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(Received for publication, February 6, 1996)
From the Departments of Cellular and Molecular Pharmacology, and
Biochemistry and Biophysics, University of California, San
Francisco, San Francisco, California 94143-0448
Steroid hormones bind and activate intracellular
receptors that are ligand-regulated transcription factors. Mammalian
steroid receptors can confer hormone-dependent
transcriptional enhancement when expressed in yeast, thereby enabling
the genetic identification of nonreceptor proteins that function in the
hormone signal transduction pathway. Pdr5p (Lem1/Sts1/Ydr1p), a yeast
ATP-binding cassette transporter, selectively decreases the
intracellular levels of particular steroid hormones, indicating that
active processes can affect the passage of steroids across biological
membranes. In yeast, the immunosuppressive drug FK506 inhibited Pdr5p,
thereby potentiating activation of the glucocorticoid receptor by
dexamethasone, a ligand that is exported by Pdr5p. In mammalian L929
cells but not in HeLa cells, FK506 potentiated dexamethasone
responsiveness and increased dexamethasone accumulation, without
altering the hormone-binding properties of the glucocorticoid receptor.
We suggest that an FK506-sensitive transporter in L929 cells
selectively decreases intracellular hormone levels and, consequently,
the potency of particular steroids. Thus, steroid transporters may
modulate, in a cell-specific manner, an initial step in signaling, the
availability of hormone to the receptor.
Cells use intracellular receptors to sense and respond to
extracellular signals such as steroid, thyroid, retinoid, and vitamin D
hormones (1). Responses to these hormones can be modulated by other
signals and by cellular context. For example, activators of kinases can
potentiate or abrogate glucocorticoid action in a cell-type specific
manner (2, 3). Resistance to glucocorticoids or mineralocorticoids in
patients with wild-type receptors for these steroids further
demonstrates that factors in addition to the receptors can determine
hormone response (4, 5, 6, 7). One step at which response could be modulated
is the availability of hormone to the intracellular receptors. The
levels of hormone that can interact with and activate the receptors
could be determined by proteins affecting hormone influx, efflux, or
metabolism.
The existence of proteins that actively affect the influx or efflux of
steroid ligands has been controversial. Although steroids are small
lipophilic molecules that can diffuse freely in lipid bilayers,
corticosterone uptake into liver membrane vesicles is saturable and
produces elevated hormone concentration inside the vesicles (8).
Studies in yeast, where expression of the mammalian glucocorticoid
receptor (GR)1 supports
hormone-dependent transcriptional regulation of appropriate
reporter constructs (9, 10), have revealed a yeast
In yeast, the ABC transporter Pdr5p selectively decreases the
responsiveness of the mammalian GR to specific ligands (11). Do
mammalian cells express Pdr5-like proteins that can modulate cellular
responses to glucocorticoids? The distinct substrate selectivity of ABC
transporters that display sequence similarities in their transmembrane
regions (21, 22) discouraged us from probing for Pdr5-like proteins on
the basis of sequence homology. We therefore took a pharmacological
approach and sought drugs that might affect the function of a mammalian
steroid exporter. Given the precedent of drugs that inhibit a given
process in both mammalian and yeast cells (e.g. brefeldin A
(23)), we identified an inhibitor of the yeast Pdr5p, and then examined
its effects on steroid accumulation and response in mammalian
cells.
The macrocyclic lactone FK506 is used clinically as a potent
immunosuppressant. It interacts with a family of cellular proteins,
termed FK506-binding proteins (FKBP), and interferes with T-cell
activation by inhibiting the signaling phosphatase calcineurin
(reviewed in Ref. 24). Interestingly, FK506 has also been shown to
interact with Mdr1 (25), to be a substrate of Mdr1 (26), and to inhibit
Mdr-mediated transport, thereby reversing multidrug resistance (27,
28). We therefore tested whether FK506 might inhibit Pdr5p in yeast,
and then determined the effect of FK506 on hormone responsiveness in
mammalian cells.
