Entry and pathogenesis of two human coronaviruses

两种人类冠状病毒的进入和发病机制

基本信息

批准号：
8055141
负责人：
Thomas Miller Gallagher
金额：
$ 33.86万
依托单位：
UNIVERSITY OF IOWA
依托单位国家：
美国
项目类别：
财政年份：
2011
资助国家：
美国
起止时间：
2011-07-19 至 2016-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8055141
关键词：
Address Affinity Animal Model Animals Antiviral Agents Attenuated Vaccines Avidity Bacterial Artificial Chromosomes Binding Biological Biological Assay Bronchiolitis Cathepsin L Cell Communication Cell Line Cells Collaborations Complementary DNA Coronavirus Cytokine Gene Cytokine Receptors Data Dependence Disease Elements Engineering Epidemic Epithelium Event Gene Expression Genetic Growth HIV Human Immune response In Vitro Infection Inflammatory Instruction Integrins Ligands Ligation Lung Lung Inflammation Lung diseases Maps Measures Mediating Membrane Membrane Fusion Modeling Mus Mutation Pathogenesis Pathogenicity Pathology Pathway interactions Peptide Hydrolases Peptidyl-Dipeptidase A Process Protein S Proteins Receptor Cell Recombinants Renin-Angiotensin System Reporter Reporting Rodent Role Severe Acute Respiratory Syndrome Severity of illness Signal Transduction Signaling Molecule Staging Technology Temperature Vaccines Variant Vertebral column Viral Virulence Virus Virus Activation Virus Receptors Virus-like particle cofactor cytokine expectation human coronavirus human disease improved in vivo in vivo Model injured lung injury member mutant novel particle receptor recombinant virus research study response tissue culture transmission process vaccine development virus tropism

项目摘要

Human coronaviruses (H-CoVs) cause respiratory diseases ranging from self-limiting bronchiolitis to severe acute respiratory syndrome (SARS). This proposal aims to compare NL63-CoV, which causes mild disease, with SARS-CoV, focusing on distinctions between the viral spike (S) proteins that mediate the entry of these H-CoVs into host cells. The SARS and NL63 S proteins are divergent yet bind to the same host cell receptor, angiotensin converting enzyme 2 (ACE2). Subsequent to ACE2 interaction, these two S proteins may differentially direct virus entry and uncoating, in ways that may correlate with virus pathogenicity differences. Aim 1 addresses this question of distinct H-CoV entry pathways and extends preliminary data suggesting that integrins are used during NL63 S-mediated entry. Experiments will determine whether particular integrins are used by NL63 and other H-CoVs in tissue culture models of lung epithelia, and genetic approaches will be used to identify novel additional coreceptors participating in H-CoV entry. ACE2 operates as an ectopeptidase and as a signaling molecule to regulate the renin-angiotensin system and the inflammatory state of the injured lung. S proteins may disturb these central ACE2 functions. Aim 2 experiments will construct S fragments, recombinant SARS / NL63 S proteins, and HCoV-like particles bearing complete S proteins, for use as ACE2 ligands. S:ACE2 interactions leading to proinflammatory host cell responses will be identified, and the host responses to virus entry events operating subsequent to the ACE2 interaction will also be discerned. These studies relating S : ACE2 interactions with inflammation and lung pathology comprise a key collaboration with PPG component 3. The broader biological relevance of H-CoV entry cofactors and proinflammatory responses require the use of complete infectious viruses and host animal models. Robust in vitro and in vivo models to evaluate NL63-CoV are lacking. In Aim 3, we plan to select NL63-CoV variants for vigorous growth in culture, using cells overproducing ACE2 and integrins, with expectations for informative S mutants. Variants will be evaluated for growth in mice to establish models for the human disease. We will also construct recombinant NL63 / SARS chimeric viruses in collaboration with PPG component 4 to further correlate S proteins with virus growth, coreceptor usage and host cell responsiveness.

人类冠状病毒（H-CoV）会引起从自限性细支气管炎到严重急性呼吸综合征（SARS）等呼吸系统疾病。该提案旨在将引起轻度疾病的 NL63-CoV 与 SARS-CoV 进行比较，重点关注介导这些 H-CoV 进入宿主细胞的病毒刺突 (S) 蛋白之间的区别。 SARS 和 NL63 S 蛋白是不同的，但与相同的宿主细胞受体血管紧张素转换酶 2 (ACE2) 结合。 ACE2 相互作用后，这两种 S 蛋白可能以不同的方式指导病毒进入和脱壳，其方式可能与病毒致病性差异相关。目标 1 解决了不同 H-CoV 进入途径的问题，并扩展了初步数据，表明在 NL63 S 介导的进入过程中使用了整合素。实验将确定 NL63 和其他 H-CoV 在肺上皮组织培养模型中是否使用特定的整合素，并且将使用遗传方法来识别参与 H-CoV 进入的新的额外辅助受体。 ACE2 作为外肽酶和信号分子来调节肾素-血管紧张素系统和受损肺部的炎症状态。 S 蛋白可能会干扰这些中央 ACE2 功能。 Aim 2 实验将构建 S 片段、重组 SARS / NL63 S 蛋白和带有完整 S 蛋白的 HCoV 样颗粒，用作 ACE2 配体。 S:ACE2相互作用导致促炎性宿主细胞反应将被识别，并且宿主对ACE2相互作用之后的病毒进入事件的反应也将被辨别。这些有关 S : ACE2 与炎症和肺部病理学相互作用的研究包括与 PPG 成分 3 的关键合作。 H-CoV 进入辅助因子和促炎症反应的更广泛的生物学相关性需要使用完整的感染性病毒和宿主动物模型。缺乏评估 NL63-CoV 的稳健的体外和体内模型。在目标 3 中，我们计划使用过量产生 ACE2 和整合素的细胞，选择能够在培养物中旺盛生长的 NL63-CoV 变体，并期望获得信息丰富的 S 突变体。将评估变体在小鼠体内的生长情况，以建立人类疾病模型。我们还将与 PPG 组分 4 合作构建重组 NL63 / SARS 嵌合病毒，以进一步将 S 蛋白与病毒生长、辅助受体使用和宿主细胞反应性相关联。