ROLE CD28 PI3-KINASE AND GRB-2/SOS IN T-CELL IMMUNITY

CD28 PI3 激酶和 GRB-2/SOS 在 T 细胞免疫中的作用

• 批准号: 2748780
• 负责人: Christopher E. Rudd
• 金额: $ 19.64万
• 依托单位: DANA-FARBER CANCER INST
• 依托单位国家: 美国
• 财政年份: 1995
• 资助国家: 美国
• 起止时间: 1995-09-30 至 2000-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2748780
• 关键词: CD antigens; CD28 molecule; CD3 molecule; CD4 molecule; T cell receptor; T lymphocyte; biological signal transduction; clone cells; gene mutation; genetically modified animals; human tissue; hybridomas; interleukin 2; laboratory mouse; leukocyte activation /transformation; leukocyte adhesion molecules; membrane proteins; phosphatidylinositol 3 kinase; phosphorylation; receptor binding; site directed mutagenesis; transfection

The T cell antigen CD28 provides a co-stimulatory signal that regulates the T cell response to foreign antigen. Understanding the intracellular signalling mechanisms that mediate CD28 function is an issue of great importance. We have shown that CD28 binds to two intracellular signalling molecules, phosphatidylinositol 3-kinase(Pl 3-kinase) and GRB-2/SOS. Each mediate distinct and major pathways of sign transduction. PI 3-kinase generates D-3 inositol lipids, and is an essential component in signalling by various receptors. Similarly, GRB-2/SOS activate p21ras, a central component in the growth control of mammalian cells. Site-directs mutagenesis, peptide competition and binding studies demonstrated that PI kinase and GRB-2 bind to the same site wit cytoplasmic tail of CD28 define by the motif pYMNM. Despite this, both CD28-PI 3-kinase and CD28-GRB-2/SC complexes can be found in T-cells. CD28 is therefore endowed with the capacity to engage multiple signalling pathways in T-cells. In this application, we plan further our analysis. of the nature of CD28pYMNM mediated signalling by assessing CD28 phosphorylation sites, the influence of TcR/CD3-C 4 ligation, and the identification of the kinase responsible for phosphorylating CD28. We will also examine CTLA-4 and CD7 binding to these intracellular proteins, and whether CD7 can function as an alternate co-stimulatory antigen (Aims 1-3). We propose assess the roles played by PI 3-kinase and GRB-2 in mediating co-stimulation ar IL-2 production by generating mutants of CD28 with altered abilities to bind PI kinase or GRB-2 for use in the transfection of T-cell hybridomas and clones (Aim 4). Lastly, we propose to express CD28 signalling mutants in CD28-/- gene knockout mice in an effort to dissect the role of the different CD28 signalling pathways in T-cell immunity (Aim 5).

T 细胞抗原 CD28 提供共刺激信号，调节 T 细胞对外来抗原的反应。 了解细胞内 介导CD28功能的信号机制是一个重大问题 重要性。 我们已经证明 CD28 与两个细胞内信号传导结合 分子，磷脂酰肌醇3-激酶（Pl 3-激酶）和GRB-2/SOS。每个 介导独特且主要的信号转导途径。 PI 3-激酶 产生 D-3 肌醇脂质，是信号传导的重要组成部分 通过各种受体。类似地，GRB-2/SOS 激活 p21ras，这是一个中枢 哺乳动物细胞生长控制的组成部分。站点直接 诱变、肽竞争和结合研究表明 PI 激酶和 GRB-2 与 CD28 胞质尾部的同一位点结合 由主题 pYMNM 组成。尽管如此，CD28-PI 3-激酶和 CD28-GRB-2/SC 复合物可以在 T 细胞中找到。因此，CD28 被赋予 参与 T 细胞中多种信号传导途径的能力。在这个 应用程序，我们计划进一步分析。 CD28pYMNM 的性质 通过评估 CD28 磷酸化位点介导的信号传导，影响 TcR/CD3-C 4 连接，以及负责的激酶的鉴定 用于磷酸化 CD28。我们还将检查 CTLA-4 和 CD7 与 这些细胞内蛋白，以及 CD7 是否可以作为替代蛋白 共刺激抗原（目标 1-3）。我们建议评估所扮演的角色 PI 3-激酶和 GRB-2 介导共刺激 ar IL-2 的产生 产生 CD28 突变体，其结合 PI 激酶的能力发生改变，或 GRB-2 用于 T 细胞杂交瘤和克隆的转染（目标 4）。 最后，我们建议在CD28-/-基因中表达CD28信号突变体 基因敲除小鼠试图剖析不同 CD28 的作用 T 细胞免疫中的信号通路（目标 5）。