title: Cooperative DNA binding by the B-isoform of human progesterone receptor: thermodynamic analysis reveals strongly favorable and unfavorable contributions to assembly.
abstract: Progesterone receptors (PR) play critical roles in eukaryotic gene regulation, yet the mechanisms by which they assemble at multisite promoters are poorly understood. Here we present a thermodynamic analysis of the interactions of the PR B-isoform (PR-B) with promoters containing either one or two progesterone response elements (PREs). Utilizing quantitative footprinting, we have resolved the microscopic energetics of PR-B binding, including cooperativity terms. The results of this analysis challenge a number of assumptions found in traditional models of receptor function. First, PR-B interactions at a single PRE can be equally well described by mechanisms invoking either the receptor monomer or the dimer as the active DNA binding species. If, as is commonly accepted, PR-B interacts with response elements only as a preformed dimer, then its intrinsic binding affinity is not the typically observed nanomolar but is rather picomolar. This high affinity binding is opposed, however, by a large energetic penalty. The penalty presumably pays for costly structural rearrangements of the receptor dimer and/or response element that are needed to form the protein-DNA complex. If PR-B assembles at a single response element via successive monomer binding reactions, then this penalty minimizes cooperative interactions between adjacent monomers. When binding to two response elements, the receptor exhibits strong intersite cooperativity. Although this phenomenon has been observed before, the present work demonstrates that the energetics reach levels seen in highly cooperative systems such as lambda cI repressor. This first quantitative dissection of cooperative receptor-promoter interactions suggests that PR-B function is more complex than traditionally envisioned.
authors.affiliation: Department of Pharmaceutical Sciences, University of Colorado Health Sciences Center, 4200 East 9th Avenue, Denver, Colorado 80262, USA.
authors.forename: Aaron F
authors.initials: AF
authors.lastname: Heneghan
authors.forename: Keith D
authors.initials: KD
authors.lastname: Connaghan-Jones
authors.forename: Michael T
authors.initials: MT
authors.lastname: Miura
authors.forename: David L
authors.initials: DL
authors.lastname: Bain
mesh_terms: D000495:Allosteric Site; D001483:Base Sequence; D001665:Binding Sites; D004247:DNA / Q000737:chemistry / Q000378:metabolism*; D018983:DNA Footprinting; D019281:Dimerization; D005786:Gene Expression Regulation; D006801:Humans; D007700:Kinetics; D008957:Models, Genetic; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D011980:Receptors, Progesterone / Q000737:chemistry / Q000378:metabolism*; D013329:Structure-Activity Relationship; D013816:Thermodynamics
chemical_list: D011980:Receptors, Progesterone; D004247:DNA