. This disparate degradation of the two genomes results in over- or underestimation of mtDNA copy number in degraded samples. Moreover, as analysis of defined archival tissue would allow to precise the molecular pathomechanism of mitochondrial disorders presenting with abnormal mtDNA content, we compared fresh frozen (FF) with formalin-fixed paraffin-embedded (FFPE) skeletal muscle tissue of the same sample. By extrapolation of measured decay constants for nuclear DNA ({lambda}{sub nDNA}) and mtDNA ({lambda}{sub mtDNA}) we present an approach to possibly correct measurements in

  18. Transcription profiles of mitochondrial genes correlate with mitochondrial DNA haplotypes in a natural population of Silene vulgaris

    Directory of Open Access Journals (Sweden)

    Olson Matthew S

    2010-01-01

    Full Text Available Abstract Background Although rapid changes in copy number and gene order are common within plant mitochondrial genomes, associated patterns of gene transcription are underinvestigated. Previous studies have shown that the gynodioecious plant species Silene vulgaris exhibits high mitochondrial diversity and occasional paternal inheritance of mitochondrial markers. Here we address whether variation in DNA molecular markers is correlated with variation in transcription of mitochondrial genes in S. vulgaris collected from natural populations. Results We analyzed RFLP variation in two mitochondrial genes, cox1 and atp1, in offspring of ten plants from a natural population of S. vulgaris in Central Europe. We also investigated transcription profiles of the atp1 and cox1 genes. Most DNA haplotypes and transcription profiles were maternally inherited; for these, transcription profiles were associated with specific mitochondrial DNA haplotypes. One individual exhibited a pattern consistent with paternal inheritance of mitochondrial DNA; this individual exhibited a transcription profile suggestive of paternal but inconsistent with maternal inheritance. We found no associations between gender and transcript profiles. Conclusions Specific transcription profiles of mitochondrial genes were associated with specific mitochondrial DNA haplotypes in a natural population of a gynodioecious species S. vulgaris. Our findings suggest the potential for a causal association between rearrangements in the plant mt genome and transcription product variation.

  19. Infantile encephalopathy and defective mitochondrial DNA translation in patients with mutations of mitochondrial elongation factors EFG1 and EFTu.

    Science.gov (United States)

    Valente, Lucia; Tiranti, Valeria; Marsano, Rene Massimiliano; Malfatti, Edoardo; Fernandez-Vizarra, Erika; Donnini, Claudia; Mereghetti, Paolo; De Gioia, Luca; Burlina, Alberto; Castellan, Claudio; Comi, Giacomo P; Savasta, Salvatore; Ferrero, Iliana; Zeviani, Massimo

    2007-01-01

    Mitochondrial protein translation is a complex process performed within mitochondria by an apparatus composed of mitochondrial DNA (mtDNA)-encoded RNAs and nuclear DNA-encoded proteins. Although the latter by far outnumber the former, the vast majority of mitochondrial translation defects in humans have been associated with mutations in RNA-encoding mtDNA genes, whereas mutations in protein-encoding nuclear genes have been identified in a handful of cases. Genetic investigation involving patients with defective mitochondrial translation led us to the discovery of novel mutations in the mitochondrial elongation factor G1 (EFG1) in one affected baby and, for the first time, in the mitochondrial elongation factor Tu (EFTu) in another one. Both patients were affected by severe lactic acidosis and rapidly progressive, fatal encephalopathy. The EFG1-mutant patient had early-onset Leigh syndrome, whereas the EFTu-mutant patient had severe infantile macrocystic leukodystrophy with micropolygyria. Structural modeling enabled us to make predictions about the effects of the mutations at the molecular level. Yeast and mammalian cell systems proved the pathogenic role of the mutant alleles by functional complementation in vivo. Nuclear-gene abnormalities causing mitochondrial translation defects represent a new, potentially broad field of mitochondrial medicine. Investigation of these defects is important to expand the molecular characterization of mitochondrial disorders and also may contribute to the elucidation of the complex control mechanisms, which regulate this fundamental pathway of mtDNA homeostasis.

  20. Reverse genetic studies of mitochondrial DNA-based diseases using a mouse model

    OpenAIRE

    Nakada, Kazuto; Sato, Akitsugu; Hayashi, Jun-Ichi

    2008-01-01

    In the situation that it would not be able to produce model animals for mitochondrial diseases caused by mitochondrial DNA (mtDNA) with pathogenic mutations, we succeeded in generating mice with pathogenic deletion mutant mtDNA (ΔmtDNA), named “mito-mice”, by direct introduction of mitochondria with ΔmtDNA into mouse zygotes. In the mito-mice, accumulation of ΔmtDNA induced mitochondrial respiration defects in various tissues, resulting in mitochondrial disease phenotypes, such as low body we...

  1. Mitochondrial DNA deletions are associated with non-B DNA conformations

    Science.gov (United States)

    Damas, Joana; Carneiro, João; Gonçalves, Joana; Stewart, James B.; Samuels, David C.; Amorim, António; Pereira, Filipe

    2012-01-01

    Mitochondrial DNA (mtDNA) deletions are a primary cause of mitochondrial disease and are believed to contribute to the aging process and to various neurodegenerative diseases. Despite strong observational and experimental evidence, the molecular basis of the deletion process remains obscure. In this study, we test the hypothesis that the primary cause of mtDNA vulnerability to breakage resides in the formation of non-B DNA conformations, namely hairpin, cruciform and cloverleaf-like elements. Using the largest database of human mtDNA deletions built thus far (753 different cases), we show that site-specific breakage hotspots exist in the mtDNA. Furthermore, we discover that the most frequent deletion breakpoints occur within or near predicted structures, a result that is supported by data from transgenic mice with mitochondrial disease. There is also a significant association between the folding energy of an mtDNA region and the number of breakpoints that it harbours. In particular, two clusters of hairpins (near the D-loop 3′-terminus and the L-strand origin of replication) are hotspots for mtDNA breakage. Consistent with our hypothesis, the highest number of 5′- and 3′-breakpoints per base is found in the highly structured tRNA genes. Overall, the data presented in this study suggest that non-B DNA conformations are a key element of the mtDNA deletion process. PMID:22661583

  2. The role of mitochondrial DNA mutations in aging and sarcopenia: implications for the mitochondrial vicious cycle theory of aging.

    Science.gov (United States)

    Hiona, Asimina; Leeuwenburgh, Christiaan

    2008-01-01

    Aging is associated with a progressive loss of skeletal muscle mass and strength and the mechanisms mediating these effects likely involve mitochondrial DNA (mtDNA) mutations, mitochondrial dysfunction and the activation of mitochondrial-mediated apoptosis. Because the mitochondrial genome is densely packed and close to the main generator of reactive oxygen species (ROS) in the cell, the electron transport chain (ETC), an important role for mtDNA mutations in aging has been proposed. Point mutations and deletions in mtDNA accumulate with age in a wide variety of tissues in mammals, including humans, and often coincide with significant tissue dysfunction. Here, we examine the evidence supporting a causative role for mtDNA mutations in aging and sarcopenia. We review experimental outcomes showing that mtDNA mutations, leading to mitochondrial dysfunction and possibly apoptosis, are causal to the process of sarcopenia. Moreover, we critically discuss and dispute an important part of the mitochondrial 'vicious cycle' theory of aging which proposes that accumulation of mtDNA mutations may lead to an enhanced mitochondrial ROS production and ever increasing oxidative stress which ultimately leads to tissue deterioration and aging. Potential mechanism(s) by which mtDNA mutations may mediate their pathological consequences in skeletal muscle are also discussed.

  3. Analysis of Translesion DNA Synthesis by the Mitochondrial DNA Polymerase γ.

    Science.gov (United States)

    Copeland, William C; Kasiviswanathan, Rajesh; Longley, Matthew J

    2016-01-01

    Mitochondrial DNA is replicated by the nuclear-encoded DNA polymerase γ (pol γ) which is composed of a single 140 kDa catalytic subunit and a dimeric 55 kDa accessory subunit. Mitochondrial DNA is vulnerable to various forms of damage, including several types of oxidative lesions, UV-induced photoproducts, chemical adducts from environmental sources, as well as alkylation and inter-strand cross-links from chemotherapy agents. Although many of these lesions block DNA replication, pol γ can bypass some lesions by nucleotide incorporation opposite a template lesion and further extension of the DNA primer past the lesion. This process of translesion synthesis (TLS) by pol γ can occur in either an error-free or an error-prone manner. Assessment of TLS requires extensive analysis of oligonucleotide substrates and replication products by denaturing polyacrylamide sequencing gels. This chapter presents protocols for the analysis of translesion DNA synthesis.

  4. Characterization of a Dairy Gyr herd with respect to its mitochondrial DNA (mt DNA origin

    Directory of Open Access Journals (Sweden)

    Anibal Eugênio Vercesi Filho

    2010-01-01

    Full Text Available The Zebu breeds were introduced in Brazil mainly in the last century by imports from the Indian subcontinent. When the Zebu cattle arrived, the national herd suffered a significative change by backcrossing the national cows of taurine origin with Zebu sires. These processes created a polymorphism in the mitochondrial DNA (mtDNA in the Zebu animals with are in a major part derived from backcrossing and sharing mtDNA of taurine origin. To verify the maternal origin of cows belonging to the Dairy Gyr herd of APTA, Mococa 60 females were analyzed and 33 presented mtDNA from Bos taurus origin and 27 presented mtDNA from Bos indicus origin. None of these animals presented patterns of both mtDNA origins, indicating absence of heteroplasmy for these mitochondrial genotypes.

  5. Analysis of Translesion DNA Synthesis by the Mitochondrial DNA Polymerase γ

    Science.gov (United States)

    Copeland, William C.; Kasiviswanathan, Rajesh; Longley, Matthew J.

    2016-01-01

    Summary Mitochondrial DNA is replicated by the nuclear encoded DNA polymerase γ (pol γ) which is composed of a single 140 kDa catalytic subunit and a dimeric 55 kDa accessory subunit. Mitochondrial DNA is vulnerable to various forms of damage, including several types of oxidative lesions, UV-induced photoproducts, chemical adducts from environmental sources, as well as alkylation and inter-strand crosslinks from chemotherapy agents. Although many of these lesions block DNA replication, Pol γ can bypass some lesions by nucleotide incorporation opposite a template lesion and further extension of the DNA primer past the lesion. This process of translesion synthesis (TLS) by Pol γ can occur in either an error-free or an error-prone manner. Assessment of TLS requires extensive analysis of oligonucleotide substrates and replication products by denaturing polyacrylamide sequencing gels. This chapter presents protocols for the analysis of translesion DNA synthesis. PMID:26530671

  6. Complete sequence of the mitochondrial genome of Tetrahymena pyriformis and comparison with Paramecium aurelia mitochondrial DNA.

    Science.gov (United States)

    Burger, G; Zhu, Y; Littlejohn, T G; Greenwood, S J; Schnare, M N; Lang, B F; Gray, M W

    2000-03-24

    We report the complete nucleotide sequence of the Tetrahymena pyriformis mitochondrial genome and a comparison of its gene content and organization with that of Paramecium aurelia mtDNA. T. pyriformis mtDNA is a linear molecule of 47,172 bp (78.7 % A+T) excluding telomeric sequences (identical tandem repeats of 31 bp at each end of the genome). In addition to genes encoding the previously described bipartite small and large subunit rRNAs, the T. pyriformis mitochondrial genome contains 21 protein-coding genes that are clearly homologous to genes of defined function in other mtDNAs, including one (yejR) that specifies a component of a cytochrome c biogenesis pathway. As well, T. pyriformis mtDNA contains 22 open reading frames of unknown function larger than 60 codons, potentially specifying proteins ranging in size from 74 to 1386 amino acid residues. A total of 13 of these open reading frames ("ciliate-specific") are found in P. aurelia mtDNA, whereas the remaining nine appear to be unique to T. pyriformis; however, of the latter, five are positionally equivalent and of similar size in the two ciliate mitochondrial genomes, suggesting they may also be homologous, even though this is not evident from sequence comparisons. Only eight tRNA genes encoding seven distinct tRNAs are found in T. pyriformis mtDNA, formally confirming a long-standing proposal that most T. pyriformis mitochondrial tRNAs are nucleus-encoded species imported from the cytosol. Atypical features of mitochondrial gene organization and expression in T. pyriformis mtDNA include split and rearranged large subunit rRNA genes, as well as a split nad1 gene (encoding subunit 1 of NADH dehydrogenase of respiratory complex I) whose two segments are located on and transcribed from opposite strands, as is also the case in P. aurelia. Gene content and arrangement are very similar in T. pyriformis and P. aurelia mtDNAs, the two differing by a limited number of duplication, inversion and rearrangement events

  7. Mitochondrial DNA mutations in etiopathogenesis of male infertility

    Directory of Open Access Journals (Sweden)

    Monis Bilal Shamsi

    2008-01-01

    Conclusion: In the context of male infertility, mt mutations, generation of reactive oxygen species and lowered antioxidant capacity are interlinked and constitute a unified pathogenic molecular mechanism. In the era of assisted reproduction technique (ART, it is very important to distinguish between mutations in nuclear and mitochondrial genomes in sperm, as mtDNA mutations are better diagnostic and prognostic markers in infertile men opting for ART.

  8. Reduction in mitochondrial DNA copy number in peripheral leukocytes after onset of Huntington's disease

    DEFF Research Database (Denmark)

    Petersen, Maria Hvidberg; Budtz-Jørgensen, Esben; Sørensen, Sven Asger;

    2014-01-01

    to the investigation of the mitochondrial DNA (mtDNA) copy number relative to nuclear DNA (nDNA) in leukocytes from carriers of the HD mutation compared to healthy individuals. We found significantly reduced mtDNA/nDNA in HD mutation carriers compared to controls. A longitudinal study of archive DNA sample pairs from...

  9. Adult mitochondrial DNA depletion syndrome with mild manifestations

    Directory of Open Access Journals (Sweden)

    Josef Finsterer

    2013-06-01

    Full Text Available Mitochondrial DNA depletion syndrome (MDS is usually a severe disorder of infancy or childhood, due to a reduced copy number of mtDNA molecules. MDS with only mild, non-specific clinical manifestations and onset in adulthood has not been reported. A 47-year-old Caucasian female with short stature and a history of migraine, endometriosis, Crohn’s disease, C-cell carcinoma of the thyroid gland, and a family history positive for mitochondrial disorder (2 sisters, aunt, niece, developed day-time sleepiness, exercise intolerance, and myalgias in the lower-limb muscles since age 46y. She slept 9-10 hours during the night and 2 hours after lunch daily. Clinical exam revealed sore neck muscles, bilateral ptosis, and reduced Achilles tendon reflexes exclusively. Blood tests revealed hyperlipidemia exclusively. Nerve conduction studies, needle electromyography, and cerebral and spinal magnetic resonance imaging were non-informative. Muscle biopsy revealed detached lobulated fibers with subsarcolemmal accentuation of the NADH and SDH staining. Real-time polymerase chain reaction revealed depletion of the mtDNA down to 9% of normal. MDS may be associated with a mild phenotype in adults and may not significantly progress during the first year after onset. In an adult with hypersomnia, severe tiredness, exercise intolerance, and a family history positive for mitochondrial disorder, a MDS should be considered.

  10. Association Between Mitochondrial DNA Haplogroup Variation and Autism Spectrum Disorders.

    Science.gov (United States)

    Chalkia, Dimitra; Singh, Larry N; Leipzig, Jeremy; Lvova, Maria; Derbeneva, Olga; Lakatos, Anita; Hadley, Dexter; Hakonarson, Hakon; Wallace, Douglas C

    2017-08-23

    Autism spectrum disorders (ASD) are characterized by impairments in social interaction, communication, and repetitive or restrictive behavior. Although multiple physiologic and biochemical studies have reported defects in mitochondrial oxidative phosphorylation in patients with ASD, the role of mitochondrial DNA (mtDNA) variation has remained relatively unexplored. To assess what impact mitochondrial lineages encompassing ancient mtDNA functional polymorphisms, termed haplogroups, have on ASD risk. In this cohort study, individuals with autism and their families were studied using the Autism Genetic Resource Exchange cohort genome-wide association studies data previously generated at the Children's Hospital of Philadelphia. From October 2010 to January 2017, we analyzed the data and used the mtDNA single-nucleotide polymorphisms interrogated by the Illumina HumanHap 550 chip to determine the mtDNA haplogroups of the individuals. Taking into account the familial structure of the Autism Genetic Resource Exchange data, we then determined whether the mtDNA haplogroups correlate with ASD risk. Odds ratios of mitochondrial haplogroup as predictors of ASD risk. Of 1624 patients with autism included in this study, 1299 were boys (80%) and 325 were girls (20%). Families in the Autism Genetic Resource Exchange collection (933 families, encompassing 4041 individuals: 1624 patients with ASD and 2417 healthy parents and siblings) had been previously recruited in the United States with no restrictions on age, sex, race/ethnicity, or socioeconomic status. Relative to the most common European haplogroup HHV, European haplogroups I, J, K, O-X, T, and U were associated with increased risk of ASD, as were Asian and Native American haplogroups A and M, with odds ratios ranging from 1.55 (95% CI, 1.16-2.06) to 2.18 (95% CI, 1.59-3) (adjusted P < .04). Hence, mtDNA haplogroup variation is an important risk factor for ASD. Because haplogroups I, J, K, O-X, T, and U encompass 55% of the

  11. Mitochondrial DNA damage associated with lipid peroxidation of the mitochondrial membrane induced by Fe2+-citrate

    OpenAIRE

    2006-01-01

    Iron imbalance/accumulation has been implicated in oxidative injury associated with many degenerative diseases such as hereditary hemochromatosis, beta-thalassemia, and Friedreich's ataxia. Mitochondria are particularly sensitive to iron-induced oxidative stress - high loads of iron cause extensive lipid peroxidation and membrane permeabilization in isolated mitochondria. Here we detected and characterized mitochondrial DNA damage in isolated rat liver mitochondria exposed to a Fe2+-citrate c...

  12. Fatal dilated cardiomyopathy associated with a mitochondrial DNA deletion.

    Science.gov (United States)

    Moslemi, A R; Selimovic, N; Bergh, C H; Oldfors, A

    2000-01-01

    A 27-year-old man was admitted to hospital because of severe cardiac failure. Investigation revealed dilated cardiomyopathy with a left ventricular ejection fraction of 15-20%. During adolescence the patient had been investigated for growth retardation and he also had progressive external ophthalmoplegia. There had been no symptoms of cardiac disease until 2 weeks before admittance. An endomyocardial biopsy showed cardiomyocytes deficient in cytochrome c oxidase (COX) in a mosaic pattern. A skeletal muscle biopsy showed mitochondrial myopathy with COX-deficient ragged-red fibers. Molecular genetic analysis revealed a heteroplasmic, 3.8-kb, mitochondrial DNA (mtDNA) deletion in heart and muscle. PCR-based quantification of the proportion of mtDNA with deletion showed 47% mutated mtDNA in the myocardial biopsy and 68% in muscle. In spite of treatment, the condition deteriorated and the patient died 5 days after admittance. This case demonstrates that mtDNA deletions may occasionally be the cause of severe dilated cardiomyopathy, and that morphological and molecular genetic diagnosis may be obtained by endomyocardial biopsy. Copyright 2000 S. Karger AG, Basel.

  13. Mitochondrial DNA Variation in Southeastern Pre-Columbian Canids.

    Science.gov (United States)

    Brzeski, Kristin E; DeBiasse, Melissa B; Rabon, David R; Chamberlain, Michael J; Taylor, Sabrina S

    2016-05-01

    The taxonomic status of the red wolf (Canis rufus) is heavily debated, but could be clarified by examining historic specimens from the southeastern United States. We analyzed mitochondrial DNA (mtDNA) from 3 ancient (350-1900 year olds) putative wolf samples excavated from middens and sinkholes within the historic red wolf range. We detected 3 unique mtDNA haplotypes, which grouped with the coyote mtDNA clade, suggesting that the canids inhabiting southeastern North America prior to human colonization from Europe were either coyotes, which would vastly expand historic coyote distributions, an ancient coyote-wolf hybrid, or a North American evolved red wolf lineage related to coyotes. Should the red wolf prove to be a distinct species, our results support the idea of either an ancient hybrid origin for red wolves or a shared common ancestor between coyotes and red wolves.

  14. Mitochondrial glutathione oxidation correlates with age-associated oxidative damage to mitochondrial DNA.

    Science.gov (United States)

    de la Asuncion, J G; Millan, A; Pla, R; Bruseghini, L; Esteras, A; Pallardo, F V; Sastre, J; Viña, J

    1996-02-01

    Mitochondria may be primary targets of free radical damage associated with aging. We have found that mitochondrial glutathione is markedly oxidized with aging in rats and mice. The oxidized to reduced glutathione ratio rises with aging in the liver, kidney, and brain. The magnitude of these changes is much higher than that previously found in whole cells of any species previously studied. In the liver, this ratio (expressing GSSG as a percent of GSH) changed from 0.77 +/- 0.19% (n=5) in young rats to 2.47 +/- 1.25% (n=5) in old ones, i.e., 320% of the controls. In the brain and kidney, values for old rats were, respectively, 600 and 540% higher than those of young rats. A marked oxidation of mitochondrial glutathione also occurred in mice. Aging also caused an increase in 8-oxo-7,8-dihydro-2'-deoxyguanosine levels in mtDNA in rats and mice. Oral antioxidant administration protected against both glutathione oxidation and mtDNA damage in rats and mice. Finally, we have found a direct relationship between mtDNA damage and mitochondrial glutathione oxidation. This occurs both in rats (r=0.95) and in mice (r=0.98). This relationship, which has been observed for the first time in these studies, underscores the role of glutathione in the protection against free radical damage that occurs upon aging.

  15. Infantile Encephalopathy and Defective Mitochondrial DNA Translation in Patients with Mutations of Mitochondrial Elongation Factors EFG1 and EFTu

    Science.gov (United States)

    Valente, Lucia; Tiranti, Valeria; Marsano, René Massimiliano; Malfatti, Edoardo; Fernandez-Vizarra, Erika; Donnini, Claudia; Mereghetti, Paolo; De Gioia, Luca; Burlina, Alberto; Castellan, Claudio; Comi, Giacomo P.; Savasta, Salvatore; Ferrero, Iliana; Zeviani, Massimo

    2007-01-01

    Mitochondrial protein translation is a complex process performed within mitochondria by an apparatus composed of mitochondrial DNA (mtDNA)–encoded RNAs and nuclear DNA–encoded proteins. Although the latter by far outnumber the former, the vast majority of mitochondrial translation defects in humans have been associated with mutations in RNA-encoding mtDNA genes, whereas mutations in protein-encoding nuclear genes have been identified in a handful of cases. Genetic investigation involving patients with defective mitochondrial translation led us to the discovery of novel mutations in the mitochondrial elongation factor G1 (EFG1) in one affected baby and, for the first time, in the mitochondrial elongation factor Tu (EFTu) in another one. Both patients were affected by severe lactic acidosis and rapidly progressive, fatal encephalopathy. The EFG1-mutant patient had early-onset Leigh syndrome, whereas the EFTu-mutant patient had severe infantile macrocystic leukodystrophy with micropolygyria. Structural modeling enabled us to make predictions about the effects of the mutations at the molecular level. Yeast and mammalian cell systems proved the pathogenic role of the mutant alleles by functional complementation in vivo. Nuclear-gene abnormalities causing mitochondrial translation defects represent a new, potentially broad field of mitochondrial medicine. Investigation of these defects is important to expand the molecular characterization of mitochondrial disorders and also may contribute to the elucidation of the complex control mechanisms, which regulate this fundamental pathway of mtDNA homeostasis. PMID:17160893

  16. Genetics Home Reference: MPV17-related hepatocerebral mitochondrial DNA depletion syndrome

    Science.gov (United States)

    ... on PubMed Central Spinazzola A, Santer R, Akman OH, Tsiakas K, Schaefer H, Ding X, Karadimas CL, ... Zeviani M. MPV17 encodes an inner mitochondrial membrane protein and is mutated in infantile hepatic mitochondrial DNA ...

  17. An increase in mitochondrial DNA promotes nuclear DNA replication in yeast.

    Directory of Open Access Journals (Sweden)

    Heidi M Blank

    2008-04-01

    Full Text Available Coordination between cellular metabolism and DNA replication determines when cells initiate division. It has been assumed that metabolism only plays a permissive role in cell division. While blocking metabolism arrests cell division, it is not known whether an up-regulation of metabolic reactions accelerates cell cycle transitions. Here, we show that increasing the amount of mitochondrial DNA accelerates overall cell proliferation and promotes nuclear DNA replication, in a nutrient-dependent manner. The Sir2p NAD+-dependent de-acetylase antagonizes this mitochondrial role. We found that cells with increased mitochondrial DNA have reduced Sir2p levels bound at origins of DNA replication in the nucleus, accompanied with increased levels of K9, K14-acetylated histone H3 at those origins. Our results demonstrate an active role of mitochondrial processes in the control of cell division. They also suggest that cellular metabolism may impact on chromatin modifications to regulate the activity of origins of DNA replication.

  18. Primer retention owing to the absence of RNase H1 is catastrophic for mitochondrial DNA replication.

    Science.gov (United States)

    Holmes, J Bradley; Akman, Gokhan; Wood, Stuart R; Sakhuja, Kiran; Cerritelli, Susana M; Moss, Chloe; Bowmaker, Mark R; Jacobs, Howard T; Crouch, Robert J; Holt, Ian J

    2015-07-28

    Encoding ribonuclease H1 (RNase H1) degrades RNA hybridized to DNA, and its function is essential for mitochondrial DNA maintenance in the developing mouse. Here we define the role of RNase H1 in mitochondrial DNA replication. Analysis of replicating mitochondrial DNA in embryonic fibroblasts lacking RNase H1 reveals retention of three primers in the major noncoding region (NCR) and one at the prominent lagging-strand initiation site termed Ori-L. Primer retention does not lead immediately to depletion, as the persistent RNA is fully incorporated in mitochondrial DNA. However, the retained primers present an obstacle to the mitochondrial DNA polymerase γ in subsequent rounds of replication and lead to the catastrophic generation of a double-strand break at the origin when the resulting gapped molecules are copied. Hence, the essential role of RNase H1 in mitochondrial DNA replication is the removal of primers at the origin of replication.

  19. Mitochondrial DNA copy number in peripheral blood and melanoma risk.

    Directory of Open Access Journals (Sweden)

    Jie Shen

    Full Text Available Mitochondrial DNA (mtDNA copy number in peripheral blood has been suggested as risk modifier in various types of cancer. However, its influence on melanoma risk is unclear. We evaluated the association between mtDNA copy number in peripheral blood and melanoma risk in 500 melanoma cases and 500 healthy controls from an ongoing melanoma study. The mtDNA copy number was measured using real-time polymerase chain reaction. Overall, mean mtDNA copy number was significantly higher in cases than in controls (1.15 vs 0.99, P<0.001. Increased mtDNA copy number was associated with a 1.45-fold increased risk of melanoma (95% confidence interval: 1.12-1.97. Significant joint effects between mtDNA copy number and variables related to pigmentation and history of sunlight exposure were observed. This study supports an association between increased mtDNA copy number and melanoma risk that is independent on the known melanoma risk factors (pigmentation and history of sunlight exposure.

  20. Introgression of mitochondrial DNA among Myodes voles: consequences for energetics?

    Directory of Open Access Journals (Sweden)

    Boratyński Zbyszek

    2011-12-01

    Full Text Available Abstract Background Introgression of mitochondrial DNA (mtDNA is among the most frequently described cases of reticulate evolution. The tendency of mtDNA to cross interspecific barriers is somewhat counter-intuitive considering the key function of enzymes that it encodes in the oxidative-phosphorylation process, which could give rise to hybrid dysfunction. How mtDNA reticulation affects the evolution of metabolic functions is, however, uncertain. Here we investigated how morpho-physiological traits vary in natural populations of a common rodent (the bank vole, Myodes glareolus and whether this variation could be associated with mtDNA introgression. First, we confirmed that M. glareolus harbour mtDNA introgressed from M. rutilus by analyzing mtDNA (cytochrome b, 954 bp and nuclear DNA (four markers; 2333 bp in total sequence variation and reconstructing loci phylogenies among six natural populations in Finland. We then studied geographic variation in body size and basal metabolic rate (BMR among the populations of M. glareolus and tested its relationship with mtDNA type. Results Myodes glareolus and its arctic neighbour, M. rutilus, are reciprocally monophyletic at the analyzed nuclear DNA loci. In contrast, the two northernmost populations of M. glareolus have a fixed mitotype that is shared with M. rutilus, likely due to introgressive hybridization. The analyses of phenotypic traits revealed that the body mass and whole-body, but not mass corrected, BMR are significantly reduced in M. glareolus females from northern Finland that also have the introgressed mitotype. Restricting the analysis to the single population where the mitotypes coexist, the association of mtDNA type with whole-body BMR remained but those with mass corrected BMR and body mass did not. Mitochondrial sequence variation in the introgressed haplotypes is compatible with demographic growth of the populations, but may also be a result of positive selection. Conclusion Our

  1. Mitochondrial DNA sequence variation in the Anatolian Peninsula (Turkey)

    Indian Academy of Sciences (India)

    Hatice Mergen; Reyhan Öner; Cihan Öner

    2004-04-01

    Throughout human history, the region known today as the Anatolian peninsula (Turkey) has served as a junction connecting the Middle East, Europe and Central Asia, and, thus, has been subject to major population movements. The present study is undertaken to obtain information about the distribution of the existing mitochondrial D-loop sequence variations in the Turkish population of Anatolia. A few studies have previously reported mtDNA sequences in Turks. We attempted to extend these results by analysing a cohort that is not only larger, but also more representative of the Turkish population living in Anatolia. In order to obtain a descriptive picture for the phylogenetic distribution of the mitochondrial genome within Turkey, we analysed mitochondrial D-loop region sequence variations in 75 individuals from different parts of Anatolia by direct sequencing. Analysis of the two hypervariable segments within the noncoding region of the mitochondrial genome revealed the existence of 81 nucleotide mutations at 79 sites. The neighbour-joining tree of Kimura’s distance matrix has revealed the presence of six main clusters, of which H and U are the most common. The data obtained are also compared with several European and Turkic Central Asian populations.

  2. Two potential Petunia hybrida mitochondrial DNA replication origins show structural and in vitro functional homology with the animal mitochondrial DNA heavy and light strand replication origins

    NARCIS (Netherlands)

    Haas, Jan M. de; Hille, Jacques; Kors, Frank; Meer, Bert van der; Kool, Ad J.; Folkerts, Otto; Nijkamp, H. John J.

    1991-01-01

    Four Petunia hybrida mitochondrial (mt) DNA fragments have been isolated, sequenced, localized on the physical map and analyzed for their ability to initiate specific DNA synthesis. When all four mtDNA fragments were tested as templates in an in vitro DNA synthesizing lysate system, developed from

  3. Two potential Petunia hybrida mitochondrial DNA replication origins show structural and in vitro functional homology with the animal mitochondrial DNA heavy and light strand replication origins

    NARCIS (Netherlands)

    Haas, Jan M. de; Hille, Jacques; Kors, Frank; Meer, Bert van der; Kool, Ad J.; Folkerts, Otto; Nijkamp, H. John J.

    1991-01-01

    Four Petunia hybrida mitochondrial (mt) DNA fragments have been isolated, sequenced, localized on the physical map and analyzed for their ability to initiate specific DNA synthesis. When all four mtDNA fragments were tested as templates in an in vitro DNA synthesizing lysate system, developed from p

  4. Mitochondrial DNA phylogeography of least cisco Coregonus sardinella in Alaska.

    Science.gov (United States)

    Padula, V M; Causey, D; López, J A

    2017-03-01

    This study presents the first detailed analysis of the mitochondrial DNA diversity of least cisco Coregonus sardinella in Alaska using a 678 bp segment of the control region (D-loop) of the mitochondrial genome. Findings suggest that the history of C. sardinella in Alaska differs from that of other species of Coregonus present in the state and surrounding regions. The examined populations of C. sardinella are genetically diverse across Alaska. Sixty-eight distinct mitochondrial haplotypes were identified among 305 individuals sampled from nine locations. The haplotype minimum spanning network and phylogeny showed a modest level of geographic segregation among haplotypes, suggesting high levels of on-going or recent connectivity among distant populations. Observed ΦST values and the results of homogeneity and AMOVAs indicate incipient genetic differentiation between aggregations in three broad regional groups. Sites north of the Brooks Range formed one group, sites in the Yukon and Selawik Rivers formed a second group and sites south of the Yukon drainage formed the third group. Overall, the sequence data showed that a large proportion of mtDNA genetic variation in C. sardinella is shared across Alaska, but this variation is not homogeneously distributed across all regions and for all haplotype groups.

  5. Maternal inheritance of mitochondrial DNA by diverse mechanisms to eliminate paternal mitochondrial DNA

    National Research Council Canada - National Science Library

    Sato, Miyuki; Sato, Ken

    2013-01-01

    .... 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...

  6. mtDNA/nDNA ratio in 14484 LHON mitochondrial mutation carriers.

    Science.gov (United States)

    Nishioka, Tomoki; Soemantri, Augustinus; Ishida, Takafumi

    2004-01-01

    Leber's hereditary optic neuropathy (LHON) is a maternally inherited disease caused by mitochondrial DNA (mtDNA) mutations. In this study, the mtDNA/nuclear DNA ratio was evaluated in 11 LHON patients with the 14484 mutation, 13 asymptomatic carriers and 18 non-carrier relatives as controls, to reveal possible relationships between the disease and mtDNA content. DNAs from peripheral blood lymphocytes were subjected to quantitative PCR. Gender differences and age-dependent changes in the mtDNA content were not observed. Significant increase in the mtDNA content was observed only in the asymptomatic carriers (PLHON development, whereas those whose levels had not, had developed LHON. Since the asymptomatic carriers are the stock of the future LHON patients, monitoring the mtDNA content in patients and their relatives may help to predict the prognosis of the disease.

  7. Detection of mitochondrial DNA deletion by a modified PCR method

    Institute of Scientific and Technical Information of China (English)

    汪振诚; 王学敏; 缪明永; 章卫平; 焦炳华; 倪庆桂

    2003-01-01

    Objective: To develop a simple and efficient method for detecting small populations of mitochondrial DNA deletion. Methods: Peripheral blood cell DNA was obtained from a victim who was accidently exposed to a 60Co radiation source 11 years ago. Using the DNA as template, PCR was performed to generate multiple products including true deletions and artifacts. The full length product was recovered and used as template of secondary PCR. The suspicious deletion product of mtDNA could be confirmed if it was only yielded by first PCR. Using either original primers or their nested primers, the suspicious deletion product was amplified and authenticated as true deletion product. The template was recovered and determined to be a deletion by sequencing directly. Results: A new mtDNA deletion, spanning 889 bp from nt11688 to nt12576, was detected in the peripheral blood cells of the victim. Conclusion: The new PCR-based method is more efficient in detecting small populations of mtDNA deletion than other routine methods. MtDNA deletion is found in the victim, suggesting there is relationship between the deletion and phenotypes of the disease.

  8. Discovery of cyanophage genomes which contain mitochondrial DNA polymerase.

    Science.gov (United States)

    Chan, Yi-Wah; Mohr, Remus; Millard, Andrew D; Holmes, Antony B; Larkum, Anthony W; Whitworth, Anna L; Mann, Nicholas H; Scanlan, David J; Hess, Wolfgang R; Clokie, Martha R J

    2011-08-01

    DNA polymerase γ is a family A DNA polymerase responsible for the replication of mitochondrial DNA in eukaryotes. The origins of DNA polymerase γ have remained elusive because it is not present in any known bacterium, though it has been hypothesized that mitochondria may have inherited the enzyme by phage-mediated nonorthologous displacement. Here, we present an analysis of two full-length homologues of this gene, which were found in the genomes of two bacteriophages, which infect the chlorophyll-d containing cyanobacterium Acaryochloris marina. Phylogenetic analyses of these phage DNA polymerase γ proteins show that they branch deeply within the DNA polymerase γ clade and therefore share a common origin with their eukaryotic homologues. We also found homologues of these phage polymerases in the environmental Community Cyberinfrastructure for Advanced Microbial Ecology Research and Analysis (CAMERA) database, which fell in the same clade. An analysis of the CAMERA assemblies containing the environmental homologues together with the filter fraction metadata indicated some of these assemblies may be of bacterial origin. We also show that the phage-encoded DNA polymerase γ is highly transcribed as the phage genomes are replicated. These findings provide data that may assist in reconstructing the evolution of mitochondria.

  9. Addressing RNA Integrity to Determine the Impact of Mitochondrial DNA Mutations on Brain Mitochondrial Function with Age

    Science.gov (United States)

    Wang, Wei; Scheffler, Katja; Esbensen, Ying; Strand, Janne M.; Stewart, James B.; Bjørås, Magnar; Eide, Lars

    2014-01-01

    Mitochondrial DNA (mtDNA) mutations can result in mitochondrial dysfunction, but emerging experimental data question the fundamental role of mtDNA mutagenesis in age-associated mitochondrial impairment. The multicopy nature of mtDNA renders the impact of a given mtDNA mutation unpredictable. In this study, we compared mtDNA stability and mtRNA integrity during normal aging. Seven distinct sites in mouse brain mtDNA and corresponding mtRNA were analyzed. Accumulation of mtDNA mutations during aging was highly site-specific. The variation in mutation frequencies overrode the age-mediated increase by more than 100-fold and aging generally did not influence mtDNA mutagenesis. Errors introduced by mtRNA polymerase were also site-dependent and up to two hundred-fold more frequent than mtDNA mutations, and independent of mtDNA mutation frequency. We therefore conclude that mitochondrial transcription fidelity limits the impact of mtDNA mutations. PMID:24819950

  10. Mitochondrial DNA damage: Molecular marker of vulnerable nigral neurons in Parkinson's disease

    NARCIS (Netherlands)

    L.H. Sanders (Laurie); J. McCoy (Jennifer); X. Hu (Xiaoping); P.G. Mastroberardino (Pier); B.C. Dickinson (Bryan); C.J. Chang (Christopher); C.T. Chu (Charleen); B. van Houten (Bennett); J.T. Greenamyre (Timothy)

    2014-01-01

    textabstractDNA damage can cause (and result from) oxidative stress and mitochondrial impairment, both of which are implicated in the pathogenesis of Parkinson's disease (PD). We therefore examined the role of mitochondrial DNA (mtDNA) damage in human postmortem brain tissue and in in vivo and in vi

  11. Mitochondrial DNA damage: Molecular marker of vulnerable nigral neurons in Parkinson's disease

    NARCIS (Netherlands)

    L.H. Sanders (Laurie); J. McCoy (Jennifer); X. Hu (Xiaoping); P.G. Mastroberardino (Pier); B.C. Dickinson (Bryan); C.J. Chang (Christopher); C.T. Chu (Charleen); B. van Houten (Bennett); J.T. Greenamyre (Timothy)

    2014-01-01

    textabstractDNA damage can cause (and result from) oxidative stress and mitochondrial impairment, both of which are implicated in the pathogenesis of Parkinson's disease (PD). We therefore examined the role of mitochondrial DNA (mtDNA) damage in human postmortem brain tissue and in in vivo and in

  12. Complete mitochondrial DNA genome of Pseudobagrus truncatus (Siluriformes: Bagridae).

    Science.gov (United States)

    Liang, Hong-wei; Meng, Yan; Li, Zhong; Zhang, Yan; Zou, Gui-wei

    2014-06-01

    In this study, the complete mitochondrial DNA (mtDNA) sequence of Pseudobagrus truncatus (Siluriformes: Bagridae) was determined. The complete mtDNA genome sequence of P. truncatus is 16,533 bp in size. It consists of 13 protein-coding genes, 22 transfer RNA genes, 2 ribosomal RNA genes and one non-coding control region. The gene order and genes were the same as that found in other previously reported catfishes. The overall-based composition was 31.6% A, 26.7% T, 14.9% G and 26.8% C, with a high A + T content (58.3%). This complete mitogenome of P. truncatus provides a basic data for studies on species identification, molecular systematics and conservation genetics.

  13. Mitochondrial DNA data reveal cryptic species within Taenia krabbei.

    Science.gov (United States)

    Lavikainen, Antti; Haukisalmi, Voitto; Lehtinen, Markus J; Laaksonen, Sauli; Holmström, Sauli; Isomursu, Marja; Oksanen, Antti; Meri, Seppo

    2010-06-01

    Cysticerci of Taenia sp. from two elks (Alces alces) in Finland were characterized using morphological criteria and sequences of two mitochondrial DNA regions. The host species, size, structure and location of the cysticerci indicated that they might belong to Taenia krabbei, a circumpolar species occurring in a sylvatic life cycle in wild canids and cervids. Based on the number, length and shape of the rostellar hooks, the specimens could not be unambiguously defined as belonging to T. krabbei, T. cervi, T. ovis or T. solium. In the phylogenetic analysis, based on mitochondrial nucleotide sequence data, Taenia sp. was placed as a sister species of T. solium, distant from T. krabbei isolates previously characterized from Svalbard. This indicates that the Finnish and the Svalbard isolates, resembling T. krabbei, cannot represent a single species. The results suggest that careful morphological and genetic analyses of further isolates from intermediate and definitive hosts are required to define the taxonomic status of these two cryptic species.

  14. Mitochondrial DNA copy numbers in pyramidal neurons are decreased and mitochondrial biogenesis transcriptome signaling is disrupted in Alzheimer's disease hippocampi.

    Science.gov (United States)

    Rice, Ann C; Keeney, Paula M; Algarzae, Norah K; Ladd, Amy C; Thomas, Ravindar R; Bennett, James P

    2014-01-01

    Alzheimer's disease (AD) is the major cause of adult-onset dementia and is characterized in its pre-diagnostic stage by reduced cerebral cortical glucose metabolism and in later stages by reduced cortical oxygen uptake, implying reduced mitochondrial respiration. Using quantitative PCR we determined the mitochondrial DNA (mtDNA) gene copy numbers from multiple groups of 15 or 20 pyramidal neurons, GFAP(+) astrocytes and dentate granule neurons isolated using laser capture microdissection, and the relative expression of mitochondrial biogenesis (mitobiogenesis) genes in hippocampi from 10 AD and 9 control (CTL) cases. AD pyramidal but not dentate granule neurons had significantly reduced mtDNA copy numbers compared to CTL neurons. Pyramidal neuron mtDNA copy numbers in CTL, but not AD, positively correlated with cDNA levels of multiple mitobiogenesis genes. In CTL, but not in AD, hippocampal cDNA levels of PGC1α were positively correlated with multiple downstream mitobiogenesis factors. Mitochondrial DNA copy numbers in pyramidal neurons did not correlate with hippocampal Aβ1-42 levels. After 48 h exposure of H9 human neural stem cells to the neurotoxic fragment Aβ25-35, mtDNA copy numbers were not significantly altered. In summary, AD postmortem hippocampal pyramidal neurons have reduced mtDNA copy numbers. Mitochondrial biogenesis pathway signaling relationships are disrupted in AD, but are mostly preserved in CTL. Our findings implicate complex alterations of mitochondria-host cell relationships in AD.

  15. Low abundance of mitochondrial DNA changes mitochondrial status and renders cells resistant to serum starvation and sodium nitroprusside insult.

    Science.gov (United States)

    Lee, Sung Ryul; Heo, Hye Jin; Jeong, Seung Hun; Kim, Hyoung Kyu; Song, In Sung; Ko, Kyung Soo; Rhee, Byoung Doo; Kim, Nari; Han, Jin

    2015-07-01

    Mutation or depletion of mitochondrial DNA (mtDNA) can cause severe mitochondrial malfunction, originating from the mitochondrion itself, or from the crosstalk between nuclei and mitochondria. However, the changes that would occur if the amount of mtDNA is diminished are less known. Thus, we generated rat myoblast H9c2 cells containing lower amounts of mtDNA via ethidium bromide and uridine supplementation. After confirming the depletion of mtDNA by quantitative PCR and gel electrophoresis analysis, we investigated the changes in mitochondrial physical parameters by using flow cytometry. We also evaluated the resistance of these cells to serum starvation and sodium nitroprusside. H9c2 cells with diminished mtDNA contents showed decreased mitochondrial membrane potential, mass, free calcium, and zinc ion contents as compared to naïve H9c2 cells. Furthermore, cytosolic and mitochondrial reactive oxygen species levels were significantly higher in mtDNA-lowered H9c2 cells than in the naïve cells. Although the oxygen consumption rate and cell proliferation were decreased, mtDNA-lowered H9c2 cells were more resistant to serum deprivation and nitroprusside insults than the naïve H9c2 cells. Taken together, we conclude that the low abundance of mtDNA cause changes in cellular status, such as changes in reactive oxygen species, calcium, and zinc ion levels inducing resistance to stress.

  16. CLK-1 protein has DNA binding activity specific to O(L) region of mitochondrial DNA.

    Science.gov (United States)

    Gorbunova, Vera; Seluanov, Andrei

    2002-04-10

    Mutations in the clk-1 gene of Caenorhabditis elegans extend worm life span and slow down a variety of physiological processes. Here we report that C. elegans CLK-1 as well as its mouse homologue have DNA binding activity that is specific to the O(L) region of mitochondrial DNA. DNA binding activity of CLK-1 is inhibited by ADP, and is altered by mutations that extend nematode life span. Our results suggest that, in addition to its enzymatic function in ubiquinone biosynthesis, CLK-1 is involved in the regulation of mtDNA replication or transcription.

  17. DNA Sequences Proximal to Human Mitochondrial DNA Deletion Breakpoints Prevalent in Human Disease Form G-quadruplexes, a Class of DNA Structures Inefficiently Unwound by the Mitochondrial Replicative Twinkle Helicase

    NARCIS (Netherlands)

    Bharti, S.K.; Sommers, J.A.; Zhou, J.; Kaplan, D.L.; Spelbrink, J.N.; Mergny, J.L.; Brosh, R.M., Jr.

    2014-01-01

    Mitochondrial DNA deletions are prominent in human genetic disorders, cancer, and aging. It is thought that stalling of the mitochondrial replication machinery during DNA synthesis is a prominent source of mitochondrial genome instability; however, the precise molecular determinants of defective mit

  18. Mitochondrial DNA lineages of Italian Giara and Sarcidano horses.

    Science.gov (United States)

    Morelli, L; Useli, A; Sanna, D; Barbato, M; Contu, D; Pala, M; Cancedda, M; Francalacci, P

    2014-10-20

    Giara and Sarcidano are 2 of the 15 extant native Italian horse breeds with limited dispersal capability that originated from a larger number of individuals. The 2 breeds live in two distinct isolated locations on the island of Sardinia. To determine the genetic structure and evolutionary history of these 2 Sardinian breeds, the first hypervariable segment of the mitochondrial DNA (mtDNA) was sequenced and analyzed in 40 Giara and Sarcidano horses and compared with publicly available mtDNA data from 43 Old World breeds. Four different analyses, including genetic distance, analysis of molecular variance, haplotype sharing, and clustering methods, were used to study the genetic relationships between the Sardinian and other horse breeds. The analyses yielded similar results, and the FST values indicated that a high percentage of the total genetic variation was explained by between-breed differences. Consistent with their distinct phenotypes and geographic isolation, the two Sardinian breeds were shown to consist of 2 distinct gene pools that had no gene flow between them. Giara horses were clearly separated from the other breeds examined and showed traces of ancient separation from horses of other breeds that share the same mitochondrial lineage. On the other hand, the data from the Sarcidano horses fit well with variation among breeds from the Iberian Peninsula and North-West Europe: genetic relationships among Sarcidano and the other breeds are consistent with the documented history of this breed.

  19. Development of a Model for the Teaching of Mitochondrial DNA

    Directory of Open Access Journals (Sweden)

    A.P.S. Souza

    2010-05-01

    Full Text Available The Cellular Biology and Molecular Biology are fields of Science that use very abstract concepts, because they look into microscopic and molecular aspects of the nature. The process of teaching/learning of those disciplines requires didactic material, as an alternative approach for the students, to increase the chances of understanding these issues and to become an important tool in the synthesis of this knowledge. One of the methods that can be employed is the didactic models based on multimedia, because they allow an easy and fun interaction with these subjects. On this work was created a new educational model that represents the human mitochondrial DNA molecule, mtDNA, in its circular form, using the softwares Excel 2007 and PowerPoint 2007. The model was constructed in hypertext format, which allowed a quick and interactive access to the information contained in the genes found in the L and the H strands of mtDNA, and its function in the mitochondrial processes, like themechanism of energy production that occurs inside of the mitochondria by the coupling of electron transfer and ATP synthesis or still others uses like forensic identification.

  20. Mitochondrial DNA deletion analysis: a comparison of PCR quantitative methods.

    Science.gov (United States)

    Hamblet, N S; Castora, F J

    1995-02-15

    The role of mitochondrial DNA (mtDNA) deletions in aging and in neurodegenerative diseases is often determined by measuring the amount of deleted mtDNA in the affected tissue. Upon examining brain autopsy tissue from a 59 year old individual with lung cancer we determined by serial dilution PCR and kinetic PCR that a greater ratio of deleted mtDNA was present in the caudate than in the parietal cortex. However, the magnitude difference for these two brain regions appeared to be technique dependent; by serial dilution PCR the caudate had 10 times more deleted mtDNA than the parietal cortex (0.0141 vs 0.0014) whereas kinetic PCR yielded a 4-fold difference (0.1258 vs 0.0316). These results indicate that although it is valid to compare the amount of deleted mtDNA in normal and diseased tissue and draw conclusions based on relative comparisons within one study, greater caution should be exercised when comparing absolute values from studies using different measurement techniques.

  1. A novel R-loop in mouse mitochondrial DNA

    Institute of Scientific and Technical Information of China (English)

    2003-01-01

    Mammalian mitochondrial D-loop L-strand RNA (DL-RNA) is thought functionally not important because no obvious function has been found so far. In this study, we detected a novel D-loop L-strand RNA (DL-RNA) in mouse mitochondrion by RT-PCR. The L-strand RNA spans the whole D-loop region of mouse mtDNA, and is resistant to RNase A and RNase T1 but not RNase H digestion. After binding of the L-strand RNA to D-loop, the DL-RNA complex can protect the D-loop from digestion by restriction endonuclease HaeⅢ. These results indicate that a novel RNA-DNA triplex hybrid (R-loop) can be formed in mouse mtDNA D-loop region, and that the DL-RNA structure is capable of protecting the D-loop from certain microbial restriction enzyme digestion. And the similar R-loop structure can not be found in Cyt.b gene in control experiment which confirmed this R-loop is not the fleeting structure in RNA transcription. Considering the D-loop represents the control region of mtDNA, the novel triplex DNA-RNA complex may play an important role in mtDNA replication and transcription.

  2. Historically low mitochondrial DNA diversity in koalas (Phascolarctos cinereus).

    Science.gov (United States)

    Tsangaras, Kyriakos; Ávila-Arcos, María C; Ishida, Yasuko; Helgen, Kristofer M; Roca, Alfred L; Greenwood, Alex D

    2012-10-24

    The koala (Phascolarctos cinereus) is an arboreal marsupial that was historically widespread across eastern Australia until the end of the 19th century when it suffered a steep population decline. Hunting for the fur trade, habitat conversion, and disease contributed to a precipitous reduction in koala population size during the late 1800s and early 1900s. To examine the effects of these reductions in population size on koala genetic diversity, we sequenced part of the hypervariable region of mitochondrial DNA (mtDNA) in koala museum specimens collected in the 19th and 20th centuries, hypothesizing that the historical samples would exhibit greater genetic diversity. The mtDNA haplotypes present in historical museum samples were identical to haplotypes found in modern koala populations, and no novel haplotypes were detected. Rarefaction analyses suggested that the mtDNA genetic diversity present in the museum samples was similar to that of modern koalas. Low mtDNA diversity may have been present in koala populations prior to recent population declines. When considering management strategies, low genetic diversity of the mtDNA hypervariable region may not indicate recent inbreeding or founder events but may reflect an older historical pattern for koalas.

  3. Historically low mitochondrial DNA diversity in koalas (Phascolarctos cinereus

    Directory of Open Access Journals (Sweden)

    Tsangaras Kyriakos

    2012-10-01

    Full Text Available Abstract Background The koala (Phascolarctos cinereus is an arboreal marsupial that was historically widespread across eastern Australia until the end of the 19th century when it suffered a steep population decline. Hunting for the fur trade, habitat conversion, and disease contributed to a precipitous reduction in koala population size during the late 1800s and early 1900s. To examine the effects of these reductions in population size on koala genetic diversity, we sequenced part of the hypervariable region of mitochondrial DNA (mtDNA in koala museum specimens collected in the 19th and 20th centuries, hypothesizing that the historical samples would exhibit greater genetic diversity. Results The mtDNA haplotypes present in historical museum samples were identical to haplotypes found in modern koala populations, and no novel haplotypes were detected. Rarefaction analyses suggested that the mtDNA genetic diversity present in the museum samples was similar to that of modern koalas. Conclusions Low mtDNA diversity may have been present in koala populations prior to recent population declines. When considering management strategies, low genetic diversity of the mtDNA hypervariable region may not indicate recent inbreeding or founder events but may reflect an older historical pattern for koalas.

