Transcriptional Regulation of Cyclooxygenase-2 in Mouse Skin Carcinoma Cells

Abstract

Many studies have suggested that overexpression of cyclooxygenase-2 (COX-2) contributes to the development of tumors in several tissues. COX-2 expression tends to be up-regulated in various types of tumors and transformed cell lines, and the overexpression of COX-2 is caused by enhanced transcription of the gene. In an attempt to characterize the signaling pathway leading to the overexpression of COX-2 in the mouse skin carcinoma cell line JWF2, we investigated cis-and transacting factors required for COX-2 expression and demonstrated a molecular mechanism by which COX-2 is expressed differentially in normal and neoplastic tissues. Two regions of the COX-2 promoter containing an E-box and nuclear factor IL6 site were identified as the positive regulatory elements through transient transfections with luciferase reporter vectors containing the various 5-flanking regions of the promoter. Moreover, electrophoretic mobility shift assays and cotransfection experiments showed that upstream stimulatory factors and CCAAT/enhancer-binding proteins (C/EBPs) bind to the E-box and nuclear factor IL6 site, respectively, and functionally transactivate the COX-2 promoter. We also found that C/EBP isoforms are expressed differentially during mouse skin carcinogenesis, suggesting that overexpression of COX-2 in tumors may be caused by a change in C/EBP expression levels. Prostaglandins (PGs)1 are involved in many normal and pathophysiological responses (1). As a rate-limiting enzyme in the synthesis of PGs, cyclooxygenase (COX)

Role of Mitogen-Activated Protein Kinase Cascades in Mediating Lipopolysaccharide-Stimulated Induction of Cyclooxygenase-2 and IL-1β in RAW264 Macrophages

“…The transcription factor C/EBP␤ is known to play a role in the activation of the COX-2 gene (14), and LPS was reported to increase the level of C/EBP␤ in RAW264.7 and J774 macrophages by stimulating transcription of the gene (15). We confirmed that LPS induces a striking rise in the level of C/EBP␤, and we also found that this increase was not prevented by SB 203580 and/or PD 98059 or by Ro 318220 (Fig.…”

The Transcription Factor C/EBPβ Is Essential for Inducible Expression of the cox-2 Gene in Macrophages but Not in Fibroblasts

“…Transcription factors belonging to the C/EBP family share a strong homology in their leucine zipper and DNA binding domains, and as a consequence are able to form both homo-and heterodimers and bind to the same DNA sequences (22). Different C/EBP family members can bind to the Ϫ138/Ϫ130 element, but their relative role in activating the cox-2 promoter in different cell types is unclear and sometimes contradictory (9,10,12,17,20,23). However, all these studies are based on transient transfection/overexpression experiments and do not take into account the endogenous and regulated ratio of C/EBP family members able to bind to the promoter under uninduced or induced conditions.…”

“…Interestingly, whereas in macrophages and fibroblasts the CRE moiety is responsible for transcriptional activation by members of the CRE-binding protein (CREB) and activating protein-1 (AP-1) family (12,15), in rat granulosa and skin carcinoma cells the E box is predominantly involved through the interaction with the upstream stimulating factors 1 and 2 (16,17). Finally, the nuclear factor for IL-6/CCAAT enhancer-binding protein (NF-IL6/C/EBP) element at position Ϫ138/Ϫ130 was reported to play an important role in mediating signal-dependent transcriptional induction in macrophages, osteoblastic cells, pancreatic islet cells, skin carcinoma cells, and chondrocytes (6,9,10,12,13,17,18,20) but not in rat granulosa cells or in fibroblasts (15,16). Transcription factors belonging to the C/EBP family share a strong homology in their leucine zipper and DNA binding domains, and as a consequence are able to form both homo-and heterodimers and bind to the same DNA sequences (22).…”

“…We first analyzed the Ϫ138/Ϫ130 C/EBP1 element, a canonical C/EBP-binding site that has been shown to bind members of the C/EBP family (6,9,13,17,18,20,23). EMSA analysis using nuclear extracts from untreated wild type W2 cells revealed at least four distinct complexes (Fig.…”

Conditional ablation of C/EBPβ demonstrates its keratinocyte-specific requirement for cell survival and mouse skin tumorigenesis

“…As a result of the high degree of conservation in their bZIP domains, C/EBP family members form both homodimers and heterodimers (Rask et al, 2000). C/EBPd, another member of the C/EBP family, is upregulated in mouse skin papillomas and squamous cell carcinomas (Kim and Fischer, 1998). On the other hand, C/EBPd might act as a tumor suppressor by virtue of its role in genome maintenance (Huang et al, 2004).…”