title: Muscular dystrophy with marked Dysferlin deficiency is consistently caused by primary dysferlin gene mutations.
abstract: Dysferlin is a 237-kDa transmembrane protein involved in calcium-mediated sarcolemma resealing. Dysferlin gene mutations cause limb-girdle muscular dystrophy (LGMD) 2B, Miyoshi myopathy (MM) and distal myopathy of the anterior tibialis. Considering that a secondary Dysferlin reduction has also been described in other myopathies, our original goal was to identify cases with a Dysferlin deficiency without dysferlin gene mutations. The dysferlin gene is huge, composed of 55 exons that span 233 140 bp of genomic DNA. We performed a thorough mutation analysis in 65 LGMD/MM patients with ≤20% Dysferlin. The screening was exhaustive, as we sequenced both genomic DNA and cDNA. When required, we used other methods, including real-time PCR, long PCR and array CGH. In all patients, we were able to recognize the primary involvement of the dysferlin gene. We identified 38 novel mutation types. Some of these, such as a dysferlin gene duplication, could have been missed by conventional screening strategies. Nonsense-mediated mRNA decay was evident in six cases, in three of which both alleles were only detectable in the genomic DNA but not in the mRNA. Among a wide spectrum of novel gene defects, we found the first example of a 'nonstop' mutation causing a dysferlinopathy. This study presents the first direct and conclusive evidence that an amount of Dysferlin ≤20% is pathogenic and always caused by primary dysferlin gene mutations. This demonstrates the high specificity of a marked reduction of Dysferlin on western blot and the value of a comprehensive molecular approach for LGMD2B/MM diagnosis.
authors.affiliation: Dipartimento di Patologia Generale, Telethon Institute of Genetics and Medicine (TIGEM), Seconda Università degli Studi di Napoli,S. Andrea delle Dame, via L. De Crecchio 7, Naples, Italy.
authors.forename: Mafalda
authors.initials: M
authors.lastname: Cacciottolo
authors.forename: Gelsomina
authors.initials: G
authors.lastname: Numitone
authors.forename: Stefania
authors.initials: S
authors.lastname: Aurino
authors.forename: Imma Rosaria
authors.initials: IR
authors.lastname: Caserta
authors.forename: Marina
authors.initials: M
authors.lastname: Fanin
authors.forename: Luisa
authors.initials: L
authors.lastname: Politano
authors.forename: Carlo
authors.initials: C
authors.lastname: Minetti
authors.forename: Enzo
authors.initials: E
authors.lastname: Ricci
authors.forename: Giulio
authors.initials: G
authors.lastname: Piluso
authors.forename: Corrado
authors.initials: C
authors.lastname: Angelini
authors.forename: Vincenzo
authors.initials: V
authors.lastname: Nigro
mesh_terms: D000328:Adult; D000368:Aged; D055028:Comparative Genomic Hybridization; D004252:DNA Mutational Analysis; D049310:Distal Myopathies / Q000235:genetics*; D000073939:Dysferlin; D006801:Humans; D008565:Membrane Proteins / Q000172:deficiency / Q000235:genetics*; D008875:Middle Aged; D009124:Muscle Proteins / Q000172:deficiency / Q000235:genetics*; D009133:Muscular Atrophy / Q000235:genetics*; D049288:Muscular Dystrophies, Limb-Girdle / Q000235:genetics*; D059365:Nonsense Mediated mRNA Decay
chemical_list: C114839:DYSF protein, human; D000073939:Dysferlin; D008565:Membrane Proteins; D009124:Muscle Proteins