RESUMEN
BACKGROUND: Autophagy is the major intracellular degradation route in mammalian cells. Systemic ablation of core autophagy-related (ATG) genes in mice leads to embryonic or perinatal lethality, and conditional models show neurodegeneration. Impaired autophagy has been associated with a range of complex human diseases, yet congenital autophagy disorders are rare. METHODS: We performed a genetic, clinical, and neuroimaging analysis involving five families. Mechanistic investigations were conducted with the use of patient-derived fibroblasts, skeletal muscle-biopsy specimens, mouse embryonic fibroblasts, and yeast. RESULTS: We found deleterious, recessive variants in human ATG7, a core autophagy-related gene encoding a protein that is indispensable to classical degradative autophagy. Twelve patients from five families with distinct ATG7 variants had complex neurodevelopmental disorders with brain, muscle, and endocrine involvement. Patients had abnormalities of the cerebellum and corpus callosum and various degrees of facial dysmorphism. These patients have survived with impaired autophagic flux arising from a diminishment or absence of ATG7 protein. Although autophagic sequestration was markedly reduced, evidence of basal autophagy was readily identified in fibroblasts and skeletal muscle with loss of ATG7. Complementation of different model systems by deleterious ATG7 variants resulted in poor or absent autophagic function as compared with the reintroduction of wild-type ATG7. CONCLUSIONS: We identified several patients with a neurodevelopmental disorder who have survived with a severe loss or complete absence of ATG7, an essential effector enzyme for autophagy without a known functional paralogue. (Funded by the Wellcome Centre for Mitochondrial Research and others.).
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Anomalías Múltiples/genética , Ataxia/genética , Proteína 7 Relacionada con la Autofagia/genética , Autofagia/genética , Discapacidades del Desarrollo/genética , Mutación Missense , Adolescente , Adulto , Autofagia/fisiología , Proteína 7 Relacionada con la Autofagia/fisiología , Células Cultivadas , Cerebelo/anomalías , Simulación por Computador , Cara/anomalías , Femenino , Fibroblastos , Genes Recesivos , Humanos , Lactante , Masculino , Músculo Esquelético/metabolismo , Músculo Esquelético/patología , Malformaciones del Sistema Nervioso/genética , Linaje , FenotipoRESUMEN
BCS1L encodes a homolog of the Saccharomyces cerevisiae bcs1 protein, which has a known role in the assembly of Complex III of the mitochondrial respiratory chain. Phenotypes reported in association with pathogenic BCS1L variants include growth retardation, aminoaciduria, cholestasis, iron overload, lactic acidosis and early death (GRACILE syndrome), and Björnstad syndrome, characterized by abnormal flattening and twisting of hair shafts (pili torti) and hearing problems. Here we describe two patients harbouring biallelic variants in BCS1L; the first with a heterozygous variant c.166C>T, p.(Arg56*) together with a novel heterozygous variant c.205C>T, p.(Arg69Cys) and a second patient with a novel homozygous c.325C>T, p.(Arg109Trp) variant. The two patients presented with different phenotypes; the first patient presented as an adult with aminoaciduria, seizures, bilateral sensorineural deafness and learning difficulties. The second patient was an infant who presented with a classical GRACILE syndrome leading to death at 4 months of age. A decrease in BCS1L protein levels was seen in both patients, and biochemical analysis of Complex III revealed normal respiratory chain enzyme activities in the muscle of both patients. A decrease in Complex III assembly was detected in the adult patient's muscle, whilst the paediatric patient displayed a combined mitochondrial respiratory chain defect in cultured fibroblasts. Yeast complementation studies indicate that the two missense variants, c.205C>T, p.(Arg69Cys) and c.325C>T, p.(Arg109Trp), impair the respiratory capacity of the cell. Together, these data support the pathogenicity of the novel BCS1L variants identified in our patients.
