RESUMEN
Mitochondrial fission and fusion are vital dynamic processes for mitochondrial quality control and for the maintenance of cellular respiration; they also play an important role in the formation and maintenance of cells with high energy demand including cardiomyocytes and neurons. The DNM1L (dynamin-1 like) gene encodes for the DRP1 protein, an evolutionary conserved member of the dynamin family that is responsible for the fission of mitochondria; it is ubiquitous but highly expressed in the developing neonatal heart. De novo heterozygous pathogenic variants in the DNM1L gene have been previously reported to be associated with neonatal or infantile-onset encephalopathy characterized by hypotonia, developmental delay and refractory epilepsy. However, cardiac involvement has been previously reported only in one case. Next-Generation Sequencing (NGS) was used to genetically assess a baby girl characterized by developmental delay with spastic-dystonic, tetraparesis and hypertrophic cardiomyopathy of the left ventricle. Histochemical analysis and spectrophotometric determination of electron transport chain were performed to characterize the muscle biopsy; moreover, the morphology of mitochondria and peroxisomes was evaluated in cultured fibroblasts as well. Herein, we expand the phenotype of DNM1L-related disorder, describing the case of a girl with a heterozygous mutation in DNM1L and affected by progressive infantile encephalopathy, with cardiomyopathy and fatal paroxysmal vomiting correlated with bulbar transitory abnormal T2 hyperintensities and diffusion-weighted imaging (DWI) restriction areas, but without epilepsy. In patients with DNM1L mutations, careful evaluation for cardiac involvement is recommended.
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Cardiomiopatías , Dinaminas , Mutación , Humanos , Femenino , Dinaminas/genética , Cardiomiopatías/genética , Mutación/genética , Lactante , Resultado Fatal , Encefalopatías/genética , Encefalopatías/patología , GTP Fosfohidrolasas/genéticaRESUMEN
INTRODUCTION: COXPD23 is a rare mitochondrial disease caused by biallelic pathogenic variants in GTPBP3. We report on two siblings with a mild phenotype. CASE REPORTS: The young boy presented with global developmental delay, ataxic gait and upper limbs tremor, and the older sister with absence seizures and hypertrophic cardiomyopathy. Respiratory chain impairment was confirmed in muscle. DISCUSSION: Reviewed cases point toward clustering around two prevalent phenotypes: an early-onset presentation with severe fatal encephalopathy and a late milder presentation with global developmental delay/ID and cardiopathy, with the latter as, is the main feature. Our patients showed an intermediate phenotype with intrafamilial variability.
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Enfermedades Mitocondriales , Convulsiones , Masculino , Humanos , Mitocondrias , Fenotipo , Proteínas de Unión al GTPRESUMEN
The increased availability of genetic technologies has significantly improved the detection of novel germline variants conferring a predisposition to tumor development in patients with malignant disease. The identification of variants of uncertain significance (VUS) represents a challenge for the clinician, leading to difficulties in decision-making regarding medical management, the surveillance program, and genetic counseling. Moreover, it can generate confusion and anxiety for patients and their family members. Herein, we report a 5-year-old girl carrying a VUS in the Succinate Dehydrogenase Complex Subunit C (SHDC) gene who had been previously treated for high-risk neuroblastoma and subsequently followed by the development of secondary acute myeloid leukemia. In this context, we describe how functional studies can provide additional insight on gene function determining whether the variant interferes with normal protein function or stability.
RESUMEN
Genetic defect in the nuclear encoded subunits of cytochrome c oxidase are very rare. To date, most deleterious variants affect the mitochondrially encoded subunits of complex IV and the nuclear genes encoded for assembly factors. A biallelic pathogenic variant in the mitochondrial complex IV subunit COX5A was previously reported in a couple of sibs with failure to thrive, lactic acidosis and pulmonary hypertension and a lethal phenotype. Here, we describe a second family with a 11-year-old girl presenting with failure to thrive, lactic acidosis, hypoglycemia and short stature. Clinical exome revealed the homozygous missense variant c.266 T > G in COX5A, which produces a drop of the corresponding protein and a reduction of the COX activity. Compared to the previous observation, this girl showed an attenuated metabolic derangement without involvement of the cardiovascular system and neurodevelopment. Our observation confirms that COX5A recessive variants may cause mitochondrial disease and expands the associated phenotype to less severe presentations.
