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Department of Molecular and Integrative Physiology (G.L., L.N.P., A.M.N., B.S.K.) and Department of Cell and Structural Biology (T.R.E., B.S.K.) University of Illinois Urbana, Illinois 61801
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ABSTRACT |
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 TOP ABSTRACT INTRODUCTIONRESULTSDISCUSSIONMATERIALS AND METHODSREFERENCES |
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INTRODUCTION |
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TOPABSTRACTINTRODUCTIONRESULTSDISCUSSIONMATERIALS AND METHODSREFERENCES |
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Numerous studies have focused on different aspects of the process by which the receptor is transformed from an inactive state in the absence of ligand to an activated state upon ligand exposure. However, this process is still not fully understood. In its inactive form, the receptor is associated with a number of other proteins (including at least hsp90, hsp70, and p23), forming a multiprotein complex with a sedimentation constant of 8–9 S (9, 10, 11). In the unliganded state, receptors for thyroid hormone (TR) or for retinoids [retinoic acid receptor (RAR) and retinoid X receptor (RXR)] can have an inhibitory effect on transcription and are associated with corepressors (12, 13, 14, 15, 16, 17). Upon hormone exposure, most of the associated proteins are released, and the receptor appears as a 4–5 S sedimenting complex (18, 19). In this state, the receptor is able to dimerize and to associate with coactivators (20, 21, 22, 23, 24, 25, 26, 27). The receptor is also known to undergo changes in its phosphorylation state (28, 29) and conformation (30, 31).
The aim of this study is to understand what kind of changes in function and conformation the receptor undergoes when it is converted to the active state. To address these questions, we have used the wild type ER and a set of three constitutive ER mutants that correspond to amino acid substitutions at residue 380 or residue 537 (32, 33). These mutants show constitutive activity in the absence of estradiol (E2) ranging from 20 to nearly 100% of the activity of wild type receptor in the presence of E2. By comparing the properties of wild type and these mutant receptors in the absence and in the presence of estrogens or antiestrogens, we have analyzed their ability to interact with several known coactivators and members of the preinitiation complex (PIC). Moreover, the conformation of these receptors was studied by limited proteolytic digest experiments. Finally, we analyzed the ability of these receptors to bend ERE-containing DNA, which is thought to reflect in part the transcriptional ability of transcription factors (34, 35, 36). Altogether, these results demonstrate that, upon hormone exposure, the wild type receptor undergoes major changes in its conformation and in its properties, whereas no effect or only limited effects of hormone are observed with the constitutively active receptors, as these appear to have already undergone those changes that render them ligand-independent transcriptional activators.
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RESULTS |
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TOPABSTRACTINTRODUCTIONRESULTSDISCUSSIONMATERIALS AND METHODSREFERENCES |
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Transcriptional activity of the wild type or mutant ERs was monitored
in the presence and absence of hormone with estrogen-responsive CAT
reporter genes containing a minimal (TATA) or a complex (pS2) promoter.
As shown in Fig. 1
, in the absence of ligand, the Y537S
ER exhibited constitutive activity 65–90% that of the
estrogen-occupied ER, the level of activity being dependent on the
promoter used, while the Y537A and E380Q mutant ERs exhibited
constitutive activity in the absence of ligand about 15% to 30% that
of wild type activity in the presence of E2. Moreover, all
three receptors were as potent as wild type ER in activating
transcription in the presence of E2. The magnitude of
constitutive activity of the ERs shown in Fig. 1 is consistent with our
earlier findings (32, 33). Referring to the structure of the related
nuclear receptors hRAR
and rTR
1, for which crystallographic
information is available (37, 38), the Y537 residue would be in helix
12, containing the hormone-dependent activation function (AF-2) of
nuclear receptors, and the E380 residue would be at the end of helix 4
of the ligand-binding domain. [Note: The helix numbering is from the
RAR structure (37).]
