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. 1997 Feb;17(2):644–655. doi: 10.1128/mcb.17.2.644

Retinoid X receptor (RXR) within the RXR-retinoic acid receptor heterodimer binds its ligand and enhances retinoid-dependent gene expression.

S Minucci 1, M Leid 1, R Toyama 1, J P Saint-Jeannet 1, V J Peterson 1, V Horn 1, J E Ishmael 1, N Bhattacharyya 1, A Dey 1, I B Dawid 1, K Ozato 1
PMCID: PMC231790  PMID: 9001218

Abstract

Retinoic acid receptor (RAR) and retinoid X receptor (RXR) form heterodimers and regulate retinoid-mediated gene expression. We studied binding of RXR- and RAR-selective ligands to the RXR-RAR heterodimer and subsequent transcription. In limited proteolysis analyses, both RXR and RAR in the heterodimer bound their respective ligands and underwent a conformational change in the presence of a retinoic acid-responsive element. In reporter analyses, the RAR ligand (but not the RXR ligand), when added singly, activated transcription, but coaddition of the two ligands led to synergistic activation of transcription. This activation required the AF-2 domain of both RXR and RAR. Genomic footprinting analysis was performed with P19 embryonal carcinoma cells, in which transcription of the RARbeta gene is induced upon retinoid addition. Paralleling the reporter activation data, only the RAR ligand induced in vivo occupancy of the RARbeta2 promoter when added singly. However, at suboptimal concentrations of RAR ligand, coaddition of the RXR ligand increased the stability of promoter occupancy. Thus, liganded RXR and RAR both participate in transcription. Finally, when these ligands were tested for teratogenic effects on zebra fish and Xenopus embryos, we found that coadministration of the RXR and RAR ligands caused more severe abnormalities in these embryos than either ligand alone, providing biological support for the synergistic action of the two ligands.

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Selected References

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  1. Allan G. F., Leng X., Tsai S. Y., Weigel N. L., Edwards D. P., Tsai M. J., O'Malley B. W. Hormone and antihormone induce distinct conformational changes which are central to steroid receptor activation. J Biol Chem. 1992 Sep 25;267(27):19513–19520. [PubMed] [Google Scholar]
  2. Allan G. F., Tsai S. Y., Tsai M. J., O'Malley B. W. Ligand-dependent conformational changes in the progesterone receptor are necessary for events that follow DNA binding. Proc Natl Acad Sci U S A. 1992 Dec 15;89(24):11750–11754. doi: 10.1073/pnas.89.24.11750. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Allegretto E. A., McClurg M. R., Lazarchik S. B., Clemm D. L., Kerner S. A., Elgort M. G., Boehm M. F., White S. K., Pike J. W., Heyman R. A. Transactivation properties of retinoic acid and retinoid X receptors in mammalian cells and yeast. Correlation with hormone binding and effects of metabolism. J Biol Chem. 1993 Dec 15;268(35):26625–26633. [PubMed] [Google Scholar]
  4. Allenby G., Bocquel M. T., Saunders M., Kazmer S., Speck J., Rosenberger M., Lovey A., Kastner P., Grippo J. F., Chambon P. Retinoic acid receptors and retinoid X receptors: interactions with endogenous retinoic acids. Proc Natl Acad Sci U S A. 1993 Jan 1;90(1):30–34. doi: 10.1073/pnas.90.1.30. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Angulo A., Suto C., Heyman R. A., Ghazal P. Characterization of the sequences of the human cytomegalovirus enhancer that mediate differential regulation by natural and synthetic retinoids. Mol Endocrinol. 1996 Jul;10(7):781–793. doi: 10.1210/mend.10.7.8813719. [DOI] [PubMed] [Google Scholar]
