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Sample records for hiv-1 ltr transcription

  1. Nuclear Matrix protein SMAR1 represses HIV-1 LTR mediated transcription through chromatin remodeling

    International Nuclear Information System (INIS)

    Sreenath, Kadreppa; Pavithra, Lakshminarasimhan; Singh, Sandeep; Sinha, Surajit; Dash, Prasanta K.; Siddappa, Nagadenahalli B.; Ranga, Udaykumar; Mitra, Debashis; Chattopadhyay, Samit

    2010-01-01

    Nuclear Matrix and MARs have been implicated in the transcriptional regulation of host as well as viral genes but their precise role in HIV-1 transcription remains unclear. Here, we show that > 98% of HIV sequences contain consensus MAR element in their promoter. We show that SMAR1 binds to the LTR MAR and reinforces transcriptional silencing by tethering the LTR MAR to nuclear matrix. SMAR1 associated HDAC1-mSin3 corepressor complex is dislodged from the LTR upon cellular activation by PMA/TNFα leading to an increase in the acetylation and a reduction in the trimethylation of histones, associated with the recruitment of RNA Polymerase II on the LTR. Overexpression of SMAR1 lead to reduction in LTR mediated transcription, both in a Tat dependent and independent manner, resulting in a decreased virion production. These results demonstrate the role of SMAR1 in regulating viral transcription by alternative compartmentalization of LTR between the nuclear matrix and chromatin.

  2. Intracellular high mobility group B1 protein (HMGB1) represses HIV-1 LTR-directed transcription in a promoter- and cell-specific manner

    International Nuclear Information System (INIS)

    Naghavi, Mojgan H.; Nowak, Piotr; Andersson, Jan; Soennerborg, Anders; Yang Huan; Tracey, Kevin J.; Vahlne, Anders

    2003-01-01

    We investigated whether the high mobility group B 1 (HMGB1), an abundant nuclear protein in all mammalian cells, affects HIV-1 transcription. Intracellular expression of human HMGB1 repressed HIV-1 gene expression in epithelial cells. This inhibitory effect of HMGB1 was caused by repression of long terminal repeat (LTR)-mediated transcription. Other viral promoters/enhancers, including simian virus 40 or cytomegalovirus, were not inhibited by HMGB1. In addition, HMGB1 inhibition of HIV-1 subtype C expression was dependent on the number of NFκB sites in the LTR region. The inhibitory effect of HMGB1 on viral gene expression observed in HeLa cells was confirmed by an upregulation of viral replication in the presence of antisense HMGB1 in monocytic cells. In contrast to what was found in HeLa cells and monocytic cells, endogenous HMGB1 expression did not affect HIV-1 replication in unstimulated Jurkat cells. Thus, intracellular HMGB1 affects HIV-1 LTR-directed transcription in a promoter- and cell-specific manner

  3. Alterations in HIV-1 LTR promoter activity during AIDS progression

    International Nuclear Information System (INIS)

    Hiebenthal-Millow, Kirsten; Greenough, Thomas C.; Bretttler, Doreen B.; Schindler, Michael; Wildum, Steffen; Sullivan, John L.; Kirchhoff, Frank

    2003-01-01

    HIV-1 variants evolving in AIDS patients frequently show increased replicative capacity compared to those present during early asymptomatic infection. It is known that late stage HIV-1 variants often show an expanded coreceptor tropism and altered Nef function. In the present study we investigated whether enhanced HIV-1 LTR promoter activity might also evolve during disease progression. Our results demonstrate increased LTR promoter activity after AIDS progression in 3 of 12 HIV-1-infected individuals studied. Further analysis revealed that multiple alterations in the U3 core-enhancer and in the transactivation-response (TAR) region seem to be responsible for the enhanced functional activity. Our findings show that in a subset of HIV-1-infected individuals enhanced LTR transcription contributes to the increased replicative potential of late stage virus isolates and might accelerate disease progression

  4. Cocaine promotes both initiation and elongation phase of HIV-1 transcription by activating NF-κB and MSK1 and inducing selective epigenetic modifications at HIV-1 LTR

    International Nuclear Information System (INIS)

    Sahu, Geetaram; Farley, Kalamo; El-Hage, Nazira; Aiamkitsumrit, Benjamas; Fassnacht, Ryan; Kashanchi, Fatah; Ochem, Alex; Simon, Gary L.; Karn, Jonathan; Hauser, Kurt F.; Tyagi, Mudit

