Regulating peripheral T cell tolerance

描述(申请人提供)：大多数健康人的T细胞处于幼稚的静止或静止状态。静止期通过防止不受控制的多克隆扩增，确保在特定空间内维持庞大的T细胞保存库。最近的证据表明，T细胞的静止是主动编程和维持的。然而，控制T细胞静止的机制却知之甚少。稳定的初始CD8 T细胞的维持依赖于IL-7受体和T细胞抗原受体(TCR)的协调信号。TCR信号是在TCR与装载有自体多肽的主要组织相容性复合体I分子结合后启动的。这种紧张性TCR信号是如何被严格控制以防止T细胞完全激活的，目前还知之甚少。二酰基甘油激酶(DGKs)是将二酰基甘油(DAG)转化为磷脂酸(PA)的酶，既是重要的第二信使，又参与多种信号通路的激活，调节多种细胞过程和功能。在哺乳动物中，存在10种DGK亚型，但人们对它们的生理功能知之甚少。我们发现，T细胞表达的主要亚型DGKα和ζ的缺失导致NA�ve T细胞失去静止，导致获得效应器功能。此外，DGKα和ζ双缺陷小鼠发生严重的自身免疫性疾病，T细胞起关键作用。本应用的目的是以条件性DGKα和ζ缺陷小鼠为模型，研究控制CD8T细胞静止的机制，并利用新生的有条件表达WT和突变型GFP-DGKζ融合蛋白的小鼠，对DGKζ在原代T细胞中的结构/功能进行彻底的分析。我们将验证这样的假设，即DGKα和ζ调制包括mTOR信号在内的多个信号通路以确保T细胞耐受和静止，以及DGKζ通过不同的结构域/基序指导其定位和相互作用，使其能够在多个亚细胞室中作为T细胞的关键调节因子发挥作用。我们将追求以下两个具体目标。在目标1中，我们将研究由DGKα和ζ控制的确保CD8T细胞静止的机制。在目标2中，我们将确定DGKζ作为T细胞发育和耐受的关键调节因子的关键结构特征。本申请中提出的研究将为T细胞发育和信号调控提供新的见解 静止期以及DGKζ如何在T细胞中发挥其关键的调节作用。

DESCRIPTION (provided by applicant): Most T cells in healthy individuals are in a naïve resting or quiescent state. Quiescence ensures the maintenance of the vast repertoire of T cells in a defined space by preventing uncontrolled polyclonal expansion. Recent evidence indicates that T cell quiescence is actively programmed and maintained. However, the mechanisms that control T cell quiescence are poorly understood. The maintenance of steady-state naïve CD8 T cells relies on orchestrated signaling from the IL-7 receptor and the T cell antigen receptor (TCR). TCR signaling is initiated after the engagement of the TCR with major histocompatibility complex I molecules loaded with self-peptides. How such tonic TCR signaling is tightly controlled to prevent full T cell activation is poorly understood. Diacylglycerol kinases (DGKs) are enzymes that convert diacylglycerol (DAG) to phosphatidic acid (PA), both important second messengers involved in activating in multiple signaling pathways and regulate diverse cellular processes and functions. In mammals, ten DGK isoforms exist yet their physiological functions are poorly understood. We have found that deficiency of both DGKα and ζ, the major isoforms expressed in T cells, causes na�ve T cells to lose quiescence, leading to acquisition of effector function. Furthermore, DGKα and ζ double deficient mice develop severe autoimmune diseases with T cells play critical roles. The objectives of this application are to investigate mechanisms that control CD8 T cell quiescence using conditional DGKα and ζ deficient mice as a model and to perform thorough structure/function analysis of DGKζ in primary T cells using newly generated mice that conditionally express WT and mutant GFP-DGKζ fusion proteins. We will test the hypotheses that DGKα and ζ modulate multiple signaling pathways including mTOR signaling to ensure T cell tolerance and quiescence and that DGKζ directs its localizations and interactions via distinct structural domains/motifs, enabling it to function in multiple subcellular compartments as a critical regulator in T cells. We will pursue the following two specific aims. In Aim 1, we will investigate mechanisms that are controlled by DGKα and ζ to ensure CD8 T cell quiescence. In Aim 2, we will determine structural features critical for DGKζ to function as a critical regulator for T cell development and tolerance. Studies proposed in this application will provide novel insight into the signaling control of T cell development and quiescence and how DGKζ fulfills its critical regulatory roles in T cells.