Research Project 3: Role of Posttranslational Protein Modifications in the Pathogenesis of Ebola Virus Disease

研究项目3：翻译后蛋白修饰在埃博拉病毒疾病发病机制中的作用

• 批准号: 10394322
• 负责人: Ricardo Rajsbaum
• 金额: $ 36.12万
• 依托单位: UNIVERSITY OF TEXAS MED BR GALVESTON
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-04-15 至 2026-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10394322
• 关键词: Address; Affect; Africa; Animal Model; Antibody Therapy; Antiviral Response; Bioinformatics; Biological Assay; Cells; Cessation of life; Co-Immunoprecipitations; Collaborations; Complex; Data; Dendritic Cells; Deubiquitination; Development; Disease; Ebola Hemorrhagic Fever; Ebola virus; Endothelial Cells; Enzymes; Epidemic; Equilibrium; Funding; Gene Expression; Genetic Transcription; Goals; Hepatocyte; Human; Immune; Immune response; Immune signaling; Immunological Models; In Vitro; Infection; Inflammation; Inflammatory; Inflammatory Response; Injury; Interferons; Knock-out; Knowledge; Laboratories; Ligase; Link; Liver; Mass Spectrum Analysis; Mediating; Modeling; Modification; Molecular; Molecular Target; Monoclonal Antibodies; Outcome; Outcome Study; Pathogenesis; Pathogenicity; Pathway interactions; Pattern; Phosphoric Monoester Hydrolases; Phosphorylation; Phosphotransferases; Physiological; Post-Translational Protein Processing; Post-Translational Regulation; Primary Infection; Process; Production; Protein Dephosphorylation; Proteomics; Regulation; Research Project Grants; Role; Sampling; Signal Pathway; Signal Transduction; System; T-Lymphocyte; Ubiquitin; Ubiquitination; Validation; Vascular Endothelium; Viral; Viral Proteins; Virus Diseases; Virus Replication; Work; antiviral drug development; cell type; cytokine; cytokine release syndrome; in vivo; insight; interferon antagonist; kidney cell; kidney vascular structure; knock-down; monocyte; mutant; nonhuman primate; novel strategies; overexpression; pathogenic virus; proteogenomics; response; transforming virus; ubiquitin ligase; virus core

RESEARCH PROJECT 3 (RP3): PROJECT SUMMARY/ABSTRACT The epidemic of Ebola virus (EBOV) in Africa in 2013-2016 and its reappearance in subsequent years have reached alarming numbers of infections and deaths, prompting extensive efforts to develop antiviral strategies; however, none have thus far been licensed for use in humans. Pathogenesis of EBOV infection derives from a combination of the viral suppression of the host's antiviral protective immune responses and induction of hyper- inflammation, including a “cytokine storm”. Cytokine production and inflammatory responses require activation of multiple signaling pathways that are extensively regulated at the posttranslational level to maintain a balance between efficient antiviral responses and excessive inflammation. Posttranslational modifications (PTMs) of pro- teins serve as molecular regulatory mechanisms, a process which requires the activity of specific enzymes (i.e., kinases, ubiquitin ligases, etc.). Thus, the expression and activation patterns of these enzymes in specific cell types determines the complex regulation that balances an effective immune physiological response. The goal of Research Project 3 (RP3) is to elucidate how EBOV transforms the PTM cell-type specific landscape leading to immune dysregulation and disease, with the long-term goal of applying this knowledge in the development of effective approaches to treat the disease caused by EBOV. Our preliminary data indicate that interferon antagonist domains of EBOV proteins can hijack the host ubiquitin system to increase virus replication while causing cell-type specific dysregulation of signaling pathways that control hyperinflammation. However, it is not known which PTM enzymes or what changes in host protein mod- ifications drive immune dysregulation and disease. Our hypothesis is that EBOV targets specific enzymes that regulate PTM in a cell-type specific manner resulting in uncontrolled inflammation, while also blocking protective innate immune signaling. We will use novel approaches to assess how EBOV infection dysregulates immune signaling by targeting unconventional PTMs, in vitro and in vivo. With information obtained from the BSL-4 Core (Core B), Proteogenomics Core (Core C), and Bioinformatics and Modeling Core (Core D), and from RP1 and RP2 we will build a complete model of inflammatory pathways activated during EBOV infection. In Aim 1 we will characterize the PTM landscape during EBOV infection in primary human immune and nonimmune cells, and in Aim 2 in cells isolated from infected nonhuman primates, using mass spectrometry analysis and validation assays. In Aim 3, we will validate PTM enzymes and their modifications, and elucidate cell-type specific PTM- mediated mechanisms that result in dysregulated immune responses to EBOV. The outcome of these studies is significant because it will provide fundamental knowledge on the function of specific PTMs during infection and uncover molecular targets to treat pathogenic inflammation. Our work on posttranslational regulation together with information derived from changes in the transcriptional (RP1) and post- transcriptional (RP2) landscapes will provide a comprehensive model of immune dysregulation.

研究项目3（RP 3）：项目总结/摘要 2013-2016年埃博拉病毒（EBOV）在非洲的流行及其在随后几年的重现， 感染和死亡人数达到惊人的水平，促使人们做出广泛努力，制定抗病毒战略; 然而，迄今为止，还没有一种被许可用于人类。EBOV感染的发病机制源自一种 病毒抑制宿主的抗病毒保护性免疫应答和诱导超- 炎症，包括“细胞因子风暴”。细胞因子的产生和炎症反应需要激活 在翻译后水平广泛调节以维持平衡的多种信号通路 有效的抗病毒反应和过度炎症之间的联系翻译后修饰（PTMs） 蛋白作为分子调节机制，这是一个需要特定酶活性的过程（即， 激酶、泛素连接酶等）。因此，这些酶在特定细胞中的表达和激活模式 类型决定了平衡有效免疫生理反应的复杂调节。的目标 研究项目3（RP 3）是阐明EBOV如何转化PTM细胞类型特异性景观， 免疫失调和疾病，长期目标是将这些知识应用于发展 治疗EBOV引起的疾病的有效方法。 我们的初步数据表明，EBOV蛋白的干扰素拮抗剂结构域可以劫持宿主泛素， 系统，以增加病毒复制，同时引起细胞类型特异性的信号传导途径的失调， 控制过度炎症。然而，目前还不知道哪些PTM酶或宿主蛋白质中的哪些变化是调制的。 影响导致免疫失调和疾病。我们的假设是EBOV靶向特定的酶， 以细胞类型特异性方式调节PTM，导致不受控制的炎症，同时也阻断保护性 先天免疫信号我们将使用新的方法来评估EBOV感染如何失调免疫系统， 通过靶向非常规PTM，在体外和体内进行信号传导。从BSL-4核心获得的信息 (Core B）、蛋白质基因组学核心（核心C）和生物信息学和建模核心（核心D），以及RP 1 和RP 2，我们将建立一个完整的模型，炎症途径激活EBOV感染。在目标1中， 将表征EBOV感染期间在原代人免疫和非免疫细胞中的PTM景观， 在从感染的非人灵长类动物分离的细胞中，使用质谱分析和验证， 分析。在目标3中，我们将验证PTM酶及其修饰，并阐明细胞类型特异性PTM- 介导的机制导致对EBOV的免疫应答失调。 这些研究的结果是重要的，因为它将提供有关功能的基本知识， 在感染期间特异性PTM和发现分子靶点以治疗致病性炎症。我们的工作 翻译后调节与来自转录（RP 1）和翻译后 转录（RP 2）景观将提供免疫失调的综合模型。