    2015-01-01

    Cocaine accelerates human immunodeficiency virus (HIV-1) replication by altering specific cell-signaling and epigenetic pathways. We have elucidated the underlying molecular mechanisms through which cocaine exerts its effect in myeloid cells, a major target of HIV-1 in central nervous system (CNS). We demonstrate that cocaine treatment promotes HIV-1 gene expression by activating both nuclear factor-kappa B (NF-ĸB) and mitogen- and stress-activated kinase 1 (MSK1). MSK1 subsequently catalyzes the phosphorylation of histone H3 at serine 10, and p65 subunit of NF-ĸB at 276th serine residue. These modifications enhance the interaction of NF-ĸB with P300 and promote the recruitment of the positive transcription elongation factor b (P-TEFb) to the HIV-1 LTR, supporting the development of an open/relaxed chromatin configuration, and facilitating the initiation and elongation phases of HIV-1 transcription. Results are also confirmed in primary monocyte derived macrophages (MDM). Overall, our study provides detailed insights into cocaine-driven HIV-1 transcription and replication. - Highlights: • Cocaine induces the initiation phase of HIV transcription by activating NF-ĸB. • Cocaine induced NF-ĸB phosphorylation promotes its interaction with P300. • Cocaine enhances the elongation phase of HIV transcription by stimulating MSK1. • Cocaine activated MSK1 catalyzes the phosphorylation of histone H3 at its Ser10. • Cocaine induced H3S10 phosphorylation facilitates the recruitment of P-TEFb at LTR

  5. Cocaine promotes both initiation and elongation phase of HIV-1 transcription by activating NF-κB and MSK1 and inducing selective epigenetic modifications at HIV-1 LTR

    Energy Technology Data Exchange (ETDEWEB)

    Sahu, Geetaram; Farley, Kalamo [Division of Infectious Diseases, Department of Medicine, George Washington University, Washington, DC (United States); El-Hage, Nazira [Virginia Commonwealth University, Richmond, VA (United States); Aiamkitsumrit, Benjamas; Fassnacht, Ryan [Division of Infectious Diseases, Department of Medicine, George Washington University, Washington, DC (United States); Kashanchi, Fatah [George Mason University, Manassas, VA (United States); Ochem, Alex [ICGEB, Wernher and Beit Building, Anzio Road, Observatory, 7925 Cape Town (South Africa); Simon, Gary L. [Division of Infectious Diseases, Department of Medicine, George Washington University, Washington, DC (United States); Karn, Jonathan [Case Western Reserve University, Cleveland, OH (United States); Hauser, Kurt F. [Virginia Commonwealth University, Richmond, VA (United States); Tyagi, Mudit, E-mail: tmudit@email.gwu.edu [Division of Infectious Diseases, Department of Medicine, George Washington University, Washington, DC (United States); Department of Microbiology, Immunology and Tropical Medicine, George Washington University, Washington, DC 20037 (United States)

    2015-09-15

    Cocaine accelerates human immunodeficiency virus (HIV-1) replication by altering specific cell-signaling and epigenetic pathways. We have elucidated the underlying molecular mechanisms through which cocaine exerts its effect in myeloid cells, a major target of HIV-1 in central nervous system (CNS). We demonstrate that cocaine treatment promotes HIV-1 gene expression by activating both nuclear factor-kappa B (NF-ĸB) and mitogen- and stress-activated kinase 1 (MSK1). MSK1 subsequently catalyzes the phosphorylation of histone H3 at serine 10, and p65 subunit of NF-ĸB at 276th serine residue. These modifications enhance the interaction of NF-ĸB with P300 and promote the recruitment of the positive transcription elongation factor b (P-TEFb) to the HIV-1 LTR, supporting the development of an open/relaxed chromatin configuration, and facilitating the initiation and elongation phases of HIV-1 transcription. Results are also confirmed in primary monocyte derived macrophages (MDM). Overall, our study provides detailed insights into cocaine-driven HIV-1 transcription and replication. - Highlights: • Cocaine induces the initiation phase of HIV transcription by activating NF-ĸB. • Cocaine induced NF-ĸB phosphorylation promotes its interaction with P300. • Cocaine enhances the elongation phase of HIV transcription by stimulating MSK1. • Cocaine activated MSK1 catalyzes the phosphorylation of histone H3 at its Ser10. • Cocaine induced H3S10 phosphorylation facilitates the recruitment of P-TEFb at LTR.

  6. Cocaine promotes both initiation and elongation phase of HIV-1 transcription by activating NF-κB and MSK1 and inducing selective epigenetic modifications at HIV-1 LTR

    Science.gov (United States)

    Sahu, Geetaram; Farley, Kalamo; El-Hage, Nazira; Aiamkitsumrit, Benjamas; Fassnacht, Ryan; Kashanchi, Fatah; Ochem, Alex; Simon, Gary L.; Karn, Jonathan; Hauser, Kurt F.; Tyagi, Mudit

    2015-01-01

    Cocaine accelerates human immunodeficiency virus (HIV-1) replication by altering specific cell-signaling and epigenetic pathways. We have elucidated the underlying molecular mechanisms through which cocaine exerts its effect in myeloid cells, a major target of HIV-1 in central nervous system (CNS). We demonstrate that cocaine treatment promotes HIV-1 gene expression by activating both nuclear factor-kappa B (NF-κB) and mitogen- and stress-activated kinase 1 (MSK1). MSK1 subsequently catalyzes the phosphorylation of histone H3 at serine 10, and p65 subunit of NF-κB at 276th serine residue. These modifications enhance the interaction of NF-κB with P300 and promote the recruitment of the positive transcription elongation factor b (P-TEFb) to the HIV-1 LTR, supporting the development of an open/relaxed chromatin configuration, and facilitating the initiation and elongation phases of HIV-1 transcription. Results are also confirmed in primary monocyte derived macrophages (MDM). Overall, our study provides detailed insights into cocaine-driven HIV-1 transcription and replication. PMID:25980739

