title: Novel molecular diagnostic approaches for X-linked centronuclear (myotubular) myopathy reveal intronic mutations.
abstract: X-linked centronuclear myopathy (XLMTM), also called myotubular myopathy, is a severe congenital myopathy characterized by generalized hypotonia and weakness at birth and the typical histological finding of centralization of myo-nuclei. It is caused by mutations in the MTM1 gene encoding the 3-phosphoinositides phosphatase myotubularin. Mutations in dynamin 2 and amphiphysin 2 genes lead to autosomal forms of centronuclear myopathy (CNM). While XLMTM is the most frequent and severe form of CNM, no mutations are found in about 30% of patients by sequencing all MTM1 exons. Moreover, the impact of MTM1 sequence variants is sometimes difficult to assess. It is thus important to devise a complete molecular diagnostic strategy that includes analysis of the myotubularin transcript and protein expression. We therefore developed novel antibodies against human myotubularin and showed that they are able to detect the endogenous protein by direct Western blot from muscle samples and from cultured cells. In conjunction with RT-PCR analysis we validated the consequences of missense and splice mutations on transcript integrity and protein level. We also detected and characterized a novel deep intronic mutation consisting of a single nucleotide change that induces exonisation of a conserved intronic sequence. Patients with centronuclear myopathy and no molecular diagnosis should be investigated for MTM1 defects at the cDNA and protein level.
authors.affiliation: Department of Neurobiology and Genetics, Institut de Génétique et de Biologie Moléculaire et Cellulaire, Illkirch, France.
authors.forename: Valérie
authors.initials: V
authors.lastname: Tosch
authors.forename: Nasim
authors.initials: N
authors.lastname: Vasli
authors.forename: Christine
authors.initials: C
authors.lastname: Kretz
authors.forename: Anne-Sophie
authors.initials: AS
authors.lastname: Nicot
authors.forename: Claire
authors.initials: C
authors.lastname: Gasnier
authors.forename: Nicolas
authors.initials: N
authors.lastname: Dondaine
authors.forename: Denis
authors.initials: D
authors.lastname: Oriot
authors.forename: Magalie
authors.initials: M
authors.lastname: Barth
authors.forename: Hugues
authors.initials: H
authors.lastname: Puissant
authors.forename: Norma B
authors.initials: NB
authors.lastname: Romero
authors.forename: Carsten G
authors.initials: CG
authors.lastname: Bönnemann
authors.forename: Betty
authors.initials: B
authors.lastname: Heller
authors.forename: Gilles
authors.initials: G
authors.lastname: Duval
authors.forename: Valérie
authors.initials: V
authors.lastname: Biancalana
authors.forename: Jocelyn
authors.initials: J
authors.lastname: Laporte
mesh_terms: D001483:Base Sequence; D015153:Blotting, Western; D002478:Cells, Cultured; D004247:DNA / Q000235:genetics; D005091:Exons / Q000235:genetics; D050172:Genes, X-Linked / Q000235:genetics*; D006801:Humans; D007438:Introns / Q000235:genetics*; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics / Q000502:physiology; D020914:Myopathies, Structural, Congenital / Q000175:diagnosis* / Q000235:genetics*; D054558:Protein Tyrosine Phosphatases, Non-Receptor / Q000235:genetics; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction
chemical_list: D012333:RNA, Messenger; D004247:DNA; D054558:Protein Tyrosine Phosphatases, Non-Receptor; C099588:myotubularin