Phenotypic analyses of HSV-1 miRNAs: H1, H6, & H8

项目摘要 大多数人群潜伏感染1型单纯疱疹病毒（HSV-1），但也有一些人感染HSV-1。 没有清除潜伏病毒的有效疗法。在潜伏期，病毒抑制裂解基因的表达， 表达选择的几种非编码RNA，包括潜伏相关转录本（LAT）和病毒miRNA。的 大多数HSV-1miRNA在病毒感染中的生物学功能在很大程度上是未知的。推定 HSV-1miRNA H1、H6和H8的靶包括病毒和宿主转录物。然而，这些证据 机制主要依赖于瞬时表达测定。重组蛋白的严格表型分析 在与HSV-1生物学相关的细胞中缺乏单个miRNA的病毒，在病毒感染的情况下， 将阐明这些病毒miRNA相互作用在HSV-1发病机制中的生物学意义。 假设：将发现HSV-1 miRNAs H1、H6和H8促进HSV-1裂解、潜伏和/或复发 通过微调病毒和宿主转录物的翻译来防止感染。由于miRNAs抑制翻译， 缺乏靶向1）裂解基因的miRNA，将增加毒力或2）宿主免疫反应，将 降低毒性。此外，我可能会发现这些miRNAs有多个宿主或病毒靶点， 协同地或拮抗地调节病毒潜伏期的动态状态。 目的1：miR-H1、H6和H8在细胞培养物中急性感染期间的表型和机制分析。 目的2：确定miR-H1、H6和H8在小鼠急性感染过程中对发病机制的影响 足垫感染模型。 目的3：研究miR-H1、H6和H8在小鼠背根神经节潜伏期和再激活中的作用。

Project Summary The majority of the human population is latently infected by herpes simplex virus type-1 (HSV-1), yet there are no effective therapies for clearing latent virus. During latency, the virus curbs lytic gene expression and only express a select few noncoding RNAs, including the latency associated transcript (LAT) and viral miRNAs. The biological functions of most HSV-1 miRNAs, in the context of viral infection, are largely unknown. Putative targets for HSV-1 miRNAs H1, H6, and H8 include viral and host transcripts. However, evidence for these mechanisms rely primarily on transient expression assays. Rigorous phenotypic analyses of recombinant viruses lacking individual miRNAs in cells relevant to HSV-1 biology and in vivo, in the context of viral infection, will elucidate the biological significance of these viral miRNA interactions in HSV-1 pathogenesis. Hypothesis: HSV-1 miRNAs H1, H6, & H8 will be found to facilitate HSV-1 lytic, latent, and/or recrudescent infections by fine-tuning the translation of viral and host transcripts. Since miRNAs repress translation, a virus lacking a miRNA that targets 1) a lytic gene, will increase virulence or 2) the host immune response, will decrease virulence. In addition, I may find that these miRNAs have multiple host or viral targets, that work synergistically or antagonistically to regulate the dynamic state of viral latency. Aim 1: Phenotypic & mechanistic analyses of miR-H1, H6, and H8 during acute infection in cell culture. Aim 2: Determine the effects of miR-H1, H6, and H8 on pathogenesis during acute infection in the mouse footpad infection model. Aim 3: Characterize the roles of miR-H1, H6, and H8 in latency and reactivation in murine dorsal root ganglia.