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BACKGROUND: The late-onset cerebellar ataxias (LOCAs) have largely resisted molecular diagnosis. METHODS: We sequenced the genomes of six persons with autosomal dominant LOCA who were members of three French Canadian families and identified a candidate pathogenic repeat expansion. We then tested for association between the repeat expansion and disease in two independent case-control series - one French Canadian (66 patients and 209 controls) and the other German (228 patients and 199 controls). We also genotyped the repeat in 20 Australian and 31 Indian index patients. We assayed gene and protein expression in two postmortem cerebellum specimens and two induced pluripotent stem-cell (iPSC)-derived motor-neuron cell lines. RESULTS: In the six French Canadian patients, we identified a GAA repeat expansion deep in the first intron of FGF14, which encodes fibroblast growth factor 14. Cosegregation of the repeat expansion with disease in the families supported a pathogenic threshold of at least 250 GAA repeats ([GAA]≥250). There was significant association between FGF14 (GAA)≥250 expansions and LOCA in the French Canadian series (odds ratio, 105.60; 95% confidence interval [CI], 31.09 to 334.20; P<0.001) and in the German series (odds ratio, 8.76; 95% CI, 3.45 to 20.84; P<0.001). The repeat expansion was present in 61%, 18%, 15%, and 10% of French Canadian, German, Australian, and Indian index patients, respectively. In total, we identified 128 patients with LOCA who carried an FGF14 (GAA)≥250 expansion. Postmortem cerebellum specimens and iPSC-derived motor neurons from patients showed reduced expression of FGF14 RNA and protein. CONCLUSIONS: A dominantly inherited deep intronic GAA repeat expansion in FGF14 was found to be associated with LOCA. (Funded by Fondation Groupe Monaco and others.).
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Ataxia Cerebelosa , Expansión de las Repeticiones de ADN , Intrones , Humanos , Australia , Canadá , Ataxia Cerebelosa/genética , Ataxia Cerebelosa/patología , Ataxia de Friedreich/genética , Ataxia de Friedreich/patología , Intrones/genética , Expansión de las Repeticiones de ADN/genéticaRESUMEN
Biallelic SORD mutations cause one of the most frequent forms of recessive hereditary neuropathy, estimated to affect approximately 10,000 patients in North America and Europe alone. Pathogenic SORD loss-of-function changes in the encoded enzyme sorbitol dehydrogenase result in abnormally high sorbitol levels in cells and serum. How sorbitol accumulation leads to peripheral neuropathy remains to be elucidated. A reproducible animal model for SORD neuropathy is essential to illuminate the pathogenesis of SORD deficiency and for preclinical studies of potential therapies. Therefore, we have generated a Sord knockout (KO), Sord-/-, Sprague Dawley rat, to model the human disease and to investigate the pathophysiology underlying SORD deficiency. We have characterized the phenotype in these rats with a battery of behavioral tests as well as biochemical, physiological, and comprehensive histological examinations. Sord-/- rats had remarkably increased levels of sorbitol in serum, cerebrospinal fluid (CSF), and peripheral nerve. Moreover, serum from Sord-/- rats contained significantly increased levels of neurofilament light chain, NfL, an established biomarker for axonal degeneration. Motor performance significantly declined in Sord-/- animals starting at â¼7 months of age. Gait analysis evaluated with video motion tracking confirmed abnormal gait patterns in the hindlimbs. Motor nerve conduction velocities of the tibial nerves were slowed. Light and electron microscopy of the peripheral nervous system revealed degenerating myelinated axons, de- and remyelinated axons, and a likely pathognomonic finding - enlarged "ballooned" myelin sheaths. These findings mainly affected myelinated motor axons; myelinated sensory axons were largely spared. In summary, Sord-/- rats develop a motor-predominant neuropathy that closely resembles the human phenotype. Our studies revealed novel significant aspects of SORD deficiency, and this model will lead to an improved understanding of the pathophysiology and the therapeutic options for SORD neuropathy.
