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Journal of Virology, September 2000, p. 8159-8165, Vol. 74, No. 17
0022-538X/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.
Human Immunodeficiency Virus Type 1 Vpr Contains
Two Leucine-Rich Helices That Mediate Glucocorticoid Receptor
Coactivation Independently of Its Effects on G2 Cell
Cycle Arrest
Michael P.
Sherman,1,2
Carlos M. C.
de
Noronha,1
David
Pearce,3 and
Warner C.
Greene1,3,4,*
Gladstone Institute of Virology and
Immunology1 and Departments of
Medicine,3 Microbiology and
Immunology,4 and Hematology and
Oncology,2 University of California, San
Francisco, California
Received 23 March 2000/Accepted 31 May 2000
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ABSTRACT |
Human immunodeficiency virus type 1 (HIV-1) Vpr participates in
nuclear targeting of the viral preintegration complex in nondividing cells and induces G2 cell cycle arrest in proliferating
cells, which creates an intracellular milieu favorable for viral
replication. Vpr also activates the transcription of several promoters
and enhancers by a poorly understood mechanism. Vpr enhances
glucocorticoid receptor (GR) signaling and may mediate the effects of
steroids on HIV replication. More specifically, recombinant Vpr can
potentiate virion production from U937 cells, downregulate NF-
B
induction, and enhance programmed cell death, all effects also mediated
by glucocorticoids. Vpr has been proposed to act as a GR coactivator, although other studies suggest that these enhancing effects are merely
a consequence of G2 cell cycle arrest. We now demonstrate that Vpr functions as a GR coactivator and that this activity is
independent of cell cycle arrest. In addition, we show that the
Vpr-induced coactivation requires an intact glucocorticoid response
element, that it is dependent on the presence of hormone and the
corresponding receptor, and that it is mediated by the two highly
conserved leucine-rich domains within Vpr that resemble the GR
coactivator signature motif.
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INTRODUCTION |
The genomes of human
immunodeficiency virus types 1 and 2 (HIV-1 and HIV-2) encode several
accessory proteins that are highly conserved in vivo and likely
function to accelerate virus production. The vpr gene of
HIV-1 encodes a 96-amino-acid, 14-kDa viral protein R (Vpr) that is
expressed in infected cells and packaged into the virion through its
interaction with the p6 region of the Gag precursor (6, 40,
62). Structural analysis of Vpr by nuclear magnetic resonance
(48) and circular dichroism (35) predicts two
-helices, one located at the amino terminus between amino acids 17 and 34, and the other between amino acids 53 and 78. These helices
likely play a role in Vpr dimerization (64) and perhaps
binding to other proteins (63), likely through a leucine zipper-like mechanism (58). The carboxy-terminal
region of Vpr corresponds to a less-well-characterized basic
amino acid stretch between amino acids 79 and 96.
While Vpr is not required for viral replication in transformed cell
lines or even cultured peripheral blood mononuclear cells (PBMCs)
(39, 46), the open reading frame is maintained in vivo
(17, 61), reflecting its importance in the HIV replicative life cycle. Vpr facilitates virus production in macrophages, perhaps by
cooperating with integrase and matrix proteins to promote nuclear import of the viral preintegration complex (8, 15, 16, 21, 42,
56). In this regard, Vpr contains at least two distinct import
signals that facilitate nuclear entry by a novel mechanism that may
involve direct docking to the nuclear pore complex (25).
Vpr also induces G2 cell cycle arrest in proliferating
human cells (4, 17, 19, 26, 43). In fact, each virion
contains sufficient quantities of packaged Vpr to arrest infected T
cells in the G2 phase of the cell cycle (24,
41). Such arrest during the initial phase of infection may create
an intracellular environment conducive to improved transcription of the
long terminal repeat (LTR) (17). However, at a later point
in HIV replication, the G2 arrest induced by Vpr may lead
to increased apoptosis even in cells infected with
replication-defective forms of HIV (41, 51, 52, 59). The
G2 arrest property of HIV-1 Vpr has been dissociated from
the nuclear import properties by mutagenesis experiments (14, 36,
53). In the related HIV-2 and simian immunodeficiency virus (SIV)
lentiviruses, the nuclear import and G2 cell cycle arrest
properties are segregated between Vpx and Vpr2/VprSIV
(13, 55).
In addition to increasing HIV LTR transcription, Vpr also upregulates
the activity of several heterologous promoters and enhancers (7). It is possible that this effect is a byproduct of
G2 cell cycle arrest. Increased transcription is observed
in arrested cells following expression of Vpr by transfection or by
infection (18, 53). However, it has been suggested that Vpr
may increase transcription by physically interacting with various host
factors, such as Sp1 (57), TFIIB (1, 2, 28), or
an activated glucocorticoid receptor (GR) (28, 44). If Vpr
binds DNA or other transcription factors directly in vivo, it likely
enhances transcription through improved recruitment of p300/CBP
(11).
Receptors for steroid hormones, thyroid hormones, and retinoic acid all
belong to a superfamily of ligand-dependent transcription factors (for
a review, see reference 54). These factors have sequence homology that includes an amino-terminal transcriptional activation domain (AD or AF-1), a central DNA-binding motif, and a
carboxy-terminal, ligand-binding domain containing a second activation
domain termed AF-2. Mutations in AF-2 can abolish signal transduction
without affecting ligand binding or dimerization. Recently, steroid
receptor coactivators (SRCs) have been identified as important members
of the transcriptional activation complex that mediates nuclear hormone
signaling. The recruitment of gene transcription machinery by the
activated nuclear hormone receptor is dependent on this new class of
coactivator proteins. The SRCs bind the hydrophobic cleft within the
AF-2 domain through physical interaction of their signature LXXLL
motifs, which are required for their function (9, 12, 20,
54).
It has been suggested that glucocorticoids and recombinant Vpr
influence the HIV life cycle in an analogous fashion (44). Specifically, the Vpr-mediated enhancement of virus production demonstrated in cultured macrophages was mimicked by substitution of
glucocorticoids and, correspondingly, inhibited by the addition of the
GR antagonist RU486 (mifepristone). Moreover, exogenous addition of
recombinant Vpr to cells appears to stimulate glucocorticoid activity
in cultured cells with respect to apoptosis, increased virion
production from U937 cells, and downregulation of NF-B (3). Treatment with RU486 inhibits all of these effects. The observation that SRCs contained the signature motif LXXLL and the
presence within Vpr of an LQQLL element at amino acids 64 to 68 raised
the possibility that Vpr functions as a GR coactivator. Kino and
colleagues have described such an activity (28). However, others have directly disputed this finding and suggest that the G2 cell cycle arresting properties account for these
coactivator-like effects of Vpr (14). In this regard,
replacement of L64 with alanine in the LQQLL motif (AQQLL or L64A) both
inhibits the transcriptional activating properties of Vpr and reduces
the degree of G2 cell cycle arrest (28, 36).
