CD28 Triggering

CD28触发

• 批准号: 7963606
• 负责人: JIM F Miller
• 金额: $ 19.14万
• 依托单位: UNIVERSITY OF ROCHESTER
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-06-15 至 2012-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7963606
• 关键词: Abbreviations; Accounting; Amino Acids; Antigen-Presenting Cells; Binding Proteins; Biological Assay; CD28 gene; Cell membrane; Disulfides; Event; Fluorescence; Fluorescence Resonance Energy Transfer; Goals; ITAM; Integrins; Left; Ligand Binding; Link; Mediating; Membrane; Modeling; Molecular; Molecular Conformation; Monoclonal Antibodies; Mutation; Phosphorylation; Phosphotransferases; Plant Leaves; Proteins; Recruitment Activity; Regulation; Research; Rest; Role; Signal Transduction; Signaling Molecule; T cell regulation; T-Cell Activation; T-Cell Receptor; T-Lymphocyte; TCR Activation; Tail; Testing; dimer; immunological synapse; interest; pathogen; public health relevance

DESCRIPTION (provided by applicant): The two signal model and the concept of costimulation are well engrained in our understanding of T cell regulation. It is well established that costimulation through CD28 can have a dramatic effect on T cell activation, survival, tolerance, and differentiation. However, in spite of considerable interest and effort, the regulation and mechanism of CD28 costimulation are still poorly understood. CD28 signaling is mediated through the recruitment of cytosolic signaling molecules to specific motifs within the cytosolic tail (CT) domain of CD28. In an effort to understand how ligand binding to the lumenal domain of CD28 could transduce changes to the cytosolic domains to initiate signaling, we established a FRET assay. CFP and YFP were fused to the end of the CT of CD28. CD28 is a disulfide-linked dimer and when CD28-CFP and CD28-YFP were co-expressed, high level of FRET was directed, indicating that the ends of the CT domains are in close proximity within the CD28 dimer. When CD28 was recruited to the immunological synapse, the level of FRET was reduced, indicating that there was some structural change in the orientation of the CT domains within the CD28 dimer. Surprisingly, this change in FRET was not mediated by CD28 ligand binding, but instead was mediated through TCR signaling, even in the absence of CD28 ligand binding. This suggests that TCR signaling can activate CD28, possibly impacting on ligand binding (in analogy to inside-out signaling for integrin activation), immunological synapse recruitment, or signal transduction. The overall goal of this R21 application is to identify the mechanism and functional consequence of this TCR- mediated change in the orientation of the CD28 CT domains. We will do this through two Specific Aims. 1, Determine the molecular events associated with TCR induced changes in the orientation of the CD28 CT domains. 2, Determine whether TCR signaling regulates CD28 ligand binding. PUBLIC HEALTH RELEVANCE: T cell activation requires the specific recognition of pathogen-specific proteins by the TCR and co-signaling through costimulatory molecules, most notably CD28. Although the function of CD28 is well described, the molecular events associated with CD28 function as not well understood. We have made the surprising finding that the TCR may modify the conformation of CD28 and so regulate the function of CD28.

描述（由申请人提供）：两个信号模型和共刺激的概念深深植根于我们对 T 细胞调节的理解中。众所周知，通过 CD28 进行的共刺激可以对 T 细胞的激活、存活、耐受和分化产生巨大影响。然而，尽管有相当大的兴趣和努力，CD28共刺激的调控和机制仍然知之甚少。 CD28 信号传导是通过将胞质信号分子募集到 CD28 胞质尾 (CT) 结构域内的特定基序来介导的。为了了解配体与 CD28 腔结构域的结合如何转导胞质结构域的变化以启动信号传导，我们建立了 FRET 测定。 CFP和YFP融合到CD28的CT末端。 CD28是二硫键连接的二聚体，当CD28-CFP和CD28-YFP共表达时，定向高水平的FRET，表明CT结构域的末端在CD28二聚体内非常接近。当 CD28 被招募到免疫突触时，FRET 水平降低，表明 CD28 二聚体内 CT 结构域的方向发生了一些结构变化。令人惊讶的是，FRET 的这种变化不是由 CD28 配体结合介导的，而是通过 TCR 信号传导介导的，即使在没有 CD28 配体结合的情况下也是如此。这表明 TCR 信号传导可以激活 CD28，可能影响配体结合（类似于整合素激活的内向外信号传导）、免疫突触募集或信号转导。 R21 应用的总体目标是确定 TCR 介导的 CD28 CT 结构域方向变化的机制和功能结果。我们将通过两个具体目标来实现这一目标。 1、确定与 TCR 诱导的 CD28 CT 结构域方向变化相关的分子事件。 2、确定TCR信号是否调节CD28配体结合。 公共健康相关性：T 细胞激活需要 TCR 特异性识别病原体特异性蛋白，并通过共刺激分子（尤其是 CD28）进行协同信号传导。尽管 CD28 的功能已得到很好的描述，但与 CD28 功能相关的分子事件尚不清楚。我们得到了令人惊讶的发现，TCR可能会修饰CD28的构象，从而调节CD28的功能。