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. 1993 Feb 15;90(4):1503–1507. doi: 10.1073/pnas.90.4.1503

Postproliferative transcription of the rat osteocalcin gene is reflected by vitamin D-responsive developmental modifications in protein-DNA interactions at basal and enhancer promoter elements.

T A Owen 1, R Bortell 1, V Shalhoub 1, A Heinrichs 1, J L Stein 1, G S Stein 1, J B Lian 1
PMCID: PMC45902  PMID: 8381969

Abstract

In the osteocalcin (OC) gene promoter, both independent positive and negative regulatory elements, as well as others with contiguous [TATA/glucocorticoid-responsive elements (GRE)] or overlapping [TATA/GRE, vitamin D-responsive enhancer elements (VDRE)/AP-1, and OC box/AP-1] domains, are sites for modifications in protein-DNA interactions. In the present studies, we have examined nuclear protein extracts from fetal rat calvarial cells that undergo a developmental sequence of bone cell differentiation. Our results demonstrate modifications in protein-DNA interactions that relate to the developmental stages of the osteoblast and support developmental regulation of OC gene transcription. Basal expression of the OC gene is associated with sequence-specific protein-DNA interactions at the OC box, VDRE, and TATA/GRE box. Distinct differences are observed in proliferating osteoblasts, where the OC gene is not transcribed compared to postproliferative, differentiated osteoblasts that transcribe the OC gene. Furthermore, the protein-DNA complexes that reflect hormonal control are also developmentally regulated, mediating both the transcriptionally active and repressed states of the OC gene. For example, in proliferating osteoblasts, a vitamin D receptor-antibody-sensitive complex is formed that is different from the DNA binding complex induced by vitamin D postproliferatively when the OC gene is transcribed. Mutational analysis of the steroid hormone binding domain and the overlapping AP-1 site at the VDRE supports mutually exclusive occupancy by Fos-Jun heterodimers and vitamin D receptor. Such protein-DNA interactions at the VDRE are consistent with repression of competency for vitamin D-mediated transcriptional enhancement in proliferating osteoblasts expressing high levels of Fos and Jun.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Aronow M. A., Gerstenfeld L. C., Owen T. A., Tassinari M. S., Stein G. S., Lian J. B. Factors that promote progressive development of the osteoblast phenotype in cultured fetal rat calvaria cells. J Cell Physiol. 1990 May;143(2):213–221. doi: 10.1002/jcp.1041430203. [DOI] [PubMed] [Google Scholar]
  2. Baumbach L. L., Marashi F., Plumb M., Stein G., Stein J. Inhibition of DNA replication coordinately reduces cellular levels of core and H1 histone mRNAs: requirement for protein synthesis. Biochemistry. 1984 Apr 10;23(8):1618–1625. doi: 10.1021/bi00303a006. [DOI] [PubMed] [Google Scholar]
  3. Baumbach L. L., Stein G. S., Stein J. L. Regulation of human histone gene expression: transcriptional and posttranscriptional control in the coupling of histone messenger RNA stability with DNA replication. Biochemistry. 1987 Sep 22;26(19):6178–6187. doi: 10.1021/bi00393a034. [DOI] [PubMed] [Google Scholar]
  4. Bellows C. G., Aubin J. E., Heersche J. N., Antosz M. E. Mineralized bone nodules formed in vitro from enzymatically released rat calvaria cell populations. Calcif Tissue Int. 1986 Mar;38(3):143–154. doi: 10.1007/BF02556874. [DOI] [PubMed] [Google Scholar]
  5. Bhargava U., Bar-Lev M., Bellows C. G., Aubin J. E. Ultrastructural analysis of bone nodules formed in vitro by isolated fetal rat calvaria cells. Bone. 1988;9(3):155–163. doi: 10.1016/8756-3282(88)90005-1. [DOI] [PubMed] [Google Scholar]
