Glucocorticoid receptor condensates link DNA-dependent receptor dimerization and transcriptional transactivation.
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ABSTRACT:
SUBMITTER: Frank F
PROVIDER: S-EPMC8325269 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:PurposeGlucocorticoid (GC)-induced ocular hypertension (GC-OHT) is a serious side effect of prolonged GC therapy that can lead to glaucoma and permanent vision loss. GCs cause a plethora of changes in the trabecular meshwork (TM), an ocular tissue that regulates intraocular pressure (IOP). GCs act through the glucocorticoid receptor (GR), and the GR regulates transcription both through transactivation and transrepression. Many of the anti-inflammatory properties of GCs are mediated by GR transrepression, while GR transactivation largely accounts for GC metabolic effects and side effects of GC therapy. There is no evidence showing which of the two mechanisms plays a role in GC-OHT.MethodsGRdim transgenic mice (which have active transrepression and impaired transactivation) and wild-type (WT) C57BL/6J mice received weekly periocular dexamethasone acetate (DEX-Ac) injections. IOP, outflow facilities, and biochemical changes to the TM were determined.ResultsGRdim mice did not develop GC-OHT after continued DEX treatment, while WT mice had significantly increased IOP and decreased outflow facilities. Both TM tissue in eyes of DEX-treated GRdim mice and cultured TM cells isolated from GRdim mice had reduced or no change in the expression of fibronectin, myocilin, collagen type I, and α-smooth muscle actin (α-SMA). GRdim mouse TM (MTM) cells also had a significant reduction in DEX-induced cytoskeletal changes, which was clearly seen in WT MTM cells.ConclusionsWe provide the first evidence for the role of GR transactivation in regulating GC-mediated gene expression in the TM and in the development of GC-OHT. This discovery suggests a novel therapeutic approach for treating ocular inflammation without causing GC-OHT and glaucoma.
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Project description:Glucocorticoid receptor (GR) levels vary between tissues and individuals and are altered by physiological and pharmacological effectors. However, the effects and implications of differences in GR concentration have not been fully elucidated. Using three statistically different GR concentrations in transiently transfected COS-1 cells, we demonstrate, using co-immunoprecipitation (CoIP) and fluorescent resonance energy transfer (FRET), that high levels of wild type GR (wtGR), but not of dimerization deficient GR (GRdim), display ligand-independent dimerization. Whole-cell saturation ligand-binding experiments furthermore establish that positive cooperative ligand-binding, with a concomitant increased ligand-binding affinity, is facilitated by ligand-independent dimerization at high concentrations of wtGR, but not GRdim. The down-stream consequences of ligand-independent dimerization at high concentrations of wtGR, but not GRdim, are shown to include basal priming of the system as witnessed by ligand-independent transactivation of both a GRE-containing promoter-reporter and the endogenous glucocorticoid (GC)-responsive gene, GILZ, as well as ligand-independent loading of GR onto the GILZ promoter. Pursuant to the basal priming of the system, addition of ligand results in a significantly greater modulation of transactivation potency than would be expected solely from the increase in ligand-binding affinity. Thus ligand-independent dimerization of the GR at high concentrations primes the system, through ligand-independent DNA loading and transactivation, which together with positive cooperative ligand-binding increases the potency of GR agonists and shifts the bio-character of partial GR agonists. Clearly GR-levels are a major factor in determining the sensitivity to GCs and a critical factor regulating transcriptional programs.
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