CpG protects human monocytic cells against HIV-Vpr-induced apoptosis by cellular inhibitor of apoptosis-2 through the calcium-activated JNK pathway in a TLR9-independent manner

Monocytic cells survive HIV replication and consequent cytopathic effects because of their decreased sensitivity to HIV-induced apoptosis. However, the mechanism underlying this resistance to apoptosis remains poorly understood. Lymphocytic cells are exposed to microbial products because of their translocation from the gut in persons with chronic HIV infections or following coinfections. We hypothesized that activation of monocytic cells by such microbial products through interaction with corresponding TLRs may confer antiapoptotic signals. Using HIV-viral protein R (Vpr)(52-96) peptide as a model apoptosis-inducing agent, we demonstrated that unlike monocyte-derived macrophages, undifferentiated primary human monocytes and promonocytic THP-1 cells are highly susceptible to Vpr(52-96)-induced apoptosis. Interestingly, monocytes and THP-1 cells stimulated with TLR9 agonist CpG induced almost complete resistance to Vpr(52-96)-induced apoptosis, albeit through a TLR9-independent signaling pathway. Moreover, CpG selectively induced the antiapoptotic cellular inhibitor of apoptosis (c-IAP)-2 protein and inhibition of the c-IAP-2 gene by either specific small interfering RNA or synthetic second mitochondrial activator of caspases mimetic reversed CpG-induced resistance against Vpr(52-96)-mediated apoptosis. We demonstrated that c-IAP-2 is regulated by the JNK and calcium signaling pathway, in particular calmodulin-dependent protein kinase-II. Furthermore, inhibition of JNK and the calcium signaling including the calmodulin-dependent protein kinase-II by either pharmacological inhibitors or their specific small interfering RNAs reversed CpG-induced protection against Vpr(52-96)-mediated apoptosis. We also show that CpG induced JNK phosphorylation through activation of the calcium signaling pathway. Taken together, our results suggest that CpG-induced protection may be mediated by c-IAP-2 through the calcium-activated JNK pathway via what appeared to be TLR9-independent signaling pathways.     

Depletion of Wee-1 kinase is necessary for both human immunodeficiency virus type 1 Vpr- and gamma irradiation-induced apoptosis

Human immunodeficiency virus (HIV) protein R (Vpr) induces G2 arrest, and prolonged G2 arrest leads to apoptosis. We find that in HeLa cells the cell cycle regulatory kinase, Wee-1, is depleted following prolonged G2 arrest induced by Vpr. Of note, small interfering RNAs directed to Wee-1 triggered apoptosis, suggesting a direct role for Wee-1 in apoptosis. In support of this hypothesis, overexpression of Wee-1 suppressed Vpr-mediated apoptosis. Importantly, similar results were observed with cells induced to undergo apoptosis gamma irradiation. Thus, Wee-1 may serve as a key regulator of both HIV type 1 Vpr- and gamma irradiation-mediated apoptosis and possibly serve as a general regulator linking the cell cycle to some pathways of apoptosis.     

The cationic amphipathic alpha-helix of HIV-1 viral protein R (Vpr) binds to nucleic acids,permeabilizes membranes,and efficiently transfects cells

Viral protein R (Vpr) is a small protein of 96 amino acids that is conserved among the lentiviruses human immunodeficiency virus type 1 (HIV-1), HIV-2, and simian immunodeficiency virus. We recently sought to determine whether the karyophilic properties of Vpr, as well as its ability to bind nucleic acids, could be used to deliver DNA into cells. We have found that the C-terminal domain of Vpr-(52-96) is able to efficiently transfect various cell lines. Here, we show that the shortest active sequence for gene transfer corresponds to the domain that adopts a alpha-helix conformation. DNA binding studies and permeabilization assays performed on cells demonstrated that the peptides that are efficient in transfection condense plasmid DNA and are membranolytic. Electron microscopy studies and transfection experiments performed in the presence of inhibitors of the endocytic processes indicated that the major entry pathway of Vpr-DNA complexes is through endocytosis. Taken together, the results show that the cationic C-terminal alpha-helix of Vpr has DNA-condensing as well as membrane-destabilizing capabilities, both properties that are indispensable for efficient DNA transfection.     

