A Testis-specific Androgen Receptor Coregulator That Belongs to a
Novel Family of Nuclear Proteins*
Anu-Maarit
Moilanen
,
Ulla
Karvonen
,
Hetti
Poukka
,
Wei
Yan§,
Jorma
Toppari§,
Olli A.
Jänne
¶, and
Jorma J.
Palvimo
From the
Department of Physiology, Institute of
Biomedicine, and the ¶ Department of Clinical Chemistry,
University of Helsinki, FIN-00014 Helsinki, Finland and the
§ Departments of Physiology and Pediatrics, University
of Turku, Kiinamyllynkatu 10, FIN-20520 Turku, Finland
 |
ABSTRACT |
We have characterized a novel partner for
androgen receptor (AR), termed ARIP3, that interacts with the
DNA-binding domain/zinc finger region of AR and is predominantly
expressed in the testis. Rat ARIP3 is a nuclear protein comprising 572 amino acids. It modulates AR-dependent but not basal
transcription, suggesting that ARIP3 acts as an AR transcriptional
coregulator. Except for the C-terminal AR-interacting domain, ARIP3
contains distinct regions that are also present in two recently
described proteins, a protein inhibitor of activated Stat3 and an RNA
helicase II-interacting protein (Gu/RH-II binding protein). Conserved
structural features of these proteins indicate the existence of a gene
family involved in the regulation of various transcription factors.
Collectively, ARIP3 belongs to a novel nuclear protein family and is
perhaps the first tissue-specific coregulator of androgen receptor.
 |
INTRODUCTION |
Receptors for androgens, glucocorticoids, progesterone, and
mineralocorticoids all recognize the same DNA response elements, and
very little is known about the mechanisms governing the
hormone-specific transcription of target genes in vivo by
these receptors (1, 2). Obvious ways to achieve steroid-specific gene
activation include cell-specific expression of the receptor protein and
differential availability of the ligand. In many instances, these may
not suffice, and tissue-specific coregulatory proteins have been
suggested to add an important level of control to ensure that
appropriate responses to hormonal signals are achieved. It has been
somewhat unexpected that most of the numerous nuclear hormone receptor coactivators identified to date are expressed in a ubiquitous fashion
and bind to the same conserved activation function-2
(AF-2)1 located in the C
terminus of the ligand-binding domain (LBD) (3, 4). There is, however,
increasing evidence for the DNA-binding domain (DBD)/zinc finger region
(ZFR) of nuclear receptors to function as a specific interaction
interface for coregulatory proteins (5-11). We report herein the
identification and characterization of a novel, testis-specific
androgen receptor (AR)-interacting protein that recognizes the ZFR of
AR and modulates AR-dependent transcription.
 |
EXPERIMENTAL PROCEDURES |
Materials--
pCMV
and the following mammalian two-hybrid
system vectors were purchased from CLONTECH: pM for
the DNA-binding domain of the Saccharomyces cerevisiae Gal4,
pVP16 for the transcriptional activation domain (VP16 AD) of VP16, and
pVP16-CP for a fusion of VP16 AD to polyoma virus coat protein.
pARE2-TATA-LUC, pPB(
285/+32)-LUC, pSG5rAR, and Gal4 DBD
fusion vectors of AR have been described (8, 12, 13). The yeast
two-hybrid system vectors, pVP16 (14) and pLex-a, which is based on
pBTM116 (15), were kindly provided by Dr. Stanley M. Hollenberg (Oregon
Health Sciences University, Portland, OR).
Yeast Two-hybrid System--
pLex-DBD expressing LexA fused to
hAR ZFR (residues 554-644) was used as a bait to screen a mouse embryo
E10.5 cDNA library (a gift from S. M. Hollenberg) for
interacting proteins as described previously (7, 8, 14).