In view of these uncertainties, we further explored the possibility
that Vpr acts as a GR coactivator. Because it is known that multiple
LXXLL interaction domains function synergistically in SRCs (9, 38,
47), we also examined the possible cooperation between the two
leucine-rich motifs of Vpr with respect to G2 cell cycle
arrest as well as GR coactivation.
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MATERIALS AND METHODS |
Plasmids.
The plasmid containing the glucocorticoid response
element (GRE) derived from the tyrosine aminotransferase (TAT) 5'
regulatory region and the control plasmid lacking the TAT3 repeats have
been described (33). The rat GR encoded by the 6RGR plasmid
(32) was used in coimmunoprecipitation assays and in
transfection of CV1-B cells, which lack an endogenous cortisol
receptor. The GRIP1-encoding plasmid (23) was kindly
provided by Michael Stallcup (University of Southern California). All
Vpr constructs were inserted into the pCMV4 vector containing three
hemagglutinin (HA) repeats at the amino terminus. Mutations were
generated by oligonucleotide-directed PCR and confirmed by sequencing.
Expression was verified by Western blot analysis. All constructs that
induced G2 arrest as well as the Vpx construct expressed
proteins at nearly equivalent levels (data not shown).
Cell lines and transfections.
All transfections were
performed using calcium phosphate for precipitation of DNA. Cells were
plated at 300,000/well in six-well plates containing Dulbecco's
modified Eagle's medium (Gibco-BRL, Gaithersburg, Md.) with 5%
"stripped" fetal bovine serum (charcoal treated at 10 g/liter for
1 h to remove endogenous steroids), penicillin G at 100 U/ml, and
streptomycin at 100 µg/ml. Transfections were performed 18 h
after plating at a constant DNA concentration of 4 µg per well. Cells
were washed 24 h after transfection. At 48 h, the medium was
replaced with medium containing the indicated steroid and analyzed
3 h later. Transfected cells were assayed for luciferase activity
after lysis with 1× lysis buffer (Promega), adding reagents A and B
(Amersham) to 20 µl of lysate, and measuring light output on a
Microbeta 1450 Trilux luminescence counter (Wallac Company).
Coimmunoprecipitation.
293T cells were plated in 100-mm
dishes, transfected, and cultured for 48 h. Cells were washed with
phosphate-buffered saline (PBS) and lysed for 20 min, and the
supernatant was incubated with monoclonal mouse antibody HA.11
immobilized on Sepharose Fast Flow beads (BabCO) for 1 h at 4°C
and then washed three times. The beads were then boiled for 5 min to
dissociate any bound proteins. Lysis and wash buffer consisted of 50 mM
HEPES, 150 mM NaCl, 5 mM EDTA, and 0.5% NP-40 detergent. Western
blotting of the proteins that had bound the immunoprecipitation beads
or proteins from the lysates was performed using polyclonal antibodies
(Affinity Bioreagents) specific for the GR.
Cell cycle analysis.
Cell cycle analyses were performed by
using green fluorescent protein (GFP) as a marker to distinguish
transfected and untransfected cells. Experiments were performed with
pEGFP expression vector (Clontech) cotransfected at a 1:8 ratio with an
HA expression vector cloned in frame with Vpr or mutants of Vpr. 293T
cells were prepared for cell cycle analysis by initial trypsinization followed by fixation for 30 min in 2% formaldehyde. The cells were
washed with PBS and treated with RNase A (1 mg/ml) and propidium iodide
(10 µg/ml) in PBS for 30 min. Cellular DNA content in the transfected
(GFP+) and untransfected (GFP
) cells was
assessed with a FACScan flow cytometer (Becton Dickinson).
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RESULTS |
Vpr functions as an SRC.
The potential action of Vpr as an SRC
was compared to coactivation by GR-interacting protein 1 (GRIP1), a
well-characterized SRC (22, 23). Both GRIP1 and Vpr
significantly enhanced luciferase activity driven by the GRE in the
presence of increasing amounts of dexamethasone, with a constant amount
of transfected GRIP1 or Vpr expression plasmid together with the
endogenous GR expressed in 293T cells (Fig.
1). These proteins induced a similar
shift in the coactivation curve, although Vpr exhibited roughly 50% of
the activity of GRIP1. In several replicates, Vpr produced a three- to
sixfold greater GR-mediated response than was obtained in cells treated
with similar concentrations of dexamethasone and control vector. The
weak induction of GRE obtained in the latter condition presumably
reflects the action of an endogenous coactivator.
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FIG. 1.
Vpr displays SRC activity. DNAs (2.7 µg in a total of
4 µg) encoding the HA-tagged known SRC GRIP1, wild-type HIV-1 Vpr, or
HA alone (vector) were transfected into 293T cells and incubated in the
presence of increasing concentrations of dexamethasone. Luciferase
activity from a GRE-driven reporter, expressed as relative light units
(RLU), was normalized to activity obtained with a cotransfected RSV
 -galactosidase reporter to account for differences in gene transfer.
Vpr produces effects similar to though less marked than those of GRIP1,
shifting the dose-response curve up and to the left.
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Subsequently, we examined the ability of Vpr to produce a
dose-dependent coactivation effect and the capacity of RU486 to
inhibit
such a response (Fig.
2). Transfecting
increasing amounts
of Vpr in the presence of a constant amount of total
DNA induced
a dose-dependent increase in endogenous GR coactivation.
Furthermore,
this coactivation was specifically inhibited by the
introduction
of the RU486 antagonist when used at a concentration
higher than
that of dexamethasone. We controlled for nonspecific Vpr
transactivation
through induction of G
2 cell cycle arrest
(
7,
14,
59) by
including a Rous sarcoma virus (RSV)
promoter-driven,
-galactosidase
reporter in all cell transfections.
