scholarly journals Human Immunodeficiency Virus Type 1-Induced Macrophage Gene Expression Includes the p21 Gene, a Target for Viral Regulation

2005 ◽  
Vol 79 (7) ◽  
pp. 4479-4491 ◽  
Author(s):  
Nancy Vázquez ◽  
Teresa Greenwell-Wild ◽  
Nancy J. Marinos ◽  
William D. Swaim ◽  
Salvador Nares ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Viral Replication ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Kinase Inhibitor ◽  
Peroxisome Proliferator ◽  
Small Interfering Rnas ◽  
Immunodeficiency Virus ◽  
Hiv 1

ABSTRACT In contrast to CD4+ T cells, human immunodeficiency virus type 1 (HIV-1)-infected macrophages typically resist cell death, support viral replication, and consequently, may facilitate HIV-1 transmission. To elucidate how the virus commandeers the macrophage's intracellular machinery for its benefit, we analyzed HIV-1-infected human macrophages for virus-induced gene transcription by using multiple parameters, including cDNA expression arrays. HIV-1 infection induced the transcriptional regulation of genes associated with host defense, signal transduction, apoptosis, and the cell cycle, among which the cyclin-dependent kinase inhibitor 1A (CDKN1A/p21) gene was the most prominent. p21 mRNA and protein expression followed a bimodal pattern which was initially evident during the early stages of infection, and maximum levels occurred concomitant with active HIV-1 replication. Mechanistically, viral protein R (Vpr) independently regulates p21 expression, consistent with the reduced viral replication and lack of p21 upregulation by a Vpr-negative virus. Moreover, the treatment of macrophages with p21 antisense oligonucleotides or small interfering RNAs reduced HIV-1 infection. In addition, the synthetic triterpenoid and peroxisome proliferator-activated receptor γ ligand, 2-cyano-3,12-dioxooleana-1,9-dien-28-oic acid (CDDO), which is known to influence p21 expression, suppressed viral replication. These data implicate p21 as a pivotal macrophage facilitator of the viral life cycle. Moreover, regulators of p21, such as CDDO, may provide an interventional approach to modulate HIV-1 replication.

scholarly journals The CDK Inhibitor p21Cip1/WAF1 Is Induced by FcγR Activation and Restricts the Replication of Human Immunodeficiency Virus Type 1 and Related Primate Lentiviruses in Human Macrophages

2009 ◽  
Vol 83 (23) ◽  
pp. 12253-12265 ◽  
Author(s):  
Anna Bergamaschi ◽  
Annie David ◽  
Erwann Le Rouzic ◽  
Sébastien Nisole ◽  
Françoise Barré-Sinoussi ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Viral Replication ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Kinase Inhibitor ◽  
Specific Interaction ◽  
Immunodeficiency Virus ◽  
Hiv 1 ◽  
Reverse Transcript

ABSTRACT Macrophages are major targets of human immunodeficiency virus type 1 (HIV-1). We have previously shown that aggregation of activating immunoglobulin G Fc receptors (FcγR) by immune complexes inhibits reverse transcript accumulation and integration of HIV-1 and related lentiviruses in monocyte-derived macrophages. Here, we show that FcγR-mediated restriction of HIV-1 is not due to enhanced degradation of incoming viral proteins or cDNA and is associated to the induction of the cyclin-dependent kinase inhibitor p21Cip1/WAF1 (p21). Small interfering RNA-mediated p21 knockdown rescued viral replication in FcγR-activated macrophages and enhanced HIV-1 infection in unstimulated macrophages by increasing reverse transcript and integrated DNA levels. p21 induction by other stimuli, such as phorbol myristate acetate and the histone deacetylase inhibitor MS-275, was also associated with preintegrative blocks of HIV-1 replication in macrophages. Binding of p21 to reverse transcription/preintegration complex-associated HIV-1 proteins was not detected in yeast two-hybrid, pulldown, or coimmunoprecipitation assays, suggesting that p21 may affect viral replication independently of a specific interaction with an HIV-1 component. Consistently, p21 silencing rescued viral replication from the FcγR-mediated restriction also in simian immunodeficiency virus SIVmac- and HIV-2-infected macrophages. Our results point to a role of p21 as an inhibitory factor of lentiviral infection in macrophages and to its implication in FcγR-mediated restriction.