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Acidosis Láctica , Enanismo , Hipoglucemia , Acidosis Láctica/genética , Acidosis Láctica/patología , Complejo IV de Transporte de Electrones/genética , Complejo IV de Transporte de Electrones/metabolismo , Insuficiencia de Crecimiento/genética , Homocigoto , Humanos , Hipoglucemia/genética , FenotipoRESUMEN
Germline-activating mutations in HRAS cause Costello syndrome (CS), a cancer prone multisystem disorder characterized by reduced postnatal growth. In CS, poor weight gain and growth are not caused by low caloric intake. Here, we show that constitutive plasma membrane translocation and activation of the GLUT4 glucose transporter, via reactive oxygen species-dependent AMP-activated protein kinase α and p38 hyperactivation, occurs in primary fibroblasts of CS patients, resulting in accelerated glycolysis and increased fatty acid synthesis and storage as lipid droplets. An accelerated autophagic flux was also identified as contributing to the increased energetic expenditure in CS. Concomitant inhibition of p38 and PI3K signaling by wortmannin was able to rescue both the dysregulated glucose intake and accelerated autophagic flux. Our findings provide a mechanistic link between upregulated HRAS function, defective growth and increased resting energetic expenditure in CS, and document that targeting p38 and PI3K signaling is able to revert this metabolic dysfunction.
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Síndrome de Costello , Síndrome de Costello/genética , Síndrome de Costello/metabolismo , Fibroblastos/metabolismo , Humanos , Oxidación-Reducción , Fosfatidilinositol 3-Quinasas/genética , Fosfatidilinositol 3-Quinasas/metabolismo , Proteínas Proto-Oncogénicas p21(ras)/genética , Transducción de Señal/genéticaRESUMEN
Isolated biochemical deficiency of mitochondrial complex I is the most frequent signature among mitochondrial diseases and is associated with a wide variety of clinical symptoms. Leigh syndrome represents the most frequent neuroradiological finding in patients with complex I defect and more than 80 monogenic causes have been involved in the disease. In this report, we describe seven patients from four unrelated families harboring novel NDUFA12 variants, with six of them presenting with Leigh syndrome. Molecular genetic characterization was performed using next-generation sequencing combined with the Sanger method. Biochemical and protein studies were achieved by enzymatic activities, blue native gel electrophoresis, and western blot analysis. All patients displayed novel homozygous mutations in the NDUFA12 gene, leading to the virtual absence of the corresponding protein. Surprisingly, despite the fact that in none of the analyzed patients, NDUFA12 protein was detected, they present a different onset and clinical course of the disease. Our report expands the array of genetic alterations in NDUFA12 and underlines phenotype variability associated with NDUFA12 defect.
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Enfermedad de Leigh/genética , Enfermedades Mitocondriales/genética , NADPH Deshidrogenasa/genética , Adolescente , Niño , Preescolar , Estudios de Cohortes , Consanguinidad , Complejo I de Transporte de Electrón/genética , Familia , Femenino , Predisposición Genética a la Enfermedad , Humanos , Italia , Enfermedad de Leigh/complicaciones , Enfermedad de Leigh/patología , Masculino , Enfermedades Mitocondriales/complicaciones , Enfermedades Mitocondriales/patología , Fenotipo , Polimorfismo de Nucleótido SimpleRESUMEN
Mitochondria do not exist as individual entities in the cell-conversely, they constitute an interconnected community governed by the constant and opposite process of fission and fusion. The mitochondrial fission leads to the formation of smaller mitochondria, promoting the biogenesis of new organelles. On the other hand, following the fusion process, mitochondria appear as longer and interconnected tubules, which enhance the communication with other organelles. Both fission and fusion are carried out by a small number of highly conserved guanosine triphosphatase proteins and their interactors. Disruption of this equilibrium has been associated with several pathological conditions, ranging from cancer to neurodegeneration, and mutations in genes involved in mitochondrial fission and fusion have been reported to be the cause of a subset of neurogenetic disorders.