This reporter is activated
by G
2 arrest and modestly
induced by Vpr (
7). Accordingly,
differences occurring
secondary to the G
2 arrest or transfection
efficiency could
be accounted for by normalizing the GRE-driven
luciferase activity to
overall galactosidase activity. Together,
these data suggest that Vpr
exerts a dose-dependent effect on
the GRE independent of changes
related to G
2 cell cycle arrest.
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FIG. 2.
Vpr mediates dose-dependent enhancement of coactivation
of endogenous GR by dexamethasone, and the GR antagonist RU486 blocks
the effects. 293T cells were transfected with the indicated amounts of
HA-Vpr in a constant background of 4 µg of total DNA. Luciferase
activity from a GRE-driven reporter was normalized using RSV
-galactosidase (-gal) reporter activity. Glucocorticoid
activation increased with increasing amounts of transfected Vpr in
cells incubated with 0.1 µM dexamethasone. This response was
inhibited when cells were incubated with 0.1 µM dexamethasone and 1 µM RU486.
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We further established the specificity of the Vpr-mediated GR
coactivation by examining the dependence on a GRE. In the presence
of
three GRE repeats, HIV-1 Vpr caused an almost 3-fold induction
over
similarly hormone-treated cells transfected with a background
vector
and a 16-fold increase over non-hormone-treated cells (Fig.
3A). The small amount of basal
coactivation seen may have been
due to residual hormones remaining in
the charcoal-treated serum
supplement or more likely represents
oscillations of basal expression,
as there was no coactivation above
the vector control. When the
hormone response element was absent,
parallel experiments revealed
that coactivation was abolished in the
presence and absence of
the agonist. We next examined whether the
analogous proteins of
HIV-2 were likewise able to coactivate the GR.
Vpx has no identifiable
leucine-rich domain, while Vpr from HIV-2
(Vpr2), which can induce
G
2 cell cycle arrest, contains a
single domain homologous to HIV-1
Vpr that is centered on the pentamer
LQRAL. Vpr2 displayed about
50% of the coactivator activity of HIV-1
Vpr, while Vpx had no
coactivation potential despite being expressed at
higher levels
(data not shown). Together, these findings indicate that
Vpr2
but not Vpx has a moderate GR coactivation function that is both
hormone and GRE dependent.
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FIG. 3.
Vpr coactivation of GR is dependent on the presence of
ligand, an intact GRE, and GR. (A) DNAs encoding HA, HA-tagged Vpr from
HIV-1 or HIV-2, and Vpx were transfected into 293T cells in the
presence (+) or absence ( ) of dexamethasone (Dex) or an intact GRE.
(B) CVI-B cells, which lack the machinery to respond to cortisol
stimulation, were transfected with HA-tagged Vpr or an HA vector with
(+) or without () transfected GR in the presence (+) or absence ()
of the glucocorticoid cortisol. Vpr coactivation required the presence
of both cotransfected GR and ligand stimulation.
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As a final control, we tested whether the Vpr coactivation was receptor
dependent (Fig.
3B). The monkey kidney cell line CV-1B
lacks endogenous
cortisol receptors (
33). Using these cells,
the Vpr
coactivator response did not occur unless GR was also
provided during
transfection. In the presence of GR, a 2.5- to
3-fold enhancement of
GRE-luciferase activity was observed. Notably,
Vpr is unable to induce
G
2 cell cycle arrest in CV1-B cells, again
providing strong
evidence that cell cycle alterations are not
solely responsible for the
induction of the
GRE.
Vpr-induced GR coactivation involves two leucine-rich motifs and
occurs independently of cell cycle arrest.
We next sought to
confirm that the LQQLL motif present in HIV-1 Vpr at residues 64 to 68 was responsible for mediating Vpr-induced glucocorticoid coactivation
and to further distinguish the G2 arrest phenotype of
Vpr from its coactivation properties. We also examined the
possibility that a more amino-terminal LLEEL pentamer might be involved
in coactivation. While this latter motif is "reversed" in
orientation from the classical SRC signature motif, such determinants
have been implicated in coactivation by known SRCs (Beatrice Darimont,
personal communication). Furthermore, SRCs such as GRIP1 contain
multiple LXXLL motifs that function in concert and may serve as a model
for Vpr-mediated functions. Table 1
summarizes the ability of transfected Vpr to induce G2 cell
cycle arrest when these two leucine-rich -helices were mutated individually or in tandem. While no leucine in the amino-terminal helix
was involved in G2 arrest, the L64A
(A64QQLL) mutation alone disrupted the ability of Vpr
to delay cells at the G2 checkpoint. These findings are
consistent with data from Mahalingam et al. (36) but
disagree with the results described by Kino et al. (28), who
found that this mutation abolished coactivation without
affecting G2 arrest. The single L67A
(LQQA67L) and L68A (LQQLA68) substitution
mutations did not affect the arrest phenotype, but the composite
LQQA67A68 mutation did. Vpr2
contains an analogous leucine-rich motif (LQRAL) that is similar to
that of the LQQA67L mutant of HIV-1 Vpr, and the wild-type
form is able to cause G2 arrest. Of special note, Vpr
mutants that retained the ability to induce G2 arrest were
consistently expressed at much higher levels than those that failed to
induce arrest. In the analysis of coactivation, these different levels
of Vpr expression must be carefully considered. We chose to compare Vpr
mutants that retained G2-arresting properties.
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TABLE 1.
G2 cell cycle arrest of 293T cells induced by
HIV-1 and HIV-2 Vpr homologues and corresponding
helix mutantsa
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The LQQA
67L and LQQLA
68 mutations of Vpr in the
carboxy leucine-rich motif (H2) only partially reduced coactivation
potential
(approximately 50%) (Fig.
4A).
Therefore, we had further evidence
prompting us to examine the
potential contribution of the amino-terminal
leucine-rich motif (H1) to
the coactivation response. Mutation
of all three leucines in the first
helix (A
22A
23EEA
26) did not
compromise the ability of Vpr to induce G
2 arrest but
did
reduce GR coactivation by approximately 50%. When the
A
22A
23EEA
26 and LQQA
67L
mutations were combined
(A
22A
23EEA
26/LQQA
67L),
the G
2 arrest phenotype was maintained but coactivation was
abolished (Fig.
4B). Thus, both leucine-rich domains appear to
cooperate to facilitate GR coactivation. A similar coactivation
pattern
is seen with GRIP1, in which multiple LXXLL motifs contribute
to SRC
function (
9,
47).
