Genetic studies linking LSP1 function in T cells to Inflammatory Bowel Disease

T 细胞中 LSP1 功能与炎症性肠病相关的遗传学研究

基本信息

批准号：
10636526
负责人：
BENJAMIN JOACHIM SCHMIEDEL
金额：
$ 40.26万
依托单位：
LA JOLLA INSTITUTE FOR IMMUNOLOGY
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-04-01 至 2028-01-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10636526
关键词：
Address Affect Apoptosis Arthritis Autoimmune Diseases Automobile Driving Binding Biological Assay Biological Sciences CD4 Positive T Lymphocytes CRISPR interference Cell physiology Cells Clinical Clustered Regularly Interspaced Short Palindromic Repeats Colitis Collaborations Colonic inflammation Crohn&apos s disease DNA Sequence Databases Delayed Hypersensitivity Disease Disease Progression Enhancers F-actin-binding proteins Gene Expression Genes Genetic Genetic Polymorphism Genetic study Human Immune Immunology Impairment Infiltration Inflammation Inflammatory Inflammatory Bowel Diseases Inflammatory Response Inherited Knowledge Leukocytes Link Luciferases Mediating Modeling Molecular Morbidity - disease rate Mus Outcome Pathogenesis Pathogenicity Pathologic Pathway interactions Patients Play Production Proliferating Proteins Quantitative Trait Loci RNA Interference Regulation Reporter Reporting Rest Risk Role Science Signal Pathway Small Interfering RNA T cell therapy T-Cell Activation T-Lymphocyte T-Lymphocyte Subsets Testing Translating Ulcerative Colitis Untranslated RNA Variant Work causal variant cell motility cell type chromatin immunoprecipitation cytokine cytotoxicity disorder risk epigenomics genetic association genetic variant genome wide association study improved insight loss of function mortality mouse model murine colitis new therapeutic target overexpression phase 3 study prevent programs promoter response restraint risk variant therapeutic target transcription factor tumor

项目摘要

PROJECT SUMMARY/ABSTRACT Inflammatory bowel diseases (IBD) cause substantial mortality and morbidity. Current treatments that block pathological inflammatory responses have improved clinical outcomes in some patients but they are not highly effective in modifying disease progression or preventing relapses. Hence, there is a large unmet need to develop novel therapeutic targets. Genome-wide association studies (GWAS) offer an unbiased approach to identify therapeutic targets in relevant immune cell types such as CD4+ T cells that play key roles in IBD pathogenesis. Because T cells are quiescent in the absence of extrinsic stimulation, it is not possible to fully examine the effects of disease-risk variants on functionally relevant effector genes under resting conditions. To identify IBD-risk genes in activated CD4+ T cells, we performed the first large-scale single-cell eQTL study on activated CD4+ T cells. We found that reduced expression of Leukocyte-specific protein 1 (LSP1), specifically in activated CD4+ T-cell subsets such as TH1 and TH17 cells, was associated with the risk of IBD. In this R01 proposal, we will investigate how reduced levels of LSP1 influences the differentiation and function of CD4+ T cells to drive disease pathogenesis, and will test the hypothesis that LSP1 plays a key role in restraining the re-programming of CD4+ T cells into a more pathogenic cell state in IBD patients. In Aim 1, we will determine the functional IBD-risk variants that reduce LSP1 expression in CD4+ T cells. We will employ luciferase reporter assays to determine functional variants in the enhancers and promoter of LSP1, perform CRISPR-mediated editing of prioritized IBD-risk eQTLs to define causal variants, and perform CRISPRi and ChIP assays to determine functional enhancers, relevant up-stream regulators and whether the functional LSP1 eQTLs directly perturb the binding of key transcription factors that modulate LSP1 expression. In Aim 2, we will determine the role of LSP1 in reprogramming of CD4+ T cells into the pathogenic state observed in IBD. To determine whether LSP1 influences the differentiation and pathogenic function of CD4+ T cells from healthy and IBD donors, we will reduce and increase LSP1 levels and assess the effects on CD4+ T-cell activation, apoptosis, proliferation and proinflammatory cytokine production. In an adoptive T cell transfer model of colitis, we will compare the ability of Lsp1- sufficient (wild-type) and Lsp1-deficient CD4+ T cells in driving colonic inflammation and pathogenic TH1 and TH17 differentiation. We will determine whether reducing Lsp1 expression in CD4+ T cells enhances their pathogenicity in mouse models of colitis, thus implicating an important T cell-intrinsic role for LSP1 in IBD pathogenesis. Overall, this study, examining the function, expression, activation and regulation of LSP1 in CD4+ T cells, will provide important mechanistic insights into the genetic basis of risk for inflammatory bowel disease.

项目摘要/摘要炎症性肠道疾病（IBD）导致大量死亡率和发病率。当前的治疗方法病理炎症反应改善了某些患者的临床结局，但并不高有效改变疾病进展或预防复发。因此，有很大的未满足需要发展新型治疗靶标。全基因组协会研究（GWAS）提供了一种公正的方法来识别相关免疫细胞类型的治疗靶标，例如在IBD发病机理中起关键作用的CD4+ T细胞。由于在没有外部刺激的情况下T细胞是静止的，因此无法完全检查影响疾病风险在静止条件下功能相关效应基因的变体的变体。识别IBD风险激活的CD4+ T细胞中的基因，我们对激活的CD4+ T进行了首个大型单细胞EQTL研究细胞。我们发现白细胞特异性蛋白1（LSP1）的表达降低，特别是在活化的CD4+中 T细胞子集（例如Th1和Th17细胞）与IBD的风险有关。在此R01提案中，我们将研究LSP1水平降低如何影响CD4+ T细胞的分化和功能驱动疾病发病机理，并将检验以下假设：LSP1在限制CD4+的重新编程中起关键作用在IBD患者中，T细胞进入更致病的细胞状态。在AIM 1中，我们将确定降低CD4+ T细胞中LSP1表达的功能性IBD风险变体。我们将采用荧光素酶报告基因测定法确定LSP1的增强子和启动子中的功能变体，进行CRISPR介导的优先级IBD风险EQTL的编辑以定义因果变体，并执行CRISPRI 和CHIP分析以确定功能增强器，相关的上流调节器以及功能是否功能 LSP1 EQTL直接扰动调节LSP1表达的关键转录因子的结合。在AIM 2中，我们将确定LSP1在CD4+ T细胞重编程中的作用在IBD中。确定LSP1是否影响CD4+ T细胞的分化和致病功能健康和IBD供体，我们将降低和增加LSP1水平，并评估对CD4+ T细胞的影响激活，凋亡，增殖和促炎细胞因子产生。在收养T细胞转移模型中结肠炎，我们将比较LSP1足够（野生型）和LSP1缺乏的CD4+ T细胞的能力结肠炎症以及致病性TH1和Th17分化。我们将确定是否减少LSP1 在CD4+ T细胞中的表达增强了结肠炎小鼠模型中的致病性，因此暗示了重要的 LSP1在IBD发病机理中的T细胞中性作用。总体而言，这项研究，检查了CD4+ T细胞中LSP1的功能，表达，激活和调节为炎症性肠病风险的遗传基础提供重要的机械见解。