Molecular Determinants of Usual Interstitial Pneumonia (UIP)

摘要：本提案的总体目标是了解获得功能的MUC5B启动子是如何 变异会影响转录调节和基因表达，这在临床上是不同类型的 间质性肺炎(UIP)。UIP最初被描述为一个形态实体，然而最近被描述为 使用特定的病理和放射学标准来定义。而UIP是特发性肺病的特征 纤维化(IPF)，这些慢性纤维性间质性肺炎的病理和放射学模式也是 典型的慢性过敏性肺炎(CHP)，类风湿性关节炎相关间质性肺疾病 (RA-ILD)、石棉肺和几种药物引起的肺部疾病。MUC5B的功能增强启动子变异体 (Rs35705950)是IPF的主要危险因素，存在于50%的受影响患者中，也 据报道，这是CHP和RA-ILD发生的主要遗传风险变异。然而，虽然IPF是 根据定义，CHP是在反复接触有机抗原后发生的，而RA-ILD是一种 类风湿关节炎的并发症。我们通过了解MUC5B之间的关系提出 在IPF、CHP和RA-ILD中的启动子变异、转录调控和基因表达，我们将能够 确定对uIP的发展至关重要的共同分子元素。两个关键问题 我们计划解决的问题是：1)无论临床情况如何，uIP的共同分子特征是什么， 2)MUC5B启动子变异体是否具有与UIP相同的独特分子特征？我们的假设是 计划测试的是功能增强的MUC5B启动子变体驱动细胞特异性染色质 可获得性和定义uIP的基因表达。在目标1中，我们将使用单核RNA测序 (SnRNA-seq)以鉴定IPF、CHP和RA-ILD的细胞特异性转录图谱，并确定 UIP常见的细胞特异性转录图谱与MUC5B启动子变异的关系。在目标2中， 我们将使用转座酶可及染色质的组合指标化单核分析(snatac-seq)。 确定IPF、CHP和RA-ILD的细胞特定染色质可及性配置文件，并确定 UIP常见细胞特异性染色质可及性谱与MUC5B启动子变异的关系。 在目标3中，我们将对MUC5B启动子基因、snrna-seq和snatac-seq进行综合分析。 数据采用单细胞表达QTL、多组体和网络推理方法鉴定uIP特异性基因 在特定细胞类型中具有UIP关键驱动因素的监管网络。在目标4中，我们将验证转录 通过确定这些关键基因与MUC5B相关的uIP的关系来确定这些共同的特征 UIP的致病异质性及相关的体外生物学研究。总的来说，我们将描述监管机构的特征 MUC5B对UIP细胞特异性转录图谱和网络的影响，启动了对新的研究 这些不治之症的致病机制和药物靶点。

ABSTRACT: The overall goal of this proposal is to understand how the gain-of-function MUC5B promoter variant affects transcriptional regulation and gene expression that are common to clinically distinct types of usual interstitial pneumonia (UIP). UIP was initially described as a morphologic entity, however, recently has been defined using specific pathologic and radiographic criteria. While UIP is characteristic of idiopathic pulmonary fibrosis (IPF), these pathologic and radiographic patterns of chronic fibrosing interstitial pneumonia are also typical of chronic hypersensitivity pneumonitis (CHP), rheumatoid arthritis-associated interstitial lung disease (RA-ILD), asbestosis, and several drug-induced lung diseases. The gain-of-function promoter variant in MUC5B (rs35705950) is the dominant risk factor for IPF, is present in >50% of affected patients, and has also been reported to be the dominant genetic risk variant for the development of CHP and RA-ILD. However, while IPF is by definition idiopathic, CHP develops following repeated exposure to organic antigens, and RA-ILD is a complication of rheumatoid arthritis. We proposed by understanding the relationship between the MUC5B promoter variant, transcriptional regulation, and gene expression in IPF, CHP, and RA-ILD, we will be able to identify the common molecular elements that are critical to the development of UIP. The two critical questions that we plan to address are: 1) what are the common molecular features of UIP, irrespective of clinical context, and 2) does MUC5B promoter variant have a unique molecular signature common to UIP? The hypothesis we plan to test is that the gain-of-function MUC5B promoter variant drives cell-specific chromatin accessibility and gene expression that define UIP. In Aim 1, we will use single nucleus RNA sequencing (snRNA-seq) to identify the cell-specific transcriptional profiles for IPF, CHP, and RA-ILD, and determine the relationship of the common cell-specific transcriptional profiles of UIP to the MUC5B promoter variant. In Aim 2, we will use a combinatorial indexing single nucleus assay for transposase-accessible chromatin (snATAC-seq) to identify the cell-specific chromatin accessibility profiles for IPF, CHP, and RA-ILD, and determine the relationship of the common cell-specific chromatin accessibility profiles of UIP to the MUC5B promoter variant. In Aim 3, we will perform integrative analyses of the MUC5B promoter genotype, snRNA-seq, and snATAC-seq data using single-cell expression QTL, multi-omic and network inference methods to identify UIP-specific gene regulatory networks with key drivers of UIP in specific cell types. In Aim 4, we will validate the transcriptional features that are common to MUC5B-associated UIP by determining the relationship of these key genes to the pathogenic heterogeneity of UIP and relevant in vitro biology. In aggregate, we will characterize regulatory effects of MUC5B on cell-specific transcriptional profiles and networks in UIP, launching investigation of novel pathogenic mechanisms and drug targets for these incurable diseases.