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Updated: Jul 14, 2026

Detecting the Ligand-binding Domain Dimerization Activity of Estrogen Receptor Alpha Using the Mammalian Two-Hybrid Assay
09:07

Detecting the Ligand-binding Domain Dimerization Activity of Estrogen Receptor Alpha Using the Mammalian Two-Hybrid Assay

Published on: December 19, 2018

Direct interaction between estrogen receptor alpha and NF-kappaB in the nucleus of living cells

Monique E Quaedackers1, Christina E van den Brink, Paul T van der Saag

  • 1Hubrecht Laboratory, Netherlands Institute for Developmental Biology, Uppsalalaan 8, 3584 CT Utrecht, The Netherlands. m.quaedackers@erasmusmc.nl

Molecular and Cellular Endocrinology
|June 26, 2007
PubMed

Insights

Estrogen receptor alpha (ERalpha) directly interacts with NF-kappaB proteins in the cell nucleus. This protein-protein interaction is crucial for the anti-inflammatory effects of estrogen signaling.

Area of Science:

  • Molecular Biology
  • Cell Biology
  • Endocrinology

Background:

  • Steroid receptors, like estrogen receptor alpha (ERalpha), inhibit NF-kappaB transcriptional activity, contributing to anti-inflammatory effects.
  • The molecular mechanism for this cross-talk is hypothesized to involve direct protein-protein interactions.

Purpose of the Study:

  • To investigate and confirm the direct in vivo interaction between ERalpha and NF-kappaB.
  • To elucidate the cellular localization and conditions favoring this interaction.

Main Methods:

  • Utilized fluorescence resonance energy transfer (FRET) and co-immunoprecipitation in U2-OS cells.
  • Employed fluorescent fusion proteins of ERalpha and NF-kappaB subunits (p50, p65).
  • Assessed interactions in a stable ERalpha-expressing U2-OS clone and with NF-kappaB deletion mutants.

Main Results:

  • Demonstrated direct in vivo interaction between ERalpha and NF-kappaB subunits within the cell nucleus.
  • Maximal interaction observed in the nucleus upon estrogen stimulation and co-expression of ERalpha with both NF-kappaB subunits.
  • The Rel homology domain of p65 was identified as critical for ERalpha interaction.

Conclusions:

  • Provided strong evidence supporting a direct protein-protein interaction model for ERalpha and NF-kappaB cross-talk.
  • This interaction is a key molecular mechanism underlying the anti-inflammatory actions mediated by ERalpha.
  • Findings highlight the importance of nuclear localization and specific protein domains in mediating receptor-transcription factor cross-talk.

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