Replication of influenza virus

流感病毒的复制

• 批准号: 7924259
• 负责人: Robert M Krug
• 金额: $ 8.56万
• 依托单位: UNIVERSITY OF TEXAS AT AUSTIN
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-26 至 2010-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7924259
• 关键词: Accounting; Amino Acids; Animal Model; Antiviral Agents; Antiviral Response; Applications Grants; Attenuated; Attenuated Live Virus Vaccine; Benchmarking; Binding; Binding Sites; Biochemical; Biological Assay; Birds; Cell physiology; Cells; Cercopithecine Herpesvirus 1; Complex; Defect; Development; Double-Stranded RNA; Enzymes; Event; Future; Generations; Genes; Genetic; Goals; Grant; Hemagglutinin; Human; Human Influenza A Virus; Inactivated Vaccines; Infection; Influenza; Influenza A Virus, H1N1 Subtype; Influenza A Virus, H3N2 Subtype; Influenza A Virus, H5N1 Subtype; Influenza A virus; Influenza B virus; Interferon Activation; Interferons; Laboratories; Lead; Life; Ligase; Lung diseases; Mediating; Messenger RNA; Molecular; Mus; N-terminal; National Institute of Allergy and Infectious Disease; Neuraminidase; Nuclear Magnetic Resonance; Oligonucleotides; PTPN11 gene; Pathway interactions; Play; Polymerase; Population; Principal Investigator; Process; Production; Protein Binding; Proteins; RNA Binding; RNA Recognition Motif; RNA chemical synthesis; Reagent; Recombinants; Regulation; Research; Ribonucleases; Roentgen Rays; Role; Simian B disease; Site; Structure; Surface; Testing; Transfection; Ubiquitin; Vaccines; Vero Cells; Viral; Viral Physiology; Viral Proteins; Virulence; Virulence Factors; Virus; Virus Diseases; Virus Replication; Work; attenuation; base; cellular targeting; human G1P2 protein; immunogenic; influenzavirus; insight; mRNA Precursor; mortality; mutant; pandemic disease; prevent; programs; recombinant virus; research study; vaccine-induced immunity; viral RNA

DESCRIPTION (provided by applicant): Influenza A and B viruses cause a highly contagious respiratory disease in humans. Influenza B viruses infect only humans, whereas influenza A viruses infect many avian and mammalian species. Influenza A viruses are responsible for the periodic wide-spread pandemics that result in high mortality rates. The 1918 pandemic was the most devastating. Highly pathogenic H5N1 influenza A viruses are prime candidates for causing the next pandemic. The NS1 proteins of influenza A virus (NS1A protein) and of influenza B virus (NS1B protein) are multifunctional proteins that suppress cellular antiviral responses and are virulence factors. Many of the functions of the NS1A protein differ from those of the NS1B protein. The overall aim is to elucidate how multiple functions of the NS1A and NS1B proteins play important roles during infection. The NS1A protein has a binding site for double-stranded RNA (dsRNA) and binding sites for several cellular proteins, including CPSF30, a cellular factor required for the processing of cellular pre-mRNAs. CPSF30 binding, which results in the inhibition of the production of all cellular mRNAs, including interferon-2 (IFN-2) mRNA, is the primary, if not the only, mechanism by which the NS1A protein of a human influenza A virus (isolated in 1972) inhibits IFN-2 mRNA production. One aim is to determine whether this mechanism is shared by all other human influenza A viruses, including H5N1 and 1918 viruses. This research will entail the generation of multiple recombinant viruses expressing NS1A proteins containing specific amino acid changes. Recent structural results on the dsRNA- and CPSF30-binding sites of the NS1A protein described in this grant application may lead to the development of new antivirals directed at these binding sites and also to the development of better live attenuated virus vaccines against H5N1 viruses. For the latter purpose, one aim is to generate viruses whose attenuation is fine-tuned by site-specific amino acid changes in these two NS1A binding sites. Motivated by genetic experiments that showed an unexpected functional interaction between the NS1A protein and the viral polymerase, another aim is to use biochemical approaches to elucidate the molecular mechanisms underlying this interaction. The IFN-induced ISG15 protein, which is conjugated to multiple cellular proteins, has antiviral activity against both influenza A and B viruses. One aim is to identify the viral and/or cellular proteins whose conjugation to ISG15 accounts for the antiviral action of ISG15 conjugation against influenza A virus. Because the NS1B protein, unlike the NS1A protein, binds ISG15 and prevents its conjugation to target proteins, research on the antiviral function of ISG15 conjugation against influenza B virus will analyze the defects in replication of a recombinant influenza B virus that expresses a NS1B protein lacking an ISG15 binding site. As an integral part of this research, the role of the dsRNA-binding activity and other functions of the NS1B protein will also be determined. Elucidation of the molecular mechanisms by which ISG15 conjugation inhibits influenza A and B viruses may lead to therapies that exploit these mechanisms.