Isolation of Full-length ARIP3 cDNA and Construction of ARIP3
Expression Plasmids--
A rat testis
ZapII cDNA library
(Stratagene) was screened with 32P-labeled ARIP3 cDNA
corresponding to residues2
443-548 using standard methods (16). The BLAST program (17) was used
to search for sequence homologies in the data bases at the National
Center for Biotechnology Information (National Institutes of Health).
pFLAG-ARIP3(1-572), pFLAG-ARIP3(11-572), and pFLAG-ARIP3
467-542 were constructed by cloning PCR-generated cDNA fragments into pFLAG-CMV-2 (Kodak). pH6-ARIP3-ID was obtained by inserting
a PCR-generated fragment corresponding to ARIP3(443-548) that encodes the interaction domain of ARIP3 into pQE31 (Qiagen). The mammalian two-hybrid expression vectors pVP16-ARIP3(443-548),
pVP16-ARIP3(1-442), and pVP16-ARIP3(1-572) were created by cloning
appropriate PCR-generated cDNA inserts into pVP16. Correctness of
all constructs was verified by sequencing using the ALFExpress system
(Amersham Pharmacia Biotech).
RNA Preparation and Northern Blot Analysis--
RNA was isolated
from adult rat tissues, enriched for poly(A)+ RNA and
processed as described previously (8). Membrane was hybridized to
32P-labeled ARIP3 cDNA fragment corresponding to
residues 443-548 (ARIP3-ID), washed at high stringency (0.2 × SSC, 0.1% SDS, 52 °C), and subjected to autoradiography. Human
multiple tissue Northern blot and human RNA master blot (both from
CLONTECH) were hybridized to the same probe and
processed according to the manufacturer's instructions.
Cell Culture and Transfections--
CV-1 and COS-1 cells were
maintained as described (8, 12). The cells (2.3 × 105
or 6.6 × 104) were plated on 6- or 12-well plates,
respectively, and transfected 24 h later using indicated amounts
of vectors and FuGene transfection reagent (Boehringer Mannheim).
18 h after transfection, the medium was changed to one containing
charcoal-stripped 2% (v/v) fetal bovine serum with or without
testosterone. Luciferase and
-galactosidase activities were assayed
as described previously (12), and the light units were normalized by
the
-galactosidase activity. For each experiment, the activity
indicated in the figure legend was set as 100, and other data were
calculated relative to this value.
Protein-Protein Interaction in Vitro--
Protein-protein
affinity chromatography using purified GST-ZFR (21) or GST alone bound
to glutathione-Sepharose (Amersham Pharmacia Biotech), and
[35S]methionine-labeled in vitro translated
ARIP3 was performed as described (7, 18). Bound proteins were released
in SDS-polyacrylamide gel electrophoresis sample buffer, resolved by
electrophoresis under denaturing conditions, and visualized by fluorography.
Antibody Production, Immunocytofluorescence, and
Immunohistochemistry--
Bacterially expressed
H6-ARIP3(443-548) was purified by chromatography on
nickel-nitrilotriacetic acid-agarose (Qiagen), and polyclonal antisera
were raised in rabbits. To study subcellular distribution of ARIP3,
CV-1 cells were transfected with 1 µg of pFLAG-ARIP3(11-572) as
described (7, 8). Cells were processed, and ARIP3 was visualized using
anti-ARIP3 antiserum (1:500 dilution) or anti-FLAG M2 monoclonal
antibody as described previously (7, 8). For immunolocalization of
ARIP3 in paraffin-embedded, formalin-fixed sections of adult rat testis
(19), anti-ARIP3 antibody (1:1000), purified by affinity chromatography
on HiTrap-Sepharose (Amersham Pharmacia Biotech) containing covalently
attached antigen, was used along with Vectastain Elite-Kit (Vector
Laboratories) according to the manufacturer's instructions (8).
 |
RESULTS |
Isolation of cDNAs for ARIP3, a Testis-specific Androgen
Receptor-interacting Protein--
The yeast two-hybrid system was used
to identify proteins that interact with the zinc finger region of AR.
