MOLECULAR MECHANISMS OF REGULATORY T CELL DEVELOPMENT

调节 T 细胞发育的分子机制

• 批准号: 6896917
• 负责人: Alexander Y Rudensky
• 金额: $ 37.1万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-06-01 至 2009-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6896917
• 关键词: biomarker; cell differentiation; chimeric proteins; developmental immunology; gene expression; gene expression profiling; genetic regulation; genetically modified animals; green fluorescent proteins; helper T lymphocyte; immune tolerance /unresponsiveness; laboratory mouse; messenger RNA; organ culture; protein structure function; thymus; transcription factor

DESCRIPTION (provided by applicant): Naturally arising regulatory CD25+ CD4 T cells play an essential role in regulation of immune homeostasis. However, specific molecular mechanisms regulating their differentiation and activity have been largely unknown. We have found that forkhead transcription factor Foxp3 is specifically expressed in regulatory CD4 T cells and is required for their development and function. Although identification of Foxp3 as a critical molecular mechanism establishing regulatory T cell differentiation program is a major breakthrough, it remains unknown what are Foxp3 target genes essential for this program. In addition, our findings raise an important question as to why there is such a discrepancy in the severity and onset of the autoimmune syndrome in a genetic model of CD25+ CD4 regulatory T cell deficiency in Foxp3 deficient mice and in murine models relying on elimination of CD25+ CD4 T cells. We propose several possible non-mutually exclusive hypotheses to explain this apparent controversy: a) In addition to CD25+ CD4 T cells, Foxp3 is expressed by an unidentified subset(s) of CD25- CD4 or CD8 T cells with the regulatory function capable of providing some measure of protection from autoimmunity in mice subjected to acute depletion of CD25+ CD4 T cells; b) In the absence of Foxp3, T cells with an increased TCR affinity for self peptide/MHC class II complexes that normally develop into regulatory CD25+ CD4 T cells instead become pathogenic and largely responsible for the rapid disease progression; c) A subset of CD25+ CD4 T cells with autoreactive TCR lack Foxp3 and may represent a major population of autoreactive T cells normally controlled by Foxp3 expressing CD25+ CD4 regulatory T cells; d) Foxp3 expressed at a low level in nonregulatory CD4 T cells controls thresholds of activation, dependence on co-stimulation, or anergy induction. In this proposal we will address the aforementioned questions using genetic approaches including Foxp3 protein and mRNA reporter mice, conditional deletion and induced expression of Foxp3 gene in vivo and in vitro in combination with functional and molecular analyses. The following specific Aims will be pursued: 1. To investigate Foxp3 protein and mRNA expression and their regulation during thymic development (Aim 1a) and a role for Foxp3 in negative selection of T cells in the thymus and to evaluate autoreactivity of T cells with the Foxp3 deletion (Aim 1b). 2. To investigate Foxp3 protein and mRNA expression and to assess regulatory function of distinct Foxp3+ T cell subsets (Aim2a); to investigate Foxp3 protein and mRNA regulation in peripheral T cells and to explore, thereby, possible generation of Foxp3+ T cells in the periphery (Aim 2b); to test a requirement for Foxp3 expression in maintaining regulatory T cell function (Aim 2c). 3. To investigate potential cell-intrinsic role of Foxp3 in controlling activation, central and peripheral tolerance induction in non-regulatory CD25- CD4 T cells. 4. To identify molecular targets of the Foxp3 gene involved in regulation of regulatory T cell development and function.

描述（由申请人提供）：自然出现的调节性CD25+ CD4 T细胞在调节免疫稳态中起着至关重要的作用。但是，调节其分化和活性的特定分子机制在很大程度上未知。我们发现，叉子转录因子FOXP3在调节性CD4 T细胞中特异性表达，其发育和功能是必需的。尽管将FOXP3鉴定为建立调节T细胞分化程序的关键分子机制是一个主要突破，但对于该程序来说，FOXP3靶基因是什么是未知的。此外，我们的发现提出了一个重要的问题，即为什么在FOXP3不足小鼠的CD25+ CD4调节T细胞缺乏症的遗传模型以及依赖CD225+ CD4 T细胞的鼠模型中，自身免疫综合征的严重程度和发作差异如此之大。我们提出了几种可能的非完全排斥的假设来解释这一明显的争议：a）除CD25+ CD4 T细胞外，FOXP3还通过CD25- CD2-CD4或CD8 T细胞的未鉴定子集表达，具有调节功能，具有能够提供某种量度的CD25+ CD25+ CD25+ CD25+ CD25+ CD4 TID的调节功能。 b）在没有FOXP3的情况下，对自肽/MHC II类配合物的TCR亲和力增加的T细胞通常会发展为调节性CD25+ CD4 T细胞，而是变为致病性，并在很大程度上造成了快速疾病进展。 c）具有自动反应性TCR的CD25+ CD4 T细胞的子集缺乏FOXP3，并且可能代表通常由FOXP3控制CD25+ CD4调节性T细胞的主要自动反应性T细胞； d）在非调节CD4 T细胞中以低水平表达的FOXP3控制激活，依赖于共刺激或厌食诱导的阈值。在此提案中，我们将使用包括FOXP3蛋白和mRNA报告小鼠在内的遗传方法，有条件的缺失以及诱导Foxp3基因在体内和体外结合功能和分子分析的诱导表达来解决上述问题。将追求以下具体目的：1。研究FOXP3蛋白和mRNA表达及其在胸腺发育过程中的调节（AIM 1A），以及FOXP3在胸腺中T细胞负选择的作用，并用FOXP3缺失评估T细胞的自动反应性（AIM 1B）。 2。研究FOXP3蛋白和mRNA表达并评估不同FOXP3+ T细胞子集的调节功能（AIM2A）；研究周围T细胞中FOXP3蛋白和mRNA调节，并探索周围可能产生的Foxp3+ T细胞（AIM 2B）；在维持调节性T细胞功能时测试FOXP3表达的需求（AIM 2C）。 3。研究FOXP3在非调节CD25-CD4 T细胞中控制激活，中心和外周耐受诱导中的潜在细胞中性作用。 4。确定与调节T细胞发育和功能调节有关的FOXP3基因的分子靶标。