Netrin-4 inhibits angiogenesis via binding to neogenin and recruitment of Unc5B.
Netrins are secreted molecules with roles in axon guidance and angiogenesis. We identified Netrin-4 as a gene specifically overexpressed in VEGF-stimulated endothelial cells (EC) in vitro as well as in vivo. Knockdown of Netrin-4 expression in EC increased their ability to form tubular structures on Matrigel. To identify which receptor is involved, we showed by quantitative RT-PCR that EC express three of the six Netrin-1 cognate receptors: neogenin, Unc5B, and Unc5C. In contrast to Netrin-1, Netrin-4 bound only to neogenin but not to Unc5B or Unc5C receptors. Neutralization of Netrin-4 binding to neogenin by blocking antibodies abolished the chemotactic effect of Netrin-4. Furthermore, the silencing of either neogenin or Unc5B abolished Netrin-4 inhibitory effect on EC migration, suggesting that both receptors are essential for its function in vitro. Coimmunoprecipitation experiments demonstrated that Netrin-4 increased the association between Unc5B and neogenin on VEGF- or FGF-2-stimulated EC. Finally, we showed that Netrin-4 significantly reduced pathological angiogenesis in Matrigel and laser-induced choroidal neovascularization models. Interestingly, Netrin-4, neogenin, and Unc5B receptor expression was up-regulated in choroidal neovessel EC after laser injury. Moreover, Netrin-4 overexpression delayed tumor angiogenesis in a model of s.c. xenograft. We propose that Netrin-4 acts as an antiangiogenic factor through binding to neogenin and recruitment of Unc5B.
Lejmi
Esma
E
Institut National de la Santé et de la Recherche Médicale, Unité 689/Institut des Vaisseaux et du Sang, F-75010 Paris, France.
Leconte
Laurence
L
Pédron-Mazoyer
Sandrine
S
Ropert
Stanislas
S
Raoul
William
W
Lavalette
Sophie
S
Bouras
Ilyes
I
Feron
Jean-Guillaume
JG
Maitre-Boube
Martine
M
Assayag
Franck
F
Feumi
Charles
C
Alemany
Monica
M
Jie
Tan Xian
TX
Merkulova
Tatyana
T
Poupon
Marie-France
MF
Ruchoux
Marie-Magdeleine
MM
Tobelem
Gérard
G
Sennlaub
Florian
F
Plouët
Jean
J
D000818:Animals; D002417:Cattle; D045744:Cell Line, Tumor; D002478:Cells, Cultured; D002633:Chemotaxis; D042783:Endothelial Cells / Q000166:cytology*; D005260:Female; D006801:Humans; D007834:Lasers / Q000009:adverse effects; D008297:Male; D008565:Membrane Proteins / Q000378:metabolism*; D051379:Mice; D008807:Mice, Inbred BALB C; D008819:Mice, Nude; D009374:Neoplasms, Experimental / Q000098:blood supply; D009389:Neovascularization, Pathologic; D009414:Nerve Growth Factors / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D000075386:Netrin Receptors; D000075385:Netrins; D011471:Prostatic Neoplasms / Q000473:pathology; D011485:Protein Binding / Q000502:physiology; D011956:Receptors, Cell Surface / Q000378:metabolism*; D011994:Recombinant Proteins / Q000494:pharmacology; D014183:Transplantation, Heterologous; D015854:Up-Regulation / Q000235:genetics
D008565:Membrane Proteins; C498815:NTN4 protein, human; D009414:Nerve Growth Factors; D000075386:Netrin Receptors; D000075385:Netrins; C416214:Ntn4 protein, mouse; D011956:Receptors, Cell Surface; D011994:Recombinant Proteins; C532445:UNC5B protein, human; C091279:neogenin