The Endoplasmic Reticulum Chaperone BiP/GRP78 Is Important in the Structure and Function of the Human Cytomegalovirus Assembly Compartment
The Endoplasmic Reticulum Chaperone BiP/GRP78 Is Important in the Structure and Function of the Human Cytomegalovirus Assembly Compartment
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DOI：
10.1128/jvi.00762-09
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发表时间：
2009-11-15
影响因子：
5.4
通讯作者：
Alwine, James C.
中科院分区：
文献类型：
--
作者：
Buchkovich, Nicholas J.;Maguire, Tobi G.;Alwine, James C.
我们先前已经证明内质网(ER)伴侣蛋白Bip在人巨细胞病毒(HCMV)的组装和输出中发挥作用。在这里，我们发现Bip定位于感染细胞的两个细胞质结构中。针对极端C末端的抗体，包括Bip的KDEL ER定位序列，在细胞外围附近的浓缩内质网区域检测Bip。针对全长、N末端或C末端较大部分的抗体可检测组装隔间中的BIP。这种C-末端抗体不能检测到组装间的Bip，这表明Bip的KDEL序列被封闭在组装间。Bip的耗尽导致浓缩的内质网和组装隔间解离，表明Bip对它们的完整性很重要。BiP和pp28在组装室中相互关联，因为在组装室中检测BiP的抗体共免疫沉淀pp28，反之亦然。此外，Bip和pp28在蔗糖梯度上与其他组装室组件共同作用。BIP还能免疫共沉淀TRS1。先前的数据表明，感染了TRS1缺陷病毒的细胞具有细胞质和组装间隔室的缺陷，就像BiP耗尽时看到的那样。我们表明，TRS1的一部分与组装室一起纯化。这些发现表明，Bip和TRS1在装配舱的维持中具有共同的功能。总而言之，BiP从ER转移到与pp28和TRS1相关联，有助于组装隔间的完整性和功能。
We previously demonstrated that the endoplasmic reticulum (ER) chaperone BiP functions in human cytomegalovirus (HCMV) assembly and egress. Here, we show that BiP localizes in two cytoplasmic structures in infected cells. Antibodies to the extreme C terminus, which includes BiP's KDEL ER localization sequence, detect BiP in regions of condensed ER near the periphery of the cell. Antibodies to the full length, N terminus, or larger portion of the C terminus detect BiP in the assembly compartment. This inability of C-terminal antibodies to detect BiP in the assembly compartment suggests that BiP's KDEL sequence is occluded in the assembly compartment. Depletion of BiP causes the condensed ER and assembly compartments to dissociate, indicating that BiP is important for their integrity. BiP and pp28 are in association in the assembly compartment, since antibodies that detect BiP in the assembly compartment coimmunoprecipitate pp28 and vice versa. In addition, BiP and pp28 copurify with other assembly compartment components on sucrose gradients. BiP also coimmunoprecipitates TRS1. Previous data show that cells infected with a TRS1-deficient virus have cytoplasmic and assembly compartment defects like those seen when BiP is depleted. We show that a fraction of TRS1 purifies with the assembly compartment. These findings suggest that BiP and TRS1 share a function in assembly compartment maintenance. In summary, BiP is diverted from the ER to associate with pp28 and TRS1, contributing to the integrity and function of the assembly compartment.