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Dynamic tracking and mobility analysis of single GLUT4 storage vesicle in live 3T3-L1 cells

机译:3T3-L1活细胞中单个GLUT4储存囊泡的动态跟踪和迁移性分析

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Glucose transporter 4 (GLUT4) is responsible foi insulin-stimulated glucose transporting into the insulin-sensitive fat and muscle cells. The dynamics of GLUT4 storage vesicles (GSVs) remains to be explored and it is unclear how GSVs are arranged based on their mobility We examined this issue in 3T3-L1 cells via investigating the three-dimensional mobility of single GSV labeled with EGFP-fused GLUT4. A thin layer of cytosol right adjacent to the plasma membrane was illuminated and successively imagedat 5 Hz under a total internal reflection fluorescence microscope with a penetration depth of 136 nm Employing single particle tracking, the three-dimensional subpixel displacement of single GSV was tracked at a spatial precision of 22 nm. Both the meansquare displacement and the diffusion coefficient were calculated for each vesicle. Tracking results revealed that vesicles moved as if restricted within a cage that has a mean radius of 160 nm, suggesting the presence of some intracellular tethering matrix. By constructing the histogram of the diffusion coefficients of GSVs, we observed a smooth distribution instead of the existence of distinct groups. The result indicates that GSVs are dynamically retained in a continuous and wide range of mobility rather than into separate. classes.
机译:葡萄糖转运蛋白4(GLUT4)负责将胰岛素刺激的葡萄糖转运到对胰岛素敏感的脂肪和肌肉细胞中。 GLUT4储存囊泡(GSV)的动力学仍有待探索,并且尚不清楚如何根据其迁移率排列GSV。我们通过研究用EGFP融合的GLUT4标记的单个GSV的三维迁移率,研究了3T3-L1细胞中的这个问题。 。照射紧邻质膜的一层薄薄的胞质溶胶,并在全内反射荧光显微镜下以5 Hz的频率连续成像,穿透深度为136 nm。使用单粒子跟踪,在G处跟踪单个GSV的三维亚像素位移。 22 nm的空间精度。计算每个囊泡的均方位移和扩散系数。跟踪结果显示,囊泡似乎在受限于平均半径为160 nm的笼子中移动,表明存在一些细胞内束缚基质。通过构造GSV扩散系数的直方图,我们观察到了平滑的分布,而不是存在明显的基团。结果表明,GSV被动态保留在连续而又广泛的移动性范围内,而不是单独存在。类。

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