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FIG. 4.
Mutations of the glucocorticoid coactivator motif
64LXXLL68 in the carboxy-terminal helix (H2)
and the noncanonical motif (22LLEEL26) in the
amino-terminal helix (H1) of HIV-1 Vpr attenuate the glucocorticoid
response. (A) DNAs encoding wild-type HIV-1 Vpr (WT), mutations of
HIV-1 Vpr in the 64LXXLL68 motif as indicated,
or HIV-2 Vpr (Vpr2) were transfected into 293T cells in the presence or
absence of 0.1 µM dexamethasone, and fold induction of GRE was
compared. Mutations of the 64LXXLL68 sequence
that maintained G2 cell cycle arrest are shown. Vpr2
facilitated G2 arrest yet only induced 50% of the
coactivation activity. (B) Wild-type (WT) HIV-1 Vpr and the indicated
H1, H2, and H1 + H2 mutants all maintain the arrest phenotype, but
coactivation of GRE was reduced by 50% when either helix was mutated
and lost completely when both helices were altered. Luciferase activity
from a GRE-driven reporter was normalized to an RSV -galactosidase
reporter and is represented as fold induction for both A and B. (C)
Expression of all mutants was mostly equivalent or superior to
expression of wild-type (WT) Vpr.
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Vpr physically interacts with the GR.
Subsequently, we
assessed the physical interaction of Vpr and GR to further
address the role of each leucine-rich motif in coactivation.
Wild-type Vpr and GRIP1 can bind to the GR under similar
conditions, whereas the protein phosphatase 2A regulatory subunit A, a control protein, did not bind. Mutations that
replaced key leucines in both motifs interfered with the ability of Vpr to bind to GR (Fig. 5). Of note, the
addition of exogenous glucocorticoid was not required for
coimmunoprecipitation of either Vpr or GRIP1 with GR. These results
suggest either that sufficient endogenous hormone is present in
serum-supplemented medium or that this binding occurs in the absence of
hormone when these proteins are expressed at high levels.
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FIG. 5.
Mutations in either the first (H1) or second (H2)
leucine-rich motif of HIV-1 Vpr abolished binding to the GR. HA-Vpr and
GR were cotransfected into 293T cells, and the lysates were
immunoprecipitated (IP) with monoclonal mouse antibody HA.11
immobilized onto Sepharose Fast Flow beads. Western blots (IB) of the
beads or the lysates were probed with anti-GR antibodies. Wild-type
HA-tagged Vpr (HA-Vpr) bound GR under the same conditions as HA-tagged
GRIP1 (HA-GRIP1). No binding was seen with the HA-tagged protein
phosphatase 2A (HA-PP2A), and reduced binding was seen when the first
leucine-rich motif (22LLEEL26) was mutated
(A22A23EEA26), when the second
motif (64LXXLL68) was mutated
(A64QQA67A68), or when both these
motif mutations were combined in a double helix mutation (H1 + H2).
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Vpr alleles derived from primary viral isolates retain the ability
to coactivate the GR.
We next addressed whether the Vpr proteins
encoded by patient-derived viruses shared the GR coactivation
properties observed with NL4-3-derived Vpr. Vpr was amplified from
PBMCs isolated from two HIV-infected patients and inserted into pUC-19,
whereupon multiple clones were sequenced (Fig.
6A). Both leucine-rich motifs were found
to be highly conserved among the quasispecies obtained from these two
patients. One clone (2.28) had a single leucine mutation with a
phenylalanine at position 68, but the coding sequence of this clone was
interrupted by a stop codon (Z) at amino acid 9, thereby removing the
pressure for sequence conservation. Another clone from the same patient
(2.07) had a deletion in the first helix, but expression of this clone
was not detectable by Western blot analysis. Four Vpr alleles (1.06, 1.23, 2.10, and 2.12) containing conserved leucine motifs were tested
for the ability to coactivate endogenous GR in 293T cells by
cotransfection with the GRE-luciferase reporter. They displayed nearly
the same degree of coactivation as the NL4-3 Vpr clone (4- to 6-fold,
magnified by the addition of dexamethasone to 40- to 60-fold) (Fig.
6B), despite lower levels of protein expression (Fig. 6C).
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FIG. 6.
Leucine-rich coactivator domains are conserved among
patients, and wild-type isolates retain SRC function. (A) Ten
quasispecies from two HIV-infected patients were sequenced, and the
conserved coactivator domains are highlighted. Isolate 2.28 had a
phenylalanine substitution at amino acid (aa) position 26, but this
clone was terminated early by a stop codon (Z). Isolate 2.07 lacked the
first leucine-rich domain due to a deletion, but this clone was not
expressed in a stable manner. (B) Coactivation of four randomly
selected patient isolates compared to that of the NL43-derived clone.
(C) Expression of the patient-derived clones was reduced compared to
that of the NL-43 clone but still retained coactivation potential, as
seen in panel B.
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DISCUSSION |
HIV-1 contains several accessory proteins that play various roles
in the viral life cycle. Vpr is highly conserved in vivo and has been
implicated in accelerating virus production by facilitating entry of
the preintegration complex into the nuclei of nondividing cells and by
causing G2 cell cycle arrest of infected T cells, leading
to increased LTR transcription. Although transactivation by Vpr
correlates with its ability to cause G2 arrest, we now show
that Vpr-mediated coactivation of the GR is a distinct phenotype, confirming and extending the work of Kino and colleagues
(28). In addition to the LQQLL motif present in the
carboxy-terminal helix, we find a contribution of the amino-terminal
LLEEL motif to the coactivation response. A search of the GenBank
database revealed that both of the leucine-rich domains are highly
conserved among various patient isolates. In fact, we have now
sequenced over 100 intact Vpr quasispecies from 10 patients and found
that both helices are completely conserved in all but one intact isolate.
