RESUMEN
Mutations in profilin 1 (PFN1) have been identified in rare familial cases of Amyotrophic Lateral Sclerosis (ALS). PFN1 is involved in multiple pathways that could intervene in ALS pathology. However, the specific pathogenic role of PFN1 mutations in ALS is still not fully understood. We hypothesized that PFN1 could play a role in regulating autophagy pathways and that PFN1 mutations could disrupt this function. We used patient cells (lymphoblasts) or tissue (post-mortem) carrying PFN1 mutations (M114T and E117G), and designed experimental models expressing wild-type or mutant PFN1 (cell lines and novel PFN1 mice established by lentiviral transgenesis) to study the effects of PFN1 mutations on autophagic pathway markers. We observed no accumulation of PFN1 in the spinal cord of one E117G mutation carrier. Moreover, in patient lymphoblasts and transfected cell lines, the M114T mutant PFN1 protein was unstable and deregulated the RAB9-mediated alternative autophagy pathway involved in the clearance of damaged mitochondria. In vivo, motor neurons expressing M114T mutant PFN1 showed mitochondrial abnormalities. Our results demonstrate that the M114T PFN1 mutation is more deleterious than the E117G variant in patient cells and experimental models and suggest a role for the RAB9-dependent autophagic pathway in ALS.
Asunto(s)
Esclerosis Amiotrófica Lateral , Profilinas , Proteínas de Unión al GTP rab , Esclerosis Amiotrófica Lateral/metabolismo , Animales , Autofagia/genética , Homeostasis , Humanos , Ratones , Mitocondrias/metabolismo , Mutación , Profilinas/genética , Profilinas/metabolismo , Proteínas de Unión al GTP rab/metabolismoRESUMEN
The nuclear envelope (NE) separates the nucleus from the cytoplasm in all eukaryotic cells. A disruption of the NE structure compromises normal gene regulation and leads to severe human disorders collectively classified as nuclear envelopathies and affecting skeletal muscle, heart, brain, skin, and bones. The ubiquitous NE component LAP1B is encoded by TOR1AIP1, and the use of an alternative start codon gives rise to the shorter LAP1C isoform. TOR1AIP1 mutations have been identified in patients with diverging clinical presentations such as muscular dystrophy, progressive dystonia with cerebellar atrophy, and a severe multi-systemic disorder, but the correlation between the mutational effect and the clinical spectrum remains to be determined. Here, we describe a novel TOR1AIP1 patient manifesting childhood-onset muscle weakness and contractures, and we provide clinical, histological, ultrastructural, and genetic data. We demonstrate that the identified TOR1AIP1 frameshift mutation leads to the selective loss of the LAP1B isoform, while the expression of LAP1C was preserved. Through comparative review of all previously reported TOR1AIP1 cases, we delineate a genotype/phenotype correlation and conclude that LAP1B-specific mutations cause a progressive skeletal muscle phenotype, while mutations involving a loss of both LAP1B and LAP1C isoforms induce a syndromic disorder affecting skeletal muscle, brain, eyes, ear, skin, and bones.
Asunto(s)
Fosfatidilinositol 3-Quinasa Clase I/genética , Mutación/genética , Membrana Nuclear/genética , Isoformas de Proteínas/genética , Niño , Femenino , Mutación del Sistema de Lectura/genética , Humanos , Masculino , Músculos/metabolismo , Músculos/patología , Membrana Nuclear/metabolismo , Membrana Nuclear/ultraestructura , Proteínas Nucleares/genética , FenotipoRESUMEN
BACKGROUND: Congenital myopathies are severe genetic diseases with a strong impact on patient autonomy and often on survival. A large number of patients do not have a genetic diagnosis, precluding genetic counseling and appropriate clinical management. Our objective was to find novel pathogenic variants and genes associated with congenital myopathies and to decrease diagnostic odysseys and dead-end. METHODS: To identify pathogenic variants and genes implicated in congenital myopathies, we established and conducted the MYOCAPTURE project from 2009 to 2018 to perform exome sequencing in a large cohort of 310 families partially excluded for the main known genes. RESULTS: Pathogenic variants were identified in 156 families (50%), among which 123 families (40%) had a conclusive diagnosis. Only 44 (36%) of the resolved cases were linked to a known myopathy gene with the corresponding phenotype, while 55 (44%) were linked to pathogenic variants in a known myopathy gene with atypical signs, highlighting that most genetic diagnosis could not be anticipated based on clinical-histological assessments in this cohort. An important phenotypic and genetic heterogeneity was observed for the different genes and for the different congenital myopathy subtypes, respectively. In addition, we identified 14 new myopathy genes not previously associated with muscle diseases (20% of all diagnosed cases) that we previously reported in the literature, revealing novel pathomechanisms and potential therapeutic targets. CONCLUSIONS: Overall, this approach illustrates the importance of massive parallel gene sequencing as a comprehensive tool for establishing a molecular diagnosis for families with congenital myopathies. It also emphasizes the contribution of clinical data, histological findings on muscle biopsies, and the availability of DNA samples from additional family members to the diagnostic success rate. This study facilitated and accelerated the genetic diagnosis of congenital myopathies, improved health care for several patients, and opened novel perspectives for either repurposing of existing molecules or the development of novel treatments.
