RESUMO
BACKGROUND: Variants in the ring finger protein 213 (RNF213) gene are known to be associated with increased predisposition to cerebrovascular diseases development. Genomic studies have identified RNF213 as a major risk factor of Moyamoya disease in East Asian descendants. However, little is known about the RNF213 (ring finger protein 213) biological functions or its associated pathogenic mechanisms underlying Moyamoya disease. METHODS: To investigate RNF213 loss-of-function effect in endothelial cell, stable RNF213-deficient human cerebral endothelial cells were generated using the CRISPR-Cas9 genome editing technology. RESULTS: In vitro assays, using RNF213 knockout brain endothelial cells, showed clear morphological changes and increased blood-brain barrier permeability. Downregulation and delocalization of essential interendothelial junction proteins involved in the blood-brain barrier maintenance, such as PECAM-1 (platelet endothelial cell adhesion molecule-1), was also observed. Brain endothelial RNF213-deficient cells also showed an abnormal potential to transmigration of leukocytes and secreted high amounts of proinflammatory cytokines. CONCLUSIONS: Taken together, these results indicate that RNF213 could be a key regulator of cerebral endothelium integrity, whose disruption could be an early pathological mechanism leading to Moyamoya disease. This study also further reinforces the importance of blood-brain barrier integrity in the development of Moyamoya disease and other RNF213-associated diseases.
Assuntos
Adenosina Trifosfatases , Doença de Moyamoya , Ubiquitina-Proteína Ligases , Adenosina Trifosfatases/genética , Células Endoteliais/metabolismo , Endotélio , Predisposição Genética para Doença , Humanos , Doença de Moyamoya/patologia , Fatores de Transcrição , Ubiquitina-Proteína Ligases/genéticaRESUMO
BACKGROUND: Although the typical inheritance of spastic paraplegia 7 is recessive, several reports have suggested that SPG7 variants may also cause autosomal dominant hereditary spastic paraplegia (HSP). OBJECTIVES: We aimed to conduct an exome-wide genetic analysis on a large Canadian cohort of HSP patients and controls to examine the association of SPG7 and HSP. METHODS: We analyzed 585 HSP patients from 372 families and 1175 controls, including 580 unrelated individuals. Whole-exome sequencing was performed on 400 HSP patients (291 index cases) and all 1175 controls. RESULTS: The frequency of heterozygous pathogenic/likely pathogenic SPG7 variants (4.8%) among unrelated HSP patients was higher than among unrelated controls (1.7%; OR 2.88, 95% CI 1.24-6.66, P = 0.009). The heterozygous SPG7 p.(Ala510Val) variant was found in 3.7% of index patients versus 0.85% in unrelated controls (OR 4.42, 95% CI 1.49-13.07, P = 0.005). Similar results were obtained after including only genetically-undiagnosed patients. We identified four heterozygous SPG7 variant carriers with an additional pathogenic variant in known HSP genes, compared to zero in controls (OR 19.58, 95% CI 1.05-365.13, P = 0.0031), indicating potential digenic inheritance. We further identified four families with heterozygous variants in SPG7 and SPG7-interacting genes (CACNA1A, AFG3L2, and MORC2). Of these, there is especially compelling evidence for epistasis between SPG7 and AFG3L2. The p.(Ile705Thr) variant in AFG3L2 is located at the interface between hexamer subunits, in a hotspot of mutations associated with spinocerebellar ataxia type 28 that affect its proteolytic function. CONCLUSIONS: Our results provide evidence for complex inheritance in SPG7-associated HSP, which may include recessive and possibly dominant and digenic/epistasis forms of inheritance. © 2021 International Parkinson and Movement Disorder Society.
Assuntos
Paraplegia Espástica Hereditária , Proteases Dependentes de ATP , ATPases Associadas a Diversas Atividades Celulares/genética , Canadá , Humanos , Metaloendopeptidases/genética , Mutação/genética , Paraplegia , Paraplegia Espástica Hereditária/genética , Fatores de TranscriçãoRESUMO
Childhood-onset schizophrenia (COS) is a rare and severe form of schizophrenia defined as onset before age of 13. Here we report on two unrelated cases diagnosed with both COS and alternating hemiplegia of childhood (AHC), and for whom two distinct pathogenic de novo variants were identified in the ATP1A3 gene. ATP1A3 encodes the α-subunit of a neuron-specific ATP-dependent transmembrane sodium-potassium pump. Using whole exome sequencing data derived from a cohort of 17 unrelated COS cases, we also examined ATP1A3 and all of its interactors known to be expressed in the brain to establish if variants could be identified. This led to the identification of a third case with a possibly damaging missense mutation in ATP1A3 and three others cases with predicted pathogenic missense variants in the FXYD gene family (FXYD1, FXYD6, and FXYD6-FXYD2 readthrough). FXYD genes encode proteins that modulate the ATP-dependant pump function. This report is the first to identify variants in the same pathway for COS. Our COS study illustrates the interest of stratifying a complex condition according to the age of onset for the identification of deleterious variants. Whereas ATP1A3 is a replicated gene in rare neuropediatric diseases, this gene has previously been linked with COS in only one case report. The association with rare variants in FXYD gene family is novel and highlights the interest of exploring these genes in COS as well as in pediatric neurodevelopmental disorders.
