Enhancer-based Immune and Beta Cell Dysregulation Underlying T1D Risk

基于增强剂的免疫和 β 细胞失调是 T1D 风险的基础

• 批准号: 10442605
• 负责人: Dieter Meinrad Egli
• 金额: $ 110.06万
• 依托单位: COLUMBIA UNIVERSITY HEALTH SCIENCES
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-15 至 2024-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10442605
• 关键词: Affect; Alleles; Area; Autoimmune Diseases; Base Pairing; Beta Cell; Binding; Biological Assay; Biology; CRISPR/Cas technology; Cell Nucleus; Cells; Chromatin; Clinical Research; Communities; Computing Methodologies; Coupled; Data; Data Set; Defect; Detection; Development; Disease; Enhancers; Event; Gene Expression; Genes; Genetic Risk; Genetic Transcription; Genomic Segment; Genomics; Homeostasis; Immune; Individual Differences; Inflammation; Inflammatory; Insulin; Insulin-Dependent Diabetes Mellitus; Islets of Langerhans; Lead; Maps; Measures; Mediating; Molecular; Mutation; Organ; Output; Pancreas; Pathologic; Patients; Phenotype; Play; Point Mutation; Predisposition; Preventive; RNA; Regulator Genes; Regulatory Element; Research; Research Design; Resolution; Resources; Response Elements; Role; Seminal; Signal Pathway; Signal Transduction; System; T-Lymphocyte; Technology; Therapeutic; Transcription Initiation Site; Translating; Ultrafine; Untranslated RNA; Validation; Variant; base; causal variant; cell type; clinical application; cytokine; diabetes risk; genetic association; genome editing; genome wide association study; genome-wide; monocyte; promoter; response; risk variant; skills; stem cells; transcription factor; transcriptome sequencing

Abstract Type 1 diabetes (T1D) is an organ-specific autoimmune disease, whereby immune cell-mediated and inflammatory cytokines lead to loss of the insulin-producing β cells in the pancreas. Genome-wide association studies (GWAS) have identified ~60 genomic regions associated with T1D risk. The vast majority of GWAS risk variants associated with T1D reside in non-coding regions, particularly enhancers, suggesting that gene regulatory changes substantially contribute to inter-individual differences in susceptibility to T1D. Driven by our T1D GWAS annotation data, highlighting the importance of a systematic analysis of both immune and β cell systems of both T1D patients and controls, we propose to identify causal enhancer variants and causal target genes using contemporary computational methods and cutting-edge global genomics. We will perform our PRO-cap and PRO-seq assays to comprehensively identify active enhancers harboring T1D-associated variants in T cells, monocytes and stem cell derived β cells (sc-β cells) from T1D patients and controls, followed by validation of active enhancers harboring T1D-associated variants using our eSTARR-seq assays (Aim 1). We will perform our Tri-HiC assays to profile the enhancer-promoter interactomes at unprecedented high resolution in primary T cells, primary monocytes and sc-β cells from T1D patients and controls as well as pancreatic islets to comprehensively identify target genes of T1D-associated variants, and refine targets of T1D-associated variants with T1D-sepcific alteration in target gene expression in each cell type using our coupled single cell nucleus (sn) RNA-seq and snATAC-seq assays (Aim 2). We will validate targets of T1D-associated variants in T cells, monocytes and β cells using CRISPR/Cas9 endogenous enhancer mutational strategies in the context of the endogenous chromatin landscape in each cell type. Our analytical and experimental framework represents an exciting new paradigm for studying T1D, and the subsequent clinical research based on our results has the potential to develop preventive and therapeutic strategies against T1D. The data sets generated by these studies will represent important, but currently lacking, resources for the T1D research community.

摘要 1型糖尿病（T1 D）是一种器官特异性自身免疫性疾病，其中免疫细胞介导和 炎症细胞因子导致胰腺中产生胰岛素的β细胞损失。全基因组关联 GWAS研究已经确定了约60个与T1 D风险相关的基因组区域。绝大多数GWAS风险 与T1 D相关的变异存在于非编码区，特别是增强子，这表明基因 调节变化实质上促成了T1 D易感性的个体间差异。感召下 T1 D GWAS注释数据，强调了免疫和β-半乳糖苷酶系统分析的重要性。 T1 D患者和对照的细胞系统，我们建议鉴定因果增强子变体和因果增强子变体。 利用当代计算方法和尖端的全球基因组学来靶向基因。我们将执行我们的 PRO-cap和PRO-seq测定法用于全面鉴定携带T1 D相关变体的活性增强子 在来自T1 D患者和对照的T细胞、单核细胞和干细胞衍生的β细胞（sc-β细胞）中，随后 使用我们的eSTARR-seq测定验证携带T1 D相关变体的活性增强子（Aim 1）。我们 将进行我们的Tri-HiC分析，以前所未有的高分辨率分析增强子-启动子相互作用组 在来自T1 D患者和对照以及胰岛的原代T细胞、原代单核细胞和sc-β细胞中， 全面鉴定T1 D相关变异的靶基因，并完善T1 D相关变异的靶基因， 使用我们的偶联单细胞，在每种细胞类型中靶基因表达具有T1 D特异性改变的变体 核（sn）RNA-seq和snATAC-seq测定（Aim 2）。我们将验证T1 D相关变体的靶点， 在背景下使用CRISPR/Cas9内源性增强子突变策略的T细胞、单核细胞和β细胞 每种细胞类型的内源性染色质景观。我们的分析和实验框架 一个令人兴奋的研究T1 D的新范例，以及基于我们结果的后续临床研究， 开发针对T1 D的预防和治疗策略的潜力。这些研究产生的数据集 将是T1 D研究界重要但目前缺乏的资源。