Suppression of RNA Interference by Herpes Simplex Virus

单纯疱疹病毒对 RNA 干扰的抑制

• 批准号: 6984353
• 负责人: Deborah S. Parris
• 金额: $ 18.69万
• 依托单位: OHIO STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-07-01 至 2007-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6984353
• 关键词: RNA interference; RNase protection assay; Vero cells; cell line; double stranded RNA; flow cytometry; gene expression; gene induction /repression; green fluorescent proteins; host organism interaction; microRNAs; molecular cloning; plasmids; small interfering RNA; virus genetics; virus infection mechanism; virus protein

DESCRIPTION (provided by applicant): Eukaryotic cells have developed innate mechanisms to protect them from invasion by viruses and foreign nucleic acids, most of which involve silencing of gene expression. One such adaptive innate response is RNA interference (RNAi). The RNAi pathway is highly conserved in almost all eukaryotes, and is involved in a variety of phenomena that negatively impact gene expression, including RNA silencing, microRNA processing, RNA-directed DNA methylation, histone H3K9 methylation, and the establishment of heterochromatin. In plants, RNAi is an important adaptive defense which is induced by and targets viruses. Plant viruses counter this attack by producing proteins that suppress components of the pathway, and these silencing suppressors were first identified as pathogenicity determinants. It is likely that mammalian viruses must also suppress RNAi, and their recently demonstrated susceptibility to RNAi-based approaches supports this view. Viral silencing suppressors represent potent targets for the development of antiviral and vaccine therapeutics, but the presence of such suppressors among mammalian viruses is unknown. Herpes simplex virus (HSV) is effective in establishing its transcription program in cells in which it replicates productively, but viral gene transcription is largely silenced during latency. The effectiveness with which HSV avoids being silenced in permissive cells makes HSV a strong candidate for encoding a silencing suppressor. It is hypothesized that HSV encodes one or more proteins that suppress the RNAi pathway in order to productively replicate. The ability of wild-type and mutant HSV to induce RNAi will be determined by the appearance of virus-specific small interfering RNA (siRNA) using a candidate screening approach and a global approach to clone small dsRNAs from infected cells. In aim 2, a transient silencing system will be used to identify silencing suppressors encoded by HSV.

描述（由申请人提供）：真核细胞已经发展出先天机制来保护它们免受病毒和外源核酸的入侵，其中大部分涉及基因表达的沉默。其中一种适应性先天反应是 RNA 干扰 (RNAi)。 RNAi 通路在几乎所有真核生物中高度保守，并参与多种对基因表达产生负面影响的现象，包括 RNA 沉默、microRNA 加工、RNA 指导的 DNA 甲基化、组蛋白 H3K9 甲基化和异染色质的建立。在植物中，RNAi 是一种重要的适应性防御，由病毒诱导并针对病毒。植物病毒通过产生抑制该途径成分的蛋白质来对抗这种攻击，这些沉默抑制子首先被鉴定为致病性决定因素。哺乳动物病毒很可能也必须抑制 RNAi，并且它们最近表现出的对基于 RNAi 的方法的敏感性支持了这一观点。病毒沉默抑制因子代表了抗病毒和疫苗疗法开发的有效靶点，但哺乳动物病毒中是否存在此类抑制因子尚不清楚。单纯疱疹病毒（HSV）可以有效地在细胞中有效地建立其转录程序，并在其中进行高效复制，但病毒基因转录在潜伏期基本上被沉默。 HSV 在允许的细胞中避免沉默的有效性使得 HSV 成为编码沉默抑制子的有力候选者。据推测，HSV 编码一种或多种抑制 RNAi 途径的蛋白质，以便有效复制。野生型和突变型 HSV 诱导 RNAi 的能力将取决于病毒特异性小干扰 RNA (siRNA) 的出现，使用候选筛选方法和从受感染细胞克隆小 dsRNA 的全局方法。在目标 2 中，将使用瞬态沉默系统来识别 HSV 编码的沉默抑制子。