RESUMO
BACKGROUND: The mitochondrial DNA m.3243A>G mutation can affect mitochondrial function and lead to a wide phenotypic spectrum, including mitochondrial encephalopathy with lactic acidosis and stroke-like episodes (MELAS) syndrome, diabetes mellitus, hearing impairment, cardiac involvement, epilepsy, migraine, myopathy, and cerebellar ataxia. However, m.3243A>G has been rarely reported in patients with cerebellar ataxia as their predominant manifestation. The aim of this study is to investigate the prevalence and clinical features of m.3243A>G in a Taiwanese cohort of cerebellar ataxia with unknown genetic diagnosis. METHODS: This retrospective cohort study conducted the mutation analysis of m.3243A>G by polymerase chain reaction and restriction fragment length polymorphism (PCR-RFLP) in 232 unrelated Han Chinese patients with genetically-undetermined cerebellar ataxia. The clinical presentation and neuroimaging features of patients with m.3243A>G mutation-related cerebellar ataxia were characterized. RESULTS: We identified two patients harboring m.3243A>G mutation. These patients have suffered from apparently sporadic and slowly progressive cerebellar ataxia since age 52 and 35 years, respectively. Both patients had diabetes mellitus and/or hearing impairment. The neuroimaging studies revealed generalized brain atrophy with predominantly cerebellar involvement in both individuals and bilateral basal ganglia calcifications in one of the patients. CONCLUSION: Mitochondrial m.3243A>G mutation accounted for 0.9% (2/232) of genetically-undetermined cerebellar ataxia in the Han Chinese cohort in Taiwan. These findings highlight the importance of investigating m.3243A>G in patients with genetically-undetermined cerebellar ataxia.
Assuntos
Ataxia Cerebelar , Diabetes Mellitus , Perda Auditiva , Humanos , Estudos Retrospectivos , Ataxia Cerebelar/genética , Mutação , DNA Mitocondrial/genéticaRESUMO
BACKGROUND: Mutations in the neurofilament light polypeptide gene (NEFL) are an uncommon cause of Charcot-Marie-Tooth disease (CMT). The aim of this study is to elucidate the clinical characteristics and genetic spectrum of NEFL-related neuropathy in a Taiwanese CMT cohort. METHODS: Mutational analysis of the coding regions of NEFL was performed by Sanger sequencing or targeted resequencing. Twenty-one patients from nine CMT pedigrees, identified from a cohort of 508 unrelated CMT patients, were found to have a NEFL mutation. Genetic, clinical and electrophysiological features were analyzed. RESULTS: Six NEFL mutations were identified, including two novel ones (p.P8S, p.N98Y). NEFL p.E396K was the most common mutation, accounting for 33.3% of the patients in our cohort. All patients manifested sensorimotor polyneuropathy with a mean age of disease onset of 13.5 ± 9.6 (1-40) years. Their motor nerve conduction velocities (MNCVs) of the ulnar nerve ranged from 22.1 to 48.7 m/s. Seventy percent of the patients could be classified as intermediate CMT with ulnar MNCVs between 25 and 45 m/s. Six of the 21 patients (28.6%) had additional features of central nervous system (CNS) involvement, including motor developmental delay, spasticity, cerebellar signs, neuropathic pain and scoliosis. CONCLUSION: NEFL mutations account for 1.8% (9/508) of the CMT patients in Taiwan. The present study delineates the clinical and genetic characteristics of NEFL-related neuropathy in Taiwan, and highlights that ulnar MNCV above 25 m/s and CNS involvement may serve as diagnostic clues for NEFL-related neuropathy.