Plasmids
Yeast plasmids pG1F620S and pTCA/PDR5 (previously denoted as
pTCA/LEM1) express, respectively, a point mutant (F620S) rat GR from
the constitutive yeast GPD promoter (11, 29) and Pdr5p from
its own promoter (11). Reporter plasmid p Yeast Strains and Methods
The following strains were used: YPH252 ( Liquid Hormone Binding
Yeast cells expressing GR (1.5 ml of a
culture at A600 =~ 0.8) were incubated with
[3H]Dex (1.05 Ci/mmol) at 30 °C for 2 h, in the
absence or presence of 300-fold excess unlabeled Dex, and in the
presence of 0.1% carrier ethanol or 10 µM FK506. Cells
were harvested by centrifugation (16,000 × g, 5 min at
4 °C), washed 3 times with cold PBS containing 2% glucose,
re-centrifuged, suspended in 50 µl of PBS, and counted by liquid
scintillation. Mammalian cells (1-4 × 106
cells/60-mm dish) were incubated for 2 h with labeled ligand
([3H]Dex, 10.5 Ci/mmol; [3H]TA, 11 Ci/mmol), in the absence or presence of 150-fold excess unlabeled
ligand, and in the presence of 0.1% carrier ethanol or 10 µM FK506. Cells were washed once with PBS, treated with
0.3 ml of trypsin followed by 0.5 ml of charcoal-stripped serum,
transferred to Eppendorf tubes, washed twice with PBS, harvested by
centrifugation, suspended in 50 µl of PBS, and counted by liquid
scintillation. Specific binding was determined as the counts bound in
the absence of excess unlabeled ligand minus the counts bound in its
presence.
Extracts were prepared from L929 cells or
YPH252 yeast expressing the rat F620S GR as described (11, 29) except
that: (a) cells were treated with either 0.1% carrier
ethanol or 10 µM FK506 for 2 h prior to lysis, and
(b) lysis and incubation with [3H]Dex were in
the presence of 0.1% carrier ethanol or 10 µM FK506.
Binding in yeast extracts was as described (11). L929 cell extracts
were incubated with 0.2-18 nM [3H]Dex for
6 h, in a final volume of 75 µl and at a protein concentration
of 2 mg/ml. Binding data were analyzed using a nonlinear least-squares
curve fitting program (33) with a three-parameter model (one specific
and one nonspecific binding site) for binding in yeast extracts and a
two-parameter model (one specific binding site) for binding in L929
extracts.
Immunoblots
The yeast cell extracts that were used for hormone binding were
also used to assay GR protein levels. Pdr5p, a membrane protein, was
not detectable in these soluble extracts. Thus, crude membranes were
prepared from YPH252 cells that had been treated for 2 h with
either 0.1% carrier ethanol or 10 µM FK506. Cells (15 ml
at A600 =~ 0.8) were washed with cold PBS,
resuspended in 50 µl of lysis buffer (50 mM Tris·HCl,
pH 7.5, 50 mM mannitol, 1.5 mM
MgCl2, 2.5 mM EGTA, 0.5 mM
phenylmethylsulfonyl fluoride, 1 µg each of leupeptin, aprotinin, and
pepstatin A per ml), and lysed by vortex mixing with glass beads at
4 °C for 30 min. The lysates were cleared by low speed
centrifugation (3,000 × g for 10 min), crude membranes
were pelleted by high speed centrifugation (150,000 × g for 30 min at 4 °C) and resuspended in 50 µl of lysis
buffer. Thirty µg of soluble cell extract (for GR) or crude membranes
(for Pdr5p) were separated on 7% SDS-polyacrylamide gels and
transferred to Immobilon-P membranes (Millipore). GR was detected with
the monoclonal antibody BUGR2 (34); Pdr5p was detected with a rabbit
polyclonal antibody raised against Sts1p (Pdr5p) (35) and kindly
provided by K. Kuchler. Blots were developed with alkaline
phosphatase-conjugated secondary antibodies (Bio-Rad).
