Selcen Duygu, Muntoni Francesco, Burton Barbara K, Pegoraro Elena, Sewry Caroline, Bite Anna V, Engel Andrew G
Department of Neurology and Neuromuscular Research Laboratory, Mayo Clinic, Rochester, NY, USA.
Ann Neurol. 2009 Jan;65(1):83-9. doi: 10.1002/ana.21553.
Myofibrillar myopathies (MFMs) are morphologically distinct but genetically heterogeneous muscular dystrophies in which disintegration of Z disks and then of myofibrils is followed by ectopic accumulation of multiple proteins. Cardiomyopathy, neuropathy, and dominant inheritance are frequent associated features. Mutations in alphaB-crystallin, desmin, myotilin, Zasp, or filamin-C can cause MFMs and were detected in 32 of 85 patients of the Mayo MFM cohort. Bag3, another Z-disk-associated protein, has antiapoptotic properties, and its targeted deletion in mice causes fulminant myopathy with early lethality. We therefore searched for mutations in BAG3 in 53 unrelated MFM patients.
We searched for mutations in BAG3 by direct sequencing. We analyzed structural changes in muscle by histochemistry, immunocytochemistry, and electron microscopy, examined mobility of the mutant Bag3 by nondenaturing electrophoresis, and searched for abnormal aggregation of the mutant protein in COS-7 (SV-40 transformed monkey kidney fibroblast-7) cells.
We identified a heterozygous p.Pro209Leu mutation in three patients. All presented in childhood, had progressive limb and axial muscle weakness, and experienced development of cardiomyopathy and severe respiratory insufficiency in their teens; two had rigid spines, and one a peripheral neuropathy. Electron microscopy showed disintegration of Z disks, extensive accumulation of granular debris and larger inclusions, and apoptosis of 8% of the nuclei. On nondenaturing electrophoresis of muscle extracts, the Bag3 complex migrated faster in patient than control extracts, and expression of FLAG-labeled mutant and wild-type Bag3 in COS cells showed abnormal aggregation of the mutant protein.
We conclude mutation in Bag3 defines a novel severe autosomal dominant childhood muscular dystrophy.
肌原纤维肌病（MFMs）是形态学上有明显特征但基因异质性的肌营养不良症，其中Z盘随后是肌原纤维解体，接着多种蛋白质异位积聚。心肌病、神经病变和显性遗传是常见的相关特征。αB-晶状体蛋白、结蛋白、肌联蛋白、Zasp或细丝蛋白C的突变可导致MFMs，在梅奥MFMs队列的85名患者中有32名检测到这些突变。Bag3是另一种与Z盘相关的蛋白质，具有抗凋亡特性，其在小鼠中的靶向缺失会导致暴发性肌病并早期致死。因此，我们在53名无亲缘关系的MFMs患者中寻找BAG3的突变。
我们通过直接测序寻找BAG3的突变。我们通过组织化学、免疫细胞化学和电子显微镜分析肌肉的结构变化，通过非变性电泳检测突变型Bag3的迁移率，并在COS-7（SV-40转化的猴肾成纤维细胞-7）细胞中寻找突变蛋白的异常聚集。
我们在三名患者中鉴定出杂合的p.Pro209Leu突变。所有患者均在儿童期发病，有进行性肢体和躯干肌无力，在十几岁时出现心肌病和严重呼吸功能不全；两名患者有脊柱僵硬，一名患者有周围神经病变。电子显微镜显示Z盘解体，颗粒碎片和较大包涵体广泛积聚，8%的细胞核发生凋亡。在肌肉提取物的非变性电泳中，患者提取物中的Bag3复合物迁移速度比对照提取物快，FLAG标记的突变型和野生型Bag3在COS细胞中的表达显示突变蛋白有异常聚集。
我们得出结论，Bag3突变定义了一种新型严重常染色体显性遗传性儿童肌营养不良症。
