Molecular Mechanisms for the High Pathogenicity of 1918 Influenza Virus

描述(申请人提供)：我们的长期目标是了解大流行性流感病毒的出现，例如导致有记录以来最具破坏性的传染病爆发的1918年流感病毒。我们特别问了两个问题：(I)出现可与1918年病毒相媲美的大流行毒株的可能性有多大？(Ii)导致1918年病毒空前毒力的分子特征是什么？我们的研究表明，编码1918病毒样蛋白的禽流感病毒基因在自然界中仍然存在，这表明未来可能会出现1918类大流行病毒。因此，在目标1中，我们将评估1918年出现类似病毒的大流行性流感病毒的风险。为了评估这种风险，我们产生了一种类似1918年的禽流感病毒，它包括编码1918年类似蛋白的禽流感病毒片段。我们将在该病毒中引入1918个病毒氨基酸，以确定对致病性至关重要的变化。在一种平行的方法中，我们产生了突变的1918年病毒，在这种病毒中，一次一个基因被其最接近的禽类亲属(由蛋白质同源性确定)所取代。将1918个氨基酸引入‘禽类’基因/蛋白将揭示对1918年病毒致病性至关重要的氨基酸。我们和其他人已经确定HA基因是1918年流感病毒空前毒力的重要决定因素。HA蛋白在病毒生命周期中有两个重要的功能--受体结合和病毒与内体膜的融合。目前，尚不清楚1918年的HA蛋白在其受体结合特性和/或融合动力学方面是否与当代人类病毒不同，以及是否潜在的差异导致了1918年病毒的高毒力。因此，在目标2中，我们将测试1918 HA蛋白在其受体结合特性方面是否与当代人类病毒的不同。这些特征可能会受到HA糖基化模式的影响，促使我们测试1918年非典型HA糖基化模式对毒力的意义。此外，我们将表征1918年HA蛋白的融合动力学，特别强调1918年HA蛋白中先前未被识别的被认为影响融合的组氨酸补丁的特征。除了HA基因，病毒复制复合体已被确定为宿主范围和致病性的决定因素。我们的初步数据还表明，1918年病毒复制复合体在1918年病毒的毒力中发挥了作用。然而，复制复合体的组件对提高毒力的确切贡献和潜在的机制(包括对宿主细胞反应的影响)目前尚不清楚，我们建议在目标3中填补这一知识空白。总的来说，这些研究旨在解决流感病毒研究中的两个关键问题--为什么1918年的病毒具有高致病性，以及出现另一种类似1918年的病毒的风险有多大？公共卫生相关性：我们的长期目标是了解出现类似1918年的大流行性流感病毒的可能性，以及导致1918年大流行性流感病毒空前毒力的机制。这些知识将有助于我们为未来的流感大流行做好准备，并迅速识别具有大流行潜力的流感病毒株。

DESCRIPTION (provided by applicant): Our long-term goal is to understand the emergence of pandemic influenza viruses, such as the 1918 influenza virus that caused the most devastating outbreak of an infectious disease on record. In particular, we are asking two questions: (i) how likely is the emergence of a pandemic strain comparable to the 1918 virus?; and (ii) what are the molecular features that accounted for the unprecedented virulence of the 1918 virus? Our studies have revealed that avian influenza virus genes that encode 1918 virus-like proteins still exist in nature, suggesting that 1918-like pandemic viruses may emerge in the future. In Aim 1, we will therefore assess the risk for the emergence of 1918 virus-like pandemic influenza viruses. To assess this risk, we have generated a 1918-like avian influenza virus comprising avian influenza viral segments that encode 1918-like proteins. We will introduce 1918 viral amino acids into this virus to identify changes that are critical for pathogenicity. In a parallel approach, we have generated mutant 1918 viruses in which one gene at a time is replaced with its closest avian relative (as determined by protein homology). The introduction of 1918 amino acids into the 'avian' gene/protein will reveal amino acids that are critical for 1918 virus pathogenicity. We and others have identified the HA gene as an important determinant of the unprecedented virulence of the 1918 influenza virus. The HA protein carries out two critical functions in the viral life cycle - receptor-binding and fusion of the viral and endosomal membranes. Currently, it is not known if the 1918 HA protein differs in its receptor-binding properties and/or fusion kinetics from contemporary human viruses, and if potential differences contributed to the high virulence of the 1918 virus. In Aim 2, we will therefore test whether the 1918 HA protein differs from that of contemporary human viruses in its receptor-binding properties. These features may be affected by the HA glycosylation pattern, prompting us to test the significance of the atypical 1918 HA glycosylation pattern for virulence. In addition, we will characterize the fusion kinetics of the 1918 HA protein, with particular emphasis on the characterization of a previously unrecognized histidine patch in the 1918 HA protein that is thought to affect fusion. In addition to the HA gene, the viral replication complex has been identified as a determinant of host range and pathogenicity. Our preliminary data also suggest a role for the 1918 virus replication complex in the virulence of the 1918 virus. However, the exact contributions of the components of the replication complex to increased virulence and the underlying mechanisms (including effects on host cell responses) are currently unknown, a gap in knowledge that we propose to fill in Aim 3. Collectively, these studies are designed to address two critical questions in influenza virus research - why was the 1918 virus highly pathogenic, and what is the risk for emergence of another 1918-like virus? PUBLIC HEALTH RELEVANCE: Our long-term goal is to understand the likelihood of the emergence of 1918-like pandemic influenza viruses, and the mechanisms that accounted for the unprecedented virulence of the 1918 pandemic influenza virus. Such knowledge will help us to prepare for future influenza pandemics and to quickly recognize influenza virus strains with pandemic potential.