Endoplasmic reticulum (ER)-associated degradation of misfolded N-linked glycoproteins is suppressed upon inhibition of ER mannosidase I
Endoplasmic reticulum (ER)-associated degradation of misfolded N-linked glycoproteins is suppressed upon inhibition of ER mannosidase I
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DOI：
10.1074/jbc.m001073200
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发表时间：
2000-12-29
影响因子：
4.8
通讯作者：
Arvan, P
中科院分区：
文献类型：
--
作者：
Tokunaga, F;Brostrom, C;Arvan, P
为了研究早期碳水化合物识别/修剪反应在靶向内质网 (ER) 保留的错误折叠糖蛋白以进行 ER 相关降解 (ERAD) 中的作用，我们在中国仓鼠卵巢细胞中稳定表达了 cog 甲状腺球蛋白 (Tg) 突变体 cDNA。我们发现 ER 甘露糖苷酶 I（但不是其他糖苷酶）的抑制剂强烈抑制 Cog Tg 降解，并且还扰乱二硫苏糖醇还原的 Tg 以及华法林处理的幼仓鼠肾细胞中表达的 γ-羧化缺陷蛋白 C 的 ERAD 过程。 Kifunensine 抑制 ER 甘露糖苷酶 I 也抑制栗精胺处理的细胞中的 ERAD；因此，抑制 ERAD 不需要 ER 伴侣钙连接蛋白和钙网蛋白与单葡糖基化寡糖的凝集素样结合。值得注意的是，未降解的蛋白质部分仍然完全与微粒体相关。在脉冲追踪研究中，即使在 Tg 合成后 1 小时，kifunensine 敏感的降解仍然是可抑制的。有趣的是，kifunensine 的长期治疗导致中国仓鼠卵巢细胞中 Cog Tg 的积累增加了 3 倍，但并未导致 ER 未折叠蛋白反应的显着诱导。我们假设，以不需要钙连接蛋白/钙网蛋白的凝集素样活性的方式，特定支链N-连接甘露糖结构的识别或加工增强了糖蛋白从内质网腔逆向转位的效率。
To examine the role of early carbohydrate recognition/trimming reactions in targeting endoplasmic reticulum (ER)-retained, misfolded glycoproteins for ER-associated degradation (ERAD), we have stably expressed the cog thyroglobulin (Tg) mutant cDNA in Chinese hamster ovary cells. We found that inhibitors of ER mannosidase I (but not other glycosidases) acutely suppressed Cog Tg degradation and also perturbed the ERAD process for Tg reduced with dithiothreitol as well as for gamma -carboxylation-deficient protein C expressed in warfarin-treated baby hamster kidney cells. Kifunensine inhibition of ER mannosidase I also suppressed ERAD in castanospermine-treated cells; thus, suppression of ERAD does not require lectin-like binding of ER chaperones calnexin and calreticulin to monoglucosylated oligosaccharides. Notably, the undegraded protein fraction remained completely microsome-associated. In pulse-chase studies, kifunensine-sensitive degradation was still inhibitable even 1 h after Tg synthesis. Intriguingly, chronic treatment with kifunensine caused a 3-fold accumulation of Cog Tg in Chinese hamster ovary cells and did not lead to significant induction of the ER unfolded protein response. We hypothesize that, in a manner not requiring lectin-like activity of calnexin/ calreticulin, the recognition or processing of a specific branched N-linked mannose structure enhances the efficiency of glycoprotein retrotranslocation from the ER lumen.