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Thyroid Unit, Department of Medicine Beth Israel Deaconess Medical Center and Harvard Medical School Boston, Massachusetts 02215
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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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Full-length NCoR is a ubiquitously expressed 270 kDa protein (6). A second shorter isoform, RIP13a, of unknown significance also exists (16). NCoR is a modular protein and likely possesses four separate N-terminal repressing domains, two of which appear to interact with mSin3 and the histone deacetylase RPD3, HDAC1, to mediate repression (17, 18). It has been proposed that repression results from changes in chromatin structure mediated by this complex.
NCoR binds a portion of the TR hinge region termed the CoR box (6). Mutations in this region block the ability of unliganded TR to mediate repression. Two C-terminal NCoR domains, termed the interaction domains (ID I and ID II), appear to be involved in TR binding (16, 19). NCoR also appears to mediate repression by the orphan receptor RevErb (19). Protein interaction assays revealed that the NCoR interaction domains that mediate NCoR-RevErb binding are similar to those that mediate NCoR-TR effects; however, the domain of RevErb that binds NCoR is not homologous to the CoR box. The NCoR interaction domains have been identified using yeast two-hybrid and mammalian two-hybrid systems, but their functional significance on native TREs has not yet been defined. These interactions are critical as NCoR may be able to bind a number of NHRs in solution [peroxisome proliferator-activated receptors (PPARs) and RXR] but the presence of DNA response elements appears to functionally limit the interactions between NCoR and NHRs (20).
SMRT (TRAC2) is also a modular protein. It shares close to 40% amino-acid-identity with NCoR (21). It also contains N-terminal-repressing domains that appear to function through mSin3, and C-terminal interacting domains that interact with the TR, retinoic acid receptor (RAR), and RXR in a yeast two-hybrid system (7, 17). The interacting domains also share significant homology with those of NCoR. Recently, an endogenous inhibitor of SMRT, TRAC1, has been identified (8). It appears to be an alternatively spliced variant that contains the interacting domains but lacks the repressing domains.
We have identified a human homolog of NCoR that contains the C-terminal IDs, but not the N-terminal repressor domains (22). In transient transfection experiments, this construct inhibits endogenous NCoR function and has thus been termed NCoR inhibitor, or NCoRI. This truncated inhibitor of NCoR, which may or may not be expressed endogenously, provides an ideal tool to study the functional significance of the NCoR IDs on endogenous TREs. In this study, deletion constructs of hNCoRI were made that lack one or both of the known IDs. The functional characteristics of these constructs were assessed in transient transfection experiments using both positive (direct repeat, palindromic, and inverted palindromic) and negative (TRH) reporter elements. Electrophoretic mobility shift assays were also performed to characterize the binding of NCoRI deletion constructs to TR on DNA response elements and to compare structural binding with functional interactions.
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RESULTS |
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TOPABSTRACTINTRODUCTIONRESULTSDISCUSSIONMATERIALS AND METHODSREFERENCES |
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). It contains the C-terminal
interacting domains of NCoR, but lacks the N-terminal-repressing
domains (6). Recently, others have shown that a region present in NCoRI
may, in fact, bind mSin3 and repress (18). However, we have
demonstrated that hNCoRI has little repressing activity when linked to
the Gal4 DNA-binding domain (data not shown). In transient transfection
experiments, cotransfection of hNCoRI reverses ligand-independent
repression on direct repeat (DR+4), palindromic (PAL), and inverted
palindromic (LYS) reporters, presumably by interfering with endogenous
NCoR function (22). To ensure that NCoRI possesses separate activity
from the full-length moiety, we analyzed the function of NCoRI and NCoR
on a positive and negative reporter in an identical transfection
paradigm. As is shown in Fig. 2, NCoRI
inhibits ligand-independent repression on the palindromic positive TRE,
while full-length NCoR functions to enhance ligand-independent
repression. In contrast, on the negatively regulated TRH reporter NCoRI
is a strong activator of ligand-independent activation while NCoR has
only a slight effect. However, the activation function on the negative
reporter appears to be located downstream of the repressing domains as
NCoRI is far more effective than NCoR. Thus, NCoRI is a potent
antirepressor on both the positive and negative reporters. NCoRI has no
effect on reporter gene activity in the absence of cotransfected TRß1
(data not shown).
