In eukaryotic cells, degradation of most intracellular proteins is carried out 
by the ubiquitin-proteasome pathway. Recent investigations suggest that bone 
metabolism is also regulated by this pathway. The clinical efficacy of 
bortezomib, a 26S proteasome inhibitor used as an anticancer drug, has been 
linked to an increase in bone formation. In this study, we show that proteasome 
inhibitors induce expression of osteoblastic differentiation-related genes such 
as osteocalcin and alkaline phosphatase in C2C12 cells. In contrast, myogenic 
differentiation is inhibited. Among the proteasome inhibitors tested, bortezomib 
induced the greatest increase in osteocalcin expression. Although these effects 
were similar to that of bone morphogenetic protein (BMP) 2, proteasome 
inhibitors did not induce transcriptional activity of Smad1/4-dependent reporter 
or BMP2 signaling target gene expression. Transient transfection of osteocalcin 
promoter-luciferase constructs with bortezomib resulted in an increase in 
luciferase activity. Mutation of OSE2, but not OSE1, sites of the osteocalcin 
promoter diminished the bortezomib-induced activity. Also, Runx2 binding 
activity and protein levels were induced by bortezomib treatment. These results 
suggest that the bortezomib induces osteoblastic differentiation by modifying 
the activity of Runx2 and that the function of the proteasome in controlling 
degradation of differentiation-related transcription factors plays an important 
role in osteoblast differentiation.