  7. Cellular specificity of HIV-1 replication can be controlled by LTR sequences

    International Nuclear Information System (INIS)

    Reed-Inderbitzin, Edward; Maury, Wendy

    2003-01-01

    Two well-established determinants of retroviral tropism are envelope sequences that regulate entry and LTR sequences that can regulate viral expression in a cell-specific manner. Studies with human immunodeficiency virus-1 (HIV-1) have demonstrated that tropism of this virus maps primarily to variable envelope sequences. Studies have demonstrated that T cell and macrophage-specific transcription factor binding motifs exist in the upstream region of the LTR U3; however, the ability of the core enhancer/promoter proximal elements (two NF-κB and three Sp1 sites) to function well in macrophages and T cells have led many to conclude that HIV LTR sequences are not primary determinants of HIV tropism. To determine if cellular specificity could be imparted to HIV by the core enhancer elements, the enhancer/promoter proximal region of the HIV LTR was substituted with motifs that control gene expression in a myeloid-specific manner. The enhancer region from equine infectious anemia virus (EIAV) when substituted for the HIV enhancer/promoter proximal region was found to drive expression in a macrophage-specific manner and was responsive to HIV Tat. The addition of a 5' methylation-dependent binding site (MDBP) and a promoter proximal Sp1 motif increased expression without altering cellular specificity. Spacing between the promoter proximal region and the TATA box was also found to influence LTR activity. Infectivity studies using chimeric LTRs within the context of a dual-tropic infectious molecular clone established that these LTRs directed HIV replication and production of infectious virions in macrophages but not primary T cells or T cell lines. This investigation demonstrates that cellular specificity can be imparted onto HIV-1 replication at the level of viral transcription and not entry

  8. Divergent transcriptional regulation of HIV-1: Structural and Functional Analysis of the LTR Promoter Activity of Group M, N and O

    OpenAIRE

    Somogyi, Sybille

    2010-01-01

    HIV-1 is classified into three distinct genetic groups: M (major), N (non-M; non-O) and O (outlier). Within the group M actually nine subtypes and an increasing number of circulating recombinant forms have been identified. The global spread of subtypes and CRF (circulating recombinant form) differs with respect to geographic regions and the routes of transmission. Besides sociodemographic factors, biological properties of the host and the virus may influence the different prevalences of the s...

  9. The dual action of poly(ADP-ribose polymerase -1 (PARP-1 inhibition in HIV-1 infection: HIV-1 LTR inhibition and diminution in Rho GTPase activity

    Directory of Open Access Journals (Sweden)

    Slava eRom

    2015-08-01

    Full Text Available The transcription of HIV-1 (HIV is regulated by complex mechanisms involving various cellular factors and virus-encoded transactivators. Poly(ADP-ribose polymerase 1 (PARP-1 inhibition has emerged recently as a potent anti-inflammatory tool, since PARP-1 is involved in the regulation of some genes through its interaction with various transcription factors. We propose a novel approach to diminish HIV replication via PARP-1 inhibition using human primary monocyte-derived macrophages (MDM as an in vitro model system. PARP-1 inhibitors were able to reduce HIV replication in MDM by 60-80% after 7 days infection. Long Terminal Repeat (LTR acts as a switch in virus replication and can be triggered by several agents such as: Tat, tumor necrosis factor α (TNFα, and phorbol 12-myristate 13-acetate (PMA. Overexpression of Tat in MDM transfected with an LTR reporter plasmid led to a 4.2-fold increase in LTR activation; PARP inhibition resulted in 70% reduction of LTR activity. LTR activity, which increased 3-fold after PMA or TNFα treatment, was reduced by PARP inhibition (by 85-95%. MDM treated with PARP inhibitors showed 90% reduction in NFκB activity (known to mediate PMA- and TNFα-induced HIV LTR activation. Cytoskeleton rearrangements are important in effective HIV-1 infection. PARP inactivation reduced actin cytoskeleton rearrangements by affecting Rho GTPase machinery. These findings suggest that HIV replication in MDM could be suppressed by PARP inhibition via NFκB suppression, diminution of LTR activation and its effects on the cytoskeleton. PARP appears to be essential for HIV replication and its inhibition may provide a potent approach to treatment of HIV infection.

