Modulation of the circadian clock by glucocorticoid receptor isoforms in the H295R cell line.

Abstract

Peripheral clocks are set by different nervous, hormonal and metabolic stimuli, and regulate the circadian expression of several genes. We investigated whether a peripheral clock could be induced in the human adrenocortical cell line H295R and whether glucocorticoid receptor isoforms (GRalpha and GRss) are involved in this clock system. After synchronization of cells with serum shock, the rhythmic oscillation of clock genes PER1, PER2, REV-ERBalpha, and ARNTL was confirmed. In addition, H295R cells even without serum shock showed rhythmic expression of PER1, PER2, CRY1 and ARNTL. Glucocorticoid treatment induced a rapid response of PER1, PER2 and CRY1 in a GRalpha-dependent manner. Continuous glucocorticoid stimulation after 6h caused suppression of REV-ERBalpha. Administration of a GR antagonist, RU486, disrupted the circadian oscillation of clock genes and prevented the acute changes in PER1, PER2 and CRY1 levels. Overexpression of the GRss isoform alone did not alter the expression of the examined clock genes, but did prevent the GRalpha-related suppression of REV-ERBalpha. These alterations occurred independently from ACTH and CRH. Our data demonstrate that a peripheral clock system is present in a human adrenocortical cell line and that periodic oscillations of clock genes are influenced by glucocorticoids, mainly through GRalpha.