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 induction, we examined the effect of osteogenic protein (OP)-1/bone morphogenetic 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 analysis localizes the OP-1 response region to 33 bp (-310/-278), which confers OP-1 responsiveness to both the minimal homologous and heterologous Rous sarcoma virus promoter. Transforming growth factor-beta2 or activin, which up-regulates the expression of a transforming growth factor-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 enhancer factor (MEF)-2 like sequence (-310/-298, TTAAAAATAAAAA) in the 33-bp region are necessary for the OP-1 effect. Gel shift assays show interaction of distinct nuclear proteins from C5.18 cells with the AP-1-like and the MEF-2-like sequences. OP-1 rapidly induces nuclear protein 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 from the MEF-2 family proteins present in C2C12 myoblasts, in which OP-1 does not induce collagen X mRNA or up-regulate its promoter activity. In conclusion, we identified a specific response region for OP-1 in the 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-regulates collagen X promoter activity, which also requires an AP-1 like sequence.
Harada
S
S
Department of Bone Biology and Osteoporosis Research, Merck Research Laboratories, West Point, Pennsylvania 19486, USA.
Sampath
T K
TK
Aubin
J E
JE
Rodan
G A
GA
D028341:Activins; D000818:Animals; D055419:Bone Morphogenetic Protein 7; D019485:Bone Morphogenetic Proteins / Q000235:genetics / Q000502:physiology*; D002460:Cell Line; D003094:Collagen / Q000187:drug effects / Q000235:genetics*; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000502:physiology; D005333:Fetus; D007265:Inhibins / Q000494:pharmacology; D064326:MEF2 Transcription Factors; D051379:Mice; D018006:Myogenic Regulatory Factors; D009687:Nuclear Proteins / Q000187:drug effects / Q000235:genetics; D011401:Promoter Regions, Genetic / Q000187:drug effects; D011485:Protein Binding / Q000235:genetics; D051381:Rats; D012886:Skull / Q000166:cytology; D018808:Transcription Factor AP-1 / Q000235:genetics*; D014157:Transcription Factors / Q000235:genetics* / Q000502:physiology; D014162:Transfection; D016212:Transforming Growth Factor beta / Q000502:physiology*
C525821:Bmp7 protein, rat; D055419:Bone Morphogenetic Protein 7; D019485:Bone Morphogenetic Proteins; D004268:DNA-Binding Proteins; D064326:MEF2 Transcription Factors; D018006:Myogenic Regulatory Factors; D009687:Nuclear Proteins; D018808:Transcription Factor AP-1; D014157:Transcription Factors; D016212:Transforming Growth Factor beta; D028341:Activins; D007265:Inhibins; D003094:Collagen