Asunto(s)
ATPasas Asociadas con Actividades Celulares Diversas/genética , Complejo III de Transporte de Electrones/genética , Enfermedades Mitocondriales/genética , ATPasas Asociadas con Actividades Celulares Diversas/metabolismo , Acidosis Láctica/genética , Adulto , Secuencia de Aminoácidos , Colestasis/genética , Complejo III de Transporte de Electrones/metabolismo , Femenino , Retardo del Crecimiento Fetal/genética , Fibroblastos/metabolismo , Hemosiderosis/genética , Humanos , Lactante , Masculino , Errores Innatos del Metabolismo/genética , Enfermedades Mitocondriales/congénito , Músculo Esquelético/citología , Músculo Esquelético/metabolismo , Mutación , Fenotipo , Aminoacidurias Renales/genéticaRESUMEN
Recessively inherited variants in AARS2 (NM_020745.2) encoding mitochondrial alanyl-tRNA synthetase (mt-AlaRS) were first described in patients presenting with fatal infantile cardiomyopathy and multiple oxidative phosphorylation defects. To date, all described patients with AARS2-related fatal infantile cardiomyopathy are united by either a homozygous or compound heterozygous c.1774C>T (p.Arg592Trp) missense founder mutation that is absent in patients with other AARS2-related phenotypes. We describe the clinical, biochemical and molecular investigations of two unrelated boys presenting with fatal infantile cardiomyopathy, lactic acidosis and respiratory failure. Oxidative histochemistry showed cytochrome c oxidase-deficient fibres in skeletal and cardiac muscle. Biochemical studies showed markedly decreased activities of mitochondrial respiratory chain complexes I and IV with a mild decrease of complex III activity in skeletal and cardiac muscle. Using next-generation sequencing, we identified a c.1738C>T (p.Arg580Trp) AARS2 variant shared by both patients that was in trans with a loss-of-function heterozygous AARS2 variant; a c.1008dupT (p.Asp337*) nonsense variant or an intragenic deletion encompassing AARS2 exons 5-7. Interestingly, our patients did not harbour the p.Arg592Trp AARS2 founder mutation. In silico modelling of the p.Arg580Trp substitution suggested a deleterious impact on protein stability and folding. We confirmed markedly decreased mt-AlaRS protein levels in patient fibroblasts, skeletal and cardiac muscle, although mitochondrial protein synthesis defects were confined to skeletal and cardiac muscle. In vitro data showed that the p.Arg580Trp variant had a minimal effect on activation, aminoacylation or misaminoacylation activities relative to wild-type mt-AlaRS, demonstrating that instability of mt-AlaRS is the biological mechanism underlying the fatal cardiomyopathy phenotype in our patients.
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Alanina-ARNt Ligasa/metabolismo , Cardiomiopatías/enzimología , Alanina-ARNt Ligasa/genética , Cardiomiopatías/genética , Enfermedades en Gemelos/genética , Estabilidad de Enzimas , Fibroblastos/metabolismo , Genes Recesivos , Humanos , Lactante , Ácido Láctico , Masculino , Mitocondrias/metabolismo , Proteínas Mitocondriales/biosíntesis , Músculo Esquelético/metabolismo , Miocardio/metabolismo , Linaje , Insuficiencia Respiratoria/enzimologíaRESUMEN
The tRNA synthetases catalyze the first step of protein synthesis and have increasingly been studied for their nuclear and extra-cellular ex-translational activities. Human genetic conditions such as Charcot-Marie-Tooth have been attributed to dominant gain-of-function mutations in some tRNA synthetases. Unlike dominantly inherited gain-of-function mutations, recessive loss-of-function mutations can potentially elucidate ex-translational activities. We present here five individuals from four families with a multi-system disease associated with bi-allelic mutations in FARSB that encodes the beta chain of the alpha2beta2 phenylalanine-tRNA synthetase (FARS). Collectively, the mutant alleles encompass a 5'-splice junction non-coding variant (SJV) and six missense variants, one of which is shared by unrelated individuals. The clinical condition is characterized by interstitial lung disease, cerebral aneurysms and brain calcifications, and cirrhosis. For the SJV, we confirmed exon skipping leading to a frameshift associated with noncatalytic activity. While the bi-allelic combination of the SJV with a p.Arg305Gln missense mutation in two individuals led to severe disease, cells from neither the asymptomatic heterozygous carriers nor the compound heterozygous affected individual had any defect in protein synthesis. These results support a disease mechanism independent of tRNA synthetase activities in protein translation and suggest that this FARS activity is essential for normal function in multiple organs.
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Aminoacil-ARNt Sintetasas/genética , Enfermedades Pulmonares/genética , Mutación/genética , Adolescente , Alelos , Enfermedad de Charcot-Marie-Tooth/genética , Preescolar , Femenino , Genes Recesivos/genética , Heterocigoto , Humanos , Lactante , Masculino , Biosíntesis de Proteínas/genéticaRESUMEN
ATP synthase, H+ transporting, mitochondrial F1 complex, δ subunit (ATP5F1D; formerly ATP5D) is a subunit of mitochondrial ATP synthase and plays an important role in coupling proton translocation and ATP production. Here, we describe two individuals, each with homozygous missense variants in ATP5F1D, who presented with episodic lethargy, metabolic acidosis, 3-methylglutaconic aciduria, and hyperammonemia. Subject 1, homozygous for c.245C>T (p.Pro82Leu), presented with recurrent metabolic decompensation starting in the neonatal period, and subject 2, homozygous for c.317T>G (p.Val106Gly), presented with acute encephalopathy in childhood. Cultured skin fibroblasts from these individuals exhibited impaired assembly of F1FO ATP synthase and subsequent reduced complex V activity. Cells from subject 1 also exhibited a significant decrease in mitochondrial cristae. Knockdown of Drosophila ATPsynδ, the ATP5F1D homolog, in developing eyes and brains caused a near complete loss of the fly head, a phenotype that was fully rescued by wild-type human ATP5F1D. In contrast, expression of the ATP5F1D c.245C>T and c.317T>G variants rescued the head-size phenotype but recapitulated the eye and antennae defects seen in other genetic models of mitochondrial oxidative phosphorylation deficiency. Our data establish c.245C>T (p.Pro82Leu) and c.317T>G (p.Val106Gly) in ATP5F1D as pathogenic variants leading to a Mendelian mitochondrial disease featuring episodic metabolic decompensation.