Assuntos
Doença de Charcot-Marie-Tooth , Humanos , Pré-Escolar , Criança , Adolescente , Adulto Jovem , Adulto , Taiwan , Doença de Charcot-Marie-Tooth/genética , Mutação , Proteínas de Neurofilamentos/genéticaRESUMO
The ClC-2 channel plays a critical role in maintaining ion homeostasis in the brain and the testis. Loss-of-function mutations in the ClC-2-encoding human CLCN2 gene are linked to the white matter disease leukodystrophy. Clcn2-deficient mice display neuronal myelin vacuolation and testicular degeneration. Leukodystrophy-causing ClC-2 mutant channels are associated with anomalous proteostasis manifesting enhanced endoplasmic reticulum (ER)-associated degradation. The molecular nature of the ER quality control system for ClC-2 protein remains elusive. In mouse testicular tissues and Leydig cells, we demonstrated that endogenous ClC-2 co-existed in the same protein complex with the molecular chaperones heat shock protein 90ß (Hsp90ß) and heat shock cognate protein (Hsc70), as well as the associated co-chaperones Hsp70/Hsp90 organizing protein (HOP), activator of Hsp90 ATPase homolog 1 (Aha1), and FK506-binding protein 8 (FKBP8). Further biochemical analyses revealed that the Hsp90ß-Hsc70 chaperone/co-chaperone system promoted mouse and human ClC-2 protein biogenesis. FKBP8 additionally facilitated membrane trafficking of ClC-2 channels. Interestingly, treatment with the Hsp90-targeting small molecule 17-allylamino-17-demethoxygeldanamycin (17-AAG) substantially boosted ClC-2 protein expression. Also, 17-AAG effectively increased both total and cell surface protein levels of leukodystrophy-causing loss-of-function ClC-2 mutant channels. Our findings highlight the therapeutic potential of 17-AAG in correcting anomalous ClC-2 proteostasis associated with leukodystrophy.
Assuntos
Encéfalo/metabolismo , Canais de Cloreto/genética , Células Intersticiais do Testículo/metabolismo , Neurônios/metabolismo , Doença de Pelizaeus-Merzbacher/genética , Proteostase/genética , Animais , Benzoquinonas/farmacologia , Encéfalo/efeitos dos fármacos , Encéfalo/patologia , Células CHO , Canais de Cloro CLC-2 , Canais de Cloreto/deficiência , Cricetulus , Modelos Animais de Doenças , Degradação Associada com o Retículo Endoplasmático/efeitos dos fármacos , Regulação da Expressão Gênica , Células HEK293 , Proteínas de Choque Térmico HSP70/genética , Proteínas de Choque Térmico HSP70/metabolismo , Proteínas de Choque Térmico HSP90/genética , Proteínas de Choque Térmico HSP90/metabolismo , Humanos , Lactamas Macrocíclicas/farmacologia , Células Intersticiais do Testículo/efeitos dos fármacos , Células Intersticiais do Testículo/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Chaperonas Moleculares/genética , Chaperonas Moleculares/metabolismo , Neurônios/efeitos dos fármacos , Neurônios/patologia , Doença de Pelizaeus-Merzbacher/tratamento farmacológico , Doença de Pelizaeus-Merzbacher/metabolismo , Doença de Pelizaeus-Merzbacher/patologia , Isoformas de Proteínas/deficiência , Isoformas de Proteínas/genética , Transdução de Sinais , Proteínas de Ligação a Tacrolimo/genética , Proteínas de Ligação a Tacrolimo/metabolismoRESUMO
KCND3 encodes the voltage-gated potassium channel KV4.3 that is highly expressed in the cerebellum, where it regulates dendritic excitability and calcium influx. Loss-of-function KV4.3 mutations have been associated with dominant spinocerebellar ataxia (SCA19/22). By targeted NGS sequencing, we identified two novel KCND3 missense variants of the KV4.3 channel: p.S347W identified in a patient with adult-onset pure cerebellar syndrome and p.W359G detected in a child with congenital nonprogressive ataxia. Neuroimaging showed mild cerebellar atrophy in both patients. We performed a two-electrode voltage-clamp recording of KV4.3 currents in Xenopus oocytes: both the p.G345V (previously reported in a SCA19/22 family) and p.S347W mutants exhibited reduced peak currents by 50%, while no K+ current was detectable for the p.W359G mutant. We assessed the effect of the mutations on channel gating by measuring steady-state voltage-dependent activation and inactivation properties: no significant alterations were detected in p.G345V and p.S347W disease-associated variants, compared to controls. KV4.3 expression studies in HEK293T cells showed 53% (p.G345V), 45% (p.S347W) and 75% (p.W359G) reductions in mutant protein levels compared with the wildtype. The present study broadens the spectrum of the known phenotypes and identifies additional variants for KCND3-related disorders, outlining the importance of SCA gene screening in early-onset and congenital ataxia.