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Regulación de la Expresión Génica/efectos de los fármacos , Mitocondrias/efectos de los fármacos , Enfermedades Mitocondriales/tratamiento farmacológico , Neoplasias/tratamiento farmacológico , Enfermedades Neurodegenerativas/tratamiento farmacológico , Sustancias Protectoras/uso terapéutico , ATPasas Asociadas con Actividades Celulares Diversas/genética , ATPasas Asociadas con Actividades Celulares Diversas/metabolismo , Dinaminas/genética , Dinaminas/metabolismo , GTP Fosfohidrolasas/genética , GTP Fosfohidrolasas/metabolismo , Humanos , Proteínas de la Membrana/genética , Proteínas de la Membrana/metabolismo , Metaloendopeptidasas/genética , Metaloendopeptidasas/metabolismo , Mitocondrias/metabolismo , Mitocondrias/patología , Enfermedades Mitocondriales/genética , Enfermedades Mitocondriales/metabolismo , Enfermedades Mitocondriales/patología , Dinámicas Mitocondriales/efectos de los fármacos , Proteínas Mitocondriales/genética , Proteínas Mitocondriales/metabolismo , Neoplasias/genética , Neoplasias/metabolismo , Neoplasias/patología , Enfermedades Neurodegenerativas/genética , Enfermedades Neurodegenerativas/metabolismo , Enfermedades Neurodegenerativas/patología , Biogénesis de Organelos , Factores de Elongación de Péptidos/genética , Factores de Elongación de Péptidos/metabolismo , Transducción de SeñalRESUMEN
Dysfunction of the endolysosomal system is often associated with neurodegenerative disease because postmitotic neurons are particularly reliant on the elimination of intracellular aggregates. Adequate function of endosomes and lysosomes requires finely tuned luminal ion homeostasis and transmembrane ion fluxes. Endolysosomal CLC Cl-/H+ exchangers function as electric shunts for proton pumping and in luminal Cl- accumulation. We now report three unrelated children with severe neurodegenerative disease, who carry the same de novo c.1658A>G (p.Tyr553Cys) mutation in CLCN6, encoding the late endosomal Cl-/H+-exchanger ClC-6. Whereas Clcn6-/- mice have only mild neuronal lysosomal storage abnormalities, the affected individuals displayed severe developmental delay with pronounced generalized hypotonia, respiratory insufficiency, and variable neurodegeneration and diffusion restriction in cerebral peduncles, midbrain, and/or brainstem in MRI scans. The p.Tyr553Cys amino acid substitution strongly slowed ClC-6 gating and increased current amplitudes, particularly at the acidic pH of late endosomes. Transfection of ClC-6Tyr553Cys, but not ClC-6WT, generated giant LAMP1-positive vacuoles that were poorly acidified. Their generation strictly required ClC-6 ion transport, as shown by transport-deficient double mutants, and depended on Cl-/H+ exchange, as revealed by combination with the uncoupling p.Glu200Ala substitution. Transfection of either ClC-6Tyr553Cys/Glu200Ala or ClC-6Glu200Ala generated slightly enlarged vesicles, suggesting that p.Glu200Ala, previously associated with infantile spasms and microcephaly, is also pathogenic. Bafilomycin treatment abrogated vacuole generation, indicating that H+-driven Cl- accumulation osmotically drives vesicle enlargement. Our work establishes mutations in CLCN6 associated with neurological diseases, whose spectrum of clinical features depends on the differential impact of the allele on ClC-6 function.
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Canales de Cloruro/genética , Mutación con Ganancia de Función , Enfermedades Neurodegenerativas/genética , Alelos , Animales , Células CHO , Niño , Cricetulus , Electrofisiología , Endosomas/metabolismo , Femenino , Células HeLa , Heterocigoto , Homeostasis , Humanos , Concentración de Iones de Hidrógeno , Lactante , Transporte Iónico , Iones , Proteínas de Membrana de los Lisosomas/metabolismo , Lisosomas/metabolismo , Macrólidos/farmacología , Imagen por Resonancia Magnética , Masculino , Ratones , Ratones Noqueados , Microscopía por Video , TransfecciónRESUMEN
Mitochondrial ribosomal protein large 24 (MRPL24) is 1 of the 82 protein components of mitochondrial ribosomes, playing an essential role in the mitochondrial translation process. We report here on a baby girl with cerebellar atrophy, choreoathetosis of limbs and face, intellectual disability and a combined defect of complexes I and IV in muscle biopsy, caused by a homozygous missense mutation identified in MRPL24. The variant predicts a Leu91Pro substitution at an evolutionarily conserved site. Using human mutant cells and the zebrafish model, we demonstrated the pathological role of the identified variant. In fact, in fibroblasts we observed a significant reduction of MRPL24 protein and of mitochondrial respiratory chain complex I and IV subunits, as well a markedly reduced synthesis of the mtDNA-encoded peptides. In zebrafish we demonstrated that the orthologue gene is expressed in metabolically active tissues, and that gene knockdown induced locomotion impairment, structural defects and low ATP production. The motor phenotype was complemented by human WT but not mutant cRNA. Moreover, sucrose density gradient fractionation showed perturbed assembly of large subunit mitoribosomal proteins, suggesting that the mutation leads to a conformational change in MRPL24, which is expected to cause an aberrant interaction of the protein with other components of the 39S mitoribosomal subunit.
