Ligand-selective interactions of ER detected in living cells by fluorescence resonance energy transfer.

Journal Article (Journal Article)

Some aspects of ligand-regulated transcription activation by the estrogen receptor (ER) are associated with the estrogen-dependent formation of a hydrophobic cleft on the receptor surface. At least in vitro, this cleft is required for direct interaction of ER with an alpha helix, containing variants of the sequence LXXLL, found in many coactivators. In cells, it is unknown whether ER interactions with the different LXXLL-containing helices are uniformly similar or whether they vary with LXXLL sequence or activating ligand. Using fluorescence resonance energy transfer (FRET), we confirm in the physiological environment a direct interaction between the estradiol (E2)-bound ER and LXXLL peptides expressed in living cells as fusions with spectral variants of the green fluorescent protein. This interaction was blocked by a single amino acid mutation in the hydrophobic cleft. No FRET was detected when cells were incubated with the antiestrogenic ligands tamoxifen and ICI 182,780. E2, diethylstilbestrol, ethyl indenestrol A, and 6,4'-dihydroxyflavone all promoted FRET and activated ER-dependent transcription. Measurement of the level of FRET of ER with different LXXLL-containing peptides suggested that the orientations or affinities of the LXXLL interactions with the hydrophobic cleft were globally similar but slightly different for some activating ligands.

Full Text

Duke Authors

Cited Authors

  • Weatherman, RV; Chang, C-Y; Clegg, NJ; Carroll, DC; Day, RN; Baxter, JD; McDonnell, DP; Scanlan, TS; Schaufele, F

Published Date

  • March 2002

Published In

Volume / Issue

  • 16 / 3

Start / End Page

  • 487 - 496

PubMed ID

  • 11875107

International Standard Serial Number (ISSN)

  • 0888-8809

Digital Object Identifier (DOI)

  • 10.1210/mend.16.3.0813


  • eng

Conference Location

  • United States