title: The bZIP dimer localizes at DNA full-sites where each basic region can alternately translocate and bind to subsites at the half-site.
abstract: Crystal structures of the GCN4 bZIP (basic region/leucine zipper) with the AP-1 or CRE site show how each GCN4 basic region binds to a 4 bp cognate half-site as a single DNA target; however, this may not always fully describe how bZIP proteins interact with their target sites. Previously, we showed that the GCN4 basic region interacts with all 5 bp in half-site TTGCG (termed 5H-LR) and that 5H-LR comprises two 4 bp subsites, TTGC and TGCG, which individually are also target sites of the basic region. In this work, we explore how the basic region interacts with 5H-LR when the bZIP dimer localizes to full-sites. Using AMBER molecular modeling, we simulated GCN4 bZIP complexes with full-sites containing 5H-LR to investigate in silico the interface between the basic region and 5H-LR. We also performed in vitro investigation of bZIP-DNA interactions at a number of full-sites that contain 5H-LR versus either subsite: we analyzed results from DNase I footprinting and electrophoretic mobility shift assay (EMSA) and from EMSA titrations to quantify binding affinities. Our computational and experimental results together support a highly dynamic DNA-binding model: when a bZIP dimer localizes to its target full-site, the basic region can alternately recognize either subsite as a distinct target at 5H-LR and translocate between the subsites, potentially by sliding and hopping. This model provides added insights into how α-helical DNA-binding domains of transcription factors can localize to their gene regulatory sequences in vivo.
authors.affiliation: Department of Chemistry, University of Toronto, 3359 Mississauga Road, Mississauga, Ontario L5L 1C6, Canada.
authors.forename: I-San
authors.initials: IS
authors.lastname: Chan
authors.forename: Taufik
authors.initials: T
authors.lastname: Al-Sarraj
authors.forename: S Hesam
authors.initials: SH
authors.lastname: Shahravan
authors.forename: Anna V
authors.initials: AV
authors.lastname: Fedorova
authors.forename: Jumi A
authors.initials: JA
authors.lastname: Shin
mesh_terms: D001483:Base Sequence; D050976:Basic-Leucine Zipper Transcription Factors / Q000737:chemistry / Q000378:metabolism*; D001665:Binding Sites / Q000235:genetics; D022762:CCAAT-Enhancer-Binding Proteins / Q000378:metabolism; D018983:DNA Footprinting; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism; D006860:Hydrogen Bonding; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism
chemical_list: D050976:Basic-Leucine Zipper Transcription Factors; D022762:CCAAT-Enhancer-Binding Proteins; D004268:DNA-Binding Proteins; C501391:GCN4 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins