SARS-CoV reverse genetics: expression of small ORFs

SARS-CoV 反向遗传学：小 ORF 的表达

• 批准号: 7028256
• 负责人: Susan R Weiss
• 金额: $ 23.22万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-03-15 至 2008-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7028256
• 关键词: Coronaviridae; SARS virus; antiserum; biotechnology; emerging infectious disease; gene expression; laboratory rabbit; open reading frames; plasmids; protein structure function; recombinant virus; severe acute respiratory syndrome; site directed mutagenesis; tissue /cell culture; virulence; virus genetics; virus protein; virus replication

DESCRIPTION (provided by applicant): Within the last year a new coronavirus (SARS-CoV) has been identified as the etiological agent of severe acute respiratory syndrome (SARS). Given the high mortality, the lack of effective treatment and the uncertainties of when the disease might return and at what frequency, a SARS vaccine and the development of antivirals are of high priority. In order to develop these strategies, we need to understand the basic biology of replication as well as the function of individual viral genes in the viral life cycle. All coronaviruses contain the structural proteins, spike (S), nucleocapsid (N), membrane (M) and small envelope (E). In addition, each group of coronaviruses expresses proteins encoded in small ORFs, unique to that group of viruses. These are, in some cases, nonessential for replication, but may have important accessory roles. Thus, our goals here include establishing a reverse genetics system for SARS-CoV and selecting viruses to be used for more detailed studies of viral replication and pathogenesis. The specific aims of this project are: 1) to establish a reverse genetics system for SARS-CoV, using targeted recombination and 2) to identify the gene products encoded in the small ORFs, unique to SARS-CoV, and to probe the role of these proteins in replication and, in the long term, in pathogenesis. The reverse genetics system will provide the basis for long-term studies on the structure/function analysis of viral structural proteins as well as the small proteins encoded in the small ORFs unique to SARS-CoV. We have a long-term interest in the replication and pathogenesis of coronaviruses. Much of the work we propose here is based on the knowledge and expertise we have accumulated in working with the murine coronavirus, mouse hepatitis virus (MHV). The targeted recombination system has greatly expanded our ability to understand replication and pathogenesis of MHV and we expect it will do the same for SARS-CoV.

描述(由申请人提供)： 在过去的一年里，一种新的冠状病毒(SARS-CoV)被确定为严重急性呼吸综合征(SARS)的病原体。鉴于死亡率高，缺乏有效的治疗，以及疾病何时复发和以何种频率复发的不确定性，SARS疫苗和抗病毒药物的开发是高度优先的。为了开发这些策略，我们需要了解复制的基本生物学以及单个病毒基因在病毒生命周期中的功能。所有冠状病毒都含有结构蛋白：刺突蛋白(S)、核衣壳蛋白(N)、膜蛋白(M)和小包膜蛋白(E)。此外，每组冠状病毒都表达在小ORF中编码的蛋白质，这是该组病毒所独有的。在某些情况下，它们对复制不是必需的，但可能具有重要的辅助作用。因此，我们在这里的目标包括建立SARS冠状病毒的反向遗传学系统，并选择用于更详细的病毒复制和发病机制研究的病毒。该项目的具体目标是：1)建立SARS-CoV的反向遗传学系统，利用靶向重组；2)鉴定SARS-CoV特有的小ORF中编码的基因产物，并探讨这些蛋白在复制和长期致病中的作用。 反向遗传学系统将为病毒结构蛋白以及SARS-CoV特有的小ORF中编码的小蛋白的结构/功能分析的长期研究提供基础。我们对冠状病毒的复制和发病机制有着长期的兴趣。我们在这里提出的许多工作都是基于我们在处理小鼠冠状病毒、小鼠肝炎病毒(MHV)方面积累的知识和专业技能。靶向重组系统极大地扩展了我们了解MHV复制和致病机制的能力，我们预计它将对SARS冠状病毒起到同样的作用。