Cis Regulatory Elements and Systemic Lupus Erythematosus

项目总结/摘要 系统性红斑狼疮（SLE，lupus）是一种慢性炎症性自身免疫性疾病， 遗传成分。全基因组关联研究（GWAS）已经确定了许多相关的遗传 SLE患者中的变异。几乎所有的都在编码区之外，在可能富含监管和 转录功能变体。T滤泡辅助细胞（Tfh）和生殖中心B（GCB）细胞是主要的 SLE中系统性自身免疫的贡献者，通过它们协同产生致病性 自身抗体和随后的免疫复合物介导的组织损伤。我们的初步和已发表的研究 表明Tfh和GCB细胞中增强子和Bcl 6占据的位点（B细胞淋巴瘤6）， 这些细胞类型的分化和功能所必需的典型转录抑制因子，共定位 与SLE相关的基因变异本修订申请的目标之一是确定增强剂 Tfh和GCB细胞中控制基因表达程序的元件。我们将定义和关联染色质 结构，RNA聚合酶II占有率和基因组组织与转录组分析，以确定 在原代人Tfh和GCB细胞中的增强子。在增强子识别和验证之后，我们将整合 我们的数据，以确定和相关的细胞类型特异性增强基因表达的程序。的目标 目的二是鉴定由Bcl 6及其转录阻遏物介导的共同调控网络， Blimp 1（B淋巴细胞诱导成熟蛋白-1），在Tfh和GCB细胞中。整合转录组数据， 基因组组织、组蛋白结构以及Bcl 6和Blimp 1的占有率将为我们提供详细的知识 Tfh和GCB细胞中由这些转录抑制因子控制的基因结构、功能和调节。 这些数据集中的基因组调控将被比较和对比，使我们能够识别 并表征由Blimp 1-Bcl 6轴控制的常见调节网络。目标三的目标是 Tfh和GCB细胞中与SLE相关的功能性遗传变异体的鉴定和表征。 SLE连锁遗传变异将与转录组分析、增强子图谱、Bcl 6和Blimp 1整合 占据、染色质可及性测定和基因型特异性基因表达来鉴定功能性SNP。 将进行相关增强子的功能研究，包括使用以下技术在小鼠中进行体内基因编辑研究： CRISPR-Cas9技术对鼠基因组中的候选区域进行遗传修饰，以评估 这些元件在Tfh和GCB细胞分化和功能中的调节作用。功能识别 遗传变异将促进SLE患者新治疗策略的开发。

Project Summary/Abstract Systemic lupus erythematosus (SLE, lupus) is a chronic inflammatory autoimmune disorder with a significant genetic component. Genome wide association studies (GWAS) have identified many associated genetic variants in SLE patients. Almost all are outside coding regions, in areas likely enriched for regulatory and transcriptionally functional variants. T follicular helper cells (Tfh) and germinal center B (GCB) cells are major contributors to systemic autoimmunity in SLE through their collaborative production of pathogenic autoantibodies and subsequent immune-complex mediated tissue injury. Our preliminary and published studies indicate that enhancers in Tfh and GCB cells and sites of occupancy by Bcl6 (B cell lymphoma 6), the canonical transcriptional repressor necessary for differentiation and function of these cell types, co-localize with gene variants associated with SLE. The goal of aim one of this revised application is to identify enhancer elements controlling gene expression programs in Tfh and GCB cells. We will define and correlate chromatin architecture, RNA polymerase II occupancy, and genomic organization with transcriptome analyses to identify enhancers in primary human Tfh and GCB cells. After enhancer identification and validation, we will integrate our data to identify and correlate cell-type specific enhancers with programs of gene expression. The goal of aim two is the identification of common regulatory networks mediated by Bcl6 and its transcriptional repressor, Blimp1 (B lymphocyte-induced maturation protein-1), in Tfh and GCB cells. Integration of transcriptome data, genomic organization, histone architecture, and Bcl6 and Blimp1 occupancy will provide us detailed knowledge of gene structure, function, and regulation controlled by these transcriptional repressors in Tfh and GCB cells. The regulation of groups of genes in these data sets will be compared and contrasted, allowing us to identify and characterize common regulatory networks controlled by the Blimp1-Bcl6 axis. The goal of aim three is the identification and characterization of functional genetic variants associated with SLE in Tfh and GCB cells. SLE-linked genetic variants will be integrated with transcriptome analyses, enhancer maps, Bcl6 and Blimp1 occupancy, chromatin accessibility assays, and genotype-specific gene expression to identify functional SNPs. Functional studies of relevant enhancers will be performed including in vivo gene editing studies in mice using CRISPR-Cas9 technology to genetically modify candidate regions in the murine genome to assess the regulatory effects of these elements in Tfh and GCB cell differentiation and function. Identification of functional genetic variants should facilitate development of novel therapeutic strategies for use in SLE patients.