Regulation of epidermal growth factor receptor signaling by endocytosis and intracellular trafficking.

Ligand activation of the epidermal growth factor receptor (EGFR) leads to its rapid internalization and eventual delivery to lysosomes. This process is thought to be a mechanism to attenuate signaling, but signals could potentially be generated after endocytosis. To directly evaluate EGFR signaling during receptor trafficking, we developed a technique ...
to rapidly and selectively isolate internalized EGFR and associated molecules with the use of reversibly biotinylated anti-EGFR antibodies. In addition, we developed antibodies specific to tyrosine-phosphorylated EGFR. With the use of a combination of fluorescence imaging and affinity precipitation approaches, we evaluated the state of EGFR activation and substrate association during trafficking in epithelial cells. We found that after internalization, EGFR remained active in the early endosomes. However, receptors were inactivated before degradation, apparently due to ligand removal from endosomes. Adapter molecules, such as Shc, were associated with EGFR both at the cell surface and within endosomes. Some molecules, such as Grb2, were primarily found associated with surface EGFR, whereas others, such as Eps8, were found only with intracellular receptors. During the inactivation phase, c-Cbl became EGFR associated, consistent with its postulated role in receptor attenuation. We conclude that the association of the EGFR with different proteins is compartment specific. In addition, ligand loss is the proximal cause of EGFR inactivation. Thus, regulated trafficking could potentially influence the pattern as well as the duration of signal transduction.
Mesh Terms:
Adaptor Proteins, Signal Transducing, Adaptor Proteins, Vesicular Transport, Cell Line, Endocytosis, Endosomes, Enzyme Activation, Epithelial Cells, Humans, Intracellular Signaling Peptides and Proteins, Ligands, Microscopy, Fluorescence, Models, Biological, Phosphorylation, Phosphotyrosine, Protein Binding, Proteins, Receptor, Epidermal Growth Factor, Shc Signaling Adaptor Proteins, Signal Transduction, Temperature, Time Factors, Tyrosine
Mol. Biol. Cell
Date: Jun. 01, 2001
Download Curated Data For This Publication
141935
Switch View:
  • Interactions 5
  • Chemical Interactions 1