Asunto(s)
Alelos , Enfermedades Metabólicas/genética , ATPasas de Translocación de Protón Mitocondriales/genética , Mutación/genética , Subunidades de Proteína/genética , Secuencia de Aminoácidos , Secuencia de Bases , Niño , Preescolar , Femenino , Humanos , Lactante , Recién Nacido , Mutación con Pérdida de Función/genética , Masculino , Mitocondrias/metabolismo , Mitocondrias/ultraestructura , ATPasas de Translocación de Protón Mitocondriales/química , Subunidades de Proteína/químicaRESUMEN
LonP1 is a mitochondrial matrix protease whose selective substrate specificity is essential for maintaining mitochondrial homeostasis. Recessively inherited, pathogenic defects in LonP1 have been previously reported to underlie cerebral, ocular, dental, auricular and skeletal anomalies (CODAS) syndrome, a complex multisystemic and developmental disorder. Intriguingly, although classical mitochondrial disease presentations are well-known to exhibit marked clinical heterogeneity, the skeletal and dental features associated with CODAS syndrome are pathognomonic. We have applied whole exome sequencing to a patient with congenital lactic acidosis, muscle weakness, profound deficiencies in mitochondrial oxidative phosphorylation associated with loss of mtDNA copy number and MRI abnormalities consistent with Leigh syndrome, identifying biallelic variants in the LONP1 (NM_004793.3) gene; c.1693T > C predicting p.(Tyr565His) and c.2197G > A predicting p.(Glu733Lys); no evidence of the classical skeletal or dental defects observed in CODAS syndrome patients were noted in our patient. In vitro experiments confirmed the p.(Tyr565His) LonP1 mutant alone could not bind or degrade a substrate, consistent with the predicted function of Tyr565, whilst a second missense [p.(Glu733Lys)] variant had minimal effect. Mixtures of p.(Tyr565His) mutant and wild-type LonP1 retained partial protease activity but this was severely depleted when the p.(Tyr565His) mutant was mixed with the p.(Glu733Lys) mutant, data consistent with the compound heterozygosity detected in our patient. In summary, we conclude that pathogenic LONP1 variants can lead to a classical mitochondrial disease presentations associated with severe biochemical defects in oxidative phosphorylation in clinically relevant tissues.
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Proteasas ATP-Dependientes/genética , Anomalías Craneofaciales/genética , Anomalías del Ojo/genética , Trastornos del Crecimiento/genética , Luxación Congénita de la Cadera/genética , Enfermedad de Leigh/genética , Enfermedades Mitocondriales/genética , Proteínas Mitocondriales/genética , Osteocondrodisplasias/genética , Anomalías Dentarias/genética , Biopsia , Línea Celular , Anomalías Craneofaciales/metabolismo , Anomalías Craneofaciales/fisiopatología , Exoma/genética , Anomalías del Ojo/metabolismo , Anomalías del Ojo/fisiopatología , Trastornos del Crecimiento/metabolismo , Trastornos del Crecimiento/fisiopatología , Luxación Congénita de la Cadera/metabolismo , Luxación Congénita de la Cadera/fisiopatología , Humanos , Lactante , Enfermedad de Leigh/metabolismo , Enfermedad de Leigh/fisiopatología , Masculino , Mitocondrias/genética , Mitocondrias/patología , Enfermedades Mitocondriales/metabolismo , Enfermedades Mitocondriales/fisiopatología , Músculo Esquelético/fisiopatología , Mutación , Osteocondrodisplasias/metabolismo , Osteocondrodisplasias/fisiopatología , Fosforilación Oxidativa , Anomalías Dentarias/metabolismo , Anomalías Dentarias/fisiopatología , Secuenciación del ExomaRESUMEN
Complement component 1 Q subcomponent-binding protein (C1QBP; also known as p32) is a multi-compartmental protein whose precise function remains unknown. It is an evolutionary conserved multifunctional protein localized primarily in the mitochondrial matrix and has roles in inflammation and infection processes, mitochondrial ribosome biogenesis, and regulation of apoptosis and nuclear transcription. It has an N-terminal mitochondrial targeting peptide that is proteolytically processed after import into the mitochondrial matrix, where it forms a homotrimeric complex organized in a doughnut-shaped structure. Although C1QBP has been reported to exert pleiotropic effects on many cellular processes, we report here four individuals from unrelated families where biallelic mutations in C1QBP cause a defect in mitochondrial energy metabolism. Infants presented with cardiomyopathy accompanied by multisystemic involvement (liver, kidney, and brain), and children and adults presented with myopathy and progressive external ophthalmoplegia. Multiple mitochondrial respiratory-chain defects, associated with the accumulation of multiple deletions of mitochondrial DNA in the later-onset myopathic cases, were identified in all affected individuals. Steady-state C1QBP levels were decreased in all individuals' samples, leading to combined respiratory-chain enzyme deficiency of complexes I, III, and IV. C1qbp-/- mouse embryonic fibroblasts (MEFs) resembled the human disease phenotype by showing multiple defects in oxidative phosphorylation (OXPHOS). Complementation with wild-type, but not mutagenized, C1qbp restored OXPHOS protein levels and mitochondrial enzyme activities in C1qbp-/- MEFs. C1QBP deficiency represents an important mitochondrial disorder associated with a clinical spectrum ranging from infantile lactic acidosis to childhood (cardio)myopathy and late-onset progressive external ophthalmoplegia.
