Ligand-stimulated VEGFR2 signaling is regulated by co-ordinated trafficking and proteolysis

Traffic. 2010 Jan;11(1):161-74. doi: 10.1111/j.1600-0854.2009.01001.x.


Vascular endothelial growth factor A (VEGF-A)-induced signaling through VEGF receptor 2 (VEGFR2) regulates both physiological and pathological angiogenesis in mammals. However, the temporal and spatial mechanism underlying VEGFR2-mediated intracellular signaling is not clear. Here, we define a pathway for VEGFR2 trafficking and proteolysis that regulates VEGF-A-stimulated signaling and endothelial cell migration. Ligand-stimulated VEGFR2 activation and ubiquitination preceded proteolysis and cytoplasmic domain removal associated with endosomes. A soluble VEGFR2 cytoplasmic domain fragment displayed tyrosine phosphorylation and activation of downstream intracellular signaling. Perturbation of endocytosis by the depletion of either clathrin heavy chain or an ESCRT-0 subunit caused differential effects on ligand-stimulated VEGFR2 proteolysis and signaling. This novel VEGFR2 proteolysis was blocked by the inhibitors of 26S proteasome activity. Inhibition of proteasome activity prolonged VEGF-A-induced intracellular signaling to c-Akt and endothelial nitric oxide synthase (eNOS). VEGF-A-stimulated endothelial cell migration was dependent on VEGFR2 and VEGFR tyrosine kinase activity. Inhibition of proteasome activity in this assay stimulated VEGF-A-mediated endothelial cell migration. VEGFR2 endocytosis, ubiquitination and proteolysis could also be stimulated by a protein kinase C-dependent pathway. Thus, removal of the VEGFR2 carboxyl terminus linked to phosphorylation, ubiquitination and trafficking is necessary for VEGF-stimulated endothelial signaling and cell migration.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Cell Line
  • Cell Movement / drug effects
  • Cyclic AMP-Dependent Protein Kinases / metabolism
  • Cytoplasm / drug effects
  • Cytoplasm / enzymology
  • Cytoplasm / metabolism
  • Electrophoresis, Polyacrylamide Gel
  • Endosomes / drug effects
  • Endosomes / enzymology
  • Endosomes / metabolism
  • Endothelial Cells / drug effects*
  • Endothelial Cells / enzymology
  • Endothelial Cells / metabolism
  • Endothelial Cells / pathology
  • Endothelium, Vascular / drug effects
  • Endothelium, Vascular / enzymology
  • Endothelium, Vascular / metabolism
  • Endothelium, Vascular / pathology
  • Humans
  • Ligands
  • Lysosomes / drug effects
  • Lysosomes / enzymology
  • Lysosomes / metabolism
  • Microscopy, Fluorescence
  • Neovascularization, Pathologic / enzymology
  • Neovascularization, Pathologic / metabolism
  • Proteasome Endopeptidase Complex / metabolism
  • Protein Binding
  • Protein Transport
  • Signal Transduction / drug effects*
  • Vascular Endothelial Growth Factor A / metabolism*
  • Vascular Endothelial Growth Factor Receptor-2 / metabolism*


  • Ligands
  • VEGFA protein, human
  • Vascular Endothelial Growth Factor A
  • Vascular Endothelial Growth Factor Receptor-2
  • Cyclic AMP-Dependent Protein Kinases
  • Proteasome Endopeptidase Complex