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1.
PLoS Biol ; 16(1): e2003992, 2018 01.
Artículo en Inglés | MEDLINE | ID: mdl-29370167

RESUMEN

In endothermic species, heat released as a product of metabolism ensures stable internal temperature throughout the organism, despite varying environmental conditions. Mitochondria are major actors in this thermogenic process. Part of the energy released by the oxidation of respiratory substrates drives ATP synthesis and metabolite transport, but a substantial proportion is released as heat. Using a temperature-sensitive fluorescent probe targeted to mitochondria, we measured mitochondrial temperature in situ under different physiological conditions. At a constant external temperature of 38 °C, mitochondria were more than 10 °C warmer when the respiratory chain (RC) was fully functional, both in human embryonic kidney (HEK) 293 cells and primary skin fibroblasts. This differential was abolished in cells depleted of mitochondrial DNA or treated with respiratory inhibitors but preserved or enhanced by expressing thermogenic enzymes, such as the alternative oxidase or the uncoupling protein 1. The activity of various RC enzymes was maximal at or slightly above 50 °C. In view of their potential consequences, these observations need to be further validated and explored by independent methods. Our study prompts a critical re-examination of the literature on mitochondria.


Asunto(s)
Mitocondrias/fisiología , Termogénesis/fisiología , Fibroblastos/fisiología , Colorantes Fluorescentes , Células HEK293 , Calor , Humanos , Membranas Mitocondriales/fisiología , Proteínas Mitocondriales/metabolismo , Oxidorreductasas/metabolismo , Proteínas de Plantas/metabolismo , Cultivo Primario de Células , Piel , Temperatura , Proteína Desacopladora 1/metabolismo
2.
Am J Hum Genet ; 101(2): 283-290, 2017 Aug 03.
Artículo en Inglés | MEDLINE | ID: mdl-28757203

RESUMEN

Lipoate serves as a cofactor for the glycine cleavage system (GCS) and four 2-oxoacid dehydrogenases functioning in energy metabolism (α-oxoglutarate dehydrogenase [α-KGDHc] and pyruvate dehydrogenase [PDHc]), or amino acid metabolism (branched-chain oxoacid dehydrogenase, 2-oxoadipate dehydrogenase). Mitochondrial lipoate synthesis involves three enzymatic steps catalyzed sequentially by lipoyl(octanoyl) transferase 2 (LIPT2), lipoic acid synthetase (LIAS), and lipoyltransferase 1 (LIPT1). Mutations in LIAS have been associated with nonketotic hyperglycinemia-like early-onset convulsions and encephalopathy combined with a defect in mitochondrial energy metabolism. LIPT1 deficiency spares GCS deficiency and has been associated with a biochemical signature of combined 2-oxoacid dehydrogenase deficiency leading to early death or Leigh-like encephalopathy. We report on the identification of biallelic LIPT2 mutations in three affected individuals from two families with severe neonatal encephalopathy. Brain MRI showed major cortical atrophy with white matter abnormalities and cysts. Plasma glycine was mildly increased. Affected individuals' fibroblasts showed reduced oxygen consumption rates, PDHc, α-KGDHc activities, leucine catabolic flux, and decreased protein lipoylation. A normalization of lipoylation was observed after expression of wild-type LIPT2, arguing for LIPT2 requirement in intramitochondrial lipoate synthesis. Lipoic acid supplementation did not improve clinical condition nor activities of PDHc, α-KGDHc, or leucine metabolism in fibroblasts and was ineffective in yeast deleted for the orthologous LIP2.


Asunto(s)
Aciltransferasas/genética , Atrofia/patología , Encefalopatías/genética , Encéfalo/patología , Lipoilación/genética , Mitocondrias/metabolismo , Aminoácidos/metabolismo , Encéfalo/diagnóstico por imagen , Encefalopatías/patología , Mapeo Encefálico/métodos , Células Cultivadas , Metabolismo Energético/genética , Metabolismo Energético/fisiología , Glicina/sangre , Humanos , Recién Nacido , Imagen por Resonancia Magnética , Mitocondrias/genética , Consumo de Oxígeno/genética , Unión Proteica/genética , Ácido Tióctico/metabolismo
3.
Int J Mol Sci ; 21(6)2020 Mar 16.
Artículo en Inglés | MEDLINE | ID: mdl-32188137

RESUMEN

UDP-glucose (UDP-Glc) is synthesized by UGP2-encoded UDP-Glc pyrophosphorylase (UGP) and is required for glycoconjugate biosynthesis and galactose metabolism because it is a uridyl donor for galactose-1-P (Gal1P) uridyltransferase. Chinese hamster lung fibroblasts harboring a hypomrphic UGP(G116D) variant display reduced UDP-Glc levels and cannot grow if galactose is the sole carbon source. Here, these cells were cultivated with glucose in either the absence or presence of galactose in order to investigate glycoconjugate biosynthesis and galactose metabolism. The UGP-deficient cells display < 5% control levels of UDP-Glc/UDP-Gal and > 100-fold reduction of [6-3H]galactose incorporation into UDP-[6-3H]galactose, as well as multiple deficits in glycoconjugate biosynthesis. Cultivation of these cells in the presence of galactose leads to partial restoration of UDP-Glc levels, galactose metabolism and glycoconjugate biosynthesis. The Vmax for recombinant human UGP(G116D) with Glc1P is 2000-fold less than that of the wild-type protein, and UGP(G116D) displayed a mildly elevated Km for Glc1P, but no activity of the mutant enzyme towards Gal1P was detectable. To conclude, although the mechanism behind UDP-Glc/Gal production in the UGP-deficient cells remains to be determined, the capacity of this cell line to change its glycosylation status as a function of extracellular galactose makes it a useful, reversible model with which to study different aspects of galactose metabolism and glycoconjugate biosynthesis.


