Integrin-mediated Adhesion and Soluble Ligand Binding Stabilize COX-2 Protein Levels in Endothelial Cells by Inducing Expression and Preventing Degradation

Jelena Zaric,Curzio Rüegg

doi:10.1074/jbc.m410006200

Abstract

Cyclooxygenase-2 (COX-2), a key enzyme in prostaglandin synthesis, is highly expressed during inflammation and cellular transformation and promotes tumor progression and angiogenesis. We have previously demonstrated that endothelial cell COX-2 is required for integrin alphaVbeta3-dependent activation of Rac-1 and Cdc-42 and for endothelial cell spreading, migration, and angiogenesis (Dormond, O., Foletti, A., Paroz, C., and Ruegg, C. (2001) Nat. Med. 7, 1041-1047; Dormond, O., Bezzi, M., Mariotti, A., and Ruegg, C. (2002) J. Biol. Chem. 277, 45838-45846). In this study, we addressed the question of whether integrin-mediated cell adhesion may regulate COX-2 expression in endothelial cells. We report that cell detachment from the substrate caused rapid degradation of COX-2 protein in human umbilical vein endothelial cells (HUVEC) independent of serum stimulation. This effect was prevented by broad inhibition of cellular proteinases and by neutralizing lysosomal activity but not by inhibiting the proteasome. HUVEC adhesion to laminin, collagen I, fibronectin, or vitronectin induced rapid COX-2 protein expression with peak levels reached within 2 h and increased COX-2-dependent prostaglandin E2 production. In contrast, nonspecific adhesion to poly-L-lysine was ineffective in inducing COX-2 expression. Furthermore, the addition of matrix proteins in solution promoted COX-2 protein expression in suspended or poly-L-lysine-attached HUVEC. Adhesion-induced COX-2 expression was strongly suppressed by pharmacological inhibition of c-Src, phosphatidylinositol 3-kinase, p38, extracellular-regulated kinase 1/2, and, to a lesser extent, protein kinase C and by the inhibition of mRNA or protein synthesis. In conclusion, this work demonstrates that integrin-mediated cell adhesion and soluble integrin ligands contribute to maintaining COX-2 steady-state levels in endothelial cells by the combined prevention of lysosomal-dependent degradation and the stimulation of mRNA synthesis involving multiple signaling pathways.

Highlights

Tumor angiogenesis, i.e. the formation of new blood vessels in response to angiogenic stimuli, promotes tumor progression by stimulating tumor cell survival, tumor invasion, and metastasis formation [1]
We report that COX-2 protein is rapidly degraded through a lysosomal-dependent pathway upon human umbilical vein endothelial cells (HUVEC) detachment from the substrate and that integrin-mediated adhesion to extracellular matrix (ECM) proteins induced de novo COX-2 synthesis involving signaling through multiple pathways (i.e. p38, mitogen-activated protein kinase, phosphatidylinositol 3-kinase (PI3-K), protein kinase C (PKC), and c-Src)
Endothelial Cell Detachment Causes COX-2 Degradation—We have previously shown that endothelial cell COX-2 activity is required for ␣V␤3 integrin-mediated endothelial cell spreading and migration in vitro and fibroblast growth factor 2-induced angiogenesis in vivo [26, 27]

Summary

Introduction

I.e. the formation of new blood vessels in response to angiogenic stimuli, promotes tumor progression by stimulating tumor cell survival, tumor invasion, and metastasis formation [1]. HUVEC adhesion to laminin, collagen I, fibronectin, or vitronectin induced rapid COX-2 protein expression with peak levels reached within 2 h and increased COX-2-dependent prostaglandin E2 production.

Results

Conclusion