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).
In contrast, the NCoR interacting domains are very well conserved
between species as is shown in Fig. 3B.
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); however, when equal
amounts of the full-length protein are used in EMSA on a DR+4 element,
NCoR is preferred by the TR, and little SMRT binds to either the TR
homo or heterodimer (Fig. 4B). In addition, neither NCoR nor SMRT
interacts with DNA in the absence of TR. These data suggest that either
the DNA-binding region or other regions of the TR outside its
ligand-binding domain alter the interaction between NCoR and SMRT.
Further analysis of the interaction between the TR and NCoR and SMRT
was performed in CV-1 cells. Using a mammalian two-hybrid system, where
the ligand-binding domain of TRß1 was fused to the GAL4 DNA-binding
domain and either corepressor to the VP-16 activation domain, we found
that the interaction between TR and NCoR (Fig. 4C) was extremely strong
and in fact stronger than the interaction between TR and RXR (data not
shown). The interaction between TR and SMRT was much weaker in
comparison. In contrast, Gal4-RAR
preferred to interact with SMRT
rather than NCoR, ensuring that the VP-16 SMRT construct was adequately
expressed and localized to the nucleus. Finally, we compared the
ability of a dominant inhibitor of SMRT activity termed SMRTI (TRAC 1)
(8) and NCoRI to function as antirepressors on the DR+4 reporter in
CV-1 cells as shown in Fig. 4D. SMRTI was a consistently weaker
inhibitor at all amounts transfected, in keeping with its poorer
ability to interact with the TR. Unlike what has been reported for
full-length mNCoR (24), hNCoRI acts as a potent antirepressor at all
concentrations examined on the DR+4 reporter construct and other
positive TREs (Fig. 4D).
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) so
that their function could be ascertained both functionally and
structurally on endogenous TREs. Sequence alignment between these two
regions in the hNCoRI protein disclosed only remote similarity (Fig. 3B).
Both IDs Reverse Ligand-Independent Repression on Positive TREs
To assess the function of the interacting domains on the
ligand-independent activity of the TR, hNCoRI deletion constructs and
TRß1 were transfected into CV-1 cells at a 1:1 DNA ratio. As shown in
Fig. 5A, when cotransfected with two
copies of the LYS element linked to the luciferase reporter,
transfection of TRß1 causes approximately 3-fold repression of
basal activity. Cotransfection of hNCoRI fully reverses this
repression. Likewise, when a construct lacking ID I (hNCoRI 
ID I) is
cotransfected, there is also full reversal of ligand-independent
repression. Similarly, hNCoRI ID II reverses basal repression when
cotransfected with TRß1. In contrast, hNCoRI ID I/II, which lacks
both IDs, does not reverse ligand-independent repression on the LYS
element. Fig. 5B shows similar data, using the palindromic reporter.
Again, hNCoRI reverses basal repression, as do constructs lacking
either ID. In contrast to the LYS reporter, however, deletion of either
ID I or ID II did impair somewhat the ability of the construct to
reverse TR-mediated repression. In addition, the construct lacking both
IDs again did not reverse repression.
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), hNCoRI reverses basal repression, as do
constructs lacking ID I or ID II. Unlike the PAL and LYS elements,
however, hNCoRI ID I/II is not fully deficient in its ability to
reverse basal repression, as there is still 2-fold repression. Deletion
of another 168 amino acids (hNCoR 1–302), which removes a portion of
the mSin3-binding site (18) [mNCoR amino acids (AA) 1829–1940; hNCoRI
AA 240–351], causes virtually all ligand-independent activity to be
lost.
To rule out defective nuclear localization when IDs were deleted from
NCoR, hNCoRI and the dual ID deletion construct were placed into a
green fluorescent protein (GFP) expression vector and transfected into
CV-1 cells. The cells were viewed under fluorescence microscopy 24
h after transfection to identify cellular localization of the fusion
proteins. GFP vector alone exhibits diffuse staining in both the
nuclear and cytoplasmic compartments. In contrast, hNCoRI ID
I/II-GFP was restricted to the nucleus (Fig. 5D), suggesting that lack
of functional activity in transient transfection assays was not the
result of impaired nuclear localization. In addition, hNCoRI-GFP also
exhibited nuclear localization.