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Cardiomiopatías/genética , Proteínas Portadoras/genética , Transporte de Electrón/fisiología , Enfermedades Mitocondriales/genética , Proteínas Mitocondriales/genética , Mutación , Adulto , Edad de Inicio , Anciano , Alelos , Secuencia de Aminoácidos , Animales , Cardiomiopatías/complicaciones , Cardiomiopatías/patología , Proteínas Portadoras/química , Proteínas Portadoras/metabolismo , Células Cultivadas , Preescolar , Estudios de Cohortes , ADN Mitocondrial , Embrión de Mamíferos/metabolismo , Embrión de Mamíferos/patología , Femenino , Fibroblastos/metabolismo , Fibroblastos/patología , Humanos , Recién Nacido , Masculino , Ratones , Persona de Mediana Edad , Enfermedades Mitocondriales/complicaciones , Enfermedades Mitocondriales/patología , Proteínas Mitocondriales/química , Proteínas Mitocondriales/metabolismo , Fosforilación Oxidativa , Linaje , Conformación Proteica , Homología de Secuencia , Índice de Severidad de la Enfermedad , Adulto JovenRESUMEN
Mitochondrial disease is hugely diverse with respect to associated clinical presentations and underlying genetic causes, with pathogenic variants in over 300 disease genes currently described. Approximately half of these have been discovered in the last decade due to the increasingly widespread application of next generation sequencing technologies, in particular unbiased, whole exome-and latterly, whole genome sequencing. These technologies allow more genetic data to be collected from patients with mitochondrial disorders, continually improving the diagnostic success rate in a clinical setting. Despite these significant advances, some patients still remain without a definitive genetic diagnosis. Large datasets containing many variants of unknown significance have become a major challenge with next generation sequencing strategies and these require significant functional validation to confirm pathogenicity. This interface between diagnostics and research is critical in continuing to expand the list of known pathogenic variants and concomitantly enhance our knowledge of mitochondrial biology. The increasing use of whole exome sequencing, whole genome sequencing and other "omics" techniques such as transcriptomics and proteomics will generate even more data and allow further interrogation and validation of genetic causes, including those outside of coding regions. This will improve diagnostic yields still further and emphasizes the integral role that functional assessment of variant causality plays in this process-the overarching focus of this review.
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Secuenciación del Exoma/métodos , Genoma Mitocondrial , Secuenciación de Nucleótidos de Alto Rendimiento/métodos , Enfermedades Mitocondriales/diagnóstico , Técnicas de Diagnóstico Molecular , Humanos , Enfermedades Mitocondriales/genética , Análisis de Secuencia de ARN , TranscriptomaRESUMEN
The assembly of mitochondrial oxidative phosphorylation (OXPHOS) complexes is an intricate process, which-given their dual-genetic control-requires tight co-regulation of two evolutionarily distinct gene expression machineries. Moreover, fine-tuning protein synthesis to the nascent assembly of OXPHOS complexes requires regulatory mechanisms such as translational plasticity and translational activators that can coordinate mitochondrial translation with the import of nuclear-encoded mitochondrial proteins. The intricacy of OXPHOS complex biogenesis is further evidenced by the requirement of many tightly orchestrated steps and ancillary factors. Early-stage ancillary chaperones have essential roles in coordinating OXPHOS assembly, whilst late-stage assembly factors-also known as the LYRM (leucine-tyrosine-arginine motif) proteins-together with the mitochondrial acyl carrier protein (ACP)-regulate the incorporation and activation of late-incorporating OXPHOS subunits and/or co-factors. In this review, we describe recent discoveries providing insights into the mechanisms required for optimal OXPHOS biogenesis, including the coordination of mitochondrial gene expression with the availability of nuclear-encoded factors entering via mitochondrial protein import systems.