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DNA-based therapeutics have emerged as a revolutionary approach for addressing the treatment gap in rare inherited conditions by targeting the fundamental genetic causes of disease. Charcot-Marie-Tooth (CMT) disease, a group of inherited neuropathies, represents one of the most prevalent Mendelian disease groups in neurology and is characterized by diverse genetic etiology. Axonal forms of CMT, known as CMT2, are caused by dominant mutations in over 30 different genes which lead to degeneration of lower motor neuron axons. Recent advances in antisense oligonucleotide (ASO) therapeutics have shown promise in targeting neurodegenerative disorders. Here we elucidate pathomechanistic changes contributing to variant specific molecular phenotypes in CMT2E, caused by a single nucleotide substitution (p.N98S) in the neurofilament light chain gene (NEFL). We used a patient-derived pluripotent stem cell (iPSC)-induced motor neuron model, which recapitulates several cellular and biomarker phenotypes associated with CMT2E. Using an ASO treatment strategy targeting a heterozygous gain-of-function variant, we aimed to resolve molecular phenotypic changes observed in the CMT2E p.N98S subtype. To determine ASO therapeutic potential, we employed our treatment strategy in iPSC-derived motor neurons and used established as well as novel biomarkers of peripheral nervous system axonal degeneration. Our findings have demonstrated a significant decrease in clinically relevant biomarkers of axonal degeneration, presenting the first clinically viable genetic therapeutic for CMT2E. Similar strategies could be used to develop precision medicine approaches for otherwise untreatable gain of function inherited disorders.
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Charcot-Marie-Tooth (CMT) disease is a neuromuscular disorder affecting the peripheral nervous system. The diagnostic yield in demyelinating CMT (CMT1) is typically â¼80-95%, of which at least 60% is due to the PMP22 gene duplication. The remainder of CMT1 is more genetically heterogeneous. We used whole exome and whole genome sequencing data included in the GENESIS database to investigate novel causal genes and mutations in a cohort of â¼2,670 individuals with CMT neuropathy. A recurrent heterozygous missense variant p.Thr1424Met in the recently described CMT gene ITPR3, encoding IP3R3 (inositol 1,4,5-trisphosphate receptor 3) was identified. This previously reported p.Thr1424Met change was present in 33 affected individuals from nine unrelated families from multiple populations, representing an unusual recurrence rate at a mutational hotspot, strengthening the gene-disease relationship (GnomADv4 allele frequency 1.76e-6). Sanger sequencing confirmed the co-segregation of the CMT phenotype with the presence of the mutation in autosomal dominant and de novo inheritance patterns, including a four-generation family with multiple affected second-degree cousins. Probands from all families presented with slow nerve conduction velocities, matching the diagnostic category of CMT1. Remarkably, we observed a uniquely variable clinical phenotype for age at onset and phenotype severity in p.Thr1424Met carrying patients, even within families. Finally, we present data supportive of a dominant-negative effect of the p.Thr1424Met mutation with associated changes in protein expression in patient-derived cells.
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BACKGROUND AND PURPOSE: Mutations in the alpha-B-crystallin (CRYAB) gene have initially been associated with myofibrillar myopathy, dilated cardiomyopathy and cataracts. For the first time, peripheral neuropathy is reported here as a novel phenotype associated with CRYAB. METHODS: Whole-exome sequencing was performed in two unrelated families with genetically unsolved axonal Charcot-Marie-Tooth disease (CMT2), assessing clinical, neurophysiological and radiological features. RESULTS: The pathogenic CRYAB variant c.358A>G;p.Arg120Gly was segregated in all affected patients from two unrelated families. The disease presented as late onset CMT2 (onset over 40 years) with distal sensory and motor impairment and congenital cataracts. Muscle involvement was probably associated in cases showing mild axial and diaphragmatic weakness. In all cases, nerve conduction studies demonstrated the presence of an axonal sensorimotor neuropathy along with chronic neurogenic changes on needle examination. DISCUSSION: In cases with late onset autosomal dominant CMT2 and congenital cataracts, it is recommended that CRYAB is considered for genetic testing. The identification of CRYAB mutations causing CMT2 further supports a continuous spectrum of expressivity, from myopathic to neuropathic and mixed forms, of a growing number of genes involved in protein degradation and chaperone-assisted autophagy.
