title: Binding of disparate transcriptional activators to nucleosomal DNA is inherently cooperative.
abstract: To investigate mechanisms by which multiple transcription factors access complex promoters and enhancers within cellular chromatin, we have analyzed the binding of disparate factors to nucleosome cores. We used a purified in vitro system to analyze binding of four activator proteins, two GAL4 derivatives, USF, and NF-kappa B (KBF1), to reconstituted nucleosome cores containing different combinations of binding sites. Here we show that binding of any two or all three of these factors to nucleosomal DNA is inherently cooperative. Thus, the binuclear Zn clusters of GAL4, the helix-loop-helix/basic domains of USF, and the rel domain of NF-kappa B all participated in cooperative nucleosome binding, illustrating that this effect is not restricted to a particular DNA-binding domain. Simultaneous binding by two factors increased the affinity of individual factors for nucleosomal DNA by up to 2 orders of magnitude. Importantly, cooperative binding resulted in efficient nucleosome binding by factors (USF and NF-kappa B) which independently possess little nucleosome-binding ability. The participation of GAL4 derivatives in cooperative nucleosome binding required only DNA-binding and dimerization domains, indicating that disruption of histone-DNA contacts by factor binding was responsible for the increased affinity of additional factors. Cooperative nucleosome binding required sequence-specific binding of all transcription factors, appeared to have spatial constraints, and was independent of the orientation of the binding sites on the nucleosome. These results indicate that cooperative nucleosome binding is a general mechanism that may play a significant role in loading complex enhancer and promoter elements with multiple diverse factors in chromatin and contribute to the generation of threshold responses and transcriptional synergy by multiple activator sites in vivo.
authors.affiliation: Department of Biochemistry and Molecular Biology, Pennsylvania State University, University Park 16802-4500.
authors.forename: C C
authors.initials: CC
authors.lastname: Adams
authors.forename: J L
authors.initials: JL
authors.lastname: Workman
mesh_terms: D001483:Base Sequence; D001665:Binding Sites; D002843:Chromatin / Q000378:metabolism*; D003001:Cloning, Molecular; D004268:DNA-Binding Proteins; D004742:Enhancer Elements, Genetic; D004926:Escherichia coli; D005656:Fungal Proteins / Q000378:metabolism; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D016328:NF-kappa B / Q000378:metabolism; D009707:Nucleosomes / Q000378:metabolism*; D009838:Oligodeoxyribonucleotides; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000378:metabolism; D015183:Restriction Mapping; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors / Q000096:biosynthesis / Q000378:metabolism*; D051720:Upstream Stimulatory Factors
chemical_list: D002843:Chromatin; D004268:DNA-Binding Proteins; D005656:Fungal Proteins; C052130:GAL4 protein, S cerevisiae; D016328:NF-kappa B; D009707:Nucleosomes; D009838:Oligodeoxyribonucleotides; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; D051720:Upstream Stimulatory Factors