scholarly journals Human Immunodeficiency Virus Type 1 Escapes from RNA Interference-Mediated Inhibition

2004 ◽  
Vol 78 (5) ◽  
pp. 2601-2605 ◽  
Author(s):  
Atze T. Das ◽  
Thijn R. Brummelkamp ◽  
Ellen M. Westerhout ◽  
Monique Vink ◽  
Mandy Madiredjo ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Rna Interference ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Antiviral Drug ◽  
Target Sequence ◽  
Small Interfering Rnas ◽  
Immunodeficiency Virus ◽  
Hiv 1

ABSTRACT Short-term assays have suggested that RNA interference (RNAi) may be a powerful new method for intracellular immunization against human immunodeficiency virus type 1 (HIV-1) infection. However, RNAi has not yet been shown to protect cells against HIV-1 in long-term virus replication assays. We stably introduced vectors expressing small interfering RNAs (siRNAs) directed against the HIV-1 genome into human T cells by retroviral transduction. We report here that an siRNA directed against the viral Nef gene (siRNA-Nef) confers resistance to HIV-1 replication. This block in replication is not absolute, and HIV-1 escape variants that were no longer inhibited by siRNA-Nef appeared after several weeks of culture. These RNAi-resistant viruses contained nucleotide substitutions or deletions in the Nef gene that modified or deleted the siRNA-Nef target sequence. These results demonstrate that efficient inhibition of HIV-1 replication through RNAi is possible in stably transduced cells. Therefore, RNAi could become a realistic gene therapy approach with which to overcome the devastating effect of HIV-1 on the immune system. However, as is known for antiviral drug therapy against HIV-1, antiviral approaches involving RNAi should be used in a combined fashion to prevent the emergence of resistant viruses.

scholarly journals Impact of Natural Polymorphism within the gp41 Cytoplasmic Tail of Human Immunodeficiency Virus Type 1 on the Intracellular Distribution of Envelope Glycoproteins and Viral Assembly

2006 ◽  
Vol 81 (1) ◽  
pp. 125-140 ◽  
Author(s):  
Marie Lambelé ◽  
Béatrice Labrosse ◽  
Emmanuelle Roch ◽  
Alain Moreau ◽  
Bernard Verrier ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Viral Replication ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Intracellular Trafficking ◽  
Intracellular Distribution ◽  
Immunodeficiency Virus ◽  
Natural Polymorphism ◽  
Hiv 1

ABSTRACT The motifs involved in the various functions of the human immunodeficiency virus type 1 (HIV-1) gp41 cytoplasmic tail (CT), particularly those related to the intracellular trafficking and assembly of envelope glycoproteins (Env) onto core particles, have generally been assessed with a restricted panel of T-cell laboratory-adapted virus strains. Here, we investigated gp41 CT sequences derived from individuals infected with HIV-1 viruses of various subtypes. We identified four patients harboring HIV variants with a natural polymorphism in the membrane-proximal tyrosine-based signal Y712SPL or the Y802W803 diaromatic motif, which are two major determinants of Env intracellular trafficking. Confocal microscopy showed that the intracellular distribution of Env with a mutation in the tyrosine or diaromatic motif differed from that of Env with no mutation in these motifs. Surprisingly, the gp41 CTs of the primary viruses also had differential effects on the intracellular distribution of Env, independently of mutations in the tyrosine or diaromatic motifs, suggesting the involvement of additional determinants. Furthermore, analyses of virus replication kinetics indicated that the effects of mutations in the tyrosine or diaromatic motifs on viral replication depended on the gp41 CT context. These effects were at least partly due to differences in the efficiency of Env incorporation into virions. Thus, polymorphisms in primary HIV-1 gp41 CTs at the quasispecies or subtype level can influence the intracellular distribution of Env, its incorporation into virions, and viral replication capacity.