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The Y537S receptor showed a distinct interaction with each of these
three coactivators. In the absence of E2, this mutant
interacted with SRC-1 as strongly as did wild type receptor in the
presence of E2; this interaction in the absence of
E2 was not significantly enhanced by the addition of
E2, but was completely suppressed by TOT. The Y537S ER
showed a moderate but readily detected interaction with TIF-1 in the
absence of E2, and again this interaction was suppressed by
TOT. However, as opposed to the interaction of Y537S ER with SRC-1,
which is already maximal in the absence of E2, the binding
of TIF-1 to the Y537S ER was markedly enhanced by addition of
E2. Y537S ER shows yet a third pattern of interaction with
the coactivator ERAP-140. There is a small interaction in the absence
of E2, which was easily seen with longer times of
autoradiography (data not shown), whereas for the same exposure times,
no interaction was observed with either the unliganded wild type
ER or any of the other constitutively active receptors (Fig. 2
). Moreover, as with TIF-1, the binding of ERAP-140 to the Y537S
ER was greatly increased by addition of E2.
Therefore, the fully constitutively active mutant ER, Y537S, showed a different extent of ligand-independent interaction with the three coactivators: full interaction with SRC-1, some interaction with TIF-1, and little interaction with ERAP-140. Occupancy with E2 was needed to achieve maximal interaction with TIF-1 and ERAP-140.
Concerning Y537A and E380Q mutant ERs, both receptors showed the same type of interaction with SRC-1, namely a weak interaction in the absence of E2, which was greatly enhanced by addition of E2. The interaction in the absence of ligand (control vehicle, C) was abolished by incubation in the presence of TOT. Both the Y537A and E380Q receptors required E2 for interaction with TIF-1 or ERAP-140, but even with E2 treatment, the interaction of the E380Q mutant with ERAP-140 was much lower than that observed for E2-occupied wild type receptor.
Mutant and Wild Type ERs Interact in a Ligand-Independent Manner
with the PIC
The interaction of the ER with the PIC could be another step of
the transcriptional process involved in the activation. Transcription
factor II B (TFIIB) and TATA-box binding protein (TBP) are two members
of the PIC that have been shown to be the target of numerous
transcription factors. We therefore expressed these factors by in
vitro translation and used them in pull-down experiments to test
their interactions with our ER proteins. The wild type ER and the three
mutants showed a much stronger interaction with TBP compared with
TFIIB, but for both the mutant ERs and the wild type ER, interaction
with TFIIB and with TBP occurred in the absence or presence of
E2 (Fig. 3). Among the three mutant
receptors, the Y537S ER displayed the greatest ability to interact with
TBP, being comparable to that of wild type ER. There was less
difference in the degree of interaction of all four receptors with
TFIIB.
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The Y537S ER mutant, which shows nearly full constitutive activity in the absence of ligand, showed a proteolytic digestion pattern in both its unliganded or liganded state (compare lanes 9–12 with 13–16), which closely resembled the pattern seen for wild type ER treated with E2. Also, the transiently stabilized band at 35 kDa (arrow C), which in wild type ER is stabilized only in the presence of E2 (lane 7), is strongly stabilized in Y537S ER without or with E2 (lanes 11 and 15). There is, however, a small amount of protein present in the lower/inactive form (arrow A) in the absence of hormone (lanes 11 and 12), which shifts to the upper/active form (arrow B) when E2 is added. Thus, the proteolytic digest pattern for the Y537S receptor reflects its constitutively active state.
The unoccupied E380Q and Y537A ERs, which possess partial constitutive activity, showed a slight enhancement of the transiently stabilized, high molecular mass bands, including the approximately 40-kDa band D and the 35-kDa band C, relative to the unoccupied wild type ER, especially noticeable at a trypsin concentration of 5 µg/ml (compare lanes 11 and 19 vs. 3). Perhaps these subtle differences are indicative that the unliganded E380Q and Y537A receptors have taken on a partially active conformation. However, E2 treatment resulted in a predominance of the B species after increasing trypsin treatment, as observed with the wild type E2-occupied receptor.
