Search

We studied nine infant patients with a combination of progressive neurological and hepatic failure. Eight children, including two sibling pairs and four singletons, were affected by Alpers' hepatopathic poliodystrophy. A ninth baby patient suffered of a severe floppy infant syndrome associated with liver failure. Analysis of POLG1, the gene encoding the catalytic subunit of mitochondrial DNA polymerase, revealed that all the patients carried different allelic mutations in this gene. POLG1 is a major disease gene in mitochondrial disorders. Mutations in this gene can be associated with multiple deletions, depletion or point mutations of mitochondrial DNA (mtDNA). In turn, these different molecular phenotypes dictate an extremely heterogeneous spectrum of clinical outcomes, ranging from adult-onset progressive ophthalmoplegia to juvenile ataxic syndromes with epilepsy, to rapidly fatal hepatocerebral presentations, including Alpers' syndrome.

Using the Mpv17-negative mouse strain, which developed inner ear and kidney dysfunction, we confirm a strong relationship between the kidney and the inner ear. Both organs have specialized epithelia involved in active ion transport, which are separated from the vessels by a basement membrane of similar composition. Our recent results indicate that the glomerular and the stria vascularis basement membrane are simultaneously affected in early stages. Concomitant deposits of IgG during the progressive development of the disease support the idea of a shared antigen. Understanding the pattern of the development of the degeneration will provide a basis towards understanding the essential role of the Mpv17 protein in the structures of both organs and may provide a basis for future therapeutic intervention.

SUMMARY: Hepatocerebral MPV17 -MDS is quite rare (

Multiple deletions in mitochondrial DNA (mtDNA) are associated with mutations in nuclear genes which encode proteins either directly or indirectly involved in the replication or maintenance of mtDNA. To date, mutations in twelve nuclear genes, POLG, POLG2, C10orf2, SLC25A4, RRM2B, TK2, MPV17, DGUOK, OPA1, MFN2, MGME1, and DNA2 have been identified in patients with multiple deletions in mtDNA, yet the genetic cause of multiple mtDNA deletions still remains undefined in a large number of patients. In addition to patients with a classic mitochondrial myopathy, multiple mtDNA deletions are also found in patients with myopathies such as sporadic IBM. Mitochondrial changes have also been identified in skeletal muscle from patients with myofibrillar myopathy, with histochemical changes such as rubbed out fibres observed with COX and NADH staining, and focal clustering and depletion of mitochondria. Mitochondrial pathology, including COX negative and ragged red fibres, and paracrystalline inclusions have also been described in patients with centronuclear myopathy due to mutations in DNM2, which encodes Dynamin 2, a GTPase which is involved in membrane trafficking. Here we describe our findings in two patients with multiple mtDNA deletions. In one patient, a 70 year old male who presented with a history of exercise induced muscle pain and distal myopathy we identified a novel pArg2364His mutation in the gene FLNC which encodes the actin binding protein filamin-C. In the second patient, a 40 year old male who presented at age 3 years with ataxia and later developed facial myopathy, proximal muscle weakness, and CPEO, histochemical investigation indicated centronuclear myopathy, and a p.Glu560Lys mutation in DNM2 was identified. Our findings suggest that FLNC and DNM2 should be added to the list of genes associated with multiple deletions in mtDNA.

