New insights into the skeletal muscle circadian clock in ruminants

The core circadian clock components are expressed and functionally active in peripheral tissues, such as the intestine, liver, and skeletal muscle, where their molecular mechanisms remain highly conserved [26–28]. The light‒dark cycle acts as the principal zeitgeber (ZT), synchronizing the central clock in the SCN via photic input through the retina. This central timing signal is then transmitted to peripheral tissues via neural, hormonal, and behavioral pathways, establishing a system-wide circadian network [29, 30]. Importantly, peripheral clocks are entrained by non-photic cues, including feeding-fasting cycles, temperature variations, and physical activity [29]. Desynchronization between the central and peripheral clocks can disrupt metabolic homeostasis, immune function, and tissue development and is linked to pathologies in skeletal muscle, including muscle atrophy, sarcopenia, and muscular dystrophy, underscoring the importance of rhythmicity in muscle maintenance and function [31, 32].

The shift from SMSC proliferation to differentiation is characterized by the downregulation of Pax7 and sustained upregulation of MyoD [14]. MYOD activates MYOG and myogenic regulatory factor 4 (MRF4) to initiate myotube formation [14]. Ultimately, MYOD and MYOG cooperate to induce terminal differentiation markers, including Mrf4 and myosin heavy chain (MyHC), thereby driving the assembly of multinucleated myofibers [44, 45]. BMAL1 is a key regulator of this process, enhancing myogenic differentiation by directly upregulating Myf5 and MyoD; modulating Wnt pathway components such as Dishevelled 2, β-catenin, and transcription factor 3; and amplifying Wnt–MRF synergy [46, 47]. Non-ruminant animal models and empirical evidence in goats show that BMAL1-driven transcription of miR-27b suppresses PAX3, thereby inhibiting SMSC proliferation [48, 49]. CLOCK:BMAL1-activated MYOD binds to E-box elements in the Tcap promoter, reinforcing differentiation [50]. Although CLOCK:BMAL1 regulates miR-455, which targets Clock mRNA, the functional relevance of this feedback mechanism in myogenesis remains unclear [51]. PER1/2 supports myoblast differentiation by binding to the enhancer-promoter regions of insulin-like growth factor 2 (IGF-2), thereby increasing Igf2 expression and MyoG [38]. CRY2 facilitates differentiation and cell fusion by regulating cyclin D1 and Tmem176b, thereby preventing premature cell cycle exit and maintaining MyoG levels [52]. In contrast, REV-ERBα acts as a negative regulator; its deletion upregulates Myf5, MyoD, MyoG, and Wnt3a, accelerating myotube formation (Fig. 2d) [42]. Several non-coding RNAs influence the fate of SMSCs through specific molecular mechanisms. For example, lncMD1 competitively binds to miR-133a-3p and miR-361-3p, thereby relieving the repression of the target genes DCTN1/DCTN2, inhibiting proliferation, and promoting differentiation [53]. However, whether the circadian clock modulates SMSCs through non-coding RNA networks remains predominantly unknown. In summary, the core circadian clock negative feedback loop promotes skeletal muscle growth and development by directly or indirectly regulating myogenic differentiation and cell fusion, providing a molecular basis for enhanced meat production.

Heat stress considerably hinders global meat production, affecting approximately 80% of cattle annually, with > 80% of goats and nearly 60% of sheep reared in hot, arid zones [74, 75]. As a significant ZT, high temperatures can disrupt circadian rhythms, compromising meat production [76]. Mechanistically, heat stress activates heat shock factor 1 (HSF1), which binds to heat shock response elements near the E-box sequences in the Per2 promoter, potentially desynchronizing the central and peripheral clocks [77, 78]. This dysregulation may inhibit SMSC proliferation and differentiation via CCGs, consistent with the heat-induced downregulation of PAX7, MYOD, MYF5, MYOG, and MYHC in ovine myoblasts [79]. Heat stress causes direct clock interference and induces reactive oxygen species accumulation, which promotes BMAL1 phosphorylation via casein kinase 2 and disrupts the EZH2–CLOCK/BMAL1 interaction, collectively suppressing MYOD activity and myoblast differentiation [80–82]. Antioxidants, such as resveratrol and vitamin A, mitigate these effects and improve SMSC viability and production performance [83–86]. Cold stress may also impair SMSC function through circadian pathways, potentially via HSF1-PER2-mediated MyoD suppression or peroxisome proliferator activated receptor γ coactivator-1α (PGC-1α)-mediated Rev-erbα upregulation, which dampens MyoD rhythmicity [87–92]. Notably, heat/cold stress disrupts the circadian oscillation of body temperature in mammals, with lamb studies documenting fluctuations reaching 4 °C [93, 94]. In vitro studies employing square-wave temperature cycles to simulate circadian oscillations in body temperature have demonstrated that such thermal fluctuations can reset the circadian clock in myocytes [95]. However, whether extreme climatic conditions affect meat production in ruminants by modulating the circadian clock remains largely unknown. In summary, extreme climatic conditions may disrupt core body temperature and circadian clock oscillations, impair SMSC proliferation and differentiation, and reduce meat production in ruminants. These findings underscore the importance of precisely controlling the thermoneutral environment in ruminant husbandry.