Ferris R L, Hall C, Sipsas N V, Safrit J T, Trocha A, Koup R A, Johnson R P, Siliciano R F
Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA.
J Immunol. 1999 Feb 1;162(3):1324-32.
Processing of viral proteins for recognition by CTL involves degradation of the proteins in the cytosol of an infected cell followed by transport of the resulting peptides into the endoplasmic reticulum (ER) by the TAP1/2 complex. Uncertainty exists over the site of processing of viral envelope (env) proteins since the extracellular domains of env proteins are not present in the cytosol where the class I Ag-processing pathway begins. Rather, the ectodomains of env proteins are cotranslationally translocated into the ER during biosynthesis. To analyze env protein processing, we used the herpes simplex virus protein ICP47 to block peptide transport by TAP1/2 and examined the effects of TAP blockade on the processing of the HIV-1 env protein. For the majority of env-specific CD8+ CTL, the processing pathway required TAP1/2-mediated transport of cytosolic peptides into the ER. To determine how env peptides are generated in the cytosol, we analyzed the processing of two TAP1/2-dependent epitopes containing N-linked glycosylation sites. In each case, processing involved glycosylation-dependent posttranslational modification of asparagine residues to aspartic acid. These results are consistent with cotranslational translocation of env into the ER, where glycosylation occurs. This is followed by export of a fraction of the newly synthesized protein into the cytosol, where it is deglycosylated, with conversion of the asparagines to aspartic acid residues. Following cytoplasmic proteolysis, env peptides are retransported by TAP1/2 into the ER, where association with class I occurs. Thus, the env protein can enter the class I pathway through multiple distinct processing mechanisms.
病毒蛋白经加工后供细胞毒性T淋巴细胞（CTL）识别，这一过程包括在被感染细胞的胞质溶胶中降解蛋白，随后通过TAP1/2复合体将产生的肽转运至内质网（ER）。由于env蛋白的细胞外结构域不存在于I类抗原加工途径起始的胞质溶胶中，因此病毒包膜（env）蛋白的加工位点尚不确定。相反，env蛋白的胞外结构域在生物合成过程中通过共翻译转运至内质网。为了分析env蛋白的加工过程，我们使用单纯疱疹病毒蛋白ICP47阻断TAP1/2介导的肽转运，并研究TAP阻断对HIV-1 env蛋白加工的影响。对于大多数env特异性CD8+ CTL而言，加工途径需要TAP1/2介导的胞质肽转运至内质网。为了确定env肽如何在胞质溶胶中产生，我们分析了两个含有N-连接糖基化位点的TAP1/2依赖性表位的加工过程。在每种情况下，加工过程都涉及天冬酰胺残基糖基化依赖性的翻译后修饰为天冬氨酸。这些结果与env共翻译转运至内质网一致，糖基化在内质网中发生。随后，一部分新合成的蛋白输出至胞质溶胶，在那里它被去糖基化，天冬酰胺转化为天冬氨酸残基。胞质蛋白水解后，env肽通过TAP1/2重新转运至内质网，在那里与I类分子结合。因此，env蛋白可通过多种不同的加工机制进入I类途径。