Asunto(s)
Secuenciación del Exoma , Estudios de Asociación Genética , Fenotipo , Humanos , Masculino , Femenino , Predisposición Genética a la Enfermedad , Mutación , Exoma/genética , Linaje , Miopatías Estructurales Congénitas/genética , Miopatías Estructurales Congénitas/diagnóstico , Enfermedades Musculares/genética , Enfermedades Musculares/diagnóstico , Enfermedades Musculares/congénito , Niño , AdultoRESUMEN
Variants in MYH7 cause cardiomyopathies as well as myosin storage myopathy and Laing early-onset distal myopathy (MPD1). MPD1 is characterized by muscle weakness and atrophy usually beginning in the lower legs. Here, we generated iPSC lines from lymphoblastoid cells of three unrelated individuals heterozygous for the most common MPD1-causing variant; p.Lys1617del. iPSC lines showed typical morphology, expressed pluripotency markers, demonstrated trilineage differentiation potential, and had a normal karyotype. These lines represent the first iPSCs derived from MPD1 patients and complement existing MPD1 animal models. They can provide in vitro platforms to better understand and model MPD1 pathomechanisms and test therapies.
Asunto(s)
Miosinas Cardíacas , Miopatías Distales , Células Madre Pluripotentes Inducidas , Cadenas Pesadas de Miosina , Humanos , Células Madre Pluripotentes Inducidas/metabolismo , Células Madre Pluripotentes Inducidas/patología , Cadenas Pesadas de Miosina/genética , Cadenas Pesadas de Miosina/metabolismo , Miopatías Distales/genética , Miopatías Distales/patología , Miopatías Distales/metabolismo , Miosinas Cardíacas/genética , Miosinas Cardíacas/metabolismo , Masculino , Femenino , Línea Celular , Diferenciación Celular , AdultoRESUMEN
RYR1 variants are a common cause of congenital myopathies, including multi-minicore disease (MmD) and central core disease (CCD). Here, we generated iPSC lines from two CCD patients with dominant RYR1 missense variants that affect the transmembrane (pore) and SPRY3 protein domains (p.His4813Tyr and p.Asn1346Lys, respectively). Both lines had typical iPSC morphology, expressed canonical pluripotency markers, exhibited trilineage differentiation potential, and had normal karyotypes. Together with existing RYR1 iPSC lines, these represent important tools to study and develop treatments for RYR1-related myopathies.
Asunto(s)
Células Madre Pluripotentes Inducidas , Mutación Missense , Canal Liberador de Calcio Receptor de Rianodina , Humanos , Canal Liberador de Calcio Receptor de Rianodina/genética , Canal Liberador de Calcio Receptor de Rianodina/metabolismo , Células Madre Pluripotentes Inducidas/metabolismo , Miopatía del Núcleo Central/genética , Miopatía del Núcleo Central/patología , Miopatía del Núcleo Central/metabolismo , Adulto , Línea Celular , Masculino , Diferenciación Celular , FemeninoRESUMEN
RYR1 variants are the most common genetic cause of congenital myopathies, and typically cause central core disease (CCD) and/or malignant hyperthermia (MH). Here, we generated iPSC lines from two patients with CCD and MH caused by dominant RYR1 variants within the central region of the protein (p.Val2168Met and p.Arg2508Cys). Both lines displayed typical iPSC morphology, uniform expression of pluripotency markers, trilineage differentiation potential, and had normal karyotypes. These are the first RYR1 iPSC lines from patients with both CCD and MH. As these are common CCD/MH variants, these lines should be useful to study these conditions and test therapeutics.