scholarly journals Human Immunodeficiency Virus Type 1 Pathogenesis in SCID-hu Mice Correlates with Syncytium-Inducing Phenotype and Viral Replication

2000 ◽  
Vol 74 (7) ◽  
pp. 3196-3204 ◽  
Author(s):  
David Camerini ◽  
Hua-Poo Su ◽  
Graciela Gamez-Torre ◽  
Michael L. Johnson ◽  
Jerome A. Zack ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Viral Replication ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Threshold Level ◽  
Cytopathic Effects ◽  
Immunodeficiency Virus ◽  
Hiv 1 ◽  
Entry Efficiency

ABSTRACT Human immunodeficiency virus type 1 (HIV-1) patient isolates and molecular clones were used to analyze the determinants responsible for human CD4+ thymocyte depletion in SCID-hu mice. Non-syncytium-inducing, R5 or R3R5 HIV-1 isolates from asymptomatic infected people showed little or no human CD4+ thymocyte depletion in SCID-hu mice, while syncytium-inducing (SI), R5X4 or R3R5X4 HIV-1 isolates from the same individuals, isolated just prior to the onset of AIDS, rapidly and efficiently eliminated CD4-bearing human thymocytes. We have mapped the ability of one SI HIV-1 isolate to eliminate CD4+ human cells in SCID-hu mice to a region of the env gene including the three most amino-terminal variable regions (V1 to V3). We find that for all of the HIV-1 isolates that we studied, a nonlinear relationship exists between viral replication and the depletion of CD4+ cells. This relationship can best be described mathematically with a Hill-type plot indicating that a threshold level of viral replication, at which cytopathic effects begin to be seen, exists for HIV-1 infection of thymus/liver grafts in SCID-hu mice. This threshold level is 1 copy of viral DNA for every 11 cells (95% confidence interval = 1 copy of HIV-1 per 67 cells to 1 copy per 4 cells). Furthermore, while SI viruses more frequently achieve this level of replication, replication above this threshold level correlates best with cytopathic effects in this model system. We used GHOST cells to map the coreceptor specificity and relative entry efficiency of these early- and late-stage patient isolates of HIV-1. Our studies show that coreceptor specificity and entry efficiency are critical determinants of HIV-1 pathogenesis in vivo.

scholarly journals Effective Downregulation of HLA-A*2 and HLA-B*57 by Primary Human Immunodeficiency Virus Type 1 Isolates Cultured from Elite Suppressors

2009 ◽  
Vol 83 (13) ◽  
pp. 6941-6946 ◽  
Author(s):  
Eric Nou ◽  
Yan Zhou ◽  
Damaris D. Nou ◽  
Joel N. Blankson
Keyword(s):  
Human Immunodeficiency Virus ◽  
Viral Replication ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Hla Class I ◽  
Elite Controllers ◽  
Hla Class I Molecules ◽  
Immunodeficiency Virus ◽  
Hiv 1

ABSTRACT Elite controllers or suppressors (ES) are human immunodeficiency virus type 1 (HIV-1)-infected patients who control viral replication to <50 copies/ml without antiretroviral therapy. Downregulation of HLA class I molecules is an important mechanism used by HIV-1 to evade the immune system. In this study, we showed that primary isolates from ES are as effective as isolates obtained from patients with progressive HIV-1 disease at downregulating HLA-A*2 and HLA-B*57 molecules on primary CD4+ T cells. Thus, a diminished ability of viral isolates from ES to evade HIV-specific immune responses probably does not contribute to the control of viral replication in these patients.

scholarly journals Human Immunodeficiency Virus Type 1 cDNA Integration: New Aromatic Hydroxylated Inhibitors and Studies of the Inhibition Mechanism

