TY - JOUR
T1 - Concerted action of Smad and CREB-binding protein regulates bone morphogenetic protein-2-stimulated osteoblastic colony-stimulating factor-1 expression
AU - Ghosh-Choudhury, Nandini
AU - Singha, Prajjal K.
AU - Woodruff, Kathleen
AU - St Clair, Patricia
AU - Bsoul, Sameer
AU - Werner, Sherry L.
AU - Choudhury, Goutam Ghosh
N1 - Copyright:
Copyright 2008 Elsevier B.V., All rights reserved.
PY - 2006/7/21
Y1 - 2006/7/21
N2 - Bone remodeling depends upon proper osteoblast and osteoclast function. Bone morphogenetic protein-2 (BMP-2) stimulates differentiation of osteoblasts from pluripotent precursors. Osteoclast formation depends on the concerted action of osteoblast-derived receptor activator of NF-κB ligand and colony-stimulating factor-1 (CSF-1). BMP-2 stimulates receptor activator of NF-κB ligand expression. However, the effect of BMP-2 on CSF-1 expression has not been studied. We investigated the role of BMP-2 in CSF-1 expression in osteogenic C2C12 cells. Incubation of C2C12 cells with BMP-2 supported osteoclastogenesis of spleen cells with a concomitant increase in expression of CSF-1 mRNA and protein. To determine the mechanism, we identified a BMP-responsive element between -627 bp and -509 bp in the CSF-1 promoter. DNase I footprint analysis revealed the presence of consensus Smad binding motif in this region. Electrophoretic mobility shift assay showed BMP-2-stimulated binding of proteins to this motif. Mutation of core sequence as well as its 5′- and 3′-flanking sequences abolished the DNA-protein interaction resulting in inhibition of CSF-1 transcription. Supershift analysis detects the presence of Smads 1, 5, and 4 and the transcriptional coactivator CREB-binding protein in the BMP-responsive element-protein complex. In addition, Smads 1 and 5 alone or in combination with Smad 4 increased CSF-1 transcription. Furthermore, CREB-binding protein markedly increased transcription of CSF-1. These data represent the first evidence that BMP-2 increases the osteoclastogenic CSF-1 expression by a transcriptional mechanism using the canonical Smad pathway and provide a mechanism for BMP-2-induced osteoclast differentiation.
AB - Bone remodeling depends upon proper osteoblast and osteoclast function. Bone morphogenetic protein-2 (BMP-2) stimulates differentiation of osteoblasts from pluripotent precursors. Osteoclast formation depends on the concerted action of osteoblast-derived receptor activator of NF-κB ligand and colony-stimulating factor-1 (CSF-1). BMP-2 stimulates receptor activator of NF-κB ligand expression. However, the effect of BMP-2 on CSF-1 expression has not been studied. We investigated the role of BMP-2 in CSF-1 expression in osteogenic C2C12 cells. Incubation of C2C12 cells with BMP-2 supported osteoclastogenesis of spleen cells with a concomitant increase in expression of CSF-1 mRNA and protein. To determine the mechanism, we identified a BMP-responsive element between -627 bp and -509 bp in the CSF-1 promoter. DNase I footprint analysis revealed the presence of consensus Smad binding motif in this region. Electrophoretic mobility shift assay showed BMP-2-stimulated binding of proteins to this motif. Mutation of core sequence as well as its 5′- and 3′-flanking sequences abolished the DNA-protein interaction resulting in inhibition of CSF-1 transcription. Supershift analysis detects the presence of Smads 1, 5, and 4 and the transcriptional coactivator CREB-binding protein in the BMP-responsive element-protein complex. In addition, Smads 1 and 5 alone or in combination with Smad 4 increased CSF-1 transcription. Furthermore, CREB-binding protein markedly increased transcription of CSF-1. These data represent the first evidence that BMP-2 increases the osteoclastogenic CSF-1 expression by a transcriptional mechanism using the canonical Smad pathway and provide a mechanism for BMP-2-induced osteoclast differentiation.
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U2 - 10.1074/jbc.M511071200
DO - 10.1074/jbc.M511071200
M3 - Article
C2 - 16707491
AN - SCOPUS:33745975802
VL - 281
SP - 20160
EP - 20170
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
SN - 0021-9258
IS - 29
ER -