Cell Culture and Transient Transfections
L929 (mouse fibroblasts) or HeLa (human cervical carcinoma)
cells grown in Dulbecco's modified Eagle's medium (DME-H21; Life
Technologies, Inc.) supplemented with 8% charcoal-stripped fetal calf
serum were plated at ~1.5 × 105 cells/35-mm dish,
12-18 h before they were transfected by the calcium phosphate
precipitation method. Briefly, 1 µg of TAT3-luc, 1 µg of 6RZ, and 3 µg of pBluescript (Stratagene) were mixed with 20 µl of 2.5 M CaCl2, 200 µl of 2 × HBS (50 mM Hepes, pH 6.95, 1.5 mM
Na2HPO4, 280 mM NaCl) and
H20 to a total volume of 400 µl, and added to the cells.
Twelve h later, the transfected HeLa cells were shocked by exposure to
15% glycerol in DME-H21 for 1 min, washed twice with PBS lacking
calcium and magnesium (L929 cells were not glycerol shocked, and were
washed three times with PBS lacking calcium and magnesium), and
incubated for an additional 24 h with fresh DME-H21 media
supplemented with 5% charcoal-stripped serum and containing varying
concentrations of hormone (Dex or TA), and 0.1% carrier ethanol or 10 µM FK506. Cells were then harvested, and luciferase and
To determine whether FK506 can inhibit the yeast Pdr5p, we tested
the effect of FK506 on GR responsiveness to Dex, an agonist that is
exported by Pdr5p, and to deoxycorticosterone, an agonist that is
unaffected by Pdr5p (11), in yeast that do or do not have functional
Pdr5p. Wild-type yeast (Pdr5p+) and pdr5-101
cells (Pdr5
A prediction of Pdr5 inhibition by FK506 is that FK506 treatment should
increase the intracellular accumulation of Dex in cells that express
Pdr5p, without altering the Dex-binding properties of GR. We therefore
assayed the effect of 10 µM FK506 on Dex binding in whole
cells and in extracts of yeast that express GR. FK506 increased the
levels of Dex binding in wild-type cells, rendering them similar to the
levels seen in cells that lack Pdr5p (Fig.
2A). The increased levels of Dex binding
could not be attributed to altered GR properties, as FK506 did not
increase the levels of GR protein (Fig. 2B) or the affinity
of GR for Dex (Table I). Moreover, FK506 treatment had no
effect on Dex accumulation in pdr5-101 cells that express
GR (Fig. 2A). In summary, FK506 treatment of wild-type yeast
rendered the cells phenotypically Pdr5
Dex-binding properties of GR, in the absence or presence of FK506
Volume 271, Number 29,
Issue of July 19, 1996
pp. 17152-17156
©1996 by The American Society for Biochemistry and Molecular Biology, Inc.
and
ABSTRACT
INTRODUCTION
EXPERIMENTAL PROCEDURES
RESULTS AND DISCUSSION
FOOTNOTES
Acknowledgments
REFERENCES
TP-
inding
assette (ABC)
transporter, Pdr5p (Lem1/Sts1/Ydr1p), that decreases intracellular
accumulation of particular steroids (11), and of other non-steroid
drugs (12, 13, 14). The identification of a steroid exporter in yeast
raised the possibility that similar transporters might modulate steroid
potencies in mammalian cells. In support of this notion, L929 mouse
fibroblasts have been reported to export cortisol and dexamethasone
(Dex) by a saturable, energy-dependent,
temperature-sensitive process (15). Furthermore, several studies in
mammalian cells suggest that overexpressed Mdr1, a mammalian
ABC-transporter that can confer multidrug resistance (reviewed in Ref.
16), can also transport particular steroids (17, 18, 19, 20). The nature of the
activity that exports steroids from L929 cells and the role of putative
mammalian steroid transporters, including Mdr1, in hormonal regulation
have not been yet determined. Indeed, control of steroid transport
across the cell membrane has not been considered as a regulatory step
that, together with other mechanisms (e.g. ligand metabolism
and transcription factor interactions), may contribute to the cell-type
specificity of steroid hormone action.
s26x (29) contains three
glucocorticoid response elements upstream of a minimal yeast
CYC1 promoter driving the Escherichia coli lacZ
gene. The mammalian reporter TAT3-Luc contains three glucocorticoid
response elements upstream of a minimal alcohol dehydrogenase promoter
driving the luciferase gene (30). Plasmid 6RZ contains the E. coli lacZ gene under the control of the Rous sarcoma virus
LTR.
