Coreceptor Modification of TCR Tyrosine Kinase Signals

脂筏膜区室化和MAGUK家族分子支架作为关键组织者发挥作用， 神经元和上皮细胞中蛋白质和膜运输、细胞骨架重组和信号转导 交叉点在这里，我们考虑了Dlghl MAGUK家族成员在组织细胞骨架、脂质代谢和细胞凋亡中的潜在作用。 T细胞上的筏和信号转导：APC突触连接。在过去的融资周期中，我们展示了 在介导TCR/共刺激诱导中对LckSHS结构域的需求：突触筏簇集;进行性和 持续的信号传导;减少的TCR接合所需的持续时间; Erk活化;和IL-2产生。我们成 我们对Dlghl作为潜在的LckSH 3效应物感兴趣，因为我们鉴定了它作为LckSH 3配体，并证明了它是LckSH 3的一种配体。 LckSH 3：Dlghl相互作用对于Dlghl筏微结构域膜定位是必需的。而且我们 最近鉴定了WASp、Erk-1和p38作为额外的Dlghl配体，并表明它们可能与Dlghl（rLck）起作用。 效应器在这个提议中，我们设计了旨在评估Dlghl在TCR/共刺激分子中的潜在作用的实验。 诱导的信号转导和阐明Dlghl活性的分子基础。此外，我们认为， 不同的T细胞亚群差异性地利用Dlghl活性以产生独特适合的突触的可能性 以实现特定功能。为了解决这些问题，我们利用了三管齐下的方法， 介导的Dlghl敲低、Dlghl过表达/再表达和dlghl基因敲除。我们分析了 BI-141 T杂交瘤、CD 4 + 5CC 7和CD 8 + OT-1 TCR转基因T细胞和发育中的T细胞中的Dlghl活性 子集我们预测，直接比较Dlghl支架在不同的发育和T效应细胞群体中， 将阐明新的TCR信号转导机制和分子细节参与专门的突触。 具体而言，我们提出以下建议：1）研究Dlghl在抗原诱导的T细胞免疫中的潜在作用。 信号转导、免疫突触组装和效应器功能;目的2）确定 3）为了确定发育中的T细胞和效应T细胞是否（差异地）依赖于Dlghl活性， Dlghl脚手架活动。 我们的研究可能会导致更好地了解TCR信号是如何调节介导功能性 这是设计旨在可预测地调节特定TCR应答的治疗剂所必需的。 事实上，阐明突触组织和信号转导事件的分子介质将提供新的 用于治疗的靶点，其目的在于抑制不需要的T细胞活化应答， 排斥和T细胞转化。相反，在设计中可以使用个体激活事件的促进者 肿瘤或其他疫苗，旨在加强对次优抗原的反应。

Lipid raft membrane compartmentalization and MAGUK-family molecular scaffolds function as key organizers of protein and membrane trafficking, cytoskeletal reorganization and signal transduction in neuronal and epithelial cell junctions. Here we consider a potential role for the Dlghl MAGUK family member in organizing the cytoskeleton, lipid rafts and signal transducers at the T cell:APC synapticjunction. During the past funding cycle we demonstrated a requirement for the LckSHS domain in mediating TCR/costimulation induced: synaptic raft clustering; processive and sustained signaling; reduced required duration of TCR engagement; Erk activation; and IL-2 production. We became interested in Dlghl as a potential LckSH3 effector because we identified it as a LckSH3 ligand and demonstrated LckSH3:Dlghl interactions as essential for Dlghl raft microdomain membrane localization. Moreover, we have recently identified WASp, Erk-1 and p38 as additional Dlghl ligands and suggest they may function as Dlghl(rLck) effectors. In this proposal, we design experiments aimed at assessing a potential role for Dlghl in TCR/costimulator induced signal transduction and elucidating the molecular basis of Dlghl activity. Furthermore, we consider the possibility that distinct T cell subsets differentially capitalize on Dlghl activities to generate synapses uniquely suited for effecting particular functions. To address these issues we capitalize on a three pronged approach involving siRNA mediated Dlghl knockdown, Dlghl over-expression/re-expression and dlghl gene knockout. We include analysisof Dlghl activity in the BI-141 T hybridoma, CD4+ 5CC7 and CD8+ OT-1 TCR transgenic T cells and in developing T cell subsets. We predict that direct comparison of Dlghl scaffolds withindistinct developing and T effector populations will elucidate novel TCR signal transduction mechanisms and molecular details involved in specializing synapses. Specifically, we propose the following: Arm 1) To investigate potential roles for Dlghl in antigen-induced T cell signal transduction, immune synapse assembly, and effector function; Aim 2) To determine the molecular basis of Dlghl activity in T cells; and Aim 3) To determine whether developing and effector T cells (differentially)rely on Dlghl scaffolding activities. Our studies will likely lead to a better understanding of how TCR signals are regulated to mediate functional outcome, which is essential to the design of therapeutics aimed at predictably modulating particular TCR responses. Indeed, elucidation of molecular mediators of synaptic organization and signal transduction events will provide novel targets for therapeutics aimed at inhibiting unwanted T cell activation responses including autoimmunity, graft rejection, and T cell transformation. Conversely, facilitators of individual activation events could be used in the design of tumor or other vaccines aimed at potentiating responses against suboptimally presented antigens.