Sequences flanking the core-binding site modulate glucocorticoid receptor structure and activity.

Stefanie Schöne,Remo Rohs,Martin Vingron,Isabelle Lebars,Petra Imhof,Iris Dror,Bruno Kieffer,Morgane Thomas-Chollier,Marcel Jurk,Mahdi Bagherpoor Helabad,Sebastiaan H Meijsing

doi:10.1038/ncomms12621

Abstract

The glucocorticoid receptor (GR) binds as a homodimer to genomic response elements, which have particular sequence and shape characteristics. Here we show that the nucleotides directly flanking the core-binding site, differ depending on the strength of GR-dependent activation of nearby genes. Our study indicates that these flanking nucleotides change the three-dimensional structure of the DNA-binding site, the DNA-binding domain of GR and the quaternary structure of the dimeric complex. Functional studies in a defined genomic context show that sequence-induced changes in GR activity cannot be explained by differences in GR occupancy. Rather, mutating the dimerization interface mitigates DNA-induced changes in both activity and structure, arguing for a role of DNA-induced structural changes in modulating GR activity. Together, our study shows that DNA sequence identity of genomic binding sites modulates GR activity downstream of binding, which may play a role in achieving regulatory specificity towards individual target genes.

Highlights

The glucocorticoid receptor (GR) binds as a homodimer to genomic response elements, which have particular sequence and shape characteristics
Our studies suggest that modulation of GR activity and structure by GR-binding sequence (GBS) variation at positions directly adjacent to the core recognition sequence plays a role in fine-tuning the expression of endogenous target genes
When we compared the spectra of the complexes between GR DNA-binding domain (DBD) and G/C and A477T DBD (A/T) flanked Cgt oligonucleotides, we found a number of differences between spectra (Supplementary Fig. 5B)

Summary

Introduction

The glucocorticoid receptor (GR) binds as a homodimer to genomic response elements, which have particular sequence and shape characteristics. We first studied if GBS variants can modulate GR activity in a chromosomal context and found that GBS variants can modulate GR activity when integrated at a defined genomic locus This modulation appears to occur downstream of GR binding as the differences in transcriptional responses cannot be explained by differences in occupancy levels based on chromatin immunoprecipitation (ChIP) experiments. Using a combination of experiments with atomic resolution and functional studies, we found that the base pairs directly flanking the core 15-bp GBS modulate GR activity and induce structural changes in both DNA and the associated DNA-binding domain of GR. Our studies suggest that modulation of GR activity and structure by GBS variation at positions directly adjacent to the core recognition sequence plays a role in fine-tuning the expression of endogenous target genes

Methods

Results

Conclusion