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Proteínas Mitocondriales/genética , Trastornos del Movimiento/genética , Proteínas Ribosómicas/genética , Animales , Cerebelo/patología , Femenino , Humanos , Lactante , Leviviridae , Masculino , Trastornos del Movimiento/patología , Músculo Cuádriceps/patología , Pez CebraRESUMEN
OBJECTIVE: To investigate the molecular cause(s) underlying a severe form of infantile-onset parkinsonism and characterize functionally the identified variants. METHODS: A trio-based whole exome sequencing (WES) approach was used to identify the candidate variants underlying the disorder. In silico modeling, and in vitro and in vivo studies were performed to explore the impact of these variants on protein function and relevant cellular processes. RESULTS: WES analysis identified biallelic variants in WARS2, encoding the mitochondrial tryptophanyl tRNA synthetase (mtTrpRS), a gene whose mutations have recently been associated with multiple neurological phenotypes, including childhood-onset, levodopa-responsive or unresponsive parkinsonism in a few patients. A substantial reduction of mtTrpRS levels in mitochondria and reduced OXPHOS function was demonstrated, supporting their pathogenicity. Based on the infantile-onset and severity of the phenotype, additional variants were considered as possible genetic modifiers. Functional assessment of a selected panel of candidates pointed to a de novo missense mutation in CHRNA6, encoding the α6 subunit of neuronal nicotinic receptors, which are involved in the cholinergic modulation of dopamine release in the striatum, as a second event likely contributing to the phenotype. In silico, in vitro (Xenopus oocytes and GH4C1 cells) and in vivo (C. elegans) analyses demonstrated the disruptive effects of the mutation on acetylcholine receptor structure and function. CONCLUSION: Our findings consolidate the association between biallelic WARS2 mutations and movement disorders, and suggest CHRNA6 as a genetic modifier of the phenotype.
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Trastornos Parkinsonianos/genética , Receptores Nicotínicos/genética , Triptófano-ARNt Ligasa/genética , Edad de Inicio , Niño , Humanos , Masculino , Mutación , Índice de Severidad de la Enfermedad , Secuenciación del ExomaRESUMEN
Mitochondria are highly dynamic organelles, undergoing continuous fission and fusion. The DNM1L (dynamin-1 like) gene encodes for the DRP1 protein, an evolutionary conserved member of the dynamin family, responsible for fission of mitochondria, and having a role in the division of peroxisomes, as well. DRP1 impairment is implicated in several neurological disorders and associated with either de novo dominant or compound heterozygous mutations. In five patients presenting with severe epileptic encephalopathy, we identified five de novo dominant DNM1L variants, the pathogenicity of which was validated in a yeast model. Fluorescence microscopy revealed abnormally elongated mitochondria and aberrant peroxisomes in mutant fibroblasts, indicating impaired fission of these organelles. Moreover, a very peculiar finding in our cohort of patients was the presence, in muscle biopsy, of core like areas with oxidative enzyme alterations, suggesting an abnormal distribution of mitochondria in the muscle tissue.
