Identification of an osteogenic protein-1 (bone morphogenetic protein-7)-responsive element in the promoter of the rat insulin-like growth factor-binding protein-5 gene.
Osteogenic protein-1 (OP-1), a member of the bone morphogenetic protein subfamily of the transforming growth factor-beta superfamily, induces new bone formation in vivo and regulates the expression of numerous growth factors. We previously showed that OP-1 down-regulates the transcription of the insulin-like growth factor-binding protein-5 (IGFBP-5) in primary cultures of fetal rat calvaria (FRC) cells. In the present study we identified, within the IGFBP-5 promoter, a 21-bp region that confers OP-1 responsiveness in FRC cells. Within this region lie three putative cis-acting regulatory elements, viz. a CAAT-like sequence, a CCAAT/enhancer-binding protein (C/EBPalpha)-like element, and a c-Myb or E-box-like motif. Mutations in the CAAT-like sequence reduced the promoter activity in both control and OP-1-treated cells, but did not abrogate the OP-1-induced down-regulation. Mutations in the C/EBPalpha-like element reduced the promoter activity in both control and OP-1-treated cells without significantly affecting the extent of down-regulation. Mutations in the putative c-Myb or E-box-like motif reduced the promoter activity in both the OP-1-treated and control cells and completely abolished the inhibitory effect of OP-1 on the IGFBP-5 promoter activity. Gel mobility shift analyses further showed specific interaction between nuclear protein(s) in FRC cells and the 21-bp region. OP-1 down-regulates the nuclear regulatory protein interaction with the 21-bp region by reducing either the cellular concentration of the regulatory protein(s) or the affinity of the regulatory protein(s) for the OP-1 responsive element. In conclusion, we identified an OP-1 response region in the rat IGFBP-5 promoter and further showed that OP-1 down-regulates the nuclear protein interaction with the response element(s).
Yeh
L C
LC
Department of Biochemistry, University of Texas Health Science Center, San Antonio 78229-3900, USA. carolyeh@biochem.uthscsa.edu
Lee
J C
JC
D000818:Animals; D001483:Base Sequence; D055419:Bone Morphogenetic Protein 7; D019485:Bone Morphogenetic Proteins / Q000502:physiology*; D002478:Cells, Cultured; D005260:Female; D006801:Humans; D018975:Insulin-Like Growth Factor Binding Protein 5 / Q000235:genetics*; D008156:Luciferases / Q000235:genetics; D051379:Mice; D009154:Mutation / Q000235:genetics; D011247:Pregnancy; D011401:Promoter Regions, Genetic / Q000235:genetics*; D051381:Rats; D017207:Rats, Sprague-Dawley; D020218:Response Elements / Q000235:genetics*; D014162:Transfection; D016212:Transforming Growth Factor beta
C525819:BMP7 protein, human; C525821:Bmp7 protein, rat; D055419:Bone Morphogenetic Protein 7; D019485:Bone Morphogenetic Proteins; D018975:Insulin-Like Growth Factor Binding Protein 5; D016212:Transforming Growth Factor beta; D008156:Luciferases