Identification of the circadian transcriptome in adult mouse skeletal muscle

Abstract

Circadian rhythms are approximate 24-h behavioral and physiological cycles that function to prepare an organism for daily environmental changes. The basic clock mechanism is a network of transcriptional-translational feedback loops that drive rhythmic expression of genes over a 24-h period. The objectives of this study were to identify transcripts with a circadian pattern of expression in adult skeletal muscle and to determine the effect of the Clock mutation on gene expression. Expression profiling on muscle samples collected every 4 h for 48 h was performed. Using COSOPT, we identified a total of 215 transcripts as having a circadian pattern of expression. Real-time PCR results verified the circadian expression of the core clock genes, Bmal1, Per2, and Cry2. Annotation revealed cycling genes were involved in a range of biological processes including transcription, lipid metabolism, protein degradation, ion transport, and vesicular trafficking. The tissue specificity of the skeletal muscle circadian transcriptome was highlighted by the presence of known muscle-specific genes such as Myod1, Ucp3, Atrogin1 (Fbxo32), and Myh1 (myosin heavy chain IIX). Expression profiling was also performed on muscle from the Clock mutant mouse and sarcomeric genes such as actin and titin, and many mitochondrial genes were significantly downregulated in the muscle of Clock mutant mice. Defining the circadian transcriptome in adult skeletal muscle and identifying the significant alterations in gene expression that occur in muscle of the Clock mutant mouse provide the basis for understanding the role of circadian rhythms in the daily maintenance of skeletal muscle.

CLOCK and BMAL1 regulate MyoD and are necessary for maintenance of skeletal muscle phenotype and function

“…The loss of circadian expression of MyoD in skeletal muscle from both Clock Δ19 and Bmal1 −/− mice is very similar to what has been previously observed with other genes known to be regulated by BMAL1 and CLOCK, such as Per2 and Dbp (referred to as clock-controlled genes) (7,8). Results from reporter gene assays and ChIP experiments confirmed that MyoD is a direct target of CLOCK and BMAL1.…”

“…MyoD is a wellestablished skeletal muscle-specific transcription factor that directly regulates expression of the myogenic program (9). In addition to MyoD, we found that the core components of the molecular clock, including Bmal1 and Per2 expression, were oscillating as is seen in liver and other peripheral tissues (7). Interestingly, MyoD expression, like Per2 mRNA, did not oscillate in muscle of the Clock Δ19 mutant mouse, suggesting the possibility that MyoD is a clock-controlled gene.…”

“…To date, however, no genetic studies have examined what role circadian rhythms play in adult skeletal muscle. Our earlier results from an expression profiling study identified circadian expression of the components of the molecular clock (e.g., Bmal1, Per2) and showed that expression of MyoD, a master regulator of skeletal muscle differentiation, followed a circadian pattern (7,8).…”

“…We recently reported that myogenic differentiation 1 (MyoD) mRNA is expressed in a circadian fashion in adult skeletal muscle (7). The identification of MyoD as a circadian gene was of interest because MyoD is well-established as a master regulator of the skeletal muscle transcriptional program (9).…”

“…These mRNAs exhibit significant oscillation in gene expression with a repeating period length of 24 h. One of the intriguing observations from the array data was the finding that MyoD mRNA exhibited a circadian pattern (7,8). MyoD is a wellestablished skeletal muscle-specific transcription factor that directly regulates expression of the myogenic program (9).…”