Characterization of the ligand-dependent transactivation domain of thyroid hormone receptor.
Transcriptional activation by nuclear receptors is achieved through autonomous activation functions (AFs), a constitutive N-terminal AF-1 and a C-terminal, ligand-dependent AF-2 that comprises a motif conserved between nuclear receptors. We have performed an extensive mutational analysis of the putative AF-2 domain of chicken thyroid hormone receptor alpha (cT3R alpha). We show that the AF-2 region mediates transactivation as well as transcriptional interference (squelching), not only between the thyroid hormone and vitamin (type II) receptors, but also between type II and steroid hormone (type I) receptors. Transcriptional activation and interference require equivalent doses of the cognate ligand, and mutations in the conserved motif that reduce ligand-induced transactivation also impair transcriptional interference. When fused to the Gal4 DNA binding domain, a 35 amino acid long fragment containing the conserved motif is able to transactivate and squelch, albeit in a ligand-independent manner. Our results define the AF-2 of cT3R alpha as an autonomous transactivation domain that, in its natural context, is governed by ligand. We propose that AF-2 is probably part of a surface for interaction with either a general transcription factor or a putative bridging factor, that might be utilized by type I and II receptors.
Gene Expression Programme, EMBL, Heidelberg, Germany.
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Barettino
M M
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Vivanco Ruiz
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Stunnenberg
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002460:Cell Line; D002645:Chickens; D017124:Conserved Sequence; D008024:Ligands; D008969:Molecular Sequence Data; D009154:Mutation; D011401:Promoter Regions, Genetic; D011988:Receptors, Thyroid Hormone / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism*; D014158:Transcription, Genetic; D014284:Triiodothyronine / Q000378:metabolism*
D008024:Ligands; D011988:Receptors, Thyroid Hormone; D011993:Recombinant Fusion Proteins; D015534:Trans-Activators; D014284:Triiodothyronine