• Extracellular matrix stiffness modulates bone marrow stromal stem cell proliferation via a novel YAP/primary cilia mechanism.
• Rigid substrates enhance proliferation and activate Wnt/β-catenin signaling, while inducing primary cilia shortening and YAP nuclear accumulation.
• YAP knockdown abolishes stiffness-dependent differences in proliferation and cilia length, confirming its central role.
• These findings provide insights for designing tissue engineering scaffolds with optimal mechanical properties.