The target sequence (residues 554-644 of human AR) also includes
one-third of the hinge region. Northern blot analysis of mRNAs from
human and rat tissues revealed that one of the cDNA clones
hybridized to a ~2-kilobase transcript expressed predominantly or
solely in the testis (Fig. 1). The
protein encoded by this transcript was termed ARIP3
(androgen receptor-interacting
protein 3.

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Fig. 1.
High level of ARIP3 mRNA expression in
adult rat and human tissues is limited to testis. A,
Northern blot of poly(A)+ RNA (5 µg/lane) from various
rat tissues. B, Northern blot of poly(A)+ RNA (2 µg/lane) from various human tissues. s. intestine, small
intestine. Both blots were probed with a 32P-labeled
cDNA fragment of ARIP3 as described under "Experimental
Procedures." C, human mRNA blot containing
poly(A)+ RNA from 50 different human tissues was hybridized
to 32P-labeled ARIP3 cDNA as above. Row A,
columns 1-8, whole brain, amygdala, caudate nucleus,
cerebellum, cerebral cortex, frontal lobe, hippocampus, and medulla
oblongata, respectively; row B, columns 1-7,
occipital lobe, putamen, substantia nigra, temporal lobe, thalamus,
subthalamic nucleus, and spinal cord, respectively; row C,
columns 1-8, heart, aorta, skeletal muscle, colon, bladder,
uterus, prostate, and stomach, respectively; row D,
columns 1-8, testis, ovary, pancreas, pituitary gland,
adrenal gland, thyroid gland, salivary gland, and mammary gland,
respectively; row E, columns 1-8, kidney, liver,
small intestine, spleen, thymus, peripheral leukocyte, lymph node, and
bone marrow, respectively; row F, columns 1-4,
appendix, lung, trachea, and placenta, respectively; row G,
columns 1-7, fetal brain, fetal heart, fetal kidney, fetal
liver, fetal thymus, and fetal lung, respectively.
|
|
The complete cDNA sequence of ARIP3 isolated from rat testis
cDNA library contains an open reading frame of 572 amino acids with
a calculated molecular mass of 63.3 kDa (Fig.
2). The 1860-base pair sequence has a
115-nucleotide 5'-untranslated region with stop codons in front of a
consensus translation start site and a polyadenylation signal
downstream of the translation termination codon (GenBankTM
accession number AF044058). ARIP3 is particularly rich in Ser/Thr
(18%), Leu/Ile (15%), proline (9%), and charged amino acids (22%).
On the basis of ARIP3 sequences isolated by the two-hybrid screen,
C-terminal residues 443-548 appear to be sufficient for the
interaction with AR.

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Fig. 2.
Amino acid sequence comparison of rat ARIP3,
mouse PIAS3, and human GBP. The numbers depict amino
acid positions; gaps in sequence are shown by dashes. The AR
interaction domain corresponds to residues 443-548. Black
boxes and gray shadings depict amino acids that are
identical or conserved, respectively, among the three sequences.
|
|
ARIP3 shows extensive sequence identity with a recently cloned protein
termed Miz1 that interacts with the homeodomain protein Msx2 (20).
Residues 1-419 of murine Miz1 are 99.5% identical with amino acids
132-550 of rat ARIP3, suggesting that Miz1 and ARIP3 are encoded by
mRNAs that are splice variants of the same gene. ARIP3 is also
highly homologous to two recently identified proteins, mouse PIAS3 and
human GBP (21, 22). Sequence alignment revealed distinct regions that
are well conserved in all three proteins, with amino acid identities
ranging from 60 to 80% (Fig. 2). Most of the C-terminal region of
ARIP3 that harbors the AR ID (residues 443-548) is unique to ARIP3
(Fig. 2).
ARIP3 Interacts with AR in Intact Mammalian Cells and in
Vitro--
A mammalian two-hybrid system in COS-1 cells was used to
confirm that AR-ARIP3 interaction is not a feature peculiar to yeast cells. Transfection of VP16-ARIP3 ID together with Gal4-rAR fusion protein yielded a strong reporter activity, indicating an efficient interaction between AR and ARIP3 ID (Fig.
