Circadian phase-dependent myoblast differentiation and muscle regeneration

Circadian rhythms regulate cell differentiation and tissue regeneration; however, their implications in tissue regeneration remain elusive. Here, we present evidence that tissue regeneration efficiency is dependent on the circadian timing of injury affliction in a mouse model of muscle injury. We found that depletion of Per1 or Per2, members of the core circadian regulators in vertebrates, impaired mouse muscle regeneration and myoblast differentiation. Both Per1 and Per2 were necessary to maintain the expression level of Igf2, one of the critical regulators for myoblast differentiation, through epigenetic regulation at the Igf2 promoter and enhancer. Reflecting this, myoblast differentiation was promoted when differentiation was initiated while the level of Pers and Igf2 were high, compared with the time when their expression levels were low. Furthermore, muscle regeneration in the mouse was more efficient when chemical injury was inflicted at night when Pers were highly expressed, compared with morning injury. This study uncovers circadian timing-based control of myoblast differentiation and muscle regeneration. Overall design: Per1 and Per2 genes were knocked out in mouse myoblast C2C12 cells with CRISPR-Cas9. The cells were induced to differentiate into myotubes. RNA from undifferentiated (day 0) and differentiating (day 3 and day 5) cells were applied to RNA-seq. n=1. WT is control treated with non-target sgRNA.