Assuntos
Proteínas de Membrana/genética , Fosfoproteínas/genética , Esquizofrenia Infantil/genética , ATPase Trocadora de Sódio-Potássio/genética , Pré-Escolar , Feminino , Humanos , Lactente , Masculino , Proteínas de Membrana/metabolismo , Mutação/genética , Mutação de Sentido Incorreto/genética , Fosfoproteínas/metabolismo , Esquizofrenia Infantil/fisiopatologia , ATPase Trocadora de Sódio-Potássio/metabolismoRESUMO
Restless legs syndrome is a common complex disorder with different genetic and environmental risk factors. Here we used human cell lines to conduct an RNA-Seq study and observed how the gene showing the most significant association with RLS, MEIS1, acts as a regulator of the expression of many other genes. Some of the genes affected by its expression level are linked to pathways previously reported to be associated with RLS. We found that in cells where MEIS1 expression was either increased or prevented, mineral absorption is the principal dysregulated pathway. The mineral absorption pathway genes, HMOX1 and VDR are involved in iron metabolism and response to vitamin D, respectively. This shows a strong functional link to the known RLS pathways. We observed the same enrichment of the mineral absorption pathway in postmortem brain tissues of RLS patients showing a reduced expression of MEIS1. The expression of genes encoding metallothioneins (MTs) was observed to be dysregulated across the RNA-Seq datasets generated from both human cells and tissues. MTs are highly relevant to RLS as they bind intracellular metals, protect against oxidative stress and interact with ferritins which manage iron level in the central nervous system. Overall, our study suggests that in a subset of RLS patients, the contribution of MEIS1 appears to be associated to its downstream regulation of genes that are more directly involved in pathways that are relevant to RLS. While MTs have been implicated in the pathogenesis of neurodegenerative diseases such as Parkinson's diseases, this is a first report to propose that they have a role in RLS.
Assuntos
Encéfalo/metabolismo , Regulação da Expressão Gênica , Minerais/metabolismo , Proteína Meis1/genética , Síndrome das Pernas Inquietas/genética , Síndrome das Pernas Inquietas/metabolismo , Biomarcadores , Linhagem Celular Tumoral , Humanos , Redes e Vias MetabólicasRESUMO
BACKGROUND: Amyotrophic lateral sclerosis [1] is a genetically heterogeneous neurodegenerative disorder, characterized by late-onset degeneration of motor neurons leading to progressive limb and bulbar weakness, as well as of the respiratory muscles, which is the primary cause of disease fatality. To date, over 25 genes have been implicated as causative in ALS with C9orf72, SOD1, FUS, and TARDBP accounting for the majority of genetically positive cases. RESULTS: We identified two patients of Italian and French ancestry with a clinical diagnosis of juvenile-onset ALS who were mutation-negative in any of the known ALS causative genes. Starting with the index case, a consanguineous family of Italian origin, we performed whole-exome sequencing and identified candidate pathogenic mutations in 35 genes, 27 of which were homozygous. We next parsed all candidates against a cohort of 3641 ALS cases; only ATP13A2 was found to harbor recessive changes, in a patient with juvenile-onset ALS, similar to the index case. In vivo complementation of ATP13A2 using a zebrafish surrogate model that focused on the assessment of motor neuron morphology and cerebellar integrity confirmed the role of this gene in central and peripheral nervous system maintenance and corroborated the damaging direction of effect of the change detected in the index case of this study. CONCLUSIONS: We here expand the phenotypic spectrum associated with genetic variants in ATP13A2 that previously comprised Kufor-Rakeb syndrome, spastic paraplegia 78, and neuronal ceroid lipofuscinosis type 12 (CLN12), to also include juvenile-onset ALS, as supported by both genetic and functional data. Our findings highlight the importance of establishing a complete genetic profile towards obtaining an accurate clinical diagnosis.