  4. Mesencephalic complex I deficiency does not correlate with parkinsonism in mitochondrial DNA maintenance disorders.

    Science.gov (United States)

    Palin, Eino J H; Paetau, Anders; Suomalainen, Anu

    2013-08-01

    Genetic evidence from recessively inherited Parkinson's disease has indicated a clear causative role for mitochondrial dysfunction in Parkinson's disease. This role has long been discussed based on findings that toxic inhibition of mitochondrial respiratory complex I caused parkinsonism and that tissues of patients with Parkinson's disease show complex I deficiency. Disorders of mitochondrial DNA maintenance are a common cause of inherited neurodegenerative disorders, and lead to mitochondrial DNA deletions or depletion and respiratory chain defect, including complex I deficiency. However, parkinsonism associates typically with defects of catalytic domain of mitochondrial DNA polymerase gamma. Surprisingly, however, not all mutations affecting DNA polymerase gamma manifest as parkinsonism, but, for example, spacer region mutations lead to spinocerebellar ataxia and/or severe epilepsy. Furthermore, defective Twinkle helicase, a close functional companion of DNA polymerase gamma in mitochondrial DNA replication, results in infantile-onset spinocerebellar ataxia, epilepsy or adult-onset mitochondrial myopathy, but not typically parkinsonism. Here we sought for clues for this specificity in the neurological manifestations of mitochondrial DNA maintenance disorders by studying mesencephalic neuropathology of patients with DNA polymerase gamma or Twinkle defects, with or without parkinsonism. We show here that all patients with mitochondrial DNA maintenance disorders had neuronopathy in substantia nigra, most severe in DNA polymerase gamma-associated parkinsonism. The oculomotor nucleus was also affected, but less severely. In substantia nigra, all patients had a considerable decrease of respiratory chain complex I, but other respiratory chain enzymes were not affected. Complex I deficiency did not correlate with parkinsonism, age, affected gene or inheritance. We conclude that the cell number in substantia nigra correlated well with parkinsonism in DNA polymerase gamma

  5. Protection from palmitate-induced mitochondrial DNA damage prevents from mitochondrial oxidative stress, mitochondrial dysfunction, apoptosis, and impaired insulin signaling in rat L6 skeletal muscle cells.

    Science.gov (United States)

    Yuzefovych, Larysa V; Solodushko, Viktoriya A; Wilson, Glenn L; Rachek, Lyudmila I

    2012-01-01

    Saturated free fatty acids have been implicated in the increase of oxidative stress, mitochondrial dysfunction, apoptosis, and insulin resistance seen in type 2 diabetes. The purpose of this study was to determine whether palmitate-induced mitochondrial DNA (mtDNA) damage contributed to increased oxidative stress, mitochondrial dysfunction, apoptosis, impaired insulin signaling, and reduced glucose uptake in skeletal muscle cells. Adenoviral vectors were used to deliver the DNA repair enzyme human 8-oxoguanine DNA glycosylase/(apurinic/apyrimidinic) lyase (hOGG1) to mitochondria in L6 myotubes. After palmitate exposure, we evaluated mtDNA damage, mitochondrial function, production of mitochondrial reactive oxygen species, apoptosis, insulin signaling pathways, and glucose uptake. Protection of mtDNA from palmitate-induced damage by overexpression of hOGG1 targeted to mitochondria significantly diminished palmitate-induced mitochondrial superoxide production, restored the decline in ATP levels, reduced activation of c-Jun N-terminal kinase (JNK) kinase, prevented cells from entering apoptosis, increased insulin-stimulated phosphorylation of serine-threonine kinase (Akt) (Ser473) and tyrosine phosphorylation of insulin receptor substrate-1, and thereby enhanced glucose transporter 4 translocation to plasma membrane, and restored insulin signaling. Addition of a specific inhibitor of JNK mimicked the effect of mitochondrial overexpression of hOGG1 and partially restored insulin sensitivity, thus confirming the involvement of mtDNA damage and subsequent increase of oxidative stress and JNK activation in insulin signaling in L6 myotubes. Our results are the first to report that mtDNA damage is the proximal cause in palmitate-induced mitochondrial dysfunction and impaired insulin signaling and provide strong evidence that targeting DNA repair enzymes into mitochondria in skeletal muscles could be a potential therapeutic treatment for insulin resistance.

  6. Condensation of Plasmid DNA Enhances Mitochondrial Association in Skeletal Muscle Following Hydrodynamic Limb Vein Injection

    Directory of Open Access Journals (Sweden)

    Yukari Yasuzaki

    2014-08-01

    Full Text Available Mitochondrial gene therapy and diagnosis have the potential to provide substantial medical benefits. However, the utility of this approach has not yet been realized because the technology available for mitochondrial gene delivery continues to be a bottleneck. We previously reported on mitochondrial gene delivery in skeletal muscle using hydrodynamic limb vein (HLV injection. HLV injection, a useful method for nuclear transgene expression, involves the rapid injection of a large volume of naked plasmid DNA (pDNA. Moreover, the use of a condensed form of pDNA enhances the nuclear transgene expression by the HLV injection. The purpose of this study was to compare naked pDNA and condensed pDNA for mitochondrial association in skeletal muscle, when used in conjunction with HLV injection. PCR analysis showed that the use of condensed pDNA rather than naked pDNA resulted in a more effective mitochondrial association with pDNA, suggesting that the physicochemical state of pDNA plays a key role. Moreover, no mitochondrial toxicities in skeletal muscle following the HLV injection of condensed pDNA were confirmed, as evidenced by cytochrome c oxidase activity and mitochondrial membrane potential. These findings have the potential to contribute to the development for in vivo mitochondrial gene delivery system.

  7. Prolonged decay of molecular rate estimates for metazoan mitochondrial DNA

    Directory of Open Access Journals (Sweden)

    Martyna Molak

    2015-03-01

    Full Text Available Evolutionary timescales can be estimated from genetic data using the molecular clock, often calibrated by fossil or geological evidence. However, estimates of molecular rates in mitochondrial DNA appear to scale negatively with the age of the clock calibration. Although such a pattern has been observed in a limited range of data sets, it has not been studied on a large scale in metazoans. In addition, there is uncertainty over the temporal extent of the time-dependent pattern in rate estimates. Here we present a meta-analysis of 239 rate estimates from metazoans, representing a range of timescales and taxonomic groups. We found evidence of time-dependent rates in both coding and non-coding mitochondrial markers, in every group of animals that we studied. The negative relationship between the estimated rate and time persisted across a much wider range of calibration times than previously suggested. This indicates that, over long time frames, purifying selection gives way to mutational saturation as the main driver of time-dependent biases in rate estimates. The results of our study stress the importance of accounting for time-dependent biases in estimating mitochondrial rates regardless of the timescale over which they are inferred.

  8. Sperm mitochondrial DNA deletion in Iranian infertiles with asthenozoospermia.

    Science.gov (United States)

    Bahrehmand Namaghi, I; Vaziri, H

    2017-04-01

    Asthenozoospermia is an important cause of male infertility. The mutations in sperm mitochondrial DNA (mtDNA) result in either functionless or malfunctioning some proteins, subsequently affecting sperm motility leading to asthenozoospermia. The purpose of this study was to investigate sperm mtDNA 4,977-bp deletion in infertile men with low sperm motility/immotile spermatozoa compared to healthy subjects with high sperm motility. Semen samples of 256 asthenozoospermic infertiles and 200 controls from northern Iran were collected. After extraction of spermatozoa total DNA, Gap-polymerase chain reaction (Gap-PCR) was performed. The deletion was observed in 85.93% of patients with asthenozoospermia compared with 14% in controls [OR = 37.5397, 95% confidence interval = 12.937-108.9276, p asthenozoospermia-induced infertility in the population examined. Large-scale mtDNA deletions in spermatozoa may induce bioenergetic disorders. Nevertheless, to validate our results broader research may be needed.

  9. Translesion synthesis past acrolein-derived DNA adducts by human mitochondrial DNA polymerase γ.

    Science.gov (United States)

    Kasiviswanathan, Rajesh; Minko, Irina G; Lloyd, R Stephen; Copeland, William C

    2013-05-17

    Acrolein, a mutagenic aldehyde, is produced endogenously by lipid peroxidation and exogenously by combustion of organic materials, including tobacco products. Acrolein reacts with DNA bases forming exocyclic DNA adducts, such as γ-hydroxy-1,N(2)-propano-2'-deoxyguanosine (γ-HOPdG) and γ-hydroxy-1,N(6)-propano-2'-deoxyadenosine (γ-HOPdA). The bulky γ-HOPdG adduct blocks DNA synthesis by replicative polymerases but can be bypassed by translesion synthesis polymerases in the nucleus. Although acrolein-induced adducts are likely to be formed and persist in mitochondrial DNA, animal cell mitochondria lack specialized translesion DNA synthesis polymerases to tolerate these lesions. Thus, it is important to understand how pol γ, the sole mitochondrial DNA polymerase in human cells, acts on acrolein-adducted DNA. To address this question, we investigated the ability of pol γ to bypass the minor groove γ-HOPdG and major groove γ-HOPdA adducts using single nucleotide incorporation and primer extension analyses. The efficiency of pol γ-catalyzed bypass of γ-HOPdG was low, and surprisingly, pol γ preferred to incorporate purine nucleotides opposite the adduct. Pol γ also exhibited ∼2-fold lower rates of excision of the misincorporated purine nucleotides opposite γ-HOPdG compared with the corresponding nucleotides opposite dG. Extension of primers from the termini opposite γ-HOPdG was accomplished only following error-prone purine nucleotide incorporation. However, pol γ preferentially incorporated dT opposite the γ-HOPdA adduct and efficiently extended primers from the correctly paired terminus, indicating that γ-HOPdA is probably nonmutagenic. In summary, our data suggest that acrolein-induced exocyclic DNA lesions can be bypassed by mitochondrial DNA polymerase but, in the case of the minor groove γ-HOPdG adduct, at the cost of unprecedented high mutation rates.

  10. Mitochondrial DNA analysis of two southern African elephant populations

    Directory of Open Access Journals (Sweden)

    M.F. Essop

    1996-01-01

    Full Text Available The modern view is that there are at most only two valid forms of the African elephant namely Loxodonta qfricana africana, the bush elephant, and L.a. cyclotis, the forest elephant (Ansell 1974; Meester et al. 1986. The Knysna elephant which was also described as a separate sub-species is now almost extinct. Plans to augment the remnant population by introducing other animals must take into account the taxonomic questions and issue of conserving elephant gene pools (Greig 1982a. Mitochondrial DNA (mtDNA restriction fragment-size comparisons were performed on specimens from the Kruger National Park and the Addo Elephant National Park. If the Addo population's results are extrapolated to the Knysna population, it may be concluded that there is no genetic evidence for the Kruger and Knysna elephant populations to be considered as different sub-species.

  11. What cost mitochondria? The maintenance of functional mitochondrial DNA within and across generations

    NARCIS (Netherlands)

    Aanen, D.K.; Spelbrink, J.N.; Beekman, M.

    2014-01-01

    The peculiar biology of mitochondrial DNA (mtDNA) potentially has detrimental consequences for organismal health and lifespan. Typically, eukaryotic cells contain multiple mitochondria, each with multiple mtDNA genomes. The high copy number of mtDNA implies that selection on mtDNA functionality is r

  12. A mitochondrial DNA SNP multiplex assigning Caucasians into 36 haplo- and subhaplogroups

    DEFF Research Database (Denmark)

    Mikkelsen, Martin; Rockenbauer, Eszter; Sørensen, Erik;

    2008-01-01

    Mitochondrial DNA (mtDNA) is maternally inherited without recombination events and has a high copy number, which makes mtDNA analysis feasible even when genomic DNA is sparse or degraded. Here, we present a SNP typing assay with 33 previously described mtDNA coding region SNPs for haplogroup assi...

  13. What cost mitochondria? The maintenance of functional mitochondrial DNA within and across generations

    NARCIS (Netherlands)

    Aanen, D.K.; Spelbrink, J.N.; Beekman, M.

    2014-01-01

    The peculiar biology of mitochondrial DNA (mtDNA) potentially has detrimental consequences for organismal health and lifespan. Typically, eukaryotic cells contain multiple mitochondria, each with multiple mtDNA genomes. The high copy number of mtDNA implies that selection on mtDNA functionality is

  14. Enhanced base excision repair capacity in carotid atherosclerosis may protect nuclear DNA but not mitochondrial DNA

    DEFF Research Database (Denmark)

    Skarpengland, Tonje; B. Dahl, Tuva; Skjelland, Mona

    2016-01-01

    Lesional and systemic oxidative stress has been implicated in the pathogenesis of atherosclerosis, potentially leading to accumulation of DNA base lesions within atherosclerotic plaques. Although base excision repair (BER) is a major pathway counteracting oxidative DNA damage, our knowledge on BER...... and accumulation of DNA base lesions in clinical atherosclerosis is scarce. Here, we evaluated the transcriptional profile of a wide spectrum of BER components as well as DNA damage accumulation in atherosclerotic and non-atherosclerotic arteries. BER gene expression levels were analyzed in 162 carotid plaques, 8...... genes in atherosclerosis may contribute to lesional nuclear DNA stability but appears insufficient to maintain mtDNA integrity, potentially influencing mitochondrial function in cells within the atherosclerotic lesion....

  15. Mitochondrial DNA haplogroup H structure in North Africa

    Directory of Open Access Journals (Sweden)

    Dzimiri Nduna

    2009-02-01

    Full Text Available Abstract Background The Strait of Gibraltar separating the Iberian Peninsula from North Africa is thought to be a stronger barrier to gene flow for male than for female lineages. However, the recent subdivision of the haplogroup H at mitochondrial DNA (mtDNA level has revealed greater genetic differentiation among geographic regions than previously detected. The dissection of the mtDNA haplogroup H in North Africa, and its comparison with the Iberian Peninsula and Near-East profiles would help clarify the relative affinities among these regions. Results Like the Iberian Peninsula, the dominant mtDNA haplogroup H subgroups in North Africa are H1 (42% and H3 (13%. The similarity between these regions is stronger in the North-West edge affecting mainly Moroccan Arabs, West Saharans and Mauritanians, and decreases eastwards probably due to gene flow from Near East as attested for the higher frequencies of H4, H5, H7, H8 and H11 subgroups. Moroccan Berbers show stronger affinities with Tunisian and Tunisian Berbers than with Moroccan Arabs. Coalescence ages for H1 (11 ± 2 ky and H3 (11 ± 4 ky in North Africa point to the possibility of a late Palaeolithic settlement for these lineages similar to those found for other mtDNA haplogroups. Total and partial mtDNA genomic sequencing unveiled stronger mtDNA differentiation among regions than previously found using HVSI mtDNA based analysis. Conclusion The subdivision of the mtDNA haplogroup H in North Africa has confirmed that the genetic differentiation found among Western and Eastern populations is mainly due to geographical rather than cultural barriers. It also shows that the historical Arabian role on the region had more a cultural than a demic effect. Whole mtDNA sequencing of identical H haplotypes based on HVSI and RFLP information has unveiled additional mtDNA differences between North African and Iberian Peninsula lineages, pointing to an older mtDNA genetic flow between regions than previously

  16. Statistical analysis of post mortem DNA damage-derived miscoding lesions in Neandertal mitochondrial DNA

    OpenAIRE

    Vives, Sergi; Gilbert, Thomas; Arenas, Conchita; Gigli, Elena, 1978-; Lao Grueso, Oscar; Lalueza-Fox, Carles

    2008-01-01

    textabstractBackground. We have analysed the distribution of post mortem DNA damage derived miscoding lesions from the datasets of seven published Neandertal specimens that have extensive cloned sequence coverage over the mitochondrial DNA (mtDNA) hypervariable region 1 (HVS1). The analysis was restricted to C → T and G → A miscoding lesions (the predominant manifestation of post mortem damage) that are seen at a frequency of more than one clone among sequences from a single PCR, but do not r...

  17. Complete genome sequence of mitochondrial DNA (mtDNA) of Chlorella sorokiniana.

    Science.gov (United States)

    Orsini, Massimiliano; Costelli, Cristina; Malavasi, Veronica; Cusano, Roberto; Concas, Alessandro; Angius, Andrea; Cao, Giacomo

    2016-01-01

    The complete sequence of mitochondrial genome of the Chlorella sorokiniana strain (SAG 111-8 k) is presented in this work. Within the Chlorella genus, it represents the second species with a complete sequenced and annotated mitochondrial genome (GenBank accession no. KM241869). The genome consists of circular chromosomes of 52,528 bp and encodes a total of 31 protein coding genes, 3 rRNAs and 26 tRNAs. The overall AT contents of the C. sorokiniana mtDNA is 70.89%, while the coding sequence is of 97.4%.

  18. ER-mitochondria contacts couple mtDNA synthesis with mitochondrial division in human cells.

    Science.gov (United States)

    Lewis, Samantha C; Uchiyama, Lauren F; Nunnari, Jodi

    2016-07-15

    Mitochondrial DNA (mtDNA) encodes RNAs and proteins critical for cell function. In human cells, hundreds to thousands of mtDNA copies are replicated asynchronously, packaged into protein-DNA nucleoids, and distributed within a dynamic mitochondrial network. The mechanisms that govern how nucleoids are chosen for replication and distribution are not understood. Mitochondrial distribution depends on division, which occurs at endoplasmic reticulum (ER)-mitochondria contact sites. These sites were spatially linked to a subset of nucleoids selectively marked by mtDNA polymerase and engaged in mtDNA synthesis--events that occurred upstream of mitochondrial constriction and division machine assembly. Our data suggest that ER tubules proximal to nucleoids are necessary but not sufficient for mtDNA synthesis. Thus, ER-mitochondria contacts coordinate licensing of mtDNA synthesis with division to distribute newly replicated nucleoids to daughter mitochondria.

  19. The elusive nature of adaptive mitochondrial DNA evolution of an arctic lineage prone to frequent introgression

    National Research Council Canada - National Science Library

    Melo-Ferreira, José; Vilela, Joana; Fonseca, Miguel M; da Fonseca, Rute R; Boursot, Pierre; Alves, Paulo C

    2014-01-01

    Mitochondria play a fundamental role in cellular metabolism, being responsible for most of the energy production of the cell in the oxidative phosphorylation (OXPHOS) pathway. Mitochondrial DNA (mtDNA...

  20. Detection of novel polymorphisms in the mitochondrial DNA D-Loop ...

    African Journals Online (AJOL)

    aghomotsegin

    2015-04-08

    Apr 8, 2015 ... The mitochondrial DNA (mtDNA) is a small circular genome located within the .... to you for choosing the project, your enthusiasm for helping us ...... Comparisons of the characteristics across D-loop region in different human.

  1. Oxidative phosphorylation differences between mitochondrial DNA haplogroups modify the risk of Leber's hereditary optic neuropathy.

    Science.gov (United States)

    Gómez-Durán, Aurora; Pacheu-Grau, David; Martínez-Romero, Iñigo; López-Gallardo, Ester; López-Pérez, Manuel J; Montoya, Julio; Ruiz-Pesini, Eduardo

    2012-08-01

    Leber's hereditary optic neuropathy is a maternally inherited optic atrophy caused by mitochondrial DNA point mutations. Previous epidemiological studies have shown that individuals from mitochondrial genetic backgrounds (haplogroups) J/Uk and H have a higher and a lower risk, respectively, of suffering this disorder. To analyze the bases of these associations at cellular and molecular levels, functional studies with cybrids provide high quality evidence. Cybrids from haplogroup J contain less mitochondrial deoxyribonucleic acid (DNA) and ribonucleic acid (RNA) and synthesize a smaller amount of mitochondrial DNA-encoded polypeptides than those from haplogroup H. Haplogroup J cybrids also display lower oxygen consumption, mitochondrial inner membrane potential and total adenosine-5'-triphosphate (ATP) levels. Moreover, mitochondrial DNA levels correlate with many parameters of the oxidative phosphorylation system. These results suggest that the mitochondrial DNA amount determines oxidative phosphorylation capacity and, along with other recently published observations, support the possibility that mitochondrial DNA levels may be responsible for the bias of the disorder toward males, for the incomplete penetrance of mutations causing Leber's hereditary optic neuropathy and for the association of the disease with particular mitochondrial DNA haplogroups.

  2. Human mitochondrial DNA complete amplification and sequencing: a new validated primer set that prevents nuclear DNA sequences of mitochondrial origin co-amplification.

    Science.gov (United States)

    Ramos, Amanda; Santos, Cristina; Alvarez, Luis; Nogués, Ramon; Aluja, Maria Pilar

    2009-05-01

    To date, there are no published primers to amplify the entire mitochondrial DNA (mtDNA) that completely prevent the amplification of nuclear DNA (nDNA) sequences of mitochondrial origin. The main goal of this work was to design, validate and describe a set of primers, to specifically amplify and sequence the complete human mtDNA, allowing the correct interpretation of mtDNA heteroplasmy in healthy and pathological samples. Validation was performed using two different approaches: (i) Basic Local Alignment Search Tool and (ii) amplification using isolated nDNA obtained from sperm cells by differential lyses. During the validation process, two mtDNA regions, with high similarity with nDNA, represent the major problematic areas for primer design. One of these could represent a non-published nuclear DNA sequence of mitochondrial origin. For two of the initially designed fragments, the amplification results reveal PCR artifacts that can be attributed to the poor quality of the DNA. After the validation, nine overlapping primer pairs to perform mtDNA amplification and 22 additional internal primers for mtDNA sequencing were obtained. These primers could be a useful tool in future projects that deal with mtDNA complete sequencing and heteroplasmy detection, since they represent a set of primers that have been tested for the non-amplification of nDNA.

  3. Characterization of nucleotide misincorporation patterns in the iceman's mitochondrial DNA.

    Directory of Open Access Journals (Sweden)

    Cristina Olivieri

    Full Text Available BACKGROUND: The degradation of DNA represents one of the main issues in the genetic analysis of archeological specimens. In the recent years, a particular kind of post-mortem DNA modification giving rise to nucleotide misincorporation ("miscoding lesions" has been the object of extensive investigations. METHODOLOGY/PRINCIPAL FINDINGS: To improve our knowledge regarding the nature and incidence of ancient DNA nucleotide misincorporations, we have utilized 6,859 (629,975 bp mitochondrial (mt DNA sequences obtained from the 5,350-5,100-years-old, freeze-desiccated human mummy popularly known as the Tyrolean Iceman or Otzi. To generate the sequences, we have applied a mixed PCR/pyrosequencing procedure allowing one to obtain a particularly high sequence coverage. As a control, we have produced further 8,982 (805,155 bp mtDNA sequences from a contemporary specimen using the same system and starting from the same template copy number of the ancient sample. From the analysis of the nucleotide misincorporation rate in ancient, modern, and putative contaminant sequences, we observed that the rate of misincorporation is significantly lower in modern and putative contaminant sequence datasets than in ancient sequences. In contrast, type 2 transitions represent the vast majority (85% of the observed nucleotide misincorporations in ancient sequences. CONCLUSIONS/SIGNIFICANCE: This study provides a further contribution to the knowledge of nucleotide misincorporation patterns in DNA sequences obtained from freeze-preserved archeological specimens. In the Iceman system, ancient sequences can be clearly distinguished from contaminants on the basis of nucleotide misincorporation rates. This observation confirms a previous identification of the ancient mummy sequences made on a purely phylogenetical basis. The present investigation provides further indication that the majority of ancient DNA damage is reflected by type 2 (cytosine

  4. Advances in Human Mitochondrial Diseases Molecular Genetic Analysis of Pathogenic mtDNA Mutations.

    Science.gov (United States)

    Davidson, E; King, M P

    1997-01-01

    The mitochondrial diseases are a heterogeneous group of disorders that have been defined by specific morphological alterations in muscle and by deficits of the mitochondrial respiratory chain. The morphological hallmarks of these diseases include ragged-red fibers (an extensive proliferation of mitochondria in muscle fibers) and abnormal paracrystalline inclusions and membrane structures in mitochondria. The identification of pathogenic mutations in mitochondrial DNA (mtDNA) has resulted in a genetic classification of mitochondrial diseases. Investigations are being conducted to understand the molecular basis for the biochemical and morphological alterations of mitochondria associated with mtDNA mutations. © 1997, Elsevier Science Inc. (Trends Cardiovasc Med 1997;7:16-24).

  5. Mitochondrial DNA variation in the Viking age population of Norway.

    Science.gov (United States)

    Krzewińska, Maja; Bjørnstad, Gro; Skoglund, Pontus; Olason, Pall Isolfur; Bill, Jan; Götherström, Anders; Hagelberg, Erika

    2015-01-19

    The medieval Norsemen or Vikings had an important biological and cultural impact on many parts of Europe through raids, colonization and trade, from about AD 793 to 1066. To help understand the genetic affinities of the ancient Norsemen, and their genetic contribution to the gene pool of other Europeans, we analysed DNA markers in Late Iron Age skeletal remains from Norway. DNA was extracted from 80 individuals, and mitochondrial DNA polymorphisms were detected by next-generation sequencing. The sequences of 45 ancient Norwegians were verified as genuine through the identification of damage patterns characteristic of ancient DNA. The ancient Norwegians were genetically similar to previously analysed ancient Icelanders, and to present-day Shetland and Orkney Islanders, Norwegians, Swedes, Scots, English, German and French. The Viking Age population had higher frequencies of K*, U*, V* and I* haplogroups than their modern counterparts, but a lower proportion of T* and H* haplogroups. Three individuals carried haplotypes that are rare in Norway today (U5b1b1, Hg A* and an uncommon variant of H*). Our combined analyses indicate that Norse women were important agents in the overseas expansion and settlement of the Vikings, and that women from the Orkneys and Western Isles contributed to the colonization of Iceland.

  6. Holes influence the mutation spectrum of human mitochondrial DNA

    Science.gov (United States)

    Villagran, Martha; Miller, John

    Mutations drive evolution and disease, showing highly non-random patterns of variant frequency vs. nucleotide position. We use computational DNA hole spectroscopy [M.Y. Suarez-Villagran & J.H. Miller, Sci. Rep. 5, 13571 (2015)] to reveal sites of enhanced hole probability in selected regions of human mitochondrial DNA. A hole is a mobile site of positive charge created when an electron is removed, for example by radiation or contact with a mutagenic agent. The hole spectra are quantum mechanically computed using a two-stranded tight binding model of DNA. We observe significant correlation between spectra of hole probabilities and of genetic variation frequencies from the MITOMAP database. These results suggest that hole-enhanced mutation mechanisms exert a substantial, perhaps dominant, influence on mutation patterns in DNA. One example is where a trapped hole induces a hydrogen bond shift, known as tautomerization, which then triggers a base-pair mismatch during replication. Our results deepen overall understanding of sequence specific mutation rates, encompassing both hotspots and cold spots, which drive molecular evolution.

  7. Amplification of Mitochondrial DNA for detection of Plasmodiumvivax in Balochistan.

    Science.gov (United States)

    Shahwani, Muhammad Naeem; Nisar, Samia; Aleem, Abdul; Panezai, Marina; Afridi, Sarwat; Malik, Shaukat Iqbal

    2017-05-01

    To access a new step using PCR to amplify the targeted mtDNA sequence for detecting specifically Plasmodium vivax and its co-infections, false positive and false negative results with Plasmodium falciparum. In this study we have standardized a new technical approach in which the target mitochondrial DNA sequence (mtDNA) was amplified by using a PCR technique as a tool to detect Plasmodium spp. Species specific primers were designed to hybridize with cytochrome c oxidase gene of P. vivax (cox I) and P. falciparum (cox III). Two hundred blood samples were collected on the basis of clinical symptoms which were initially examined through microscopic analysis after preparing Giemsa stained thick and thin blood smears. Afterwards genomic DNA was extracted from all samples and was then subjected to PCR amplification by using species specific primers and amplified segments were sequenced for confirmation of results. One-hundred and thirty-two blood samples were detected as positive for malaria by PCR, out of which 64 were found to be positive by PCR and 53 by both microscopy and PCR for P.vivax infection. Nine samples were found to be false negative, one P.vivax mono infection was declared as co infection by PCR and 3 samples identified as having P.falciparum gametes were confirmed as P.vivax by PCR amplification. Sensitivity and specificity were found to be 85% and 92% respectively. Results obtained through PCR method were comparatively better and reliable than microscopy.

  8. Segregation of naturally occurring mitochondrial DNA variants in a mini-pig model

    Science.gov (United States)

    Within cells and tissues, the maternally inherited mitochondrial genome (mtDNA) is present in multimeric form and can harbour naturally occurring variants. Whilst high variant load can cause mitochondrial disease, naturally occurring mtDNA variants likely persist at low levels across generations of ...

  9. Complete DNA sequence of the linear mitochondrial genome of the pathogenic yeast Candida parapsilosis

    DEFF Research Database (Denmark)

    Nosek, J.; Novotna, M.; Hlavatovicova, Z.

    2004-01-01

    The complete sequence of the mitochondrial DNA of the opportunistic yeast pathogen Candida parapsilosis was determined. The mitochondrial genome is represented by linear DNA molecules terminating with tandem repeats of a 738-bp unit. The number of repeats varies, thus generating a population...

  10. Complete DNA sequence of the linear mitochondrial genome of the pathogenic yeast Candida parapsilosis

    DEFF Research Database (Denmark)

    Nosek, J.; Novotna, M.; Hlavatovicova, Z.

    2004-01-01

    The complete sequence of the mitochondrial DNA of the opportunistic yeast pathogen Candida parapsilosis was determined. The mitochondrial genome is represented by linear DNA molecules terminating with tandem repeats of a 738-bp unit. The number of repeats varies, thus generating a population...

  11. Mitochondrial DNA deletion and impairment of mitochondrial biogenesis by reactive oxygen species in ionizing radiation-induced premature senescence

    Energy Technology Data Exchange (ETDEWEB)

    Eom, Hyeon Soo; Jung, U Hee; Jo, Sung Kee [Korea Atomic Energy Research Institute, Daejeon (Korea, Republic of)

    2011-10-15

    The aim of this study was to determine whether an increase of ROS level in cellular senescence induced by IR could mediate mtDNA deletion via impairment of mitochondria biogenesis in IMR-90 human lung fibroblast cells. Our results showed that IR induced cellular senescence, intracellular ROS, and mtDNA deletion, and in particular, suppressed the expression of mitochondrial biogenesis genes (NRF-1, TFAM). Furthermore, these IR-induced events were abolished using a potent antioxidant, NAC, which suggests that ROS is a key cause of mtDNA deletion in IR-induced cellular senescence, and that the alteration of mitochondrial biogenesis may mediate these processes

  12. Presequence-Independent Mitochondrial Import of DNA Ligase Facilitates Establishment of Cell Lines with Reduced mtDNA Copy Number.

    Directory of Open Access Journals (Sweden)

    Domenico Spadafora

    Full Text Available Due to the essential role played by mitochondrial DNA (mtDNA in cellular physiology and bioenergetics, methods for establishing cell lines with altered mtDNA content are of considerable interest. Here, we report evidence for the existence in mammalian cells of a novel, low- efficiency, presequence-independent pathway for mitochondrial protein import, which facilitates mitochondrial uptake of such proteins as Chlorella virus ligase (ChVlig and Escherichia coli LigA. Mouse cells engineered to depend on this pathway for mitochondrial import of the LigA protein for mtDNA maintenance had severely (up to >90% reduced mtDNA content. These observations were used to establish a method for the generation of mouse cell lines with reduced mtDNA copy number by, first, transducing them with a retrovirus encoding LigA, and then inactivating in these transductants endogenous Lig3 with CRISPR-Cas9. Interestingly, mtDNA depletion to an average level of one copy per cell proceeds faster in cells engineered to maintain mtDNA at low copy number. This makes a low-mtDNA copy number phenotype resulting from dependence on mitochondrial import of DNA ligase through presequence-independent pathway potentially useful for rapidly shifting mtDNA heteroplasmy through partial mtDNA depletion.

  13. Presequence-Independent Mitochondrial Import of DNA Ligase Facilitates Establishment of Cell Lines with Reduced mtDNA Copy Number.

    Science.gov (United States)

    Spadafora, Domenico; Kozhukhar, Natalia; Alexeyev, Mikhail F

    2016-01-01

    Due to the essential role played by mitochondrial DNA (mtDNA) in cellular physiology and bioenergetics, methods for establishing cell lines with altered mtDNA content are of considerable interest. Here, we report evidence for the existence in mammalian cells of a novel, low- efficiency, presequence-independent pathway for mitochondrial protein import, which facilitates mitochondrial uptake of such proteins as Chlorella virus ligase (ChVlig) and Escherichia coli LigA. Mouse cells engineered to depend on this pathway for mitochondrial import of the LigA protein for mtDNA maintenance had severely (up to >90%) reduced mtDNA content. These observations were used to establish a method for the generation of mouse cell lines with reduced mtDNA copy number by, first, transducing them with a retrovirus encoding LigA, and then inactivating in these transductants endogenous Lig3 with CRISPR-Cas9. Interestingly, mtDNA depletion to an average level of one copy per cell proceeds faster in cells engineered to maintain mtDNA at low copy number. This makes a low-mtDNA copy number phenotype resulting from dependence on mitochondrial import of DNA ligase through presequence-independent pathway potentially useful for rapidly shifting mtDNA heteroplasmy through partial mtDNA depletion.

  14. Nuclear Expression of a Mitochondrial DNA Gene: Mitochondrial Targeting of Allotopically Expressed Mutant ATP6 in Transgenic Mice

    Directory of Open Access Journals (Sweden)

    David A. Dunn

    2012-01-01

    Full Text Available Nuclear encoding of mitochondrial DNA transgenes followed by mitochondrial targeting of the expressed proteins (allotopic expression; AE represents a potentially powerful strategy for creating animal models of mtDNA disease. Mice were created that allotopically express either a mutant (A6M or wildtype (A6W mt-Atp6 transgene. Compared to non-transgenic controls, A6M mice displayed neuromuscular and motor deficiencies (wire hang, pole, and balance beam analyses; P0.05. This study illustrates a mouse model capable of circumventing in vivo mitochondrial mutations. Moreover, it provides evidence supporting AE as a tool for mtDNA disease research with implications in development of DNA-based therapeutics.

  15. An Overview of Ten Italian Horse Breeds through Mitochondrial DNA

    Science.gov (United States)

    Capodiferro, Marco Rosario; Capomaccio, Stefano; Buttazzoni, Luca; Biggio, Giovanni Paolo; Cherchi, Raffaele; Albertini, Emidio; Olivieri, Anna; Cappelli, Katia; Achilli, Alessandro; Silvestrelli, Maurizio

    2016-01-01

    Background The climatic and cultural diversity of the Italian Peninsula triggered, over time, the development of a great variety of horse breeds, whose origin and history are still unclear. To clarify this issue, analyses on phenotypic traits and genealogical data were recently coupled with molecular screening. Methodology To provide a comprehensive overview of the horse genetic variability in Italy, we produced and phylogenetically analyzed 407 mitochondrial DNA (mtDNA) control-region sequences from ten of the most important Italian riding horse and pony breeds: Bardigiano, Esperia, Giara, Lipizzan, Maremmano, Monterufolino, Murgese, Sarcidano, Sardinian Anglo-Arab, and Tolfetano. A collection of 36 Arabian horses was also evaluated to assess the genetic consequences of their common use for the improvement of some local breeds. Conclusions In Italian horses, all previously described domestic mtDNA haplogroups were detected as well as a high haplotype diversity. These findings indicate that the ancestral local mares harbored an extensive genetic diversity. Moreover, the limited haplotype sharing (11%) with the Arabian horse reveals that its impact on the autochthonous mitochondrial gene pools during the final establishment of pure breeds was marginal, if any. The only significant signs of genetic structure and differentiation were detected in the geographically most isolated contexts (i.e. Monterufolino and Sardinian breeds). Such a geographic effect was also confirmed in a wider breed setting, where the Italian pool stands in an intermediate position together with most of the other Mediterranean stocks. However, some notable exceptions and peculiar genetic proximities lend genetic support to historical theories about the origin of specific Italian breeds. PMID:27054850

  16. DNA double-strand breaks activate ATM independent of mitochondrial dysfunction in A549 cells.

    Science.gov (United States)

    Kalifa, Lidza; Gewandter, Jennifer S; Staversky, Rhonda J; Sia, Elaine A; Brookes, Paul S; O'Reilly, Michael A

    2014-10-01

    Excessive nuclear or mitochondrial DNA damage can lead to mitochondrial dysfunction, decreased energy production, and increased generation of reactive oxygen species (ROS). Although numerous cell signaling pathways are activated when cells are injured, the ataxia telangiectasia mutant (ATM) protein has emerged as a major regulator of the response to both mitochondrial dysfunction and nuclear DNA double-strand breaks (DSBs). Because mitochondrial dysfunction is often a response to excessive DNA damage, it has been difficult to determine whether nuclear and/or mitochondrial DNA DSBs activate ATM independent of mitochondrial dysfunction. In this study, mitochondrial and nuclear DNA DSBs were generated in the A549 human lung adenocarcinoma cell line by infecting with retroviruses expressing the restriction endonuclease PstI fused to a mitochondrial targeting sequence (MTS) or nuclear localization sequence (NLS) and a hemagglutinin antigen epitope tag (HA). Expression of MTS-PstI-HA or NLS-PstI-HA activated the DNA damage response defined by phosphorylation of ATM, the tumor suppressor protein p53 (TP53), KRAB-associated protein (KAP)-1, and structural maintenance of chromosomes (SMC)-1. Phosphorylated ATM and SMC1 were detected in nuclear fractions, whereas phosphorylated TP53 and KAP1 were detected in both mitochondrial and nuclear fractions. PstI also enhanced expression of the cyclin-dependent kinase inhibitor p21 and inhibited cell growth. This response to DNA damage occurred in the absence of detectable mitochondrial dysfunction and excess production of ROS. These findings reveal that DNA DSBs are sufficient to activate ATM independent of mitochondrial dysfunction and suggest that the activated form of ATM and some of its substrates are restricted to the nuclear compartment, regardless of the site of DNA damage.

  17. The Mitochondrial DNA (mtDNA)-Associated Protein SWIB5 Influences mtDNA Architecture and Homologous Recombination

    KAUST Repository

    Blomme, Jonas

    2017-04-19

    In addition to the nucleus, mitochondria and chloroplasts in plant cells also contain genomes. Efficient DNA repair pathways are crucial in these organelles to fix damage resulting from endogenous and exogenous factors. Plant organellar genomes are complex compared with their animal counterparts, and although several plant-specific mediators of organelle DNA repair have been reported, many regulators remain to be identified. Here, we show that a mitochondrial SWI/SNF (nucleosome remodeling) complex B protein, SWIB5, is capable of associating with mitochondrial DNA (mtDNA) in Arabidopsis thaliana. Gainand loss-of-function mutants provided evidence for a role of SWIB5 in influencing mtDNA architecture and homologous recombination at specific intermediate-sized repeats both under normal and genotoxic conditions. SWIB5 interacts with other mitochondrial SWIB proteins. Gene expression and mutant phenotypic analysis of SWIB5 and SWIB family members suggests a link between organellar genome maintenance and cell proliferation. Taken together, our work presents a protein family that influences mtDNA architecture and homologous recombination in plants and suggests a link between organelle functioning and plant development.

  18. Mitochondrial DNA damage associated with lipid peroxidation of the mitochondrial membrane induced by Fe2+-citrate

    Directory of Open Access Journals (Sweden)

    Andréa M. Almeida

    2006-09-01

    Full Text Available Iron imbalance/accumulation has been implicated in oxidative injury associated with many degenerative diseases such as hereditary hemochromatosis, beta-thalassemia, and Friedreich's ataxia. Mitochondria are particularly sensitive to iron-induced oxidative stress - high loads of iron cause extensive lipid peroxidation and membrane permeabilization in isolated mitochondria. Here we detected and characterized mitochondrial DNA damage in isolated rat liver mitochondria exposed to a Fe2+-citrate complex, a small molecular weight complex. Intense DNA fragmentation was induced after the incubation of mitochondria with the iron complex. The detection of 3' phosphoglycolate ends at the mtDNA strand breaks by a 32P-postlabeling assay, suggested the involvement of hydroxyl radical in the DNA fragmentation induced by Fe2+-citrate. Increased levels of 8-oxo-7,8-dihydro-2'-deoxyguanosine also suggested that Fe2+-citrate-induced oxidative stress causes mitochondrial DNA damage. In conclusion, our results show that iron-mediated lipid peroxidation was associated with intense mtDNA damage derived from the direct attack of reactive oxygen species.Desequilíbrio/acúmulo de ferro tem sido implicado em injúria oxidativa associada a diversas doenças degenerativas tais como, hemocromatose hereditária, beta-talassemia e ataxia de Friedreich. As mitocôndrias são particularmente sensíveis a estresse oxidativo induzido por ferro - um carregamento alto de ferro em mitocôndrias isoladas pode causar uma extensiva peroxidação lipídica e a permeabilização de membrana. Nesse estudo, nós detectamos e caracterizamos danos do DNA mitocondrial em mitocôndrias isoladas de fígado de rato, expostas ao complexo Fe2+-citrato, um dos complexos de baixo peso molecular. A intensa fragmentação do DNA foi induzida após a incubação das mitocôndrias com o complexo de ferro. A detecção de finais 3' de fosfoglicolato nas quebras de fitas de DNA mitocondrial pelo ensaio 32

  19. Mutations in the SPG7 gene cause chronic progressive external ophthalmoplegia through disordered mitochondrial DNA maintenance.

    Science.gov (United States)

    Pfeffer, Gerald; Gorman, Gráinne S; Griffin, Helen; Kurzawa-Akanbi, Marzena; Blakely, Emma L; Wilson, Ian; Sitarz, Kamil; Moore, David; Murphy, Julie L; Alston, Charlotte L; Pyle, Angela; Coxhead, Jon; Payne, Brendan; Gorrie, George H; Longman, Cheryl; Hadjivassiliou, Marios; McConville, John; Dick, David; Imam, Ibrahim; Hilton, David; Norwood, Fiona; Baker, Mark R; Jaiser, Stephan R; Yu-Wai-Man, Patrick; Farrell, Michael; McCarthy, Allan; Lynch, Timothy; McFarland, Robert; Schaefer, Andrew M; Turnbull, Douglass M; Horvath, Rita; Taylor, Robert W; Chinnery, Patrick F

    2014-05-01

    Despite being a canonical presenting feature of mitochondrial disease, the genetic basis of progressive external ophthalmoplegia remains unknown in a large proportion of patients. Here we show that mutations in SPG7 are a novel cause of progressive external ophthalmoplegia associated with multiple mitochondrial DNA deletions. After excluding known causes, whole exome sequencing, targeted Sanger sequencing and multiplex ligation-dependent probe amplification analysis were used to study 68 adult patients with progressive external ophthalmoplegia either with or without multiple mitochondrial DNA deletions in skeletal muscle. Nine patients (eight probands) were found to carry compound heterozygous SPG7 mutations, including three novel mutations: two missense mutations c.2221G>A; p.(Glu741Lys), c.2224G>A; p.(Asp742Asn), a truncating mutation c.861dupT; p.Asn288*, and seven previously reported mutations. We identified a further six patients with single heterozygous mutations in SPG7, including two further novel mutations: c.184-3C>T (predicted to remove a splice site before exon 2) and c.1067C>T; p.(Thr356Met). The clinical phenotype typically developed in mid-adult life with either progressive external ophthalmoplegia/ptosis and spastic ataxia, or a progressive ataxic disorder. Dysphagia and proximal myopathy were common, but urinary symptoms were rare, despite the spasticity. Functional studies included transcript analysis, proteomics, mitochondrial network analysis, single fibre mitochondrial DNA analysis and deep re-sequencing of mitochondrial DNA. SPG7 mutations caused increased mitochondrial biogenesis in patient muscle, and mitochondrial fusion in patient fibroblasts associated with the clonal expansion of mitochondrial DNA mutations. In conclusion, the SPG7 gene should be screened in patients in whom a disorder of mitochondrial DNA maintenance is suspected when spastic ataxia is prominent. The complex neurological phenotype is likely a result of the clonal

  20. Catecholamine metabolism drives generation of mitochondrial DNA deletions in dopaminergic neurons.

    Science.gov (United States)

    Neuhaus, Johannes F G; Baris, Olivier R; Hess, Simon; Moser, Natasha; Schröder, Hannsjörg; Chinta, Shankar J; Andersen, Julie K; Kloppenburg, Peter; Wiesner, Rudolf J

    2014-02-01

    Accumulation of mitochondrial DNA deletions is observed especially in dopaminergic neurons of the substantia nigra during ageing and even more in Parkinson's disease. The resulting mitochondrial dysfunction is suspected to play an important role in neurodegeneration. However, the molecular mechanisms involved in the preferential generation of mitochondrial DNA deletions in dopaminergic neurons are still unknown. To study this phenomenon, we developed novel polymerase chain reaction strategies to detect distinct mitochondrial DNA deletions and monitor their accumulation patterns. Applying these approaches in in vitro and in vivo models, we show that catecholamine metabolism drives the generation and accumulation of these mitochondrial DNA mutations. As in humans, age-related accumulation of mitochondrial DNA deletions is most prominent in dopaminergic areas of mouse brain and even higher in the catecholaminergic adrenal medulla. Dopamine treatment of terminally differentiated neuroblastoma cells, as well as stimulation of dopamine turnover in mice over-expressing monoamine oxidase B both induce multiple mitochondrial DNA deletions. Our results thus identify catecholamine metabolism as the driving force behind mitochondrial DNA deletions, probably being an important factor in the ageing-associated degeneration of dopaminergic neurons.

  1. Mitochondrial DNA disease—molecular insights and potential routes to a cure

    Energy Technology Data Exchange (ETDEWEB)

    Russell, Oliver; Turnbull, Doug, E-mail: doug.turnbull@newcastle.ac.uk

    2014-07-01

    Mitochondrial DNA diseases are common neurological conditions caused by mutations in the mitochondrial genome or nuclear genes responsible for its maintenance. Current treatments for these disorders are focussed on the management of the symptoms, rather than the correction of biochemical defects caused by the mutation. This review focuses on the molecular effects of mutations, the symptoms they cause and current work focusing on the development of targeted treatments for mitochondrial DNA disease. - Highlights: • We discuss several common disease causing mtDNA mutations. • We highlight recent work linking pathogenicity to deletion size and heteroplasmy. • We discuss recent advances in the development of targeted mtDNA disease treatments.

  2. Overexpression of DNA ligase III in mitochondria protects cells against oxidative stress and improves mitochondrial DNA base excision repair

    DEFF Research Database (Denmark)

    Akbari, Mansour; Keijzers, Guido; Maynard, Scott

    2014-01-01

    slower than the preceding mitochondrial BER steps. Overexpression of DNA ligase III in mitochondria improved the rate of overall BER, increased cell survival after menadione induced oxidative stress and reduced autophagy following the inhibition of the mitochondrial electron transport chain complex I...... by rotenone. Our results suggest that the amount of DNA ligase III in mitochondria may be critical for cell survival following prolonged oxidative stress, and demonstrate a functional link between mitochondrial DNA damage and repair, cell survival upon oxidative stress, and removal of dysfunctional......Base excision repair (BER) is the most prominent DNA repair pathway in human mitochondria. BER also results in a temporary generation of AP-sites, single-strand breaks and nucleotide gaps. Thus, incomplete BER can result in the generation of DNA repair intermediates that can disrupt mitochondrial...

  3. Unveiling the mystery of mitochondrial DNA replication in yeasts.

    Science.gov (United States)

    Chen, Xin Jie; Clark-Walker, George Desmond

    2017-08-01

    Conventional DNA replication is initiated from specific origins and requires the synthesis of RNA primers for both the leading and lagging strands. In contrast, the replication of yeast mitochondrial DNA is origin-independent. The replication of the leading strand is likely primed by recombinational structures and proceeded by a rolling circle mechanism. The coexistent linear and circular DNA conformers facilitate the recombination-based initiation. The replication of the lagging strand is poorly understood. Re-evaluation of published data suggests that the rolling circle may also provide structures for the synthesis of the lagging-strand by mechanisms such as template switching. Thus, the coupling of recombination with rolling circle replication and possibly, template switching, may have been selected as an economic replication mode to accommodate the reductive evolution of mitochondria. Such a replication mode spares the need for conventional replicative components, including those required for origin recognition/remodelling, RNA primer synthesis and lagging-strand processing. Copyright © 2017 Elsevier B.V. and Mitochondria Research Society. All rights reserved.

  4. Analysis of mitochondrial DNA sequences in patients with isolated or combined oxidative phosphorylation system deficiency.

    NARCIS (Netherlands)

    Hinttala, R.; Smeets, R.; Moilanen, J.S.; Ugalde, C.; Uusimaa, J.; Smeitink, J.A.M.; Majamaa, K.

    2006-01-01

    BACKGROUND: Enzyme deficiencies of the oxidative phosphorylation (OXPHOS) system may be caused by mutations in the mitochondrial DNA (mtDNA) or in the nuclear DNA. OBJECTIVE: To analyse the sequences of the mtDNA coding region in 25 patients with OXPHOS system deficiency to identify the underlying g

  5. Genetic variability of Taenia saginata inferred from mitochondrial DNA sequences.

    Science.gov (United States)

    Rostami, Sima; Salavati, Reza; Beech, Robin N; Babaei, Zahra; Sharbatkhori, Mitra; Harandi, Majid Fasihi

    2015-04-01

    Taenia saginata is an important tapeworm, infecting humans in many parts of the world. The present study was undertaken to identify inter- and intraspecific variation of T. saginata isolated from cattle in different parts of Iran using two mitochondrial CO1 and 12S rRNA genes. Up to 105 bovine specimens of T. saginata were collected from 20 slaughterhouses in three provinces of Iran. DNA were extracted from the metacestode Cysticercus bovis. After PCR amplification, sequencing of CO1 and 12S rRNA genes were carried out and two phylogenetic analyses of the sequence data were generated by Bayesian inference on CO1 and 12S rRNA sequences. Sequence analyses of CO1 and 12S rRNA genes showed 11 and 29 representative profiles respectively. The level of pairwise nucleotide variation between individual haplotypes of CO1 gene was 0.3-2.4% while the overall nucleotide variation among all 11 haplotypes was 4.6%. For 12S rRNA sequence data, level of pairwise nucleotide variation was 0.2-2.5% and the overall nucleotide variation was determined as 5.8% among 29 haplotypes of 12S rRNA gene. Considerable genetic diversity was found in both mitochondrial genes particularly in 12S rRNA gene.

  6. Complete elimination of maternal mitochondrial DNA during meiosis resulting in the paternal inheritance of the mitochondrial genome in Chlamydomonas species.

    Science.gov (United States)

    Aoyama, H; Hagiwara, Y; Misumi, O; Kuroiwa, T; Nakamura, S

    2006-09-01

    The non-Mendelian inheritance of organellar DNA is common in most plants and animals. In the isogamous green alga Chlamydomonas species, progeny inherit chloroplast genes from the maternal parent, as paternal chloroplast genes are selectively eliminated in young zygotes. Mitochondrial genes are inherited from the paternal parent. Analogically, maternal mitochondrial DNA (mtDNA) is thought to be selectively eliminated. Nevertheless, it is unclear when this selective elimination occurs. Here, we examined the behaviors of maternal and paternal mtDNAs by various methods during the period between the beginning of zygote formation and zoospore formation. First, we observed the behavior of the organelle nucleoids of living cells by specifically staining DNA with the fluorochrome SYBR Green I and staining mitochondria with 3,3'-dihexyloxacarbocyanine iodide. We also examined the fate of mtDNA of male and female parental origin by real-time PCR, nested PCR with single zygotes, and fluorescence in situ hybridization analysis. The mtDNA of maternal origin was completely eliminated before the first cell nuclear division, probably just before mtDNA synthesis, during meiosis. Therefore, the progeny inherit the remaining paternal mtDNA. We suggest that the complete elimination of maternal mtDNA during meiosis is the primary cause of paternal mitochondrial inheritance.

  7. Linguistic isolates in Portugal: insights from the mitochondrial DNA pattern.

    Science.gov (United States)

    Mairal, Quim; Santos, Cristina; Silva, Marina; Marques, Sofia L; Ramos, Amanda; Aluja, Maria Pilar; Amorim, Antonio; Prata, Maria João; Alvarez, Luis

    2013-12-01

    Miranda do Douro, located in the northeastern region of Portugal, has notable characteristics not only from a geographic or naturalistic point of view, but also from a cultural perspective. A remarkable one is the coexistence of two different languages: Portuguese and Mirandese, the second being an Astur-Leonese dialect. The current persistence of the Astur-Leonese dialect in this population falls on the singularity of the region: relative isolation, implying difficulties to communicate with other Portuguese regions, while the same location facilitated the establishment of social and commercial relationships with adjacent Spanish territories, origin of the Astur-Leonese language. The objective of this study was to characterize the population from Miranda through the analysis of maternal lineages in order to evaluate whether its mitochondrial DNA diversity fitted the patterns previously reported for other populations from the Iberian Peninsula. Viewing that, the entire control region of mitochondrial DNA from 121 individuals was examined. Miranda showed a haplogroup composition usual for a Western European population, in the sense that as high as 63.6% of sequences belonged to macro-haplogroup R0. Lineages ascribed to have an African (L2a and L1b) origin, were detected, but reaching an amount commonly found in Portugal. Miranda also presented a few haplogroups typically found in Jewish populations, while rarely observed in other Iberian populations. The finding can be explained by gene flow with crypto-Jew communities that since long are known to be established in the region where Miranda is located. In Miranda, both genetic and nucleotide diversities presented low values (0.9292 ± 0.0180 and 0.01101 ± 0.00614 respectively) when compared to populations from its micro-geographical framework, which constitute a sign of population isolation that certainly provided conditions for the survival of the Astur-Leonese dialect in the region. Copyright © 2013 Elsevier

  8. Statistical analysis of post mortem DNA damage-derived miscoding lesions in Neandertal mitochondrial DNA

    DEFF Research Database (Denmark)

    Vives, Sergi; Gilbert, M Thomas; Arenas, Conchita

    2008-01-01

    ABSTRACT: BACKGROUND: We have analysed the distribution of post mortem DNA damage derived miscoding lesions from the datasets of seven published Neandertal specimens that have extensive cloned sequence coverage over the mitochondrial DNA (mtDNA) hypervariable region 1 (HVS1). The analysis...... was restricted to C-->T and G-->A miscoding lesions (the predominant manifestation of post mortem damage) that are seen at a frequency of more than one clone among sequences from a single PCR, but do not represent the true endogenous sequence. FINDINGS: The data indicates an extreme bias towards C-->T over G......-->A miscoding lesions (observed ratio of 67:2 compared to an expected ratio of 7:2), implying that the mtDNA Light strand molecule suffers proportionally more damage-derived miscoding lesions than the Heavy strand. CONCLUSION: The clustering of Cs in the Light strand as opposed to the singleton pattern of Cs...

