Division of Cell and Molecular Biology, Imperial College London, London, United Kingdom.
PLoS Biol. 2011 Sep;9(9):e1001152. doi: 10.1371/journal.pbio.1001152. Epub 2011 Sep 13.
Natural Killer (NK) cells are innate immune cells that secrete lytic granules to directly kill virus-infected or transformed cells across an immune synapse. However, a major gap in understanding this process is in establishing how lytic granules pass through the mesh of cortical actin known to underlie the NK cell membrane. Research has been hampered by the resolution of conventional light microscopy, which is too low to resolve cortical actin during lytic granule secretion. Here we use two high-resolution imaging techniques to probe the synaptic organisation of NK cell receptors and filamentous (F)-actin. A combination of optical tweezers and live cell confocal microscopy reveals that microclusters of NKG2D assemble into a ring-shaped structure at the centre of intercellular synapses, where Vav1 and Grb2 also accumulate. Within this ring-shaped organisation of NK cell proteins, lytic granules accumulate for secretion. Using 3D-structured illumination microscopy (3D-SIM) to gain super-resolution of ~100 nm, cortical actin was detected in a central region of the NK cell synapse irrespective of whether activating or inhibitory signals dominate. Strikingly, the periodicity of the cortical actin mesh increased in specific domains at the synapse when the NK cell was activated. Two-colour super-resolution imaging revealed that lytic granules docked precisely in these domains which were also proximal to where the microtubule-organising centre (MTOC) polarised. Together, these data demonstrate that remodelling of the cortical actin mesh occurs at the central region of the cytolytic NK cell immune synapse. This is likely to occur for other types of cell secretion and also emphasises the importance of emerging super-resolution imaging technology for revealing new biology.
自然杀伤 (NK) 细胞是先天免疫细胞，它们通过免疫突触分泌裂解颗粒，直接杀死病毒感染或转化的细胞。然而，在理解这个过程方面，一个主要的差距是如何建立裂解颗粒如何穿过基底膜下的皮质肌动蛋白网格。由于传统的光学显微镜分辨率太低，无法在裂解颗粒分泌过程中解析皮质肌动蛋白，因此研究受到了阻碍。在这里，我们使用两种高分辨率成像技术来探测 NK 细胞受体和丝状（F）-肌动蛋白的突触组织。光学镊子和活细胞共聚焦显微镜的组合表明，NKG2D 微簇在细胞间突触的中心组装成一个环形结构，Vav1 和 Grb2 也在此处积累。在 NK 细胞蛋白的这种环形结构中，裂解颗粒聚集起来准备分泌。使用 3D 结构照明显微镜（3D-SIM）获得约 100nm 的超分辨率，在 NK 细胞突触的中心区域检测到皮质肌动蛋白，无论激活或抑制信号占主导地位。引人注目的是，当 NK 细胞被激活时，皮质肌动蛋白网格的周期性在突触的特定区域增加。双色超分辨率成像显示，裂解颗粒精确地停靠在这些区域，这些区域也靠近微管组织中心 (MTOC) 极化的位置。总之，这些数据表明，细胞毒性 NK 细胞免疫突触的中央区域发生了皮质肌动蛋白网格的重塑。这可能发生在其他类型的细胞分泌中，也强调了新兴的超分辨率成像技术对于揭示新生物学的重要性。
