Role of Two Interferon Regulatory Genes in Lupus

摘要 系统性红斑狼疮(SLE或狼疮)是I型干扰素(IFN-I)介导的自身免疫的原型 疾病，以无数的临床表现为特征，包括炎症、致病性自身抗体 生产和不可逆转的终末器官损害(例如肾脏)。系统性红斑狼疮对女性的影响不成比例(九倍 比男性更高)和非白人族裔。亚洲人和非裔美国人的SLE发病率更高，更多 与高加索人相比，严重的疾病表现，以及器官损害(如狼疮性肾炎)的风险更大。其他 我们已经报道了几个干扰素调节基因，包括干扰素调节因子8(IRF8)和溶质 携带者家族15 Non 4(SLC15A4)与SLE易感性有关。IFN是一类细胞因子家族，具有 在感染、癌症和自身免疫中发挥重要作用。体外证据表明，浆细胞样树突状细胞 细胞(PDCs)是产生干扰素-I的主要免疫细胞，与SLE的发生密切相关。最近 使用两种小鼠模型(IRF8-/-基因敲除和SLC15A4突变)的研究提供了pDC的直接证据 通过过量产生干扰素-I促进系统性红斑狼疮的发生。因此，IRF8和SLC15A4可能参与人类 系统性红斑狼疮进展。我们的遗传关联数据确定了几个可能的系统性红斑狼疮IRF8易感变异 (最佳p=1.2x10-22)和SLC15A4(最佳p=1.5x10-21)。然而，尽管表现强劲， 相关、特定的致病变异及其潜在的分子机制尚未确定。vbl.使用 生物信息学，我们预测了几个相关的变异体是cis-eQTL，具有调控基因的作用。 表情。利用荧光素酶和芯片定量聚合酶链式反应，我们从实验上验证了几种 预测的功能变异体。由于干扰素基因特征是系统性红斑狼疮的显著特征，我们假设 全面的跨种族图谱(TEM)，随后是功能遗传学的实验验证， 包括相关免疫细胞的遗传和表观遗传编辑，将确定因果变异及其功能 干扰素-I生产的后果。我们的研究团队拥有专业知识、资源和跟踪记录来发现 并描述了SLE的功能变异。在目标1中，我们将本地化这些易患SLE的变体 通过在不同种族的人群中执行基于综合归因的透射电子显微镜来检测基因(N&GT；30,000 来自亚洲人、非裔美国人、欧洲裔美国人、埃及人和西班牙裔美国人)。前景看好的变种， 特别是输入和低频率变异，将通过额外的验证性基因分型进行验证。我们 将通过评估与临床亚型和临床表型的相关性来阐明遗传和临床的异质性 自身抗体。在目标2中，我们将通过实验验证假定的变体作为调节器的功能相关性 基因表达和干扰素-I的产生。我们将同时使用培养细胞(THP-1)和原代免疫细胞 (pDC，单核细胞)来自SLE患者和正常对照。从这个项目中获得的见解将有助于定义 风险等位基因如何易患系统性红斑狼疮的分子机制，并可能定义新的药物/治疗 未来系统性红斑狼疮的靶点。

ABSTRACT Systemic lupus erythematosus (SLE or lupus) is a prototype of type-I interferon (IFN-I)-mediated autoimmune disease, characterized by a myriad of clinical manifestations including inflammation, pathogenic autoantibody production, and irreversible end-organ damage (e.g. kidneys). SLE disproportionately afflicts women (nine-fold higher than men) and non-white ethnicities. Asians and African-Americans have higher SLE incidence, more severe disease manifestations, and greater risk of organ damage (e.g. lupus nephritis) than Caucasians. Others and we have reported that several IFN regulatory genes, including IFN regulatory factor 8 (IRF8), and solute carrier family 15 number 4 (SLC15A4) are involved in SLE susceptibility. IFNs are a family of cytokines with important roles in infection, cancer, and autoimmunity. In vitro evidence suggests that plasmacytoid dendritic cells (pDCs), the principal IFN-I producing immune cells, are intimately involved in SLE development. A recent study using two mouse models (Irf8-/- knockout and Slc15a4 mutant) provided direct evidence that pDCs contribute to SLE via hyper-production of IFN-I. Thus, IRF8 and SLC15A4 could potentially be involved in human SLE progression. Our genetic association data identified several potential SLE predisposing variants for IRF8 (best p=1.2x10-22) and for SLC15A4 (best p=1.5x10-21) across multiple ethnicities. However, despite strong association, specific pathogenic variants and their underlying molecular mechanisms are not yet defined. Using bioinformatics, we predicted that several associated variants are cis-eQTLs, with roles in regulating gene expression. Using luciferase and ChIP-qPCR, we experimentally validated allele-specific effects of several predicted functional variants. Since IFN gene signatures are a prominent feature in SLE, we hypothesize that comprehensive, trans-ethnic mapping (TEM) followed by experimental validation with functional genetics, including genetic and epigenetic editing in relevant immune cells, will identify causal variants and their functional consequences in IFN-I production. Our research team has the expertise, resources, and track records to discover and characterize functional variants for SLE. In Aim 1, we will localize SLE-predisposing variants from these genes by performing comprehensive imputation-based TEM across ethnically diverse populations (N>30,000 from Asians, African-Americans, European-Americans, Egyptians, and Hispanics). Promising variants, especially imputed and low-frequency variants, will be validated through additional confirmatory genotyping. We will elucidate genetic and clinical heterogeneity by assessing associations with clinical sub-phenotypes and autoantibodies. In Aim 2, we will experimentally validate functional relevance of putative variants as regulators of gene expression and IFN-I production. We will use both cultured cells (THP-1) and primary immune cells (pDCs, monocytes) from SLE patients and controls. Insights gained from this project will help to define the molecular mechanisms underlying how risk alleles predispose to SLE, and may define novel drug/therapeutic targets for SLE in the future.