Assuntos
Ativação do Canal Iônico , Mutação/genética , Canais de Potássio Shal/genética , Ataxias Espinocerebelares/genética , Ataxias Espinocerebelares/fisiopatologia , Sequência de Aminoácidos , Animais , Criança , Feminino , Células HEK293 , Humanos , Imageamento por Ressonância Magnética , Masculino , Pessoa de Meia-Idade , Proteostase , Ataxias Espinocerebelares/diagnóstico por imagem , Xenopus laevisRESUMO
Loss-of-function mutations in the KV4.3 channel-encoding KCND3 gene are linked to neurodegenerative cerebellar ataxia. Patients suffering from neurodegeneration associated with iron deposition may also present with cerebellar ataxia. The mechanism underlying brain iron accumulation remains unclear. Here, we aim to ascertain the potential pathogenic role of KCND3 variant in iron accumulation-related cerebellar ataxia. We presented a patient with slowly progressive cerebellar ataxia, parkinsonism, cognitive impairment, and iron accumulation in the basal ganglia and the cerebellum. Whole exome sequencing analyses identified in the patient a heterozygous KCND3 c.1256G>A (p.R419H) variant predicted to be disease-causing by multiple bioinformatic analyses. In vitro biochemical and immunofluorescence examinations revealed that, compared to the human KV4.3 wild-type channel, the p.R419H variant exhibited normal protein abundance and subcellular localization pattern. Electrophysiological investigation, however, demonstrated that the KV4.3 p.R419H variant was associated with a dominant increase in potassium current amplitudes, as well as notable changes in voltage-dependent gating properties leading to enhanced potassium window current. These observations indicate that, in direct contrast with the loss-of-function KCND3 mutations previously reported in cerebellar ataxia patients, we identified a rare gain-of-function KCND3 variant that may expand the clinical and molecular spectra of neurodegenerative cerebellar disorders associated with brain iron accumulation.
Assuntos
Disfunção Cognitiva/genética , Mutação com Ganho de Função , Ferro/metabolismo , Transtornos Parkinsonianos/genética , Canais de Potássio Shal/genética , Ataxias Espinocerebelares/genética , Potenciais de Ação , Idoso , Encéfalo/metabolismo , Disfunção Cognitiva/patologia , Células HEK293 , Humanos , Masculino , Transtornos Parkinsonianos/patologia , Domínios Proteicos , Canais de Potássio Shal/química , Canais de Potássio Shal/metabolismo , Ataxias Espinocerebelares/patologiaRESUMO
Mutations in the human voltage-gated K+ channel subunit KV 4.3-encoding KCND3 gene have been associated with the autosomal dominant neurodegenerative disorder spinocerebellar ataxia types 19 and 22 (SCA19/22). The precise pathophysiology underlying the dominant inheritance pattern of SCA19/22 remains elusive. Using cerebellar ataxia-specific targeted next-generation sequencing technology, we identified two novel KCND3 mutations, c.950 G>A (p.C317Y) and c.1123 C>T (p.P375S) from a cohort with inherited cerebellar ataxias in Taiwan. The patients manifested notable phenotypic heterogeneity that includes cognitive impairment. We employed in vitro heterologous expression systems to inspect the biophysical and biochemical properties of human KV 4.3 harboring the two novel mutations, as well as two previously reported but uncharacterized disease-related mutations, c.1013 T>A (p.V338E) and c.1130 C>T (p.T377M). Electrophysiological analyses revealed that all of these SCA19/22-associated KV 4.3 mutant channels manifested loss-of-function phenotypes. Protein chemistry and immunofluorescence analyses further demonstrated that these mutants displayed enhanced protein degradation and defective membrane trafficking. By coexpressing KV 4.3 wild-type with the disease-related mutants, we provided direct evidence showing that the mutants instigated anomalous protein biosynthesis and channel gating of KV 4.3. We propose that the dominant inheritance pattern of SCA19/22 may be explained by the dominant-negative effects of the mutants on protein biosynthesis and voltage-dependent gating of KV 4.3 wild-type channel.