Transcriptionally Active Wild Type and Mutant ERs Induce Similar
DNA-Bending Angles
Wild type and mutant ERs were expressed in a reticulocyte lysate
system in the presence or absence of estrogen and used in DNA-phasing
analysis experiments to determine the orientation and magnitude of
ER-induced DNA bending (Fig. 5). The unoccupied and estrogen-occupied
in vitro translated ERs were incubated with
32P-labeled DNA fragments, each of which contained an
intrinsic DNA bend separated from a consensus ERE by either 26, 28, 30,
32, 34, or 36 bp (41). Thus, when the ER bound to the ERE and induced
DNA bending, the intrinsic and ER-induced DNA bends would either be in
phase and form a larger DNA bend or be out of phase and have the effect
of straightening the DNA fragment. When wild type and mutant ERs were
incubated with DNA fragments containing 26 or 36 bp between the
intrinsic and ER-induced DNA bends, there was an increase in the
mobilities of the receptor-DNA complexes through the acrylamide gel,
indicating that the ER-induced and intrinsic DNA bends were in phase.
When the wild type and mutant ERs were incubated with DNA fragments
containing 32 bp between the intrinsic and ER-induced DNA bends, there
was an increase in the mobilities of the receptor-DNA complexes
indicating that the ER induced and intrinsic DNA bends were out of
phase. Because the intrinsic and ER-induced DNA bends were on the same
side of the DNA helix when they were out of phase (assuming 10.5
bp/helical turn), these findings demonstrate that the ER-induced DNA
bend opposes the intrinsic DNA bend, which is directed toward the minor
groove of the DNA helix (42). Therefore, the unoccupied and
estrogen-occupied wild type and mutant ERs induced DNA bends that were
directed toward the major groove of the DNA helix. The magnitudes of
the ER-induced DNA bends were determined from replicate phasing
analysis experiments (see Materials and Methods) and are
summarized in Table 1. The unoccupied wild type ER
induced a DNA bend of 15.6°, the largest bending angle measured. In
contrast, the estrogen-occupied wild type ER induced a much smaller
bend of 7.3°. Interestingly, the three mutant receptors, Y537S,
Y537A, and E380Q, which are active in the presence and in the absence
of hormone, had statistically similar directed DNA-bending angles of
approximately 7.5°-9°, in the absence or presence of hormone. Thus,
the mutant receptors induced directed bending angles that were similar
in magnitude to the bend induced by the estrogen-occupied ER. These
findings are in good agreement with previous studies carried out with
estrogen-occupied wild type and mutant ERs that had been expressed in
COS cells (43) and support the idea that transcriptionally active wild
type and mutant ERs induce directed bending angles of similar
magnitude.
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DISCUSSION |
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TOPABSTRACTINTRODUCTIONRESULTSDISCUSSIONMATERIALS AND METHODSREFERENCES |
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Figure 6 presents a model summarizing our findings
regarding the interaction of several coregulators and basal
transcription factors with the wild type and constitutively active
Y537S ER and the effects of E2 on these interactions and on
DNA bending by these receptors. The figure emphasizes that occupancy of
the wild type ER by E2 elicits a reduction in the extent of
DNA bending and an increase in the association with coactivators. The
Y537S ER associates strongly with some, but not all, coactivators in
the ligand-unoccupied state, and it exhibits the same DNA bend angle in
its ligand-free or E2-occupied state as does the
E2-occupied wild type receptor.
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). The partially constitutively active receptors
showed liganded ER character in some assays but not others. Of the
three ER coregulators evaluated, SRC-1 interaction correlated best with
the degree of transcriptional activity displayed by the mutant ERs.