  10. Rapid turnover of 2-LTR HIV-1 DNA during early stage of highly active antiretroviral therapy.

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    Weijun Zhu

    Full Text Available BACKGROUND: Despite prolonged treatment with highly active antiretroviral therapy (HAART, the infectious HIV-1 continues to replicate and resides latently in the resting memory CD4+ T lymphocytes, which blocks the eradication of HIV-1. The viral persistence of HIV-1 is mainly caused by its proviral DNA being either linear nonintegrated, circular nonintegrated, or integrated. Previous reports have largely focused on the dynamics of HIV-1 DNA from the samples collected with relatively long time intervals during the process of disease and HAART treatment, which may have missed the intricate changes during the intervals in early treatment. METHODOLOGY/PRINCIPAL FINDINGS: In this study, we investigated the dynamics of HIV-1 DNA in patients during the early phase of HARRT treatment. Using optimized real time PCR, we observed significant changes in 2-LTR during the first 12-week of treatment, while total and integrated HIV-1 DNA remained stable. The doubling time and half-life of 2-LTR were not correlated with the baseline and the rate of changes in plasma viral load and various CD4+ T-cell populations. Longitudinal analyses on 2-LTR sequences and plasma lipopolysaccharide (LPS levels did not reveal any significant changes in the same treatment period. CONCLUSIONS/SIGNIFICANCE: Our study revealed the rapid changes in 2-LTR concentration in a relatively large number of patients during the early HAART treatment. The rapid changes indicate the rapid infusion and clearance of cells bearing 2-LTR in the peripheral blood. Those changes are not expected to be caused by the blocking of viral integration, as our study did not include the integrase inhibitor raltegravir. Our study helps better understand the dynamics of HIV-DNA and its potential role as a biomarker for the diseases and for the treatment efficacy of HAART.

  11. Effects of integration and replication on transcription of the HIV-1 long terminal repeat

    NARCIS (Netherlands)

    Jeang, K. T.; Berkhout, B.; Dropulic, B.

    1993-01-01

    The activity of a promoter is influenced by chromosomal and cell cycle/replication context. We analyzed the influences of integration and replication on transcription of the human immunodeficiency virus (HIV)-1 long terminal repeat (LTR). We found that one requirement for Tat trans-activated

  12. Interaction of the phospholipid scramblase 1 with HIV-1 Tat results in the repression of Tat-dependent transcription

    International Nuclear Information System (INIS)

    Kusano, Shuichi; Eizuru, Yoshito

    2013-01-01

    Highlights: •PLSCR1 specifically interacted with HIV-1 Tat in vitro and in vivo. •PLSCR1 repressed Tat-dependent transactivation of the HIV-1 LTR. •Suppression of PLSCR1 expression enhanced the levels of HIV-1 transcripts. •PLSCR1 reduced the nuclear localization of Tat. -- Abstract: Human phospholipid scramblase 1 (PLSCR1) is an interferon (IFN)-stimulated gene and possesses an IFN-mediated antiviral function. We show here that PLSCR1 directly interacts with human immunodeficiency virus type-1 (HIV-1) Tat. This interaction occurs both in vitro and in vivo through amino acids 160–250 of PLSCR1. Overexpression of PLSCR1 efficiently represses the Tat-dependent transactivation of the HIV-1 long terminal repeat (LTR) and reduces the nuclear translocation of Tat. In addition, shRNA-mediated suppression of endogenous PLSCR1 expression enhances the levels of gag mRNA in an HIV-1-infected T-cell line. These findings indicate that PLSCR1 negatively regulates the Tat-dependent transactivation of the HIV-1 LTR during HIV-1 infection

  13. Receptor-targeted aptamer-siRNA conjugate-directed transcriptional regulation of HIV-1

    Science.gov (United States)

    Zhou, Jiehua; Lazar, Daniel; Li, Haitang; Xia, Xin; Satheesan, Sangeetha; Charlins, Paige; O'Mealy, Denis; Akkina, Ramesh; Saayman, Sheena; Weinberg, Marc S.; Rossi, John J.; Morris, Kevin V.

    2018-01-01

    Gene-based therapies represent a promising therapeutic paradigm for the treatment of HIV-1, as they have the potential to maintain sustained viral inhibition with reduced treatment interventions. Such an option may represent a long-term treatment alternative to highly active antiretroviral therapy. Methods: We previously described a therapeutic approach, referred to as transcriptional gene silencing (TGS), whereby small noncoding RNAs directly inhibit the transcriptional activity of HIV-1 by targeting sites within the viral promoter, specifically the 5' long terminal repeat (LTR). TGS differs from traditional RNA interference (RNAi) in that it is characterized by concomitant silent-state epigenetic marks on histones and DNA. To deliver TGS-inducing RNAs, we developed functional RNA conjugates based on the previously reported dual function of the gp120 (A-1) aptamer conjugated to 27-mer Dicer-substrate anti-HIV-1 siRNA (dsiRNA), LTR-362. Results: We demonstrate here that high levels of processed guide RNAs localize to the nucleus in infected T lymphoblastoid CEM cell line and primary human CD4+ T-cells. Treatment of the aptamer-siRNA conjugates induced TGS with an ~10-fold suppression of viral p24 levels as measured at day 12 post infection. To explore the silencing efficacy of aptamer-siRNA conjugates in vivo, HIV-1-infected humanized NOD/SCID/IL2 rγnull mice (hu-NSG) were treated with the aptamer-siRNA conjugates. Systemic delivery of the A-1-stick-LTR-362 27-mer siRNA conjugates suppressed HIV-1 infection and protected CD4+ T cell levels in viremia hu-NSG mice. Principle conclusions: Collectively these data suggest that the gp120 aptamer-dsiRNA conjugate design is suitable for systemic delivery of small RNAs that can be used to suppress HIV-1. PMID:29556342