Assuntos
Ativação do Canal Iônico , Mutação , Biossíntese de Proteínas , Canais de Potássio Shal/metabolismo , Degenerações Espinocerebelares/genética , Degenerações Espinocerebelares/metabolismo , Adulto , Idoso , Alelos , Sequência de Aminoácidos , Animais , Linhagem Celular , Análise Mutacional de DNA , Feminino , Predisposição Genética para Doença , Genótipo , Humanos , Imageamento por Ressonância Magnética , Masculino , Pessoa de Meia-Idade , Modelos Moleculares , Linhagem , Fenótipo , Domínios Proteicos , Canais de Potássio Shal/química , Canais de Potássio Shal/genética , Degenerações Espinocerebelares/diagnóstico , Relação Estrutura-Atividade , Adulto JovemRESUMO
Distal hereditary motor neuropathy is a heterogeneous group of inherited neuropathies characterized by distal limb muscle weakness and atrophy. Although at least 15 genes have been implicated in distal hereditary motor neuropathy, the genetic causes remain elusive in many families. To identify an additional causal gene for distal hereditary motor neuropathy, we performed exome sequencing for two affected individuals and two unaffected members in a Taiwanese family with an autosomal dominant distal hereditary motor neuropathy in which mutations in common distal hereditary motor neuropathy-implicated genes had been excluded. The exome sequencing revealed a heterozygous mutation, c.770A > G (p.His257Arg), in the cytoplasmic tryptophanyl-tRNA synthetase (TrpRS) gene (WARS) that co-segregates with the neuropathy in the family. Further analyses of WARS in an additional 79 Taiwanese pedigrees with inherited neuropathies and 163 index cases from Australian, European, and Korean distal hereditary motor neuropathy families identified the same mutation in another Taiwanese distal hereditary motor neuropathy pedigree with different ancestries and one additional Belgian distal hereditary motor neuropathy family of Caucasian origin. Cell transfection studies demonstrated a dominant-negative effect of the p.His257Arg mutation on aminoacylation activity of TrpRS, which subsequently compromised protein synthesis and reduced cell viability. His257Arg TrpRS also inhibited neurite outgrowth and led to neurite degeneration in the neuronal cell lines and rat motor neurons. Further in vitro analyses showed that the WARS mutation could potentiate the angiostatic activities of TrpRS by enhancing its interaction with vascular endothelial-cadherin. Taken together, these findings establish WARS as a gene whose mutations may cause distal hereditary motor neuropathy and alter canonical and non-canonical functions of TrpRS.
Assuntos
Predisposição Genética para Doença/genética , Neuropatia Hereditária Motora e Sensorial/genética , Triptofano-tRNA Ligase/genética , Animais , Sobrevivência Celular , Células Cultivadas , Exoma/genética , Feminino , Humanos , Masculino , Camundongos , Mutação , Neuritos/patologia , Neuritos/fisiologia , Linhagem , Biossíntese de Proteínas/genética , Proteínas , Análise de Sequência de DNA , Triptofano-tRNA Ligase/metabolismoRESUMO
Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) is the most prevalent monogenic cerebral small vessel diseases caused by a mutation in the NOTCH3 gene. The clinical manifestations of CADASIL range from single or multiple lacunar infarcts, transient ischemic attacks, dementia, migraine with aura to psychiatric disorders. The features of brain MRI of CADASIL include multiple lacunar infarcts and diffuse leukoencephalopathy, which frequently involves external capsules and anterior temporal regions. Almost all patients with CADASIL harbor cysteine-involving mutations in NOTCH3. In Taiwan, two thirds of CADASIL patients carry NOTCH3 p.R544C mutations, and only approximately 56% of patients with CADASIL have leukoencephalopathy with anterior temporal regions involvement.
