Abstract

Cyclooxygenase-2 (COX-2) is frequently expressed in cancer cells, contributing to tumor development. Most studies of COX-2 expression have examined artificially induced expression in noncancer cells rather than basal expression in cancer cells. Therefore, basal COX-2 expression and its regulation were examined in cell lines derived from a murine model of lung adenocarcinoma. The presence of COX-2 protein in these cells was demonstrated by Western analysis. COX-2 promoter activity was repressed by U0126 [1,4-diamino-2,3-dicyano-1,4-bis(2-aminophenylthio)butadiene], a mitogen-activated protein kinase kinase inhibitor, as well as SB202190 [4-(4-fluorophenyl)-2-(4-hydroxyphenyl)-5-(4-pyridyl)-1H-imidazole], an inhibitor of p38 mitogen-activated protein kinase, substantiating the involvement of these signal transduction pathways in the regulation of basal COX-2 expression. Retinoic acid also repressed promoter activity, yet increased activity significantly in one cell line after 18 and 30 h of treatment. Deletions of the murine COX-2 promoter revealed that the 5′ transcription factor binding sites were not required for basal expression, including the only nuclear factor-κB sites of the promoter. Site-directed mutagenesis of the 3′ C/EBP (CCAAT/enhancer-binding protein) sites inhibited promoter activity by 20 to 55%, while mutation of the 3′ ATF/CREB/AP-1 (activating transcription factor/cAMP response element-binding protein/activator protein-1) site inhibited activity by 70%. Mutation of the 3′ upstream stimulatory factor site did not affect promoter activity. Electrophoretic mobility shift assays indicated that the AP-1 transcription factor does not bind to the 3′ ATF/CREB/AP-1 site, leaving C/EBP and ATF/CREB as the major transcriptional regulators of basal expression of COX-2 in these lung tumor-derived cell lines and identifying new targets for the prevention/treatment of lung cancer through the modulation of COX-2 expression.