3A). The full-length ARIP3 and
AR also associated in intact cells, and in agreement with the above
data, deletion of the ID diminished markedly this interaction. Together
these results demonstrate that ARIP3 residues 443-548 are critical for
the AR-ARIP3 interaction to occur (Fig. 3A). Interestingly,
association of AR to ARIP3 ID was attenuated by the presence of
androgen, whereas the interaction of AR with full-length ARIP3 was
slightly enhanced by the hormone. The amount of immunoreactive Gal4-rAR
protein was not influenced by the presence of
androgen.3 AR LBD alone did
not associate with ARIP3 ID (Fig. 3A), and the N-terminal
half of AR encompassing AF-1 (residues 5-538) failed to recognize
either ARIP3 ID or full-length ARIP3,3 indicating that AR
interacts with ARIP3 ID primarily through the ZFR. However, AR LBD
exhibited a weak hormone-enhanced interaction with full-length ARIP3
(Fig. 3A).

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Fig. 3.
Interaction between ARIP3 and AR in mammalian
cells and in vitro. A, interaction of ARIP3 with
AR as assessed by a mammalian two-hybrid system. The ability of rAR
(residues 3-902) or LBD (residues 640-902) fused to Gal4 DBD (Gal4-AR
and Gal4-LBD, respectively) to interact with ARIP3 residues 443-548
(VP16-ARIP3 ID), residues 1-442 (VP16-ARIP3(1-442)), full-length
ARIP3 (VP16-ARIP3(1-572)), or polyoma virus coat protein (VP16-CP)
fused to VP16 AD was examined in COS-1 cells by assaying luciferase
activity from pG5LUC reporter containing five Gal4-binding sites and a
TATA sequence. Cells cultured on 12-well plates were transfected with
200 ng of each chimeric expression vector, 200 ng of pG5LUC and 40 ng
of pCMV using the FuGene reagent. 18 h after transfection, the
medium was changed to one containing charcoal-stripped 2% (v/v) fetal
bovine serum with (+) or without ( ) 100 nM testosterone
(Test), and the cells were incubated for additional 30 h. The mean ± S.E. values for at least three separate experiments
are shown. Reporter gene activities were corrected for transfection
efficiency and are expressed relative to that of Gal4-AR + VP16-ARIP3(1-572) in the presence of testosterone, which was set as
100. Inset, ARIP3 binds AR ZFR in vitro.
35S-Labeled ARIP3 was incubated with GST alone (lane
2) or GST-ZFR (lane 3) adsorbed to
glutathione-Sepharose, after which the matrix was washed, and bound
proteins were analyzed. Lane 1 represent 15% of the amount
of 35S-ARIP3 incubated with the matrix. The doublet band
migrating ~45 kDa represents most likely ARIP3 degradation products.
B, coimmunoprecipitation of ARIP3 with AR.
35S-Labeled ARIP3 or luciferase was incubated with an
extract from COS-1 cells transfected with pSG5rAR (lanes 5 and 6) or empty pSG5 (lanes 3 and 4)
as described previously (12). The extract was subjected to
immunoprecipitation with anti-AR antibody (8), followed by
SDS-polyacrylamide gel electrophoresis and fluorography. Lanes
1 and 2 show 10% of the input samples.
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GST pull-down experiments were used to examine the interaction of AR
ZFR with ARIP3 in vitro. As shown in Fig. 3A
(inset), 35S-labeled ARIP3 produced by
translation in vitro binds specifically to GST-ZFR
immobilized to glutathione-Sepharose. These results thus indicate that
ARIP3 is capable of a direct physical interaction with AR. This
conclusion was strengthened by results that 35S-labeled
full-length ARIP3 was immunoprecipitated with anti-AR antiserum (K183;
Refs. 7 and 8) from an extract of COS-1 cells transfected with pSG5rAR
but not from an extract of cells without AR expression (Fig.