Assuntos
CADASIL/diagnóstico , CADASIL/etiologia , CADASIL/terapia , Humanos , Receptor Notch3 , Receptores Notch/genéticaRESUMO
OBJECTIVE: TFG mutations have previously been implicated in autosomal recessive hereditary spastic paraplegia (HSP), also known as SPG57. This study aimed to investigate the clinical and molecular features of TFG mutations in a Taiwanese HSP cohort. METHODS: Genetic analysis of TFG was conducted in 242 unrelated Taiwanese HSP patients using a targeted resequencing panel covering the entire coding regions of TFG. Functional assays were performed using an in vitro cell model to assess the impact of TFG variants on protein function. Additionally, other representative TFG mutant proteins were examined to understand the broader implications of TFG mutations in HSP. RESULTS: The study identified a novel homozygous TFG c.177A>C (p.(Lys59Asn)) variant in a family with adolescent-onset, pure form HSP. Functional analysis revealed that the Lys59Asn TFG variant, similar to other HSP-associated TFG mutants, exhibited a low affinity between TFG monomers and abnormal assembly of TFG homo-oligomers. These structural alterations led to aberrant intracellular distribution, compromising TFG's protein secretion function and resulting in decreased cellular viability. INTERPRETATION: These findings confirm that the homozygous TFG c.177A>C (p.(Lys59Asn)) variant is a novel cause of SPG57. The study expands our understanding of the clinical and mutational spectrum of TFG-associated diseases, highlighting the functional defects associated with this specific TFG variant. Overall, this research contributes to the broader comprehension of the genetic and molecular mechanisms underlying HSP.
Assuntos
Paraplegia Espástica Hereditária , Humanos , Paraplegia Espástica Hereditária/genética , Masculino , Feminino , Linhagem , Adulto , Adolescente , Mutação , Adulto Jovem , Proteínas de Transporte VesicularRESUMO
PAR3/INSC/LGN form an evolutionarily conserved complex required for asymmetric cell division in the developing brain, but its post-developmental function and disease relevance in the peripheral nervous system (PNS) remains unknown. We mapped a new locus for axonal Charcot-Marie-Tooth disease (CMT2) and identified a missense mutation c.209 T > G (p.Met70Arg) in the INSC gene. Modeling the INSCM70R variant in Drosophila, we showed that it caused proprioceptive defects in adult flies, leading to gait defects resembling those in CMT2 patients. Cellularly, PAR3/INSC/LGN dysfunction caused tubulin aggregation and necrotic neurodegeneration, with microtubule-stabilizing agents rescuing both morphological and functional defects of the INSCM70R mutation in the PNS. Our findings underscore the critical role of the PAR3/INSC/LGN machinery in the adult PNS and highlight a potential therapeutic target for INSC-associated CMT2.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal , Doença de Charcot-Marie-Tooth , Proteínas do Citoesqueleto , Mutação de Sentido Incorreto , Animais , Humanos , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Doença de Charcot-Marie-Tooth/genética , Doença de Charcot-Marie-Tooth/patologia , Modelos Animais de Doenças , Drosophila/genética , Proteínas Nucleares , Doenças do Sistema Nervoso Periférico/genética , Doenças do Sistema Nervoso Periférico/patologia , Tubulina (Proteína)/genética , Tubulina (Proteína)/metabolismo , Proteínas do Citoesqueleto/genética , Proteínas do Citoesqueleto/metabolismo , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismoRESUMO
BACKGROUND AND OBJECTIVES: The GGC repeat expansion in the 5' untranslated region of NOTCH2NLC was recently identified as the cause of neuronal intranuclear inclusion disease (NIID), which may manifest with peripheral neuropathy. The aim of this study is to investigate its contribution to inherited neuropathy. METHODS: This cohort study screened patients with molecularly undiagnosed Charcot-Marie-Tooth disease (CMT) and healthy controls for the GGC repeat expansion in NOTCH2NLC using repeat-primed PCR and fragment analysis. The clinical and electrophysiologic features of the patients harboring the GGC repeat expansion were scrutinized. Skin biopsy with immunohistochemistry staining and electric microscopic imaging were performed. RESULTS: One hundred twenty-seven unrelated patients with CMT, including 66 cases with axonal CMT (CMT2), and 200 healthy controls were included. Among them, 7 patients with CMT carried a variant NOTCH2NLC allele with GGC repeat expansion, but it was absent in controls. The sizes of the expanded GGC repeats ranged from 80 to 104 repeats. All 7 patients developed sensory predominant neuropathy with an average age at disease onset of 37.1 years (range 21-55 years). Electrophysiologic studies revealed mild axonal sensorimotor polyneuropathy. Leukoencephalopathy was absent in the 5 patients who received a brain MRI. Skin biopsy from 2 patients showed eosinophilic, ubiquitin- and p62-positive intranuclear inclusions in the sweat gland cells and dermal fibroblasts. Two of the 7 patients had a family history of NIID. DISCUSSION: The NOTCH2NLC GGC repeat expansions are an underdiagnosed and important cause of inherited neuropathy. The expansion accounts for 10.6% (7 of 66) of molecularly unassigned CMT2 cases in the Taiwanese CMT cohort. CLASSIFICATION OF EVIDENCE: This study provides Class III evidence that in Taiwanese patients with genetically undiagnosed CMT, 10.6% of the CMT2 cases have the GGC repeat expansion in NOTCH2NLC.
Assuntos
Peptídeos e Proteínas de Sinalização Intercelular , Proteínas do Tecido Nervoso , Doenças Neurodegenerativas , Doenças do Sistema Nervoso Periférico , Adulto , Estudos de Casos e Controles , Estudos de Coortes , Humanos , Peptídeos e Proteínas de Sinalização Intercelular/genética , Corpos de Inclusão Intranuclear/patologia , Pessoa de Meia-Idade , Proteínas do Tecido Nervoso/genética , Doenças Neurodegenerativas/patologia , Doenças do Sistema Nervoso Periférico/patologia , Expansão das Repetições de Trinucleotídeos , Adulto JovemRESUMO
BACKGROUND: Adrenoleukodystrophy (ALD) is an X-linked peroxisomal disorder caused by mutations in the ABCD1 gene. The clinical manifestations of ALD vary widely with some patients presenting with adrenomyeloneuropathy (AMN) that resembles the phenotype of hereditary spastic paraplegia (HSP). The aim of this study is to investigate the frequency, spectrum, and clinical features of ABCD1 mutations in Taiwanese patients with HSP phenotype. METHODS: Mutational analysis of the ABCD1 gene was performed in 230 unrelated Taiwanese patients with clinically suspected HSP by targeted resequencing. Clinical, electrophysiological, and neuroimaging features of the patients carrying an ABCD1 pathogenic mutation were characterized. RESULTS: Ten different ABCD1 mutations were identified in eleven patients, including two novel mutations (p.Q177Pfs*17 and p.Y357*) and eight ever reported in ALD cases of other ethnicities. All patients were male and exhibited slowly progressive spastic paraparesis with onset ages ranging from 21 to 50 years. Most of them had additional non-motor symptoms, including autonomic dysfunction in nine patients, sensory deficits in seven, premature baldness in seven, skin hyperpigmentation in five, psychiatric symptoms in one and cerebellar ataxia in one. Seven of the ten patients who ever received nerve conduction studies showed axonal polyneuropathy. Magnetic resonance imaging (MRI) revealed diffuse spinal cord atrophy in seven patients, cerebral white matter hyperintensity in one patient, and cerebellar involvement in one patient. CONCLUSIONS: ABCD1 mutations account for 4.8% (11/230) of the cases with HSP phenotype in Taiwan. This study highlights the importance to consider ABCD1 mutations in patients with clinically suspected HSP of unknown genetic causes.
Assuntos
Membro 1 da Subfamília D de Transportadores de Cassetes de Ligação de ATP/genética , Adrenoleucodistrofia/genética , Povo Asiático/genética , Paraplegia Espástica Hereditária/genética , Adulto , Idade de Início , Estudos de Coortes , Análise Mutacional de DNA , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Mutação/genética , Fenótipo , Taiwan , Adulto JovemRESUMO
The voltage-dependent ClC-1 chloride channel, whose open probability increases with membrane potential depolarization, belongs to the superfamily of CLC channels/transporters. ClC-1 is almost exclusively expressed in skeletal muscles and is essential for stabilizing the excitability of muscle membranes. Elucidation of the molecular structures of human ClC-1 and several CLC homologs provides important insight to the gating and ion permeation mechanisms of this chloride channel. Mutations in the human CLCN1 gene, which encodes the ClC-1 channel, are associated with a hereditary skeletal muscle disease, myotonia congenita. Most disease-causing CLCN1 mutations lead to loss-of-function phenotypes in the ClC-1 channel and thus increase membrane excitability in skeletal muscles, consequently manifesting as delayed relaxations following voluntary muscle contractions in myotonic subjects. The inheritance pattern of myotonia congenita can be autosomal dominant (Thomsen type) or recessive (Becker type). To date over 200 myotonia-associated ClC-1 mutations have been identified, which are scattered throughout the entire protein sequence. The dominant inheritance pattern of some myotonia mutations may be explained by a dominant-negative effect on ClC-1 channel gating. For many other myotonia mutations, however, no clear relationship can be established between the inheritance pattern and the location of the mutation in the ClC-1 protein. Emerging evidence indicates that the effects of some mutations may entail impaired ClC-1 protein homeostasis (proteostasis). Proteostasis of membrane proteins comprises of biogenesis at the endoplasmic reticulum (ER), trafficking to the surface membrane, and protein turn-over at the plasma membrane. Maintenance of proteostasis requires the coordination of a wide variety of different molecular chaperones and protein quality control factors. A number of regulatory molecules have recently been shown to contribute to post-translational modifications of ClC-1 and play critical roles in the ER quality control, membrane trafficking, and peripheral quality control of this chloride channel. Further illumination of the mechanisms of ClC-1 proteostasis network will enhance our understanding of the molecular pathophysiology of myotonia congenita, and may also bring to light novel therapeutic targets for skeletal muscle dysfunction caused by myotonia and other pathological conditions.
