Morphological changes of plasma membrane and protein assembly during clathrin-mediated endocytosis
| Autor: | Shige H. Yoshimura, Yoshitsuna Itagaki, Aiko Yoshida, Yuka Imaoka, Yuki Suzuki, Nobuaki Sakai, Yoshitsugu Uekusa |
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| Rok vydání: | 2017 |
| Předmět: |
0301 basic medicine
Fluorescence-lifetime imaging microscopy Cell Membranes Microscopy Atomic Force Biochemistry Contractile Proteins Chlorocebus aethiops Biology (General) Microscopy Microscopy Confocal biology Chemistry Atomic force microscopy General Neuroscience Vesicle Clathrin-Coated Vesicles Endocytosis Atomic Force Microscopy Cell biology Membrane Cell Processes COS Cells Cellular Structures and Organelles General Agricultural and Biological Sciences Research Article Dynamins Imaging Techniques QH301-705.5 Confocal Research and Analysis Methods Green Fluorescent Protein Clathrin General Biochemistry Genetics and Molecular Biology 03 medical and health sciences Fluorescence Imaging Animals Vesicles Actin General Immunology and Microbiology Scanning Probe Microscopy Biology and Life Sciences Membrane Proteins Proteins Cell Biology Plasma Receptor-mediated endocytosis Actins Cytoskeletal Proteins Luminescent Proteins 030104 developmental biology Membrane protein biology.protein Biophysics Actin Polymerization |
| Zdroj: | PLoS Biology, Vol 16, Iss 5, p e2004786 (2018) PLoS Biology |
| Popis: | 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. Author summary 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. |
| Databáze: | OpenAIRE |
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