  6. Apfel C. M., Kamber M., Klaus M., Mohr P., Keidel S., LeMotte P. K. Enhancement of HL-60 differentiation by a new class of retinoids with selective activity on retinoid X receptor. J Biol Chem. 1995 Dec 22;270(51):30765–30772. doi: 10.1074/jbc.270.51.30765. [DOI] [PubMed] [Google Scholar]
  7. Barettino D., Vivanco Ruiz M. M., Stunnenberg H. G. Characterization of the ligand-dependent transactivation domain of thyroid hormone receptor. EMBO J. 1994 Jul 1;13(13):3039–3049. doi: 10.1002/j.1460-2075.1994.tb06603.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Bhat M. K., Parkison C., McPhie P., Liang C. M., Cheng S. Y. Conformational changes of human beta 1 thyroid hormone receptor induced by binding of 3,3',5-triiodo-L-thyronine. Biochem Biophys Res Commun. 1993 Aug 31;195(1):385–392. doi: 10.1006/bbrc.1993.2055. [DOI] [PubMed] [Google Scholar]
  9. Blanco J. C., Dey A., Leid M., Minucci S., Park B. K., Jurutka P. W., Haussler M. R., Ozato K. Inhibition of ligand induced promoter occupancy in vivo by a dominant negative RXR. Genes Cells. 1996 Feb;1(2):209–221. doi: 10.1046/j.1365-2443.1996.d01-229.x. [DOI] [PubMed] [Google Scholar]
  10. Brooks S. C., 3rd, Kazmer S., Levin A. A., Yen A. Myeloid differentiation and retinoblastoma phosphorylation changes in HL-60 cells induced by retinoic acid receptor- and retinoid X receptor-selective retinoic acid analogs. Blood. 1996 Jan 1;87(1):227–237. [PubMed] [Google Scholar]
  11. Chambon P. A decade of molecular biology of retinoic acid receptors. FASEB J. 1996 Jul;10(9):940–954. [PubMed] [Google Scholar]
  12. Chen J. D., Evans R. M. A transcriptional co-repressor that interacts with nuclear hormone receptors. Nature. 1995 Oct 5;377(6548):454–457. doi: 10.1038/377454a0. [DOI] [PubMed] [Google Scholar]
  13. Chen J. Y., Clifford J., Zusi C., Starrett J., Tortolani D., Ostrowski J., Reczek P. R., Chambon P., Gronemeyer H. Two distinct actions of retinoid-receptor ligands. Nature. 1996 Aug 29;382(6594):819–822. doi: 10.1038/382819a0. [DOI] [PubMed] [Google Scholar]
  14. Clifford J., Chiba H., Sobieszczuk D., Metzger D., Chambon P. RXRalpha-null F9 embryonal carcinoma cells are resistant to the differentiation, anti-proliferative and apoptotic effects of retinoids. EMBO J. 1996 Aug 15;15(16):4142–4155. [PMC free article] [PubMed] [Google Scholar]
  15. Davis K. D., Berrodin T. J., Stelmach J. E., Winkler J. D., Lazar M. A. Endogenous retinoid X receptors can function as hormone receptors in pituitary cells. Mol Cell Biol. 1994 Nov;14(11):7105–7110. doi: 10.1128/mcb.14.11.7105. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Dey A., Thornton A. M., Lonergan M., Weissman S. M., Chamberlain J. W., Ozato K. Occupancy of upstream regulatory sites in vivo coincides with major histocompatibility complex class I gene expression in mouse tissues. Mol Cell Biol. 1992 Aug;12(8):3590–3599. doi: 10.1128/mcb.12.8.3590. