Functional Genomics of IL-33 expression and asthma risk

描述（由申请人提供）：哮喘影响着全球约3亿人，是基础科学造福临床医学的重要挑战。最近，两个独立的全基因组关联研究的荟萃分析GWAS数据已经确定了编码IL-1家族细胞因子IL-33及其受体IL 1 RL 1（ST 2）的基因是哮喘的易感基因座。IL-33具有多种功能，包括激活2型先天淋巴细胞、扩增Th 2细胞和增强CD 8 T细胞的抗病毒功能。随着IL-33与哮喘病因学的细胞和分子机制成为密切关注的焦点，一些基本问题仍然没有答案。本申请集中于阐明人IL 33表达在转录水平以及生物体水平上的调节。虽然已知IL-33在上皮细胞和结构细胞中组成型表达，但我们和其他人最近发现IL-33由鼠造血细胞产生。我们现在证明IL 33在人肺白细胞中表达，并且过敏原诱导IL 33在人CD 14+单核细胞中表达。这些发现表明了IL 33在肺中表达的新范例，并且代表了理解该细胞因子在气道Th 2炎症中的影响组织特异性表达的创新机会。使用ENCODE和1000个基因组数据库，我们已经进行了IL 33基因座的调控表观遗传景观的计算机询问，并确定了两个高度关注的区域;一个区域具有较强的增强子活性，第二个区域包含两个定义的CTCF结合位点，提示绝缘子活性。我们现在证明了第一个区域可以作为增强子发挥作用，重要的是，这种活性被非洲血统个体中特异性发现的SNP修饰。然而有用的，这些公共数据库未能突出调控元件，只有在诱导基因表达的环境刺激后变得明显。因此，过敏原刺激后原代细胞中IL 33表达的调节将可能阐明在静态细胞系中未实现的新的调节元件。我们研究的中心假设是，调控元件内的非编码遗传变异改变了IL 33的时空表达模式，这是IL 33基因座SNP与哮喘风险相关的核心机制。为了验证这一假设，提出了以下目标：1）鉴定在刺激的造血细胞和支气管上皮细胞中IL 33基因表达后发生的表观遗传学变化; 2)确定角色 增强子元件在IL 33表达的调节中起作用;和3）确定CTCF在IL 33表达中的作用。

DESCRIPTION (provided by applicant): Asthma affects an estimated 300 million people worldwide and represents an important challenge for basic science to benefit clinical medicine. Recently, two independent meta-analysis of genome wide association studies GWAS data have identified the genes encoding the IL-1¿ family cytokine, IL-33, and its receptor, IL1RL1 (ST2), as susceptibility loci for asthma. A plethora of functions have been attributed to IL-33 including activation of type-2 innate lymphoid cells, expansion of Th2 cells, and augmentation of anti-viral function of CD8 T cells. As the cellular and molecular mechanisms connecting IL-33 to asthma etiology become the focus of intense scrutiny, several basic questions remain unanswered. This application focuses on elucidating the regulation of human IL33 expression, both at the transcriptional level as well as the organismal level. While IL33 is known to be constitutively expressed in epithelial cells and structural cells, we and others have recently found IL-33 produced by murine hematopoietic cells. We now demonstrate that IL33 is expressed in human lung leukocytes and that allergens induce IL33 expression in human CD14+ monocytes. These findings suggest a new paradigm of IL33 expression in the lungs, and represent an innovative opportunity to understand the impact tissue specific expression of this cytokine in airway Th2 inflammation. Using ENCODE and 1000 Genomes databases, we have performed in silico interrogation of the regulatory epigenetic landscape of the IL33 locus and have identified two regions of high interest; one region has strong enhancer activity, and a second region contains two defined CTCF binding sites, suggestive of insulator activity. We now demonstrate that the first region can function as an enhancer, and importantly, this activity is modified by a SNP found specifically in individuals of African ancestry. However useful, these public databases fail to highlight regulatory elements that only become evident after induction of gene expression by environmental stimulation. Thus, regulation of IL33 expression in primary cells after allergen stimulation will likely elucidate novel regulatory elements not realized in static cell lines. The central hypothesis of our study posits that noncoding genetic variants within regulatory elements alter the temporal and spatial expression patterns of IL33, and that this is a central mechanism behind the association of SNPs at the IL33 locus with asthma risk. To test this hypothesis the following aims are proposed: 1) identify the epigenetic changes that occur upon IL33 gene expression in stimulated hematopoietic cells and bronchial epithelial cells.; 2) determine the role the enhancer element(s) play in the regulation of IL33 expression; and 3) determine the role of CTCF in IL33 expression.