Disruption of actin cytoskeleton with cytochalasin D has been known to induce chondrogenic differentiation of chick embryo Limb bud mesenchymal cells. However, the mechanism(s) for the induction of chondrogenesis by cytochalasin D is not yet-clearly known, In the present study, we examined possible involvement of protein kinase C (PKC) and extracellular signal-regulated protein kinase (Erk-1) in chondrogenesis of mesenchymal cells induced by disruption of actin cytoskeleton. Disruption of actin cytoskeleton with cytochalasin D or latrunculin B induced chondrogenesis of mesenchymal cells cultured at subconfluent cell density, as determined by type II collagen expression. Among the expressed PKC isoforms, cytochalasin D dramatically increased expression and activation of PKC alpha in a dose-dependent manner, and inhibition or downregulation of PKC alpha blocked cytochalasin D-induced chondrogenesis. Cytochalasin D also downregulated Erk-1 phosphorylation that is associated with chondrogenesis. Our results, therefore, suggest that disruption of actin cytoskeleton induces chondrogenesis of mesenchymal cells by activating PKC alpha and by inhibiting Erk-1 signaling.