Src kinase becomes preferentially associated with the VEGFR, KDR/Flk-1, following VEGF stimulation of vascular endothelial cells

BMC Biochem. 2002 Dec 31;3:32. doi: 10.1186/1471-2091-3-32. Epub 2002 Dec 31.

Abstract

Background: The cytoplasmic tyrosine kinase, Src, has been found to play a crucial role in VEGF (vascular endothelial growth factor) - dependent vascular permeability involved in angiogenesis. The two main VEGFRs present on vascular endothelial cells are KDR/Flk-1 (kinase insert domain-containing receptor/fetal liver kinase-1) and Flt-1 (Fms-like tyrosine kinase-1). However, to date, it has not been determined which VEGF receptor (VEGFR) is involved in binding to and activating Src kinase following VEGF stimulation of the receptors.

Results: In this report, we demonstrate that Src preferentially associates with KDR/Flk-1 rather than Flt-1 in human umbilical vein endothelial cells (HUVECs), and that VEGF stimulation resulted in an increase of Src activity associated with activated KDR/Flk-1. These findings were determined through immunoprecipitation-kinase experiments and coimmunoprecipitation studies, and were further confirmed by GST-pull-down assays and Far Western studies. However, Fyn and Yes, unlike Src, were found to associate preferentially with Flt-1.

Conclusions: Thus, Src preferentially associates with KDR/Flk-1, rather than with Flt-1, upon VEGF stimulation in endothelial cells. Our findings further highlight the potential significance of upregulated KDR/Flk-1-associated Src activity in the process of angiogenesis, and help to elucidate more clearly the specific roles and mechanisms involving Src family tyrosine kinase in VEGF-stimulated signal transduction events.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • CSK Tyrosine-Protein Kinase
  • Cells, Cultured
  • Cytoplasm / enzymology
  • Endothelial Growth Factors / isolation & purification
  • Endothelial Growth Factors / metabolism*
  • Endothelium, Vascular / chemistry
  • Endothelium, Vascular / cytology
  • Endothelium, Vascular / enzymology*
  • Endothelium, Vascular / metabolism*
  • Enzyme Activation / physiology
  • Glutathione Transferase / biosynthesis
  • Glutathione Transferase / genetics
  • Humans
  • Intercellular Signaling Peptides and Proteins / isolation & purification
  • Intercellular Signaling Peptides and Proteins / metabolism*
  • Lymphokines / isolation & purification
  • Lymphokines / metabolism*
  • Peptide Mapping
  • Precipitin Tests
  • Protein-Tyrosine Kinases / biosynthesis
  • Protein-Tyrosine Kinases / genetics
  • Protein-Tyrosine Kinases / metabolism*
  • Protein-Tyrosine Kinases / physiology
  • Proto-Oncogene Proteins / isolation & purification
  • Proto-Oncogene Proteins c-fyn
  • Proto-Oncogene Proteins c-yes
  • Recombinant Fusion Proteins / biosynthesis
  • Recombinant Fusion Proteins / genetics
  • Time Factors
  • Umbilical Veins
  • Vascular Endothelial Growth Factor A
  • Vascular Endothelial Growth Factor Receptor-1 / immunology
  • Vascular Endothelial Growth Factor Receptor-1 / isolation & purification
  • Vascular Endothelial Growth Factor Receptor-1 / metabolism*
  • Vascular Endothelial Growth Factor Receptor-1 / physiology
  • Vascular Endothelial Growth Factor Receptor-2 / isolation & purification
  • Vascular Endothelial Growth Factor Receptor-2 / metabolism*
  • Vascular Endothelial Growth Factor Receptor-2 / physiology
  • Vascular Endothelial Growth Factors
  • src Homology Domains / physiology
  • src-Family Kinases*

Substances

  • Endothelial Growth Factors
  • Intercellular Signaling Peptides and Proteins
  • Lymphokines
  • Proto-Oncogene Proteins
  • Recombinant Fusion Proteins
  • Vascular Endothelial Growth Factor A
  • Vascular Endothelial Growth Factors
  • Glutathione Transferase
  • Protein-Tyrosine Kinases
  • Vascular Endothelial Growth Factor Receptor-1
  • Vascular Endothelial Growth Factor Receptor-2
  • CSK Tyrosine-Protein Kinase
  • FYN protein, human
  • Proto-Oncogene Proteins c-fyn
  • Proto-Oncogene Proteins c-yes
  • src-Family Kinases
  • CSK protein, human