CELL BIOLOGY OF IMMUNE INTERACTIONS

抗原（Ag）特异性T细胞和抗原呈递细胞（APC）之间的相互作用启动 生产性免疫应答，但这种相互作用也可能导致灭活或诱导 相互作用细胞的耐受性。揭示了多种细胞和分子的复杂整合 决定相互作用细胞命运并导致这些不同生物学结果的机制是 这些研究的长期目标。 使用T-APC缀合物的新型3-D数字成像，表明信号传导和粘附受体 和细胞内蛋白质募集到细胞接触，并形成空间分离的超分子 激活簇（SMAC），构成免疫突触（IS）。的职能作用 SMAC不清楚。一些TCR介导的信号甚至在SMAC形成之前产生。 然而，TCR信号随后位于c-SMAC内，并且SMAC形成似乎是细胞内的一部分。 生产性T细胞激活所需的。这个应用程序的重点是测试一个新的综合模型的T 细胞激活该模型提出，成功的T细胞活化需要TCR的有序进展 信号，这是在形成SMAC之前启动，通过几个空间和时间上不同的 在SMAC中。这种连续的空间信号传导的任何中断都将导致T细胞失活。 该模型将使用新的多维成像的活的和固定的T-APC缀合物，耦合 进行基因、生化和功能研究。这些研究将识别和定位信号事件， 发生在T细胞激活的不同阶段。最先进的成像工具，包括Single 分子跟踪、FRET和活化特异性生化试剂将用于追踪关键 酪氨酸激酶Lck和Zap-70在T细胞活化的不同阶段，并将研究 这些激酶与CD 3/TCR、CD 4、CD 28和衔接蛋白的相互作用。药理学抑制剂 基因操作将被用来破坏和定位选择性的时空阶段， 将确定活化过程以及结构和功能后果。 这些新的研究可能会更好地理解调节T细胞的分子事件， 激活，并可能在未来设计更好的免疫监视方案，免疫抑制， 药物和新疫苗。

The interactions between antigen (Ag)-specific T cells and antigen-presenting cells (APCs) initiate productive immune responses, but such interactions can also result in the inactivation or induction of tolerance in the interacting cells. Uncovering the complex integration of the multiple cellular and molecular mechanisms that determine the fates of the interacting cells and cause these diverse biologic outcomes is the long term goal of these studies. Using novel 3-D digital imaging of T-APC conjugates it was shown that signaling and adhesion receptors and intracellular proteins are recruited to the cell contacts and form spatially segregated Supra-Molecular Activation Clusters (SMACs), which constitute the Immunological Synapse (IS). The functional role of the SMACs is unclear. Some TCR mediated signals are generated even before the formation of SMACs. However, TCR signals are subsequently located within the c-SMACs and SMAC formation appears to be required for productive T cell activation. This application focuses on testing a novel integrative model of T cell activation. The model proposes that successful T cell activation requires the ordered progression of TCR signals, which are initiated before the formation of SMACs, through several spatially and temporally distinct stages in the SMACs. Any disruption of this sequential spatial signaling would result in T cell inactivation. The model will be tested using novel multi-dimensional imaging of live and fixed T-APC conjugates, coupled with genetic, biochemical and functional studies. The studies would identify and localize the signaling events that take place at the different stages of T cells activation. State of the art imaging tools including Single Molecule Tracking, FRET and activation-specific biochemical reagents would be used to follow the key tyrosine kinases Lck and Zap-70 during the different stages of T cell activation and would study the interactions of these kinases with CD3/TCR, CD4, CD28 and adapter proteins. Pharmacological inhibitors and genetic manipulation would be used to disrupt and localize selective spatial-temporal stages of the activation processes and the structural and functional consequences would be determined. These novel studies are likely to achieve better understanding of the molecular events that regulate T cell activation and may be useful in future designs of better immune surveillance protocols, immunosuppression drugs and new vaccines.