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Negative regulation of thyroid stimulating hormone α and β subunit genes by thyroid hormone and its receptor

甲状腺激素及其受体对促甲状腺激素α和β亚基基因的负调控

基本信息

批准号：
15590973
负责人：
SASAKI Shigekazu
金额：
$ 0.96万
依托单位：
Hamamatsu University School of Medicine
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2003
资助国家：
日本
起止时间：
2003 至 2004
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-15590973/
关键词：
TSH thyroid hormone thyroid hormone receptor GATA2 Pit1 pituitary nuclear hormone receptor transcriptional regulation

项目摘要

The negative regulation of thyrotropin (thyroid stimulating hormone, TSH,) production by the thyroid hormone (T3) and its receptor (TR) is the central mechanism of the pituitary-thyroid axis. We previously reported that such negative regulation is able to be observed in kidney derived CV1 cells in the presence of GATA2 and Pit1, which are the thyrotroph specific transcription factors. Using CAT-based reporter assays in CV1 cells, we have analyzed the function of the putative negative regulatory element (NRE) which has been reported to mediate the negative regulation of TSHβ gene by T3-bound TR (T3/TR). Unexpectedly, the negative regulation was maintained even in the deletion or the mutation of NRE. Further analysis revealed that the negative regulation was detected in the construct, TSHβ-D4-CAT, which possesses only Pit1 and GATA2 binding sites. Interestingly, TSHβ-D4-CAT is activated by the presence of GATA2 only and such a transactivation is specifically inhibited by T3/TR. We found that the Zn-finger of GATA2 interacts with the DNA binding domain (DBD) of TR in a T3 independent fashion, suggesting that the negative regulation of TSHβ promoter is mediated by the cross-talk between IR and GATA2. The suppression by T3/TR was recovered by the overexpression of dominant negative-type TR associated protein (TRAP) 220 indicating an essential participation of TRAP220 in the negative regulation of TSHβ gene. Chromatin immunoprecipitation assay with thyrotroph cell line, TαT1, showed that the acetylation of histone H4 is reduced by the addition of T3. In conclusion, the present study suggests (1) GATA2 is the major transcriptional activator for TSHβ promoter (2) T3/TR interacts and interferes with GATA2 (3) TRAP220 that lacks intrinsic histone acetyltransferase activity and the alteration of histone acetylation are involved in the mechanism of the negative regulation of TSHβ gene.

甲状腺激素（T3）及其受体（TR）对促甲状腺激素（促甲状腺激素，TSH）产生的负调节是垂体-甲状腺轴的中心机制。我们以前报道过，这种负调控是能够观察到在肾脏衍生的CV1细胞的存在下，GATA 2和Pit 1，这是促甲状腺细胞特异性转录因子。利用CAT报告基因分析技术，对T3结合TR（T3/TR）介导TSHβ基因负调控的负调控元件（NRE）的功能进行了研究。出乎意料的是，即使在NRE的缺失或突变中，负调控也保持不变。进一步的分析表明，在只有Pit 1和GATA 2结合位点的构建体TSHβ-D4-CAT中检测到负调控。有趣的是，TSHβ-D4-CAT仅被GATA 2激活，这种反式激活被T3/TR特异性抑制。我们发现GATA 2的锌指与TR的DNA结合域（DBD）以T3非依赖性方式相互作用，提示TSHβ启动子的负调控是由IR和GATA 2之间的相互作用介导的。T3/TR的抑制可通过显性负型TR相关蛋白（TRAP）220的过度表达而恢复，表明TRAP 220参与了TSHβ基因的负调控。用促甲状腺细胞系TαT1进行的染色质免疫沉淀试验表明，组蛋白H4的乙酰化被T3的加入所减少。结论：（1）GATA 2是TSHβ启动子的主要转录激活因子;（2）T3/TR与GATA 2相互作用并干扰;（3）缺乏内源性组蛋白乙酰转移酶活性的TRAP 220和组蛋白乙酰化改变参与了TSHβ基因负调控的机制。