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Complejo IV de Transporte de Electrones/metabolismo , Proteínas Mitocondriales/metabolismo , Proteínas Nucleares/metabolismo , Biogénesis de Organelos , Fosforilación Oxidativa , Transporte de Proteínas , Activación Transcripcional , Secuencias de Aminoácidos , Animales , Núcleo Celular/metabolismo , Citosol/metabolismo , ADN Mitocondrial/metabolismo , Regulación de la Expresión Génica , Humanos , Ratones , Mitocondrias/metabolismo , Biosíntesis de Proteínas , Dominios Proteicos , Especificidad de la EspecieRESUMEN
Complex I deficiency is the most common biochemical phenotype observed in individuals with mitochondrial disease. With 44 structural subunits and over 10 assembly factors, it is unsurprising that complex I deficiency is associated with clinical and genetic heterogeneity. Massively parallel sequencing (MPS) technologies including custom, targeted gene panels or unbiased whole-exome sequencing (WES) are hugely powerful in identifying the underlying genetic defect in a clinical diagnostic setting, yet many individuals remain without a genetic diagnosis. These individuals might harbor mutations in poorly understood or uncharacterized genes, and their diagnosis relies upon characterization of these orphan genes. Complexome profiling recently identified TMEM126B as a component of the mitochondrial complex I assembly complex alongside proteins ACAD9, ECSIT, NDUFAF1, and TIMMDC1. Here, we describe the clinical, biochemical, and molecular findings in six cases of mitochondrial disease from four unrelated families affected by biallelic (c.635G>T [p.Gly212Val] and/or c.401delA [p.Asn134Ilefs(∗)2]) TMEM126B variants. We provide functional evidence to support the pathogenicity of these TMEM126B variants, including evidence of founder effects for both variants, and establish defects within this gene as a cause of complex I deficiency in association with either pure myopathy in adulthood or, in one individual, a severe multisystem presentation (chronic renal failure and cardiomyopathy) in infancy. Functional experimentation including viral rescue and complexome profiling of subject cell lines has confirmed TMEM126B as the tenth complex I assembly factor associated with human disease and validates the importance of both genome-wide sequencing and proteomic approaches in characterizing disease-associated genes whose physiological roles have been previously undetermined.
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Alelos , Complejo I de Transporte de Electrón/deficiencia , Proteínas de la Membrana/genética , Enfermedades Mitocondriales/genética , Mutación/genética , Fenotipo , Adolescente , Adulto , Edad de Inicio , Secuencia de Aminoácidos , Niño , Complejo I de Transporte de Electrón/genética , Femenino , Humanos , Lactante , Masculino , Proteínas de la Membrana/química , Persona de Mediana Edad , Linaje , Adulto JovenRESUMEN
In this issue of Molecular Cell, Molin et al. (2011) reveal that caloric restriction alleviates PKA-dependent inhibition of sulfiredoxin translation, maintaining the thioredoxin peroxidase activity of a peroxiredoxin and increasing the hydrogen peroxide resistance and replicative life span of Saccharomyces cerevisiae.
RESUMEN
PURPOSE: To understand the role of the mitochondrial oxodicarboxylate carrier (SLC25A21) in the development of spinal muscular atrophy-like disease. METHODS: We identified a novel pathogenic variant in a patient by whole-exome sequencing. The pathogenicity of the mutation was studied by transport assays, computer modeling, followed by targeted metabolic testing and in vitro studies in human fibroblasts and neurons. RESULTS: The patient carries a homozygous pathogenic variant c.695A>G; p.(Lys232Arg) in the SLC25A21 gene, encoding the mitochondrial oxodicarboxylate carrier, and developed spinal muscular atrophy and mitochondrial myopathy. Transport assays show that the mutation renders SLC25A21 dysfunctional and 2-oxoadipate cannot be imported into the mitochondrial matrix. Computer models of central metabolism predicted that impaired transport of oxodicarboxylate disrupts the pathways of lysine and tryptophan degradation, and causes accumulation of 2-oxoadipate, pipecolic acid, and quinolinic acid, which was confirmed in the patient's urine by targeted metabolomics. Exposure to 2-oxoadipate and quinolinic acid decreased the level of mitochondrial complexes in neuronal cells (SH-SY5Y) and induced apoptosis. CONCLUSION: Mitochondrial oxodicarboxylate carrier deficiency leads to mitochondrial dysfunction and the accumulation of oxoadipate and quinolinic acid, which in turn cause toxicity in spinal motor neurons leading to spinal muscular atrophy-like disease.
