Tuffery-Giraud Sylvie, Saquet Céline, Chambert Sylvie, Claustres Mireille
Laboratoire de Génétique Moleculaire, CNRS UPR 1042, Institut Universitaire de Recherche Clinique and CHU, Montpellier, France.
Hum Mutat. 2003 Jun;21(6):608-14. doi: 10.1002/humu.10214.
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.
我们报告了两名无关的贝克型肌营养不良症（BMD）患者的杜氏肌营养不良症（DMD）基因中两个内含子深处突变的特征，这些突变导致成熟转录本中异常包含一个假外显子。这两个突变是通过对从肌肉中分离的转录本进行逆转录聚合酶链反应（RT-PCR）鉴定出来的。在一名患有智力障碍的BMD患者中，发现了第一个异常大的转录本，它是由外显子62和外显子63之间插入58个碱基对产生的。该转录本的起源是内含子62中的一个突变（IVS62-285A>G），这导致出现了一个高质量的供体剪接位点。在一名肌酸激酶（CK）水平高的亚临床BMD患者中，鉴定出了内含子25中的IVS25+2036A>G突变。该突变增强了一个预先存在的受体剪接位点的强度，导致激活了一个95个碱基对的内含子假外显子。通过变性高效液相色谱（DHPLC）分析，发现患者的母亲是体细胞嵌合体。这些新识别的额外外显子的插入导致了过早的终止密码子，但我们可以观察到两名患者中都发生了一定程度的正常剪接。这些残留全长转录本的检测与临床表现和肌营养不良蛋白分析结果一致。这是首次报道假外显子激活作为贝克型肌营养不良症的一种机制，并进一步揭示了导致BMD的遗传异常的多样性。
