Characterization of the SARSCoV frameshift signal

SARSCoV 移码信号的表征

• 批准号: 7884348
• 负责人: Jonathan D Dinman
• 金额: $ 37.52万
• 依托单位: UNIV OF MARYLAND, COLLEGE PARK
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-07-01 至 2011-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7884348
• 关键词: Affect; Animals; Antiviral Agents; Base Sequence; Biochemistry; Biological Assay; Biological Models; Budgets; Cell Line; Complementary DNA; Complex; Coronavirus; Dimerization; Epithelial Cells; Family; Genes; Germ; Hepatitis; Human; Laboratories; Link; Maps; Messenger RNA; Methodology; Molecular; Molecular Genetics; Molecular Models; Monitor; Mus; Mutation; Nucleic acid sequencing; Oryctolagus cuniculus; Phylogenetic Analysis; Positioning Attribute; Property; RNA; Recommendation; Regulation; Regulatory Element; Relative (related person); Research; Resolution; Reticulocytes; Ribosomal Frameshifting; Ribosomes; Saccharomyces; Sequence Analysis; Severe Acute Respiratory Syndrome; Signal Transduction; Structure; Study Section; System; Therapeutic; Totivirus; Viral; Viral Pathogenesis; Viral Proteins; Virus; Wheat; Yeasts; base; comparative; genetic analysis; in vivo; interest; molecular modeling; mouse model; mutant; nuclease; programs; research study; small molecule; stem; tissue culture; vaccine development; virology

DESCRIPTION (provided by applicant): Viruses utilize programmed ribosomal frameshifting (PRF) to post-transcriptionally regulate the expression of multiple genes that are encoded on monocistronic viral mRNAs. Studies in Retro- and Totiviruses have shown that maintaining correct PRF efficiencies is critical for virus propagation, thus identifying this mechanism as a potential target for antiviral therapeutics. PRF has previously been shown to be utilized by the Coronaviruses (CoV), and primary sequence analysis of the rapidly emerging SARS-CoV, the etiological agent of Severe Acute Respiratory Syndrome (SARS), reveals the presence of a putative PRF signal that is predicted to shift elongating ribosomes by one base in the -1 or 5' direction (-1 PRF). Initial comparative, structural and functional analysis of this sequence in our laboratory suggests that the -1 PRF signal of SARS-CoV (and possibly of the entire Coronavirus family) utilizes a complex, heretofore unknown mRNA pseudoknot structure that contains three stem loops as opposed to the usual two-stem loop variety. Further, our findings suggest that the function of the third stem-loop may be to regulate -1 PRF efficiency, and hence the expression of viral proteins. Thus, small molecules that interact with this structure may potentially have antiviral properties. The broad aim of the proposed research is to characterize important features of the SARS-CoV -1 PRF signal using a combination of phylogenetic, molecular, structural, and viral assays. Specifically, we will 1) characterize how changes in the mRNA pseudoknots of the SARS and Mouse Hepatitis CoV's affect frameshifting efficiency using an in vivo human epithelial cell based assay system, 2) determine the effects of changes in frameshift efficiency on propagation of the SARS-CoV using an infectious cDNA clone in tissue culture and in a mouse model system, and 3) structurally characterize the SARS-CoV -1 PRF signal using nuclease mapping, high-resolution NMR and calorimetric methodologies.

描述（由申请人提供）：病毒利用编程的核糖体框架（PRF）在转录后调节在单科传播病毒mRNA上编码的多个基因的表达。在复古病毒和小说中的研究表明，保持正确的PRF效率对于病毒繁殖至关重要，因此将这种机制确定为抗病毒疗法的潜在靶标。先前已证明PRF被冠状病毒（COV）使用，以及对严重急性呼吸综合征（SARS）的病因的快速新兴SARS-COV的主要序列分析，揭示了存在的假定PRF信号的存在，该信号预计将通过-1或5'方向（-1或5'prf）移动核糖体。该序列中该序列的最初比较，结构和功能分析表明，SARS-COV的-1 PRF信号（可能是整个冠状病毒家族的）使用了一个复杂的，迄今未知的mRNA伪诺特结构，其中包含三个与通常的两肺发套件相反的三个茎。此外，我们的发现表明，第三个茎环的功能可能是调节-1 prf效率，从而调节病毒蛋白的表达。因此，与该结构相互作用的小分子可能具有抗病毒特性。拟议研究的广泛目的是使用系统发育，分子，结构和病毒测定的组合来表征SARS -COV -1 PRF信号的重要特征。 Specifically, we will 1) characterize how changes in the mRNA pseudoknots of the SARS and Mouse Hepatitis CoV's affect frameshifting efficiency using an in vivo human epithelial cell based assay system, 2) determine the effects of changes in frameshift efficiency on propagation of the SARS-CoV using an infectious cDNA clone in tissue culture and in a mouse model system, and 3) structurally characterize the SARS-CoV -1 PRF信号使用核酸酶映射，高分辨率NMR和量热方法。

项目成果

RNA dimerization plays a role in ribosomal frameshifting of the SARS coronavirus.

• DOI: 10.1093/nar/gks1361
• 发表时间: 2013-02-01
• 期刊: Nucleic acids research
• 影响因子: 14.9
• 作者: Ishimaru D;Plant EP;Sims AC;Yount BL Jr;Roth BM;Eldho NV;Pérez-Alvarado GC;Armbruster DW;Baric RS;Dinman JD;Taylor DR;Hennig M
• 通讯作者: Hennig M