The human immunodeficiency virus type 1 Vpr protein and its carboxy-terminally truncated form induce apoptosis in tumor cells

The human immunodeficiency virus type 1 (HIV-1) accessory protein Vpr induces apoptosis after cell cycle arrest at the G₂ phase in primate cells. We have reported previously that C81, a carboxy-terminally truncated form of Vpr, interferes with cell proliferation and results in apoptosis without G₂ arrest. Here, we investigated whether this property of Vpr and C81 could be exploited for use as a potential anticancer agent. First, we demonstrated that C81 induced G₁ arrest and apoptosis in all tumor cells tested. In contrast, Vpr resulted in G₂ arrest and apoptosis in HeLa and 293 T cells. Vpr also suppressed the damaged-DNA-specific binding protein 1 (DDB1) in HepG2 cells, thereby inducing apoptosis without G₂ arrest. G₂ arrest was restored when DDB1 was overexpressed in cells that also expressed Vpr. Surprisingly, C81 induced G₂ arrest when DDB1 was overexpressed in HepG2 cells, but not in HeLa or 293 T cells. Thus, the induction of Vpr- and C81-mediated cell cycle arrest appears to depend on the cell type, whereas apoptosis was observed in all tumor cells tested. Overall, Vpr and C81 have potential as novel therapeutic agents for treatment of cancer.     

A competition model for viral inhibition of host cell proliferation

Some viruses encode proteins that promote cell proliferation while others, such as the human immunodeficiency virus (HIV), encode proteins that prevent cell division. It has been hypothesized that the selective advantage determining which strategy evolves depends on the ability of the virus to induce a cellular environment which maximizes both virus production and cell life span. In HIV, the protein that causes cell cycle arrest is Vpr. In this paper, we develop a mathematical model, based on difference equations, to study the competition between two genotypes of HIV - one that encodes this protein (Vpr+) and one that does not (Vpr-). In particular, we are interested in parameters that could be different between the in vitro condition, where the Vpr- genotype dominates, and the in vivo condition, where the Vpr+ genotype dominates. Our model indicates that the infected cell death-rate, the viral half-life, and the dynamics of the target cell population all effect the competition dynamics between the Vpr+ and Vpr- viral genotypes. Perturbing any of these parameters from the in vitro estimates while holding the others fixed has no affect on the competition outcome, i. e., the Vpr- genotype dominates. Perturbing the infected cell death-rate and the target cell source causes a switch in competitive outcome, although not necessarily at values, which represent the in vivo condition. Adding a perturbation in the viral half-life from in vitro to in vivo condition results in a switch of the competitive advantage from the Vpr- genotype to the Vpr+ genotype with parameters for all three mechanisms set to estimated in vivo values.     

Viral protein R regulates nuclear import of the HIV-1 pre-integration complex.

Replication of human immunodeficiency virus type 1 (HIV-1) in non-dividing cells critically depends on import of the viral pre-integration complex into the nucleus. Genetic evidence suggests that viral protein R (Vpr) and matrix antigen (MA) are directly involved in the import process. An in vitro assay that reconstitutes nuclear import of HIV-1 pre-integration complexes in digitonin-permeabilized cells was used to demonstrate that Vpr is the key regulator of the viral nuclear import process. Mutant HIV-1 pre-integration complexes that lack Vpr failed to be imported in vitro, whereas mutants that lack a functional MA nuclear localization sequence (NLS) were only partially defective. Strikingly, the import defect of the Vpr- mutant was rescued when recombinant Vpr was re-added. In addition, import of Vpr- virus was rescued by adding the cytosol of HeLa cells, where HIV-1 replication had been shown to be Vpr-independent. In a solution binding assay, Vpr associated with karyopherin alpha, a cellular receptor for NLSs. This association increased the affinity of karyopherin alpha for basic-type NLSs, including that of MA, thus explaining the positive effect of Vpr on nuclear import of the HIV-1 pre-integration complex and BSA-NLS conjugates. These results identify the biochemical mechanism of Vpr function in transport of the viral pre-integration complex to, and across, the nuclear membrane.     