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The receptor proteolytic digestion pattern and receptor coregulator
association profiles only appear to clearly indicate the "active"
character of the most constitutively active mutant ER Y537S; the other
two receptors showed subtle differences in these assays consistent with
some change toward the active state observed with the
E2-liganded wild type ER, but overall the E380Q and Y537A
receptors resembled wild type ER. Although DNA bending did not
distinguish between partially and fully constitutively active
receptors, it appears to be very sensitive to the propensity of the
receptor to be in the active conformation. By contrast, the protease
digestion pattern differences may require that receptor be very
strongly in the active conformation, as the hormone-occupied wild
type-like ER pattern was only observed with the most constitutively
active ER, Y537S. The fact that both partially active and fully
constitutively active receptors gave the same directed DNA-bending
angle suggests that the DNA may form a scaffold for the accumulation of
other protein factors and coregulators important in determining
receptor transcriptional activity, with DNA bending being the first of
several important steps leading to full receptor transcriptional
effectiveness. Interaction of the ER with basal factors did not require
ligand, and it is therefore not surprising that interaction with TFIIB
and TBP does not discriminate wild type from constitutively active
ERs. The wild type ER was able to interact with the three coactivators tested (ERAP140, TIF-1, and SRC-1) in the presence of E2, but not in the presence of antiestrogens such as TOT. The strongest interaction was observed with SRC-1 and TIF-1 and to a lesser extent with ERAP140. In the presence of E2, the three constitutive mutants displayed the same ability to interact with these coactivators as the wild type ER. However, in the the absence of E2, these mutants exhibited different abilities to interact with the coactivators. Indeed, the Y537S mutant was able to interact with SRC-1 in the absence of E2 to the same extent as the wild type receptor in the presence of E2. Moreover, this interaction was not enhanced by addition of E2. With the two other coactivators, the interaction with Y537S ER was still markedly enhanced by (TIF-1), or dependent on, ligand (ERAP 140). Concerning E380Q and Y537A mutants, they showed basically a common pattern of interaction with the coactivators: no interaction in the absence of E2 with ERAP140 or TIF-1 and only a weak interaction with SRC-1 in the absence of ligand. Our observations suggest that SRC-1, TIF-1, and ERAP-140 play somewhat different roles in ER activity and may be involved to different degrees in the process of receptor-regulated transcription. That the activity of the mutants correlated well with binding to SRC-1 implies that SRC-1 is likely a functional mediator of ER transcriptional activity. The interaction of the three mutant receptors with coactivator in the absence of ligand was abolished by addition of trans-hydroxytamoxifen, suggesting that antiestrogen induces a different conformation of the receptor that is not compatible with the interaction with coactivator. Our data are in agreement with the model proposed by Chen and Evans (13), i.e. that the receptor bound to DNA could be in three states: in the absence of hormone, the receptor is inactive (or might be repressive as in the case of TR and RAR, which interact with corepressors). Once the ligand is added, the receptor is in an intermediary state, and basal transcription can take place. And finally, the liganded receptor is able to interact with coactivators, which would be a link with the PIC, and to activate strongly the transcription process.
The mechanism by which the nuclear receptors activate transcription remains unclear, but it is proposed that the receptors could stimulate PIC formation either by recruiting the different members of the PIC or by positioning a preformed PIC on the DNA. The PIC consists of at least seven basal transcription factors, namely TFIIA, TFIIB, TFIID [comprised of the TBP interacting with TBP-associated factors, TFIIE, TFIIF, TFIIH, and TFIIJ (for review, see Ref. 44)]. A number of transcription factors, including nuclear receptors and, in particular, the ER, have been shown to interact in vitro with TFIID (45, 46, 47, 48) or TFIIB (49, 50, 51, 52, 53). We have now investigated the interaction of the wild type and mutant ERs with members of the PIC. Basically, all the ERs tested, whether wild type or constitutively active, interacted with TBP and TFIIB in the absence or presence of hormone. However, of note was the fact that TBP interaction, which was substantial in the absence of estrogen but was enhanced by E2 in the case of wild type receptor, was not increased by E2 for mutant receptors. These data suggest that the ability of a receptor to interact with the PIC is not a specific feature of activated receptors. However, it is possible that the interaction of inactive receptors (i.e. unoccupied wild type ER) with the PIC could be nonproductive. The transformation of the receptor into an active state would then not lead to an increase of the interactions but rather to a modification of the nature of these interactions.