  14. HIV-1, HTLV-I and the interleukin-2 receptor: insights into transcriptional control.

    Science.gov (United States)

    Böhnlein, E; Lowenthal, J W; Wano, Y; Franza, B R; Ballard, D W; Greene, W C

    1989-01-01

    In this study, we present direct evidence for the binding of the inducible cellular protein, HIVEN86A, to a 12-bp element present in the IL-2R alpha promoter. This element shares significant sequence similarity with the NF-kappa B binding sites present in the HIV-1 and kappa immunoglobulin enhancers. Transient transfection studies indicate that this kappa B element is both necessary and sufficient to confer tax or mitogen inducibility to a heterologous promoter. As summarized schematically in Fig. 5, the findings suggest that the HIVEN86A protein may play a central role in the activation of cellular genes required for T-cell growth, specifically the IL-2R alpha gene. In addition, the induced HIVEN86A protein also binds to a similar sequence present in the HIV-1 LTR leading to enhanced viral gene expression and ultimately T-cell death. Thus, mitogen activation of the HIV-1 LTR appears to involve the same inducible transcription factor(s) that normally regulates IL-2R alpha gene expression and T-cell growth. These findings further underscore the importance of the state of T-cell activation in the regulation of HIV-1 replication. Our results also demonstrate that HIVEN86A is induced by the tax protein of HTLV-I. Thus, in HTLV-I infected cells, normally the tight control of the transient expression of the IL-2R alpha gene is lost. The constitutive high-level display of IL-2 receptors may play a role in leukemic transformation mediated by HTLV-I (ATL). Apparently by the same mechanism, the tax protein also activates the HIV-1 LTR through the induction of HIVEN86A.(ABSTRACT TRUNCATED AT 250 WORDS)

  15. Overexpression of octamer transcription factors 1 or 2 alone has no effect on HIV-1 transcription in primary human CD4 T cells

    International Nuclear Information System (INIS)

    Zhang Mingce; Genin, Anna; Cron, Randy Q.

    2004-01-01

    We explored the binding of octamer (Oct) transcription factors to the HIV-1 long terminal repeat (LTR) by gel shift assays and showed none of the previously identified four potential Oct binding sites bound Oct-1 or Oct-2. Overexpression of Oct-1 or Oct-2 had no effect on HIV-1 LTR activity in transiently transfected primary human CD4 T cells. Next, primary human CD4 T cells were co-transfected with a green fluorescent protein (GFP)-expression vector and an Oct-1 or Oct-2 expression plasmid. The transfected cells were stimulated for 2 days and then infected with the NL4-3 strain of HIV-1. After 3 days of infection, there were no differences in HIV-1 p24 supernatant levels. Apoptosis of infected or bystander cells overexpressing Oct-1 or Oct-2 compared to control was also unaffected. Our studies demonstrate that Oct-1 and Oct-2 fail to bind to the HIV-1 LTR and have no effect on HIV-1 transcription in primary human CD4 T cells

  16. Iron chelators ICL670 and 311 inhibit HIV-1 transcription

    International Nuclear Information System (INIS)

    Debebe, Zufan; Ammosova, Tatyana; Jerebtsova, Marina; Kurantsin-Mills, Joseph; Niu, Xiaomei; Charles, Sharroya; Richardson, Des R.; Ray, Patricio E.; Gordeuk, Victor R.; Nekhai, Sergei

    2007-01-01

    HIV-1 replication is induced by an excess of iron and iron chelation by desferrioxamine (DFO) inhibits viral replication by reducing proliferation of infected cells. Treatment of cells with DFO and 2-hydroxy-1-naphthylaldehyde isonicotinoyl hydrazone (311) inhibit expression of proteins that regulate cell-cycle progression, including cycle-dependent kinase 2 (CDK2). Our recent studies showed that CDK2 participates in HIV-1 transcription and viral replication suggesting that inhibition of CDK2 by iron chelators might also affect HIV-1 transcription. Here we evaluated the effect of a clinically approved orally effective iron chelator, 4-[3,5-bis-(hydroxyphenyl)-1,2,4-triazol-1-yl]-benzoic acid (ICL670) and 311 on HIV-1 transcription. Both ICL670 and 311 inhibited Tat-induced HIV-1 transcription in CEM-T cells, 293T and HeLa cells. Neither ICL670 nor 311 induced cytotoxicity at concentrations that inhibited HIV-1 transcription. The chelators decreased cellular activity of CDK2 and reduced HIV-1 Tat phosphorylation by CDK2. Neither ICL670A or 311 decreased CDK9 protein level but significantly reduced association of CDK9 with cyclin T1 and reduced phosphorylation of Ser-2 residues of RNA polymerase II C-terminal domain. In conclusion, our findings add to the evidence that iron chelators can inhibit HIV-1 transcription by deregulating CDK2 and CDK9. Further consideration should be given to the development of iron chelators for future anti-retroviral therapeutics