s, 7 Annual UK Neuromuscular Translational Research Conference, 2014 /Neuromuscular Disorders 24S1 (2014) S7–S27 S15 to the age-dependent manifestation or spontaneous recovery of infantile reversible COX deficiency myopathy. We performed immunohistochemistry and immunoblotting with antibodies against COXVI and COXVII isoforms in mice, in human skeletal muscle of controls and patients in different ages (0–6 months, >1 year, adults) and in human muscle cells. Both in mice and humans, the liver type isoforms gradually decreased, and the heart/muscle-type isoforms increased through development in the first weeks of life, confirming an age-dependent isoform switch. In skeletal muscle of a patient with reversible COX deficiency myopathy we proved the existence of the COXVI and COXVII isoform switch, although we could not confirm an association with the clinical recovery. However, understanding developmental changes of the COX isoforms may have implications for other mitochondrial diseases. P31 Genotypic and phenotypic heterogeneity in adult-onset progressive external ophthalmoplegia (PEO) with mitochondrial DNA instability: a systematic review E.W. Sommerville, P.F. Chinnery, G.S. Gorman, R.W. Taylor. Wellcome Trust Centre for Mitochondrial Research, Cookson Building, Framlington Place, Newcastle University, Newcastle upon Tyne, Tyne and Wear, NE2 4HH, UK; Institute of Genetic Medicine, International Centre for Life, Newcastle University, Central Parkway, Newcastle upon Tyne NE1 3BZ, UK Background: Progressive external ophthalmoplegia (PEO) is an eye movement disorder characterised by extraocular muscle paresis and muscle restricted mitochondrial DNA (mtDNA) deletions. Patient classification is difficult due to overlapping clinical phenotypes and poor genotype-phenotype correlates. This is compounded by the fact that approximately half of PEO patients do not have a genetic diagnosis. Aims: To review the phenotypic and genotypic manifestations of adult-onset PEO and to identify possible novel candidate genes. Patients: Patients were identified in the literature using electronic searches from Scopus, Medline via PubMed and Genetics Abstracts databases (1 January 1970 to 8 November 2013). Adult patients presenting PEO (≥16 years) presenting with PEO and mtDNA instability and with a confirmed genetic diagnosis were selected. The criterion was extended when searching candidate novel genes. Results: We identified 575 PEO patients, harbouring 12 known nuclear encoded genes (TYMP, SLC25A4, POLG, C10ORF2, OPA1, POLG2, RRM2B, TK2, DGUOK , MPV17, MGME1, and DNA2). Additional novel candidate genes (twenty in total), including several encoding proteins not predicted to localise to mitochondria, were also identified. Conclusion: We propose to use the findings of this systematic review coupled to whole exome and targeted next-generation sequencing technology, to help direct the investigation of a large cohort of clinically well-defined, genetically undetermined adult patients with PEO and mtDNA instability. P32 Mutations in SPG7 cause chronic progressive external ophthalmoplegia through disordered mtDNA maintenance G.S. Gorman, G. Pfeffer, H. Griffin, M. Kurzawa-Akanbi, E.L. Blakely, I. Wilson, K. Sitarz, D. Moore, J.L. Murphy, C.L. Alston, A. Pyle, J. Coxhead, B. Payne, G.H. Gorrie, C. Longman, M. Hadjivassiliou, J. McConville, D. Dick, I. Imam, D. Hilton, F. Norwood, M.R. Baker, S.R. Jaiser, P. Yu-Wai-Man, M. Farrell, A. McCarthy, T. Lynch, R. McFarland, A.M. Schaefer, D.M. Turnbull, R. Horvath, R.W. Taylor, P.F. Chinnery. Wellcome Centre for Mitochondrial Research, Newcastle University, Newcastle upon Tyne, NE2 4HH, UK Background: Despite being a canonical presenting feature of mitochondrial (mt) disease, the genetic basis of progressive external ophthalmoplegia (PEO) remains unknown in a large proportion of patients. Aims: To identify the causative gene in patients with genetically undetermined mtDNA maintenance disorders. Methods: Whole exome sequencing, targeted Sanger sequencing and MLPA analysis were used to study 68 adult patients with PEO either with or without multiple mtDNA deletions in skeletal muscle. Functional studies included transcript analysis, proteomics, mitochondrial network analysis, single fibre mtDNA analysis and deep re-sequencing of mtDNA. Results: Nine patients (eight probands) were found to carry compound heterozygous SPG7 mutations, including three novel mutations: c.2221G>A; p.(Glu741Lys), c.2224G>A; p.(Asp742Asn), and 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 PEO/ptosis and spastic ataxia, or a progressive ataxic disorder. Functional studies revealed increased mitochondrial biogenesis in patient muscle, and mitochondrial fusion in patient fibroblasts associated with the clonal expansion of mtDNA mutations. Conclusion: The SPG7 gene should be screened in patients in whom a disorder of mtDNA maintenance is suspected when spastic ataxia is prominent. The complex neurological phenotype is likely due to the clonal expansion of secondary mtDNA mutations modulating the phenotype, driven by compensatory mitochondrial biogenesis. a Joint first authors. P33 Do modulators of mitophagy select pathogenic mtDNA mutations? A. Hinks-Roberts, E. Dombi, A. Diot, C. Liao, K. Morten, J. Carver, T. Lodge, H. Mortiboys, J. Poulton. Nuffield Department of Obstetrics and Gynaecology, University of Oxford, UK; Sheffield Institute for Translational Neuroscience, University of Sheffield, UK Mitochondrial diseases that result from maternally transmitted mitochondrial DNA (mtDNA) mutations occur in 1/400 individuals. In heteroplasmic diseases, the balance between co-existing mutant and wild type mtDNA usually underlies disease progression. Cellular mechanisms for maintaining mitochondrial quality include mitophagy, and this could be a critical determinant of disease severity. Previous investigators showed that mitophagy was increased by the drug phenanthroline, a metallopeptidase inhibitor. We reasoned that activating mitophagy with phenanthroline might potentially reduce the load of pathogenic mutant mtDNA in tissue culture cells. We used a previously developed high throughput imaging for quantifying mitophagy in cultured primary fibroblasts bearing the common pathogenic A3243G mtDNA mutation, associated with the mitochondrial encephalomyopathy, lactic acidosis, and strokelike episode syndrome (MELAS) and with diabetes mellitus and deafness. We showed that phenanthroline significantly increased mitophagy in fibroblasts and reduced both the mitochondrial volume and mtDNA content. We conclude that phenanthroline activates mitophagy. However, there was little evidence that it reduced the load of mutant mtDNA. This suggests that both wild type and mutant mtDNA are turned over during mitophagy activated by phenanthroline. We conclude that phenanthroline is a poorly selective activator of mitophagy. Modulators of mitophagy need to target mitochondria enriched for mutant mtDNA if they are to benefit patients with heteroplasmic mtDNA disease.

