Molecular Human Reproduction, Vol. 6, No. 9, 771-778,
September 2000
© 2000 European Society of Human Reproduction and Embryology
Endocrinology |
NF-IL6 and CRE elements principally account for both basal and interleukin-1ß-induced transcriptional activity of the proximal 528 bp of the PGHS-2 promoter in amnion-derived AV3 cells: evidence for involvement of C/EBPß
1 Department of Pharmacology and Clinical Pharmacology, University of Auckland, Faculty of Medical and Health Sciences, Auckland, New Zealand, and 2 Department of Pathology, Yale University, School of Medicine, New Haven, CT 06520–8023, USA
Abstract
Prostaglandin H synthase (PGHS)-2 promoter fragments (–528 to +9 bp and 5' unidirectional deletions thereof) were cloned upstream of the chloramphenicol acetyl-transferase (CAT) reporter gene. These were transfected into amnion-derived AV3 cells. The region, –528 to –203, which includes NF-
B sites, had little influence on CAT expression. The region, –203 and –52, however, was responsible for most of the basal promoter activity and also conferred responsiveness to interleukin (IL)-1ß (>3-times basal). Point mutations of NF-IL6 and cAMP response element (CRE) in this region reduced both basal and IL-1ß-stimulated production of CAT; dual mutation eliminated IL-1ß responsiveness. Factors in nuclear extracts from control or IL-1ß-stimulated AV3 cells specifically complexed the NF-IL6 and CRE sequences. However, the NF-IL6 and CRE oligonucleotides cross-competed, suggesting a common factor. C/EBPß was identified by supershift assay as interacting with both sequences. To a lesser extent C/EBP
and 
also interacted with the NF-IL6 site. However, CRE binding protein (CREB), was absent from the complex with the CRE. In conclusion, NF-IL6 and CRE elements principally account in AV3 amnion cells for basal and IL-1ß-inducible transcriptional activity of the proximal 528 bp of the PGHS-2 promoter, while NF-B elements play no substantial role. C/EBPs, particularly C/EBPß, are implicated in control of PGHS-2 transcription through the NF-IL6 and CRE sites.
amnion/cytokines/interleukin 1ß/prostaglandins/PGHS-2
Notes
3 To whom correspondence should be addressed at: Department of Pharmacology and Clinical Pharmacology, University of Auckland, Faculty of Medical and Health Sciences, 85 Park Road, Grafton, Auckland, New Zealand. E-mail: k.marvin{at}auckland.ac.nz
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