Asunto(s)
Células Madre Pluripotentes Inducidas , Hipertermia Maligna , Mutación Missense , Canal Liberador de Calcio Receptor de Rianodina , Humanos , Canal Liberador de Calcio Receptor de Rianodina/genética , Canal Liberador de Calcio Receptor de Rianodina/metabolismo , Hipertermia Maligna/genética , Células Madre Pluripotentes Inducidas/metabolismo , Miopatía del Núcleo Central/genética , Miopatía del Núcleo Central/patología , Masculino , Femenino , Línea Celular , Diferenciación CelularRESUMEN
Amyotrophic lateral sclerosis (ALS) is a devastating motor neuron disease. The immunosuppressive functions of regulatory T lymphocytes (Tregs) are impaired in ALS, and correlate to disease progression. The phase 2a IMODALS trial reported an increase in Treg number in ALS patients following the administration of low-dose (ld) interleukin-2 (IL-2). We propose a pharmacometabolomics approach to decipher metabolic modifications occurring in patients treated with ld-IL-2 and its relationship with Treg response. Blood metabolomic profiles were determined on days D1, D64, and D85 from patients receiving 2 MIU of IL-2 (n = 12) and patients receiving a placebo (n = 12). We discriminated the three time points for the treatment group (average error rate of 42%). Among the important metabolites, kynurenine increased between D1 and D64, followed by a reduction at D85. The percentage increase of Treg number from D1 to D64, as predicted by the metabolome at D1, was highly correlated with the observed value. This study provided a proof of concept for metabolic characterization of the effect of ld-IL-2 in ALS. These data could present advances toward a personalized medicine approach and present pharmacometabolomics as a key tool to complement genomic and transcriptional data for drug characterization, leading to systems pharmacology.
Asunto(s)
Esclerosis Amiotrófica Lateral , Interleucina-2 , Metabolómica , Linfocitos T Reguladores , Esclerosis Amiotrófica Lateral/tratamiento farmacológico , Esclerosis Amiotrófica Lateral/metabolismo , Humanos , Interleucina-2/administración & dosificación , Interleucina-2/metabolismo , Metabolómica/métodos , Linfocitos T Reguladores/metabolismo , Linfocitos T Reguladores/efectos de los fármacos , Linfocitos T Reguladores/inmunología , Masculino , Persona de Mediana Edad , Femenino , Quinurenina/metabolismo , Anciano , Metaboloma/efectos de los fármacosRESUMEN
Central core disease (CCD) is a congenital disorder that results in hypotonia, delayed motor development, and areas of reduced oxidative activity in the muscle fibre. Two induced pluripotent stem cell (iPSC) lines were generated from the lymphoblastoid cells of a 33-year-old male with CCD, caused by a previously unreported dominant c.14145_14156delCTACTGGGACA (p.Asn4715_Asp4718del) deletion in the RYR1 gene. Both lines demonstrated typical morphology, pluripotency, trilineage differentiation, and had a normal karyotype. As the first published iPSC model of CCD caused by an RYR1 variant these lines are a potential resource for further investigation of RYR1-related myopathies in a human context.
Asunto(s)
Células Madre Pluripotentes Inducidas , Miopatía del Núcleo Central , Masculino , Humanos , Adulto , Miopatía del Núcleo Central/genética , Miopatía del Núcleo Central/metabolismo , Células Madre Pluripotentes Inducidas/metabolismo , Canal Liberador de Calcio Receptor de Rianodina/genética , Canal Liberador de Calcio Receptor de Rianodina/metabolismo , MutaciónRESUMEN
Variants in the ACTA1 gene are a common cause of nemaline myopathy (NM); a muscle disease that typically presents at birth or early childhood with hypotonia and muscle weakness. Here, we generated an induced pluripotent stem cell line (iPSC) from lymphoblastoid cells of a 3-month-old female patient with intermediate NM caused by a dominant ACTA1 variant (c.515C > A (p.Ala172Glu)). iPSCs showed typical morphology, expressed pluripotency markers, demonstrated trilineage differentiation potential, and had a normal karyotype. This line complements our previously published ACTA1 iPSC lines derived from patients with typical and severe NM.
