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: 10602495
• 负责人: Ricardo Rajsbaum
• 金额: $ 34.52万
• 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/10602495
• 关键词: Address; Affect; Africa; Animal Model; Antibody Therapy; Antiviral Response; Appearance; Bioinformatics; Biological Assay; Cell Separation; 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; Kidney; Knock-out; Knowledge; Laboratories; Licensing; 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; 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; knock-down; monocyte; mutant; nonhuman primate; novel strategies; overexpression; pathogenic virus; posttranscriptional; proteogenomics; response; transforming virus; ubiquitin ligase

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(报告3)：项目摘要/摘要 2013-2016年埃博拉病毒(EBOV)在非洲的流行以及随后几年的重新出现 感染和死亡人数达到令人震惊的数字，促使广泛努力制定抗病毒战略； 然而，到目前为止，还没有一种药物获得在人类身上使用的许可。EBOV感染的发病机制源于一种 病毒抑制宿主的抗病毒保护性免疫反应和诱导高... 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 在翻译后水平上受到广泛调控以维持平衡的多个信号通路 在有效的抗病毒反应和过度的炎症之间。PRO的翻译后修饰(PTM) Teins充当分子调节机制，这一过程需要特定酶的活性(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。的目标是 研究项目3(RP3)是为了阐明EBOV如何转变PTM细胞类型的特定景观领先 免疫失调和疾病，长期目标是将这些知识应用于 治疗EBOV引起的疾病的有效方法。 我们的初步数据表明，EBOV蛋白的干扰素拮抗域可以劫持宿主泛素 增加病毒复制，同时导致细胞类型特异性信号通路失调的系统， 控制过度炎症。然而，目前尚不清楚哪些PTM酶或宿主蛋白MOD-MOD发生了什么变化。 结果会导致免疫调节失调和疾病。我们的假设是EBOV以特定的酶为靶标 以细胞类型特异性的方式调节PTM，导致不受控制的炎症，同时也阻断保护 先天免疫信号。我们将使用新的方法来评估EBOV感染如何扰乱免疫 在体外和体内通过靶向非常规PTM的信号转导。使用从BSL-4核心获得的信息 (核心B)、蛋白质基因组学核心(核心C)、生物信息学和建模核心(核心D)，以及来自RP1 和RP2，我们将建立一个在EBOV感染期间激活的炎症通路的完整模型。在目标1中，我们 将描述在原代人类免疫和非免疫细胞中感染EBOV期间的PTM情况， 在AIM 2中，从受感染的非人灵长类动物中分离出细胞，使用质谱分析和验证 化验。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的PTM- 导致EBOV免疫应答失调的中介机制。 这些研究的结果是重要的，因为它将提供关于 感染过程中的特定PTM，并发现治疗致病性炎症的分子靶点。我们的工作是 翻译后调控以及从转录(RP1)和转录后的变化中获得的信息 转录(RP2)景观将提供一个全面的免疫失调模型。