Human immunodeficiency virus type 1 Vpr induces apoptosis in human neuronal cells

Human immunodeficiency virus type 1 (HIV-1) infection of the central nervous system (CNS) causes AIDS dementia complex (ADC) in certain infected individuals. Recent studies have suggested that patients with ADC have an increased incidence of neuronal apoptosis leading to neuronal dropout. Of note, a higher level of the HIV-1 accessory protein Vpr has been detected in the cerebrospinal fluid of AIDS patients with neurological disorders. Moreover, extracellular Vpr has been shown to form ion channels, leading to cell death of cultured rat hippocampal neurons. Based on these previous findings, we first investigated the apoptotic effects of the HIV-1 Vpr protein on the human neuronal precursor NT2 cell line at a range of concentrations. These studies demonstrated that apoptosis induced by both Vpr and the envelope glycoprotein, gp120, occurred in a dose-dependent manner compared to protein treatment with HIV-1 integrase, maltose binding protein (MBP), and MBP-Vpr in the undifferentiated NT2 cells. For mature, differentiated neurons, apoptosis was also induced in a dose-dependent manner by both Vpr and gp120 at concentrations ranging from 1 to 100 ng/ml, as demonstrated by both the terminal deoxynucleotidyltransferase (Tdt)-mediated dUTP-biotin nick end labeling and Annexin V assays for apoptotic cell death. In order to clarify the intracellular pathways and molecular mechanisms involved in Vpr- and gp120-induced apoptosis in the NT2 cell line and differentiated mature human neurons, we then examined the cellular lysates for caspase-8 activity in these studies. Vpr and gp120 treatments exhibited a potent increase in activation of caspase-8 in both mature neurons and undifferentiated NT2 cells. This suggests that Vpr may be exerting selective cytotoxicity in a neuronal precursor cell line and in mature human neurons through the activation of caspase-8. These data represent a characterization of Vpr-induced apoptosis in human neuronal cells, and suggest that extracellular Vpr, along with other lentiviral proteins, may increase neuronal apoptosis in the CNS. Also, identification of the intracellular activation of caspase-8 in Vpr-induced apoptosis of human neuronal cells may lead to therapeutic approaches which can be used to combat HIV-1-induced neuronal apoptosis in AIDS patients with ADC.     

Critical role for antiapoptotic Bcl-xL and Mcl-1 in human macrophage survival and cellular IAP1/2 (cIAP1/2) in resistance to HIV-Vpr-induced apoptosis

Macrophages are resistant to HIV cytopathic effects, which contributes to viral persistence and reservoir formation. HIV viral protein R (Vpr) is a potent apoptosis-inducing agent for primary monocytes. Because the biologically active Vpr is found in serum and cerebrospinal fluid of HIV-infected patients, we investigated the apoptotic effect of Vpr on monocyte-derived macrophages and phorbol 12-myristate 13-acetate-activated THP1 macrophages. Our results show that primary monocytes and THP1 cells develop resistance to Vpr-induced apoptosis following differentiation into macrophages. To determine the effect of Vpr on the expression of antiapoptotic proteins, we show that in contrast to the undifferentiated cells, Vpr did not down-regulate the expression of antiapoptotic inhibitors of apoptosis (IAPs) and Bcl2 family members in macrophages, suggesting their involvement in resistance to Vpr-induced apoptosis. However, knocking down Bcl-xL and Mcl-1 proteins induced spontaneous apoptosis with no impact on susceptibility to Vpr-induced apoptosis. In contrast, down-regulation of cellular IAP1 (cIAP1) and cIAP2 by using siRNAs and SMAC (second mitochondria-derived activator of caspases) mimetic sensitized macrophages to Vpr-induced apoptosis. Overall, our results suggest that resistance to Vpr-induced apoptosis is specifically mediated by cIAP1/2 genes independent of Bcl-xL and Mcl-1, which play a key role in maintaining cell viability. Moreover, IAP modulation may be a potential strategy to eliminate HIV persistence in macrophages.     

Requirement of the Pr55gag precursor for incorporation of the Vpr product into human immunodeficiency virus type 1 viral particles.