s, 7 Annual UK Neuromuscular Translational Research Conference, 2014 /Neuromuscular Disorders 24S1 (2014) S7–S27 S15 to the age-dependent manifestation or spontaneous recovery of infantile reversible COX deficiency myopathy. We performed immunohistochemistry and immunoblotting with antibodies against COXVI and COXVII isoforms in mice, in human skeletal muscle of controls and patients in different ages (0–6 months, >1 year, adults) and in human muscle cells. Both in mice and humans, the liver type isoforms gradually decreased, and the heart/muscle-type isoforms increased through development in the first weeks of life, confirming an age-dependent isoform switch. In skeletal muscle of a patient with reversible COX deficiency myopathy we proved the existence of the COXVI and COXVII isoform switch, although we could not confirm an association with the clinical recovery. However, understanding developmental changes of the COX isoforms may have implications for other mitochondrial diseases. P31 Genotypic and phenotypic heterogeneity in adult-onset progressive external ophthalmoplegia (PEO) with mitochondrial DNA instability: a systematic review E.W. Sommerville, P.F. Chinnery, G.S. Gorman, R.W. Taylor. Wellcome Trust Centre for Mitochondrial Research, Cookson Building, Framlington Place, Newcastle University, Newcastle upon Tyne, Tyne and Wear, NE2 4HH, UK; Institute of Genetic Medicine, International Centre for Life, Newcastle University, Central Parkway, Newcastle upon Tyne NE1 3BZ, UK Background: Progressive external ophthalmoplegia (PEO) is an eye movement disorder characterised by extraocular muscle paresis and muscle restricted mitochondrial DNA (mtDNA) deletions. Patient classification is difficult due to overlapping clinical phenotypes and poor genotype-phenotype correlates. This is compounded by the fact that approximately half of PEO patients do not have a genetic diagnosis. Aims: To review the phenotypic and genotypic manifestations of adult-onset PEO and to identify possible novel candidate genes. Patients: Patients were identified in the literature using electronic searches from Scopus, Medline via PubMed and Genetics Abstracts databases (1 January 1970 to 8 November 2013). Adult patients presenting PEO (≥16 years) presenting with PEO and mtDNA instability and with a confirmed genetic diagnosis were selected. The criterion was extended when searching candidate novel genes. Results: We identified 575 PEO patients, harbouring 12 known nuclear encoded genes (TYMP, SLC25A4, POLG, C10ORF2, OPA1, POLG2, RRM2B, TK2, DGUOK , MPV17, MGME1, and DNA2). Additional novel candidate genes (twenty in total), including several encoding proteins not predicted to localise to mitochondria, were also identified. Conclusion: We propose to use the findings of this systematic review coupled to whole exome and targeted next-generation sequencing technology, to help direct the investigation of a large cohort of clinically well-defined, genetically undetermined adult patients with PEO and mtDNA instability. P32 Mutations in SPG7 cause chronic progressive external ophthalmoplegia through disordered mtDNA maintenance G.S. Gorman, G. Pfeffer, H. Griffin, M. Kurzawa-Akanbi, E.L. Blakely, I. Wilson, K. Sitarz, D. Moore, J.L. Murphy, C.L. Alston, A. Pyle, J. Coxhead, B. Payne, G.H. Gorrie, C. Longman, M. Hadjivassiliou, J. McConville, D. Dick, I. Imam, D. Hilton, F. Norwood, M.R. Baker, S.R. Jaiser, P. Yu-Wai-Man, M. Farrell, A. McCarthy, T. Lynch, R. McFarland, A.M. Schaefer, D.M. Turnbull, R. Horvath, R.W. Taylor, P.F. Chinnery. Wellcome Centre for Mitochondrial Research, Newcastle University, Newcastle upon Tyne, NE2 4HH, UK Background: Despite being a canonical presenting feature of mitochondrial (mt) disease, the genetic basis of progressive external ophthalmoplegia (PEO) remains unknown in a large proportion of patients. Aims: To identify the causative gene in patients with genetically undetermined mtDNA maintenance disorders. Methods: Whole exome sequencing, targeted Sanger sequencing and MLPA analysis were used to study 68 adult patients with PEO either with or without multiple mtDNA deletions in skeletal muscle. Functional studies included transcript analysis, proteomics, mitochondrial network analysis, single fibre mtDNA analysis and deep re-sequencing of mtDNA. Results: Nine patients (eight probands) were found to carry compound heterozygous SPG7 mutations, including three novel mutations: c.2221G>A; p.(Glu741Lys), c.2224G>A; p.(Asp742Asn), and 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 PEO/ptosis and spastic ataxia, or a progressive ataxic disorder. Functional studies revealed increased mitochondrial biogenesis in patient muscle, and mitochondrial fusion in patient fibroblasts associated with the clonal expansion of mtDNA mutations. Conclusion: The SPG7 gene should be screened in patients in whom a disorder of mtDNA maintenance is suspected when spastic ataxia is prominent. The complex neurological phenotype is likely due to the clonal expansion of secondary mtDNA mutations modulating the phenotype, driven by compensatory mitochondrial biogenesis. a Joint first authors. P33 Do modulators of mitophagy select pathogenic mtDNA mutations? A. Hinks-Roberts, E. Dombi, A. Diot, C. Liao, K. Morten, J. Carver, T. Lodge, H. Mortiboys, J. Poulton. Nuffield Department of Obstetrics and Gynaecology, University of Oxford, UK; Sheffield Institute for Translational Neuroscience, University of Sheffield, UK Mitochondrial diseases that result from maternally transmitted mitochondrial DNA (mtDNA) mutations occur in 1/400 individuals. In heteroplasmic diseases, the balance between co-existing mutant and wild type mtDNA usually underlies disease progression. Cellular mechanisms for maintaining mitochondrial quality include mitophagy, and this could be a critical determinant of disease severity. Previous investigators showed that mitophagy was increased by the drug phenanthroline, a metallopeptidase inhibitor. We reasoned that activating mitophagy with phenanthroline might potentially reduce the load of pathogenic mutant mtDNA in tissue culture cells. We used a previously developed high throughput imaging for quantifying mitophagy in cultured primary fibroblasts bearing the common pathogenic A3243G mtDNA mutation, associated with the mitochondrial encephalomyopathy, lactic acidosis, and strokelike episode syndrome (MELAS) and with diabetes mellitus and deafness. We showed that phenanthroline significantly increased mitophagy in fibroblasts and reduced both the mitochondrial volume and mtDNA content. We conclude that phenanthroline activates mitophagy. However, there was little evidence that it reduced the load of mutant mtDNA. This suggests that both wild type and mutant mtDNA are turned over during mitophagy activated by phenanthroline. We conclude that phenanthroline is a poorly selective activator of mitophagy. Modulators of mitophagy need to target mitochondria enriched for mutant mtDNA if they are to benefit patients with heteroplasmic mtDNA disease.

