Ewers Helge, Smith Alicia E, Sbalzarini Ivo F, Lilie Hauke, Koumoutsakos Petros, Helenius Ari
Institute of Biochemistry, Swiss Federal Institute of Technology, CH-8093 Zurich, Switzerland.
Proc Natl Acad Sci U S A. 2005 Oct 18;102(42):15110-5. doi: 10.1073/pnas.0504407102. Epub 2005 Oct 11.
The lateral mobility of individual murine polyoma virus-like particles (VLPs) bound to live cells and artificial lipid bilayers was studied by single fluorescent particle tracking using total internal reflection fluorescence microscopy. The particle trajectories were analyzed in terms of diffusion rates and modes of motion as described by the moment scaling spectrum. Although VLPs bound to their ganglioside receptor in lipid bilayers exhibited only free diffusion, analysis of trajectories on live 3T6 mouse fibroblasts revealed three distinct modes of mobility: rapid random motion, confined movement in small zones (30-60 nm in diameter), and confined movement in zones with a slow drift. After binding to the cell surface, particles typically underwent free diffusion for 5-10 s, and then they were confined in an actin filament-dependent manner without involvement of clathrin-coated pits or caveolae. Depletion of cholesterol dramatically reduced mobility of VLPs independently of actin, whereas inhibition of tyrosine kinases had no effect on confinement. The results suggested that clustering of ganglioside molecules by the multivalent VLPs induced transmembrane coupling that led to confinement of the virus/receptor complex by cortical actin filaments.
利用全内反射荧光显微镜通过单荧光粒子追踪技术研究了结合在活细胞和人工脂质双层上的单个鼠多瘤病毒样颗粒（VLPs）的侧向流动性。根据矩量缩放谱描述的扩散速率和运动模式对粒子轨迹进行了分析。尽管脂质双层中结合到其神经节苷脂受体的VLPs仅表现出自由扩散，但对活的3T6小鼠成纤维细胞上轨迹的分析揭示了三种不同的流动性模式：快速随机运动、在小区域（直径30 - 60 nm）内的受限运动以及在具有缓慢漂移的区域内的受限运动。结合到细胞表面后，粒子通常进行5 - 10秒的自由扩散，然后它们以肌动蛋白丝依赖的方式被限制，且不涉及网格蛋白包被小窝或小窝。胆固醇的耗尽显著降低了VLPs的流动性，且与肌动蛋白无关，而酪氨酸激酶的抑制对限制作用没有影响。结果表明，多价VLPs对神经节苷脂分子的聚集诱导了跨膜偶联，导致病毒/受体复合物被皮质肌动蛋白丝限制。