The human immunodeficiency virus type 1 (HIV-1) particles consists of two molecules of genomic RNA as well as molecules originating from gag, pol, and env products, all synthesized as precursor proteins. The 96-amino-acid Vpr protein, the only virion-associated HIV-1 regulatory protein, is not part of the virus polyprotein precursors, and its incorporation into virus particles must occur by way of an interaction with a component normally found in virions. To investigate the mechanism of incorporation of Vpr into the HIV-1 virion, Vpr- proviral DNA constructs harboring mutations or deletions in specific virion-associated gene products were cotransfected with Vpr expressor plasmids in COS cells. Virus released from the transfected cells was tested for the presence of Vpr by immunoprecipitation with Vpr-specific antibodies. The results of these experiments show that Vpr is trans-incorporated into virions but at a lower efficiency than when Vpr is expressed from a proviral construct. The minimal viral genetic information necessary for Vpr incorporation was a deleted provirus encoding only the pr55gag polyprotein precursor. Incorporation of Vpr requires the expression but not the processing of gag products and is independent of pol and env expression. Direct interaction of Vpr with the Pr55gag precursor protein was demonstrated by coprecipitation experiments with gag product-specific antibodies. Overall, these results indicate that HIV-1 Vpr is incorporated into the nascent virion through an interaction with the Gag precursor polyprotein and demonstrate a novel mechanism by which viral protein can be incorporated into virus particles.     

Efficient DNA transfection mediated by the C-terminal domain of human immunodeficiency virus type 1 viral protein R

Viral protein R (Vpr) of human immunodeficiency virus type 1 is produced late in the virus life cycle and is assembled into the virion through binding to the Gag protein. It is known to play a significant role early in the viral life cycle by facilitating the nuclear import of the preintegration complex in nondividing cells. Vpr is also able to interact with nucleic acids, and we show here that it induces condensation of plasmid DNA. We have explored the possibility of using these properties in DNA transfection experiments. We report that the C-terminal half of the protein (Vpr(52-96)) mediates DNA transfection in a variety of human and nonhuman cell lines with efficiencies comparable to those of the best-known transfection agents. Compared with polylysine, a standard polycationic transfection reagent, Vpr(52-96) was 10- to 1,000-fold more active. Vpr(52-96)-DNA complexes were able to reach the cell nucleus through a pH-independent mechanism. These observations possibly identify an alternate pathway for DNA transfection.     

HIV-1 Vpr induces apoptosis through caspase 9 in T cells and peripheral blood mononuclear cells

Human immunodeficiency virus, type 1 (HIV-1), vpr gene encodes a 14-kDa virion-associated protein, which exhibits significant effects on human cells. One important property of Vpr is its ability to induce apoptosis during infection. Apoptotic induction is likely to play a role in the pathogenesis of AIDS. However, the pathway of apoptosis is not clearly defined. In this report we investigate the mechanism of apoptosis induced by HIV-1 Vpr using a Vpr pseudotype viral infection system or adeno delivery of Vpr in primary human lymphoid cells and T-cells. With either vector, HIV-1 Vpr induced cell cycle arrest at the G(2)/M phase and apoptosis in lymphoid target cells. Furthermore, we observed that with both vectors, caspase 9, but not caspase 8, was activated following infection of human peripheral blood mononuclear cell with either Vpr-positive HIV virions or adeno-delivered Vpr. Activation of the caspase 9 pathway resulted in caspase 3 activation and apoptosis in human primary cells. These effects were coincident with the disruption of the mitochondrial transmembrane potential and induction of cytochrome c release by Vpr. The Vpr-induced signaling pathway did not induce CD95 or CD95L expression. Bcl-2 overexpressing cells succumb to Vpr-induced apoptosis. These studies illustrate that Vpr induces a mitochondria-dependent apoptotic pathway that is distinct from apoptosis driven by the Fas-FasL pathway.     

Curcumin inhibits 19-kDa lipoprotein of Mycobacterium tuberculosis induced macrophage apoptosis via regulation of the JNK pathway

Recently, synthetic curcumin analogs are reported as potential active compounds against Mycobacterium tuberculosis (MTB). During the process of MTB infection, macrophages show increased apoptosis. The candidate virulence factors such as 19-kDa lipoprotein secreted by the MTB (P19) strongly influences macrophages by activation of Toll-like receptor 2 (TLR2) and mitogen-activated protein kinases (MAPKs). It has been reported that curcumin could affect the apoptosis of tumor cells via regulation of MAPKs. However, its effect on the P19-induced apoptosis of macrophages is unclear. This study investigates the effect of curcumin on the MAPKs signaling and apoptosis in human macrophages. The results showed that curcumin and P19 induced macrophage apoptosis in a time- and dose-dependent manner Low doses of curcumin (10 and 20 μM) protected macrophages from P19 induced apoptosis, accompanied by decreased production of cytokines and reduced activation of the c-Jun amino-terminal kinase (JNK) and p38 MAPK. The protective effect of curcumin on P19 induced apoptosis of macrophages were enhanced by treatment with the JNK-specific inhibitors, whereas SB203580, the inhibitor of p38 MAPK had no effect. Curcumin had no effect on the activity of extracellular signal-regulated protein kinase (ERK). Taken together, our data show that the JNK pathway, but not the p38 or ERK pathway, plays an important role in the protective effect of curcumin against P19 induced macrophage apoptosis, and regulation of the JNK pathway may partially elucidate the anti-tuberculosis activity of curcumin.     

