title: Transcription of brain creatine kinase in U87-MG glioblastoma is modulated by factor AP2.
abstract: Our previous studies established in U87-MG glioblastoma cells that elevated cAMP increased transcription of the endogenous as well as a transiently-transfected brain creatine kinase (CKB) gene, despite the absence of a cAMP response element (CRE) in the CKB proximal promoter. This report employed transfection to show that the transcription of CKB in U87 cells is induced by transcription factor AP2alpha, which is known to be activated by cAMP. Dominant-negative forms of AP2alpha not only prevented the AP2alpha-mediated activation of CKB but also blocked the cAMP-mediated increase in CKB transcription caused by forskolin treatment. The mutation of the four potential AP2 elements within the CKB proximal promoter showed that induction of CKB by AP2 was mediated principally through the AP2 element located at -50 bp in the promoter. Electromobility shift assays revealed a protein in U87 nuclear extracts that bound to a consensus AP2alpha element as well as to the (-50) AP2 element in CKB. Interestingly, the CKB (-50) AP2 element contains GCCAATGGG which also bound NF-Y, the CCAAT-binding protein, suggesting that interplay between AP2 and NF-Y may modulate CKB transcription. This is the first report of a role for AP2 in the regulation of CKB transcription and of an AP2 element within which an NF-Y site is located.
authors.affiliation: Department of Biological Sciences, University of Delaware, 117 Wolf Hall, Newark, DE 19716, USA.
authors.forename: Dianna
authors.initials: D
authors.lastname: Willis
authors.forename: Yanping
authors.initials: Y
authors.lastname: Zhang
authors.forename: George R
authors.initials: GR
authors.lastname: Molloy
mesh_terms: D001483:Base Sequence; D001921:Brain / Q000378:metabolism*; D023081:CCAAT-Binding Factor / Q000235:genetics / Q000378:metabolism; D015500:Chloramphenicol O-Acetyltransferase / Q000378:metabolism; D005576:Colforsin; D003402:Creatine Kinase / Q000235:genetics / Q000378:metabolism*; D052276:Creatine Kinase, BB Form; D000242:Cyclic AMP / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D024202:Electrophoretic Mobility Shift Assay; D006801:Humans; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009841:Oligonucleotides; D010957:Plasmids / Q000235:genetics; D014158:Transcription, Genetic / Q000235:genetics*; D014407:Tumor Cells, Cultured; D001616:beta-Galactosidase / Q000378:metabolism
chemical_list: D023081:CCAAT-Binding Factor; D004268:DNA-Binding Proteins; D007527:Isoenzymes; D009841:Oligonucleotides; C083115:activator protein-2 binding element; D005576:Colforsin; D000242:Cyclic AMP; D015500:Chloramphenicol O-Acetyltransferase; D003402:Creatine Kinase; D052276:Creatine Kinase, BB Form; D001616:beta-Galactosidase