Asunto(s)
Células Madre Pluripotentes Inducidas , Miopatías Nemalínicas , Actinas/genética , Actinas/metabolismo , Preescolar , Femenino , Humanos , Células Madre Pluripotentes Inducidas/metabolismo , Lactante , Recién Nacido , Músculo Esquelético/metabolismo , Mutación , Miopatías Nemalínicas/genéticaRESUMEN
Nemaline myopathy (NM) is a congenital skeletal muscle disorder that typically results in muscle weakness and the presence of rod-like structures (nemaline bodies) in the sarcoplasma and/or in the nuclei of myofibres. Two induced pluripotent stem cell (iPSC) lines were generated from the lymphoblastoid cells of a 1-month-old male with severe NM caused by a homozygous recessive mutation in the ACTA1 gene (c.121C > T, p.Arg39Ter). The iPSC lines demonstrated typical morphology, expressed pluripotency markers, exhibited trilineage differentiation potential and displayed a normal karyotype. These isogenic lines represent a potential resource to investigate and model recessive ACTA1 disease in a human context.
Asunto(s)
Células Madre Pluripotentes Inducidas , Miopatías Nemalínicas , Actinas/genética , Actinas/metabolismo , Homocigoto , Humanos , Células Madre Pluripotentes Inducidas/metabolismo , Lactante , Masculino , Músculo Esquelético/metabolismo , Mutación , Miopatías Nemalínicas/genética , Miopatías Nemalínicas/metabolismoRESUMEN
Nemaline myopathy (NM) is a congenital myopathy typically characterized by skeletal muscle weakness and the presence of nemaline bodies in myofibres. Approximately 25% of NM cases are caused by variants in ACTA1. We generated two induced pluripotent stem cell lines from lymphoblastoid cells of a 10-year-old female with typical NM harbouring a dominant pathogenic variant in ACTA1 (c.541C>A). The isogenic lines displayed typical iPSC morphology, expressed pluripotency markers, and could differentiate into each of the three germ layers. Although the lines have partial or complete X chromosome duplication, they may still prove useful as models of human ACTA1 disease.
Asunto(s)
Células Madre Pluripotentes Inducidas , Miopatías Nemalínicas , Actinas/genética , Niño , Femenino , Humanos , Músculo Esquelético , Mutación , Miopatías Nemalínicas/genéticaRESUMEN
Nemaline myopathy (NM) is a congenital myopathy typically characterized by skeletal muscle weakness and the presence of abnormal thread- or rod-like structures (nemaline bodies) in myofibres. Pathogenic variants in the skeletal muscle alpha actin gene, ACTA1, cause approximately 25% of all NM cases. We generated two induced pluripotent stem cell lines from lymphoblastoid cells of a 4-month-old female with severe NM harbouring a dominant variant in ACTA1 (c.553C > A). The isogenic lines displayed characteristic iPSC morphology, expressed pluripotency markers, differentiated into cells of all three germ layers, and possessed normal karyotypes. These lines could be useful models of human ACTA1 disease.
Asunto(s)
Células Madre Pluripotentes Inducidas , Miopatías Nemalínicas , Actinas/genética , Femenino , Humanos , Lactante , Músculo Esquelético , Mutación , Miopatías Nemalínicas/genéticaRESUMEN
ANXA11 mutations have previously been discovered in amyotrophic lateral sclerosis (ALS) motor neuron disease. To confirm the contribution of ANXA11 mutations to ALS, a large exome data set obtained from 330 French patients, including 150 familial ALS index cases and 180 sporadic ALS cases, was analyzed, leading to the identification of 3 rare ANXA11 variants in 5 patients. The novel p.L254V variant was associated with early onset sporadic ALS. The novel p.D40Y mutation and the p.G38R variant concerned patients with predominant pyramidal tract involvement and cognitive decline. Neuropathologic findings in a p.G38R carrier associated the presence of ALS typical inclusions within the spinal cord, massive degeneration of the lateral tracts, and type A frontotemporal lobar degeneration. This mutant form of annexin A11 accumulated in various brain regions and in spinal cord motor neurons, although its stability was decreased in patients' lymphoblasts. Because most ANXA11 inclusions were not colocalized with transactive response DNA-binding protein 43 or p62 deposits, ANXA11 aggregation does not seem mandatory to trigger neurodegeneration with additional participants/partner proteins that could intervene.