Platycodin D induces anoikis and caspase‐mediated apoptosis via p38 MAPK in AGS human gastric cancer cells

Mitogen‐activated protein kinases (MAPKs) cascades play important roles in cell proliferation, death, and differentiation in response to external stimuli. However, the precise role of MAPKs in platycodin D (PD)‐induced cytotoxicity remains unclear. In this study, we investigated the anticancer effect of PD and its underlying mechanism on AGS human gastric cancer cells. PD significantly inhibited cell proliferation and induced anoikis, which is a form of apoptosis in which cells detach from the substrate. It showed phosphatidylserine externalization, DNA fragmentation, increase of sub‐G1 phase, and activation of caspases in a dose‐ and time‐dependent manner. This apoptosis has been associated with the extrinsic pathway via Fas‐L and the intrinsic pathway via mitochondrial Bcl‐2 family members. Moreover, PD led to the phosphorylation of stresses‐activated protein kinases such as JNK and p38, followed by the activation of AP‐1. However, pretreatment with SB203580 (a p38 specific inhibitor) suppressed PD‐induced p38 and AP‐1 activation, and subsequently attenuated the PD‐induced apoptosis in AGS cells. These results suggest that p38 activation is responsible for PD‐induced apoptosis in AGS cells and PD might be useful for the development as the anticancer agent of gastric cancer. J. Cell. Biochem. 114: 456–470, 2013. © 2012 Wiley Periodicals, Inc.     

MAPKs and NF-κB-mediated acrylamide-induced neuropathy in rat striatum and human neuroblastoma cells SY5Y

Acrylamide (ACR) is a potent neurotoxin that can be produced during high-temperature food processing, but the underlying toxicological mechanism remains unclear. In this study, the detrimental effects of ACR on the striatal dopaminergic neurons and the roles of mitogen-activated protein kinases (MAPKs) and nuclear factor κB (NF-κB) in ACR-induced neuronal apoptosis were investigated. Acute ACR exposure caused dopaminergic neurons loss and apoptosis as revealed by decreased tyrosine hydroxylase (TH)-positive cells and TH protein level and increased terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL)-positive cells in the striatum. ACR-decreased glutathione content, increased levels of malondialdehyde, proinflammatory cytokines tumor necrosis factor α, and interleukin 6. In addition, nuclear NF-κB and MAPKs signaling pathway with c-Jun N-terminal kinase (JNK) and p38 were activated by ACR. Specific inhibitors were used to explore the roles of MAPKs and NF-κB pathways in ACR-induced apoptosis in SH-SY5Y cells. Pretreatment with JNK-specific inhibitors SP600125 markedly upregulated the reduced B-cell lymphoma 2 (Bcl-2) content and downregulated the increased Bcl-2-associated X protein (Bax) level and thereby eventually reduced the proportions of early and late apoptotic cells induced by ACR, while p38 suppression by SB202190 only reversed the decrease in Bcl-2 expression. Inhibition of NF-κB by BAY 11-7082 markedly upregulated Bax level and decreased Bcl-2 expression, and eventually increasing the proportions of neuronal apoptosis compared with that in ACR alone. These results suggested that JNK contributed to ACR-induced apoptosis, while NF-κB acted as a protective regulator in response to ACR-induced neuropathy. This study helps to offer a deeper insight into the mechanism of ACR-induced neuropathy.     