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Durand B., Saunders M., Gaudon C., Roy B., Losson R., Chambon P. Activation function 2 (AF-2) of retinoic acid receptor and 9-cis retinoic acid receptor: presence of a conserved autonomous constitutive activating domain and influence of the nature of the response element on AF-2 activity. EMBO J. 1994 Nov 15;13(22):5370–5382. doi: 10.1002/j.1460-2075.1994.tb06872.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Durston A. J., Timmermans J. P., Hage W. J., Hendriks H. F., de Vries N. J., Heideveld M., Nieuwkoop P. D. Retinoic acid causes an anteroposterior transformation in the developing central nervous system. Nature. 1989 Jul 13;340(6229):140–144. doi: 10.1038/340140a0. [DOI] [PubMed] [Google Scholar]
  19. Forman B. M., Goode E., Chen J., Oro A. E., Bradley D. J., Perlmann T., Noonan D. J., Burka L. T., McMorris T., Lamph W. W. Identification of a nuclear receptor that is activated by farnesol metabolites. Cell. 1995 Jun 2;81(5):687–693. doi: 10.1016/0092-8674(95)90530-8. [DOI] [PubMed] [Google Scholar]
  20. Forman B. M., Umesono K., Chen J., Evans R. M. Unique response pathways are established by allosteric interactions among nuclear hormone receptors. Cell. 1995 May 19;81(4):541–550. doi: 10.1016/0092-8674(95)90075-6. [DOI] [PubMed] [Google Scholar]
  21. Fritsch M., Leary C. M., Furlow J. D., Ahrens H., Schuh T. J., Mueller G. C., Gorski J. A ligand-induced conformational change in the estrogen receptor is localized in the steroid binding domain. Biochemistry. 1992 Jun 16;31(23):5303–5311. doi: 10.1021/bi00138a009. [DOI] [PubMed] [Google Scholar]
  22. Glass C. K. Differential recognition of target genes by nuclear receptor monomers, dimers, and heterodimers. Endocr Rev. 1994 Jun;15(3):391–407. doi: 10.1210/edrv-15-3-391. [DOI] [PubMed] [Google Scholar]
  23. Heyman R. A., Mangelsdorf D. J., Dyck J. A., Stein R. B., Eichele G., Evans R. M., Thaller C. 9-cis retinoic acid is a high affinity ligand for the retinoid X receptor. Cell. 1992 Jan 24;68(2):397–406. doi: 10.1016/0092-8674(92)90479-v. [DOI] [PubMed] [Google Scholar]
  24. Horn V., Minucci S., Ogryzko V. V., Adamson E. D., Howard B. H., Levin A. A., Ozato K. RAR and RXR selective ligands cooperatively induce apoptosis and neuronal differentiation in P19 embryonal carcinoma cells. FASEB J. 1996 Jul;10(9):1071–1077. doi: 10.1096/fasebj.10.9.8801169. [DOI] [PubMed] [Google Scholar]
  25. Hörlein A. J., När A. M., Heinzel T., Torchia J., Gloss B., Kurokawa R., Ryan A., Kamei Y., Söderström M., Glass C. K. Ligand-independent repression by the thyroid hormone receptor mediated by a nuclear receptor co-repressor. Nature. 1995 Oct 5;377(6548):397–404. doi: 10.1038/377397a0. [DOI] [PubMed] [Google Scholar]
  26. Kastner P., Mark M., Chambon P. Nonsteroid nuclear receptors: what are genetic studies telling us about their role in real life? Cell. 1995 Dec 15;83(6):859–869. doi: 10.1016/0092-8674(95)90202-3. [DOI] [PubMed] [Google Scholar]
  27. Keidel S., LeMotte P., Apfel C. Different agonist- and antagonist-induced conformational changes in retinoic acid receptors analyzed by protease mapping. Mol Cell Biol. 1994 Jan;14(1):287–298. doi: 10.1128/mcb.14.1.287. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Kersten S., Dawson M. I., Lewis B. A., Noy N. Individual subunits of heterodimers comprised of retinoic acid and retinoid X receptors interact with their ligands independently. Biochemistry. 1996 Mar 26;35(12):3816–3824. doi: 10.1021/bi952737k. [DOI] [PubMed] [Google Scholar]
