Interaction of Sindbis Virus with Cellular Processes

描述(申请人提供)：Togaviridae家族的甲型病毒属包含一些重要的人类和动物病原体，广泛分布于除南极地区以外的所有大陆。一些甲型病毒，例如委内瑞拉(VEEV)、东部(EEEV)和西部(WeEV)马脑炎病毒，会导致严重的人类疾病。其他病毒，如辛德比斯(SINV)和塞姆利基森林(SFV)病毒，对人类的致病性较低，然而，它们采用类似的复制策略，并用于研究病毒RNA合成的基本问题。在上一次赠款期间，我们在了解SINV基因组复制方面取得了一些重要发现，包括：i)揭示了RNA启动子元件的结构和功能；ii)展示了旧世界甲型病毒特有的nsP2和新世界特有的衣壳蛋白在病毒与宿主细胞相互作用中的关键作用；在脊椎动物起源的细胞中，转录阻断和细胞病变效应的发生；以及iii)鉴定了SINV特异蛋白复合体的病毒和细胞成分。最重要的是，我们最近的研究表明，SINV在脊椎动物细胞中的复制导致两种类型的含有NSP3的蛋白质复合体的形成。根据我们的数据，我们假设第一种类型的NSP3特异性复合体最初形成于细胞膜上，并在内吞作用之后与内吞体样的含膜细胞器保持联系。这些复合体在病毒特异性RNA合成中作为复制复合体(RC)。第二类复合体与细胞骨架有关，具有类似的病毒成分，但细胞成分不同，通常在细胞应激颗粒(SGS)中检测到。后一种复合体要么作为SG形成的竞争者，要么服务于病毒蛋白的优先合成。拟议的研究计划集中于两个广泛的具体目标：i)研究SINV复制过程中病毒特异性蛋白复合体的形成，ii)分析细胞和病毒蛋白在SINV复制中的功能。与公共卫生相关：甲型病毒包括一组广泛分布的人类和动物病原体；其中一些会导致高度衰弱的疾病，并在美国构成严重的公共卫生威胁。这项建议的目标是了解细胞环境在病毒感染中的作用。识别对病毒生长至关重要的细胞蛋白将导致新的抗病毒治疗策略的开发。

DESCRIPTION (provided by applicant): The Alphavirus genus in the Togaviridae family contains a number of significant human and animal pathogens that are widely distributed on all continents, excluding the Antarctic regions. Some of the alphaviruses, e.g., the Venezuelan (VEEV), eastern (EEEV) and western (WEEV) equine encephalitis viruses, cause severe human disease. Others, such as Sindbis (SINV) and Semliki Forest (SFV) viruses, are less pathogenic for humans, however, they employ similar replication strategy and are used for studying fundamental issues of the viral RNA synthesis. In the previous grant period, we made a number of important findings toward understanding the SINV genome replication that include i) uncovering the structure and functioning of the RNA promoter elements; ii) demonstration of the critical roles of the Old World alphavirus-specific nsP2 and the New World-specific capsid protein in virus-host cell interactions, in the development of transcriptional shutoff and cytopathic effect in the cells of vertebrate origin, and iii) identification of the viral and cellular components of the SINV-specific protein complexes. Most importantly, our recent studies demonstrated that SINV replication in the cells of vertebrate origin leads to formation of two types of nsP3-containing protein complexes. Based on our data, we hypothesize that the first type of nsP3-specific complexes is initially formed on the cellular membrane and, following endocytosis, remains associated with endosome-like, membrane-containing organelles. These complexes serve as replication complexes (RCs) in virus-specific RNA synthesis. The second type of complexes is associated with the cytoskeleton and has similar viral, but different cellular components, which are normally detected in cellular stress granules (SGs). The latter complexes either function as competitors for SG formation and/or serve in the preferential synthesis of viral proteins. The proposed research plan is focused on two broad Specific Aims: i) to study virus-specific protein complexes formation during SINV replication, and ii) to analyze functioning of cellular and viral proteins in SINV replication. PUBLIC HEALTH RELEVANCE: Alphaviruses comprise a group of widely distributed human and animal pathogens; some of them induce highly debilitating diseases and represent a serious public health threat in the US. The goal of this proposal is to understand the role of the cellular environment in viral infection. Identification of the cellular proteins that are essential for virus growth will lead to development of new antiviral therapeutic strategies.