Morphological changes of plasma membrane and protein assembly during clathrin-mediated endocytosis.
Morphological changes of plasma membrane and protein assembly during clathrin-mediated endocytosis.
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DOI：
10.1371/journal.pbio.2004786
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发表时间：
2018-05
期刊：
影响因子：
9.8
通讯作者：
Yoshimura SH
中科院分区：
文献类型：
--
作者：
Yoshida A;Sakai N;Uekusa Y;Imaoka Y;Itagaki Y;Suzuki Y;Yoshimura SH
网格蛋白介导的胞吞作用（CME）是通过一系列蛋白组分诱导的质膜形态变化进行的。虽然这些蛋白质的时空组装已经通过基于荧光的技术阐明，但在活细胞中蛋白质诱导的质膜形态变化尚未完全澄清。在这里，我们利用高速原子力显微镜（HS-AFM）结合共聚焦激光扫描装置，在CME过程中可视化膜形态和蛋白质定位。在网格蛋白开始组装后约30 s，质膜开始内陷，孔径随着网格蛋白的积累而增大。肌动蛋白在窝周围迅速积聚，引起小的膜肿胀，在30秒内迅速不可逆地覆盖窝。抑制肌动蛋白的转换可消除肿胀，并诱导坑的可逆开合运动，表明肌动蛋白动力学对于CME结束时有效和不可逆的坑闭合是必要的。细胞通过质膜和各种膜蛋白与环境进行交流。网格蛋白介导的胞内吞噬（CME）在这种通讯中起着核心作用，并通过一系列多蛋白组装、质膜变形和膜泡的产生将细胞外信号分子传递到细胞质中。在本研究中，我们利用自制的高速原子力显微镜（HS-AFM）和共聚焦荧光显微镜组成的相关成像系统，同时对活细胞CME过程中质膜的形态变化和蛋白质定位进行了成像。结果显示，坑的大小和网格蛋白的数量紧密相关。肌动蛋白动力学在该过程的组装、成熟和关闭阶段发挥多种作用，并影响膜形态，表明内吞作用与细胞皮层的动态事件密切相关。动力蛋白的敲低也影响着坑的闭合运动，并与肌动蛋白表现出功能相关性。
Clathrin-mediated endocytosis (CME) proceeds through a series of morphological changes of the plasma membrane induced by a number of protein components. Although the spatiotemporal assembly of these proteins has been elucidated by fluorescence-based techniques, the protein-induced morphological changes of the plasma membrane have not been fully clarified in living cells. Here, we visualize membrane morphology together with protein localizations during CME by utilizing high-speed atomic force microscopy (HS-AFM) combined with a confocal laser scanning unit. The plasma membrane starts to invaginate approximately 30 s after clathrin starts to assemble, and the aperture diameter increases as clathrin accumulates. Actin rapidly accumulates around the pit and induces a small membrane swelling, which, within 30 s, rapidly covers the pit irreversibly. Inhibition of actin turnover abolishes the swelling and induces a reversible open–close motion of the pit, indicating that actin dynamics are necessary for efficient and irreversible pit closure at the end of CME. Cells communicate with their environments via the plasma membrane and various membrane proteins. Clathrin-mediated endocytosis (CME) plays a central role in such communication and proceeds with a series of multiprotein assembly, deformation of the plasma membrane, and production of a membrane vesicle that delivers extracellular signaling molecules into the cytoplasm. In this study, we utilized our home-built correlative imaging system comprising high-speed atomic force microscopy (HS-AFM) and confocal fluorescence microscopy to simultaneously image morphological changes of the plasma membrane and protein localization during CME in a living cell. The results revealed a tight correlation between the size of the pit and the amount of clathrin assembled. Actin dynamics play multiple roles in the assembly, maturation, and closing phases of the process, and affects membrane morphology, suggesting a close relationship between endocytosis and dynamic events at the cell cortex. Knock down of dynamin also affected the closing motion of the pit and showed functional correlation with actin.
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影响因子：
21.3
作者：
通讯作者：
--
DOI：
10.1111/tra.12152
发表时间：
2014-04
期刊：
Traffic (Copenhagen, Denmark)
影响因子：
--
作者：
Chandrasekar I;Goeckeler ZM;Turney SG;Wang P;Wysolmerski RB;Adelstein RS;Bridgman PC
通讯作者：
Bridgman PC
DOI：
10.1016/j.biocel.2006.12.004
发表时间：
2007-01-01
影响因子：
4
作者：
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通讯作者：
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影响因子：
9.2
作者：
Collins, Agnieszka;Warrington, Anthony;Taylor, Kenneth A.;Svitkina, Tatyana
通讯作者：
Svitkina, Tatyana
影响因子：
4
作者：
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