Regulation of the human P-selectin promoter by Bcl-3 and specific homodimeric members of the NF-kappa B/Rel family.
P-selectin, an adhesion receptor for leukocytes, is constitutively expressed by megakaryocytes and endothelial cells. Synthesis of P-selectin is also increased by some inflammatory mediators. We characterized a previously identified kappa B site (-218GGGGGTGACCCC-207) in the promoter of the human P-selectin gene. The kappa B site was unique in that it bound constitutive nuclear protein complexes containing p50 or p52, but not inducible nuclear protein complexes containing p65. Furthermore, the element bound recombinant p50 or p52 homodimers, but not p65 homodimers. Methylation interference analysis indicated that p50 or p52 homodimers contacted the guanines at positions -218 to -214 on the coding strand and at -210 to -207 on the noncoding strand. Changes in the three central residues at -213 to -211 altered binding specificity for members of the NF-kappa B/Rel family. Mutations that eliminated binding to NF-kappa B/Rel proteins reduced by approximately 40% the expression of a reporter gene driven by the P-selectin promoter in transfected bovine aortic endothelial cells. Overexpression of p52 enhanced P-selectin promoter activity, and co-overexpression of Bcl-3 further induced promoter activity in a kappa B site-dependent manner. In contrast, overexpression of p50 repressed promoter activity; this repression was prevented by co-overexpression of Bcl-3. Similar phenomena were observed with reporter gene constructs driven by two tandem P-selectin kappa B sequences linked to the SV40 minimal promoter. These data suggest that Bcl-3 differentially regulates the effects of p50 and p52 homodimers bound to the kappa B site of the P-selectin promoter. This site may be a prototype for kappa B elements in other genes that bind specifically to p50 and/or p52 homodimers.
Pan
J
J
Department of Medicine, W.K. Warren Medical Research Institute, University of Oklahoma, Oklahoma City, USA.
McEver
R P
RP
D000818:Animals; D001011:Aorta; D000081242:B-Cell Lymphoma 3 Protein; D001483:Base Sequence; D001665:Binding Sites; D002417:Cattle; D002460:Cell Line; D002478:Cells, Cultured; D004730:Endothelium, Vascular / Q000378:metabolism*; D005786:Gene Expression Regulation; D018922:HL-60 Cells; D006801:Humans; D008156:Luciferases / Q000378:metabolism; D046911:Macromolecular Substances; D008745:Methylation; D008969:Molecular Sequence Data; D016328:NF-kappa B / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D020569:Proto-Oncogene Proteins c-rel; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism*; D014162:Transfection; D014471:Umbilical Veins
D000081242:B-Cell Lymphoma 3 Protein; C000631344:BCL3 protein, human; D046911:Macromolecular Substances; D016328:NF-kappa B; D011518:Proto-Oncogene Proteins; D020569:Proto-Oncogene Proteins c-rel; D011993:Recombinant Fusion Proteins; D014157:Transcription Factors; D008156:Luciferases