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Systematic Mapping of the Functional Common Noncoding Variants in the TNFAIP3 Locus

TNFAIP3 基因座功能性常见非编码变异的系统作图

基本信息

批准号：
9451927
负责人：
John Philip Ray
金额：
$ 6.12万
依托单位：
BROAD INSTITUTE, INC.
依托单位国家：
美国
项目类别：
财政年份：
2017
资助国家：
美国
起止时间：
2017-02-01 至 2020-01-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9451927
关键词：
A20 protein Affect Alleles Animal Disease Models Autoimmune Diseases Autoimmune Process Autoimmunity B-Lymphocytes Binding Biochemical Biological Assay CRISPR/Cas technology Cells Chromatin Complex Data Diagnostic Factor Disease Disease model Enhancers Environment Epigenetic Process Future Gene Expression Gene Expression Profile Genes Genetic Genetic Engineering Genetic Predisposition to Disease Genetic Variation Genome Genome engineering Genomics Goals Haplotypes Heritability Human Human Genetics Immune Knowledge Learning Linkage Disequilibrium Maps Methods Mus Mutation Nature Nucleic Acid Regulatory Sequences Pathogenesis Pathogenicity Patients Population Protein Isoforms RNA Splicing Regulation Regulator Genes Reporter Research Risk Site Syndrome Systemic Lupus Erythematosus T-Lymphocyte Testing Therapeutic United States Untranslated RNA Variant Woman Work base cell type epigenetic variation genetic variant genome wide association study genome-wide genotyped patients histone modification improved in vivo insight monocyte mouse model new therapeutic target personalized medicine repaired trait transcription factor transcriptomics

项目摘要

Project Summary/Abstract The mechanisms for induction of autoimmunity remain enigmatic, in part due to the unconfirmed activity of many autoimmune-associated noncoding common genetic variants. To dissect the specific mechanisms of how all common and disease-associated noncoding variation modulates gene expression, I propose to employ scalable assays in three cell types that will assess 2708 noncoding common variants for gene-modulatory function in an important autoimmune-associated locus containing the negative regulatory gene TNFAIP3/A20. One assay will test ~2700 variants for their effects on reporter expression pre- and post-stimulation. A second assay will evaluate ~43,000 systematic locus-wide deletions in the genome to determine regulatory regions (and potentially active variants within) in cell types. I will then map the active disease-associated variants from these assays to the open chromatin of autoimmune patient cell types to create a ranked list of those that putatively operate in disease. I will then determine the mechanism of action for these active, open chromatin localized variants through genetically engineering risk and non-risk alleles using CRISPR-Cas9 homology directed repair and performing biochemical assays. This method is scalable to study many loci in future work, which will contribute to our knowledge of disease gene networks, accurate animal disease models, and more efficacious therapeutics.

项目总结/摘要诱导自身免疫的机制仍然是个谜，部分原因是许多免疫调节剂的活性未经证实。自身免疫相关的非编码常见遗传变异。为了剖析所有的常见的和疾病相关的非编码变异调节基因表达，我建议采用可扩展的在三种细胞类型中进行的检测，将评估2708种非编码常见变异体的基因调节功能，重要的自身免疫相关基因座，包含负调控基因TNFAIP 3/A20。一种测定将测试约2700种变体对刺激前和刺激后报告基因表达的影响。第二次检测将评估基因组中约43，000个系统性基因座范围缺失，以确定调控区域（和潜在活性区域）细胞类型中的变异）。然后，我将从这些检测中绘制出与疾病相关的活跃变异体，自身免疫患者细胞类型的染色质，以创建在疾病中起作用的那些染色质的排名列表。我会然后通过遗传学方法确定这些活跃的开放染色质定位变体的作用机制，使用CRISPR-Cas9同源定向修复工程化风险和非风险等位基因，并进行生物化学分析。这种方法是可扩展的，以研究许多基因座在未来的工作，这将有助于我们的知识，疾病基因网络、精确的动物疾病模型和更有效的治疗方法。