Bone-specific transcription factor Runx2 interacts with the 1α,25-dihydroxyvitamin D3 receptor to up-regulate rat osteocalcin gene expression in osteoblastic cells

Roberto Paredes, Gloria Arriagada, Fernando Cruzat, Alejandro Villagra, Juan Olate, Kaleem Zaidi, Andre J van Wijnen, Jane B. Lian, Gary S. Stein, Janet L. Stein, Martin Montecino

Research output: Contribution to journalArticle

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Abstract

Bone-specific transcription of the osteocalcin (OC) gene is regulated principally by the Runx2 transcription factor and is further stimulated in response to 1α,25-dihydroxyvitamin D3 via its specific receptor (VDR). The rat OC gene promoter contains three recognition sites for Runx2 (sites A, B, and C). Mutation of sites A and B, which flank the 1α,25-dihydroxyvitamin D3-responsive element (VDRE), abolishes 1α,25-dihydroxyvitamin D3-dependent enhancement of OC transcription, indicating a tight functional relationship between the VDR and Runx2 factors. In contrast to most of the members of the nuclear receptor family, VDR possesses a very short N-terminal A/B domain, which has led to the suggestion that its N-terminal region does not contribute to transcriptional enhancement. Here, we have combined transient-overexpression, coimmunoprecipitation, in situ colocalization, chromatin immunoprecipitation, and glutathione S-transferase pull-down analyses to demonstrate that in osteoblastic cells expressing OC, VDR interacts directly with Runx2 bound to site B, which is located immediately adjacent to the VDRE. This interaction contributes significantly to 1α,25-dihydroxyvitamin D3- dependent enhancement of the OC promoter and requires a region located C terminal to the runt homology DNA binding domain of Runx2 and the N-terminal region of VDR. Together, our results indicate that Runx2 plays a key role in the 1α,25-dihydroxyvitamin D3-dependent stimulation of the OC promoter in osteoblastic cells by further stabilizing the interaction of the VDR with the VDRE. These studies demonstrate a novel mechanism for combinatorial control of bone tissue-specific gene expression. This mechanism involves the intersection of two major pathways: Runx2, a "master" transcriptional regulator of osteoblast differentiation, and 1α,25-dihydroxyvitamin D 3, a hormone that promotes expression of genes associated with these terminally differentiated bone cells.

Original languageEnglish (US)
Pages (from-to)8847-8861
Number of pages15
JournalMolecular and Cellular Biology
Volume24
Issue number20
DOIs
StatePublished - Oct 2004
Externally publishedYes

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Calcitriol Receptors
Osteocalcin
Calcitriol
Transcription Factors
Up-Regulation
Gene Expression
Bone and Bones
Chromatin Immunoprecipitation
Cytoplasmic and Nuclear Receptors
Glutathione Transferase
Osteoblasts
Nuclear Family
Genes
Hormones
Mutation
DNA

ASJC Scopus subject areas

  • Molecular Biology
  • Genetics
  • Cell Biology

Cite this

Bone-specific transcription factor Runx2 interacts with the 1α,25-dihydroxyvitamin D3 receptor to up-regulate rat osteocalcin gene expression in osteoblastic cells. / Paredes, Roberto; Arriagada, Gloria; Cruzat, Fernando; Villagra, Alejandro; Olate, Juan; Zaidi, Kaleem; van Wijnen, Andre J; Lian, Jane B.; Stein, Gary S.; Stein, Janet L.; Montecino, Martin.

In: Molecular and Cellular Biology, Vol. 24, No. 20, 10.2004, p. 8847-8861.

Research output: Contribution to journalArticle

Paredes, R, Arriagada, G, Cruzat, F, Villagra, A, Olate, J, Zaidi, K, van Wijnen, AJ, Lian, JB, Stein, GS, Stein, JL & Montecino, M 2004, 'Bone-specific transcription factor Runx2 interacts with the 1α,25-dihydroxyvitamin D3 receptor to up-regulate rat osteocalcin gene expression in osteoblastic cells', Molecular and Cellular Biology, vol. 24, no. 20, pp. 8847-8861. https://doi.org/10.1128/MCB.24.20.8847-8861.2004
Paredes, Roberto ; Arriagada, Gloria ; Cruzat, Fernando ; Villagra, Alejandro ; Olate, Juan ; Zaidi, Kaleem ; van Wijnen, Andre J ; Lian, Jane B. ; Stein, Gary S. ; Stein, Janet L. ; Montecino, Martin. / Bone-specific transcription factor Runx2 interacts with the 1α,25-dihydroxyvitamin D3 receptor to up-regulate rat osteocalcin gene expression in osteoblastic cells. In: Molecular and Cellular Biology. 2004 ; Vol. 24, No. 20. pp. 8847-8861.
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abstract = "Bone-specific transcription of the osteocalcin (OC) gene is regulated principally by the Runx2 transcription factor and is further stimulated in response to 1α,25-dihydroxyvitamin D3 via its specific receptor (VDR). The rat OC gene promoter contains three recognition sites for Runx2 (sites A, B, and C). Mutation of sites A and B, which flank the 1α,25-dihydroxyvitamin D3-responsive element (VDRE), abolishes 1α,25-dihydroxyvitamin D3-dependent enhancement of OC transcription, indicating a tight functional relationship between the VDR and Runx2 factors. In contrast to most of the members of the nuclear receptor family, VDR possesses a very short N-terminal A/B domain, which has led to the suggestion that its N-terminal region does not contribute to transcriptional enhancement. Here, we have combined transient-overexpression, coimmunoprecipitation, in situ colocalization, chromatin immunoprecipitation, and glutathione S-transferase pull-down analyses to demonstrate that in osteoblastic cells expressing OC, VDR interacts directly with Runx2 bound to site B, which is located immediately adjacent to the VDRE. This interaction contributes significantly to 1α,25-dihydroxyvitamin D3- dependent enhancement of the OC promoter and requires a region located C terminal to the runt homology DNA binding domain of Runx2 and the N-terminal region of VDR. Together, our results indicate that Runx2 plays a key role in the 1α,25-dihydroxyvitamin D3-dependent stimulation of the OC promoter in osteoblastic cells by further stabilizing the interaction of the VDR with the VDRE. These studies demonstrate a novel mechanism for combinatorial control of bone tissue-specific gene expression. This mechanism involves the intersection of two major pathways: Runx2, a {"}master{"} transcriptional regulator of osteoblast differentiation, and 1α,25-dihydroxyvitamin D 3, a hormone that promotes expression of genes associated with these terminally differentiated bone cells.",
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AU - Villagra, Alejandro

AU - Olate, Juan

AU - Zaidi, Kaleem

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