Asunto(s)
Galactosa/biosíntesis , Glicoconjugados/biosíntesis , UTP-Glucosa-1-Fosfato Uridililtransferasa/genética , Animales , Encefalopatías/metabolismo , Línea Celular , Trastornos Congénitos de Glicosilación/genética , Trastornos Congénitos de Glicosilación/metabolismo , Cricetinae , Medios de Cultivo/química , Glicoesfingolípidos , Glicosilación , Humanos , Cinética , Pulmón , UTP-Glucosa-1-Fosfato Uridililtransferasa/metabolismo , Uridina Difosfato Glucosa/biosíntesis
4.
Am J Hum Genet ; 95(6): 708-20, 2014 Dec 04.
Artículo en Inglés | MEDLINE | ID: mdl-25434004

RESUMEN

Respiratory chain deficiencies exhibit a wide variety of clinical phenotypes resulting from defective mitochondrial energy production through oxidative phosphorylation. These defects can be caused by either mutations in the mtDNA or mutations in nuclear genes coding for mitochondrial proteins. The underlying pathomechanisms can affect numerous pathways involved in mitochondrial physiology. By whole-exome and candidate gene sequencing, we identified 11 individuals from 9 families carrying compound heterozygous or homozygous mutations in GTPBP3, encoding the mitochondrial GTP-binding protein 3. Affected individuals from eight out of nine families presented with combined respiratory chain complex deficiencies in skeletal muscle. Mutations in GTPBP3 are associated with a severe mitochondrial translation defect, consistent with the predicted function of the protein in catalyzing the formation of 5-taurinomethyluridine (τm(5)U) in the anticodon wobble position of five mitochondrial tRNAs. All case subjects presented with lactic acidosis and nine developed hypertrophic cardiomyopathy. In contrast to individuals with mutations in MTO1, the protein product of which is predicted to participate in the generation of the same modification, most individuals with GTPBP3 mutations developed neurological symptoms and MRI involvement of thalamus, putamen, and brainstem resembling Leigh syndrome. Our study of a mitochondrial translation disorder points toward the importance of posttranscriptional modification of mitochondrial tRNAs for proper mitochondrial function.


Asunto(s)
Acidosis Láctica/genética , Encefalopatías/genética , Cardiomiopatía Hipertrófica/genética , Proteínas de Unión al GTP/genética , Procesamiento Proteico-Postraduccional , Acidosis Láctica/fisiopatología , Secuencia de Aminoácidos , Encéfalo/patología , Encefalopatías/fisiopatología , Cardiomiopatía Hipertrófica/fisiopatología , Línea Celular , Niño , Preescolar , Consanguinidad , Femenino , Fibroblastos , Proteínas de Unión al GTP/metabolismo , Humanos , Lactante , Recién Nacido , Masculino , Datos de Secuencia Molecular , Mutación , Linaje , Biosíntesis de Proteínas , Interferencia de ARN , ARN de Transferencia/genética , ARN de Transferencia/metabolismo , Alineación de Secuencia
5.
Hum Mutat ; 37(12): 1340-1353, 2016 12.
Artículo en Inglés | MEDLINE | ID: mdl-27528516

RESUMEN

Next-generation sequencing (NGS) has an established diagnostic value for inherited ataxia. However, the need of a rigorous process of analysis and validation remains challenging. Moreover, copy number variations (CNV) or dynamic expansions of repeated sequence are classically considered not adequately detected by exome sequencing technique. We applied a strategy of mini-exome coupled to read-depth based CNV analysis to a series of 33 patients with probable inherited ataxia and onset <50 years. The mini-exome consisted of the capture of 4,813 genes having associated clinical phenotypes. Pathogenic variants were found in 42% and variants of uncertain significance in 24% of the patients. These results are comparable to those from whole exome sequencing and better than previous targeted NGS studies. CNV and dynamic expansions of repeated CAG sequence were identified in three patients. We identified both atypical presentation of known ataxia genes (ATM, NPC1) and mutations in genes very rarely associated with ataxia (ERCC4, HSD17B4). We show that mini-exome bioinformatics data analysis allows the identification of CNV and dynamic expansions of repeated sequence. Our study confirms the diagnostic value of the proposed genetic analysis strategy. We also provide an algorithm for the multidisciplinary process of analysis, interpretation, and validation of NGS data.


