Dominant negative activity of mutant thyroid hormone alpha 1 receptors from patients with hepatocellular carcinoma
Dominant negative activity of mutant thyroid hormone alpha 1 receptors from patients with hepatocellular carcinoma
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DOI：
10.1210/en.138.12.5308
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发表时间：
1997-12-01
期刊：
影响因子：
4.8
通讯作者：
Cheng, SY
中科院分区：
文献类型：
--
作者：
Lin, KH;Zhu, XG;Cheng, SY
从两名患者的肝细胞癌中分离出两种突变型甲状腺激素α 1受体（TR α 1）的互补DNA。互补DNA的序列分析表明，突变体H和L分别具有一个瓦尔（390）Ala和两个Pro（398）Ser/Glu（350）Lys突变。我们表征了它们的结合酶、DNA结合酶和显性负活性。突变体H和L不结合激素T-3。使用三种类型的甲状腺激素反应元件（TREs），其中半位点结合基序排列在一个外翻重复序列（Lys），一个反向重复序列（DR 4），或由四个核苷酸（DR 4）分开的直接重复序列中，分析了它们的DNA结合活性。与野生型TR α 1（w-TR α 1）相比，其以不同的同二聚体/单体比率结合这些TREs，突变体L与作为同二聚体的三种TREs的结合减少了类似于90%。然而，突变体H与这些TREs的结合更为复杂。虽然它通常以同源二聚体的形式与DR 4结合，但其以同源二聚体的形式与Lys的结合减少了约80%。令人惊讶的是，与w-TR α 1相比，其与α 1的结合显著增强。这两种突变体与作为具有类维生素A X受体（RXR α和-β）的异二聚体的三种TREs的结合没有受到显著影响。与缺乏T-3结合活性一致，两种突变体都失去了其反式激活能力。突变体H和L表现出显性负活性，但不同的TRE依赖性。突变体H的显性负效应大小顺序为DR 4> DR 4> Lys，而突变体L则不存在TRE依赖性。本研究表明，TR α基因突变确实发生在患者中，这些新的TR α 1突变体提供了一个有价值的工具，以进一步了解突变TR的显性负作用的分子基础。
Complementary DNAs for two mutant thyroid hormone (yl receptors (TR alpha 1) were isolated from hepatocellular carcinomas of two patients. Sequence analyses of the complementary DNAs showed a single Val(390)Ala and double Pro(398)Ser/Glu(350)Lys mutations in mutants H and L, respectively. We characterized their hormone-binding, DNA-binding, and dominant negative activities. Mutants H and L did not bind the hormone T-3,. Their DNA-binding activities were analyzed using three types of thyroid hormone response elements (TREs) in which the half-site binding motifs are arranged in an everted repeat (Lys), an inverted repeat (Pal), or a direct repeat separated by four nucleotides (DR4). Compared with wild-type TR alpha 1 (w-TR alpha 1), which bound these TREs with different homodimer/monomer ratios, binding of mutant L to the three TREs as homodimers was reduced by similar to 90%. However, binding of mutant H to these TREs was more complex. Although it bound normally to DR4 as homodimers, its binding to Lys as homodimers was reduced by similar to 80%. Surprisingly, its binding to Pal was markedly enhanced compared with w-TR alpha 1. The binding of these two mutants to the three TREs as heterodimers with retinoid X receptors (RXR alpha and -beta) was not significantly affected. Consistent with the lack of T-3-binding activity, both mutants had lost their trans-activation capacity. Mutants H and L exhibited dominant negative activity, but differed in their TRE dependency. The dominant negative potency of mutant H was in the rank order of Pal > DR4 > Lys, whereas no TRE dependency was observed for mutant L. The present study indicates that mutations of the TR alpha gene do occur in patients and that these novel TR alpha 1 mutants provide a valuable tool to further understand the molecular basis of the dominant negative action of mutant TRs.