Host Determinants of Human Metapneumovirus Immunity and Pathogenesis

HMPV是全球儿童下呼吸道感染的主要原因，仅次于呼吸道感染。 合胞病毒(RSV)。虽然几乎所有的人都在童年时感染了HMPV，但对HMPV的免疫力 是不完整的，再感染在一生中都会发生。感染HMPV的患者更有可能住院治疗 潜在疾病，如哮喘、慢性阻塞性肺疾病、艾滋病毒或早产。确实有 没有批准的抗病毒疗法或疫苗。宿主或病毒的毒力决定因素对于HMPV来说是未知的。 许多先前研究的局限性之一是，大多数HMPV研究使用的是少数几种实验室适应的毒株中的一种 对小鼠造成最小的疾病。我们已经确定了导致严重和致命的HMPV临床分离株 为阐明发病机制提供了工具。我们的初步数据显示， 强毒HMPV能有效地诱导型和III型干扰素，并且这些细胞因子表现出不一致 角色。干扰素是启动和形成适应性免疫反应的关键，但也可能导致疾病；我们的目标是 明确I型和III型干扰素在HMPV中的作用。HMPV抑制包括STAT1在内的I型干扰素反应 和STAT2的磷酸化，机制未知，HMPV缺乏副粘病毒V蛋白或 RSV的NS1/NS2。几种HMPV蛋白已被发现，包括G、M2-1、P和SH。已发布的数据 来自我们小组和其他人的研究表明，SH介导免疫抑制，但机制或特定的SH宿主 蛋白质之间的相互作用是未知的。其他HMPV蛋白对毒力的贡献尚未确定。 我们建议使用体外和体内方法来解决这些知识差距。在目标1中，我们将定义 用全局性和条件性方法研究III型干扰素(λ)在人乳头瘤病毒免疫和致病机制中的作用 基因敲除老鼠。目标2建议识别SH的细胞靶点(S)，定义相互作用的结构域，并发现 HMPV先天免疫抑制的分子机制。我们将使用标记的瞬时转染法 突变的SH蛋白和带有SH突变的病毒在细胞和小鼠实验中。在目标3中，将使用反转 在实验室开发的遗传学，用于产生嵌合病毒并识别毒力的病毒蛋白决定因素 使用已建立的小鼠模型。我们的初步数据表明，干扰素-λ在人乳头瘤病毒免疫中起着重要作用， 确认HMPV SH对先天免疫的抑制，并展示无毒和强毒的强度 以确定毒力的病毒决定因素。拟议中的研究结果将阐明机制。 HMPV致病机制的研究，为病毒的潜在治疗途径和策略提供了蓝图 疫苗的衰减率。这一发现很可能与其他呼吸道病毒相关。

HMPV is a major cause of lower respiratory infection (LRI) in children worldwide, second only to respiratory syncytial virus (RSV). Although nearly all people are infected with HMPV during childhood, immunity to HMPV is incomplete and re-infections occur throughout life. Hospitalization with HMPV is more likely in persons with underlying conditions such as asthma, chronic obstructive pulmonary disease, HIV, or prematurity. There are no approved antiviral therapies or vaccines. Host or viral determinants of virulence are not known for HMPV. One limitation of many prior studies is that most HMPV research uses one of a few lab-adapted strains that cause minimal disease in mice. We have identified clinical isolates of HMPV that cause severe and fatal disease in mice, providing tools to elucidate mechanisms of pathogenesis. Our preliminary data show that virulent HMPV potently induces types I and III interferon (IFN), and that these cytokines exhibit discordant roles. IFN is critical to initiate and shape adaptive immune responses but can contribute to disease; we aim to define the contribution of types I and III IFN in HMPV. HMPV inhibits type I IFN responses including STAT1 and STAT2 phosphorylation by an unknown mechanism, and HMPV lacks the paramyxovirus V protein or NS1/NS2 of RSV. Several HMPV proteins have been implicated, including G, M2-1, P, and SH. Published data from our group and others suggest SH mediates immune inhibition but a mechanism or specific SH-host protein interaction is unknown. The contribution of other HMPV proteins to virulence has not been defined. We propose to use in vitro and in vivo approaches to address these knowledge gaps. In Aim 1, we will define the contributions of type III IFN (IFN-λ) to HMPV immunity and pathogenesis using global and conditional knockout mice. Aim 2 proposes to identify the cellular target(s) of SH, define interacting domains, and discover the molecular mechanisms of innate immune inhibition by HMPV. We will use transient transfection of tagged mutant SH proteins and viruses with SH mutations in cell and mouse experiments. In Aim 3, will use reverse genetics developed in the lab to generate chimeric viruses and identify viral protein determinants of virulence using established mouse models. Our preliminary data suggest an important role for IFN-λ in HMPV immunity, confirm innate immune inhibition by HMPV SH, and demonstrate the strength of the avirulent and virulent strains to identify viral determinants of virulence. The results of the proposed research will clarify mechanisms of HMPV pathogenesis and provide a blueprint for potential therapeutic avenues and strategies for viral attenuation for vaccines. The findings are likely to be relevant to other respiratory viruses.