Peripheral CLOCK Regulates Target-Tissue Glucocorticoid Receptor Transcriptional Activity in a Circadian Fashion in Man

Abstract

Context and ObjectiveCirculating cortisol fluctuates diurnally under the control of the “master” circadian CLOCK, while the peripheral “slave” counterpart of the latter regulates the transcriptional activity of the glucocorticoid receptor (GR) at local glucocorticoid target tissues through acetylation. In this manuscript, we studied the effect of CLOCK-mediated GR acetylation on the sensitivity of peripheral tissues to glucocorticoids in humans.Design and ParticipantsWe examined GR acetylation and mRNA expression of GR, CLOCK-related and glucocorticoid-responsive genes in peripheral blood mononuclear cells (PBMCs) obtained at 8 am and 8 pm from 10 healthy subjects, as well as in PBMCs obtained in the morning and cultured for 24 hours with exposure to 3-hour hydrocortisone pulses every 6 hours. We used EBV-transformed lymphocytes (EBVLs) as non-synchronized controls.ResultsGR acetylation was higher in the morning than in the evening in PBMCs, mirroring the fluctuations of circulating cortisol in reverse phase. All known glucocorticoid-responsive genes tested responded as expected to hydrocortisone in non-synchronized EBVLs, however, some of these genes did not show the expected diurnal mRNA fluctuations in PBMCs in vivo. Instead, their mRNA oscillated in a Clock- and a GR acetylation-dependent fashion in naturally synchronized PBMCs cultured ex vivo in the absence of the endogenous glucocorticoid, suggesting that circulating cortisol might prevent circadian GR acetylation-dependent effects in some glucocorticoid-responsive genes in vivo.ConclusionsPeripheral CLOCK-mediated circadian acetylation of the human GR may function as a target-tissue, gene-specific counter regulatory mechanism to the actions of diurnally fluctuating cortisol, effectively decreasing tissue sensitivity to glucocorticoids in the morning and increasing it at night.

Endocrine regulation of circadian physiology

“…Not only GC production but also GC action has been found to be gated by the molecular clock. GC receptors (GRs) are rhythmically expressed in various tissues and subject to acetylation (and subsequent inactivation) by CLOCK (Charmandari et al 2011). Recently, CRY1/2 proteins have been shown to directly repress the transactivational function of GR in a ligand-dependent manner , providing another layer of complexity to the circadian regulation of GR signaling.…”

Circadian Clock and Stress Interactions in the Molecular Biology of Psychiatric Disorders

“…In the adrenal, the availability of the steroid precursor cholesterol, the storage of cholesterol and the activity of steroidogenic enzymes all show circadian rhythms [26]. At the protein level, BMAL1/CLOCK complexes interact with the glucocorticoid receptor (GR), modulating GR sensitivity in a circadian fashion, through acetylation of the GR by CLOCK during the day [27]. Accordingly, Bmal1 -/-mice suffer from low CORT levels, reduced CORT sensitivity and a disturbed stress response [28].…”

Circadian clock mechanisms in adrenal adenomas are not related to osteopenia and osteoporosis

“…Further on, it is known that humans live under the strong influence of light/dark cycles associated with the day/night changes created by the 24-hour rotation of the earth 17,18 . This cycling has as a result the adaptation of the body physiology in the light/dark cycles.…”

“…This cycling has as a result the adaptation of the body physiology in the light/dark cycles. Through this system, human beings synchronize their physical activities, such as motivational behaviors, food intake, energy metabolism, sleep, reproductive activity and immune function [17][18][19][20] . The circadian system consists of central and peripheral components, which are located respectively in the suprachiasmatic nuclei (SCN) of the hypothalamus and virtually all remaining organs and tissues, including the extra-SCN brain 17,18.…”

“…Through this system, human beings synchronize their physical activities, such as motivational behaviors, food intake, energy metabolism, sleep, reproductive activity and immune function [17][18][19][20] . The circadian system consists of central and peripheral components, which are located respectively in the suprachiasmatic nuclei (SCN) of the hypothalamus and virtually all remaining organs and tissues, including the extra-SCN brain 17,18. The SCN acts as a "master" clock under the strong influence of light/dark input from the eyes, whereas the peripheral system behaves as a "slave" circadian rhythm generator subjugated by the SCN through as yet unclear neural and humoral connections 21 .…”