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Adipatos/metabolismo , ADN Mitocondrial/genética , Transportadores de Ácidos Dicarboxílicos/genética , Proteínas de Transporte de Membrana Mitocondrial/genética , Atrofia Muscular Espinal/genética , Adipatos/farmacología , Apoptosis/efectos de los fármacos , Línea Celular , ADN Mitocondrial/metabolismo , Transportadores de Ácidos Dicarboxílicos/metabolismo , Fibroblastos/efectos de los fármacos , Homocigoto , Humanos , Mitocondrias/genética , Mitocondrias/metabolismo , Proteínas de Transporte de Membrana Mitocondrial/metabolismo , Neuronas Motoras/efectos de los fármacos , Atrofia Muscular Espinal/metabolismo , Atrofia Muscular Espinal/fisiopatología , Mutación , Ácidos Pipecólicos/metabolismo , Ácido Quinolínico/metabolismoRESUMEN
Mitochondrial diseases collectively represent one of the most heterogeneous group of metabolic disorders. Symptoms can manifest at any age, presenting with isolated or multiple-organ involvement. Advances in next-generation sequencing strategies have greatly enhanced the diagnosis of patients with mitochondrial disease, particularly where a mitochondrial aetiology is strongly suspected yet OXPHOS activities in biopsied tissue samples appear normal. We used whole exome sequencing (WES) to identify the molecular basis of an early-onset mitochondrial syndrome-pathogenic biallelic variants in the HTRA2 gene, encoding a mitochondria-localised serine protease-in five subjects from two unrelated families characterised by seizures, neutropenia, hypotonia and cardio-respiratory problems. A unifying feature in all affected children was 3-methylglutaconic aciduria (3-MGA-uria), a common biochemical marker observed in some patients with mitochondrial dysfunction. Although functional studies of HTRA2 subjects' fibroblasts and skeletal muscle homogenates showed severely decreased levels of mutant HTRA2 protein, the structural subunits and complexes of the mitochondrial respiratory chain appeared normal. We did detect a profound defect in OPA1 processing in HTRA2-deficient fibroblasts, suggesting a role for HTRA2 in the regulation of mitochondrial dynamics and OPA1 proteolysis. In addition, investigated subject fibroblasts were more susceptible to apoptotic insults. Our data support recent studies that described important functions for HTRA2 in programmed cell death and confirm that patients with genetically-unresolved 3-MGA-uria should be screened by WES with pathogenic variants in the HTRA2 gene prioritised for further analysis.
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Variación Genética/genética , Serina Peptidasa A2 que Requiere Temperaturas Altas/genética , Errores Innatos del Metabolismo/genética , Mitocondrias/genética , Enfermedades Mitocondriales/genética , Muerte Celular/genética , Células Cultivadas , Niño , Exoma/genética , Femenino , Fibroblastos/metabolismo , Humanos , Masculino , Proteínas Mitocondriales/genética , Músculo Esquelético/metabolismo , Serina Proteasas/genética , SíndromeRESUMEN
BACKGROUND: Isolated Complex I deficiency is the most common paediatric mitochondrial disease presentation, associated with poor prognosis and high mortality. Complex I comprises 44 structural subunits with at least 10 ancillary proteins; mutations in 29 of these have so far been associated with mitochondrial disease but there are limited genotype-phenotype correlations to guide clinicians to the correct genetic diagnosis. METHODS: Patients were analysed by whole-exome sequencing, targeted capture or candidate gene sequencing. Clinical phenotyping of affected individuals was performed. RESULTS: We identified a cohort of 10 patients from 8 families (7 families are of unrelated Irish ancestry) all of whom have short stature (<9th centile) and similar facial features including a prominent forehead, smooth philtrum and deep-set eyes associated with a recurrent homozygous c.64T>C, p.Trp22Arg NDUFB3 variant. Two sibs presented with primary short stature without obvious metabolic dysfunction. Analysis of skeletal muscle from three patients confirmed a defect in Complex I assembly. CONCLUSIONS: Our report highlights that the long-term prognosis related to the p.Trp22Arg NDUFB3 mutation can be good, even for some patients presenting in acute metabolic crisis with evidence of an isolated Complex I deficiency in muscle. Recognition of the distinctive facial features-particularly when associated with markers of mitochondrial dysfunction and/or Irish ancestry-should suggest screening for the p.Trp22Arg NDUFB3 mutation to establish a genetic diagnosis, circumventing the requirement of muscle biopsy to direct genetic investigations.
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Enanismo/genética , Complejo I de Transporte de Electrón/genética , Mitocondrias/genética , Enfermedades Mitocondriales/genética , Mutación/genética , Niño , Preescolar , Exoma/genética , Facies , Femenino , Estudios de Asociación Genética/métodos , Homocigoto , Humanos , Lactante , Masculino , Linaje , FenotipoRESUMEN
Mitochondrial Complex IV [cytochrome c oxidase (COX)] deficiency is one of the most common respiratory chain defects in humans. The clinical phenotypes associated with COX deficiency include liver disease, cardiomyopathy and Leigh syndrome, a neurodegenerative disorder characterized by bilateral high signal lesions in the brainstem and basal ganglia. COX deficiency can result from mutations affecting many different mitochondrial proteins. The French-Canadian variant of COX-deficient Leigh syndrome is unique to the Saguenay-Lac-Saint-Jean region of Québec and is caused by a founder mutation in the LRPPRC gene. This encodes the leucine-rich pentatricopeptide repeat domain protein (LRPPRC), which is involved in post-transcriptional regulation of mitochondrial gene expression. Here, we present the clinical and molecular characterization of novel, recessive LRPPRC gene mutations, identified using whole exome and candidate gene sequencing. The 10 patients come from seven unrelated families of UK-Caucasian, UK-Pakistani, UK-Indian, Turkish and Iraqi origin. They resemble the French-Canadian Leigh syndrome patients in having intermittent severe lactic acidosis and early-onset neurodevelopmental problems with episodes of deterioration. In addition, many of our patients have had neonatal cardiomyopathy or congenital malformations, most commonly affecting the heart and the brain. All patients who were tested had isolated COX deficiency in skeletal muscle. Functional characterization of patients' fibroblasts and skeletal muscle homogenates showed decreased levels of mutant LRPPRC protein and impaired Complex IV enzyme activity, associated with abnormal COX assembly and reduced steady-state levels of numerous oxidative phosphorylation subunits. We also identified a Complex I assembly defect in skeletal muscle, indicating different roles for LRPPRC in post-transcriptional regulation of mitochondrial mRNAs between tissues. Patient fibroblasts showed decreased steady-state levels of mitochondrial mRNAs, although the length of poly(A) tails of mitochondrial transcripts were unaffected. Our study identifies LRPPRC as an important disease-causing gene in an early-onset, multisystem and neurological mitochondrial disease, which should be considered as a cause of COX deficiency even in patients originating outside of the French-Canadian population.
