Genetics and Gene Expression Profiling in Asthma

描述（由申请人提供）：哮喘是最常见的慢性呼吸道肺部疾病，在美国影响超过1700万人；鉴定其分子决定因素仍然是转化科学的一个重要优先事项。四年前，我们提出了一种整合基因组方法，将全基因组遗传和表达研究（所谓的表达数量性状位点（eQTL）定位）结合起来，以促进鉴定与哮喘发病机制和自然史有关的新型调控遗传变异。使用这种方法，我们发现了在四个不同的基因组区域赋予哮喘易感性的新的调控遗传变异：染色体17q位点（包括基因ZPBP2， GSDMB和ORMDL3）；脂肪酸去饱和酶位点（FADS1和FADS2）；n -乙酰半乳糖胺酶（NAGA）位点和因子XIII A亚基（F13A1）位点。我们现在建议扩展这些研究，目标是：(1)绘制导致哮喘发病机制的其他调节变异；(2)确定与哮喘遗传相关的特定功能调控变异；(3)表征这些候选基因在哮喘发展中的功能作用。在具体目标1中，我们将扩展我们之前在整合基因组学方面的工作，使用来自四个大型现有数据集的数据进行首次eQTL荟萃分析，这些数据集包含4000多名不同祖先的受试者。这项工作的结果将用于重新解释哮喘全基因组关联研究的荟萃分析。将在另外三个哮喘队列（n E12,000）中重复上述关联。在Specific Aim 2中，我们将通过等位基因失衡筛选、基于染色质的DNA-DNA和dna -蛋白质相互作用研究以及等位基因特异性报告基因测定，对人类支气管上皮细胞和相关细胞类型中的三个新型哮喘基因位点（FADS2、NAGA和F13A1）进行功能精细定位。确认的功能变异将在EVE联盟队列中测试与哮喘的遗传关联。在Specific Aim 3中，我们将在已建立的小鼠模型中对FADS2基因进行功能评估，并在小鼠和人源性人支气管上皮细胞的体外细胞实验中确认其分子机制。该项目具有很高的潜力，可以精确定位哮喘发病机制的功能性遗传变异，更好地了解这些基因在哮喘发病机制中的作用，从而为开发旨在修饰其异常基因表达的新药奠定坚实的基础。

DESCRIPTION (provided by applicant): Asthma is the most common chronic respiratory lung disease, affecting more than 17 million people in the US; identification of its molecular determinants remains an important priority in translational science. Four years ago, we proposed an integrative genomic approach that combined genome-wide genetic and expression studies (so-called expression quantitative trait locus (eQTL) mapping) to facilitate the identification of novel regulatory genetic variation contributing to the pathogenesis and natural history of asthma. Using this approach, we identified novel regulatory genetic variants that confer susceptibility to asthma at four distinct genomic regions: the chromosome 17q locus (that includes the genes ZPBP2, GSDMB and ORMDL3); the Fatty Acid Desaturase locus (FADS1 & FADS2); the N-acetyl galactosaminidase (NAGA) locus, and the factor XIII A subunit (F13A1) locus. We now propose to extend these studies with the goals of (1) mapping additional regulatory variants that contribute to the pathogenesis of asthma; (2) identifying the specific functional regulatory variants underlying the genetic associations with asthma; and (3) characterizing the functional role of these candidate genes in the development of asthma. In Specific Aim 1 we will extend our previous work in the integrative genomics by performing a first of its kind eQTL meta-analysis using data from four large existing datasets of more than 4,000 subjects of diverse ancestry. Results from this effort will be used to reinterpret a meta-analysis of asthma genome-wide association studies. Associations will be replicated in three additional asthma cohorts (n E12,000). In Specific Aim 2, we will perform functional fine-mapping of three of the novel asthma loci (FADS2, NAGA and F13A1) in human bronchial epithelial cells and related cell types by means of allelic imbalance screens, chromatin-based DNA-DNA and DNA-protein interaction studies, and allele specific reporter assays. Confirmed functional variants will be tested for genetic association with asthma in the EVE Consortium cohorts. In Specific Aim 3, we will conduct a functional evaluation of the FADS2 gene in an established murine model, with confirmation of the molecular mechanisms using in vitro cellular assays in both murine- and human-derived human bronchial epithelial cells. This project has high potential to precisely localize functional genetic variants contributing to the pathogenesis of asthma, and provide a better understanding of the role of these genes in the pathogenesis of asthma, thereby establishing a solid foundation for new drug development aimed at modifying their aberrant gene expression. PUBLIC HEALTH RELEVANCE: Asthma is the most common chronic respiratory lung disease; identification of its molecular determinants remains an important priority in translational science. The asthma-associated regulatory genetic variants identified through this application will provide novel insights into the pathogenesis of asthma. The studies of FADS2 will confirm it as a potential target for therapeutic development or environmental modification through dietary modification of fatty acid intake.