NCoR IDs Bind TRß1 on TREs and Preferentially Bind TR
Homodimer
We next tested the ability of these hNCoRI constructs to interact
with the TR on DNA response elements using EMSAs. As shown in Fig. 6A, wild-type TRß1 homodimer interacts
strongly with hNCoRI. The addition of RXR (or RXRß) causes NCoRI
binding to be reduced without altering the position of the band. The
addition of an RXR antibody supershifts the TR-RXR heterodimer. The
NCoR-TR complex is not supershifted by the anti-RXR antibody,
however, suggesting that this complex does not contain RXR. In
contrast, an anti-TR antibody supershifts all complexes, including
TR-NCoR (Fig. 6B). An antibody against C/EBP does not shift any
complex, indicating that these antibody supershift assays are specific.
These data suggest that the NCoRI complex contains TR, but not RXR, and
that NCoRI preferably binds to the TR homodimer over the TR-RXR
heterodimer in this assay. Similar studies were performed using
bacterially expressed NCoR-interacting domains and, even in the
presence of limiting TRß1, the homodimer preferred to interact with
NCoR (Fig. 6C). When the amount of GST-NCoR was increased by up to
50-fold (20 ng to 1000 ng) there was no difference found in the
preference of binding.
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construct to affect both silencing by GAL4-TR and its
interactions with NCoR. Cotransfection of GAL4-RXR with GAL4-TR will
cause a heterodimer to form on the upstream activating sequence (UAS)
response element in contrast to the TR homodimer when GAL4-TR is
transfected alone. As is shown in Fig. 6D, cotransfected GAL4-RXR
causes both a loss of silencing (GAL4-TR alone causes between 5- and
10-fold repression of basal activity) and diminishes considerably the
ability of the TR to interact with the VP-16 NCoR construct. These
results are consistent with the GAL4-TR homodimers avidly binding
corepressor to mediate silencing. As the TR homodimer is shifted by
NCoR in EMSA, we next investigated whether overexpressed NCoR could
promote homodimerization of the TR in solution. Collingwood et
al. (25) have previously shown that TR homodimers do not form in
solution in the mammalian two-hybrid system. This is demonstrated in
Fig. 6E as GAL4-TR is unable to interact with a VP-16-TR chimeric
protein in CV-1 cells. We next asked whether increased amounts of NCoR
could enhance the homodimer interaction. Indeed, cotransfected NCoRI
allowed the TR homodimer to form in solution, suggesting that the
homodimer may be stabilized by corepressor binding. In contrast,
cotransfected SMRTI had no affect on TR homodimer formation, again
supporting the preference of the TR for NCoR.
Since NCoRI appears to preferentially interact with the TR homodimer,
the ability of the NCoRI deletion constructs to bind TRß1 on DNA
response elements was assessed principally using the homodimer in EMSA.
As shown in Fig. 7A, on the LYS element,
TRß1 bound well as a homodimer. The addition of hNCoRI results in a
supershift, representing TRß1-hNCoRI binding. Constructs lacking
either ID alone bind TR on the LYS element, although these complexes
migrate slightly further than TR-NCoRI, due to their smaller size. In
contrast, constructs lacking both IDs do not bind TRß1 on the LYS
element. However, on the PAL (Fig. 7B) and DR+4 (Fig. 7C) elements,
although either interacting domain promoted TR binding, there appeared
to be a preference for the ID II-containing construct. Constructs
lacking either ID were unable to bind these elements.
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as
discussed above and shown in Fig. 8A.
Similar results are seen with constructs lacking either ID I or ID II,
suggesting that these IDs do not differentially bind TR-RXR
vs. TR-TR on DNA response elements. Fig. 8B shows similar
data in the context of 10 nM T3. As expected,
there is loss of NCoR binding to TR in the
presence of ligand, as well as a decrease in the homodimeric complex
(26, 27). There is no significant binding of NCoRI to TR-RXR
heterodimers in the presence of T3.
