Concerted action of Smad and CREB-binding protein regulates bone morphogenetic protein-2-stimulated osteoblastic colony-stimulating factor-1 expression

Nandini Ghosh-Choudhury, Prajjal K. Singha, Kathleen Woodruff, Patricia St Clair, Sameer Bsoul, Sherry L. Werner, Goutam Ghosh Choudhury

Research output: Contribution to journalArticlepeer-review

39 Scopus citations

Abstract

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.

Original languageEnglish (US)
Pages (from-to)20160-20170
Number of pages11
JournalJournal of Biological Chemistry
Volume281
Issue number29
DOIs
StatePublished - Jul 21 2006

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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