权益分类	功能权益	普通用户	{{item.name}}会员
{{category.name}}	{{benefitItem.name}}

Integrated high-throughput functional assays to identify ulcerative colitis causal risk variants

综合高通量功能测定，以确定溃疡性结肠炎的因果风险变异

基本信息

批准号：
10391446
负责人：
ROBERT J. KLEIN
金额：
$ 65.71万
依托单位：
CORNELL UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2018
资助国家：
美国
起止时间：
2018-08-01 至 2024-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10391446
关键词：
Affect Automobile Driving Binding Biological Assay Biological Models CRISPR/Cas technology Chromosomes Code Colon Colorectal Complex Crohn&apos s disease Data Diagnosis Disease Enhancers Epigenetic Process Epithelial Epithelial Cells Family Gene Expression Gene Frequency Genes Genetic Genetic Diseases Genetic Risk Genetic Transcription Genotype Goals Haplotypes Human Individual Inflammatory Bowel Diseases Inherited Knowledge Lead Letters Libraries Linkage Disequilibrium Luciferases Mediating Meta-Analysis Methodology Missense Mutation Mutagenesis Mutation National Institute of Diabetes and Digestive and Kidney Diseases Natural Immunity Nucleic Acid Regulatory Sequences Organoids Parents Pathway Analysis Patients Persons Promoter Regions Proteins Recording of previous events Regulator Genes Regulatory Element Relative Risks Resources Risk Risk Factors Single Nucleotide Polymorphism Structure Susceptibility Gene Testing Transcript Ulcerative Colitis Untranslated RNA Variant Work base causal variant epigenomics follow-up genetic variant genome editing genome wide association study improved in vivo evaluation innovation insight interest novel protein protein interaction risk variant screening therapy development tool trait transcription factor transcriptome sequencing transcriptomics

项目摘要

SUMMARY Ulcerative colitis (UC) affects over 1 million people worldwide. Relative risk is increased by >13.8 fold for close relatives of affected individuals. Genome-wide association studies (GWAS), including a meta-analysis lead by Co-I Cho, have identified 133 UC- or shared Crohn’s Disease (CD)/UC-associated loci. Due to haplotype structure, GWAS usually nominate clusters of large numbers of single nucleotide polymorphisms (SNPs) in linkage disequilibrium (LD), making it difficult to distinguish causal vs neutral flanking SNPs in LD. Furthermore, most of these SNPs are in non-coding regions, making functional interpretation even more difficult due to incomplete knowledge of non-coding regulatory elements. Here, we test the hypothesis that genetic risk to UC is mediated through colon-intrinsic mechanisms. The overall goal of this study is to rationally select the causal UC-associated non-coding and coding variants active in normal colon epithelial cells. We have computationally identified 1,407 GWAS UC-associated non-coding variants mapping to enhancer/promoter regions active in the colon, and 248 GWAS UC-associated missense variants. In Aim 1, we will experimentally examine each of the 1,407 candidate non-coding SNPs in colon organoids through an innovative high-throughput mutagenesis transcriptional readout pipeline integrating a novel massively parallel chromosome-integrated self-transcribing active regulatory region sequencing assay (iSTARR-seq) with the high-throughput quantitative dual luciferase assay to nominate causal UC non-coding risk variants. In Aim 2, we will experimentally examine each of the 248 candidate missense SNPs through our INtegrated PrOtein INteractome perTurbation screening (InPOINT) pipeline combining six high-throughput mutagenesis functional assays to quantify the impact of coding variants on protein stability and specific protein-protein interactions. Based on our experimental results, we will perform integrated network analysis to nominate 10 causal variant candidates (7 non-coding and 3 coding variants) for functional evaluation in vivo using CRISPR/Cas9 genome editing in colon organoids in Aim 3. Successful completion of these aims will provide important insights into the genetic mechanisms driving Ulcerative Colitis and establish a strategy broadly applicable for identifying causal variants underlying complex traits for other diseases.

概括溃疡性结肠炎 (UC) 影响着全球超过 100 万人。关闭时相对风险增加 >13.8 倍受影响个人的亲属。全基因组关联研究 (GWAS)，包括由 Co-I Cho 已鉴定出 133 个 UC 或共有克罗恩病 (CD)/UC 相关位点。由于单倍型根据结构，GWAS 通常会提名大量单核苷酸多态性 (SNP) 的簇连锁不平衡 (LD)，使得很难区分 LD 中的因果侧翼 SNP 和中性侧翼 SNP。此外，大多数 SNP 位于非编码区域，这使得功能解释更加困难由于对非编码调控元件的不完全了解，这很困难。在这里，我们检验以下假设： UC 的遗传风险是通过结肠内在机制介导的。本研究的总体目标是合理地选择在正常结肠上皮细胞中活跃的致病 UC 相关非编码和编码变体。我们通过计算确定了 1,407 个 GWAS UC 相关非编码变体映射到结肠中活跃的增强子/启动子区域，以及 248 个 GWAS UC 相关错义变异。在目标 1 中，我们将通过实验检查结肠类器官中 1,407 个候选非编码 SNP 创新的高通量诱变转录读出管道集成了新颖的大规模并行染色体整合自转录活性调控区测序分析 (iSTARR-seq) 高通量定量双荧光素酶测定可提名致病性 UC 非编码风险变异。在目标 2 中，我们将通过我们的集成蛋白对 248 个候选错义 SNP 中的每一个进行实验检查 INteractome perTurbation 筛选 (InPOINT) 流程结合了六种高通量诱变功能量化编码变体对蛋白质稳定性和特定蛋白质-蛋白质相互作用的影响的测定。根据我们的实验结果，我们将进行综合网络分析以提名 10 个因果变异使用 CRISPR/Cas9 基因组进行体内功能评估的候选者（7 个非编码变体和 3 个编码变体）目标 3 中的结肠类器官编辑。这些目标的成功完成将为我们提供重要的见解驱动溃疡性结肠炎的遗传机制，并建立广泛适用于识别因果关系的策略其他疾病复杂特征的变异。

项目成果

期刊论文数量（5）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

A comparison of experimental assays and analytical methods for genome-wide identification of active enhancers.

DOI：
10.1038/s41587-022-01211-7
发表时间：
2022-07
期刊：
NATURE BIOTECHNOLOGY
影响因子：
46.9
作者：
Yao, Li;Liang, Jin;Ozer, Abdullah;Leung, Alden King-Yung;Lis, John T.;Yu, Haiyuan
通讯作者：
Yu, Haiyuan

Advancing discovery of risk-altering variants for complex diseases by functionally informed fine-mapping.

通过功能知情的精细绘图，推进复杂疾病风险改变变异的发现。