Endorepellin Affects Angiogenesis by Antagonizing Diverse Vascular Endothelial Growth Factor Receptor 2 (VEGFR2)-evoked Signaling Pathways

• Published in: The Journal of biological chemistry Vol. 287; no. 52; pp. 43543 - 43556
• Main Authors: Goyal, Atul, Poluzzi, Chiara, Willis, Chris D., Smythies, James, Shellard, Adam, Neill, Thomas, Iozzo, Renato V.
• Format: Journal Article
• Language: English
• Published: Elsevier Inc 01.12.2012
• Subjects: Angiogenesis; AP1; Calcineurin; Endothelial Cell; NFAT1; Perlecan; Proteoglycan; Vascular Biology; Vascular Endothelial Growth Factor (VEGF); VEGF Receptor 2; Proteoglycan; Angiogenesis; Vascular Endothelial Growth Factor (VEGF); AP1; VEGF Receptor 2; Vascular Biology; Calcineurin; Perlecan; Endothelial Cell; NFAT1
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M112.401786

Endorepellin, the angiostatic C-terminal domain of the heparan sulfate proteoglycan perlecan, inhibits angiogenesis by simultaneously binding to the α2β1 integrin and the vascular endothelial growth factor (VEGF) receptor 2 (VEGFR2) on endothelial cells. This interaction triggers the down-regulation of both receptors and the concurrent activation of the tyrosine phosphatase SHP-1, which leads to a signaling cascade resulting in angiostasis. Here, we provide evidence that endorepellin is capable of attenuating both the PI3K/PDK1/Akt/mTOR and the PKC/JNK/AP1 pathways. We show that hypoxia-inducible factor 1α (HIF-1α) transcriptional activity induced by VEGFA was inhibited by endorepellin independent of oxygen concentration and that only a combination of both PI3K and calcineurin inhibitors completely blocked the suppressive activity evoked by endorepellin on HIF1A and VEGFA promoter activity. Moreover, endorepellin inhibited the PKC/JNK/AP1 axis induced by the recruitment of phospholipase γ and attenuated the VEGFA-induced activation of NFAT1, a process dependent on calcineurin activity. Finally, endorepellin inhibited VEGFA-evoked nuclear translocation of NFAT1 and promoted NFAT1 stability. Thus, we provide evidence for a novel downstream signaling axis for an angiostatic fragment and for the key components involved in the dual antagonistic activity of endorepellin, highlighting its potential use as a therapeutic agent. Endorepellin specifically targets endothelial cells via dual-receptor antagonism of α2β1 and VEGFR2 to inhibit angiogenesis. Endorepellin attenuates two major signaling branches of VEGFR2 to transcriptionally repress HIF-1α concurrent with stabilized and cytoplasmically localized NFAT1. Endorepellin suppresses signaling of VEGFR2 independent of oxygen tension to inhibit angiogenesis. Endorepellin via dual-receptor antagonism provides novel mechanisms applicable to similar angiostatic fragments.