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, cotransfection of hNCoRI with TRß1
resulted in an approximately 7- to 8-fold increase in
ligand-independent activation. Cotransfection of hNCoRI ID I or
ID II resulted in a similar stimulation of activity. In contrast,
cotransfection of hNCoR ID I/II only stimulated activity 2-fold
above baseline. These data suggest that either ID is able to
functionally interact with TR on negative TREs.
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DISCUSSION |
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TOPABSTRACTINTRODUCTIONRESULTSDISCUSSIONMATERIALS AND METHODSREFERENCES |
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We have initially compared the ability of the TR to interact with each of the corepressors. Our data demonstrate that the TR prefers to interact with NCoR on TREs and in mammalian cells. The EMSA data suggest that conformation on a DNA-response element may be important for this specificity. While others have shown interaction between SMRT and TR homodimers in EMSA, these studies were performed in the presence of significantly greater amounts of corepressor (20). The contrast between the GST-TR pull-down results, which showed equal binding to SMRT and NCoR, and the mammalian two-hybrid experiments, which showed an overwhelming preference by the TR for NCoR, is striking as both systems employed only the TRß1 ligand-binding domain. These experiments suggest that other nuclear proteins may influence the interactions between nuclear receptors and corepressors. Given the recent cloning of a novel corepressor, SunCoR (36), it is reasonable to hypothesize that other such proteins may influence the formation of corepressor complexes in cells. The preference of the TR for NCoR is supported by recent antibody microinjection studies, which demonstrate that inhibition of NCoR blocks silencing by the TR while anti-SMRT antibodies had no effect on TR-mediated silencing (37).
In transient transfection experiments, NCoR enhances the repressing function of TR. However, when increasing amounts of NCoR are transfected, paradoxical activation has been reported to occur with localization of transfected NCoR, but not endogenous NCoR, to nuclear dot structures (24). This complexity was addressed in the present study using NCoRI, which inhibits endogenous NCoR function, and thus provides an ideal tool for examining the function of the NCoR-interacting domains. Moreover, NCoRI progressively reversed ligand-independent repression on a positive TRE without any paradoxical effects on TR action. Furthermore, full-length NCoR did not activate when transfected in an identical paradigm.
When tested in cotransfection experiments, deletion of either
interacting domain in the context of NCoRI had a negligible effect on
the inhibitory activity of NCoRI on each of the three positive TREs
tested. Furthermore, constructs containing a single ID retained the
ability to bind to the TR on these TREs in EMSA, although ID II
appeared to promote a stronger interaction. Comparison of the NCoR- and
SMRT-interacting domains shows that ID I is quite similar while ID II
is more dissimilar. Thus, the increased avidity of ID II for the TR is
consistent with the preference of NCoR for the TR. Functional activity
of NCoRI was abolished on the PAL and LYS reporters when both IDs were
deleted. This NCoRI construct (hNCoRI ID I/II) localized to the
nucleus when fused to GFP, implying that this functional defect was not
due to defective nuclear transport. When examined in EMSA, hNCoRI ID
I/II was unable to bind to TR on any of the response elements studied.
Therefore, the lack of functional activity of this construct on the PAL
and LYS elements results from its inability to bind to the TR on DNA.
In contrast, hNCoRI ID I/II retained a small but consistent
inhibitory function on the DR+4 reporter without evidence of DNA
binding, raising the possibility that the region remaining of NCoR may
interact with TR to mediate repression on DR+4, or that the
mSin3-binding site present in this construct may be important in
mediating repression on this TRE. In fact, deletion of a portion of the
mSin3 ID resulted in further loss of NCoRI function on the DR+4
element.
In contrast to ligand-independent repression on positive TREs, the TR
causes ligand-independent activation in tissue culture on negative
TREs. While the mechanism underlying negative regulation has not been
defined, it is clear that TR-binding sites exist in the regulatory
regions of genes negatively regulated by the TR (28). We have
previously demonstrated that cotransfected NCoRI enhances
ligand-independent activation on the TRH promoter in the presence of
TRß1 and TR1 (22). Deletion of either interacting domain does not
impair the ability of hNCoRI to activate the TRH promoter. However,
once both interacting domains are deleted, enhancement of
ligand-independent activation is all but lost. Interestingly
full-length NCoR also slightly activated the TRH promoter, as has been
reported previously (38), but the effect was far less than that seen
with NCoRI. These differences may be the result of the different cell
types used as well as the amounts of cotransfected NCoR or NCoRI.