  9. Peripheral Blood Mitochondrial DNA as a Biomarker of Cerebral Mitochondrial Dysfunction following Traumatic Brain Injury in a Porcine Model.

    Directory of Open Access Journals (Sweden)

    Todd J Kilbaugh

    Full Text Available Traumatic brain injury (TBI has been shown to activate the peripheral innate immune system and systemic inflammatory response, possibly through the central release of damage associated molecular patterns (DAMPs. Our main purpose was to gain an initial understanding of the peripheral mitochondrial response following TBI, and how this response could be utilized to determine cerebral mitochondrial bioenergetics. We hypothesized that TBI would increase peripheral whole blood relative mtDNA copy number, and that these alterations would be associated with cerebral mitochondrial bioenergetics triggered by TBI.Blood samples were obtained before, 6 h after, and 25 h after focal (controlled cortical impact injury: CCI and diffuse (rapid non-impact rotational injury: RNR TBI. PCR primers, unique to mtDNA, were identified by aligning segments of nuclear DNA (nDNA to mtDNA, normalizing values to nuclear 16S rRNA, for a relative mtDNA copy number. Three unique mtDNA regions were selected, and PCR primers were designed within those regions, limited to 25-30 base pairs to further ensure sequence specificity, and measured utilizing qRT-PCR.Mean relative mtDNA copy numbers increased significantly at 6 and 25 hrs after following both focal and diffuse traumatic brain injury. Specifically, the mean relative mtDNA copy number from three mitochondrial-specific regions pre-injury was 0.84 ± 0.05. At 6 and 25 h after diffuse non-impact TBI, mean mtDNA copy number was significantly higher: 2.07 ± 0.19 (P < 0.0001 and 2.37 ± 0.42 (P < 0.001, respectively. Following focal impact TBI, relative mtDNA copy number was also significantly higher, 1.35 ± 0.12 (P < 0.0001 at 25 hours. Alterations in mitochondrial respiration in the hippocampus and cortex post-TBI correlated with changes in the relative mtDNA copy number measured in peripheral blood.Alterations in peripheral blood relative mtDNA copy numbers may be a novel biosignature of cerebral mitochondrial bioenergetics

  10. Mitochondrial replacement therapies can circumvent mtDNA based disease transmission

    Science.gov (United States)

    Wolf, Don P.; Mitalipov, Shoukhrat

    2014-01-01

    Mitochondrial DNA diseases are relatively common, sometimes devastating and transmitted exclusively through the egg to children of carrier mothers. The study by Wang et al. (2014) adds the exciting possibility of a new therapy for preventing mitochondrial disease transmission predicated on the use of polar body genomes in mice. Mitochondrial DNA diseases are relatively common, sometimes devastating and transmitted exclusively through the egg to children of carrier mothers. The study by Wang et al. (2014) adds the exciting possibility of a new therapy for preventing mitochondrial disease transmission predicated on the use of polar body genomesin mice. PMID:24988456

  11. Iron deficiency and iron excess damage mitochondria and mitochondrial DNA in rats.

    Science.gov (United States)

    Walter, Patrick B; Knutson, Mitchell D; Paler-Martinez, Andres; Lee, Sonia; Xu, Yu; Viteri, Fernando E; Ames, Bruce N

    2002-02-19

    Approximately two billion people, mainly women and children, are iron deficient. Two studies examined the effects of iron deficiency and supplementation on rats. In study 1, mitochondrial functional parameters and mitochondrial DNA (mtDNA) damage were assayed in iron-deficient (mitochondrial respiratory control ratios and increased levels of oxidants in polymorphonuclear-leukocytes, as assayed by dichlorofluorescein (P mitochondrial malfunction. Although excess iron has been known to cause oxidative damage, the observation of oxidant-induced damage to mitochondria from iron deficiency has been unrecognized previously. Untreated iron deficiency, as well as excessive-iron supplementation, are deleterious and emphasize the importance of maintaining optimal iron intake.

  12. Human mitochondrial mTERF wraps around DNA through a left-handed superhelical tandem repeat.

    Science.gov (United States)

    Jiménez-Menéndez, Nereida; Fernández-Millán, Pablo; Rubio-Cosials, Anna; Arnan, Carme; Montoya, Julio; Jacobs, Howard T; Bernadó, Pau; Coll, Miquel; Usón, Isabel; Solà, Maria

    2010-07-01

    The regulation of mitochondrial DNA (mtDNA) processes is slowly being characterized at a structural level. We present here crystal structures of human mitochondrial regulator mTERF, a transcription termination factor also implicated in replication pausing, in complex with double-stranded DNA oligonucleotides containing the tRNA(Leu)(UUR) gene sequence. mTERF comprises nine left-handed helical tandem repeats that form a left-handed superhelix, the Zurdo domain.

  13. Mitochondrial DNA response to high altitude: a new perspective on high-altitude adaptation.

    Science.gov (United States)

    Luo, Yongjun; Yang, Xiaohong; Gao, Yuqi

    2013-08-01

    Mitochondria are the energy metabolism centers of the cell. More than 95% of cellular energy is produced by mitochondrial oxidative phosphorylation. Hypoxia affects a wide range of energy generation and consumption processes in animals. The most important mechanisms limiting ATP consumption increase the efficiency of ATP production and accommodate the reduced production of ATP by the body. All of these mechanisms relate to changes in mitochondrial function. Mitochondrial function can be affected by variations in mitochondrial DNA, including polymorphisms, content changes, and deletions. These variations play an important role in acclimatization or adaptation to hypoxia. In this paper, the association between mitochondrial genome sequences and high-altitude adaptation is reviewed.

  14. Concise Review: Heteroplasmic Mitochondrial DNA Mutations and Mitochondrial Diseases: Toward iPSC-Based Disease Modeling, Drug Discovery, and Regenerative Therapeutics.

    Science.gov (United States)

    Hatakeyama, Hideyuki; Goto, Yu-Ichi

    2016-04-01

    Mitochondria contain multiple copies of their own genome (mitochondrial DNA; mtDNA). Once mitochondria are damaged by mutant mtDNA, mitochondrial dysfunction is strongly induced, followed by symptomatic appearance of mitochondrial diseases. Major genetic causes of mitochondrial diseases are defects in mtDNA, and the others are defects of mitochondria-associating genes that are encoded in nuclear DNA (nDNA). Numerous pathogenic mutations responsible for various types of mitochondrial diseases have been identified in mtDNA; however, it remains uncertain why mitochondrial diseases present a wide variety of clinical spectrum even among patients carrying the same mtDNA mutations (e.g., variations in age of onset, in affected tissues and organs, or in disease progression and phenotypic severity). Disease-relevant induced pluripotent stem cells (iPSCs) derived from mitochondrial disease patients have therefore opened new avenues for understanding the definitive genotype-phenotype relationship of affected tissues and organs in various types of mitochondrial diseases triggered by mtDNA mutations. In this concise review, we briefly summarize several recent approaches using patient-derived iPSCs and their derivatives carrying various mtDNA mutations for applications in human mitochondrial disease modeling, drug discovery, and future regenerative therapeutics.

  15. Complete DNA sequences of the mitochondrial genomes of the pathogenic yeasts Candida orthopsilosis and Candida metapsilosis: insight into the evolution of linear DNA genomes from mitochondrial telomere mutants.

    Science.gov (United States)

    Kosa, Peter; Valach, Matus; Tomaska, Lubomir; Wolfe, Kenneth H; Nosek, Jozef

    2006-01-01

    We determined complete mitochondrial DNA sequences of the two yeast species, Candida orthopsilosis and Candida metapsilosis, and compared them with the linear mitochondrial genome of their close relative, C.parapsilosis. Mitochondria of all the three species harbor compact genomes encoding the same set of genes arranged in the identical order. Differences in the length of these genomes result mainly from the presence/absence of introns. Multiple alterations were identified also in the sequences of the ribosomal and transfer RNAs, and proteins. However, the most striking feature of C.orthopsilosis and C.metapsilosis is the existence of strains differing in the molecular form of the mitochondrial genome (circular-mapping versus linear). Their analysis opens a unique window for understanding the role of mitochondrial telomeres in the stability and evolution of molecular architecture of the genome. Our results indicate that the circular-mapping mitochondrial genome derived from the linear form by intramolecular end-to-end fusions. Moreover, we suggest that the linear mitochondrial genome evolved from a circular-mapping form present in a common ancestor of the three species and, at the same time, the emergence of mitochondrial telomeres enabled the formation of linear monomeric DNA forms. In addition, comparison of isogenic C.metapsilosis strains differing in the form of the organellar genome suggests a possibility that, under some circumstances, the linearity and/or the presence of telomeres provide a competitive advantage over a circular-mapping mitochondrial genome.

  16. The exonuclease activity of DNA polymerase γ is required for ligation during mitochondrial DNA replication

    Science.gov (United States)

    Macao, Bertil; Uhler, Jay P.; Siibak, Triinu; Zhu, Xuefeng; Shi, Yonghong; Sheng, Wenwen; Olsson, Monica; Stewart, James B.; Gustafsson, Claes M.; Falkenberg, Maria

    2015-01-01

    Mitochondrial DNA (mtDNA) polymerase γ (POLγ) harbours a 3′–5′ exonuclease proofreading activity. Here we demonstrate that this activity is required for the creation of ligatable ends during mtDNA replication. Exonuclease-deficient POLγ fails to pause on reaching a downstream 5′-end. Instead, the enzyme continues to polymerize into double-stranded DNA, creating an unligatable 5′-flap. Disease-associated mutations can both increase and decrease exonuclease activity and consequently impair DNA ligation. In mice, inactivation of the exonuclease activity causes an increase in mtDNA mutations and premature ageing phenotypes. These mutator mice also contain high levels of truncated, linear fragments of mtDNA. We demonstrate that the formation of these fragments is due to impaired ligation, causing nicks near the origin of heavy-strand DNA replication. In the subsequent round of replication, the nicks lead to double-strand breaks and linear fragment formation. PMID:26095671

  17. Cellular models for mitochondrial DNA-based diseases: lymphoblastoid cell lines and transmitochondrial cybrids.

    Science.gov (United States)

    Jiji, Sun; Xiaoxu, Zhao; Lihua, Qiao; Shuang, Mei; Zhipeng, Nie; Qinghai, Zhang; Yanchun, Ji; Pingping, Jiang; Min-Xin, Guan

    2016-07-20

    Mitochondrial DNA (mtDNA) mutations cause a variety of mitochondrial DNA-based diseases which have been studied using Lymphoblastoid cell lines (LCLs) and transmitochondrial cybrids. Individual genetic information is preserved permanently in LCLs while the development of transmitochondrial cybrids provide ex-vivo cellular platform to study molecular mechanism of mitochondrial DNA-based diseases. The cytoplasmic donor cells for previous transmitochondrial cybrids come from patient's tissue or platelet directly. Here, we depicted in details the principle, methods and techniques to establish LCLs from frozen peripheral bloods harboring mitochondrial 4401G > A mutation by infection of Epstein Barr virus, and then to generate cybrids using ρ(0) 206 and LCLs. The process of establishing these two cellular models was summarized into four steps as follows: (1) Generation of LCLs; (2) Transformation; (3) Selection; (4) Verification. To faithfully represent the function of mtDNA mutation, we analyzed and identified the sites of mtDNA mutations and copy numbers of each cellular models as well as the karyotype of transmitochondrial cybrids. Those clones with consistent parameters were selected for preservation and future analysis of the function of point mutations of mtDNA. Although these two cellular models play important roles in understanding molecular mechanism of mitochondrial DNA-based diseases on the cellular level, their limitations should be considered when elucidating the character of tissue specificity of mitochondrial DNA-based diseases.

  18. Inhibiting Mitochondrial DNA Ligase IIIα Activates Caspase 1-Dependent Apoptosis in Cancer Cells.

    Science.gov (United States)

    Sallmyr, Annahita; Matsumoto, Yoshihiro; Roginskaya, Vera; Van Houten, Bennett; Tomkinson, Alan E

    2016-09-15

    Elevated levels of DNA ligase IIIα (LigIIIα) have been identified as a biomarker of an alteration in DNA repair in cancer cells that confers hypersensitivity to a LigIIIα inhibitor, L67, in combination with a poly (ADP-ribose) polymerase inhibitor. Because LigIIIα functions in the nucleus and mitochondria, we examined the effect of L67 on these organelles. Here, we show that, although the DNA ligase inhibitor selectively targets mitochondria, cancer and nonmalignant cells respond differently to disruption of mitochondrial DNA metabolism. Inhibition of mitochondrial LigIIIα in cancer cells resulted in abnormal mitochondrial morphology, reduced levels of mitochondrial DNA, and increased levels of mitochondrially generated reactive oxygen species that caused nuclear DNA damage. In contrast, these effects did not occur in nonmalignant cells. Furthermore, inhibition of mitochondrial LigIIIα activated a caspase 1-dependent apoptotic pathway, which is known to be part of inflammatory responses induced by pathogenic microorganisms in cancer, but not nonmalignant cells. These results demonstrate that the disruption of mitochondrial DNA metabolism elicits different responses in nonmalignant and cancer cells and suggests that the abnormal response in cancer cells may be exploited in the development of novel therapeutic strategies that selectively target cancer cells. Cancer Res; 76(18); 5431-41. ©2016 AACR.

  19. Limited clinical relevance of mitochondrial DNA mutation and gene expression analyses in ovarian cancer

    Directory of Open Access Journals (Sweden)

    Rachinger Andrea

    2008-10-01

    Full Text Available Abstract Background In recent years, numerous studies have investigated somatic mutations in mitochondrial DNA in various tumours. The observed high mutation rates might reflect mitochondrial deregulation; consequently, mutation analyses could be clinically relevant. The purpose of this study was to determine if mutations in the mitochondrial D-loop region and/or the level of mitochondrial gene expression could influence the clinical course of human ovarian carcinomas. Methods We sequenced a 1320-base-pair DNA fragment of the mitochondrial genome (position 16,000-750 in 54 cancer samples and in 44 corresponding germline control samples. In addition, six transcripts (MT-ATP6, MT-CO1, MT-CYB, MT-ND1, MT-ND6, and MT-RNR1 were quantified in 62 cancer tissues by real-time RT-PCR. Results Somatic mutations in the D-loop sequence were found in 57% of ovarian cancers. Univariate analysis showed no association between mitochondrial DNA mutation status or mitochondrial gene expression and any of the examined clinicopathologic parameters. A multivariate logistic regression model revealed that the expression of the mitochondrial gene RNR1 might be used as a predictor of tumour sensitivity to chemotherapy. Conclusion In contrast to many previously published papers, our study indicates rather limited clinical relevance of mitochondrial molecular analyses in ovarian carcinomas. These discrepancies in the clinical utility of mitochondrial molecular tests in ovarian cancer require additional large, well-designed validation studies.

  20. Intracellular evolution of mitochondrial DNA (mtDNA) and the tragedy of the cytoplasmic commons.

    Science.gov (United States)

    Haig, David

    2016-06-01

    Mitochondria exist in large numbers per cell. Therefore, the strength of natural selection on individual mtDNAs for their contribution to cellular fitness is weak whereas the strength of selection in favor of mtDNAs that increase their own replication without regard for cellular functions is strong. This problem has been solved for most mitochondrial genes by their transfer to the nucleus but a few critical genes remain encoded by mtDNA. Organisms manage the evolution of mtDNA to prevent mutational decay of essential services mitochondria provide to their hosts. Bottlenecks of mitochondrial numbers in female germlines increase the homogeneity of mtDNAs within cells and allow intraorganismal selection to eliminate cells with low quality mitochondria. Mechanisms of intracellular "quality control" allow direct selection on the competence of individual mtDNAs. These processes maintain the integrity of mtDNAs within the germline but are inadequate to indefinitely maintain mitochondrial function in somatic cells. © 2016 WILEY Periodicals, Inc.

  1. Overexpression of DNA ligase III in mitochondria protects cells against oxidative stress and improves mitochondrial DNA base excision repair.

    Science.gov (United States)

    Akbari, Mansour; Keijzers, Guido; Maynard, Scott; Scheibye-Knudsen, Morten; Desler, Claus; Hickson, Ian D; Bohr, Vilhelm A

    2014-04-01

    Base excision repair (BER) is the most prominent DNA repair pathway in human mitochondria. BER also results in a temporary generation of AP-sites, single-strand breaks and nucleotide gaps. Thus, incomplete BER can result in the generation of DNA repair intermediates that can disrupt mitochondrial DNA replication and transcription and generate mutations. We carried out BER analysis in highly purified mitochondrial extracts from human cell lines U2OS and HeLa, and mouse brain using a circular DNA substrate containing a lesion at a specific position. We found that DNA ligation is significantly slower than the preceding mitochondrial BER steps. Overexpression of DNA ligase III in mitochondria improved the rate of overall BER, increased cell survival after menadione induced oxidative stress and reduced autophagy following the inhibition of the mitochondrial electron transport chain complex I by rotenone. Our results suggest that the amount of DNA ligase III in mitochondria may be critical for cell survival following prolonged oxidative stress, and demonstrate a functional link between mitochondrial DNA damage and repair, cell survival upon oxidative stress, and removal of dysfunctional mitochondria by autophagy.

  2. The importance of mitochondrial DNA in aging and cancer

    DEFF Research Database (Denmark)

    Madsen, Claus Desler; Espersen, Maiken Lise Marcker; Singh, Keshav K

    2011-01-01

    Mitochondrial dysfunction has been implicated in premature aging, age-related diseases, and tumor initiation and progression. Alterations of the mitochondrial genome accumulate both in aging tissue and tumors. This paper describes our contemporary view of mechanisms by which alterations...... of the mitochondrial genome contributes to the development of age- and tumor-related pathological conditions. The mechanisms described encompass altered production of mitochondrial ROS, altered regulation of the nuclear epigenome, affected initiation of apoptosis, and a limiting effect on the production...

  3. Mitochondrial DNA in CSF distinguishes LRRK2 from idiopathic Parkinson's disease.

    Science.gov (United States)

    Podlesniy, Petar; Vilas, Dolores; Taylor, Peggy; Shaw, Leslie M; Tolosa, Eduard; Trullas, Ramon

    2016-10-01

    Mitochondrial DNA regulates mitochondrial function which is altered in both idiopathic and familial forms of Parkinson's disease. To investigate whether these two disease forms exhibit an altered regulation of mitochondrial DNA we measured cell free mitochondrial DNA content in cerebrospinal fluid (CSF) from idiopathic and LRRK2-related Parkinson's disease patients. The concentration of mitochondrial DNA was measured using a digital droplet polymerase chain reaction technique in a total of 98 CSF samples from a cohort of subjects including: 20 LRRK2(G2019S) mutation carriers with Parkinson's disease, 26 asymptomatic LRRK2(G2019S) mutation carriers, 31 patients with idiopathic Parkinson's disease and 21 first-degree relatives of LRRK2 Parkinson's disease patients without the mutation. Here we report that LRRK2(G2019S) mutation carriers with Parkinson's disease exhibit a high concentration of mitochondrial DNA in CSF compared with asymptomatic LRRK2(G2019S) mutation carriers and with idiopathic Parkinson's disease patients. In addition, idiopathic, but not LRRK2 Parkinson's disease is associated with low CSF concentration of α-synuclein. These results show that high mitochondrial DNA content in CSF distinguishes idiopathic from LRRK2-related Parkinson's disease suggesting that different biochemical pathways underlie neurodegeneration in these two disorders.

  4. Mitochondrial DNA polymerase gamma variants in idiopathic sporadic Parkinson disease.

    Science.gov (United States)

    Luoma, P T; Eerola, J; Ahola, S; Hakonen, A H; Hellström, O; Kivistö, K T; Tienari, P J; Suomalainen, A

    2007-09-11

    Dysfunction of mitochondrial DNA polymerase gamma (POLG) has been recently recognized as an important cause of inherited neurodegenerative diseases. We have reported dominant and recessive inheritance of parkinsonism, mitochondrial myopathy, and premature amenorrhea in five ethnically distinct families with POLG1 mutations. This prompted us to carry out a detailed analysis of the coding region and intron-exon boundaries of POLG1 in Finnish patients with idiopathic sporadic Parkinson disease (PD) and in nonparkinsonian controls. The coding region of POLG1 was analyzed in 140 Finnish patients with PD and their 127 spouses as age- and ethnically matched controls. Further, we analyzed the intragenic CAG-repeat region of POLG1 in 126 additional patients with nonparkinsonian neurologic disorders and in 516 Finnish population controls. We found clustering of rare variants of the POLG1 CAG-repeat, encoding a polyglutamine tract, in Finnish patients with idiopathic PD as compared to their spouses (p = 0.003; OR 3.01, 95% CI 1.35 to 6.71), population controls (p = 0.001; OR 2.45, 95% CI 1.45 to 4.14), and patients with nonparkinsonian neurologic disorders (p = 0.05, OR 1.98, 95% CI 0.97 to 4.05). We found several amino acid substitutions, none of them associating with PD. These included a previously parkinsonism-associated POLG variant Y831C, found in one patient with PD, but also in five controls, suggesting that it is a neutral amino acid polymorphism. Our results suggest that POLG polyglutamine tract variants should be considered as a predisposing genetic factor in idiopathic sporadic Parkinson disease.

  5. Biparental inheritance of organelles in Pelargonium: evidence for intergenomic recombination of mitochondrial DNA.

    Science.gov (United States)

    Apitz, Janina; Weihe, Andreas; Pohlheim, Frank; Börner, Thomas

    2013-02-01

    While uniparental transmission of mtDNA is widespread and dominating in eukaryotes leaving mutation as the major source of genotypic diversity, recently, biparental inheritance of mitochondrial genes has been demonstrated in reciprocal crosses of Pelargonium zonale and P. inquinans. The thereby arising heteroplasmy carries the potential for recombination between mtDNAs of different descent, i.e. between the parental mitochondrial genomes. We have analyzed these Pelargonium hybrids for mitochondrial intergenomic recombination events by examining differences in DNA blot hybridization patterns of the mitochondrial genes atp1 and cob. Further investigation of these genes and their flanking regions using nucleotide sequence polymorphisms and PCR revealed DNA segments in the progeny, which contained both P. zonale and P. inquinans sequences suggesting an intergenomic recombination in hybrids of Pelargonium. This turns Pelargonium into an interesting subject for studies of recombination and evolutionary dynamics of mitochondrial genomes.

  6. Psychiatric symptoms of patients with primary mitochondrial DNA disorders

    Directory of Open Access Journals (Sweden)

    Inczedy-Farkas Gabriella

    2012-02-01

    Full Text Available Abstract Background The aim of our study was to assess psychiatric symptoms in patients with genetically proven primary mutation of the mitochondrial DNA. Methods 19 adults with known mitochondrial mutation (MT have been assessed with the Stanford Health Assessment Questionnaire 20-item Disability Index (HAQ-DI, the Symptom Check List-90-Revised (SCL-90-R, the Beck Depression Inventory-Short Form (BDI-SF, the Hamilton Depression Rating Scale (HDRS and the clinical version of the Structured Clinical Interview for the the DSM-IV (SCID-I and SCID-II As control, 10 patients with hereditary sensorimotor neuropathy (HN, harboring the peripheral myelin protein-22 (PMP22 mutation were examined with the same tools. Results The two groups did not differ significantly in gender, age or education. Mean HAQ-DI score was 0.82 in the MT (range: 0-1.625 and 0.71 in the HN group (range: 0-1.625. Level of disability between the two groups did not differ significantly (p = 0.6076. MT patients scored significantly higher on the BDI-SF and HDRS than HN patients (12.85 versus 4.40, p = 0.031, and 15.62 vs 7.30, p = 0.043, respectively. The Global Severity Index (GSI of SCL-90-R also showed significant difference (1.44 vs 0.46, p = 0.013 as well as the subscales except for somatization. SCID-I interview yielded a variety of mood disorders in both groups. Eight MT patient (42% had past, 6 (31% had current, 5 (26% had both past and current psychiatric diagnosis, yielding a lifetime prevalence of 9/19 (47% in the MT group. In the HN group, 3 patients had both past and current diagnosis showing a lifetime prevalence of 3/10 (30% in this group. SCID-II detected personality disorder in 8 MT cases (42%, yielding 3 avoidant, 2 obsessive-compulsive and 3 personality disorder not otherwise specified (NOS diagnosis. No personality disorder was identified in the HN group. Conclusions Clinicians should be aware of the high prevalence of psychiatric symptoms in patients with

  7. Involvement of DNA ligase III and ribonuclease H1 in mitochondrial DNA replication in cultured human cells.

    Science.gov (United States)

    Ruhanen, Heini; Ushakov, Kathy; Yasukawa, Takehiro

    2011-12-01

    Recent evidence suggests that coupled leading and lagging strand DNA synthesis operates in mammalian mitochondrial DNA (mtDNA) replication, but the factors involved in lagging strand synthesis are largely uncharacterised. We investigated the effect of knockdown of the candidate proteins in cultured human cells under conditions where mtDNA appears to replicate chiefly via coupled leading and lagging strand DNA synthesis to restore the copy number of mtDNA to normal levels after transient mtDNA depletion. DNA ligase III knockdown attenuated the recovery of mtDNA copy number and appeared to cause single strand nicks in replicating mtDNA molecules, suggesting the involvement of DNA ligase III in Okazaki fragment ligation in human mitochondria. Knockdown of ribonuclease (RNase) H1 completely prevented the mtDNA copy number restoration, and replication intermediates with increased single strand nicks were readily observed. On the other hand, knockdown of neither flap endonuclease 1 (FEN1) nor DNA2 affected mtDNA replication. These findings imply that RNase H1 is indispensable for the progression of mtDNA synthesis through removing RNA primers from Okazaki fragments. In the nucleus, Okazaki fragments are ligated by DNA ligase I, and the RNase H2 is involved in Okazaki fragment processing. This study thus proposes that the mitochondrial replication system utilises distinct proteins, DNA ligase III and RNase H1, for Okazaki fragment maturation.

  8. Nuclear transfer to prevent mitochondrial DNA disorders : revisiting the debate on reproductive cloning

    NARCIS (Netherlands)

    Bredenoord, A. L.; Dondorp, W.; Pennings, G.; De Wert, G.

    2011-01-01

    Preclinical experiments are currently performed to examine the feasibility of several types of nuclear transfer to prevent mitochondrial DNA (mtDNA) disorders. Whereas the two most promising types of nuclear transfer to prevent mtDNA disorders, spindle transfer and pronuclear transfer, do not amount

  9. Distinct structural features of TFAM drive mitochondrial DNA packaging versus transcriptional activation.

    Science.gov (United States)

    Ngo, Huu B; Lovely, Geoffrey A; Phillips, Rob; Chan, David C

    2014-01-01

    TFAM (transcription factor A, mitochondrial) is a DNA-binding protein that activates transcription at the two major promoters of mitochondrial DNA (mtDNA)--the light strand promoter (LSP) and the heavy strand promoter 1 (HSP1). Equally important, it coats and packages the mitochondrial genome. TFAM has been shown to impose a U-turn on LSP DNA; however, whether this distortion is relevant at other sites is unknown. Here we present crystal structures of TFAM bound to HSP1 and to nonspecific DNA. In both, TFAM similarly distorts the DNA into a U-turn. Yet, TFAM binds to HSP1 in the opposite orientation from LSP explaining why transcription from LSP requires DNA bending, whereas transcription at HSP1 does not. Moreover, the crystal structures reveal dimerization of DNA-bound TFAM. This dimerization is dispensable for DNA bending and transcriptional activation but is important in DNA compaction. We propose that TFAM dimerization enhances mitochondrial DNA compaction by promoting looping of the DNA.

  10. Simultaneous DNA and RNA mapping of somatic mitochondrial mutations across diverse human cancers

    DEFF Research Database (Denmark)

    Stewart, James B.; Alaei-Mahabadi, Babak; Radhakrishnan, Sabarinathan;

    2015-01-01

    Somatic mutations in the nuclear genome are required for tumor formation, but the functional consequences of somatic mitochondrial DNA (mtDNA) mutations are less understood. Here we identify somatic mtDNA mutations across 527 tumors and 14 cancer types, using an approach that takes advantage of e...

  11. Heterology of mitochondrial DNA from mammals detected by electron microscopic heteroduplex analyses

    DEFF Research Database (Denmark)

    Christiansen, Gunna; Christiansen, C

    1983-01-01

    Heteroduplex analysis of mitochondrial DNA (mtDNA) from evolutionary closely related mammals (rat vs. mouse, man vs. monkey) are analyzed and compared to heteroduplex analysis of mt-DNA from more distantly related mammals (rat vs. man, rat vs. monkey, mouse vs. man, mouse vs. monkey and man vs. c...

  12. Simultaneous DNA and RNA mapping of somatic mitochondrial mutations across diverse human cancers

    DEFF Research Database (Denmark)

    Stewart, James B.; Alaei-Mahabadi, Babak; Radhakrishnan, Sabarinathan

    2015-01-01

    Somatic mutations in the nuclear genome are required for tumor formation, but the functional consequences of somatic mitochondrial DNA (mtDNA) mutations are less understood. Here we identify somatic mtDNA mutations across 527 tumors and 14 cancer types, using an approach that takes advantage of e...

  13. Nuclear transfer to prevent mitochondrial DNA disorders : revisiting the debate on reproductive cloning

    NARCIS (Netherlands)

    Bredenoord, A. L.; Dondorp, W.; Pennings, G.; De Wert, G.

    2011-01-01

    Preclinical experiments are currently performed to examine the feasibility of several types of nuclear transfer to prevent mitochondrial DNA (mtDNA) disorders. Whereas the two most promising types of nuclear transfer to prevent mtDNA disorders, spindle transfer and pronuclear transfer, do not amount

  14. Genetics Home Reference: RRM2B-related mitochondrial DNA depletion syndrome, encephalomyopathic form with renal ...

    Science.gov (United States)

    ... Munnich A, Rötig A. Mutation of RRM2B, encoding p53-controlled ribonucleotide reductase (p53R2), causes severe mitochondrial DNA depletion. Nat Genet. 2007 Jun;39(6):776-80. Epub 2007 May 7. Citation on PubMed GeneReview: RRM2B-Related Mitochondrial Disease Pontarin G, Ferraro P, Bee L, Reichard P, ...

  15. Characterization of mitochondrial DNA in various Candida species: isolation, restriction endonuclease analysis, size, and base composition.

    Science.gov (United States)

    Su, C S; Meyer, S A

    1991-01-01

    A practical and effective method for the extraction of mitochondrial DNA from Candida species was developed. Zymolyase was used to induce yeast protoplasts, and mitochondrial DNA was extracted from DNase I-treated mitochondrial preparations. Restriction endonuclease analyses of mitochondrial DNAs from 19 isolates representing seven species of Candida (C. albicans, C. kefyr, C. lusitaniae, C. maltosa, C. parapsilosis, C. shehatae, and C. tropicalis) and Lodderomyces elongisporus revealed different cleavage patterns that appeared to be specific for the species. Few common restriction fragments were evident. The genome sizes of the mitochondrial DNAs ranged from 26.4 to 51.4 kilobase pairs, and the guanine-plus-cytosine contents ranged from 20.7 to 36.8 mol%. There was no correlation between the base compositions of nuclear and mitochondrial DNAs. Eight isolates of C. parapsilosis, including the type culture, and an ascosporogenous strain of L. elongisporus, which was once proposed as the teleomorph of C. parapsilosis, had similar mitochondrial DNA molecular sizes (30.2 and 28.8 kilobase pairs); however, restriction endonuclease patterns of these organisms were distinct. These data provide additional support for discrimination of these two species. The results of our experiments demonstrate that mitochondrial DNA analyses may provide useful criteria for the differentiation of yeast species.

  16. Mitochondrial replacement therapies can circumvent mtDNA based disease transmission

    OpenAIRE

    Wolf, Don P; Mitalipov, Shoukhrat

    2014-01-01

    Mitochondrial DNA diseases are relatively common, sometimes devastating and transmitted exclusively through the egg to children of carrier mothers. The study by Wang et al. (2014) adds the exciting possibility of a new therapy for preventing mitochondrial disease transmission predicated on the use of polar body genomes in mice.

  17. Mitochondrial replacement therapies can circumvent mtDNA-based disease transmission.

    Science.gov (United States)

    Wolf, Don P; Mitalipov, Shoukhrat

    2014-07-01

    Mitochondrial DNA diseases are relatively common, sometimes devastating, and transmitted exclusively through the egg to children of carrier mothers. A study in Cell by Wang et al. (2014) adds the exciting possibility of a new therapy for preventing mitochondrial disease transmission predicated on the use of polar body genomes in mice.

  18. Mitochondrial Inverted Repeats Strongly Correlate with Lifespan: mtDNA Inversions and Aging

    Science.gov (United States)

    Yang, Jiang-Nan; Seluanov, Andrei; Gorbunova, Vera

    2013-01-01

    Mitochondrial defects are implicated in aging and in a multitude of age-related diseases, such as cancer, heart failure, Parkinson’s disease, and Huntington’s disease. However, it is still unclear how mitochondrial defects arise under normal physiological conditions. Mitochondrial DNA (mtDNA) deletions caused by direct repeats (DRs) are implicated in the formation of mitochondrial defects, however, mitochondrial DRs show relatively weak (Pearson’s r = −0.22, p<0.002; Spearman’s ρ = −0.12, p = 0.1) correlation with maximum lifespan (MLS). Here we report a stronger correlation (Pearson’s r = −0.55, p<10–16; Spearman’s ρ = −0.52, p<10–14) between mitochondrial inverted repeats (IRs) and lifespan across 202 species of mammals. We show that, in wild type mice under normal conditions, IRs cause inversions, which arise by replication-dependent mechanism. The inversions accumulate with age in the brain and heart. Our data suggest that IR-mediated inversions are more mutagenic than DR-mediated deletions in mtDNA, and impose stronger constraint on lifespan. Our study identifies IR-induced mitochondrial genome instability during mtDNA replication as a potential cause for mitochondrial defects. PMID:24069185

  19. Mitochondrial inverted repeats strongly correlate with lifespan: mtDNA inversions and aging.

    Directory of Open Access Journals (Sweden)

    Jiang-Nan Yang

    Full Text Available Mitochondrial defects are implicated in aging and in a multitude of age-related diseases, such as cancer, heart failure, Parkinson's disease, and Huntington's disease. However, it is still unclear how mitochondrial defects arise under normal physiological conditions. Mitochondrial DNA (mtDNA deletions caused by direct repeats (DRs are implicated in the formation of mitochondrial defects, however, mitochondrial DRs show relatively weak (Pearson's r = -0.22, p<0.002; Spearman's ρ = -0.12, p = 0.1 correlation with maximum lifespan (MLS. Here we report a stronger correlation (Pearson's r = -0.55, p<10(-16; Spearman's ρ = -0.52, p<10(-14 between mitochondrial inverted repeats (IRs and lifespan across 202 species of mammals. We show that, in wild type mice under normal conditions, IRs cause inversions, which arise by replication-dependent mechanism. The inversions accumulate with age in the brain and heart. Our data suggest that IR-mediated inversions are more mutagenic than DR-mediated deletions in mtDNA, and impose stronger constraint on lifespan. Our study identifies IR-induced mitochondrial genome instability during mtDNA replication as a potential cause for mitochondrial defects.

  20. Effects of Fcj1-Mos1 and mitochondrial division on aggregation of mitochondrial DNA nucleoids and organelle morphology.

    Science.gov (United States)

    Itoh, Kie; Tamura, Yasushi; Iijima, Miho; Sesaki, Hiromi

    2013-06-01

    Mitochondrial DNA (mtDNA) is packaged into DNA-protein complexes called nucleoids, which are distributed as many small foci in mitochondria. Nucleoids are crucial for the biogenesis and function of mtDNA. Here, using a yeast genetic screen for components that control nucleoid distribution and size, we identify Fcj1 and Mos1, two evolutionarily conserved mitochondrial proteins that maintain the connection between the cristae and boundary membranes. These two proteins are also important for establishing tubular morphology of mitochondria, as mitochondria lacking Fcj1 and Mos1 form lamellar sheets. We find that nucleoids aggregate, increase in size, and decrease in number in fcj1 and mos1 cells. In addition, Fcj1 form punctate structures and localized adjacent to nucleoids. Moreover, connecting mitochondria by deleting the DNM1 gene required for organelle division enhances aggregation of mtDNA nucleoids in fcj1 and mos1 cells, whereas single deletion of DNM1 does not affect nucleoids. Conversely, deleting F1Fo-ATP synthase dimerization factors generates concentric ring-like cristae, restores tubular mitochondrial morphology, and suppresses nucleoid aggregation in these mutants. Our findings suggest an unexpected role of Fcj1-Mos1 and organelle division in maintaining the distribution and size of mtDNA nucleoids.

  1. Laser controlled singlet oxygen generation in mitochondria to promote mitochondrial DNA replication in vitro.

    Science.gov (United States)

    Zhou, Xin; Wang, Yupei; Si, Jing; Zhou, Rong; Gan, Lu; Di, Cuixia; Xie, Yi; Zhang, Hong

    2015-11-18

    Reports have shown that a certain level of reactive oxygen species (ROS) can promote mitochondrial DNA (mtDNA) replication. However, it is unclear whether it is the mitochondrial ROS that stimulate mtDNA replication and this requires further investigation. Here we employed a photodynamic system to achieve controlled mitochondrial singlet oxygen ((1)O2) generation. HeLa cells incubated with 5-aminolevulinic acid (ALA) were exposed to laser irradiation to induce (1)O2 generation within mitochondria. Increased mtDNA copy number was detected after low doses of 630 nm laser light in ALA-treated cells. The stimulated mtDNA replication was directly linked to mitochondrial (1)O2 generation, as verified using specific ROS scavengers. The stimulated mtDNA replication was regulated by mitochondrial transcription factor A (TFAM) and mtDNA polymerase γ. MtDNA control region modifications were induced by (1)O2 generation in mitochondria. A marked increase in 8-Oxoguanine (8-oxoG) level was detected in ALA-treated cells after irradiation. HeLa cell growth stimulation and G1-S cell cycle transition were also observed after laser irradiation in ALA-treated cells. These cellular responses could be due to a second wave of ROS generation detected in mitochondria. In summary, we describe a controllable method of inducing mtDNA replication in vitro.

  2. Mitochondrial bioenergetics and drug-induced toxicity in a panel of mouse embryonic fibroblasts with mitochondrial DNA single nucleotide polymorphisms

    Energy Technology Data Exchange (ETDEWEB)

    Pereira, Claudia V.; Oliveira, Paulo J. [CNC—Center for Neuroscience and Cell Biology, University of Coimbra (Portugal); Will, Yvonne [Compound Safety Prediction, Pfizer Global Research and Development, Groton, CT (United States); Nadanaciva, Sashi, E-mail: sashi.nadanaciva@pfizer.com [Compound Safety Prediction, Pfizer Global Research and Development, Groton, CT (United States)

    2012-10-15

    Mitochondrial DNA (mtDNA) variations including single nucleotide polymorphisms (SNPs) have been proposed to be involved in idiosyncratic drug reactions. However, current in vitro and in vivo models lack the genetic diversity seen in the human population. Our hypothesis is that different cell strains with distinct mtDNA SNPs may have different mitochondrial bioenergetic profiles and may therefore vary in their response to drug-induced toxicity. Therefore, we used an in vitro system composed of four strains of mouse embryonic fibroblasts (MEFs) with mtDNA polymorphisms. We sequenced mtDNA from embryonic fibroblasts isolated from four mouse strains, C57BL/6J, MOLF/EiJ, CZECHII/EiJ and PERA/EiJ, with the latter two being sequenced for the first time. The bioenergetic profile of the four strains of MEFs was investigated at both passages 3 and 10. Our results showed that there were clear differences among the four strains of MEFs at both passages, with CZECHII/EiJ having a lower mitochondrial robustness when compared to C57BL/6J, followed by MOLF/EiJ and PERA/EiJ. Seven drugs known to impair mitochondrial function were tested for their effect on the ATP content of the four strains of MEFs in both glucose- and galactose-containing media. Our results showed that there were strain-dependent differences in the response to some of the drugs. We propose that this model is a useful starting point to study compounds that may cause mitochondrial off-target toxicity in early stages of drug development, thus decreasing the number of experimental animals used. -- Highlights: ► mtDNA SNPs may be linked to individual predisposition to drug-induced toxicity. ► CZECHII/EiJ and PERA/EiJ mtDNA was sequenced for the first time in this study. ► Strain-dependent mitochondrial capacity differences were measured. ► Strain-dependent differences in response to mitochondrial toxicants were observed.

  3. Cloning and molecular genetics analyses of Deschampsia antarctica Desv. chloroplast and mitochondrial DNA sequence

    Directory of Open Access Journals (Sweden)

    O.P. Savchuk

    2012-03-01

    Full Text Available Chloroplast and mitochondrial DNA sequences of Deschampsia antarctica were studied. We had made comparison analysis with completely sequenced genomes of other temperateness plants to find homology.

  4. Mitochondrial DNA and Functional Investigations into the Radiosensitivity of Four Mouse Strains

    Directory of Open Access Journals (Sweden)

    Steven B. Zhang

    2014-01-01

    Full Text Available We investigated whether genetic radiosensitivity-related changes in mtDNA/nDNA ratios are significant to mitochondrial function and if a material effect on mtDNA content and function exists. BALB/c (radiosensitive, C57BL/6 (radioresistant, and F1 hybrid mouse strains were exposed to total body irradiation. Hepatic genomic DNA was extracted, and mitochondria were isolated. Mitochondrial oxygen consumption, ROS, and calcium-induced mitochondrial swelling were measured. Radiation influenced strain-specific survival in vivo. F1 hybrid survival was influenced by maternal input. Changes in mitochondrial content corresponded to survival in vivo among the 4 strains. Calcium-induced mitochondrial swelling was strain dependent. Isolated mitochondria from BALB/c mice were significantly more sensitive to calcium overload than mitochondria from C57BL/6 mice. Maternal input partially influenced the recovery effect of radiation on calcium-induced mitochondrial swelling in F1 hybrids; the hybrid with a radiosensitive maternal lineage exhibited a lower rate of recovery. Hybrids had a survival rate that was biased toward maternal input. mtDNA content and mitochondrial permeability transition pores (MPTP measured in these strains before irradiation reflected a dominant input from the parent. After irradiation, the MPTP opened sooner in radiosensitive and hybrid strains, likely triggering intrinsic apoptotic pathways. These findings have important implications for translation into predictors of radiation sensitivity/resistance.

  5. In vivo levels of mitochondrial hydrogen peroxide increase with age in mtDNA mutator mice

    OpenAIRE

    Logan, Angela; Shabalina, Irina G.; Prime, Tracy A.; Rogatti, Sebastian; Kalinovich, Anastasia V.; Hartley, R. C.; Budd, Ralph C.; Cannon, Barbara; Murphy, Michael P.

    2014-01-01

    In mtDNA mutator mice, mtDNA mutations accumulate leading to a rapidly aging phenotype. However, there is little evidence of oxidative damage to tissues, and when analyzed ex vivo, no change in production of the reactive oxygen species (ROS) superoxide and hydrogen peroxide by mitochondria has been reported, undermining the mitochondrial oxidative damage theory of aging. Paradoxically, interventions that decrease mitochondrial ROS levels in vivo delay onset of aging. To reconcile these findin...

  6. Mitochondrial DNA Variability in Populations of Alectoris rufa: A Single-Stranded Conformation Polymorphism (SSCP Approach.

    Directory of Open Access Journals (Sweden)

    Martínez-Fresno, M.

    2006-06-01

    Full Text Available A variable domain of the mitochondrial DNA of the red-legged partridge Alectoris rufa was analysed by single-stranded DNA polymorphism (SSCP, in animals of different populations. Ten mitochondrial types were detected unevenly distributed among samples. A preserved natural population in Northern Spain, Fuentes Carrionas, showed the highest degree of polymorphism. Farm bred animals seem to be less variable and show some genotypes not usually found in the natural sites, suggesting an alien origin of many breeders.

  7. Paternal transmission of mitochondrial DNA as an integral part of mitochondrial inheritance in metapopulations of Drosophila simulans.

    Science.gov (United States)

    Wolff, J N; Nafisinia, M; Sutovsky, P; Ballard, J W O

    2013-01-01

    Maternal inheritance is one of the hallmarks of animal mitochondrial DNA (mtDNA) and central to its success as a molecular marker. This mode of inheritance and subsequent lack of heterologous recombination allows us to retrace evolutionary relationships unambiguously down the matriline and without the confounding effects of recombinant genetic information. Accumulating evidence of biparental inheritance of mtDNA (paternal leakage), however, challenges our current understanding of how this molecule is inherited. Here, using Drosophila simulans collected from an East African metapopulation exhibiting recurring mitochondrial heteroplasmy, we conducted single fly matings and screened F1 offspring for the presence of paternal mtDNA using allele-specific PCR assays (AS-PCR). In all, 27 out of 4092 offspring were identified as harboring paternal mtDNA, suggesting a frequency of 0.66% paternal leakage in this species. Our findings strongly suggest that recurring mtDNA heteroplasmy as observed in natural populations of Drosophila simulans is most likely caused by repeated paternal leakage. Our findings further suggest that this phenomenon to potentially be an integral part of mtDNA inheritance in these populations and consequently of significance for mtDNA as a molecular marker.

  8. Peripheral blood mitochondrial DNA/nuclear DNA (mtDNA/nDNA) ratio as a marker of mitochondrial toxicities of stavudine containing antiretroviral therapy in HIV-infected Malawian patients.

    Science.gov (United States)

    Kampira, Elizabeth; Dzobo, Kevin; Kumwenda, Johnstone; van Oosterhout, Joep J; Parker, M Iqbal; Dandara, Collet

    2014-07-01

    Mitochondrial toxicity is a major concern related to nucleoside reverse transcriptase inhibitors. Common manifestations are peripheral neuropathy and lipodystrophy. Depletion of mitochondria has been associated with mitochondrial dysfunction. We investigated whether mitochondria DNA (mtDNA) levels in peripheral blood can be used as biomarker of stavudine-associated mitochondrial toxicities. We enrolled 203 HIV-infected Malawian adult patients on stavudine-containing ART and 64 healthy controls of Bantu origin in a cross-sectional study. Total DNA was extracted from whole blood.The glyceraldehyde-3-phosphate dehydrogenase gene was used to estimate nuclear DNA (nDNA) levels and the ATP synthase-8 mitochondrial DNA gene to estimate mtDNA levels, from which mtDNA/nDNA ratios were determined. MtDNA subhaplogroups were established by sequencing. Among patients, peripheral neuropathy was present in 21% (43/203), lipodystrophy in 18% (20/112), elevated lactate level (>2.5 mmol/L) in 17% (19/113). Healthy controls had a higher median mtDNA/nDNA ratio when compared to HIV/AIDS patients (6.64 vs. 5.08; p=0.05), patients presenting with peripheral neuropathy (6.64 vs. 3.40, p=0.039), and patients with high lactate levels (6.64 vs. 0.68, p=0.024), respectively. Significant differences in median mtDNA/nDNA ratios were observed between patients with high and normal lactate levels (5.88 vs. 0.68, p=0.018). The median mtDNA/nDNA ratio of patients in subhaplogroup L0a2 was much lower (0.62 vs. 8.50, p=0.01) than that of those in subhaplogroup L2a. Our data indicate that peripheral blood mtDNA/nDNA ratio is a marker of mitochondrial toxicities of stavudine and is associated with elevated lactate levels and mtDNA subhaplogroups. This could open the prospect to select a substantial group of patients who will not have problematic side effects from stavudine, an affordable and effective antiretroviral drug that is being phased out in Africa due to its toxicity.

  9. The Role of Mitochondrial DNA in Mediating Alveolar Epithelial Cell Apoptosis and Pulmonary Fibrosis

    Directory of Open Access Journals (Sweden)

    Seok-Jo Kim

    2015-09-01

    Full Text Available Convincing evidence has emerged demonstrating that impairment of mitochondrial function is critically important in regulating alveolar epithelial cell (AEC programmed cell death (apoptosis that may contribute to aging-related lung diseases, such as idiopathic pulmonary fibrosis (IPF and asbestosis (pulmonary fibrosis following asbestos exposure. The mammalian mitochondrial DNA (mtDNA encodes for 13 proteins, including several essential for oxidative phosphorylation. We review the evidence implicating that oxidative stress-induced mtDNA damage promotes AEC apoptosis and pulmonary fibrosis. We focus on the emerging role for AEC mtDNA damage repair by 8-oxoguanine DNA glycosylase (OGG1 and mitochondrial aconitase (ACO-2 in maintaining mtDNA integrity which is important in preventing AEC apoptosis and asbestos-induced pulmonary fibrosis in a murine model. We then review recent studies linking the sirtuin (SIRT family members, especially SIRT3, to mitochondrial integrity and mtDNA damage repair and aging. We present a conceptual model of how SIRTs modulate reactive oxygen species (ROS-driven mitochondrial metabolism that may be important for their tumor suppressor function. The emerging insights into the pathobiology underlying AEC mtDNA damage and apoptosis is suggesting novel therapeutic targets that may prove useful for the management of age-related diseases, including pulmonary fibrosis and lung cancer.

  10. Suppression of a defect in mitochondrial protein import identifies cytosolic proteins required for viability of yeast cells lacking mitochondrial DNA.

    Science.gov (United States)

    Dunn, Cory D; Jensen, Robert E

    2003-01-01

    The TIM22 complex, required for the insertion of imported polytopic proteins into the mitochondrial inner membrane, contains the nonessential Tim18p subunit. To learn more about the function of Tim18p, we screened for high-copy suppressors of the inability of tim18Delta mutants to live without mitochondrial DNA (mtDNA). We identified several genes encoding cytosolic proteins, including CCT6, SSB1, ICY1, TIP41, and PBP1, which, when overproduced, rescue the mtDNA dependence of tim18Delta cells. Furthermore, these same plasmids rescue the petite-negative phenotype of cells lacking other components of the mitochondrial protein import machinery. Strikingly, disruption of the genes identified by the different suppressors produces cells that are unable to grow without mtDNA. We speculate that loss of mtDNA leads to a lowered inner membrane potential, and subtle changes in import efficiency can no longer be tolerated. Our results suggest that increased amounts of Cct6p, Ssb1p, Icy1p, Tip41p, and Pbp1p help overcome the problems resulting from a defect in protein import. PMID:14504216

  11. Comparison of mitochondrial mutation spectra in ageing human colonic epithelium and disease: absence of evidence for purifying selection in somatic mitochondrial DNA point mutations

    NARCIS (Netherlands)

    Greaves, L.C.; Elson, J.L.; Nooteboom, M.; Grady, J.P.; Taylor, G.A.; Taylor, R.W.; Mathers, J.C.; Kirkwood, T.B.; Turnbull, D.M.

    2012-01-01

    Human ageing has been predicted to be caused by the accumulation of molecular damage in cells and tissues. Somatic mitochondrial DNA (mtDNA) mutations have been documented in a number of ageing tissues and have been shown to be associated with cellular mitochondrial dysfunction. It is unknown whethe

  12. Statistical analysis of post mortem DNA damage-derived miscoding lesions in Neandertal mitochondrial DNA

    Directory of Open Access Journals (Sweden)

    Gigli Elena

    2008-07-01

    Full Text Available Abstract Background We have analysed the distribution of post mortem DNA damage derived miscoding lesions from the datasets of seven published Neandertal specimens that have extensive cloned sequence coverage over the mitochondrial DNA (mtDNA hypervariable region 1 (HVS1. The analysis was restricted to C→T and G→A miscoding lesions (the predominant manifestation of post mortem damage that are seen at a frequency of more than one clone among sequences from a single PCR, but do not represent the true endogenous sequence. Findings The data indicates an extreme bias towards C→T over G→A miscoding lesions (observed ratio of 67:2 compared to an expected ratio of 7:2, implying that the mtDNA Light strand molecule suffers proportionally more damage-derived miscoding lesions than the Heavy strand. Conclusion The clustering of Cs in the Light strand as opposed to the singleton pattern of Cs in the Heavy strand could explain the observed bias, a phenomenon that could be further tested with non-PCR based approaches. The characterization of the HVS1 hotspots will be of use to future Neandertal mtDNA studies, with specific regards to assessing the authenticity of new positions previously unknown to be polymorphic.

  13. Deep sequencing unearths nuclear mitochondrial sequences under Leber's hereditary optic neuropathy-associated false heteroplasmic mitochondrial DNA variants.

    Science.gov (United States)

    Petruzzella, Vittoria; Carrozzo, Rosalba; Calabrese, Claudia; Dell'Aglio, Rosa; Trentadue, Raffaella; Piredda, Roberta; Artuso, Lucia; Rizza, Teresa; Bianchi, Marzia; Porcelli, Anna Maria; Guerriero, Silvana; Gasparre, Giuseppe; Attimonelli, Marcella

    2012-09-01

    Leber's hereditary optic neuropathy (LHON) is associated with mitochondrial DNA (mtDNA) ND mutations that are mostly homoplasmic. However, these mutations are not sufficient to explain the peculiar features of penetrance and the tissue-specific expression of the disease and are believed to be causative in association with unknown environmental or other genetic factors. Discerning between clear-cut pathogenetic variants, such as those that appear to be heteroplasmic, and less penetrant variants, such as the homoplasmic, remains a challenging issue that we have addressed here using next-generation sequencing approach. We set up a protocol to quantify MTND5 heteroplasmy levels in a family in which the proband manifests a LHON phenotype. Furthermore, to study this mtDNA haplotype, we applied the cybridization protocol. The results demonstrate that the mutations are mostly homoplasmic, whereas the suspected heteroplasmic feature of the observed mutations is due to the co-amplification of Nuclear mitochondrial Sequences.

