Modulation of DNA topoisomerase II alpha promoter activity by members of the Sp (specificity protein) and NF-Y (nuclear factor Y) families of transcription factors.
Topo IIalpha (topoisomerase IIalpha) is a major target of several commonly used anticancer drugs and is subject to down-regulation at the transcriptional level in some drug-resistant cell lines and tumours in response to chemotherapy. Clinical resistance to such drugs has been correlated with down-regulation of topo IIalpha at transcription in some drug-resistant cell lines and tumours. Putative binding sites for a variety of transcription factors, including Sp1 (specificity protein 1) and NF-Y (nuclear factor Y) have previously been identified in the topo IIalpha promoter, but their functional significance and interactions have not been described following exposure to anti-cancer drugs. The binding of these factors to specific putative regulatory elements in the topo IIalpha promoter was studied using electrophoretic-mobility-shift assays. Sp1 was found to bind strongly to both distal and proximal GC-rich elements and NF-Y to ICB1 (the first inverted CCAAT box). The functional significance of transcription-factor binding was studied using transient transfection of HeLa cells using a luciferase reporter driven by a 617-bp minimal promoter containing point mutations in putative regulatory elements. Sp1 and NF-Y were both found to be transcriptional modulators with activator or repressor functions depending on protein/DNA context. Moreover, a functional interaction between Sp1 and NF-Y bound at proximal elements was observed.
Magan
Natisha
N
Institute of Molecular BioSciences, Massey University, Private Bag 11-222, Palmerston North, New Zealand.
Szremska
Agnieszka P
AP
Isaacs
Richard J
RJ
Stowell
Kathryn M
KM
D000951:Antigens, Neoplasm; D023081:CCAAT-Binding Factor / Q000235:genetics / Q000502:physiology*; D004250:DNA Topoisomerases, Type II / Q000235:genetics* / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D024202:Electrophoretic Mobility Shift Assay; D015971:Gene Expression Regulation, Enzymologic; D017930:Genes, Reporter / Q000235:genetics; D006367:HeLa Cells; D006801:Humans; D008156:Luciferases / Q000235:genetics; D016297:Mutagenesis, Site-Directed; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins / Q000235:genetics; D016329:Sp1 Transcription Factor / Q000235:genetics / Q000502:physiology*; D051704:Sp2 Transcription Factor; D014157:Transcription Factors / Q000235:genetics / Q000502:physiology*; D014162:Transfection; D014407:Tumor Cells, Cultured
D000951:Antigens, Neoplasm; D023081:CCAAT-Binding Factor; D004268:DNA-Binding Proteins; D011993:Recombinant Fusion Proteins; D016329:Sp1 Transcription Factor; D014157:Transcription Factors; C402244:nuclear factor Y; D051704:Sp2 Transcription Factor; D008156:Luciferases; D004250:DNA Topoisomerases, Type II