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ABSTRACT: Mitochondrial complex I (CI) is a multi-subunit enzyme that forms the major entry point of nicotinamide adenine dinucleotide (NADH) electrons into the respiratory chain. Mutations in the NDUFS4 gene, encoding an accessory subunit of this complex, cause a Leigh-like phenotype in humans. To study the nature and penetrance of the CI defect in different tissues, we investigated the role of NDUFS4 in mice with fatal mitochondrial encephalomyopathy, caused by a systemic inactivation of the Ndufs4 gene. We report that the absence of NDUFS4 in different mouse tissues results in decreased activity and stability of CI. This CI instability leads to an increased disconnection of electron influx of the NADH dehydrogenase module from the holo-complex. However, the formation of respiratory supercomplexes still allows formation of active CI in these Ndufs4 knock-out mice. These results reveal the importance of these supramolecular interactions not only for stabilization but also for the assembly of CI, which becomes especially relevant in pathological conditions.
Human Molecular Genetics 09/2011; 21(1):115-20. · 7.64 Impact Factor
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ABSTRACT: We report a new mutation in m.12146 A>G in the mt-tRNA(His) in a family with a remarkable clinical history having different degrees of lactic acidosis and stroke-like episodes. Biochemical measurements of a muscle biopsy established an isolated complex IV deficiency, while similar analysis of fibroblasts showed a combined complex I,III and IV deficiency. Transmitochondrial cybrid analysis proved that this tRNA(His) mutation causes the enzymatic deficiency. This family illustrates the complexity of the clinical, biochemical and genetic characteristics of a novel mtDNA encoded disorder, as well as the challenge to prove its pathogenicity.
Mitochondrion 06/2011; 11(5):778-82. · 3.62 Impact Factor
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ABSTRACT: This study investigated a girl with Leigh syndrome born to first-cousin parents of Pakistani descent with an isolated respiratory chain complex I deficiency in muscle and fibroblasts. Her early development was delayed, and from age 2 years she started losing motor abilities. Cerebral MRI showed basal ganglia lesions typical of Leigh syndrome.
A genome-wide search for homozygosity was performed with the Affymetrix GeneChip 50K Xba array. The analysis revealed several homozygous regions. Three candidate genes were identified, and in one of the genes, NDUFA12, a homozygous c.178C→T mutation leading to a premature stop codon (p.Arg60X) was found. Western blot analysis showed absence of NDUFA12 protein in patient fibroblasts and functional complementation by a baculovirus system showed restoration of complex I activity.
NDUFA12 mutations are apparently not a frequent cause of complex I deficiency, since mutations were not found by screening altogether 122 complex I deficient patients in two different studies. NDUFA12 encodes an accessory subunit of complex I and is a paralogue of NDUFAF2. Despite the complete absence of NDUFA12 protein, a fully assembled and enzymatically active complex I could be found, albeit in reduced amounts. This suggests that NDUFA12 is required either at a late step in the assembly of complex I, or in the stability of complex I.
Journal of Medical Genetics 05/2011; 48(11):737-40. · 6.36 Impact Factor
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Mateusz Kolanczyk,
Markus Pech,
Tomasz Zemojtel,
Hiroshi Yamamoto,
Ivan Mikula,
Maria-Antonietta Calvaruso, Mariël van den Brand,
Ricarda Richter,
Bjoern Fischer,
Anita Ritz, [......],
Jan Smeitink,
Uwe Kornak,
Danny Chan,
Martin Vingron,
Pavel Martasek,
Robert N Lightowlers,
Leo Nijtmans,
Markus Schuelke,
Knud H Nierhaus,
Stefan Mundlos
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ABSTRACT: Nitric oxide associated-1 (NOA1) is an evolutionarily conserved guanosine triphosphate (GTP) binding protein that localizes predominantly to mitochondria in mammalian cells. On the basis of bioinformatic analysis, we predicted its possible involvement in ribosomal biogenesis, although this had not been supported by any experimental evidence. Here we determine NOA1 function through generation of knockout mice and in vitro assays. NOA1-deficient mice exhibit midgestation lethality associated with a severe developmental defect of the embryo and trophoblast. Primary embryonic fibroblasts isolated from NOA1 knockout embryos show deficient mitochondrial protein synthesis and a global defect of oxidative phosphorylation (OXPHOS). Additionally, Noa1⁻/⁻ cells are impaired in staurosporine-induced apoptosis. The analysis of mitochondrial ribosomal subunits from Noa1⁻/⁻ cells by sucrose gradient centrifugation and Western blotting showed anomalous sedimentation, consistent with a defect in mitochondrial ribosome assembly. Furthermore, in vitro experiments revealed that intrinsic NOA1 GTPase activity was stimulated by bacterial ribosomal constituents. Taken together, our data show that NOA1 is required for mitochondrial protein synthesis, likely due to its yet unidentified role in mitoribosomal biogenesis. Thus, NOA1 is required for such basal mitochondrial functions as adenosine triphosphate (ATP) synthesis and apoptosis.
