Transforming growth factor-β1 responsiveness of the rat osteocalcin gene is mediated by an activator protein-1 binding site

Chaitali Banerjee, Janet L. Stein, Andre J van Wijnen, Baruch Frenkel, Jane B. Lian, Gary S. Stein

Research output: Contribution to journalArticle

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Abstract

Osteocalcin (OC), a bone specific protein expressed during differentiation and mineralization of the bone extracellular matrix, is down-regulated upon treatment with transforming growth factor (TGF)-β1. To address the potential role of OC gene expression in relation to TGF-β1 regulation of bone formation and resorption, we examined the transcriptional activity of the rat OC promoter after TGF-β1 treatment. 5' deletion analysis of rat OC promoter- chloramphenicol acetyltransferase constructs demonstrated that TGF-β1 treatment repressed chloramphenicol acetyltransferase activity by 2.4-fold in transient transfections of ROS 17/2.8 cells. A 29-bp region between -162 and -134 identified as the TGF-β1 response domain, conferred TGF-β1 responsiveness to the -108 to +24 rat OC basal promoter in an orientation dependent manner. Mutation of an activator protein-1/cAMP-response element- like motif (-146 to -139) abolished TGF-β1 responsiveness of the construct. In vitro gel-mobility shift and competition assays using wild-type and mutated oligonucleotides and antibodies indicate that Fra-2, a Fos related transcription factor, binds to this motif. We show that Fra-2 is an activator of the OC promoter, and TGF-β1 inhibits this activation. Our results demonstrate that Fra-2 is hyperphosphorylated upon TGF-β1 treatment of ROS 17/2.8 cells. Additionally, treatment of cells with a staurosporine protein kinase C inhibitor abrogates TGF-β1 mediated down-regulation of the OC promoter activity. Together, these results demonstrate that TGF-β1 responsiveness of the rat osteocalcin gene in ROS 17/2.8 cells is mediated through an activator protein-1 like cis-acting element that interacts with Fra-2. Furthermore, our results are consistent with a critical role for TGF- β1 induced phosphorylation of Fra-2 in the repression of OC gene transcription.

Original languageEnglish (US)
Pages (from-to)1991-2000
Number of pages10
JournalEndocrinology
Volume137
Issue number5
DOIs
StatePublished - 1996
Externally publishedYes

Fingerprint

Osteocalcin
Transcription Factor AP-1
Transforming Growth Factors
Protein Binding
Binding Sites
Genes
Chloramphenicol O-Acetyltransferase
Bone Matrix
Staurosporine
Protein C Inhibitor
Response Elements
Electrophoretic Mobility Shift Assay
Protein Kinase Inhibitors
Bone Resorption
Osteogenesis
Oligonucleotides
Protein Kinase C
Extracellular Matrix
Transfection
Transcription Factors

ASJC Scopus subject areas

  • Endocrinology
  • Endocrinology, Diabetes and Metabolism

Cite this

Banerjee, C., Stein, J. L., van Wijnen, A. J., Frenkel, B., Lian, J. B., & Stein, G. S. (1996). Transforming growth factor-β1 responsiveness of the rat osteocalcin gene is mediated by an activator protein-1 binding site. Endocrinology, 137(5), 1991-2000. https://doi.org/10.1210/en.137.5.1991

Transforming growth factor-β1 responsiveness of the rat osteocalcin gene is mediated by an activator protein-1 binding site. / Banerjee, Chaitali; Stein, Janet L.; van Wijnen, Andre J; Frenkel, Baruch; Lian, Jane B.; Stein, Gary S.

In: Endocrinology, Vol. 137, No. 5, 1996, p. 1991-2000.

Research output: Contribution to journalArticle

Banerjee, C, Stein, JL, van Wijnen, AJ, Frenkel, B, Lian, JB & Stein, GS 1996, 'Transforming growth factor-β1 responsiveness of the rat osteocalcin gene is mediated by an activator protein-1 binding site', Endocrinology, vol. 137, no. 5, pp. 1991-2000. https://doi.org/10.1210/en.137.5.1991
Banerjee, Chaitali ; Stein, Janet L. ; van Wijnen, Andre J ; Frenkel, Baruch ; Lian, Jane B. ; Stein, Gary S. / Transforming growth factor-β1 responsiveness of the rat osteocalcin gene is mediated by an activator protein-1 binding site. In: Endocrinology. 1996 ; Vol. 137, No. 5. pp. 1991-2000.
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