Asunto(s)
Ataxia Cerebelosa/genética , Variaciones en el Número de Copia de ADN , Exoma , Secuenciación de Nucleótidos de Alto Rendimiento/métodos , Análisis de Secuencia de ADN/métodos , Adolescente , Adulto , Edad de Inicio , Proteínas de la Ataxia Telangiectasia Mutada/genética , Proteínas Portadoras/genética , Ataxia Cerebelosa/etiología , Niño , Preescolar , Proteínas de Unión al ADN/genética , Femenino , Predisposición Genética a la Enfermedad , Humanos , Péptidos y Proteínas de Señalización Intracelular , Masculino , Glicoproteínas de Membrana/genética , Proteína Niemann-Pick C1 , Proteína-2 Multifuncional Peroxisomal/genética , Adulto Joven
6.
Clin Sci (Lond) ; 130(6): 393-407, 2016 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-26846578

RESUMEN

As with other mitochondrial respiratory chain components, marked clinical and genetic heterogeneity is observed in patients with a cytochrome c oxidase deficiency. This constitutes a considerable diagnostic challenge and raises a number of puzzling questions. So far, pathological mutations have been reported in more than 30 genes, in both mitochondrial and nuclear DNA, affecting either structural subunits of the enzyme or proteins involved in its biogenesis. In this review, we discuss the possible causes of the discrepancy between the spectacular advances made in the identification of the molecular bases of cytochrome oxidase deficiency and the lack of any efficient treatment in diseases resulting from such deficiencies. This brings back many unsolved questions related to the frequent delay of clinical manifestation, variable course and severity, and tissue-involvement often associated with these diseases. In this context, we stress the importance of studying different models of these diseases, but also discuss the limitations encountered in most available disease models. In the future, with the possible exception of replacement therapy using genes, cells or organs, a better understanding of underlying mechanism(s) of these mitochondrial diseases is presumably required to develop efficient therapy.


Asunto(s)
Deficiencia de Citocromo-c Oxidasa/genética , Animales , Células Cultivadas , Deficiencia de Citocromo-c Oxidasa/terapia , Modelos Animales de Enfermedad , Complejo IV de Transporte de Electrones/química , Complejo IV de Transporte de Electrones/genética , Complejo IV de Transporte de Electrones/metabolismo , Humanos , Estructura Molecular
7.
Nat Genet ; 39(6): 776-80, 2007 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-17486094

RESUMEN

Mitochondrial DNA (mtDNA) depletion syndrome (MDS; MIM 251880) is a prevalent cause of oxidative phosphorylation disorders characterized by a reduction in mtDNA copy number. The hitherto recognized disease mechanisms alter either mtDNA replication (POLG (ref. 1)) or the salvage pathway of mitochondrial deoxyribonucleosides 5'-triphosphates (dNTPs) for mtDNA synthesis (DGUOK (ref. 2), TK2 (ref. 3) and SUCLA2 (ref. 4)). A last gene, MPV17 (ref. 5), has no known function. Yet the majority of cases remain unexplained. Studying seven cases of profound mtDNA depletion (1-2% residual mtDNA in muscle) in four unrelated families, we have found nonsense, missense and splice-site mutations and in-frame deletions of the RRM2B gene, encoding the cytosolic p53-inducible ribonucleotide reductase small subunit. Accordingly, severe mtDNA depletion was found in various tissues of the Rrm2b-/- mouse. The mtDNA depletion triggered by p53R2 alterations in both human and mouse implies that p53R2 has a crucial role in dNTP supply for mtDNA synthesis.


Asunto(s)
Proteínas de Ciclo Celular/genética , ADN Mitocondrial/genética , Eliminación de Gen , Enfermedades Mitocondriales/etiología , Mutación/genética , Ribonucleótido Reductasas/genética , Proteína p53 Supresora de Tumor/metabolismo , Animales , Proteínas de Ciclo Celular/fisiología , Células Cultivadas , Análisis Mutacional de ADN , Femenino , Fibroblastos , Homocigoto , Humanos , Recién Nacido , Escala de Lod , Masculino , Ratones , Ratones Noqueados , Mitocondrias Musculares , Enfermedades Mitocondriales/patología , Datos de Secuencia Molecular , Linaje , Ribonucleótido Reductasas/fisiología , Proteína p53 Supresora de Tumor/genética
8.
J Med Genet ; 51(12): 834-8, 2014 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-25351951

RESUMEN

BACKGROUND: Inherited optic neuropathy has been ascribed to mutations in mitochondrial fusion/fission dynamics genes, nuclear and mitochondrial DNA-encoded respiratory enzyme genes or nuclear genes of poorly known mitochondrial function. However, the disease causing gene remains unknown in many families. METHODS: We used exome sequencing in order to identify the gene responsible for isolated or syndromic optic atrophy in five patients from three independent families. RESULTS: We found homozygous or compound heterozygous missense and frameshift mutations in the gene encoding mitochondrial aconitase (ACO2), a tricarboxylic acid cycle enzyme, catalysing interconversion of citrate into isocitrate. Unlike wild type ACO2, all mutant ACO2 proteins failed to complement the respiratory growth of a yeast aco1-deletion strain. Retrospective studies using patient-derived cultured skin fibroblasts revealed various degrees of deficiency in ACO2 activity, but also in ACO1 cytosolic activity. CONCLUSIONS: Our study shows that autosomal recessive ACO2 mutations can cause either isolated or syndromic optic neuropathy. This observation identifies ACO2 as the second gene responsible for non-syndromic autosomal recessive optic neuropathies and provides evidence for a genetic overlap between isolated and syndromic forms, giving further support to the view that optic atrophy is a hallmark of defective mitochondrial energy supply.


