Determining the role of trogocytosis-mediated signaling on CD4 T cell phenotype and effector functions

 描述(申请人提供)：CD4T细胞对同源MHC II类的识别：抗原提呈细胞(APC)上的多肽复合体(APC)触发T-APC界面上的大规模分子重排，形成一种称为免疫突触的结构。在突触处，T细胞从APC捕获大的膜碎片和相关蛋白，这一过程称为巨噬细胞增多。虽然这种现象有可能显著改变单个T细胞的生物学特性，但我们目前对这一过程的生物学后果了解有限。由于巨噬细胞增多，CD4T细胞在其表面显示免疫突触成分，包括同源的MHC：肽和共刺激分子，如CD80。我们之前已经证明，这些分子可能通过与T细胞上的受体结合来调节T细胞内的细胞内信号。在这项建议中，我们将研究这一信号对单个T细胞的影响。我们的初步数据表明，这种巨噬细胞增多相关的信号维持效应细胞因子(IL-4)的产生，并可能在体外介导T细胞向TH2表型的转换。这一建议的中心假设是，巨噬细胞增多介导的持续的TCR信号支持效应细胞因子的产生，并介导trog T细胞向TH2表型的转化。这一假设将通过两个特定的目标来检验：目标1)确定观察到的trog IL-4和GATA-3hi细胞频率的增加是否是来自巨噬细胞分子的信号结果。目的#2)确定观察到的trog细胞IL-4和GATA-3表达的增加是否反映了TH1和TH2能力的内在差异 进行巨噬细胞增多症或选择性存活和/或转换为TH2的结果。使用流式细胞术、qrt-pcr和3d广域去卷积显微镜，这些实验将涉及到体外和体内方法来监测效应细胞因子的产生。 细胞自主信号和通过巨噬细胞增多症获得APC膜碎片的细胞的表型检查。这一建议的结果将为T细胞增多症在T细胞效应器功能和效应器表型中的作用提供重要的见解，并将有助于进一步探讨巨噬细胞增多症在体内的功能。

 DESCRIPTION (provided by applicant): CD4+ T cell recognition of cognate MHC class II:peptide complexes on antigen presenting cells (APC) triggers large-scale molecular rearrangements at the T-APC interface forming a structure called the immune synapse. At the synapse, T cells capture large membrane fragments and associated proteins from the APC in a process termed trogocytosis. While this phenomenon has the potential to significantly alter the biology of the individual T cell, we currently have only limited understanding of the biological consequences of this process. As a result of trogocytosis, CD4+ T cells display immunological synapse components on their surface including cognate MHC:peptide and costimulatory molecules, such as CD80. We have previously shown that these molecules mediate intracellular signaling within the T cell, presumably by engaging their receptors on the cell. In this proposal we will examine the impact that this signaling has on the individual T cell. Our preliminary data suggests that this trogocytosis-associated signaling maintains effector cytokine (IL-4) production and may mediate the conversion of T cells to a TH2 phenotype in vitro. The central hypothesis of this proposal is that is that trogocytosis-mediated, sustained TCR signaling sustains effector cytokine production and mediates the conversion of trog+ T cells to a TH2 phenotype. This hypothesis will be examined using 2 specific aims: Aim #1) Determine whether the the observed increase in the frequency of trog+ IL-4+ and GATA-3hi cells is the result of signaling from the trogocytosed molecules. Aim #2) Determine whether the observed increase in IL-4 and GATA-3 expression by trog+ cells is reflective of an intrinsic difference in the ability of TH1 and TH2 to perform trogocytosis or the result of selective survival and/or conversion to TH2. Using flow cytometry, qRT-PCR and 3D wide-field deconvolution microscopy, these experiments will involve both in vitro and in vivo approaches to monitor effector cytokine production as a result of cell- autonomous signaling and examination of the phenotype of the cells that have acquired APC membrane fragments via trogocytosis. The results from this proposal will provide important insight into the role of trogycytosis in T cell effector function and effector phenotype and will lad to additional lines of inquiry to elucidate the in vivo functions of trogocytosis.