Immunological Synapse Restricted Metabolic Reprogramming Drives Driectional Cytokine Synthesis

项目概要/摘要 T细胞在遇到抗原呈递细胞时被激活，并接受三种信号： 通过肽-MHC的信号传导、共刺激信号传导和细胞因子。激活驱动大量的代谢 要求高的过程，如增殖、分化、迁移和效应子功能。因此，T细胞 从氧化磷酸化（OXPHOS），当幼稚时，到激活后的有氧糖酵解，以产生足够的 ATP以适应这些过程。我们最近发现，T细胞参与有氧糖酵解， 分钟的TCR信号传导，独立于共刺激信号传导。我们发现这种机制是通过 丙酮酸脱氢酶激酶1（PDHK 1），一种与LCK相关的线粒体酶， 与T细胞突触相连这些数据表明有氧糖酵解可能是空间调控的 在T细胞突触上的作用。使用pH敏感的荧光系统，我们能够生成数据， 可见有氧糖酵解仅限于T细胞突触。这些数据表明， 定位在T细胞中可以调节免疫突触处有氧糖酵解的起始。此外，我们的 实验室和其他人已经表明，糖酵解酶GAPDH和LDH是mRNA结合蛋白， 它们在幼稚T细胞中的细胞因子翻译，以及通过TCR刺激激活糖酵解促进解离 糖酵解酶从细胞因子mRNA。因此，我们假设线粒体迁移到 IS受线粒体LCK调节，使局部有氧糖酵解和随后的突触形成成为可能 用于定向效应子功能的限制性细胞因子翻译。为了解决这个问题，我们将：（1）识别 线粒体LCK在驱动线粒体迁移到T细胞突触和启动有氧运动中的作用 糖。使用pH敏感的荧光系统，以及线粒体和LCK报告基因，我们将可视化细胞内的蛋白质。 当TCR被APC刺激时，T细胞突触处的有氧糖酵解、线粒体和LCK的动力学。 此外，我们将（2）研究位点限制性有氧糖酵解在促进局部细胞因子中的作用 T细胞突触的翻译。利用糖酵解代谢的调节剂，我们将确定有氧代谢是否 糖酵解可增强活化T细胞中效应细胞因子的产生，有氧糖酵解是否 预测细胞因子被翻译的位点。通过更好地理解促进 代谢重编程和驱动效应器功能，我们可以开发治疗靶点， 我们代谢调节T细胞活性。

Project Summary/Abstract T cells become activated when they encounter an antigen presenting cell and receive three signals: TCR signaling via peptide-MHC, co-stimulatory signaling and cytokines. Activation drives a host of metabolically demanding processes such as proliferation, differentiation, migration, and effector functions. Thus, T cells switch from oxidative phosphorylation (OXPHOS), when naïve, to aerobic glycolysis upon activation to generate enough ATP to accommodate these processes. We recently showed that T cells engage aerobic glycolysis within minutes of TCR signaling, independent of co-stimulatory signaling. We found that this mechanism was mediated by pyruvate dehydrogenase kinase 1 (PDHK1), a mitochondrial enzyme that associated with LCK and migrated to the T cell synapse upon activation. These data suggested that aerobic glycolysis could be spatially regulated at the T cell synapse during activation. Using pH sensitive fluorescent systems, we were able to generate data visualizing aerobic glycolysis restricted to the T cell synapse. These data suggest that the mitochondrial positioning in T cells could regulate the initiation of aerobic glycolysis at the immunological synapse. Further, our lab and others have shown that glycolytic enzymes, GAPDH and LDH, are mRNA binding proteins and repress their cytokine translation in naïve T cells, and activation of glycolysis via TCR stimulus promotes the dissociation of glycolytic enzymes from cytokines mRNA. Therefore, we hypothesize that mitochondrial migration to the IS, regulated by mitochondrial LCK, enables localized aerobic glycolysis and subsequent synapse restricted cytokine translation for directed effector functions. To address this hypothesis we will, (1) Identify the role of mitochondrial LCK in driving mitochondrial migration to the T cell synapse and the initiation of aerobic glycolis. Using pH sensitive fluorescence systems, and mitochondrial and LCK reporters, we will visualize the dynamics of aerobic glycolysis, mitochondria and LCK at the T cell synapse when TCR is stimulated by an APC. Moreover, we will (2) Investigate the role of site restricted aerobic glycolysis in promoting localized cytokine translation at the T cell synapse. Using modulators of glycolytic metabolism, we will determine whether aerobic glycolysis could enhance the production of effector cytokines in activated T cells, and whether aerobic glycolysis predicts sites where cytokines are being translated. By better understanding the early signals that promote metabolic reprogramming and drive effector functions we can develop therapeutic targets that will allow us to metabolically modulate T cell activity.