Asunto(s)
Aconitato Hidratasa/genética , Mutación , Enfermedades del Nervio Óptico/genética , Aconitato Hidratasa/metabolismo , Adulto , Encéfalo/patología , Preescolar , Ciclo del Ácido Cítrico , Activación Enzimática , Exoma , Resultado Fatal , Femenino , Expresión Génica , Genes Recesivos , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Imagen por Resonancia Magnética , Masculino , Oftalmoscopios , Atrofia Óptica/diagnóstico , Atrofia Óptica/genética , Enfermedades del Nervio Óptico/diagnóstico , Enfermedades del Nervio Óptico/metabolismo , Evaluación del Resultado de la Atención al Paciente , Hermanos
9.
Biochim Biophys Acta ; 1832(8): 1304-12, 2013 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-23603806

RESUMEN

Multiple respiratory chain deficiencies represent a common cause of mitochondrial diseases and are associated with a wide range of clinical symptoms. We report a subject, born to consanguineous parents, with growth retardation and neurological deterioration. Multiple respiratory chain deficiency was found in muscle and fibroblasts of the subject as well as abnormal assembly of complexes I and IV. A microsatellite genotyping of the family members detected only one region of homozygosity on chromosome 17q24.2-q25.3 in which we focused our attention to genes involved in mitochondrial translation. We sequenced MRPL12, encoding the mitochondrial ribosomal protein L12 and identified a c.542C>T transition in exon 5 changing a highly conserved alanine into a valine (p.Ala181Val). This mutation resulted in a decreased steady-state level of MRPL12 protein, with altered integration into the large ribosomal subunit. Moreover, an overall mitochondrial translation defect was observed in the subject's fibroblasts with a significant reduction of synthesis of COXI, COXII and COXIII subunits. Modeling of MRPL12 shows Ala181 positioned in a helix potentially involved in an interface of interaction suggesting that the p.Ala181Val change might be predicted to alter interactions with the elongation factors. These results contrast with the eubacterial orthologues of human MRPL12, where L7/L12 proteins do not appear to have a selective effect on translation. Therefore, analysis of the mutated version found in the subject presented here suggests that the mammalian protein does not function in an entirely analogous manner to the eubacterial L7/L12 equivalent.


Asunto(s)
Proteínas de Ciclo Celular/genética , Trastornos del Crecimiento/genética , Enfermedades Mitocondriales/genética , Proteínas Mitocondriales/genética , Mutación , Enfermedades del Sistema Nervioso/genética , Proteínas Nucleares/genética , Proteínas Ribosómicas/genética , Secuencia de Aminoácidos , Proteínas de Ciclo Celular/química , Proteínas de Ciclo Celular/metabolismo , Células Cultivadas , Preescolar , Fibroblastos/metabolismo , Genotipo , Trastornos del Crecimiento/metabolismo , Humanos , Masculino , Mitocondrias/genética , Mitocondrias/metabolismo , Enfermedades Mitocondriales/metabolismo , Proteínas Mitocondriales/metabolismo , Modelos Moleculares , Datos de Secuencia Molecular , Enfermedades del Sistema Nervioso/metabolismo , Proteínas Nucleares/química , Proteínas Nucleares/metabolismo , Extensión de la Cadena Peptídica de Translación , Proteínas Ribosómicas/química , Proteínas Ribosómicas/metabolismo , Alineación de Secuencia
10.
Mol Genet Metab ; 109(1): 28-32, 2013 May.
Artículo en Inglés | MEDLINE | ID: mdl-23478190

RESUMEN

The causes of Reye-like syndrome are not completely understood. Dihydrolipoamide dehydrogenase (DLD or E3) deficiency is a rare metabolic disorder causing neurological or liver impairment. Specific changes in the levels of urinary and plasma metabolites are the hallmark of the classical form of the disease. Here, we report a consanguineous family of Algerian origin with DLD deficiency presenting without suggestive clinical laboratory and anatomopathological findings. Two children died at birth from hepatic failure and three currently adult siblings had recurrent episodes of hepatic cytolysis associated with liver failure or Reye-like syndrome from infancy. Biochemical investigation (lactate, pyruvate, aminoacids in plasma, organic acids in urine) was normal. Histologic examination of liver and muscle showed mild lipid inclusions that were only visible by electron microscopy. The diagnosis of DLD deficiency was possible only after genome-wide linkage analysis, confirmed by a homozygous mutation (p.G229C) in the DLD gene, previously reported in patients with the same geographic origin. DLD and pyruvate dehydrogenase activities were respectively reduced to 25% and 70% in skin fibroblasts of patients and were unresponsive to riboflavin supplementation. In conclusion, this observation clearly supports the view that DLD deficiency should be considered in patients with Reye-like syndrome or liver failure even in the absence of suggestive biochemical findings, with the p.G229C mutation screening as a valuable test in the Arab patients because of its high frequency. It also highlights the usefulness of genome-wide linkage analysis for decisive diagnosis advance in inherited metabolic disorders.


