Fluorescent resonance energy transfer imaging of VEGFR dimerization.
Activation of vascular endothelial growth factor receptor 1 (VEGFR1/FLT1) and 2 (VEGFR2/KDR) involves receptor dimerization. Formation of VEGFR dimer has so far not been visualized in single intact cells. In the present study we describe different optical assays which can be used to observe dimerization of VEGFR1 and VEGFR2. Bimolecular fluorescence complementation (BIFC) assays confirmed homo,- and heterodimerization of transfected receptors. Fluorescence resonance energy transfer (FRET) techniques in living and fixed CHO-K1 cells allowed observation of VEGFR1 homodimer,- and VEGFR1 and VEGFR2 heterodimer formation after ligand stimulation. After inhibition of ligand binding by the VEGFA JH121 antibody VEGFR1 homodimerization was completely abolished. Under the same conditions, cells transfected by VEGFR1 and VEGFR2 maintained relevant receptor heterodimerization. These techniques to monitor VEGFR1 and VEGFR2 homo- and heterodimerization in living and fixed cells may help in the search for new angiogenesis-directed inhibitors of VEGFR dimerization.
Clinic for Vascular and Endovascular Surgery, University Hospital Frankfurt, Frankfurt, Germany. Michael.keese@kgu.de.
Zhala
Z
Ahmadova
Vugar
V
Yagublu
Tassilo
T
Förg
Yegana
Y
Hajiyeva
Ralf
R
Jesenofsky
Mathias
M
Hafner
Michael
M
Keese
D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D031541:Fluorescence Resonance Energy Transfer / Q000379:methods*; D006801:Humans; D055503:Protein Multimerization / Q000502:physiology*; D040281:Vascular Endothelial Growth Factor Receptor-1 / Q000737:chemistry* / Q000378:metabolism; D040301:Vascular Endothelial Growth Factor Receptor-2 / Q000737:chemistry* / Q000378:metabolism
D040281:Vascular Endothelial Growth Factor Receptor-1; D040301:Vascular Endothelial Growth Factor Receptor-2
BIFC; FLT1; FRET; KDR; VEGF; VEGFR1; VEGFR2; dimerization; vascular endothelial growth factor receptors