RESUMO
OBJECTIVE: Dysferlin (DYSF) gene mutations cause limb girdle muscular dystrophy type 2B and Miyoshi's myopathy. The consequences of DYSF mutations on protein structure are poorly understood. METHODS: The gene encoding dysferlin was sequenced in patients with suspected dysferlin-deficient muscular dystrophy. Muscle biopsy specimens were analyzed by histochemistry, immunohistochemistry, and electron microscopy. Antibodies against N-terminal dysferlin-peptides were raised. RESULTS: We found three families with muscular dystrophy caused by homozygous or compound heterozygous DYSF mutations featuring sarcolemmal and interstitial amyloid deposits. These mutations were all located in the N-terminal region of the protein. Dysferlin was a constituent of the amyloid deposits. INTERPRETATION: Limb girdle muscular dystrophy type 2B is the first muscular dystrophy associated with amyloidosis. Molecular treatment strategies will necessarily have to consider the presence of amyloidogenesis.
Assuntos
Amiloidose/genética , Amiloidose/metabolismo , Proteínas de Membrana/deficiência , Proteínas de Membrana/genética , Proteínas Musculares/deficiência , Proteínas Musculares/genética , Distrofia Muscular do Cíngulo dos Membros/genética , Distrofia Muscular do Cíngulo dos Membros/metabolismo , Adulto , Idoso , Sequência de Aminoácidos , Substituição de Aminoácidos/genética , Amiloidose/diagnóstico , Disferlina , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Dados de Sequência Molecular , Distrofia Muscular do Cíngulo dos Membros/diagnóstico , Estrutura Terciária de Proteína/genéticaRESUMO
Mutations in the gene encoding dysferlin (DYSF) cause the allelic autosomal recessive disorders limb girdle muscular dystrophy 2B and Miyoshi myopathy. It encompasses 55 exons spanning 150 kb of genomic DNA. Dysferlin is involved in membrane repair in skeletal muscle. We identified three families with novel sequence variants in DYSF. All affected family members showed limb girdle weakness and had reduced or absent dysferlin protein on immunohistochemistry. All exons of DYSF were screened by genomic sequencing. Five novel variants in DYSF were found: two missense mutations (c.895G>A and c.4022T>C), one 5' donor splice-site variant (c.855+1delG), one nonsense mutation (c.1448C>A), and a variant in the 3'UTR of DYSF (c.*107T>A). All alterations were confirmed by restriction enzyme analysis and not found in 400 control alleles. Nonsense mediated RNA decay or changes in the three-dimensional protein structure resulting in intracellular dysferlin aggregates and finally the lack of dysferlin protein were identified as consequences of the novel DYSF variants.
Assuntos
Proteínas de Membrana/química , Proteínas de Membrana/genética , Proteínas Musculares/química , Proteínas Musculares/genética , Distrofias Musculares/genética , Mutação , Adulto , Sequência de Aminoácidos , Análise Mutacional de DNA , Disferlina , Feminino , Heterozigoto , Homozigoto , Humanos , Masculino , Proteínas de Membrana/deficiência , Modelos Moleculares , Dados de Sequência Molecular , Proteínas Musculares/deficiência , Músculo Esquelético/patologia , Distrofias Musculares/diagnóstico , Linhagem , Dobramento de Proteína , Estrutura Terciária de Proteína , Sítios de Splice de RNA , Estabilidade de RNA , RNA Mensageiro/metabolismo , Alinhamento de SequênciaRESUMO
Dysferlin is expressed in skeletal and cardiac muscles. However, dysferlin deficiency results in skeletal muscle weakness, but spares the heart. We compared intraindividual mRNA expression profiles of cardiac and skeletal muscle in dysferlin-deficient SJL/J mice and found down-regulation of the complement inhibitor, decay-accelerating factor/CD55, in skeletal muscle only. This finding was confirmed on mRNA and protein levels in two additional dysferlin-deficient mouse strains, A/J mice and Dysf-/- mice, as well as in patients with dysferlin-deficient muscular dystrophy. In vitro, the absence of CD55 led to an increased susceptibility of human myotubes to complement attack. Evidence is provided that decay-accelerating factor/CD55 is regulated via the myostatin-SMAD pathway. In conclusion, a novel mechanism of muscle fiber injury in dysferlin-deficient muscular dystrophy is demonstrated, possibly opening therapeutic avenues in this to date untreatable disorder.