Genetic dissection of the signaling domain of a mammalian steroid receptor in yeast

MJ Garabedian and KR Yamamoto

Department of Biochemistry and Biophysics, University of California, San Francisco 94143-0448.

The mechanism of signal transduction by steroid receptor proteins is complex and not yet understood. We describe here a facile genetic strategy for dissection of the rat glucocorticoid receptor "signaling domain," a region of the protein that binds and transduces the hormonal signal. We found that the characteristics of signal transduction by the receptor expressed in yeast were similar to those of endogenous receptors in mammalian cells. Interestingly, the rank order of particular ligands differed between species with respect to receptor binding and biological efficacy. This suggests that factors in addition to the receptor alone must determine or influence ligand efficacy in vivo. To obtain a collection of receptors with distinct defects in signal transduction, we screened in yeast an extensive series of random point mutations introduced in that region in vitro. Three phenotypic classes were obtained: one group failed to bind hormone, a second displayed altered ligand specificity, and a third bound hormone but lacked regulatory activity. Our results demonstrate that analysis of glucocorticoid receptor action in yeast provides a general approach for analyzing the mechanism of signaling by the nuclear receptor family and may facilitate identification of non-receptor factors that participate in this process.

Volume 3, Issue 11, pp. 1245-1257, 11/01/1992
Copyright © 1992 by The American Society for Cell Biology

This article has been cited by other articles:

				N. Yoshikawa, K. Yamamoto, N. Shimizu, S. Yamada, C. Morimoto, and H. Tanaka The Distinct Agonistic Properties of the Phenylpyrazolosteroid Cortivazol Reveal Interdomain Communication within the Glucocorticoid Receptor Mol. Endocrinol., May 1, 2005; 19(5): 1110 - 1124. [Abstract] [Full Text] [PDF]

				N. Ismaili, R. Blind, and M. J. Garabedian Stabilization of the Unliganded Glucocorticoid Receptor by TSG101 J. Biol. Chem., March 25, 2005; 280(12): 11120 - 11126. [Abstract] [Full Text] [PDF]

				G. M. Nelson, V. Prapapanich, P. E. Carrigan, P. J. Roberts, D. L. Riggs, and D. F. Smith The Heat Shock Protein 70 Cochaperone Hip Enhances Functional Maturation of Glucocorticoid Receptor Mol. Endocrinol., July 1, 2004; 18(7): 1620 - 1630. [Abstract] [Full Text] [PDF]

				B. Kauppi, C. Jakob, M. Farnegardh, J. Yang, H. Ahola, M. Alarcon, K. Calles, O. Engstrom, J. Harlan, S. Muchmore, et al. The Three-dimensional Structures of Antagonistic and Agonistic Forms of the Glucocorticoid Receptor Ligand-binding Domain: RU-486 INDUCES A TRANSCONFORMATION THAT LEADS TO ACTIVE ANTAGONISM J. Biol. Chem., June 13, 2003; 278(25): 22748 - 22754. [Abstract] [Full Text] [PDF]

				S. M. Markus, S. S. Taneja, S. K. Logan, W. Li, S. Ha, A. B. Hittelman, I. Rogatsky, and M. J. Garabedian Identification and Characterization of ART-27, a Novel Coactivator for the Androgen Receptor N Terminus Mol. Biol. Cell, February 1, 2002; 13(2): 670 - 682. [Abstract] [Full Text] [PDF]

				J. G. A. Savory, G. G. Préfontaine, C. Lamprecht, M. Liao, R. F. Walther, Y. A. Lefebvre, and R. J. G. Haché Glucocorticoid Receptor Homodimers and Glucocorticoid-Mineralocorticoid Receptor Heterodimers Form in the Cytoplasm through Alternative Dimerization Interfaces Mol. Cell. Biol., February 1, 2001; 21(3): 781 - 793. [Abstract] [Full Text]

				R. Sitcheran, R. Emter, A. Kralli, and K. R. Yamamoto A Genetic Analysis of Glucocorticoid Receptor Signaling: Identification and Characterization of Ligand-Effect Modulators in Saccharomyces cerevisiae Genetics, November 1, 2000; 156(3): 963 - 972. [Abstract] [Full Text]

				J. DiRenzo, Y. Shang, M. Phelan, S. Sif, M. Myers, R. Kingston, and M. Brown BRG-1 Is Recruited to Estrogen-Responsive Promoters and Cooperates with Factors Involved in Histone Acetylation Mol. Cell. Biol., October 15, 2000; 20(20): 7541 - 7549. [Abstract] [Full Text]

