Tyrosine Residues 951 and 1059 of Vascular Endothelial Growth Factor Receptor-2 (KDR) Are Essential for Vascular Permeability Factor/Vascular Endothelial Growth Factor-induced Endothelium Migration and Proliferation, Respectively

Huiyan Zeng, Sohini Sanyal, Debabrata Mukhopadhyay

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124 Citations (Scopus)

Abstract

Vascular permeability factor/vascular endothelial growth factor (VPF/VEGF) exerts its multiple functions by activating two receptor tyrosine kinases, Flt-1 (VEGFR-1) and KDR (VEGFR-2), both of which are selectively expressed on primary vascular endothelium. To dissect the respective signaling pathways and biological functions mediated by these receptors in primary endothelial cells with two receptors intact, we, recently developed chimeric receptors (EGDR and EGLT) in which the extracellular domain of the epidermal growth factor receptor was fused to the transmembrane domain and intracellular domain of KDR and Flt-1, respectively. With these fusion receptors, we have shown that KDR is solely responsible for VPF/VEGF-induced human umbilical vein endothelial cell (HUVEC) proliferation and migration, whereas Flt-1 showed an inhibitory effect on KDR-mediated proliferation but not migration. To further characterize the VPF/VEGF-stimulated HUVEC proliferation and migration here, we have created several EGDR mutants by site-directed mutagenesis. We show that tyrosine residues 1059 and 951 of KDR are essential for VPF/VEGF-induced HUVEC proliferation and migration, respectively. Furthermore, the mutation of tyrosine 1059 to phenylanaline results in the complete loss of KDR/ EGDR-mediated intracellular Ca2+ mobilization and MAPK phosphorylation, but the mutation of tyrosine 951 to phenylanaline did not affect these events. Our results suggest that KDR mediates different signaling pathways for HUVEC proliferation and migration and, moreover, intracellular Ca2+ mobilization and MAPK phosphorylation are not essential for VPF/VEGF-induced HUVEC migration.

Original languageEnglish (US)
Pages (from-to)32714-32719
Number of pages6
JournalJournal of Biological Chemistry
Volume276
Issue number35
DOIs
StatePublished - Aug 31 2001
Externally publishedYes

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Vascular Endothelial Growth Factor Receptor-2
Vascular Endothelial Growth Factor A
Endothelium
Tyrosine
Endothelial cells
Human Umbilical Vein Endothelial Cells
Cell Movement
Cell proliferation
Cell Proliferation
Phosphorylation
Vascular Endothelial Growth Factor Receptor-1
Mutagenesis
Mutation
Vascular Endothelium
Receptor Protein-Tyrosine Kinases
Site-Directed Mutagenesis
Epidermal Growth Factor Receptor
Fusion reactions
Endothelial Cells

ASJC Scopus subject areas

  • Biochemistry

Cite this

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title = "Tyrosine Residues 951 and 1059 of Vascular Endothelial Growth Factor Receptor-2 (KDR) Are Essential for Vascular Permeability Factor/Vascular Endothelial Growth Factor-induced Endothelium Migration and Proliferation, Respectively",
abstract = "Vascular permeability factor/vascular endothelial growth factor (VPF/VEGF) exerts its multiple functions by activating two receptor tyrosine kinases, Flt-1 (VEGFR-1) and KDR (VEGFR-2), both of which are selectively expressed on primary vascular endothelium. To dissect the respective signaling pathways and biological functions mediated by these receptors in primary endothelial cells with two receptors intact, we, recently developed chimeric receptors (EGDR and EGLT) in which the extracellular domain of the epidermal growth factor receptor was fused to the transmembrane domain and intracellular domain of KDR and Flt-1, respectively. With these fusion receptors, we have shown that KDR is solely responsible for VPF/VEGF-induced human umbilical vein endothelial cell (HUVEC) proliferation and migration, whereas Flt-1 showed an inhibitory effect on KDR-mediated proliferation but not migration. To further characterize the VPF/VEGF-stimulated HUVEC proliferation and migration here, we have created several EGDR mutants by site-directed mutagenesis. We show that tyrosine residues 1059 and 951 of KDR are essential for VPF/VEGF-induced HUVEC proliferation and migration, respectively. Furthermore, the mutation of tyrosine 1059 to phenylanaline results in the complete loss of KDR/ EGDR-mediated intracellular Ca2+ mobilization and MAPK phosphorylation, but the mutation of tyrosine 951 to phenylanaline did not affect these events. Our results suggest that KDR mediates different signaling pathways for HUVEC proliferation and migration and, moreover, intracellular Ca2+ mobilization and MAPK phosphorylation are not essential for VPF/VEGF-induced HUVEC migration.",
author = "Huiyan Zeng and Sohini Sanyal and Debabrata Mukhopadhyay",
year = "2001",
month = "8",
day = "31",
doi = "10.1074/jbc.M103130200",
language = "English (US)",
volume = "276",
pages = "32714--32719",
journal = "Journal of Biological Chemistry",
issn = "0021-9258",
publisher = "American Society for Biochemistry and Molecular Biology Inc.",
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T1 - Tyrosine Residues 951 and 1059 of Vascular Endothelial Growth Factor Receptor-2 (KDR) Are Essential for Vascular Permeability Factor/Vascular Endothelial Growth Factor-induced Endothelium Migration and Proliferation, Respectively

