Genetic Control of Airway Epithelium Gene Expression in Childhood Asthmatics

 描述(申请人提供)：哮喘是儿童中最常见的慢性病。对包括哮喘在内的复杂人类疾病的基因图谱研究结果表明，大多数相关变异是非编码的，而不是蛋白质编码的。许多这些非编码风险变体可能存在于影响基因表达的调控元件中。这表明在相关组织中发现的表达数量性状基因座(EQTL)变异将呈现一个丰富的哮喘疾病变异的变异库。支气管呼吸道上皮通过影响气道免疫反应、产生堵塞呼吸道的粘液以及向影响气道重塑的气道间充质发出信号而参与哮喘的发病。我们的目标是确定在不同人群中与儿童哮喘风险相关的呼吸道上皮细胞eQTL。鼻腔呼吸道上皮是一种容易获得的替代支气管气道上皮的方法，鼻腔气道转录组与支气管气道转录组非常接近。我们假设许多哮喘遗传易感基因的功能基础涉及对呼吸道上皮基因表达的顺式调节。我们将通过三个特定的目标来验证我们的假设：目标1：从波多黎各儿童哮喘患者和对照组的初级呼吸道上皮细胞中确定顺式调节基因及其eQTL。我们将使用靶向rna-seq来确定所有159个基因的总/等位基因表达水平，这些基因定位于21个已确定的哮喘。 750例波多黎各哮喘患者和正常对照鼻道上皮细胞GWAs基因座研究顺式调节基因将通过eQTL分析确定，eQTL分析使用全基因组的基因型数据和等位基因不平衡分析。EQTL的最终定位将利用来自所有受试者的目标DNA序列的完整变异数据来完成。目的：通过对波多黎各儿童哮喘患者和对照组的原代呼吸道上皮进行eQTL全转录组分析，寻找新的哮喘相关eQTL。所有750名受试者的全部转录组表达将被用于识别哮喘中差异表达和顺式调控的基因，如目标1所示。DNA-SEQ将用于在哮喘相关顺式调控基因中生成完整的变异数据，以执行eQTL精细定位。目的3：研究不同哮喘人群中已识别的鼻哮喘eQTL在下呼吸道、临床和生物学上的意义。我们将在100名哮喘患者的支气管呼吸道上皮细胞中鉴定最强的鼻哮喘eQTL。我们将在7200名波多黎各、墨西哥和非洲裔美国儿童中确定AIMS 1和AIMS 2中确定的最强的细纹呼吸道eQTL是否易患哮喘及相关特征。改变的生物机制将通过对哮喘相关基因的通路分析来确定，这些基因由呼吸道eQTL调节。

 DESCRIPTION (provided by applicant): Asthma is the most common chronic disease among children. Results from gene mapping studies of complex human diseases including asthma have indicated that most associated variants are non-coding, rather than protein coding. Many of these non-coding risk variants likely reside in regulatory elements that affect gene expression. This suggests expression quantitative trait loci (eQTL) variants identified in a relevant tissue will present a variant pool enriched for asthma disease variants. The bronchial airway epithelium contributes to asthma pathogenesis by affecting airway immune responses, through the production of mucus that plugs airways, and by signaling to the airway mesenchyme which influences airway remodeling. Our goal is to identify airway epithelium eQTLs which confer risk of childhood asthma in diverse populations. The nasal airway epithelium is an easily accessible alternative to bronchial airway epithelium and the nasal airway transcriptome closely approximates the bronchial airway transcriptome. We hypothesize the functional underpinnings of many asthma genetic susceptibility loci involve cis-regulation of airway epithelium gene expression. We will test our hypothesis with three specific aims: Aim 1: Identify cis-regulated genes and their eQTLs within asthma GWAS loci in primary airway epithelium from Puerto Rican childhood asthmatics and controls. We will use targeted RNA-seq to determine total/allelic expression levels of all 159 genes localized to the 21 identified asthma GWAS loci in the nasal airway epithelium of 750 Puerto Rican asthmatics and controls. Cis-regulated genes will be determined by eQTL analyses using genome-wide genotype data and allelic imbalance analyses. Finemapping of eQTLs will be accomplished with complete variant data from targeted DNA-seq of all subjects. Aim 2: Identify novel asthma-related eQTLs by whole transcriptome eQTL analysis of primary airway epithelium from Puerto Rican childhood asthmatics and controls. Whole transcriptome expression on all 750 subjects will be used to identify genes that are both differentially expressed in asthma and are cis-regulated, as in Aim 1. DNA-seq will be used to generate complete variant data at the asthma-related, cis-regulated genes to perform eQTL finemapping. Aim 3: Characterize the lower airway, clinical, and biological significance of identified nasal asthma eQTLs in diverse populations of asthmatics. We will characterize the strongest nasal asthma eQTLs in the bronchial airway epithelium of 100 asthmatics. We will determine if the strongest finemapped airway eQTLs identified in Aims 1 and 2 predispose to asthma and related traits in 7200 Puerto Rican, Mexican, and African American children. Altered biologic mechanisms will be identified by pathway analysis of asthma-associated genes regulated by airway eQTLs.