Reovirus nonstructural protein μNS recruits viral core surface proteins and entering core particles to factory-like inclusions
Reovirus nonstructural protein μNS recruits viral core surface proteins and entering core particles to factory-like inclusions
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DOI：
10.1128/jvi.78.4.1882-1892.2004
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发表时间：
2004-02-01
影响因子：
5.4
通讯作者：
Parker, JSL
中科院分区：
文献类型：
--
作者：
Broering, TJ;Kim, J;Parker, JSL
哺乳动物呼肠孤病毒被认为在称为病毒工厂的细胞质非膜结构中组装和复制。在体外实验中，病毒非结构蛋白muNS在没有其他病毒蛋白的情况下表达时形成工厂样的球状包涵体，并结合到病毒核心颗粒的表面。鉴于这些先前的观察结果，我们假设一种或多种核心表面蛋白可能通过与muNS的特定关联而被招募到病毒工厂。我们发现所有这三种蛋白-lambda1， lambda2和sigma2-都定位于感染细胞中的工厂，但当它们在没有其他病毒蛋白的情况下单独表达时，它们分散分布在细胞质和细胞核中。另一方面，当与muNS单独共表达时，每个核心表面蛋白都与muNS在球状包涵体中共定位，这支持了最初的假设。我们还发现lambda1、lambda2和sigma2都定位于muNS和mu2（一种与微管相关的结构较小的核心蛋白）共表达形成的丝状内含物上。muNS的前40个残基是与mu2和rna结合的非结构蛋白sigmaNS结合所必需的，而与三种核心表面蛋白中的任何一种结合都不需要。当在没有muNS的情况下与mu2共表达时，每个核心表面蛋白弥散分布，仅在细丝上与mu2表现出零星的弱关联。经过环己亚胺处理的细胞进入细胞质并部分脱壳后，许多核心颗粒被募集到已在这些细胞中形成的muNS内含物中，这证明muNS可以在体内与核心表面结合。这些发现扩展了病毒和细胞成分如何被招募到感染细胞中的病毒工厂的模型，并为病毒蛋白muNS和mu2在这一过程中的核心但独特的作用提供了进一步的证据。
Mammalian reoviruses are thought to assemble and replicate within cytoplasmic, nonmembranous structures called viral factories. The viral nonstructural protein muNS forms factory-like globular inclusions when expressed in the absence of other viral proteins and binds to the surfaces of the viral core particles in vitro. Given these previous observations, we hypothesized that one or more of the core surface proteins may be recruited to viral factories through specific associations with muNS. We found that all three of these proteins-lambda1, lambda2, and sigma2-localized to factories in infected cells but were diffusely distributed through the cytoplasm and nucleus when each was separately expressed in the absence of other viral proteins. When separately coexpressed with muNS, on the other hand, each core surface protein colocalized with muNS in globular inclusions, supporting the initial hypothesis. We also found that lambda1, lambda2, and sigma2 each localized to filamentous inclusions formed upon the coexpression of muNS and mu2, a structurally minor core protein that associates with micro-tubules. The first 40 residues of muNS, which are required for association with mu2 and the RNA-binding nonstructural protein sigmaNS, were not required for association with any of the three core surface proteins. When coexpressed with mu2 in the absence of muNS, each of the core surface proteins was diffusely distributed and displayed only sporadic, weak associations with mu2 on filaments. Many of the core particles that entered the cytoplasm of cycloheximide-treated cells following entry and partial uncoating were recruited to inclusions of muNS that had been preformed in those cells, providing evidence that muNS can bind to the surfaces of cores in vivo. These findings expand a model for how viral and cellular components are recruited to the viral factories in infected cells and provide further evidence for the central but distinct roles of viral proteins muNS and mu2 in this process.