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Dinaminas/genética , Estudios de Asociación Genética , Predisposición Genética a la Enfermedad , Encefalomiopatías Mitocondriales/diagnóstico , Encefalomiopatías Mitocondriales/genética , Músculos/metabolismo , Músculos/patología , Biomarcadores , Encéfalo/diagnóstico por imagen , Encéfalo/metabolismo , Encéfalo/patología , Análisis Mutacional de ADN , Dinaminas/química , Fibroblastos/metabolismo , Estudios de Asociación Genética/métodos , Humanos , Imagen por Resonancia Magnética/métodos , Modelos Biológicos , Músculos/ultraestructura , Mutación , Conformación Proteica , Relación Estructura-ActividadRESUMEN
OBJECTIVE: To characterize clinically and molecularly an early-onset, variably progressive neurodegenerative disorder characterized by a cerebellar syndrome with severe ataxia, gaze palsy, dyskinesia, dystonia, and cognitive decline affecting 11 individuals from 3 consanguineous families. METHODS: We used whole-exome sequencing (WES) (families 1 and 2) and a combined approach based on homozygosity mapping and WES (family 3). We performed in vitro studies to explore the effect of the nontruncating SQSTM1 mutation on protein function and the effect of impaired SQSTM1 function on autophagy. We analyzed the consequences of sqstm1 down-modulation on the structural integrity of the cerebellum in vivo using zebrafish as a model. RESULTS: We identified 3 homozygous inactivating variants, including a splice site substitution (c.301+2T>A) causing aberrant transcript processing and accelerated degradation of a resulting protein lacking exon 2, as well as 2 truncating changes (c.875_876insT and c.934_936delinsTGA). We show that loss of SQSTM1 causes impaired production of ubiquitin-positive protein aggregates in response to misfolded protein stress and decelerated autophagic flux. The consequences of sqstm1 down-modulation on the structural integrity of the cerebellum in zebrafish documented a variable but reproducible phenotype characterized by cerebellum anomalies ranging from depletion of axonal connections to complete atrophy. We provide a detailed clinical characterization of the disorder; the natural history is reported for 2 siblings who have been followed up for >20 years. CONCLUSIONS: This study offers an accurate clinical characterization of this recently recognized neurodegenerative disorder caused by biallelic inactivating mutations in SQSTM1 and links this phenotype to defective selective autophagy.
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Alelos , Progresión de la Enfermedad , Mutación/genética , Enfermedades Neurodegenerativas/diagnóstico por imagen , Enfermedades Neurodegenerativas/genética , Proteína Sequestosoma-1/genética , Adolescente , Adulto , Edad de Inicio , Animales , Femenino , Humanos , Masculino , Linaje , Secuenciación del Exoma/métodos , Adulto Joven , Pez CebraRESUMEN
Multiple mitochondrial dysfunction syndromes (MMDS) comprise a group of severe autosomal recessive diseases characterized by impaired respiration and lipoic acid metabolism, resulting in infantile-onset mitochondrial encephalopathy, non-ketotic hyperglycinemia, myopathy, lactic acidosis and early death. Four different MMDS have been analyzed in detail according to the genes involved in the disease, MMDS1 (NFU1), MMDS2 (BOLA3), MMDS3 (IBA57) and MMDS4 (ISCA2). MMDS5 has recently been described in a clinical case report of patients carrying a mutation in ISCA1, but with no further functional analysis. ISCA1 encodes a mitochondrial protein essential for the assembly of [4Fe-4S] clusters in key metabolic and respiratory enzymes. Here, we describe a patient with a severe early onset leukodystrophy, multiple defects of respiratory complexes and a severe impairment of lipoic acid synthesis. A homozygous missense mutation in ISCA1 (c.29T>G; p.V10G) identified by targeted MitoExome sequencing resulted in dramatic reduction of ISCA1 protein level. The mutation located in the uncleaved presequence severely affected both mitochondrial import and stability of ISCA1. Down-regulation of ISCA1 in HeLa cells by RNAi impaired the biogenesis of mitochondrial [4Fe-4S] proteins, yet could be complemented by expression of wild-type ISCA1. In contrast, the ISCA1 p.V10G mutant protein only partially complemented the defects, closely resembling the biochemical phenotypes observed for ISCA1 patient fibroblasts. Collectively, our comprehensive clinical and biochemical investigations show that the ISCA1 p.V10G mutation functionally impaired mitochondrial [4Fe-4S] protein assembly and hence was causative for the observed clinical defects.
