Regulation of T cell anergy by palmitoyl acyl transferases

棕榈酰酰基转移酶对 T 细胞无反应性的调节

• 批准号: 7571748
• 负责人: AMNON ALTMAN
• 金额: $ 22.88万
• 依托单位: LA JOLLA INSTITUTE FOR IMMUNOLOGY
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-01-01 至 2010-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7571748
• 关键词: Accounting; Acylation; Adaptor Signaling Protein; Address; Antibodies; Antigens; Area; Autoimmunity; Biological; Cell membrane; Cell physiology; Cysteine; Defect; Development; Drug Delivery Systems; Ectopic Expression; Enzymes; Event; Family; Family member; Future; Immune System Diseases; Immune Tolerance; Immune response; Immune system; Interleukin-2; Label; Mediating; Membrane Microdomains; Metabolic; Methods; Mitogen-Activated Protein Kinases; Molecular Profiling; Mus; Palmitates; Pathway interactions; Peripheral; Play; Process; Proteins; Receptor Signaling; Regulation; Relative (related person); Rest; Role; Signal Pathway; Signal Transduction; Signaling Molecule; Signaling Protein; Small Interfering RNA; Substrate Specificity; System; T cell regulation; T-Cell Activation; T-Cell Development; T-Cell Receptor; T-Lymphocyte; Testing; Time; Transcription Factor AP-1; Transferase; Transplantation; Tumor Immunity; anergy; base; clinically relevant; clinically significant; driving force; expression vector; functional status; knock-down; mRNA Expression; novel; palmitoylation; prevent; public health relevance; receptor

DESCRIPTION (provided by applicant): T cell anergy has important implications in autoimmunity, transplantation and tumor immunity. Recently, we identified a novel TCR signaling defect in antigen-specific anergic T cells, namely, impaired palmitoylation and, consequently, lipid raft localization and function, of linker for activation of T cells (LAT). Proper localization and function of other T cell signaling proteins is also critically dependent on their palmitoylation. Although protein palmitoylation has been known for >30 years, its mechanistic basis and regulation has, until very recently, been poorly understood. The recent discovery of a novel, large (23 member) family of mammalian protein palmitoyl acyl transferases (PATs) represents a major breakthrough in this area. This advance and our more recent preliminary evidence that defective LAT palmitoylation may actually cause anergy serve as a driving force for this project. We propose to conduct exploratory/developmental studies in order to characterize the expression of PATs in T lymphocytes, identify LAT-reactive PAT(s), and analyze their potential role in T cell responsiveness and anergy. In Aim 1, we will use quantitative real-time PCR to determine the mRNA expression profile of 23 known mouse PATs in resting, activated and anergic T cells, and confirm expression of T cell-expressed PAT mRNAs at the protein level. In Aim 2, we will use expression vectors of all T cell-expressed PATs, and employ an established ectopic expression system in order to screen the ability of these PATs to enhance the palmitoylation of LAT. Identified LAT-reactive PATs will be further tested for their ability to palmitoylate a limited set of other palmitoylation substrates as a test of their relative substrate specificity. In Aim 3, we will select the best candidates PATs identified in previous aims for further functional analysis. Specifically, we will use a novel, highly efficient method for siRNA delivery in order to knock-down the expression of selected PATs, and analyze the effects of PAT silencing on the functional status of T cells, with emphasis on determining whether silencing of LAT-reactive PATs will induce a functional state resembling T cell anergy. Understanding the mechanism that underlies defective LAT palmitoylation and its association with T cell anergy may provide critical clues on how reversible protein palmitoylation regulates key aspects of T cell fate and function, and potentially implicate substrate-selective PATs as future drug targets in autoimmunity and other immunological diseases. PUBLIC HEALTH RELEVANCE: Anergy is an important form of immune tolerance, in which T lymphocytes of the immune system, despite being able to recognize an antigen, cannot generate an effective immune response to it. Strategies aimed at inducing or preventing T cell anergy have significant implications in autoimmunity, transplantation, and tumor immunity. Here we will investigate a novel pathway for anergy induction discovered by us, which consists of defective processing and function of a key molecule, LAT, which is essential for the development and proper function of T lymphocytes.

描述（由申请人提供）：T细胞消极对自身免疫，移植和肿瘤免疫具有重要意义。最近，我们确定了抗原特异性厌氧T细胞中的一种新型TCR信号缺陷，即棕榈酰化受损，因此，脂质筏定位​​和功能，用于激活T细胞（LAT）的链接器。其他T细胞信号蛋白的适当定位和功能也取决于其棕榈酰化。尽管蛋白质棕榈酰化已有30年了，但其机械基础和调节直到最近才尚未了解。最近发现了一个新颖的大型（23个成员）哺乳动物棕榈酰酰基转移酶（PATS）家族，这是该地区的重大突破。这一进步和我们最近的初步证据表明，有缺陷的拉棕榈酰化实际上可能导致反应成为该项目的驱动力。我们建议进行探索性/发育研究，以表征PAT在T淋巴细胞中的表达，鉴定LAT反应性PAT（S），并分析其在T细胞反应性和不良反应性中的潜在作用。在AIM 1中，我们将使用定量的实时PCR来确定在静止，激活和厌氧的T细胞中23个已知小鼠PAT的mRNA表达谱，并确认T细胞表达的PAT mRNA在蛋白质水平上的表达。在AIM 2中，我们将使用所有T细胞表达PAT的表达向量，并采用已建立的异位表达系统，以筛选这些PAT增强LAT的棕榈酰化的能力。确定的LAT反应性PAT将进一步测试其棕榈酰基含有有限的其他棕榈酰化底物的能力，以测试其相对底物特异性。在AIM 3中，我们将选择以前目标中确定的最佳候选人PATS进行进一步的功能分析。具体而言，我们将使用一种新型的高效方法进行siRNA递送，以敲除选定的PAT的表达，并分析PAT沉默对T细胞功能状态的影响，重点是确定LAT反应性PATS的沉默是否会引起功能性状态状态状态，类似于T细胞的功能状态。了解底层有缺陷的LAT棕榈酰化的机制及其与T细胞的关联可能会为可逆蛋白质棕榈酰化如何调节T细胞命运和功能的关键方面提供关键的线索，并可能将底物选择性PAT作为自身免疫性和其他免疫学疾病中未来的药物靶标。公共卫生相关性：反对是免疫耐受性的一种重要形式，其中免疫系统的T淋巴细胞尽管能够识别抗原，但无法产生对其有效的免疫反应。旨在诱导或预防T细胞厌食的策略对自身免疫，移植和肿瘤免疫具有重要意义。在这里，我们将研究我们发现的一种新颖的无反抗诱导途径，该途径包括关键分子LAT的处理和功能有缺陷，这对于T淋巴细胞的发育和正常功能至关重要。