Shaking culture attenuates circadian rhythms in induced pluripotent stem cells during osteogenic differentiation through the YAP-TEAD axis

Circadian rhythms synchronize cellular activities with the Earth's 24-hour cycle, primarily regulated by clock genes that influence metabolism, physiology, and osteogenesis. While genetic regulation of circadian rhythms is well characterized, how extracellular mechanical forces modulate these rhythms during iPSC osteogenic differentiation remains unclear. Here, we show that shaking culture, which promotes three-dimensional iPSC-EB formation, disrupts clock gene oscillations by activating YAP-TEAD signaling. Inhibition of YAP-TEAD with verteporfin restores circadian oscillations and enhances osteogenesis, highlighting a biomechanical-circadian interplay with implications for regenerative medicine. Overall design: Total RNA was extracted from adherent and shaking cultures two days after osteogenic induction, with two biological replicates per group. To analyze transcriptome profiles, RNA sequencing reads were mapped to the mm10 reference genome.