OSTEOGENIC PROTEIN-1 UP-REGULATION OF THE COLLAGEN X PROMOTER ACTIVITY IS MEDIATED BY A MEF-2-LIKE SEQUENCE AND REQUIRES AN ADJACENT AP-1 SEQUENCE

Bone morphogenetic proteins induce chondrogenesis and osteogenesis in vivo. To investigate molecular mechanisms involved in chondrocyte indu ction, we examined the effect of osteogenic protein (OP)-1/bone morpho genetic protein-7 on the collagen X promoter. In rat calvaria-derived chondrogenic C5.18 cells, OP-1 up-regulates collagen X mRNA levels and its promoter activity in a cell type-specific manner. Deletion analys is localizes the OP-1 response region to 33 bp (-310/-278), which conf ers OP-1 responsiveness to both the minimal homologous and heterologou s Rous sarcoma virus promoter. Transforming growth factor-beta 2 or ac tivin, which up-regulates the expression of a transforming growth fact or-beta-inducible p3TP-Lux construct, has little effect on collagen X mRNA and on this 33-bp region. Mutational analysis shows that both an AP-1 like sequence (-294/-285, TGAATCATCA) and an A/T-rich myocyte enh ancer factor (MEF)-2 like sequence (-310/-298, TTAAAAATAAAAA) in the 3 3-bp region are necessary for the OR-1 effect, Gel shift assays show i nteraction of distinct nuclear proteins from 05.18 cells with the AP-1 -like and the MEF-2-like sequences. OR-1 rapidly induces nuclear prote in interaction with the MEF-2-like sequence but not with the AP-1 like sequence. MEF-2-like binding activity induced by OP-1 is distinct fro m the MEF-2 family proteins present in C2C12 myoblasts, in which OR-1 does not induce collagen X mRNA or up-regulate its promoter activity. In conclusion, we identified a specific response region for OP-l in th e mouse collagen X promoter. Mutational and gel shift analyses suggest that OP-1 induces nuclear protein interaction with an A/T-rich MEF-2 like sequence, distinct from the MEF-2 present in myoblasts, and up-re gulates collagen X promoter activity, which also requires an AP-1 like sequence.