Vpr likely multimerizes through interactions at the amino-terminal
leucine-rich domain (64) and binds heterologous proteins through interactions at the carboxy-terminal leucine domain
(63). Vpr is a relatively small protein, and mutations
throughout its 96 amino acids interfere with its various phenotypes
differentially (36). While each of the two leucine-rich
domains may promote an interaction with a specific binding partner, our
finding that the L68A (LQQLA68) mutant of Vpr retained
G2-arresting properties except when it also had mutations
in the first leucine-rich helix implies that the two helices may
cooperate to promote arrest. While mutations of each -helix
interfered with binding of Vpr to GR (Fig. 5) and partially compromised
the coactivation function, we cannot distinguish between the
possibilities that the mutations interfere with direct binding to GR
and that multimerization of Vpr is required for coactivation. In this
regard, it has been shown that the binding domain of Vpr for the p6
region of the Gag precursor resides in the amino terminus but is
affected by mutations in the carboxy terminus (60). It
should be noted that Vpr binding to Gag also involves an LXXLF motif in
p6 (29, 34).
Vpr can be detected in the serum of HIV-1-infected patients, and
exogenously applied, recombinant Vpr has been shown to enhance virus
production from various latently infected cell lines and PBMCs
(30, 31). Extracellular Vpr is thought to have several potential effects in vivo, including enhancement of virion production. Even in replication-defective forms of the virus, there is sufficient Vpr in the virion to induce G2 arrest in the absence of de
novo Vpr synthesis within the infected cell. This finding has led some to hypothesize that Vpr may play a role in immune suppression involving
non-productively infected cells (41). Of note, a similar function has been ascribed to glucocorticoids. Since Vpr in the virion
can induce G2 cell cycle arrest, which in turn may lead to
programmed cell death, it may be that Vpr depletes cell populations by
inducing apoptosis, an effect also known to be mediated by steroids.
Recombinant Vpr possesses glucocorticoid-like activity that can be
inhibited by antibodies to Vpr or by the glucocorticoid antagonist
RU486 (30, 31, 44). Specifically, recombinant Vpr can act as
a glucocorticoid on cultured cells by inducing apoptosis,
increasing virion production from U937 cells, and downregulating NF-B through an upregulation of IB synthesis. However, it is now
becoming clear that glucocorticoids may not modulate IB expression in order to exert their effects (5, 10, 27, 45).
Nevertheless, the parallels between glucocorticoid and recombinant Vpr
action are intriguing given the fact that Vpr can function as an SRC. This is particularly interesting with respect to increased virus production in HIV-infected cultured H9 T cells in the presence of Vpr
and glucocorticoids (37, 49). In fact, the HIV-1 provirus contains an intact and functional GRE within the vif open
reading frame (49).
These observations have led to the proposal that glucocorticoid
antagonists be used as antiretroviral agents (3, 44). However, Soudenyns and Wainberg (50) have been quick to
point out that the effects of glucocorticoids on HIV replication in vitro and in vivo are diverse and varied and should be further explored
at the molecular level before any such agents are used clinically. We
have been unable to show any effect of glucocorticoids or RU486 on
HIV-1 replication in cultured H9 cells, PBMCs, or primary macrophages
(data not shown). Likewise, isogenic viruses lacking only the Vpr open
reading frame replicated with kinetics similar to those of the
wild-type constructs despite the addition of exogenous dexamethasone or
RU486. Since Vpr may also coactivate hormone receptors for estradiol
and progesterone (28), perhaps another agonist or antagonist
hormone ligand might have more profound effects on virus production.
Alternatively, it is likely that the plethora of effects exerted by
glucocorticoids in vivo on T-cell function and virus-host cell
interactions as well as the effects of circulating Vpr cannot be
accurately recapitulated with in vitro culture systems. While it is
clear that Vpr can function as an SRC, further studies are necessary to
determine whether this function contributes to the spread or production of HIV or to the pathogenesis of AIDS.
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ACKNOWLEDGMENTS |
We thank Michael Stallcup for providing the GRIP1 plasmid and
Beatrice Darimont for sharing unpublished results. We also thank J. Lo and M. Schambelan for providing clinical specimens.
This work was supported by the UCSF-GIVI Center for AIDS Research,
grant NIH P30 MH59037, and SFGH GCRC grant M01 RR00083.
| |
FOOTNOTES |
*
Corresponding author. Mailing address: Gladstone
Institute of Virology and Immunology, P.O. Box 419100, San Francisco,
CA 94141-9100. Phone: (415) 826-3800. Fax: (415) 826-1817. E-mail: wgreene{at}gladstone.ucsf.edu.
| |
REFERENCES |
| 1.
|
Agostini, I.,
J. M. Navarro,
M. Bouhamdan,
K. Willetts,
F. Rey,
B. Spire,
R. Vigne,
R. Pomerantz, and J. Sire.
1999.
The HIV-1 Vpr co-activator induces a conformational change in TFIIB.
FEBS Lett.
450:235-239[CrossRef][Medline].
|
| 2.
|
Agostini, I.,
J. M. Navarro,
F. Rey,
M. Bouhamdan,
B. Spire,
R. Vigne, and J. Sire.
1996.
The human immunodeficiency virus type 1 Vpr transactivator: cooperation with promoter-bound activator domains and binding to TFIIB.
J. Mol. Biol.
261:599-606[CrossRef][Medline].
|
| 3.
|
Ayyavoo, V.,
A. Mahboubi,
S. Mahalingam,
R. Ramalingam,
S. Kudchodkar,
W. V. Williams,
D. R. Green, and D. B. Weiner.
1997.
HIV-1 Vpr suppresses immune activation and apoptosis through regulation of nuclear factor kappa B.
Nat. Med.
3:1117-1123[CrossRef][Medline].
|
| 4.
|
Bartz, S. R.,
M. E. Rogel, and M. Emerman.
1996.
Human immunodeficiency virus type 1 cell cycle control: Vpr is cytostatic and mediates G2 accumulation by a mechanism which differs from DNA damage checkpoint control.
J. Virol.
70:2324-2331[Abstract].
|
| 5.
|
Brostjan, C.,
J. Anrather,
V. Csizmadia,
D. Stroka,
M. Soares,
F. H. Bach, and H. Winkler.
1996.
Glucocorticoid-mediated repression of NFkappaB activity in endothelial cells does not involve induction of IkappaBalpha synthesis.
J. Biol. Chem.
271:19612-19616[Abstract/Free Full Text].
|
| 6.
|
Cohen, E. A.,
G. Dehni,
J. G. Sodroski, and W. A. Haseltine.
1990.
Human immunodeficiency virus vpr product is a virion-associated regulatory protein.
J. Virol.
64:3097-3099[Abstract/Free Full Text].
|
| 7.
|
Cohen, E. A.,
E. F. Terwilliger,
Y. Jalinoos,
J. Proulx,
J. G. Sodroski, and W. A. Haseltine.
1990.
