KDR activation is crucial for VEGF165-mediated Ca2+ mobilization in human umbilical vein endothelial cells.
We have prepared a polyclonal mouse antibody directed against the first three immunoglobulin-like domains of the kinase insert domain-containing receptor (KDR) tyrosine kinase. It possesses the ability to inhibit binding of the 165-amino acid splice variant of vascular endothelial cell growth factor (VEGF165) to recombinant KDR in vitro as well as to reduce VEGF165 binding to human umbilical vein endothelial cells (HUVEC). These results confirm that the first three immunoglobulin-like domains of KDR are involved in VEGF165 interactions. The anti-KDR antibody is able to completely block VEGF165-mediated intracellular Ca2+ mobilization in HUVEC. Therefore, it appears that binding of VEGF165 to the fms-like tyrosine kinase (Flt-1) in these cells does not translate into a Ca2+ response. This is further exemplified by the lack of response to placental growth factor (PlGF), an Flt-1-specific ligand. Additionally, PlGF is unable to potentiate the effects of submaximal concentrations of VEGF165. Surprisingly, the VEGF-PlGF heterodimer was also very inefficient at eliciting a Ca2+ signaling event in HUVEC. We conclude that KDR activation is crucial for mobilization of intracellular Ca2+ in HUVEC in response to VEGF165.
Cunningham
S A
SA
Department of Pharmacology, Texas Biotechnology Corporation, Houston, 77030, Texas.
Tran
T M
TM
Arrate
M P
MP
Bjercke
R
R
Brock
T A
TA
D000818:Animals; D001692:Biological Transport / Q000502:physiology; D002118:Calcium / Q000378:metabolism*; D002478:Cells, Cultured; D016228:Endothelial Growth Factors / Q000502:physiology*; D004730:Endothelium, Vascular / Q000166:cytology / Q000378:metabolism*; D004789:Enzyme Activation / Q000502:physiology; D006801:Humans; D007158:Immunologic Techniques; D008222:Lymphokines / Q000502:physiology*; D051379:Mice; D020794:Receptor Protein-Tyrosine Kinases / Q000378:metabolism*; D017978:Receptors, Growth Factor / Q000378:metabolism*; D040262:Receptors, Vascular Endothelial Growth Factor; D011994:Recombinant Proteins; D014471:Umbilical Veins / Q000166:cytology / Q000378:metabolism*; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors
D016228:Endothelial Growth Factors; D008222:Lymphokines; D017978:Receptors, Growth Factor; D011994:Recombinant Proteins; C467484:VEGFA protein, human; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors; D020794:Receptor Protein-Tyrosine Kinases; D040262:Receptors, Vascular Endothelial Growth Factor; D002118:Calcium