Assuntos
Esclerose Lateral Amiotrófica/genética , Predisposição Genética para Doença , ATPases Translocadoras de Prótons/genética , Adulto , Esclerose Lateral Amiotrófica/patologia , Animais , Modelos Animais de Doenças , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Neurônios Motores/patologia , Mutação/genética , Lipofuscinoses Ceroides Neuronais/genética , Lipofuscinoses Ceroides Neuronais/patologia , Transtornos Parkinsonianos/genética , Transtornos Parkinsonianos/patologia , Linhagem , Fenótipo , Sequenciamento do Exoma , Peixe-ZebraRESUMO
Intracranial Aneurysm (IA) is a common disease with a worldwide prevalence of 1-3%. In the French-Canadian (FC) population, where there is an important founder effect, the incidence of IA is higher and is frequently seen in families. In this study, we genotyped a cohort of 257 mostly familial FC IA patients and 1,992 FC controls using the Illumina NeuroX SNP-chip. The most strongly associated loci were tested in 34 Inuit IA families and in 32 FC IA patients and 106 FC controls that had been exome sequenced (WES). After imputation, one locus at 3p14.2 (FHIT, rs1554600, p = 4.66 × 10-9) reached a genome-wide significant level of association and a subsequent validation in Nunavik Inuit cohort further confirmed the significance of the FHIT variant association (rs780365, FBAT-O, p = 0.002839). Additionally, among the other promising loci (p < 5 × 10-6), the one at 3q13.2 (rs78125721, p = 4.77 × 10-7), which encompasses CCDC80, also showed an increased mutation burden in the WES data (CCDC80, SKAT-O, p = 0.0005). In this study, we identified two new potential IA loci in the FC population: FHIT, which is significantly associated with hypertensive IA, and CCDC80, which has potential genetic and functional relevance to IA pathogenesis, providing evidence on the additional risk loci for familial IA. We also replicated the previous IA GWAS risk locus 18q11.2, and suggested a potential locus at 8p23.1 that warrants further study.
Assuntos
Hidrolases Anidrido Ácido/genética , Loci Gênicos , Predisposição Genética para Doença , Glicoproteínas/genética , Peptídeos e Proteínas de Sinalização Intercelular/genética , Aneurisma Intracraniano/genética , Proteínas de Neoplasias/genética , Adulto , Idoso , Canadá , Estudos de Coortes , Proteínas da Matriz Extracelular , Feminino , Efeito Fundador , Estudo de Associação Genômica Ampla , Humanos , Masculino , Pessoa de Meia-Idade , Mutação , Polimorfismo de Nucleotídeo ÚnicoRESUMO
Loss-of-function mutations in the potassium-chloride cotransporter KCC3 lead to Andermann syndrome, a severe sensorimotor neuropathy characterized by areflexia, amyotrophy and locomotor abnormalities. The molecular events responsible for axonal loss remain poorly understood. Here, we establish that global or neuron-specific KCC3 loss-of-function in mice leads to early neuromuscular junction (NMJ) abnormalities and muscular atrophy that are consistent with the pre-synaptic neurotransmission defects observed in patients. KCC3 depletion does not modify chloride handling, but promotes an abnormal electrical activity among primary motoneurons and mislocalization of Na+/K+-ATPase α1 in spinal cord motoneurons. Moreover, the activity-targeting drug carbamazepine restores Na+/K+-ATPase α1 localization and reduces NMJ denervation in Slc12a6-/- mice. We here propose that abnormal motoneuron electrical activity contributes to the peripheral neuropathy observed in Andermann syndrome.