,PDR5)
(31), YNK100 (
,pdr5-101 (previously referred to as
lem1-1)) (11), and YNK102 (
,
pdr5::LEU2) (11). Cells were transformed by a
standard lithium acetate protocol (32).
-Gal Assays
-gal assays were performed as described (29). Briefly,
yeast carrying the plasmids pG1F620S and p
s26x were grown to
saturation in selective media, diluted 1:10 in fresh media containing
hormone, FK506 (kindly provided by Fujisawa U. S. A.) or carrier
ethanol, and grown for an additional 12 h at 30 °C. Enzyme
activity was determined as described (29). FK506 was not toxic to cells
under any of the experimental conditions tested.
-Gal activity were assayed as described (30).
) were transformed with a GR expression
plasmid and a GR-responsive LacZ-reporter plasmid. As shown in Fig.
1A, FK506 potentiated the response of GR to
Dex in wild-type cells, rendering it similar to the response in
pdr5-101 cells. FK506 had no effect on GR response to Dex
in pdr5-101 cells or to deoxycorticosterone in either
wild-type or pdr5-101 cells (Fig. 1B),
indicating that FK506 altered neither the transcriptional regulatory
properties of GR nor the general permeability properties of yeast
membranes. Dosage studies revealed that 1 µM FK506
increased Dex responsiveness to ~50% of that seen in cells lacking
Pdr5p, and that 10 µM FK506 produced almost maximal
induction by 1 µM Dex (Fig. 1C). At all
concentrations, FK506 had no effect on the GR response to Dex in
pdr5-101 cells. Importantly, introduction of a plasmid
expressing Pdr5p in pdr5-101 cells restored both reduced
responsiveness to Dex in the absence of FK506 and potentiation of the
Dex response by FK506 (Fig. 1C). We conclude from these
results that FK506 acts by counteracting the function of Pdr5p,
i.e. export of Dex.
Fig. 1.
Effect of FK506 on GR function in yeast.
A and B, effect of 10 µM FK506 on
GR responsiveness to hormone in wild-type and pdr5-101
yeast. YPH252 (wild-type) and YNK100 (pdr5-101) cells
carrying the GR expression plasmid pG1F620S and the reporter plasmid
p
s26x were treated for 12 h with the indicated concentrations
of Dex (A) or deoxycorticosterone (DOC)
(B) in the presence of 0.1% carrier ethanol (open
symbols) or 10 µM FK506 (closed symbols),
and assayed for
-Gal activity, as described under ``Experimental
Procedures.'' Data are the mean and range of results from two
independent transformants from a given experiment and are
representative of three or more experiments. Squares,
wild-type; circles, pdr5-101 yeast.
C, effects of various doses of FK506 on Dex responsiveness.
YPH252 (wild type) and YNK100 (pdr5-101) cells carrying
pG1F620S, p
s26x, and a control vector, and YNK100 cells carrying
pG1F620S, p
s26x, and a Pdr5p expression plasmid
(pdr5-101 + Pdr5p) cells were treated for 12 h with
the indicated concentrations of FK506 and 1 µM Dex, and
assayed for
-Gal activity, as described under ``Experimental
Procedures.''
, suggesting that
FK506 inhibits export of Dex by Pdr5p. Interestingly, FK506 modestly
increased Pdr5p protein levels, suggesting that it inhibits the
activity and not the expression of Pdr5p (Fig. 2B).
Fig. 2.
Effect of FK506 on Dex binding in
vivo, and on protein levels of GR and Pdr5p. A, FK506
effect on Dex accumulation in wild-type and pdr5-101 yeast.
Specific binding of Dex in whole cells was measured, as described under
``Experimental Procedures,'' in yeast that carried the GR expression
plasmid pG1F620S and that had been preincubated for 2 h with
either 0.1% carrier ethanol (no FK506, black bars) or 10 µM FK506 (10 µM FK506, hatched
bars). No specific binding was seen in cells that do not express
GR. B, protein levels of GR and Pdr5p. GR (indicated by the
arrow) and Pdr5p (indicated by the bracket) were
detected in 30 µg of total protein of either soluble extracts (GR) or
crude membranes (Pdr5p) from yeast that expressed GR and that were
treated for 2 h with either 0.1% carrier ethanol (lanes 1, 3, and 5) or 10 µM FK506 (lanes
2 and 4). Lanes 1, 2, 4, and 5,
YPH252 (wild-type) yeast; lane 3, YNK102
(Pdr5
) yeast.
