The ubiquitin- and proteasome-dependent degradation of COX-2 is regulated by the COP9 signalosome and differentially influenced by coxibs

Abstract

The cyclooxygenase-2 (COX-2) enzyme is induced upon inflammation and in neoplastic tissues. It produces prostaglandins that stimulate tumor angiogenesis and tumor growth. Therefore, destruction and/or specific inhibition of COX-2 should be an important aspect of future tumor therapy. Recently, clinical application of specific COX-2 inhibitors called coxibs became doubtfully because they produce serious renal and cardiovascular complications under long term application. The exact underlying mechanisms are poorly understood and the different effects of diverse coxibs are not explained. It has been demonstrated before that COX-2 is degraded by the ubiquitin (Ub) proteasome system (UPS). However, how ubiquitination is accomplished and regulated was unclear. An important regulator of the UPS is the COP9 signalosome (CSN), which controls the stability of many proteins. Here we show that the proteasome-dependent degradation of COX-2 in HeLa cell lysate and in HeLa cells was stimulated by curcumin, an inhibitor of CSN-associated kinases. These data suggest a function of the CSN in the degradation of COX-2. In addition, proteolysis of COX-2 was significantly accelerated by parecoxib, but not by celecoxib or rofecoxib. By density gradient centrifugation and immunoprecipitation we demonstrate that COX-2 physically interacts with the CSN. Moreover, COX-2 is associated with large complexes consisting of the CSN, cullin-RING Ub ligases and the 26S proteasome. Pulldown experiments with Flag-COX-2 revealed cullin 1 and cullin 4 as components of the large super-complexes. Cullin 1 and 4 are scaffolding proteins of Ub ligases that presumably ubiquitinate COX-2. Treatment of HeLa cells with parecoxib results in an accelerated degradation of endogenous COX-2 accompanied by an increase of COX-2-Ub conjugates. In HeLa cells parecoxib is converted to the selective COX-2 inhibitor valdecoxib. Addition of valdecoxib also stimulates COX-2 degradation in HeLa cells. We therefore conclude that valdecoxib specifically interacts with COX-2 and induces a conformation accessible for ubiquitination and degradation.

Prostaglandin EP1 Receptor Down-regulates Expression of Cyclooxygenase-2 by Facilitating Its Proteasomal Degradation

“…In accordance with our own results, inhibition of the proteasome in neuronal cells causes the accumulation of high molecular weight ubiquitin-COX-2 (22). Furthermore, COX-2 ubiquitination in HeLa cells was found to depend on its association with large complexes COP9 signalsomes consisting of cullin proteins that scaffold unidentified E3 ligases (8). Given our evidence for the existence of an EP 1 -COX-2 complex as well as the fact that they share similar cellular locations (2, 3), it is tempting to speculate that the EP 1 receptor or associated accessory proteins may scaffold a currently unrecognized E3 ligase or other proteins that modulate COX-2 degradation.…”

“…Application of a mannosidase inhibitor or deletion of a 19 amino acid sequence at the C-tail, which contains an N-glycosylation site unique to COX-2, stabilizes the enzyme and prolongs its half-life (6,7). From the endoplasmic reticulum, the enzyme is transported to the cytosol, where it is ubiquitinated (8) and degraded by the proteasome (7) in a partially understood pathway.…”

USP22 acts as an oncogene by regulating the stability of cyclooxygenase-2 in non-small cell lung cancer

“…COX-2 is a membrane-bound protein that resides in the endoplasmic reticulum (ER) and its degradation by the UPS is preceded by its translocation from the ER to the cytoplasm via the ER-associated degradation system [25]. The stability of COX-2 was shown to be regulated by the COP9 signalosome, a multi-subunit complex that regulates the stability of proteins by affecting their ubiquitination [26]. The modulation of COX-2 levels by ubiquitination and the importance of COX-2/PGE2 signaling in lung cancer have been demonstrated; therefore, our findings indicating the modulation of COX-2 levels by USP22 are relevant, although additional experiments are necessary to further examine the relationship between USP22 upregulation and COX-2 in NSCLC.…”

A genome-wide association study identifies a gene network of ADAMTS genes in the predisposition to pediatric stroke

“…The TRIM29 block is a cassette of genes involved in regulation of transcription. COPS8 (as part of the protein complex COP9) is a regulator of cyclooxogenase-2 25 and might therefore influence coagulation . It has been argued that variants identified in GWAS explain only a small proportion of the genetic variance underlying most complex traits.…”