Assuntos
Agenesia do Corpo Caloso/metabolismo , Neurônios Motores/metabolismo , Junção Neuromuscular/metabolismo , Doenças do Sistema Nervoso Periférico/metabolismo , Terminações Pré-Sinápticas/metabolismo , Simportadores/deficiência , Transmissão Sináptica/fisiologia , Agenesia do Corpo Caloso/tratamento farmacológico , Agenesia do Corpo Caloso/patologia , Animais , Carbamazepina/farmacologia , Células Cultivadas , Cloretos/metabolismo , Modelos Animais de Doenças , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Neurônios Motores/efeitos dos fármacos , Neurônios Motores/patologia , Junção Neuromuscular/efeitos dos fármacos , Junção Neuromuscular/patologia , Neurotransmissores/farmacologia , Doenças do Sistema Nervoso Periférico/tratamento farmacológico , Doenças do Sistema Nervoso Periférico/patologia , Terminações Pré-Sinápticas/efeitos dos fármacos , Terminações Pré-Sinápticas/patologia , ATPase Trocadora de Sódio-Potássio/metabolismo , Medula Espinal/efeitos dos fármacos , Medula Espinal/metabolismo , Medula Espinal/patologia , Simportadores/genética , Transmissão Sináptica/efeitos dos fármacosRESUMO
Intracranial aneurysms (IAs) are the result of focal weakness in the artery wall and have a complex genetic makeup. To date, genome-wide association and sequencing studies have had limited success in identifying IA risk factors. Distinct populations, such as the French-Canadian (FC) population, have increased IA prevalence. In our study, we used exome sequencing to prioritize risk variants in a discovery cohort of six FC families affected by IA, and the analysis revealed an increased variation burden for ring finger protein 213 (RNF213). We resequenced RNF213 in a larger FC validation cohort, and association tests on further identified variants supported our findings (SKAT-O, p = 0.006). RNF213 belongs to the AAA+ protein family, and two variants (p.Arg2438Cys and p.Ala2826Thr) unique to affected FC individuals were found to have increased ATPase activity, which could lead to increased risk of IA by elevating angiogenic activities. Common SNPs in RNF213 were also extracted from the NeuroX SNP-chip genotype data, comprising 257 FC IA-affected and 1,988 control individuals. We discovered that the non-ancestral allele of rs6565666 was significantly associated with the affected individuals (p = 0.03), and it appeared as though the frequency of the risk allele had changed through genetic drift. Although RNF213 is a risk factor for moyamoya disease in East Asians, we demonstrated that it might also be a risk factor for IA in the FC population. It therefore appears that the function of RNF213 can be differently altered to predispose distinct populations to dissimilar neurovascular conditions, highlighting the importance of a population's background in genetic studies of heterogeneous disease.
Assuntos
Adenosina Trifosfatases/genética , Aneurisma Intracraniano/genética , Ubiquitina-Proteína Ligases/genética , População Branca/genética , Adulto , Idoso , Alelos , Canadá , Estudos de Casos e Controles , Estudos de Coortes , Feminino , Estudo de Associação Genômica Ampla , Genótipo , Técnicas de Genotipagem , Humanos , Aneurisma Intracraniano/diagnóstico , Masculino , Pessoa de Meia-Idade , Linhagem , Polimorfismo de Nucleotídeo Único , Reprodutibilidade dos Testes , Análise de Sequência de DNARESUMO
Hereditary motor and sensory neuropathy associated with agenesis of the corpus callosum (HMSN/ACC) is an autosomal recessive disease of the central and peripheral nervous system that presents as early-onset polyneuropathy. Patients are hypotonic and areflexic from birth, with abnormal facial features and atrophic muscles. Progressive peripheral neuropathy eventually confines them to a wheelchair in the second decade of life, and death occurs by the fourth decade. We here define the neuropathologic features of the disease in autopsy tissues from eight cases. Both developmental and neurodegenerative features were found. Hypoplasia or absence of the major telencephalic commissures and a hypoplasia of corticospinal tracts to half the normal size, were the major neurodevelopmental defects we observed. Despite being a neurodegenerative disease, preservation of brain weight and a conspicuous absence of neuronal or glial cell death were signal features of this disease. Small tumor-like overgrowths of axons, termed axonomas, were found in the central and peripheral nervous system, indicating attempted axonal regeneration. We conclude that the neurodegenerative deficits in HMSN/ACC are primarily caused by an axonopathy superimposed upon abnormal development, affecting peripheral but also central nervous system axons, all ultimately because of a genetic defect in the axonal cotransporter KCC3.
Assuntos
Agenesia do Corpo Caloso/patologia , Axônios/patologia , Encéfalo/patologia , Doenças do Sistema Nervoso Periférico/patologia , Sistema Nervoso Periférico/patologia , Simportadores/genética , Adulto , Agenesia do Corpo Caloso/genética , Agenesia do Corpo Caloso/metabolismo , Agenesia do Corpo Caloso/fisiopatologia , Autopsia , Axônios/metabolismo , Encéfalo/metabolismo , Encéfalo/fisiopatologia , Feminino , Predisposição Genética para Doença , Humanos , Masculino , Degeneração Neural , Sistema Nervoso Periférico/metabolismo , Sistema Nervoso Periférico/fisiopatologia , Doenças do Sistema Nervoso Periférico/genética , Doenças do Sistema Nervoso Periférico/metabolismo , Doenças do Sistema Nervoso Periférico/fisiopatologia , Fenótipo , Prognóstico , Simportadores/metabolismo , Adulto JovemRESUMO
The MC1R gene, suggested to be involved in Parkinson disease (PD) and melanoma, was sequenced in PD patients (n = 539) and controls (n = 265) from New York, and PD patients (n = 551), rapid eye movement sleep behavior disorder (RBD) patients (n = 351), and controls (n = 956) of European ancestry. Sixty-eight MC1R variants were identified, including 7 common variants with frequency > 0.01. None of the common variants was associated with PD or RBD in the different regression models. In a meta-analysis with fixed-effect model, the p.R160W variant was associated with an increased risk for PD (odds ratio = 1.22, 95% confidence interval = 1.02-1.47, p = 0.03) but with significant heterogeneity (p = 0.048). Removing one study that introduced the heterogeneity resulted in nonsignificant association (odds ratio = 1.11, 95% confidence interval, 0.92-1.35, p = 0.27, heterogeneity p = 0.57). Rare variants had similar frequencies in patients and controls (10.54% and 10.15%, respectively, p = 0.75), and no cumulative effect of carrying more than one MC1R variant was found. The present study does not support a role for the MC1R p.R160W and other variants in susceptibility for PD or RBD.