To further investigate the conformational changes that could arise from
treatment with estrogen, we performed limited proteolytic digestion
experiments. Using the wild type receptor, we present evidence that the
conformation of the receptor in the absence and in the presence of
E2 are different, which is in agreement with previous work
(30, 54). Indeed, in the absence of ligand, the receptor was highly
sensitive to trypsin and gives a proteolytic digestion pattern in
which, for high concentrations of trypsin, two stable bands (
25 and
28 kDa) of approximately equal intensity appeared. Treatment with
E2 stabilized strongly the upper, 28-kDa band. The
transformation of the receptor into a distinct, active conformation
upon hormone exposure has been reported not only for ER but also for
progesterone receptor and RAR/RXR (30, 31), suggesting that this is a
phenomenon common to the ligand-activatable nuclear receptors. Of note,
the fully constitutively active Y537S ER exhibited a proteolytic
digestion pattern in the absence of E2, which was very
similar to the pattern of the active wild type estrogen-occupied ER,
suggesting that this mutant receptor was in the conformationally active
state.
If the receptor can undergo conformational changes after activation, it
is possible that these changes could modify the nature of its
interactions with DNA. It has been reported by several groups that the
binding of transcription factors to DNA can induce bending of DNA
(34, 35, 36, 41, 43, 55, 56, 57). A change in the bending state of the DNA
could reflect a difference in the ability of the receptor to activate
transcription. Our results show that the unliganded wild type receptor
induced a larger bend (16°) and that the E2-occupied
wild type receptor induced a dramatically smaller DNA bend (
7°). Of particular interest is the fact that the three consitutively
active unoccupied and estrogen-occupied ER mutants induced DNA bends
that were similar to the bending angle induced by the
E2-occupied wild type receptor. This smaller DNA bend would
be a characteristic of the active state. We propose that the distinct
conformations induced by active and inactive receptors could be
interpreted as activating and silencing signals. We should emphasize
that the difference in DNA bending observed for the active and inactive
wild type ER could be obtained using a totally cell-free system,
suggesting that cell context is not a factor in the effect of receptor
on DNA bending.
Altogether, the data from studies with these constitutively active ERs provide evidence that the transformation of receptor into an active state involves a complex set of events that include conformational changes as well as distinct alterations in the ability of the receptor to interact with coregulator proteins and with DNA. Collectively, these changes in conformation and interactions "mark" the receptor as being transcriptionally productive.
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MATERIALS AND METHODS |
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TOPABSTRACTINTRODUCTIONRESULTSDISCUSSIONMATERIALS AND METHODSREFERENCES |
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Plasmid Construction
The plasmid encoding SRC-1 (26) was kindly provided by Drs. Ming
Tsai and Bert O’Malley. The plasmid encoding TIF-1 (amino acids
434–750) (39) was a gift from Dr. Pierre Chambon. The plasmid
pGEX-2TK-ER, which contains the human ER spanning amino acids 282–595,
and the plasmid encoding ERAP140 (23) were kindly provided by Dr. Myles
Brown. The two expression vectors encoding TBP and TFIIB were kindly
provided by Dr. Danny Reinberg. The GST-ER mutant plasmids for Y537A
and Y537S were previously described (33). PCR was used to generate the
E380Q ER fragment from amino acids 282 to 595 bearing BamHI
and EcoRI sites and cloned into PGEX-2TK.
