title: VEGF binding to NRP1 is essential for VEGF stimulation of endothelial cell migration, complex formation between NRP1 and VEGFR2, and signaling via FAK Tyr407 phosphorylation.
abstract: In endothelial cells, neuropilin-1 (NRP1) binds vascular endothelial growth factor (VEGF)-A and is thought to act as a coreceptor for kinase insert domain-containing receptor (KDR) by associating with KDR and enhancing VEGF signaling. Here we report mutations in the NRP1 b1 domain (Y297A and D320A), which result in complete loss of VEGF binding. Overexpression of Y297A and D320A NRP1 in human umbilical vein endothelial cells reduced high-affinity VEGF binding and migration toward a VEGF gradient, and markedly inhibited VEGF-induced angiogenesis in a coculture cell model. The Y297A NRP1 mutant also disrupted complexation between NRP1 and KDR and decreased VEGF-dependent phosphorylation of focal adhesion kinase at Tyr407, but had little effect on other signaling pathways. Y297A NRP1, however, heterodimerized with wild-type NRP1 and NRP2 indicating that nonbinding NRP1 mutants can act in a dominant-negative manner through formation of NRP1 dimers with reduced binding affinity for VEGF. These findings indicate that VEGF binding to NRP1 has specific effects on endothelial cell signaling and is important for endothelial cell migration and angiogenesis mediated via complex formation between NRP1 and KDR and increased signaling to focal adhesions. Identification of key residues essential for VEGF binding and biological functions provides the basis for a rational design of antagonists of VEGF binding to NRP1.
authors.affiliation: BHF Laboratories, University College London, London WC1E 6JJ, UK.
authors.forename: Birger
authors.initials: B
authors.lastname: Herzog
authors.forename: Caroline
authors.initials: C
authors.lastname: Pellet-Many
authors.forename: Gary
authors.initials: G
authors.lastname: Britton
authors.forename: Basil
authors.initials: B
authors.lastname: Hartzoulakis
authors.forename: Ian C
authors.initials: IC
authors.lastname: Zachary
mesh_terms: D000818:Animals; D001665:Binding Sites; D002448:Cell Adhesion; D002465:Cell Movement; D018920:Coculture Techniques; D019281:Dimerization; D042783:Endothelial Cells / Q000166:cytology / Q000378:metabolism*; D004730:Endothelium, Vascular / Q000166:cytology / Q000378:metabolism*; D051417:Focal Adhesion Kinase 1 / Q000235:genetics / Q000378:metabolism*; D022001:Focal Adhesions / Q000378:metabolism*; D006801:Humans; D009154:Mutation; D018919:Neovascularization, Physiologic; D039942:Neuropilin-1 / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D010957:Plasmids; D011485:Protein Binding; D015398:Signal Transduction / Q000502:physiology; D014162:Transfection; D014471:Umbilical Veins / Q000166:cytology / Q000378:metabolism; D042461:Vascular Endothelial Growth Factor A / Q000235:genetics / Q000378:metabolism*; D040301:Vascular Endothelial Growth Factor Receptor-2 / Q000235:genetics / Q000378:metabolism*
chemical_list: C467484:VEGFA protein, human; D042461:Vascular Endothelial Growth Factor A; D039942:Neuropilin-1; D040301:Vascular Endothelial Growth Factor Receptor-2; D051417:Focal Adhesion Kinase 1