Proteomic analysis of tyrosine phosphorylations in vascular endothelial growth factor- and reactive oxygen species-mediated signaling pathway.
Vascular endothelial growth factor (VEGF) mediates angiogenic signaling by activating tyrosine kinase receptors. Endothelial cells treated with VEGF are known to increase reactive oxygen species (ROS) production and activate the MAPK pathway. To identify the target proteins of the VEGF receptor, we treated human umbilical vein endothelial cells (HUVECs) with VEGF or H2O2, and identified and semiquantified tyrosine-phosphorylated proteins, combining 2D-gel electrophoresis, Western analysis using antibody against phospho-tyrosine, and mass spectrometry. We detected 95 proteins that were differentially phosphorylated; some were specifically phosphorylated by VEGF but not by H2O2. 2D-gel electrophoresis revealed that heterogeneous populations of the same protein responded differently to H2O2 and VEGF. Bioinformatic studies examining the nature of the differential phosphorylation in various subpopulations of proteins should provide new insights into VEGF- and H2O2-induced signaling pathways.
Kim
Young Mee
YM
The Center for Cell Signaling & Drug Discovery Research, College of Pharmacy and Division of Life & Pharmaceutical Sciences, Ewha Womans University, Seoul 120-750, Korea.
Song
Eun Joo
EJ
Seo
Jawon
J
Kim
Hee-Jung
HJ
Lee
Kong-Joo
KJ
D015180:Electrophoresis, Gel, Two-Dimensional; D004730:Endothelium, Vascular / Q000378:metabolism / Q000502:physiology*; D005982:Glutathione Transferase / Q000378:metabolism; D006801:Humans; D006861:Hydrogen Peroxide / Q000378:metabolism; D013058:Mass Spectrometry; D010766:Phosphorylation; D019000:Phosphotyrosine / Q000032:analysis* / Q000737:chemistry; D020543:Proteome / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D017382:Reactive Oxygen Species / Q000378:metabolism*; D015398:Signal Transduction / Q000502:physiology*; D014404:Tubulin / Q000378:metabolism; D014471:Umbilical Veins; D042461:Vascular Endothelial Growth Factor A / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism
D020543:Proteome; D017382:Reactive Oxygen Species; D014404:Tubulin; D042461:Vascular Endothelial Growth Factor A; D019000:Phosphotyrosine; D006861:Hydrogen Peroxide; D005982:Glutathione Transferase; D048051:p38 Mitogen-Activated Protein Kinases