High mobility group B proteins facilitate strong estrogen receptor binding to classical and half-site estrogen response elements and relax binding selectivity.
The estrogen receptor alpha (ER) is a ligand-dependent transcription factor that regulates the expression of estrogen-responsive genes. A key step in the activation process is the initial binding of the ER dimer to the estrogen response element (ERE). We examined the effect of the coactivator proteins, HMGB1 and HMGB2, in enhancing ER binding affinity to single and tandem EREs. Using EMSAs, both HMGB proteins are shown to enhance ER binding and induce cooperative ER binding on tandem ERE elements. We demonstrate that HMGB proteins facilitate strong ER binding to ERE consensus half-sites, exhibiting binding affinities comparable with ER binding to consensus ERE in the absence of HMGB proteins. These findings reveal that although HMGB proteins enhance binding affinity, they also relax ER binding specificity. Deoxyribonuclease I footprinting demonstrates that ER binds very differently to consensus ERE and ERE consensus half-sites, whereas both deoxyribonuclease I and exonuclease III digestions show that the presence of HMGB1/2 does not alter the DNA protection in ER/ERE complexes. Protease digestions of the complexes support this conclusion and show that a global conformation change occurs in ER when bound to the different ER binding sites. Models for these interactions are discussed, together with a hit-and-run mechanism that HMGB proteins may utilize to produce these effects.
Das
Dweepanita
D
Department of Chemistry, Bowling Green State University, Overman Hall, Bowling Green, Ohio 43403, USA.
Peterson
Ronald C
RC
Scovell
William M
WM
D000818:Animals; D001483:Base Sequence; D016384:Consensus Sequence; D018983:DNA Footprinting; D024202:Electrophoretic Mobility Shift Assay; D047628:Estrogen Receptor alpha / Q000378:metabolism*; D024243:HMGB1 Protein / Q000378:metabolism*; D024261:HMGB2 Protein / Q000378:metabolism*; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D020218:Response Elements / Q000235:genetics*
D047628:Estrogen Receptor alpha; D024243:HMGB1 Protein; D024261:HMGB2 Protein