Asunto(s)
Esclerosis Amiotrófica Lateral/genética , Anexinas/genética , Estudios de Asociación Genética , Mutación , Bases de Datos Genéticas , Conjuntos de Datos como Asunto , Exoma/genética , Femenino , Francia , Degeneración Lobar Frontotemporal/genética , Humanos , MasculinoRESUMEN
Mutations in UBQLN2 have been associated with rare cases of X-linked juvenile and adult forms of amyotrophic lateral sclerosis (ALS) and ALS linked to frontotemporal dementia (FTD). Here, we report 1 known (c.1489C>T, p.Pro497Ser, P497S) and 3 novel (c.1481C>T, p.Pro494Leu, P494L; c.1498C>T, p.Pro500Ser, P500S; and c.1516C>G, p.Pro506Ala, P506A) missense mutations in the PXX domain of UBQLN2 in familial motor neuron diseases including ALS and spastic paraplegia (SP). A novel missense mutation (c.1462G>A, p.Ala488Thr, A488T) adjacent to this hotspot UBQLN2 domain was identified in a sporadic case of ALS. These mutations are conserved in mammals, are absent from ExAC and gnomAD browsers, and are predicted to be deleterious by SIFT in silico analysis. Patient lymphoblasts carrying a UBQLN2 mutation showed absence of ubiquilin-2 accumulation, disrupted binding with HSP70, and impaired autophagic pathway. Our results confirm the role of PXX repeat in ALS pathogenesis, show that UBQLN2-linked disease can manifest like a SP phenotype, evidence a highly reduced disease penetrance in females carrying UBQLN2 mutations, which is important information for genetic counseling, and underline the pivotal role of ubiquilin-2 in proteolysis regulation pathways.
Asunto(s)
Esclerosis Amiotrófica Lateral/genética , Proteínas de Ciclo Celular/genética , Demencia Frontotemporal/genética , Estudios de Asociación Genética , Proteínas HSP70 de Choque Térmico/genética , Proteínas HSP70 de Choque Térmico/metabolismo , Mutación Missense/genética , Fenotipo , Proteolisis , Paraplejía Espástica Hereditaria/genética , Ubiquitinas/genética , Proteínas Adaptadoras Transductoras de Señales , Anciano , Anciano de 80 o más Años , Proteínas Relacionadas con la Autofagia , Proteínas de Ciclo Celular/química , Proteínas de Ciclo Celular/metabolismo , Dimerización , Femenino , Humanos , Masculino , Persona de Mediana Edad , Dominios Proteicos/genética , Ubiquitinas/química , Ubiquitinas/metabolismo , Inactivación del Cromosoma XRESUMEN
The EuroBioBank (EBB) network (www.eurobiobank.org) is the first operating network of biobanks in Europe to provide human DNA, cell and tissue samples as a service to the scientific community conducting research on rare diseases (RDs). The EBB was established in 2001 to facilitate access to RD biospecimens and associated data; it obtained funding from the European Commission in 2002 (5th framework programme) and started operation in 2003. The set-up phase, during the EC funding period 2003-2006, established the basis for running the network; the following consolidation phase has seen the growth of the network through the joining of new partners, better network cohesion, improved coordination of activities, and the development of a quality-control system. During this phase the network participated in the EC-funded TREAT-NMD programme and was involved in planning of the European Biobanking and Biomolecular Resources Research Infrastructure. Recently, EBB became a partner of RD-Connect, an FP7 EU programme aimed at linking RD biobanks, registries, and bioinformatics data. Within RD-Connect, EBB contributes expertise, promotes high professional standards, and best practices in RD biobanking, is implementing integration with RD patient registries and 'omics' data, thus challenging the fragmentation of international cooperation on the field.