Glucocorticoid Receptor-Tethered Mineralocorticoid Receptors Increase Glucocorticoid-Induced Transcriptional Responses.

Caroline A Rivers,Stafford L Lightman,John R Pooley,Felicity E Stubbs,Becky L Conway-Campbell,Mark F Rogers

doi:10.1210/en.2018-00819

Abstract

Mineralocorticoid and glucocorticoid receptors (MRs and GRs) constitute a functionally important dual receptor system detecting and transmitting circulating corticosteroid signals. High expression of MRs and GRs occurs in the same cells in the limbic system, the primary site of glucocorticoid action on cognition, behavior, and mood; however, modes of interaction between the receptors are poorly characterized. We used chromatin immunoprecipitation with nucleotide resolution using exonuclease digestion, unique barcode, and single ligation (ChIP-nexus) for high-resolution genome-wide characterization of MR and GR DNA binding profiles in neuroblastoma cells and demonstrate recruitment to highly similar DNA binding sites. Expressed MR or GR showed differential regulation of endogenous gene targets, including Syt2 and Ddc, whereas coexpression produced augmented transcriptional responses even when MRs were unable to bind DNA (MR-XDBD). ChIP confirmed that MR-XDBD could be tethered to chromatin by GR. Our data demonstrate that MR can interact at individual genomic DNA sites in multiple modes and suggest a role for MR in increasing the transcriptional response to glucocorticoids.

Highlights

Mineralocorticoid and glucocorticoid receptors (MRs and GRs) constitute a functionally important dual receptor system detecting and transmitting circulating corticosteroid signals
The EGFP tag on GR and the mCherry tag on mineralocorticoid receptor (MR) did not significantly alter transactivation potential in Neuro 2A (N2A) cells, subtle changes in response amplitude prompted the use of untagged MR and GR in all experiments when tags were not essential for chromatin immunoprecipitation (ChIP)
We propose a new mode of action for MR in which it can alter transcription when tethered to the DNA by GR

Summary

Introduction

Mineralocorticoid and glucocorticoid receptors (MRs and GRs) constitute a functionally important dual receptor system detecting and transmitting circulating corticosteroid signals. High expression of MRs and GRs occurs in the same cells in the limbic system, the primary site of glucocorticoid action on cognition, behavior, and mood; modes of interaction between the receptors are poorly characterized. High CORT levels circulate during the active phase of the day, with significantly lower levels in the inactive phase establishing a characteristic circadian profile. Underlying this circadian profile is an ultradian profile with approximately hourly pulses of endogenous glucocorticoid detected in rat and human peripheral blood [1, 2], dialysates of subcutaneous extracellular fluid [3], and dialysates of extracellular fluid from discrete brain regions, including the hippocampus, of experimental rats [4]. Specific binding sites have been identified for MRs and GRs in rat hippocampus [14], with overlapping hippocampal binding sites reported to range from 20% of total GR binding sites (475 out of 1925) [14] to 77% (10 out of 13) GR binding sites tested [15], these studies cannot conclusively say that MR and GR binding occurs in the same cell types

Methods

Results

Conclusion