INTRACELLULAR MANIPULATION OF DISULFIDE BOND FORMATION IN ROTAVIRUS PROTEINS DURING ASSEMBLY
INTRACELLULAR MANIPULATION OF DISULFIDE BOND FORMATION IN ROTAVIRUS PROTEINS DURING ASSEMBLY
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DOI：
10.1128/jvi.68.8.5204-5215.1994
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发表时间：
1994-08-01
影响因子：
5.4
通讯作者：
GREENBERG, HB
中科院分区：
文献类型：
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作者：
SVENSSON, L;DORMITZER, PR;GREENBERG, HB
轮状病毒在感染细胞的粗面内质网（RER）中经历独特的组装模式。管腔粗面内质网蛋白进行显着的共翻译和翻译后修饰，包括二硫键的形成。向轮状病毒感染的细胞中添加还原剂（二硫苏糖醇[DTT]）不会显着抑制翻译或破坏轮状病毒蛋白中已建立的二硫键，但可以防止新二硫键和感染性病毒后代的形成。在DTT处理，轮状病毒感染的细胞中，所有的VP4，VP6，和NS 28表位，但没有VP7表位的单克隆抗体组通过免疫组化染色检测。当DTT处理的细胞中重新建立氧化条件时，VP7中的分子内二硫键迅速而正确地建立，并恢复抗原性，尽管长时间的DTT处理导致了永久性错误折叠的VP7的积累。电子显微镜显示，单壳颗粒和出芽到ER的胞质组装不受DTT治疗的影响，但外部衣壳组装被阻断，导致单壳和包膜中间亚病毒颗粒在RER管腔中的积累。
Rotavirus undergoes a unique mode of assembly in the rough endoplasmic reticulum (RER) of infected cells. Luminal RER proteins undergo significant cotranslational and posttranslational modifications, including disulfide bond formation. Addition of a reducing agent (dithiothreitol [DTT]) to rotavirus-infected cells did not significantly inhibit translation or disrupt established disulfide bonds in rotavirus proteins but prevented the formation of new disulfide bonds and infectious viral progeny. In DTT-treated, rotavirus-infected cells, all vp4, vp6, and ns28 epitopes but no vp7 epitopes were detected by immunohistochemical staining with a panel of monoclonal antibodies. When oxidizing conditions were reestablished in DTT-treated cells, intramolecular disulfide bonds in vp7 were rapidly and correctly established with the restoration of antigenicity, although prolonged DTT treatment led to the accumulation of permanently misfolded vp7. Electron microscopy revealed that cytosolic assembly of single-shelled particles and budding into the ER was not affected by DTT treatment but that outer capsid assembly was blocked, leading to the accumulation of single-shelled and enveloped intermediate subviral particles in the RER lumen.