AU - Zeng, Huiyan

AU - Sanyal, Sohini

AU - Mukhopadhyay, Debabrata

PY - 2001/8/31

Y1 - 2001/8/31

N2 - Vascular permeability factor/vascular endothelial growth factor (VPF/VEGF) exerts its multiple functions by activating two receptor tyrosine kinases, Flt-1 (VEGFR-1) and KDR (VEGFR-2), both of which are selectively expressed on primary vascular endothelium. To dissect the respective signaling pathways and biological functions mediated by these receptors in primary endothelial cells with two receptors intact, we, recently developed chimeric receptors (EGDR and EGLT) in which the extracellular domain of the epidermal growth factor receptor was fused to the transmembrane domain and intracellular domain of KDR and Flt-1, respectively. With these fusion receptors, we have shown that KDR is solely responsible for VPF/VEGF-induced human umbilical vein endothelial cell (HUVEC) proliferation and migration, whereas Flt-1 showed an inhibitory effect on KDR-mediated proliferation but not migration. To further characterize the VPF/VEGF-stimulated HUVEC proliferation and migration here, we have created several EGDR mutants by site-directed mutagenesis. We show that tyrosine residues 1059 and 951 of KDR are essential for VPF/VEGF-induced HUVEC proliferation and migration, respectively. Furthermore, the mutation of tyrosine 1059 to phenylanaline results in the complete loss of KDR/ EGDR-mediated intracellular Ca2+ mobilization and MAPK phosphorylation, but the mutation of tyrosine 951 to phenylanaline did not affect these events. Our results suggest that KDR mediates different signaling pathways for HUVEC proliferation and migration and, moreover, intracellular Ca2+ mobilization and MAPK phosphorylation are not essential for VPF/VEGF-induced HUVEC migration.

AB - Vascular permeability factor/vascular endothelial growth factor (VPF/VEGF) exerts its multiple functions by activating two receptor tyrosine kinases, Flt-1 (VEGFR-1) and KDR (VEGFR-2), both of which are selectively expressed on primary vascular endothelium. To dissect the respective signaling pathways and biological functions mediated by these receptors in primary endothelial cells with two receptors intact, we, recently developed chimeric receptors (EGDR and EGLT) in which the extracellular domain of the epidermal growth factor receptor was fused to the transmembrane domain and intracellular domain of KDR and Flt-1, respectively. With these fusion receptors, we have shown that KDR is solely responsible for VPF/VEGF-induced human umbilical vein endothelial cell (HUVEC) proliferation and migration, whereas Flt-1 showed an inhibitory effect on KDR-mediated proliferation but not migration. To further characterize the VPF/VEGF-stimulated HUVEC proliferation and migration here, we have created several EGDR mutants by site-directed mutagenesis. We show that tyrosine residues 1059 and 951 of KDR are essential for VPF/VEGF-induced HUVEC proliferation and migration, respectively. Furthermore, the mutation of tyrosine 1059 to phenylanaline results in the complete loss of KDR/ EGDR-mediated intracellular Ca2+ mobilization and MAPK phosphorylation, but the mutation of tyrosine 951 to phenylanaline did not affect these events. Our results suggest that KDR mediates different signaling pathways for HUVEC proliferation and migration and, moreover, intracellular Ca2+ mobilization and MAPK phosphorylation are not essential for VPF/VEGF-induced HUVEC migration.

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