Phosphorylation of Human Thyroid Hormone Receptor beta-1 by Casein Kinase II

酪蛋白激酶 II 磷酸化人甲状腺激素受体 beta-1

基本信息

批准号：
09671018
负责人：
SUGAWARA Akira
金额：
$ 1.98万
依托单位：
Tohoku University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
1997
资助国家：
日本
起止时间：
1997 至 1998
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-09671018/
关键词：
Thyroid Hormone Receptor Phosphorylation

项目摘要

Bacterially-expressed human thyroid hormone receptor beta-1 (hTRbeta1) recently has been shown to be phosphorylated in vitro by HeLa cytoso1ic extract. In the present study, we first demonstrated that hTRbeta1 also could be phosphorylated in vitro b purified casein kinase II (CKII). Phosphoamino acid analysis revealed hTrbeta1 was phosphorylated on both serine and threonine residues. In vitro CKII phosphorylation of glutathione S-transferase (GST)-hTRbeta1 fusion proteins whose predicted CKII phosphorylation sites were mutated demonstrated that a threonine residue located in the hinge region(Thr-210) could be phosphorylated by CKII. In order to elucidate the functional significance of CKfl phosphorylation of Thr-210 in hTRbeta1, we next performed transient transfection studies using either wild type or Thr-210 mutated hTRbeta1.In terestingly, the basal repression eve in the absence of ligand was attenuated significantly when Thr-210 mutated hTRbeta1 was used. Since Thr-210 is located within the interacting domain with nudear receptor co-repressor (N-CoR), we next performed electrophoretic mobility shifi assay (EMSA)to examine the interaction between amino terminal-truncated N-CoR (NCoRI) and wild type or Thr-210 mutated hTRbeta1. Interestingly, in contrast to retinoid OMEGA receptor beta(RXRbeta) which equally formed heterodimers with both types of hTR beta1, N CoRIp referentially formed heterodimers with wild type hTRbeta1 than Thr-210mutated hTRbeta1. Taken together, we speculate that CKII phosphorylation ofThr-210 m hTRbeta1 might modulate interaction with N-Co R, and contribute tomediate basal repression in the absence of ligand.

细菌表达的人甲状腺激素受体β-1（HTRBETA1）最近已被HELA CYTOSO1IC提取物在体外磷酸化。在本研究中，我们首先证明了HTRBETA1在体外也可以磷酸化，并可以在体外纯化的酪蛋白激酶II（CKII）。磷酸氨基酸分析表明，丝氨酸和苏氨酸残基磷酸化htrbeta1。谷胱甘肽S-转移酶（GST）-HTRBETA1融合蛋白的体外CKII磷酸化，其预测CKII磷酸化位点的突变表明，位于铰链区域（THR-210）的苏氨酸残基可以由CKII磷酸化。为了阐明THRBETA1中THR-210 CKFL磷酸化的功能重要性，我们接下来使用野生型或THR-210突变的htrbeta1.iin进行了短暂转染研究，从而使基底抑制剂在缺乏配体的基础抑制剂中显着减轻，而当Thr-210突变时使用了HTRBETATED HTRBETATED HTRBETATAINE HTRBETATAINED HTRBETASA1。由于THR-210与裸体受体共抑制剂（N-COR）位于相互作用结构域中，因此我们接下来进行了电泳迁移率SHIFI测定（EMSA），以检查氨基终端截断的N-COR（NCORI）与野生型或THR-210突变的Htrbeta1之间的相互作用。有趣的是，与类似于野生型Htrbeta1的HTR Beta1的类型异二聚体相比，与类型的HTR Beta1相同，与THR-210的HTRBETA1相比，与类型的HTR Beta1相同形成的异二聚体相反，与类型的异二聚体相反，它同样形成异二聚体。综上所述，我们推测，THR-210 M HTRBETA1的CKII磷酸化可能会调节与N-CO R的相互作用，并在没有配体的情况下导致基础抑制作用。