title: Tumor necrosis factor-alpha- or lipopolysaccharide-induced expression of the murine P-selectin gene in endothelial cells involves novel kappaB sites and a variant activating transcription factor/cAMP response element.
abstract: Tumor necrosis factor-alpha (TNF-alpha) or lipopolysaccharide (LPS) increases expression of the P-selectin gene in murine, but not in human, endothelial cells. These mediators augment expression of a reporter gene driven by the murine, but not the human, P-selectin promoter in transfected endothelial cells. The regions from -593 to -474 and from -229 to -13 in the murine P-selectin promoter are required for TNF-alpha or LPS to stimulate reporter gene expression. Within these regions, we identified two tandem kappaB elements, a reverse-oriented kappaB site and a variant activating transcription factor/cAMP response element (ATF/CRE), that participate in TNF-alpha- or LPS-induced expression. The tandem kappaB elements bound to NF-kappaB heterodimers and p65 homodimers, the reverse-oriented kappaB site bound to p65 homodimers, and the variant ATF/CRE bound to nuclear proteins that included activating transcription factor-2. Mutations in each individual element eliminated binding to nuclear proteins and decreased by 20-60% the TNF-alpha- or LPS-induced expression of a reporter gene driven by the murine P-selectin promoter in transfected endothelial cells. Simultaneous mutations of all elements further decreased, but did not abolish, induced expression. Co-overexpression of p50 and p65 enhanced murine P-selectin promoter activity in a kappaB site-dependent manner. These data indicate that the kappaB sites and the variant ATF/CRE are required for TNF-alpha or LPS to optimally induce expression of the murine P-selectin gene. The presence of these elements in the murine, but not the human, P-selectin gene may explain in part why TNF-alpha or LPS stimulates transcription of P-selectin in a species-specific manner.
authors.affiliation: Department of Medicine, Warren Medical Research Institute, University of Oklahoma Health Sciences Center, Oklahoma City, Oklahoma 73104, USA.
authors.forename: J
authors.initials: J
authors.lastname: Pan
authors.forename: L
authors.initials: L
authors.lastname: Xia
authors.forename: L
authors.initials: L
authors.lastname: Yao
authors.forename: R P
authors.initials: RP
authors.lastname: McEver
mesh_terms: D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D016466:CHO Cells; D002417:Cattle; D002478:Cells, Cultured; D016384:Consensus Sequence; D006224:Cricetinae; D017362:Cyclic AMP Response Element-Binding Protein / Q000378:metabolism*; D004730:Endothelium, Vascular / Q000187:drug effects / Q000378:metabolism*; D005786:Gene Expression Regulation / Q000187:drug effects*; D017930:Genes, Reporter; D014644:Genetic Variation; D006801:Humans; D008070:Lipopolysaccharides / Q000494:pharmacology*; D008156:Luciferases / Q000096:biosynthesis; D051379:Mice; D008969:Molecular Sequence Data; D016328:NF-kappa B / Q000378:metabolism*; D009687:Nuclear Proteins / Q000378:metabolism; D019007:P-Selectin / Q000096:biosynthesis* / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D012045:Regulatory Sequences, Nucleic Acid; D016415:Sequence Alignment; D012689:Sequence Homology, Nucleic Acid; D013045:Species Specificity; D014157:Transcription Factors / Q000378:metabolism*; D014162:Transfection; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*
chemical_list: D017362:Cyclic AMP Response Element-Binding Protein; D008070:Lipopolysaccharides; D016328:NF-kappa B; D009687:Nuclear Proteins; D019007:P-Selectin; D011993:Recombinant Fusion Proteins; D014157:Transcription Factors; D014409:Tumor Necrosis Factor-alpha; D008156:Luciferases