Relationships between circadian rhythms and modulation of gene expression by glucocorticoids in skeletal muscle

Abstract

The existence and maintenance of biological rhythms linked to the 24-h light-dark cycle are essential to the health and functioning of an organism. Although much is known concerning central clock mechanisms, much less is known about control in peripheral tissues. In this study, circadian regulation of gene expression was examined in rat skeletal muscle. A rich time series involving 54 animals euthanized at 18 distinct time points within the 24-h cycle was performed, and mRNA expression in gastrocnemius muscles was examined using Affymetrix gene arrays. Data mining identified 109 genes that were expressed rhythmically, which could be grouped into eight distinct temporal clusters within the 24-h cycle. These genes were placed into 11 functional categories, which were examined within the context of temporal expression. Transcription factors involved in the regulation of central rhythms were examined, and eight were found to be rhythmically expressed in muscle. Because endogenous glucocorticoids are a major effector of circadian rhythms, genes identified here were compared with those identified in previous studies as glucocorticoid regulated. Of the 109 genes identified here as circadian rhythm regulated, only 55 were also glucocorticoid regulated. Examination of transcription factors involved in circadian control suggests that corticosterone may be the initiator of their rhythmic expression patterns in skeletal muscle.

Fig. 1.

A nonlinear curve fit using MATLAB was conducted which fitted a sinusoid function [A·sin(Bt + c)] to the data, including the replicates. Genes that could be curve fitted with a R² correlation of greater than 0.8 were kept.

Fig. 2.

QT clustering of genes in muscle measured over 24 h. Each probe set has greater than a 0.75 Pearson's correlation with the centroid of the cluster.

Fig. 3.

Plasma corticosterone (CST) as a function of circadian time as measured by HPLC. Unshaded areas indicate light period and shaded areas indicate dark period.

Fig. 4.

Expression patterns of 8 clock-related transcription factors in skeletal muscle as a function of circadian time. Unshaded areas indicate light periods, and shaded areas indicate dark periods.

Fig. 5.

Expression patterns of 5 clock-related transcription factors in skeletal muscle as a function of time after methylprednisolone (MPL) administration to adrenalectomized animals. Left: data from acute (bolus 50 mg/kg) MPL dosing. Right: data from chronic (0.3 mg·kg⁻¹·h⁻¹) MPL infusion.

Fig. 6.

Expression patterns of 3 clock-related transcription factors in skeletal muscle as a function of time after MPL administration to adrenalectomized animals. Left: data from acute (bolus 50 mg/kg) MPL dosing. Right: data from chronic (0.3 mg·kg⁻¹·h⁻¹) MPL infusion.

Fig. 7.

Expression patterns of GS (left) and Eif4ebp1 (right) following chronic infusion of MPL (0.3 mg·kg⁻¹·h⁻¹) over 168 h (top), and within the 24 h circadian cycle (bottom).

Fig. 8.

Circadian pattern of expression of glutamine synthetase measured by gene arrays (•) and by kinetic-based quantitative RT PCR (○).