Bending of the estrogen response element by polyamines and estrogen receptors α and β:: A fluorescence resonance energy transfer study

Estrogenic regulation of gene expression is mediated by the binding of the hormone to its receptors (ER alpha and ER beta) followed by their binding to estrogen response element (ERE). Previous studies showed that natural polyamines-putrescine, spermidine, and spermine-facilitated ER alpha(ERE)-E-. recognition. We determined the effects of natural and synthetic polyamines on the bending of a 27-mer oligonucleotide (ODN) harboring the ERE (ERE-ODN), using fluorescence resonance energy transfer (FRET) technique. Complementary strands of the ERE-ODN were labeled with fluorescein and tetramethylrhodamine, as donor and acceptor, respectively. The ERE-ODN was intrinsically bent with an end-to-end distance of 76 +/- 2 angstrom, compared to a theoretical value of 98 angstrom. The end-to-end distance of the ERE-ODN was reduced to 64 angstrom in the presence of 250 mu M spermine. A control ODN with scrambled sequence did not show intrinsic bending or spermine-induced bending. Alkyl substitution at the pendant amino groups reduced the ability of spermine to bend the ERE-ODN. Both ER alpha and ER beta decreased the end-to-end distance of the ERE-ODN, although ER alpha. was more efficient than ER beta in inducing ERE bending. Spermine-induced bending of the ERE-ODN was significantly increased by ER alpha. Fluorescence anisotropy measurement showed that the equilibrium association constant of ER alpha-ERE binding increased by 12-fold in the presence of 250 mu M spermine compared to control. The free energy change (AG) of ER alpha-ERE complex formation was -13.1 kcal/mol at 22 degrees C in the presence of spermine. Our results suggest that polyamine-induced bending of the ERE might be a mechanism for enhancing ER alpha-ERE binding affinity and thereby fine-tuning the transcriptional response of estrogen-responsive genes. (c) 2006 Elsevier Ltd. All rights reserved.