Molecular biology of the cell 01/2011; 22(1):1-11. · 5.98 Impact Factor
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Jessica Nouws,
Leo Nijtmans,
Sander M Houten, Mariël van den Brand,
Martijn Huynen,
Hanka Venselaar,
Saskia Hoefs,
Jolein Gloerich,
Jonathan Kronick,
Timothy Hutchin,
Peter Willems,
Richard Rodenburg,
Ronald Wanders,
Lambert van den Heuvel,
Jan Smeitink,
Rutger O Vogel
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ABSTRACT: Acyl-CoA dehydrogenase 9 (ACAD9) is a recently identified member of the acyl-CoA dehydrogenase family. It closely resembles very long-chain acyl-CoA dehydrogenase (VLCAD), involved in mitochondrial beta oxidation of long-chain fatty acids. Contrary to its previously proposed involvement in fatty acid oxidation, we describe a role for ACAD9 in oxidative phosphorylation. ACAD9 binds complex I assembly factors NDUFAF1 and Ecsit and is specifically required for the assembly of complex I. Furthermore, ACAD9 mutations result in complex I deficiency and not in disturbed long-chain fatty acid oxidation. This strongly contrasts with its evolutionary ancestor VLCAD, which we show is not required for complex I assembly and clearly plays a role in fatty acid oxidation. Our results demonstrate that two closely related metabolic enzymes have diverged at the root of the vertebrate lineage to function in two separate mitochondrial metabolic pathways and have clinical implications for the diagnosis of complex I deficiency.
Cell metabolism 09/2010; 12(3):283-94. · 17.35 Impact Factor
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Paulien Smits,
Sandy Mattijssen,
Eva Morava, Mariël van den Brand,
Frans van den Brandt,
Frits Wijburg,
Ger Pruijn,
Jan Smeitink,
Leo Nijtmans,
Richard Rodenburg,
Lambert van den Heuvel
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ABSTRACT: Combined oxidative phosphorylation (OXPHOS) system deficiencies are a group of mitochondrial disorders that are associated with a range of clinical phenotypes and genetic defects. They occur in approximately 30% of all OXPHOS disorders and around 4% are combined complex I, III and IV deficiencies. In this study we present two mutations in the mitochondrial tRNA(Trp) (MT-TW) and tRNA(Arg) (MT-TR) genes, m.5556G>A and m.10450A>G, respectively, which were detected in two unrelated patients showing combined OXPHOS complex I, III and IV deficiencies and progressive multisystemic diseases. Both mitochondrial tRNA mutations were almost homoplasmic in fibroblasts and muscle tissue of the two patients and not present in controls. Patient fibroblasts showed a general mitochondrial translation defect. The mutations resulted in lowered steady-state levels and altered conformations of the tRNAs. Cybrid cell lines showed similar tRNA defects and impairment of OXPHOS complex assembly as patient fibroblasts. Our results show that these tRNA(Trp) and tRNA(Arg) mutations cause the combined OXPHOS deficiencies in the patients, adding to the still expanding group of pathogenic mitochondrial tRNA mutations.
European journal of human genetics: EJHG 10/2009; 18(3):324-9. · 3.56 Impact Factor
