Biology of Animal Nuclear Mononegaviruses

博尔纳病病毒（Borna disease virus，BDV）是研究病毒在中枢神经系统持续存在的重要模型 中枢神经系统（CNS）是一个与健康相关的主题，因为病毒可导致人类神经系统疾病。 BDV是一个新的单负病毒科的原型， 可能会照亮MNV生物学的新方面。我们的长期目标是阐明 控制BDV的复制和基因表达是了解BDV分子基础的前提 持续存在于中枢神经系统和病毒引起的脑细胞功能紊乱。实验重点 该建议是在功能上表征有助于控制以下的病毒和细胞蛋白： BDV复制和基因表达程序。我们的具体目标是： 1. BDV预测多肽p8.3、p8.4和p165的检测和功能表征。 BDV利用可变RNA剪接和转录终止信号产生三种mRNA 它可以编码三种新的预测病毒多肽，这些多肽在病毒生命中的表达和功能 循环没有被研究过。我们将：1）确定这些多肽是否在BDV期间产生 感染2)表征它们在BDV感染期间的亚细胞和时间表达模式。第三章 检测它们在控制BDV RNA复制和基因表达中的作用。 2.检测BDV p10和Np 38多肽在BDV生命周期中的作用。我们将确定： 1)p10抑制病毒RNA合成的机制。2)p10如何调节其抑制性 BDV感染细胞中的活性以及p10是否是病毒繁殖所需的。3)的机制 由此Np 38有助于最佳的BDV聚合酶活性，以及Np 38是否是BDV聚合酶活性所需的。 感染性病毒的产生及其繁殖。 3. BDV聚合酶复合物-宿主细胞蛋白的生物化学和功能鉴定 交互.我们将研究BDV聚合酶复合物-hCLE的机制和后果 我们发现的互动。我们将使用串联亲和纯化（TAP）和质谱法 鉴定BDV聚合酶复合物相互作用配偶体的完整阵列的方法。作为 作为一种补充方法，我们将使用基于siRNA的筛选来鉴定BDV的细胞抑制剂 聚合酶复合体通过蛋白质组学方法和基于RNAi的筛选鉴定候选分子 将使用生物化学和遗传学方法进行验证。经验证病毒细胞的生物学相关性 蛋白质相互作用将在BDV MG拯救系统和BDV感染的背景下进行检查。 使用生物化学、遗传学和细胞生物学方法的组合。

Borna disease virus (BDV) is an important model for the study of viral persistence in the central nervous system (CNS), a subject relevant to health because viruses can contribute to human neurologic disease. BDV is the prototype of a new family of mononegaviruses, and information obtained from studies on BDV might illuminate novel aspects of MNV biology. Our long-term goal is to elucidate the mechanisms that control BDV replication and gene expression as a prerequisite to understand the molecular bases of BDV persistence in the CNS and virally induced disturbances in brain cell functions. The experimental focus of this proposal is to functionally characterize viral and cellular proteins that contribute to the control of replication and gene expression program of BDV. Our specific aims are: 1. Detection and functional characterization of BDV predicted polypeptides p8.3, p8.4 and p165. BDV uses alternative RNA splicing and transcription termination signals to generate three mRNA species that can code for three novel predicted viral polypeptides whose expression and functions in the virus life cycle have not been studied. We will: 1) Determine whether these polypeptides are produced during BDV infection. 2) Characterize their sub-cellular and temporal expression patterns during BDV infection. 3) Examine their roles in the control of BDV RNA replication and gene expression. 2. Examine the roles of BDV p10 and Np38 polypeptides in the life cycle of BDV. We will determine: 1) The mechanisms whereby p10 inhibits viral RNA synthesis. 2) How p10 modulates its inhibitory activity in BDV-infected cells and whether p10 is required for virus propagation. 3) The mechanisms whereby Np38 contributes to optimal BDV polymerase activity, and whether Np38 is required for the generation of infectious virus and its propagation. 3. Biochemical and functional characterization of the BDV polymerase complex-host cell protein interactions. We will investigate the mechanisms and consequences of BDV polymerase complex-hCLE interaction we have identified. We will use Tandem Affinity Purification (TAP) and Mass Spectrometry approaches to identify the full array of BDV polymerase complex interacting partners. As a complementary approach we will use siRNA-based screenings to identify cellular inhibitors of the BDV polymerase complex. Candidate molecules identified by proteomic approaches and RNAi-based screens will be validated using biochemical and genetic approaches. The biological relevance of validated viruscell protein interactions will be examined in the context of the BDV MG rescue system and BDV-infected cells using a combination of biochemical, genetic and cell biology approaches.