Further studies will be needed to define the mechanism by which the
corepressor influences negative regulation. The data presented here
indicate that a direct interaction between the TR and NCoR is needed to
enhance ligand-independent activity.
In the present study, we have also addressed the ability of hNCoRI to
interact with either the TR homodimer of TR/RXR heterodimer. It is
unclear in vivo what is the preferred TR configuration in
the absence of ligand. Previous work by Zamir et al. (20)
has demonstrated that the TR homodimer and TR/RXR heterodimer can both
bind nuclear corepressor on a DR+4 element (20). These studies did not
address the relative preference of homo- or heterodimer for the
corepressors. In addition, it is also clear from another study (39)
that RXR/TR1 heterodimers can interact in cells with NCoR. The data
presented here also show an interaction between NCoR and RXR/TRß1
heterodimers in the mammalian two-hybrid assay. However, this
interaction is considerably weaker than the interaction with TR
homodimers. Furthermore, in the mammalian two-hybrid system, RXR-TR
heterodimers also silence less well. In addition, NCoRI appears to
allow the TR homodimer to form in solution through its interacting
domains. Further support for the preference of the homodimer for NCoR
comes from gel-mobility shift assays on TREs, which demonstrate that
the TRß1 homodimer binds in vitro translated NCoR and
bacterially expressed NCoR, while the heterodimer demonstrates little
avidity for NCoR. These data support the concept that the homodimer may
be able to play a role in thyroid hormone action through its strong
interaction with nuclear corepressors. While our data do not rule out
that the heterodimer is an active silencer, they suggest that the
homodimer preferentially interacts with each of the NCoR interacting
domains. An alternative explanation could be that the gel-mobility
shift assay is too insensitive to detect in vivo
interactions between the TR/RXR heterodimer and NCoR and that
interaction studies are cell specific. Further work will be necessary
to clearly define the role of the in vivo configuration of
TR when ligand-independent repression occurs and to determine the exact
stoichiometry of the homodimer and heterodimer interactions.
In summary, we have demonstrated that the two separate NCoR domains interact with the TR on both positive and negative TREs. Furthermore, individual IDs can mediate dominant inhibition of endogenous NCoR activity, suggesting that the separate IDs are powerful mediators of ligand-independent interaction with the TR. The IDs are remarkably conserved across species; however, the two IDs themselves share only remote homology with each other. While the two IDs may be present to ensure strong interactions with TR, our data suggest that either ID alone allows NCoR-TR interactions to occur on native TREs. Further work on these modular domains will help delineate the specific amino acids necessary for ligand-independent interactions with the TR and will help identify the important differences that allow the TR to interact much more strongly with NCoR. This may allow for the identification of other such proteins that can interact with the TR to mediate its ligand-independent functions.
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MATERIALS AND METHODS |
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TOPABSTRACTINTRODUCTIONRESULTSDISCUSSIONMATERIALS AND METHODSREFERENCES |
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encompasses AA 201–462 of
human RXR in frame with the GAL4 DNA-binding domain. GST-TR and
AASV-TR were created by ligating the ligand-binding domain of the TR
from Gal4-TR in frame with the GST moiety in PGEX4T1 and the VP-16
moiety in AASV, respectively. GST-NCoR was created by ligating the two
interacting domains of hNCoR (AA 2063–2300) into PGEX 4T1. Constructs
were cloned into pKCR2 for transient transfection experiments and pEGFP
(CLONTECH, Palo Alto, CA) for cellular localization studies. The
integrity of each plasmid was confirmed by restriction endonuclease
digestion and dideoxy sequencing. All plasmids for transfection were
purified using column (Qiagen, Chatsworth, CA) purification.
Cell Culture and Transient Transfection
CV-1 cells were maintained in DMEM supplemented with
L-glutamine, 10% FCS, 100 µg/ml penicillin, 0.25 µg/ml
streptomycin, and amphotericin.