1998 ◽  
Vol 42 (9) ◽  
pp. 2245-2253 ◽  
Author(s):  
C. M. Farnet ◽  
B. Wang ◽  
M. Hansen ◽  
J. R. Lipford ◽  
L. Zalkow ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Viral Replication ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Catalytic Domain ◽  
Restriction Enzymes ◽  
Inhibition Mechanism ◽  
Immunodeficiency Virus ◽  
Hiv 1

ABSTRACT Integration of the human immunodeficiency virus type 1 (HIV-1) cDNA is a required step for viral replication. Integrase, the virus-encoded enzyme important for integration, has not yet been exploited as a target for clinically useful inhibitors. Here we report on the identification of new polyhydroxylated aromatic inhibitors of integrase including ellagic acid, purpurogallin, 4,8,12-trioxatricornan, and hypericin, the last of which is known to inhibit viral replication. These compounds and others were characterized in assays with subviral preintegration complexes (PICs) isolated from HIV-1-infected cells. Hypericin was found to inhibit PIC assays, while the other compounds tested were inactive. Counterscreening of these and other integrase inhibitors against additional DNA-modifying enzymes revealed that none of the polyhydroxylated aromatic compounds are active against enzymes that do not require metals (methylases, a pox virus topoisomerase). However, all were cross-reactive with metal-requiring enzymes (restriction enzymes, a reverse transcriptase), implicating metal atoms in the inhibitory mechanism. In mechanistic studies, we localized binding of some inhibitors to the catalytic domain of integrase by assaying competition of binding by labeled nucleotides. These findings help elucidate the mechanism of action of the polyhydroxylated aromatic inhibitors and provide practical guidance for further inhibitor development.

scholarly journals N-Myristoyltransferase isozymes exhibit differential specificity for human immunodeficiency virus type 1 Gag and Nef

2008 ◽  
Vol 89 (1) ◽  
pp. 288-296 ◽  
Author(s):  
Kelly E. Seaton ◽  
Charles D. Smith
Keyword(s):  
Human Immunodeficiency Virus ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Catalytic Efficiency ◽  
Selective Inhibition ◽  
Competitive Inhibitor ◽  
Small Interfering Rnas ◽  
Immunodeficiency Virus ◽  
Hiv 1

Myristoylation of the human immunodeficiency virus type 1 (HIV-1) proteins Gag and Nef by N-myristoyltransferase (NMT) is a key process in retroviral replication and virulence, yet remains incompletely characterized. Therefore, the roles of the two isozymes, NMT1 and NMT2, in myristoylating Gag and Nef were examined using biochemical and molecular approaches. Fluorescently labelled peptides corresponding to the N terminus of HIV-1 Gag or Nef were myristoylated by recombinant human NMT1 and NMT2. Kinetic analyses indicated that NMT1 and NMT2 had 30- and 130-fold lower K m values for Nef than Gag, respectively. Values for K cat indicated that, once Gag or Nef binds to the enzyme, myristoylation by NMT1 and NMT2 proceeds at comparable rates. Furthermore, the catalytic efficiencies for the processing of Gag by NMT1 and NMT2 were equivalent. In contrast, NMT2 had approximately 5-fold higher catalytic efficiency for the myristoylation of Nef than NMT1. Competition experiments confirmed that the Nef peptide acts as a competitive inhibitor for the myristoylation of Gag. Experiments using full-length recombinant Nef protein also indicated a lower K m for Nef myristoylation by NMT2 than NMT1. Small interfering RNAs were used to selectively deplete NMT1 and/or NMT2 from HEK293T cells expressing a recombinant Nef–sgGFP fusion protein. Depletion of NMT1 had minimal effect on the intracellular distribution of Nef–sgGFP, whereas depletion of NMT2 altered distribution to a diffuse, widespread pattern, mimicking that of a myristoylation-deficient mutant of Nef–sgGFP. Together, these findings indicate that Nef is preferentially myristoylated by NMT2, suggesting that selective inhibition of NMT2 may provide a novel means of blocking HIV virulence.

