Oxidant stress impaired DNA-binding of estrogen receptor from human breast cancer

Xiaoshan Liang, Biao Lu, Gary K. Scott, Chuan Hsiung Chang, Michael A. Baldwin, Christopher C. Benz*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

55 Scopus citations

Abstract

Full-length (67 kDa) immunoreactive estrogen receptor (ER) extracted from a third of untreated ER-positive primary breast tumors appears unable to bind to its cognate estrogen response element (ERE). We have observed partial reversibility of this ER DNA-binding defect upon treatment of these tumor extracts with excess thiol reducing agent (DTT), suggesting that ER DNA- binding is subject to redox modulation as is reported for other zinc-finger proteins and transcriptional activators. Treatment of recombinant ER DNA- binding domain (ER-DBD) or ER-enriched extracts from CHO(ER) and MCF-7 γ cells with thiol-reacting oxidants (diamide, iodosobenzoate, H2O2) or alkylator (iodoacetamide) produces a dose-dependent loss in ER DNA-binding capacity. Thiol-specific oxidative loss in ER DNA-binding is fully reversible by DTT reduction, unlike the defect caused by thiol-specific alkylation. Circular dichroism spectrometry shows that both forms of treatment substantially modify ER secondary structure, inducing loss of β-helical content within the ER-DBD that is reversible after thiol oxidation but not after thiol alkylation. Oxidant (H2O2, menadione) exposure of cultured CHO(ER) or MCF-7 cells impairs the ability of endogenous ER to bind DNA and transactivate an ER-responsive reporter gene (ERE-tk-CAT), demonstrating that extracellular redox stress can modulate intracellular ER function. Since these thiol-specific oxidant and alkylator treatments have no significant effect on either recombinant ER ligand-binding or intracellular immunoreactive ER content, our findings suggest that DNA-binding and transactivation are the most sensitive intracellular ER functions impaired by oxidant stress in some ER-positive human breast tumors.

Original languageEnglish
Pages (from-to)151-161
Number of pages11
JournalMolecular and Cellular Endocrinology
Volume146
Issue number1-2
DOIs
StatePublished - 25 Nov 1998

Keywords

  • Cell lines
  • ER DNA-binding
  • Estrogen receptor (ER)
  • Human breast tumors
  • Oxidant stress

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