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Voltage-gated ClC-2 channels are essential for chloride homeostasis. Complete knockout of mouse ClC-2 leads to testicular degeneration and neuronal myelin vacuolation. Gain-of-function and loss-of-function mutations in the ClC-2-encoding human CLCN2 gene are linked to the genetic diseases aldosteronism and leukodystrophy, respectively. The protein homeostasis (proteostasis) mechanism of ClC-2 is currently unclear. Here, we aimed to identify the molecular mechanism of endoplasmic reticulum-associated degradation of ClC-2, and to explore the pathophysiological significance of disease-associated anomalous ClC-2 proteostasis. In both heterologous expression system and native neuronal and testicular cells, ClC-2 is subject to significant regulation by cullin-RING E3 ligase-mediated polyubiquitination and proteasomal degradation. The cullin 4 (CUL4)-damage-specific DNA binding protein 1 (DDB1)-cereblon (CRBN) E3 ubiquitin ligase co-exists in the same complex with and promotes the degradation of ClC-2 channels. The CRBN-targeting immunomodulatory drug lenalidomide and the cullin E3 ligase inhibitor MLN4924 promotes and attenuates, respectively, proteasomal degradation of ClC-2. Analyses of disease-related ClC-2 mutants reveal that aldosteronism and leukodystrophy are associated with opposite alterations in ClC-2 proteostasis. Modifying CUL4 E3 ligase activity with lenalidomide and MLN4924 ameliorates disease-associated ClC-2 proteostasis abnormality. Our results highlight the significant role and therapeutic potential of CUL4 E3 ubiquitin ligase in regulating ClC-2 proteostasis.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Encefalopatias/metabolismo , Canais de Cloreto/metabolismo , Proteínas Culina/metabolismo , Proteínas de Ligação a DNA/metabolismo , Hiperaldosteronismo/metabolismo , Proteostase , Ubiquitina-Proteína Ligases/metabolismo , Animais , Encefalopatias/patologia , Canais de Cloro CLC-2 , Células HEK293 , Humanos , Hiperaldosteronismo/patologia , Camundongos Endogâmicos C57BL , Modelos Biológicos , Poliubiquitina/metabolismo , Proteólise , Ratos Wistar , Especificidade por Substrato , UbiquitinaçãoRESUMO
INTRODUCTION: Mutations in the PUM1 gene were recently identified to cause spinocerebellar ataxia type 47 (SCA47). However, their role in cerebellar ataxia in various populations remains elusive. The aim of this study was to elucidate the frequency and spectrum of PUM1 mutations in a cohort of Taiwanese patients with molecularly undetermined cerebellar ataxia. METHODS: Mutational analyses of PUM1 were performed by Sanger sequencing in a cohort of 248 unrelated patients with cerebellar ataxia of unknown cause, including 108 with autosomal-dominantly inherited cerebellar ataxia, 45 with autosomal-recessively inherited cerebellar ataxia, and 95 with apparently sporadic cerebellar ataxia. Among them, the genetic causes of ataxia remained unknown after excluding mutations responsible for SCA1, 2, 3, 6, 7, 8, 10, 12, 17, 19/22, 23, 26, 27, 28, 31, 35, 36, dentatorubral-pallidoluysian atrophy and Friedreich's ataxia. RESULTS: Two heterozygous missense PUM1 variants were identified in two patients with apparently sporadic cerebellar ataxia, including a known disease-causing mutation (p.R1139W) and a variant of uncertain significance (p.K151R). The patient carrying the p.R1139W mutation had a slowly progressive, relatively pure cerebellar ataxia, presenting with gait unsteadiness, limb dysmetria, ataxic dysarthria and saccadic pursuit. CONCLUSION: Our findings support the pathogenic role of PUM1 mutations in cerebellar ataxia and emphasize the importance of considering PUM1 mutations as a possible etiology of cerebellar ataxia.
