Genetic dissection of the RNAi-mediated antiviral immunity in C. elegans

线虫 RNAi 介导的抗病毒免疫的基因剖析

• 批准号: 8063606
• 负责人: Shou-Wei Ding
• 金额: $ 28.39万
• 依托单位: UNIVERSITY OF CALIFORNIA RIVERSIDE
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-05-01 至 2014-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8063606
• 关键词: Alleles; Animals; Antiviral Agents; Apoptosis; C. elegans genome; Caenorhabditis elegans; Chemicals; DNA; DNA-Directed DNA Polymerase; Dengue Virus; Dissection; Double-Stranded RNA; Drosophila genus; Enhancers; Genes; Genetic; Genetic Screening; Genome; Genomics; Goals; HIV; Hepatitis C virus; Housing; Human; Human poliovirus; Immune; Immunity; Immunologic Receptors; Individual; Influenza; Invertebrates; Mediating; Methodology; Modeling; Molecular; Mutagenesis; Mutation; Natural Immunity; Nematoda; Pathway interactions; Plants; Play; Polioviruses; Process; Production; Proteins; RNA; RNA Interference; RNA Interference Pathway; RNA Viruses; RNA-Directed DNA Polymerase; RNA-Directed RNA Polymerase; Reading; Research; Resources; Role; Specificity; System; Transgenes; Viral; Virus; Virus Replication; West Nile virus; base; design; feeding; genetic analysis; genome-wide; influenzavirus; insight; mutant; novel; pathogen; public health relevance; research study; response; viral RNA

DESCRIPTION (provided by applicant): Viruses with an RNA genome replicate and evolve via mechanisms distinct from DNA genomes and are important human, animal and plant pathogens. The mammalian immune mechanisms that defend against RNA viruses are poorly characterized. The proposed research will genetically dissect the immune pathway of Caenorhabditis elegans animals triggered by Flock house virus (FHV), one of the best characterized RNA viruses. We have shown recently that FHV replication launched from an inducible transgene integrated in the C. elegans genome triggers production of virus-derived siRNAs, which guide viral RNA clearance by RNA silencing or RNA interference (RNAi). Further genetic analyses have provided evidence for a conserved RNA silencing virus immunity (RSI) pathway in C. elegans in response to either FHV or another distinct RNA virus. We have carried out pilot feeding RNAi and chemical mutagenesis screens using the C. elegans/FHV system. Our feeding RNAi screen has identified 35 genes required for RSI including drh-1, which is dispensable for RNAi induced by exogenous dsRNA and highly homologous to the recently identified mammalian immune receptor for RNA viruses. Viable RSI-defective worm mutants recovered from the chemical screen include both RNAi-defective and RNAi-sensitive mutants and none are allelic with drh-1 or another frequent occurring RNAi-defective allele. Hence, our results have shown that C. elegans RSI requires components that are shared with, and dispensable for, exogenous RNAi, as well as those homologous to mammalian viral immunity components. We plan to further characterize the mechanism of viral RNA clearance guided by viral siRNAs, the features and roles of secondary viral siRNAs, and the specificity mechanism of RSI components dispensable for exogenous RNAi in worms. Further, we will isolate additional components of the animal RSI pathway through genome-wide feeding RNAi and chemical mutagenesis screens using the methodology already established in the lab. The proposed experiments will provide the first genome-view of the viral immunity pathway in whole animals, identify novel RNAi components that cannot be obtained in previous screens using non-viral RNAi triggers, and reveal immune mechanisms against RNA viruses that facilitate understanding mammalian viral immunity. PUBLIC HEALTH RELEVANCE: The nematode C. elegans has been a powerful model for elucidating the molecular mechanisms of both RNA interference (RNAi) and programmed cell death in humans. Many RNA viruses such as influenza, hepatitis C virus, poliovirus, dengue virus and West Nile virus are important human pathogens. Thus, it is likely that the proposed studies on the RNAi-mediated viral immunity pathway of C. elegans to RNA viruses will provide mechanistic insights into mammalian viral immunity and RNAi.

描述(申请人提供)：具有RNA基因组的病毒通过不同于DNA基因组的机制进行复制和进化，是人类、动物和植物的重要病原体。哺乳动物防御RNA病毒的免疫机制还没有得到很好的描述。这项拟议的研究将从基因上剖析由鸡舍病毒(FHV)引发的秀丽线虫动物的免疫途径，FHV是最具特征的RNA病毒之一。我们最近发现，FHV复制从整合到线虫基因组中的可诱导转基因启动，触发病毒来源的siRNAs的产生，通过RNA沉默或RNA干扰(RNAi)来引导病毒RNA的清除。进一步的遗传分析已经为线虫对FHV或另一种不同的RNA病毒的保守的RNA沉默病毒免疫(RSI)途径提供了证据。我们已经使用线虫/FHV系统进行了RNAi和化学诱变筛选的中试。我们的补给RNAi筛选发现了RSI所需的35个基因，其中包括DRH-1，它是外源dsRNA诱导的RNAi所必需的，与最近发现的哺乳动物RNA病毒免疫受体高度同源。从化学筛选中恢复的活的RSI缺陷蠕虫突变体包括RNAi缺陷突变和RNAi敏感突变，没有一个与DRH-1或另一个常见的RNAi缺陷等位基因等位。因此，我们的结果表明，线虫RSI需要与外源RNAi共享和非必需的成分，以及那些与哺乳动物病毒免疫成分同源的成分。我们计划进一步研究病毒siRNAs引导病毒RNA清除的机制，次级病毒siRNAs的特征和作用，以及蠕虫中外源RNAi所必需的RSI组分的特异性机制。此外，我们将使用实验室中已经建立的方法，通过全基因组喂养RNAi和化学诱变筛选来分离动物RSI途径的其他组件。这些拟议的实验将提供整个动物中病毒免疫途径的第一个基因组视图，识别以前使用非病毒RNAi触发器无法获得的新的RNAi成分，并揭示针对RNA病毒的免疫机制，有助于理解哺乳动物的病毒免疫。 与公共卫生相关：线虫一直是阐明RNA干扰(RNAi)和人类细胞程序性死亡的分子机制的有力模型。许多RNA病毒，如流感、丙型肝炎病毒、脊髓灰质炎病毒、登革热病毒和西尼罗河病毒都是重要的人类病原体。因此，对线虫RNAi介导的病毒免疫途径的研究很有可能为哺乳动物的病毒免疫和RNAi提供机械性的见解。