Mitochondrial DNA (mtDNA)-depletion syndromes (MDS; OMIM 251880) are phenotypically heterogeneous, autosomal-recessive disorders characterized by tissue-specific reduction in mtDNA copy number. Affected individuals with the hepatocerebral form of MDS have early progressive liver failure and neurological abnormalities, hypoglycemia and increased lactate in body fluids. Affected tissues show both decreased activity of the mtDNA-encoded respiratory chain complexes (I, III, IV, V) and mtDNA depletion. We used homozygosity mapping in three kindreds of Druze origin to map the gene causing hepatocerebral MDS to a region of 6.1 cM on chromosome 2p13, between markers D2S291 and D2S2116. This interval encompasses the gene (DGUOK) encoding the mitochondrial deoxyguanosine kinase (dGK). We identified a single-nucleotide deletion (204delA) within the coding region of DGUOK that segregates with the disease in the three kindreds studied. Western-blot analysis did not detect dGK protein in the liver of affected individuals. The main supply of deoxyribonucleotides (dNTPs) for mtDNA synthesis comes from the salvage pathway initiated by dGK and thymidine kinase-2 (TK2). The association of mtDNA depletion with mutated DGUOK suggests that the salvage-pathway enzymes are involved in the maintenance of balanced mitochondrial dNTP pools.

