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Pseudoexon activation in the DMD gene as a novel mechanism for Becker muscular dystrophy
Sylvie Tuffery-Giraud1, Céline Saquet, Sylvie Chambert
1Laboratoire de Génétique Moleculaire, CNRS UPR 1042, Institut Universitaire de Recherche Clinique and CHU, Montpellier, France. tuffery@igh.cnrs.fr
Abstract:
We report the characterization of two deep intronic mutations in the Duchenne muscular dystrophy (DMD) gene of two unrelated Becker muscular dystrophy (BMD) patients, causing the aberrant inclusion of a pseudoexon in the mature transcripts. These two mutations were identified by the use of RT-PCR on transcripts isolated from muscle. The first abnormally large transcript resulting from a 58-bp insertion between exon 62 and exon 63 was identified in a BMD patient with mental retardation. The origin of this transcript was a mutation in intron 62 (IVS62-285A>G), which resulted in the occurrence of a high quality donor splice site. The IVS25+2036A>G in intron 25 was identified in a subclinical BMD patient with high CK levels. The mutation reinforces the strength of a pre-existing acceptor splice site, resulting in activation of an intronic pseudoexon of 95 bp. By using DHPLC, the patient's mother was found to be a somatic mosaic. The insertion of these newly recognized extra exons leads to premature termination codons, but we could observe that some degree of normal splicing was taking place in both patients. The detection of these residual full length transcripts is consistent with the clinical presentation and dystrophin analyses. This is the first report of pseudoexon activation as a mechanism for Becker muscular dystrophy, and this reveals further the diversity of genetic abnormalities causing BMD.
Insights
Deep intronic mutations in the Duchenne muscular dystrophy (DMD) gene activate pseudoexons, leading to Becker muscular dystrophy (BMD). This study identifies novel mutations causing aberrant splicing and reveals pseudoexon activation as a BMD mechanism.
Area of Science:
- Genetics
- Molecular Biology
- Neuromuscular Disorders
Background:
- Duchenne muscular dystrophy (DMD) and Becker muscular dystrophy (BMD) are allelic disorders caused by mutations in the DMD gene.
- Genetic mutations can lead to aberrant RNA splicing, altering protein function.
Purpose of the Study:
- To characterize deep intronic mutations in the DMD gene in two unrelated Becker muscular dystrophy patients.
- To investigate the mechanism of pseudoexon activation and its role in BMD pathogenesis.
Main Methods:
- Reverse transcription polymerase chain reaction (RT-PCR) was used to analyze muscle transcripts.
- Denaturing High Performance Liquid Chromatography (DHPLC) was employed to detect mutations and mosaicism.
Main Results:
- Two deep intronic mutations (IVS62-285A>G and IVS25+2036A>G) were identified, causing the inclusion of pseudoexons in DMD transcripts.
- One mutation (IVS62-285A>G) led to a 58-bp insertion in a BMD patient with mental retardation.
- The other mutation (IVS25+2036A>G) resulted in a 95-bp pseudoexon activation in a subclinical BMD patient, with the mother identified as a somatic mosaic.
Conclusions:
- Pseudoexon activation is a newly identified mechanism contributing to Becker muscular dystrophy.
- These findings expand the understanding of genetic abnormalities underlying BMD and highlight the diversity of DMD gene mutations.