Asunto(s)
Acidosis Láctica/patología , Dihidrolipoamida Deshidrogenasa , Fallo Hepático Agudo/genética , Enfermedad de la Orina de Jarabe de Arce/patología , Síndrome de Reye/genética , Acidosis Láctica/sangre , Acidosis Láctica/genética , Acidosis Láctica/mortalidad , Acidosis Láctica/orina , Adulto , Argelia , Niño , Dihidrolipoamida Deshidrogenasa/genética , Dihidrolipoamida Deshidrogenasa/metabolismo , Femenino , Humanos , Lactante , Hígado/patología , Fallo Hepático Agudo/sangre , Fallo Hepático Agudo/mortalidad , Fallo Hepático Agudo/patología , Fallo Hepático Agudo/orina , Masculino , Enfermedad de la Orina de Jarabe de Arce/sangre , Enfermedad de la Orina de Jarabe de Arce/genética , Enfermedad de la Orina de Jarabe de Arce/mortalidad , Enfermedad de la Orina de Jarabe de Arce/orina , Músculos/patología , Mutación , Síndrome de Reye/metabolismo , Síndrome de Reye/mortalidad , Síndrome de Reye/fisiopatología
11.
J Hepatol ; 56(1): 294-7, 2012 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-21741925

RESUMEN

BACKGROUND & AIMS: Multiple respiratory chain deficiencies represent a common cause of mitochondrial diseases and often result in hepatic failure. A significant fraction of patients present mitochondrial DNA depletion but a number of cases remain unexplained. The aim of our study was to identify the disease causing gene in a kindred with intrauterine growth retardation, neonatal lactic acidosis, liver dysfunction and multiple respiratory chain deficiency in muscle. METHODS: Homozygosity mapping was performed by 50K SNP genotyping and candidate genes were successively analyzed by direct sequencing on genomic DNA of the family members. RESULTS: SNP genotyping detected several regions of homozygosity in which we focused our attention to genes involved in mitochondrial translation. We sequenced the TSFM gene, encoding the mitochondrial translation factor EFTs and identified a homozygous mutation changing a highly conserved arginine into a tryptophan (R312W). CONCLUSIONS: This mutation has been previously reported in two unrelated kindred presenting two distinct syndromes (fatal mitochondrial encephalomyopathy and hypertrophic cardiomyopathy respectively). The description of a third syndrome associated with a same TSFM mutation gives support to the broad clinical and genetic heterogeneity of mitochondrial translation deficiencies in human. It suggests that mitochondrial translation deficiency represents a growing cause of hepatic failure of mitochondrial origin in infants.


Asunto(s)
Fallo Hepático Agudo/genética , Mutación Missense , Factores de Elongación de Péptidos/genética , Sustitución de Aminoácidos , Secuencia de Bases , ADN Mitocondrial/genética , Femenino , Homocigoto , Humanos , Lactante , Recién Nacido , Fallo Hepático Agudo/etiología , Masculino , Enfermedades Mitocondriales/etiología , Enfermedades Mitocondriales/genética , Linaje
12.
Am J Hum Genet ; 84(4): 493-8, 2009 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-19327736

RESUMEN

Nonsyndromic autosomal-recessive optic neuropathies are rare conditions of unknown genetic and molecular origin. Using an approach of whole-genome homozygosity mapping and positional cloning, we have identified the first gene, to our knowledge, responsible for this condition, TMEM126A, in a large multiplex inbred Algerian family and subsequently in three other families originating from the Maghreb. TMEM126A is conserved in higher eukaryotes and encodes a transmembrane mitochondrial protein of unknown function, supporting the view that mitochondrial dysfunction may be a hallmark of inherited optic neuropathies including isolated autosomal-recessive forms.


Asunto(s)
Proteínas Mitocondriales/genética , Mutación , Atrofias Ópticas Hereditarias/genética , Argelia , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Células COS , Chlorocebus aethiops , Codón sin Sentido , Femenino , Expresión Génica , Genes Recesivos , Haplotipos , Humanos , Masculino , Ratones , Proteínas Mitocondriales/química , Proteínas Mitocondriales/metabolismo , Datos de Secuencia Molecular , Estructura Terciaria de Proteína , ARN Mensajero/genética , ARN Mensajero/metabolismo , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/metabolismo , Retina/metabolismo , Homología de Secuencia de Aminoácido , Transfección
13.
Hum Mutat ; 32(11): 1225-31, 2011 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-21786366

RESUMEN

By combining exome sequencing in conjunction with genetic mapping, we have identified the first mutation in large mitochondrial ribosomal protein MRPL3 in a family of four sibs with hypertrophic cardiomyopathy, psychomotor retardation, and multiple respiratory chain deficiency. Affected sibs were compound heterozygotes for a missense MRPL3 mutation (P317R) and a large-scale deletion, inherited from the mother and the father, respectively. These mutations were shown to alter ribosome assembly and cause a mitochondrial translation deficiency in cultured skin fibroblasts resulting in an abnormal assembly of several complexes of the respiratory chain. This observation gives support to the view that exome sequencing combined with genetic mapping is a powerful approach for the identification of new genes of mitochondrial disorders.


