Nonstructural roles of capsid proteins in virus replication

衣壳蛋白在病毒复制中的非结构作用

• 批准号: 7532260
• 负责人: TERYL K FREY
• 金额: $ 18.06万
• 依托单位: GEORGIA STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-06-10 至 2010-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7532260
• 关键词: Antiviral Agents; Autoimmune Diseases; Binding; Binding Sites; Capsid; Capsid Proteins; Complement; Complex; Congenital Abnormality; Exhibits; Family; Financial compensation; Flavivirus; Genes; Genome; Genomics; Glycoproteins; Goals; Human; Infection; Maps; Mediating; Modeling; Molecular Chaperones; Molecular Conformation; Mutation; Penetration; Play; Protein C; Proteins; Public Health; RNA; RNA Binding; RNA Sequences; RNA Viruses; RNA chemical synthesis; RNA replication; RNA-Directed RNA Polymerase; Rate; Recruitment Activity; Replicon; Reporter Genes; Research; Role; Rubella virus; Rubivirus; Site; Structural Genes; Structural Protein; Structure; Testing; Togaviridae; Translations; Untranslated Regions; Viral; Virion; Virus; Virus Replication; West Nile virus; cis acting element; drug development; human TFRC protein; insight; mutant; novel; pathogen; protein function; replicase; tyrosine kinase ABL1; viral RNA

DESCRIPTION (provided by applicant): Rubella virus (RUBV) is a major human pathogen that causes birth defects and is associated with autoimmune disease. RUBV is a simple, plus-strand RNA virus (rubivirus genus, Togavirus family) that has five genes: two replicase genes and three structural protein genes that comprise the virus particle. We have made the unexpected discovery that one of the virion proteins, the capsid protein (C), is involved in an early step in virus RNA synthesis. Using replicons, constructs in which the structural genes have been replaced with a reporter gene, we have found that C enhances the rate of wild type replicon RNA accumulation and rescues deletions in three regions of the genome, at the 5' and 3' ends and internally within the gene of one of the replicase proteins. Our overarching hypothesis is that C executes or compensates for functions normally associated with the structure of the genomic RNA. In the proposed research, four Specific Aims will be pursued. In the first Specific Aim, we will test the hypothesis that C-mediated rescue of the internal deletion is through compensation for RNA structure rather than compensation for protein function. In the second Specific Aim, we will test the hypothesis that C binding to the genomic RNA is necessary for enhancement of wt replicon RNA replication and mutant rescue. In the third Specific Aim we will test whether C stabilizes replicon RNAs or serves as an RNA chaperone, both potential mechanisms by which C executes wt enhancement or mutant rescue. Finally, we have found that C associates with one of the replicase proteins, a novel finding among plus-strand RNA viruses, suggesting a model for involvement of C in viral RNA synthesis by which the C recruits both the viral RNA and the replicase proteins, facilitating formation of the replication complex and/or initiation of RNA synthesis. To test this model, the binding sites on C for the replicase protein will be mapped and mutants that fail to bind the replicase will be identified and used to see if they abrogate wt replicon enhancement and mutant rescue. We have evidence that the capsid protein of West Nile virus (Flavivirus family) enhances the replication of a West Nile virus replicon, indicating that C involvement in viral RNA synthesis that we are studying with RUBV may represent a paradigm among plus-strand RNA virus families. The function of capsid protein in viral RNA synthesis appears to at a step between penetration and unencapsidation of the virion and establishment of viral RNA replication complexes and thus the proposed research should yield insight into this poorly defined part of the virus replication cycle. PUBLIC HEALTH RELEVANCE: The proposed research will specifically explore how the capsid protein of rubella virus, a protein that makes up part of the virus particle, is also involved in viral RNA synthesis. Capsid proteins of other medically important viruses may play a similar role. A major hypothesis to be tested in the proposed research is that capsid protein involvement in viral RNA synthesis is through direct interaction of the capsid and replicase proteins and this interaction is an attractive target for antiviral drug development.

描述（由申请人提供）：风疹病毒（RUBV）是引起先天缺陷并与自身免疫性疾病有关的主要人类病原体。 RUBV是一种简单的，链的RNA病毒（Rubivirus Genus，Togavirus家族），具有五个基因：两个复制酶基因和三个组成病毒颗粒的结构蛋白基因。我们已经意外发现的是，一种病毒蛋白（C）的一种病毒蛋白参与了病毒RNA合成的早期步骤。使用复制子，结构基因已被报告基因代替的构建体，我们发现C增强了野生型复制子RNA积累的速率，并在基因组的三个区域，在5'和3'末端，内部以及内部内部的三个区域的缺失率在其中一种重复酶蛋白的基因内。我们的总体假设是C执行或补偿与基因组RNA结构通常相关的功能。在拟议的研究中，将追求四个具体目标。在第一个特定目的中，我们将检验以下假设：C介导的内部缺失是通过补偿RNA结构而不是补偿蛋白质功能。在第二个特定目的中，我们将检验以下假设：C与基因组RNA的结合对于增强WT复制子RNA复制和突变挽救是必要的。在第三个特定目的中，我们将测试C是稳定复制子RNA还是用作RNA伴侣，这两种潜在的机制是C通过其执行WT增强或突变救援的潜在机制。最后，我们发现C与一种复制酶蛋白之一的结合，这是链链RNA病毒中的一种新发现，这表明C参与病毒RNA合成的模型，C c在该模型中C募集了病毒RNA和复制酶蛋白，并促进了复制复制复合物和/或/或RNA综合启动的复制形成。为了测试该模型，将映射复制酶蛋白的C上的结合位点，并且将识别未结合复制酶的突变体，并用于查看它们是否消除了WT复制子增强和突变体救援。我们有证据表明，西尼罗河病毒（Flavivivirus家族）的衣壳蛋白增强了西尼罗河病毒复制子的复制，这表明我们正在使用RUBV研究的病毒RNA合成的参与可能代表加大史的RNA病毒家族中的范式。衣菌蛋白在病毒RNA合成中的功能似乎是在渗透与病毒RNA复制复制复合物的渗透与不兼容的一步之间，因此拟议的研究应洞悉病毒复制循环的这一定义较差的部分。公共卫生相关性：拟议的研究将专门探讨风疹病毒的衣壳蛋白（一种构成病毒颗粒一部分的蛋白质）如何参与病毒RNA合成。其他重要病毒的衣壳蛋白可能起着相似的作用。在拟议的研究中要检验的一个主要假设是，衣壳蛋白参与病毒RNA合成是通过直接相互作用的衣壳和复制酶蛋白的直接相互作用，而这种相互作用是抗病毒药物开发的有吸引力的靶标。