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Catarata , Enfermedad de Charcot-Marie-Tooth , Cristalinas , Humanos , Enfermedad de Charcot-Marie-Tooth/complicaciones , Enfermedad de Charcot-Marie-Tooth/genética , Enfermedad de Charcot-Marie-Tooth/diagnóstico , Mutación/genética , Pruebas Genéticas , Fenotipo , Cristalinas/genética , Catarata/genética , LinajeRESUMEN
BACKGROUND: Caused by duplications of the gene encoding peripheral myelin protein 22 (PMP22), Charcot-Marie-Tooth disease type 1A (CMT1A) is the most common hereditary neuropathy. Despite this shared genetic origin, there is considerable variability in clinical severity. It is hypothesized that genetic modifiers contribute to this heterogeneity, the identification of which may reveal novel therapeutic targets. In this study, we present a comprehensive analysis of clinical examination results from 1564 CMT1A patients sourced from a prospective natural history study conducted by the RDCRN-INC (Inherited Neuropathy Consortium). Our primary objective is to delineate extreme phenotype profiles (mild and severe) within this patient cohort, thereby enhancing our ability to detect genetic modifiers with large effects. METHODS: We have conducted large-scale statistical analyses of the RDCRN-INC database to characterize CMT1A severity across multiple metrics. RESULTS: We defined patients below the 10th (mild) and above the 90th (severe) percentiles of age-normalized disease severity based on the CMT Examination Score V2 and foot dorsiflexion strength (MRC scale). Based on extreme phenotype categories, we defined a statistically justified recruitment strategy, which we propose to use in future modifier studies. INTERPRETATION: Leveraging whole genome sequencing with base pair resolution, a future genetic modifier evaluation will include single nucleotide association, gene burden tests, and structural variant analysis. The present work not only provides insight into the severity and course of CMT1A, but also elucidates the statistical foundation and practical considerations for a cost-efficient and straightforward patient enrollment strategy that we intend to conduct on additional patients recruited globally.
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Enfermedad de Charcot-Marie-Tooth , Enfermedad de Charcot-Marie-Tooth/genética , Enfermedad de Charcot-Marie-Tooth/fisiopatología , Humanos , Adulto , Masculino , Femenino , Persona de Mediana Edad , Adolescente , Adulto Joven , Índice de Severidad de la Enfermedad , Niño , Proteínas de la Mielina/genética , Selección de Paciente , Fenotipo , Anciano , Genes Modificadores , PreescolarRESUMEN
COQ7 encodes a hydroxylase responsible for the penultimate step of coenzyme Q10 (CoQ10) biosynthesis in mitochondria. CoQ10 is essential for multiple cellular functions, including mitochondrial oxidative phosphorylation, lipid metabolism, and reactive oxygen species homeostasis. Mutations in COQ7 have been previously associated with primary CoQ10 deficiency, a clinically heterogeneous multisystemic mitochondrial disorder. We identified COQ7 biallelic variants in nine families diagnosed with distal hereditary motor neuropathy with upper neuron involvement, expending the clinical phenotype associated with defects in this gene. A recurrent p.Met1? change was identified in five families from Brazil with evidence of a founder effect. Fibroblasts isolated from patients revealed a substantial depletion of COQ7 protein levels, indicating protein instability leading to loss of enzyme function. High-performance liquid chromatography assay showed that fibroblasts from patients had reduced levels of CoQ10, and abnormal accumulation of the biosynthetic precursor DMQ10. Accordingly, fibroblasts from patients displayed significantly decreased oxygen consumption rates in patients, suggesting mitochondrial respiration deficiency. Induced pluripotent stem cell-derived motor neurons from patient fibroblasts showed significantly increased levels of extracellular neurofilament light protein, indicating axonal degeneration. Our findings indicate a molecular pathway involving CoQ10 biosynthesis deficiency and mitochondrial dysfunction in patients with distal hereditary motor neuropathy. Further studies will be important to evaluate the potential benefits of CoQ10 supplementation in the clinical outcome of the disease.