Transient transfections using NCoRI alone were performed using the calcium phosphate technique in six-well plates with each well receiving 1.7 µg of reporter and 80 ng of pKCR2-TR and/or equal amounts of pKCR2 alone and/or equal amounts of pKCR2-NCoRI construct, except as noted. Each well also received 20 ng of a ß-galactosidase expression vector to control for transfection efficiency. Studies involving the full-length murine NCoR included 330 ng of expression vector per well. Fifteen to 18 h after transfection, the cells were washed in PBS, and refed with DMEM with 10% steroid hormone-depleted FBS. To remove steroid and thyroid hormones, FBS was treated for 24 h at 4 C with 50 mg/ml activated charcoal (Sigma Chemical Co., St. Louis, MO) and 30 mg/ml anion exchange resin (type AGX-8, analytical grade, Bio-Rad, Richmond, CA). Forty to 44 h after transfection, the cells were harvested in extraction buffer and assayed for both luciferase and ß-galactosidase (Tropix, Bedford, MA) activity. Experiments were performed in triplicate and repeated two to three times. The data are the pooled results ± SEM.
Transient transfections to assess cellular localization of hNCoR constructs were performed in 100-mm tissue culture dishes. Twenty micrograms of pEGFP-hNCoRI construct or pEGFP alone were transfected using the calcium phosphate technique. Twenty four hours later, cells were washed in PBS and refed with DMEM. Cells were then visualized under fluorescence microscopy.
EMSAs
Gel-mobility shift assays were performed with proteins derived
either from a coupled in vitro translation/transcription
reaction in reticulocyte lysate (Promega, Madison WI) or from
Escherichia coli as GST-fusion proteins. Either TRß1 or
RXR (0.5 to 4 µl) with equal amounts of NCoRI or SMRTI construct
were used, as well as 50,000–150,000 cpm of a
32P-radiolabeled DR+4 (5'-AGGTCACAGGAGGTCA-3'), palindromic
(5'-AGGTCATGACCT-3'), F2-LYS (5'-TGACCCCAGCTGAGGTCA-3'),
oligonucleotide probe. When employed, 20–1000 ng of GST-NCoR were
used. Incubations were carried out at room temperature, and complexes
were resolved on 5% nondenaturing acrylamide gels, followed by
autoradiography. The concentration of T3 used was 10
nM. EMSA with antibody supershifts (anti-RXR, Santa Cruz
Biotechnology, Santa Cruz, CA; anti-TR, Affinity Bioreagents, Inc.,
Golden, CO; anti-C/EBP, Santa Cruz Biotechnology) were carried out
similarly, except that in vitro translated proteins and
probe were incubated at room temperature for 20 min, followed by
addition of 1 µl antibody. The resulting complexes were incubated for
another 20 min at room temperature and then resolved on a 5%
nondenaturing gel, followed by autoradiography.
GST-Protein Binding Assays
GST-TR was expressed in BL21 E. coli expressing
thioredoxin (41) by induction with 0.1 MM
isopropylthio-ß-D-galactosidase (IPTG) at 30 C. GST-NCoR
was expressed in DH5 E. coli under similar conditions.
The proteins were isolated by lysis with lysozyme and purified on
Sepharose beads. Verification of protein synthesis was obtained on
SDS-PAGE. GST-TR (50 µl) and equivalent amounts of GST alone were
incubated with 6 µl of [35S]methionine-labeled in
vitro translated NCoRI or SMRTI. The concentration of
T3 used was 10-6 M. After
extensive washing, the bound proteins were eluted by boiling in
SDS-PAGE loading buffer and run on SDS-PAGE.
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ACKNOWLEDGMENTS |
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FOOTNOTES |
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This work was supported by NIH Grants to A.N.H. (5DK 02354) and F.E.W. (3DK49126).
Received for publication April 28, 1998. Revision received June 9, 1998. Accepted for publication July 10, 1998.