Assuntos
Estudos de Associação Genética , Predisposição Genética para Doença/genética , Variação Genética , Melanoma/genética , Doença de Parkinson/genética , Transtorno do Comportamento do Sono REM/genética , Receptor Tipo 1 de Melanocortina/genética , Adulto , Idoso , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Adulto JovemRESUMO
Amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) are overlapping, fatal neurodegenerative disorders in which the molecular and pathogenic basis remains poorly understood. Ubiquitinated protein aggregates, of which TDP-43 is a major component, are a characteristic pathological feature of most ALS and FTD patients. Here we use genome-wide linkage analysis in a large ALS/FTD kindred to identify a novel disease locus on chromosome 16p13.3. Whole-exome sequencing identified a CCNF missense mutation at this locus. Interrogation of international cohorts identified additional novel CCNF variants in familial and sporadic ALS and FTD. Enrichment of rare protein-altering CCNF variants was evident in a large sporadic ALS replication cohort. CCNF encodes cyclin F, a component of an E3 ubiquitin-protein ligase complex (SCF(Cyclin F)). Expression of mutant CCNF in neuronal cells caused abnormal ubiquitination and accumulation of ubiquitinated proteins, including TDP-43 and a SCF(Cyclin F) substrate. This implicates common mechanisms, linked to protein homeostasis, underlying neuronal degeneration.
Assuntos
Esclerose Lateral Amiotrófica/genética , Ciclinas/genética , Demência Frontotemporal/genética , Predisposição Genética para Doença/genética , Mutação de Sentido Incorreto , Adulto , Idoso , Sequência de Aminoácidos , Animais , Linhagem Celular Tumoral , Mapeamento Cromossômico , Cromossomos Humanos Par 16/genética , Saúde da Família , Feminino , Estudo de Associação Genômica Ampla , Humanos , Masculino , Pessoa de Meia-Idade , Linhagem , Análise de Sequência de DNA/métodos , Homologia de Sequência de AminoácidosRESUMO
BACKGROUND: The exact genetic causes within each of the known restless legs syndrome (RLS) loci are still unknown. Recently, it was suggested that an intronic protein tyrosine phosphatase, receptor type δ (PTPRD) single-nucleotide polymorphism (SNP) (reference SNP no. rs2381970) is associated with its expression, which may lead to RLS and other related phenotypes. Another study identified 3 nonsynonymous PTPRD variants in familial RLS cases: p.Q447E (a residue change from glutamine to glutamic acid at position 447), p.T781A (a residue change from threonine to alanine at position 781), and p.R995C (a residue change from arginine to cysteine at position 995). METHODS: Two cohorts of sporadic RLS, a French-Canadian cohort and a cohort from the United States, with a total of 577 patients and 455 controls, and an additional familial RLS cohort with a total of 635 individuals (140 families) were genotyped for these 4 variants (rs2381970, p.Q447E, p.T781A, and p.R995C) by using specific TaqMan probes, and the effects of each variant as well as haplotypes were analyzed. RESULTS: None of the 4 PTPRD-specific variants or haplotypes that were tested were associated with RLS in the case-control cohorts or in the familial cohort. The frequencies of the rs2381970 variant in the French-Canadian and US cohorts were 0.07 and 0.04, respectively, and their frequencies in the respective control populations were 0.06 and 0.04, respectively (P > 0.4 for both). Similar results were obtained for the 3 nonsynonymous variants. CONCLUSIONS: Although the PTPRD gene is well established as an RLS-associated locus, the rs2381970 SNP and the 3 nonsynonymous PTPRD variants are not likely to cause or affect the risk for developing RLS in the study population. More studies in other populations are needed to determine their potential role in RLS.
