Isosomppi Juha, Vesa Jouni, Jalanko Anu, Peltonen Leena
Department of Molecular Medicine, National Public Health Institute and Department of Medical Genetics, University of Helsinki, Finland.
Hum Mol Genet. 2002 Apr 15;11(8):885-91. doi: 10.1093/hmg/11.8.885.
The Finnish variant late infantile neuronal ceroid lipofuscinosis (vLINCL) belongs to the neuronal ceroid lipofuscinosis group of common recessively inherited neurodegenerative disorders. The CLN 5 gene responsible for this brain disorder codes for a novel protein with no homology to previously reported proteins. In this study, we have investigated the biosynthesis and intracellular localization of this protein in transiently transfected BHK-21 cells using a CLN5-specific peptide antibody. Confocal immunofluorescence microscopy showed that wild-type CLN5 is predominantly targeted to lysosomes and immunoprecipitation analysis recognized a 60 kDa polypeptide. The molecular weight of this protein was reduced to 40 kDa by deglycosylation with Endo H and to 38 kDa with PNGase F. The same-sized glycosylated polypeptides were also observed in the media, suggesting that the 60 kDa glycosylated CLN5 polypeptide represents a soluble lysosomal glycoprotein, not an integral transmembrane protein as predicted earlier. The most common human vLINCL mutation blocked the lysosomal targeting of expressed polypeptides. This would imply that the pathogenesis of vLINCL would be associated with the defective lysosomal trafficking, preventing the normal biological function of the corresponding polypeptide.
芬兰型晚发性婴儿神经元蜡样脂褐质沉积症（vLINCL）属于常见的隐性遗传神经退行性疾病中的神经元蜡样脂褐质沉积症组。导致这种脑部疾病的CLN 5基因编码一种与先前报道的蛋白质无同源性的新型蛋白质。在本研究中，我们使用CLN5特异性肽抗体，在瞬时转染的BHK - 21细胞中研究了该蛋白质的生物合成和细胞内定位。共聚焦免疫荧光显微镜显示，野生型CLN5主要靶向溶酶体，免疫沉淀分析识别出一条60 kDa的多肽。用内切糖苷酶H去糖基化后，该蛋白质的分子量降至40 kDa，用肽-N-糖苷酶F处理后降至38 kDa。在培养基中也观察到了相同大小的糖基化多肽，这表明60 kDa的糖基化CLN5多肽代表一种可溶性溶酶体糖蛋白，而不是如先前预测的整合跨膜蛋白。最常见的人类vLINCL突变阻断了表达多肽的溶酶体靶向。这意味着vLINCL的发病机制可能与溶酶体运输缺陷有关，从而阻止了相应多肽的正常生物学功能。