描述（由申请人提供）：流感和B病毒引起人类具有高度传染性的呼吸道疾病。流感B病毒仅感染人类，而流感A病毒感染了许多禽和哺乳动物。流感病毒是导致导致高死亡率的周期性广泛大流行病的原因。 1918年的大流行是最毁灭性的。高致病性的H5N1流感病毒是导致下一个大流行的主要候选者。流感A病毒（NS1A蛋白）和流感B病毒（NS1B蛋白）的NS1蛋白是抑制细胞抗病毒反应的多功能蛋白，是毒力因素。 NS1A蛋白的许多功能与NS1B蛋白的功能不同。总体目的是阐明在感染过程中NS1A和NS1B蛋白的多个功能如何在感染过程中起重要作用。 NS1A蛋白具有用于双链RNA（DSRNA）的结合位点和几种细胞蛋白的结合位点，包括CPSF30，这是细胞前MRNA处理所需的细胞因子。 CPSF30结合导致抑制所有细胞mRNA的产生，包括干扰素2（IFN-2）mRNA，是主要的（如果不是唯一的，即使不是人类流感病毒的NS1A蛋白）的主要机制（1972年分离出来的），则IFN-2 mRNA的产生。一个目的是确定该机制是否由所有其他人类流感病毒共享，包括H5N1和1918年病毒。这项研究将需要产生多种表达含有特定氨基酸变化的NS1A蛋白的重组病毒。在本赠款应用中描述的NS1A蛋白的DSRNA和CPSF30结合位点的最新结构结果可能导致针对这些结合位点的新抗病毒药的发展，也导致开发更好的活体减毒病毒疫苗针对H5N1病毒。出于后一个目的，一个目的是生成病毒，其衰减受到这两个NS1A结合位点位点特异性氨基酸变化的微调。由遗传实验激发的，该实验表明NS1A蛋白与病毒聚合酶之间存在意外的功能相互作用，另一个目的是使用生化方法来阐明这种相互作用的分子机制。 IFN诱导的ISG15蛋白与多种细胞蛋白共轭具有抗病毒活性，对流感和B病毒均具有抗病毒活性。一个目的是鉴定其与ISG15的偶联的病毒和/或细胞蛋白，解释了ISG15与流感病毒的抗病毒作用。因为NS1B蛋白与NS1A蛋白不同，它结合ISG15并防止其与靶蛋白的结合，因此对ISG15抗病毒功能的研究对影响流感B病毒的抗病毒功能将分析复制的缺陷，以表达AS1b proting Ans1b proting Ans1b proting a ns1b proting Ans1b proting Ans1b proting a ns1b proting Ans1b proting a ns1b proting a ns1b proting a ns1b proting a proting a proting a ns15 ns15。作为这项研究不可或缺的一部分，还将确定DSRNA结合活性和其他功能的作用。阐明ISG15结合抑制流感和B病毒的分子机制可能导致利用这些机制的疗法。