title: DNA binding and interaction with the nuclear receptor corepressor of thyroid hormone receptor are required for ligand-independent stimulation of the mouse preprothyrotropin-releasing hormone gene.
abstract: A negative thyroid hormone response element (TRE) in the mouse preprothyrotropin-releasing hormone (TRH) gene was previously mapped within the proximal promoter element between -83 and +53 that contained a TRE half-site motif at -57 (-57TGACCT-51). In transfection experiments, the promoter activity is stimulated by unliganded thyroid hormone receptor (TR) and T3 reverses the basal promoter stimulation. In this study, we determined whether the direct binding of TR to the TRE half-site in the mouse TRH gene is required for the ligand-independent stimulation using a transient transfection assay into CV-1 cells and electrophoretic mobility shift assays (EMSA). In addition, the role of a corepressor protein for the ligand-independent stimulation was examined using a putative splicing variant of the nuclear receptor corepressor (N-CoRI). Point mutations introduced into the TRE half-site at -57 eliminated the binding of TR and the stimulatory effect of unliganded TR. Two mutant TRs lacking DNA-binding activity and two CoR box mutant TRs showed no stimulation in the wild-type TRH promoter. The cotransfected N-CoRI potentiated the ligand-independent stimulation by the wild-type TR, but did not compensate for the impaired function of the CoR box mutant TR. In EMSA, TR strongly bound as homodimers and weakly as heterodimers with retinoid X receptor (RXR) to the element containing the TRE half-site at -57. Binding of TR to the TRE half-site was essential to form homo- and heterodimers, and the RXR binding site appeared to be located downstream of the TRE half-site. In vitro translated N-CoRI preferentially bound TR homodimers over TR/RXR heterodimers. These results collectively suggest that the DNA-bound TR/corepressor complex might be directly involved in the ligand-independent stimulation of the mouse TRH gene promoter.
authors.affiliation: First Department of Internal Medicine, Gunma University School of Medicine, Maebashi, Japan. tsato@sb.gunma-u.ac.jp
authors.forename: T
authors.initials: T
authors.lastname: Satoh
authors.forename: T
authors.initials: T
authors.lastname: Monden
authors.forename: T
authors.initials: T
authors.lastname: Ishizuka
authors.forename: T
authors.initials: T
authors.lastname: Mitsuhashi
authors.forename: M
authors.initials: M
authors.lastname: Yamada
authors.forename: M
authors.initials: M
authors.lastname: Mori
mesh_terms: D000818:Animals; D002465:Cell Movement; D004247:DNA / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000235:genetics; D019281:Dimerization; D004847:Epithelial Cells / Q000166:cytology; D005786:Gene Expression Regulation; D051379:Mice; D009154:Mutation; D009687:Nuclear Proteins / Q000235:genetics* / Q000378:metabolism; D056971:Nuclear Receptor Co-Repressor 1; D011401:Promoter Regions, Genetic; D011498:Protein Precursors / Q000235:genetics*; D011988:Receptors, Thyroid Hormone / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D012097:Repressor Proteins / Q000235:genetics* / Q000378:metabolism; D020218:Response Elements / Q000235:genetics; D013963:Thyroid Hormones / Q000378:metabolism; D013973:Thyrotropin-Releasing Hormone / Q000235:genetics*; D014162:Transfection
chemical_list: D004268:DNA-Binding Proteins; C539128:Ncor1 protein, mouse; D009687:Nuclear Proteins; D056971:Nuclear Receptor Co-Repressor 1; D011498:Protein Precursors; D011988:Receptors, Thyroid Hormone; D012097:Repressor Proteins; D013963:Thyroid Hormones; C063596:prepro-thyrotropin-releasing hormone; D013973:Thyrotropin-Releasing Hormone; D004247:DNA