3,5,3'-triiodothyronine receptor auxiliary protein (TRAP) enhances receptor binding by interactions within the thyroid hormone response element.
We have previously demonstrated that binding of in vitro synthesized thyroid hormone receptor (TR) to thyroid hormone response elements (TREs) is enhanced by the addition of nuclear extracts from several different cell types, suggesting that binding of TR is partially dependent on a T3 receptor auxiliary protein (TRAP). We have used the avidin-biotin complex DNA-binding assay to discriminate between regions of TREs that bind TR alone and sites that are influenced by interactions with TRAP. Mutations in the TREs from rat GH and glycoprotein hormone alpha-subunit genes show that a specific DNA sequence is required for TRAP-mediated enhancement of TR binding. Mutations in the B half-site of the rat GH TRE or in similar sequences [(T/A)GGGA] in the alpha-subunit TRE ablate the enhancement of TR binding by TRAP. Furthermore, binding of TR to a natural half-site in the TSH beta-subunit gene (bases -16 to 6), which lacks an additional AGGGA-like sequence, is not enhanced by the addition of TRAP. Binding of TR to TREs was also tested at physiological salt concentrations in the avidin-biotin complex DNA-binding assay. Binding of human TR beta to TREs decreases dramatically at 140 mM KCl compared to binding at 50 mM KCl; however, the addition of TRAP enhances the binding to almost 4-fold of basal binding, suggesting that TRAP may be important for stabilization of TR binding to TREs in the cell.
Beebe
J S
JS
Department of Medicine, Brigham and Women's Hospital, Boston, Massachusetts 02115.
Darling
D S
DS
Chin
W W
WW
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D001710:Biotin; D002460:Cell Line; D004247:DNA / Q000378:metabolism*; D015292:Glycoprotein Hormones, alpha Subunit / Q000235:genetics; D013006:Growth Hormone / Q000235:genetics; D008969:Molecular Sequence Data; D016296:Mutagenesis; D009687:Nuclear Proteins / Q000494:pharmacology*; D009994:Osmolar Concentration; D011189:Potassium Chloride / Q000494:pharmacology; D011518:Proto-Oncogene Proteins / Q000235:genetics; D051381:Rats; D011988:Receptors, Thyroid Hormone / Q000378:metabolism*; D013972:Thyrotropin / Q000235:genetics; D014284:Triiodothyronine / Q000494:pharmacology
D015292:Glycoprotein Hormones, alpha Subunit; D009687:Nuclear Proteins; D011518:Proto-Oncogene Proteins; D011988:Receptors, Thyroid Hormone; D014284:Triiodothyronine; D011189:Potassium Chloride; D001710:Biotin; D013972:Thyrotropin; D013006:Growth Hormone; D004247:DNA