Disruption of circadian rhythms is related to disorders of glucose metabolism, and the molecular clock also exists in skeletal muscle. The ChIP-derived repressor of network oscillator (Chrono) and brain and muscle ARNT-like 1 (Bmal1) are core circadian components. Chrono is considered to be the repressor of Bmal1, and the Chrono–Bmal1 pathway is important in regulating the circadian rhythm; it has been speculated that this pathway could be a new mechanism for regulating glucose metabolism. The purpose of this study was to investigate the effects of Chrono on glucose metabolism in skeletal muscle and exercise capacity by using mice with skeletal-muscle-specific overexpression of Chrono (Chrono TG) and wild-type (WT) mice as the animal models. The results of this cross-sectional study indicated that the Chrono TG mice had an impaired glucose tolerance, lower exercise capacity, and higher levels of nonfasted blood glucose and glycogen content in skeletal muscle compared to WT mice. In addition, the Chrono TG mice also showed a significant increase in the amount of Chrono bound to Bmal1 according to a co-IP analysis; a remarkable decrease in mRNA expression of Tbc1d1, Glut4, Hk2, Pfkm, Pdp1, Gbe1, and Phka1, as well as in activity of Hk and protein expression of Ldhb; but higher mRNA expression of Pdk4 and protein expression of Ldha compared with those of WT mice. These data suggested the skeletal-muscle-specific overexpression of Chrono led to a greater amount of Chrono bound to Bmal1, which then could affect the glucose transporter, glucose oxidation, and glycogen utilization in skeletal muscle, as well as exercise capacity.

Brown adipose tissue (BAT) is a critical target for obesity treatment, and exercise can enhance BAT function through the activation of ADRβ3. However, the molecular mechanisms underlying BAT metabolism following the exercise-induced activation of ADRβ3 remain unclear. This study utilized RNA sequencing, Western blotting, Oil Red O staining, weighted gene co-expression network analysis (WGCNA), and machine learning to investigate the role of the ADRβ3-COX2 pathway in lipid metabolism in brown adipocytes. We identified Ywhah as a key gene and validated our findings using external datasets. Our results demonstrate that exercise significantly enhances brown adipose tissue metabolism in mice, with ADRβ3 activation promoting metabolic activity in brown adipocytes. In contrast, COX2 inhibition notably reduced the lipolytic effect and thermogenic gene expression induced by ADRβ3 activation. WGCNA and machine learning identified Ywhah as the most important feature variable in the downstream signaling of the ADRβ3-COX2 pathway. External microarray data further confirmed that 8 weeks of aerobic exercise significantly upregulated Ywhah expression. Additionally, Ywhah displayed strong binding affinity to cytoskeletal proteins in affinity purification–mass spectrometry experiments, and its expression was highly correlated with cytoskeletal GSVA scores. In summary, this study reveals the potential role of the ADRβ3-COX2-Ywhah-cytoskeleton axis in regulating brown adipocyte metabolism, providing new insights into obesity treatment mechanisms.