Interactions of Ebola virus glycoproteins with host cells

描述(申请人提供)：埃博拉病毒是最致命的人类病原体之一，扎伊尔亚型的死亡率接近90%。他们也是潜在的生物恐怖主义代理人。埃博拉病毒感染会导致人类患上一种严重的出血性疾病，目前既没有治疗方法，也没有保护性疫苗。出于这些原因，埃博拉病毒被美国国立卫生研究院列为“A类优先病原体”。埃博拉糖蛋白表达在病毒和感染细胞表面，促进病毒进入宿主细胞，并以其他重要方式与宿主细胞相互作用。这项研究计划的总体目标是彻底描述埃博拉糖蛋白与宿主细胞的相互作用，以确定潜在的治疗干预目标，并了解埃博拉复制的基本生物学。这项研究的重点是识别和分析与埃博拉GP相互作用的宿主因素，以及表征埃博拉糖蛋白如何影响宿主细胞。埃博拉病毒的表面糖蛋白(GP)在病毒进入宿主的过程中起着关键作用，并被认为负责附着、受体识别和膜融合。确定进入埃博拉病毒所需的宿主因素对于理解埃博拉病毒的复制和发病机制至关重要。内体组织蛋白是目前已知的唯一进入埃博拉病毒所需的细胞因子。具体目标1利用一系列旨在确定重要的细胞埃博拉进入因素的遗传和生化实验。除了在病毒进入期间的功能外，埃博拉GP似乎还以其他重要方式与宿主细胞相互作用。Tetherin/BST-2是最近被描述的一种内在的细胞抗病毒因子，它对许多被包裹的病毒具有活性，并阻止成熟病毒颗粒的释放。我们最近的PNAS手稿表明，埃博拉GP拮抗细胞抗病毒因子Tetherin，促进埃博拉颗粒的释放。具体目标2的研究将检查埃博拉GP抑制这种细胞抗病毒因子的要求，并将分析GP对Tetherin的影响。除了在进入和病毒释放中发挥作用外，埃博拉gp在宿主细胞中的表达还导致表面蛋白的明显下调，导致细胞黏附丧失，MHC和其他几种宿主蛋白的表面水平降低。初步数据表明，GP中的粘蛋白区域起到了立体屏障的作用，阻止了表面的可及性。测试这一模型，明确GP介导的表面蛋白下调的机制，并分析下调对免疫识别的影响，是本提案的具体目标3。 公共卫生相关性：埃博拉病毒感染与高死亡率有关，对埃博拉感染的恐惧使该病毒成为生物恐怖问题。目前还没有治疗埃博拉病毒的有效方法。实现这一建议的目标将大大加强我们对埃博拉病毒糖蛋白(GP)及其与宿主相互作用的了解，这些知识可用于设计阻断GP功能或干扰埃博拉GP-受体相互作用的治疗方法。

DESCRIPTION (provided by applicant): Ebola viruses are among the most lethal human pathogens with mortality rates approaching 90% for the Zaire subtype. They are also a potential bioterrorism agent. Ebola virus infection causes a severe hemorrhagic disease in humans for which there are no therapeutic treatments nor protective vaccines currently available. For these reasons, Ebola virus is classified as a "category A priority pathogen" by NIH. Expressed on the virus and infected cell surface, the Ebola glycoproteins facilitate entry of the virus into host cells and also interact with the host cells in other important ways. The overall goal of this research program is to thoroughly characterize Ebola glycoprotein-host cell interactions to identify potential targets for therapeutic intervention and also to inform about the basic biology of Ebola replication. This research proposal focuses upon identification and analysis of host factors that interact in with Ebola GP and characterization of how Ebola glycoproteins affect the host cell. The surface glycoprotein (GP) of Ebola plays a key role in entry of the virus into the host and is believed to be responsible for attachment, receptor recognition, and membrane fusion. Definition of the host factors that are required for Ebola entry is essential for comprehending Ebola virus replication and pathogenesis. Endosomal cathepsins are the only currently known cellular factor required for Ebola entry. Specific aim 1 utilizes a series of genetic and biochemical experiments designed to identify important cellular Ebola entry factors. In addition to its function during viral entry, Ebola GP appears to interact with host cells in other important ways. Tetherin/BST-2 is a recently described intrinsic cellular antiviral factor that is active against a number of enveloped viruses and blocks release of mature viral particles. Our recent PNAS manuscript demonstrates that Ebola GP antagonizes the cellular anti-viral factor Tetherin to promote release of Ebola particles. Studies in Specific Aim 2 will examine the requirements for Ebola GP inhibition of this cellular anti-viral factor and will analyze the effects of GP upon Tetherin. In addition to roles in entry and virus release, Ebola GP expression in host cells causes apparent dramatic down modulation of surface proteins leading to loss of cellular adhesion and diminished surface levels of MHC and several other host proteins. Preliminary data suggests a model in which the mucin region in GP acts as a steric shield to block surface accessibility. Testing this model, defining the mechanism of GP-mediated surface protein down-regulation and analyzing the consequences of down- modulation on immune recognition comprise Specific Aim 3 of this proposal. PUBLIC HEALTH RELEVANCE: Ebola viral infection is associated with a high mortality rate and the fear of Ebola infection makes this virus a bioterror concern. There are no effective treatments for Ebola infection. Achieving the Aims of this proposal will significantly enhance our understanding of the Ebola virus glycoproteins (GP) and their interactions with the host and this knowledge can be applied to the design of therapeutics that block GP functions or interfere with Ebola GP-receptor interactions.