title: Interaction of CBF alpha/AML/PEBP2 alpha transcription factors with nucleosomes containing promoter sequences requires flexibility in the translational positioning of the histone octamer and exposure of the CBF alpha site.
abstract: Chromatin remodeling at eukaryotic gene promoter sequences accompanies transcriptional activation. Both molecular events rely on specific protein-DNA interactions that occur within these promoter sequences. Binding of CBFalpha/AML/PEBP2alpha (core binding factor alpha/acute myelogenous leukemia/polyoma enhancer binding protein 2alpha) proteins is a key event in both tissue-specific and developmentally regulated osteocalcin (OC) promoter activity. To address linkage between chromatin organization and transcription factor binding, we reconstituted segments of the rat OC gene proximal promoter into mononucleosomes and studied binding of CBFalpha proteins. We analyzed binding of bacterially produced Cbfalpha2Alpha and Cbfalpha2B, two splice variants of the human CBFalpha2 gene, and determined the effect of heterodimerization with the Cbfbeta subunit on binding activity. Our results indicate that binding of the truncated Cbfalpha2A protein to naked DNA is independent of Cbfbeta whereas Cbfalpha2A binding to nucleosomal DNA was enhanced by Cbfbeta. In contrast, the Cbfalpha2B interaction with either naked or nucleosomal DNA was strongly dependent on heterodimerization with the Cbfbeta subunit. Additionally, our results demonstrate that both Cbfalpha2A alone and Cbfalpha2B complexed with Cbfbeta can interact with nucleosomal DNA only if there is a degree of flexibility in the positioning of the histone octamer on the DNA fragment and exposure of the CBFalpha site. This situation was achieved with a DNA segment of 182 bp from the rat OC promoter that preferentially positions mononucleosomes upstream of the CBFalpha binding site and leaves this element partially exposed. Taken together, these results suggest that nucleosomal translational positioning is a major determinant of the binding of CBFalpha factors to nucleosomal DNA.
authors.affiliation: Departamento de Biología Molecular, Facultad de Ciencias Biológicas, Universidad de Concepción, Casilla 160-C, Concepción, Chile.
authors.forename: J
authors.initials: J
authors.lastname: Gutiérrez
authors.forename: J
authors.initials: J
authors.lastname: Sierra
authors.forename: R
authors.initials: R
authors.lastname: Medina
authors.forename: M
authors.initials: M
authors.lastname: Puchi
authors.forename: M
authors.initials: M
authors.lastname: Imschenetzky
authors.forename: A
authors.initials: A
authors.lastname: van Wijnen
authors.forename: J
authors.initials: J
authors.lastname: Lian
authors.forename: G
authors.initials: G
authors.lastname: Stein
authors.forename: J
authors.initials: J
authors.lastname: Stein
authors.forename: M
authors.initials: M
authors.lastname: Montecino
mesh_terms: D000818:Animals; D001482:Base Composition; D001665:Binding Sites / Q000235:genetics; D002645:Chickens; D050659:Core Binding Factor Alpha 1 Subunit; D050657:Core Binding Factor alpha Subunits; D050660:Core Binding Factors; D004247:DNA / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D006657:Histones / Q000235:genetics / Q000378:metabolism*; D009363:Neoplasm Proteins; D009707:Nucleosomes / Q000235:genetics / Q000378:metabolism*; D015675:Osteocalcin / Q000235:genetics / Q000378:metabolism; D011401:Promoter Regions, Genetic; D014176:Protein Biosynthesis; D051381:Rats; D012689:Sequence Homology, Nucleic Acid; D050656:Transcription Factor AP-2; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*
chemical_list: D050659:Core Binding Factor Alpha 1 Subunit; D050657:Core Binding Factor alpha Subunits; D050660:Core Binding Factors; D004268:DNA-Binding Proteins; D006657:Histones; D009363:Neoplasm Proteins; D009707:Nucleosomes; C493717:Runx2 protein, rat; D050656:Transcription Factor AP-2; D014157:Transcription Factors; D015675:Osteocalcin; D004247:DNA