Colocalization and interaction of cyclooxygenase-2 with caveolin-1 in human fibroblasts

J Biol Chem. 2001 Sep 14;276(37):34975-82. doi: 10.1074/jbc.M105946200. Epub 2001 Jun 29.

Abstract

Results from our previous study suggest that cyclooxygenase-2 (COX-2) induced by phorbol 12-myristate 13-acetate (PMA) may be localized to caveolae-like structures (Liou, J.-Y., Shyue, S.-K., Tsai, M.-J., Chung, C.-L., Chu, K.-Y., and Wu, K. K. (2000) J. Biol. Chem. 275, 15314-15320). In this study, we determined subcellular localization of COX-2 and caveolin-1 by confocal microscopy. COX-2 in human foreskin fibroblasts stimulated by PMA (100 nm) or interleukin-1beta (1 ng/ml) for 6 h was localized to plasma membrane in addition to endoplasmic reticulum and nuclear envelope. Caveolin-1 was localized to plasma membrane, and image overlay showed colocalization of COX-2 with caveolin-1. This was confirmed by the presence of COX-2 and caveolin-1 in the detergent-insoluble membrane fraction of cells stimulated by PMA. Immunoprecipitation showed complex formation of COX-2 with caveolin-1 in a time-dependent manner. A larger quantity of COX-2 was complexed with caveolin-1 in PMA-treated than in interleukin-1beta-treated cells. Purified COX-2 complexed with glutathione S-transferase-fused caveolin-1, which was not inhibited by the scaffolding domain peptide. Caveolin-1-bound COX-2 was catalytically active, and its activity was not inhibited by the scaffolding domain peptide. These results suggest that COX-2 induced by PMA and interleukin-1beta is colocalized with caveolin-1 in the segregated caveolae compartment. Because caveolae are rich in signaling molecules, this COX-2 compartment may play an important role in diverse pathophysiological processes.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Blotting, Western
  • Caveolae / chemistry
  • Caveolin 1
  • Caveolins / analysis*
  • Caveolins / chemistry
  • Caveolins / physiology
  • Cells, Cultured
  • Cyclooxygenase 2
  • Fibroblasts / chemistry
  • Humans
  • Interleukin-1 / pharmacology
  • Isoenzymes / analysis*
  • Isoenzymes / chemistry
  • Isoenzymes / physiology
  • Membrane Proteins
  • Microscopy, Confocal
  • Prostaglandin-Endoperoxide Synthases / analysis*
  • Prostaglandin-Endoperoxide Synthases / chemistry
  • Prostaglandin-Endoperoxide Synthases / physiology
  • Tetradecanoylphorbol Acetate / pharmacology

Substances

  • CAV1 protein, human
  • Caveolin 1
  • Caveolins
  • Interleukin-1
  • Isoenzymes
  • Membrane Proteins
  • Cyclooxygenase 2
  • PTGS2 protein, human
  • Prostaglandin-Endoperoxide Synthases
  • Tetradecanoylphorbol Acetate