Signal Transduction in T Lymphocyte Activation

描述(由申请人提供)：这笔赠款之前的工作主要集中在两个 互补区域，即蛋白激酶C-α的功能和调节 (PKCalpha)，以及Syk家族PTK与其相互作用 它们在T细胞中的底物和靶点。最近的研究，包括我们自己的研究， 证明PKCtheta是免疫突触的关键成分 (IS)。PKCtheta在成熟T细胞活化中发挥重要作用 整合TCR/CD28共刺激信号导致核因子-kB的激活 信令级联和AP-1。诱导这些转录因子，这是 对产生IL-2和预防T细胞无能是必不可少的，在 无能T细胞。鉴于PKCtheta在T细胞中的重要性日益显现 激活和我们对这一领域的广泛承诺，我们将把我们的 PKCtheta在抗原特异性T细胞中的作用和调控研究进展 细胞，重点是它在CD28共刺激和无能中的潜在作用。 首先，我们将阐明分子和生化机制(S) 命令有选择地将PKCtheta招募到IS并激活它， 包括推测的支架蛋白(S)的身份，PI3K的作用，以及 PKCtheta本地化的功能重要性。第二， 将PKCtheta连接到核因子-kB级联的中间信号蛋白是 未知。我们将使用抗原特异的野生型或PKCtheta缺陷的T细胞 Jurkat T细胞与生化和遗传学方法相结合 确定这些蛋白质，重点是已知的核因子-kB和 MAPK信号传递级联。第三，我们将研究PKCtheta在 转导CD28共刺激信号预防T细胞无能 使用选择性的PKCtheta干扰策略，或者相反，确定 结构性激活的PKCtheta能否挽救T细胞的激活 CD28缺失的T细胞。这些研究将产生对 这种关键T细胞酶的结构-功能关系，建立 它在T细胞激活、共刺激和无能中的作用，并提供一种声音 为将来将其用作T细胞的药物靶点奠定了基础。

DESCRIPTION (provided by applicant): Previous work on this grant focused on two complementary areas, i.e.,the function and regulation of protein kinase C-alpha (PKCalpha), which we isolated in 1993, and interactions of Syk-family PTKs with their substrates and targets in T cells. Recent studies, including our own, demonstrated that PKCtheta is a key component of the immunological synapse (IS). PKCtheta plays an essential role in mature T cell activation by integrating TCR/CD28 costimulatory signals leading to activation of the NF-kB signaling cascade and AP-1. Induction of these transcription factors, which is essential for IL-2 production and prevention of T cell anergy, is defective in anergic T cells. In view of this emerging importance of PKCtheta in T cell activation and our extensive commitment to this area, we will focus our proposed studies on the function and regulation of PKCtheta in Ag-specific T cells, with emphasis on its potential role in CD28 costimulation and anergy. First, we will elucidate the molecular and biochemical mechanism(s) that dictate the selective recruitment of PKCtheta to the IS and its activation, including the identity of putative scaffold protein(s), the role of PI3K, and the functional importance of the IS localization of PKCtheta. Second, the intermediate signaling proteins that couple PKCtheta to the NF-kB cascade are unknown. We will use Ag-specific wild type or PKCtheta-deficient T cells and Jurkat T cells in conjunction with biochemical and genetic approaches to identify these proteins, with emphasis on known components of the NF-kB and MAPK signaling cascades. Third, we will study the role of PKCtheta in transducing CD28 costimulatory signals and preventing T cell anergy by employing selective PKCtheta interfering strategies or, conversely, determining whether constitutively active PKCtheta can rescue T cell activation in CD28-deficient T cells. These studies will generate a detailed understanding of the structure-function relationship of this critical T cell enzyme, establish its role in T cell activation, costimulation, and anergy, and provide a sound basis for its future use as a drug target in T cells.