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Enfermedades Mitocondriales , Humanos , Mitocondrias/genética , Mitocondrias/metabolismo , Enfermedades Mitocondriales/metabolismo , Neuronas Motoras/metabolismo , Mutación/genética , Ubiquinona/genéticaRESUMEN
The diagnostic gap for rare neurodegenerative diseases is still considerable, despite continuous advances in gene identification. Many novel Mendelian genes have only been identified in a few families worldwide. Here we report the identification of an autosomal-dominant gene for hereditary spastic paraplegia (HSP) in 10 families that are of diverse geographic origin and whose affected members all carry unique truncating changes in a circumscript region of UBAP1 (ubiquitin-associated protein 1). HSP is a neurodegenerative disease characterized by progressive lower-limb spasticity and weakness, as well as frequent bladder dysfunction. At least 40% of affected persons are currently undiagnosed after exome sequencing. We identified pathological truncating variants in UBAP1 in affected persons from Iran, USA, Germany, Canada, Spain, and Bulgarian Roma. The genetic support ranges from linkage in the largest family (LOD = 8.3) to three confirmed de novo mutations. We show that mRNA in the fibroblasts of affected individuals escapes nonsense-mediated decay and thus leads to the expression of truncated proteins; in addition, concentrations of the full-length protein are reduced in comparison to those in controls. This suggests either a dominant-negative effect or haploinsufficiency. UBAP1 links endosomal trafficking to the ubiquitination machinery pathways that have been previously implicated in HSPs, and UBAP1 provides a bridge toward a more unified pathophysiology.
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Proteínas Portadoras/genética , Mutación , Paraplejía Espástica Hereditaria/genética , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Animales , Niño , Preescolar , Bases de Datos Factuales , Modelos Animales de Enfermedad , Endosomas/metabolismo , Salud de la Familia , Femenino , Fibroblastos/metabolismo , Genes Dominantes , Ligamiento Genético , Predisposición Genética a la Enfermedad , Genómica , Células HEK293 , Haploinsuficiencia , Humanos , Masculino , Persona de Mediana Edad , Linaje , Isoformas de Proteínas , Adulto Joven , Pez CebraRESUMEN
PURPOSE: The chaperone protein BiP is the master regulator of the unfolded protein response in the endoplasmic reticulum. BiP chaperone activity is regulated by the post-translational modification AMPylation, exclusively provided by FICD. We investigated whether FICD variants identified in patients with motor neuron disease could interfere with BiP activity regulation. METHODS: Exome sequencing was performed to identify causative pathogenic variants associated with motor neuron diseases. Functional studies were conducted on fibroblasts from patients to explore the molecular mechanism of the disease. RESULTS: We identified biallelic variants in FICD causing a neurodegenerative disease of upper and lower motor neurons. Affected individuals harbor a specific missense variant, Arg374His, positioned in the catalytic motif of the enzyme and important for adenosine triphosphate binding. The mutated residue abolishes intramolecular interaction with the regulatory residue Glu234, essential to inhibit AMPylation and to promote de-AMPylation by FICD. Consequently, fibroblasts from patients with FICD variants have abnormally increased levels of AMPylated and thus inactivated BiP. CONCLUSION: Loss of BiP chaperone activity in patients likely results in a chronic impairment of the protein quality control system in the endoplasmic reticulum. These findings will guide the development of therapeutic strategies for motoneuron and related diseases linked to proteotoxic stress.