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Proteínas Hierro-Azufre/metabolismo , Leucoencefalopatías/genética , Enfermedades Mitocondriales/etiología , Proteínas Mitocondriales/metabolismo , Mutación , Edad de Inicio , Encéfalo/diagnóstico por imagen , Encéfalo/patología , Niño , Femenino , Prueba de Complementación Genética , Células HeLa , Homocigoto , Humanos , Proteínas Hierro-Azufre/genética , Enfermedades Mitocondriales/genética , Proteínas Mitocondriales/genéticaRESUMEN
Microtubules participate in fundamental cellular processes, including chromosomal segregation and cell division, migration and intracellular trafficking. Their proper function is required for correct central nervous system development and operative preservation, and mutations in genes coding tubulins, the constituting units of microtubules, underlie a family of neurodevelopmental and neurodegenerative diseases, collectively known as 'tubulinopathies', characterized by a wide range of neuronal defects resulting from defective proliferation, migration and function. Here, we causally link a previously unreported missense mutation in TUBB2A (c.1249G>A, p.D417N), encoding one of the neuron-specific ß-tubulin isotype II, to a disorder characterized by progressive spastic paraplegia, peripheral sensory-motor polyneuropathy and ataxia. Asp417 is a highly conserved solvent-exposed residue at the site mediating binding of kinesin superfamily motors. Impaired binding to KIF1A, a neuron-specific kinesin required for transport of synaptic vesicle precursors of the disease-associated TUBB2A mutant, was predicted by structural analyses and confirmed experimentally in vitro. We show that overexpression of TUBB2AD417N disrupts the mitotic spindle bipolarity and morphology and affects the M phase entry and length. Differently from the TUBB2AN247K and TUBB2AA248V, two mutants previously identified to affect neurodevelopment, TUBB2AD417N retains the ability to assemble into microtubules. Consistent with the differential clinical and structural impact, TUBB2AA248V does not drastically affect TUBB2A binding to KIF1A, nor mitotic spindle bipolarity. Overall, our data demonstrate a pathogenic role of the p.D417N substitution that is different from previously reported TUBB2A mutations and expand the phenotypic spectrum associated with mutations in this gene.
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Discapacidad Intelectual/genética , Cinesinas/genética , Espasticidad Muscular/genética , Atrofia Óptica/genética , Paraplejía/genética , Ataxias Espinocerebelosas/genética , Degeneraciones Espinocerebelosas/genética , Tubulina (Proteína)/genética , Adolescente , Adulto , Movimiento Celular/genética , Proliferación Celular/genética , Niño , Femenino , Humanos , Discapacidad Intelectual/diagnóstico por imagen , Discapacidad Intelectual/fisiopatología , Masculino , Microtúbulos/genética , Microtúbulos/patología , Espasticidad Muscular/diagnóstico por imagen , Espasticidad Muscular/fisiopatología , Neuronas/metabolismo , Neuronas/patología , Atrofia Óptica/diagnóstico por imagen , Atrofia Óptica/fisiopatología , Paraplejía/fisiopatología , Polineuropatías/genética , Polineuropatías/fisiopatología , Unión Proteica , Corteza Sensoriomotora/metabolismo , Corteza Sensoriomotora/fisiopatología , Huso Acromático/genética , Ataxias Espinocerebelosas/diagnóstico por imagen , Ataxias Espinocerebelosas/fisiopatología , Degeneraciones Espinocerebelosas/fisiopatologíaRESUMEN
BACKGROUND: Heterozygous mutations in OPA1 are a common cause of autosomal dominant optic atrophy, sometimes associated with extra-ocular manifestations. Few cases harboring compound heterozygous OPA1 mutations have been described manifesting complex neurodegenerative disorders in addition to optic atrophy. RESULTS: We report here three patients: one boy showing an early-onset mitochondrial disorder with hypotonia, ataxia and neuropathy that was severely progressive, leading to early death because of multiorgan failure; two unrelated sporadic girls manifesting a spastic ataxic syndrome associated with peripheral neuropathy and, only in one, optic atrophy. Using a targeted resequencing of 132 genes associated with mitochondrial disorders, in two probands we found compound heterozygous mutations in OPA1: in the first a 5 nucleotide deletion, causing a frameshift and insertion of a premature stop codon (p.Ser64Asnfs*7), and a missense change (p.Ile437Met), which has recently been reported to have clinical impact; in the second, a novel missense change (p.Val988Phe) co-occurred with the p.Ile437Met substitution. In the third patient a homozygous mutation, c.1180G > A (p.Ala394Thr) in OPA1 was detected by a trio-based whole exome sequencing approach. One of the patients presented also variants in mitochondrial DNA that may have contributed to the peculiar phenotype. The deleterious effect of the identified missense changes was experimentally validated in yeast model. OPA1 level was reduced in available patients' biological samples, and a clearly fragmented mitochondrial network was observed in patients' fibroblasts. CONCLUSIONS: This report provides evidence that bi-allelic OPA1 mutations may lead to complex and severe multi-system recessive mitochondrial disorders, where optic atrophy might not represent the main feature.