  17. Novel RNA Duplex Locks HIV-1 in a Latent State via Chromatin-mediated Transcriptional Silencing

    Directory of Open Access Journals (Sweden)

    Chantelle Ahlenstiel

    2015-01-01

    Full Text Available Transcriptional gene silencing (TGS of mammalian genes can be induced by short interfering RNA (siRNA targeting promoter regions. We previously reported potent TGS of HIV-1 by siRNA (PromA, which targets tandem NF-κB motifs within the viral 5′LTR. In this study, we screened a siRNA panel with the aim of identifying novel 5′LTR targets, to provide multiplexing potential with enhanced viral silencing and application toward developing alternate therapeutic strategies. Systematic examination identified a novel siRNA target, si143, confirmed to induce TGS as the silencing mechanism. TGS was prolonged with virus suppression >12 days, despite a limited ability to induce post- TGS. Epigenetic changes associated with silencing were suggested by partial reversal by histone deacetylase inhibitors and confirmed by chromatin immunoprecipitation analyses, which showed induction of H3K27me3 and H3K9me3, reduction in H3K9Ac, and recruitment of argonaute-1, all characteristic marks of heterochromatin and TGS. Together, these epigenetic changes mimic those associated with HIV-1 latency. Further, robust resistance to reactivation was observed in the J-Lat 9.2 cell latency model, when transduced with shPromA and/or sh143. These data support si/shRNA-mediated TGS approaches to HIV-1 and provide alternate targets to pursue a functional cure, whereby the viral reservoir is locked in latency following antiretroviral therapy cessation.

  18. Targeted HIV-1 Latency Reversal Using CRISPR/Cas9-Derived Transcriptional Activator Systems.

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    Julia K Bialek

    Full Text Available CRISPR/Cas9 technology is currently considered the most advanced tool for targeted genome engineering. Its sequence-dependent specificity has been explored for locus-directed transcriptional modulation. Such modulation, in particular transcriptional activation, has been proposed as key approach to overcome silencing of dormant HIV provirus in latently infected cellular reservoirs. Currently available agents for provirus activation, so-called latency reversing agents (LRAs, act indirectly through cellular pathways to induce viral transcription. However, their clinical performance remains suboptimal, possibly because reservoirs have diverse cellular identities and/or proviral DNA is intractable to the induced pathways. We have explored two CRISPR/Cas9-derived activator systems as targeted approaches to induce dormant HIV-1 proviral DNA. These systems recruit multiple transcriptional activation domains to the HIV 5' long terminal repeat (LTR, for which we have identified an optimal target region within the LTR U3 sequence. Using this target region, we demonstrate transcriptional activation of proviral genomes via the synergistic activation mediator complex in various in culture model systems for HIV latency. Observed levels of induction are comparable or indeed higher than treatment with established LRAs. Importantly, activation is complete, leading to production of infective viral particles. Our data demonstrate that CRISPR/Cas9-derived technologies can be applied to counteract HIV latency and may therefore represent promising novel approaches in the quest for HIV elimination.

  19. Large-scale transcriptome data reveals transcriptional activity of fission yeast LTR retrotransposons

    DEFF Research Database (Denmark)

    Mourier, Tobias; Willerslev, Eske

    2010-01-01

    of transcriptional activity are observed from both strands of solitary LTR sequences. Transcriptome data collected during meiosis suggests that transcription of solitary LTRs is correlated with the transcription of nearby protein-coding genes. CONCLUSIONS: Presumably, the host organism negatively regulates...

  20. APOBEC3G inhibits elongation of HIV-1 reverse transcripts.

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    Kate N Bishop

    2008-12-01

    Full Text Available APOBEC3G (A3G is a host cytidine deaminase that, in the absence of Vif, restricts HIV-1 replication and reduces the amount of viral DNA that accumulates in cells. Initial studies determined that A3G induces extensive mutation of nascent HIV-1 cDNA during reverse transcription. It has been proposed that this triggers the degradation of the viral DNA, but there is now mounting evidence that this mechanism may not be correct. Here, we use a natural endogenous reverse transcriptase assay to show that, in cell-free virus particles, A3G is able to inhibit HIV-1 cDNA accumulation not only in the absence of hypermutation but also without the apparent need for any target cell factors. We find that although reverse transcription initiates in the presence of A3G, elongation of the cDNA product is impeded. These data support the model that A3G reduces HIV-1 cDNA levels by inhibiting synthesis rather than by inducing degradation.

