Stress Granule Components G3BP1 and G3BP2 Play a Proviral Role Early in Chikungunya Virus Replication
Stress Granule Components G3BP1 and G3BP2 Play a Proviral Role Early in Chikungunya Virus Replication
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DOI：
10.1128/jvi.03612-14
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发表时间：
2015-04-01
影响因子：
5.4
通讯作者：
van Hemert, Martijn J.
中科院分区：
文献类型：
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作者：
Scholte, Florine E. M.;Tas, Ali;van Hemert, Martijn J.
应激颗粒(SGS)是一种蛋白质-信使核糖核酸的聚合体，它是对环境胁迫的反应，导致翻译抑制。SGS通常被认为具有抗病毒作用，并被许多病毒操纵，包括各种甲型病毒。GTP酶激活蛋白(SH3结构域)结合蛋白1(G3BP1)是SGS的关键成分和常用的标志物。它的同系物G3BP2是一个研究较少的SG成分。在这里，我们证明了基孔肯雅病毒(CHIKV)感染诱导了含有G3BP1和G3BP2的细胞质颗粒，这些颗粒在形态、组成和行为上不同于真正的SGS。对于几种旧大陆甲型病毒，已经证明非结构蛋白3(NSP3)与G3BP相互作用，可能是为了抑制SG的形成，我们已经在CHIKV感染的细胞中证实了这种相互作用。令人惊讶的是，CHIKV也依赖G3BPs进行有效复制，因为G3BP1和G3BP2同时缺失会降低病毒RNA水平、CHIKV蛋白表达和病毒后代滴度。G3BPs与CHIKV nsP2和NSP3共定位于胞浆内，但未与nsP1、nsp4或dsRNA共定位。此外，在富含CHIKV复制/转录复合体的细胞部分中不能检测到G3BPs，这表明它们不直接参与CHIKV RNA的合成。G3BP的耗尽不会影响病毒进入、传入基因组的翻译或非结构多蛋白加工，但会导致负链(从而也是正链)RNA水平严重降低。这表明G3BP在从翻译到基因组扩增的转换中发挥了作用，尽管它们的确切作用机制仍有待探索。重要的基孔肯雅病毒(CHIKV)导致严重的多发性关节炎，自2004年重新出现以来，已影响到数百万人。缺乏经批准的疫苗或治疗选择，以及加勒比地区持续爆发的疫情，突显了更好地了解CHIKV复制的重要性。应激颗粒(SGS)是对包括病毒感染在内的各种应激反应所形成的细胞质蛋白-mRNA聚集体。RNA结合蛋白G3BP1和G3BP2是SG的重要组成部分。SG的形成和由此产生的翻译抑制通常被认为是一种抗病毒反应，许多病毒操纵或阻止这一过程。在感染后期，我们和其他人在含有G3BP1和G3BP2的细胞质颗粒中观察到CHIKV非结构蛋白3。这些病毒诱导的病灶不同于真正的SGS，似乎不代表复制复合体。令人惊讶的是，我们发现G3BP1和G3BP2也是有效复制CHIKV所必需的，可能是通过促进感染早期从翻译到基因组扩增的转换。
Stress granules (SGs) are protein-mRNA aggregates that are formed in response to environmental stresses, resulting in translational inhibition. SGs are generally believed to play an antiviral role and are manipulated by many viruses, including various alphaviruses. GTPase-activating protein (SH3 domain)-binding protein 1 (G3BP1) is a key component and commonly used marker of SGs. Its homolog G3BP2 is a less extensively studied SG component. Here, we demonstrate that Chikungunya virus (CHIKV) infection induces cytoplasmic G3BP1- and G3BP2-containing granules that differ from bona fide SGs in terms of morphology, composition, and behavior. For several Old World alphaviruses it has been shown that nonstructural protein 3 (nsP3) interacts with G3BPs, presumably to inhibit SG formation, and we have confirmed this interaction in CHIKV-infected cells. Surprisingly, CHIKV also relied on G3BPs for efficient replication, as simultaneous depletion of G3BP1 and G3BP2 reduced viral RNA levels, CHIKV protein expression, and viral progeny titers. The G3BPs colocalized with CHIKV nsP2 and nsP3 in cytoplasmic foci, but no colocalization with nsP1, nsP4, or dsRNA was observed. Furthermore, G3BPs could not be detected in a cellular fraction enriched for CHIKV replication/transcription complexes, suggesting that they are not directly involved in CHIKV RNA synthesis. Depletion of G3BPs did not affect viral entry, translation of incoming genomes, or nonstructural polyprotein processing but resulted in severely reduced levels of negative-stranded (and consequently also positive-stranded) RNA. This suggests a role for the G3BPs in the switch from translation to genome amplification, although the exact mechanism by which they act remains to be explored.IMPORTANCEChikungunya virus (CHIKV) causes a severe polyarthritis that has affected millions of people since its reemergence in 2004. The lack of approved vaccines or therapeutic options and the ongoing explosive outbreak in the Caribbean underline the importance of better understanding CHIKV replication. Stress granules (SGs) are cytoplasmic protein-mRNA aggregates formed in response to various stresses, including viral infection. The RNA-binding proteins G3BP1 and G3BP2 are essential SG components. SG formation and the resulting translational inhibition are generally considered an antiviral response, and many viruses manipulate or block this process. Late in infection, we and others have observed CHIKV nonstructural protein 3 in cytoplasmic G3BP1- and G3BP2-containing granules. These virally induced foci differed from true SGs and did not appear to represent replication complexes. Surprisingly, we found that G3BP1 and G3BP2 were also needed for efficient CHIKV replication, likely by facilitating the switch from translation to genome amplification early in infection.