Glucocorticoid Receptor Quaternary Structure Drives Chromatin Occupancy and Transcriptional Outcome

Most transcription factors, including nuclear receptors, are widely modeled as binding regulatory elements as monomers, homodimers, or heterodimers. Recent findings in live cells show that the glucocorticoid receptor (GR) forms tetramers on enhancers, due to an allosteric alteration induced by DNA binding, and suggest that higher oligomerization states are important for the gene regulatory responses of GR. Using a variant (GRtetra) that mimics this allosteric transition, we performed genome-wide studies using a GR knock-out cell line with reintroduced wild-type GR or reintroduced GRtetra. GRtetra acts as a super receptor by binding to response elements not accessible to wild-type receptor, and both induces and represses more genes than GRwt. These results argue that DNA binding induces a structural transition to the tetrameric state, forming a transient higher order structure that drives both the activating and repressive actions of glucocorticoids. Overall design: Genome-wide occupancy profiling of GR and active histone marks by ChIP-seq from mouse mammary adenocarcinoma cell lines (3617) knock-out of endogenous GR (3617-KOGR) and stably integrated with GFP-tagged GR mutants, in biological duplicates, using Illumina NextSeq. GR binding data from the 3617-KOGR cells was used as control for GR and input sample from 3617-KOGR cells were used as controls for the rest. Chromatin accessibility examined by ATAC-seq from GR mutant cell lines, in biological duplicates, using Illumina HiSeq4000. Gene expression profiling from GR mutant cell lines by RNA-seq, in biological triplicates, using Illumina HiSeq2500. Genome-wide occupancy profiling of GR by ChIP-seq from mouse embryonic fibroblast (MEF) cell lines knock-out of endogenous GR (MEF-KOGR) and stably integrated with GFP-tagged GR mutants, in biological duplicates, using Illumina NextSeq. GR binding data from the MEF-KOGR cells was used as control.