Infectious bronchitis virus generates spherules from zippered endoplasmic reticulum membranes.
Infectious bronchitis virus generates spherules from zippered endoplasmic reticulum membranes.
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DOI：
10.1128/mbio.00801-13
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发表时间：
2013-10-22
期刊：
影响因子：
6.4
通讯作者：
Britton P
中科院分区：
文献类型：
--
作者：
Maier HJ;Hawes PC;Cottam EM;Mantell J;Verkade P;Monaghan P;Wileman T;Britton P
正义RNA病毒的复制与细胞膜的重排有关。先前关于β冠状病毒小鼠肝炎病毒和严重急性呼吸综合征冠状病毒（SARS-CoV）感染组织培养细胞系的研究表明，它们产生双膜囊泡（dmv）和卷曲膜，作为网状膜网络的一部分。在这里，我们描述了由禽γ冠状病毒传染性支气管炎病毒（IBV）在哺乳动物细胞系、原代禽细胞和离体气管器官培养的上皮细胞中诱导的膜重排的详细研究。在所有细胞类型中，发现了IBV感染的新结构，我们称之为拉链内质网（ER）和球粒。拉链内质网缺乏腔隙，提示内质网池有拉链，小球呈均匀内陷。电子断层扫描显示，ibv诱导的小球体被拴在拉链内质网上，并且有一个通道将小球体的内部与细胞质连接起来，这一特征被认为是RNA合成位点所必需的，但在冠状病毒诱导的膜重排中却没有发现。我们还在ibv感染的细胞中发现了dmv，这些dmv被观察为单个的dmv，或者通过外膜连接到内质网，但不连接到拉链内质网。有趣的是，IBV诱导的球体与甲病毒、诺达病毒和溴病毒的RNA合成位点非常相似，这可能表明IBV和这些不同的病毒在RNA复制复合物的组装方面具有相似的策略。所有正义单链RNA病毒诱导细胞膜重排，为病毒复制复合体组装提供平台，并保护病毒RNA免受细胞防御。我们研究了一种重要的家禽病原体——γ冠状病毒传染性支气管炎病毒（IBV）诱导的膜重排。先前研究密切相关的冠状病毒的工作鉴定了感染细胞中源自内质网（ER）的双膜囊泡（DMVs）和卷曲膜（CMs）。然而，dmv和CMs在病毒RNA合成中的作用尚不清楚，因为这些密封的囊泡缺乏将病毒RNA传递到细胞质的手段。在这里，我们描述了IBV感染的新结构：拉链内质网和系在拉链内质网上的小泡，称为球粒。值得注意的是，小球体包含一个将其内部连接到细胞质的通道，并且与其他正意义RNA病毒的RNA合成位点非常相似，使其成为IBV RNA合成位点的理想候选者。所有正义单链RNA病毒诱导细胞膜重排，为病毒复制复合体组装提供平台，保护病毒RNA免受细胞防御。我们研究了一种重要的家禽病原体——γ冠状病毒传染性支气管炎病毒（IBV）诱导的膜重排。先前研究密切相关的冠状病毒的工作鉴定了感染细胞中源自内质网（ER）的双膜囊泡（DMVs）和卷曲膜（CMs）。然而，dmv和CMs在病毒RNA合成中的作用尚不清楚，因为这些密封的囊泡缺乏将病毒RNA传递到细胞质的手段。在这里，我们描述了IBV感染的新结构：拉链内质网和系在拉链内质网上的小泡，称为球粒。值得注意的是，小球体包含一个将其内部连接到细胞质的通道，并且与其他正意义RNA病毒的RNA合成位点非常相似，使其成为IBV RNA合成位点的理想候选者。
Replication of positive-sense RNA viruses is associated with the rearrangement of cellular membranes. Previous work on the infection of tissue culture cell lines with the betacoronaviruses mouse hepatitis virus and severe acute respiratory syndrome coronavirus (SARS-CoV) showed that they generate double-membrane vesicles (DMVs) and convoluted membranes as part of a reticular membrane network. Here we describe a detailed study of the membrane rearrangements induced by the avian gammacoronavirus infectious bronchitis virus (IBV) in a mammalian cell line but also in primary avian cells and in epithelial cells of ex vivo tracheal organ cultures. In all cell types, structures novel to IBV infection were identified that we have termed zippered endoplasmic reticulum (ER) and spherules. Zippered ER lacked luminal space, suggesting zippering of ER cisternae, while spherules appeared as uniform invaginations of zippered ER. Electron tomography showed that IBV-induced spherules are tethered to the zippered ER and that there is a channel connecting the interior of the spherule with the cytoplasm, a feature thought to be necessary for sites of RNA synthesis but not seen previously for membrane rearrangements induced by coronaviruses. We also identified DMVs in IBV-infected cells that were observed as single individual DMVs or were connected to the ER via their outer membrane but not to the zippered ER. Interestingly, IBV-induced spherules strongly resemble confirmed sites of RNA synthesis for alphaviruses, nodaviruses, and bromoviruses, which may indicate similar strategies of IBV and these diverse viruses for the assembly of RNA replication complexes. IMPORTANCE All positive-sense single-stranded RNA viruses induce rearranged cellular membranes, providing a platform for viral replication complex assembly and protecting viral RNA from cellular defenses. We have studied the membrane rearrangements induced by an important poultry pathogen, the gammacoronavirus infectious bronchitis virus (IBV). Previous work studying closely related betacoronaviruses identified double-membrane vesicles (DMVs) and convoluted membranes (CMs) derived from the endoplasmic reticulum (ER) in infected cells. However, the role of DMVs and CMs in viral RNA synthesis remains unclear because these sealed vesicles lack a means of delivering viral RNA to the cytoplasm. Here, we characterized structures novel to IBV infection: zippered ER and small vesicles tethered to the zippered ER termed spherules. Significantly, spherules contain a channel connecting their interior to the cytoplasm and strongly resemble confirmed sites of RNA synthesis for other positive-sense RNA viruses, making them ideal candidates for the site of IBV RNA synthesis. All positive-sense single-stranded RNA viruses induce rearranged cellular membranes, providing a platform for viral replication complex assembly and protecting viral RNA from cellular defenses. We have studied the membrane rearrangements induced by an important poultry pathogen, the gammacoronavirus infectious bronchitis virus (IBV). Previous work studying closely related betacoronaviruses identified double-membrane vesicles (DMVs) and convoluted membranes (CMs) derived from the endoplasmic reticulum (ER) in infected cells. However, the role of DMVs and CMs in viral RNA synthesis remains unclear because these sealed vesicles lack a means of delivering viral RNA to the cytoplasm. Here, we characterized structures novel to IBV infection: zippered ER and small vesicles tethered to the zippered ER termed spherules. Significantly, spherules contain a channel connecting their interior to the cytoplasm and strongly resemble confirmed sites of RNA synthesis for other positive-sense RNA viruses, making them ideal candidates for the site of IBV RNA synthesis.