title: In situ analysis of interleukin-1-induced transcription of cox-2 and il-8 in cultured human myometrial cells.
abstract: The specific binding of transcription factors to DNA has been shown to be inhibited by chromatin structure and increased by cooperative interactions with other proteins. Consequently, in situ analysis using chromatin immunoprecipitation offers the most accurate view of transcriptional control. Transient transfection studies and in vitro analyses of IL-1-induced cox-2 transcription in a number of cell types have indicated regulation by either nuclear factor kappa B (NF-kappa B) or CCAAT/enhancer binding protein (C/EBP beta), or both acting cooperatively. To determine the mechanisms of COX-2 (cyclooxygenase or prostaglandin endoperoxide synthase) induction in cultured human myometrial cells in situ, we examined the cross-linking of the RelA subunit of NF-kappa B and C/EBP beta to the cox-2 promoter and flanking sequences. As a control, we inspected the interaction of these transcription factors with the IL-8 gene, which has been shown in other cell types to be activated by the cooperative interaction of NF-kappa B and C/EBP beta. Indeed, both transcription factors were cross-linked to the il-8 promoter after IL-1 treatment, but only RelA was cross-linked to cox-2 DNA. The il-8 promoter was also found to physically interact with proteins cross-linked to sites further upstream. IL-1 treatment also increased polymerase II cross-linking to both promoters and increased histone H4 acetylation at specific sites. These results indicate that modification of chromatin structure is part of the response to IL-1 stimulation. Chromatin immunoprecipitation thus provides critical insight into the mechanisms of COX-2 and IL-8 expression in human myometrial cells.
authors.affiliation: Department of Obstetrics and Gynecology, and Sealy Center for Molecular Science, University of Texas Medical Branch, Galveston, Texas 77555-1062, USA. msoloff@utmb.edu
authors.forename: Melvyn S
authors.initials: MS
authors.lastname: Soloff
authors.forename: Dennis L
authors.initials: DL
authors.lastname: Cook
authors.forename: Yow-Jiun
authors.initials: YJ
authors.lastname: Jeng
authors.forename: Garland D
authors.initials: GD
authors.lastname: Anderson
mesh_terms: D000107:Acetylation; D000818:Animals; D022782:CCAAT-Enhancer-Binding Protein-beta / Q000378:metabolism; D002478:Cells, Cultured; D002843:Chromatin / Q000378:metabolism; D003432:Cross-Linking Reagents / Q000378:metabolism; D051546:Cyclooxygenase 2; D005260:Female; D015870:Gene Expression / Q000187:drug effects; D006657:Histones / Q000378:metabolism; D020672:I-kappa B Proteins / Q000378:metabolism; D017403:In Situ Hybridization; D007375:Interleukin-1 / Q000494:pharmacology*; D016209:Interleukin-8 / Q000235:genetics*; D007527:Isoenzymes / Q000235:genetics*; D009215:Myometrium / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D000072000:NF-KappaB Inhibitor alpha; D016328:NF-kappa B / Q000235:genetics / Q000378:metabolism; D011233:Precipitin Tests / Q000379:methods; D011451:Prostaglandin-Endoperoxide Synthases / Q000235:genetics*; D012333:RNA, Messenger / Q000032:analysis; D051996:Transcription Factor RelA; D014158:Transcription, Genetic / Q000187:drug effects*
chemical_list: D022782:CCAAT-Enhancer-Binding Protein-beta; D002843:Chromatin; D003432:Cross-Linking Reagents; D006657:Histones; D020672:I-kappa B Proteins; D007375:Interleukin-1; D016209:Interleukin-8; D007527:Isoenzymes; D016328:NF-kappa B; C000606204:NFKBIA protein, human; D012333:RNA, Messenger; D051996:Transcription Factor RelA; D000072000:NF-KappaB Inhibitor alpha; D051546:Cyclooxygenase 2; D011451:Prostaglandin-Endoperoxide Synthases