An Active Nuclear Retention Signal in the Glucocorticoid Receptor Functions as a Strong Inducer of Transcriptional Activation

The glucocorticoid receptor (GR) cycles between a naive chaperone-complexed form in the cytoplasm and a transcriptionally active steroid-bound nuclear form. Nuclear import of GR occurs rapidly and is mediated through the importin α/β karyopherin import pathway. By contrast, nuclear export of GR occurs only slowly under most conditions, despite a dependence on active signaling. In this study we have defined a nuclear retention signal (NRS) in the hinge region of GR that actively opposes the nuclear export of GR as well as the nuclear export mediated through an ectopic CRM1-dependent nuclear export signal (NES). The GR NRS overlaps closely with the basic NL1 nuclear localization signal (NLS) but can be distinguished from NL1 by targeted mutagenesis. Substitution of the classical NLS from SV40 T antigen for the GR NL1 results in a receptor in which nuclear export is accelerated. Remarkably, although the SV40-modified GR remains predominantly nuclear in the presence of steroid and is recruited to transcriptional regulatory regions indistinguishably from wild-type GR, the substitution dramatically weakens the ability of GR to activate transcription of a mouse mammary tumor virus reporter gene. These results suggest that active nuclear retention of GR plays an integral role in glucocorticoid signaling. The glucocorticoid receptor (GR) cycles between a naive chaperone-complexed form in the cytoplasm and a transcriptionally active steroid-bound nuclear form. Nuclear import of GR occurs rapidly and is mediated through the importin α/β karyopherin import pathway. By contrast, nuclear export of GR occurs only slowly under most conditions, despite a dependence on active signaling. In this study we have defined a nuclear retention signal (NRS) in the hinge region of GR that actively opposes the nuclear export of GR as well as the nuclear export mediated through an ectopic CRM1-dependent nuclear export signal (NES). The GR NRS overlaps closely with the basic NL1 nuclear localization signal (NLS) but can be distinguished from NL1 by targeted mutagenesis. Substitution of the classical NLS from SV40 T antigen for the GR NL1 results in a receptor in which nuclear export is accelerated. Remarkably, although the SV40-modified GR remains predominantly nuclear in the presence of steroid and is recruited to transcriptional regulatory regions indistinguishably from wild-type GR, the substitution dramatically weakens the ability of GR to activate transcription of a mouse mammary tumor virus reporter gene. These results suggest that active nuclear retention of GR plays an integral role in glucocorticoid signaling. The glucocorticoid receptor (GR) 5The abbreviations used are: GR, glucocorticoid receptor; NRS, nuclear retention signal; NES, nuclear export signal; NLS, nuclear localization signal; DBD, DNA binding domain; LBD, ligand binding domain; MMTV, mouse mammary tumor virus; WT, wild type; ChIP, chromatin immunoprecipitation; GFP, green fluorescent protein; FRAP, fluorescence recovery after photobleaching; GST, glutathione S-transferase; CMV, cytomegalovirus. 5The abbreviations used are: GR, glucocorticoid receptor; NRS, nuclear retention signal; NES, nuclear export signal; NLS, nuclear localization signal; DBD, DNA binding domain; LBD, ligand binding domain; MMTV, mouse mammary tumor virus; WT, wild type; ChIP, chromatin immunoprecipitation; GFP, green fluorescent protein; FRAP, fluorescence recovery after photobleaching; GST, glutathione S-transferase; CMV, cytomegalovirus. is a nuclear hormone receptor of the steroid receptor subfamily whose activity is tightly regulated by glucocorticoids (1Beato M. Herrlich P. Schutz G. Cell. 1995; 83: 851-857Abstract Full Text PDF PubMed Scopus (1634) Google Scholar). GR is a modular protein that features a central DNA binding domain (DBD) and a C-terminal ligand binding domain (LBD). Two activation functions (AF-1 and AF-2Gustafsson J.A. Carlstedt-Duke J. Wrange O. Okret S. Wikstrom A.C. J. Steroid Biochem. 1986; 24: 63-68Crossref PubMed Scopus (13) Google Scholar) are located in the N terminus and the LBD of the receptor, respectively (2Gustafsson J.A. Carlstedt-Duke J. Wrange O. Okret S. Wikstrom A.C. J. Steroid Biochem. 