  29. Kliewer S. A., Umesono K., Noonan D. J., Heyman R. A., Evans R. M. Convergence of 9-cis retinoic acid and peroxisome proliferator signalling pathways through heterodimer formation of their receptors. Nature. 1992 Aug 27;358(6389):771–774. doi: 10.1038/358771a0. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Kruyt F. A., Folkers G., van den Brink C. E., van der Saag P. T. A cyclic AMP response element is involved in retinoic acid-dependent RAR beta 2 promoter activation. Nucleic Acids Res. 1992 Dec 11;20(23):6393–6399. doi: 10.1093/nar/20.23.6393. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Kurokawa R., DiRenzo J., Boehm M., Sugarman J., Gloss B., Rosenfeld M. G., Heyman R. A., Glass C. K. Regulation of retinoid signalling by receptor polarity and allosteric control of ligand binding. Nature. 1994 Oct 6;371(6497):528–531. doi: 10.1038/371528a0. [DOI] [PubMed] [Google Scholar]
  32. Kurokawa R., Söderström M., Hörlein A., Halachmi S., Brown M., Rosenfeld M. G., Glass C. K. Polarity-specific activities of retinoic acid receptors determined by a co-repressor. Nature. 1995 Oct 5;377(6548):451–454. doi: 10.1038/377451a0. [DOI] [PubMed] [Google Scholar]
  33. La Vista-Picard N., Hobbs P. D., Pfahl M., Dawson M. I., Pfahl M. The receptor-DNA complex determines the retinoid response: a mechanism for the diversification of the ligand signal. Mol Cell Biol. 1996 Aug;16(8):4137–4146. doi: 10.1128/mcb.16.8.4137. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Leblanc B. P., Stunnenberg H. G. 9-cis retinoic acid signaling: changing partners causes some excitement. Genes Dev. 1995 Aug 1;9(15):1811–1816. doi: 10.1101/gad.9.15.1811. [DOI] [PubMed] [Google Scholar]
  35. Lehmann J. M., Jong L., Fanjul A., Cameron J. F., Lu X. P., Haefner P., Dawson M. I., Pfahl M. Retinoids selective for retinoid X receptor response pathways. Science. 1992 Dec 18;258(5090):1944–1946. doi: 10.1126/science.1335166. [DOI] [PubMed] [Google Scholar]
  36. Leid M. Ligand-induced alteration of the protease sensitivity of retinoid X receptor alpha. J Biol Chem. 1994 May 13;269(19):14175–14181. [PubMed] [Google Scholar]
  37. Leng X., Blanco J., Tsai S. Y., Ozato K., O'Malley B. W., Tsai M. J. Mouse retinoid X receptor contains a separable ligand-binding and transactivation domain in its E region. Mol Cell Biol. 1995 Jan;15(1):255–263. doi: 10.1128/mcb.15.1.255. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Leng X., Tsai S. Y., O'Malley B. W., Tsai M. J. Ligand-dependent conformational changes in thyroid hormone and retinoic acid receptors are potentially enhanced by heterodimerization with retinoic X receptor. J Steroid Biochem Mol Biol. 1993 Dec;46(6):643–661. doi: 10.1016/0960-0760(93)90306-h. [DOI] [PubMed] [Google Scholar]
  39. Levin A. A., Sturzenbecker L. J., Kazmer S., Bosakowski T., Huselton C., Allenby G., Speck J., Kratzeisen C., Rosenberger M., Lovey A. 9-cis retinoic acid stereoisomer binds and activates the nuclear receptor RXR alpha. Nature. 1992 Jan 23;355(6358):359–361. doi: 10.1038/355359a0. [DOI] [PubMed] [Google Scholar]
  40. Lotan R., Dawson M. I., Zou C. C., Jong L., Lotan D., Zou C. P. Enhanced efficacy of combinations of retinoic acid- and retinoid X receptor-selective retinoids and alpha-interferon in inhibition of cervical carcinoma cell proliferation. Cancer Res. 1995 Jan 15;55(2):232–236. [PubMed] [Google Scholar]