Asunto(s)
Cardiomiopatía Hipertrófica/genética , Mitocondrias/metabolismo , Enfermedades Mitocondriales/genética , Proteínas Mitocondriales/genética , Mutación , Proteínas Ribosómicas/genética , Secuencia de Bases , Cardiomiopatía Hipertrófica/patología , Análisis Mutacional de ADN , ADN Mitocondrial/química , Exoma , Femenino , Humanos , Lactante , Recién Nacido , Masculino , Enfermedades Mitocondriales/patología , Datos de Secuencia Molecular , Proteínas Ribosómicas/metabolismo , Eliminación de Secuencia
14.
Hum Mutat ; 32(9): 1046-52, 2011 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-21560188

RESUMEN

Fumarase deficiency (FD), caused by biallelic alteration of the Fumarase Hydratase gene (FH), and a rare metabolic disorder that affects the Krebs cycle, causes severe neurological impairment and fumaric aciduria. Less than 30 unrelated cases are known to date. In addition, heterozygous mutations of the FH gene are responsible for hereditary leiomyomatosis and renal cell cancer (HLRCC). We report three additional patients with dramatically different clinical presentations of FD and novel missense mutations in the FH gene. One patient had severe neonatal encephalopathy, polymicrogyria, <1% enzyme activity, and mildly increased levels of urinary fumarate. The second patient had microcephaly, mental retardation, 20% of fumarase activity, and intermediate levels of urinary fumarate. The third patient had mild mental retardation, polymicrogyria, 42-61% enzyme activity in different cell types and massive amounts of urinary fumarate. In silico analysis predicted minor yet significant structural changes in the encoded proteins. The nuclear translocation of hypoxia-inducible factor (HIF)-1alpha (HIF1A) in cultured fibroblasts was similar to controls. These results extend the range of clinical and biochemical variation associated with FD, supporting the notion that patients with moderate increases in fumarate excretion should be investigated for this disease. The tumoral risk in the patients and their relatives requires adequate screening protocols.


Asunto(s)
Fumarato Hidratasa/deficiencia , Fumarato Hidratasa/metabolismo , Malformaciones del Desarrollo Cortical/enzimología , Malformaciones del Desarrollo Cortical/patología , Hipoxia de la Célula , Niño , Preescolar , Simulación por Computador , Femenino , Fumarato Hidratasa/química , Fumaratos/orina , Humanos , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Masculino , Mutación/genética , Transducción de Señal
15.
Am J Hum Genet ; 82(3): 623-30, 2008 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-18319072

RESUMEN

Coenzyme Q(10) (CoQ(10)) plays a pivotal role in oxidative phosphorylation (OXPHOS) in that it distributes electrons between the various dehydrogenases and the cytochrome segments of the respiratory chain. Primary coenzyme Q(10) deficiency represents a clinically heterogeneous condition suggestive of genetic heterogeneity, and several disease genes have been previously identified. The CABC1 gene, also called COQ8 or ADCK3, is the human homolog of the yeast ABC1/COQ8 gene, one of the numerous genes involved in the ubiquinone biosynthesis pathway. The exact function of the Abc1/Coq8 protein is as yet unknown, but this protein is classified as a putative protein kinase. We report here CABC1 gene mutations in four ubiquinone-deficient patients in three distinct families. These patients presented a similar progressive neurological disorder with cerebellar atrophy and seizures. In all cases, enzymological studies pointed to ubiquinone deficiency. CoQ(10) deficiency was confirmed by decreased content of ubiquinone in muscle. Various missense mutations (R213W, G272V, G272D, and E551K) modifying highly conserved amino acids of the protein and a 1 bp frameshift insertion c.[1812_1813insG] were identified. The missense mutations were introduced into the yeast ABC1/COQ8 gene and expressed in a Saccharomyces cerevisiae strain in which the ABC1/COQ8 gene was deleted. All the missense mutations resulted in a respiratory phenotype with no or decreased growth on glycerol medium and a severe reduction in ubiquinone synthesis, demonstrating that these mutations alter the protein function.


Asunto(s)
Ataxia Cerebelosa/genética , Convulsiones/genética , Ubiquinona/deficiencia , Adolescente , Adulto , Secuencia de Aminoácidos , Benzoquinonas/análisis , Encéfalo/enzimología , Encéfalo/patología , Femenino , Haplotipos , Humanos , Imagen por Resonancia Magnética , Masculino , Datos de Secuencia Molecular , Músculo Esquelético/química , Mutación Missense , Linaje , Ubiquinona/análisis , Ubiquinona/genética
16.
J Clin Invest ; 117(3): 765-72, 2007 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-17332895

RESUMEN

Coenzyme Q10 (CoQ10) plays a pivotal role in oxidative phosphorylation (OXPHOS), as it distributes electrons among the various dehydrogenases and the cytochrome segments of the respiratory chain. We have identified 2 novel inborn errors of CoQ10 biosynthesis in 2 distinct families. In both cases, enzymologic studies showed that quinone-dependent OXPHOS activities were in the range of the lowest control values, while OXPHOS enzyme activities were normal. CoQ10 deficiency was confirmed by restoration of normal OXPHOS activities after addition of quinone. A genome-wide search for homozygosity in family 1 identified a region of chromosome 10 encompassing the gene prenyldiphosphate synthase, subunit 1 (PDSS1), which encodes the human ortholog of the yeast COQ1 gene, a key enzyme of CoQ10 synthesis. Sequencing of PDSS1 identified a homozygous nucleotide substitution modifying a conserved amino acid of the protein (D308E). In the second family, direct sequencing of OH-benzoate polyprenyltransferase (COQ2), the human ortholog of the yeast COQ2 gene, identified a single base pair frameshift deletion resulting in a premature stop codon (c.1198delT, N401fsX415). Transformation of yeast Deltacoq1 and Deltacoq2 strains by mutant yeast COQ1 and mutant human COQ2 genes, respectively, resulted in defective growth on respiratory medium, indicating that these mutations are indeed the cause of OXPHOS deficiency.