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Enfermedad de la Neurona Motora , Enfermedades Neurodegenerativas , Humanos , Proteínas de Choque Térmico/química , Proteínas de Choque Térmico/metabolismo , Chaperón BiP del Retículo Endoplásmico , Retículo Endoplásmico/genética , Retículo Endoplásmico/metabolismo , Enfermedad de la Neurona Motora/genética , Enfermedad de la Neurona Motora/metabolismoRESUMEN
The CADM family of proteins consists of four neuronal specific adhesion molecules (CADM1, CADM2, CADM3 and CADM4) that mediate the direct contact and interaction between axons and glia. In the peripheral nerve, axon-Schwann cell interaction is essential for the structural organization of myelinated fibres and is primarily mediated by the binding of CADM3, expressed in axons, to CADM4, expressed by myelinating Schwann cells. We have identified-by whole exome sequencing-three unrelated families, including one de novo patient, with axonal Charcot-Marie-Tooth disease (CMT2) sharing the same private variant in CADM3, Tyr172Cys. This variant is absent in 230 000 control chromosomes from gnomAD and predicted to be pathogenic. Most CADM3 patients share a similar phenotype consisting of autosomal dominant CMT2 with marked upper limb involvement. High resolution mass spectrometry analysis detected a newly created disulphide bond in the mutant CADM3 potentially modifying the native protein conformation. Our data support a retention of the mutant protein in the endoplasmic reticulum and reduced cell surface expression in vitro. Stochastic optical reconstruction microscopy imaging revealed decreased co-localization of the mutant with CADM4 at intercellular contact sites. Mice carrying the corresponding human mutation (Cadm3Y170C) showed reduced expression of the mutant protein in axons. Cadm3Y170C mice showed normal nerve conduction and myelin morphology, but exhibited abnormal axonal organization, including abnormal distribution of Kv1.2 channels and Caspr along myelinated axons. Our findings indicate the involvement of abnormal axon-glia interaction as a disease-causing mechanism in CMT patients with CADM3 mutations.
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Moléculas de Adhesión Celular/genética , Enfermedad de Charcot-Marie-Tooth/genética , Inmunoglobulinas/genética , Adulto , Axones/patología , Enfermedad de Charcot-Marie-Tooth/metabolismo , Enfermedad de Charcot-Marie-Tooth/patología , Niño , Femenino , Humanos , Masculino , Persona de Mediana Edad , Mutación , Neuroglía/patología , Linaje , FenotipoRESUMEN
Peroxiredoxin 3 (PRDX3) belongs to a superfamily of peroxidases that function as protective antioxidant enzymes. Among the six isoforms (PRDX1-PRDX6), PRDX3 is the only protein exclusively localized to the mitochondria, which are the main source of reactive oxygen species. Excessive levels of reactive oxygen species are harmful to cells, inducing mitochondrial dysfunction, DNA damage, lipid and protein oxidation and ultimately apoptosis. Neuronal cell damage induced by oxidative stress has been associated with numerous neurodegenerative disorders including Alzheimer's and Parkinson's diseases. Leveraging the large aggregation of genomic ataxia datasets from the PREPARE (Preparing for Therapies in Autosomal Recessive Ataxias) network, we identified recessive mutations in PRDX3 as the genetic cause of cerebellar ataxia in five unrelated families, providing further evidence for oxidative stress in the pathogenesis of neurodegeneration. The clinical presentation of individuals with PRDX3 mutations consists of mild-to-moderate progressive cerebellar ataxia with concomitant hyper- and hypokinetic movement disorders, severe early-onset cerebellar atrophy, and in part olivary and brainstem degeneration. Patient fibroblasts showed a lack of PRDX3 protein, resulting in decreased glutathione peroxidase activity and decreased mitochondrial maximal respiratory capacity. Moreover, PRDX3 knockdown in cerebellar medulloblastoma cells resulted in significantly decreased cell viability, increased H2O2 levels and increased susceptibility to apoptosis triggered by reactive oxygen species. Pan-neuronal and pan-glial in vivo models of Drosophila revealed aberrant locomotor phenotypes and reduced survival times upon exposure to oxidative stress. Our findings reveal a central role for mitochondria and the implication of oxidative stress in PRDX3 disease pathogenesis and cerebellar vulnerability and suggest targets for future therapeutic approaches.