Production of GST Fusion Proteins
Bacteria expressing GST fusion proteins were grown at 37 C in
500 ml of LB (Luria Bertani) broth until the absorbance (600 nm)
reached 0.8. Then the induction was performed for 3 h at 30 C with
1 mM isopropyl ß-D-thiogalactopyranoside
(IPTG). Cells were collected by centrifugation at 4 C at 3000 x
g for 15 min. The supernatant was discarded, and the pellet
was rapidly frozen in liquid nitrogen and then kept for 1 h at
-80 C. Frozen pellets were then thawed on ice and resuspended in 0.01
volume of NET buffer (20 mM Tris, pH 8.0/100 mM
NaCl/1 mM EDTA) and sonicated twice for 30 sec at maximum
level. The suspension was centrifuged for 10 min at 12,000 x
g, and the supernatant was then transferred to other tubes
and centrifuged at 105,000 x g (30 min, 4 C). Protein
concentration was estimated by the Bradford method. The levels of
expressed fusion proteins were determined by in vitro
binding assays followed by Western analysis with H222 monoclonal
antibody.
In Vitro Translation of Receptor-Associated Proteins
and Human ER Proteins
In vitro translation was performed using the TNT
Promega kit (Promega, Madison, WI). Briefly, 1 µg of expression
vector was mixed with 25 µl TNT rabbit reticulocyte lysate, 2 µl
TNT buffer, 1 µl of mix containing all amino acids except methionine,
1 µl RNAsin (50 U/µl), 1 µl T3 RNA polymerase (20 U/µl), and 4
µl of [35S]methionine (15 µCi/µl). The final
reaction volume was 50 µl. The reaction was performed for 1.5 h
at 30 C. The translation efficiency was checked by loading 1 µl of
lysate on an SDS-PAGE gel.
For gel mobility shift assays, the translation was performed in the presence of control vehicle (0.1% ethanol) or 1 µM E2 as above, except that labeled methionine was replaced with unlabeled methionine.
In Vitro Binding Assays with Glutathione
Sepharose
Glutathione Sepharose (Pharmacia, Piscataway, NJ) was
equilibrated with IP binding buffer (25 mM Tris-HCl (pH
7.9), 10% vol/vol glycerol, 0.1% NP-40, 0.5 mM
dithiothreitol, 100 mM KCl). The in vitro
translated products were first precleared for 2 h by incubation
with 100 µl of beads and 300 µg of GST (which does not contain any
insert). Crude bacterial extract (500 µg) containing GST fusion
proteins was incubated at 4 C with 25 µl of beads for 2.5 h in
the presence of vehicle (0.1% ethanol) or hormone (E2 or
TOT, at 1 µM concentration). After three washes, the
beads were incubated with 5 µl of in vitro translated
product for 2.5 h in the presence of vehicle or hormone at 4 C.
The beads were washed three times with 1 ml of IP buffer and two times
with 1 ml of IP buffer containing 300 mM KCl. After
washing, beads were boiled in SDS sample buffer, and a quarter of the
proteins were run on SDS-PAGE. The gel was fixed, dried, and submitted
to autoradiography.
Protease Digestion Assays
Bluescript vector (Stratagene, La Jolla, Ca) was used for
insertion of cDNA sequences of wild type ER, E380Q, Y537A, and Y537S ER
mutants. Aliquots of in vitro translated,
[35S]-labeled proteins (25 µl) were treated with
control (0.1% EtOH) vehicle or ligand at a final concentration of
9 x 10-6 M for 20 min at room
temperature. Aliquots (5 µl) of the ligand-treated receptor were
incubated without trypsin or with trypsin to a final concentration of
1.5, 5, 15, or 25 µg/ml (Worthington Biochemicals, Freehold, NJ)
After a 10-min incubation at room temperature, the digestion was
stopped with 20 µl of Laemmli buffer, and the samples were boiled for
5 min and then separated on a 12% SDS-PAGE gel. The radiolabeled
products were visualized by autoradiography.
