Molecular Determinants of Usual Interstitial Pneumonia (UIP)

摘要：本研究的总体目标是了解功能获得性MUC 5 B启动子 变异影响转录调节和基因表达，这是常见的临床不同类型的常见 间质性肺炎（UIP）。UIP最初被描述为一种形态学实体，然而，最近已被 使用特定的病理学和放射学标准定义。而UIP是特发性肺动脉高压的特征， 虽然慢性纤维化间质性肺炎的病理学和放射学模式与肺间质纤维化（IPF）有关， 典型的慢性过敏性肺炎（CHP）、类风湿性关节炎相关的间质性肺病 （RA-ILD）、石棉肺和几种药物引起的肺病。MUC 5 B中的功能获得性启动子变体 （rs35705950）是IPF的主要风险因素，存在于>50%的受影响患者中， 据报道，是CHP和RA-ILD发生的主要遗传风险变体。然而，虽然IPF 根据定义，特发性CHP是在反复暴露于有机抗原后发生的，RA-ILD是一种 类风湿关节炎的并发症我们通过了解MUC 5 B之间的关系， IPF、CHP和RA-ILD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）UIP的共同分子特征是什么，无论临床背景如何， 和2）MUC 5 B启动子变体是否具有与UIP共同的独特分子标记？假设我们 一个测试计划是，功能获得性MUC 5 B启动子变体驱动细胞特异性染色质 定义UIP的可及性和基因表达。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的细胞特异性转录谱，并确定IPF、CHP和RA-ILD的细胞特异性转录谱。 UIP的常见细胞特异性转录谱与MUC 5 B启动子变体的关系。在目标2中， 我们将使用转座酶可接近染色质的组合索引单核测定（snATAC-seq）， 确定IPF、CHP和RA-ILD的细胞特异性染色质可及性谱，并确定 UIP与MUC 5 B启动子变体的常见细胞特异性染色质可及性谱的关系。 在目标3中，我们将对MUC 5 B启动子基因型、snRNA-seq和snATAC-seq进行整合分析 数据使用单细胞表达QTL，多组学和网络推理方法来鉴定UIP特异性基因 调控网络与特定细胞类型中UIP的关键驱动因素。在目标4中，我们将验证转录 通过确定这些关键基因与MUC 5 B相关UIP的关系， UIP的致病异质性和相关的体外生物学。总的来说，我们将描述监管 MUC 5 B对UIP中细胞特异性转录谱和网络的影响，启动了新的研究。 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.