Flavivirus NS3-mediated Innate Immune Escape

黄病毒 NS3 介导的先天免疫逃逸

• 批准号: 9219701
• 负责人: Michaela Ulrike Gack
• 金额: $ 52.72万
• 依托单位: UNIVERSITY OF CHICAGO
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-09-23 至 2021-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9219701
• 关键词: Adaptor Signaling Protein; Amino Acids; Antiviral Agents; Antiviral Response; Architecture; Attenuated; Attenuated Live Virus Vaccine; Binding; Biochemical; Biological; Cell Culture System; Cells; Complex; Congenital Abnormality; Culicidae; Cytosol; Data; Dengue; Dengue Virus; Disease; Disease Outcome; Encephalitis; FDA approved; Fever; Flavivirus; Flavivirus Infections; Foundations; Genetic; Growth; Host Defense; Human; Immune; Immune response; Immune system; Immunity; Immunology; In Vitro; Individual; Infection; Interferon Type I; Interferon-beta; Interferons; Laboratories; Mediating; Meningitis; Microcephaly; Mitochondria; Molecular; Morbidity - disease rate; Natural Immunity; Nature; Outcome; Paralysed; Pathogenesis; Physiological; Pregnant Women; Proteins; Proteolysis; Recombinants; Role; Series; Severity of illness; Shock; Signal Pathway; Signal Transduction; Syndrome; T cell response; T-Cell Activation; Therapeutic Intervention; Vaccine Design; Vaccines; Viral; Viral Hemorrhagic Fevers; Viral Pathogenesis; Virulence; Virus; Virus Diseases; Virus Replication; West Nile virus; Work; Zika Virus; adaptive immunity; base; cell type; cytokine; defense response; design; global health; in utero; in vivo; insight; member; mortality; mouse model; mutant; novel; novel therapeutics; novel vaccines; pathogen; protein transport; recombinant virus; response; sensor; viral RNA; virus host interaction; virus pathogenesis

PROJECT SUMMARY Flaviviruses that are transmitted by mosquitoes, including dengue (DV) and West Nile (WNV) viruses as well as the recently emerging Zika virus (ZIKV), represent a significant global health concern. Despite the high morbidity and mortality resulting from infection by these viral pathogens, there are currently no FDA-approved therapies. Hence, there is a pressing need to understand better the pathogenesis of these viruses to aid the design of vaccines and antivirals. Disease severity and pathogenesis of viral infections in humans depend on many factors, including pre-existing immunity, strain virulence, host genetics and virus-host interactions. Among the virus-host interactions that modulate pathogenesis, virus-mediated suppression of innate immune signaling pathways has a critical role. However, the precise mechanisms by which flaviviruses evade host innate immunity are not well characterized. The proposed study builds on a recent discovery by the Gack laboratory that the NS3 protein of DV interacts with the mitochondrial-targeting trafficking protein 14-3-3ε to block the cytosol-to- mitochondria translocation of the viral RNA sensor RIG-I, thereby suppressing antiviral signaling and type I interferon (IFN) induction. We have identified the precise motif in DV NS3 for 14-3-3ε interaction, a four-amino-acid phosphomimetic 64RxEP67 motif, and also generated a recombinant mutant DV. This recombinant DV, encoding a 14-3-3ε-binding-deficient mutant NS3 protein, is growth-attenuated compared to wild-type DV in cells with intact innate host defense and elicits robust innate immune and T cell responses in vitro. WNV and ZIKV NS3 proteins encode a similar phosphomimetic motif, 64RLDP67, and our preliminary results show that WNV NS3 also targets 14-3-3ε to block RIG-I-mediated innate immunity. Using molecular, biochemical and cell biological approaches combined with infection studies with recombinant NS3 mutant viruses, we will define in precise detail how DV and WNV NS3 inhibits the host IFN response. This study also will yield insight into the mechanism(s) by which ZIKV NS3 protein modulates IFN-mediated host defense responses (Aim 1). Finally, we will determine the physiological relevance of the 14-3-3ε-NS3 interaction for viral pathogenesis and escape from host innate and adaptive immunity using in vitro cell culture systems and mouse models of WNV infection (Aim 2). Our studies will provide a molecular understanding of the immune escape mechanisms of DV, WNV and ZIKV, which may guide the rational design of new vaccines and antivirals.

项目摘要 蚊子传播的黄病毒，包括登革热（DV）和西尼罗河（WNV） 病毒以及最近新兴的寨卡病毒（ZIKV）代表了重要的全球健康 忧虑。尽管这些病毒感染引起的发病率和死亡率很高 病原体，目前没有FDA批准的疗法。因此，有迫切需要 更好地了解这些病毒的发病机理，以帮助设计疫苗和抗病毒药。 人类病毒感染的疾病严重程度和发病机理取决于许多因素， 包括预先存在的免疫力，菌株病毒，宿主遗传学和病毒宿主相互作用。 在调节发病机理的病毒宿主相互作用中，病毒介导的抑制 先天免疫信号通路具有关键作用。但是，确切的机制 哪种黄病毒逃避宿主先天免疫的特征没有很好地表征。 拟议的研究是基于Gack实验室最近发现的，即NS3蛋白 DV与线粒体靶向运输蛋白14-3-3ε相互作用，以阻止细胞质至 病毒RNA传感器RIG-I的线粒体易位，从而抑制抗病毒信号传导 和类型I干扰素（IFN）诱导。我们已经确定了14-3-3ε的DV NS3中的精确基序 相互作用，一种四氨基酸磷酸化的64rxep67基序，也产生了 重组突变体DV。该重组DV，编码14-3-3ε结合的缺陷突变体 NS3蛋白，与具有完整宿主的细胞中的野生型DV相比，生长稳定 防御和在体外引起稳健的先天免疫和T细胞反应。 WNV和ZIKV NS3 蛋白质编码类似的磷光基序64RLDP67，我们的初步结果显示 WNV NS3还靶向14-3-3ε，以阻止RIG-I介导的先天免疫。 使用分子，生化和细胞生物学方法与感染研究结合 使用重组NS3突变病毒，我们将精确地定义DV和WNV NS3 抑制宿主IFN响应。这项研究还将深入了解该机制 ZIKV NS3蛋白调节IFN介导的宿主防御反应（AIM 1）。最后，我们会的 确定14-3-3ε-NS3相互作用在病毒发病机理和 使用体外细胞培养系统和小鼠逃脱宿主先天和适应性免疫学 WNV感染的模型（AIM 2）。我们的研究将为分子理解 家庭暴力，WNV和ZIKV的免疫逃生机制，可以指导合理设计 新的疫苗和抗病毒药。