Identification of nuclear factors that enhance binding of the thyroid hormone receptor to a thyroid hormone response element.
Using a gel shift assay, we analyzed the binding of in vitro translated alpha- and beta-thyroid hormone (T3) receptors to a T3-response element (TRE) derived from the rat GH gene. No receptor-TRE complexes were observed when translated receptor alone was incubated with the TRE. However, addition of a nuclear extract from liver to the translational products resulted in the formation of two receptor-DNA complexes for both the alpha- and beta-receptors. These complexes were shown to contain translated receptor by comigration of 32P-labeled TRE and 35S-labeled receptor in the gel shift assay. A competition experiment demonstrated that formation of the complexes was sequence specific. Preincubation of the liver nuclear extract at 60 C abolished formation of both complexes indicating that receptor-TRE binding required a heat-labile nuclear factor. Phosphocellulose chromatography of the nuclear extract resulted in separation of the activities required for formation of the two complexes. Analysis of nuclear extracts from different tissues revealed that one complex formed in the presence of all extracts, whereas the second complex appeared predominantly with a nuclear extract from liver. Addition of T3 to the binding reaction had no effect on receptor-TRE complex formation. We suggest that nuclear factors interact with the T3 receptor to enhance hormone-independent binding to a TRE.
Murray
M B
MB
Department of Biochemistry, University of Minnesota Medical School, Minneapolis 55455.
Towle
H C
HC
D001667:Binding, Competitive; D002352:Carrier Proteins / Q000378:metabolism*; D002845:Chromatography; D008099:Liver / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism*; D009687:Nuclear Proteins / Q000502:physiology; D014176:Protein Biosynthesis; D011988:Receptors, Thyroid Hormone / Q000502:physiology*; D013963:Thyroid Hormones; D014158:Transcription, Genetic; D014284:Triiodothyronine / Q000378:metabolism*; D000097810:Thyroid Hormone-Binding Proteins
D002352:Carrier Proteins; D008565:Membrane Proteins; D009687:Nuclear Proteins; D011988:Receptors, Thyroid Hormone; D013963:Thyroid Hormones; D014284:Triiodothyronine; D000097810:Thyroid Hormone-Binding Proteins