Hadzic E, Desai-Yajnik V, Helmer E, Guo S, Wu S, Koudinova N, Casanova J, Raaka B M, Samuels H H
Department of Cell Biology, New York University Medical Center, New York 10016, USA.
Mol Cell Biol. 1995 Aug;15(8):4507-17. doi: 10.1128/MCB.15.8.4507.
The effects of the thyroid hormone (3,5,3'-triiodo-L-thyronine [T3]) on gene transcription are mediated by nuclear T3 receptors (T3Rs). alpha- and beta-isoform T3Rs (T3R alpha and -beta) are expressed from different genes and are members of a superfamily of ligand-dependent transcription factors that also includes the receptors for steroid hormones, vitamin D, and retinoids. Although T3 activates transcription by mediating a conformational change in the C-terminal approximately 220-amino-acid ligand-binding domain (LBD), the fundamental mechanisms of T3R-mediated transcriptional activation remain to be determined. We found that deletion of the 50-amino-acid N-terminal A/B domain of chicken T3R alpha (cT3R alpha) decreases T3-dependent stimulation of genes regulated by native thyroid hormone response elements about 10- to 20-fold. The requirement of the A/B region for transcriptional activation was mapped to amino acids 21 to 30, which contain a cluster of five basic amino acids. The A/B region of cT3R alpha is not required for T3 binding or for DNA binding of the receptor as a heterodimer with retinoid X receptor. In vitro binding studies indicate that the N-terminal region of cT3R alpha interacts efficiently with TFIIB and that this interaction requires amino acids 21 to 30 of the A/B region. In contrast, the LBD interacts poorly with TFIIB. The region of TFIIB primarily involved in the binding of cT3R alpha includes an amphipathic alpha helix contained within residues 178 to 201. Analysis using a fusion protein containing the DNA-binding domain of GAL4 and the entire A/B region of cT3R alpha suggests that this region does not contain an intrinsic activation domain. These and other studies indicate that cT3R alpha mediates at least some of its effects through TFIIB in vivo and that the N-terminal region of DNA-bound cT3R alpha acts to recruit and/or stabilize the binding of TFIIB to the transcription complex. T3 stimulation could then result from ligand-mediated changes in the LBD which may lead to the interaction of other factors with cT3R alpha, TFIIB, and/or other components involved in the initiation of transcription.
甲状腺激素（3,5,3'-三碘-L-甲状腺原氨酸 [T3]）对基因转录的影响是由核T3受体（T3Rs）介导的。α-和β-异构体T3Rs（T3Rα和-β）由不同基因表达，是配体依赖性转录因子超家族的成员，该超家族还包括类固醇激素、维生素D和类视黄醇的受体。尽管T3通过介导C末端约220个氨基酸的配体结合域（LBD）的构象变化来激活转录，但T3R介导的转录激活的基本机制仍有待确定。我们发现，缺失鸡T3Rα（cT3Rα）的50个氨基酸的N末端A/B结构域会使天然甲状腺激素反应元件调控的基因的T3依赖性刺激降低约10至20倍。转录激活对A/B区域的需求被定位到氨基酸21至30，其中包含一簇五个碱性氨基酸。cT3Rα的A/B区域对于T3结合或受体作为与视黄酸X受体的异二聚体的DNA结合不是必需的。体外结合研究表明，cT3Rα的N末端区域与TFIIB有效相互作用，并且这种相互作用需要A/B区域的氨基酸21至30。相反，LBD与TFIIB的相互作用较差。TFIIB中主要参与cT3Rα结合的区域包括178至201位残基内的一个两亲性α螺旋。使用包含GAL4的DNA结合结构域和cT3Rα的整个A/B区域的融合蛋白进行的分析表明，该区域不包含内在激活结构域。这些研究和其他研究表明，cT3Rα在体内至少通过TFIIB介导其一些作用，并且DNA结合的cT3Rα的N末端区域起到招募和/或稳定TFIIB与转录复合物结合的作用。然后，T3刺激可能源于LBD中配体介导的变化，这可能导致其他因子与cT3Rα、TFIIB和/或参与转录起始的其他成分相互作用。