RESUMO
INTRODUCTION: Several studies proposed that Restless Legs Syndrome (RLS) and Parkinson disease (PD) may be clinically and/or etiologically related. To examine this hypothesis, we aimed to determine whether the known RLS genetic markers may be associated with PD risk, as well as with PD subtype. METHODS: Two case-control cohorts from Tel-Aviv and New-York, including 1133 PD patients and 867 controls were genotyped for four RLS-related SNPs in the genes MEIS1, BTBD9, PTPRD and MAP2K5/SKOR1. The association between genotype, PD risk and phenotype was tested using multivariate regression models. RESULTS: None of the tested SNPs was significantly associated with PD risk, neither in any individual cohort nor in the combined analysis after correction for multiple comparisons. The MAP2K5/SKOR1 marker rs12593813 was associated with higher frequency of tremor in the Tel-Aviv cohort (61.0% vs. 46.5%, p = 0.001, dominant model). However, the risk allele for tremor in this gene has been associated with reduced RLS risk. Moreover, this association did not replicate in Tremor-dominant PD patients from New-York. CONCLUSION: RLS genetic risk markers are not associated with increased PD risk or subtype in the current study. Together with previous genetic, neuropathological and epidemiologic studies, our results further strengthen the notion that RLS and PD are likely to be distinct entities.
Assuntos
Marcadores Genéticos/genética , Doença de Parkinson/genética , Síndrome das Pernas Inquietas/genética , Adulto , Idoso , Estudos de Casos e Controles , Estudos de Coortes , Feminino , Predisposição Genética para Doença , Proteínas de Homeodomínio/genética , Humanos , MAP Quinase Quinase 5/genética , Masculino , Pessoa de Meia-Idade , Proteína Meis1 , Proteínas de Neoplasias/genética , Proteínas do Tecido Nervoso , Polimorfismo de Nucleotídeo Único , Proteínas Tirosina Fosfatases Classe 2 Semelhantes a Receptores/genética , Fatores de Risco , Fatores de Transcrição/genéticaRESUMO
Amyotrophic lateral sclerosis (ALS) is a devastating neurological disorder characterized by the degeneration of motor neurons and typically results in death within 3-5 years from onset. Familial ALS (FALS) comprises 5%-10% of ALS cases, and the identification of genes associated with FALS is indispensable to elucidating the molecular pathogenesis. We identified a Japanese family affected by late-onset, autosomal-dominant ALS in which mutations in genes known to be associated with FALS were excluded. A whole- genome sequencing and parametric linkage analysis under the assumption of an autosomal-dominant mode of inheritance with incomplete penetrance revealed the mutation c.2780G>A (p. Arg927Gln) in ERBB4. An extensive mutational analysis revealed the same mutation in a Canadian individual with familial ALS and a de novo mutation, c.3823C>T (p. Arg1275Trp), in a Japanese simplex case. These amino acid substitutions involve amino acids highly conserved among species, are predicted as probably damaging, and are located within a tyrosine kinase domain (p. Arg927Gln) or a C-terminal domain (p. Arg1275Trp), both of which mediate essential functions of ErbB4 as a receptor tyrosine kinase. Functional analysis revealed that these mutations led to a reduced autophosphorylation of ErbB4 upon neuregulin-1 (NRG-1) stimulation. Clinical presentations of the individuals with mutations were characterized by the involvement of both upper and lower motor neurons, a lack of obvious cognitive dysfunction, and relatively slow progression. This study indicates that disruption of the neuregulin-ErbB4 pathway is involved in the pathogenesis of ALS and potentially paves the way for the development of innovative therapeutic strategies such using NRGs or their agonists to upregulate ErbB4 functions.