  1. NF90 una proteína celular que se une a RNAds inhibe la transactivación del LTR del HIV-1 mediada por TAT

    Directory of Open Access Journals (Sweden)

    Silvio Urcuqui Inchima

    2001-04-01

    Full Text Available

    La transactivación del LTR de HIV-1 requiere la interacción de la proteína Tat ( trans-activation transcription con la estructura TAR ( trans-activation response, que se encuentra localizada en el extremo 5’ de todos los transcriptos virales. La interacción de la proteína Tat con TAR forma un precomplejo transcripcional indispensable para la eficiente transcripción del genoma viral.
    La interacción de diferentes factores celulares (cdk9 y ciclina T, entre otros con el precomplejo Tat-TAR, constituye un complejo estable que facilita la fosforilación de la subunidad mayor de la RNA polimerasa II, asegurando una eficiente elongación de los transcriptos virales. No se han descripto proteínas capaces de regular negativamente la función de Tat con resultados nefastos para la replicación viral. También se ha descrito otro tipo de proteínas celulares con capacidad de interactuar con TAR. El ejemplo mejor caracterizado es la proteína-kinasa dependiente de RNA (PKR, se autofosforila y mediante fosforilación del factor eIF2, inhibe la síntesis de proteínas. Con el presente trabajo se
    pretendió aislar y caracterizar otras proteínas celulares capaces de interactuar con la estructura TAR y estudiar su efecto en la función de Tat.

     

  2. Human macrophages support persistent transcription from unintegrated HIV-1 DNA

    International Nuclear Information System (INIS)

    Kelly, Jeremy; Beddall, Margaret H.; Yu Dongyang; Iyer, Subashini R.; Marsh, Jon W.; Wu Yuntao

    2008-01-01

    Retroviruses require integration of their RNA genomes for both stability and productive viral replication. In HIV infection of non-dividing, resting CD4 T cells, where integration is greatly impeded, the reverse transcribed HIV DNA has limited biological activity and a short half-life. In metabolically active and proliferating T cells, unintegrated DNA rapidly diminishes with cell division. HIV also infects the non-dividing but metabolically active macrophage population. In an in vitro examination of HIV infection of macrophages, we find that unintegrated viral DNA not only has an unusual stability, but also maintains biological activity. The unintegrated linear DNA, 1-LTR, and 2-LTR circles are stable for at least 30 days. Additionally, there is persistent viral gene transcription, which is selective and skewed towards viral early genes such as nef and tat with highly diminished rev and vif. One viral early gene product Nef was measurably synthesized. We also find that independent of integration, the HIV infection process in macrophages leads to generation of numerous chemokines

  3. FoxA1 binding to the MMTV LTR modulates chromatin structure and transcription

    International Nuclear Information System (INIS)

    Holmqvist, Per-Henrik; Belikov, Sergey; Zaret, Kenneth S.; Wrange, Oerjan

    2005-01-01

    Novel binding sites for the forkhead transcription factor family member Forkhead box A (FoxA), previously referred to as Hepatocyte Nuclear Factor 3 (HNF3), were found within the mouse mammary tumor virus long terminal repeat (MMTV LTR). The effect of FoxA1 on MMTV LTR chromatin structure, and expression was evaluated in Xenopus laevis oocytes. Mutagenesis of either of the two main FoxA binding sites showed that the distal site, -232/-221, conferred FoxA1-dependent partial inhibition of glucocorticoid receptor (GR) driven MMTV transcription. The proximal FoxA binding segment consisted of two individual FoxA sites at -57/-46 and -45/-34, respectively, that mediated an increased basal MMTV transcription. FoxA1 binding altered the chromatin structure of both the inactive- and the hormone-activated MMTV LTR. Hydroxyl radical foot printing revealed FoxA1-mediated changes in the nucleosome arrangement. Micrococcal nuclease digestion showed the hormone-dependent sub-nucleosome complex, containing ∼120 bp of DNA, to be expanded by FoxA1 binding to the proximal segment into a larger complex containing ∼200 bp. The potential function of the FoxA1-mediated expression of the MMTV provirus for maintenance of expression in different tissues is discussed

  4. Inhibition of both HIV-1 reverse transcription and gene expression by a cyclic peptide that binds the Tat-transactivating response element (TAR RNA.

    Directory of Open Access Journals (Sweden)

