1α,25-dihydroxyvitamin D3 stimulates human SOST gene expression and sclerostin secretion

Asiri R. Wijenayaka, Dongqing Yang, Matthew Prideaux, Nobuaki Ito, Masakazu Kogawa, Paul H. Anderson, Howard A. Morris, Lucian B. Solomon, Gabriela G. Loots, David M. Findlay, Gerald J. Atkins

Research output: Contribution to journalArticle

21 Citations (Scopus)

Abstract

Sclerostin, the SOST gene product, is a negative regulator of bone formation and a positive regulator of bone resorption. In this study, treatment of human primary osteoblasts, including cells differentiated to an osteocyte-like stage, with 1α,25-dihydroxyvitaminD3 (1,25D) resulted in the dose-dependent increased expression of SOST mRNA. A similar effect was observed in human trabecular bone samples cultured ex vivo, and in osteocyte-like cultures of differentiated SAOS2 cells. Treatment of SAOS2 cells with 1,25D resulted in the production and secretion of sclerostin protein. In silico analysis of the human SOST gene revealed a single putative DR3-type vitamin D response element (VDRE) at position -6216 bp upstream of the transcription start site (TSS). This sequence was confirmed to have strong VDRE activity by luciferase reporter assays and electrophoretic mobility shift analysis (EMSA). Sequence substitution in the VDR/RXR half-sites abolished VDRE reporter activity and binding of nuclear proteins. A 6.3 kb fragment of the human proximal SOST promoter demonstrated responsiveness to 1,25D. The addition of the evolutionary conserved region 5 (ECR5), a known bone specific enhancer region, ahead of the 6.3 kb fragment increased basal promoter activity but did not increase 1,25D responsiveness. Site-specific mutagenesis abolished the responsiveness of the 6.3 kb promoter to 1,25D. We conclude that 1,25D is a direct regulator of human SOST gene and sclerostin protein expression, extending the pathways of control of sclerostin expression. At least some of this responsiveness is mediated by the identified classical VDRE however the nature of the transcriptional regulation by 1,25D warrants further investigation.

Original languageEnglish (US)
Pages (from-to)157-167
Number of pages11
JournalMolecular and Cellular Endocrinology
Volume413
DOIs
StatePublished - Sep 5 2015
Externally publishedYes

Fingerprint

Vitamin D Response Element
Calcitriol
Gene expression
Bone
Gene Expression
Osteocytes
Genes
Electrophoretic mobility
Mutagenesis
Transcription Initiation Site
Osteoblasts
Electrophoretic Mobility Shift Assay
Bone Resorption
Nuclear Proteins
Site-Directed Mutagenesis
Luciferases
Osteogenesis
Computer Simulation
Assays
Carrier Proteins

Keywords

  • Gene regulation
  • Osteocyte
  • Promoter
  • Sclerostin
  • SOST
  • Vitamin D
  • Vitamin D response element

ASJC Scopus subject areas

  • Endocrinology
  • Molecular Biology
  • Biochemistry
  • Medicine(all)

Cite this

Wijenayaka, A. R., Yang, D., Prideaux, M., Ito, N., Kogawa, M., Anderson, P. H., ... Atkins, G. J. (2015). 1α,25-dihydroxyvitamin D3 stimulates human SOST gene expression and sclerostin secretion. Molecular and Cellular Endocrinology, 413, 157-167. https://doi.org/10.1016/j.mce.2015.06.021

1α,25-dihydroxyvitamin D3 stimulates human SOST gene expression and sclerostin secretion. / Wijenayaka, Asiri R.; Yang, Dongqing; Prideaux, Matthew; Ito, Nobuaki; Kogawa, Masakazu; Anderson, Paul H.; Morris, Howard A.; Solomon, Lucian B.; Loots, Gabriela G.; Findlay, David M.; Atkins, Gerald J.

In: Molecular and Cellular Endocrinology, Vol. 413, 05.09.2015, p. 157-167.

Research output: Contribution to journalArticle

Wijenayaka, AR, Yang, D, Prideaux, M, Ito, N, Kogawa, M, Anderson, PH, Morris, HA, Solomon, LB, Loots, GG, Findlay, DM & Atkins, GJ 2015, '1α,25-dihydroxyvitamin D3 stimulates human SOST gene expression and sclerostin secretion', Molecular and Cellular Endocrinology, vol. 413, pp. 157-167. https://doi.org/10.1016/j.mce.2015.06.021
Wijenayaka, Asiri R. ; Yang, Dongqing ; Prideaux, Matthew ; Ito, Nobuaki ; Kogawa, Masakazu ; Anderson, Paul H. ; Morris, Howard A. ; Solomon, Lucian B. ; Loots, Gabriela G. ; Findlay, David M. ; Atkins, Gerald J. / 1α,25-dihydroxyvitamin D3 stimulates human SOST gene expression and sclerostin secretion. In: Molecular and Cellular Endocrinology. 2015 ; Vol. 413. pp. 157-167.
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abstract = "Sclerostin, the SOST gene product, is a negative regulator of bone formation and a positive regulator of bone resorption. In this study, treatment of human primary osteoblasts, including cells differentiated to an osteocyte-like stage, with 1α,25-dihydroxyvitaminD3 (1,25D) resulted in the dose-dependent increased expression of SOST mRNA. A similar effect was observed in human trabecular bone samples cultured ex vivo, and in osteocyte-like cultures of differentiated SAOS2 cells. Treatment of SAOS2 cells with 1,25D resulted in the production and secretion of sclerostin protein. In silico analysis of the human SOST gene revealed a single putative DR3-type vitamin D response element (VDRE) at position -6216 bp upstream of the transcription start site (TSS). This sequence was confirmed to have strong VDRE activity by luciferase reporter assays and electrophoretic mobility shift analysis (EMSA). Sequence substitution in the VDR/RXR half-sites abolished VDRE reporter activity and binding of nuclear proteins. A 6.3 kb fragment of the human proximal SOST promoter demonstrated responsiveness to 1,25D. The addition of the evolutionary conserved region 5 (ECR5), a known bone specific enhancer region, ahead of the 6.3 kb fragment increased basal promoter activity but did not increase 1,25D responsiveness. Site-specific mutagenesis abolished the responsiveness of the 6.3 kb promoter to 1,25D. We conclude that 1,25D is a direct regulator of human SOST gene and sclerostin protein expression, extending the pathways of control of sclerostin expression. At least some of this responsiveness is mediated by the identified classical VDRE however the nature of the transcriptional regulation by 1,25D warrants further investigation.",
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AU - Ito, Nobuaki

AU - Kogawa, Masakazu

AU - Anderson, Paul H.

AU - Morris, Howard A.

AU - Solomon, Lucian B.

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