				J. L. Johnson and E. A. Craig A Role for the Hsp40 Ydj1 in Repression of Basal Steroid Receptor Activity in Yeast Mol. Cell. Biol., May 1, 2000; 20(9): 3027 - 3036. [Abstract] [Full Text]

				B. Panaretou, K. Sinclair, C. Prodromou, J. Johal, L. Pearl, and P. W. Piper The Hsp90 of Candida albicans can confer Hsp90 functions in Saccharomyces cerevisiae: a potential model for the processes that generate immunogenic fragments of this molecular chaperone in C. albicans infections Microbiology, December 1, 1999; 145(12): 3455 - 3463. [Abstract] [Full Text]

				M. Maggiolini, O. Donze, E. Jeannin, S. Ando, and D. Picard Adrenal Androgens Stimulate the Proliferation of Breast Cancer Cells as Direct Activators of Estrogen Receptor {{alpha}} Cancer Res., October 1, 1999; 59(19): 4864 - 4869. [Abstract] [Full Text] [PDF]

				H. Hong, L. Yang, and M. R. Stallcup Hormone-independent Transcriptional Activation and Coactivator Binding by Novel Orphan Nuclear Receptor ERR3 J. Biol. Chem., August 6, 1999; 274(32): 22618 - 22626. [Abstract] [Full Text] [PDF]

				R. Knoblauch and M. J. Garabedian Role for Hsp90-Associated Cochaperone p23 in Estrogen Receptor Signal Transduction Mol. Cell. Biol., May 1, 1999; 19(5): 3748 - 3759. [Abstract] [Full Text] [PDF]

				J. G. A. Savory, B. Hsu, I. R. Laquian, W. Giffin, T. Reich, R. J. G. Hache, and Y. A. Lefebvre Discrimination between NL1- and NL2-Mediated Nuclear Localization of the Glucocorticoid Receptor Mol. Cell. Biol., February 1, 1999; 19(2): 1025 - 1037. [Abstract] [Full Text] [PDF]

				S. P. Bohen Genetic and Biochemical Analysis of p23 and Ansamycin Antibiotics in the Function of Hsp90-Dependent Signaling Proteins Mol. Cell. Biol., June 1, 1998; 18(6): 3330 - 3339. [Abstract] [Full Text]

				N. Dzeletovic, J. McGuire, M. Daujat, J. Tholander, M. Ema, Y. Fujii-Kuriyama, J. Bergman, P. Maurel, and L. Poellinger Regulation of Dioxin Receptor Function by Omeprazole J. Biol. Chem., May 9, 1997; 272(19): 12705 - 12713. [Abstract] [Full Text] [PDF]

				A. A. Duina, H.-C. J. Chang, J. A. Marsh, S. Lindquist, and R. F. Gaber A Cyclophilin Function in Hsp90-Dependent Signal Transduction Science, December 6, 1996; 274(5293): 1713 - 1715. [Abstract] [Full Text]

				E. J. Licitra and J. O. Liu A three-hybrid system for detecting small ligand-protein receptor interactions PNAS, November 12, 1996; 93(23): 12817 - 12821. [Abstract] [Full Text] [PDF]

				B R Cairns, R S Levinson, K R Yamamoto, and R D Kornberg Essential role of Swp73p in the function of yeast Swi/Snf complex. Genes & Dev., September 1, 1996; 10(17): 2131 - 2144. [Abstract] [PDF]

				A. Kralli and K. R. Yamamoto An FK506-sensitive Transporter Selectively Decreases Intracellular Levels and Potency of Steroid Hormones J. Biol. Chem., July 19, 1996; 271(29): 17152 - 17156. [Abstract] [Full Text] [PDF]

				D B Starr, W Matsui, J R Thomas, and K R Yamamoto Intracellular receptors use a common mechanism to interpret signaling information at response elements. Genes & Dev., May 15, 1996; 10(10): 1271 - 1283. [Abstract] [PDF]

				S. P. Bohen and S. P. Bohen Hsp90 Mutants Disrupt Glucocorticoid Receptor Ligand Binding and Destabilize Aporeceptor Complexes J. Biol. Chem., December 8, 1995; 270(49): 29433 - 29438. [Abstract] [Full Text] [PDF]

				J A Lefstin, J R Thomas, and K R Yamamoto Influence of a steroid receptor DNA-binding domain on transcriptional regulatory functions. Genes & Dev., December 1, 1994; 8(23): 2842 - 2856. [Abstract] [PDF]