Two Pathways for Cyclooxygenase-2 Protein Degradation in Vivo

COX-2, formally known as prostaglandin endoperoxide H synthase-2 (PGHS-2), catalyzes the committed step in prostaglandin biosynthesis. COX-2 is induced during inflammation and is overexpressed in colon cancer. In vitro, an 18-amino acid segment, residues 595–612, immediately upstream of the C-terminal endoplasmic reticulum targeting sequence is required for N-glycosylation of Asn594, which permits COX-2 protein to enter the endoplasmic reticulum-associated protein degradation system. To determine the importance of this COX-2 degradation pathway in vivo, we engineered a del595–612 PGHS-2 (Δ18 COX-2) knock-in mouse lacking this 18-amino acid segment. Δ18 COX-2 knock-in mice do not exhibit the renal or reproductive abnormalities of COX-2 null mice. Δ18 COX-2 mice do have elevated urinary prostaglandin E2 metabolite levels and display a more pronounced and prolonged bacterial endotoxin-induced febrile response than wild type (WT) mice. Normal brain tissue, cultured resident peritoneal macrophages, and cultured skin fibroblasts from Δ18 COX-2 mice overexpress Δ18 COX-2 relative to WT COX-2 expression in control mice. These results indicate that COX-2 can be degraded via the endoplasmic reticulum-associated protein degradation pathway in vivo. Treatment of cultured cells from WT or Δ18 COX-2 mice with flurbiprofen, which blocks substrate-dependent degradation, attenuates COX-2 degradation, and treatment of normal mice with ibuprofen increases the levels of COX-2 in brain tissue. Thus, substrate turnover-dependent COX-2 degradation appears to contribute to COX-2 degradation in vivo. Curiously, WT and Δ18 COX-2 protein levels are similar in kidneys and spleens from WT and Δ18 COX-2 mice. There must be compensatory mechanisms to maintain constant COX-2 levels in these tissues.