Strain Dependent Structure and Function of the Influenza NS1 Protein

流感 NS1 蛋白的菌株依赖性结构和功能

• 批准号: 10291406
• 负责人: Chad Petit
• 金额: $ 37.13万
• 依托单位: UNIVERSITY OF ALABAMA AT BIRMINGHAM
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-11-16 至 2024-10-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10291406
• 关键词: Affect; Allosteric Regulation; Amino Acids; Animals; Antiviral Agents; Antiviral Response; Automobile Driving; Binding; Cells; Cleavage And Polyadenylation Specificity Factor; Crystallization; Data; Development; Disease; Drug Design; Drug Targeting; Epidemic; Future; Genes; Genetic; Goals; Health; Human; Immune; Immune response; Impairment; Infection; Influenza; Innate Immune Response; Interferon-beta; Interferons; Investigation; Knowledge; Measures; Mission; Molecular; Morbidity - disease rate; Motion; NMR Spectroscopy; Nonstructural Protein; Pathogenicity; Play; Production; Property; Protein Dynamics; Proteins; Public Health; RNA Recognition Motif; Recombinants; Relaxation; Research; Resolution; Role; Sampling; Signal Transduction; Structural Models; Structure; Structure-Activity Relationship; Testing; Tretinoin; United States National Institutes of Health; Variant; Viral; Virulence; Virus Replication; anti-influenza; antiviral drug development; base; experimental study; influenza infection; influenza virulence; influenza virus strain; influenzavirus; inhibitor; innovation; millisecond; mortality; mutant; novel; nucleocytoplasmic transport; pandemic disease; pathogen; resistant strain; response; small molecule

Developing effective antiviral drugs requires a detailed understanding of the molecular mechanisms underlying the targeted host-pathogen interaction. Specifically, precise structural models of these interactions can provide mechanistic details with atomic resolution to assist in the efficient development of novel compounds against pathogens of significant health relevance. The influenza virus is a prime example of one such pathogen. Novel strains of the influenza virus develop annually via infection and replication in a number of animal hosts, including humans. The relative ability of these strains to cause disease, or virulence, is determined by a number of interactions between viral and cellular proteins. Although influenza non-structural protein 1 (NS1) is known to play a critical role in virulence, there is a fundamental gap in our knowledge of the genetic and structural determinants that facilitate the multiple strain-dependent functions attributed to NS1 in the host cell. It is therefore our long-term goal to understand the molecular mechanisms that underlie strain-dependent function of NS1. The objective of this application is to structurally characterize interactions with two cellular proteins (CPSF30 and RIG-I) that are important for the activation of the innate immune response. Our central hypothesis is that structural and dynamic features unique to certain NS1 variants account for the diverse array of functions attributed to NS1. The rationale that underlies the proposed research is that elucidating structure- function relationships between NS1 and its cellular interaction partners will aid in the development of antiviral drugs that target these critical interactions known to modulate virulence. Our central hypothesis will be tested by pursuing three specific aims: 1) structurally and functionally characterize the multiple interactions between NS1 and RIG-I, 2) determine the role of microsecond-millisecond (µs-ms) motions in proper function of the NS1 effector domain (NS1ED), and 3) determine the mechanism of action by which JJ3297 suppresses influenza replication. In Aim 1, NMR spectroscopy and mutant recombinant influenza viruses will be used to structurally and functionally characterize the multiple interactions between NS1 and RIG-I. In Aim 2, relaxation dispersion experiments will be used to determine the role that protein dynamics play in facilitating the interaction between the NS1ED and CPSF30 and intracellular localization of NS1. In Aim 3, NMR spectroscopy will be used to determine the mechanism of action by which JJ3297 suppresses influenza replication. Our innovative approach will be the first investigation into how protein dynamics and strain specific structural variations facilitate proper function of NS1 in the context of viral replication and pathogenicity. This will also be the first systematic study to determine functional variations in NS1 between multiple strains of influenza. The proposed research is significant because it will define the molecular mechanisms underlying NS1 functions shown to modulate influenza virulence. By defining these molecular mechanisms, this proposal will inform efforts in developing influenza antiviral drugs targeting NS1, thereby supporting the overall mission of the NIH.

开发有效的抗病毒药物需要详细了解目标宿主与病原体相互作用的分子机制，这些相互作用的精确结构模型可以提供具有原子分辨率的机制细节，以帮助有效开发针对具有重要健康相关性的病原体的新型化合物。流感病毒是此类病原体的一个主要例子，每年都会通过在包括人类在内的许多动物宿主中感染和复制而产生新的流感病毒株。这些病毒株引起疾病或毒力的相对能力由下式决定。一些尽管已知流感非结构蛋白 1 (NS1) 在毒力中发挥关键作用，但我们对促进多种毒株依赖性功能的遗传和结构决定因素的了解存在根本差距。因此，我们的长期目标是了解 NS1 菌株依赖性功能的分子机制，该应用的目的是从结构上表征与两种细胞蛋白（CPSF30 和 RIG-I）的相互作用。那我们的中心假设是，某些 NS1 变体独特的结构和动态特征解释了 NS1 的多种功能，该研究的基本原理是阐明结构与功能的关系。 NS1 与其细胞相互作用伙伴之间的相互作用将有助于开发针对这些已知调节毒力的关键相互作用的抗病毒药物，我们的中心假设将通过追求三个特定目标进行检验：1）在结构和功能上表征之间的多重相互作用。 NS1 和 RIG-I，2) 确定微秒-毫秒 (μs-ms) 运动在 NS1 效应器结构域 (NS1ED) 正常功能中的作用，3) 确定 JJ3297 抑制流感病毒复制的作用机制。 1，核磁共振波谱和突变重组流感病毒将用于结构和功能表征 NS1 和 RIG-I 之间的多重相互作用。分散实验将用于确定蛋白质动力学在促进 NS1ED 和 CPSF30 之间的相互作用以及 NS1 的细胞内定位中所起的作用。在目标 3 中，NMR 波谱将用于确定 JJ3297 抑制流感病毒复制的作用机制。我们的创新方法将首次研究蛋白质动力学和菌株特异性结构变异如何促进 NS1 在病毒复制和致病性背景下的正常功能。确定多种流感病毒株之间 NS1 的功能变异，这项研究意义重大，因为它将定义 NS1 调节流感毒力的分子机制。通过定义这些分子机制，该提案将为开发针对 NS1 的流感抗病毒药物的努力提供信息。 ，从而支持 NIH 的总体使命。

项目成果

NETSeq reveals heterogeneous nucleotide incorporation by RNA polymerase I.

• DOI: 10.1073/pnas.1809421115
• 发表时间: 2018-12-11
• 期刊: Proceedings of the National Academy of Sciences of the United States of America
• 影响因子: 11.1
• 作者: Clarke AM;Engel KL;Giles KE;Petit CM;Schneider DA
• 通讯作者: Schneider DA

Structural Investigations of Interactions between the Influenza a Virus NS1 and Host Cellular Proteins.

• DOI: 10.3390/v15102063
• 发表时间: 2023-10-07
• 期刊: Viruses
• 作者: Blake ME;Kleinpeter AB;Jureka AS;Petit CM
• 通讯作者: Petit CM