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Encefalopatías/genética , GTP Fosfohidrolasas/genética , Atrofia Óptica Autosómica Dominante/genética , Atrofia Óptica/genética , Western Blotting , Encefalopatías/metabolismo , Preescolar , Electrofisiología , GTP Fosfohidrolasas/metabolismo , Humanos , Lactante , Masculino , Microscopía Fluorescente , Mutación , Atrofia Óptica/metabolismo , Atrofia Óptica Autosómica Dominante/metabolismo , Tomografía de Coherencia Óptica , Secuenciación del ExomaRESUMEN
The mitochondrial Elongation Factor Tu (EF-Tu), encoded by the TUFM gene, is a highly conserved GTPase, which is part of the mitochondrial protein translation machinery. In its activated form it delivers the aminoacyl-tRNAs to the A site of the mitochondrial ribosome. We report here on a baby girl with severe infantile macrocystic leukodystrophy with micropolygyria and a combined defect of complexes I and IV in muscle biopsy, caused by a novel mutation identified in TUFM. Using human mutant cells and the yeast model, we demonstrate the pathological role of the novel variant. Moreover, results of a molecular modeling study suggest that the mutant is inactive in mitochondrial polypeptide chain elongation, probably as a consequence of its reduced ability to bind mitochondrial aa-tRNAs. Four patients have so far been described with mutations in TUFM, and, following the first description of the disease in a single patient, we describe similar clinical and neuroradiological features in an additional patient.
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Secuencia de Bases , ADN Mitocondrial/genética , Leucoencefalopatías/genética , Mitocondrias/genética , Proteínas Mitocondriales/genética , Extensión de la Cadena Peptídica de Translación , Factor Tu de Elongación Peptídica/genética , Eliminación de Secuencia , ADN Mitocondrial/metabolismo , Femenino , Humanos , Leucoencefalopatías/metabolismo , Masculino , Mitocondrias/patología , Proteínas Mitocondriales/metabolismo , Factor Tu de Elongación Peptídica/metabolismo , Ribosomas/genética , Ribosomas/metabolismo , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismoRESUMEN
Defects of the Fe/S cluster biosynthesis represent a subgroup of diseases affecting the mitochondrial energy metabolism. In the last years, mutations in four genes (NFU1, BOLA3, ISCA2 and IBA57) have been related to a new group of multiple mitochondrial dysfunction syndromes characterized by lactic acidosis, hyperglycinemia, multiple defects of the respiratory chain complexes, and impairment of four lipoic acid-dependent enzymes: α-ketoglutarate dehydrogenase complex, pyruvic dehydrogenase, branched-chain α-keto acid dehydrogenase complex and the H protein of the glycine cleavage system. Few patients have been reported with mutations in IBA57 and with variable clinical phenotype. Herein, we describe four unrelated patients carrying novel mutations in IBA57. All patients presented with combined or isolated defect of complex I and II. Clinical features varied widely, ranging from fatal infantile onset of the disease to acute and severe psychomotor regression after the first year of life. Brain MRI was characterized by cavitating leukodystrophy. The identified mutations were never reported previously and all had a dramatic effect on IBA57 stability. Our study contributes to expand the array of the genotypic variation of IBA57 and delineates the leukodystrophic pattern of IBA57 deficient patients.
