Dislocation of an endoplasmic reticulum membrane glycoprotein involves the formation of partially dislocated ubiquitinated polypeptides
Dislocation of an endoplasmic reticulum membrane glycoprotein involves the formation of partially dislocated ubiquitinated polypeptides
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DOI：
10.1074/jbc.m704315200
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发表时间：
2007-09-14
影响因子：
4.8
通讯作者：
Tortorella, Domenico
中科院分区：
文献类型：
--
作者：
Baker, Brooke M.;Tortorella, Domenico
内质网（ER）中不正确折叠的多肽的积累可以触发应激反应，导致异常蛋白质输出到细胞质中并最终导致蛋白酶体降解。人类巨细胞病毒编码一种I型糖蛋白US11，它与新生的MHC I类重链分子结合，并导致它们从内质网脱位到细胞质中，在那里它们被蛋白酶体降解。对us11介导的I类降解的研究已经确定了许多参与位错反应的细胞蛋白，包括胞质AAA atp酶p97、膜蛋白Derlin-1和E3泛素连接酶Sel1L。然而，发生在位错的起始和错误折叠的底物完全提取到胞质溶胶之间的中间步骤尚不清楚。我们证明了US11本身经历ER输出和蛋白酶体降解，并利用该系统定义了US11位错的多个步骤。用蛋白酶体抑制剂处理表达US11的细胞导致糖基化和泛素化物种的积累，以及去糖基化的US11中间体。蛋白酶体抑制的US11细胞的亚细胞分离表明，去糖基化的中间体继续在内质网膜内整合，这表明蛋白酶体在位错的后期步骤中起作用。这些数据支持US11被泛素修饰的模型，而跨膜区域则整合在内质网膜中，并且在完全错位之前发生去糖基化。
Accumulation of improperly folded polypeptides in the endoplasmic reticulum ( ER) can trigger a stress response that leads to the export of aberrant proteins into the cytosol and their ultimate proteasomal degradation. Human cytomegalovirus encodes a type I glycoprotein, US11, that binds to nascent MHC class I heavy chain molecules and causes their dislocation from the ER to the cytosol where they are degraded by the proteasome. Examination of US11-mediated class I degradation has identified a host of cellular proteins involved in the dislocation reaction, including the cytosolic AAA ATPase p97, the membrane protein Derlin-1, and the E3 ubiquitin ligase Sel1L. However, the intermediate steps occurring between the initiation of dislocation and full extraction of the misfolded substrate into the cytosol are not known. We demonstrate that US11 itself undergoes ER export and proteasomal degradation and utilize this system to define multiple steps of US11 dislocation. Treatment of US11-expressing cells with proteasome inhibitor resulted in the accumulation of glycosylated and ubiquitinated species as well as a deglycosylated US11 intermediate. Subcellular fractionation of proteasome-inhibited US11 cells demonstrated that deglycosylated intermediates continued to be integrated within the ER membrane, suggesting that the proteasome functions in the latter steps of dislocation. The data supports a model in which US11 is modified with ubiquitin, whereas the transmembrane region is integrated in the ER membrane, and deglycosylation occurs before complete dislocation.