  6. Bortell R., Owen T. A., Bidwell J. P., Gavazzo P., Breen E., van Wijnen A. J., DeLuca H. F., Stein J. L., Lian J. B., Stein G. S. Vitamin D-responsive protein-DNA interactions at multiple promoter regulatory elements that contribute to the level of rat osteocalcin gene expression. Proc Natl Acad Sci U S A. 1992 Jul 1;89(13):6119–6123. doi: 10.1073/pnas.89.13.6119. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Demay M. B., Gerardi J. M., DeLuca H. F., Kronenberg H. M. DNA sequences in the rat osteocalcin gene that bind the 1,25-dihydroxyvitamin D3 receptor and confer responsiveness to 1,25-dihydroxyvitamin D3. Proc Natl Acad Sci U S A. 1990 Jan;87(1):369–373. doi: 10.1073/pnas.87.1.369. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Dignam J. D., Lebovitz R. M., Roeder R. G. Accurate transcription initiation by RNA polymerase II in a soluble extract from isolated mammalian nuclei. Nucleic Acids Res. 1983 Mar 11;11(5):1475–1489. doi: 10.1093/nar/11.5.1475. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Dworetzky S. I., Wright K. L., Fey E. G., Penman S., Lian J. B., Stein J. L., Stein G. S. Sequence-specific DNA-binding proteins are components of a nuclear matrix-attachment site. Proc Natl Acad Sci U S A. 1992 May 1;89(9):4178–4182. doi: 10.1073/pnas.89.9.4178. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Ecarot-Charrier B., Glorieux F. H., van der Rest M., Pereira G. Osteoblasts isolated from mouse calvaria initiate matrix mineralization in culture. J Cell Biol. 1983 Mar;96(3):639–643. doi: 10.1083/jcb.96.3.639. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Ecarot-Charrier B., Shepard N., Charette G., Grynpas M., Glorieux F. H. Mineralization in osteoblast cultures: a light and electron microscopic study. Bone. 1988;9(3):147–154. doi: 10.1016/8756-3282(88)90004-x. [DOI] [PubMed] [Google Scholar]
  12. Grimes S., Weisz-Carrington P., Daum H., 3rd, Smith J., Green L., Wright K., Stein G., Stein J. A rat histone H4 gene closely associated with the testis-specific H1t gene. Exp Cell Res. 1987 Dec;173(2):534–545. doi: 10.1016/0014-4827(87)90293-x. [DOI] [PubMed] [Google Scholar]
  13. Gundberg C. M., Hauschka P. V., Lian J. B., Gallop P. M. Osteocalcin: isolation, characterization, and detection. Methods Enzymol. 1984;107:516–544. doi: 10.1016/0076-6879(84)07036-1. [DOI] [PubMed] [Google Scholar]
  14. Holthuis J., Owen T. A., van Wijnen A. J., Wright K. L., Ramsey-Ewing A., Kennedy M. B., Carter R., Cosenza S. C., Soprano K. J., Lian J. B. Tumor cells exhibit deregulation of the cell cycle histone gene promoter factor HiNF-D. Science. 1990 Mar 23;247(4949 Pt 1):1454–1457. doi: 10.1126/science.247.4949.1454. [DOI] [PubMed] [Google Scholar]
  15. Lee W., Haslinger A., Karin M., Tjian R. Activation of transcription by two factors that bind promoter and enhancer sequences of the human metallothionein gene and SV40. Nature. 1987 Jan 22;325(6102):368–372. doi: 10.1038/325368a0. [DOI] [PubMed] [Google Scholar]
  16. Lian J., Stewart C., Puchacz E., Mackowiak S., Shalhoub V., Collart D., Zambetti G., Stein G. Structure of the rat osteocalcin gene and regulation of vitamin D-dependent expression. Proc Natl Acad Sci U S A. 1989 Feb;86(4):1143–1147. doi: 10.1073/pnas.86.4.1143. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Markose E. R., Stein J. L., Stein G. S., Lian J. B. Vitamin D-mediated modifications in protein-DNA interactions at two promoter elements of the osteocalcin gene. Proc Natl Acad Sci U S A. 1990 Mar;87(5):1701–1705. doi: 10.1073/pnas.87.5.1701. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Morrison N. A., Shine J., Fragonas J. C., Verkest V., McMenemy M. L., Eisman J. A. 1,25-dihydroxyvitamin D-responsive element and glucocorticoid repression in the osteocalcin gene. Science. 1989 Dec 1;246(4934):1158–1161. doi: 10.1126/science.2588000. [DOI] [PubMed] [Google Scholar]
  19. Owen T. A., Aronow M. S., Barone L. M., Bettencourt B., Stein G. S., Lian J. B. Pleiotropic effects of vitamin D on osteoblast gene expression are related to the proliferative and differentiated state of the bone cell phenotype: dependency upon basal levels of gene expression, duration of exposure, and bone matrix competency in normal rat osteoblast cultures. Endocrinology. 1991 Mar;128(3):1496–1504. doi: 10.1210/endo-128-3-1496. [DOI] [PubMed] [Google Scholar]
  20. Owen T. A., Aronow M., Shalhoub V., Barone L. M., Wilming L., Tassinari M. S., Kennedy M. B., Pockwinse S., Lian J. B., Stein G. S. Progressive development of the rat osteoblast phenotype in vitro: reciprocal relationships in expression of genes associated with osteoblast proliferation and differentiation during formation of the bone extracellular matrix. J Cell Physiol. 1990 Jun;143(3):420–430. doi: 10.1002/jcp.1041430304. [DOI] [PubMed] [Google Scholar]
