Endocytosis by random initiation and stabilization of clathrin-coated pits

Cell. 2004 Sep 3;118(5):591-605. doi: 10.1016/j.cell.2004.08.017.

Abstract

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.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • 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
  • Reoviridae / metabolism
  • Transferrin / metabolism

Substances

  • Adaptor Protein Complex 2
  • Lipoproteins, LDL
  • Luminescent Proteins
  • Recombinant Fusion Proteins
  • Transferrin
  • red fluorescent protein
  • Green Fluorescent Proteins