Imaging and Function of the Immunological Synapse

描述（由申请人提供）：T淋巴细胞将抗原识别为与抗原呈递细胞（APC）表面MHC分子结合的小肽。除了T细胞抗原受体和多肽负载的MHC外，许多其他细胞表面分子在决定T细胞/APC相互作用的结果中起着必要的或调节作用。抗原识别伴随着这些分子的大规模细胞骨架依赖性重排，形成T细胞和APC之间有组织的接触界面，称为“免疫突触”。许多实验室正在研究突触如何提供或调节T细胞完全激活所必需的信号。在这项拨款申请中，重点是免疫突触在将效应功能从CD4 T细胞传递到APC中的作用，使用FasL （CD178）和CD40L （CD154）作为膜结合效应细胞因子的原型。由于体内大多数相互作用可能太短，无法允许效应细胞因子的重新合成，因此提出的实验研究了预先形成的细胞内FasL和CD40L储存在T细胞/APC相互作用中发挥重要作用的可能性。CD40L首先被确定为细胞接触依赖性CD4 T细胞帮助B细胞的介质，并且已被证明是巨噬细胞杀死树突状细胞内病原体的有效激活剂，用于启动CD4和CD8 T细胞的效应功能和记忆。FasL是CD4和CD8 T细胞接触依赖性杀伤的主要介质，并在激活诱导的细胞死亡过程中通过T细胞自杀或自相残杀来调节T细胞反应。

DESCRIPTION (provided by applicant): T lymphocytes recognize antigen as small peptides bound to MHC molecules on the surface of antigen presenting cells (APC). In addition to the T cell antigen receptor and peptide-loaded MHC, dozens of other cell surface molecules play necessary or modulating roles in determining the outcome of the T cell/APC interaction. Antigen recognition is accompanied by large-scale, cytoskeleton-dependent rearrangements of these molecules to form an organized contact interface between the T cell and the APC termed the "immunological synapse". Many laboratories are investigating the ways in which the synapse may provide or regulate the signals necessary for full T cell activation. In this grant application, the focus is on the role of the immunological synapse in the delivery of effector functions from CD4 T cells to APC, using FasL (CD178) and CD40L (CD154) as prototypes of membrane-bound effector cytokines. Since most interactions in vivo may be too short to allow for de novo synthesis of effector cytokines, the proposed experiments examine the possibility that preformed, intracellular stores of FasL and CD40L play important roles in T cell/APC interactions. CD40L was first identified as the mediator of cell contact-dependent CD4 T cell help for B cells, and has since been shown to be a potent activator of macrophages for killing of intracellular pathogens of dendritic cells for priming effector functions and memory in CD4 and CD8 T cells. FasL is a principal mediator of contact-dependent killing by CD4 and CD8 T cells, and regulates T cell responses through T cell suicide or fratricide during activation-induced cell death. Using a combination of state-of-the-art microscopy and functional assays in vitro and in vivo, this proposal will test the hypothesis that pre-formed CD40L and FasL are stored in secretory lysosomes in most primed CD4 effector and memory T cells, are delivered rapidly through the immunological synapse to B cells and other APC in transient antigen-specific interactions, and have important functional consequences distinct from those of newly synthesized CD40L and FasL. It will also investigate structural difference between synapses formed by type 1 and type 2 helper T cells, and test the hypothesis that the function of the ring of adhesion molecules in the mature immunological synapse formed by CD4 T cells is to ensure antigen-specific delivery of effector molecules to the APC and prevent delivery to surrounding bystander cells. PUBLIC HEALTH RELEVANCE: T lymphocytes recognize antigen only on the surface of other cells, called antigen presenting cells (APC), and exert their effects by secreting potent cytokines and other effector molecules. The goal of this application is to understand how various kinds of CD4 T cells deliver effector cytokines to APC. The application is focused on FasL (CD178) and CD40L (CD154), key membrane-bound cytokines which control the adaptive immune response in health and disease. The proposed experiments will contribute to a basic understanding of the regulation CD4 T cell effector functions, which in turn will have public health-related applications in transplantation, autoimmunity, allergy, cancer immunotherapy, and control of infectious diseases.