Magnitude of Ubiquitination Determines the Fate of Epidermal Growth Factor Receptor Upon Ligand Stimulation

Receptor tyrosine kinases (RTK) bind growth factors and are critical for cell proliferation and differentiation. Their dysregulation leads to a loss of growth control, often resulting in cancer. Epidermal growth factor receptor (EGFR) is the prototypic RTK and can bind several ligands exhibiting distinct mitogenic potentials. Whereas the phosphorylation on individual EGFR sites and their roles for downstream signaling have been extensively studied, less is known about ligand-specific ubiquitination events on EGFR, which are crucial for signal attenuation and termination. We used a proteomics-based workflow for absolute quantitation combined with mathematical modeling to unveil potentially decisive ubiquitination events on EGFR from the first 30 seconds to 15 minutes of stimulation. Four ligands were used for stimulation: epidermal growth factor (EGF), heparin-binding-EGF like growth factor, transforming growth factor-a and epiregulin. Whereas only little differences in the order of individual ubiquitination sites were observed, the overall amount of modified receptor differed depending on the used ligand, indicating that absolute magnitude of EGFR ubiquitination, and not distinctly regulated ubiquitination sites, is a major determinant for signal attenuation and the subsequent cellular outcomes. (C) 2021 The Author(s). Published by Elsevier Ltd.

Automated summary

Receptor tyrosine kinases (RTK) play crucial roles in cell proliferation and differentiation by binding growth factors, with EGFR being a prototypic RTK that can bind multiple ligands. A proteomics-based workflow combined with mathematical modeling revealed that ligand-specific ubiquitination events on EGFR play a significant role in signal attenuation and termination. The absolute magnitude of EGFR ubiquitination, rather than distinctly regulated ubiquitination sites, is a major determinant for signal attenuation and subsequent cellular outcomes.