Cell
type
GR fmol/mg
Kd(nM)
Wild-type
yeast
3.96 ± 0.18
73 ± 19
Wild-type yeast + FK506
4.60 ± 0.80
83 ± 23
L929
1.40
± 0.21
276 ± 33
L929 + FK506
1.52 ± 0.01
329
± 32
We next determined the effect of FK506 on glucocorticoid responsiveness
in mammalian cells. L929 and HeLa cells were transiently transfected
with a reporter plasmid containing a GR-responsive luciferase gene, and
treated with the GR agonists Dex or triamcinolone acetonide (TA), in
the absence or presence of FK506. Luciferase activity in extracts from
transfected cells was used to monitor hormone response. In agreement
with a previous report (36), FK506 treatment of L929 cells potentiated
the response of the endogenous GR to Dex (Fig.
3A). However, we found that potentiation was
both ligand specific and cell-type specific: FK506 affected Dex but not
TA activity in L929 cells (Fig. 3A), whereas it altered
neither Dex nor TA activity in HeLa cells (Fig. 3B).
Furthermore, FK506 did not potentiate response to deoxycorticosterone,
a weak GR agonist in mammalian cells, in L929 or HeLa cells (data not
shown).
-Gal expression plasmid 6RZ for
monitoring transfection efficiency. Transfected cells were treated for
24 h with the indicated concentrations of Dex or TA, in the
presence of 0.1% carrier ethanol (no FK506, filled bars) or
10 µM FK506 (hatched bars), in 5%
charcoal-stripped serum. Luciferase activity was determined in extracts
from transfected cells, as described (30). Data are the mean and range
of two to five independent experiments, with two or more replicates
each.
If FK506 potentiates GR responsiveness to Dex by inhibiting Dex export,
then FK506 should increase Dex accumulation in L929 cells without
affecting GR protein levels or affinity for Dex. Indeed, FK506 induced
substantially higher levels of [3H]Dex binding in L929
cells (Fig. 4A): FK506 treatment decreased
from ~60 nM to ~3 nM the concentration of
extracellular Dex required to achieve 50% occupancy of the endogenous
GR. At the same time, FK506 altered neither the affinity of GR for Dex
in extracts of L929 cells nor the levels of GR protein (Table I).
Interestingly, the Kd of the GR-Dex interaction in
L929 extracts (1.5 nM) was similar to the Dex concentration
required for 50% occupancy of GR in the presence of FK506 in whole
cells (~3 nM), suggesting that FK506 inhibits an activity
that reduces Dex availability to GR in whole cells, but not in cell
extracts. Consistent with the view that FK506 increases the
intracellular accumulation of Dex in L929 cells and not the affinity of
GR for Dex, we also observed that nonspecific retention of Dex in L929
cells was increased 5-10-fold by FK506 (data not shown). Finally, as
predicted from the hormone response profiles in Fig. 3, FK506 had no
affect on accumulation of [3H]Dex in HeLa (Fig.
4B; 50% GR occupancy in HeLa cells was achieved at ~3
nM Dex) or on accumulation of [3H]TA in L929
and HeLa cells (Fig. 4C).
Taken together, our results indicate that L929 but not HeLa cells contain an FK506-sensitive activity that reduces the intracellular accumulation of Dex, thereby decreasing its potency, but does not affect TA. Formally, FK506 might be stimulating a Dex import activity, or triggering metabolism of Dex to a more active species. However, based on the functional similarity of the L929 activity to the yeast Pdr5p transporter, we suggest that a Pdr5p-like ABC transporter exports Dex from L929 cells, and that the mammalian exporter, like Pdr5p, is inhibited by FK506. Furthermore, our results imply that mammalian ABC transporter(s) may function in modulating cellular responsiveness to steroids.
