Role of immunoproteasome in airway viral infection

项目概要/摘要 该提案的目标是确定免疫蛋白酶体（IP）在过程中的调节和功能。 气道中鼻病毒感染伴有 2 型炎症，这是 2 型炎症的标志（例如白细胞介素 13 [IL-13]）高 哮喘。鼻病毒 (RV) 感染由于会导致病情加重，因此仍然是一个重大的医疗负担 哮喘和其他肺部疾病。由于气道RV感染尚无有效治疗方法，因此势在必行 定义从受感染的气道细胞（例如上皮细胞）中清除 RV 的新机制（例如 IP）。我们发现 患有气道 2 型炎症的严重哮喘患者气道上皮细胞中 IP 表达减少。 重要的是，我们的初步数据强烈表明 IP 在人原代气道上皮细胞中的表达 RV（RV-A 和 RV-C）感染显着上调，并且关键 IP 的敲除或抑制 成分（例如 LMP2 和 LMP7）增加气道上皮病毒载量。 2 型细胞因子 IL-13 显着 RV 减少了 IP 感应。感染 RV 的 LMP2 敲除小鼠（与野生型小鼠相比）无法清除病毒 有效地增加肺部炎症。我们将检验 RV 期间气道 IP 诱导的假设 感染是消除病毒感染的关键宿主防御机制，但在气道中受到损害 2型炎症，导致持续感染和过度炎症。在目标 1 中，我们将确定 干扰素 (IFN)/干扰素受体 (IFNR)/STAT1 轴在 RV 和 2 型炎症 IP 调节中的作用 在人原代气道上皮细胞中进行基因敲除实验，并利用人体精准- 切割暴露于 IL-13、过敏原和/或 RV 的肺切片或基因（例如 IFNAR 或 STAT1）缺陷小鼠。在目标 2 中， 我们将通过使用 IP 确定 IP 在 RV 介导的哮喘恶化中的作用及其潜在机制 缺陷小鼠（全身或气道上皮条件性基因敲除）和人类初级气道上皮 IP基因敲除的细胞。我们将测试IP的抗病毒和抗炎功能是否依赖于它的调节 RV 3C 蛋白酶和 A20（一种负性免疫调节剂）。在目标 3 中，我们将确定知识产权作为 患有 2 型炎症的气道中的抗病毒药物。通过提供单次低剂量的 IFN-1 或病毒模拟物（即， PolyI:C) 进入培养的人气道上皮或患有 2 型炎症的小鼠气道，我们将测试是否 增强 IP 表达可促进气道急性 RV 感染和炎症的消退。研究结果 将显着提高我们对宿主针对 RV 感染的防御机制的理解，并且可能 为治疗哮喘和其他疾病患者的 RV 感染提供了新的机会。

Project Summary/Abstract The goal of this proposal is to determine the regulation and functions of immunoproteasome (IP) during rhinovirus infection in airways with type 2 inflammation, a hallmark of the type 2 (e.g., interleukin 13 [IL-13]) high asthma. Rhinovirus (RV) infection remains a significant healthcare burden due to its contribution to exacerbations of asthma and other lung diseases. Because there is no effective therapy for airway RV infection, it is imperative to define novel mechanisms (e.g., IP) that clear RV from the infected airway cells (e.g., epithelial cells). We found reduced IP expression in airway epithelial cells of severe asthma patients with airway type 2 inflammation. Importantly, our preliminary data strongly suggest that IP expression in human primary airway epithelial cells was significantly up-regulated by RV (RV-A and RV-C) infection, and that knockout or inhibition of key IP components (e.g., LMP2 and LMP7) increased airway epithelial viral load. Type 2 cytokine IL-13 significantly reduced IP induction by RV. RV-infected LMP2 knockout (vs. wild-type) mice were unable to clear the viruses effectively, and increased lung inflammation. We will test the hypothesis that airway IP induction during RV infection serves as a critical host defense mechanism to eliminate viral infection, but is impaired in airways with type 2 inflammation, leading to persistent infection and excessive inflammation. In Aim 1, we will determine the role of the interferons (IFNs)/IFN receptor (IFNR)/STAT1 axis in IP regulation by RV and type 2 inflammation by performing gene knockout experiments in human primary airway epithelial cells, and using the human precision- cut lung slices or gene (e.g., IFNAR or STAT1) deficient mice exposed to IL-13, allergen and/or RV. In Aim 2, we will determine the role of IP in RV-mediated asthma exacerbation and underlying mechanisms by using IP deficient mice (whole body or airway epithelial conditional gene knockout) and human primary airway epithelial cells with IP gene knockout. We will test if IP’s antiviral and anti-inflammatory functions depend on its regulation of RV 3C protease and A20, a negative immune regulator. In Aim 3, we will determine the efficacy of IP as an antiviral agent in airways with type 2 inflammation. By delivering a single low-dose of IFN-l or a viral mimic (i.e., polyI:C) into cultured human airway epithelium or mouse airways with type 2 inflammation, we will test if enhancing IP expression promotes resolution of airway acute RV infection and inflammation. Research findings will significantly improve our understanding about host defense mechanisms against RV infection, and likely provide a new opportunity to treat RV infections in patients with asthma and other diseases.