Assuntos
Esclerose Lateral Amiotrófica/genética , Receptores ErbB/genética , Mutação , Neurregulinas/genética , Idoso , Sequência de Aminoácidos , Substituição de Aminoácidos , Esclerose Lateral Amiotrófica/patologia , Povo Asiático/genética , Canadá , Análise Mutacional de DNA , Receptores ErbB/metabolismo , Feminino , Estudo de Associação Genômica Ampla , Humanos , Masculino , Pessoa de Meia-Idade , Dados de Sequência Molecular , Neurônios Motores/metabolismo , Neurônios Motores/patologia , Neurregulinas/metabolismo , Linhagem , Fosforilação , Receptor ErbB-4 , Análise de Sequência de DNA , Transdução de SinaisRESUMO
Loss-of-function of the potassium-chloride cotransporter 3 (KCC3) causes hereditary motor and sensory neuropathy with agenesis of the corpus callosum (HMSN/ACC), a severe neurodegenerative disease associated with defective midline crossing of commissural axons in the brain. Conversely, KCC3 over-expression in breast, ovarian and cervical cancer is associated with enhanced tumor cell malignancy and invasiveness. We identified a highly conserved proline-rich sequence within the C-terminus of the cotransporter which when mutated leads to loss of the KCC3-dependent regulatory volume decrease (RVD) response in Xenopus Laevis oocytes. Using SH3 domain arrays, we found that this poly-proline motif is a binding site for SH3-domain containing proteins in vitro. This approach identified the guanine nucleotide exchange factor (GEF) Vav2 as a candidate partner for KCC3. KCC3/Vav2 physical interaction was confirmed using GST-pull down assays and immuno-based experiments. In cultured cervical cancer cells, KCC3 co-localized with the active form of Vav2 in swelling-induced actin-rich protruding sites and within lamellipodia of spreading and migrating cells. These data provide evidence of a molecular and functional link between the potassium-chloride co-transporters and the Rho GTPase-dependent actin remodeling machinery in RVD, cell spreading and cell protrusion dynamics, thus providing new insights into KCC3's involvement in cancer cell malignancy and in corpus callosum agenesis in HMSN/ACC.
Assuntos
Tamanho Celular , Extensões da Superfície Celular/metabolismo , Oócitos/citologia , Proteínas Proto-Oncogênicas c-vav/metabolismo , Simportadores/metabolismo , Actinas/metabolismo , Motivos de Aminoácidos , Sequência de Aminoácidos , Animais , Tamanho Celular/efeitos dos fármacos , Extensões da Superfície Celular/efeitos dos fármacos , Sequência Conservada , Células HeLa , Humanos , Soluções Hipotônicas/farmacologia , Dados de Sequência Molecular , Proteínas Mutantes/metabolismo , Oócitos/efeitos dos fármacos , Oócitos/metabolismo , Ligação Proteica/efeitos dos fármacos , Estrutura Terciária de Proteína , Simportadores/química , Xenopus laevisRESUMO
Hereditary sensory and autonomic neuropathy type 2 (HSNAII) is a rare pathology characterized by an early onset of severe sensory loss (all modalities) in the distal limbs. It is due to autosomal recessive mutations confined to exon "HSN2" of the WNK1 (with-no-lysine protein kinase 1) serine-threonine kinase. While this kinase is well studied in the kidneys, little is known about its role in the nervous system. We hypothesized that the truncating mutations present in the neural-specific HSN2 exon lead to a loss-of-function of the WNK1 kinase, impairing development of the peripheral sensory system. To investigate the mechanisms by which the loss of WNK1/HSN2 isoform function causes HSANII, we used the embryonic zebrafish model and observed strong expression of WNK1/HSN2 in neuromasts of the peripheral lateral line (PLL) system by immunohistochemistry. Knocking down wnk1/hsn2 in embryos using antisense morpholino oligonucleotides led to improper PLL development. We then investigated the reported interaction between the WNK1 kinase and neuronal potassium chloride cotransporter KCC2, as this transporter is a target of WNK1 phosphorylation. In situ hybridization revealed kcc2 expression in mature neuromasts of the PLL and semi-quantitative RT-PCR of wnk1/hsn2 knockdown embryos showed an increased expression of kcc2 mRNA. Furthermore, overexpression of human KCC2 mRNA in embryos replicated the wnk1/hsn2 knockdown phenotype. We validated these results by obtaining double knockdown embryos, both for wnk1/hsn2 and kcc2, which alleviated the PLL defects. Interestingly, overexpression of inactive mutant KCC2-C568A, which does not extrude ions, allowed a phenocopy of the PLL defects. These results suggest a pathway in which WNK1/HSN2 interacts with KCC2, producing a novel regulation of its transcription independent of KCC2's activation, where a loss-of-function mutation in WNK1 induces an overexpression of KCC2 and hinders proper peripheral sensory nerve development, a hallmark of HSANII.