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Ataxia Cerebelosa/genética , Estrés Oxidativo/genética , Peroxiredoxina III/genética , Adulto , Animales , Ataxia Cerebelosa/metabolismo , Ataxia Cerebelosa/patología , Drosophila , Femenino , Humanos , Mutación con Pérdida de Función , Masculino , Persona de Mediana Edad , LinajeRESUMEN
Although mutations in more than 90 genes are known to cause CMT, the underlying genetic cause of CMT remains unknown in more than 50% of affected individuals. The discovery of additional genes that harbor CMT2-causing mutations increasingly depends on sharing sequence data on a global level. In this way-by combining data from seven countries on four continents-we were able to define mutations in ATP1A1, which encodes the alpha1 subunit of the Na+,K+-ATPase, as a cause of autosomal-dominant CMT2. Seven missense changes were identified that segregated within individual pedigrees: c.143T>G (p.Leu48Arg), c.1775T>C (p.Ile592Thr), c.1789G>A (p.Ala597Thr), c.1801_1802delinsTT (p.Asp601Phe), c.1798C>G (p.Pro600Ala), c.1798C>A (p.Pro600Thr), and c.2432A>C (p.Asp811Ala). Immunostaining peripheral nerve axons localized ATP1A1 to the axolemma of myelinated sensory and motor axons and to Schmidt-Lanterman incisures of myelin sheaths. Two-electrode voltage clamp measurements on Xenopus oocytes demonstrated significant reduction in Na+ current activity in some, but not all, ouabain-insensitive ATP1A1 mutants, suggesting a loss-of-function defect of the Na+,K+ pump. Five mutants fall into a remarkably narrow motif within the helical linker region that couples the nucleotide-binding and phosphorylation domains. These findings identify a CMT pathway and a potential target for therapy development in degenerative diseases of peripheral nerve axons.
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Enfermedad de Charcot-Marie-Tooth/genética , Genes Dominantes , Mutación/genética , ATPasa Intercambiadora de Sodio-Potasio/genética , Adulto , Anciano , Anciano de 80 o más Años , Secuencia de Aminoácidos , Niño , Familia , Femenino , Humanos , Masculino , Persona de Mediana Edad , Linaje , ATPasa Intercambiadora de Sodio-Potasio/química , Adulto JovenRESUMEN
Dominant mutations in ATP1A1, encoding the alpha-1 isoform of the Na+ /K+ -ATPase, have been recently reported to cause an axonal to intermediate type of Charcot-Marie-Tooth disease (ie, CMT2DD) and a syndrome with hypomagnesemia, intractable seizures and severe intellectual disability. Here, we describe the first case of hereditary spastic paraplegia (HSP) caused by a novel de novo (p.L337P) variant in ATP1A1. We provide evidence for the causative role of this variant with functional and homology modeling studies. This finding expands the phenotypic spectrum of the ATP1A1-related disorders, adds a piece to the larger genetic puzzle of HSP, and increases knowledge on the molecular mechanisms underlying inherited axonopathies (ie, CMT and HSP).
