Clock Gene Control of Viral Infection and Asthma

项目总结 哮喘是一种常见的肺部疾病，其活动受时间的影响很大。哮喘症状因人而异 季节(反映呼吸道病毒感染率)，而且它们也因一天中的不同时间而不同，因此许多 患者在半夜出现最严重的症状。后者表明生物钟， 一组产生昼夜节律的基因与哮喘有关。而病毒和昼夜节律 节律都会影响哮喘症状，这两个方面的疾病从未出现过 机械连接在一起。在这里，我们发现一种名为BMal1的生物钟基因可能通过以下方式影响哮喘 调节肺部对常见呼吸道病毒的反应。我们发现时钟基因BMal1的破坏 对副流感和甲型流感病毒产生异常的抗病毒反应，导致严重的 下呼吸道感染。BMal1的抗病毒作用似乎局限于呼吸道上皮细胞，其中 这种基因在急性病毒性疾病期间协调干扰素反应。在急性感染痊愈后，我们展示了 在小鼠中，BMal1缺乏会加剧病毒后慢性呼吸道疾病，包括 以哮喘肺为特征的。最后，我们发现BMal1在呼吸道样本中的表达下调 来自哮喘患者。根据这些数据，我们假设生物钟调节抗病毒药物。 呼吸道上皮细胞通过BMal1的反应，从而控制急性和慢性病毒的严重程度 肺部病理。我们进一步假设，人呼吸道上皮细胞的生物钟功能存在缺陷。 哮喘患者，导致BMal1表达减少，并潜在地解释了 这些患者已经感染了呼吸道病毒。为了确定BMal1介导抗病毒的机制 呼吸系统中的防御我们提出了以下具体目标：1)确定BMal1如何在呼吸道中 上皮细胞控制干扰素基因表达和急性呼吸道病毒病的严重程度 2)确定BMal1缺乏如何促进小鼠的病毒后慢性呼吸道疾病；以及，3) 哮喘患者呼吸道上皮昼夜节律紊乱的性质探讨 病人。我们将通过使用呼吸道条件BMal1基因敲除小鼠和BMal1-1体外培养来解决Aim1问题。 缺失小鼠气管上皮细胞以描述该基因如何调节干扰素对呼吸的反应 病毒(仙台病毒和甲型流感病毒)。我们将通过使用他莫昔芬诱导的BMal1基因敲除来解决目标2 小鼠确定在感染过程中何时需要BMal1来调节病毒后肺部疾病。我们 我将通过使用荧光素酶报告系统来分析生物钟功能和BMal1的表达来解决目标3 在培养的人类呼吸道细胞中，来自正常和哮喘受试者。

PROJECT SUMMARY Asthma is a common lung disorder whose activity is strongly influenced by time. Asthma symptoms vary with the season (reflecting rates of respiratory viral infection), and they also vary by the time of day, such that many patients experience their worst symptoms in the middle of the night. The latter suggests that the circadian clock, a collection of genes that produce circadian rhythms, is connected to asthma. While viruses and circadian rhythms are both known to affect asthma symptoms, these two aspects of disease have never been mechanistically linked. Here, we discovered that a circadian clock gene called bmal1 may impact asthma by regulating the lung’s response to common respiratory viruses. We show that disruption of the clock gene bmal1 in mice produces an aberrant antiviral response to parainfluenza and influenza A viruses, resulting in severe lower respiratory tract infection. The antiviral actions of bmal1 appear localized to airway epithelial cells, where this gene orchestrates interferon responses during acute viral illness. After the acute infection resolves, we show in mice that bmal1 deficiency exacerbates post-viral chronic airway disease, including features that are characteristic of asthmatic lungs. Finally, we found that bmal1 expression is down-regulated in airway samples from asthma patients. Based on these data, we hypothesize that the circadian clock regulates the antiviral responses of airway epithelial cells through bmal1, and thereby controls the severity of acute and chronic viral lung pathology. We further hypothesize that circadian clock function is defective in the airway epithelial cells of asthmatics, leading to reduced bmal1 expression, and potentially explaining the heightened susceptibility that these patients have to respiratory viruses. To determine the mechanism by which bmal1 mediates antiviral defenses in the respiratory system we propose the following Specific Aims: 1) Determine how bmal1 in airway epithelial cells controls interferon gene expression and the severity of acute respiratory viral illness in mice; 2) Determine how bmal1 deficiency promotes post-viral chronic airway disease in mice; and, 3) Determine the nature of the circadian clock dysfunction in the airway epithelium of asthmatic patients. We will address Aim1 by using airway-conditional bmal1 knockout mice and in vitro culture of bmal1- null mouse tracheal epithelial cells to delineate how this gene regulates the interferon response to respiratory viruses (Sendai and Influenza A viruses). We will address Aim 2 by using tamoxifen-inducible bmal1 knockout mice to define when in the course of infection bmal1 is needed for the regulation of post-viral lung disease. We will address Aim 3 by using a luciferase reporter system to analyze circadian clock function and bmal1 expression in cultured human airway cells, derived from normal and asthmatic subjects.