Exploring the dynamics of nsp1 and RNA interaction in SARS-CoV with undergraduate researchers

与本科生研究人员一起探索 SARS-CoV 中 nsp1 和 RNA 相互作用的动态

• 批准号: 10730676
• 负责人: Anita Nag
• 金额: $ 40.65万
• 依托单位: UNIVERSITY OF SOUTH CAROLINA
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-06-03 至 2026-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10730676
• 关键词: 2019-nCoV; Amino Acids; Antiviral Agents; Antiviral Therapy; Binding; Binding Proteins; Binding Sites; Biochemical; C-terminal; Career Choice; Cells; Charge; Complex; Computer Analysis; Computer Models; Computers; Coronavirus; Ensure; Future; G3BP1 gene; Gene Expression; Goals; Heat shock proteins; Hepatitis C virus; Immunologist; Knowledge; Life; Maps; Mediating; Mentors; Messenger RNA; Middle East Respiratory Syndrome Coronavirus; Modeling; Mutation; Nonstructural Protein; Pathway interactions; Poly A; Polyadenylation; Process; Protein Biosynthesis; Proteins; Proteomics; Publishing; RNA; RNA Binding; RNA Sequences; RNA Stability; RNA analysis; Research; Research Personnel; Ribosomes; Role; SARS coronavirus; Science; Scientist; Specificity; Structural Models; Structure; Training; Translating; Translational Repression; Translations; Viral; Viral Proteins; cricket paralysis virus; design; experience; flexibility; inhibitor; innovation; mRNA Stability; mRNA Transcript Degradation; mRNA Translation; multidisciplinary; stem; stress granule; undergraduate student; viral RNA; zoonotic coronavirus

Project Summary Viral host shutoff proteins selectively hijack cellular machinery to aid viral propagation. In SARS coronaviruses (SARS-CoV-1 and SARS-CoV-2), nonstructural protein 1 (nsp1) serves as the host shutoff factor by dampening host gene expression through ribosomal stalling on host mRNAs followed by their cleavage and decay. However, how nsp1 selectively targets host mRNAs over the viral RNA remains an unsolved puzzle. The long-term goal of this research is to study the steps of nsp1-mediated host shutoff to clearly understand how it specifically selects host mRNAs but not capped and polyadenylated viral RNA that resembles host mRNAs. The rationale for this project is based on the observation that the deletion of stem-loop 1 (SL1) from the viral RNA leader sequence eliminates its ability to escape host shutoff. A critical factor in differentiating viral RNA from host mRNA lies in the interaction between nsp1 and SL1. Based on our previously published results that nsp1 associates with stress granule proteins and disengages G3BP1 protein from the stress granule, we propose to examine the protein and RNA composition of stress granules to determine if viral RNA is protected from decay because of its specific localization connected to the interaction between nsp1 and viral RNA. In Aim 1, we propose to conduct an in-depth study of the interaction between nsp1 and SL1 that will map the sequence responsible for binding, characterize the proteomic profile of the host proteins bound to SL1, and analyze the effect of nsp1 on host mRNA translation and stability. In Aim 2, we propose to examine the disassembly of stress granules in the presence of nsp1 and study its effect on selective cleavage and decay of host mRNA. This multidisciplinary collaborative research will engage undergraduate students to be trained in a highly transformative experience with a team of biochemists, computer scientists, and immunologists to explore a current and relevant topic in biomedical sciences. Overall, our research will connect nsp1’s ability to bind viral RNA sequence to RNA stability and localization and will allow us to examine the mechanism that leads to host mRNA cleavage and decay. Finally, this project will identify interactors of nsp1 and pave the way for designing anti-viral therapeutics.

项目摘要 病毒宿主关闭蛋白有选择地劫持细胞机械，以帮助病毒传播。在SARS冠状病毒中 （SARS-COV-1和SARS-COV-2），非结构蛋白1（NSP1）作为宿主关闭因子 通过核糖体在宿主mRNA上衰减的宿主基因表达，然后进行裂解和 衰变。但是，NSP1如何在病毒RNA上选择性地靶向宿主mRNA仍然是一个未解决的拼图。 这项研究的长期目标是研究NSP1介导的宿主关闭的步骤，以清楚地了解 它如何特异性选择宿主mRNA，但不限制和类似于宿主的聚腺苷酸化病毒RNA mrnas。该项目的基本原理是基于以下观察结果： 病毒RNA领导者序列消除了其逃脱宿主关闭的能力。区分病毒的关键因素 来自宿主mRNA的RNA在于NSP1和SL1之间的相互作用。根据我们先前发表的结果 NSP1与应激颗粒蛋白相关并脱离应激颗粒的G3BP1蛋白，我们 提出检查应激颗粒的蛋白质和RNA组成，以确定是否保护病毒RNA 由于其特定的定位与NSP1与病毒RNA之间的相互作用有关。在 AIM 1，我们建议对NSP1和SL1之间的相互作用进行深入研究，以绘制 负责结合的序列，表征与SL1结合的宿主蛋白的蛋白质组学分布，并且 分析NSP1对宿主mRNA翻译和稳定性的影响。在AIM 2中，我们建议检查 在存在NSP1的情况下应力颗粒的拆卸，并研究其对选择性裂解和衰变的影响 宿主mRNA。这项多学科合作研究将吸引本科生 与生物化学家，计算机科学家和免疫学家团队的高度变革性经验 探索生物医学科学中当前和相关的主题。总体而言，我们的研究将连接NSP1的能力 将病毒RNA序列结合到RNA稳定性和定位，将使我们能够检查以下机制 导致托管mRNA裂解和衰减。最后，该项目将识别NSP1的交互者，并为方式铺平道路 用于设计抗病毒疗法。