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Enfermedad de Charcot-Marie-Tooth/genética , Polineuropatías/genética , ATPasa Intercambiadora de Sodio-Potasio/genética , Paraplejía Espástica Hereditaria/genética , Enfermedad de Charcot-Marie-Tooth/patología , Preescolar , Predisposición Genética a la Enfermedad , Mutación de Línea Germinal/genética , Humanos , Masculino , Linaje , Fenotipo , Polineuropatías/complicaciones , Polineuropatías/patología , Paraplejía Espástica Hereditaria/complicaciones , Paraplejía Espástica Hereditaria/patologíaRESUMEN
OBJECTIVE: Genetic modifiers in rare disease have long been suspected to contribute to the considerable variance in disease expression, including Charcot-Marie-Tooth disease type 1A (CMT1A). To address this question, the Inherited Neuropathy Consortium collected a large standardized sample of such rare CMT1A patients over a period of 8 years. CMT1A is caused in most patients by a uniformly sized 1.5 Mb duplication event involving the gene PMP22. METHODS: We genotyped DNA samples from 971 CMT1A patients on Illumina BeadChips. Genome-wide analysis was performed in a subset of 330 of these patients, who expressed the extremes of a hallmark symptom: mild and severe foot dorsiflexion strength impairment. SIPA1L2 (signal-induced proliferation-associated 1 like 2), the top identified candidate modifier gene, was expressed in the peripheral nerve, and our functional studies identified and confirmed interacting proteins using coimmunoprecipitation analysis, mass spectrometry, and immunocytochemistry. Chromatin immunoprecipitation and in vitro siRNA experiments were used to analyze gene regulation. RESULTS: We identified significant association of 4 single nucleotide polymorphisms (rs10910527, rs7536385, rs4649265, rs1547740) in SIPA1L2 with foot dorsiflexion strength (p < 1 × 10-7 ). Coimmunoprecipitation and mass spectroscopy studies identified ß-actin and MYH9 as SIPA1L2 binding partners. Furthermore, we show that SIPA1L2 is part of a myelination-associated coexpressed network regulated by the master transcription factor SOX10. Importantly, in vitro knockdown of SIPA1L2 in Schwannoma cells led to a significant reduction of PMP22 expression, hinting at a potential strategy for drug development. INTERPRETATION: SIPA1L2 is a potential genetic modifier of CMT1A phenotypic expressions and offers a new pathway to therapeutic interventions. ANN NEUROL 2019;85:316-330.
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Enfermedad de Charcot-Marie-Tooth/genética , Pie/fisiopatología , Proteínas Activadoras de GTPasa/genética , Genes Modificadores/genética , Debilidad Muscular/genética , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Animales , Línea Celular Tumoral , Enfermedad de Charcot-Marie-Tooth/fisiopatología , Niño , Preescolar , Femenino , Regulación de la Expresión Génica , Técnicas de Silenciamiento del Gen , Redes Reguladoras de Genes , Humanos , Técnicas In Vitro , Masculino , Persona de Mediana Edad , Debilidad Muscular/fisiopatología , Proteínas de la Mielina/genética , Neurilemoma/genética , Fenotipo , Polimorfismo de Nucleótido Simple , Ratas , Índice de Severidad de la Enfermedad , Adulto JovenRESUMEN
Abnormal protein aggregation is observed in an expanding number of neurodegenerative diseases. Here, we describe a mechanism for intracellular toxic protein aggregation induced by an unusual mutation event in families affected by axonal neuropathy. These families carry distinct frameshift variants in NEFH (neurofilament heavy), leading to a loss of the terminating codon and translation of the 3' UTR into an extra 40 amino acids. In silico aggregation prediction suggested the terminal 20 residues of the altered NEFH to be amyloidogenic, which we confirmed experimentally by serial deletion analysis. The presence of this amyloidogenic motif fused to NEFH caused prominent and toxic protein aggregates in transfected cells and disrupted motor neurons in zebrafish. We identified a similar aggregation-inducing mechanism in NEFL (neurofilament light) and FUS (fused in sarcoma), in which mutations are known to cause aggregation in Charcot-Marie-Tooth disease and amyotrophic lateral sclerosis, respectively. In summary, we present a protein-aggregation-triggering mechanism that should be taken into consideration during the evaluation of stop-loss variants.
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Regiones no Traducidas 3'/genética , Axones/patología , Filamentos Intermedios/genética , Neuronas Motoras/patología , Secuencia de Aminoácidos , Esclerosis Amiotrófica Lateral/genética , Animales , Línea Celular , Enfermedad de Charcot-Marie-Tooth/genética , Mutación del Sistema de Lectura , Humanos , Filamentos Intermedios/metabolismo , Ratones , Datos de Secuencia Molecular , Neuronas Motoras/metabolismo , Mutación , Linaje , Pez Cebra/genéticaRESUMEN
We report on two patients, with different POLG mutations, in whom axonal neuropathy dominated the clinical picture. One patient presented with late onset sensory axonal neuropathy caused by a homozygous c.2243G>C (p.Trp748Ser) mutation that resulted from uniparental disomy of the long arm of chromosome 15. The other patient had a complex phenotype that included early onset axonal Charcot-Marie-Tooth disease (CMT) caused by compound heterozygous c.926G>A (p.Arg309His) and c.2209G>C (p.Gly737Arg) mutations.
