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Article: Characterization of the human topoisomerase IIβ (TOP2B) promoter activity: Essential roles of the nuclear factor-Y (NF-Y)- and specificity protein-1 (Sp1)-binding sites

TitleCharacterization of the human topoisomerase IIβ (TOP2B) promoter activity: Essential roles of the nuclear factor-Y (NF-Y)- and specificity protein-1 (Sp1)-binding sites
Authors
Issue Date2002
PublisherPortland Press Ltd. The Journal's web site is located at http://www.biochemj.org
Citation
Biochemical Journal, 2002, v. 368 n. 3, p. 741-751 How to Cite?
AbstractEukaryotic topoisomerase II (topo II) catalyses topological genomic changes essential for chromosome segregation, chromatin reorganization, DNA replication and transcription. Mammalian topo II exists as two isoforms, designated α and β. Human topo IIα is an important cancer drug target, and an established determinant of drug sensitivity and resistance. Human topo IIβ is also the target of anticancer drugs but its role in drug resistance is less clear. The two human topo II proteins are encoded by the TOP2A and TOP2B genes, respectively, which despite their highly conserved structural organization, are subject to distinctly different modes of regulation. In the present study, we have cloned and characterized the human TOP2B promoter containing a 1.3 kb fragment of the 5′-flanking and untranslated region (-1067 to +193). We found that the promoter activity of this TOP2B fragment was constant throughout the cell cycle, in contrast to the activity of the proximal promoter of TOP2A which was low in resting cells and enhanced during proliferation. Analyses of 5′-serially and internally deleted luciferase reporter constructs revealed that 80% of the TOP2B promoter activity could be attributed to the region between -533 and -481. Mutational analyses of putative regulatory elements indicated that two inverted CCAAT boxes (ICBs) within this region were essential for TOP2B promoter activity and gel mobility-shift assays indicated these sites bound the transcription factor nuclear factor-Y (NF-Y). Co-transfection experiments using a dominant-negative form of subunit A of NF-Y suggested that TOP2B promoter activity required direct interaction of NF-Y with the ICBs. In addition, a specificity protein-1 (Sp1)-binding GC box located just upstream of the ICBs was shown to contribute to TOP2B promoter activity in a synergistic manner with the ICBs. Our results suggest that the binding sites for NF-Y and Sp1 are critical for TOP2B transcription.
Persistent Identifierhttp://hdl.handle.net/10722/167756
ISSN
2015 Impact Factor: 3.562
2015 SCImago Journal Rankings: 2.582
PubMed Central ID
References

 