  14. Mitochondrial DNA paradox: sex-specific genetic structure in a marine mussel--despite maternal inheritance and passive dispersal

    National Research Council Canada - National Science Library

    Teske, Peter R; Papadopoulos, Isabelle; Barker, Nigel P; McQuaid, Christopher D

    2012-01-01

    When genetic structure is identified using mitochondrial DNA (mtDNA), but no structure is identified using biparentally-inherited nuclear DNA, the discordance is often attributed to differences in dispersal potential between the sexes...

  15. Analysis of sequence variation in Gnathostoma spinigerum mitochondrial DNA by single-strand conformation polymorphism analysis and DNA sequence.

    Science.gov (United States)

    Ngarmamonpirat, Charinthon; Waikagul, Jitra; Petmitr, Songsak; Dekumyoy, Paron; Rojekittikhun, Wichit; Anantapruti, Malinee T

    2005-03-01

    Morphological variations were observed in the advance third stage larvae of Gnathostoma spinigerum collected from swamp eel (Fluta alba), the second intermediate host. Larvae with typical and three atypical types were chosen for partial cytochrome c oxidase subunit I (COI) gene sequence analysis. A 450 bp polymerase chain reaction product of the COI gene was amplified from mitochondrial DNA. The variations were analyzed by single-strand conformation polymorphism and DNA sequencing. The nucleotide variations of the COI gene in the four types of larvae indicated the presence of an intra-specific variation of mitochondrial DNA in the G. spinigerum population.

  16. Troglitazone, but not rosiglitazone, damages mitochondrial DNA and induces mitochondrial dysfunction and cell death in human hepatocytes.

    Science.gov (United States)

    Rachek, Lyudmila I; Yuzefovych, Larysa V; Ledoux, Susan P; Julie, Neil L; Wilson, Glenn L

    2009-11-01

    Thiazolidinediones (TZDs), such as troglitazone (TRO) and rosiglitazone (ROSI), improve insulin resistance by acting as ligands for the nuclear receptor peroxisome proliferator-activated receptor-gamma (PPARgamma). TRO was withdrawn from the market because of reports of serious hepatotoxicity. A growing body of evidence suggests that TRO caused mitochondrial dysfunction and induction of apoptosis in human hepatocytes but its mechanisms of action remain unclear. We hypothesized that damage to mitochondrial DNA (mtDNA) is an initiating event involved in TRO-induced mitochondrial dysfunction and hepatotoxicity. Primary human hepatocytes were exposed to TRO and ROSI. The results obtained revealed that TRO, but not ROSI at equimolar concentrations, caused a substantial increase in mtDNA damage and decreased ATP production and cellular viability. The reactive oxygen species (ROS) scavenger, N-acetyl cystein (NAC), significantly diminished the TRO-induced cytotoxicity, suggesting involvement of ROS in TRO-induced hepatocyte cytotoxicity. The PPARgamma antagonist (GW9662) did not block the TRO-induced decrease in cell viability, indicating that the TRO-induced hepatotoxicity is PPARgamma-independent. Furthermore, TRO induced hepatocyte apoptosis, caspase-3 cleavage and cytochrome c release. Targeting of a DNA repair protein to mitochondria by protein transduction using a fusion protein containing the DNA repair enzyme Endonuclease III (EndoIII) from Escherichia coli, a mitochondrial translocation sequence (MTS) and the protein transduction domain (PTD) from HIV-1 TAT protein protected hepatocytes against TRO-induced toxicity. Overall, our results indicate that significant mtDNA damage caused by TRO is a prime initiator of the hepatoxicity caused by this drug.

  17. DNA packaging proteins Glom and Glom2 coordinately organize the mitochondrial nucleoid of Physarum polycephalum.

    Science.gov (United States)

    Itoh, Kie; Izumi, Akiko; Mori, Toshiyuki; Dohmae, Naoshi; Yui, Ryoko; Maeda-Sano, Katsura; Shirai, Yuki; Kanaoka, Masahiro M; Kuroiwa, Tsuneyoshi; Higashiyama, Tetsuya; Sugita, Mamoru; Murakami-Murofushi, Kimiko; Kawano, Shigeyuki; Sasaki, Narie

    2011-07-01

    Mitochondrial DNA (mtDNA) is generally packaged into the mitochondrial nucleoid (mt-nucleoid) by a high-mobility group (HMG) protein. Glom is an mtDNA-packaging HMG protein in Physarum polycephalum. Here we identified a new mtDNA-packaging protein, Glom2, which had a region homologous with yeast Mgm101. Glom2 could bind to an entire mtDNA and worked synergistically with Glom for condensation of mtDNA in vitro. Down-regulation of Glom2 enhanced the alteration of mt-nucleoid morphology and the loss of mtDNA induced by down-regulation of Glom, and impaired mRNA accumulation of some mtDNA-encoded genes. These data suggest that Glom2 may organize the mt-nucleoid coordinately with Glom. © Elsevier B.V. and Mitochondria Research Society. All rights reserved.

  18. The interplay between SUCLA2, SUCLG2, and mitochondrial DNA depletion

    DEFF Research Database (Denmark)

    Miller, Chaya; Wang, Liya; Ostergaard, Elsebet;

    2011-01-01

    SUCLA2-related mitochondrial DNA (mtDNA) depletion syndrome is a result of mutations in the β subunit of the ADP-dependent isoform of the Krebs cycle succinyl-CoA synthase (SCS). The mechanism of tissue specificity and mtDNA depletion is elusive but complementation by the GDP-dependent isoform...... encoded by SUCLG2, and the association with mitochondrial nucleoside diphosphate kinase (NDPK), is a plausible link. We have investigated this relationship by studying SUCLA2 deficient fibroblasts derived from patients and detected normal mtDNA content and normal NDPK activity. However, knockdown of SUCLG...... and that mitochondrial NDPK is involved. Although results pertain to a cell culture system, the findings might explain the pathomechanism and tissue specificity in mtDNA depletion caused by defective SUCLA2....

  19. ER-associated mitochondrial division links the distribution of mitochondria and mitochondrial DNA in yeast.

    Science.gov (United States)

    Murley, Andrew; Lackner, Laura L; Osman, Christof; West, Matthew; Voeltz, Gia K; Walter, Peter; Nunnari, Jodi

    2013-05-14

    Mitochondrial division is important for mitochondrial distribution and function. Recent data have demonstrated that ER-mitochondria contacts mark mitochondrial division sites, but the molecular basis and functions of these contacts are not understood. Here we show that in yeast, the ER-mitochondria tethering complex, ERMES, and the highly conserved Miro GTPase, Gem1, are spatially and functionally linked to ER-associated mitochondrial division. Gem1 acts as a negative regulator of ER-mitochondria contacts, an activity required for the spatial resolution and distribution of newly generated mitochondrial tips following division. Previous data have demonstrated that ERMES localizes with a subset of actively replicating mitochondrial nucleoids. We show that mitochondrial division is spatially linked to nucleoids and that a majority of these nucleoids segregate prior to division, resulting in their distribution into newly generated tips in the mitochondrial network. Thus, we postulate that ER-associated division serves to link the distribution of mitochondria and mitochondrial nucleoids in cells. DOI:http://dx.doi.org/10.7554/eLife.00422.001.

  20. Mitochondrial DNA hyperdiversity and its potential causes in the marine periwinkle Melarhaphe neritoides (Mollusca: Gastropoda)

    OpenAIRE

    2016-01-01

    We report the presence of mitochondrial DNA (mtDNA) hyperdiversity in the marine periwinkle Melarhaphe neritoides (Linnaeus, 1758), the first such case among marine gastropods. Our dataset consisted of concatenated 16S-COI-Cytb gene fragments. We used Bayesian analyses to investigate three putative causes underlying genetic variation, and estimated the mtDNA mutation rate, possible signatures of selection and the effective population size of the species in the Azores archipelago. The mtDNA hy...

  1. Differential chromosomal and mitochondrial DNA synthesis in temperature-sensitive mutants of Ustilago maydis

    Energy Technology Data Exchange (ETDEWEB)

    Unrau, P.

    1977-01-01

    The amount and type of residual DNA synthesis was determined in eight temperature-sensitive mutants of the smut fungus Ustilago maydis after incubation at the restrictive temperature (32/sup 0/C) for eight hours. Mutants ts-220, ts-207, ts-432 and ts-346 were found to have an overall reduction in the synthesis of both nuclear and mitochondrial DNA in comparison to the wild-type. In mutants ts-20, tsd 1-1, ts-84 and pol 1-1 nuclear DNA synthesis was depressed relative to mitochondrial synthesis. The DNA-polymerase mutant pol 1-1 had persistent nuclear synthesis at about 50% of the rate of synthesis of mitochondrial DNA and similar behavior was observed in a diploid homozygous strain. Mutant ts-84 had an initial burst of DNA synthesis which was reduced for nuclear but not mitochondrial synthesis after three hours preincubation at 32/sup 0/C. tsd 1-1 and ts-20 had nuclear residual synthesis amounting to about 25% of the relative rate of mitochondrial synthesis which correlates to increasing UV sensitivity of these strains on incubation at 32/sup 0/C. A pol 1-1 ts-84 double mutant had an additive loss of nuclear DNA synthesis which indicates that the steps of replication involved may be sequential.

  2. Silencing of PINK1 expression affects mitochondrial DNA and oxidative phosphorylation in dopaminergic cells.

    Directory of Open Access Journals (Sweden)

    Matthew E Gegg

    Full Text Available BACKGROUND: Mitochondrial dysfunction has been implicated in the pathogenesis of Parkinson's disease (PD. Impairment of the mitochondrial electron transport chain (ETC and an increased frequency in deletions of mitochondrial DNA (mtDNA, which encodes some of the subunits of the ETC, have been reported in the substantia nigra of PD brains. The identification of mutations in the PINK1 gene, which cause an autosomal recessive form of PD, has supported mitochondrial involvement in PD. The PINK1 protein is a serine/threonine kinase localized in mitochondria and the cytosol. Its precise function is unknown, but it is involved in neuroprotection against a variety of stress signalling pathways. METHODOLOGY/PRINCIPAL FINDINGS: In this report we have investigated the effect of silencing PINK1 expression in human dopaminergic SH-SY5Y cells by siRNA on mtDNA synthesis and ETC function. Loss of PINK1 expression resulted in a decrease in mtDNA levels and mtDNA synthesis. We also report a concomitant loss of mitochondrial membrane potential and decreased mitochondrial ATP synthesis, with the activity of complex IV of the ETC most affected. This mitochondrial dysfunction resulted in increased markers of oxidative stress under basal conditions and increased cell death following treatment with the free radical generator paraquat. CONCLUSIONS: This report highlights a novel function of PINK1 in mitochondrial biogenesis and a role in maintaining mitochondrial ETC activity. Dysfunction of both has been implicated in sporadic forms of PD suggesting that these may be key pathways in the development of the disease.

  3. Repair of 8-oxodeoxyguanosine lesions in mitochondrial DNA depends on the oxoguanine DNA glycosylase (OGG1) gene and 8- oxoguanine accumulates in the mitochondrial DNA of OGG1- defective mice

    DEFF Research Database (Denmark)

    Souza-Pinto, N.C.; Eide, L.; Hogue, B.A.

    2001-01-01

    encodes for the mitochondrial 8-oxodG glycosylase because these extracts have no incision activity toward an oligonucleotide containing a single 8-oxodG DNA base lesion, Consistent with an important role for the OGG1 protein in the removal of 8-oxodC from the mitochondrial genome, we found that mtDNA...... DNA, The main pathway for the repair of 8-oxodG is the base excision repair pathway initiated by oxoguanine DNA glycosylase (OGG1), We previously demonstrated that mammalian mitochondria from mice efficiently remove 8-oxodG from their genomes and isolated a protein from rat liver mitochondria with 8......Mitochondria are not only the major site for generation of reactive oxygen species, but also one of the main targets of oxidative damage. One of the major products of DNA oxidation, 8-oxodeoxyguanosine (8-oxodC), accumulates in mitochondrial DNA (mtDNA) at levels three times higher than in nuclear...

  4. Mitochondrial polymerase gamma dysfunction and aging cause cardiac nuclear DNA methylation changes.

    Science.gov (United States)

    Koczor, Christopher A; Ludlow, Ivan; Fields, Earl; Jiao, Zhe; Ludaway, Tomika; Russ, Rodney; Lewis, William

    2016-04-01

    Cardiomyopathy (CM) is an intrinsic weakening of myocardium with contractile dysfunction and congestive heart failure (CHF). CHF has been postulated to result from decreased mitochondrial energy production and oxidative stress. Effects of decreased mitochondrial oxygen consumption also can accelerate with aging. We previously showed DNA methylation changes in human hearts with CM. This was associated with mitochondrial DNA depletion, being another molecular marker of CM. We examined the relationship between mitochondrial dysfunction and cardiac epigenetic DNA methylation changes in both young and old mice. We used genetically engineered C57Bl/6 mice transgenic for a cardiac-specific mutant of the mitochondrial polymerase-γ (termed Y955C). Y955C mice undergo left ventricular hypertrophy (LVH) at a young age (∼ 94 days old), and LVH decompensated to CHF at old age (∼ 255 days old). Results found 95 genes differentially expressed as a result of Y955C expression, while 4,452 genes were differentially expressed as a result of aging hearts. Moreover, cardiac DNA methylation patterns differed between Y955C (4,506 peaks with 68.5% hypomethylation) and aged hearts (73,286 peaks with 80.2% hypomethylated). Correlatively, of the 95 Y955C-dependent differentially expressed genes, 30 genes (31.6%) also displayed differential DNA methylation; in the 4,452 age-dependent differentially expressed genes, 342 genes (7.7%) displayed associated DNA methylation changes. Both Y955C and aging demonstrated significant enrichment of CACGTG-associated E-box motifs in differentially methylated regions. Cardiac mitochondrial polymerase dysfunction alters nuclear DNA methylation. Furthermore, aging causes a robust change in cardiac DNA methylation that is partially associated with mitochondrial polymerase dysfunction. Copyright © 2016 the American Physiological Society.

  5. Implementation of DNA Mitochondrial Analysis in Rhinoclemmys nasuta (Testudines: Geoemydidae

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    Yherson Franchesco Molina Henao

    2014-05-01

    Full Text Available Rhinoclemmys nasuta (Testudines:Geoemydidae is considered an almost endemic specie to Colombia and the most primitive species of Rhynoclemmys. However, it is classified data deficient by IUCN because the available information is not enough to make a direct or indirect assessment of its extinction risk. Here, we describe the implementation of the method to analyze the mitochondrial DNA control sequence (mtDNA of R. nasuta in order to generate tools for future studies in systematics and population conservation. Genomic mtDNA was extracted by salting-out from blood samples from Isla Palma and Playa Chucheros (Bahía Málaga - Colombian Pacific Coast. The polymerase reaction chain (PCR was performed using a pair of degenerate primers (reported for Chrysemys picta, Testudines:Emydidae in polyacrylamide gel stained with silver nitrate. Fragments of 800pb were obtained and the sequencing reaction was effective.High homology percentage (> 92 % was established between the obtained sequences with mtDNA sequences from two species of Geoemydidae, Sacalia quadriocellata and Cuora aurocapitata which are deposited in the GenBank. This demonstrate an effective sequencing ofthe mtDNA control region of R. nasuta.IMPLEMENTACIÓN DE LA METODOLOGÍA DE ANÁLISIS DE ADN MITOCONDRIAL EN Rhinoclemmys nasuta (TESTUDINES:GEOEMYDIDAERhinoclemmys nasuta (Testudines: Geoemydidae es considerada una especie casi endémica de Colombia y la más primitiva del género, sin embargo, se encuentra clasificada por la IUCN como deficiente de datos, ya que la información disponible no es suficiente para hacer una evaluación directa o indirecta de su riesgo de extinción. Con el propósito de generar herramientas de análisis poblacional se describe la metodología de secuenciación de la región control del ADN mitocondrial (ADNmt para el análisis genético de poblaciones de Rhinoclemmys nasuta, que permita establecer bases para futuros estudios evolutivos y de conservación. Se

  6. Altered Mitochondrial DNA Methylation Pattern in Alzheimer Disease-Related Pathology and in Parkinson Disease.

    Science.gov (United States)

    Blanch, Marta; Mosquera, Jose Luis; Ansoleaga, Belén; Ferrer, Isidre; Barrachina, Marta

    2016-02-01

    Mitochondrial dysfunction is linked with the etiopathogenesis of Alzheimer disease and Parkinson disease. Mitochondria are intracellular organelles essential for cell viability and are characterized by the presence of the mitochondrial (mt)DNA. DNA methylation is a well-known epigenetic mechanism that regulates nuclear gene transcription. However, mtDNA methylation is not the subject of the same research attention. The present study shows the presence of mitochondrial 5-methylcytosine in CpG and non-CpG sites in the entorhinal cortex and substantia nigra of control human postmortem brains, using the 454 GS FLX Titanium pyrosequencer. Moreover, increased mitochondrial 5-methylcytosine levels are found in the D-loop region of mtDNA in the entorhinal cortex in brain samples with Alzheimer disease-related pathology (stages I to II and stages III to IV of Braak and Braak; n = 8) with respect to control cases. Interestingly, this region shows a dynamic pattern in the content of mitochondrial 5-methylcytosine in amyloid precursor protein/presenilin 1 mice along with Alzheimer disease pathology progression (3, 6, and 12 months of age). Finally, a loss of mitochondrial 5-methylcytosine levels in the D-loop region is found in the substantia nigra in Parkinson disease (n = 10) with respect to control cases. In summary, the present findings suggest mtDNA epigenetic modulation in human brain is vulnerable to neurodegenerative disease states.

  7. Mitochondrial DNA heteroplasmy dynamics in a kindred harboring a novel pathogenic mutation in the mitochondrial tRNA glutamate gene

    Energy Technology Data Exchange (ETDEWEB)

    Moraes, C.T.; Hao, H. [Univ. of Miami, FL (United States); Bonilla, E.; DiMauro, S.

    1994-09-01

    We have identified a novel mitochondrial DNA (mtDNA) mutation in a 32-year-old male with a myopathy (without progressive external ophthalmoplegia) and mild pyramidal involvement. This A{yields}G transition at mtDNA position 14709 alters an evolutionary conserved nucleotide in a region coding for the anticodon loop of the mitcohondrial tRNA{sup Glu}. The 14709 mtDNA mutation was heteroplasmic but present at very high levels in the patient`s muscle (95%), white blood cells (81%) and hair follicles (90%). The same mutant mtDNA population was observed in white blood cells and hair follicles of all maternal relatives, but a lesser percentage (25-80%). The patient`s muscle showed many ragged-red fibers and a severe focal defect in cytochrome c oxidase activity, accompanied by the absence of cross-reacting material for mitochondrially synthesized polypeptides (ND 1 and COX II). The percentage of mutant mtDNA was not preferentially increased over two generations. Rather, the percentage of mutant mtDNA observed in siblings seemed to follow a normal distribution around the percentage observed in their mothers. Single hair PCR/RFLP analysis showed that the intercellular fluctuation in the percentage of mutant mtDNA differs among family members. Younger generations tend to have a more homogeneous distribution of mutant mtDNA in different hair follicles. The highest degree of variability between individual hair follicles was observed in the patient`s grandmother. These results suggest that the intercellular distribution of the mutant and wild-type mtDNA populations may drift towards homogeneity in subsequent generations.

  8. Large heterogeneity of mitochondrial DNA transcription and initiation of replication exposed by single-cell imaging.

    Science.gov (United States)

    Chatre, Laurent; Ricchetti, Miria

    2013-02-15

    Mitochondrial DNA (mtDNA) replication and transcription are crucial for cell function, but these processes are poorly understood at the single-cell level. We describe a novel fluorescence in situ hybridization protocol, called mTRIP (mitochondrial transcription and replication imaging protocol), that reveals simultaneously mtDNA and RNA, and that can also be coupled to immunofluorescence for in situ protein examination. mTRIP reveals mitochondrial structures engaged in initiation of DNA replication by identification of a specific sequence in the regulatory D-loop, as well as unique transcription profiles in single human cells. We observe and quantify at least three classes of mitochondrial structures: (i) replication initiation active and transcript-positive (Ia-Tp); (ii) replication initiation silent and transcript-positive (Is-Tp); and (iii) replication initiation silent and transcript-negative (Is-Tn). Thus, individual mitochondria are dramatically heterogeneous within the same cell. Moreover, mTRIP exposes a mosaic of distinct nucleic acid patterns in the D-loop, including H-strand versus L-strand transcripts, and uncoupled rRNA transcription and mtDNA initiation of replication, which might have functional consequences in the regulation of the mtDNA. Finally, mTRIP identifies altered mtDNA processing in cells with unbalanced mtDNA content and function, including in human mitochondrial disorders. Thus, mTRIP reveals qualitative and quantitative alterations that provide additional tools for elucidating the dynamics of mtDNA processing in single cells and mitochondrial dysfunction in diseases.

  9. Prevalence of mitochondrial DNA mutations in sporadic patients with nonsyndromic sensorineural hearing loss

    Directory of Open Access Journals (Sweden)

    Hua Jiang

    Full Text Available ABSTRACT INTRODUCTION: Several mitochondrial DNA mutations have been reported to be associated with nonsyndromic hearing loss in several families. However, little is known about the prevalence of these mutations in sporadic patients with nonsyndromic sensorineural hearing loss. OBJECTIVE: The purpose of our study was to investigate the incidence of these mitochondrial DNA mutations in such population. METHODS: A total of 178 sporadic patients with nonsyndromic sensorineural hearing loss were enrolled in this study. Genomic DNA was extracted from the peripheral blood sample. We employed the SNaPshot(r sequencing method to detect five mitochondrial DNA mutations, including A1555G and A827G in 12S rRNA gene and A7445G, 7472insC, and T7511C in tRNASerUCN gene. Meanwhile, we used polymerase chain reaction and sequenced the products to screen GJB2 gene mutations in patients carrying mitochondrial DNA mutations. RESULTS: We failed to detect the presence of A1555G mutation in 12S rRNA gene, and of A7445G, 7472insC, T7511C mutations in tRNASerUCN gene in our population. However, we found that 6 patients (3.37% were carriers of a homozygous A827G mutation and one of them also carried homozygous GJB2 235delC mutation. CONCLUSION: Our findings in the present study indicate that even in sporadic patients with nonsyndromic sensorineural hearing loss, mitochondrial DNA mutations might also contribute to the clinical phenotype.

  10. Base composition at mtDNA boundaries suggests a DNA triple helix model for human mitochondrial DNA large-scale rearrangements.

    Science.gov (United States)

    Rocher, Christophe; Letellier, Thierry; Copeland, William C; Lestienne, Patrick

    2002-06-01

    Different mechanisms have been proposed to account for mitochondrial DNA (mtDNA) instability based on the presence of short homologous sequences (direct repeats, DR) at the potential boundaries of mtDNA rearrangements. Among them, slippage-mispairing of the replication complex during the asymmetric replication cycle of the mammalian mitochondrial DNA has been proposed to account for the preferential localization of deletions. This mechanism involves a transfer of the replication complex from the first neo-synthesized heavy (H) strand of the DR1, to the DR2, thus bypassing the intervening sequence and producing a deleted molecule. Nevertheless, the nature of the bonds between the DNA strands remains unknown as the forward sequence of DR2, beyond the replication complex, stays double-stranded. Here, we have analyzed the base composition of the DR at the boundaries of mtDNA deletions and duplications and found a skewed pyrimidine content of about 75% in the light-strand DNA template. This suggests the possible building of a DNA triple helix between the G-rich neo-synthesized DR1 and the base-paired homologous G.C-rich DR2. In vitro experiments with the purified human DNA polymerase gamma subunits enabled us to show that the third DNA strand may be used as a primer for DNA replication, using a template with the direct repeat forming a hairpin, with which the primer could initiate DNA replication. These data suggest a novel molecular basis for mitochondrial DNA rearrangements through the distributive nature of the DNA polymerase gamma, at the level of the direct repeats. A general model accounting for large-scale mitochondrial DNA deletion and duplication is proposed. These experiments extend to a DNA polymerase from an eucaryote source the use of a DNA triple helix strand as a primer, like other DNA polymerases from phage and bacterial origins.

  11. The effect of chronic alcohol consumption on mitochondrial DNA mutagenesis in human blood.

    Science.gov (United States)

    von Wurmb-Schwark, N; Ringleb, A; Schwark, T; Broese, T; Weirich, S; Schlaefke, D; Wegener, R; Oehmichen, M

    2008-01-01

    The 4977bp deletion of mitochondrial DNA (mtDNA) is known to accumulate with increasing age in post mitotic tissues. Recently, studies came out detecting this specific alteration also in fast replicating cells, e.g. in blood or skin tissue, often in correlation to specific diseases or -- specifically in skin -- external stressors such as UV radiation. In this study, we investigated mitochondrial mutagenesis in 69 patients with a chronic alcoholic disease and 46 age matched controls with a moderate drinking behavior. Two different fragments, specific for total and for deleted mtDNA (dmtDNA) were amplified in a duplex-PCR. A subsequent fragment analysis was performed and for relative quantification, the quotient of the peak areas of amplification products specific for deleted and total mtDNA was determined. Additionally, a real time PCR was performed to quantify mtDNA copy number. The relative amount of 4977bp deleted mtDNA in alcoholics was significantly increased compared to controls. On the other hand, no difference regarding the mtDNA/nuclear DNA ratio in both investigated groups was detected. Additionally, no age dependence could be found nor in alcoholics, neither in the control group. These findings indicate that mtDNA mutagenesis in blood can be influenced by stressors such as alcohol. Ethanol seems to be a significant factor to alter mitochondrial DNA in blood and might be an additional contributor for the cellular aging process.

  12. The effect of chronic alcohol consumption on mitochondrial DNA mutagenesis in human blood

    Energy Technology Data Exchange (ETDEWEB)

    Wurmb-Schwark, N. von [Institute of Legal Medicine, Christian Albrecht University of Kiel, Arnold-Heller-Str. 12, 24105 Kiel (Germany)], E-mail: nvonwurmb@rechtsmedizin.uni-kiel.de; Ringleb, A.; Schwark, T. [Institute of Legal Medicine, Christian Albrecht University of Kiel, Arnold-Heller-Str. 12, 24105 Kiel (Germany); Broese, T.; Weirich, S.; Schlaefke, D. [Clinic of Psychiatry and Psychotherapy, University of Rostock, Gehlsheimer Str. 20, Rostock (Germany); Wegener, R. [Institute of Legal Medicine, St-Georg-Str. 108, University of Rostock, 18055 Rostock (Germany); Oehmichen, M. [Institute of Legal Medicine, Christian Albrecht University of Kiel, Arnold-Heller-Str. 12, 24105 Kiel (Germany)

    2008-01-01

    The 4977 bp deletion of mitochondrial DNA (mtDNA) is known to accumulate with increasing age in post mitotic tissues. Recently, studies came out detecting this specific alteration also in fast replicating cells, e.g. in blood or skin tissue, often in correlation to specific diseases or - specifically in skin - external stressors such as UV radiation. In this study, we investigated mitochondrial mutagenesis in 69 patients with a chronic alcoholic disease and 46 age matched controls with a moderate drinking behavior. Two different fragments, specific for total and for deleted mtDNA (dmtDNA) were amplified in a duplex-PCR. A subsequent fragment analysis was performed and for relative quantification, the quotient of the peak areas of amplification products specific for deleted and total mtDNA was determined. Additionally, a real time PCR was performed to quantify mtDNA copy number. The relative amount of 4977 bp deleted mtDNA in alcoholics was significantly increased compared to controls. On the other hand, no difference regarding the mtDNA/nuclear DNA ratio in both investigated groups was detected. Additionally, no age dependence could be found nor in alcoholics, neither in the control group. These findings indicate that mtDNA mutagenesis in blood can be influenced by stressors such as alcohol. Ethanol seems to be a significant factor to alter mitochondrial DNA in blood and might be an additional contributor for the cellular aging process.

  13. DNA Amplification and Nucleotide Sequence Determination of a Region of Mitochondrial DNA in the Sea Snake, Laticauda Semifasciata

    OpenAIRE

    Eguchi, Tomoko; Eguchi, Yukinori; Oshiro, Minoru; Asato, Tsuyoshi; Takei, Hiroshi; Nakashima, Yasutsugu

    1993-01-01

    We determined the nucleotide sequence of a region of the 12S ribosomal RNA (rRNA) gene in the mitochondrial DNA (mtDNA) of the sea snake, Laticauda semifasciata, using the polymerase chain reaction (PCR). We synthesized oligonucleotide primers according to the nucleotide sequence of human mt DNA 12S rRNA gene and found that the target sequence (386bp) of the sea snake mtDNA could be amplified with these primers. The nucleotide sequence of the amplified region of the sea snake mt DNA was deter...

  14. Recent mitochondrial DNA mutations increase the risk of developing common late-onset human diseases.

    Directory of Open Access Journals (Sweden)

    Gavin Hudson

    2014-05-01

    Full Text Available Mitochondrial DNA (mtDNA is highly polymorphic at the population level, and specific mtDNA variants affect mitochondrial function. With emerging evidence that mitochondrial mechanisms are central to common human diseases, it is plausible that mtDNA variants contribute to the "missing heritability" of several complex traits. Given the central role of mtDNA genes in oxidative phosphorylation, the same genetic variants would be expected to alter the risk of developing several different disorders, but this has not been shown to date. Here we studied 38,638 individuals with 11 major diseases, and 17,483 healthy controls. Imputing missing variants from 7,729 complete mitochondrial genomes, we captured 40.41% of European mtDNA variation. We show that mtDNA variants modifying the risk of developing one disease also modify the risk of developing other diseases, thus providing independent replication of a disease association in different case and control cohorts. High-risk alleles were more common than protective alleles, indicating that mtDNA is not at equilibrium in the human population, and that recent mutations interact with nuclear loci to modify the risk of developing multiple common diseases.

  15. Clinical expression of Leber hereditary optic neuropathy is affected by the mitochondrial DNA-haplogroup background.

    NARCIS (Netherlands)

    Hudson, G.; Carelli, V.; Spruijt, L.; Gerards, M.; Mowbray, C.; Achilli, A.; Pyle, A.; Elson, J.; Howell, N.; Morgia, C. La; Valentino, M.L.; Huoponen, K.; Savontaus, M.L.; Nikoskelainen, E.; Sadun, A.A.; Salomao, S.R.; Belfort Jr, R.; Griffiths, P.; Man, P.Y.; Coo, R.F. de; Horvath, R.; Zeviani, M.; Smeets, H.J.M.; Torroni, A.; Chinnery, P.F.

    2007-01-01

    Leber hereditary optic neuropathy (LHON) is due primarily to one of three common point mutations of mitochondrial DNA (mtDNA), but the incomplete penetrance implicates additional genetic or environmental factors in the pathophysiology of the disorder. Both the 11778G-->A and 14484T-->C LHON mutation

  16. The elusive nature of adaptive mitochondrial DNA evolution of an Arctic lineage prone to frequent introgression

    DEFF Research Database (Denmark)

    Melo-Ferreira, Jose; Vilela, Joana; Fonseca, Miguel M.;

    2014-01-01

    Mitochondria play a fundamental role in cellular metabolism, being responsible for most of the energy production of the cell in the oxidative phosphorylation (OXPHOS) pathway. Mitochondrial DNA (mtDNA) encodes for key components of this process, but its direct role in adaptation remains far from...

  17. Introducing Human Population Biology through an Easy Laboratory Exercise on Mitochondrial DNA

    Science.gov (United States)

    Pardinas, Antonio F.; Dopico, Eduardo; Roca, Agustin; Garcia-Vazquez, Eva; Lopez, Belen

    2010-01-01

    This article describes an easy and cheap laboratory exercise for students to discover their own mitochondrial haplogroup. Students use buccal swabs to obtain mucosa cells as noninvasive tissue samples, extract DNA, and with a simple polymerase chain reaction-restriction fragment length polymorphism analysis they can obtain DNA fragments of…

  18. In vitro-reconstituted nucleoids can block mitochondrial DNA replication and transcription

    NARCIS (Netherlands)

    Farge, Géraldine; Mehmedovic, Majda; Baclayon, Marian; van den Wildenberg, Siet M J L; Roos, Wouter H; Gustafsson, Claes M; Wuite, Gijs J L; Falkenberg, Maria

    2014-01-01

    The mechanisms regulating the number of active copies of mtDNA are still unclear. A mammalian cell typically contains 1,000-10,000 copies of mtDNA, which are packaged into nucleoprotein complexes termed nucleoids. The main protein component of these structures is mitochondrial transcription factor A

  19. Sequencing strategy of mitochondrial HV1 and HV2 DNA with length heteroplasmy

    DEFF Research Database (Denmark)

    Rasmussen, Erik Michael; Sørensen, E; Eriksen, Birthe

    2002-01-01

    We describe a method to obtain reliable mitochondrial DNA (mtDNA) sequences downstream of the homopolymeric stretches with length heteroplasmy in the sequencing direction. The method is based on the use of junction primers that bind to a part of the homopolymeric stretch and the first 2-4 bases...

  20. Effects of Wolbachia on mitochondrial DNA variation in populations of Athetis lepigone (Lepidoptera: Noctuidae) in China

    Science.gov (United States)

    Wolbachia are endosymbiotic bacteria that infect arthropods and incompatibility among strains can affect gene flow within host insect populations, that can result in significant host mitochondrial DNA (MtD) variation. The effects of Wolbachia infection on mtDNA variation was studied in Athetis lepi...

  1. In vitro-reconstituted nucleoids can block mitochondrial DNA replication and transcription

    NARCIS (Netherlands)

    Farge, Géraldine; Mehmedovic, Majda; Baclayon, Marian; van den Wildenberg, Siet M J L; Roos, Wouter H; Gustafsson, Claes M; Wuite, Gijs J L; Falkenberg, Maria

    2014-01-01

    The mechanisms regulating the number of active copies of mtDNA are still unclear. A mammalian cell typically contains 1,000-10,000 copies of mtDNA, which are packaged into nucleoprotein complexes termed nucleoids. The main protein component of these structures is mitochondrial transcription factor A

  2. Introducing Human Population Biology through an Easy Laboratory Exercise on Mitochondrial DNA

    Science.gov (United States)

    Pardinas, Antonio F.; Dopico, Eduardo; Roca, Agustin; Garcia-Vazquez, Eva; Lopez, Belen

    2010-01-01

    This article describes an easy and cheap laboratory exercise for students to discover their own mitochondrial haplogroup. Students use buccal swabs to obtain mucosa cells as noninvasive tissue samples, extract DNA, and with a simple polymerase chain reaction-restriction fragment length polymorphism analysis they can obtain DNA fragments of…

  3. The Restriction Fragment Map of Rat-Liver Mitochondrial DNA : A Reconsideration

    NARCIS (Netherlands)

    Pepe, G.; Bakker, H.; Holtrop, M.; Bollen, J.E.; Bruggen, E.F.J. van; Cantatore, P.; Terpstra, P.; Saccone, C.

    1977-01-01

    1. Rat-liver mitochondrial DNA (mtDNA) contains at least 8 cleavage sites for the restriction endonuclease Eco RI, 6 for the restriction endonuclease Hind III, 2 for the restriction endonuclease Bam HI and 11 for the restriction endonuclease Hap II. 2. The physical map of the restriction fragments o

  4. Patterns of linkage disequilibrium in mitochondrial DNA of 16 ruminant populations.

    Science.gov (United States)

    Slate, J; Phua, S H

    2003-03-01

    Mitochondrial DNA (mtDNA) is a widely employed molecular tool in phylogeography, in the inference of human evolutionary history, in dating the domestication of livestock and in forensic science. In humans and other vertebrates the popularity of mtDNA can be partially attributed to an assumption of strict maternal inheritance, such that there is no recombination between mitochondrial lineages. The recent demonstration that linkage disequilibrium (LD) declines as a function of distance between polymorphic sites in hominid mitochondrial genomes has been interpreted as evidence of recombination between mtDNA haplotypes, and hence nonclonal inheritance. However, critics of mtDNA recombination have suggested that this association is an artefact of an inappropriate measure of LD or of sequencing error, and subsequent studies of other populations have failed to replicate the initial finding. Here we report the analysis of 16 ruminant populations and present evidence that LD significantly declines with distance in five of them. A meta-analysis of the data indicates a nonsignificant trend of LD declining with distance. Most of the earlier criticisms of patterns between LD and distance in hominid mtDNA are not applicable to this data set. Our results suggest that either ruminant mtDNA is not strictly clonal or that compensatory selection has influenced patterns of variation at closely linked sites within the mitochondrial control region. The potential impact of these processes should be considered when using mtDNA as a tool in vertebrate population genetic, phylogenetic and forensic studies.

  5. An improved method of mitochondrial DNA isolation for XL-PCR

    Institute of Scientific and Technical Information of China (English)

    SHI Duo; ZHU Ke-jun; WANG Xue-min; WANG Zhen-cheng; ZHENG Jian-ming; MIAO Ming-yong; JIAO Bing-hua

    2006-01-01

    Objective: To obtain high quality of mitochondrial DNA (mtDNA) and carry out extra-long PCR (XL-PCR). Methods: Mitochondria were isolated by differential centrifugation, and membranes were disrupted using 10%SDS (pH 7.0). mtDNA was then extracted using phenol and chloroform. Results: The mtDNA obtained by using our improved method can be used as effective template for XL-PCR,and total mtDNA (16 kb) can be amplified easily. Conclusion: Our improved method is effective in preparing high quality of mtDNA, which can be used as template for XL-PCR.

  6. The mitochondrial and plastid genomes of Volvox carteri: bloated molecules rich in repetitive DNA

    Directory of Open Access Journals (Sweden)

    Lee Robert W

    2009-03-01

    Full Text Available Abstract Background The magnitude of noncoding DNA in organelle genomes can vary significantly; it is argued that much of this variation is attributable to the dissemination of selfish DNA. The results of a previous study indicate that the mitochondrial DNA (mtDNA of the green alga Volvox carteri abounds with palindromic repeats, which appear to be selfish elements. We became interested in the evolution and distribution of these repeats when, during a cursory exploration of the V. carteri nuclear DNA (nucDNA and plastid DNA (ptDNA sequences, we found palindromic repeats with similar structural features to those of the mtDNA. Upon this discovery, we decided to investigate the diversity and evolutionary implications of these palindromic elements by sequencing and characterizing large portions of mtDNA and ptDNA and then comparing these data to the V. carteri draft nuclear genome sequence. Results We sequenced 30 and 420 kilobases (kb of the mitochondrial and plastid genomes of V. carteri, respectively – resulting in partial assemblies of these genomes. The mitochondrial genome is the most bloated green-algal mtDNA observed to date: ~61% of the sequence is noncoding, most of which is comprised of short palindromic repeats spread throughout the intergenic and intronic regions. The plastid genome is the largest (>420 kb and most expanded (>80% noncoding ptDNA sequence yet discovered, with a myriad of palindromic repeats in the noncoding regions, which have a similar size and secondary structure to those of the mtDNA. We found that 15 kb (~0.01% of the nuclear genome are homologous to the palindromic elements of the mtDNA, and 50 kb (~0.05% are homologous to those of the ptDNA. Conclusion Selfish elements in the form of short palindromic repeats have propagated in the V. carteri mtDNA and ptDNA, resulting in the distension of these genomes. Copies of these same repeats are also found in a small fraction of the nucDNA, but appear to be inert in this

  7. Comprehensive scanning of somatic mitochondrial DNA alterations in acute leukemia developing from myelodysplastic syndromes.

    Science.gov (United States)

    Linnartz, Bjoern; Anglmayer, Roswitha; Zanssen, Stefanie

    2004-03-15

    Myelodysplastic syndromes (MDS) are clonal myeloid disorders characterized by ineffective hematopoiesis resulting in refractory cytopenias. Transformation resulting in acute myeloblastic leukemia is the final stage in the multistep process of MDS evolution. Functional relevant mutations of mitochondrial DNA (mtDNA) have been related to sideroblastic anemia and MDS. To investigate the role of mtDNA in malignant transformation to acute leukemia, we used high-resolution techniques such as single-strand conformational polymorphism and fluorescence sequencing for investigation of the whole mitochondrial genome from blood cells of 10 patients with MDS. Functionally relevant point mutations in mitochondrial RNA and polypeptide-encoding genes were detected in 50% of patients with MDS. Their increasing mutation load connects MDS and the developing acute myeloid leukemias. Several point mutations of mtDNA, including secondary point mutations for Leber's hereditary optic neuropathy, occur in one bone marrow and may synergically affect bone marrow stem cells by an apoptotic pathway.

  8. Genetic characteristics of mitochondrial DNA was associated with colorectal carcinogenesis and its prognosis.

    Directory of Open Access Journals (Sweden)

    Jae-Ho Lee

    Full Text Available Clinical value of mitochondrial DNA has been described in colorectal cancer (CRC. To clarify its role in colorectal carcinogenesis, mitochondrial microsatellite instability (mtMSI and other markers were investigated in CRCs and their precancerous lesions, as a multitier genetic study. DNA was isolated from paired normal and tumoral tissues in 78 tubular adenomas (TAs, 34 serrated polyps (SPs, and 100 CRCs. mtMSI, nucleus microsatellite instability (nMSI, KRAS mutation, and BRAF mutation were investigated in these tumors and their statistical analysis was performed. mtMSI was found in 30% of CRCs and 21.4% of precancerous lesions. Mitochondrial copy number was higher in SPs than TAs and it was associated with mtMSI in low grade TAs. KRAS and BRAF mutations were mutually exclusive in TAs and SPs. CRCs with mtMSI showed shorter overall survival times than the patients without mtMSI. In CRCs without nMSI or BRAF mutation, mtMSI was a more accurate marker for predicting prognosis. The genetic change of mitochondrial DNA is an early and independent event in colorectal precancerous lesions and mtMSI and mitochondrial contents are associated with the tubular adenoma-carcinoma sequence, resulting in poor prognosis. This result suggested that the genetic change in mitochondrial DNA appears to be a possible prognosis marker in CRC.

  9. Interference of Co-Amplified Nuclear Mitochondrial DNA Sequences on the Determination of Human mtDNA Heteroplasmy by Using the SURVEYOR Nuclease and the WAVE HS System

    OpenAIRE

    Hsiu-Chuan Yen; Shiue-Li Li; Wei-Chien Hsu; Petrus Tang

    2014-01-01

    High-sensitivity and high-throughput mutation detection techniques are useful for screening the homoplasmy or heteroplasmy status of mitochondrial DNA (mtDNA), but might be susceptible to interference from nuclear mitochondrial DNA sequences (NUMTs) co-amplified during polymerase chain reaction (PCR). In this study, we first evaluated the platform of SURVEYOR Nuclease digestion of heteroduplexed DNA followed by the detection of cleaved DNA by using the WAVE HS System (SN/WAVE-HS) for detectin...

  10. Increased plasma levels of circulating cell-free mitochondrial DNA in suicide attempters: associations with HPA-axis hyperactivity

    OpenAIRE

    Lindqvist, D.; Fernström, J; Grudet, C; Ljunggren, L; Träskman-Bendz, L; Ohlsson, L; Westrin, Å

    2016-01-01

    Preclinical data suggest that chronic stress may cause cellular damage and mitochondrial dysfunction, potentially leading to the release of mitochondrial DNA (mtDNA) into the bloodstream. Major depressive disorder has been associated with an increased amount of mtDNA in leukocytes from saliva samples and blood; however, no previous studies have measured plasma levels of free-circulating mtDNA in a clinical psychiatric sample. In this study, free circulating mtDNA was quantified in plasma samp...

  11. Effect of ethidium bromide on transmission of mitochondrial genomes and DNA synthesis in the petite negative yeast Schizosaccharomyces pomhe.

    Science.gov (United States)

    Wolf, K; Del Giudice, L

    1980-04-01

    Treatment of haploid strains of the petite negative yeast Schizosaccharomyces pomhe with ethidium bromide prior to mating with untreated cells reduces transmission of mitochondrial markers from the treated strains. This effect is fully reversible after 20 generations of growth in drug free medium before mating. In contrast to the petite positive yeast Saccharomyces cerevisiae, where nuclear DNA synthesis is not affected but mitochondrial DNA is degraded in the presence of 20 μg/ml ethidium bromide, the same concentration decreases both nuclear and mitochondrial DNA synthesis in Schizosaccharomyces pomhe. After removal of the drug, nuclear DNA synthesis increases faster than its mitochondrial counterpart in Schizosaccharomyces pomhe.

  12. How do human cells react to the absence of mitochondrial DNA?

    Directory of Open Access Journals (Sweden)

    Rossana Mineri

    Full Text Available BACKGROUND: Mitochondrial biogenesis is under the control of two different genetic systems: the nuclear genome (nDNA and the mitochondrial genome (mtDNA. The mtDNA is a circular genome of 16.6 kb encoding 13 of the approximately 90 subunits that form the respiratory chain, the remaining ones being encoded by the nDNA. Eukaryotic cells are able to monitor and respond to changes in mitochondrial function through alterations in nuclear gene expression, a phenomenon first defined in yeast and known as retrograde regulation. To investigate how the cellular transcriptome is modified in response to the absence of mtDNA, we used Affymetrix HG-U133A GeneChip arrays to study the gene expression profile of two human cell lines, 143BTK(- and A549, which had been entirely depleted of mtDNA (rho(o cells, and compared it with that of corresponding undepleted parental cells (rho(+ cells. RESULTS: Our data indicate that absence of mtDNA is associated with: i a down-regulation of cell cycle control genes and a reduction of cell replication rate, ii a down-regulation of nuclear-encoded subunits of complex III of the respiratory chain and iii a down-regulation of a gene described as the human homolog of ELAC2 of E. coli, which encodes a protein that we show to also target to the mitochondrial compartment. CONCLUSIONS: Our results indicate a strong correlation between mitochondrial biogenesis and cell cycle control and suggest that some proteins could have a double role: for instance in controlling both cell cycle progression and mitochondrial functions. In addition, the finding that ELAC2 and maybe other transcripts that are located into mitochondria, are down-regulated in rho(o cells, make them good candidates for human disorders associated with defective replication and expression of mtDNA.

  13. Maternal inheritance and mitochondrial DNA variants in familial Parkinson's disease

    National Research Council Canada - National Science Library

    Simon, David K; Pankratz, Nathan; Kissell, Diane K; Pauciulo, Michael W; Halter, Cheryl A; Rudolph, Alice; Pfeiffer, Ronald F; Nichols, William C; Foroud, Tatiana

    2010-01-01

    .... We examined the possibility of a maternal inheritance bias as well as the association between mitochondrial haplogroups and maternal inheritance and disease risk in a case-control study of 168...

  14. Nucleotide pools dictate the identity and frequency of ribonucleotide incorporation in mitochondrial DNA

    Science.gov (United States)

    Hoberg, Emily; Szilagyi, Zsolt; Taylor, Robert W.; Gustafsson, Claes M.; Falkenberg, Maria

    2017-01-01

    Previous work has demonstrated the presence of ribonucleotides in human mitochondrial DNA (mtDNA) and in the present study we use a genome-wide approach to precisely map the location of these. We find that ribonucleotides are distributed evenly between the heavy- and light-strand of mtDNA. The relative levels of incorporated ribonucleotides reflect that DNA polymerase γ discriminates the four ribonucleotides differentially during DNA synthesis. The observed pattern is also dependent on the mitochondrial deoxyribonucleotide (dNTP) pools and disease-causing mutations that change these pools alter both the absolute and relative levels of incorporated ribonucleotides. Our analyses strongly suggest that DNA polymerase γ-dependent incorporation is the main source of ribonucleotides in mtDNA and argues against the existence of a mitochondrial ribonucleotide excision repair pathway in human cells. Furthermore, we clearly demonstrate that when dNTP pools are limiting, ribonucleotides serve as a source of building blocks to maintain DNA replication. Increased levels of embedded ribonucleotides in patient cells with disturbed nucleotide pools may contribute to a pathogenic mechanism that affects mtDNA stability and impair new rounds of mtDNA replication. PMID:28207748

  15. Mitochondrial DNA acquires immunogenicity on exposure to nitrosative stress in patients with vitiligo.

    Science.gov (United States)

    Al-Shobaili, Hani A; Rasheed, Zafar

    2014-10-01

    Vitiligo is a common pigmentary skin disorder of unknown etiology. Many studies show the defective mitochondrial functionality in vitiligo patients, but the potential role of mitochondrial DNA (mtDNA) in the pathogenesis of vitiligo remains to be investigated. Recent evidences demonstrate that mitochondria possess their own nitric-oxide-synthase and can produce endogenous peroxynitrite (ONOO(-)). This study was undertaken to investigate the role of ONOO(-)-modified-mitochondrial-DNA (ONOO(-)-mtDNA) in vitiligo autoimmunity. Our data revealed that ONOO(-)-induced modifications in mtDNA caused structural alterations. Specificity of immunoglobulin G (IgG) from vitiligo patients (n=26) and controls (n=25) were analysed towards ONOO(-)-mtDNA. Vitligo-IgG samples (Vt-IgG) show preferential binding to ONOO(-)-mtDNA in comparison with native mtDNA (pvitiligo patients. Furthermore, levels of anti-ONOO(-)-mtDNA-IgG, inducible-nitric-oxide-synthase (iNOS), nitric oxide (NO) and nitrotyrosine were higher among vitiligo patients whose disease durations (DD) were ⩾5 years as compared to patients with lower DD (DDvitiligo patients. Our data provide an important insight into the immunological mechanisms occur in vitiligo. The ONOO(-)-mtDNA may be useful in elucidating the mechanisms of disease pathogenesis.

  16. Gene therapy of mitochondrial DNA mutations: a brief, biased history of allotopic expression in mammalian cells.

    Science.gov (United States)

    Zullo, S J

    2001-09-01

    Successful treatment of mitochondrial DNA (mtDNA) mutations might be possible by construction of mtDNA-encoded protein genes so that they can be inserted into the nuclear genome and the protein expressed in the mitochondria (allotopic expression). This technique would require individual assembly of all 13 mtDNA-encoded protein genes with an aminoterminal leader peptide that directs the cytoplasmic translated protein to the mitochondrial membrane. The 13 allotopic genes could be inserted into the nuclear genome of a patient's stem cell that had been "cured" of its nascent mtDNA via ethidium bromide treatment (rho-zero cell). The rho-zero cell would be a uridine auxotroph, and recovery from uridine auxotrophy would indicate successful transformation. The patient's own cells could then be returned to the patient's body. With a selective advantage of recovered oxidative phosphorylation, the transformed cells could replace cells with mtDNA mutations. Results of experiments by us on allotopically expressed CHO ATPase6 and of experiments by other workers suggest that there might be competition with endogenous mtDNA-encoded proteins if the particular protein gene is not removed from the endogenous mitochondrial genomes. Thus, it is likely that all 13 mtDNA-encoded protein genes will need to be allotopically expressed, with concomitant removal of all mtDNA genomes, in order for this form of mtDNA gene therapy to be successful.

  17. Complete mitochondrial DNA sequence of the Qianshao spotted pig.

    Science.gov (United States)

    Xu, Dong; Chai, Yu-Lan; Jiang, Juan; He, Chang-Qing; Ma, Hai-Ming

    2015-01-01

    The complete mitochondrial genome sequence of Qianshao spotted pig was first determined in this study. The mitogenome (16,700 bp) consists of 22 tRNA genes, 2 ribosomal RNA genes, 13 protein-coding genes and 1 control region (D-loop region). The complete mitochondrial genome sequence of the Qianshao spotted pig enriches data resource for further study in genetic mechanism.

  18. The potential role for use of mitochondrial DNA copy number as predictive biomarker in presbycusis

    Directory of Open Access Journals (Sweden)

    Falah M

    2016-10-01

    Full Text Available Masoumeh Falah,1,2 Massoud Houshmand,3 Mohammad Najafi,2 Maryam Balali,1 Saeid Mahmoudian,1 Alimohamad Asghari,4 Hessamaldin Emamdjomeh,1 Mohammad Farhadi1 1ENT and Head & Neck Research Center and Department, Iran University of Medical Sciences, Tehran, Iran; 2Cellular and Molecular Research Center, Biochemistry Department, Iran University of Medical Sciences, Tehran, Iran; 3Department of Medical Genetics, National Institute for Genetic Engineering and Biotechnology, Tehran, Iran; 4Skull base research center, Iran University of Medical Sciences, Tehran, Iran Objectives: Age-related hearing impairment, or presbycusis, is the most common communication disorder and neurodegenerative disease in the elderly. Its prevalence is expected to increase, due to the trend of growth of the elderly population. The current diagnostic test for detection of presbycusis is implemented after there has been a change in hearing sensitivity. Identification of a pre-diagnostic biomarker would raise the possibility of preserving hearing sensitivity before damage occurs. Mitochondrial dysfunction, including the production of reactive oxygen species and induction of expression of apoptotic genes, participates in the progression of presbycusis. Mitochondrial DNA sequence variation has a critical role in presbycusis. However, the nature of the relationship between mitochondrial DNA copy number, an important biomarker in many other diseases, and presbycusis is undetermined.Methods: Fifty-four subjects with presbycusis and 29 healthy controls were selected after ear, nose, throat examination and pure-tone audiometry. DNA was extracted from peripheral blood samples. The copy number of mitochondrial DNA relative to the nuclear genome was measured by quantitative real-time polymerase chain reaction.Results: Subjects with presbycusis had a lower median mitochondrial DNA copy number than healthy subjects and the difference was statistically significant (P=0.007. Mitochondrial DNA

  19. Epstein-Barr Virus Immediate-Early Protein Zta Co-Opts Mitochondrial Single-Stranded DNA Binding Protein To Promote Viral and Inhibit Mitochondrial DNA Replication▿

    Science.gov (United States)

    Wiedmer, Andreas; Wang, Pu; Zhou, Jing; Rennekamp, Andrew J.; Tiranti, Valeria; Zeviani, Massimo; Lieberman, Paul M.