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Enfermedad de Charcot-Marie-Tooth/diagnóstico , ADN Polimerasa gamma/genética , Mutación , Conducción Nerviosa/fisiología , Enfermedades del Sistema Nervioso Periférico/diagnóstico , Adolescente , Enfermedad de Charcot-Marie-Tooth/genética , Diagnóstico Diferencial , Electrodiagnóstico , Femenino , Humanos , Persona de Mediana Edad , Linaje , Enfermedades del Sistema Nervioso Periférico/genética , FenotipoRESUMEN
Recessive mutations in the mitochondrial copper-binding protein SCO2, cytochrome c oxidase (COX) assembly protein, have been reported in several cases with fatal infantile cardioencephalomyopathy with COX deficiency. Significantly expanding the known phenotypic spectrum, we identified compound heterozygous variants in SCO2 in two unrelated patients with axonal polyneuropathy, also known as Charcot-Marie-Tooth disease type 4. Different from previously described cases, our patients developed predominantly motor neuropathy, they survived infancy, and they have not yet developed the cardiomyopathy that causes death in early infancy in reported patients. Both of our patients harbour missense mutations near the conserved copper-binding motif (CXXXC), including the common pathogenic variant E140K and a novel change D135G. In addition, each patient carries a second mutation located at the same loop region, resulting in compound heterozygote changes E140K/P169T and D135G/R171Q. Patient fibroblasts showed reduced levels of SCO2, decreased copper levels and COX deficiency. Given that another Charcot-Marie-Tooth disease gene, ATP7A, is a known copper transporter, our findings further underline the relevance of copper metabolism in Charcot-Marie-Tooth disease.
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Proteínas Portadoras/genética , Enfermedad de Charcot-Marie-Tooth/complicaciones , Enfermedad de Charcot-Marie-Tooth/genética , Cobre/deficiencia , Proteínas Mitocondriales/genética , Mutación/genética , Adenosina Trifosfato/metabolismo , Adulto , Animales , Axones/patología , Proteínas Portadoras/metabolismo , Células Cultivadas , Enfermedad de Charcot-Marie-Tooth/diagnóstico por imagen , Enfermedad de Charcot-Marie-Tooth/patología , Niño , Análisis Mutacional de ADN , Complejo IV de Transporte de Electrones/metabolismo , Femenino , Fibroblastos/metabolismo , Fibroblastos/ultraestructura , Humanos , Imagen por Resonancia Magnética , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Proteínas Mitocondriales/metabolismo , Modelos Moleculares , Chaperonas Moleculares , Consumo de Oxígeno/genética , Nervio Ciático/metabolismo , Nervio Ciático/patología , Nervio Ciático/ultraestructuraRESUMEN
Recessive SLC25A46 mutations cause a spectrum of neurodegenerative disorders with optic atrophy as a core feature. We report a patient with optic atrophy, peripheral neuropathy, ataxia, but not cerebellar atrophy, who is on the mildest end of the phenotypic spectrum. By studying seven different nontruncating mutations, we found that the stability of the SLC25A46 protein inversely correlates with the severity of the disease and the patient's variant does not markedly destabilize the protein. SLC25A46 belongs to the mitochondrial transporter family, but it is not known to have transport function. Apart from this possible function, SLC25A46 forms molecular complexes with proteins involved in mitochondrial dynamics and cristae remodeling. We demonstrate that the patient's mutation directly affects the SLC25A46 interaction with MIC60. Furthermore, we mapped all of the reported substitutions in the protein onto a 3D model and found that half of them fall outside of the signature carrier motifs associated with transport function. We thus suggest that there are two distinct molecular mechanisms in SLC25A46-associated pathogenesis, one that destabilizes the protein while the other alters the molecular interactions of the protein. These results have the potential to inform clinical prognosis of such patients and indicate a pathway to drug target development.