3B).
Subcellular Localization of ARIP3 in Transfected Cells and
Distribution of ARIP3 Antigen in Rat Testis--
In concert with the
presence of putative nuclear localization signals in the ARIP3
sequence, transiently expressed ARIP3 antigen resided exclusively in
CV-1 cell nuclei and presented a speckled pattern of distribution (Fig.
4). In adult rat testis, ARIP3 antigen was detected in the nuclei of Sertoli cells, in spermatogonia, and in
primary spermatocytes up to late pachytene stage of development (Fig.
5A). The immunogen sequence
(ARIP3(443-548)) used to raise anti-ARIP3 antibodies is shared with
Miz1 (20), and thus, the staining of testicular cells may not be
completely specific for ARIP3.

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Fig. 4.
Nuclear localization of ARIP3 in CV-1
cells. CV-1 cells seeded on glass slips on 10-cm plastic plates
were transfected using DOTAP reagent and 1 µg of FLAG-ARIP3
expression vector as described under "Experimental Procedures."
A, ARIP3 was visualized using a rabbit antiserum against
H6-ARIP3 ID. B, anti-FLAG M2 antibody was used
to detect ARIP3. Anti-ARIP3 antiserum neutralized with purified
H6-ARIP3 ID fusion protein showed no specific staining
(data not shown).
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Fig. 5.
Distribution of ARIP3 immunoreactivity in
different cell types of the seminiferous epithelium of rat testis.
Immunoperoxidase technique was applied to visualize ARIP3 antigen using
polyclonal antiserum as described under "Experimental Procedures."
A, ARIP3 can be detected in the nuclei of Sertoli cells
(arrowheads), spermatogonia (double arrowhead)
and in primary spermatocytes (arrows). B, control
staining with normal rabbit IgG used as a primary antibody.
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ARIP3 Is Able to Modulate AR-dependent Transcriptional
Activation--
COS-1 cells devoid of endogenous AR and ARIP3 were
transfected with expression vectors encoding ARIP3 and rat AR along
with a reporter driven by two AREs in front of E1b TATA sequence. Low amounts of coexpressed ARIP3 enhanced AR-dependent
transactivation up to 4-fold, but the effect vanished gradually with
increasing amounts of ARIP3 expression plasmid (Fig.
6A). Deletion of
AR-interacting region (ARIP3
467-547) rendered the protein inactive,
implying that direct interaction between AR and ARIP3 is indeed
required for ARIP3 to influence receptor function (Fig. 6A).
ARIP3 exhibited a similar biphasic dose-response curve with the natural
probasin promoter. In this latter case, however, the activating effect of ARIP3 was weaker, and the highest amount of ARIP3 repressed significantly AR-dependent transcription (Fig.
6B). Ectopic ARIP3 expression did not influence either
promoter in the absence of AR or in the presence of AR without
androgen, and no effect was measured on reporters driven by
constitutively active viral promoters such as CMV and
SV-40.3 In addition, empty pFLAG-CMV-2 (100-800 ng)
modified AR function by less than ± 10%.3

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Fig. 6.
ARIP3 influences AR-dependent
transcription and interaction between N and C termini of AR.
A, effect of ARIP3 coexpression on AR-dependent
activation of a minimal ARE-containing promoter. COS-1 cells cultured
on 6-well plates were transfected with 400 ng of
pARE2-TATA-LUC reporter, 40 ng of pSG5rAR, 40 ng of
pCMV , and increasing amounts of pFLAG-ARIP3(1-572) or
pFLAG-ARIP3 467-547 (in ng) with (+) or without ( ) 100 nM testosterone (T). Total amount of DNA was
kept constant by adding empty pFLAG-CMV-2 as needed. The two ARIP3
forms were expressed to equivalent levels as shown by immunoblot
analysis using anti-FLAG M2 antibody.4 B, effect
of ARIP3 on AR-dependent transactivation of the probasin
promoter. COS-1 cells were transfected as described in the legend to
A except that pPB( 285/+32)-LUC was used as the reporter.