Identification of HIV-1 vpr product and function.
J. Acquir. Immune Defic. Syndr.
3:11-18.
|
| 8.
|
Connor, R. I.,
B. K. Chen,
S. Choe, and N. R. Landau.
1995.
Vpr is required for efficient replication of human immunodeficiency virus type-1 in mononuclear phagocytes.
Virology
206:935-944[CrossRef][Medline].
|
| 9.
|
Darimont, B. D.,
R. L. Wagner,
J. W. Apriletti,
M. R. Stallcup,
P. J. Kushner,
J. D. Baxter,
R. J. Fletterick, and K. R. Yamamoto.
1998.
Structure and specificity of nuclear receptor-coactivator interactions.
Genes Dev.
12:3343-3356[Abstract/Free Full Text].
|
| 10.
|
De Bosscher, K.,
M. L. Schmitz,
W. Vanden Berghe,
S. Plaisance,
W. Fiers, and G. Haegeman.
1997.
Glucocorticoid-mediated repression of nuclear factor-kappaB-dependent transcription involves direct interference with transactivation.
Proc. Natl. Acad. Sci. USA
94:13504-13509[Abstract/Free Full Text].
|
| 11.
|
Felzien, L. K.,
C. Woffendin,
M. O. Hottiger,
R. A. Subbramanian,
E. A. Cohen, and G. J. Nabel.
1998.
HIV transcriptional activation by the accessory protein, VPR, is mediated by the p300 co-activator.
Proc. Natl. Acad. Sci. USA
95:5281-5286[Abstract/Free Full Text].
|
| 12.
|
Feng, W.,
R. C. Ribeiro,
R. L. Wagner,
H. Nguyen,
J. W. Apriletti,
R. J. Fletterick,
J. D. Baxter,
P. J. Kushner, and B. L. West.
1998.
Hormone-dependent coactivator binding to a hydrophobic cleft on nuclear receptors.
Science
280:1747-1749[Abstract/Free Full Text].
|
| 13.
|
Fletcher, T. M., 3rd,
B. Brichacek,
N. Sharova,
M. A. Newman,
G. Stivahtis,
P. M. Sharp,
M. Emerman,
B. H. Hahn, and M. Stevenson.
1996.
Nuclear import and cell cycle arrest functions of the HIV-1 Vpr protein are encoded by two separate genes in HIV-2/SIV(SM).
EMBO J.
15:6155-6165[Medline].
|
| 14.
|
Forget, J.,
X. J. Yao,
J. Mercier, and E. A. Cohen.
1998.
Human immunodeficiency virus type 1 vpr protein transactivation function: mechanism and identification of domains involved.
J. Mol. Biol.
284:915-923[CrossRef][Medline].
|
| 15.
|
Gallay, P.,
T. Hope,
D. Chin, and D. Trono.
1997.
HIV-1 infection of nondividing cells through the recognition of integrase by the importin/karyopherin pathway.
Proc. Natl. Acad. Sci. USA
94:9825-9830[Abstract/Free Full Text].
|
| 16.
|
Gallay, P.,
V. Stitt,
C. Mundy,
M. Oettinger, and D. Trono.
1996.
Role of the karyopherin pathway in human immunodeficiency virus type 1 nuclear import.
J. Virol.
70:1027-1032[Abstract].
|
| 17.
|
Goh, W. C.,
M. E. Rogel,
C. M. Kinsey,
S. F. Michael,
P. N. Fultz,
M. A. Nowak,
B. H. Hahn, and M. Emerman.
1998.
HIV-1 Vpr increases viral expression by manipulation of the cell cycle: a mechanism for selection of Vpr in vivo.
Nat. Med.
4:65-71[CrossRef][Medline].
|
| 18.
|
Gummuluru, S., and M. Emerman.
1999.
Cell cycle- and Vpr-mediated regulation of human immunodeficiency virus type 1 expression in primary and transformed T-cell lines.
J. Virol.
73:5422-5430[Abstract/Free Full Text].
|
| 19.
|
He, J.,
S. Choe,
R. Walker,
P. Di Marzio,
D. O. Morgan, and N. R. Landau.
1995.
Human immunodeficiency virus type 1 viral protein R (Vpr) arrests cells in the G2 phase of the cell cycle by inhibiting p34cdc2 activity.
J. Virol.
69:6705-6711[Abstract].
|
| 20.
|
Heery, D. M.,
E. Kalkhoven,
S. Hoare, and M. G. Parker.
1997.
A signature motif in transcriptional co-activators mediates binding to nuclear receptors.
Nature
387:733-736[CrossRef][Medline].
|
| 21.
|
Heinzinger, N. K.,
M. I. Bukinsky,
S. A. Haggerty,
A. M. Ragland,
V. Kewalramani,
M. A. Lee,
H. E. Gendelman,
L. Ratner,
M. Stevenson, and M. Emerman.
1994.
The Vpr protein of human immunodeficiency virus type 1 influences nuclear localization of viral nucleic acids in nondividing host cells.
Proc. Natl. Acad. Sci. USA
91:7311-7315[Abstract/Free Full Text].
|
| 22.
|
Hong, H.,
K. Kohli,
M. J. Garabedian, and M. R. Stallcup.
1997.
GRIP1, a transcriptional coactivator for the AF-2 transactivation domain of steroid, thyroid, retinoid, and vitamin D receptors.
Mol. Cell. Biol.
17:2735-2744[Abstract].
|
| 23.
|
Hong, H.,
K. Kohli,
A. Trivedi,
D. L. Johnson, and M. R. Stallcup.
1996.
GRIP1, a novel mouse protein that serves as a transcriptional coactivator in yeast for the hormone binding domains of steroid receptors.
Proc. Natl. Acad. Sci. USA
93:4948-4952[Abstract/Free Full Text].
|
| 24.
|
Hrimech, M.,
X. J. Yao,
F. Bachand,
N. Rougeau, and E. A. Cohen.
1999.
Human immunodeficiency virus type 1 (HIV-1) Vpr functions as an immediate-early protein during HIV-1 infection.
J. Virol.
73:4101-4109[Abstract/Free Full Text].
|
| 25.
|
Jenkins, Y.,
M. McEntee,
K. Weis, and W. C. Greene.
1998.
Characterization of HIV-1 vpr nuclear import: analysis of signals and pathways.
