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Chemosphere. 2012 May;87(7):782-8. doi: 10.1016/j.chemosphere.2011.12.080. Epub 2012 Jan 25.

Tetrabromobisphenol-A disrupts thyroid hormone receptor alpha function in vitro: use of fluorescence polarization to assay corepressor and coactivator peptide binding.

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  • 1Univ Paris-Sud, Public Health-Environment, UMR 8079, France.

Abstract

Thyroid hormone receptors (TRs) recruit corepressor or coactivator factors to the promoters of target genes to regulate their transcription. Corepressors such as nuclear hormone receptor corepressor (NCoR) are recruited by unliganded TRs, whereas coactivators such as steroid receptor coactivator-2 (SRC2) are recruited when triiodothyronine (T3) is bound to TRs. These coregulator proteins interact with the ligand binding domain (LBD) of TRs via short, conserved peptide sequences that can be used to probe the conformational changes induced in TR LBD by TR ligands. Recombinant LBD of the human TRα1 isoform (hTRα1 LBD) was produced as a fusion with glutathione S-transferase, and used to develop assays based on fluorescence polarization to quantify the binding of either NCoR- or SRC2-derived fluorescent peptides to the hTRα1 LBD. The optimum concentrations of recombinant hTRα1 LBD, and of peptide probes were adjusted in order to produce the greatest possible T3-dependent signal variations in fluorescence polarization. Under these conditions, T3 induced a dose-dependent decrease in NCoR peptide binding, and a reciprocal dose-dependent increase in SRC2 peptide binding, in both cases at similar 50%-effective doses. The TR agonists triiodothyroacetic acid and thyroxine were also effective in preventing NCoR peptide binding and increasing SRC2 peptide binding, whereas reverse-triiodothyronine was less efficient and the biologically inactive thyronine had no effect on either process. These experiments validate cell-free assays based on the use of binding of corepressor or coactivator peptide probes, as measured by fluorescence polarization, for investigating the conformational changes of TRα1 LBD induced by potentially TR-interfering compounds. Both these methods were used to elucidate the mechanism of the disrupting effects of tetrabromobisphenol-A (TBBPA) on the hTRα1 LBD conformation related to the transcriptional activity of the receptor. TBBPA is a flame retardant that is released into the environment, and is a suspected disrupter of thyroid homeostasis. The present results indicate that TBBPA did indeed interfere with the ability of the hTRα1 LBD to bind both NCoR and SRC2. TBBPA behaved similarly to T3 in promoting the release of NCoR from LBD, whereas it failed to promote LBD interactions with SRC2. However, it did reduce the T3-induced interactions between LBD and the coactivator peptide. This study therefore suggests that TBBPA in the micromolar range can affect the regulation of transcription by both the apo- and the holo-TRα1, with potential disruption of the expression of genes that are either up- or down-regulated by T3.

Copyright © 2012 Elsevier Ltd. All rights reserved.

PMID:
22277881
[PubMed - indexed for MEDLINE]
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