Transcriptional regulation of cyclooxygenase-2 in the human microvascular endothelial cell line, HMEC-1: control by the combinatorial actions of AP2, NF-IL-6 and CRE elements.
Interleukin-1beta (IL-1) is a potent inducer of cyclooxygenase-2 (COX-2) and prostaglandin biosynthesis in many types of cells, yet little is known about the molecular mechanisms regulating IL-1 mediated prostanoid biosynthesis in the endothelium of the microvasculature. Therefore, we examined the cis- and trans-acting factors regulating IL-1-induced COX-2 expression in the human microvascular endothelial cell line, HMEC-1. IL-1 enhanced steady state levels of COX-2 protein and mRNA synthesis by approximately 2-fold which preceded a 2-fold increase in PGF(alpha) biosynthesis. Expression of a series of COX-2 promoter-luciferase constructs in IL-1 treated HMEC-1 cells revealed that the 'full length' (-1432/+59 bp) promoter was 10 times more active than the SV-40 promoter/enhancer and that it could be further activated by IL-1. Surprisingly however, all except for the shortest COX-2 promoter construct retained the ability to respond to IL-1 and luciferase activity driven by -191/+59 bp COX-2 promoter was as responsive to IL-1 as the full-length promoter. Moreover, site-directed promoter mutagenesis and electophoretic mobility shift assays (EMSA) indicate that the combinatorial actions of AP2, NF-IL6, and CRE elements are critical for both constitutive and IL-1-inducible COX-2 promoter activity. Understanding the mechanism(s) regulating COX-2 gene expression and prostaglandin biosynthesis in the microvasculature has important implications with regard to inflammation and angiogenesis in vivo.
Kirtikara
K
K
Department of Medicine, The University of Tennessee, Memphis, USA.
Raghow
R
R
Laulederkind
S J
SJ
Goorha
S
S
Kanekura
T
T
Ballou
L R
LR
D001483:Base Sequence; D051743:CCAAT-Enhancer-Binding Protein-delta; D022762:CCAAT-Enhancer-Binding Proteins; D002460:Cell Line; D000242:Cyclic AMP / Q000378:metabolism*; D051546:Cyclooxygenase 2; D017931:DNA Primers; D004268:DNA-Binding Proteins / Q000235:genetics*; D015237:Dinoprost / Q000096:biosynthesis; D004730:Endothelium, Vascular / Q000166:cytology / Q000201:enzymology* / Q000378:metabolism; D015971:Gene Expression Regulation, Enzymologic / Q000235:genetics*; D006801:Humans; D007527:Isoenzymes / Q000235:genetics*; D008565:Membrane Proteins; D009154:Mutation; D009687:Nuclear Proteins / Q000235:genetics*; D011401:Promoter Regions, Genetic; D011451:Prostaglandin-Endoperoxide Synthases / Q000235:genetics*; D050656:Transcription Factor AP-2; D014157:Transcription Factors / Q000235:genetics*; D014158:Transcription, Genetic / Q000235:genetics*
D022762:CCAAT-Enhancer-Binding Proteins; C497252:CEBPD protein, human; D017931:DNA Primers; D004268:DNA-Binding Proteins; D007527:Isoenzymes; D008565:Membrane Proteins; D009687:Nuclear Proteins; D050656:Transcription Factor AP-2; D014157:Transcription Factors; D051743:CCAAT-Enhancer-Binding Protein-delta; D015237:Dinoprost; D000242:Cyclic AMP; D051546:Cyclooxygenase 2; C496540:PTGS2 protein, human; D011451:Prostaglandin-Endoperoxide Synthases