Ikeya K, Saito K, Hayashi K, Tanaka H, Hagiwara Y, Yoshida M, Yamauchi A, Fukuyama Y, Ishiguro T, Eguchi C
Division of Cell Biology, National Institute of Neuroscience, NCNP, Kodaira, Japan.
Am J Med Genet. 1992 Jun 1;43(3):580-7. doi: 10.1002/ajmg.1320430315.
We examined the nucleotide sequence of deleted part of dystrophin mRNA and its translational product with immunoblot and immunohistochemical methods in a 6-year-old boy with a deleted DMD/BMD gene. On Southern blot analysis of his genomic DNA, we found a deletion of exons 10 to 37 in the DMD/BMD gene, which was expected to preserve the translational open reading frame (ORF). Dystrophin mRNA from his biopsy sample was amplified by polymerase chain reaction (PCR) and sequenced. The mRNA lacked the sequence corresponding to the gene from exons 10-37, and the translational ORF was preserved. The transcript was expected to code a 260 kDa protein. Dystrophin expressed in this patient was investigated with immunological methods. A 260 kDa protein was detected by immunoblot analysis with antidystrophin antiserum against nondeleted regions. These observations confirmed the preservation of the reading frame and the 260 kDa protein was produced as a mutant dystrophin. All these are compatible with the diagnosis of BMD. However, the immunohistochemical pattern of his muscle cells was peculiar. With deleted-region-directed antiserum, the membrane was not stained at all as in DMD patients. In contrast, with nondeleted-region-directed antiserum, all the muscle cell membrane was stained continuously as in non-DMD/BMD individuals. These are quite different from the staining pattern in most BMD patients where muscles are stained patchily or discontinuously.
我们运用免疫印迹法和免疫组织化学方法，对一名患有DMD/BMD基因缺失的6岁男孩的抗肌萎缩蛋白mRNA缺失部分的核苷酸序列及其翻译产物进行了检测。对其基因组DNA进行Southern印迹分析时，我们发现DMD/BMD基因中外显子10至37缺失，预计这将保留翻译开放阅读框（ORF）。通过聚合酶链反应（PCR）扩增并测序了来自其活检样本的抗肌萎缩蛋白mRNA。该mRNA缺少对应于外显子第10 - 37位基因的序列，但翻译ORF得以保留。预计该转录本编码一种260 kDa的蛋白质。我们用免疫学方法研究了该患者体内表达的抗肌萎缩蛋白。用抗非缺失区域的抗肌萎缩蛋白抗血清进行免疫印迹分析，检测到了一种260 kDa的蛋白质。这些观察结果证实了阅读框得以保留，并且产生了一种作为突变抗肌萎缩蛋白的260 kDa蛋白质。所有这些均与贝克型肌营养不良（BMD）的诊断相符。然而，其肌肉细胞的免疫组织化学模式却很特殊。使用针对缺失区域的抗血清时，细胞膜完全未被染色，如同杜氏肌营养不良（DMD）患者一样。相反，使用针对非缺失区域的抗血清时，所有肌肉细胞膜均连续染色，如同非DMD/BMD个体一样。这些与大多数BMD患者的染色模式截然不同，在大多数BMD患者中，肌肉染色呈斑驳状或不连续。