1986; 24: 63-68Crossref PubMed Scopus (13) Google Scholar, 3Giguere V. Hollenberg S.M. Rosenfeld M.G. Evans R.M. Cell. 1986; 46: 645-652Abstract Full Text PDF PubMed Scopus (678) Google Scholar). GR is directly and indirectly involved in both the activation and suppression of genes that are involved in cell differentiation, glucose uptake and homeostasis, inflammation, response to stress, neuronal function, lipid metabolism, and cancer (4Charmandari E. Kino T. Chrousos G.P. Ann. N. Y. Acad. Sci. 2004; 1024: 1-8Crossref PubMed Scopus (51) Google Scholar, 5Chrousos G.P. Charmandari E. Kino T. J. Clin. Endocrinol. Metab. 2004; 89: 563-564Crossref PubMed Scopus (59) Google Scholar). Transcriptional regulation by GR is tightly controlled. The regulation of function by segregation in subcellular compartment has been proposed to act as an important regulatory checkpoint for a number of transcription factors, including steroid receptors (6Nguyen T. Sherratt P.J. Nioi P. Yang C.S. Pickett C.B. J. Biol. Chem. 2005; 280: 32485-32492Abstract Full Text Full Text PDF PubMed Scopus (299) Google Scholar, 7Ghisletti S. Meda C. Maggi A. Vegeto E. Mol. Cell. Biol. 2005; 25: 2957-2968Crossref PubMed Scopus (332) Google Scholar). Unlike other steroid receptors, such as estrogen receptor and progesterone receptor, that are nuclear in the absence of steroid, naive GR is largely cytoplasmic (8Picard D. Kumar V. Chambon P. Yamamoto K.R. Cell Regul. 1990; 1: 291-299Crossref PubMed Scopus (211) Google Scholar). In the cytoplasm GR is maintained in an inactive hsp90 cochaperone complex that includes hsp70, immunophilins, p23, and other factors (9Pratt W.B. Toft D.O. Endocr. Rev. 1997; 18: 306-360Crossref PubMed Scopus (1534) Google Scholar). Upon ligand binding, GR dissociates from the chaperone complex, homodimerizes, and rapidly translocates to the nucleus to regulate target gene expression (10Madan A.P. DeFranco D.B. Proc. Natl. Acad. Sci. U. S. A. 1993; 90: 3588-3592Crossref PubMed Scopus (144) Google Scholar, 11Sackey F.N. Hache R.J. Reich T. Kwast-Welfeld J. Lefebvre Y.A. Mol. Endocrinol. 1996; 10: 1191-1205PubMed Google Scholar, 12Yang J. Liu J. DeFranco D.B. J. Cell Biol. 1997; 137: 523-538Crossref PubMed Scopus (86) Google Scholar). Nuclear import of GR is accomplished through two nuclear localization signals as follows: NL1, located in a hinge-like region of GR that separates the DBD from the LBD; and NL2, which is within the LBD (13Picard D. Yamamoto K.R. EMBO J. 1987; 6: 3333-3340Crossref PubMed Scopus (722) Google Scholar). NL1 is a basic NLS that mediates the nuclear import of GR through interaction with importin α and importin 7 (14Jewell C.M. Webster J.C. Burnstein K.L. Sar M. Bodwell J.E. Cidlowski J.A. J. Steroid Biochem. Mol. Biol. 1995; 55: 135-146Crossref PubMed Scopus (55) Google Scholar, 15Savory J.G. Hsu B. Laquian I.R. Giffin W. Reich T. Hache R.J. Lefebvre Y.A. Mol. Cell. Biol. 1999; 19: 1025-1037Crossref PubMed Scopus (181) Google Scholar, 16Freedman N.D. Yamamoto K.R. Mol. Biol. Cell. 2004; 15: 2276-2286Crossref PubMed Scopus (181) Google Scholar). NL2 is strictly steroid-dependent, and both the sequence within the LBD that includes NL2 and the karyopherins that determine NL2-mediated nuclear import remain to be identified. Using a mutant GR that lacks NL1 (GRNL1–), we have previously demonstrated that NL2 is an agonist-specific NLS that mediates the incomplete localization of GR to the nucleus in cells treated with cortisol or the synthetic steroid dexamethasone (15Savory J.G. Hsu B. Laquian I.R. Giffin W. Reich T. Hache R.J. Lefebvre Y.A. Mol. Cell. Biol. 1999; 19: 1025-1037Crossref PubMed Scopus (181) Google Scholar). Furthermore, this reduced nuclear occupancy correlates with a strong decrease in GR transcriptional regulatory potential. Upon hormone withdrawal, GR reassociates rapidly into a cochaperone complex, but only slowly redistributes to the cytoplasm over periods that can extend over 12–24 h (11Sackey F.N. Hache R.J. Reich T. Kwast-Welfeld J. Lefebvre Y.A. Mol. Endocrinol. 1996; 10: 1191-1205PubMed Google Scholar, 17Hache R.J. Tse R. Reich T. Savory J.G. Lefebvre Y.A. J. Biol. Chem. 1999; 274: 1432-1439Abstract Full Text Full Text PDF PubMed Scopus (98) Google Scholar, 18Qi M. Hamilton B.J. DeFranco D. Mol. Endocrinol. 