Department of Neurology, Neuromuscular Center Ruhrgebiet, University Hospital Bergmannsheil, Ruhr-University Bochum, Germany.
Mol Cell Proteomics. 2013 Jan;12(1):215-27. doi: 10.1074/mcp.M112.023176. Epub 2012 Oct 31.
Filaminopathy is a subtype of myofibrillar myopathy caused by mutations in FLNC, the gene encoding filamin C, and histologically characterized by pathologic accumulation of several proteins within skeletal muscle fibers. With the aim to get new insights in aggregate composition, we collected aggregates and control tissue from skeletal muscle biopsies of six myofibrillar myopathy patients harboring three different FLNC mutations by laser microdissection and analyzed the samples by a label-free mass spectrometry approach. A total of 390 proteins were identified, and 31 of those showed significantly higher spectral indices in aggregates compared with patient controls with a ratio >1.8. These proteins included filamin C, other known myofibrillar myopathy associated proteins, and a striking number of filamin C binding partners. Across the patients the patterns were extremely homogeneous. Xin actin-binding repeat containing protein 2, heat shock protein 27, nebulin-related-anchoring protein, and Rab35 could be verified as new filaminopathy biomarker candidates. In addition, further experiments identified heat shock protein 27 and Xin actin-binding repeat containing protein 2 as novel filamin C interaction partners and we could show that Xin actin-binding repeat containing protein 2 and the known interaction partner Xin actin-binding repeat containing protein 1 simultaneously associate with filamin C. Ten proteins showed significant lower spectral indices in aggregate samples compared with patient controls (ratio <0.56) including M-band proteins myomesin-1 and myomesin-2. Proteomic findings were consistent with previous and novel immunolocalization data. Our findings suggest that aggregates in filaminopathy have a largely organized structure of proteins also interacting under physiological conditions. Different filamin C mutations seem to lead to almost identical aggregate compositions. The finding that filamin C was detected as highly abundant protein in aggregates in filaminopathy indicates that our proteomic approach may be suitable to identify new candidate genes among the many MFM patients with so far unknown mutation.
纤维蛋白病是一种由 FLNC 基因突变引起的肌原纤维肌病亚型，该基因编码纤维蛋白 C，组织学特征为骨骼肌纤维内几种蛋白质的病理性积累。为了深入了解聚集体的组成，我们通过激光微切割收集了六位肌原纤维肌病患者的骨骼肌活检中的聚集体和对照组织，这些患者携带三种不同的 FLNC 突变，并通过无标记质谱方法分析了这些样本。共鉴定出 390 种蛋白质，其中 31 种蛋白质在聚集体中的光谱指数明显高于患者对照，比值>1.8。这些蛋白质包括纤维蛋白 C、其他已知的肌原纤维肌病相关蛋白，以及大量的纤维蛋白 C 结合伴侣。在所有患者中，这些模式都非常一致。Xin 肌动蛋白结合重复蛋白 2、热休克蛋白 27、nebuilin 相关锚定蛋白和 Rab35 可被验证为新的纤维蛋白病生物标志物候选物。此外，进一步的实验还鉴定出热休克蛋白 27 和 Xin 肌动蛋白结合重复蛋白 2 是新的纤维蛋白 C 相互作用伙伴，并且我们可以证明 Xin 肌动蛋白结合重复蛋白 2 和已知的相互作用伙伴 Xin 肌动蛋白结合重复蛋白 1 同时与纤维蛋白 C 结合。与患者对照相比，10 种蛋白质在聚集体样本中的光谱指数明显较低（比值<0.56），包括 M 带蛋白肌联蛋白-1 和肌联蛋白-2。蛋白质组学发现与以前和新的免疫定位数据一致。我们的发现表明，纤维蛋白病中的聚集体具有很大程度上组织化的蛋白质结构，这些蛋白质在生理条件下也相互作用。不同的纤维蛋白 C 突变似乎导致几乎相同的聚集体组成。在纤维蛋白病中，纤维蛋白 C 被检测为聚集体中高度丰富的蛋白质，这表明我们的蛋白质组学方法可能适合在许多 MFM 患者中鉴定新的候选基因，这些患者的突变迄今未知。
