Scheper Wiep, Thaminy Safia, Kais Sanja, Stagljar Igor, Römisch Karin
University of Cambridge, Cambridge Institute for Medical Research, Hills Road, Cambridge CB2 2XY, United Kingdom.
J Biol Chem. 2003 Sep 26;278(39):37998-8003. doi: 10.1074/jbc.M300176200. Epub 2003 Jul 14.
Secretory proteins are translocated across the endoplasmic reticulum (ER) membrane through a channel formed by three proteins, namely Sec61p, Sbh1p, and Sss1p (Johnson, A. E., and van Waes, M. A. (1999) Annu. Rev. Cell Dev. Biol. 15, 799-842). Sec61p and Sss1p are essential for translocation (Esnault, Y., Blondel, M. O., Deshaies, R. J., Schekman, R., and Kepes, F. (1993) EMBO J. 12, 4083-4093). Sec61p is a polytopic membrane protein that lines the protein translocation channel. The role of Sss1p is unknown. During import into the ER through the Sec61p channel, many proteins are N-glycosylated before translocation is completed. In addition, both the Sec61 channel and oligosaccharyl transferase (OST) copurify with ribosomes from rough ER, suggesting that OST is located in close proximity to the Sec61 channel (Gorlich, D., Prehn, S., Hartmann, E., Kalies, K.-U., and Rapoport, T. A. (1992) Cell 71, 489-503 and Wang, L., and Dobberstein, B. (1999) FEBS Lett. 457, 316-322). Here, we demonstrate a direct interaction between Sss1p and a subunit of OST, Wbp1p, using the split-ubiquitin system and co-immunoprecipitation. We generated mutants in the cytoplasmic domain of Sss1p that disturb the interaction with OST and are viable but display a translocation defect specific for proteins with glycosylation acceptor sites. Our data suggest that Sss1p coordinates translocation across the ER membrane and N-linked glycosylation of secretory proteins.
分泌蛋白通过由三种蛋白质Sec61p、Sbh1p和Sss1p形成的通道跨内质网（ER）膜转运（约翰逊，A.E.，和范·瓦伊斯，M.A.（1999年）《细胞与发育生物学年度评论》15，799 - 842）。Sec61p和Sss1p对于转运是必需的（埃斯纳尔，Y.，布隆代尔，M.O.，德谢伊斯，R.J.，谢克曼，R.，和凯佩斯，F.（1993年）《欧洲分子生物学组织杂志》12，4083 - 4093）。Sec61p是一种多结构域膜蛋白，构成蛋白质转运通道的内衬。Sss1p的作用尚不清楚。在通过Sec61p通道导入内质网的过程中，许多蛋白质在转运完成之前就进行了N - 糖基化。此外，Sec61通道和寡糖基转移酶（OST）都与来自糙面内质网的核糖体共纯化，这表明OST位于Sec61通道附近（戈利希，D.，普雷恩，S.，哈特曼，E.，卡利斯，K.-U.，和拉波波特，T.A.（1992年）《细胞》71，489 - 503；以及王，L.，和多伯斯坦，B.（1999年）《欧洲生物化学学会联合会快报》457，316 - 322）。在这里，我们使用分裂泛素系统和免疫共沉淀证明了Sss1p与OST的一个亚基Wbp1p之间存在直接相互作用。我们在Sss1p的胞质结构域中产生了突变体，这些突变体扰乱了与OST的相互作用，并且是可行的，但表现出对具有糖基化受体位点的蛋白质具有特异性的转运缺陷。我们的数据表明，Sss1p协调分泌蛋白跨内质网膜的转运和N - 连接糖基化。