    2008-01-01

    Disruption of cellular metabolic processes and usurpation of host proteins are hallmarks of herpesvirus lytic infection. Epstein-Barr virus (EBV) lytic replication is initiated by the immediate-early protein Zta. Zta is a multifunctional DNA binding protein that stimulates viral gene transcription, nucleates a replication complex at the viral origin of lytic replication, and inhibits cell cycle proliferation. To better understand these functions and identify cellular collaborators of Zta, we purified an epitope-tagged version of Zta in cells capable of supporting lytic replication. FLAG-tagged Zta was purified from a nuclear fraction using FLAG antibody immunopurification and peptide elution. Zta-associated proteins were isolated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and identified by mass spectrometry. The Zta-associated proteins included members of the HSP70 family and various single-stranded DNA and RNA binding proteins. The nuclear replication protein A subunits (RPA70 and RPA32) and the human mitochondrial single-stranded DNA binding protein (mtSSB) were confirmed by Western blotting to be specifically enriched in the FLAG-Zta immunopurified complex. mtSSB coimmunoprecipitated with endogenous Zta during reactivation of EBV-positive Burkitt lymphoma and lymphoblastoid cell lines. Small interfering RNA depletion of mtSSB reduced Zta-induced lytic replication of EBV but had only a modest effect on transcription activation function. A point mutation in the Zta DNA binding domain (C189S), which is known to reduce lytic cycle replication, eliminated mtSSB association with Zta. The predominantly mitochondrial localization of mtSSB was shifted to partly nuclear localization in cells expressing Zta. Mitochondrial DNA synthesis and genome copy number were reduced by Zta-induced EBV lytic replication. We conclude that Zta interaction with mtSSB serves the dual function of facilitating viral and blocking mitochondrial DNA replication. PMID:18305033

  20. Epstein-Barr virus immediate-early protein Zta co-opts mitochondrial single-stranded DNA binding protein to promote viral and inhibit mitochondrial DNA replication.

    Science.gov (United States)

    Wiedmer, Andreas; Wang, Pu; Zhou, Jing; Rennekamp, Andrew J; Tiranti, Valeria; Zeviani, Massimo; Lieberman, Paul M

    2008-05-01

    Disruption of cellular metabolic processes and usurpation of host proteins are hallmarks of herpesvirus lytic infection. Epstein-Barr virus (EBV) lytic replication is initiated by the immediate-early protein Zta. Zta is a multifunctional DNA binding protein that stimulates viral gene transcription, nucleates a replication complex at the viral origin of lytic replication, and inhibits cell cycle proliferation. To better understand these functions and identify cellular collaborators of Zta, we purified an epitope-tagged version of Zta in cells capable of supporting lytic replication. FLAG-tagged Zta was purified from a nuclear fraction using FLAG antibody immunopurification and peptide elution. Zta-associated proteins were isolated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and identified by mass spectrometry. The Zta-associated proteins included members of the HSP70 family and various single-stranded DNA and RNA binding proteins. The nuclear replication protein A subunits (RPA70 and RPA32) and the human mitochondrial single-stranded DNA binding protein (mtSSB) were confirmed by Western blotting to be specifically enriched in the FLAG-Zta immunopurified complex. mtSSB coimmunoprecipitated with endogenous Zta during reactivation of EBV-positive Burkitt lymphoma and lymphoblastoid cell lines. Small interfering RNA depletion of mtSSB reduced Zta-induced lytic replication of EBV but had only a modest effect on transcription activation function. A point mutation in the Zta DNA binding domain (C189S), which is known to reduce lytic cycle replication, eliminated mtSSB association with Zta. The predominantly mitochondrial localization of mtSSB was shifted to partly nuclear localization in cells expressing Zta. Mitochondrial DNA synthesis and genome copy number were reduced by Zta-induced EBV lytic replication. We conclude that Zta interaction with mtSSB serves the dual function of facilitating viral and blocking mitochondrial DNA replication.

  1. Interaction of human mitochondrial transcription factor A in mitochondria: its involvement in the dynamics of mitochondrial DNA nucleoids.

    Science.gov (United States)

    Kasashima, Katsumi; Endo, Hitoshi

    2015-12-01

    Mitochondrial transcription factor A (TFAM) is a key regulator of mitochondrial DNA (mtDNA). TFAM interacts with itself and forms dimers; however, the precise interaction domain in vivo has not yet been determined. We herein showed that human TFAM formed oligomers in mitochondria by in situ chemical cross-linking. We used the separated fluorescent protein, monomeric Kusabira-Green, as a reporter to monitor their self-association in mitochondria. This reporter successfully detected the TFAM-TFAM interaction in cells as fluorescent signals on mitochondria. We also found that the N-terminal high-mobility group box domain was sufficient for this interaction. The expression of the dimer-defective mutant induced enlarged mtDNA nucleoids, suggesting the importance of dimerization in the distribution of mtDNA. The reporter system also supported the association and mixture between independent nucleoids through TFAM by a cell fusion assay using hemagglutinating virus of Japan. We here, for the first time, visualized the interaction of TFAM molecules in mitochondria and proposed its implications for the dynamics of mtDNA nucleoids.

  2. Mitochondrial DNA T4216C and A4917G variations in multiple sclerosis

    DEFF Research Database (Denmark)

    Andalib, Sasan; Talebi, Mahnaz; Sakhinia, Ebrahim;

    2015-01-01

    BACKGROUND: Multiple sclerosis (MS) affects the brain and spinal cord and long has been the topic of global research; yet there is no commonly accepted cause and no cure for the disease. Mounting evidence supports the role of genetics in susceptibility to MS. From this perspective, a current effort...... focuses on the neurogenetics of the complex pathogenesis of MS in relation to factors such as mitochondrial DNA (mtDNA) variations. T4216C and A4917G are common mitochondrial gene variations associated with MS. The present study tested whether mtDNA T4216C variation in the NADH Dehydrogenase 1 (ND1) mtDNA...... gene and A4917G variation in the mtDNA NADH Dehydrogenase 2 (ND2) gene are associated with MS in an Iranian population. MATERIAL AND METHODS: Blood samples were collected from 100 patients with MS and 100 unrelated healthy controls, and DNA extraction was performed by salting-out. By means...

  3. Aspects of Ancient Mitochondrial DNA Analysis in Different Populations for Understanding Human Evolution

    Directory of Open Access Journals (Sweden)

    D.V. Nesheva

    2014-06-01

    Full Text Available The evolution of modern humans is a long and difficult process which started from their first appearance and continues to the present day. The study of the genetic origin of populations can help to determine population kinship and to better understand the gradual changes of the gene pool in space and time. Mitochondrial DNA (mtDNA is a proper tool for the determination of the origin of populations due to its high evolutionary importance. Ancient mitochondrial DNA retrieved from museum specimens, archaeological finds and fossil remains can provide direct evidence for population origins and migration processes. Despite the problems with contaminations and authenticity of ancient mitochondrial DNA, there is a developed set of criteria and platforms for obtaining authentic ancient DNA. During the last two decades, the application of different methods and techniques for analysis of ancient mitochondrial DNA gave promising results. Still, the literature is relatively poor with information for the origin of human populations. Using comprehensive phylogeographic and population analyses we can observe the development and formation of the contemporary populations. The aim of this study was to shed light on human migratory processes and the formation of populations based on available ancient mtDNA data.

  4. Visualization of mitochondrial DNA replication in individual cells by EdU signal amplification.

    Science.gov (United States)

    Haines, Kristine M; Feldman, Eva L; Lentz, Stephen I

    2010-11-15

    Mitochondria are key regulators of cellular energy and mitochondrial biogenesis is an essential component of regulating mitochondria numbers in healthy cells. One approach for monitoring mitochondrial biogenesis is to measure the rate of mitochondrial DNA (mtDNA) replication. We developed a sensitive technique to label newly synthesized mtDNA in individual cells in order to study mtDNA biogenesis. The technique combines the incorporation of 5-ethynyl-2'-deoxyuridine (EdU) with a tyramide signal amplification (TSA) protocol to visualize mtDNA replication within subcellular compartments of neurons. EdU is superior to other thymidine analogs, such as 5-bromo-2-deoxyuridine (BrdU), because the initial click reaction to label EdU does not require the harsh acid treatments or enzyme digests that are required for exposing the BrdU epitope. The milder labeling of EdU allows for direct comparison of its incorporation with other cellular markers. The ability to visualize and quantify mtDNA biogenesis provides an essential tool for investigating the mechanisms used to regulate mitochondrial biogenesis and would provide insight into the pathogenesis associated with drug toxicity, aging, cancer and neurodegenerative diseases. Our technique is applicable to sensory neurons as well as other cell types. The use of this technique to measure mtDNA biogenesis has significant implications in furthering the understanding of both normal cellular physiology as well as impaired disease states.

  5. Mitochondrial DNA Unwinding Enzyme Required for Liver Regeneration | Center for Cancer Research

    Science.gov (United States)

    The liver has an exceptional capacity to proliferate. This ability allows the liver to regenerate its mass after partial surgical removal or injury and is the key to successful partial liver transplants. Liver cells, called hepatocytes, are packed with mitochondria, and regulating mitochondrial DNA (mtDNA) copy number is crucial to mitochondrial function, including energy production, during proliferation. Yves Pommier, M.D., Ph.D., of CCR’s Developmental Therapeutics Branch, and his colleagues recently showed that the vertebrate mitochondrial topoisomerase, Top1mt, was critical in maintaining mitochondrial function in the heart after doxorubicin-induced damage. The group wondered whether Top1mt might play a similar role in liver regeneration.

  6. Mitochondrial DNA (mtDNA) biogenesis: visualization and duel incorporation of BrdU and EdU into newly synthesized mtDNA in vitro.

    Science.gov (United States)

    Lentz, Stephen I; Edwards, James L; Backus, Carey; McLean, Lisa L; Haines, Kristine M; Feldman, Eva L

    2010-02-01

    Mitochondria are key regulators of cellular energy and are the focus of a large number of studies examining the regulation of mitochondrial dynamics and biogenesis in healthy and diseased conditions. One approach to monitoring mitochondrial biogenesis is to measure the rate of mitochondrial DNA (mtDNA) replication. We developed a sensitive technique to visualize newly synthesized mtDNA in individual cells to study mtDNA replication within subcellular compartments of neurons. The technique combines the incorporation of 5-bromo-2-deoxyuridine (BrdU) and/or 5-ethynyl-2'-deoxyuridine (EdU) into mtDNA, together with a tyramide signal amplification protocol. Employing this technique, we visualized and measured mtDNA biogenesis in individual cells. The labeling procedure for EdU allows for more comprehensive results by allowing the comparison of its incorporation with other intracellular markers, because it does not require the harsh acid or enzyme digests necessary to recover the BrdU epitope. In addition, the utilization of both BrdU and EdU permits sequential pulse-chase experiments to follow the intracellular localization of mtDNA replication. The ability to quantify mitochondrial biogenesis provides an essential tool for investigating the alterations in mitochondrial dynamics involved in the pathogenesis of multiple cellular disorders, including neuropathies and neurodegenerative diseases.

  7. Kearns-Sayre syndrome: different amounts of deleted mitochondrial DNA are present in several autoptic tissues.

    Science.gov (United States)

    Ponzetto, C; Bresolin, N; Bordoni, A; Moggio, M; Meola, G; Bet, L; Prelle, A; Scarlato, G

    1990-05-01

    A population of deleted mitochondrial DNA (mtDNA) was found in different amounts in autoptic muscle, heart, cortex, cerebellum, liver and kidney of a patient who died of Kearn-Sayre Syndrome (KSS). The widespread occurrence of the deletion correlates with the multisystem nature of KSS and supports the hypothesis that this is a genetic disease due to an alteration of mtDNA presumably arising in the oocyte or early embryo.

  8. Decreased Integrity, Content, and Increased Transcript Level of Mitochondrial DNA Are Associated with Keratoconus

    Science.gov (United States)

    Hao, Xiao-Dan; Chen, Zhao-Li; Qu, Ming-Li; Zhao, Xiao-Wen; Li, Su-Xia; Chen, Peng

    2016-01-01

    Oxidative stress may play an important role in the pathogenesis of keratoconus (KC). Mitochondrial DNA (mtDNA) is involved in mitochondrial function, and the mtDNA content, integrity, and transcript level may affect the generation of reactive oxygen species (ROS) and be involved in the pathogenesis of KC. We designed a case-control study to research the relationship between KC and mtDNA integrity, content and transcription. One-hundred ninety-eight KC corneas and 106 normal corneas from Chinese patients were studied. Quantitative real-time PCR was used to measure the relative mtDNA content, transcript levels of mtDNA and related genes. Long-extension PCR was used to detect mtDNA damage. ROS, mitochondrial membrane potential and ATP were measured by respective assay kit, and Mito-Tracker Green was used to label the mitochondria. The relative mtDNA content of KC corneas was significantly lower than that of normal corneas (P = 9.19×10−24), possibly due to decreased expression of the mitochondrial transcription factor A (TFAM) gene (P = 3.26×10−3). In contrast, the transcript levels of mtDNA genes were significantly increased in KC corneas compared with normal corneas (NADH dehydrogenase subunit 1 [ND1]: P = 1.79×10−3; cytochrome c oxidase subunit 1 [COX1]: P = 1.54×10−3; NADH dehydrogenase subunit 1, [ND6]: P = 4.62×10−3). The latter may be the result of increased expression levels of mtDNA transcription-related genes mitochondrial RNA polymerase (POLRMT) (P = 2.55×10−4) and transcription factor B2 mitochondrial (TFB2M) (P = 7.88×10−5). KC corneas also had increased mtDNA damage (P = 3.63×10−10), higher ROS levels, and lower mitochondrial membrane potential and ATP levels compared with normal corneas. Decreased integrity, content and increased transcript level of mtDNA are associated with KC. These changes may affect the generation of ROS and play a role in the pathogenesis of KC. PMID:27783701

  9. Mitochondrial DNA dynamics during in vitro culture and pluripotency induction of a bovine Rho0 cell line.

    Science.gov (United States)

    Pessôa, L V F; Bressan, F F; Chiaratti, M R; Pires, P R L; Perecin, F; Smith, L C; Meirelles, F V

    2015-10-30

    Large number of cellular changes and diseases are related to mutations in the mitochondrial DNA copy number. Cell culture in the presence of ethidium bromide is a known way of depleting mitochondrial DNA and is a useful model for studying such conditions. Interestingly, the morphology of these depleted cells resembles that of pluripotent cells, as they present larger and fragmented mitochondria with poorly developed cristae. Herein, we aimed to study the mechanisms responsible for the control of mitochondrial DNA replication during mitochondrial DNA depletion mediated by ethidium bromide and during the in vitro induction of cellular pluripotency with exogenous transcription factor expression in a bovine model. This article reports the generation of a bovine Rho0 mesenchymal cell line and describes the analysis of mitochondrial DNA copy number in a time-dependent manner. The expression of apoptosis and mitochondrial-related genes in the cells during mitochondrial DNA repletion were also analyzed. The dynamics of mitochondrial DNA during both the depletion process and in vitro reprogramming are discussed. It was possible to obtain bovine mesenchymal cells almost completely depleted of their mitochondrial DNA content (over 90%). However, the production of induced pluripotent stem cells from the transduction of both control and Rho0 bovine mesenchymal cells with human reprograming factors was not successful.

  10. Glom Is a Novel Mitochondrial DNA Packaging Protein in Physarum polycephalum and Causes Intense Chromatin Condensation without Suppressing DNA Functions

    Science.gov (United States)

    Sasaki, Narie; Kuroiwa, Haruko; Nishitani, Chikako; Takano, Hiroyoshi; Higashiyama, Tetsuya; Kobayashi, Tamaki; Shirai, Yuki; Sakai, Atsushi; Kawano, Shigeyuki; Murakami-Murofushi, Kimiko; Kuroiwa, Tsuneyoshi

    2003-01-01

    Mitochondrial DNA (mtDNA) is packed into highly organized structures called mitochondrial nucleoids (mt-nucleoids). To understand the organization of mtDNA and the overall regulation of its genetic activity within the mt-nucleoids, we identified and characterized a novel mtDNA packaging protein, termed Glom (a protein inducing agglomeration of mitochondrial chromosome), from highly condensed mt-nucleoids of the true slime mold, Physarum polycephalum. This protein could bind to the entire mtDNA and package mtDNA into a highly condensed state in vitro. Immunostaining analysis showed that Glom specifically localized throughout the mt-nucleoid. Deduced amino acid sequence revealed that Glom has a lysine-rich region with proline-rich domain in the N-terminal half and two HMG boxes in C-terminal half. Deletion analysis of Glom revealed that the lysine-rich region was sufficient for the intense mtDNA condensation in vitro. When the recombinant Glom proteins containing the lysine-rich region were expressed in Escherichia coli, the condensed nucleoid structures were observed in E. coli. Such in vivo condensation did not interfere with transcription or replication of E. coli chromosome and the proline-rich domain was essential to keep those genetic activities. The expression of Glom also complemented the E. coli mutant lacking the bacterial histone-like protein HU and the HMG-boxes region of Glom was important for the complementation. Our results suggest that Glom is a new mitochondrial histone-like protein having a property to cause intense DNA condensation without suppressing DNA functions. PMID:12960433

  11. Mitochondrial DNA variant discovery and evaluation in human Cardiomyopathies through next-generation sequencing.

    Directory of Open Access Journals (Sweden)

    Michael V Zaragoza

    Full Text Available Mutations in mitochondrial DNA (mtDNA may cause maternally-inherited cardiomyopathy and heart failure. In homoplasmy all mtDNA copies contain the mutation. In heteroplasmy there is a mixture of normal and mutant copies of mtDNA. The clinical phenotype of an affected individual depends on the type of genetic defect and the ratios of mutant and normal mtDNA in affected tissues. We aimed at determining the sensitivity of next-generation sequencing compared to Sanger sequencing for mutation detection in patients with mitochondrial cardiomyopathy. We studied 18 patients with mitochondrial cardiomyopathy and two with suspected mitochondrial disease. We "shotgun" sequenced PCR-amplified mtDNA and multiplexed using a single run on Roche's 454 Genome Sequencer. By mapping to the reference sequence, we obtained 1,300x average coverage per case and identified high-confidence variants. By comparing these to >400 mtDNA substitution variants detected by Sanger, we found 98% concordance in variant detection. Simulation studies showed that >95% of the homoplasmic variants were detected at a minimum sequence coverage of 20x while heteroplasmic variants required >200x coverage. Several Sanger "misses" were detected by 454 sequencing. These included the novel heteroplasmic 7501T>C in tRNA serine 1 in a patient with sudden cardiac death. These results support a potential role of next-generation sequencing in the discovery of novel mtDNA variants with heteroplasmy below the level reliably detected with Sanger sequencing. We hope that this will assist in the identification of mtDNA mutations and key genetic determinants for cardiomyopathy and mitochondrial disease.

  12. Genotype-Phenotype Correlation of Maternally Inherited Disorders due to Mutations in Mitochondrial DNA

    Directory of Open Access Journals (Sweden)

    Peterus Thajeb

    2006-09-01

    Full Text Available Mitochondrial disorders are heterogeneous systemic ailments that are most often caused by maternal inheritance of a variety of mutations of the mitochondrial (mt DNA. Paternal inheritance and somatic mutation are rare. The disorders are well recognized not only for the genotypic heterogeneity, but also the phenotypic variation among the affected members of a single family. The genotype-phenotype correlation of the diversity of the syndromic and non-syndromic features of mitochondrial disorders are discussed. Some aspects of the molecular mechanisms of this heterogeneity, and the histopathologic findings are highlighted.

  13. Energy, ageing, fidelity and sex: oocyte mitochondrial DNA as a protected genetic template.

    Science.gov (United States)

    de Paula, Wilson B M; Lucas, Cathy H; Agip, Ahmed-Noor A; Vizcay-Barrena, Gema; Allen, John F

    2013-07-19

    Oxidative phosphorylation couples ATP synthesis to respiratory electron transport. In eukaryotes, this coupling occurs in mitochondria, which carry DNA. Respiratory electron transport in the presence of molecular oxygen generates free radicals, reactive oxygen species (ROS), which are mutagenic. In animals, mutational damage to mitochondrial DNA therefore accumulates within the lifespan of the individual. Fertilization generally requires motility of one gamete, and motility requires ATP. It has been proposed that oxidative phosphorylation is nevertheless absent in the special case of quiescent, template mitochondria, that these remain sequestered in oocytes and female germ lines and that oocyte mitochondrial DNA is thus protected from damage, but evidence to support that view has hitherto been lacking. Here we show that female gametes of Aurelia aurita, the common jellyfish, do not transcribe mitochondrial DNA, lack electron transport, and produce no free radicals. In contrast, male gametes actively transcribe mitochondrial genes for respiratory chain components and produce ROS. Electron microscopy shows that this functional division of labour between sperm and egg is accompanied by contrasting mitochondrial morphology. We suggest that mitochondrial anisogamy underlies division of any animal species into two sexes with complementary roles in sexual reproduction. We predict that quiescent oocyte mitochondria contain DNA as an unexpressed template that avoids mutational accumulation by being transmitted through the female germ line. The active descendants of oocyte mitochondria perform oxidative phosphorylation in somatic cells and in male gametes of each new generation, and the mutations that they accumulated are not inherited. We propose that the avoidance of ROS-dependent mutation is the evolutionary pressure underlying maternal mitochondrial inheritance and the developmental origin of the female germ line.

  14. Do All Hemochromatosis Patients Have the Same Origin? A Pilot Study of Mitochondrial DNA and Y-DNA

    Directory of Open Access Journals (Sweden)

    Caitlin J Symonette

    2011-01-01

    Full Text Available BACKGROUND: Mitochondrial DNA (mtDNA and Y-DNA analysis have been widely used to predict ancestral origin. Genetic anthropologists predict that human civilizations may have originated in central Africa one to two million years previously. Primary iron overload is not a common diagnosis among indigenous people of northern Africa, but hereditary hemochromatosis is present in approximately one in 200 people in northern Europe. MtDNA analysis has the potential to determine whether contemporary hemochromatosis patients have an ancient ancestral linkage.

  15. Mitochondrial DNA G13708A variation and multiple sclerosis: Is there an association?

    DEFF Research Database (Denmark)

    Andalib, S.; Talebi, M.; Sakhinia, E.

    2017-01-01

    Background Multiple sclerosis (MS) is considered a pathogenetic enigma. Recently, efforts to implicate genetics in human susceptibility to MS have identified an important role of mitochondrial DNA (mtDNA). G13708A is a common mtDNA variation associated with MS in specific populations. This study...... tested the hypothesis that the mtDNA G13708A variation is associated with MS in an Iranian population. Materials and methods Blood samples were collected from 100 MS patients and 100 unrelated healthy controls. DNA was extracted using a salting-out method, followed by polymerase chain reaction (PCR...... in the selected Iranian population....

  16. Mutations in the D-loop region of mitochondrial DNA in gastric cancer

    Institute of Scientific and Technical Information of China (English)

    Yibing Zhao; Hongyu Yang; Guoyu Chen

    2005-01-01

    Objective: To investigate the mutations in the D-loop region of mitochondrial DNA (mtDNA) in gastric cancer.Methods: The mtDNA of D-loop region was amplified by PCR and sequenced in 20 samples from gastric cancer tissue and adjacent normal membrane. Results: There were 7/20(35% ) mutations in the mtDNA of D-loop region in gastric cancer patients. There were four microsatellite instabilities among the 18 mutations. Nine new polymorphisms were identified in 20 patients. Conclusion: The mtDNA of Dloop region might be highly polymorphoric and the mutation rate is high in patients with gastric cancer.

  17. Mitochondrial capture enriches mito-DNA 100 fold, enabling PCR-free mitogenomics biodiversity analysis

    DEFF Research Database (Denmark)

    Liu, Shanlin; Wang, Xin; Xie, Lin

    2016-01-01

    . The major impediment of such a method is the lack of appropriate mitochondrial DNA enrichment ways. Because mitochondrial genomes (mitogenomes) make up only a small proportion of total DNA, PCR-free methods will inevitably result in a huge excess of data (>99%). Furthermore, the massive volume of sequence...... data is highly demanding on computing resources. Here, we present a mitogenome enrichment pipeline via a gene capture chip that was designed by virtue of the mitogenome sequences of the 1000 Insect Transcriptome Evolution project (1KITE, www.1kite.org). A mock sample containing 49 species was used...... to evaluate the efficiency of the mitogenome capture method. We demonstrate that the proportion of mitochondrial DNA can be increased by approximately 100-fold (from the original 0.47% to 42.52%). Variation in phylogenetic distances of target taxa to the probe set could in principle result in bias...

  18. OPA1-related dominant optic atrophy is not strongly influenced by mitochondrial DNA background

    Directory of Open Access Journals (Sweden)

    Amati-Bonneau Patrizia

    2009-07-01

    Full Text Available Abstract Background Leber's hereditary optic neuropathy (LHON and autosomal dominant optic atrophy (ADOA are the most frequent forms of hereditary optic neuropathies. LHON is associated with mitochondrial DNA (mtDNA mutations whereas ADOA is mainly due to mutations in the OPA1 gene that encodes a mitochondrial protein involved in the mitochondrial inner membrane remodeling. A striking influence of mtDNA haplogroup J on LHON expression has been demonstrated and it has been recently suggested that this haplogroup could also influence ADOA expression. In this study, we have tested the influence of mtDNA backgrounds on OPA1 mutations. Methods To define the relationships between OPA1 mutations and mtDNA backgrounds, we determined the haplogroup affiliation of 41 French patients affected by OPA1-related ADOA by control-region sequencing and RFLP survey of their mtDNAs. Results The comparison between patient and reference populations did not revealed any significant difference. Conclusion Our results argue against a strong influence of mtDNA background on ADOA expression. These data allow to conclude that OPA1 could be considered as a "severe mutation", directly responsible of the optic atrophy, whereas OPA1-negative ADOA and LHON mutations need an external factor(s to express the pathology (i.e. synergistic interaction with mitochondrial background.

  19. In Situ Labeling of Mitochondrial DNA Replication in Drosophila Adult Ovaries by EdU Staining.

    Science.gov (United States)

    Chen, Zhe; Xu, Hong

    2016-10-15

    The mitochondrial genome is inherited exclusively through the maternal line. Understanding of how the mitochondrion and its genome are proliferated and transmitted from one generation to the next through the female oocyte is of fundamental importance. Because of the genetic tractability, and the elegant, ordered simplicity by which oocyte development proceeds, Drosophila oogenesis has become an invaluable system for mitochondrial study. An EdU (5-ethynyl-2´-deoxyuridine) labeling method was utilized to detect mitochondrial DNA (mtDNA) replication in Drosophila ovaries. This method is superior to the BrdU (5-bromo-2'-deoxyuridine) labeling method in that it allows for good structural preservation and efficient fluorescent dye penetration of whole-mount tissues. Here we describe a detailed protocol for labeling replicating mitochondrial DNA in Drosophila adult ovaries with EdU. Some technical solutions are offered to improve the viability of the ovaries, maintain their health during preparation, and ensure high-quality imaging. Visualization of newly synthesized mtDNA in the ovaries not only reveals the striking temporal and spatial pattern of mtDNA replication through oogenesis, but also allows for simple quantification of mtDNA replication under various genetic and pharmacological perturbations.

  20. Ribosomal and Mitochondrial DNA Analyses of Xiphinema americanum-Group Populations.

    Science.gov (United States)

    Lazarova, Stela S; Malloch, Gaynor; Oliveira, Claudio M G; Hübschen, Judith; Neilson, Roy

    2006-12-01

    The 18S ribosomal DNA (rDNA) and cytochrome oxidase I region of mitochondrial DNA (mtDNA) were sequenced for 24 Xiphinema americanum-group populations sourced from a number of geographically disparate locations. Sequences were subjected to phylogenetic analysis and compared. 18S rDNA strongly suggested that only X. pachtaicum, X. simile (two populations) and a X. americanum s.l. population from Portugal were different from the other 20 populations studied, whereas mtDNA indicated some heterogeneity between populations. Phylogenetically, based on mtDNA, an apparent dichotomy existed amongst X. americanum-group populations from North America and those from Asia, South America and Oceania. Analyses of 18S rDNA and mtDNA sequences underpin the classical taxonomic issues of the X. americanum-group and cast doubt on the degree of speciation within the X. americanum-group.

  1. Retrospective assessment of the most common mitochondrial DNA mutations in a large Hungarian cohort of suspect mitochondrial cases.

    Science.gov (United States)

    Remenyi, Viktoria; Inczedy-Farkas, Gabriella; Komlosi, Katalin; Horvath, Rita; Maasz, Anita; Janicsek, Ingrid; Pentelenyi, Klara; Gal, Aniko; Karcagi, Veronika; Melegh, Bela; Molnar, Maria Judit

    2015-08-01

    Prevalence estimations for mitochondrial disorders still vary widely and only few epidemiologic studies have been carried out so far. With the present work we aim to give a comprehensive overview about frequencies of the most common mitochondrial mutations in Hungarian patients. A total of 1328 patients were tested between 1999 and 2012. Among them, 882 were screened for the m.3243A > G, m.8344A > G, m.8993T > C/G mutations and deletions, 446 for LHON primary mutations. The mutation frequency in our cohort was 2.61% for the m.3243A > G, 1.47% for the m.8344A > G, 17.94% for Leber's Hereditary Optic Neuropathy (m.3460G > A, m.11778G > A, m.14484T > C) and 0.45% for the m.8993T > C/G substitutions. Single mtDNA deletions were detected in 14.97%, while multiple deletions in 6.01% of the cases. The mutation frequency in Hungarian patients suggestive of mitochondrial disease was similar to other Caucasian populations. Further retrospective studies of different populations are needed in order to accurately assess the importance of mitochondrial diseases and manage these patients.

  2. 常见线粒体DNA病的分子遗传学研究进展%Molecular genetics of common mitochondrial DNA disorders

    Institute of Scientific and Technical Information of China (English)

    Lee-Jun C. WONG

    2005-01-01

    SUMMARY Diagnosis of mitochondrial disorders has been difficult due to the clinical and genetic heterogeneity, as well as unique features of mitochondrial genetics. Definitive diagnosis requires the identification of molecular defects in either the mitochondrial or the nuclear genome. We describe the clinical and molecular characteristic of some common mitochondrial syndromes and molecular methodologies available for the detection of mitochondrial DNA mutations. This review provides overview of current molecular diagnosis of mitochondrial DNA disorders that is useful in patient care and genetic counseling.

  3. Cardiometabolic phenotypes and mitochondrial DNA copy number in two cohorts of UK women.

    Science.gov (United States)

    Guyatt, Anna L; Burrows, Kimberley; Guthrie, Philip A I; Ring, Sue; McArdle, Wendy; Day, Ian N M; Ascione, Raimondo; Lawlor, Debbie A; Gaunt, Tom R; Rodriguez, Santiago

    2017-08-15

    The mitochondrial genome is present at variable copy number between individuals. Mitochondria are vulnerable to oxidative stress, and their dysfunction may be associated with cardiovascular disease. The association of mitochondrial DNA copy number with cardiometabolic risk factors (lipids, glycaemic traits, inflammatory markers, anthropometry and blood pressure) was assessed in two independent cohorts of European origin women, one in whom outcomes were measured at mean (SD) age 30 (4.3) years (N=2278) and the second at 69.4 (5.5) years (N=2872). Mitochondrial DNA copy number was assayed by quantitative polymerase chain reaction. Associations were adjusted for smoking, sociodemographic status, laboratory factors and white cell traits. Out of a total of 12 outcomes assessed in both cohorts, mitochondrial DNA copy number showed little or no association with the majority (point estimates were close to zero and nearly all p-values were >0.01). The strongest evidence was for an inverse association in the older cohort with insulin (standardised beta [95%CI]: -0.06, [-0.098, -0.022], p=0.002), but this association did not replicate in the younger cohort. Our findings do not provide support for variation in mitochondrial DNA copy number having an important impact on cardio-metabolic risk factors in European origin women. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

  4. Human REV3 DNA Polymerase Zeta Localizes to Mitochondria and Protects the Mitochondrial Genome.

    Science.gov (United States)

    Singh, Bhupendra; Li, Xiurong; Owens, Kjerstin M; Vanniarajan, Ayyasamy; Liang, Ping; Singh, Keshav K

    2015-01-01

    To date, mitochondrial DNA polymerase γ (POLG) is the only polymerase known to be present in mammalian mitochondria. A dogma in the mitochondria field is that there is no other polymerase present in the mitochondria of mammalian cells. Here we demonstrate localization of REV3 DNA polymerase in the mammalian mitochondria. We demonstrate localization of REV3 in the mitochondria of mammalian tissue as well as cell lines. REV3 associates with POLG and mitochondrial DNA and protects the mitochondrial genome from DNA damage. Inactivation of Rev3 leads to reduced mitochondrial membrane potential, reduced OXPHOS activity, and increased glucose consumption. Conversely, inhibition of the OXPHOS increases expression of Rev3. Rev3 expression is increased in human primary breast tumors and breast cancer cell lines. Inactivation of Rev3 decreases cell migration and invasion, and localization of Rev3 in mitochondria increases survival and the invasive potential of cancer cells. Taken together, we demonstrate that REV3 functions in mammalian mitochondria and that mitochondrial REV3 is associated with the tumorigenic potential of cells.

  5. An alternative model for the early peopling of southern South America revealed by analyses of three mitochondrial DNA haplogroups

    National Research Council Canada - National Science Library

    de Saint Pierre, Michelle; Bravi, Claudio M; Motti, Josefina M B; Fuku, Noriyuki; Tanaka, Masashi; Llop, Elena; Bonatto, Sandro L; Moraga, Mauricio

    2012-01-01

    ... seven complete mitochondrial DNA sequences. We identified two novel mtDNA monophyletic clades, preliminarily designated B2l and C1b13, which together with the recently described D1g sub-haplogroup have locally high frequencies...

  6. Mitochondrial DNA backgrounds might modulate diabetes complications rather than T2DM as a whole.

    Directory of Open Access Journals (Sweden)

    Alessandro Achilli

    Full Text Available Mitochondrial dysfunction has been implicated in rare and common forms of type 2 diabetes (T2DM. Additionally, rare mitochondrial DNA (mtDNA mutations have been shown to be causal for T2DM pathogenesis. So far, many studies have investigated the possibility that mtDNA variation might affect the risk of T2DM, however, when found, haplogroup association has been rarely replicated, even in related populations, possibly due to an inadequate level of haplogroup resolution. Effects of mtDNA variation on diabetes complications have also been proposed. However, additional studies evaluating the mitochondrial role on both T2DM and related complications are badly needed. To test the hypothesis of a mitochondrial genome effect on diabetes and its complications, we genotyped the mtDNAs of 466 T2DM patients and 438 controls from a regional population of central Italy (Marche. Based on the most updated mtDNA phylogeny, all 904 samples were classified into 57 different mitochondrial sub-haplogroups, thus reaching an unprecedented level of resolution. We then evaluated whether the susceptibility of developing T2DM or its complications differed among the identified haplogroups, considering also the potential effects of phenotypical and clinical variables. MtDNA backgrounds, even when based on a refined haplogroup classification, do not appear to play a role in developing T2DM despite a possible protective effect for the common European haplogroup H1, which harbors the G3010A transition in the MTRNR2 gene. In contrast, our data indicate that different mitochondrial haplogroups are significantly associated with an increased risk of specific diabetes complications: H (the most frequent European haplogroup with retinopathy, H3 with neuropathy, U3 with nephropathy, and V with renal failure.

  7. Mitochondrial DNA variation and HIV-associated sensory neuropathy in CHARTER.

    Science.gov (United States)

    Holzinger, Emily R; Hulgan, Todd; Ellis, Ronald J; Samuels, David C; Ritchie, Marylyn D; Haas, David W; Kallianpur, Asha R; Bloss, Cinnamon S; Clifford, David B; Collier, Ann C; Gelman, Benjamin B; Marra, Christina M; McArthur, Justin C; McCutchan, J Allen; Morgello, Susan; Simpson, David M; Franklin, Donald R; Rosario, Debralee; Selph, Doug; Letendre, Scott; Grant, Igor

    2012-12-01

    HIV-associated sensory neuropathy remains an important complication of combination antiretroviral therapy and HIV infection. Mitochondrial DNA haplogroups and single nucleotide polymorphisms (SNPs) have previously been associated with symptomatic neuropathy in clinical trial participants. We examined associations between mitochondrial DNA variation and HIV-associated sensory neuropathy in CNS HIV Antiretroviral Therapy Effects Research (CHARTER). CHARTER is a USA-based longitudinal observational study of HIV-infected adults who underwent a structured interview and standardized examination. HIV-associated sensory neuropathy was determined by trained examiners as ≥1 sign (diminished vibratory and sharp-dull discrimination or ankle reflexes) bilaterally. Mitochondrial DNA sequencing was performed and haplogroups were assigned by published algorithms. Multivariable logistic regression of associations between mitochondrial DNA SNPs, haplogroups, and HIV-associated sensory neuropathy were performed. In analyses of associations of each mitochondrial DNA SNP with HIV-associated sensory neuropathy, the two most significant SNPs were at positions A12810G [odds ratio (95 % confidence interval) = 0.27 (0.11-0.65); p = 0.004] and T489C [odds ratio (95 % confidence interval) = 0.41 (0.21-0.80); p = 0.009]. These synonymous changes are known to define African haplogroup L1c and European haplogroup J, respectively. Both haplogroups were associated with decreased prevalence of HIV-associated sensory neuropathy compared with all other haplogroups [odds ratio (95 % confidence interval) = 0.29 (0.12-0.71); p = 0.007 and odds ratio (95 % confidence interval) = 0.42 (0.18-1.0); p = 0.05, respectively]. In conclusion, in this cohort of mostly combination antiretroviral therapy-treated subjects, two common mitochondrial DNA SNPs and their corresponding haplogroups were associated with a markedly decreased prevalence of HIV-associated sensory neuropathy.

  8. One-way sequencing of multiple amplicons from tandem repetitive mitochondrial DNA control region.

    Science.gov (United States)

    Xu, Jiawu; Fonseca, Dina M

    2011-10-01

    Repetitive DNA sequences not only exist abundantly in eukaryotic nuclear genomes, but also occur as tandem repeats in many animal mitochondrial DNA (mtDNA) control regions. Due to concerted evolution, these repetitive sequences are highly similar or even identical within a genome. When long repetitive regions are the targets of amplification for the purpose of sequencing, multiple amplicons may result if one primer has to be located inside the repeats. Here, we show that, without separating these amplicons by gel purification or cloning, directly sequencing the mitochondrial repeats with the primer outside repetitive region is feasible and efficient. We exemplify it by sequencing the mtDNA control region of the mosquito Aedes albopictus, which harbors typical large tandem DNA repeats. This one-way sequencing strategy is optimal for population surveys.

  9. Maternal age effect and severe germ-line bottleneck in the inheritance of human mitochondrial DNA

    DEFF Research Database (Denmark)

    Rebolledo-Jaramillo, Boris; Su, Marcia Shu-Wei; Stoler, Nicholas

    2014-01-01

    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......, 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....

  10. Lack of founding Amerindian mitochondrial DNA lineages in extinct aborigines from Tierra del Fuego-Patagonia.

    Science.gov (United States)

    Lalueza, C; Pérez-Pérez, A; Prats, E; Cornudella, L; Turbón, D

    1997-01-01

    Ancient DNA from bones and teeth of 60 individuals from four extinct human populations from Tierra del Fuego-Patagonia (Selknam, Yamana, Kaweskar and Aonikenk) has been extracted and the mitochondrial DNA (mtDNA) amplified by using the polymerase chain reaction. High-resolution analysis of endonuclease restriction site variation in the mtDNA and sequencing of its hypervariable non-coding control region, revealed complete absence of two of the four primary mitochondrial haplotype groups present in contemporary Amerinds, namely A and B. In contrast, haplogroups C and D were found in all but one sample with frequencies of approximately 38% and 60%. These results, together with the decreasing incidence of group A in more southerly latitudes in the American continent and the absence of cluster B above 55 degrees North in America and Asia, argue that the first settlers entering America 21000-14000 years ago already lacked both mtDNA lineages.

  11. Do Alterations in Mitochondrial DNA Play a Role in Breast Carcinogenesis?

    Directory of Open Access Journals (Sweden)

    Thomas E. Rohan

    2010-01-01

    Full Text Available A considerable body of evidence supports a role for oxidative stress in breast carcinogenesis. Due to their role in producing energy via oxidative phosphorylation, the mitochondria are a major source of production of reactive oxygen species, which may damage DNA. The mitochondrial genome may be particularly susceptible to oxidative damage leading to mitochondrial dysfunction. Genetic variants in mtDNA and nuclear DNA may also contribute to mitochondrial dysfunction. In this review, we address the role of alterations in mtDNA in the etiology of breast cancer. Several studies have shown a relatively high frequency of mtDNA mutations in breast tumor tissue in comparison with mutations in normal breast tissue. To date, several studies have examined the association of genetic variants in mtDNA and breast cancer risk. The G10398A mtDNA polymorphism has received the most attention and has been shown to be associated with increased risk in some studies. Other variants have generally been examined in only one or two studies. Genome-wide association studies may help identify new mtDNA variants which modify breast cancer risk. In addition to assessing the main effects of specific variants, gene-gene and gene-environment interactions are likely to explain a greater proportion of the variability in breast cancer risk.

  12. Do alterations in mitochondrial DNA play a role in breast carcinogenesis?

    Science.gov (United States)

    Rohan, Thomas E; Wong, Lee-Jun; Wang, Tao; Haines, Jonathan; Kabat, Geoffrey C

    2010-01-01

    A considerable body of evidence supports a role for oxidative stress in breast carcinogenesis. Due to their role in producing energy via oxidative phosphorylation, the mitochondria are a major source of production of reactive oxygen species, which may damage DNA. The mitochondrial genome may be particularly susceptible to oxidative damage leading to mitochondrial dysfunction. Genetic variants in mtDNA and nuclear DNA may also contribute to mitochondrial dysfunction. In this review, we address the role of alterations in mtDNA in the etiology of breast cancer. Several studies have shown a relatively high frequency of mtDNA mutations in breast tumor tissue in comparison with mutations in normal breast tissue. To date, several studies have examined the association of genetic variants in mtDNA and breast cancer risk. The G10398A mtDNA polymorphism has received the most attention and has been shown to be associated with increased risk in some studies. Other variants have generally been examined in only one or two studies. Genome-wide association studies may help identify new mtDNA variants which modify breast cancer risk. In addition to assessing the main effects of specific variants, gene-gene and gene-environment interactions are likely to explain a greater proportion of the variability in breast cancer risk.

  13. Telomere length is correlated with mitochondrial DNA copy number in intestinal, but not diffuse, gastric cancer.

    Science.gov (United States)

    Jung, Soo-Jung; Cho, Ji-Hyoung; Park, Won-Jin; Heo, Yu-Ran; Lee, Jae-Ho

    2017-07-01

    A positive correlation between telomere length and mitochondrial DNA (mtDNA) copy number has previously been observed in healthy individuals, and in patients with psychiatric disorders. In the present study, telomere length and mtDNA copy number were evaluated in gastric cancer (GC) tissue samples. DNA was extracted from 109 GC samples (including 82 intestinal, and 27 diffuse cases), and the telomere length and mtDNA copy number were analyzed using a quantitative-polymerase chain reaction assay. The relative telomere length and mtDNA copy number in tumor tissue, as compared with in normal tissue, (mean ± standard deviation) in all GC samples were 11.48±1.14 and 14.86±1.35, respectively. Telomere length and mtDNA copy number were not identified as exhibiting clinical or prognostic value for GC. However, positive correlations between telomere length and mitochondrial DNA copy number were identified in GC (r=0.408, P<0.001) and in the adjacent normal mucosa (r=0.363; P<0.001). When stratified by Lauren classification, the correlation was identified in intestinal type GC samples (r=0.461; P<0.001), but not in diffuse type GC samples (r=0.225; P=0.260). This result indicated that loss of the correlation of telomeres and mitochondrial function may induce the initiation or progression of GC pathogenesis.

  14. Mitochondrial DNA from archived tissue samples kept in formalin for forensic odontology studies.

    Science.gov (United States)

    Pandey, Rahul; Mehrotra, Divya; Kowtal, Pradnya; Mahdi, Abbas Ali; Sarin, Rajiv

    2014-01-01

    Samples used for DNA isolation to be used for forensic odontology studies are often limited. The possibility to use tissue samples stored in formalin for a prolonged period, which contains nucleic acids of questionable quality, opens exciting possibilities for genetic and molecular biology studies useful in speciality of forensic odontology. The present study defines substantial modification of existing protocols for total genomic isolation including mitochondrial DNA and proves the utility of such obtained mitochondrial DNA in microsatellite analyses. 50 dental tissue samples which were kept in neutral buffered formalin liquid bottles were taken for DNA isolation and subsequent analysis. For the isolation of total genomic DNA from tissue samples, a new protocol with substantial modifications from routine ones was adopted by us. Total genomic DNA from matched blood samples were extracted using standard phenol-chloroform extraction method. Polymerase Chain Reaction and Sequencing of such extracted DNA samples for mitochondrial D loop region were successful and the results were comparable with DNA extracted from normal sources of samples. The present study reports for the first time that nucleic acids extracted from human dental tissue samples under prolonged formalin fixation times can be used for forensic odontology studies using the described methodology.

  15. Mitonuclear coevolution as the genesis of speciation and the mitochondrial DNA barcode gap.

    Science.gov (United States)

    Hill, Geoffrey E

    2016-08-01

    Mitochondrial genes are widely used in taxonomy and systematics because high mutation rates lead to rapid sequence divergence and because such changes have long been assumed to be neutral with respect to function. In particular, the nucleotide sequence of the mitochondrial gene cytochrome c oxidase subunit 1 has been established as a highly effective DNA barcode for diagnosing the species boundaries of animals. Rarely considered in discussions of mitochondrial evolution in the context of systematics, speciation, or DNA barcodes, however, is the genomic architecture of the eukaryotes: Mitochondrial and nuclear genes must function in tight coordination to produce the complexes of the electron transport chain and enable cellular respiration. Coadaptation of these interacting gene products is essential for organism function. I extend the hypothesis that mitonuclear interactions are integral to the process of speciation. To maintain mitonuclear coadaptation, nuclear genes, which code for proteins in mitochondria that cofunction with the products of mitochondrial genes, must coevolve with rapidly changing mitochondrial genes. Mitonuclear coevolution in isolated populations leads to speciation because population-specific mitonuclear coadaptations create between-population mitonuclear incompatibilities and hence barriers to gene flow between populations. In addition, selection for adaptive divergence of products of mitochondrial genes, particularly in response to climate or altitude, can lead to rapid fixation of novel mitochondrial genotypes between populations and consequently to disruption in gene flow between populations as the initiating step in animal speciation. By this model, the defining characteristic of a metazoan species is a coadapted mitonuclear genotype that is incompatible with the coadapted mitochondrial and nuclear genotype of any other population.

  16. Paternal inheritance of mitochondrial DNA in the sheep (Ovine aries)

    National Research Council Canada - National Science Library

    Zhao, Xingbo; Chu, Mingxing; Li, Ning; Wu, Changxin

    2001-01-01

    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...

  17. Decreased skeletal muscle mitochondrial DNA in patients with statin-induced myopathy.

    Science.gov (United States)

    Stringer, Henry A J; Sohi, Gurmeet K; Maguire, John A; Côté, Hélène C F

    2013-02-15

    Statins are widely used to treat hyperlipidemia and lower cardiovascular disease risk. While statins are generally well tolerated, some patients experience statin-induced myopathy (SIM). Statin treatment has been associated with mitochondrial dysfunction and mitochondrial DNA (mtDNA) depletion. In this retrospective study, skeletal muscle biopsies from patients diagnosed with SIM were studied. These were compared with biopsies from patients clinically assessed as having statin-unrelated myopathy but whose biopsy showed no or negligible pathology. For each biopsy sample, mtDNA was quantified relative to nuclear DNA (mtDNA content) by qPCR, mtDNA deletions were investigated by long-template PCR followed by gel densitometry, and mtDNA oxidative damage was quantified using a qPCR-based assay. For a subset of matched samples, mtDNA heteroplasmy and mutations were investigated by cloning/sequencing. Skeletal muscle mtDNA content was significantly lower in SIM patients (N=23, mean±SD, 2036±1146) than in comparators (N=24, 3220±1594), p=0.006. There was no difference in mtDNA deletion score or oxidative mtDNA damage between the two groups, and no evidence of increased mtDNA heteroplasmy or somatic mutations was detected. The significant difference in skeletal muscle mtDNA suggests that SIM or statin treatments are associated with depletion of skeletal muscle mtDNA or that patients with an underlying predisposition to SIM have lower mtDNA levels. If statins induce mtDNA depletion, this would likely reflect decreased mitochondria biogenesis and/or increased mitochondria autophagy. Further work is necessary to distinguish between the lower mtDNA as a predisposition to SIM or an effect of SIM or statin treatment.

  18. Oxidative stress is not a major contributor to somatic mitochondrial DNA mutations.

    Directory of Open Access Journals (Sweden)

    Leslie S Itsara

    2014-02-01

    Full Text Available The accumulation of somatic mitochondrial DNA (mtDNA mutations is implicated in aging and common diseases of the elderly, including cancer and neurodegenerative disease. However, the mechanisms that influence the frequency of somatic mtDNA mutations are poorly understood. To develop a simple invertebrate model system to address this matter, we used the Random Mutation Capture (RMC assay to characterize the age-dependent frequency and distribution of mtDNA mutations in the fruit fly Drosophila melanogaster. Because oxidative stress is a major suspect in the age-dependent accumulation of somatic mtDNA mutations, we also used the RMC assay to explore the influence of oxidative stress on the somatic mtDNA mutation frequency. We found that many of the features associated with mtDNA mutations in vertebrates are conserved in Drosophila, including a comparable somatic mtDNA mutation frequency (∼10(-5, an increased frequency of mtDNA mutations with age, and a prevalence of transition mutations. Only a small fraction of the mtDNA mutations detected in young or old animals were G∶C to T∶A transversions, a signature of oxidative damage, and loss-of-function mutations in the mitochondrial superoxide dismutase, Sod2, had no detectable influence on the somatic mtDNA mutation frequency. Moreover, a loss-of-function mutation in Ogg1, which encodes a DNA repair enzyme that removes oxidatively damaged deoxyguanosine residues (8-hydroxy-2'-deoxyguanosine, did not significantly influence the somatic mtDNA mutation frequency of Sod2 mutants. Together, these findings indicate that oxidative stress is not a major cause of somatic mtDNA mutations. Our data instead suggests that somatic mtDNA mutations arise primarily from errors that occur during mtDNA replication. Further studies using Drosophila should aid in the identification of factors that influence the frequency of somatic mtDNA mutations.

  19. What cost mitochondria? The maintenance of functional mitochondrial DNA within and across generations.

    Science.gov (United States)

    Aanen, Duur K; Spelbrink, Johannes N; Beekman, Madeleine

    2014-07-05

    The peculiar biology of mitochondrial DNA (mtDNA) potentially has detrimental consequences for organismal health and lifespan. Typically, eukaryotic cells contain multiple mitochondria, each with multiple mtDNA genomes. The high copy number of mtDNA implies that selection on mtDNA functionality is relaxed. Furthermore, because mtDNA replication is not strictly regulated, within-cell selection may favour mtDNA variants with a replication advantage, but a deleterious effect on cell fitness. The opportunities for selfish mtDNA mutations to spread are restricted by various organism-level adaptations, such as uniparental transmission, germline mtDNA bottlenecks, germline selection and, during somatic growth, regular alternation between fusion and fission of mitochondria. These mechanisms are all hypothesized to maintain functional mtDNA. However, the strength of selection for maintenance of functional mtDNA progressively declines with age, resulting in age-related diseases. Furthermore, organismal adaptations that most probably evolved to restrict the opportunities for selfish mtDNA create secondary problems. Owing to predominantly maternal mtDNA transmission, recombination among mtDNA from different individuals is highly restricted or absent, reducing the scope for repair. Moreover, maternal inheritance precludes selection against mtDNA variants with male-specific effects. We finish by discussing the consequences of life-history differences among taxa with respect to mtDNA evolution and make a case for the use of microorganisms to experimentally manipulate levels of selection.

  20. Comparison of the fine structure of mitochondrial DNA from Saccharomyces cerevisiae and S. carlsbergensis: electron microscopy of partially denatured molecules

    DEFF Research Database (Denmark)

    Christiansen, Gunna; Christiansen, C

    1976-01-01

    Denaturation-maps of mitochondrial DNA from Saccharomyces cerevisiae and S. carlsbergensis have been derived from electron microscopic observations of partially denatured complete circular molecules and large fragments of these circles. The mitochondrial DNA from the two species differ by 6...

  1. Mitochondrial DNA deletion and impairment of mitochondrial biogenesis are mediated by reactive oxygen species in ionizing radiation-induced premature senescence

    Energy Technology Data Exchange (ETDEWEB)

    Eom, Hyeon Soo; Jung, U Hee; Jo, Sung Kee [Radiation Biotechnology Research Division, Korea Atomic Energy Research Institute, Daejeon (Korea, Republic of); Kim, Young Sang [College of Natural Sciences, Chungnam National University, Daejeon (Korea, Republic of)

    2011-09-15

    Mitochondrial DNA (mtDNA) deletion is a well-known marker for oxidative stress and aging, and contributes to harmful effects in cultured cells and animal tissues. mtDNA biogenesis genes (NRF-1, TFAM) are essential for the maintenance of mtDNA, as well as the transcription and replication of mitochondrial genomes. Considering that oxidative stress is known to affect mitochondrial biogenesis, we hypothesized that ionizing radiation (IR)-induced reactive oxygen species (ROS) causes mtDNA deletion by modulating the mitochondrial biogenesis, thereby leading to cellular senescence. Therefore, we examined the effects of IR on ROS levels, cellular senescence, mitochondrial biogenesis, and mtDNA deletion in IMR-90 human lung fibroblast cells. Young IMR-90 cells at population doubling (PD) 39 were irradiated at 4 or 8 Gy. Old cells at PD55, and H2O2-treated young cells at PD 39, were compared as a positive control. The IR increased the intracellular ROS level, senescence-associated {beta}-galactosidase (SA-{beta}-gal) activity, and mtDNA common deletion (4977 bp), and it decreased the mRNA expression of NRF-1 and TFAM in IMR-90 cells. Similar results were also observed in old cells (PD 55) and H{sub 2}O{sub 2}-treated young cells. To confirm that a increase in ROS level is essential for mtDNA deletion and changes of mitochondrial biogenesis in irradiated cells, the effects of N-acetylcysteine (NAC) were examined. In irradiated and H{sub 2}O{sub 2}-treated cells, 5 mM NAC significantly attenuated the increases of ROS, mtDNA deletion, and SA-{beta}-gal activity, and recovered from decreased expressions of NRF-1 and TFAM mRNA. These results suggest that ROS is a key cause of IR-induced mtDNA deletion, and the suppression of the mitochondrial biogenesis gene may mediate this process.

