Interplay between coronaviruses and nonsense-mediated mRNA decay pathway

冠状病毒(CoV)携带一个大的、单链的、阳性的RNA基因组，导致各种 人类和家畜中的疾病。人类冠状病毒(HCoV)通常感染呼吸道和 引起一系列症状，从轻微的，如普通感冒，到更严重的呼吸道疾病 如严重急性呼吸综合征(SARS)和中东呼吸综合征(MERS)，由两个 高致病性HCoV、SARS冠状病毒和MERS冠状病毒。SARS冠状病毒在2002年引发了一场全球疫情- 2003年，导致8,000多例病例，死亡率约为10%，而MERS-CoV在#年出现 沙特阿拉伯于2012年推出，并已传播到中东、北非、欧洲、 和东亚。HCoV是对公共健康的主要威胁，并有可能导致重大的 对经济的负面影响。目前，还没有获得批准的针对HCoV的疫苗和治疗剂。 制定有效的冠状病毒控制措施需要对病毒有全面的了解 基因表达策略和宿主-冠状病毒相互作用。过多的研究集中在调查冠状病毒 生物学，并对我们理解冠状病毒复制机制做出了重大贡献，包括 病毒RNA元件以及与病毒有关的病毒蛋白的结构-功能分析 复制和组装。然而，我们对转录后调控的认识仍然存在空白。 病毒基因表达的研究，因为只有有限数量的研究涉及这一领域的冠状病毒研究。 特别是，对顺式作用的病毒rna元件和反式作用的宿主和病毒因子知之甚少。 调节冠状病毒mRNA转录稳定性的基因。病毒学中一个新兴的研究领域是理解 病毒与宿主mRNA监测通路之间的相互作用，以防止产生/积累 不想要的基因产品。我们已经证明，冠状病毒mRNAs是无意义介导的 信使核糖核酸衰变(NMD)通路--宿主信使核糖核酸监测通路之一，病毒N蛋白保护 NMD的冠状病毒mRNAs。我们的数据表明N介导的NMD抑制对于有效病毒的重要性 复制。本应用将通过测试NMD途径和冠状病毒之间的相互作用来研究 以下假设：NMD的主要协调者UPF1与冠状病毒mRNAs的3‘非编码区结合，具有 特定的基序，经历磷酸化，并招募内切酶SMG6，导致内切核溶解 核糖核酸切割；N与NMD靶标的3‘UTRs结合，并阻止NMD因子(S)访问这些UTRs 靶和/或N与NMD因子(S)相互作用并将其隔离在NMD途径之外；以及冠状病毒 突变株对NMD的易感性增加，不能像亲本病毒那样有效复制。这个 从这些研究中获得的数据将提供对冠状病毒mRNAs和N介导的NMD的机械性见解 NMD途径的抑制，并将揭示通过以下途径减弱CoV的新策略的可行性 增加对NMD途径的敏感性。

Coronaviruses (CoVs), which carry a large, single-stranded, positive-sense RNA genome, cause a variety of diseases in humans and domestic animals. Human CoVs (HCoVs) usually infect the respiratory tract and cause a range of symptoms varying from mild, such as the common cold, to more serious respiratory illnesses like severe acute respiratory syndrome (SARS) and Middle East respiratory syndrome (MERS), caused by two highly pathogenic HCoVs, SARS-CoV and MERS-CoV. SARS-CoV caused a worldwide epidemic in 2002- 2003, resulting in more than 8,000 cases with an approximate mortality of 10%, while MERS-CoV emerged in Saudi Arabia in 2012 and has been disseminated into other countries in the Middle East, North Africa, Europe, and East Asia. HCoVs represent a major threat to public health and have the potential to cause a significant negative economic impact. Currently, there are no approved vaccines and therapeutic agents against HCoVs. The development of effective control measures against CoVs requires a comprehensive understanding of viral gene expression strategies and host-CoV interactions. A plethora of studies have focused on investigating CoV biology and have significantly contributed to our understanding of CoV replication mechanisms, including the structure-function analyses of viral RNA elements as well as the viral proteins that are involved in viral replication and assembly. However, there are still gaps in our knowledge of the post-transcriptional regulation of viral gene expression, as only a limited number of studies have addressed this area of CoV research. Particularly, very little is known about the cis-acting viral RNA elements and trans-acting host and viral factors that regulate CoV mRNA transcript stability. One newly emerging research area in virology is understanding interactions between viruses and host mRNA surveillance pathways that prevent generation/accumulation of unwanted gene products. We have demonstrated that CoV mRNAs are the targets of the nonsense-mediated mRNA decay (NMD) pathway, one of the host mRNA surveillance pathways, and that viral N protein protects CoV mRNAs from NMD. Our data suggest the importance of N-mediated NMD suppression for efficient virus replication. The present application will study the interplay between the NMD pathway and CoVs by testing the following hypotheses: UPF1, the principal orchestrator of NMD, binds to the 3’ UTR of CoV mRNAs having specific motifs, undergoes phosphorylation, and recruits SMG6, an endonuclease, leading to endonucleolytic RNA cleavage; N binds to the 3’ UTRs of NMD targets and prevents an NMD factor(s) from accessing these targets and/or N interacts with an NMD factor(s) and sequesters it away from the NMD pathway; and CoV mutants having an increased susceptibility to NMD, cannot replicate as efficiently as the parental viruses. The data obtained from these studies will provide mechanistic insights into NMD of CoV mRNAs and N-mediated NMD pathway suppression, and will reveal the feasibility of novel strategies for attenuating CoVs through increased susceptibility to the NMD pathway.