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

基本信息

批准号：
10263319
负责人：
Dieter Meinrad Egli
金额：
$ 111.5万
依托单位：
COLUMBIA UNIVERSITY HEALTH SCIENCES
依托单位国家：
美国
项目类别：
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/10263319
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

项目摘要

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.

抽象的 1型糖尿病（T1D）是一种特定器官特异性的自身免疫性疾病，通过免疫细胞介导并炎性细胞因子导致胰腺中产生胰岛素的β细胞的损失。全基因组关联研究（GWAS）已经确定了与T1D风险相关的约60个基因组区域。绝大多数GWAS风险与T1D相关的变体位于非编码区域，尤其是增强子中，表明基因监管变化极大地导致了个体间的T1D敏感性差异。由我们的驱动 T1D GWAS注释数据，强调了对免疫和β进行系统分析的重要性 T1D患者和对照组的细胞系统，我们建议鉴定因果增强子变体和因果使用当代计算方法和尖端的全球基因组学靶向基因。我们将执行我们的亲帽和pro-seq分析，以全面识别具有T1D相关变体的活跃增强剂在T细胞中，来自T1D患者和对照的单核细胞和干细胞衍生的β细胞（SC-β细胞），然后是使用我们的ESTARR-SEQ分析（AIM 1）验证具有T1D相关变体的活性增强剂。我们将执行我们的Tri-HIC测定法以介绍前所未有的高分辨率的增强子促销相互作用在原代T细胞中，来自T1D患者和对照组的原代单核细胞和SC-β细胞以及胰岛全面识别与T1D相关变体的靶基因，并完善与T1D相关的靶标使用我们的耦合单细胞在每个细胞类型中具有T1D-SEPCIFIC变化的变体核（SN）RNA-SEQ和SNATAC-SEQ分析（AIM 2）。我们将验证与T1D相关变体的靶标 T细胞，单核细胞和β细胞在情况下使用CRISPR/CAS9内源性增强子突变策略每种细胞类型中的内源性染色质景观。我们的分析和实验框架代表一个令人兴奋的新范式用于研究T1D，以及根据我们的结果进行的随后的临床研究具有针对T1D制定预防和治疗策略的潜力。这些研究产生的数据集对于T1D研究社区来说，将代表重要但目前缺乏资源。