1989; 3: 1279-1288Crossref PubMed Scopus (73) Google Scholar). Furthermore, fluorescence recovery after photobleaching (FRAP), digitonin cell permeabilization, and other assays have shown that this slow export from the nucleus occurs for both liganded and steroid-withdrawn receptor (17Hache R.J. Tse R. Reich T. Savory J.G. Lefebvre Y.A. J. Biol. Chem. 1999; 274: 1432-1439Abstract Full Text Full Text PDF PubMed Scopus (98) Google Scholar, 19Walther R.F. Lamprecht C. Ridsdale A. Groulx I. Lee S. Lefebvre Y.A. Hache R.J. J. Biol. Chem. 2003; 278: 37858-37864Abstract Full Text Full Text PDF PubMed Scopus (44) Google Scholar). The mechanism through which this slow transfer of GR from the cytoplasm to the nucleus is accomplished is not understood. For other transcription factors and nuclear proteins, export from the nucleus is accomplished when nuclear export signals (NESs) interact with karyopherins, collectively referred to as exportins (20Mosammaparast N. Pemberton L.F. Trends Cell Biol. 2004; 14: 547-556Abstract Full Text Full Text PDF PubMed Scopus (269) Google Scholar). Most of the known NESs are composed of hydrophobic amino acid; however, they are highly diverse with only a loose consensus (21Fried H. Kutay U. Cell. Mol. Life Sci. 2003; 60: 1659-1688Crossref PubMed Scopus (404) Google Scholar, 22Weis K. Cell. 2003; 112: 441-451Abstract Full Text Full Text PDF PubMed Scopus (575) Google Scholar). The prototypical exportin for nuclear protein export is exportin1 or CRM1. CRM1 binds to or CRM1 of an the binding of and is by M. M. M. Cell. 1997; 90: Full Text Full Text PDF PubMed Scopus Google Scholar, K. C.S. C. K. Cell. 1997; 90: Full Text Full Text PDF PubMed Scopus Google Scholar). from and other suggest that nuclear export of GR be accomplished by both CRM1-dependent and (15Savory J.G. Hsu B. Laquian I.R. Giffin W. Reich T. Hache R.J. Lefebvre Y.A. Mol. Cell. Biol. 1999; 19: 1025-1037Crossref PubMed Scopus (181) Google Scholar, J. DeFranco D.B. Mol. Endocrinol. 14: PubMed Scopus Google Scholar). of steroid-withdrawn GR from the nucleus has been to be to and is export of liganded GR to of CRM1 (15Savory J.G. Hsu B. Laquian I.R. Giffin W. Reich T. Hache R.J. Lefebvre Y.A. Mol. Cell. Biol. 1999; 19: 1025-1037Crossref PubMed Scopus (181) Google Scholar). we that a substitution of amino of GR within NL1 not only NL1 activity but to an of GR to the cytoplasm the of steroid (15Savory J.G. Hsu B. Laquian I.R. Giffin W. Reich T. Hache R.J. Lefebvre Y.A. Mol. Cell. Biol. 1999; 19: 1025-1037Crossref PubMed Scopus (181) Google Scholar). that the GR NL1 directly or an active nuclear retention signal (NRS) for the retention of GR within the nucleus been proposed previously to be an regulatory for the receptor, and export of GR has been shown to transcriptional T. E. M. Y. Chrousos G.P. J. Biol. Chem. 2003; 278: Full Text Full Text PDF PubMed Scopus Google Scholar). such as hsp90 and the protein have been to nuclear and cytoplasmic retention of GR, respectively T. E. M. Y. Chrousos G.P. J. Biol. Chem. 2003; 278: Full Text Full Text PDF PubMed Scopus Google Scholar, K. M. T. K. Mol. Cell. Endocrinol. 2004; PubMed Scopus (44) Google Scholar). on GR for retention have not been In this study we have to the NRS activity in GR that overlaps with the receptor NL1 and to determine to GR that the GR NRS overlaps closely with NL1 and can be in of the hinge region of GR NL1 with the NLS region from the SV40 T antigen NRS activity in this substitution the activation of a mouse mammary tumor virus reporter gene by GR the of GR to the active retention of GR in the nucleus nuclear export and to an important role in the transcriptional regulatory of the Cell cells maintained in with amino and the to with or with to the from a from S. GR and between the and an and by of into the and previously (15Savory J.G. Hsu B. Laquian I.R. Giffin W. Reich T. Hache R.J. Lefebvre Y.A. Mol. Cell. Biol. 1999; 19: 1025-1037Crossref PubMed Scopus (181) Google Scholar). by amino in GR and the SV40 T antigen sequence amino into a by SV40 T antigen amino into the the GR N The substitution in the GR by and a from I. W. M. M. M. Cell. 1997; 90: Full Text Full Text PDF PubMed Scopus Google Scholar, P. J. J. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google of the SV40 hinge transcriptional activity of cells with of GR from an h after with of is to the activation of the wild-type receptor and is to for activation over with with wild-type GR, of cells a and with or of the factors in the absence or presence of of with The results shown are of transcriptional activity of cells with of wild-type and mutant GR, h after with dexamethasone as in A. with to activation of of wild-type and cells on with and the with of DNA and of after h by of h cells from h the cells treated with cortisol to nuclear localization of GR, and h to of protein over the of the FRAP, with cells in a and maintained in and over the of the on a with an and of for of cells in with as the of GR expression of as an and to the DNA to treated with dexamethasone for h and in and the a to the and with of a of and cells on in h after the cells with of DNA from for h as in Steroid accomplished by the cells for in and cells to and in and on with cells in a with a to a of in In and of and with the in the presence of for as the of the in with of in for h by in binding CRM1 and and under as previously M. M. M. Cell. 1997; 90: Full Text Full Text PDF PubMed Scopus Google Scholar, P. J. J. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). For binding of CRM1 and of with in binding for h The with binding and in cells from cells in in and by the on to and with For the of CRM1 binding to the on to a and with The and by as previously N. D. C. Hache R.J. EMBO J. 2003; PubMed Scopus Google Scholar). treated with dexamethasone for and with for with and with and Cell and and by with of of protein for h with to DBD, GR and of protein and of DNA and the for and in of to DNA and by the for the and and The of a NLS to the to GR Nuclear have previously shown that the of GR to the cytoplasm from the nucleus in cells the of steroid occurs only over h for (15Savory J.G. Hsu B. Laquian I.R. Giffin W. Reich T. Hache R.J. Lefebvre Y.A. Mol. Cell. Biol. 1999; 19: 1025-1037Crossref PubMed Scopus (181) Google Scholar). By contrast, the of a GR a substitution that NL1 within from a in which the receptor only nuclear (15Savory J.G. Hsu B. Laquian I.R. Giffin W. Reich T. Hache R.J. Lefebvre Y.A. Mol. Cell. Biol. 1999; 19: 1025-1037Crossref PubMed Scopus (181) Google Scholar). for this is that the GR NL1 nuclear although this is a that has not been previously for other the to of a strong NLS be to GR nuclear we the classical SV40 T antigen NLS sequence into a between the and the GR cytoplasmic in naive to steroid and rapidly and to the nucleus with the steroid cortisol or the synthetic steroid dexamethasone not By contrast, of steroid which to of ligand and of GR with the protein cochaperone complex (15Savory J.G. Hsu B. Laquian I.R. Giffin W. Reich T. Hache R.J. Lefebvre Y.A. Mol. Cell. Biol. 1999; 19: 1025-1037Crossref PubMed Scopus (181) Google of to the cytoplasm with the of for directly determine this of of an of nuclear we directly the nuclear export of in cells and which we have previously in R.F. Lamprecht C. Ridsdale A. Groulx I. Lee S. Lefebvre Y.A. Hache R.J. J. Biol. Chem. 2003; 278: 37858-37864Abstract Full Text Full Text PDF PubMed Scopus (44) Google to study the of between in cells to of cells in a fluorescent nuclear protein with a strong NLS that is nuclear export be to transfer from nucleus to with the of transfer of the of export in the the protein be into shown previously R.F. Lamprecht C. Ridsdale A. Groulx I. Lee S. Lefebvre Y.A. Hache R.J. J. Biol. Chem. 2003; 278: 37858-37864Abstract Full Text Full Text PDF PubMed Scopus (44) Google liganded only slowly from the nucleus to the nucleus in the presence of such that transfer h photobleaching By contrast, the liganded which as to the nucleus as an of with that of we the for the in the nucleus when to cortisol not results that the GR NL1 NLS overlaps with a nuclear retention activity that is from a strong NLS, but which the export of GR from the Furthermore, they that the NRS activity both the CRM1-dependent and nuclear export of of GR an NRS with GR NL1 is to the hinge region of the receptor that separates the DBD and LBD of the receptor (13Picard D. Yamamoto K.R. EMBO J. 1987; 6: 3333-3340Crossref PubMed Scopus (722) Google Scholar). determine this region of GR for NRS activity and to determine this activity actively nuclear we of amino of the GR hinge NL1 in the of a in which nuclear import by the SV40 NLS and nuclear export mediated by the virus CRM1-dependent nuclear export signal In the a protein the of GR sequence that hinge amino to rapidly between in the of of from that of a a not shown and 19Walther R.F. Lamprecht C. Ridsdale A. Groulx I. Lee S. Lefebvre Y.A. Hache R.J. J. Biol. Chem. 2003; 278: 37858-37864Abstract Full Text Full Text PDF PubMed Scopus (44) Google Scholar). the substitution in NL1 between with By contrast, of the protein the GR hinge of nuclear export not from the presence of two strong on this as an the SV40 NLS nuclear but in nuclear export not The reduced nuclear export of the GR to a of the interaction between the and as when in the ability to interact with in a in an to between NL1 and the NRS within the GR we within the GR hinge and the of on nuclear export within the of Two of the and to have a on NRS activity only the substitution is in which the most transfer a of transfer with the GR hinge These results that substitution NRS activity in GR, which to the of NRS activity for GR cytoplasmic in the presence of steroid and rapidly to the nucleus cortisol or dexamethasone in a that from GR, that this substitution not on the GR NL1 activity not the of nuclear export in for from that of The of to the cytoplasm of cortisol by the substitution not of GR NL1 with the SV40 NLS NRS and Transcriptional GR hinge region is an region of the GR that of the receptor DNA and ligand binding R. J. Steroid Biochem. Mol. Biol. 2005; PubMed Scopus Google Scholar). results with that NRS activity be to from NL1 by within we that a substitution the GR NL1 with strong NLS NRS activity the of a GR with nuclear localization activity to receptor and and of the DBD and the SV40 NLS in to NRS we an of the amino of GR with the sequence from SV40 in a that the of the basic amino of the The hinge region of GR between amino and a that not DNA or GR DNA binding Yamamoto K.R. PubMed Scopus Google and have shown that of the terminus of GR to not DNA binding S.M. V. P. Evans R.M. Cell. 1987; Full Text PDF PubMed Scopus Google Scholar). the as and and to the nucleus as as in response to cortisol between in a results with dexamethasone not SV40 substitution the NRS activity by the not determine this decrease in NRS activity the ability of the receptor to activate we the ability of and to activate transcription from the mouse mammary tumor virus when the in 7 cells transcriptional activation from the has been shown previously to be on a complex of that between and we shown that the substitution the GR transcriptional activation the DNA binding of GR (15Savory J.G. Hsu B. Laquian I.R. Giffin W. Reich T. Hache R.J. Lefebvre Y.A. Mol. Cell. Biol. 1999; 19: 1025-1037Crossref PubMed Scopus (181) Google Scholar). with the and incomplete nuclear transfer of to the nucleus in response to Remarkably, despite the nuclear localization of in response to steroid, this substitution reduced the transcriptional activation of the in response to dexamethasone to of the activity of GR of transcriptional activity to that with despite the reduced nuclear localization of the decrease in the of transcriptional activity not of a decrease in the of GR to the of the into the cell of GR and to the steroid with the decrease in transcriptional activation mediated by the of to the dramatically reduced in the presence of this mutant with only GR of the that the substitution in the hinge region not the for GR to interact with in the but that of the nuclear export of GR through NRS activity an important to of the of the GR in and binds to we the of of GR or on the activation of transcription In the the of GR from to an in transcription from to the of from to the activation of transcription in response to steroid from to not of of with of GR, a that in a of and a of which is the that have been the of expression of a of the two not These results suggest that GR and largely transcriptional activation when transcription as with activity over GR, which between an active nuclear form and an inactive cytoplasmic has been to have the to be regulated through of between nucleus and GR has previously been shown to be actively in the and of an that the of GR nuclear export has been to decrease GR transcriptional activation T. E. M. Y. Chrousos G.P. J. Biol. Chem. 2003; 278: Full Text Full Text PDF PubMed Scopus Google Scholar, K. M. T. K. Mol. Cell. Endocrinol. 