scholarly journals Nef Binds p6* in GagPol during Replication of Human Immunodeficiency Virus Type 1

2004 ◽  
Vol 78 (10) ◽  
pp. 5311-5323 ◽  
Author(s):  
Luciana J. Costa ◽  
Yong-Hui Zheng ◽  
Jerica Sabotic ◽  
Johnson Mak ◽  
Oliver T. Fackler ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Viral Replication ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Virion Production ◽  
Immunodeficiency Virus ◽  
Intermediate Compartment ◽  
Hiv 1

ABSTRACT The atypical Nef protein (NefF12) from human immunodeficiency virus type 1 strain F12 (HIV-1F12) interferes with virion production and infectivity via a mysterious mechanism. The correlation of these effects with the unusual perinuclear subcellular localization of NefF12 suggested that the wild-type Nef protein could bind to assembly intermediates in late stages of viral replication. To test this hypothesis, Nef from HIV-1NL4-3 was fused to an endoplasmic reticulum (ER) retention signal (NefKKXX). This mutant NefKKXX protein recapitulated fully the effects of NefF12 on Gag processing and virion production, either alone or as a CD8 fusion protein. Importantly, the mutant NefKKXX protein also localized to the intermediate compartment, between the ER and the trans-Golgi network. Furthermore, Nef bound the GagPol polyprotein in vitro and in vivo. This binding mapped to the C-terminal flexible loop in Nef and the transframe p6* protein in GagPol. The significance of this interaction was demonstrated by a genetic assay in which the release of a mutant HIV-1 provirus lacking the PTAP motif in the late domain that no longer binds Tsg101 was rescued by a Nef.Tsg101 chimera. Importantly, this rescue as well as incorporation of Nef into HIV-1 virions correlated with the ability of Nef to interact with GagPol. Our data demonstrate that the retention of Nef in the intermediate compartment interferes with viral replication and suggest a new role for Nef in the production of HIV-1.

scholarly journals High Rates of Human Immunodeficiency Virus Type 1 Recombination: Near-Random Segregation of Markers One Kilobase Apart in One Round of Viral Replication

2003 ◽  
Vol 77 (20) ◽  
pp. 11193-11200 ◽  
Author(s):  
Terence Rhodes ◽  
Heather Wargo ◽  
Wei-Shau Hu
Keyword(s):  
Human Immunodeficiency Virus ◽  
Viral Replication ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Leukemia Virus ◽  
Viral Population ◽  
Recombination Rates ◽  
Immunodeficiency Virus ◽  
Hiv 1

ABSTRACT One of the genetic consequences of packaging two copies of full-length viral RNA into a single retroviral virion is frequent recombination during reverse transcription. Many of the currently circulating strains of human immunodeficiency virus type 1 (HIV-1) are recombinants. Recombination can also accelerate the generation of multidrug-resistant HIV-1 and therefore presents challenges to effective antiviral therapy. In this study, we determined that HIV-1 recombination rates with markers 1.0, 1.3, and 1.9 kb apart were 42.4, 50.4, and 47.4% in one round of viral replication. Because the predicted recombination rate of two unlinked markers is 50%, we conclude that markers 1 kb apart segregated in a manner similar to that for two unlinked markers in one round of retroviral replication. These recombination rates are exceedingly high even among retroviruses. Recombination rates of markers separated by 1 kb are 4 and 4.7% in one round of spleen necrosis virus and murine leukemia virus replication, respectively. Therefore, HIV-1 recombination can be 10-fold higher than that of other retroviruses. Recombination can be observed only in the proviruses derived from heterozygous virions that contain two genotypically different RNAs. The high rates of HIV-1 recombination observed in our studies also indicate that heterozygous virions are formed efficiently during HIV-1 replication and most HIV-1 virions are capable of undergoing recombination. Our results demonstrate that recombination is an effective mechanism to break the genetic linkage between neighboring sequences, thereby reassorting the HIV-1 genome and increasing the diversity in the viral population.

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