title: Glucocorticoid receptor ligand binding domain is sufficient for the modulation of glucocorticoid induction properties by homologous receptors, coactivator transcription intermediary factor 2, and Ubc9.
abstract: Several factors modulate the position of the dose-response curve of steroid receptor-agonist complexes and the partial agonist activity of antagonist complexes, thereby causing differential gene activation by circulating hormones and unequal gene repression during endocrine therapies with antisteroids. We now ask whether the modulatory activity of three factors (homologous receptor, coactivator transcription intermediary factor 2, and Ubc9) requires the same or different domains of glucocorticoid receptors (GRs). In all cases, we find that neither the amino terminal half of the receptor, which contains the activation function-1 activation domain, nor the DNA binding domain is required. This contrasts with the major role of activation function-1 in determining the amount of gene expression and partial agonist activity of antisteroids with most steroid receptors. However, the situation is more complicated with Ubc9, where GR N-terminal sequences prevent the actions of Ubc9, but not added GR or transcription intermediary factor 2, at low GR concentrations. Inhibition is relieved by deletion of these sequences or by replacement with the comparable region of progesterone receptors but not by overexpression of the repressive sequences. These results plus the binding of C-terminal GR sequences to the suppressive N-terminal domain implicate an intramolecular mechanism for the inhibition of Ubc9 actions at low GR concentrations. A shift from noncooperative to cooperative steroid binding at high GR concentrations suggests that conformational changes reposition the inhibitory N-terminal sequence to allow Ubc9 interaction with elements of the ligand binding domain. Collectively, these results indicate a dominant role of GR C-terminal sequences in the modulation of the dose-response curve and partial agonist activity of GR complexes. They also reveal mechanistic differences both among individual modulators and between the ability of the same factors to regulate the total amount of gene expression.
authors.affiliation: Steriod Hormones Section, National Institute of Diabetes and Digestive and Kidney Diseases/LMCB, National Institutes of Health, Bethesda, Maryland 20892, USA.
authors.forename: Sehyung
authors.initials: S
authors.lastname: Cho
authors.forename: Benjamin L
authors.initials: BL
authors.lastname: Kagan
authors.forename: John A
authors.initials: JA
authors.lastname: Blackford
authors.forename: Daniele
authors.initials: D
authors.lastname: Szapary
authors.forename: S Stoney
authors.initials: SS
authors.lastname: Simons
mesh_terms: D000818:Animals; D015153:Blotting, Western; D019556:COS Cells; D002460:Cell Line; D002474:Cell-Free System; D004247:DNA / Q000378:metabolism; D003907:Dexamethasone / Q000494:pharmacology; D019281:Dimerization; D004305:Dose-Response Relationship, Drug; D005786:Gene Expression Regulation; D005799:Genes, Dominant; D005938:Glucocorticoids / Q000378:metabolism*; D005978:Glutathione / Q000737:chemistry; D005982:Glutathione Transferase / Q000378:metabolism; D007700:Kinetics; D008024:Ligands; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D051958:Nuclear Receptor Coactivator 2; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D014176:Protein Biosynthesis; D017434:Protein Structure, Tertiary; D051381:Rats; D011965:Receptors, Glucocorticoid / Q000737:chemistry* / Q000378:metabolism; D012685:Sepharose / Q000737:chemistry; D013256:Steroids / Q000737:chemistry / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism*; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D014162:Transfection; D044763:Ubiquitin-Conjugating Enzymes / Q000378:metabolism*; D000098843:Ubiquitin-Conjugating Enzyme UBC9
chemical_list: D005938:Glucocorticoids; D008024:Ligands; D051958:Nuclear Receptor Coactivator 2; D011965:Receptors, Glucocorticoid; D013256:Steroids; D014157:Transcription Factors; D003907:Dexamethasone; D004247:DNA; D012685:Sepharose; D044763:Ubiquitin-Conjugating Enzymes; D005982:Glutathione Transferase; D000098843:Ubiquitin-Conjugating Enzyme UBC9; D005978:Glutathione