Progressive external ophthalmoplegias (PEO) characterized by accumulation of large-scale mitochondrial DNA (mtDNA) deletions are rare human diseases. We mapped a new locus for dominant PEO at 15q22-q26 in a Belgian pedigree and identified a heterozygous mutation (Y955C) in the polymerase motif B of the mtDNA polymerase gamma (POLG). We identified three additional POLG missense mutations compatible with recessive PEO In two nuclear families. POLG is the only DNA polymerase responsible for mtDNA replication.

The gene products involved in mammalian mitochondrial DNA (mtDNA) maintenance and organization remain largely unknown. We report here a novel mitochondrial protein, Twinkle, with structural similarity to phage T7 gene 4 primase/helicase and other hexameric ring helicases. Twinkle colocalizes with mtDNA in mitochondrial nucleoids. Screening of the gene encoding Twinkle in individuals with autosomal dominant progressive external ophthalmoplegia (adPEO), associated with multiple mtDNA deletions, identified 11 different coding-region mutations co-segregating with the disorder in 12 adPEO pedigrees of various ethnic origins. The mutations cluster in a region of the protein proposed to be involved in subunit interactions. The function of Twinkle is inferred to be critical for lifetime maintenance of human mtDNA integrity.

The mutant mouse strain Mpv17−/−, carries a retroviral germline integration that inactivates the Mpv17 gene. Mpv17-deficient mice develop progressive glomerulosclerosis and sensineural deafness at early age. Characteristic basement membrane alterations are found in both sites of pathology. Mpv17 is a peroxisomal protein involved in the metabolism of reactive oxygen species, yet its molecular function is unknown. Dysregulation of antioxidant enzymes and basal membrane components has been established in this model and successful therapeutic intervention with antioxidants prove the causal role of reactive oxygen species in the development of the disease phenotype. We here investigated if the Mpv17−/ − mice might be hypertensive. Indeed, our study revealed that Mpv17−/ − mice developed significant systemic hypertension and tachycardia between 4 weeks and 5 months of age, accompanied by polyuria and elevated natriuresis. Judging from serum and urine parameters, the hypertensive condition develops concommittantly with the renal disease. Biochemical and pharmacological studies that used the endothelin receptor antagonist bosentan and the angiotensin converting enzyme inhibitor cilazapril indicated no involvement of the endothelin and renin-angiotensin systems in this hypertension, suggesting a potential novel mechanism of blood pressure regulation in this new murine hypertension model. Thus, Mpv17−/− mice unravel an intriguing new association between a defect in reactive oxygen metabolism and the age-dependent development of hypertension.

Objective: To identify the cause of an adult-onset multisystemic disease with multiple deletions of mitochondrial DNA (mtDNA). Design: Case report. Setting: University hospitals. Patient: A 65-year-old man with axonal sensorimotor peripheral neuropathy, ptosis, ophthalmoparesis, diabetes mellitus, exercise intolerance, steatohepatopathy, depression, parkinsonism, and gastrointestinal dysmotility. Results: Skeletal muscle biopsy revealed ragged-red and cytochrome-c oxidase-deficient fibers, and Southern blot analysis showed multiple mtDNA deletions. No deletions were detected in fibroblasts, and the results of quantitative polymerase chain reaction showed that the amount of mtDNA was normal in both muscle and fibroblasts. Exome sequencing using a mitochondrial library revealed compound heterozygous MPV17 mutations (p.LysMet88-89MetLeu and p.Leu143*), a novel cause of mtDNA multiple deletions. Conclusions: In addition to causing juvenile-onset disorders with mtDNA depletion, MPV17 mutations can cause adult-onset multisystemic disease with multiple mtDNA deletions. ©2012 American Medical Association. All rights reserved.

