Evaluation of SARS-CoV 2'O Methyltransferase Mutants

SARS-CoV 2O 甲基转移酶突变体的评估

• 批准号: 8525763
• 负责人: VINEET D MENACHERY
• 金额: $ 5.22万
• 依托单位: UNIV OF NORTH CAROLINA CHAPEL HILL
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-01-01 至 2015-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8525763
• 关键词: Acute; Adult Respiratory Distress Syndrome; Age; Alveolar; Amino Acid Motifs; Animal Model; Antiviral Agents; Attenuated; Attenuated Vaccines; Body Weight decreased; Coronavirus; Coronavirus Infections; DNA Viruses; Data; Diffuse; Disease Outbreaks; Evaluation; Event; Family; Family member; Genes; Guanine; Human; Human Virus; Immune; Immune response; In Vitro; Infection; Interferon Type I; Interferons; Kinetics; Laboratories; Lung diseases; Mammalian Cell; Methylation; Methyltransferase; Modeling; Morbidity - disease rate; Mus; Nucleosides; Pathogenesis; Pathway interactions; Pharmaceutical Preparations; Phase; Play; Process; Proteins; Publishing; RNA; RNA Caps; RNA Viruses; Research; Role; S-Adenosylmethionine; SH2D3A gene; Severe Acute Respiratory Syndrome; Severities; Signal Transduction; Testing; Therapeutic; Time; Vaccination; Vaccine Design; Vaccines; Vertebral column; Viral; Viral Genes; Virulence; Virus; Virus Diseases; Virus Replication; Work; aged; attenuation; atypical pneumonia; base; gene function; in vivo; insight; interest; mortality; mutant; public health relevance; recombinant virus; response; scaffold; small hairpin RNA; success; therapeutic target; transmission process; treatment strategy; viral DNA

DESCRIPTION (provided by applicant): Coronaviruses (CoV) are important emerging human viruses, with multiple cross-species transmission events identified over the past 200 years. Human infections with severe acute respiratory syndrome coronavirus (SARS-CoV), the first newly emerged virus of the 21st century, results in acute and organizing phase diffuse alveolar damage, atypical pneumonia, and acute respiratory distress syndrome (ARDS), leading to mortality rates of 10-50 percent, dependent on age. A major component in the success of the SARS-CoV is its ability to manipulate and subvert the host immune response. Studies by our laboratory and others have revealed numerous genes that antagonize the type I interferon (IFN) response including NSP1, ORF3b, and ORF6. In addition, several groups have generated SARS-CoV deletion mutants and demonstrated various levels of attenuation, possibly due to increased type I IFN sensitivity. A central hypothesis in this application is that highly conserved viral gene functions that target type I IFN responses can serve to provide a universal platform for the rational design of vaccine and drug therapeutics affording rapid responses in outbreak settings. Recently, research has focused on viral components involved in type I RNA capping utilized by SARS, other CoV, and many RNA and DNA viruses. In an ordered process, several CoV proteins contribute to the capping process including NSP13 (RTPase), NSP14 (N7-guanine methylation), and NSP10 (scaffold). However, interest has focused on NSP16, a S-adenosylmethionine (SAM) dependent nucleoside 2'-O-methyltransferase (2'-O-MTase) and its critical role in subverting the type I IFN response. Recent works have implicated 2'O- methylation in distinguishing between self and non-self RNA by MDA-5, a RIG-I like recognition molecule, and the IFIT family of interferon stimulated genes (ISGs). These results suggest that 2'-O-MTases like NSP16 play a critical role in immune antagonism during viral infection. Based on these recent findings, we sought to evaluate the impact of 2'O-methylation on SARS-CoV replication and pathogenesis by generating mutants lacking NSP16 2'O-MTase activity. We hypothesize that deltaNSP16 mutant viruses will be exquisitely sensitive to and attenuated in the presence of type I IFN both in vitro and in vivo. We anticipate the absence of type I IFN signaling or specific ISGs including MDA5 and IFIT family members will restore replication and possibly virulence. Due to broad conservation of this activity across RNA and DNA virus families, targeting the 2'O methylation pathways with either vaccine or drug strategies may provide unique, broadly applicable treatment options that can protect against current and emerging viruses.

描述(申请人提供)：冠状病毒(CoV)是重要的新兴人类病毒，在过去200年中发现了多次跨物种传播事件。人类感染严重急性呼吸综合征冠状病毒(SARS-CoV)是21世纪第一个新出现的病毒，可导致急性和组织期弥漫性肺泡损伤、非典型肺炎和急性呼吸窘迫综合征(ARDS)，死亡率为10%-50%，取决于年龄。SARS冠状病毒成功的一个主要组成部分是它操纵和颠覆宿主免疫反应的能力。我们实验室和其他实验室的研究揭示了许多拮抗I型干扰素(干扰素)反应的基因，包括NSP1、ORF3b和ORF6。此外，有几个组产生了SARS-CoV缺失突变体，并表现出不同程度的衰减，可能是由于I型干扰素敏感性增加所致。这个应用中的一个中心假设是高度保守的 针对I型干扰素反应的病毒基因功能可为合理设计疫苗和药物疗法提供一个通用平台，在暴发环境中提供快速反应。最近，研究集中在SARS、其他冠状病毒以及许多RNA和DNA病毒所利用的I型RNA帽所涉及的病毒成分上。在一个有序的过程中，几个冠状病毒蛋白参与了封顶过程，包括NSP13(RTPase)、NSP14(N7-鸟氨酸甲基化)和NSP10(支架)。然而，人们对NSP16的兴趣主要集中在NSP16，它是一种依赖S腺苷蛋氨酸的核苷2‘-O-甲基转移酶(2’-O-MTase)，及其在逆转I型干扰素应答中的关键作用。最近的研究表明，2‘O-甲基化可以通过类似RIG-I的识别分子MDA-5和干扰素刺激基因(ISGs)的IFIT家族来区分自体和异体RNA。这些结果表明，像NSP16这样的2‘-O-MTase在病毒感染的免疫拮抗中起着关键作用。基于这些最新发现，我们试图通过产生缺乏NSP16 2‘O-MTase活性的突变体来评估2’O-甲基化对SARS-CoV复制和致病的影响。我们推测，DeltaNSP16突变病毒在体外和体内都对I型干扰素非常敏感和减弱。我们预计，缺少I型干扰素信号或特定的ISG，包括MDA5和IFIT家族成员，将恢复复制，并可能恢复毒力。由于这种活性在RNA和DNA病毒家族中具有广泛的保守性，以2‘O甲基化途径为靶点的疫苗或药物策略可能提供独特的、广泛适用的治疗选择，可以预防当前和新出现的病毒。