Human immunodeficiency virus type 1 Vpr interacts with antiapoptotic mitochondrial protein HAX-1

Human immunodeficiency virus type 1 viral protein R (Vpr) is required for viral pathogenesis and has been implicated in T-cell apoptosis through its activation of caspase 3 and caspase 9 and perturbation of mitochondrial membrane potential. To understand better Vpr-mitochondria interaction, we report here the identification of antiapoptotic mitochondrial protein HAX-1 as a novel Vpr target. We show that Vpr and HAX-1 physically associate with each other. Overexpression of Vpr in cells dislocates HAX-1 from its normal residence in mitochondria and creates mitochondrion instability and cell death. Conversely, overexpression of HAX-1 suppressed the proapoptotic activity of Vpr.     

A growth factor attenuates HIV-1 Tat and morphine induced damage to human neurons: implication in HIV/AIDS-drug abuse cases

The neuropathological abnormalities of human immunodeficiency virus (HIV)-1 patients abusing illicit drugs suggest extensive interactions between the two agents, thereby leading to increased rate of progression to neurodegeneration. The role of HIV-1 transactivating protein, Tat has been elucidated in mediating neuronal damage via apoptosis, a hallmark of HIV-associated dementia (HAD), however the underlying mechanisms involved in enhanced neurodegeneration by illicit drugs remain elusive. In this study, we demonstrated that morphine enhances HIV-Tat induced toxicity in human neurons and neuroblastoma cells. Enhanced toxicity by Tat and morphine was accompanied by increased numbers of TUNEL positive apoptotic neurons, elevated caspase-3 levels and decreased ratio of anti- and pro-apoptotic proteins, Bcl2/Bax. Tat and morphine together elicited high levels of reactive oxygen species that were NADPH dependent. Significant alterations in mitochondrial membrane homeostasis were also observed with co-exposure of these agents. Extensive studies of mitogen activated protein kinase (MAPK) signaling pathways revealed the involvement of c-Jun N-terminal kinase (JNK) and extracellular signal-regulated kinase-1/2 (ERK1/2) pathways in enhanced toxicity of Tat and morphine. In addition to this, we found that pre-treatment of cells with platelet derived growth factor (PDGF-BB) protected neurons from HIV-Tat and morphine induced damage. PDGF-BB alleviated ROS production, maintained mitochondrial membrane potential, decreased caspase-3 activation and hence protected the cells from undergoing apoptosis. PDGF-BB mediated protection against Tat and morphine involved the phosphatidylinositol-3 kinase (PI3K) pathway, as specific inhibitor of PI3K abrogated the protection conferred by PDGF-BB. This study demonstrates the mechanism of enhanced toxicity in human neurons subjected to co-exposure of HIV protein Tat and morphine, thus implying its importance in HIV positive drug abusers, where damage to the brain is reported to be more severe than non-drug abusers. We have also showed for the first time that PDGF-BB can protect against simultaneous exposure of Tat and morphine, strengthening its role as a neuroprotective agent that could be considered for therapeutic intervention.     

Extracellular Vpr protein increases cellular permissiveness to human immunodeficiency virus replication and reactivates virus from latency.

The vpr gene product of human immunodeficiency virus (HIV) and simian immunodeficiency virus is a virion-associated regulatory protein that has been shown using vpr mutant viruses to increase virus replication, particularly in monocytes/macrophages. We have previously shown that vpr can directly inhibit cell proliferation and induce cell differentiation, events linked to the control of HIV replication, and also that the replication of a vpr mutant but not that of wild-type HIV type 1 (HIV-1) was compatible with cellular proliferation (D. N. Levy, L. S. Fernandes, W. V. Williams, and D. B. Weiner, Cell 72:541-550, 1993). Here we show that purified recombinant Vpr protein, in concentrations of < 100 pg/ml to 100 ng/ml, increases wild-type HIV-1 replication in newly infected transformed cell lines via a long-lasting increase in cellular permissiveness to HIV replication. The activity of extracellular Vpr protein could be completely inhibited by anti-Vpr antibodies. Extracellular Vpr also induced efficient HIV-1 replication in newly infected resting peripheral blood mononuclear cells. Extracellular Vpr transcomplemented a vpr mutant virus which was deficient in replication in promonocytic cells, restoring full replication competence. In addition, extracellular Vpr reactivated HIV-1 expression in five latently infected cell lines of T-cell, B-cell, and promonocytic origin which normally express very low levels of HIV RNA and protein, indicating an activation of translational or pretranslational events in the virus life cycle. Together, these results describe a novel pathway governing HIV replication and a potential target for the development of anti-HIV therapeutics.