The Hypersensitive Glucocorticoid Response Specifically Regulates Period 1 and Expression of Circadian Genes

Abstract

dGlucocorticoids regulate gene expression by binding and activating the glucocorticoid receptor (GR). While ligand affinity determines the global sensitivity of the response, additional proteins act on the genome to tune sensitivity of some genes. However, the genomic extent and specificity of dose-specific glucocorticoid responses are unknown. We show that dose-specific glucocorticoid responses are extraordinarily specific at the genomic scale, able to distinctly express a single gene, the circadian rhythm gene for Period 1 (PER1), at concentrations consistent with the nighttime nadir of human cortisol. We mapped the PER1 response to a single GR binding site. The specific GR binding sequence did not impact sensitivity, and we instead attributed the response to a combination of additional transcription factors and chromatin accessibility acting in the same locus. The PER1 hypersensitive response element is conserved in the mouse, where we found similar upregulation of Per1 in pituitary cells. Targeted and transient overexpression of PER1 led to regulation of additional circadian rhythm genes hours later, suggesting that hypersensitive expression of PER1 impacts circadian gene expression. These findings show that hypersensitive GR binding occurs throughout the genome, drives targeted gene expression, and may be important to endocrine mediation of peripheral circadian rhythms.T he glucocorticoid receptor (GR) directs the transcriptional response to the steroid hormone cortisol (reviewed in reference 64). Glucocorticoids bind the receptor with high affinity, leading to nuclear translocation, direct binding to DNA, and ultimately regulation of gene expression. The affinity of glucocorticoid for the GR is a primary determinant of activation. Many studies have characterized GR binding and transcriptional regulatory activity at saturating concentrations of hormone at which most of the GR is bound (7,38,43,48,50,56). While these studies have revealed hundreds of genes that are differentially expressed in response to treatment with high concentrations of hormone, endogenous cortisol concentrations vary throughout the day as part of circadian rhythms (70) and are often near or below the dissociation constant (K d ) for the GR (46).Low doses of glucocorticoids that are well below the K d of the GR have been shown to elicit GR-mediated gene regulation for a small number of genes (47, 55). Such findings indicate that GR activity can be tuned to different doses of glucocorticoids in a tissue-specific manner, potentially in order to regulate expression of specific genes at different points in the circadian cycle. For example, subsaturating doses of the synthetic glucocorticoid dexamethasone (DEX) can drive expression of the tyrosine aminotransferase (TAT) gene in FU5-5 cells but not in the HTC rat hepatoma cell line (44). In addition, doses of DEX as low as 0.001 nM, well below the K d of DEX for the GR (K d , ϳ3 to 5 nM [31,66]), can act in a GR-and protein synthesis-dependent manner to inhibit the activity of vasoactive inte...

Glucocorticoid receptor recruits to enhancers and drives activation by motif-directed binding

“…We reanalyzed published data sets (Supplemental Table S5) to test the updated model, including GR binding data under varied dex concentrations and in different cellular contexts. We recapitulated the finding of Reddy et al (2012) that enhancers with GR binding at low dex concentration had increased initial accessibility (dex hypersensitive vs. low-sensitive; Mann-Whitney U test, P = 1.1 × 10 −14 ); however, the difference was starker in terms of initial EP300 (dex hypersensitive vs. low-sensitive; Mann-Whitney U test, P < 10 −100 ) (Fig. 3B,C).…”

“…We reasoned that GR binding sites present at low dex dosages may similarly depend on enhancer interactions, and we confirmed this with a comparable analysis (50 nM and 100 nM, 1 h dex; Mann-Whitney U test, P < 4.07 × 10-21) (Supplemental Fig. S6E; Reddy et al 2012).…”

Cistrome-based Cooperation between Airway Epithelial Glucocorticoid Receptor and NF-κB Orchestrates Anti-inflammatory Effects

“…First, there was little to no GR occupancy detected under basal culture conditions at binding sites within the KLF15 (Fig. 3B) and PER1 loci, which others have shown are robustly occupied by GR after exposure to low nanomolar concentrations of supplemental ligand (80). Second, whereas GR occupancy was markedly reduced at some sites after dex treatment, other sites had dramatically increased occupancy, and still other regions exhibited minimal changes with the addition of supplemental ligand (see Figs.…”

Mistimed sleep disrupts circadian regulation of the human transcriptome

“…What are the mechanisms underlying persistent rhythmicity and loss of rhythmicity? The circadian rhythms of both melatonin and cortisol are known to regulate gene expression (11,43) and are driven by the SCN. The fact that the vast majority of circadian transcripts become arrhythmic when sleeping out of phase with melatonin would indicate that the SCN neural outputs and hormones driven by the SCN (i.e., melatonin and cortisol only has a limited influence on the peripheral blood).…”

“…Furthermore, physiological factors such as body temperature (10) and endocrine rhythms such as cortisol (11) can modify and reinforce these regulatory processes.…”