  21. Owen T. A., Bortell R., Yocum S. A., Smock S. L., Zhang M., Abate C., Shalhoub V., Aronin N., Wright K. L., van Wijnen A. J. Coordinate occupancy of AP-1 sites in the vitamin D-responsive and CCAAT box elements by Fos-Jun in the osteocalcin gene: model for phenotype suppression of transcription. Proc Natl Acad Sci U S A. 1990 Dec;87(24):9990–9994. doi: 10.1073/pnas.87.24.9990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Owen T. A., Holthuis J., Markose E., van Wijnen A. J., Wolfe S. A., Grimes S. R., Lian J. B., Stein G. S. Modifications of protein-DNA interactions in the proximal promoter of a cell-growth-regulated histone gene during onset and progression of osteoblast differentiation. Proc Natl Acad Sci U S A. 1990 Jul;87(13):5129–5133. doi: 10.1073/pnas.87.13.5129. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Ozono K., Liao J., Kerner S. A., Scott R. A., Pike J. W. The vitamin D-responsive element in the human osteocalcin gene. Association with a nuclear proto-oncogene enhancer. J Biol Chem. 1990 Dec 15;265(35):21881–21888. [PubMed] [Google Scholar]
  24. Schepmoes G., Breen E., Owen T. A., Aronow M. A., Stein G. S., Lian J. B. Influence of dexamethasone on the vitamin D-mediated regulation of osteocalcin gene expression. J Cell Biochem. 1991 Oct;47(2):184–196. doi: 10.1002/jcb.240470212. [DOI] [PubMed] [Google Scholar]
  25. Schüle R., Rangarajan P., Kliewer S., Ransone L. J., Bolado J., Yang N., Verma I. M., Evans R. M. Functional antagonism between oncoprotein c-Jun and the glucocorticoid receptor. Cell. 1990 Sep 21;62(6):1217–1226. doi: 10.1016/0092-8674(90)90397-w. [DOI] [PubMed] [Google Scholar]
  26. Schüle R., Umesono K., Mangelsdorf D. J., Bolado J., Pike J. W., Evans R. M. Jun-Fos and receptors for vitamins A and D recognize a common response element in the human osteocalcin gene. Cell. 1990 May 4;61(3):497–504. doi: 10.1016/0092-8674(90)90531-i. [DOI] [PubMed] [Google Scholar]
  27. Shalhoub V., Jackson M. E., Lian J. B., Stein G. S., Marks S. C., Jr Gene expression during skeletal development in three osteopetrotic rat mutations. Evidence for osteoblast abnormalities. J Biol Chem. 1991 May 25;266(15):9847–9856. [PubMed] [Google Scholar]
  28. Stein G. S., Lian J. B., Owen T. A. Bone cell differentiation: a functionally coupled relationship between expression of cell-growth- and tissue-specific genes. Curr Opin Cell Biol. 1990 Dec;2(6):1018–1027. doi: 10.1016/0955-0674(90)90151-4. [DOI] [PubMed] [Google Scholar]
  29. Strömstedt P. E., Poellinger L., Gustafsson J. A., Carlstedt-Duke J. The glucocorticoid receptor binds to a sequence overlapping the TATA box of the human osteocalcin promoter: a potential mechanism for negative regulation. Mol Cell Biol. 1991 Jun;11(6):3379–3383. doi: 10.1128/mcb.11.6.3379. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Terpening C. M., Haussler C. A., Jurutka P. W., Galligan M. A., Komm B. S., Haussler M. R. The vitamin D-responsive element in the rat bone Gla protein gene is an imperfect direct repeat that cooperates with other cis-elements in 1,25-dihydroxyvitamin D3- mediated transcriptional activation. Mol Endocrinol. 1991 Mar;5(3):373–385. doi: 10.1210/mend-5-3-373. [DOI] [PubMed] [Google Scholar]
  31. Umesono K., Murakami K. K., Thompson C. C., Evans R. M. Direct repeats as selective response elements for the thyroid hormone, retinoic acid, and vitamin D3 receptors. Cell. 1991 Jun 28;65(7):1255–1266. doi: 10.1016/0092-8674(91)90020-y. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Yang-Yen H. F., Chambard J. C., Sun Y. L., Smeal T., Schmidt T. J., Drouin J., Karin M. Transcriptional interference between c-Jun and the glucocorticoid receptor: mutual inhibition of DNA binding due to direct protein-protein interaction. Cell. 1990 Sep 21;62(6):1205–1215. doi: 10.1016/0092-8674(90)90396-v. [DOI] [PubMed] [Google Scholar]
  33. Yu V. C., Delsert C., Andersen B., Holloway J. M., Devary O. V., När A. M., Kim S. Y., Boutin J. M., Glass C. K., Rosenfeld M. G. RXR beta: a coregulator that enhances binding of retinoic acid, thyroid hormone, and vitamin D receptors to their cognate response elements. Cell. 1991 Dec 20;67(6):1251–1266. doi: 10.1016/0092-8674(91)90301-e. [DOI] [PubMed] [Google Scholar]

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