  2. Resistance training in patients with single, large-scale deletions of mitochondrial DNA.

    Science.gov (United States)

    Murphy, Julie L; Blakely, Emma L; Schaefer, Andrew M; He, Langping; Wyrick, Phil; Haller, Ronald G; Taylor, Robert W; Turnbull, Douglass M; Taivassalo, Tanja

    2008-11-01

    Dramatic tissue variation in mitochondrial heteroplasmy has been found to exist in patients with sporadic mitochondrial DNA (mtDNA) mutations. Despite high abundance in mature skeletal muscle, levels of the causative mutation are low or undetectable in satellite cells. The activation of these typically quiescent mitotic cells and subsequent shifting of wild-type mtDNA templates to mature muscle have been proposed as a means of restoring a more normal mitochondrial genotype and function in these patients. Because resistance exercise is known to serve as a stimulus for satellite cell induction within active skeletal muscle, this study sought to assess the therapeutic potential of resistance training in eight patients with single, large-scale mtDNA deletions by assessing: physiological determinants of peak muscle strength and oxidative capacity and muscle biopsy-derived measures of damage, mtDNA mutation load, level of oxidative impairment and satellite cell numbers. Our results show that 12 weeks of progressive overload leg resistance training led to: (i) increased muscle strength; (ii) myofibre damage and regeneration; (iii) increased proportion of neural cell adhesion molecule (NCAM)-positive satellite cells; (iv) improved muscle oxidative capacity. Taken together, we believe these findings support the hypothesis of resistance exercise-induced mitochondrial gene-shifting in muscle containing satellite cells which have low or absent levels of deleted mtDNA. Further investigation is warranted to refine parameters of the exercise training protocol in order to maximize the training effect on mitochondrial genotype and treatment potential for patients with selected, sporadic mutations of mtDNA in skeletal muscle.

  3. Association between Leukocyte Mitochondrial DNA Copy Number and Regular Exercise in Postmenopausal Women.

    Science.gov (United States)

    Chang, Yu Kyung; Kim, Da Eun; Cho, Soo Hyun; Kim, Jung-Ha

    2016-11-01

    Previous studies suggest that habitual exercise can improve skeletal mitochondrial function; however, to date, the association between exercise and mitochondrial function in peripheral leukocytes has not been reported. The aim of this study was to evaluate the relationship between regular exercise and mitochondrial function by measuring leukocyte mitochondrial DNA (mtDNA) copy number in postmenopausal women. This cross-sectional study included 144 relatively healthy, non-diabetic, non-smoking, postmenopausal women. Clinical parameters, including anthropometric measurements and cardio-metabolic parameters, were assessed. Regular exercise was defined as at least 150 minutes per week of moderate-intensity activity, or an equivalent combination of moderate and vigorous-intensity activity, over a duration of at least 6 months. Leukocyte mtDNA copy numbers were measured using real-time polymerase chain reaction assays, and these were normalized to the β-globin copy number to give the relative mtDNA copy number. The mtDNA copy number of peripheral leukocytes was significantly greater in the exercise group (1.33±0.02) than in the no exercise group (1.05±0.02, Pcopy number (β=0.25, Pcopy number in postmenopausal women.

  4. Reassessing the role of mitochondrial DNA mutations in autism spectrum disorder

    Directory of Open Access Journals (Sweden)

    Carracedo Ángel

    2011-04-01

    Full Text Available Abstract Background There is increasing evidence that impairment of mitochondrial energy metabolism plays an important role in the pathophysiology of autism spectrum disorders (ASD; OMIM number: 209850. A significant proportion of ASD cases display biochemical alterations suggestive of mitochondrial dysfunction and several studies have reported that mutations in the mitochondrial DNA (mtDNA molecule could be involved in the disease phenotype. Methods We analysed a cohort of 148 patients with idiopathic ASD for a number of mutations proposed in the literature as pathogenic in ASD. We also carried out a case control association study for the most common European haplogroups (hgs and their diagnostic single nucleotide polymorphisms (SNPs by comparing cases with 753 healthy and ethnically matched controls. Results We did not find statistical support for an association between mtDNA mutations or polymorphisms and ASD. Conclusions Our results are compatible with the idea that mtDNA mutations are not a relevant cause of ASD and the frequent observation of concomitant mitochondrial dysfunction and ASD could be due to nuclear factors influencing mitochondrion functions or to a more complex interplay between the nucleus and the mitochondrion/mtDNA.

  5. Mitochondrial-targeted DNA delivery using a DF-MITO-Porter, an innovative nano carrier with cytoplasmic and mitochondrial fusogenic envelopes

    Energy Technology Data Exchange (ETDEWEB)

    Yamada, Yuma; Kawamura, Eriko; Harashima, Hideyoshi, E-mail: harasima@pharm.hokudai.ac.jp [Hokkaido University, Laboratory for Molecular Design of Pharmaceutics, Faculty of Pharmaceutical Sciences (Japan)

    2012-08-15

    Mitochondrial gene therapy has the potential for curing a variety of diseases that are associated with mitochondrial DNA mutations and/or defects. To achieve this, it will be necessary to deliver therapeutic agents into the mitochondria in diseased cells. A number of mitochondrial drug delivery systems have been reported to date. However, reports of mitochondrial-targeted DNA delivery are limited. To achieve this, the therapeutic agent must be taken up by the cell (1), after which, the multi-processes associated with intracellular trafficking must be sophisticatedly regulated so as to release the agent from the endosome and deliver it to the cytosol (2) and to pass through the mitochondrial membrane (3). We report herein on the mitochondrial delivery of oligo DNA as a model therapeutic using a Dual Function (DF)-MITO-Porter, an innovative nano carrier designed for mitochondrial delivery. The critical structural elements of the DF-MITO-Porter include mitochondria-fusogenic inner envelopes and endosome-fusogenic outer envelopes, modified with octaarginine which greatly assists in cellular uptake. Inside the cell, the carrier passes through the endosomal and mitochondrial membranes via step-wise membrane fusion. When the oligo DNA was packaged in the DF-MITO-Porter, cellular uptake efficiency was strongly enhanced. Intracellular observation using confocal laser scanning microscopy showed that the DF-MITO-Porter was effectively released from endosomes. Moreover, the findings confirmed that the mitochondrial targeting activity of the DF-MITO-Porter was significantly higher than that of a carrier without outer endosome-fusogenic envelopes. These results support the conclusion that mitochondrial-targeted DNA delivery using a DF-MITO-Porter can be achieved when intracellular trafficking is optimally regulated.

  6. Simultaneous detection of human mitochondrial DNA and nuclear-inserted mitochondrial-origin sequences (NumtS) using forensic mtDNA amplification strategies and pyrosequencing technology.

    Science.gov (United States)

    Bintz, Brittania J; Dixon, Groves B; Wilson, Mark R

    2014-07-01

    Next-generation sequencing technologies enable the identification of minor mitochondrial DNA variants with higher sensitivity than Sanger methods, allowing for enhanced identification of minor variants. In this study, mixtures of human mtDNA control region amplicons were subjected to pyrosequencing to determine the detection threshold of the Roche GS Junior(®) instrument (Roche Applied Science, Indianapolis, IN). In addition to expected variants, a set of reproducible variants was consistently found in reads from one particular amplicon. A BLASTn search of the variant sequence revealed identity to a segment of a 611-bp nuclear insertion of the mitochondrial control region (NumtS) spanning the primer-binding sites of this amplicon (Nature 1995;378:489). Primers (Hum Genet 2012;131:757; Hum Biol 1996;68:847) flanking the insertion were used to confirm the presence or absence of the NumtS in buccal DNA extracts from twenty donors. These results further our understanding of human mtDNA variation and are expected to have a positive impact on the interpretation of mtDNA profiles using deep-sequencing methods in casework.

  7. Liver ultrastructural morphology and mitochondrial DNA levels in HIV/hepatitis C virus coinfection: no evidence of mitochondrial damage with highly active antiretroviral therapy.

    Science.gov (United States)

    Matsukura, Motoi; Chu, Fanny F S; Au, May; Lu, Helen; Chen, Jennifer; Rietkerk, Sonja; Barrios, Rolando; Farley, John D; Montaner, Julio S; Montessori, Valentina C; Walker, David C; Côté, Hélène C F

    2008-06-19

    Liver mitochondrial toxicity is a concern, particularly in HIV/hepatitis C virus (HCV) coinfection. Liver biopsies from HIV/HCV co-infected patients, 14 ON-highly active antiretroviral therapy (HAART) and nine OFF-HAART, were assessed by electron microscopy quantitative morphometric analyses. Hepatocytes tended to be larger ON-HAART than OFF-HAART (P = 0.05), but mitochondrial volume, cristae density, lipid volume, mitochondrial DNA and RNA levels were similar. We found no evidence of increased mitochondrial toxicity in individuals currently on HAART, suggesting that concomitant HAART should not delay HCV therapy.

  8. MPV17 encodes an inner mitochondrial membrane protein and is mutated in infantile hepatic mitochondrial DNA depletion.

    Science.gov (United States)

    Spinazzola, Antonella; Viscomi, Carlo; Fernandez-Vizarra, Erika; Carrara, Franco; D'Adamo, Pio; Calvo, Sarah; Marsano, René Massimiliano; Donnini, Claudia; Weiher, Hans; Strisciuglio, Pietro; Parini, Rossella; Sarzi, Emmanuelle; Chan, Alicia; DiMauro, Salvatore; Rötig, Agnes; Gasparini, Paolo; Ferrero, Iliana; Mootha, Vamsi K; Tiranti, Valeria; Zeviani, Massimo

    2006-05-01

    The mitochondrial (mt) DNA depletion syndromes (MDDS) are genetic disorders characterized by a severe, tissue-specific decrease of mtDNA copy number, leading to organ failure. There are two main clinical presentations: myopathic (OMIM 609560) and hepatocerebral (OMIM 251880). Known mutant genes, including TK2, SUCLA2, DGUOK and POLG, account for only a fraction of MDDS cases. We found a new locus for hepatocerebral MDDS on chromosome 2p21-23 and prioritized the genes on this locus using a new integrative genomics strategy. One of the top-scoring candidates was the human ortholog of the mouse kidney disease gene Mpv17. We found disease-segregating mutations in three families with hepatocerebral MDDS and demonstrated that, contrary to the alleged peroxisomal localization of the MPV17 gene product, MPV17 is a mitochondrial inner membrane protein, and its absence or malfunction causes oxidative phosphorylation (OXPHOS) failure and mtDNA depletion, not only in affected individuals but also in Mpv17-/- mice.

  9. Novel large-range mitochondrial DNA deletions and fatal multisystemic disorder with prominent hepatopathy

    Energy Technology Data Exchange (ETDEWEB)

    Bianchi, Marzia; Rizza, Teresa; Verrigni, Daniela [Unit of Molecular Medicine for Neuromuscular and Neurodegenerative Diseases, ' Bambino Gesu' Children' s Hospital, Rome (Italy); Martinelli, Diego [Division of Metabolism, ' Bambino Gesu' Children' s Hospital, Rome (Italy); Tozzi, Giulia; Torraco, Alessandra; Piemonte, Fiorella [Unit of Molecular Medicine for Neuromuscular and Neurodegenerative Diseases, ' Bambino Gesu' Children' s Hospital, Rome (Italy); Dionisi-Vici, Carlo [Division of Metabolism, ' Bambino Gesu' Children' s Hospital, Rome (Italy); Nobili, Valerio [Gastroenterology and Liver Unit, ' Bambino Gesu' Children' s Hospital, Rome (Italy); Francalanci, Paola; Boldrini, Renata; Callea, Francesco [Dept. Pathology, ' Bambino Gesu' Children' s Hospital, Rome (Italy); Santorelli, Filippo Maria [UOC Neurogenetica e Malattie Neuromuscolari, Fondazione Stella Maris, Pisa (Italy); Bertini, Enrico [Unit of Molecular Medicine for Neuromuscular and Neurodegenerative Diseases, ' Bambino Gesu' Children' s Hospital, Rome (Italy); and others

    2011-11-18

    Highlights: Black-Right-Pointing-Pointer Expanded array of mtDNA deletions. Black-Right-Pointing-Pointer Pearson syndrome with prominent hepatopathy associated with single mtDNA deletions. Black-Right-Pointing-Pointer Detection of deletions in fibroblasts and blood avoids muscle and liver biopsy. Black-Right-Pointing-Pointer Look for mtDNA deletions before to study nuclear genes related to mtDNA depletion. -- Abstract: Hepatic involvement in mitochondrial cytopathies rarely manifests in adulthood, but is a common feature in children. Multiple OXPHOS enzyme defects in children with liver involvement are often associated with dramatically reduced amounts of mtDNA. We investigated two novel large scale deletions in two infants with a multisystem disorder and prominent hepatopathy. Amount of mtDNA deletions and protein content were measured in different post-mortem tissues. The highest levels of deleted mtDNA were in liver, kidney, pancreas of both patients. Moreover, mtDNA deletions were detected in cultured skin fibroblasts in both patients and in blood of one during life. Biochemical analysis showed impairment of mainly complex I enzyme activity. Patients manifesting multisystem disorders in childhood may harbour rare mtDNA deletions in multiple tissues. For these patients, less invasive blood specimens or cultured fibroblasts can be used for molecular diagnosis. Our data further expand the array of deletions in the mitochondrial genomes in association with liver failure. Thus analysis of mtDNA should be considered in the diagnosis of childhood-onset hepatopathies.

  10. Entecavir for Treatment of Hepatitis B Virus Displays No In Vitro Mitochondrial Toxicity or DNA Polymerase Gamma Inhibition▿

    Science.gov (United States)

    Mazzucco, Charles E.; Hamatake, Robert K.; Colonno, Richard J.; Tenney, Daniel J.

    2008-01-01

    Therapy with nucleoside reverse transcriptase inhibitors (NRTIs) can be associated with mitochondrial toxicity. In vitro studies have been used to predict the predisposition for and characterize the mechanisms causing mitochondrial toxicity. Entecavir (ETV) is an approved deoxyguanosine nucleoside for the treatment of chronic hepatitis B virus (HBV) infection that exhibits potent activity against viral reverse transcriptase. We assessed the potential for mitochondrial toxicity of ETV in long-term cultures of HepG2 hepatoma cells by measuring mitochondrial function (through lactate secretion), levels of mitochondrial DNA (mtDNA), and levels of mitochondrial proteins COX II and COX IV. Furthermore, we tested the activity of ETV-triphosphate (ETV-TP) against mitochondrial DNA polymerase γ (Pol γ) in vitro. ETV concentrations as high as 100 times the maximal clinical exposure (Cmax) did not affect cell proliferation, levels of lactate, mitochondrial DNA, or mitochondrial proteins throughout the 15-day culture. The lack of mitochondrial toxicity was consistent with the finding that ETV-TP was not recognized by mitochondrial DNA Pol γ and failed to be incorporated into DNA or inhibit the polymerase assay at the highest levels tested, 300 μM. Combinations of ETV with each of the other HBV NRTI antivirals, adefovir, tenofovir, and lamivudine at 10 times their respective Cmax levels also failed to result in cellular or mitochondrial toxicity. In summary, cell culture and enzymatic studies yielded no evidence that would predict mitochondrial toxicity of ETV at exposure levels in excess of those expected to be achieved clinically. PMID:18056280

  11. Mitochondrial DNA haplogroups and type 2 diabetes: a study of 897 cases and 1010 controls.

    Science.gov (United States)

    Chinnery, P F; Mowbray, C; Patel, S K; Elson, J L; Sampson, M; Hitman, G A; McCarthy, M I; Hattersley, A T; Walker, M

    2007-06-01

    Mitochondria play a central role in the secretion of insulin by pancreatic beta-cells, and pathogenic mutations of mitochondrial DNA (mtDNA) can cause diabetes. The aetiology of type 2 diabetes has a strong genetic component, raising the possibility that genetic variants of mtDNA alter the risk of developing the disorder. Recent studies have produced conflicting results. By studying 897 UK cases of type 2 diabetes and 1010 population-matched controls, it is shown that European mtDNA haplogroups are unlikely to play a major role in the risk of developing the disorder.

  12. High levels of mitochondrial DNA deletions in substantia nigra neurons in aging and Parkinson disease.

    Science.gov (United States)

    Bender, Andreas; Krishnan, Kim J; Morris, Christopher M; Taylor, Geoffrey A; Reeve, Amy K; Perry, Robert H; Jaros, Evelyn; Hersheson, Joshua S; Betts, Joanne; Klopstock, Thomas; Taylor, Robert W; Turnbull, Douglass M

    2006-05-01

    Here we show that in substantia nigra neurons from both aged controls and individuals with Parkinson disease, there is a high level of deleted mitochondrial DNA (mtDNA) (controls, 43.3% +/- 9.3%; individuals with Parkinson disease, 52.3% +/- 9.3%). These mtDNA mutations are somatic, with different clonally expanded deletions in individual cells, and high levels of these mutations are associated with respiratory chain deficiency. Our studies suggest that somatic mtDNA deletions are important in the selective neuronal loss observed in brain aging and in Parkinson disease.

  13. Origin of Tetrapods Inferred from Their Mitochondrial DNA Affiliation to Lungfish

    OpenAIRE

    Meyer, Axel; Wilson, Allan C.

    1990-01-01

    This paper shows that questions of an unexpected phylogenetic depth can be addressed by the study of mitochondrial DNA (mtDNA) sequences. For decades, it has been unclear whether coelacanth fishes or lungfishes are the closest living relatives of land vertebrates (Tetrapoda). Segments of mtDNA from a lungfish, the coelacanth, and a ray-finned fish were sequenced and compared to the published sequence of a frog mtDNA. A tree based on inferred amino acid replacements, silent transversions, and ...

  14. Atypical presentation of Leigh syndrome associated with a Leber hereditary optic neuropathy primary mitochondrial DNA mutation.

    Science.gov (United States)

    Fruhman, Gary; Landsverk, Megan L; Lotze, Timothy E; Hunter, Jill V; Wangler, Michael F; Adesina, Adekunle M; Wong, Lee-Jun C; Scaglia, Fernando

    2011-06-01

    Leber hereditary optic neuropathy (LHON) is caused by point mutations in mitochondrial DNA (mtDNA), and is characterized by bilateral, painless sub-acute visual loss that develops during the second decade of life. Here we report the case of a five year old girl who presented with clinical and neuroradiological findings reminiscent of Leigh syndrome but carried a mtDNA mutation m.11778G>A (p.R340H) in the MTND4 gene usually observed in patients with LHON. This case is unusual for age of onset, gender, associated neurological findings and evolution, further expanding the clinical spectrum associated with primary LHON mtDNA mutations.

  15. Overexpression of DNA polymerase zeta reduces the mitochondrial mutability caused by pathological mutations in DNA polymerase gamma in yeast.

    Directory of Open Access Journals (Sweden)

    Enrico Baruffini

    Full Text Available In yeast, DNA polymerase zeta (Rev3 and Rev7 and Rev1, involved in the error-prone translesion synthesis during replication of nuclear DNA, localize also in mitochondria. We show that overexpression of Rev3 reduced the mtDNA extended mutability caused by a subclass of pathological mutations in Mip1, the yeast mitochondrial DNA polymerase orthologous to human Pol gamma. This beneficial effect was synergistic with the effect achieved by increasing the dNTPs pools. Since overexpression of Rev3 is detrimental for nuclear DNA mutability, we constructed a mutant Rev3 isoform unable to migrate into the nucleus: its overexpression reduced mtDNA mutability without increasing the nuclear one.

  16. Animal mitochondrial DNA polymorphism:a valuable tool for evolutionary studies

    Institute of Scientific and Technical Information of China (English)

    ZHANGYAPING; HONGLAN; 等

    1993-01-01

    Since the mitochondrial DNA(mtDNA) was discovered[1],a great amount of information have been accumulated about its sturcutres and functions[2],The complete nucleotide sequence of the mitochondrial genome has been determined for one individual in each of the following species:mouse(Mus musculus)[3],human(HOmo sapiens)[4],cow(Bos taurus)[5],Xenopus laevis[6],fruit fly(Drosophila yacuba)[7],sea urchin (Strongylocentrotus purpuratus)[8].rat(rattus novegicus)[9],fin whale(Balaenoptera physalus)[10],and harbor seal(Phoca vitulina)[11],More than 300 species of animals have been studied on mtDNA3.In the past 10 years.mtDNA has been a useful and powerful tool in the field of evolutionary biology and taxonomy.

  17. Complete mitochondrial DNA genome of tetraploid Carassius auratus gibelio.

    Science.gov (United States)

    Li, Zhong; Liang, Hong-Wei; Zou, Gui-Wei

    2016-01-01

    The complete mitochondrial genome was sequenced from the tetraploid Carassius auratus gibelio in this study. The genome sequence was 16,576 bp in length. The mitochondrial genome contains 13 protein-coding genes, 2 ribosomal RNAs, 22 transfer RNAs, and 2 non-coding regions (control region and origin of light-strand replication). All genes were encoded on the heavy strain except for ND6 and eight tRNA genes. The overall base composition is 31.61% A, 25.81% T, 26.62% G, 15.96% C, with an A+T bias of 57.42%. The complete mitogenome data provides useful genetic markers for the studies on the molecular identification, population genetics, phylogenetic analysis and conservation genetics.

  18. Complete mitochondrial DNA genome of Pseudobagrus brevicaudatus (Siluriformes: Bagridae).

    Science.gov (United States)

    Liang, Hong-Wei; Meng, Yan; Li, Zhong; Zhang, Yan; Zou, Gui-wei

    2014-06-01

    The complete mitochondrial genome of Pseudobagrus brevicaudatus (Siluriformes: Bagridae) was sequenced in this study. The total length of the mitogenome is 16,533 bp, with the base composition of 31.6% A, 26.8% T, 15.0%G, 26.6% C. The gene order and genes were the same as that found in other previously reported catfishes, including 13 protein-coding genes, 22 transfer RNA genes, 2 ribosomal RNA genes, and 1 non-coding control region. Except for ND6 gene and 8 tRNA genes, all other mitochondrial genes were encoded on the heavy strand. This complete mitogenome data provides the basis for taxonomic and conservation research of this and closely related species.

  19. Complete mitochondrial DNA genome of Zacco platypus (Cypriniformes: Cyprinidae).

    Science.gov (United States)

    Ueng, Yih-Tsong; Chen, Kun-Neng; Han, Chiao-Chuan; Cheng, Chung-Yao; Li, Yi-Min

    2015-04-01

    The complete mitochondrial genome of Zacco platypus (Cypriniformes, Cyprinidae), which has broader distribution range and diverse genetic structure than other species under the genus Zacco, was first determined in this study. The mitochondrial genome is 16,612 base pairs (bp) in length, encoding 13 protein-coding genes, 2 ribosomal RNAs, 22 transfer RNAs and 1 non-coding control region. Its gene arrangement and translation direction were identical to those of other typical vertebrate. Control region (D-Loop), of 929 bp lengths long, is located between tRNA(Pro) and tRNA(Phe). The overall base composition of the heavy strand shows T 27.02%, C 26.23%, A 28.94% and G 17.82%, with a slight AT bias of 55.95%.

  20. Complete mitochondrial DNA genome of Microphysogobio brevirostris (Cypriniformes: Cyprinidae).

    Science.gov (United States)

    Cheng, Chung-Yao; Wang, Jiang-Ping; Ho, Chuan-Wen; Cheng, Ju-Wen; Ueng, Yih-Tsong

    2015-04-01

    In this study, we sequenced the complete mitochondrial genome of Microphysogobio brevirostris (Cypriniformes, Cyprinidae), an endemic primary freshwater fish in Taiwan. This mitochondrial genome, consisting of 16,608 base pairs (bp), encoded 13 protein-coding genes, 2 ribosomal RNAs, 22 transfer RNAs, and a non-coding control region as those found in other vertebrates, with the gene synteny identical to that of typical vertebrates. Control region (D-Loop), of 929 bp lengths long, is located between tRNA(Pro) and tRNA(Phe). The overall base composition of the heavy strand shows T 26.28%, C 26.62%, A 30.26%, and G 16.85%, with a slight AT bias of 56.53%.

  1. Levels of plasma circulating cell free nuclear and mitochondrial DNA as potential biomarkers for breast tumors

    Directory of Open Access Journals (Sweden)

    Diesch Claude

    2009-11-01

    Full Text Available Abstract Background With the aim to simplify cancer management, cancer research lately dedicated itself more and more to discover and develop non-invasive biomarkers. In this connection, circulating cell-free DNA (ccf DNA seems to be a promising candidate. Altered levels of ccf nuclear DNA (nDNA and mitochondrial DNA (mtDNA have been found in several cancer types and might have a diagnostic value. Methods Using multiplex real-time PCR we investigated the levels of ccf nDNA and mtDNA in plasma samples from patients with malignant and benign breast tumors, and from healthy controls. To evaluate the applicability of plasma ccf nDNA and mtDNA as a biomarker for distinguishing between the three study-groups we performed ROC (Receiver Operating Characteristic curve analysis. We also compared the levels of both species in the cancer group with clinicopathological parameters. Results While the levels of ccf nDNA in the cancer group were significantly higher in comparison with the benign tumor group (P P P P = 0.022. The level of ccf nDNA was also associated with tumor-size (2 cmP = 0.034. Using ROC curve analysis, we were able to distinguish between the breast cancer cases and the healthy controls using ccf nDNA as marker (cut-off: 1866 GE/ml; sensitivity: 81%; specificity: 69%; P P Conclusion Our data suggests that nuclear and mitochondrial ccf DNA have potential as biomarkers in breast tumor management. However, ccf nDNA shows greater promise regarding sensitivity and specificity.

  2. Mitochondrial genome rearrangements in glomus species triggered by homologous recombination between distinct mtDNA haplotypes.

    Science.gov (United States)

    Beaudet, Denis; Terrat, Yves; Halary, Sébastien; de la Providencia, Ivan Enrique; Hijri, Mohamed

    2013-01-01

    Comparative mitochondrial genomics of arbuscular mycorrhizal fungi (AMF) provide new avenues to overcome long-lasting obstacles that have hampered studies aimed at understanding the community structure, diversity, and evolution of these multinucleated and genetically polymorphic organisms.AMF mitochondrial (mt) genomes are homogeneous within isolates, and their intergenic regions harbor numerous mobile elements that have rapidly diverged, including homing endonuclease genes, small inverted repeats, and plasmid-related DNA polymerase genes (dpo), making them suitable targets for the development of reliable strain-specific markers. However, these elements may also lead to genome rearrangements through homologous recombination, although this has never previously been reported in this group of obligate symbiotic fungi. To investigate whether such rearrangements are present and caused by mobile elements in AMF, the mitochondrial genomes from two Glomeraceae members (i.e., Glomus cerebriforme and Glomus sp.) with substantial mtDNA synteny divergence,were sequenced and compared with available glomeromycotan mitochondrial genomes. We used an extensive nucleotide/protein similarity network-based approach to investigated podiversity in AMF as well as in other organisms for which sequences are publicly available. We provide strong evidence of dpo-induced inter-haplotype recombination, leading to a reshuffled mitochondrial genome in Glomus sp. These findings raise questions as to whether AMF single spore cultivations artificially underestimate mtDNA genetic diversity.We assessed potential dpo dispersal mechanisms in AMF and inferred a robust phylogenetic relationship with plant mitochondrial plasmids. Along with other indirect evidence, our analyses indicate that members of the Glomeromycota phylum are potential donors of mitochondrial plasmids to plants.

  3. Paternal inheritance of mitochondrial DNA in the sheep (Ovine aries)

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    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.

  4. Analysis of mitochondrial DNA sequences in childhood encephalomyopathies reveals new disease-associated variants.

    Directory of Open Access Journals (Sweden)

    Aijaz A Wani

    Full Text Available BACKGROUND: Mitochondrial encephalomyopathies are a heterogeneous group of clinical disorders generally caused due to mutations in either mitochondrial DNA (mtDNA or nuclear genes encoding oxidative phosphorylation (OXPHOS. We analyzed the mtDNA sequences from a group of 23 pediatric patients with clinical and morphological features of mitochondrial encephalopathies and tried to establish a relationship of identified variants with the disease. METHODOLOGY/PRINCIPLE FINDINGS: Complete mitochondrial genomes were amplified by PCR and sequenced by automated DNA sequencing. Sequencing data was analyzed by SeqScape software and also confirmed by BLASTn program. Nucleotide sequences were compared with the revised Cambridge reference sequence (CRS and sequences present in mitochondrial databases. The data obtained shows that a number of known and novel mtDNA variants were associated with the disease. Most of the non-synonymous variants were heteroplasmic (A4136G, A9194G and T11916A suggesting their possibility of being pathogenic in nature. Some of the missense variants although homoplasmic were showing changes in highly conserved amino acids (T3394C, T3866C, and G9804A and were previously identified with diseased conditions. Similarly, two other variants found in tRNA genes (G5783A and C8309T could alter the secondary structure of Cys-tRNA and Lys-tRNA. Most of the variants occurred in single cases; however, a few occurred in more than one case (e.g. G5783A and A10149T. CONCLUSIONS AND SIGNIFICANCE: The mtDNA variants identified in this study could be the possible cause of mitochondrial encephalomyopathies with childhood onset in the patient group. Our study further strengthens the pathogenic score of known variants previously reported as provisionally pathogenic in mitochondrial diseases. The novel variants found in the present study can be potential candidates for further investigations to establish the relationship between their incidence and role

  5. Analysis of mitochondrial DNA in Tibetan gastric cancer patients at high altitude

    OpenAIRE

    Jiang, Jun; Zhao, Jun-Hui; Wang, Xue-Lian; DI, JI; Liu, Zhi-Bo; Li, Guo-Yuan; WANG, MIAO-ZHOU; Li, Yan; Chen, Rong; Ge, Ri-Li

    2015-01-01

    The highest risk areas of gastric cancer are currently Japan, Korea and China; Qinghai, a high-altitude area, has one of the highest gastric cancer rates in China. The incidence of gastric cancer is higher in the Tibetan ethnic group compared to that in the Han ethnic group in Qinghai. This study was conducted to determine the clinical characteristics of mitochondrial DNA (mtDNA) mutations and copy numbers among Tibetans with gastric cancer residing at high altitudes and investigate the assoc...

  6. Dynamical change of mitochondrial DNA induced in the living cell by perturbing the electrochemical gradient.

    OpenAIRE

    Coppey-Moisan, M; Brunet, A C; Morais, R.; Coppey, J

    1996-01-01

    Digital-imaging microscopy was used in conditions that allowed the native state to be preserved and hence fluorescence variations of specific probes to be followed in the real time of living mammalian cells. Ethidium bromide was shown to enter into living cells and to intercalate stably into mitochondrial DNA (mtDNA), giving rise to high fluorescence. When the membrane potential or the pH gradient across the inner membrane was abolished by specific inhibitors or ionophores, the ethidium fluor...

  7. Mitochondrial DNA genotypes in nuclear transfer-derived cloned sheep

    OpenAIRE

    Evans, Matthew J.; Gurer, Cagan; Loike, John D.; Wilmut, Ian; Schnieke, Angelika E.; Schon, Eric A.

    1999-01-01

    Eukaryotic cells contain two distinct genomes. One is located in the nucleus (nDNA) and is transmitted in a mendelian fashion, whereas the other is located in mitochondria (mtDNA) and is transmitted by maternal inheritance. Cloning of mammals1-6 typically has been achieved via nuclear transfer, in which a donor somatic cell is fused by electoporation with a recipient enucleated oocyte. During this whole-cell electrofusion, nDNA as well as mtDNA ought to be transferred to the oocyte7,8. Thus, ...

  8. Recurrent De Novo Dominant Mutations in SLC25A4 Cause Severe Early-Onset Mitochondrial Disease and Loss of Mitochondrial DNA Copy Number.

    Science.gov (United States)

    Thompson, Kyle; Majd, Homa; Dallabona, Christina; Reinson, Karit; King, Martin S; Alston, Charlotte L; He, Langping; Lodi, Tiziana; Jones, Simon A; Fattal-Valevski, Aviva; Fraenkel, Nitay D; Saada, Ann; Haham, Alon; Isohanni, Pirjo; Vara, Roshni; Barbosa, Inês A; Simpson, Michael A; Deshpande, Charu; Puusepp, Sanna; Bonnen, Penelope E; Rodenburg, Richard J; Suomalainen, Anu; Õunap, Katrin; Elpeleg, Orly; Ferrero, Ileana; McFarland, Robert; Kunji, Edmund R S; Taylor, Robert W

    2016-10-06

    Mutations in SLC25A4 encoding the mitochondrial ADP/ATP carrier AAC1 are well-recognized causes of mitochondrial disease. Several heterozygous SLC25A4 mutations cause adult-onset autosomal-dominant progressive external ophthalmoplegia associated with multiple mitochondrial DNA deletions, whereas recessive SLC25A4 mutations cause childhood-onset mitochondrial myopathy and cardiomyopathy. Here, we describe the identification by whole-exome sequencing of seven probands harboring dominant, de novo SLC25A4 mutations. All affected individuals presented at birth, were ventilator dependent and, where tested, revealed severe combined mitochondrial respiratory chain deficiencies associated with a marked loss of mitochondrial DNA copy number in skeletal muscle. Strikingly, an identical c.239G>A (p.Arg80His) mutation was present in four of the seven subjects, and the other three case subjects harbored the same c.703C>G (p.Arg235Gly) mutation. Analysis of skeletal muscle revealed a marked decrease of AAC1 protein levels and loss of respiratory chain complexes containing mitochondrial DNA-encoded subunits. We show that both recombinant AAC1 mutant proteins are severely impaired in ADP/ATP transport, affecting most likely the substrate binding and mechanics of the carrier, respectively. This highly reduced capacity for transport probably affects mitochondrial DNA maintenance and in turn respiration, causing a severe energy crisis. The confirmation of the pathogenicity of these de novo SLC25A4 mutations highlights a third distinct clinical phenotype associated with mutation of this gene and demonstrates that early-onset mitochondrial disease can be caused by recurrent de novo mutations, which has significant implications for the application and analysis of whole-exome sequencing data in mitochondrial disease. Copyright © 2016 The Author(s). Published by Elsevier Inc. All rights reserved.

  9. Rapid Mitochondrial DNA Segregation in Primate Preimplantation Embryos Precedes Somatic and Germline Bottleneck

    Directory of Open Access Journals (Sweden)

    Hyo-Sang Lee

    2012-05-01

    Full Text Available The 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.

  10. Rapid mitochondrial DNA segregation in primate preimplantation embryos precedes somatic and germline bottleneck.

    Science.gov (United States)

    Lee, Hyo-Sang; Ma, Hong; Juanes, Rita Cervera; Tachibana, Masahito; Sparman, Michelle; Woodward, Joy; Ramsey, Cathy; Xu, Jing; Kang, Eun-Ju; Amato, Paula; Mair, Georg; Steinborn, Ralf; Mitalipov, Shoukhrat

    2012-05-31

    The 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.

  11. High copy number of mitochondrial DNA predicts poor prognosis in patients with advanced stage colon cancer.

    Science.gov (United States)

    Wang, Yun; He, Shuixiang; Zhu, Xingmei; Qiao, Wei; Zhang, Juan

    2016-12-23

    The aim of this investigation was to determine whether alterations in mitochondrial DNA (mtDNA) copy number in colon cancer were associated with clinicopathological parameters and postsurgical outcome. By quantitative real-time PCR assay, the mtDNA copy number was detected in a cohort of colon cancer and matched adjacent colon tissues (n = 162). The majority of patients had higher mtDNA content in colon cancer tissues than matched adjacent colon tissues. Moreover, high mtDNA content in tumor tissues was associated with larger tumor size, higher serum CEA level, advanced TNM stage, vascular emboli, and liver metastases. Further survival curve analysis showed that high mtDNA content was related to the worst survival in patients with colon cancer at advanced TNM stage. High mtDNA content is a potential effective factor of poor prognosis in patients with advanced stage colon cancer.

  12. Corresponding mitochondrial DNA and niche divergence for crested newt candidate species.

    Directory of Open Access Journals (Sweden)

    Ben Wielstra

    Full Text Available Genetic divergence of mitochondrial DNA does not necessarily correspond to reproductive isolation. However, if mitochondrial DNA lineages occupy separate segments of environmental space, this supports the notion of their evolutionary independence. We explore niche differentiation among three candidate species of crested newt (characterized by distinct mitochondrial DNA lineages and interpret the results in the light of differences observed for recognized crested newt species. We quantify niche differences among all crested newt (candidate species and test hypotheses regarding niche evolution, employing two ordination techniques (PCA-env and ENFA. Niche equivalency is rejected: all (candidate species are found to occupy significantly different segments of environmental space. Furthermore, niche overlap values for the three candidate species are not significantly higher than those for the recognized species. As the three candidate crested newt species are, not only in terms of mitochondrial DNA genetic divergence, but also ecologically speaking, as diverged as the recognized crested newt species, our findings are in line with the hypothesis that they represent cryptic species. We address potential pitfalls of our methodology.

  13. Meta-Analysis of mitochondrial DNA reveals several population bottlenecks during worldwide migrations of cattle

    NARCIS (Netherlands)

    Lenstra, Johannes A.; Ajmone-Marsan, Paolo; Beja-Pereira, Albano; Bollongino, Ruth; Bradley, Daniel G.; Colli, Licia; De Gaetano, Anna; Edwards, Ceiridwen J.; Felius, Marleen; Ferretti, Luca; Ginja, Catarina; Hristov, Peter; Kantanen, Juha; Lirón, Juan Pedro; Magee, David A.; Negrini, Riccardo; Radoslavov, Georgi A.

    2014-01-01

    Several studies have investigated the differentiation of mitochondrial DNA in Eurasian, African and American cattle as well as archaeological bovine material. A global survey of these studies shows that haplogroup distributions are more stable in time than in space. All major migrations of cattle ha

  14. Genetic characterization of Phytophthora nicotianae by the analysis of polymorphic regions of the mitochondrial DNA.

    Science.gov (United States)

    A new method based on the analysis of mitochondrial intergenic regions characterized by intraspecific variation in DNA sequences was developed and applied to the study of the plant pathogen Phytophthora nicotianae. Two regions flanked by genes trny and rns and trnw and cox2 were identified by compa...

  15. Mitochondrial DNA variation of the common hippopotamus: evidence for a recent population expansion

    DEFF Research Database (Denmark)

    Okello, John Bosco A.; Nyakaana, Silvester; Masembe, C.

    2005-01-01

    Mitochondrial DNA control region sequence variation was obtained and the population history of the common hippopotamus was inferred from 109 individuals from 13 localities covering six populations in sub-Saharan Africa. In all, 100 haplotypes were defined, of which 98 were locality specific...

  16. Systematics of the Dioryctria abietella Species Group (Lepidoptera: Pyralidae) Based on Mitochondrial DNA Ann

    Science.gov (United States)

    G. Roux-Morabito; N.E. Gillette; A. Roques; L. Dormont; J. Stein; F.A.H. Sperling

    2008-01-01

    Coneworms of the genus Dioryctria Zeller include several serious pests of conifer seeds that are notoriously difficult to distinguish as species. We surveyed mitochondrial DNA variation within the abietella species group by sequencing 451 bp of cytochrome oxidase subunit 1 (COI) and 572 bp of cytochrome oxidase subunit 2 (COII...

  17. Assessing Clark's nutcracker seed-caching flights using maternally inherited mitochondrial DNA of whitebark pine

    Science.gov (United States)

    Bryce A. Richardson; Ned B. Klopfenstein; Steven J. Brunsfeld

    2002-01-01

    Maternally inherited mitochondrial DNA haplotypes in whitebark pine (Pinus albicaulis Engelm.) were used to examine the maternal genetic structure at three hierarchical spatial scales: fine scale, coarse scale, and interpopulation. These data were used to draw inferences into Clark’s nutcracker (Nucifraga columbiana Wilson)...

  18. Direct estimation of the mitochondrial DNA mutation rate in Drosophila melanogaster.

    Directory of Open Access Journals (Sweden)

    Cathy Haag-Liautard

    2008-08-01

    Full Text Available Mitochondrial DNA (mtDNA variants are widely used in evolutionary genetics as markers for population history and to estimate divergence times among taxa. Inferences of species history are generally based on phylogenetic comparisons, which assume that molecular evolution is clock-like. Between-species comparisons have also been used to estimate the mutation rate, using sites that are thought to evolve neutrally. We directly estimated the mtDNA mutation rate by scanning the mitochondrial genome of Drosophila melanogaster lines that had undergone approximately 200 generations of spontaneous mutation accumulation (MA. We detected a total of 28 point mutations and eight insertion-deletion (indel mutations, yielding an estimate for the single-nucleotide mutation rate of 6.2 x 10(-8 per site per fly generation. Most mutations were heteroplasmic within a line, and their frequency distribution suggests that the effective number of mitochondrial genomes transmitted per female per generation is about 30. We observed repeated occurrences of some indel mutations, suggesting that indel mutational hotspots are common. Among the point mutations, there is a large excess of G-->A mutations on the major strand (the sense strand for the majority of mitochondrial genes. These mutations tend to occur at nonsynonymous sites of protein-coding genes, and they are expected to be deleterious, so do not become fixed between species. The overall mtDNA mutation rate per base pair per fly generation in Drosophila is estimated to be about 10x higher than the nuclear mutation rate, but the mitochondrial major strand G-->A mutation rate is about 70x higher than the nuclear rate. Silent sites are substantially more strongly biased towards A and T than nonsynonymous sites, consistent with the extreme mutation bias towards A+T. Strand-asymmetric mutation bias, coupled with selection to maintain specific nonsynonymous bases, therefore provides an explanation for the extreme base

  19. Polymorphisms in the mitochondrial DNA control region and frailty in older adults.

    Directory of Open Access Journals (Sweden)

    Ann Z Moore

    Full Text Available BACKGROUND: Mitochondria contribute to the dynamics of cellular metabolism, the production of reactive oxygen species, and apoptotic pathways. Consequently, mitochondrial function has been hypothesized to influence functional decline and vulnerability to disease in later life. Mitochondrial genetic variation may contribute to altered susceptibility to the frailty syndrome in older adults. METHODOLOGY/PRINCIPAL FINDINGS: To assess potential mitochondrial genetic contributions to the likelihood of frailty, mitochondrial DNA (mtDNA variation was compared in frail and non-frail older adults. Associations of selected SNPs with a muscle strength phenotype were also explored. Participants were selected from the Cardiovascular Health Study (CHS, a population-based observational study (1989-1990, 1992-1993. At baseline, frailty was identified as the presence of three or more of five indicators (weakness, slowness, shrinking, low physical activity, and exhaustion. mtDNA variation was assessed in a pilot study, including 315 individuals selected as extremes of the frailty phenotype, using an oligonucleotide sequencing microarray based on the Revised Cambridge Reference Sequence. Three mtDNA SNPs were statistically significantly associated with frailty across all pilot participants or in sex-stratified comparisons: mt146, mt204, and mt228. In addition to pilot participants, 4,459 additional men and women with frailty classifications, and an overlapping subset of 4,453 individuals with grip strength measurements, were included in the study population genotyped at mt204 and mt228. In the study population, the mt204 C allele was associated with greater likelihood of frailty (adjusted odds ratio = 2.04, 95% CI = 1.07-3.60, p = 0.020 and lower grip strength (adjusted coefficient = -2.04, 95% CI = -3.33- -0.74, p = 0.002. CONCLUSIONS: This study supports a role for mitochondrial genetic variation in the frailty syndrome and later life muscle strength

  20. Persistence and protection of mitochondrial DNA in the generative cell of cucumber is consistent with its paternal transmission

    Science.gov (United States)

    Cucumber, unlike most plants, shows paternal inheritance of its mitochondrial DNA (mtDNA); however, the mechanisms regulating this unique transmission mode are unclear. Here we monitored the amounts of mtDNA through the development of cucumber microspores to pollen and observed that mtDNA decreases ...

  1. Decoupled mitochondrial and chloroplast DNA population structure reveals Holocene collapse and population isolation in a threatened Mexican-endemic conifer.

    Science.gov (United States)

    Juan P. Jaramillo-Correa; Jean Beaulieu; F. Thomas Ledig; Jean. Bousqueter

    2006-01-01

    Chihuahua spruce (Picea chihuahuana Martínez) is a montane subtropical conifer endemic to the Sierra Madre Occidental in northwestern México. Range-wide variation was investigated using maternally inherited mitochondrial (mtDNA) and paternally inherited chloroplast (cpDNA) DNA markers. Among the 16 mtDNA regions analysed, only...

  2. Import of desired nucleic acid sequences using addressing motif of mitochondrial ribosomal 5S-rRNA for fluorescent in vivo hybridization of mitochondrial DNA and RNA.

    Science.gov (United States)

    Zelenka, Jaroslav; Alán, Lukáš; Jabůrek, Martin; Ježek, Petr

    2014-04-01

    Based on the matrix-addressing sequence of mitochondrial ribosomal 5S-rRNA (termed MAM), which is naturally imported into mitochondria, we have constructed an import system for in vivo targeting of mitochondrial DNA (mtDNA) or mt-mRNA, in order to provide fluorescence hybridization of the desired sequences. Thus DNA oligonucleotides were constructed, containing the 5'-flanked T7 RNA polymerase promoter. After in vitro transcription and fluorescent labeling with Alexa Fluor(®) 488 or 647 dye, we obtained the fluorescent "L-ND5 probe" containing MAM and exemplar cargo, i.e., annealing sequence to a short portion of ND5 mRNA and to the light-strand mtDNA complementary to the heavy strand nd5 mt gene (5'-end 21 base pair sequence). For mitochondrial in vivo fluorescent hybridization, HepG2 cells were treated with dequalinium micelles, containing the fluorescent probes, bringing the probes proximally to the mitochondrial outer membrane and to the natural import system. A verification of import into the mitochondrial matrix of cultured HepG2 cells was provided by confocal microscopy colocalizations. Transfections using lipofectamine or probes without 5S-rRNA addressing MAM sequence or with MAM only were ineffective. Alternatively, the same DNA oligonucleotides with 5'-CACC overhang (substituting T7 promoter) were transcribed from the tetracycline-inducible pENTRH1/TO vector in human embryonic kidney T-REx®-293 cells, while mitochondrial matrix localization after import of the resulting unlabeled RNA was detected by PCR. The MAM-containing probe was then enriched by three-order of magnitude over the natural ND5 mRNA in the mitochondrial matrix. In conclusion, we present a proof-of-principle for mitochondrial in vivo hybridization and mitochondrial nucleic acid import.

  3. Mitochondrial DNA sequence and phylogenetic evaluation of geographically disparate Sus scrofa breeds.

    Science.gov (United States)

    Cannon, M V; Brandebourg, T D; Kohn, M C; Ðikić, D; Irwin, M H; Pinkert, C A

    2015-01-01

    Next generation sequencing of mitochondrial DNA (mtDNA) facilitates studies into the metabolic characteristics of production animals and their relation to production traits. Sequence analysis of mtDNA from pure-bred swine with highly disparate production characteristics (Mangalica Blonde, Mangalica Swallow-bellied, Meishan, Turopolje, and Yorkshire) was initiated to evaluate the influence of mtDNA polymorphisms on mitochondrial function. Herein, we report the complete mtDNA sequences of five Sus scrofa breeds and evaluate their position within the phylogeny of domestic swine. Phenotypic traits of Yorkshire, Mangalica Blonde, and Swallow-belly swine are presented to demonstrate their metabolic characteristics. Our data support the division of European and Asian breeds noted previously and confirm European ancestry of Mangalica and Turopolje breeds. Furthermore, mtDNA differences between breeds suggest function-altering changes in proteins involved in oxidative phosphorylation such as ATP synthase 6 (MT-ATP6), cytochrome oxidase I (MT-CO1), cytochrome oxidase III (MT-CO3), and cytochrome b (MT-CYB), supporting the hypothesis that mtDNA polymorphisms contribute to differences in metabolic traits between swine breeds. Our sequence data form the basis for future research into the roles of mtDNA in determining production traits in domestic animals. Additionally, such studies should provide insight into how mtDNA haplotype influences the extreme adiposity observed in Mangalica breeds.

  4. Exposure to 1800 MHz radiofrequency radiation induces oxidative damage to mitochondrial DNA in primary cultured neurons.

    Science.gov (United States)

    Xu, Shangcheng; Zhou, Zhou; Zhang, Lei; Yu, Zhengping; Zhang, Wei; Wang, Yuan; Wang, Xubu; Li, Maoquan; Chen, Yang; Chen, Chunhai; He, Mindi; Zhang, Guangbin; Zhong, Min

    2010-01-22

    Increasing evidence indicates that oxidative stress may be involved in the adverse effects of radiofrequency (RF) radiation on the brain. Because mitochondrial DNA (mtDNA) defects are closely associated with various nervous system diseases and mtDNA is particularly susceptible to oxidative stress, the purpose of this study was to determine whether radiofrequency radiation can cause oxidative damage to mtDNA. In this study, we exposed primary cultured cortical neurons to pulsed RF electromagnetic fields at a frequency of 1800 MHz modulated by 217 Hz at an average special absorption rate (SAR) of 2 W/kg. At 24 h after exposure, we found that RF radiation induced a significant increase in the levels of 8-hydroxyguanine (8-OHdG), a common biomarker of DNA oxidative damage, in the mitochondria of neurons. Concomitant with this finding, the copy number of mtDNA and the levels of mitochondrial RNA (mtRNA) transcripts showed an obvious reduction after RF exposure. Each of these mtDNA disturbances could be reversed by pretreatment with melatonin, which is known to be an efficient antioxidant in the brain. Together, these results suggested that 1800 MHz RF radiation could cause oxidative damage to mtDNA in primary cultured neurons. Oxidative damage to mtDNA may account for the neurotoxicity of RF radiation in the brain.

  5. Mitochondrial DNA evolution in the Anaxyrus boreas species group

    Science.gov (United States)

    Anna M. Goebel; Tom A. Ranker; Paul Stephen Corn; Richard G. Olmstead

    2009-01-01

    The Anaxyrus boreas species group currently comprises four species in western North America including the broadly distributed A. boreas, and three localized species, Anaxyrus nelsoni, Anaxyrus exsul and Anaxyrus canorus. Phylogenetic analyses of the mtDNA 12S rDNA, cytochrome...

  6. Rapid screening mitochondrial DNA mutation by using denaturing high-performance liquid chromatography

    Institute of Scientific and Technical Information of China (English)

    Man-Ran Liu; Kai-Feng Pan; Zhen-Fu Li; Yi Wang; Da-Jun Deng; Lian Zhang; You-Yong Lu

    2002-01-01

    AIM: To optimize conditions of DHPLC and analyze theeffectiveness of various DNA polymerases on DHPLCresolution, and evaluate the sensitivity of DHPLC in themutation screening of mitochondrial DNA (mtDNA).METHODS: Two fragments of 16s gene of mitochondrial DNA(one of them F2 is a mutant fragment) and an A3243Gmutated fragment were used to analyze the UV detectionlimit and determine the minimum percentage of mutant PCRproducts for DHPLC and evaluate effects of DNApolymerases on resolution of DHPLC. Under the optimalconditions, we analyzed the mtDNA mutations from muscletissues of mitochondrial encephalomyopathy with lacticacidosis and stroke-like episodes (NELAS) and screenedblindly for variances in D-loop region of mtDNA from humangastric tumor specimen.RESULTS: Ten A3243G variants were detected in 12 cases ofMELAS, no alterations were detected in controls and theseresults were consistent with the results obtained by analysisof RFLP with Apel. We also identified 26 D-loop variances in46 cases of human gastric cancer tissues and 38 alterationsin 13 gastric cancer cell lines. The mutation of mtDNA at 80 ngPCFI products containing a minimum of 5 % mutant sequencescould be detected by using DHPLC with UV detector.Moreover, Ampli-Taq Gold polymerase was equally as good asthe proofreading DNA polymerase (e. g., Pfu) in eliminatingthe false positive produced by Taq DNA polymerases.CONCLUSION: DHPLC is a powerful, rapid and sensitivemutation screening method for mtDNA. Proofreading DNApolymerase is more suitable for DHPLC analysis than Taqpolymerase.

  7. Mitochondrial DNA copy number in peripheral blood cells declines with age and is associated with general health among elderly

    DEFF Research Database (Denmark)

    Mengel-From, Jonas; Thinggaard, Mikael; Dalgård, Christine

    2014-01-01

    compared to nuclear DNA, i.e. the mitochondrial DNA copy number, was measured by PCR technology and used as a proxy for the content of mitochondria copies. In 1,067 Danish twins and singletons (18-93 years of age), with the majority being elderly individuals, the estimated mean mitochondrial DNA copy...... an individual was more than twice as steep as observed in the cross-sectional analysis [decline of mtDNA content: -1.27; 95 % CI (-1.71; -0.82)]. Subjects with low mitochondrial DNA copy number had poorer outcomes in terms of cognitive performance, physical strength, self-rated health, and higher all......-cause mortality than subjects with high mitochondrial DNA copy number, also when age was controlled for. The copy number mortality association can contribute to the smaller decline in a cross-sectional sample of the population compared to the individual, longitudinal decline. This study suggests that high...

  8. Mitochondrial DNA transcription levels during spermatogenesis and early development in doubly uniparental inheritance of the mitochondrial DNA system of the blue mussel Mytilus galloprovincialis.