Functional rescue of the glomerulosclerosis phenotype in Mpv17 mice by transgenesis with the human Mpv17 homologue. The germ line insertion of a defective retrovirus into the Mpv17 gene of mice is associated with a recessive phenotype. Mice homozygous for the integration develop glomerulosclerosis at a young age. The phenotype resembles human glomerulosclerosis in its physiological parameters as well as in histology. A human homologue of the Mpv17 gene has been identified, isolated and analyzed. We here show that this gene, which has a role in the production of reactive oxygen species, can rescue the phenotype of Mpv17 deficient mice when introduced by transgenesis. This provides formal proof for the hypothesis that the phenotype is caused by the loss of function of the Mpv17 gene. It also provides evidence for the functional conservation of the Mpv17 gene in mammals and points to a potential role of this gene in human kidney disease.

Corrigendum to “Lethal hepatopathy and leukodystrophy caused by a novel mutation in MPV17 gene: Description of an alternative MPV17 spliced form” [Molecular Genetics and Metabolism 94/2 (2008) 234–239] Aleix Navarro-Sastre, Elena Martin-Hernandez, Yolanda Campos, Ester Quintana, Enrique Medina, Rogelio Simon de las Heras, Montserrat Lluch, Alberto Munoz , Pilar del Hoyo, Rebeca Martin, Laura Gort , Paz Briones, Antonia Ribes⁎

The mutant mouse strain Mpv17 carries a retroviral insert in its genome which inactivates the Mpv17 gene. At a young age these mice develop glomerulosclerosis and nephrotic syndrome which resembles human disease. We show here that the Mpv17 gene product is highly conserved and encodes a peroxisomal protein. Loss of the Mpv17 protein does not impair peroxisome biogenesis but instead leads to a reduced ability to produce reactive oxygen species (ROS). In turn, overproduction of the Mpv17 gene in transfected cells results in dramatically enhanced levels of intracellular ROS indicating a direct involvement of Mpv17 in ROS production. These data reveal a role for the Mpv17 protein in peroxisomal reactive oxygen metabolism and establish a novel link between peroxisomal ROS production and glomerulosclerosis.

To determine the actual incidence of mitochondrial DNA (mtDNA) depletion syndrome in multiple respiratory chain deficiency.We carried out a real-time polymerase chain reaction quantification of mtDNA in liver or muscle tissue of 100 children with unexplained multiple oxidative phosphorylation enzyme deficiency.A reduction of mtDNA copy number to35% of control values was found in liver and/or muscle in half of the children (50/100). Most of these patients (32/50; 64%) presented with severe neonatal onset liver involvement; 7 (14%) had Alpers syndrome, and 11 (22%) exhibited various forms of neurologic involvement. Deoxyguanosine kinase or polymerase gamma (POLG) mutations could be identified in 11 of 32 patients with liver involvement, and POLG mutations were consistently found in all 7 patients with Alpers syndrome. Homozygous thymidine kinase 2 and MPV17 gene mutations were found in 2 patients.Our findings show that mtDNA depletion is a prevalent cause of multiple respiratory chain deficiency in infancy.

Alpers' syndrome is a fatal neurogenetic disorder first described more than 70 years ago. It is an autosomal recessive, developmental mitochondrial DNA depletion disorder characterized by deficiency in mitochondrial DNA polymerase γ (POLG) catalytic activity, refractory seizures, neurodegeneration, and liver disease. In two unrelated pedigrees of Alpers' syndrome, each affected child was found to carry a homozygous mutation in exon 17 of the POLG locus that led to a Glu873Stop mutation just upstream of the polymerase domain of the protein. In addition, each affected child was heterozygous for the G1681A mutation in exon 7 that led to an Ala467Thr substitution in POLG, within the linker region of the protein. Ann Neurol 2004

The mitochondrial DNA (mtDNA) depletion syndrome is a quantitative defect of mtDNA resulting from dysfunction of one of several nuclear-encoded factors responsible for maintenance of mitochondrial deoxyribonucleoside triphosphate (dNTP) pools or replication of mtDNA. Markedly decreased succinyl-CoA synthetase activity due to a deleterious mutation in SUCLA2, the gene encoding the beta subunit of the ADP-forming succinyl-CoA synthetase ligase, was found in muscle mitochondria of patients with encephalomyopathy and mtDNA depletion. Succinyl-CoA synthetase is invariably in a complex with mitochondrial nucleotide diphosphate kinase; hence, we propose that a defect in the last step of mitochondrial dNTP salvage is a novel cause of the mtDNA depletion syndrome.