Asunto(s)
Transferasas Alquil y Aril/genética , Enfermedades Mitocondriales/genética , Ubiquinona/análogos & derivados , Ubiquinona/deficiencia , Adolescente , Adulto , Secuencia de Aminoácidos , Sustitución de Aminoácidos/genética , Preescolar , Cromosomas Humanos Par 10/genética , Coenzimas , Femenino , Prueba de Complementación Genética , Homocigoto , Humanos , Masculino , Enfermedades Mitocondriales/enzimología , Datos de Secuencia Molecular , Mutación , Fosforilación Oxidativa , Linaje , Análisis de Secuencia de ADN , Ubiquinona/biosíntesis , Ubiquinona/genética , Levaduras/genética
17.
J Inherit Metab Dis ; 33 Suppl 3: S443-53, 2010 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-20978941

RESUMEN

Ethylmalonic encephalopathy (EE) is a rare metabolic disorder caused by dysfunction of ETHE1, a mitochondrial dioxygenase involved in hydrogen sulfide (H2S) detoxification. Patients present in infancy with psychomotor retardation, chronic diarrhea, orthostatic acrocyanosis and relapsing petechiae. High levels of lactic acid, ethymalonic acid (EMA) and methylsuccinic acid (MSA) are detected in body fluids. Several pathways may contribute to the pathophysiology, including isoleucine, methionine and fatty acid metabolism. We report on a 15-month-old male presenting with typical EE associated with a homozygous ETHE1 mutation. We investigated oral isoleucine (150 mg/kg), methionine (100 mg/kg), fatty acid loading tests and isoleucine-restricted diet (200 mg/day) for any effects on several metabolic parameters. Before loading tests or specific dietary interventions, EMA, C4-C5 acylcarnitines and most acylglycines were elevated, indicating functional deficiency of short chain acyl-CoA (SCAD) as well as all branched acyl-CoA dehydrogenases. Excretion of EMA and n-butyrylglycine increased following each of the loads, and isoleucine led to increased levels of derivative metabolites. An isoleucine-restricted diet for 8 days corrected some of the abnormalities but led to no obvious clinical improvement and only partial effects on EMA. A principal component analysis supports the inference that these dietary conditions have consistent effects on the global metabolic profile. Our results suggest that multiple pathways modulate EMA levels in EE. They might all interact with H2S toxicity. Prolonged dietary interventions involving the restriction for branched aminoacids, fatty acids and methionine could be discussed as auxiliary therapeutical strategies in EE.


Asunto(s)
Encefalopatías Metabólicas Innatas/enzimología , Proteínas Mitocondriales/metabolismo , Proteínas de Transporte Nucleocitoplasmático/metabolismo , Púrpura/enzimología , Aminoácidos/uso terapéutico , Biomarcadores/sangre , Biomarcadores/orina , Encefalopatías Metabólicas Innatas/diagnóstico , Encefalopatías Metabólicas Innatas/dietoterapia , Encefalopatías Metabólicas Innatas/genética , Dieta con Restricción de Proteínas , Suplementos Dietéticos , Predisposición Genética a la Enfermedad , Homocigoto , Humanos , Lactante , Masculino , Malonatos/sangre , Malonatos/orina , Proteínas Mitocondriales/genética , Mutación , Proteínas de Transporte Nucleocitoplasmático/genética , Fenotipo , Análisis de Componente Principal , Púrpura/diagnóstico , Púrpura/dietoterapia , Púrpura/genética , Resultado del Tratamiento
18.
Pediatr Res ; 66(1): 91-5, 2009 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-19342984

RESUMEN

We investigated respiratory chain (RC), tricarboxylic acid cycle (TCA) enzyme activities, and oxidative stress in the tissues of six patients with organic aciduria (OA) presenting various severe complications to further document the role of mitochondrial OXPHOS dysfunction in the development of complications. Two children with propionic acidemia (PA), presenting a severe cardiomyopathy, and four with methylmalonic aciduria (MMA), who developed a neurologic disease (3/4) and renal failure (2/4), were followed. We measured RC and TCA cycle enzyme activity in patient tissues and assessed oxidative metabolism in fibroblasts in vitro. Various RC deficiencies were found in tissues of patients with PA and MMA. TCA cycle enzyme activities were normal when investigated and reactive oxygen species were decreased as well as detoxifying systems activities in the two patients tested. In conclusion, mitochondrial dysfunction was found in all investigated tissues of six patients with organic acidemia presenting with severe complications. Reactive oxygen species production and detoxification were decreased in fibroblast primary cultures. Our results bring further support for a role of secondary respiratory deficiency in the development of late multiorgan complications of these diseases.


