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Phosphorylation of tyrosine 1214 on VEGFR2 is required for VEGF-induced activation of Cdc42 upstream of SAPK2/p38

Oncogene volume 23, pages 434–445 (2004)Cite this article

Abstract

Activation of the tyrosine kinase receptor vascular endothelial growth factor receptor 2 (VEGFR2) by VEGF leads to the activation of stress-activated protein kinase (SAPK)2/p38 and then to actin polymerization and reorganization into stress fibers in endothelial cells. In turn, this triggers endothelial cell migration. Yet, nothing is known about the molecular mechanisms that couple VEGFR2 to SAPK2/p38. Here, we found that VEGF increased by twofold the activity of the small GTPase Cdc42 and that the expression of two different constitutively active forms of Cdc42 (Cdc42 V12 and Cdc42 L61) led to a marked increase in the formation of stress fibers that was sensitive to SAPK2/p38 inhibition by SB203580. Moreover, the expression of a dominant-negative form of Cdc42 (Cdc42 N17) inhibited the activation of SAPK2/p38 and of its direct target MAP kinase-activated protein kinase 2. These results indicate that Cdc42 is upstream of SAPK2/p38 in response to the activation of VEGFR2 by VEGF. In contrast, we found that neither RhoA nor Rac was involved in the SAPK2/p38-mediated actin reorganization induced by VEGF. Using a site-specific mutant of the major autophosphorylation site Y1214 on VEGFR2, we found that the mutant Y1214F inhibited the activation of both Cdc42 and SAPK2/p38 in response to VEGF. We conclude that phosphorylation of Y1214 on VEGFR2 is required to trigger the sequential activation of Cdc42 and SAPK2/p38 and to drive the SAPK2/p38-mediated actin remodeling in stress fibers in endothelial cells exposed to VEGF.

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Abbreviations

ECGS:

endothelial cell growth supplement

FAK:

focal adhesion kinase

HSP:

heat-shock protein

HUVEC:

human umbilical vein endothelial cells

MAP kinase:

mitogen-activated protein kinase

MAPKAP K2:

MAP kinase-activated protein kinase 2

PAE:

porcine aortic endothelial cells

SAPK:

stress-activated protein kinase

VEGF:

vascular endothelial growth factor

VEGFR2:

vascular endothelial growth factor receptor 2

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Acknowledgements

We thank Drs Nathalie Lamarche and Louise Larose from McGill University and Sylvain Bourgoin (Université Laval, Québec) for providing the various GTPase constructs and the GST-CRIB protein. We also thank Dr Jacques Landry (Québec) for providing the 9E10 antibody and NIH 3T3 cells and Drs Bruce Terman (New York) and Johannes Waltenberger (Ulm, Germany) for the VEGFR2 cDNA and PAE/VEGFR2 cells. We thank Dr François Marceau and the Department of Obstetrics of l'Hôpital St-François d'Assise for providing umbilical cords. We also thank André Lévesque and Éric Pèlerin for their help with microscopy and Dr Simon Rousseau and Éric Shink for the VEGFR2-NIH 3T3 cells. This work was supported by a grant from The Canadian Institutes of Health Research.

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  1. Le Centre de recherche en cancérologie de l'Université Laval, L'Hôtel-Dieu de Québec, 9 rue McMahon, Québec, G1R 2J6, Canada

    Laurent Lamalice, François Houle, Guillaume Jourdan & Jacques Huot

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  1. Laurent Lamalice
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Correspondence to Jacques Huot.

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Lamalice, L., Houle, F., Jourdan, G. et al. Phosphorylation of tyrosine 1214 on VEGFR2 is required for VEGF-induced activation of Cdc42 upstream of SAPK2/p38. Oncogene 23, 434–445 (2004). https://doi.org/10.1038/sj.onc.1207034

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