Identification of the putative MAP kinase docking site in the thyroid hormone receptor-β1 DNA-binding domain: Functional consequences of mutations at the docking site

Hung Yun Lin, ShenLi Zhang, Brian L. West, Heng Yuan Tang, Teresa Passaretti, Faith B. Davis, Paul J. Davis

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47 引文 斯高帕斯(Scopus)

摘要

In CV-1 cells transfected with wild-type (wt) nuclear thyroid hormone receptor TRβ1 (TR), L-thyroxine (T4) causes activation and nuclear translocation of mitogen-activated protein kinase (MAPK, ERK1/2), co-immunoprecipitation of MAPK and TR, and MAPK-dependent serine phosphorylation of TR. In the present studies, we have identified (1) the likely site of TR serine phosphorylation in the TR DNA-binding domain (DBD) by T4-activated MAPK, (2) the site of MAPK docking on TR induced by T4, and (3) functional consequences of TR docking site and serine phosphorylation site mutations on co-repressor and co-activator binding and on transcriptional activation by wt and mutant receptors in T4-treated cells. Plasmids containing TRwt, serine 142-substituted TR (TRS142A or TRS142E), TRK128A, TRR132A, or TRR133A were transfected into CV-1 cells, and the cells were treated with 10-7 M T4 for 30 min. Activated MAPK was present in nuclear fractions of all T4-treated cells and co-immunoprecipitated prominently with TRwt, TRS142A, and TRS142E. TRK128A complexing with activated MAPK was minimally detectable, but no association of MAPK with TRR132A or TRR133A was seen in cells treated with T4. Serine phosphorylation of TRwt, but not of any mutants, occurred with T4. In in vitro phosphorylation studies, constitutively activated MAPK phosphorylated only TRwt. We concluded that serine 142 of the TR DBD is the likely site of phosphorylation by T4-activated MAPK and that the docking site on TR for activated MAPK includes residues 128-133 (KGFFRR), a basic amino acid-enriched motif novel for MAPK substrates. TR mutations in the proposed MAPK docking domain and at residue 142 modulated T4-conditioned shedding of co-repressor and recruitment of co-activator proteins by the receptor, and they altered transcriptional activity of TR in a thyroid hormone response element - luciferase reporter assay.
原文英語
頁(從 - 到)7571-7579
頁數9
期刊Biochemistry
42
發行號24
DOIs
出版狀態已發佈 - 六月 24 2003
對外發佈

ASJC Scopus subject areas

  • 生物化學

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