Asunto(s)
Errores Innatos del Metabolismo de los Aminoácidos/enzimología , Ciclo del Ácido Cítrico/fisiología , Transporte de Electrón/fisiología , Errores Innatos del Metabolismo Lipídico/enzimología , Estrés Oxidativo/fisiología , Adolescente , Errores Innatos del Metabolismo de los Aminoácidos/complicaciones , Errores Innatos del Metabolismo de los Aminoácidos/genética , Cardiomiopatías/etiología , Femenino , Fibroblastos , Humanos , Lactante , Recién Nacido , Riñón/metabolismo , Errores Innatos del Metabolismo Lipídico/complicaciones , Errores Innatos del Metabolismo Lipídico/genética , Hígado/metabolismo , Masculino , Metilmalonil-CoA Descarboxilasa/genética , Músculo Esquelético/metabolismo , Mutación/genética , Miocardio/metabolismo , Enfermedades del Sistema Nervioso/etiología , Fosforilación Oxidativa , Especies Reactivas de Oxígeno/metabolismo , Insuficiencia Renal/etiología
19.
Nat Commun ; 10(1): 1566, 2019 04 05.
Artículo en Inglés | MEDLINE | ID: mdl-30952952

RESUMEN

The class 3 phosphoinositide 3-kinase (PI3K) is required for lysosomal degradation by autophagy and vesicular trafficking, assuring nutrient availability. Mitochondrial lipid catabolism is another energy source. Autophagy and mitochondrial metabolism are transcriptionally controlled by nutrient sensing nuclear receptors. However, the class 3 PI3K contribution to this regulation is unknown. We show that liver-specific inactivation of Vps15, the essential regulatory subunit of the class 3 PI3K, elicits mitochondrial depletion and failure to oxidize fatty acids. Mechanistically, transcriptional activity of Peroxisome Proliferator Activated Receptor alpha (PPARα), a nuclear receptor orchestrating lipid catabolism, is blunted in Vps15-deficient livers. We find PPARα repressors Histone Deacetylase 3 (Hdac3) and Nuclear receptor co-repressor 1 (NCoR1) accumulated in Vps15-deficient livers due to defective autophagy. Activation of PPARα or inhibition of Hdac3 restored mitochondrial biogenesis and lipid oxidation in Vps15-deficient hepatocytes. These findings reveal roles for the class 3 PI3K and autophagy in transcriptional coordination of mitochondrial metabolism.


Asunto(s)
Autofagia/fisiología , Metabolismo de los Lípidos , Mitocondrias/metabolismo , PPAR alfa/metabolismo , Fosfatidilinositol 3-Quinasas/fisiología , Animales , Autofagia/efectos de los fármacos , Autofagia/genética , Fenofibrato/farmacología , Regulación de la Expresión Génica/efectos de los fármacos , Células HEK293 , Hepatocitos/citología , Hepatocitos/efectos de los fármacos , Hepatocitos/metabolismo , Histona Desacetilasas/genética , Histona Desacetilasas/metabolismo , Histona Desacetilasas/fisiología , Humanos , Metabolismo de los Lípidos/efectos de los fármacos , Masculino , Ratones , Ratones Noqueados , Co-Represor 1 de Receptor Nuclear/genética , Co-Represor 1 de Receptor Nuclear/metabolismo , Co-Represor 1 de Receptor Nuclear/fisiología , Fosfatidilinositol 3-Quinasas/genética , Fosfatidilinositol 3-Quinasas/metabolismo , Transcripción Genética/efectos de los fármacos , Proteína de Clasificación Vacuolar VPS15/genética , Proteína de Clasificación Vacuolar VPS15/metabolismo , Proteína de Clasificación Vacuolar VPS15/fisiología
20.
PLoS One ; 14(11): e0224132, 2019.
Artículo en Inglés | MEDLINE | ID: mdl-31697708

RESUMEN

Succinate dehydrogenase (SDH) inhibitors (SDHIs) are used worldwide to limit the proliferation of molds on plants and plant products. However, as SDH, also known as respiratory chain (RC) complex II, is a universal component of mitochondria from living organisms, highly conserved through evolution, the specificity of these inhibitors toward fungi warrants investigation. We first establish that the human, honeybee, earthworm and fungal SDHs are all sensitive to the eight SDHIs tested, albeit with varying IC50 values, generally in the micromolar range. In addition to SDH, we observed that five of the SDHIs, mostly from the latest generation, inhibit the activity of RC complex III. Finally, we show that the provision of glucose ad libitum in the cell culture medium, while simultaneously providing sufficient ATP and reducing power for antioxidant enzymes through glycolysis, allows the growth of RC-deficient cells, fully masking the deleterious effect of SDHIs. As a result, when glutamine is the major carbon source, the presence of SDHIs leads to time-dependent cell death. This process is significantly accelerated in fibroblasts derived from patients with neurological or neurodegenerative diseases due to RC impairment (encephalopathy originating from a partial SDH defect) and/or hypersensitivity to oxidative insults (Friedreich ataxia, familial Alzheimer's disease).


Asunto(s)
Transporte de Electrón/efectos de los fármacos , Plaguicidas/farmacología , Succinato Deshidrogenasa/antagonistas & inhibidores , Animales , Antioxidantes/metabolismo , Abejas/metabolismo , Células Cultivadas , Farmacorresistencia Fúngica/efectos de los fármacos , Proteínas Fúngicas/farmacología , Hongos/metabolismo , Humanos , Membranas Mitocondriales/efectos de los fármacos , Enfermedades Neurodegenerativas/tratamiento farmacológico , Enfermedades Neurodegenerativas/metabolismo , Oligoquetos/metabolismo , Succinato Deshidrogenasa/metabolismo
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