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REFERENCES |
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TOPABSTRACTINTRODUCTIONRESULTSDISCUSSIONMATERIALS AND METHODSREFERENCES |
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K. A. Temple, R. N. Cohen, S. R. Wondisford, C. Yu, D. Deplewski, and F. E. Wondisford An Intact DNA-binding Domain Is Not Required for Peroxisome Proliferator-activated Receptor {gamma} (PPAR{gamma}) Binding and Activation on Some PPAR Response Elements J. Biol. Chem., February 4, 2005; 280(5): 3529 - 3540. [Abstract] [Full Text] [PDF]
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Q. Wang, J. A. Blackford Jr., L.-N. Song, Y. Huang, S. Cho, and S. S. Simons Jr. Equilibrium Interactions of Corepressors and Coactivators with Agonist and Antagonist Complexes of Glucocorticoid Receptors Mol. Endocrinol., June 1, 2004; 18(6): 1376 - 1395. [Abstract] [Full Text] [PDF]
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T. Fujita, Y. Kobayashi, O. Wada, Y. Tateishi, L. Kitada, Y. Yamamoto, H. Takashima, A. Murayama, T. Yano, T. Baba, et al. Full Activation of Estrogen Receptor {alpha} Activation Function-1 Induces Proliferation of Breast Cancer Cells J. Biol. Chem., July 11, 2003; 278(29): 26704 - 26714. [Abstract] [Full Text] [PDF]
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A. Makowski, S. Brzostek, R. N. Cohen, and A. N. Hollenberg Determination of Nuclear Receptor Corepressor Interactions with the Thyroid Hormone Receptor Mol. Endocrinol., February 1, 2003; 17(2): 273 - 286. [Abstract] [Full Text] [PDF]
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N. Shibusawa, A. N. Hollenberg, and F. E. Wondisford Thyroid Hormone Receptor DNA Binding Is Required for Both Positive and Negative Gene Regulation J. Biol. Chem., January 3, 2003; 278(2): 732 - 738. [Abstract] [Full Text] [PDF]
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 N. Vasudevan, S. Ogawa, and D. Pfaff Estrogen and Thyroid Hormone Receptor Interactions: Physiological Flexibility by Molecular Specificity Physiol Rev, October 1, 2002; 82(4): 923 - 944. [Abstract] [Full Text] [PDF]
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S. Cheng, S. Brzostek, S. R. Lee, A. N. Hollenberg, and S. P. Balk Inhibition of the Dihydrotestosterone-Activated Androgen Receptor by Nuclear Receptor Corepressor Mol. Endocrinol., July 1, 2002; 16(7): 1492 - 1501. [Abstract] [Full Text] [PDF]
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 X. Li and D. P. McDonnell The Transcription Factor B-Myb Is Maintained in an Inhibited State in Target Cells through Its Interaction with the Nuclear Corepressors N-CoR and SMRT Mol. Cell. Biol., June 1, 2002; 22(11): 3663 - 3673. [Abstract] [Full Text] [PDF]
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J. C. Ghosh, X. Yang, A. Zhang, M. H. Lambert, H. Li, H. E. Xu, and J. D. Chen Interactions that determine the assembly of a retinoid X receptor/corepressor complex PNAS, April 30, 2002; 99(9): 5842 - 5847. [Abstract] [Full Text] [PDF]
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Y. Yamamoto, O. Wada, M. Suzawa, Y. Yogiashi, T. Yano, S. Kato, and J. Yanagisawa The Tamoxifen-responsive Estrogen Receptor alpha Mutant D351Y Shows Reduced Tamoxifen-dependent Interaction with Corepressor Complexes J. Biol. Chem., November 9, 2001; 276(46): 42684 - 42691. [Abstract] [Full Text] [PDF]
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R. N. Cohen, S. Brzostek, B. Kim, M. Chorev, F. E. Wondisford, and A. N. Hollenberg The Specificity of Interactions between Nuclear Hormone Receptors and Corepressors Is Mediated by Distinct Amino Acid Sequences within the Interacting Domains Mol. Endocrinol., July 1, 2001; 15(7): 1049 - 1061. [Abstract] [Full Text] [PDF]
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P. M. Yen Physiological and Molecular Basis of Thyroid Hormone Action Physiol Rev, July 1, 2001; 81(3): 1097 - 1142. [Abstract] [Full Text] [PDF]
