Translational positioning of a nucleosomal glucocorticoid response element modulates glucocorticoid receptor affinity.

Abstract

Positioned nucleosomes are often found in enhancer/promoter regions where they confer defined positioning of trans-active-factor response element(s) relative to the histone octamer. Here, we address how this affects factor/response element recognition. We used 165-bp DNA segments containing one glucocorticoid response element (GRE) and rat liver core histones to reconstitute nucleosomes in vitro. The GREs in these nucleosomes were held in identical helical settings but different translational positions. This was achieved by placing the GRE within or at the flank of a 95-bp DNA-bending sequence. Glucocorticoid receptor (GR)-binding experiments demonstrated that a GRE in free DNA has a 2.5-fold higher affinity for GR than the nucleosomal GRE when positioned 40 bp from the nucleosome dyad. A nucleosomal GRE positioned 40 bp from the nucleosome dyad, on the other hand, binds GR 4.3-fold better as compared to an identical GRE positioned 20 bp from the dyad and 1.4-fold better than a GRE positioned at the dyad. Interruption of the DNA-bending sequence, by a 5-bp AT segment next to a nucleosomal GRE positioned 20 bp from the dyad, restores GR affinity to the same level as when the GRE is placed 40 bp from the dyad. The effect on GR/GRE affinity either by different positioning within the 95-bp bending sequence or by introducing the 5-bp AT-segment is seen only in a nucleosomal context. We conclude that a translationally positioned nucleosome can modulate the affinity of a trans-active factor for its target response element.