ABC transporters comprise a superfamily of prokaryotic and eukaryotic proteins that can import or export a wide range of substrates, from small molecules like vitamins, nutrients, or drugs to polypeptides such as bacterial toxins (37). What transporter might operate on Dex in L929 cells? Potential candidates include various well characterized mammalian ABC transporters (e.g. Mdr1 (also called Mdr1b P-glycoprotein), Mdr3 (also called Mdr1a P-glycoprotein), and MRP (38)), less well characterized members such as Spgp (39) or an unknown novel transporter. Among the characterized candidates, Mdr1 and Mdr3 have been shown to transport specific steroids (17, 18, 19, 20) and overexpressed Mdr1 can export Dex efficiently enough to affect GR response (19). We thus probed L929 membranes with the anti-Mdr1 antibody C219 (Signet Laboratories, MA) that recognizes Mdr1, Mdr3, and Spgp (16, 39). We failed to detect a cross-reacting protein (data not shown), suggesting that if one or more of the above transporters are expressed, they are at levels lower than those in adrenal Y1 cells, which we used as a positive control. If indeed low levels of Mdr1, Mdr3, or Spgp cause the dramatic decrease in intracellular Dex accumulation in L929 cells, then they must export Dex very efficiently. Given that there are several candidate transporters and many steroids, it is also possible that multiple transporters act on steroids, each with a different steroid selectivity. Interestingly, the yeast Pdr5p and the putative transporter in L929 cells show overlapping but distinct substrate selectivity, as Pdr5p, but not the L929 activity, can reduce the cellular levels of TA (11).
As mammalian ABC transporters can function in yeast (40, 41), it may be
possible to clone the L929 Dex exporter by complementation of the
Pdr5
phenotype. We have expressed the human Mdr1 in yeast
and shown that it confers resistance to valinomycin as previously
reported (40). However, Mdr1 does not affect GR response to Dex in
Pdr5+ or Pdr5
cells,2 suggesting that Mdr1 may not export
Dex efficiently, at least in yeast.
The stimulation of Dex activity in L929 cells by FK506 has been reported previously (36, 42) and interpreted as a direct effect on GR, as unliganded GR resides in a hetero-oligomeric aporeceptor complex that includes FKBP-59, an FK506-binding immunophilin and peptidylprolyl isomerase (43, 44). However, FK506 has little or no effect on the affinity of GR for Dex or TA (42, 45), and the effects of FK506 analogs on Dex responsiveness do not correlate with their inhibition of peptidylprolyl isomerase (FKBP-59) or calcineurin activity (42). FK506 also potentiates the response of progesterone receptor in yeast to the steroid R5020, an effect that was attributed to enhanced steroid-induced receptor phosphorylation by sub-saturating levels of R5020 (46). If, however, FK506 inhibits R5020 export, the increase in phosphorylation might instead reflect elevated intracellular R5020 levels. It will be interesting to test whether R5020 is transported by Pdr5p or by another yeast ABC transporter. In summary, our experiments support strongly the idea that FK506 alters the availability of intracellular steroids without affecting the GR per se, and that it acts in a ligand- and cell type-specific manner.
By extension, our findings imply that responses to steroids and signals for other intracellular receptors may similarly be modulated at the level of plasma membrane transport. In an accompanying study, Ribeiro et al. (48) suggest that intracellular thyroid hormone levels and response are subject to similar controls; they find that verapamil, another inhibitor of Mdr proteins, potentiates thyroid hormone responsiveness in cells that express proteins of the Mdr family, and decreases thyroid hormone efflux from Mdr-expressing cells, primary hepatocytes, and cardiocytes (48).
In principle, hormone transporters could protect specific cells in particular developmental or physiological states from the effects of high hormone concentrations (e.g. during stress or pregnancy). If the transporter itself were under hormonal regulation, it could be used to shape the duration and magnitude of a hormone response. Deregulation of the levels or activity of such a transporter might lead to steroid hormone resistance and disease. In addition, steroid transporters may render hormone responses sensitive to non-steroid molecules, such as FK506, that can affect the activity of the transporters. These findings suggest approaches to cell- and/or ligand-selective modulation of hormone effects that could prove useful in the design of drug therapies. Notably, FK506 is commonly used in conjunction with glucocorticoids for immune suppression following organ transplants. Although high levels of FK506 are required to potentiate Dex effects in L929 or yeast cells relative to those administered to patients (47), it is conceivable that some drug interactions may occur in certain cells in this regimen. In any case, isolation and characterization of mammalian steroid transporters will allow an assessment of their role in ligand- or cell type-specificity of steroid action.