Cardiolipin (CL) is a diglycerol phospholipid mostly found in mitochondria where it optimizes numerous processes including oxidative phosphorylation (OXPHOS). To function properly CL needs to be unsaturated, which requires the acyltransferase tafazzin. Loss-of-function mutations in this protein are responsible for the Barth syndrome (BTHS), presumably because of a diminished OXPHOS capacity. Here we show that overexpressing Odc1p, a conserved oxodicarboxylic acid carrier located in the mitochondrial inner membrane, fully restores oxidative phosphorylation in a yeast model (taz1Δ) of the Barth syndrome. The rescuing activity involves the recovery of a normal expression of key components that sustain oxidative phosphorylation, including the cytochrome c and complexes IV and III, that are strongly down regulated in taz1Δ yeast. Interestingly, overexpressing Odc1p was shown previously to rescue also yeast models of mitochondrial diseases caused by defects in the assembly of ATP synthase and by mutations in the MPV17 protein that result in the hepatocerebral mitochondrial DNA depletion syndrome. These findings define the transport of oxidicarboxylic acids across the inner membrane as a potential therapeutic target for a large spectrum of mitochondrial disease including BTHS.

Background: Mitochondrial liver disease (MLD), and in particular mitochondrial DNA (mtDNA) depletion syndrome (MDS) is an important cause of acute liver failure (ALF) in infancy. Early and accurate diagnosis is important because liver transplantation (LT) is often contraindicated. It is unclear which methods are the best to diagnose MLD in the setting of ALF. Objective: The aim of the study was to determine the incidence of MLD in children younger than 2 years with ALF and the utility of routine investigations to detect MLD. Methods: Thirty-nine consecutive infants with ALF were admitted to a single unit from 2009 to 2011. All were extensively investigated using an established protocol. Genes implicated in mitochondrial DNA depletion syndrome were sequenced in all cases and tissue mtDNA copy number measured where available. Results: Five infants (17%) had genetically proven MLD: DGUOK (n ¼ 2), POLG (n ¼ 2), and MPV17 (1). Four of these died, whereas 1 recovered. Two had normal muscle mtDNA copy number and 3 had normal muscle respiratory chain enzymes. An additional 8 children had low hepatic mtDNA copy number but pathogenic mutations were not detected. One of these developed fatal multisystemic disease after LT, whereas 5 who survived remain well without evidence of multisystemic disease up to 6 years later. Magnetic resonance spectroscopy did not distinguish between those with and without MLD. Conclusions: Low liver mtDNA copy number may be a secondary phenomenon in ALF. Screening for mtDNA maintenance gene mutations may be the most efficient way to confirm MLD in ALF in the first 2 years of life.

All DNA polymerases misincorporate ribonucleotides despite their preference for deoxyribonucleotides, and analysis of cultured cells indicates that mammalian mitochondrial DNA (mtDNA) tolerates such replication errors. However, it is not clear to what extent misincorporation occurs in tissues, or whether this plays a role in human disease. Here, we show that mtDNA of solid tissues contains many more embedded ribonucleotides than that of cultured cells, consistent with the high ratio of ribonucleotide to deoxynucleotide triphosphates in tissues, and that riboadenosines account for three-quarters of them. The pattern of embedded ribonucleotides changes in a mouse model of Mpv17 deficiency, which displays a marked increase in rGMPs in mtDNA. However, while the mitochondrial dGTP is low in the Mpv17−/− liver, the brain shows no change in the overall dGTP pool, leading us to suggest that Mpv17 determines the local concentration or quality of dGTP. Embedded rGMPs are expected to distort the mtDNA and impede its replication, and elevated rGMP incorporation is associated with early-onset mtDNA depletion in liver and late-onset multiple deletions in brain of Mpv17−/− mice. These findings suggest aberrant ribonucleotide incorporation is a primary mtDNA abnormality that can result in pathology.