J. Cell Biol.
143:875-885[Abstract/Free Full Text].
|
| 26.
|
Jowett, J. B.,
V. Planelles,
B. Poon,
N. P. Shah,
M. L. Chen, and I. S. Chen.
1995.
The human immunodeficiency virus type 1 vpr gene arrests infected T cells in the G2 + M phase of the cell cycle.
J. Virol.
69:6304-6313[Abstract].
|
| 27.
|
Karin, M.
1998.
New twists in gene regulation by glucocorticoid receptor: is DNA binding dispensable?
Cell
93:487-490[CrossRef][Medline].
|
| 28.
|
Kino, T.,
A. Gragerov,
J. B. Kopp,
R. H. Stauber,
G. N. Pavlakis, and G. P. Chrousos.
1999.
The HIV-1 virion-associated protein vpr is a coactivator of the human glucocorticoid receptor.
J. Exp. Med.
189:51-62[Abstract/Free Full Text].
|
| 29.
|
Kondo, E.,
F. Mammano,
E. A. Cohen, and H. G. Gottlinger.
1995.
The p6gag domain of human immunodeficiency virus type 1 is sufficient for the incorporation of Vpr into heterologous viral particles.
J. Virol.
69:2759-2764[Abstract].
|
| 30.
|
Levy, D. N.,
Y. Refaeli,
R. R. MacGregor, and D. B. Weiner.
1994.
Serum Vpr regulates productive infection and latency of human immunodeficiency virus type 1.
Proc. Natl. Acad. Sci. USA
91:10873-10877[Abstract/Free Full Text].
|
| 31.
|
Levy, D. N.,
Y. Refaeli, and D. B. Weiner.
1995.
Extracellular Vpr protein increases cellular permissiveness to human immunodeficiency virus replication and reactivates virus from latency.
J. Virol.
69:1243-1252[Abstract].
|
| 32.
|
Liu, W.,
J. Wang,
N. K. Sauter, and D. Pearce.
1995.
Steroid receptor heterodimerization demonstrated in vitro and in vivo.
Proc. Natl. Acad. Sci. USA
92:12480-12484[Abstract/Free Full Text].
|
| 33.
|
Liu, W.,
J. Wang,
G. Yu, and D. Pearce.
1996.
Steroid receptor transcriptional synergy is potentiated by disruption of the DNA-binding domain dimer interface.
Mol. Endocrinol.
10:1399-1406[Abstract/Free Full Text].
|
| 34.
|
Lu, Y.-L.,
R. P. Bennett,
J. W. Wills,
R. Gorelick, and L. Ratner.
1995.
A leucine triplet repeat sequence (LXX)4 in p6gag is important for Vpr incorporation into human immunodeficiency virus type 1 particles.
J. Virol.
69:6873-6879[Abstract].
|
| 35.
|
Luo, Z.,
D. J. Butcher,
R. Murali,
A. Srinivasan, and Z. Huang.
1998.
Structural studies of synthetic peptide fragments derived from the HIV-1 Vpr protein.
Biochem. Biophys. Res. Commun.
244:732-736[CrossRef][Medline].
|
| 36.
|
Mahalingam, S.,
V. Ayyavoo,
M. Patel,
T. Kieber-Emmons, and D. B. Weiner.
1997.
Nuclear import, virion incorporation, and cell cycle arrest/differentiation are mediated by distinct functional domains of human immunodeficiency virus type 1 Vpr.
J. Virol.
71:6339-6347[Abstract].
|
| 37.
|
Markham, P. D.,
S. Z. Salahuddin,
K. Veren,
S. Orndorff, and R. C. Gallo.
1986.
Hydrocortisone and some other hormones enhance the expression of HTLV-III.
Int. J. Cancer
37:67-72[Medline].
|
| 38.
|
McInerney, E. M.,
D. W. Rose,
S. E. Flynn,
S. Westin,
T. M. Mullen,
A. Krones,
J. Inostroza,
J. Torchia,
R. T. Nolte,
N. Assa-Munt,
M. V. Milburn,
C. K. Glass, and M. G. Rosenfeld.
1998.
Determinants of coactivator LXXLL motif specificity in nuclear receptor transcriptional activation.
Genes Dev.
12:3357-3368[Abstract/Free Full Text].
|
| 39.
|
Nakaya, T.,
K. Fujinaga,
M. Kishi,
S. Oka,
T. Kurata,
I. M. Jones, and K. Ikuta.
1994.
Nonsense mutations in the vpr gene of HIV-1 during in vitro virus passage and in HIV-1 carrier-derived peripheral blood mononuclear cells.
FEBS Lett.
354:17-22[CrossRef][Medline].
|
| 40.
|
Paxton, W.,
R. I. Connor, and N. R. Landau.
1993.
Incorporation of Vpr into human immunodeficiency virus type 1 virions: requirement for the p6 region of gag and mutational analysis.
J. Virol.
67:7229-7237[Abstract/Free Full Text].
|
| 41.
|
Poon, B.,
K. Grovit-Ferbas,
S. A. Stewart, and I. S. Y. Chen.
1998.
Cell cycle arrest by Vpr in HIV-1 virions and insensitivity to antiretroviral agents.
Science
281:266-269[Abstract/Free Full Text].
|
| 42.
|
Popov, S.,
M. Rexach,
L. Ratner,
G. Blobel, and M. Bukrinsky.
1998.
Viral protein R regulates docking of the HIV-1 preintegration complex to the nuclear pore complex.
J. Biol. Chem.
273:13347-13352[Abstract/Free Full Text].
|
| 43.
|
Re, F.,
D. Braaten,
E. K. Franke, and J. Luban.
1995.
Human immunodeficiency virus type 1 Vpr arrests the cell cycle in G2 by inhibiting the activation of p34cdc2-cyclin B.
J. Virol.
69:6859-6864[Abstract].
|
| 44.
|
Refaeli, Y.,
D. N. Levy, and D. B. Weiner.
1995.
The glucocorticoid receptor type II complex is a target of the HIV-1 vpr gene product.
Proc. Natl. Acad. Sci. USA
92:3621-3625[Abstract/Free Full Text].
|
| 45.
|
Reichardt, H. M.,
K. H. Kaestner,
J. Tuckermann,
O. Kretz,
O. Wessely,
R. Bock,
P. Gass,
W. Schmid,
P. Herrlich,
P. Angel, and G. Schutz.
1998.
