CLOCK and BMAL1 regulate SIRT1 protein levels in mouse skeletal muscle and cultured myotubes. A, The protein levels of CLOCK, BMAL1, SIRT1, Ac-RelA, and PER2 in the muscle of C57BL/6 mice at ZT1 and ZT13 fasted for 12 hours. Ac-RelA was monitored to indicate the activity of SIRT1 (n = 4 for each group). B, SIRT1, Ac-RelA, and PER2 protein levels in the muscle of Clock^Δ19/Δ19 mice at ZT1 and ZT13 fasted for 12 hours (n = 4 for each group). C, SIRT1 and Ac-RelA protein levels in the muscle of Clock^Δ19/Δ19 mice at ZT7 (n = 4 for each group). D, Knockdown of CLOCK or BMAL1 in C2C12 myotube down-regulated SIRT1 protein level and activity as monitored by acetylated p53. E, Ectopic expression of CLOCK and BMAL1 by adenovirus (Ad-C&B) in C2C12 myotubes up-regulated SIRT1 protein level and activity. *, P < .05; **, P < .01; ***, P < .001.