    Science.gov (United States)

    Sano, Natsumi; Obata, Mayu; Komaru, Akira

    2013-08-01

    In some species of bivalve, there are two highly diverged mitochondrial genomes, one found in all individuals (F type) and the other normally in males only (M type). In Mytilus, a maternally-dependent sex ratio of the progeny has been reported. Some females almost exclusively produce daughters, while others produce a high proportion of sons. We previously reported that in M. galloprovincialis, M type mtDNA copy number may be maintained during spermatogenesis and the development of larvae of male-biased mothers to sustain the doubly uniparental inheritance system. In this study, we investigated transcription levels of M type mtDNA before and after fertilization to understand its function in the germ line. First, we quantified transcription levels of M type mtDNA in testicular cells dissected using laser-capture micro-dissection. The transcription levels of M type mtDNA were not significantly different between spermatogonia and spermatocytes versus spermatids and spermatozoa. Next, we examined differences in transcription levels of M type mtDNA between larvae from male-biased and female-biased mothers. The transcription levels of M type mtDNA significantly increased 24 and 48 h after fertilization in male-biased crosses. By contrast, transcription levels significantly decreased in female-biased crosses. These results suggest M type mtDNA may play a role in early germ line formation.

  9. Mitochondrial DNA sequence variation in Finnish patients with matrilineal diabetes mellitus

    Directory of Open Access Journals (Sweden)

    Soini Heidi K

    2012-07-01

    Full Text Available Abstract Background The genetic background of type 2 diabetes is complex involving contribution by both nuclear and mitochondrial genes. There is an excess of maternal inheritance in patients with type 2 diabetes and, furthermore, diabetes is a common symptom in patients with mutations in mitochondrial DNA (mtDNA. Polymorphisms in mtDNA have been reported to act as risk factors in several complex diseases. Findings We examined the nucleotide variation in complete mtDNA sequences of 64 Finnish patients with matrilineal diabetes. We used conformation sensitive gel electrophoresis and sequencing to detect sequence variation. We analysed the pathogenic potential of nonsynonymous variants detected in the sequences and examined the role of the m.16189 T>C variant. Controls consisted of non-diabetic subjects ascertained in the same population. The frequency of mtDNA haplogroup V was 3-fold higher in patients with diabetes. Patients harboured many nonsynonymous mtDNA substitutions that were predicted to be possibly or probably damaging. Furthermore, a novel m.13762 T>G in MTND5 leading to p.Ser476Ala and several rare mtDNA variants were found. Haplogroup H1b harbouring m.16189 T > C and m.3010 G > A was found to be more frequent in patients with diabetes than in controls. Conclusions Mildly deleterious nonsynonymous mtDNA variants and rare population-specific haplotypes constitute genetic risk factors for maternally inherited diabetes.

  10. Introgression of mitochondrial DNA promoted by natural selection in the Japanese pipistrelle bat (Pipistrellus abramus).

    Science.gov (United States)

    Dong, Ji; Mao, Xiuguang; Sun, Haijian; Irwin, David M; Zhang, Shuyi; Hua, Panyu

    2014-12-01

    Introgression of mitochondrial DNA (mtDNA) between closely related taxa can be promoted by either neutral processes or natural selection. Since mitochondrial gene-encoded proteins play critical roles in oxidative metabolism, mtDNA genes are commonly considered to experience strong selective constraint. However, metabolic requirements vary across climatic and ecological gradients, thus modifying potential selective pressures acting on mtDNA genes. Here we conducted tests to detect adaptive evolution occurring in two mtDNA genes (Cytb and ND5) in individuals of Japanese pipistrelle bat (Pipistrellus abramus) across the mainland of China and Hainan Island. Nuclear DNA markers identified two clades in both the mainland and Hainan Island populations, whereas each of these regions had a specific mtDNA clade. This cyto-nuclear discordance is most likely caused by introgression of the mtDNA by ruling out two other alternative scenarios (incomplete lineage sorting and sex-biased gene flow). Although population-based analyses revealed purifying selection acting on Cytb and neutrality in ND5, multiple nonsynonymous substitutions in both Cytb and ND5 were suggested to have been caused by positive selection by a divergence-based analysis. Our study supports the view that molecular adaptation can occur at genes under strong purifying selection if nonsynonymous substitutions cause radical changes in the physicochemical properties of amino acids.

  11. Circulating cell-free mitochondrial DNA as a novel cancer biomarker: opportunities and challenges.

    Science.gov (United States)

    Yu, Man

    2012-10-01

    The unique characteristics of the mitochondrial genome, such as short length, simple molecular structure, and high copy number, have made monitoring aberrant changes of mitochondrial DNA (mtDNA) quantity an interesting molecular tool for early tumor detection with many advantages over the nuclear genome-based methods. Recently, circulating cell-free (ccf) mtDNA in blood has emerged on the platform as a non-invasive diagnostic and prognostic biomarker for many forms of solid tumors. Accumulating evidence demonstrate that plasma or serum ccf mtDNA levels are significantly different between cancer patients and healthy individuals. Furthermore, quantification of ccf mtDNA levels in circulation may assist in identifying patients from cancer-free healthy population. This minireview attempts to summarize our recent findings in this very promising field of cancer research. The potential technical challenges that we have encountered during the quantitative analysis of ccf mtDNA and mtDNA in general are also briefly discussed. Prospective studies with a larger cohort of patients in various cancer entities are beneficial to precisely define the clinical importance of assessing the ccf mtDNA amount for diagnosing and tracking malignant diseases and their progression.

  12. A Study on the D-loop Region of Mitochondrial DNA (mtDNA) Mutation in Cervical Carcinomas

    Institute of Scientific and Technical Information of China (English)

    XUE Wen-qun; CHEN Dao-zhen

    2009-01-01

    Objective Background-study on genesis and development of tumor is mainly concentrated on gene mutation in nucleus. In recent years, however, the role of mitochondrial DNA (mtDNA) mutation in tumor genesis has been given more and more attention, which is the only extra-nucleus DNA in cells of higher animals. Carcinoma of the uterine cervix is a common tumor in gynecology, but there are few reports of mtDNA mutation in this area. The focus of this study was to investigate the mtDNA mutation in tumor tissues of cervical carcinomas and their relationship to tumorigenesis and tumor development. Methods The D-loop region of 24 cervical carcinomas together with the adjacent normal tissues were amplified by PCR and sequenced. Results Among the 24 cervical carcinomas, 30 mutations in 9 patients′ specimen were identified with the mutations rate of 37.5%(9/24). There were 8 microsatellite instabilities among the mutations and 13 new polymorphisms which were not reported previously in the Genbank. Conclusions The D-loop region of mitochondrial DNA is a highly polymorphoric and mutable region and the mutation rate is relatively high in patients with cervical carcinomas.

  13. Mitochondrial biogenesis drives a vicious cycle of metabolic insufficiency and mitochondrial DNA deletion mutation accumulation in aged rat skeletal muscle fibers.

    Directory of Open Access Journals (Sweden)

    Allen Herbst

    Full Text Available Aged muscles possess dysfunctional fibers that contain intracellular expansions of somatically derived mitochondrial DNA deletion mutations. At high abundance, these mutations disrupt the expression of mitochondrially-encoded protein subunits of the electron transport chain resulting in aerobic respiration deficient muscle fiber segments. These fiber segments atrophy and break contributing to the loss of muscle mass and function that occurs with age. By combining micro-dissection of individual muscle fibers with microarray analysis, we observed the response induced within these abnormal muscle fibers and detected an increase in many genes affecting metabolism and metabolic regulation. The transcriptional profile and subsequent protein validation suggested that a non-compensatory program of mitochondrial biogenesis was initiated. We hypothesized that this non-adaptive program of mitochondrial biogenesis was driving mtDNA deletion mutation accumulation. We tested this hypothesis by treating aged rats with β-Guanidinopropionic acid, a compound that stimulates mitochondrial biogenesis. β-Guanidinopropionic acid treatment increased muscle mitochondrial genome copy number and resulted in a 3.7 fold increase in the abundance of electron transport chain negative muscle fiber segments. We conclude that in electron transport system abnormal muscle fiber segments, a vicious cycle of metabolic insufficiency and non-compensatory mitochondrial biogenesis drive mtDNA deletion mutation accumulation.

  14. Analysis of Mitochondrial DNA from the Ancient Tombs of Turfan

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    MtDNA was successfully extracted from ten individual bones (femurs) in the tombs of ancient Jushi in Turfan basin, dated back to the year about 3 000-2 500 years ago. By means of four overlapping primers, we got nucleotide sequence of the 218bp length. Ancient mtDNA was analyzed by the sequencing of hypervariable region Ⅰ of the mtDNA control region. The result shows that 9 haplotypes with 24 polymorphic sites were obtained. The phylogenetic analysis indicated that Mongolians and Altai are the population genetically closest to the Jushi groups and Jushi mtDNA pool being an admixture of eastern Asian and European lineages. So our preliminary data imply that an ancient mingling of Euro-Asian population had existed in Turfan basin prior to the early Iron Age.

  15. CARDIOSELECTIVE OXIDATION OF MITOCHONDRIAL DNA FOLLOWING SUBCHRONIC ADMINISTRATION OF DOXORUBICIN

    Science.gov (United States)

    This preferential oxidation of cardiac mtDNA is consistent with the bioenergetic failure and the cumulative and irreversible cardiomyopathy that limits the clinical utility of this important antineoplastic drug.

  16. Nondestructive sampling of human skeletal remains yields ancient nuclear and mitochondrial DNA.

    Science.gov (United States)

    Bolnick, Deborah A; Bonine, Holly M; Mata-Míguez, Jaime; Kemp, Brian M; Snow, Meradeth H; LeBlanc, Steven A

    2012-02-01

    Museum curators and living communities are sometimes reluctant to permit ancient DNA (aDNA) studies of human skeletal remains because the extraction of aDNA usually requires the destruction of at least some skeletal material. Whether these views stem from a desire to conserve precious materials or an objection to destroying ancestral remains, they limit the potential of aDNA research. To help address concerns about destructive analysis and to minimize damage to valuable specimens, we describe a nondestructive method for extracting DNA from ancient human remains. This method can be used with both teeth and bone, but it preserves the structural integrity of teeth much more effectively than that of bone. Using this method, we demonstrate that it is possible to extract both mitochondrial and nuclear DNA from human remains dating between 300 BC and 1600 AD. Importantly, the method does not expose the remains to hazardous chemicals, allowing them to be safely returned to curators, custodians, and/or owners of the samples. We successfully amplified mitochondrial DNA from 90% of the individuals tested, and we were able to analyze 1-9 nuclear loci in 70% of individuals. We also show that repeated nondestructive extractions from the same tooth can yield amplifiable mitochondrial and nuclear DNA. The high success rate of this method and its ability to yield DNA from samples spanning a wide geographic and temporal range without destroying the structural integrity of the sampled material may make possible the genetic study of skeletal collections that are not available for destructive analysis. Copyright © 2011 Wiley Periodicals, Inc.

  17. Complete mitochondrial genome of wild aurochs (Bos primigenius) reconstructed from ancient DNA.

    Science.gov (United States)

    Zeyland, J; Wolko, L; Bocianowski, J; Szalata, M; Słomski, R; Dzieduszycki, A M; Ryba, M; Przystałowska, H; Lipiński, D

    2013-01-01

    Extinct aurochs (Bos primigenius), accepted as the ancestor of domestic cattle, was one of the largest wild animals inhabiting Europe, Asia and North Africa. The gradual process of aurochs extinction finished in Poland in 1627, were the last recorded aurochs, a female, died. Some aspects of cattle domestication history and the distribution of aurochs genetic material among modern cattle breeds still remain unclear. Analyses of ancient DNA (aDNA) from bone sample deliver new genetic information about extinct wild aurochs as well as modern cattle phylogeny. DNA was extracted from a fragment of aurochs fossil bone found in the Pisz Forest, Poland. The sample was radiocarbon-dated to about 1500 yBP. The aDNA was used for Whole Genome Amplification in order to form a DNA bank. Auroch mitochondrial DNA sequences were amplified using sets of 41 primers overlapping the whole mtDNA, cloned and sequenced. The sequence of the whole mitochondrial genome was reconstructed and deposed in GenBank [GenBank:JQ437479]. Based on the phylogenetic analyses of the Bovine mitochondrial genomes, a phylogenetic tree was created. As expected, the tree clearly shows that the mtDNA sequence of the analyzed PWA (Polish Wild Aurochs) individual belongs to haplogroup P. In the course of the comparative mtDNA analysis we identified 30 nucleotide marker positions for haplogroup P and nine unique PWA differences compared to the two remaining haplotype P representatives. Our analysis provides the next step to the reconstruction of the demographic history of this extinct but still exciting species.

  18. Mitochondrial DNA Haplogroups influence lipoatrophy after Highly Active Anti-retroviral Therapy

    Science.gov (United States)

    Hendrickson, Sher L.; Kingsley, Lawrence A.; Ruiz-Pesini, Eduardo; Poole, Jason C.; Jacobson, Lisa P.; Palella, Frank J.; Bream, Jay H.; Wallace, Douglas C.; O’Brien, Stephen J.

    2009-01-01

    Although highly active retroviral therapy (HAART) has been extremely effective in lowering AIDS incidence among patients infected with HIV, certain drugs included in HAART can cause serious mitochondrial toxicities. One of the most frequent adverse events is lipoatrophy, which is the loss of subcutaneous fat in the face, arms, buttocks and/or legs as an adverse reaction to nucleoside reverse transcriptase inhibitors (NRTIs). The clinical symptoms of lipoatrophy resemble those of inherited mitochondrial diseases, which suggests that host mitochondrial genotype may play a role in susceptibility. We analyzed the association between mitochondrial haplogroup and severity of lipoatrophy in HIV-infected European American patients on HAART in the Multicenter AIDS cohort Study (MACS) and found that mitochondrial haplogroup H was strongly associated with increased atrophy (arms: p = 0.007, OR = 1.77, 95% CI = 1.17–2.69 legs: p = 0.037, OR = 1.54 95% CI = 1.03–2.31, and buttocks: p = 0.10, OR = 1.41 95% CI = 0.94–2.12). We also saw borderline significance for haplogroup T as protective against lipoatrophy (p = 0.05, OR = 0.52, 95% CI = 0.20–1.00). These data suggest that mitochondrial DNA haplogroup may influence the propensity for lipoatrophy in patients receiving NRTIs. PMID:19339895

  19. Simultaneous DNA and RNA Mapping of Somatic Mitochondrial Mutations across Diverse Human Cancers.

    Directory of Open Access Journals (Sweden)

    James B Stewart

    2015-06-01

    Full Text Available Somatic mutations in the nuclear genome are required for tumor formation, but the functional consequences of somatic mitochondrial DNA (mtDNA mutations are less understood. Here we identify somatic mtDNA mutations across 527 tumors and 14 cancer types, using an approach that takes advantage of evidence from both genomic and transcriptomic sequencing. We find that there is selective pressure against deleterious coding mutations, supporting that functional mitochondria are required in tumor cells, and also observe a strong mutational strand bias, compatible with endogenous replication-coupled errors as the major source of mutations. Interestingly, while allelic ratios in general were consistent in RNA compared to DNA, some mutations in tRNAs displayed strong allelic imbalances caused by accumulation of unprocessed tRNA precursors. The effect was explained by altered secondary structure, demonstrating that correct tRNA folding is a major determinant for processing of polycistronic mitochondrial transcripts. Additionally, the data suggest that tRNA clusters are preferably processed in the 3' to 5' direction. Our study gives insights into mtDNA function in cancer and answers questions regarding mitochondrial tRNA biogenesis that are difficult to address in controlled experimental systems.

  20. Simultaneous DNA and RNA Mapping of Somatic Mitochondrial Mutations across Diverse Human Cancers

    Science.gov (United States)

    Stewart, James B.; Alaei-Mahabadi, Babak; Sabarinathan, Radhakrishnan; Samuelsson, Tore; Gorodkin, Jan; Gustafsson, Claes M.; Larsson, Erik

    2015-01-01

    Somatic mutations in the nuclear genome are required for tumor formation, but the functional consequences of somatic mitochondrial DNA (mtDNA) mutations are less understood. Here we identify somatic mtDNA mutations across 527 tumors and 14 cancer types, using an approach that takes advantage of evidence from both genomic and transcriptomic sequencing. We find that there is selective pressure against deleterious coding mutations, supporting that functional mitochondria are required in tumor cells, and also observe a strong mutational strand bias, compatible with endogenous replication-coupled errors as the major source of mutations. Interestingly, while allelic ratios in general were consistent in RNA compared to DNA, some mutations in tRNAs displayed strong allelic imbalances caused by accumulation of unprocessed tRNA precursors. The effect was explained by altered secondary structure, demonstrating that correct tRNA folding is a major determinant for processing of polycistronic mitochondrial transcripts. Additionally, the data suggest that tRNA clusters are preferably processed in the 3′ to 5′ direction. Our study gives insights into mtDNA function in cancer and answers questions regarding mitochondrial tRNA biogenesis that are difficult to address in controlled experimental systems. PMID:26125550

  1. VARIATION IN MITOCHONDRIAL-DNA LEVELS IN MUSCLE FROM NORMAL CONTROLS - IS DEPLETION OF MTDNA IN PATIENTS WITH MITOCHONDRIAL MYOPATHY A DISTINCT CLINICAL SYNDROME

    NARCIS (Netherlands)

    POULTON, J; SEWRY, C; POTTER, CG; BOUGERON, T; CHRETIEN, D; WIJBURG, FA; MORTEN, KJ; BROWN, G

    1995-01-01

    Recent studies have identified a group of patients with cytochrome oxidase (COX) deficiency presenting in infancy associated with a deficiency of mtDNA in muscle or other affected tissue (Moraes et al 1991). We used a navel approach to compare the level of mitochondrial (mtDNA) compared to nuclear D

  2. Aminoglycoside induced ototoxicity associated with mitochondrial DNA mutations

    Directory of Open Access Journals (Sweden)

    Joseph Foster II

    2016-07-01

    Full Text Available Despite the risk of permanent ototoxic effects, aminoglycosides remain commonly utilized antibiotics worldwide due to low cost and efficiency in treating severe infections. Over the last two decades, mitochondrial mutations have been shown to enhance the likelihood of ototoxic injury. In particular the 1555A>G mutation in the mitochondrial gene MTRNR1 has been strongly associated with the onset of aminoglycoside-induced deafness; though pinning down the exact mechanism of action has thus far been elusive. Clinically aminoglycoside-induced deafness has been characterized by variation in the degree of hearing loss, which has prompted an investigation into genetic modifiers. To date, several putative mutations have been categorized as contributing factors to the onset of deafness with no single variation being sufficient to bring about hearing loss. Meanwhile current methods to mitigate the risk of ototoxic injury are in various stages of development. Efforts to alter the molecular structure of aminoglycosides have shown a potential path to reducing ototoxicity while preserving antibacterial properties, but these drugs are not clinically available. On the other hand, application of preemptive audiometry provides the most readily available method to both monitor and reduce the extent of aminoglycoside-induced deafness.

  3. Diversity of mitochondrial DNA in three Arabian horse strains.

    Science.gov (United States)

    Almarzook, S; Reissmann, M; Brockmann, G A

    2017-05-01

    Arabian horse registries classify Arabian horses based on their dam lineages into five main strains. To test the maternal origin of Syrian Arabian horses, 192 horses representing the three major strains Saglawi, Kahlawi, and Hamdani were sequenced for 353 bp of their mitochondrial displacement loop (D-loop) region. Sequencing revealed 28 haplotypes comprising 38 sequence variations. The haplotype diversity values were 0.95, 0.91, and 0.90 in Kahlawi, Hamdani, and Saglawi strains, respectively. The pair-wise population differentiation estimates (Fst) between strains were low, ranging between 0.098 and 0.205. The haplotype diversity and the pair-wise population differentiation estimates (Fst) between strains showed high diversity within individuals of each strain and low variation between the three strains. Mitochondrial haplotypes scattered all over the neighbor-joining tree without clear separation of the three strains. In the median-joining network, the Syrian horses were grouped into seven major haplogroups. These results suggest that more than five ancestors exist that share common maternal haplotypes with other horse breeds.

  4. Prevalence of migraine in persons with the 3243A>G mutation in mitochondrial DNA

    DEFF Research Database (Denmark)

    Guo, S.; Esserlind, A-L; Andersson, Z

    2016-01-01

    BACKGROUND AND PURPOSE: Over the last three decades mitochondrial dysfunction has been postulated to be a potential mechanism in migraine pathogenesis. The lifetime prevalence of migraine in persons carrying the 3243A>G mutation in mitochondrial DNA was investigated. METHODS: In this cross......% vs. 6%; P persons with the mDNA 3243A>G mutation was found. This finding suggests a clinical association between a monogenetically inherited disorder......-sectional study, 57 mDNA 3243A>G mutation carriers between May 2012 and October 2014 were included. As a control group, a population-based cohort from our epidemiological studies on migraine in Danes was used. History of headache and migraine was obtained by telephone interview, based on a validated semi...

  5. Study on 4977-bp deletion mutation of mitochondrial DNA in lung cancer

    Institute of Scientific and Technical Information of China (English)

    DAI Ji-gang; XIAO Ying-bin; MIN Jia-xin; ZHANG Guo-qiang; YAO Ke; ZHOU Ren-jie

    2005-01-01

    Objective: To study the 4977-bp deletion of mitochondiral DNA in lung cancer, adjacent normal tissue and health lung and its significance in the development of cancer. Methods: Thirty-seven matched lung cancer/adjacent histologically normal and 20 "true" normal lung tissue samples from patients without lung cancer were analyzed by long PCR technique. Results: Mitochondrial DNA 4977-bp deletion was detected in 54. 1% (20/37) of lung cancers, 59.5% (22/37) of adjacent normal and 30.0% (6/30) of"true" normal lung tissues. The correlation of 4977-bp deletion with age and smoking factors was present in our data. Conclusion: Mitochondrial DNA 4977-bp deletion is not specific to lung cancer and unlikely to play an important role in carcinogenesis, and may only reflect the environmental and genetic influences during tumor progression.

  6. Structural Insight into Processive Human Mitochondrial DNA Synthesis and Disease-Related Polymerase Mutations

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Young-Sam; Kennedy, W. Dexter; Yin, Y. Whitney; (Texas)

    2010-09-07

    Human mitochondrial DNA polymerase (Pol {gamma}) is the sole replicase in mitochondria. Pol {gamma} is vulnerable to nonselective antiretroviral drugs and is increasingly associated with mutations found in patients with mitochondriopathies. We determined crystal structures of the human heterotrimeric Pol {gamma} holoenzyme and, separately, a variant of its processivity factor, Pol {gamma}B. The holoenzyme structure reveals an unexpected assembly of the mitochondrial DNA replicase where the catalytic subunit Pol {gamma}A interacts with its processivity factor primarily via a domain that is absent in all other DNA polymerases. This domain provides a structural module for supporting both the intrinsic processivity of the catalytic subunit alone and the enhanced processivity of holoenzyme. The Pol {gamma} structure also provides a context for interpreting the phenotypes of disease-related mutations in the polymerase and establishes a foundation for understanding the molecular basis of toxicity of anti-retroviral drugs targeting HIV reverse transcriptase.

  7. Spontaneous event of mitochondrial DNA mutation, A3243G, found in a family of identical twins.

    Science.gov (United States)

    Harihara, Shinji; Nakamura, Kennichi; Takubo, Kaiyo; Takeuchi, Fujio

    2013-04-01

    A mutation in mitochondrial DNA (mtDNA) A3243G is an important cause of some serious mitochondrial diseases, and maternal inheritance of the mutation has been reported. In order to investigate the heredity of the mutation, we measured the ratio of the mutated mtDNA molecule among 32 families of identical twins. Both twins from one family showed 20.16% and 18.49% mutated molecules, and the level is significantly high in comparison with members of other families and control subjects (0.23-0.86%). Their parents, however, showed normal level of mutated molecules (0.70% and 0.66%). The high-level mutation of the twins may be due to a spontaneous event, which occurred during development of germ line of their mother, or oogenesis of their mother, or during early stage of their development.

  8. Research advance of mitochondrial DNA%线粒体DNA研究进展

    Institute of Scientific and Technical Information of China (English)

    王艳; 刘雅诚; 唐晖

    2002-01-01

    @@ 人类细胞内存在两套基因组,一套是细胞核内的基因组,即核DNA(nuclear DNA,nDNA);另一套是位于细胞质线粒体内的基因组,即线粒体DNA(mitochondrial DNA,mtDNA).由于线粒体在生命活动中的重要作用及其基因组自身特点,使得mtDNA在细胞遗传学、分子遗传学、发育遗传学和法庭科学等领域受到了广泛重视.

  9. Mitochondrial DNA variations in ova and blastocyst: implications in assisted reproduction.

    Science.gov (United States)

    Shamsi, Monis Bilal; Govindaraj, Periyasamy; Chawla, Latika; Malhotra, Neena; Singh, Neeta; Mittal, Suneeta; Talwar, Pankaj; Thangaraj, Kumarasamy; Dada, Rima

    2013-03-01

    Mitochondrial DNA (mtDNA) of oocyte is critical for its function, embryo quality and development. Analysis of complete mtDNA of 49 oocytes and 18 blastocysts from 67 females opting for IVF revealed 437 nucleotide variations. 40.29% samples had either disease associated or non-synonymous novel or pathogenic mutation in evolutionarily conserved regions. Samples with disease associated mtDNA mutations had low fertilization rate and poor embryo quality, however no difference in implantation or clinical pregnancy rate was observed. Screening mtDNA from oocyte/blastocyst is a simple, clinically reliable method for diagnostic evaluation of female infertility and may reduce risk of mtDNA disease transmission.

  10. Mechanism of Homologous Recombination and Implications for Aging-Related Deletions in Mitochondrial DNA

    Science.gov (United States)

    2013-01-01

    SUMMARY Homologous recombination is a universal process, conserved from bacteriophage to human, which is important for the repair of double-strand DNA breaks. Recombination in mitochondrial DNA (mtDNA) was documented more than 4 decades ago, but the underlying molecular mechanism has remained elusive. Recent studies have revealed the presence of a Rad52-type recombination system of bacteriophage origin in mitochondria, which operates by a single-strand annealing mechanism independent of the canonical RecA/Rad51-type recombinases. Increasing evidence supports the notion that, like in bacteriophages, mtDNA inheritance is a coordinated interplay between recombination, repair, and replication. These findings could have profound implications for understanding the mechanism of mtDNA inheritance and the generation of mtDNA deletions in aging cells. PMID:24006472

  11. A patient with two mitochondrial DNA mutations causing PEO and LHON.

    Science.gov (United States)

    Melberg, Atle; Moslemi, Ali-Reza; Palm, Oscar; Raininko, Raili; Stålberg, Erik; Oldfors, Anders

    2009-01-01

    We report a 22-year-old man with PEO and optic atrophy. PEO developed before the onset of optic atrophy. The patient showed mitochondrial myopathy with cytochrome c oxidase deficient fibers. In skeletal muscle the patient was homoplasmic for the mtDNA G11778A Leber hereditary optic neuropathy (LHON) mutation and heteroplasmic for the mtDNA 5 kb "common" deletion mutation. In blood only the homoplasmic LHON mutation was identified. The occurrence of two pathogenic mtDNA mutations is exceedingly rare. The clinical findings in this patient indicate that the combination of the two mtDNA mutations resulted in the expected combined phenotype since the mtDNA deletion mutation accounted for the PEO and the mtDNA G11778A point mutation for the optic atrophy.

  12. High-Resolution Melting (HRM) of Hypervariable Mitochondrial DNA Regions for Forensic Science.

    Science.gov (United States)

    Dos Santos Rocha, Alípio; de Amorim, Isis Salviano Soares; Simão, Tatiana de Almeida; da Fonseca, Adenilson de Souza; Garrido, Rodrigo Grazinoli; Mencalha, Andre Luiz

    2017-08-23

    Forensic strategies commonly are proceeding by analysis of short tandem repeats (STRs); however, new additional strategies have been proposed for forensic science. Thus, this article standardized the high-resolution melting (HRM) of DNA for forensic analyzes. For HRM, mitochondrial DNA (mtDNA) from eight individuals were extracted from mucosa swabs by DNAzol reagent, samples were amplified by PCR and submitted to HRM analysis to identify differences in hypervariable (HV) regions I and II. To confirm HRM, all PCR products were DNA sequencing. The data suggest that is possible discriminate DNA from different samples by HRM curves. Also, uncommon dual-dissociation was identified in a single PCR product, increasing HRM analyzes by evaluation of melting peaks. Thus, HRM is accurate and useful to screening small differences in HVI and HVII regions from mtDNA and increase the efficiency of laboratory routines based on forensic genetics. © 2017 American Academy of Forensic Sciences.

  13. Mitochondrial genome of the moon jelly Aurelia aurita (Cnidaria, Scyphozoa): A linear DNA molecule encoding a putative DNA-dependent DNA polymerase.

    Science.gov (United States)

    Shao, Zhiyong; Graf, Shannon; Chaga, Oleg Y; Lavrov, Dennis V

    2006-10-15

    The 16,937-nuceotide sequence of the linear mitochondrial DNA (mt-DNA) molecule of the moon jelly Aurelia aurita (Cnidaria, Scyphozoa) - the first mtDNA sequence from the class Scypozoa and the first sequence of a linear mtDNA from Metazoa - has been determined. This sequence contains genes for 13 energy pathway proteins, small and large subunit rRNAs, and methionine and tryptophan tRNAs. In addition, two open reading frames of 324 and 969 base pairs in length have been found. The deduced amino-acid sequence of one of them, ORF969, displays extensive sequence similarity with the polymerase [but not the exonuclease] domain of family B DNA polymerases, and this ORF has been tentatively identified as dnab. This is the first report of dnab in animal mtDNA. The genes in A. aurita mtDNA are arranged in two clusters with opposite transcriptional polarities; transcription proceeding toward the ends of the molecule. The determined sequences at the ends of the molecule are nearly identical but inverted and lack any obvious potential secondary structures or telomere-like repeat elements. The acquisition of mitochondrial genomic data for the second class of Cnidaria allows us to reconstruct characteristic features of mitochondrial evolution in this animal phylum.

  14. Toxicity of nucleoside analogues used to treat AIDS and the selectivity of the mitochondrial DNA polymerase.

    Science.gov (United States)

    Lee, Harold; Hanes, Jeremiah; Johnson, Kenneth A

    2003-12-23

    Incorporation of nucleoside analogues by the mitochondrial DNA polymerase has been implicated as the primary cause underlying many of the toxic side effects of these drugs in HIV therapy. Recent success in reconstituting recombinant human enzyme has afforded a detailed mechanistic analysis of the reactions governing nucleotide selectivity of the polymerase and the proofreading exonuclease. The toxic side effects of nucleoside analogues are correlated with the kinetics of incorporation by the mitochondrial DNA polymerase, varying over 6 orders of magnitude in the sequence zalcitabine (ddC) > didanosine (ddI metabolized to ddA) > stavudine (d4T) > lamivudine (3TC) > tenofovir (PMPA) > zidovudine (AZT) > abacavir (metabolized to carbovir, CBV). In this review, we summarize our current efforts to examine the mechanistic basis for nucleotide selectivity by the mitochondrial DNA polymerase and its role in mitochondrial toxicity of nucleoside analogues used to treat AIDS and other viral infections. We will also discuss the promise and underlying challenges for the development of new analogues with lower toxicity.

  15. The role of mitochondrial DNA mutations and free radicals in disease and ageing

    Science.gov (United States)

    Lagouge, M; Larsson, N-G

    2013-01-01

    Considerable efforts have been made to understand the role of oxidative stress in age-related diseases and ageing. The mitochondrial free radical theory of ageing, which proposes that damage to mitochondrial DNA (mtDNA) and other macromolecules caused by the production of reactive oxygen species (ROS) during cellular respiration drives ageing, has for a long time been the central hypothesis in the field. However, in contrast with this theory, evidence from an increasing number of experimental studies has suggested that mtDNA mutations may be generated by replication errors rather than by accumulated oxidative damage. Furthermore, interventions to modulate ROS levels in humans and animal models have not produced consistent results in terms of delaying disease progression and extending lifespan. A number of recent experimental findings strongly question the mitochondrial free radical theory of ageing, leading to the emergence of new theories of how age-associated mitochondrial dysfunction may lead to ageing. These new hypotheses are mainly based on the underlying notion that, despite their deleterious role, ROS are essential signalling molecules that mediate stress responses in general and the stress response to age-dependent damage in particular. This novel view of ROS roles has a clear impact on the interpretation of studies in which antioxidants have been used to treat human age-related diseases commonly linked to oxidative stress. PMID:23432181

  16. The role of mitochondrial DNA mutations and free radicals in disease and ageing.

    Science.gov (United States)

    Lagouge, M; Larsson, N-G

    2013-06-01

    Considerable efforts have been made to understand the role of oxidative stress in age-related diseases and ageing. The mitochondrial free radical theory of ageing, which proposes that damage to mitochondrial DNA (mtDNA) and other macromolecules caused by the production of reactive oxygen species (ROS) during cellular respiration drives ageing, has for a long time been the central hypothesis in the field. However, in contrast with this theory, evidence from an increasing number of experimental studies has suggested that mtDNA mutations may be generated by replication errors rather than by accumulated oxidative damage. Furthermore, interventions to modulate ROS levels in humans and animal models have not produced consistent results in terms of delaying disease progression and extending lifespan. A number of recent experimental findings strongly question the mitochondrial free radical theory of ageing, leading to the emergence of new theories of how age-associated mitochondrial dysfunction may lead to ageing. These new hypotheses are mainly based on the underlying notion that, despite their deleterious role, ROS are essential signalling molecules that mediate stress responses in general and the stress response to age-dependent damage in particular. This novel view of ROS roles has a clear impact on the interpretation of studies in which antioxidants have been used to treat human age-related diseases commonly linked to oxidative stress. © 2013 The Association for the Publication of the Journal of Internal Medicine.

  17. Mitochondrial DNA deletion in a patient with combined features of Leigh and Pearson syndromes

    Energy Technology Data Exchange (ETDEWEB)

    Blok, R.B.; Thorburn, D.R.; Danks, D.M. [Royal Children`s Hospital, Melbourne (Australia)] [and others

    1994-09-01

    We describe a heteroplasmic 4237 bp mitochondrial DNA (mtDNA) deletion in an 11 year old girl who has suffered from progressive illness since birth. She has some features of Leigh syndrome (global developmental delay with regression, brainstem dysfunction and lactic acidosis), together with other features suggestive of Pearson syndrome (history of pancytopenia and failure to thrive). The deletion was present at a level greater than 50% in skeletal muscle, but barely detectable in skin fibroblasts following Southern blot analysis, and only observed in blood following PCR analysis. The deletion spanned nt 9498 to nt 13734, and was flanked by a 12 bp direct repeat. Genes for cytochrome c oxidase subunit III, NADH dehydrogenase subunits 3, 4L, 4 and 5, and tRNAs for glycine, arginine, histidine, serine({sup AGY}) and leucine({sup CUN}) were deleted. Southern blotting also revealed an altered Apa I restriction site which was shown by sequence analysis to be caused by G{r_arrow}A nucleotide substitution at nt 1462 in the 12S rRNA gene. This was presumed to be a polymorphism. No abnormalities of mitochondrial ultrastructure, distribution or of respiratory chain enzyme complexes I-IV in skeletal muscle were observed. Mitochondrial disorders with clinical features overlapping more than one syndrome have been reported previously. This case further demonstrates the difficulty in correlating observed clinical features with a specific mitochondrial DNA mutation.

  18. Comparative sequence analysis of the non-protein-coding mitochondrial DNA of inbred rat strains.

    Directory of Open Access Journals (Sweden)

    Avinash Abhyankar

    Full Text Available The proper function of mammalian mitochondria necessitates a coordinated expression of both nuclear and mitochondrial genes, most likely due to the co-evolution of nuclear and mitochondrial genomes. The non-protein coding regions of mitochondrial DNA (mtDNA including the D-loop, tRNA and rRNA genes form a major component of this regulated expression unit. Here we present comparative analyses of the non-protein-coding regions from 27 Rattus norvegicus mtDNA sequences. There were two variable positions in 12S rRNA, 20 in 16S rRNA, eight within the tRNA genes and 13 in the D-loop. Only one of the three neutrality tests used demonstrated statistically significant evidence for selection in 16S rRNA and tRNA-Cys. Based on our analyses of conserved sequences, we propose that some of the variable nucleotide positions identified in 16S rRNA and tRNA-Cys, and the D-loop might be important for mitochondrial function and its regulation.

  19. The Mitochondrial DNA Mutation at Position 11778 in Chinese Families with Leber's Hereditary Optic Neuropathy

    Institute of Scientific and Technical Information of China (English)

    1994-01-01

    We amplified the 340 bp of mitochondrial DMA (mtDNA) by PCR including the recognized sequence of restriction enzyme of SfaN I . After amplification and digestion of SfaN I , two bands of 190 bp and 150 bp appeared in the mtDNA of four normal individuals but only one band of 340 bp appeared in the mtDNA with the mutation of G to A at the site of the nucleotide 11778 because such mutation destroyed the recognized sequence of SfaN I . We studied the mtDNAs of the patients with Leber's hereditary optic neur...

  20. Mitochondrial DNA deletion mutations in adult mouse cardiac side population cells

    Energy Technology Data Exchange (ETDEWEB)

    Lushaj, Entela B., E-mail: lushaj@surgery.wisc.edu [Division of Cardiothoracic Surgery, Department of Surgery, School of Medicine and Public Health, University of Wisconsin, Madison, WI 53792 (United States); Lozonschi, Lucian; Barnes, Maria; Anstadt, Emily; Kohmoto, Takushi [Division of Cardiothoracic Surgery, Department of Surgery, School of Medicine and Public Health, University of Wisconsin, Madison, WI 53792 (United States)

    2012-06-01

    We investigated the presence and potential role of mitochondrial DNA (mtDNA) deletion mutations in adult cardiac stem cells. Cardiac side population (SP) cells were isolated from 12-week-old mice. Standard polymerase chain reaction (PCR) was used to screen for the presence of mtDNA deletion mutations in (a) freshly isolated SP cells and (b) SP cells cultured to passage 10. When present, the abundance of mtDNA deletion mutation was analyzed in single cell colonies. The effect of different levels of deletion mutations on SP cell growth and differentiation was determined. MtDNA deletion mutations were found in both freshly isolated and cultured cells from 12-week-old mice. While there was no significant difference in the number of single cell colonies with mtDNA deletion mutations from any of the groups mentioned above, the abundance of mtDNA deletion mutations was significantly higher in the cultured cells, as determined by quantitative PCR. Within a single clonal cell population, the detectable mtDNA deletion mutations were the same in all cells and unique when compared to deletions of other colonies. We also found that cells harboring high levels of mtDNA deletion mutations (i.e. where deleted mtDNA comprised more than 60% of total mtDNA) had slower proliferation rates and decreased differentiation capacities. Screening cultured adult stem cells for mtDNA deletion mutations as a routine assessment will benefit the biomedical application of adult stem cells.

  1. Mitochondrial DNA alterations of peripheral lymphocytes in acute lymphoblastic leukemia patients undergoing total body irradiation therapy

    Directory of Open Access Journals (Sweden)

    Ji Fuyun

    2011-10-01

    Full Text Available Abstract Background Mitochondrial DNA (mtDNA alterations, including mtDNA copy number and mtDNA 4977 bp common deletion (CD, are key indicators of irradiation-induced damage. The relationship between total body irradiation (TBI treatment and mtDNA alterations in vivo, however, has not been postulated yet. The aim of this study is to analyze mtDNA alterations in irradiated human peripheral lymphocytes from acute lymphoblastic leukemia (ALL patients as well as to take them as predictors for radiation toxicity. Methods Peripheral blood lymphocytes were isolated from 26 ALL patients 24 hours after TBI preconditioning (4.5 and 9 Gy, respectively. Extracted DNA was analyzed by real-time PCR method. Results Average 2.31 times mtDNA and 0.53 fold CD levels were observed after 4.5 Gy exposure compared to their basal levels. 9 Gy TBI produced a greater response of both mtDNA and CD levels than 4.5 Gy. Significant inverse correlation was found between mtDNA content and CD level at 4.5 and 9 Gy (P = 0.037 and 0.048. Moreover, mtDNA content of lymphocytes without irradiation was found to be correlated to age. Conclusions mtDNA and CD content may be considered as predictive factors to radiation toxicity.

  2. [Mitochondrial DNA polymorphisms shared between modern humans and neanderthals: adaptive convergence or evidence for interspecific hybridization?].

    Science.gov (United States)

    Maliarchuk, B A

    2013-09-01

    An analysis of the variability of the nucleotide sequences in the mitochondrial genome of modern humans, neanderthals, Denisovans, and other primates has shown that there are shared polymorphisms at positions 2758 and 7146 between modern Homo sapiens (in phylogenetic cluster L2'3'4'5'6) and Homo neanderthalensis (in the group of European neanderthals younger than 48000 years). It is suggested that the convergence may be due to adaptive changes in the mitochondrial genomes of modern humans and neanderthals or interspecific hybridization associated with mtDNA recombination.

  3. Accurate measurement of circulating mitochondrial DNA content from human blood samples using real-time quantitative PCR.

    Science.gov (United States)

    Ajaz, Saima; Czajka, Anna; Malik, Afshan

    2015-01-01

    We describe a protocol to accurately measure the amount of human mitochondrial DNA (MtDNA) in peripheral blood samples which can be modified to quantify MtDNA from other body fluids, human cells, and tissues. This protocol is based on the use of real-time quantitative PCR (qPCR) to quantify the amount of MtDNA relative to nuclear DNA (designated the Mt/N ratio). In the last decade, there have been increasing numbers of studies describing altered MtDNA or Mt/N in circulation in common nongenetic diseases where mitochondrial dysfunction may play a role (for review see Malik and Czajka, Mitochondrion 13:481-492, 2013). These studies are distinct from those looking at genetic mitochondrial disease and are attempting to identify acquired changes in circulating MtDNA content as an indicator of mitochondrial function. However, the methodology being used is not always specific and reproducible. As more than 95 % of the human mitochondrial genome is duplicated in the human nuclear genome, it is important to avoid co-amplification of nuclear pseudogenes. Furthermore, template preparation protocols can also affect the results because of the size and structural differences between the mitochondrial and nuclear genomes. Here we describe how to (1) prepare DNA from blood samples; (2) pretreat the DNA to prevent dilution bias; (3) prepare dilution standards for absolute quantification using the unique primers human mitochondrial genome forward primer (hMitoF3) and human mitochondrial genome reverse primer(hMitoR3) for the mitochondrial genome, and human nuclear genome forward primer (hB2MF1) and human nuclear genome reverse primer (hB2MR1) primers for the human nuclear genome; (4) carry out qPCR for either relative or absolute quantification from test samples; (5) analyze qPCR data; and (6) calculate the sample size to adequately power studies. The protocol presented here is suitable for high-throughput use.

  4. Seventeen new complete mtDNA sequences reveal extensive mitochondrial genome evolution within the Demospongiae.

    Directory of Open Access Journals (Sweden)

    Xiujuan Wang

    Full Text Available Two major transitions in animal evolution--the origins of multicellularity and bilaterality--correlate with major changes in mitochondrial DNA (mtDNA organization. Demosponges, the largest class in the phylum Porifera, underwent only the first of these transitions and their mitochondrial genomes display a peculiar combination of ancestral and animal-specific features. To get an insight into the evolution of mitochondrial genomes within the Demospongiae, we determined 17 new mtDNA sequences from this group and analyzing them with five previously published sequences. Our analysis revealed that all demosponge mtDNAs are 16- to 25-kbp circular molecules, containing 13-15 protein genes, 2 rRNA genes, and 2-27 tRNA genes. All but four pairs of sampled genomes had unique gene orders, with the number of shared gene boundaries ranging from 1 to 41. Although most demosponge species displayed low rates of mitochondrial sequence evolution, a significant acceleration in evolutionary rates occurred in the G1 group (orders Dendroceratida, Dictyoceratida, and Verticillitida. Large variation in mtDNA organization was also observed within the G0 group (order Homosclerophorida including gene rearrangements, loss of tRNA genes, and the presence of two introns in Plakortis angulospiculatus. While introns are rare in modern-day demosponge mtDNA, we inferred that at least one intron was present in cox1 of the common ancestor of all demosponges. Our study uncovered an extensive mitochondrial genomic diversity within the Demospongiae. Although all sampled mitochondrial genomes retained some ancestral features, including a minimally modified genetic code, conserved structures of tRNA genes, and presence of multiple non-coding regions, they vary considerably in their size, gene content, gene order, and the rates of sequence evolution. Some of the changes in demosponge mtDNA, such as the loss of tRNA genes and the appearance of hairpin-containing repetitive elements

  5. Seventeen New Complete mtDNA Sequences Reveal Extensive Mitochondrial Genome Evolution within the Demospongiae

    Science.gov (United States)

    Wang, Xiujuan; Lavrov, Dennis V.

    2008-01-01

    Two major transitions in animal evolution–the origins of multicellularity and bilaterality–correlate with major changes in mitochondrial DNA (mtDNA) organization. Demosponges, the largest class in the phylum Porifera, underwent only the first of these transitions and their mitochondrial genomes display a peculiar combination of ancestral and animal-specific features. To get an insight into the evolution of mitochondrial genomes within the Demospongiae, we determined 17 new mtDNA sequences from this group and analyzing them with five previously published sequences. Our analysis revealed that all demosponge mtDNAs are 16- to 25-kbp circular molecules, containing 13–15 protein genes, 2 rRNA genes, and 2–27 tRNA genes. All but four pairs of sampled genomes had unique gene orders, with the number of shared gene boundaries ranging from 1 to 41. Although most demosponge species displayed low rates of mitochondrial sequence evolution, a significant acceleration in evolutionary rates occurred in the G1 group (orders Dendroceratida, Dictyoceratida, and Verticillitida). Large variation in mtDNA organization was also observed within the G0 group (order Homosclerophorida) including gene rearrangements, loss of tRNA genes, and the presence of two introns in Plakortis angulospiculatus. While introns are rare in modern-day demosponge mtDNA, we inferred that at least one intron was present in cox1 of the common ancestor of all demosponges. Our study uncovered an extensive mitochondrial genomic diversity within the Demospongiae. Although all sampled mitochondrial genomes retained some ancestral features, including a minimally modified genetic code, conserved structures of tRNA genes, and presence of multiple non-coding regions, they vary considerably in their size, gene content, gene order, and the rates of sequence evolution. Some of the changes in demosponge mtDNA, such as the loss of tRNA genes and the appearance of hairpin-containing repetitive elements, occurred in

  6. Loss of the Association between Telomere Length and Mitochondrial DNA Copy Number Contribute to Colorectal Carcinogenesis.

    Science.gov (United States)

    Lee, Hyunsu; Cho, Ji-Hyoung; Park, Won-Jin; Jung, Soo-Jung; Choi, In-Jang; Lee, Jae-Ho

    2017-05-09

    Positive association between telomere length and mitochondrial DNA (mtDNA) copy number were introduced in healthy and patients with psychiatric disorder. Based on frequent genetic changes of telomere and mitochondria in colorectal carcinomas (CRC), we studied their clinical characteristics and their association in colorectal carcinogenesis. DNA was extracted from 109 CRCs, 64 colorectal tubular adenomas (TAs), and 28 serrated polyps (SPs), and then, telomere length and mtDNA copy number were analyzed in these legions by using a real-time PCR assay. Telomere length and mtDNA copy number (mean ± S.D) in CRCs was 1.87 ± 1.52 and 1.61 ± 1.37, respectively. In TAs and SPs, relative mtDNA copy number was 0.92 ± 0.71 and 1.84 ± 1.06, respectively, shoing statistical difference (p = 0.017). However, telomere length was similar in these precancerous legions. Telomere length and mtDNA copy number did not show clinical and prognostic values in CRCs, however, positive correlation between telomere length and mitochondrial DNA copy number were found in CRC (r = 0.408, p < 0.001). However, this association was not shown in precancerous lesions (r = -0.031, p = 0.765). This result suggests that loss of co-regulation between telomeres and mitochondrial function may induce the initiation or play a role as trigger factor of colorectal carcinogenesis.

  7. Mutations of mitochondrial DNA polymerase gammaA are a frequent cause of autosomal dominant or recessive progressive external ophthalmoplegia.

    Science.gov (United States)

    Lamantea, Eleonora; Tiranti, Valeria; Bordoni, Andreina; Toscano, Antonio; Bono, Francesco; Servidei, Serena; Papadimitriou, Alex; Spelbrink, Hans; Silvestri, Laura; Casari, Giorgio; Comi, Giacomo P; Zeviani, Massimo

    2002-08-01

    One form of familial progressive external ophthalmoplegia with multiple mitochondrial DNA deletions recently has been associated with mutations in POLG1, the gene encoding pol gammaA, the catalytic subunit of mitochondrial DNA polymerase. We screened the POLG1 gene in several PEO families and identified five different heterozygous missense mutations of POLG1 in 10 autosomal dominant families. Recessive mutations were found in three families. Our data show that mutations of POLG1 are the most frequent cause of familial progressive external ophthalmoplegia associated with accumulation of multiple mitochondrial DNA deletions, accounting for approximately 45% of our family cohort.

  8. Selective removal of deletion-bearing mitochondrial DNA in heteroplasmic Drosophila

    Science.gov (United States)

    Kandul, Nikolay P.; Zhang, Ting; Hay, Bruce A.; Guo, Ming

    2016-01-01

    Mitochondrial DNA (mtDNA) often exists in a state of heteroplasmy, in which mutant mtDNA co-exists in cells with wild-type mtDNA. High frequencies of pathogenic mtDNA result in maternally inherited diseases; maternally and somatically acquired mutations also accumulate over time and contribute to diseases of ageing. Reducing heteroplasmy is therefore a therapeutic goal and in vivo models in post-mitotic tissues are needed to facilitate these studies. Here we describe a transgene-based model of a heteroplasmic lethal mtDNA deletion (mtDNAΔ) in adult Drosophila muscle. Stimulation of autophagy, activation of the PINK1/parkin pathway or decreased levels of mitofusin result in a selective decrease in mtDNAΔ. Decreased levels of mitofusin and increased levels of ATPIF1, an inhibitor of ATP synthase reversal-dependent mitochondrial repolarization, result in a further decrease in mtDNAΔ levels. These results show that an adult post-mitotic tissue can be cleansed of a deleterious genome, suggesting that therapeutic removal of mutant mtDNA can be achieved. PMID:27841259

  9. [Distribution of foreign mitochondrial DNA during the first splittings of the transmitochondrial mouse embryos].

    Science.gov (United States)

    Kustova, M E; Sokolova, V A; Bass, M G; Zakharova, F M; Sorokin, A V; Vasil'ev, V B

    2008-01-01

    Distribution of human mitochondrial DNA (mtDNA) among separate murine blastomeres was analyzed during the splitting of embryos in which the suspension of human mitochondria had been injected at the one- or two-cell stage. Human mtDNA was detected by PCR with species specific primers. The total amount of the two- and four-cell murine embryos analyzed in the study was 339. In all embryos examined the copies of human mitochondrial genome were revealed along with murine mtDNA, which indicated the phenomenon of an artificially modeled heteroplasmy. The foreign mtDNA was not ubiquitous among the blastomeres of transmitochondrial embryos. Mathematical analysis of the results showed that in the period between the injection of human mitochondria and the subsequent splitting no equal distribution of the human mtDNA occurred in the cytoplasm. These results also point at the presence of more than 2-3 segregation units of mtDNA in the entire pool of mitochondria (about 5 x 10(2)) introduced into an embryo by microinjection.

  10. Polymorphisms in the control region of mitochondrial DNA associated with elite Japanese athlete status.

    Science.gov (United States)

    Mikami, E; Fuku, N; Takahashi, H; Ohiwa, N; Pitsiladis, Y P; Higuchi, M; Kawahara, T; Tanaka, M

    2013-10-01

    The control region of mitochondrial DNA (mtDNA) contains the main regulatory elements for mtDNA replication and transcription. Certain polymorphisms in this region would, therefore, contribute to elite athletic performance, because mitochondrial function is one of determinants of physical performance. The present study was undertaken to examine the effect of polymorphisms in this region on elite athlete status by sequencing the mtDNA control region. Subjects comprised 185 elite Japanese athletes who had represented Japan at international competitions (i.e., 100 endurance/middle-power athletes: EMA; 85 sprint/power athletes: SPA), and 672 Japanese controls (CON). The mtDNA control region was analyzed by direct sequencing. Frequency differences of polymorphisms (minor allele frequency ≥ 0.05) in the mtDNA control region between EMA, SPA, and CON were examined. EMA displayed excess of three polymorphisms [m.152T>C, m.514(CA)n repeat (n ≥ 5), and poly-C stretch at m.568-573 (C ≥ 7)] compared with CON. On the other hand, SPA showed greater frequency of the m.204T>C polymorphism compared with CON. In addition, none of the SPA had m.16278C>T polymorphism, whereas the frequencies of this polymorphism in CON and EMA were 8.3% and 10.0%, respectively. These findings imply that several polymorphisms detected in the control region of mtDNA may influence physical performance probably in a functional manner.

  11. Genetic variation of the East Balkan Swine (Sus scrofa) in Bulgaria, revealed by mitochondrial DNA and Y chromosomal DNA.