DNA-Bending Gel Mobility Shift Assays
The ERE-containing DNA-phasing vectors, ERE 26, ERE 28, ERE 30,
ERE 32, ERE 34, and ERE 36 (41), were digested with EcoRI
and HindIII, isolated on an acrylamide gel, and
electroeluted. The 281- to 291-bp DNA fragments containing the
intrinsic DNA bend and the ERE were filled in with Klenow in the
presence of [32P]dATP and [32P]dGTP and
then purified using a G-25 Sephadex Quick Spin column (Boehringer
Mannheim, Indianapolis, IN). Gel mobility shift assays were carried out
as previously described (55) with minor modifications. Briefly, 10,000
cpm of the 32P-labeled DNA phasing fragment was combined
with 4 µl (188 µg total protein) reticulocyte lysate-expressed wild
type or mutant ER and 1 µg poly(deoxyinosinic-deoxycytidylic)acid
(Sigma) in a buffer containing 10% glycerol, 50 mM KCl, 15
mM Tris, pH 7.9, 0.2 mM EDTA, and 0.4
mM dithiothreitol (20 µl final volume) for 15 min at room
temperature. Low ionic strength gels and buffers were prepared as
described (58). Twenty-centimeter gels were prerun for 1 h at 300
V. Samples were fractionated for 3 h on an 8% (75:1 acrylamide to
bis-acrylamide ratio) polyacrylamide gel. Water recirculation was used
to maintain the gels at 4 C. Radioactive bands were visualized by
autoradiography. The relative mobilities of the ER-DNA complexes and
free probes were quantitated with a Molecular Dynamics PhosphorImager
and Imagequant software (Molecular Dynamics, Sunnyvale, CA). The
magnitudes of the receptor-induced directed DNA bending angles
(B) were determined for the wild type and mutant ERs
using the empirical formula (59):
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C is the intrinsic DNA bending angle,
APH is the phasing amplitude, and k is a coefficient used
to adjust for electrophoretic conditions. A value of k = 0.991 was
determined by comparing the relative mobility of five sets of
DNA-bending standards with their known bending angles (60).
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ACKNOWLEDGMENTS |
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FOOTNOTES |
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This research was supported by NIH Grant R37 CA-18119 (to B.S.K.), a Susan G. Komen Foundation Postdoctoral Fellowship (to G.L.), and NIH Grant R29 HD-31299 (to A.M.N.).
Received for publication April 2, 1997. Accepted for publication May 21, 1997.
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REFERENCES |
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TOPABSTRACTINTRODUCTIONRESULTSDISCUSSIONMATERIALS AND METHODSREFERENCES |
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C. M. Bula, J. E. Bishop, S. Ishizuka, and A. W. Norman 25-Dehydro-1{alpha}-Hydroxyvitamin D3- 26,23S-Lactone Antagonizes the Nuclear Vitamin D Receptor by Mediating a Unique Noncovalent Conformational Change Mol. Endocrinol., November 1, 2000; 14(11): 1788 - 1796. [Abstract] [Full Text]
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F. Gay, I. Anglade, Z. Gong, and G. Salbert The LIM/Homeodomain Protein Islet-1 Modulates Estrogen Receptor Functions Mol. Endocrinol., October 1, 2000; 14(10): 1627 - 1648. [Abstract] [Full Text]
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D. M. Kraichely, J. Sun, J. A. Katzenellenbogen, and B. S. Katzenellenbogen Conformational Changes and Coactivator Recruitment by Novel Ligands for Estrogen Receptor-{alpha} and Estrogen Receptor-{beta}: Correlations with Biological Character and Distinct Differences among SRC Coactivator Family Members Endocrinology, October 1, 2000; 141(10): 3534 - 3545. [Abstract] [Full Text] [PDF]
