Regulation of CARD11 by a kinesin-like protein, GAKIN

驱动蛋白样蛋白 GAKIN 对 CARD11 的调节

• 批准号: 7406429
• 负责人: REBECCA L LAMASON
• 金额: $ 3.46万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-10-01 至 2010-09-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7406429
• 关键词: Active Sites; Aging; Antigen-Presenting Cells; Antigens; Appearance; Binding; Biological Assay; Cell Line; Cell Nucleus; Cell Proliferation; Cell physiology; Cells; Chromosome Pairing; Complex; Defect; Development; FHA Domain; Family member; Gene Expression; Glycine; Goals; Human; Image; Immune; Immune response; Immune system; Immunoprecipitation; Kinesin; Kinetics; Lead; Life; Link; Mammalian Cell; Mediating; Molecular; Motor; NF-kappa B; Pathway interactions; Phosphotransferases; Play; Point Mutation; Predisposition; Protein Kinase C; Protein Overexpression; Proteins; RNA Interference; Reagent; Receptor Signaling; Regulation; Research; Role; Signal Pathway; Signal Transduction; Signaling Protein; Synapses; System; T Cell Receptor Signaling Pathway; T-Cell Activation; T-Cell Proliferation; T-Cell Receptor; T-Lymphocyte; Testing; adapter protein; aged; cellular imaging; cofactor; design; expression cloning; immunological synapse; inhibitor/antagonist; insight; membrane-associated guanylate kinase; novel; novel therapeutics; pathogen; programs; research study; response; transcription factor; Active Sites; Aging; Antigen-Presenting Cells; Antigens; Appearance; Binding; Biological Assay; Cell Line; Cell Nucleus; Cell Proliferation; Cell physiology; Cells; Chromosome Pairing; Complex; Defect; Development; FHA Domain; Family member; Gene Expression; Glycine; Goals; Human; Image; Immune; Immune response; Immune system; Immunoprecipitation; Kinesin; Kinetics; Lead; Life; Link; Mammalian Cell; Mediating; Molecular; Motor; NF-kappa B; Pathway interactions; Phosphotransferases; Play; Point Mutation; Predisposition; Protein Kinase C; Protein Overexpression; Proteins; RNA Interference; Reagent; Receptor Signaling; Regulation; Research; Role; Signal Pathway; Signal Transduction; Signaling Protein; Synapses; System; T Cell Receptor Signaling Pathway; T-Cell Activation; T-Cell Proliferation; T-Cell Receptor; T-Lymphocyte; Testing; adapter protein; aged; cellular imaging; cofactor; design; expression cloning; immunological synapse; inhibitor/antagonist; insight; membrane-associated guanylate kinase; novel; novel therapeutics; pathogen; programs; research study; response; transcription factor

DESCRIPTION (provided by applicant): The ability of the immune system to defend against pathogens critically depends on the ability of T cells to recognize foreign antigens and respond appropriately. Upon encountering antigen, the T cell receptor (TCR) triggers intracellular signaling pathways that activate of a set of transcription factors to induce new programs of gene expression. The activation of the transcription factor NF-kB by TCR signaling is required for normal antigen-induced T cell proliferation, survival and effector function. CARD11 is a multi-domain adapter protein that is required for TCR-mediated activation of NF-kB, but its mechanisms of action are incompletely understood. To isolate CARD11 signaling cofactors, a novel expression-cloning screen was conducted to isolate modulators of CARD11 activity. From this screen, a kinesin-like protein, GAKIN, was identified as an inhibitor of CARD11, and preliminary studies suggest that GAKIN functions as an inhibitor of TCR signaling to NF-kB. The goal of the proposed research is to investigate how GAKIN functions in this important signaling pathway. The experiments are designed to test the overall hypothesis that GAKIN inhibits TCR signaling to NF-kB by binding CARD11 directly and transporting it away from the immunological synapse. The hypothesis will be tested with three specific aims. First, using GAKIN-deficient and GAKIN-overexpressing T cell lines, several steps in the TCR-to-NF-kB pathway will be assayed to determine the step at which GAKIN acts. Second, the domains of GAKIN that are required for its inhibitory activity will be determined using an RNA interference-rescue assay. Third, binding assays and T cell:APC (antigen presenting cell) imaging assays will be used to demonstrate whether or not GAKIN binds CARD11directly and influences the kinetics or extent of CARD11 recruitment to the immunological synapse. The results of these studies should expand our understanding of the molecular machinery that controls T cell activation in the adaptive immune response. In addition, this research may offer insight into the dysregulated signaling that is observed in aging T cells, which can contribute to ineffective immune responses and increased susceptibility to pathogens. Immune cell signaling defects are commonly seen in aged humans and this results in impaired responses to pathogens. The results of the proposed research have the potential to advance the understanding of the molecular machinery that immune cells use to mount an effective response. The results may also provide a target for the development of novel therapeutics that could enhance immune responses that are dysregulated in aging.

描述（由申请人提供）：免疫系统防御病原体的能力取决于T细胞识别外抗原和适当反应的能力。遇到抗原后，T细胞受体（TCR）触发细胞内信号通路，激活一组转录因子以诱导新的基因表达程序。正常抗原诱导的T细胞增殖，存活和效应子功能需要通过TCR信号传导转录因子NF-KB的激活。 Card11是TCR介导的NF-KB激活所必需的多域衔接蛋白，但其作用机理尚不完全了解。为了隔离Card11信号辅助因子，进行了一种新型的表达键合屏幕以分离Card11活性的调节剂。从这个屏幕中，将运动蛋白样蛋白Gakin鉴定为Card11的抑制剂，初步研究表明Gakin是对NF-KB的TCR信号传导的抑制剂。拟议的研究的目的是研究Gakin如何在这一重要的信号通路中发挥作用。该实验旨在测试总体假设，即Gakin通过直接结合Card11并将其从免疫学突触中运输到NF-KB抑制TCR信号传导。该假设将以三个特定目的进行检验。首先，使用Gakin缺陷和Gakin过表达的T细胞系，将分析TCR-NF-KB途径中的几个步骤，以确定Gakin作用的步骤。其次，将使用RNA干扰 - 响应测定法确定Gakin的抑制活性所需的域。第三，结合测定和T细胞：APC（抗原呈现细胞）成像测定法将用于证明Gakin是否结合Card11直接结合，并影响Card11募集的动力学或范围对免疫突触。这些研究的结果应扩展我们对控制T细胞激活的分子机械的理解。此外，这项研究可能会深入了解在衰老T细胞中观察到的失调信号传导，这可以导致无效的免疫反应并增加对病原体的敏感性。免疫细胞信号传导缺陷通常在老年人中出现，这会导致对病原体的反应受损。拟议研究的结果有可能提高对免疫细胞用于实现有效反应的分子机制的理解。结果还可能为开发新型治疗剂的发展提供了一个靶标，这些疗法可能会增强衰老失调的免疫反应。