    Science.gov (United States)

    Hirata, D; Doichev, V D; Raichev, E G; Palova, N A; Nakev, J L; Yordanov, Y M; Kaneko, Y; Masuda, R

    2015-04-01

    East Balkan Swine (EBS) Sus scrofa is the only aboriginal domesticated pig breed in Bulgaria and is distributed on the western coast of the Black Sea in Bulgaria. To reveal the breed's genetic characteristics, we analysed mitochondrial DNA (mtDNA) and Y chromosomal DNA sequences of EBS in Bulgaria. Nucleotide diversity (πn ) of the mtDNA control region, including two newly found haplotypes, in 54 EBS was higher (0.014 ± 0.007) compared with that of European (0.005 ± 0.003) and Asian (0.006 ± 0.003) domestic pigs and wild boar. The median-joining network based on the mtDNA control region showed that the EBS and wild boar in Bulgaria comprised mainly two major mtDNA clades, European clade E1 (61.3%) and Asian clade A (38.7%). The coexistence of two mtDNA clades in EBS in Bulgaria may be the relict of historical pig translocation. Among the Bulgarian EBS colonies, the geographical differences in distribution of two mtDNA clades (E1 and A) could be attributed to the source pig populations and/or historical crossbreeding with imported pigs. In addition, analysis of the Y chromosomal DNA sequences for the EBS revealed that all of the EBS had haplotype HY1, which is dominant in European domestic pigs.

  12. The African diaspora: mitochondrial DNA and the Atlantic slave trade.

    Science.gov (United States)

    Salas, Antonio; Richards, Martin; Lareu, María-Victoria; Scozzari, Rosaria; Coppa, Alfredo; Torroni, Antonio; Macaulay, Vincent; Carracedo, Angel

    2004-03-01

    Between the 15th and 19th centuries ad, the Atlantic slave trade resulted in the forced movement of approximately 13 million people from Africa, mainly to the Americas. Only approximately 11 million survived the passage, and many more died in the early years of captivity. We have studied 481 mitochondrial DNAs (mtDNAs) of recent African ancestry in the Americas and in Eurasia, in an attempt to trace them back to particular regions of Africa. Our results show that mtDNAs in America and Eurasia can, in many cases, be traced to broad geographical regions within Africa, largely in accordance with historical evidence, and raise the possibility that a greater resolution may be possible in the future. However, they also indicate that, at least for the moment, considerable caution is warranted when assessing claims to be able to trace the ancestry of particular lineages to a particular locality within modern-day Africa.

  13. The African Diaspora: Mitochondrial DNA and the Atlantic Slave Trade

    Science.gov (United States)

    Salas, Antonio; Richards, Martin; Lareu, María-Victoria; Scozzari, Rosaria; Coppa, Alfredo; Torroni, Antonio; Macaulay, Vincent; Carracedo, Ángel

    2004-01-01

    Between the 15th and 19th centuries ad, the Atlantic slave trade resulted in the forced movement of ∼13 million people from Africa, mainly to the Americas. Only ∼11 million survived the passage, and many more died in the early years of captivity. We have studied 481 mitochondrial DNAs (mtDNAs) of recent African ancestry in the Americas and in Eurasia, in an attempt to trace them back to particular regions of Africa. Our results show that mtDNAs in America and Eurasia can, in many cases, be traced to broad geographical regions within Africa, largely in accordance with historical evidence, and raise the possibility that a greater resolution may be possible in the future. However, they also indicate that, at least for the moment, considerable caution is warranted when assessing claims to be able to trace the ancestry of particular lineages to a particular locality within modern-day Africa. PMID:14872407

  14. Mitochondrial DNA Mutations in Epithelial Ovarian Tumor Progression

    Science.gov (United States)

    2007-12-01

    1648 del T1 93 98 92 72 tRNAval 1653 del T1 20 98 87 100 T1653A1 76 tRNAval 1659 del T1 89 78 95 94 COX2 8237 del A1 68 82 49 67 ATP6 A8860G 96 96 97 83...V: Identification of somatic and germline mitochon- drial DNA sequence variants in prostate cancer patients. Mutation Research 2006, 595:42-51. 7...SH, Marshall FF, Wallace DC: mtDNA mutations increase tumorigenicity in prostate can- cer. PNAS 2005, 102(3):719-24. 9. Penta JS, Johnson FM

  15. Mitochondrial DNA control region variation in Dubai, United Arab Emirates.

    Science.gov (United States)

    Alshamali, Farida; Brandstätter, Anita; Zimmermann, Bettina; Parson, Walther

    2008-01-01

    249 entire mtDNA control region sequences were generated and analyzed in a population sample from Dubai, one of the seven United Arab Emirates. The control region was amplified in one piece and sequenced with different sequencing primers. Sequence evaluation was performed twice and validated by a third senior mtDNA scientist. Phylogenetic analyses were used for quality assurance purposes and for the determination of the haplogroup affiliation of the samples. Upon publication, the population data are going to be available in the EMPOP database (www.empop.org).

  16. Inheritance of mitochondrial DNA in serially recloned pigs by somatic cell nuclear transfer (SCNT)

    Energy Technology Data Exchange (ETDEWEB)

    Do, Minhwa; Jang, Won-Gu; Hwang, Jeong Hee; Jang, Hoon; Kim, Eun-Jung; Jeong, Eun-Jeong [Regenerative Medicine Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon 305 806 (Korea, Republic of); Shim, Hosup [Department of Physiology, Dankook University School of Medicine, Cheonan 330 714 (Korea, Republic of); Hwang, Sung Soo; Oh, Keon Bong; Byun, Sung June [Animal Biotechnology Division, National Institute of Animal Science, Rural Development Administration, Suwon (Korea, Republic of); Kim, Jin-Hoi [Department of Animal Biotechnology, Konkuk University, Seoul 143 701 (Korea, Republic of); Lee, Jeong Woong, E-mail: jwlee@kribb.re.kr [Regenerative Medicine Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon 305 806 (Korea, Republic of)

    2012-08-10

    Highlights: Black-Right-Pointing-Pointer We success serial SCNT through the third generation using pig fibroblasts. Black-Right-Pointing-Pointer Donor-specific mtDNA in the recloned pigs was detected. Black-Right-Pointing-Pointer SCNT affect mtDNA mounts. -- Abstract: Somatic cell nuclear transfer (SCNT) has been established for the transmission of specific nuclear DNA. However, the fate of donor mitochondrial DNA (mtDNA) remains unclear. Here, we examined the fate of donor mtDNA in recloned pigs through third generations. Fibroblasts of recloned pigs were obtained from offspring of each generation produced by fusion of cultured fibroblasts from a Minnesota miniature pig (MMP) into enucleated oocytes of a Landrace pig. The D-loop regions from the mtDNA of donor and recipient differ at nucleotide sequence positions 16050 (A{yields}T), 16062 (T{yields}C), and 16135 (G{yields}A). In order to determine the fate of donor mtDNA in recloned pigs, we analyzed the D-loop region of the donor's mtDNA by allele-specific PCR (AS-PCR) and real-time PCR. Donor mtDNA was successfully detected in all recloned offspring (F1, F2, and F3). These results indicate that heteroplasmy that originate from donor and recipient mtDNA is maintained in recloned pigs, resulting from SCNT, unlike natural reproduction.

  17. Mitochondrial DNA sequence-based phylogenetic relationship among flesh flies of the genus Sarcophaga (Sarcophagidae: Diptera)

    Indian Academy of Sciences (India)

    Neelam Bajpai; Raghav Ram Tewari

    2010-04-01

    The phylogenetic relationships among flesh flies of the family Sarcophagidae has been based mainly on the morphology of male genitalia. However, the male genitalic character-based relationships are far from satisfactory. Therefore, in the present study mitochondrial DNA has been used as marker to unravel genetic relatedness and to construct phylogeny among five sympatric species of the genus Sarcophaga. Two mitochondrial genes viz., cytochrome oxidase subunit 1 (COI) and NAD dehydrogenase subunit 5 (ND5) were sequenced and genetic distance values were calculated on the basis of sequence differences in both the mitochondrial genes. The data revealed very few genetic difference among the five species for the COI and ND5 gene sequences.

  18. Phylogenetic Position of a Copper Age Sheep (Ovis aries) Mitochondrial DNA

    Science.gov (United States)

    Olivieri, Cristina; Ermini, Luca; Rizzi, Ermanno; Corti, Giorgio; Luciani, Stefania; Marota, Isolina; De Bellis, Gianluca; Rollo, Franco

    2012-01-01

    Background Sheep (Ovis aries) were domesticated in the Fertile Crescent region about 9,000-8,000 years ago. Currently, few mitochondrial (mt) DNA studies are available on archaeological sheep. In particular, no data on archaeological European sheep are available. Methodology/Principal Findings Here we describe the first portion of mtDNA sequence of a Copper Age European sheep. DNA was extracted from hair shafts which were part of the clothes of the so-called Tyrolean Iceman or Ötzi (5,350 - 5,100 years before present). Mitochondrial DNA (a total of 2,429 base pairs, encompassing a portion of the control region, tRNAPhe, a portion of the 12S rRNA gene, and the whole cytochrome B gene) was sequenced using a mixed sequencing procedure based on PCR amplification and 454 sequencing of pooled amplification products. We have compared the sequence with the corresponding sequence of 334 extant lineages. Conclusions/Significance A phylogenetic network based on a new cladistic notation for the mitochondrial diversity of domestic sheep shows that the Ötzi's sheep falls within haplogroup B, thus demonstrating that sheep belonging to this haplogroup were already present in the Alps more than 5,000 years ago. On the other hand, the lineage of the Ötzi's sheep is defined by two transitions (16147, and 16440) which, assembled together, define a motif that has not yet been identified in modern sheep populations. PMID:22457789

  19. Detection of Y chromosome microdeletions and mitochondrial DNA mutations in male infertility patients.

    Science.gov (United States)

    Güney, A I; Javadova, D; Kırac, D; Ulucan, K; Koc, G; Ergec, D; Tavukcu, H; Tarcan, T

    2012-04-27

    Infertility affects about 10-15% of all couples attempting pregnancy with infertility attributed to the male partner in approximately half of the cases. Proposed causes of male infertility include sperm motility disturbances, Y chromosome microdeletions, chromosomal abnormalities, single gene mutations, and sperm mitochondrial DNA (mtDNA) rearrangements. To investigate the etiology of decreased sperm fertility and motility of sperm and to develop an appropriate therapeutic strategy, the molecular basis of these defects must be elucidated. In this study, we aimed to reveal the relationships between the genetic factors including sperm mtDNA mutations, Y chromosome microdeletions, and sperm parameters that can be regarded as candidate factors for male infertility. Thirty men with a history of infertility and 30 fertile men were recruited to the study. Y chromosome microdeletions were analyzed by multiplex PCR. Mitochondrial genes ATPase6, Cytb, and ND1, were amplified by PCR and then analyzed by direct sequencing. No Y chromosome microdeletions were detected in either group. However, a total of 38 different nucleotide substitutions were identified in the examined mitochondrial genes in both groups, all of which are statistically non-significant. Fifteen substitutions caused an amino acid change and 12 were considered novel mutations. As a conclusion, mtDNA mutations and Y chromosome microdeletions in male infertility should be examined in larger numbers in order to clarify the effect of genetic factors.

  20. MITOCHONDRIAL AND MICROSATELLITE DNA ANALYSES OF AUSTROPOTAMOBIUS PALLIPES POPULATIONS IN SOUTH TYROL (ITALY AND TYROL (AUSTRIA

    Directory of Open Access Journals (Sweden)

    BARIC S.

    2005-07-01

    Full Text Available Historical data show that the native white-clawed crayfish, Austropotamobius pallipes, was once widespread throughout South Tyrol (northern Italy, whereas recent studies identified only half a dozen remaining populations. In order to implement conservation strategies based on knowledge of genetic diversity, each of the six remaining populations in South Tyrol and one population in Tyrol (Austria were investigated by (i sequencing segments of two mitochondrial DNA genes, 16S rRNA and cytochrome c oxidase subunit I, and (ii by analysing four microsatellite DNA loci. Extremely low degrees of genetic differentiation within and among the South Tyrolean populations of A. pallipes were found with mitochondrial DNA sequences. In contrast, microsatellite data displayed not only substantial genetic structure among populations, but also moderate genetic variability within four out of six populations in South Tyrol. The two remaining populations revealed a complete absence of genetic variability. Moreover, both of these populations as well as the population from Austria were fixed for single alleles at three of the four investigated microsatellite loci. Our data have important conservation implications and also show that mitochondrial DNA is not always a sufficient tool to study crayfish populations on a small geographical scale.

  1. Phylogenetic position of a copper age sheep (Ovis aries mitochondrial DNA.

    Directory of Open Access Journals (Sweden)

    Cristina Olivieri

    Full Text Available BACKGROUND: Sheep (Ovis aries were domesticated in the Fertile Crescent region about 9,000-8,000 years ago. Currently, few mitochondrial (mt DNA studies are available on archaeological sheep. In particular, no data on archaeological European sheep are available. METHODOLOGY/PRINCIPAL FINDINGS: Here we describe the first portion of mtDNA sequence of a Copper Age European sheep. DNA was extracted from hair shafts which were part of the clothes of the so-called Tyrolean Iceman or Ötzi (5,350-5,100 years before present. Mitochondrial DNA (a total of 2,429 base pairs, encompassing a portion of the control region, tRNA(Phe, a portion of the 12S rRNA gene, and the whole cytochrome B gene was sequenced using a mixed sequencing procedure based on PCR amplification and 454 sequencing of pooled amplification products. We have compared the sequence with the corresponding sequence of 334 extant lineages. CONCLUSIONS/SIGNIFICANCE: A phylogenetic network based on a new cladistic notation for the mitochondrial diversity of domestic sheep shows that the Ötzi's sheep falls within haplogroup B, thus demonstrating that sheep belonging to this haplogroup were already present in the Alps more than 5,000 years ago. On the other hand, the lineage of the Ötzi's sheep is defined by two transitions (16147, and 16440 which, assembled together, define a motif that has not yet been identified in modern sheep populations.

  2. Association of telomere length and mitochondrial DNA copy number in a community sample of healthy adults.

    Science.gov (United States)

    Tyrka, Audrey R; Carpenter, Linda L; Kao, Hung-Teh; Porton, Barbara; Philip, Noah S; Ridout, Samuel J; Ridout, Kathryn K; Price, Lawrence H

    2015-06-01

    Cellular aging plays a role in longevity and senescence, and has been implicated in medical and psychiatric conditions, including heart disease, cancer, major depression and posttraumatic stress disorder. Telomere shortening and mitochondrial dysfunction are thought to be central to the cellular aging process. The present study examined the association between mitochondrial DNA (mtDNA) copy number and telomere length in a sample of medically healthy adults. Participants (total n=392) were divided into 4 groups based on the presence or absence of early life adversity and lifetime psychopathology: No Adversity/No Disorder, n=136; Adversity/No Disorder, n=91; No Adversity/Disorder, n=46; Adversity/Disorder, n=119. Telomere length and mtDNA copy number were measured using quantitative polymerase chain reaction. There was a positive correlation between mtDNA and telomere length in the entire sample (r=0.120, ptelomere length in a large group of women and men both with and without early adversity and psychopathology, suggesting co-regulation of telomeres and mitochondrial function. The mechanisms underlying this association may be important in the pathophysiology of age-related medical conditions, such as heart disease and cancer, as well as for stress-associated psychiatric disorders.

  3. Evaluation of a multicapillary electrophoresis instrument for mitochondrial DNA typing.

    Science.gov (United States)

    Stewart, John E B; Aagaard, Patricia J; Pokorak, Eric G; Polanskey, Deborah; Budowle, Bruce

    2003-05-01

    Laser-induced detection of fluorescent labeled PCR products and multi-wavelength detection (i.e., multicolor analysis) enables rapid generation of mtDNA sequencing profiles. Traditionally, polyacrylamide slab gels have been used as the electrophoretic medium for mtDNA sequencing in forensic analyses. Replacement of slab gel electrophoresis with capillary electrophoresis (CE) can facilitate automation of the analytical process. Automation and high throughput can be further enhanced by using multicapillary electrophoretic systems. The use of the ABI Prism 3100 Genetic Analyzer (ABI 3100, Applied Biosystems, Foster City, CA) as well as the ABI Prism 310 Genetic Analyzer (ABI 310, Applied Biosystems, Foster City, CA) were evaluated for mtDNA sequencing capabilities and compared with sequencing results obtained on the platform currently in use in the FBI Laboratory (the ABI Prism 377 DNA Sequencer, ABI 377, Applied Biosystems, Foster City, CA). Various studies were performed to assess the utility of the ABI 3100, as well as the ABI 310 for mtDNA sequencing. The tests included: comparisons of results obtained among the ABI 3100, the ABI 310 and the ABI 377 instruments; comparisons of results obtained within and between capillary arrays; evaluation of capillary length; evaluation of sample injection time; evaluation of the resolution of mixtures/heteroplasmic samples; and evaluation of the sensitivity of detection of a minor component with reduced template on the ABI 3100. In addition, other studies were performed to improve sample preparation; these included: comparison of template suppression reagent (TSR, Applied Biosystems, Foster City, CA) versus formamide; the use of Performa DTR Gel Filtration Cartridges (Edge BioSystems Inc., Gaithersburg, MD) versus Centri-Sep Spin Columns (Princeton Separations, Adelphia, NJ) for product purification after cycle sequencing; and sample stability after denaturation. The data support that valid and reliable results can be obtained

  4. Matrilineal history of the endangered Cape Sable seaside sparrow inferred from mitochondrial DNA polymorphism.

    Science.gov (United States)

    Nelson, W S; Dean, T; Avise, J C

    2000-06-01

    Restriction analyses were conducted on mitochondrial DNA (mtDNA) amplified by long-PCR from an endangered bird, the Cape Sable seaside sparrow. The first of several successful mtDNA amplifications was accomplished using the partially digested tissue remains of a transmitter-monitored bird retrieved from the gut of a snake. As many as 91 mtDNA restriction fragments produced by 18 endonucleases were compared in this and four other Cape Sable specimens against mtDNA similarly amplified by long-PCR from other taxonomic forms in the seaside sparrow complex. Results indicate that the Cape Sable birds belong to an 'Atlantic' matrilineal clade, and are highly divergent from other seaside sparrows along the Gulf of Mexico.

  5. In vivo mutagenesis reveals that OriL is essential for mitochondrial DNA replication

    Science.gov (United States)

    Wanrooij, Sjoerd; Miralles Fusté, Javier; Stewart, James B; Wanrooij, Paulina H; Samuelsson, Tore; Larsson, Nils-Göran; Gustafsson, Claes M; Falkenberg, Maria

    2012-01-01

    The mechanisms of mitochondrial DNA replication have been hotly debated for a decade. The strand-displacement model states that lagging-strand DNA synthesis is initiated from the origin of light-strand DNA replication (OriL), whereas the strand-coupled model implies that OriL is dispensable. Mammalian mitochondria cannot be transfected and the requirements of OriL in vivo have therefore not been addressed. We here use in vivo saturation mutagenesis to demonstrate that OriL is essential for mtDNA maintenance in the mouse. Biochemical and bioinformatic analyses show that OriL is functionally conserved in vertebrates. Our findings strongly support the strand-displacement model for mtDNA replication. PMID:23090476

  6. Sequence analysis of three mitochondrial DNA molecules reveals interesting differences among Saccharomyces yeasts

    DEFF Research Database (Denmark)

    Langkjær, Rikke Breinhold; Casaregola, S.; Ussery, David;

    2003-01-01

    mtDNA, are not present. Surprisingly, four genes (ATP6, COX2, COX3 and COB) in the mtDNA of S. servazzii contain, in total, five + 1 frameshifts. mtDNAs of S. castellii, S. servazzii and S. cerevisiae contain all genes on the same strand, except for one tRNA gene. On the other hand, the gene order......The complete sequences of mitochondrial DNA ( mtDNA) from the two budding yeasts Saccharomyces castellii and Saccharomyces servazzii, consisting of 25 753 and 30 782 bp, respectively, were analysed and compared to Saccharomyces cerevisiae mtDNA. While some of the traits are very similar among...... Saccharomyces yeasts, others have highly diverged. The two mtDNAs are much more compact than that of S. cerevisiae and contain fewer introns and intergenic sequences, although they have almost the same coding potential. A few genes contain group I introns, but group II introns, otherwise found in S. cerevisiae...

  7. Use of sequence variation in three highly variable regions of the mitochondrial DNA for the discrimination of allogeneic platelets

    NARCIS (Netherlands)

    Warner, JB; Bruin, EJ; Hannig, H; Hellenkamp, F; Horning, A; Mittmann, K; van der Steege, G; de Leij, LFMH; Garritsen, HSP

    2006-01-01

    BACKGROUND: Human mitochondrial DNA (mtDNA) polymorphisms can be used to detect allogeneic transfused platelets. To increase the number of informative polymorphisms we investigated three hypervariable regions (HVR1, HVR2, and HVR3) within the displacement loop (D-loop) region of the mtDNA. STUDY DES

  8. mtDNA-Server: next-generation sequencing data analysis of human mitochondrial DNA in the cloud

    Science.gov (United States)

    Weissensteiner, Hansi; Forer, Lukas; Fuchsberger, Christian; Schöpf, Bernd; Kloss-Brandstätter, Anita; Specht, Günther; Kronenberg, Florian; Schönherr, Sebastian

    2016-01-01

    Next generation sequencing (NGS) allows investigating mitochondrial DNA (mtDNA) characteristics such as heteroplasmy (i.e. intra-individual sequence variation) to a higher level of detail. While several pipelines for analyzing heteroplasmies exist, issues in usability, accuracy of results and interpreting final data limit their usage. Here we present mtDNA-Server, a scalable web server for the analysis of mtDNA studies of any size with a special focus on usability as well as reliable identification and quantification of heteroplasmic variants. The mtDNA-Server workflow includes parallel read alignment, heteroplasmy detection, artefact or contamination identification, variant annotation as well as several quality control metrics, often neglected in current mtDNA NGS studies. All computational steps are parallelized with Hadoop MapReduce and executed graphically with Cloudgene. We validated the underlying heteroplasmy and contamination detection model by generating four artificial sample mix-ups on two different NGS devices. Our evaluation data shows that mtDNA-Server detects heteroplasmies and artificial recombinations down to the 1% level with perfect specificity and outperforms existing approaches regarding sensitivity. mtDNA-Server is currently able to analyze the 1000G Phase 3 data (n = 2,504) in less than 5 h and is freely accessible at https://mtdna-server.uibk.ac.at. PMID:27084948

  9. mtDNA-Server: next-generation sequencing data analysis of human mitochondrial DNA in the cloud.

    Science.gov (United States)

    Weissensteiner, Hansi; Forer, Lukas; Fuchsberger, Christian; Schöpf, Bernd; Kloss-Brandstätter, Anita; Specht, Günther; Kronenberg, Florian; Schönherr, Sebastian

    2016-07-08

    Next generation sequencing (NGS) allows investigating mitochondrial DNA (mtDNA) characteristics such as heteroplasmy (i.e. intra-individual sequence variation) to a higher level of detail. While several pipelines for analyzing heteroplasmies exist, issues in usability, accuracy of results and interpreting final data limit their usage. Here we present mtDNA-Server, a scalable web server for the analysis of mtDNA studies of any size with a special focus on usability as well as reliable identification and quantification of heteroplasmic variants. The mtDNA-Server workflow includes parallel read alignment, heteroplasmy detection, artefact or contamination identification, variant annotation as well as several quality control metrics, often neglected in current mtDNA NGS studies. All computational steps are parallelized with Hadoop MapReduce and executed graphically with Cloudgene. We validated the underlying heteroplasmy and contamination detection model by generating four artificial sample mix-ups on two different NGS devices. Our evaluation data shows that mtDNA-Server detects heteroplasmies and artificial recombinations down to the 1% level with perfect specificity and outperforms existing approaches regarding sensitivity. mtDNA-Server is currently able to analyze the 1000G Phase 3 data (n = 2,504) in less than 5 h and is freely accessible at https://mtdna-server.uibk.ac.at.

  10. The Three Genetics (Nuclear DNA, Mitochondrial DNA, and Gut Microbiome) of Longevity in Humans Considered as Metaorganisms

    Science.gov (United States)

    Candela, Marco; Brigidi, Patrizia; Luiselli, Donata; Bacalini, Maria Giulia; Salvioli, Stefano; Capri, Miriam; Collino, Sebastiano; Franceschi, Claudio

    2014-01-01

    Usually the genetics of human longevity is restricted to the nuclear genome (nDNA). However it is well known that the nDNA interacts with a physically and functionally separated genome, the mitochondrial DNA (mtDNA) that, even if limited in length and number of genes encoded, plays a major role in the ageing process. The complex interplay between nDNA/mtDNA and the environment is most likely involved in phenomena such as ageing and longevity. To this scenario we have to add another level of complexity represented by the microbiota, that is, the whole set of bacteria present in the different part of our body with their whole set of genes. In particular, several studies investigated the role of gut microbiota (GM) modifications in ageing and longevity and an age-related GM signature was found. In this view, human being must be considered as “metaorganism” and a more holistic approach is necessary to grasp the complex dynamics of the interaction between the environment and nDNA-mtDNA-GM of the host during ageing. In this review, the relationship between the three genetics and human longevity is addressed to point out that a comprehensive view will allow the researchers to properly address the complex interactions that occur during human lifespan. PMID:24868529

  11. Mitochondrial DNA copy number is maintained during spermatogenesis and in the development of male larvae to sustain the doubly uniparental inheritance of mitochondrial DNA system in the blue mussel Mytilus galloprovincialis.

    Science.gov (United States)

    Sano, Natsumi; Obata, Mayu; Ooie, Yosiyasu; Komaru, Akira

    2011-08-01

    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.

  12. Inhibition of the mitochondrial respiratory chain function abrogates quartz induced DNA damage in lung epithelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Li Hui [Institut fuer umweltmedizinische Forschung (IUF) at the Heinrich-Heine-University, Auf' m Hennekamp 50, D-40225 Duesseldorf (Germany); Haberzettl, Petra [Institut fuer umweltmedizinische Forschung (IUF) at the Heinrich-Heine-University, Auf' m Hennekamp 50, D-40225 Duesseldorf (Germany); Albrecht, Catrin [Institut fuer umweltmedizinische Forschung (IUF) at the Heinrich-Heine-University, Auf' m Hennekamp 50, D-40225 Duesseldorf (Germany); Hoehr, Doris [Institut fuer umweltmedizinische Forschung (IUF) at the Heinrich-Heine-University, Auf' m Hennekamp 50, D-40225 Duesseldorf (Germany); Knaapen, Ad M. [Department of Health Risk Analysis and Toxicology, Nutrition and Toxicology Research Institute Maastricht (NUTRIM), University of Maastricht (Netherlands); Borm, Paul J.A. [Institut fuer umweltmedizinische Forschung (IUF) at the Heinrich-Heine-University, Auf' m Hennekamp 50, D-40225 Duesseldorf (Germany); Hogeschool Zuyd Heerlen (Netherlands); Schins, Roel P.F. [Institut fuer umweltmedizinische Forschung (IUF) at the Heinrich-Heine-University, Auf' m Hennekamp 50, D-40225 Duesseldorf (Germany)]. E-mail: roel.schins@uni-duesseldorf.de

    2007-04-01

    Respirable quartz dust has been classified as a human carcinogen by the International Agency for Research on Cancer. The aim of our study was to investigate the mechanisms of DNA damage by DQ12 quartz in RLE-6TN rat lung epithelial type II cells (RLE). Transmission electron microscopy and flow-cytometry analysis showed a rapid particle uptake (30 min to 4 h) of quartz by the RLE cells, but particles were not found within the cell nuclei. This suggests that DNA strand breakage and induction of 8-hydroxydeoxyguanosine - as also observed in these cells during these treatment intervals - did not result from direct physical interactions between particles and DNA, or from short-lived particle surface-derived reactive oxygen species. DNA damage by quartz was significantly reduced in the presence of the mitochondrial inhibitors rotenone and antimycin-A. In the absence of quartz, these inhibitors did not affect DNA damage, but they reduced cellular oxygen consumption. No signs of apoptosis were observed by quartz. Flow-cytometry analysis indicated that the reduced DNA damage by rotenone was not due to a possible mitochondria-mediated reduction of particle uptake by the RLE cells. Further proof of concept for the role of mitochondria was shown by the failure of quartz to elicit DNA damage in mitochondria-depleted 143B (rho-0) osteosarcoma cells, at concentrations where it elicited DNA damage in the parental 143B cell line. In conclusion, our data show that respirable quartz particles can elicit oxidative DNA damage in vitro without entering the nuclei of type II cells, which are considered to be important target cells in quartz carcinogenesis. Furthermore, our observations indicate that such indirect DNA damage involves the mitochondrial electron transport chain function, by an as-yet-to-be elucidated mechanism.

  13. Horizontal transfer of DNA from the mitochondrial to the plastid genome and its subsequent evolution in milkweeds (Apocynaceae)

    Science.gov (United States)

    Shannon C.K. Straub; Richard C. Cronn; Christopher Edwards; Mark Fishbein; Aaron. Liston

    2013-01-01

    Horizontal gene transfer (HGT) of DNA from the plastid to the nuclear and mitochondrial genomes of higher plants is a common phenomenon; however, plastid genomes (plastomes) are highly conserved and have generally been regarded as impervious to HGT. We sequenced the 158 kb plastome and the 690 kb mitochondrial genome of common milkweed (Asclepias syriaca [Apocynaceae...

  14. Mitochondrial DNA heteroplasmy in the emerging field of massively parallel sequencing

    Science.gov (United States)

    Just, Rebecca S.; Irwin, Jodi A.; Parson, Walther

    2015-01-01

    Long an important and useful tool in forensic genetic investigations, mitochondrial DNA (mtDNA) typing continues to mature. Research in the last few years has demonstrated both that data from the entire molecule will have practical benefits in forensic DNA casework, and that massively parallel sequencing (MPS) methods will make full mitochondrial genome (mtGenome) sequencing of forensic specimens feasible and cost-effective. A spate of recent studies has employed these new technologies to assess intraindividual mtDNA variation. However, in several instances, contamination and other sources of mixed mtDNA data have been erroneously identified as heteroplasmy. Well vetted mtGenome datasets based on both Sanger and MPS sequences have found authentic point heteroplasmy in approximately 25% of individuals when minor component detection thresholds are in the range of 10–20%, along with positional distribution patterns in the coding region that differ from patterns of point heteroplasmy in the well-studied control region. A few recent studies that examined very low-level heteroplasmy are concordant with these observations when the data are examined at a common level of resolution. In this review we provide an overview of considerations related to the use of MPS technologies to detect mtDNA heteroplasmy. In addition, we examine published reports on point heteroplasmy to characterize features of the data that will assist in the evaluation of future mtGenome data developed by any typing method. PMID:26009256

  15. Complex forms of mitochondrial DNA in human B cells transformed by Epstein-Barr virus (EBV)

    DEFF Research Database (Denmark)

    Christiansen, Gunna; Christiansen, C; Zeuthen, J

    1983-01-01

    Human lymphocytes and lymphoid cell lines were analyzed for the presence of complex forms of mitochondrial DNA (mtDNA) by electron microscopy. A high frequency (9%-14.5%) of catenated dimers, circular dimers, or oligomers were found in samples from Epstein-Barr-virus-(EBV) transformed lymphoblast......Human lymphocytes and lymphoid cell lines were analyzed for the presence of complex forms of mitochondrial DNA (mtDNA) by electron microscopy. A high frequency (9%-14.5%) of catenated dimers, circular dimers, or oligomers were found in samples from Epstein-Barr-virus-(EBV) transformed...... lymphoblastoid cell lines. These complex forms of mtDNA were present in much lower frequencies in lymphocytes isolated from donor blood (1.3%-4.6%). Similar low frequencies were found with primary fibroblasts (1.1%) or freshly isolated monkey liver cells (2.1%). Samples from cultures of Burkitt lymphoma (BL......) cell lines of EBV-positive or -negative origin contained intermediate (5%-7%) frequencies of complex forms of mtDNA....

  16. Mitochondrial mosaics in the liver of 3 infants with mtDNA defects

    Directory of Open Access Journals (Sweden)

    Scalais Emmanuel

    2009-06-01

    Full Text Available Abstract Background In muscle cytochrome oxidase (COX negative fibers (mitochondrial mosaics have often been visualized. Methods COX activity staining of liver for light and electron microscopy, muscle stains, blue native gel electrophoresis and activity assays of respiratory chain proteins, their immunolocalisation, mitochondrial and nuclear DNA analysis. Results Three unrelated infants showed a mitochondrial mosaic in the liver after staining for COX activity, i.e. hepatocytes with strongly reactive mitochondria were found adjacent to cells with many negative, or barely reactive, mitochondria. Deficiency was most severe in the patient diagnosed with Pearson syndrome. Ragged-red fibers were absent in muscle biopsies of all patients. Enzyme biochemistry was not diagnostic in muscle, fibroblasts and lymphocytes. Blue native gel electrophoresis of liver tissue, but not of muscle, demonstrated a decreased activity of complex IV; in both muscle and liver subcomplexes of complex V were seen. Immunocytochemistry of complex IV confirmed the mosaic pattern in two livers, but not in fibroblasts. MRI of the brain revealed severe white matter cavitation in the Pearson case, but only slight cortical atrophy in the Alpers-Huttenlocher patient, and a normal image in the 3rd. MtDNA in leucocytes showed a common deletion in 50% of the mtDNA molecules of the Pearson patient. In the patient diagnosed with Alpers-Huttenlocher syndrome, mtDNA was depleted for 60% in muscle. In the 3rd patient muscular and hepatic mtDNA was depleted for more than 70%. Mutations in the nuclear encoded gene of POLG were subsequently found in both the 2nd and 3rd patients. Conclusion Histoenzymatic COX staining of a liver biopsy is fast and yields crucial data about the pathogenesis; it indicates whether mtDNA should be assayed. Each time a mitochondrial disorder is suspected and muscle data are non-diagnostic, a liver biopsy should be recommended. Mosaics are probably more frequent

  17. Defeating numts: semi-pure mitochondrial DNA from eggs and simple purification methods for field-collected wildlife tissues.

    Science.gov (United States)

    Ibarguchi, Gabriela; Friesen, Vicki L; Lougheed, Stephen C

    2006-11-01

    Mitochondrial DNA (mtDNA) continues to play a pivotal role in phylogeographic, phylogenetic, and population genetic studies. PCR amplification with mitochondrial primers often yields ambiguous sequences, in part because of the co-amplification of nuclear copies of mitochondrial genes (numts) and true mitochondrial heteroplasmy arising from mutations, hybridization with paternal leakage, gene duplications, and recombination. Failing to detect numts or to distinguish the origin of such homologous sequences results in the incorrect interpretation of data. However, few studies obtain purified mtDNA to confirm the mitochondrial origin of the first reference sequences for a species. Here, we demonstrate the importance and ease of obtaining semi-pure mtDNA from wildlife tissues, preserved under various typical field conditions, and investigate the success of 3 commercial extraction kits, cesium-chloride gradient mtDNA purification, long-template PCR amplification, cloning, and more species-specific degenerate primers. Using more detailed avian examples, we illustrate that unfertilized or undeveloped eggs provide the purest sources of mtDNA; that kits provide an alternative to cesium-chloride gradient methods; and that long-template PCR, cloning, and degenerate primers cannot be used to produce reliable mitochondrial reference sequences, but can be powerful tools when used in conjunction with purified mtDNA stocks to distinguish numts from true heteroplasmy.

  18. Positioning the red deer (Cervus elaphus hunted by the Tyrolean Iceman into a mitochondrial DNA phylogeny.

    Directory of Open Access Journals (Sweden)

    Cristina Olivieri

    Full Text Available In the last years several phylogeographic studies of both extant and extinct red deer populations have been conducted. Three distinct mitochondrial lineages (western, eastern and North-African/Sardinian have been identified reflecting different glacial refugia and postglacial recolonisation processes. However, little is known about the genetics of the Alpine populations and no mitochondrial DNA sequences from Alpine archaeological specimens are available. Here we provide the first mitochondrial sequences of an Alpine Copper Age Cervus elaphus. DNA was extracted from hair shafts which were part of the remains of the clothes of the glacier mummy known as the Tyrolean Iceman or Ötzi (5,350-5,100 years before present. A 2,297 base pairs long fragment was sequenced using a mixed sequencing procedure based on PCR amplifications and 454 sequencing of pooled amplification products. We analyzed the phylogenetic relationships of the Alpine Copper Age red deer's haplotype with haplotypes of modern and ancient European red deer. The phylogenetic analyses showed that the haplotype of the Alpine Copper Age red deer falls within the western European mitochondrial lineage in contrast with the current populations from the Italian Alps belonging to the eastern lineage. We also discussed the phylogenetic relationships of the Alpine Copper Age red deer with the populations from Mesola Wood (northern Italy and Sardinia.

  19. Origin and history of mitochondrial DNA lineages in domestic horses.

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    Cieslak, Michael; Pruvost, Melanie; Benecke, Norbert; Hofreiter, Michael; Morales, Arturo; Reissmann, Monika; Ludwig, Arne

    2010-12-20

    Domestic horses represent a genetic paradox: although they have the greatest number of maternal lineages (mtDNA) of all domestic species, their paternal lineages are extremely homogeneous on the Y-chromosome. In order to address their huge mtDNA variation and the origin and history of maternal lineages in domestic horses, we analyzed 1961 partial d-loop sequences from 207 ancient remains and 1754 modern horses. The sample set ranged from Alaska and North East Siberia to the Iberian Peninsula and from the Late Pleistocene to modern times. We found a panmictic Late Pleistocene horse population ranging from Alaska to the Pyrenees. Later, during the Early Holocene and the Copper Age, more or less separated sub-populations are indicated for the Eurasian steppe region and Iberia. Our data suggest multiple domestications and introgressions of females especially during the Iron Age. Although all Eurasian regions contributed to the genetic pedigree of modern breeds, most haplotypes had their roots in Eastern Europe and Siberia. We found 87 ancient haplotypes (Pleistocene to Mediaeval Times); 56 of these haplotypes were also observed in domestic horses, although thus far only 39 haplotypes have been confirmed to survive in modern breeds. Thus, at least seventeen haplotypes of early domestic horses have become extinct during the last 5,500 years. It is concluded that the large diversity of mtDNA lineages is not a product of animal breeding but, in fact, represents ancestral variability.

  20. Origin and history of mitochondrial DNA lineages in domestic horses.

    Directory of Open Access Journals (Sweden)

    Michael Cieslak

    Full Text Available Domestic horses represent a genetic paradox: although they have the greatest number of maternal lineages (mtDNA of all domestic species, their paternal lineages are extremely homogeneous on the Y-chromosome. In order to address their huge mtDNA variation and the origin and history of maternal lineages in domestic horses, we analyzed 1961 partial d-loop sequences from 207 ancient remains and 1754 modern horses. The sample set ranged from Alaska and North East Siberia to the Iberian Peninsula and from the Late Pleistocene to modern times. We found a panmictic Late Pleistocene horse population ranging from Alaska to the Pyrenees. Later, during the Early Holocene and the Copper Age, more or less separated sub-populations are indicated for the Eurasian steppe region and Iberia. Our data suggest multiple domestications and introgressions of females especially during the Iron Age. Although all Eurasian regions contributed to the genetic pedigree of modern breeds, most haplotypes had their roots in Eastern Europe and Siberia. We found 87 ancient haplotypes (Pleistocene to Mediaeval Times; 56 of these haplotypes were also observed in domestic horses, although thus far only 39 haplotypes have been confirmed to survive in modern breeds. Thus, at least seventeen haplotypes of early domestic horses have become extinct during the last 5,500 years. It is concluded that the large diversity of mtDNA lineages is not a product of animal breeding but, in fact, represents ancestral variability.

  1. Study on Mitochondrial DNA Cytochrome b Gene of Chinese sea bass,%Study on Mitochondrial DNA Cytochrome b Gene of Chinese sea bass, Lateolabrax sp.

    Institute of Scientific and Technical Information of China (English)

    高天翔; 张秀梅; 陈大刚; 张美昭; 任一平; 张亚平

    2001-01-01

    参考鳗鲡等鱼类线粒体DNA序列进行了中国花鲈线粒体DNA细胞色素b基因片断的引物设计、PCR扩增及其序列测定。得到中国花鲈的碱基序列为410bp,其A、T、G、C含量分别为101bp(24.63%)、112bp(27.32%)、72bp(17.56%)、125bp(30.49%),与鳗鲡等其他鱼类相同基因片断序列碱基含量相似。%The primers of mitochondrial DNA cytochrome b gene of C hinesesea bass were designed by referencing the sequences of Anguilla speci es. We successfully amplified and sequenced the 410 base pairs that encode the c ytochrome b gene of mitochondrial DNA. The A, T, G and C contents of the sequenc e were 101bp (24.63%), 112bp (27.32%), 72bp (17.56%) and 125bp (30 .49%) respectively, similar to the sequences of other species.

  2. Influence of seasonal migration on geographic distribution of mitochondrial DNA haplotypes in humpback whales.

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    Baker, C S; Palumbi, S R; Lambertsen, R H; Weinrich, M T; Calambokidis, J; O'Brien, S J

    1990-03-15

    Humpback whales (Megaptera novaeangliae) migrate nearly 10,000 km each year between summer feeding grounds in temperate or near-polar waters and winter breeding grounds in shallow tropical waters. Observations of marked individuals suggest that major oceanic populations of humpback whales are divided into a number of distinct seasonal subpopulations which are not separated by obvious geographic barriers. To test whether these observed patterns of distribution and migration are reflected in the genetic structure of populations, we looked for variation in the mitochondrial DNA of 84 individual humpback whales on different feeding and wintering grounds of the North Pacific and western North Atlantic oceans. On the basis of restriction-fragment analysis, we now report a marked segregation of mitochondrial DNA haplotypes among subpopulations as well as between the two oceans. We interpret this segregation to be the consequence of maternally directed fidelity to migratory destinations.

  3. Parental genetic effects in a cavefish adaptive behavior explain disparity between nuclear and mitochondrial DNA.

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    Yoshizawa, Masato; Ashida, Go; Jeffery, William R

    2012-09-01

    Epigenetic parental genetic effects are important in many biological processes but their roles in the evolution of adaptive traits and their consequences in naturally evolving populations remain to be addressed. By comparing two divergent blind cave-dwelling cavefish populations with a sighted surface-dwelling population (surface fish) of the teleost Astyanax mexicanus, we report here that convergences in vibration attraction behavior (VAB), the lateral line sensory receptors underlying this behavior, and the feeding benefits of this behavior are controlled by parental genetic effects, either maternal or paternal inheritance. From behavioral studies and mathematical evolutionary simulations, we further demonstrate that disparity in nuclear and mitochondrial DNA in one of these cavefish populations that has hybridized with surface fish can be explained by paternal inheritance of VAB. The results suggest that parental genetic effects in adaptive behaviors may be important factors in biasing mitochondrial DNA inheritance in natural populations that are subject to introgression.

  4. Modulation of Mitochondrial DNA Copy Number to Induce Hepatocytic Differentiation of Human Amniotic Epithelial Cells.

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    Vaghjiani, Vijesh; Cain, Jason E; Lee, William; Vaithilingam, Vijayaganapathy; Tuch, Bernard E; St John, Justin C

    2017-09-05

    Mitochondrial deoxyribonucleic acid (mtDNA) copy number is tightly regulated during pluripotency and differentiation. There is increased demand of cellular adenosine triphosphate (ATP) during differentiation for energy-intensive cell types such as hepatocytes and neurons to meet the cell's functional requirements. During hepatocyte differentiation, mtDNA copy number should be synchronously increased to generate sufficient ATP through oxidative phosphorylation. Unlike bone marrow mesenchymal cells, mtDNA copy number failed to increase by 28 days of differentiation of human amniotic epithelial cells (hAEC) into hepatocyte-like cells (HLC) despite their expression of some end-stage hepatic markers. This was due to higher levels of DNA methylation at exon 2 of POLGA, the mtDNA-specific replication factor. Treatment with a DNA demethylation agent, 5-azacytidine, resulted in increased mtDNA copy number, reduced DNA methylation at exon 2 of POLGA, and reduced hepatic gene expression. Depletion of mtDNA followed by subsequent differentiation did not increase mtDNA copy number, but reduced DNA methylation at exon 2 of POLGA and increased expression of hepatic and pluripotency genes. We encapsulated hAEC in barium alginate microcapsules and subsequently differentiated them into HLC. Encapsulation resulted in no net increase of mtDNA copy number but a significant reduction in DNA methylation of POLGA. RNAseq analysis showed that differentiated HLC express hepatocyte-specific genes but also increased expression of inflammatory interferon genes. Differentiation in encapsulated cells showed suppression of inflammatory genes as well as increased expression of genes associated with hepatocyte function pathways and networks. This study demonstrates that an increase in classical hepatic gene expression can be achieved in HLC through encapsulation, although they fail to effectively regulate mtDNA copy number.

  5. DNA primers for amplification of mitochondrial cytochrome c oxidase subunit I from diverse metazoan invertebrates.

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    Folmer, O; Black, M; Hoeh, W; Lutz, R; Vrijenhoek, R

    1994-10-01

    We describe "universal" DNA primers for polymerase chain reaction (PCR) amplification of a 710-bp fragment of the mitochondrial cytochrome c oxidase subunit I gene (COI) from 11 invertebrate phyla: Echinodermata, Mollusca, Annelida, Pogonophora, Arthropoda, Nemertinea, Echiura, Sipuncula, Platyhelminthes, Tardigrada, and Coelenterata, as well as the putative phylum Vestimentifera. Preliminary comparisons revealed that these COI primers generate informative sequences for phylogenetic analyses at the species and higher taxonomic levels.

  6. Mitochondrial DNA damage and oxidative damage in HL-60 cells exposed to 900MHz radiofrequency fields.

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    Sun, Yulong; Zong, Lin; Gao, Zhen; Zhu, Shunxing; Tong, Jian; Cao, Yi

    2017-03-01

    HL-60 cells, derived from human promyelocytic leukemia, were exposed to continuous wave 900MHz radiofrequency fields (RF) at 120μW/cm(2) power intensity for 4h/day for 5 consecutive days to examine whether such exposure is capable damaging the mitochondrial DNA (mtDNA) mediated through the production of reactive oxygen species (ROS). In addition, the effect of RF exposure was examined on 8-hydroxy-2'-dexoyguanosine (8-OHdG) which is a biomarker for oxidative damage and on the mitochondrial synthesis of adenosine triphosphate (ATP) which is the energy required for cellular functions. The results indicated a significant increase in ROS and significant decreases in mitochondrial transcription factor A, mtDNA polymerase gamma, mtDNA transcripts and mtDNA copy number in RF-exposed cells compared with those in sham-exposed control cells. In addition, there was a significant increase in 8-OHdG and a significant decrease in ATP in RF-exposed cells. The response in positive control cells exposed to gamma radiation (GR, which is also known to induce ROS) was similar to those in RF-exposed cells. Thus, the overall data indicated that RF exposure was capable of inducing mtDNA damage mediated through ROS pathway which also induced oxidative damage. Prior-treatment of RF- and GR-exposed the cells with melatonin, a well-known free radical scavenger, reversed the effects observed in RF-exposed cells. Copyright © 2017 Elsevier B.V. All rights reserved.

  7. Mitochondrial DNA 4977-base pair common deletion in blood leukocytes and melanoma risk.

    Science.gov (United States)

    Shen, Jie; Wan, Jie; Huff, Chad; Fang, Shenying; Lee, Jeffrey E; Zhao, Hua

    2016-05-01

    The 4977-base pair common deletion DmtDNA4977 is the most frequently observed mitochondrial DNA mutation in human tissues. Because mitochondrial DNA mutations are mainly caused by reactive oxygen species (ROS), and given that oxidative stress plays an important role in melanoma carcinogenesis, the investigation of DmtDNA4977 may be particularly relevant to the development of melanoma. In this study, we compared DmtDNA4977 levels in blood leukocytes from 206 melanoma patients and 219 healthy controls. Overall, melanoma cases had significantly higher levels of DmtDNA4977 than healthy controls (median: 0.60 vs 0.20, P = 0.008). The difference was evident among individuals who were older than 47 yrs, women, and had pigmentation risk factors (e.g., blond or red hair, blue eye, fair skin, light, or none tanning ability after prolonged sun exposure, and freckling in the sun as a child). The difference was also evident among those who had at least one lifetime sunburn with blistering and had no reported use of a sunlamp. Interestingly, among controls, DmtDNA4977 levels differed by phenotypic index and reported use of a sunlamp. In the risk assessment, increased levels of DmtDNA4977 were associated with a 1.23-fold increased risk of melanoma (odds ratio (OR): 1.23, 95% confidence interval (90% CI): 1.01, 1.50). A significant dose-response relationship was observed in quartile analysis (P = 0.001). In summary, our study suggests that high levels of DmtDNA4977 in blood leukocytes are associated with increased risk of melanoma and that association is affected by both pigmentation and personal history of sun exposure.

  8. Mitochondrial DNA deletion percentage in sun exposed and non sun exposed skin.

    Science.gov (United States)

    Powers, Julia M; Murphy, Gillian; Ralph, Nikki; O'Gorman, Susan M; Murphy, James E J

    2016-12-01

    The percentages of mitochondrial genomes carrying the mtDNA(3895) and the mtDNA(4977) (common) deletion were quantified in sun exposed and non sun exposed skin biopsies, for five cohorts of patients varying either in sun exposure profile, age or skin cancer status. Non-melanoma skin cancer diagnoses are rising in Ireland and worldwide [12] but most risk prediction is based on subjective visual estimations of sun exposure history. A quantitative objective test for pre-neoplastic markers may result in better adherence to sun protective behaviours. Mitochondrial DNA (mtDNA) is known to be subject to the loss of a significant proportion of specific sections of genetic code due to exposure to ultraviolet light in sunlight. Although one such deletion has been deemed more sensitive, another, called the mtDNA(4977) or common deletion, has proved to be a more useful indicator of possible risk in this study. Quantitative molecular analysis was carried out to determine the percentage of genomes carrying the deletion using non sun exposed and sun exposed skin biopsies in cohorts of patients with high or low sun exposure profiles and two high exposure groups undergoing treatment for NMSC. Results indicate that mtDNA deletions correlate to sun exposure; in groups with high sun exposure habits a significant increase in deletion number in exposed over non sun exposed skin occurred. An increase in deletion percentage was also seen in older cohorts compared to the younger group. The mtDNA(3895) deletion was detected in small amounts in exposed skin of many patients, the mtDNA(4977) common deletion, although present to some extent in non sun exposed skin, is suggested to be the more reliable and easily detected marker. In all cohorts except the younger group with relatively lower sun exposure, the mtDNA(4977) deletion was more frequent in sun exposed skin samples compared to non-sun exposed skin.

  9. Dynamical change of mitochondrial DNA induced in the living cell by perturbing the electrochemical gradient.

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    Coppey-Moisan, M; Brunet, A C; Morais, R; Coppey, J

    1996-11-01

    Digital-imaging microscopy was used in conditions that allowed the native state to be preserved and hence fluorescence variations of specific probes to be followed in the real time of living mammalian cells. Ethidium bromide was shown to enter into living cells and to intercalate stably into mitochondrial DNA (mtDNA), giving rise to high fluorescence. When the membrane potential or the pH gradient across the inner membrane was abolished by specific inhibitors or ionophores, the ethidium fluorescence disappeared from all mtDNA molecules within 2 min. After removal of the inhibitors or ionophores, ethidium fluorescence rapidly reappeared in mitochondria, together with the membrane potential. The fluorescence extinction did not result from an equilibrium shift caused by leakage of free ethidium out of mitochondria when the membrane potential was abolished but was most likely due to a dynamical mtDNA change that exposed intercalated ethidium to quencher, either by weakening the ethidium binding constant or by giving access of a proton acceptor (such as water) to the interior of mtDNA. Double labeling with ethidium and with a minor groove probe (4',6-diamino-2-phenylindole) indicated that mtDNA maintains a double-stranded structure. The two double-stranded DNA states, revealed by the fluorescence of mitochondrial ethidium, enhanced or quenched in the presence of ethidium, seem to coexist in mitochondria of unperturbed fibroblast cells, suggesting a spontaneous dynamical change of mtDNA molecules. Therefore, the ethidium fluorescence variation allows changes of DNA to be followed, a property that has to be taken into consideration when using this intercalator for in vivo as well as in vitro imaging studies.

  10. Mitochondrial DNA heteroplasmy in ovine fetuses and sheep cloned by somatic cell nuclear transfer

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    Müller Mathias

    2007-12-01

    Full Text Available Abstract Background The mitochondrial DNA (mtDNA of the cloned sheep "Dolly" and nine other ovine clones produced by somatic cell nuclear transfer (SCNT was reported to consist only of recipient oocyte mtDNA without any detectable mtDNA contribution from the nucleus donor cell. In cattle, mouse and pig several or most of the clones showed transmission of nuclear donor mtDNA resulting in mitochondrial heteroplasmy. To clarify the discrepant transmission pattern of donor mtDNA in sheep clones we analysed the mtDNA composition of seven fetuses and five lambs cloned from fetal fibroblasts. Results The three fetal fibroblast donor cells used for SCNT harboured low mtDNA copy numbers per cell (A: 753 ± 54, B: 292 ± 33 and C: 561 ± 88. The ratio of donor to recipient oocyte mtDNAs was determined using a quantitative amplification refractory mutation system (ARMS PCR (i.e. ARMS-qPCR. For quantification of SNP variants with frequencies below 0.1% we developed a restriction endonuclease-mediated selective quantitative PCR (REMS-qPCR. We report the first cases (n = 4 fetuses, n = 3 lambs of recipient oocyte/nuclear donor mtDNA heteroplasmy in SCNT-derived ovine clones demonstrating that there is no species-effect hindering ovine nucleus-donor mtDNA from being transmitted to the somatic clonal offspring. Most of the heteroplasmic clones exhibited low-level heteroplasmy (0.1% to 0.9%, n = 6 indicating neutral transmission of parental mtDNAs. High-level heteroplasmy (6.8% to 46.5% was observed in one case. This clone possessed a divergent recipient oocyte-derived mtDNA genotype with three rare amino acid changes compared to the donor including one substitution at an evolutionary conserved site. Conclusion Our study using state-of-the-art techniques for mtDNA quantification, like ARMS-qPCR and the novel REMS-qPCR, documents for the first time the transmission of donor mtDNA into somatic sheep clones. Mt