Research Project 2: Role of Posttranscriptional Regulatory Networks in the Pathogenesis of Ebola Virus Disease

研究项目2(RP2)：项目摘要/摘要 这一总体计划项目旨在解决有关埃博拉病毒(EBOV)的重大知识差距：如何 感染会导致致病性免疫反应，而不是保护性免疫反应。在本项目中，重点是EBOV如何 感染改变了关键免疫细胞类型和严重免疫细胞的转录后基因调控 感染(例如，肝细胞)。关于EBOV感染如何影响宿主基因表达的了解有限 在转录后水平上。的确，尽管基因转录后调控的重要性 对多种致病的人类病毒的表达是显而易见的，这种水平的调控一直没有得到充分的研究 丝状病毒。 研究项目2(RP2)的中心假设是特定细胞类型和特定基因的转录后 EBOV疾病的反应导致免疫细胞的不适应表型，最终导致 “免疫性瘫痪”和过度炎症。为了解决这一假设，我们将描述 原代人类免疫和非免疫细胞感染EBOV期间的转录后图谱。我们会 破译EBOV感染对宿主RNA加工和翻译的影响。大规模并行测序 由蛋白质组学核心(核心C)实现的方法将用于定义RNA处理 事件、RNA修饰和RNA翻译效率在幼稚和感染细胞中的转录范围内。 重要的是，BSL-4核心(核心B)中的相同实验产生的样本将被所有三个人使用 因此，我们计划将我们的数据与RP1和RP3的数据进行比较。数据分析、集成和 建模将与生物信息学和建模核心(核心D)合作进行。这些 实验将对EBOV感染对相关细胞中细胞基因表达的影响产生新的见解 类型。我们还将描述非人类体内感染EBOV期间的转录后情况 灵长类动物。使用这些实验的数据，我们将使用核心D构建一个全面的模型 调节不同免疫细胞类型中EBOV感染的调节宿主途径。此型号将识别 代表这些途径的重要节点的候选基因，因此是EBOV的潜在脆弱性。 在与Core B的合作下，该项目将验证转录后改变的基因和事件，并询问 它们是否对EBOV感染的细胞类型特异性反应具有重要的功能。 该项目的完成将揭示新的和未得到充分认识的宿主对EBOV感染的反应，以及 为该领域提供丰富的数据资源，这将导致新的假设，以解决分子基础 EBOV致病机制。此外，我们将确定关键的新宿主和/或病毒目标，以利用这些目标 破坏这种致命病毒的致病性。

RESEARCH PROJECT 2 (RP2): PROJECT SUMMARY/ABSTRACT This overall Program Project seeks to address a significant knowledge gap concerning Ebola virus (EBOV): how infection leads to a pathogenic rather than protective immune response. In this project, the focus is on how EBOV infection alters posttranscriptional gene regulation in critical immune cell types and in cells that are heavily infected (e.g., hepatocytes). There is limited understanding of how EBOV infection impacts host gene expression at the posttranscriptional level. Indeed, although the importance of posttranscriptional regulation in gene expression for multiple pathogenic human viruses is evident, this level of regulation has been understudied for filoviruses. The central hypothesis of Research Project 2 (RP2) is that cell type-specific and gene-specific posttranscriptional responses in EBOV disease contribute to the maladaptive phenotypes of immune cells, eventually leading to “immune paralysis” and hyperinflammation. To address this hypothesis, we will characterize the posttranscriptional landscape during EBOV infections of primary human immune and nonimmune cells. We will decipher the influence of EBOV infection on host RNA processing and translation. Massively parallel sequencing approaches made possible by the Proteogenomics Core (Core C) will be used to define RNA processing events, RNA modifications, and mRNA translation efficiency transcriptome-wide in naïve and infected cells. Importantly, samples generated from the same experiments in the BSL-4 Core (Core B) will be used by all three projects and thus we plan to compare our data with that of RP1 and RP3. Data analysis, integration, and modelling will be performed in collaboration with the Bioinformatics and Modeling Core (Core D). These experiments will yield new insights into the effect of EBOV infection on cellular gene expression in relevant cell types. We will also characterize the posttranscriptional landscape during EBOV infection in vivo in nonhuman primates. Using the data from these experiments, with Core D we will construct a comprehensive model of regulated host pathways that modulate EBOV infection in different immune cell types. This model will identify candidate genes that represent important nodes for these pathways and thus potential vulnerabilities for EBOV. In collaboration with Core B, this project will validate posttranscriptionally altered genes and events, and ask whether they are functionally important to cell-type specific responses to EBOV infection. Completion of this project will reveal new and underappreciated layers of host responses to EBOV infection, and provide a rich data resource for the field that will lead to new hypotheses to address the molecular basis for EBOV pathogenesis. Moreover, we will identify critical new host and/or viral targets that can be exploited to disrupt the pathogenicity of this deadly virus.