Quantitative visualization of endocytic trafficking through photoactivation of fluorescent proteins

Mol Biol Cell. 2021 Apr 19;32(9):892-902. doi: 10.1091/mbc.E20-10-0669. Epub 2021 Feb 3.

Abstract

Endocytic trafficking controls the density of molecules at the plasma membrane and by doing so, the cell surface profile, which in turn determines how cells interact with their environment. A full apprehension of any cellular process necessitates understanding how proteins associated with the plasma membrane are endocytosed, how they are sorted after internalization, and if and how they are recycled to the plasma membrane. To date, it is still difficult to experimentally gain access to this information, even more to do it in a quantitative way. Here we present a toolset based on photoactivation of fluorescent proteins that enabled us to generate quantitative information on endocytosis, incorporation into sorting and recycling endosomes, delivery from endosomes to the plasma membrane, and on the type of vesicles performing intracellular transport. We illustrate these approaches by revealing striking differences in the endocytic trafficking of T-cell receptor and CD4, which bind to the same molecule at the surface of antigen-presenting cells during T-cell activation.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Biological Transport
  • Cell Membrane / physiology
  • Endocytosis / physiology
  • Endosomes / metabolism
  • Fluorescent Antibody Technique / methods*
  • Humans
  • Jurkat Cells
  • Protein Transport / physiology*
  • Proteins / metabolism
  • Transport Vesicles / metabolism
  • Transport Vesicles / physiology*

Substances

  • Proteins