Assuntos
Ataxia Cerebelar/genética , Proteínas de Ligação a RNA/genética , Adulto , Ataxia Cerebelar/diagnóstico por imagem , Estudos de Coortes , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Mutação , TaiwanRESUMO
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RESUMO
BACKGROUND: The inositol 1,4,5-triphosphate (IP3) receptor type 1 gene (ITPR1) encodes the IP3 receptor type 1 (IP3R1), which modulates intracellular calcium homeostasis and signaling. Mutations in ITPR1 have been implicated in inherited cerebellar ataxias. The aim of this study was to investigate the role of ITPR1 mutations, including both large segmental deletion and single nucleotide mutations, in a Han Chinese cohort with inherited cerebellar ataxias in Taiwan. METHODOLOGY AND PRINCIPAL FINDINGS: Ninety-three unrelated individuals with molecularly unassigned spinocerebellar ataxia selected from 585 pedigrees with autosomal dominant cerebellar ataxias, were recruited into the study with elaborate clinical evaluations. The quantitative PCR technique was used to survey large segmental deletion of ITPR1 and a targeted sequencing approach was applied to sequence all of the 61 exons and the flanking regions of ITPR1. A novel ITPR1 mutation, c.7721T>C (p.V2574A), was identified in a family with dominantly inherited cerebellar ataxia. The proband has an adult-onset non-progressive pure cerebellar ataxia and her daughter is afflicted with a childhood onset cerebellar ataxia with intellectual sub-normalities. CONCLUSION: ITPR1 mutation is an uncommon cause of inherited cerebellar ataxia, accounting for 0.2% (1/585) of patients with dominantly inherited cerebellar ataxias in Taiwan. This study broadens the mutational spectrum of ITPR1 and also emphasizes the importance of considering ITPR1 mutations as a potential cause of inherited cerebellar ataxias.
Assuntos
Ataxia Cerebelar/genética , Receptores de Inositol 1,4,5-Trifosfato/genética , Adulto , Estudos de Coortes , Feminino , Humanos , Masculino , Linhagem , TaiwanRESUMO
Adrenoleukodystrophy (ALD) is a rare and progressive neurogenetic disease that may manifest disparate symptoms. The present study aims at investigating the role of ataxic variant of ALD (AVALD) in patients with adult-onset cerebellar ataxia, as well as characterizing their clinical features that distinguish AVALD from other cerebellar ataxias. Mutations in the ATP binding cassette subfamily D member 1 gene (ABCD1) were ascertained in 516 unrelated patients with ataxia. The patients were categorized into three groups: molecularly unassigned hereditary ataxia (n = 118), sporadic ataxia with autonomic dysfunctions (n = 296), and sporadic ataxia without autonomic dysfunctions (n = 102). Brain MRIs were scrutinized for white matter hyperintensity (WMH) in the parieto-occipital lobes, frontal lobes, corticospinal tracts, pons, middle cerebellar peduncles and cerebellar hemispheres. Two ABCD1 mutations (p.S108L and p.P623fs) previously linked to cerebral ALD and adrenomyeloneuropathy but not AVALD were identified. ALD accounts for 0.85% (1/118) of the patients with molecularly unassigned hereditary ataxia and 0.34% (1/296) of the patients with sporadic ataxia with autonomic dysfunctions. WMH in the corticospinal tracts and WMH in the cerebellar hemispheres were strongly associated with AVALD rather than other ataxias. To conclude, ALD accounts for approximately 0.39% (2/516) of adult-onset cerebellar ataxias. This study expands the mutational spectrum of AVALD and underscores the importance of considering ALD as a potential etiology of cerebellar ataxia.