title: Hormone-dependent interaction between the amino- and carboxyl-terminal domains of progesterone receptor in vitro and in vivo.
abstract: Full transcriptional activation by steroid hormone receptors requires functional synergy between two transcriptional activation domains (AF) located in the amino (AF-1) and carboxyl (AF-2) terminal regions. One possible mechanism for achieving this functional synergy is a physical intramolecular association between amino (N-) and carboxyl (C-) domains of the receptor. Human progesterone receptor (PR) is expressed in two forms that have distinct functional activities: full-length PR-B and the amino-terminally truncated PR-A. PR-B is generally a stronger activator than PR-A, whereas under certain conditions PR-A can act as a repressor in trans of other steroid receptors. We have analyzed whether separately expressed N- (PR-A and PR-B) and C-domains [hinge plus ligand-binding domain (hLBD)] of PR can functionally interact within cells by mammalian two-hybrid assay and whether this involves direct protein contact as determined in vitro with purified expressed domains of PR. A hormone agonist-dependent interaction between N-domains and the hLBD was observed functionally by mammalian two-hybrid assay and by direct protein-protein interaction assay in vitro. With both experimental approaches, N-C domain interactions were not induced by the progestin antagonist RU486. However, in the presence of the progestin agonist R5020, the N-domain of PR-B interacted more efficiently with the hLBD than the N-domain of PR-A. Coexpression of steroid receptor coactivator-1 (SRC-1) and the CREB binding protein (CBP), enhanced functional interaction between N- and C-domains by mammalian two-hybrid assay. However, addition of SRC-1 and CBP in vitro had no influence on direct interaction between purified N- and C-domains. These results suggest that the interaction between N- and C-domains of PR is direct and requires a hormone agonist-induced conformational change in the LBD that is not allowed by antagonists. Additionally, coactivators are not required for physical association between the N- and C-domains but are capable of enhancing a functionally productive interaction. In addition, the more efficient interaction of the hLBD with the N-domain of PR-B, compared with that of PR-A, suggests that distinct interactions between N- and C-terminal regions contribute to functional differences between PR-A and PR-B.
authors.affiliation: Department of Pathology and Molecular Biology Program, University of Colorado Health Sciences Center, Denver 80262, USA.
authors.forename: M J
authors.initials: MJ
authors.lastname: Tetel
authors.forename: P H
authors.initials: PH
authors.lastname: Giangrande
authors.forename: S A
authors.initials: SA
authors.lastname: Leonhardt
authors.forename: D P
authors.initials: DP
authors.lastname: McDonnell
authors.forename: D P
authors.initials: DP
authors.lastname: Edwards
mesh_terms: D000818:Animals; D001665:Binding Sites; D050882:CREB-Binding Protein; D006367:HeLa Cells / Q000187:drug effects / Q000378:metabolism; D051548:Histone Acetyltransferases; D006727:Hormone Antagonists / Q000378:metabolism / Q000494:pharmacology; D006801:Humans; D007313:Insecta / Q000821:virology; D008322:Mammals; D015735:Mifepristone / Q000378:metabolism / Q000494:pharmacology; D009687:Nuclear Proteins / Q000378:metabolism*; D056920:Nuclear Receptor Coactivator 1; D010446:Peptide Fragments / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D011374:Progesterone / Q000378:metabolism / Q000494:pharmacology; D045167:Progesterone Congeners / Q000378:metabolism / Q000494:pharmacology; D011397:Promegestone / Q000378:metabolism / Q000494:pharmacology; D011980:Receptors, Progesterone / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D015534:Trans-Activators / Q000378:metabolism*; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*
chemical_list: D006727:Hormone Antagonists; D009687:Nuclear Proteins; D010446:Peptide Fragments; D045167:Progesterone Congeners; D011980:Receptors, Progesterone; D011994:Recombinant Proteins; D015534:Trans-Activators; D014157:Transcription Factors; D015735:Mifepristone; D011374:Progesterone; D011397:Promegestone; D050882:CREB-Binding Protein; C494363:CREBBP protein, human; D051548:Histone Acetyltransferases; C538859:NCOA1 protein, human; D056920:Nuclear Receptor Coactivator 1