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X. Hu, Y. Li, and M. A. Lazar Determinants of CoRNR-Dependent Repression Complex Assembly on Nuclear Hormone Receptors Mol. Cell. Biol., March 1, 2001; 21(5): 1747 - 1758. [Abstract] [Full Text]
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P. Webb, C. M. Anderson, C. Valentine, P. Nguyen, A. Marimuthu, B. L. West, J. D. Baxter, and P. J. Kushner The Nuclear Receptor Corepressor (N-CoR) Contains Three Isoleucine Motifs (I/LXXII) That Serve as Receptor Interaction Domains (IDs) Mol. Endocrinol., December 1, 2000; 14(12): 1976 - 1985. [Abstract] [Full Text]
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U. B. Kaiser, L. M. Halvorson, and M. T. Chen Sp1, Steroidogenic Factor 1 (SF-1), and Early Growth Response Protein 1 (Egr-1) Binding Sites Form a Tripartite Gonadotropin-Releasing Hormone Response Element in the Rat Luteinizing Hormone-{beta} Gene Promoter: an Integral Role for SF-1 Mol. Endocrinol., August 1, 2000; 14(8): 1235 - 1245. [Abstract] [Full Text]
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R. N. Cohen, A. Putney, F. E. Wondisford, and A. N. Hollenberg The Nuclear Corepressors Recognize Distinct Nuclear Receptor Complexes Mol. Endocrinol., June 1, 2000; 14(6): 900 - 914. [Abstract] [Full Text]
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K. Busch, B. Martin, A. Baniahmad, J. A. Martial, R. Renkawitz, and M. Muller Silencing Subdomains of v-ErbA Interact Cooperatively with Corepressors: Involvement of Helices 5/6 Mol. Endocrinol., February 1, 2000; 14(2): 201 - 211. [Abstract] [Full Text]
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C. Oberste-Berghaus, K. Zanger, K. Hashimoto, R. N. Cohen, A. N. Hollenberg, and F. E. Wondisford Thyroid Hormone-independent Interaction between the Thyroid Hormone Receptor beta 2 Amino Terminus and Coactivators J. Biol. Chem., January 21, 2000; 275(3): 1787 - 1792. [Abstract] [Full Text] [PDF]
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 C. K. Glass and M. G. Rosenfeld The coregulator exchange in transcriptional functions of nuclear receptors Genes & Dev., January 15, 2000; 14(2): 121 - 141. [Full Text]
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V. Perissi, L. M. Staszewski, E. M. McInerney, R. Kurokawa, A. Krones, D. W. Rose, M. H. Lambert, M. V. Milburn, C. K. Glass, and M. G. Rosenfeld Molecular determinants of nuclear receptor-corepressor interaction Genes & Dev., December 15, 1999; 13(24): 3198 - 3208. [Abstract] [Full Text]
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E. A. Nillni, C. Vaslet, M. Harris, A. Hollenberg, C. Bjorbak, and J. S. Flier Leptin Regulates Prothyrotropin-releasing Hormone Biosynthesis. EVIDENCE FOR DIRECT AND INDIRECT PATHWAYS J. Biol. Chem., November 10, 2000; 275(46): 36124 - 36133. [Abstract] [Full Text] [PDF]
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Y. Wu, B. Xu, and R. J. Koenig Thyroid Hormone Response Element Sequence and the Recruitment of Retinoid X Receptors for Thyroid Hormone Responsiveness J. Biol. Chem., February 2, 2001; 276(6): 3929 - 3936. [Abstract] [Full Text] [PDF]
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S. M. Yoh and M. L. Privalsky Transcriptional Repression by Thyroid Hormone Receptors. A ROLE FOR RECEPTOR HOMODIMERS IN THE RECRUITMENT OF SMRT COREPRESSOR J. Biol. Chem., May 11, 2001; 276(20): 16857 - 16867. [Abstract] [Full Text] [PDF]
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J. C. Ghosh, X. Yang, A. Zhang, M. H. Lambert, H. Li, H. E. Xu, and J. D. Chen Interactions that determine the assembly of a retinoid X receptor/corepressor complex PNAS, April 30, 2002; 99(9): 5842 - 5847. [Abstract] [Full Text] [PDF]
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N. Nasrin, S. Ogg, C. M. Cahill, W. Biggs, S. Nui, J. Dore, D. Calvo, Y. Shi, G. Ruvkun, and M. C. Alexander-Bridges DAF-16 recruits the CREB-binding protein coactivator complex to the insulin-like growth factor binding protein 1 promoter in HepG2 cells PNAS, September 12, 2000; 97(19): 10412 - 10417. [Abstract] [Full Text] [PDF]
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