DNA binding of the glucocorticoid receptor is not essential for survival.
Cell
93:531-541[CrossRef][Medline].
|
| 46.
|
Rogel, M. E.,
L. I. Wu, and M. Emerman.
1995.
The human immunodeficiency virus type 1 vpr gene prevents cell proliferation during chronic infection.
J. Virol.
69:882-888[Abstract].
|
| 47.
|
Schmidt, S.,
A. Baniahmad,
M. Eggert,
S. Schneider, and R. Renkawitz.
1998.
Multiple receptor interaction domains of GRIP1 function in synergy.
Nucleic Acids Res.
26:1191-1197[Abstract/Free Full Text].
|
| 48.
|
Schuler, W.,
K. Wecker,
H. de Rocquigny,
Y. Baudat,
J. Sire, and B. P. Roques.
1999.
NMR structure of the (52-96) C-terminal domain of the HIV-1 regulatory protein Vpr: molecular insights into its biological functions.
J. Mol. Biol.
285:2105-2117[CrossRef][Medline].
|
| 49.
|
Soudeyns, H.,
R. Geleziunas,
G. Shyamala,
J. Hiscott, and M. A. Wainberg.
1993.
Identification of a novel glucocorticoid response element within the genome of the human immunodeficiency virus type 1.
Virology
194:758-768[CrossRef][Medline].
|
| 50.
|
Soudeyns, H., and M. A. Wainberg.
1997.
Effects of RU486 on HIV-1 replication.
Nat. Med.
3:1302-1303.
|
| 51.
|
Stewart, S. A.,
B. Poon,
J. B. Jowett,
Y. Xie, and I. S. Chen.
1999.
Lentiviral delivery of HIV-1 Vpr protein induces apoptosis in transformed cells.
Proc. Natl. Acad. Sci. USA
96:12039-12043[Abstract/Free Full Text].
|
| 52.
|
Stewart, S. A.,
B. Poon,
J. Y. Song, and I. S. Chen.
2000.
Human immunodeficiency virus type 1 vpr induces apoptosis through caspase activation.
J. Virol.
74:3105-3111[Abstract/Free Full Text].
|
| 53.
|
Subbramanian, R. A.,
A. Kessous-Elbaz,
R. Lodge,
J. Forget,
X. J. Yao,
D. Bergeron, and E. A. Cohen.
1998.
Human immunodeficiency virus type 1 Vpr is a positive regulator of viral transcription and infectivity in primary human macrophages.
J. Exp. Med.
187:1103-1111[Abstract/Free Full Text].
|
| 54.
|
Torchia, J.,
C. Glass, and M. G. Rosenfeld.
1998.
Co-activators and co-repressors in the integration of transcriptional responses.
Curr. Opin. Cell Biol.
10:373-383[CrossRef][Medline].
|
| 55.
|
Tristem, M.,
C. Marshall,
A. Karpas, and F. Hill.
1992.
Evolution of the primate lentiviruses: evidence from vpx and vpr.
EMBO J.
11:3405-3412[Medline].
|
| 56.
|
Vodicka, M. A.,
D. M. Koepp,
P. A. Silver, and M. Emerman.
1998.
HIV-1 Vpr interacts with the nuclear transport pathway to promote macrophage infection.
Genes Dev.
12:175-185[Abstract/Free Full Text].
|
| 57.
|
Wang, L.,
S. Mukherjee,
F. Jia,
O. Narayan, and L. J. Zhao.
1995.
Interaction of virion protein Vpr of human immunodeficiency virus type 1 with cellular transcription factor Sp1 and trans-activation of viral long terminal repeat.
J. Biol. Chem.
270:25564-25569[Abstract/Free Full Text].
|
| 58.
|
Wang, L.,
S. Mukherjee,
O. Narayan, and L. J. Zhao.
1996.
Characterization of a leucine-zipper-like domain in Vpr protein of human immunodeficiency virus type 1.
Gene
178:7-13[CrossRef][Medline].
|
| 59.
|
Yao, X. J.,
A. J. Mouland,
R. A. Subbramanian,
J. Forget,
N. Rougeau,
D. Bergeron, and E. A. Cohen.
1998.
Vpr stimulates viral expression and induces cell killing in human immunodeficiency virus type 1-infected dividing Jurkat T cells.
J. Virol.
72:4686-4693[Abstract/Free Full Text].
|
| 60.
|
Yao, X. J.,
R. A. Subbramanian,
N. Rougeau,
F. Boisvert,
D. Bergeron, and E. A. Cohen.
1995.
Mutagenic analysis of human immunodeficiency virus type 1 Vpr: role of a predicted N-terminal alpha-helical structure in Vpr nuclear localization and virion incorporation.
J. Virol.
69:7032-7044[Abstract].
|
| 61.
|
Yedavalli, V. R.,
C. Chappey, and N. Ahmad.
1998.
Maintenance of an intact human immunodeficiency virus type 1 vpr gene following mother-to-infant transmission.
J. Virol.
72:6937-6943[Abstract/Free Full Text].
|
| 62.
|
Yuan, X.,
Z. Matsuda,
M. Matsuda,
M. Essex, and T. H. Lee.
1990.
Human immunodeficiency virus vpr gene encodes a virion-associated protein.
AIDS Res. Hum. Retroviruses
6:1265-1271[Medline].
|
| 63.
|
Zhao, L. J.,
S. Mukherjee, and O. Narayan.
1994.
Biochemical mechanism of HIV-I Vpr function: specific interaction with a cellular protein.
J. Biol. Chem.
269:15577-15582[Abstract/Free Full Text].
|
| 64.
|
Zhao, L. J.,
L. Wang,
S. Mukherjee, and O. Narayan.
1994.
Biochemical mechanism of HIV-1 Vpr function: oligomerization mediated by the N-terminal domain.
J. Biol. Chem.
269:32131-32137[Abstract/Free Full Text].
|
Journal of Virology, September 2000, p. 8159-8165, Vol. 74, No. 17
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-
Kino, T., Slobodskaya, O., Pavlakis, G. N., Chrousos, G. P.
(2002). Nuclear Receptor Coactivator p160 Proteins Enhance the HIV-1 Long Terminal Repeat Promoter by Bridging Promoter-bound Factors and the Tat-P-TEFb Complex. J. Biol. Chem.
277: 2396-2405
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Abstract
Introduction
