Servicio de Inmunología, Hospital de la Princesa, Instituto de Investigación Sanitaria La Princesa, Madrid, Spain.
Mol Cell Biol. 2013 Sep;33(18):3644-58. doi: 10.1128/MCB.00302-13. Epub 2013 Jul 15.
In this study, we investigated the dynamics of the molecular interactions of tetraspanin CD81 in T lymphocytes, and we show that CD81 controls the organization of the immune synapse (IS) and T cell activation. Using quantitative microscopy, including fluorescence recovery after photobleaching (FRAP), phasor fluorescence lifetime imaging microscopy-Föster resonance energy transfer (phasorFLIM-FRET), and total internal reflection fluorescence microscopy (TIRFM), we demonstrate that CD81 interacts with ICAM-1 and CD3 during conjugation between T cells and antigen-presenting cells (APCs). CD81 and ICAM-1 exhibit distinct mobilities in central and peripheral areas of early and late T cell-APC contacts. Moreover, CD81-ICAM-1 and CD81-CD3 dynamic interactions increase over the time course of IS formation, as these molecules redistribute throughout the contact area. Therefore, CD81 associations unexpectedly define novel sequential steps of IS maturation. Our results indicate that CD81 controls the temporal progression of the IS and the permanence of CD3 in the membrane contact area, contributing to sustained T cell receptor (TCR)-CD3-mediated signaling. Accordingly, we find that CD81 is required for proper T cell activation, regulating CD3ζ, ZAP-70, LAT, and extracellular signal-regulated kinase (ERK) phosphorylation; CD69 surface expression; and interleukin-2 (IL-2) secretion. Our data demonstrate the important role of CD81 in the molecular organization and dynamics of the IS architecture that sets the signaling threshold in T cell activation.
在这项研究中，我们研究了四跨膜蛋白 CD81 在 T 淋巴细胞中分子相互作用的动力学，结果表明 CD81 控制着免疫突触（IS）的组织和 T 细胞的激活。我们使用定量显微镜技术，包括荧光漂白恢复（FRAP）、相荧光寿命成像显微镜-Föster 共振能量转移（phasorFLIM-FRET）和全内反射荧光显微镜（TIRFM），证明 CD81 在 T 细胞与抗原呈递细胞（APCs）之间的连接过程中与 ICAM-1 和 CD3 相互作用。在早期和晚期 T 细胞-APC 接触的中央和外周区域，CD81 和 ICAM-1 表现出不同的流动性。此外，随着 IS 形成的时间进程，CD81-ICAM-1 和 CD81-CD3 的动态相互作用增加，因为这些分子在整个接触区域重新分布。因此，CD81 的相互作用出人意料地定义了 IS 成熟的新连续步骤。我们的结果表明，CD81 控制着 IS 的时间进程和 CD3 在膜接触区的持久性，有助于持续的 T 细胞受体（TCR）-CD3 介导的信号转导。因此，我们发现 CD81 对于适当的 T 细胞激活是必需的，它调节 CD3ζ、ZAP-70、LAT 和细胞外信号调节激酶（ERK）磷酸化；CD69 表面表达；以及白细胞介素-2（IL-2）的分泌。我们的数据证明了 CD81 在 IS 结构的分子组织和动力学中的重要作用，该结构设定了 T 细胞激活中的信号转导阈值。
