on May 20, 2002
Published online before print May 20, 2002, doi: 10.1161/01.CIR.0000018127.10968.34
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Submitted on January 16, 2002
From the Vascular Biology Research Center, Institute of Molecular Medicine and Division of Hematology (Y.Z., M.S., W.-G.D., X.-M.X., K.K.W.), University of Texas Health Science Center, Houston, Tex; and Institut für Pharmakologie und Klinische Pharmakologie (K.S., J.M.-K.), Heinrich-Heine-Universtät Düsseldorf, Düsseldorf, Germany. * To whom correspondence should be addressed. E-mail: Kenneth.K.Wu{at}uth.tmc.edu.
Background—Cyclooxygenase-2 (COX-2) plays a key role in human inflammatory disorders such as vascular inflammation. COX-2 promoter activity is induced by proinflammatory mediators, but the role of cyclic adenosine monophosphate response element (CRE) in promoter stimulation remains unclear. Methods and Results—Transient transfection of a 0.9-kb COX-2 promoter fragment bearing CRE mutation abrogated COX-2 promoter activity induced by proinflammatory mediators in human endothelial cells and fibroblasts. Dual mutations of CRE and an upstream CCAAT/enhancer binding protein (C/EBP) site did not have an additional effect. Binding of CREB-2, ATF-2, USF-2, and c-Jun transactivators to a wild-type and CRE-mutated oligonucleotide was analyzed by a novel DNA-binding assay. CREB-2 and ATF-2 in nuclear extracts of unstimulated endothelial cells bound to CRE, whereas USF-2 and c-Jun or c-Fos bound to non-CRE sites. CREB-2 and c-Fos binding was increased by phorbol 12-myristate 13-acetate but not tumor necrosis factor- Conclusions—CRE plays an obligatory role in COX-2 promoter activation by diverse stimuli. CREB-2 and ATF-2 bound to CRE serve as an anchor for P300 interaction with upstream transactivators and downstream transcription machinery.
Revised on March 28, 2002
Accepted on March 28, 2002
Obligatory Role of Cyclic Adenosine Monophosphate Response Element in Cyclooxygenase-2 Promoter Induction and Feedback Regulation by Inflammatory Mediators
Karsten Schroer PhD,
. The binding assay and chromatin immunoprecipitation revealed binding of P300 coactivator to the COX-2 promoter region.
Key words: prostaglandins • interleukins • endothelium
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