DC FieldValueLanguage
dc.contributor.authorLok, CNen_US
dc.contributor.authorLang, AJen_US
dc.contributor.authorMirski, SELen_US
dc.contributor.authorCole, SPCen_US
dc.date.accessioned2012-10-08T03:11:05Z-
dc.date.available2012-10-08T03:11:05Z-
dc.date.issued2002en_US
dc.identifier.citationBiochemical Journal, 2002, v. 368 n. 3, p. 741-751en_US
dc.identifier.issn0264-6021en_US
dc.identifier.urihttp://hdl.handle.net/10722/167756-
dc.description.abstractEukaryotic topoisomerase II (topo II) catalyses topological genomic changes essential for chromosome segregation, chromatin reorganization, DNA replication and transcription. Mammalian topo II exists as two isoforms, designated α and β. Human topo IIα is an important cancer drug target, and an established determinant of drug sensitivity and resistance. Human topo IIβ is also the target of anticancer drugs but its role in drug resistance is less clear. The two human topo II proteins are encoded by the TOP2A and TOP2B genes, respectively, which despite their highly conserved structural organization, are subject to distinctly different modes of regulation. In the present study, we have cloned and characterized the human TOP2B promoter containing a 1.3 kb fragment of the 5′-flanking and untranslated region (-1067 to +193). We found that the promoter activity of this TOP2B fragment was constant throughout the cell cycle, in contrast to the activity of the proximal promoter of TOP2A which was low in resting cells and enhanced during proliferation. Analyses of 5′-serially and internally deleted luciferase reporter constructs revealed that 80% of the TOP2B promoter activity could be attributed to the region between -533 and -481. Mutational analyses of putative regulatory elements indicated that two inverted CCAAT boxes (ICBs) within this region were essential for TOP2B promoter activity and gel mobility-shift assays indicated these sites bound the transcription factor nuclear factor-Y (NF-Y). Co-transfection experiments using a dominant-negative form of subunit A of NF-Y suggested that TOP2B promoter activity required direct interaction of NF-Y with the ICBs. In addition, a specificity protein-1 (Sp1)-binding GC box located just upstream of the ICBs was shown to contribute to TOP2B promoter activity in a synergistic manner with the ICBs. Our results suggest that the binding sites for NF-Y and Sp1 are critical for TOP2B transcription.en_US
dc.languageengen_US
dc.publisherPortland Press Ltd. The Journal's web site is located at http://www.biochemj.orgen_US
dc.relation.ispartofBiochemical Journalen_US
dc.rightsCreative Commons: Attribution 3.0 Hong Kong License-
dc.subject.mesh3T3 Cellsen_US
dc.subject.meshAnimalsen_US
dc.subject.meshBase Sequenceen_US
dc.subject.meshBinding Sitesen_US
dc.subject.meshCcaat-Binding Factor - Metabolismen_US
dc.subject.meshCloning, Molecularen_US
dc.subject.meshDna Mutational Analysisen_US
dc.subject.meshDna Topoisomerases, Type Ii - Chemistry - Geneticsen_US
dc.subject.meshDna-Binding Proteinsen_US
dc.subject.meshFlow Cytometryen_US
dc.subject.meshGene Deletionen_US
dc.subject.meshGenes, Dominanten_US
dc.subject.meshHela Cellsen_US
dc.subject.meshHumansen_US
dc.subject.meshImmunoblottingen_US
dc.subject.meshLuciferases - Metabolismen_US
dc.subject.meshMiceen_US
dc.subject.meshModels, Geneticen_US
dc.subject.meshMolecular Sequence Dataen_US
dc.subject.meshPlasmids - Metabolismen_US
dc.subject.meshPromoter Regions, Geneticen_US
dc.subject.meshProtein Bindingen_US
dc.subject.meshSp1 Transcription Factor - Metabolismen_US
dc.subject.meshTime Factorsen_US
dc.subject.meshTranscription Factors - Metabolismen_US
dc.subject.meshTranscription, Geneticen_US
dc.subject.meshTransfectionen_US
dc.titleCharacterization of the human topoisomerase IIβ (TOP2B) promoter activity: Essential roles of the nuclear factor-Y (NF-Y)- and specificity protein-1 (Sp1)-binding sitesen_US
dc.typeArticleen_US
dc.identifier.emailLok, CN:cnlok@hku.hken_US
dc.identifier.authorityLok, CN=rp00752en_US
dc.description.naturepublished_or_final_versionen_US
dc.identifier.doi10.1042/BJ20020791en_US
dc.identifier.pmid12197834-
dc.identifier.pmcidPMC1223026-
dc.identifier.scopuseid_2-s2.0-0037115757en_US
dc.relation.referenceshttp://www.scopus.com/mlt/select.url?eid=2-s2.0-0037115757&selection=ref&src=s&origin=recordpageen_US
dc.identifier.volume368en_US
dc.identifier.issue3en_US
dc.identifier.spage741en_US
dc.identifier.epage751en_US
dc.publisher.placeUnited Kingdomen_US
dc.identifier.scopusauthoridLok, CN=7006410829en_US
dc.identifier.scopusauthoridLang, AJ=37044671100en_US
dc.identifier.scopusauthoridMirski, SEL=6701720297en_US
dc.identifier.scopusauthoridCole, SPC=7401503347en_US

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