Regulation of the human topoisomerase IIalpha gene promoter in confluence-arrested cells.
Expression of DNA topoisomerase IIalpha mRNA and protein reflects the proliferative state of mammalian cell lines and tissues with high levels in actively cycling cells but marked down-regulation during serum deprivation or cell density-induced growth arrest. Using stably integrated gene fusions comprising the human topoisomerase IIalpha promoter with a growth hormone reporter gene, we have localized elements required for the differential activity of the topoisomerase IIalpha promoter in proliferating and confluence-arrested cells. Deletion analysis localized the region of the promoter that responded to changes in the cellular growth state to between -101 and -144 base pairs. Mutation analysis identified an inverted CCAAT box (ICB) located at -108 to -104 as necessary for promoter down-regulation in confluence-arrested cells, while several other potential cis-acting elements, including four additional ICBs, were shown not to be required. The critical ICB was recognized in vitro by the CCAAT box binding factor, NF-Y, with levels of binding activity higher in extracts from proliferating cells than from confluence-arrested cells. We conclude that the differential regulation of topoisomerase IIalpha gene expression in cycling and confluence-arrested cells is mediated, at least in part, through proliferation-specific binding of factors to an ICB element in the gene promoter.
Isaacs
R J
RJ
Imperial Cancer Research Fund Laboratories, Institute of Molecular Medicine, University of Oxford, John Radcliffe Hospital, Oxford OX3 9DU, United Kingdom.
Harris
A L
AL
Hickson
I D
ID
D016475:3T3 Cells; D000818:Animals; D000951:Antigens, Neoplasm; D001483:Base Sequence; D022762:CCAAT-Enhancer-Binding Proteins; D002455:Cell Division / Q000235:genetics; D017931:DNA Primers; D004250:DNA Topoisomerases, Type II / Q000235:genetics* / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism; D015536:Down-Regulation; D015971:Gene Expression Regulation, Enzymologic; D017930:Genes, Reporter; D013006:Growth Hormone / Q000235:genetics; D006801:Humans; D007527:Isoenzymes / Q000235:genetics* / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D012333:RNA, Messenger / Q000235:genetics; D014157:Transcription Factors / Q000378:metabolism; D014407:Tumor Cells, Cultured
D000951:Antigens, Neoplasm; D022762:CCAAT-Enhancer-Binding Proteins; D017931:DNA Primers; D004268:DNA-Binding Proteins; D007527:Isoenzymes; D012333:RNA, Messenger; D014157:Transcription Factors; D013006:Growth Hormone; D004250:DNA Topoisomerases, Type II