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Encefalopatías/genética , Encéfalo/diagnóstico por imagen , Proteínas Portadoras/genética , Mutación , Enfermedades Neurodegenerativas/genética , Western Blotting , Encefalopatías/diagnóstico , Encefalopatías/fisiopatología , Proteínas Portadoras/metabolismo , Estudios de Cohortes , Femenino , Fibroblastos/metabolismo , Estudios de Seguimiento , Humanos , Lactante , Imagen por Resonancia Magnética , Masculino , Mitocondrias/metabolismo , Enfermedades Neurodegenerativas/diagnóstico , Enfermedades Neurodegenerativas/fisiopatología , Fenotipo , Estabilidad ProteicaRESUMEN
Tubulinopathies constitute a family of neurodevelopmental/neurodegenerative disorders caused by mutations in several genes encoding tubulin isoforms. Loss-of-function mutations in TBCE, encoding one of the five tubulin-specific chaperones involved in tubulin folding and polymerization, cause two rare neurodevelopmental syndromes, hypoparathyroidism-retardation-dysmorphism and Kenny-Caffey syndrome. Although a missense mutation in Tbce has been associated with progressive distal motor neuronopathy in the pmn/pmn mice, no similar degenerative phenotype has been recognized in humans. We report on the identification of an early-onset and progressive neurodegenerative encephalopathy with distal spinal muscular atrophy resembling the phenotype of pmn/pmn mice and caused by biallelic TBCE mutations, with the c.464T>A (p.Ile155Asn) change occurring at the heterozygous/homozygous state in six affected subjects from four unrelated families originated from the same geographical area in Southern Italy. Western blot analysis of patient fibroblasts documented a reduced amount of TBCE, suggestive of rapid degradation of the mutant protein, similarly to what was observed in pmn/pmn fibroblasts. The impact of TBCE mutations on microtubule polymerization was determined using biochemical fractionation and analyzing the nucleation and growth of microtubules at the centrosome and extracentrosomal sites after treatment with nocodazole. Primary fibroblasts obtained from affected subjects displayed a reduced level of polymerized α-tubulin, similarly to tail fibroblasts of pmn/pmn mice. Moreover, markedly delayed microtubule re-polymerization and abnormal mitotic spindles with disorganized microtubule arrangement were also documented. Although loss of function of TBCE has been documented to impact multiple developmental processes, the present findings provide evidence that hypomorphic TBCE mutations primarily drive neurodegeneration.
Asunto(s)
Encefalopatías/complicaciones , Encefalopatías/genética , Chaperonas Moleculares/genética , Atrofia Muscular Espinal/complicaciones , Atrofia Muscular Espinal/genética , Mutación/genética , Adolescente , Edad de Inicio , Animales , Niño , Femenino , Fibroblastos , Heterocigoto , Homocigoto , Humanos , Lactante , Recién Nacido , Italia , Masculino , Ratones , Microtúbulos/efectos de los fármacos , Microtúbulos/metabolismo , Microtúbulos/patología , Chaperonas Moleculares/metabolismo , Nocodazol/farmacología , Huso Acromático/metabolismo , Huso Acromático/patología , Tubulina (Proteína)/metabolismo , Adulto JovenRESUMEN
This study focused on the molecular characterization of patients with leukoencephalopathy associated with a specific biochemical defect of mitochondrial respiratory chain complex III, and explores the impact of a distinct magnetic resonance imaging pattern of leukoencephalopathy to detect biallelic mutations in LYRM7 in patients with biochemically unclassified leukoencephalopathy. 'Targeted resequencing' of a custom panel including genes coding for mitochondrial proteins was performed in patients with complex III deficiency without a molecular genetic diagnosis. Based on brain magnetic resonance imaging findings in these patients, we selected additional patients from a database of unclassified leukoencephalopathies who were scanned for mutations in LYRM7 by Sanger sequencing. Targeted sequencing revealed homozygous mutations in LYRM7, encoding mitochondrial LYR motif-containing protein 7, in four patients from three unrelated families who had a leukoencephalopathy and complex III deficiency. Two subjects harboured previously unreported variants predicted to be damaging, while two siblings carried an already reported pathogenic homozygous missense change. Sanger sequencing performed in the second cohort of patients revealed LYRM7 mutations in three additional patients, who were selected on the basis of the magnetic resonance imaging pattern. All patients had a consistent magnetic resonance imaging pattern of progressive signal abnormalities with multifocal small cavitations in the periventricular and deep cerebral white matter. Early motor development was delayed in half of the patients. All patients but one presented with subacute neurological deterioration in infancy or childhood, preceded by a febrile infection, and most patients had repeated episodes of subacute encephalopathy with motor regression, irritability and stupor or coma resulting in major handicap or death. LYRM7 protein was strongly reduced in available samples from patients; decreased complex III holocomplex was observed in fibroblasts from a patient carrying a splice site variant; functional studies in yeast confirmed the pathogenicity of two novel mutations. Mutations in LYRM7 were previously found in a single patient with a severe form of infantile onset encephalopathy. We provide new molecular, clinical, and neuroimaging data allowing us to characterize more accurately the molecular spectrum of LYRM7 mutations highlighting that a distinct and recognizable magnetic resonance imaging pattern is related to mutations in this gene. Inter- and intrafamilial variability exists and we observed one patient who was asymptomatic by the age of 6 years.