Assuntos
Neuropatias Hereditárias Sensoriais e Autônomas/genética , Peptídeos e Proteínas de Sinalização Intracelular/genética , Sistema Nervoso Periférico , Proteínas Serina-Treonina Quinases/genética , Simportadores , Peixe-Zebra , Animais , Modelos Animais de Doenças , Regulação da Expressão Gênica no Desenvolvimento , Neuropatias Hereditárias Sensoriais e Autônomas/metabolismo , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Antígenos de Histocompatibilidade Menor , Morfolinos , Mutação , Neurônios/metabolismo , Sistema Nervoso Periférico/crescimento & desenvolvimento , Sistema Nervoso Periférico/metabolismo , Fenótipo , Proteínas Serina-Treonina Quinases/metabolismo , Transdução de Sinais , Simportadores/genética , Simportadores/metabolismo , Ativação Transcricional , Proteína Quinase 1 Deficiente de Lisina WNK , Peixe-Zebra/genética , Peixe-Zebra/crescimento & desenvolvimento , Cotransportadores de K e Cl-RESUMO
Amyotrophic lateral sclerosis (ALS) is the most common motor neuron diseases (MND), while frontotemporal lobar degeneration (FTLD) is the second most common cause of early-onset dementia. Many ALS families segregating FTLD have been reported, particularly over the last decade. Recently, mutations in TARDBP, FUS/TLS, and C9ORF72 have been identified in both ALS and FTLD patients, while mutations in VCP, a FTLD associated gene, have been found in ALS families. Distinct variants located in the 3'-untranslated region (UTR) of the SIGMAR1 gene were previously reported in three unrelated FTLD or FTLD-MND families. We directly sequenced the coding and UTR regions of the SIGMAR1 gene in a targeted cohort of 25 individual familial ALS cases of Caucasian origin with a history of cognitive impairments. This screening identified one variant in the 3'-UTR of the SIGMAR1 gene in one ALS patient, but the same variant was also observed in 1 out of 380 control chromosomes. Subsequently, we screened the same samples for a C9ORF72 repeat expansion: 52% of this cohort was found expanded, including the sample with the SIGMAR1 3'-UTR variant. Consequently, coding and noncoding variants located in the 3'-UTR region of the SIGMAR1 gene are not the cause of FTLD-MND in our cohort, and more than half of this targeted cohort is genetically explained by C9ORF72 repeat expansions.
Assuntos
Esclerose Lateral Amiotrófica , Demência , Proteínas/genética , Receptores sigma/genética , Regiões 3' não Traduzidas/genética , Esclerose Lateral Amiotrófica/complicações , Esclerose Lateral Amiotrófica/genética , Esclerose Lateral Amiotrófica/fisiopatologia , Proteína C9orf72 , Expansão das Repetições de DNA/genética , Proteínas de Ligação a DNA/genética , Demência/complicações , Demência/genética , Demência/fisiopatologia , Degeneração Lobar Frontotemporal/genética , Degeneração Lobar Frontotemporal/fisiopatologia , Humanos , Neurônios Motores/metabolismo , Neurônios Motores/patologia , Proteína FUS de Ligação a RNA/genética , Receptor Sigma-1RESUMO
Essential tremor (ET) is a common neurodegenerative disorder that is characterized by a postural or motion tremor. Despite a strong genetic basis, a gene with rare pathogenic mutations that cause ET has not yet been reported. We used exome sequencing to implement a simple approach to control for misdiagnosis of ET, as well as phenocopies involving sporadic and senile ET cases. We studied a large ET-affected family and identified a FUS p.Gln290(∗) mutation as the cause of ET in this family. Further screening of 270 ET cases identified two additional rare missense FUS variants. Functional considerations suggest that the pathogenic effects of ET-specific FUS mutations are different from the effects observed when FUS is mutated in amyotrophic lateral sclerosis cases; we have shown that the ET FUS nonsense mutation is degraded by the nonsense-mediated-decay pathway, whereas amyotrophic lateral sclerosis FUS mutant transcripts are not.
Assuntos
Tremor Essencial/genética , Exoma/genética , Predisposição Genética para Doença/genética , Proteína FUS de Ligação a RNA/genética , Sequência de Bases , Humanos , Dados de Sequência Molecular , Mutação Puntual/genética , Quebeque , Análise de Sequência de DNARESUMO
Spinocerebellar ataxia type 3 is caused by the expansion of the coding CAG repeat in the ATXN3 gene. Interestingly, a -1 bp frameshift occurring within an (exp)CAG repeat would henceforth lead to translation from a GCA frame, generating polyalanine stretches instead of polyglutamine. Our results show that transgenic expression of (exp)CAG ATXN3 led to -1 frameshifting events, which have deleterious effects in Drosophila and mammalian neurons. Conversely, transgenic expression of polyglutamine-encoding (exp)CAA ATXN3 was not toxic. Furthermore, (exp)CAG ATXN3 mRNA does not contribute per se to the toxicity observed in our models. Our observations indicate that expanded polyglutamine tracts in Drosophila and mouse neurons are insufficient for the development of a phenotype. Hence, we propose that -1 ribosomal frameshifting contributes to the toxicity associated with (exp)CAG repeats.