title: Retinoid-dependent in vitro transcription mediated by the RXR/RAR heterodimer.
abstract: The effects of retinoids on gene regulation are mediated by retinoic acid receptors (RARs) and retinoid X receptors (RXRs). Here, we provide the first biochemical evidence that, in vitro, ligand governs the transcriptional activity of RXR alpha/RAR alpha by inducing conformational changes in the ligand-binding domains. Using limited proteolytic digestion we show that binding of the cognate ligand causes a conformational change in the carboxy-terminal part of the receptor. We also show that recombinant RXR alpha/RAR alpha is partially active in the absence of exogenously added ligand. Trans-activation depends critically on the ligand-dependent transcriptional activation function AF-2 of RAR alpha. Full activation by recombinant RXR alpha/RAR alpha, however, requires the addition of either all-trans RA, 9-cis RA, or other RAR-specific agonists, whereas an RAR alpha-specific antagonist abolishes trans-activation. Intriguingly, the ligand-dependent AF-2 of RXR does not contribute to the level of transcription from the RAR beta 2 promoter in vitro even when the cognate ligand (9-cis RA) is bound. Thus, the major role of RXR in trans-activation of the RAR beta 2 promoter is to serve as an auxiliary factor required for the binding of RAR which, in turn, is directly responsible for transcriptional activity.
authors.affiliation: European Molecular Biology Laboratory (EMBL), Gene Expression Program, Heidelberg, Germany.
authors.forename: R
authors.initials: R
authors.lastname: Valcárcel
authors.forename: H
authors.initials: H
authors.lastname: Holz
authors.forename: C G
authors.initials: CG
authors.lastname: Jiménez
authors.forename: D
authors.initials: D
authors.lastname: Barettino
authors.forename: H G
authors.initials: HG
authors.lastname: Stunnenberg
mesh_terms: D001483:Base Sequence; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D002846:Chromatography, Affinity; D005786:Gene Expression Regulation / Q000187:drug effects*; D005822:Genetic Vectors; D006367:HeLa Cells; D006801:Humans; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000096:biosynthesis; D009838:Oligodeoxyribonucleotides; D011401:Promoter Regions, Genetic / Q000187:drug effects; D011487:Protein Conformation / Q000187:drug effects; D018168:Receptors, Retinoic Acid / Q000096:biosynthesis* / Q000737:chemistry / Q000302:isolation & purification; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000302:isolation & purification; D047488:Retinoid X Receptors; D012176:Retinoids / Q000494:pharmacology*; D014157:Transcription Factors / Q000096:biosynthesis* / Q000737:chemistry / Q000302:isolation & purification; D014158:Transcription, Genetic / Q000187:drug effects*; D015533:Transcriptional Activation; D014162:Transfection; D014212:Tretinoin / Q000494:pharmacology; D014616:Vaccinia virus
chemical_list: D009687:Nuclear Proteins; D009838:Oligodeoxyribonucleotides; D018168:Receptors, Retinoic Acid; D011993:Recombinant Fusion Proteins; D047488:Retinoid X Receptors; D012176:Retinoids; D014157:Transcription Factors; D014212:Tretinoin