Intermolecular Interactions in the Immunological Synapse

描述（由申请人提供）：FRET显微镜是研究活细胞中蛋白质：蛋白质相互作用的非常有效的手段。该项目使用FRET显微镜来研究抗原识别期间发生的TCR和辅助受体之间的诱导相互作用。FRET显微镜将用于分析CD 8a的不同能力（3和aa作为辅助受体，并在抗原识别过程中被招募到免疫突触，以及确定内源性肽如何增强T细胞识别，包括增强TCR：辅助受体相互作用）。Lck和Fyn在抗原识别过程中的运动和相互作用将用荧光嵌合体和FRET生物传感器进行分析。在发育中的胸腺细胞和活化的T细胞中发现了CD 8的差异唾液酸化。将研究唾液酸化在CD 8募集至突触、链配对、TCR：CD 8和CD 8：Lck相互作用中的作用。表达CD 3？-的转基因小鼠CFP和CD 8？YFP将用于成像TCR：辅助受体相互作用的细胞共轭物之间的发展胸腺细胞和基质细胞，在不同的选择条件下，以了解的速度和强度的TCR：辅助受体相互作用的调制在积极和消极的选择。还将针对初始T细胞、效应T细胞和记忆T细胞分析这些因子。T细胞抗原受体、辅助受体（如CD 4）和信号蛋白之间的相互作用对免疫应答至关重要。了解这些相互作用如何发生将有助于改善对病毒或癌症的免疫反应，并减少自身免疫反应。

DESCRIPTION (provided by applicant): FRET microscopy is a very effective means to investigate protein:protein interactions in living cells. This project uses FRET microscopy to investigate the induced interactions between TCR and co-receptors that occur during antigen recognition. FRET microscopy will be used to analyze the different abilities of CD8a (3 and aa to act as coreceptors and to be recruited to the immunological synapse during antigen recognition, as well as to determine how endogenous peptides enhance T cell recognition, including enhancing the TCR:coreceptor interaction). The movement and interactions of Lck and Fyn during antigen recognition will be analyzed with fluorescent chimeras, and with a FRET biosensor. Differential sialylation of CD8 has been found in developing thymocytes and in activated T cells. The role of sialylation in recruitment of CD8 to the synapse, chain pairing, TCR:CD8 and CD8:Lck interactions will be investigated. Transgenic mice expressing CD3?-CFP and CD8 ?-YFP will be used to image TCR:coreceptor interactions in cell conjugates between developing thymocytes and stromal cells, under different selecting conditions, to understand how the speed and intensity of the TCR:coreceptor interactions are modulated in positive and negative selection. These factors will also be analyzed for naive, effector and memory T cells. The interactions between the T-cell antigen receptor, coreceptors such as CD4, and signaling proteins, are crucial to the immune response. Understanding how these interactions take place will aid in improving the immune response to viruses or cancer, and in reducing autoimmune reactions.