Endocytosis by Random Initiation and Stabilization of Clathrin-Coated Pits

• Published in: Cell Vol. 118; no. 5; pp. 591 - 605
• Main Authors: Ehrlich, Marcelo, Boll, Werner, van Oijen, Antoine, Hariharan, Ramesh, Chandran, Kartik, Nibert, Max L., Kirchhausen, Tomas
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 03.09.2004
• Subjects: Adaptor Protein Complex 2 - metabolism; Animals; Cell Line; Cell Membrane - metabolism; Cell Membrane - ultrastructure; Clathrin-Coated Vesicles - metabolism; Clathrin-Coated Vesicles - ultrastructure; Endocytosis - physiology; Endosomes - metabolism; Endosomes - ultrastructure; Green Fluorescent Proteins; Haplorhini; Lipoproteins, LDL - metabolism; Luminescent Proteins; Models, Biological; Protein Transport - physiology; Rats; Recombinant Fusion Proteins - metabolism; Red Fluorescent Protein; Reoviridae - metabolism; Reovirus; Transferrin - metabolism
• ISSN: 0092-8674; 1097-4172
• DOI: 10.1016/j.cell.2004.08.017

Clathrin-coated vesicles carry traffic from the plasma membrane to endosomes. We report here the real-time visualization of cargo sorting and endocytosis by clathrin-coated pits in living cells. We have detected the formation of coats by monitoring incorporation of fluorescently tagged clathrin or its adaptor AP-2; we have also followed clathrin-mediated uptake of transferrin and of single LDL or reovirus particles. The intensity of a cargo-loaded clathrin cluster grows steadily during its lifetime, and the time required to complete assembly is proportional to the size of the cargo particle. These results are consistent with a nucleation-growth mechanism and an approximately constant growth rate. There are no strongly preferred nucleation sites. A proportion of the nucleation events are weak and short lived. Cargo incorporation occurs primarily or exclusively in a newly formed coated pit. Our data lead to a model in which coated pits initiate randomly but collapse unless stabilized, perhaps by cargo capture.