  41. Mangelsdorf D. J., Evans R. M. The RXR heterodimers and orphan receptors. Cell. 1995 Dec 15;83(6):841–850. doi: 10.1016/0092-8674(95)90200-7. [DOI] [PubMed] [Google Scholar]
  42. Mangelsdorf D. J., Thummel C., Beato M., Herrlich P., Schütz G., Umesono K., Blumberg B., Kastner P., Mark M., Chambon P. The nuclear receptor superfamily: the second decade. Cell. 1995 Dec 15;83(6):835–839. doi: 10.1016/0092-8674(95)90199-x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Minucci S., Botquin V., Yeom Y. I., Dey A., Sylvester I., Zand D. J., Ohbo K., Ozato K., Scholer H. R. Retinoic acid-mediated down-regulation of Oct3/4 coincides with the loss of promoter occupancy in vivo. EMBO J. 1996 Feb 15;15(4):888–899. [PMC free article] [PubMed] [Google Scholar]
  44. Minucci S., Ozato K. Retinoid receptors in transcriptional regulation. Curr Opin Genet Dev. 1996 Oct;6(5):567–574. doi: 10.1016/s0959-437x(96)80085-2. [DOI] [PubMed] [Google Scholar]
  45. Minucci S., Saint-Jeannet J. P., Toyama R., Scita G., DeLuca L. M., Tiara M., Levin A. A., Ozato K., Dawid I. B. Retinoid X receptor-selective ligands produce malformations in Xenopus embryos. Proc Natl Acad Sci U S A. 1996 Mar 5;93(5):1803–1807. doi: 10.1073/pnas.93.5.1803. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Minucci S., Zand D. J., Dey A., Marks M. S., Nagata T., Grippo J. F., Ozato K. Dominant negative retinoid X receptor beta inhibits retinoic acid-responsive gene regulation in embryonal carcinoma cells. Mol Cell Biol. 1994 Jan;14(1):360–372. doi: 10.1128/mcb.14.1.360. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Nagy L., Thomázy V. A., Shipley G. L., Fésüs L., Lamph W., Heyman R. A., Chandraratna R. A., Davies P. J. Activation of retinoid X receptors induces apoptosis in HL-60 cell lines. Mol Cell Biol. 1995 Jul;15(7):3540–3551. doi: 10.1128/mcb.15.7.3540. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Perlmann T., Jansson L. A novel pathway for vitamin A signaling mediated by RXR heterodimerization with NGFI-B and NURR1. Genes Dev. 1995 Apr 1;9(7):769–782. doi: 10.1101/gad.9.7.769. [DOI] [PubMed] [Google Scholar]
  49. Rosen E. D., O'Donnell A. L., Koenig R. J. Ligand-dependent synergy of thyroid hormone and retinoid X receptors. J Biol Chem. 1992 Nov 5;267(31):22010–22013. [PubMed] [Google Scholar]
  50. Roy B., Taneja R., Chambon P. Synergistic activation of retinoic acid (RA)-responsive genes and induction of embryonal carcinoma cell differentiation by an RA receptor alpha (RAR alpha)-, RAR beta-, or RAR gamma-selective ligand in combination with a retinoid X receptor-specific ligand. Mol Cell Biol. 1995 Dec;15(12):6481–6487. doi: 10.1128/mcb.15.12.6481. [DOI] [PMC free article] [PubMed] [Google Scholar]
  51. Schulman I. G., Juguilon H., Evans R. M. Activation and repression by nuclear hormone receptors: hormone modulates an equilibrium between active and repressive states. Mol Cell Biol. 1996 Jul;16(7):3807–3813. doi: 10.1128/mcb.16.7.3807. [DOI] [PMC free article] [PubMed] [Google Scholar]
  52. Simons S. S., Jr, Sistare F. D., Chakraborti P. K. Steroid binding activity is retained in a 16-kDa fragment of the steroid binding domain of rat glucocorticoid receptors. J Biol Chem. 1989 Aug 25;264(24):14493–14497. [PubMed] [Google Scholar]