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P. G. V. Martini, R. Delage-Mourroux, D. M. Kraichely, and B. S. Katzenellenbogen Prothymosin Alpha Selectively Enhances Estrogen Receptor Transcriptional Activity by Interacting with a Repressor of Estrogen Receptor Activity Mol. Cell. Biol., September 1, 2000; 20(17): 6224 - 6232. [Abstract] [Full Text]
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S. Misiti, S. Nanni, G. Fontemaggi, Y.-S. Cong, J. Wen, H. W. Hirte, G. Piaggio, A. Sacchi, A. Pontecorvi, S. Bacchetti, et al. Induction of hTERT Expression and Telomerase Activity by Estrogens in Human Ovary Epithelium Cells Mol. Cell. Biol., June 1, 2000; 20(11): 3764 - 3771. [Abstract] [Full Text]
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A. Schlegel, C. Wang, B. S. Katzenellenbogen, R. G. Pestell, and M. P. Lisanti Caveolin-1 Potentiates Estrogen Receptor alpha (ERalpha ) Signaling. CAVEOLIN-1 DRIVES LIGAND-INDEPENDENT NUCLEAR TRANSLOCATION AND ACTIVATION OF ERalpha J. Biol. Chem., November 19, 1999; 274(47): 33551 - 33556. [Abstract] [Full Text] [PDF]
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P. S. Jones, E. Parrott, and I. N. H. White Activation of Transcription by Estrogen Receptor alpha and beta Is Cell Type- and Promoter-dependent J. Biol. Chem., November 5, 1999; 274(45): 32008 - 32014. [Abstract] [Full Text] [PDF]
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F. M. Sladek, M. D. Ruse Jr., L. Nepomuceno, S.-M. Huang, and M. R. Stallcup Modulation of Transcriptional Activation and Coactivator Interaction by a Splicing Variation in the F Domain of Nuclear Receptor Hepatocyte Nuclear Factor 4alpha 1 Mol. Cell. Biol., October 1, 1999; 19(10): 6509 - 6522. [Abstract] [Full Text] [PDF]
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S. Y. Wu, M. C. Thomas, S. Y. Hou, V. Likhite, and C. M. Chiang Isolation of Mouse TFIID and Functional Characterization of TBP and TFIID in Mediating Estrogen Receptor and Chromatin Transcription J. Biol. Chem., August 13, 1999; 274(33): 23480 - 23490. [Abstract] [Full Text] [PDF]
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 M. M. Montano, K. Ekena, R. Delage-Mourroux, W. Chang, P. Martini, and B. S. Katzenellenbogen An estrogen receptor-selective coregulator that potentiates the effectiveness of antiestrogens and represses the activity of estrogens PNAS, June 8, 1999; 96(12): 6947 - 6952. [Abstract] [Full Text] [PDF]
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F. Schaufele Regulation of Estrogen Receptor Activation of the Prolactin Enhancer/Promoter by Antagonistic Activation Function-2-Interacting Proteins Mol. Endocrinol., June 1, 1999; 13(6): 935 - 945. [Abstract] [Full Text]
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 Proceedings of the British Toxicology Society Autumn Meeting University of York 20-22 September 1998 Human and Experimental Toxicology, January 1, 1999; 18(1): 46 - 65. [PDF]
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Z. Zhang and C. T. Teng Estrogen Receptor-related Receptor alpha 1 Interacts with Coactivator and Constitutively Activates the Estrogen Response Elements of the Human Lactoferrin Gene J. Biol. Chem., June 30, 2000; 275(27): 20837 - 20846. [Abstract] [Full Text] [PDF]
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S. I. Anghel, V. Perly, G. Melancon, A. Barsalou, S. Chagnon, A. Rosenauer, W. H. Miller Jr., and S. Mader Aspartate 351 of Estrogen Receptor alpha Is Not Crucial for the Antagonist Activity of Antiestrogens J. Biol. Chem., June 30, 2000; 275(27): 20867 - 20872. [Abstract] [Full Text] [PDF]
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