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Department of Biochemistry, University of Texas Health Science Center, San Antonio, Texas 78229-3900
Address all correspondence and requests for reprints to: Lee-Chuan C. Yeh, Ph.D., Department of Biochemistry (Mail Code 7760), University of Texas Health Science Center, 7703 Floyd Curl Drive, San Antonio, Texas 78229-3900. E-mail: carolyeh{at}biochem.uthscsa.edu
Osteogenic protein-1 (OP-1), a member of the bone morphogenetic protein
subfamily of the transforming growth factor-ß 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/EBP
)-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/EBP-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).
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