Fellow of the Leukemia Society of America.
-Gal,
-galactosidase; FKBP,
FK506-binding protein.
We thank members of the Yamamoto laboratory and U. Muller for helpful discussions; K. Kuchler for sharing reagents prior to publication; I. Bekersky of Fugisawa U. S. A. for their gift of FK506; J. D. Baxter, L. Z. Benet, S. Bohen, R. Cavalieri, R. Grosschedl, I. Herskowitz, J. Lefstin, N. Lomri, D. Pearce, R. Ribeiro, B. F. Scharschmidt, and D. B. Starr for their comments on the manuscript; and R. Ribeiro for communication of results prior to publication.
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D. Knutti, A. Kaul, and A. Kralli A Tissue-Specific Coactivator of Steroid Receptors, Identified in a Functional Genetic Screen Mol. Cell. Biol., April 1, 2000; 20(7): 2411 - 2422. [Abstract] [Full Text] |
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J. J. LaPres, E. Glover, E. E. Dunham, M. K. Bunger, and C. A. Bradfield ARA9 Modifies Agonist Signaling through an Increase in Cytosolic Aryl Hydrocarbon Receptor J. Biol. Chem., February 25, 2000; 275(9): 6153 - 6159. [Abstract] [Full Text] [PDF] |
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R. R. Cavalieri, L. A. Simeoni, S. W. Park, J. D. Baxter, B. F. Scharschmidt, R. C. J. Ribeiro, and N. Lomri Thyroid Hormone Export in Rat FRTL-5 Thyroid Cells and Mouse NIH-3T3 Cells Is Carrier-Mediated, Verapamil-Sensitive, and Stereospecific Endocrinology, November 1, 1999; 140(11): 4948 - 4954. [Abstract] [Full Text] |
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L. A. Carver, J. J. LaPres, S. Jain, E. E. Dunham, and C. A. Bradfield Characterization of the Ah Receptor-associated Protein, ARA9 J. Biol. Chem., December 11, 1998; 273(50): 33580 - 33587. [Abstract] [Full Text] [PDF] |
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S. Le Bihan, V. Marsaud, C. Mercier-Bodard, E.-E. Baulieu, S. Mader, J. H. White, and J.-M. Renoir Calcium/Calmodulin Kinase Inhibitors and Immunosuppressant Macrolides Rapamycin and FK506 Inhibit Progestin- and Glucocorticosteroid Receptor-Mediated Transcription in Human Breast Cancer T47D Cells Mol. Endocrinol., July 1, 1998; 12(7): 986 - 1001. [Abstract] [Full Text] |
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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] |
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R. Egner, F. E. Rosenthal, A. Kralli, D. Sanglard, and K. Kuchler Genetic Separation of FK506 Susceptibility and Drug Transport in the Yeast Pdr5 ATP-binding Cassette Multidrug Resistance Transporter Mol. Biol. Cell, February 1, 1998; 9(2): 523 - 543. [Abstract] [Full Text] |
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S.-Y. Chen, J. Wang, W. Liu, and D. Pearce Aldosterone responsiveness of A6 cells is restored by cloned rat mineralocorticoid receptor Am J Physiol Cell Physiol, January 1, 1998; 274(1): C39 - C46. [Abstract] [Full Text] [PDF] |
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W. B. Pratt and D. O. Toft Steroid Receptor Interactions with Heat Shock Protein and Immunophilin Chaperones Endocr. Rev., June 1, 1997; 18(3): 306 - 360. [Abstract] [Full Text] |
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Y. Mahe, Y. Lemoine, and K. Kuchler The ATP Binding Cassette Transporters Pdr5 and Snq2 of Saccharomyces cerevisiae Can Mediate Transport of Steroids in Vivo J. Biol. Chem., October 11, 1996; 271(41): 25167 - 25172. [Abstract] [Full Text] [PDF] |
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