title: The effect of free DNA on the interactions of the estrogen receptor bound to hormone, partial antagonist or pure antagonist with target DNA.
abstract: Interactions between the lamb uterine estrogen receptor occupied by estradiol, 4-hydroxytamoxifen (a non-steroidal partial estrogen antagonist) or ICI 164,384 (a steroidal pure estrogen antagonist), and the vitellogenin A2 estrogen-response element (vit ERE) were compared using a biotinylated 25-base all-palindromic double-stranded oligonucleotide, containing vit ERE (b-ERE), which allowed isolation of the b-ERE.receptor.[3H]ligand assembly on streptavidin-Sepharose. The results of saturation analyses of the three receptor.[3H]ligand complexes by increasing amounts of b-ERE were quite similar for the proportion of complexes able to interact with b-ERE (which varied from 30% to 65% according to experiments) and for the equilibrium dissociation constant [Kd (0 degree C) approximately 1.2 nM, assuming that the receptor interacted as a dimer with b-ERE]. With each ligand, receptor binding to ERE did not change the rate of ligand dissociation from the receptor at 20 degrees C. The rate of estrogen receptor dissociation from b-ERE, measured at 20 degrees C in the presence of a given concentration of ERE, did not vary according to the ligand bound to the receptor; however, this dissociation rate increased linearly over the ERE concentration range (0.5-10 microM). The experimental rate constant (k-) of estrogen receptor dissociation from b-ERE appeared to be the sum of the basal dissociation-rate constant (k degrees - approximately 0.011 min-1), corresponding to spontaneous dissociation which would occur in the absence of ERE, and of the ERE-induced dissociation-rate constant, proportional to the used concentration of ERE (ki- approximately 4500 CERE M-1 min-1, where CERE is the molar concentration of ERE). Non-target DNA also induced receptor dissociation from b-ERE, but its efficiency was 6-10-fold lower than that of ERE. We conclude that, the two antiestrogens are as efficient as estradiol in promoting estrogen receptor binding to a single vit ERE; the low or nil ability of antiestrogens to induce estrogenic responses is probably not linked with the receptor DNA-binding step; DNA binding does not seem to affect the conformation of the filled hormone-binding site of the receptor at 20 degrees C; interactions of receptor dimers with DNA seems to proceed by direct transfer of receptor dimers between DNA strands.
authors.affiliation: Institut National de la Santé et de la Recherche Médicale, Montpellier, France.
authors.forename: S
authors.initials: S
authors.lastname: Aliau
authors.forename: T
authors.initials: T
authors.lastname: Groblewski
authors.forename: J L
authors.initials: JL
authors.lastname: Borgna
mesh_terms: D000818:Animals; D001426:Bacterial Proteins / Q000378:metabolism; D001483:Base Sequence; D004247:DNA / Q000378:metabolism / Q000494:pharmacology*; D004958:Estradiol / Q000031:analogs & derivatives* / Q000378:metabolism; D004965:Estrogen Antagonists / Q000378:metabolism*; D004967:Estrogens / Q000378:metabolism*; D005260:Female; D008969:Molecular Sequence Data; D053284:Polyunsaturated Alkamides; D011960:Receptors, Estrogen / Q000037:antagonists & inhibitors / Q000187:drug effects* / Q000378:metabolism; D012756:Sheep; D019809:Streptavidin; D013629:Tamoxifen / Q000031:analogs & derivatives* / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D004965:Estrogen Antagonists; D004967:Estrogens; D053284:Polyunsaturated Alkamides; D011960:Receptors, Estrogen; D013629:Tamoxifen; C016601:afimoxifene; D004958:Estradiol; C051781:ICI 164384; D004247:DNA; D019809:Streptavidin