2004; PubMed Scopus (44) Google Scholar). the signals that the active nuclear retention of such as GR remain to be In this study we that the hinge region of GR the NL1 NLS includes an active nuclear retention signal or NRS, which the retention of GR in the nucleus and nuclear protein export through the CRM1-dependent nuclear export pathway. Remarkably, of the GR NRS activity through dramatically the transcriptional activation of GR the results suggest that the GR NL1 overlaps with an NRS activity (15Savory J.G. Hsu B. Laquian I.R. Giffin W. Reich T. Hache R.J. Lefebvre Y.A. Mol. Cell. Biol. 1999; 19: 1025-1037Crossref PubMed Scopus (181) Google Scholar). we have that amino of the GR hinge region that the NLS are both and to nuclear export of reporter and to the nuclear export of nuclear the to be for NRS activity when of that NRS activity in synthetic to nuclear retention of that the GR DBD and LBD and the hinge is that the amino as the NRS to NRS activity within the of the that the substitution has a on of the NRS when into the receptor the amino in the hinge are under that the substitution within the hinge NRS activity in synthetic NL1 activity that NRS and NL1 are of Furthermore, is that most basic not with the NRS receptor or binding GR is highly in the both when liganded and ligand R.F. Lamprecht C. Ridsdale A. Groulx I. Lee S. Lefebvre Y.A. Hache R.J. J. Biol. Chem. 2003; 278: 37858-37864Abstract Full Text Full Text PDF PubMed Scopus (44) Google Scholar, M. Y. C. S. Ann. N. Y. Acad. Sci. 2004; 1024: PubMed Scopus Google Scholar, J.G. Mol. Cell. Biol. 2004; 24: PubMed Scopus Google Scholar). of the receptor mutant not that the NRS the interaction of GR with a nuclear such as the nuclear for such an interaction has been demonstrated for GR previously in cells Y. M. R. DeFranco D.B. Mol. Endocrinol. Google Scholar). the of the receptor within the nucleus not a of with such as the nuclear that form a that GR from the nuclear export is that the NRS function through interaction with a and nuclear that with the nuclear export or the of nuclear export however, the between NL1 and NRS, that NRS activity a of the through which GR with the that transfer to the The of an NRS and a that NRS activity within the the to the NRS receptor through or Substitution of an over the SV40 NLS into GR NRS but not in an export that that of be to that this substitution not the NRS that only nuclear and not be an for is that to the NRS the region or that GR NRS in the in which two The strong decrease in the transcriptional activation for have been that the localization of GR nuclear despite the in nuclear export Furthermore, the of the of transcriptional activation of GR and that of NRS activity in of GR reduced the transcriptional activation of is to that export of GR to the in the of receptor that remains predominantly results in a in the receptor that dramatically transcriptional regulatory are factors to this For the NRS an nuclear target for GR, is that this is directly important for receptor activity Yamamoto K.R. PubMed Scopus Google Scholar, J.G. Proc. Natl. Acad. Sci. U. S. A. 1990; PubMed Scopus Google Scholar). that GR and recruited to the to the that mediated through the NRS have a on the ability of GR to chromatin and transcription on the as by the decrease in and the in the presence of with has been demonstrated that of GR and factors and the of both the activation Y. Y. M. S. S. A. Y. H. M. Mol. Cell. Full Text Full Text PDF PubMed Scopus Google Scholar). that both and mutant GR recruited to the as as hormone not is that the in receptor in the of or in the of and that through the For an of active by the and the reduced be important to determine in this in GR transcriptional activation is and in of transcriptional regulation by GR, through or for are by the to the S. Lee for and

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