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Deficiencia de Citocromo-c Oxidasa/genética , Enfermedades Mitocondriales/genética , Proteínas de Neoplasias/genética , Proteínas/genética , Canadá , Células Cultivadas , Preescolar , Deficiencia de Citocromo-c Oxidasa/enzimología , Complejo IV de Transporte de Electrones/metabolismo , Femenino , Fibroblastos/metabolismo , Humanos , Lactante , Recién Nacido , Proteínas Repetidas Ricas en Leucina , Masculino , Enfermedades Mitocondriales/enzimología , Enfermedades Mitocondriales/metabolismo , Proteínas Mitocondriales/metabolismo , Músculo Esquelético/metabolismo , Mutación , Linaje , Proteínas/metabolismo , ARN Mensajero/metabolismo , ARN MitocondrialRESUMEN
Succinate dehydrogenase (SDH) is a crucial metabolic enzyme complex that is involved in ATP production, playing roles in both the tricarboxylic cycle and the mitochondrial respiratory chain (complex II). Isolated complex II deficiency is one of the rarest oxidative phosphorylation disorders with mutations described in three structural subunits and one of the assembly factors; just one case is attributed to recessively inherited SDHD mutations. We report the pathological, biochemical, histochemical and molecular genetic investigations of a male neonate who had left ventricular hypertrophy detected on antenatal scan and died on day one of life. Subsequent postmortem examination confirmed hypertrophic cardiomyopathy with left ventricular non-compaction. Biochemical analysis of his skeletal muscle biopsy revealed evidence of a severe isolated complex II deficiency and candidate gene sequencing revealed a novel homozygous c.275A>G, p.(Asp92Gly) SDHD mutation which was shown to be recessively inherited through segregation studies. The affected amino acid has been reported as a Dutch founder mutation p.(Asp92Tyr) in families with hereditary head and neck paraganglioma. By introducing both mutations into Saccharomyces cerevisiae, we were able to confirm that the p.(Asp92Gly) mutation causes a more severe oxidative growth phenotype than the p.(Asp92Tyr) mutant, and provides functional evidence to support the pathogenicity of the patient's SDHD mutation. This is only the second case of mitochondrial complex II deficiency due to inherited SDHD mutations and highlights the importance of sequencing all SDH genes in patients with biochemical and histochemical evidence of isolated mitochondrial complex II deficiency.
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Cardiomiopatía Hipertrófica Familiar/genética , Genes Recesivos , Cardiopatías Congénitas/genética , Homocigoto , Proteínas Mitocondriales/genética , Mutación Missense , Succinato Deshidrogenasa/genética , Sustitución de Aminoácidos , Cardiomiopatía Hipertrófica Familiar/enzimología , Ciclo del Ácido Cítrico/genética , Cardiopatías Congénitas/enzimología , Humanos , Recién Nacido , MasculinoRESUMEN
BACKGROUND: Phase 2 detoxification enzymes provide a vital defence against reactive oxygen species, including xenobiotic metabolites, which cause the oxidative damage involved in drug toxicity and many diseases. Hence, there is great interest in understanding how levels of these enzymes are regulated. CnC transcription factors, such as mammalian Nrf2, drive the expression of phase 2 enzymes and are activated as an important conserved response to oxidative stress and xenobiotics. For instance, the Caenorhabditis elegans Nrf2 orthologue, SKN-1, is activated in response to arsenite by the stress-activated p38-related kinase, PMK-1, leading to increased expression of phase 2 enzymes. Here we have used a genome-wide screening approach to identify other C. elegans genes that are required for stress-induced increases in phase 2 detoxification gene expression. RESULTS: Taking advantage of the elevated phase 2 gene expression in a mutant lacking the peroxidase PRDX-2, we have identified many new genes that are required for stress-induced expression of gcs-1, a phase 2 enzyme critically required for glutathione synthesis. Significantly, these include genes previously implicated in resistance to ionizing radiation, longevity and responses to pathogenic infection. Many of these new candidate activators of gcs-1 are also required for the stress-induced intestinal expression of other phase 2 genes. However, intriguingly, our data suggest other factors may be specifically required for the stress-induced expression of gcs-1. Notably, we demonstrate that the candidate activator TIR-1(SARM1) and the MAPKKK NSY-1(Ask1) are required for the arsenite-induced activation of PMK-1. However, our data suggest that the majority of candidates participate in novel mechanisms to promote gcs-1 expression. For example, the E4 ubiquitin ligase UFD-2(UBE4B) is dispensable for PMK-1 activation but important for maintaining nuclear levels of SKN-1, the stress-induced expression of multiple SKN-1-target genes and oxidative stress resistance. CONCLUSIONS: Here we present the first functional, genome-wide analysis identifying genes that are required for activation of phase 2 detoxification genes in an animal. Our study identifies potential new regulators of Nrf2, reveals that additional mechanisms promote the stress-induced expression of specific phase 2 detoxification genes and provides new insight into the relationships between these universally important stress defences, oxidative stress resistance and aging.
