Chromosome 17q21, allergic inflammation, and remodeling

这项建议的重点是ORMDL3，染色体17q21上的一个基因，它与 在几项全基因组关联研究中发现人类哮喘。由于ORMDL3在CD4+细胞中高度表达， 细胞，在这个建议中，我们试图确定ORMDL3如何影响CD4+细胞功能，过敏， 炎症和哮喘。虽然染色体17q21与哮喘的连锁关系已经非常明确， 支持肺细胞中这种相关性的生物学机制与 哮喘还没有得到很好的理解，这是本提案的重点。此外，至少有9个基因 位于染色体17q21上的这个区域， 对了解这个区域如何影响哮喘的发展很重要。作为SNP连接 ORMDL3对哮喘的作用与ORMDL3表达水平的增加有关，我们产生了普遍的 表达增加水平的人ORMDL3（hORMDL3）的ORMDL3转基因（TG）小鼠和 表明它们自发地（在没有过敏原暴露的情况下）显著发展 ASM增加和AHR增加，这是哮喘的主要特征。除了这一重要的基线效应之外， ORMDL3在ASM中的表达对AHR的影响，ORMDL3也在增强Th2反应和AHR中起重要作用 如我们对过敏原攻击的通用hORMDL3 TG小鼠的研究所证实的。因此， 这一建议是为了增加我们对ORMDL3在CD4+ T淋巴细胞中表达的理解 增强Th2对过敏原攻击的应答。在这个拨款建议中，我们建议证明， 表达增加水平的ORMDL3的CD4+细胞（小鼠和人）具有增强的Th2应答 在体外和体内对屋尘螨（HDM）变应原进行研究，并使用蛋白质组学和RNAseq方法计划 鉴定HDM四聚体阳性CD4+细胞中介导这种ORMDL3效应的下游途径。 被鉴定为ORMDL3下游的通路的靶向敲低或过表达将 证明它们对CD4细胞功能的贡献。最后，我们使用单碱基编辑来编辑 与ORMDL3连接的SNP，以确定哪些SNP在功能上调节ORMDL3、Th2 细胞因子和ORMDL3的下游通路。

The focus of this proposal is on ORMDL3 a gene on chromosome 17q21, which has been highly linked to human asthma in several genome wide association studies. As ORMDL3 is highly expressed in CD4+ cells, in this proposal we seek to determine how ORMDL3 influences CD4+ cell function, allergic inflammation, and asthma. Although the linkage of chromosome 17q21 to asthma is very well established, the biologic mechanism(s) underpinning this association in lung cells pertinent to the pathogenesis of asthma is not as well understood and is the focus of this proposal. In addition as there are at least 9 genes located in this region on chromosome 17q21 understanding the biology of each of these genes individually is important to understanding how this region influences the development of asthma. As the SNP linking ORMDL3 to asthma is associated with increased levels of ORMDL3 expression, we generated universal ORMDL3 transgenic (TG) mice that express increased levels of human ORMDL3 (hORMDL3) and demonstrated that they spontaneously (in the absence of allergen exposure) develop significantly increased ASM and increased AHR, major features of asthma. In addition to this important baseline effect of ORMDL3 in ASM on AHR, ORMDL3 also plays a significant role in enhancing Th2 responses and AHR as demonstrated in our studies of allergen challenged universal hORMDL3 TG mice. Thus, the focus of this proposal is to increase our understanding of how ORMDL3 expressed in CD4+ T lymphocytes enhances Th2 responses to allergen challenge. In this grant proposal we propose to demonstrate that CD4+ cells (mouse and human) expressing increased levels of ORMDL3 have enhanced Th2 responses in vitro and in vivo to house dust mite (HDM) allergen, and using proteomic and RNAseq approaches plan to identify downstream pathways in HDM tetramer positive CD4+ cells that mediate this ORMDL3 effect. Targeted knockdown or overexpression of pathways identified to be downstream of ORMDL3 will demonstrate their contribution to the function of CD4 cells. Finally, we are using single base editing to edit SNPs linked to ORMDL3 to determine which SNPs functionally regulate levels of ORMDL3, Th2 cytokines, and downstream pathways of ORMDL3.