C, ARIP3 enhances the interaction between N and C termini of
AR expressed as separate polypeptides. COS-1 cells on 12-well plates
were transfected with 200 ng of pG5LUC, 100 ng of Gal4-LBD and VP16-NT
(N-terminal residues 5-538 of rAR) along with pFLAG-ARIP3(1-572) (in
ng) and cultured with (+) or without ( ) T. The mean ± S.E.
values for at least three separate experiments are shown. Dose-response
experiments (not shown) revealed that 10 nM T was
sufficient to elicit a maximal response under the conditions shown in
A and C. Relative luciferase activities were
calculated as described under "Experimental Procedures" using
pSG5rAR in the presence of T as 100 in A and B,
and Gal4-LBD and VP16-NT with testosterone as 100 in
C.
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ARIP3 contains two LXXLL motifs starting at residues 18 and
304 as well as two LXXLI motifs at residues 157 and 402 (Fig. 2). LXXLL motifs are needed for the recognition of
several coregulators by AF-2 regions in nuclear receptor LBDs (23).
Some of the LXXLL motif-containing coactivator proteins were
recently shown to facilitate the interaction between N- and C-terminal
regions of AR, when these domains were expressed as separate
polypeptides (24). Likewise, coexpressed ARIP3 facilitated the
interaction between N and C termini, in that it enhanced
transcriptional activity in a modified two-hybrid assay only when the
N- and C-terminal regions were expressed concomitantly in the presence
of androgen (Fig. 6C). Empty pFLAG-CMV-2 (50-200 ng) had no
effect in this assay.4
 |
DISCUSSION |
The identification of a large number of nuclear
receptor-interacting proteins over the last several years and the
evidence from biochemical and genetic studies suggest that to regulate transcription, nuclear receptors must exist as parts of large multiprotein complexes (3, 4, 25, 26). In the present work, we have
identified a testis-specific interaction partner of AR, termed ARIP3.
ARIP3 immunoreactivity is present in all stages of seminiferous
epithelial cycle, with the early meiotic spermatocytes showing the
highest protein levels. ARIP3 interacts with AR both in intact
mammalian cells and in vitro, and it is capable of
modulating AR-dependent transcriptional activity. The biphasic responses could be interpreted to mean that ARIP3 belongs to a
multisubunit coactivator complex, and its overexpression leads to
repression of transcription when other limiting components are titrated
out of the complex. The role that the LXXLL motifs of ARIP3
play in this context requires further studies.
Similar to interactions between some nuclear receptors and the
coactivators PCAF, SNURF, ANPK, and PGC-1 (6-8, 11), the association
between AR and ARIP3 is mediated principally through the ZFR. ARIP3 is
also capable of facilitating the androgen-dependent interaction between the N- and C-terminal regions of the receptor. In
addition to proteins mentioned above, the POU domain-containing proteins Oct-1/2 and Brn-3a/3b as well as TLS/FUS have been shown to
interact with the ZFRs of nuclear receptors (5, 9, 10). These examples
support the conclusion that besides binding to DNA, the ZFR plays an
important role in the formation of heterologous protein-protein contacts.
ARIP3 represents an alternatively spliced form of the transcription
factor Msx2-interacting protein Miz1 (20). The high degree of sequence
conservation among several distinct regions of ARIP3, GBP, PIAS3,
PIAS1, and related sequences (21, 22, 27) indicates the existence of a
novel gene family. GBP was discovered in a yeast two-hybrid screen for
proteins interacting with RNA helicase II (22), and it is almost
identical with PIAS1 (27). PIAS3 and PIAS1 were identified by a similar
approach as proteins interfering with Stat3 and Stat1 function,
respectively (21, 27). Using cDNA library screening, Liu et
al. (27) also isolated a group of other PIAS sequences, namely
PIASx
, PIASx
, and PIASy. ARIP3 appears to be identical with
PIASx
, whereas Miz1 corresponds to PIASx
.