  53. Sucov H. M., Murakami K. K., Evans R. M. Characterization of an autoregulated response element in the mouse retinoic acid receptor type beta gene. Proc Natl Acad Sci U S A. 1990 Jul;87(14):5392–5396. doi: 10.1073/pnas.87.14.5392. [DOI] [PMC free article] [PubMed] [Google Scholar]
  54. Taneja R., Roy B., Plassat J. L., Zusi C. F., Ostrowski J., Reczek P. R., Chambon P. Cell-type and promoter-context dependent retinoic acid receptor (RAR) redundancies for RAR beta 2 and Hoxa-1 activation in F9 and P19 cells can be artefactually generated by gene knockouts. Proc Natl Acad Sci U S A. 1996 Jun 11;93(12):6197–6202. doi: 10.1073/pnas.93.12.6197. [DOI] [PMC free article] [PubMed] [Google Scholar]
  55. Tsai M. J., O'Malley B. W. Molecular mechanisms of action of steroid/thyroid receptor superfamily members. Annu Rev Biochem. 1994;63:451–486. doi: 10.1146/annurev.bi.63.070194.002315. [DOI] [PubMed] [Google Scholar]
  56. Valcárcel R., Holz H., Jiménez C. G., Barettino D., Stunnenberg H. G. Retinoid-dependent in vitro transcription mediated by the RXR/RAR heterodimer. Genes Dev. 1994 Dec 15;8(24):3068–3079. doi: 10.1101/gad.8.24.3068. [DOI] [PubMed] [Google Scholar]
  57. Vegeto E., Allan G. F., Schrader W. T., Tsai M. J., McDonnell D. P., O'Malley B. W. The mechanism of RU486 antagonism is dependent on the conformation of the carboxy-terminal tail of the human progesterone receptor. Cell. 1992 May 15;69(4):703–713. doi: 10.1016/0092-8674(92)90234-4. [DOI] [PubMed] [Google Scholar]
  58. Willy P. J., Umesono K., Ong E. S., Evans R. M., Heyman R. A., Mangelsdorf D. J. LXR, a nuclear receptor that defines a distinct retinoid response pathway. Genes Dev. 1995 May 1;9(9):1033–1045. doi: 10.1101/gad.9.9.1033. [DOI] [PubMed] [Google Scholar]
  59. Xiao J. H., Durand B., Chambon P., Voorhees J. J. Endogenous retinoic acid receptor (RAR)-retinoid X receptor (RXR) heterodimers are the major functional forms regulating retinoid-responsive elements in adult human keratinocytes. Binding of ligands to RAR only is sufficient for RAR-RXR heterodimers to confer ligand-dependent activation of hRAR beta 2/RARE (DR5). J Biol Chem. 1995 Feb 17;270(7):3001–3011. doi: 10.1074/jbc.270.7.3001. [DOI] [PubMed] [Google Scholar]
  60. Yang Y., Minucci S., Ozato K., Heyman R. A., Ashwell J. D. Efficient inhibition of activation-induced Fas ligand up-regulation and T cell apoptosis by retinoids requires occupancy of both retinoid X receptors and retinoic acid receptors. J Biol Chem. 1995 Aug 4;270(31):18672–18677. doi: 10.1074/jbc.270.31.18672. [DOI] [PubMed] [Google Scholar]
  61. Yao T. P., Forman B. M., Jiang Z., Cherbas L., Chen J. D., McKeown M., Cherbas P., Evans R. M. Functional ecdysone receptor is the product of EcR and Ultraspiracle genes. Nature. 1993 Dec 2;366(6454):476–479. doi: 10.1038/366476a0. [DOI] [PubMed] [Google Scholar]
  62. Zhang X. K., Lehmann J., Hoffmann B., Dawson M. I., Cameron J., Graupner G., Hermann T., Tran P., Pfahl M. Homodimer formation of retinoid X receptor induced by 9-cis retinoic acid. Nature. 1992 Aug 13;358(6387):587–591. doi: 10.1038/358587a0. [DOI] [PubMed] [Google Scholar]
  63. de Thé H., Vivanco-Ruiz M. M., Tiollais P., Stunnenberg H., Dejean A. Identification of a retinoic acid responsive element in the retinoic acid receptor beta gene. Nature. 1990 Jan 11;343(6254):177–180. doi: 10.1038/343177a0. [DOI] [PubMed] [Google Scholar]

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