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Caenorhabditis elegans/genética , Regulación de la Expresión Génica/genética , Genoma de los Helmintos , Estudio de Asociación del Genoma Completo , Fase II de la Desintoxicación Metabólica/genética , Animales , Caenorhabditis elegans/metabolismo , Estrés Oxidativo , Especies Reactivas de Oxígeno/metabolismo , Estrés Fisiológico , Xenobióticos/metabolismoRESUMEN
Efforts to understand how mitochondrial dysfunction contributes to neurodegeneration have primarily focussed on the role of mitochondria in neuronal energy metabolism. However, progress in understanding the etiological nature of emerging mitochondrial functions has yielded new ideas about the mitochondrial basis of neurological disease. Studies aimed at deciphering how mitochondria signal through interorganellar contacts, vesicular trafficking, and metabolic transmission have revealed that mitochondrial regulation of immunometabolism, cell death, organelle dynamics, and neuroimmune interplay are critical determinants of neural health. Moreover, the homeostatic mechanisms that exist to protect mitochondrial health through turnover via nanoscale proteostasis and lysosomal degradation have become integrated within mitochondrial signalling pathways to support metabolic plasticity and stress responses in the nervous system. This review highlights how these distinct mitochondrial pathways converge to influence neurological health and contribute to disease pathology.
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Mitocondrias , Enfermedades del Sistema Nervioso , Humanos , Mitocondrias/metabolismo , Orgánulos/metabolismo , Homeostasis , Transducción de Señal , Enfermedades del Sistema Nervioso/metabolismoRESUMEN
Biallelic hypomorphic variants in PRORP have been recently described as causing the autosomal recessive disorder combined oxidative phosphorylation deficiency type 54 (COXPD54). COXPD54 encompasses a phenotypic spectrum of sensorineural hearing loss and ovarian insufficiency (Perrault syndrome) to leukodystrophy. Here, we report three additional families with homozygous missense PRORP variants with pleiotropic phenotypes. Each missense variant altered a highly conserved residue within the metallonuclease domain. In vitro mitochondrial tRNA processing assays with recombinant TRMT10C, SDR5C1 and PRORP indicated two COXPD54-associated PRORP variants, c.1159A>G (p.Thr387Ala) and c.1241C>T (p.Ala414Val), decreased pre-tRNAIle cleavage, consistent with both variants impacting tRNA processing. No significant decrease in tRNA processing was observed with PRORP c.1093T>C (p.Tyr365His), which was identified in an individual with leukodystrophy. These data provide independent evidence that PRORP variants are associated with COXPD54 and that the assessment of 5' leader mitochondrial tRNA processing is a valuable assay for the functional analysis and clinical interpretation of novel PRORP variants.
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Pérdida Auditiva Sensorineural , Enfermedades Mitocondriales , Ribonucleasa P , Femenino , Humanos , Genotipo , Pérdida Auditiva Sensorineural/genética , Homocigoto , Enfermedades Mitocondriales/genética , ARN de Transferencia , Ribonucleasa P/genéticaRESUMEN
The genetic architecture of mitochondrial disease continues to expand and currently exceeds more than 350 disease-causing genes. Bi-allelic variants in RTN4IP1, also known as Optic Atrophy-10 (OPA10), lead to early-onset recessive optic neuropathy, atrophy, and encephalopathy in the afflicted patients. The gene is known to encode a mitochondrial ubiquinol oxidoreductase that interacts with reticulon 4 and is thought to be a mitochondrial antioxidant NADPH oxidoreductase. Here, we describe two unrelated consanguineous families from the northern region of Saudi Arabia harboring a missense variant (RTN4IP1:NM_032730.5; c.475GAsunto(s)
Encefalopatías
, Atrofia Óptica
, Antioxidantes
, Proteínas Portadoras/genética
, Humanos
, Proteínas Mitocondriales/genética
, Mutación/genética
, NADP/genética
, Atrofia Óptica/genética
, Oxidorreductasas/genética
, Arabia Saudita