PIAS3 and PIAS1 were demonstrated to bind to activated Stat3 and Stat1,
respectively, and thereby inhibit their DNA binding activities in
vitro (21, 27). Attenuation of STAT-DNA interaction was suggested
to be the mechanism for the repressing functions of PIAS1 and PIAS3.
This is in contrast to findings with Miz1, which enhanced DNA-binding
affinity of Msx2 in vitro (20), and with ARIP3, which
influenced minimally the binding of AR to its cognate DNA
elements.4 Interestingly,
PIASx
was not able to influence either the DNA binding activity of
Stat1 or Stat1-mediated gene activation (27). The C-terminal regions of
ARIP3/PIAS proteins are poorly conserved, suggesting that these domains
are mainly responsible for the transcription factor-specific effects.
ARIP3 was the only family member found in our yeast two-hybrid screen
of mouse embryo cDNA library, most likely due to its unique C terminus.
It is currently unclear whether ARIP3/PIAS family members indeed
utilize distinct modes of action to regulate transcription by their
interacting partners. Their further characterization, such as
identification of STAT-interacting domains of PIAS1 and PIAS3 along
with clarification of the function of LXXLL motifs conserved
among the proteins, is necessary for elucidation of the underlying
regulatory mechanisms.
 |
ACKNOWLEDGEMENTS |
We thank Leena Pietilä, Pirjo
Kilpiö, and Seija Mäki for excellent technical assistance
and Dr. S. M. Hollenberg for providing the materials used in the
yeast two-hybrid screen.
 |
FOOTNOTES |
*
This work was supported by grants from the Medical Research
Council (Academy of Finland), the Finnish Foundation for Cancer Research, the Jalmari and Rauha Ahokas Foundation, the Research and
Science Foundation of Farmos, the Sigrid Jusélius Foundation, Biocentrum Helsinki, the Helsinki University Central Hospital, and the
Turku University Central Hospital.The costs of publication of this
article were defrayed in part by the
payment of page charges. The article
must therefore be hereby marked
"advertisement" in accordance with 18 U.S.C. Section
1734 solely to indicate this fact.
The nucleotide sequence(s) reported in this paper has been submitted to the GenBankTM/EMBL Data Bank with accession number(s) AF044058.
To whom correspondence should be addressed: Dept. of
Physiology, Inst. of Biomedicine, University of Helsinki, P. O. Box 9 (Siltavuorenpenger 20J), FIN-00014 Helsinki, Finland. Tel.:
358-9-1918542; Fax: 358-9-1918681; E-mail:
jorma.palvimo{at}helsinki.fi.
The abbreviations used are:
AF, activation
function; ANPK, androgen receptor-interacting nuclear protein kinase; AR, androgen receptor; ARE, androgen response element; ARIP, androgen
receptor-interacting protein; CMV, cytomegalovirus; DBD, DNA-binding
domain; DOTAP, N-[1-(2,3-dioleoyloxy)propyl]N,N,N-trimethylammonium
methyl sulfate; GBP, Gu/RH-II binding protein; GST, glutathione
S-transferase; ID, interacting domain; LBD, ligand-binding
domain; LUC, luciferase; PCR, polymerase chain reaction; PCAF, p300/CBP-associated factor; PGC, PPAR
coactivator; PIAS, protein
inhibitor of activated STAT; ZFR, zinc finger region; SNURF, small
nuclear RING finger; STAT, signal transducer and activator of transcription.
2
The term "residues" refers always to amino acids.
3
A.-M. Moilanen, J. J. Palvimo, and O. A. Jänne, unpublished observations.
4
J. J. Palvimo, A.-M. Moilanen, H. Poukka,
and O. A. Jänne, unpublished observations.
 |
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