Cooperative binding of estrogen receptor to imperfect estrogen‐responsive DNA elements correlates with their synergistic hormone‐dependent enhancer activity.

The Xenopus vitellogenin (vit) gene B1 estrogen‐inducible enhancer is formed by two closely adjacent 13 bp imperfect palindromic estrogen‐responsive elements (EREs), i.e. ERE‐2 and ERE‐1, having one and two base substitutions respectively, when compared to the perfect palindromic consensus ERE (GGTCANNNTGACC). Gene transfer experiments indicate that these degenerated elements, on their own, have a low or no regulatory capacity at all, but in vivo act together synergistically to confer high receptor‐ and hormone‐dependent transcription activation to the heterologous HSV thymidine kinase promoter. Thus, the DNA region upstream of the vitB1 gene comprising these two imperfect EREs separated by 7 bp, was called the vitB1 estrogen‐responsive unit (vitB1 ERU). Using in vitro protein‐DNA interaction techniques, we demonstrate that estrogen receptor dimers bind cooperatively to the imperfect EREs of the vitB1 ERU. Binding of a first receptor dimer to the more conserved ERE‐2 increases approximately 4‐ to 8‐fold the binding affinity of the receptor to the adjacent less conserved ERE‐1. Thus, we suggest that the observed synergistic estrogen‐dependent transcription activation conferred by the pair of hormone‐responsive DNA elements of the vit B1 ERU is the result of cooperative binding of two estrogen receptor dimers to these two adjacent imperfect EREs.