    Matthew S Lalonde

    2011-05-01

    Full Text Available The RNA response element TAR plays a critical role in HIV replication by providing a binding site for the recruitment of the viral transactivator protein Tat. Using a structure-guided approach, we have developed a series of conformationally-constrained cyclic peptides that act as structural mimics of the Tat RNA binding region and block Tat-TAR interactions at nanomolar concentrations in vitro. Here we show that these compounds block Tat-dependent transcription in cell-free systems and in cell-based reporter assays. The compounds are also cell permeable, have low toxicity, and inhibit replication of diverse HIV-1 strains, including both CXCR4-tropic and CCR5-tropic primary HIV-1 isolates of the divergent subtypes A, B, C, D and CRF01_AE. In human peripheral blood mononuclear cells, the cyclic peptidomimetic L50 exhibited an IC(50 ∼250 nM. Surprisingly, inhibition of LTR-driven HIV-1 transcription could not account for the full antiviral activity. Timed drug-addition experiments revealed that L-50 has a bi-phasic inhibition curve with the first phase occurring after HIV-1 entry into the host cell and during the initiation of HIV-1 reverse transcription. The second phase coincides with inhibition of HIV-1 transcription. Reconstituted reverse transcription assays confirm that HIV-1 (- strand strong stop DNA synthesis is blocked by L50-TAR RNA interactions in-vitro. These findings are consistent with genetic evidence that TAR plays critical roles both during reverse transcription and during HIV gene expression. Our results suggest that antiviral drugs targeting TAR RNA might be highly effective due to a dual inhibitory mechanism.

  5. Therapeutic doses of irradiation activate viral transcription and induce apoptosis in HIV-1 infected cells

    International Nuclear Information System (INIS)

    Iordanskiy, Sergey; Van Duyne, Rachel; Sampey, Gavin C; Woodson, Caitlin M; Fry, Kelsi; Saifuddin, Mohammed; Guo, Jia; Wu, Yuntao; Romerio, Fabio; Kashanchi, Fatah

    2015-01-01

    The highly active antiretroviral therapy reduces HIV-1 RNA in plasma to undetectable levels. However, the virus continues to persist in the long-lived resting CD4"+ T cells, macrophages and astrocytes which form a viral reservoir in infected individuals. Reactivation of viral transcription is critical since the host immune response in combination with antiretroviral therapy may eradicate the virus. Using the chronically HIV-1 infected T lymphoblastoid and monocytic cell lines, primary quiescent CD4"+ T cells and humanized mice infected with dual-tropic HIV-1 89.6, we examined the effect of various X-ray irradiation (IR) doses (used for HIV-related lymphoma treatment and lower doses) on HIV-1 transcription and viability of infected cells. Treatment of both T cells and monocytes with IR, a well-defined stress signal, led to increase of HIV-1 transcription, as evidenced by the presence of RNA polymerase II and reduction of HDAC1 and methyl transferase SUV39H1 on the HIV-1 promoter. This correlated with the increased GFP signal and elevated level of intracellular HIV-1 RNA in the IR-treated quiescent CD4"+ T cells infected with GFP-encoding HIV-1. Exposition of latently HIV-1infected monocytes treated with PKC agonist bryostatin 1 to IR enhanced transcription activation effect of this latency-reversing agent. Increased HIV-1 replication after IR correlated with higher cell death: the level of phosphorylated Ser46 in p53, responsible for apoptosis induction, was markedly higher in the HIV-1 infected cells following IR treatment. Exposure of HIV-1 infected humanized mice with undetectable viral RNA level to IR resulted in a significant increase of HIV-1 RNA in plasma, lung and brain tissues. Collectively, these data point to the use of low to moderate dose of IR alone or in combination with HIV-1 transcription activators as a potential application for the “Shock and Kill” strategy for latently HIV-1 infected cells. - Highlights: • X-ray irradiation (IR) increases

  6. Therapeutic doses of irradiation activate viral transcription and induce apoptosis in HIV-1 infected cells

    Energy Technology Data Exchange (ETDEWEB)

    Iordanskiy, Sergey [School of Systems Biology, Laboratory of Molecular Virology, George Mason University, Manassas, VA 20110 (United States); Van Duyne, Rachel [School of Systems Biology, Laboratory of Molecular Virology, George Mason University, Manassas, VA 20110 (United States); Center for Cancer Research, National Cancer Institute, National Institutes of Health, Frederick, MD 21702 (United States); Sampey, Gavin C; Woodson, Caitlin M; Fry, Kelsi; Saifuddin, Mohammed; Guo, Jia; Wu, Yuntao [School of Systems Biology, Laboratory of Molecular Virology, George Mason University, Manassas, VA 20110 (United States); Romerio, Fabio [Department of Medicine, University of Maryland School of Medicine, Baltimore, MD 21201 (United States); Kashanchi, Fatah, E-mail: fkashanc@gmu.edu [School of Systems Biology, Laboratory of Molecular Virology, George Mason University, Manassas, VA 20110 (United States)

    2015-11-15

    The highly active antiretroviral therapy reduces HIV-1 RNA in plasma to undetectable levels. However, the virus continues to persist in the long-lived resting CD4{sup +} T cells, macrophages and astrocytes which form a viral reservoir in infected individuals. Reactivation of viral transcription is critical since the host immune response in combination with antiretroviral therapy may eradicate the virus. Using the chronically HIV-1 infected T lymphoblastoid and monocytic cell lines, primary quiescent CD4{sup +} T cells and humanized mice infected with dual-tropic HIV-1 89.6, we examined the effect of various X-ray irradiation (IR) doses (used for HIV-related lymphoma treatment and lower doses) on HIV-1 transcription and viability of infected cells. Treatment of both T cells and monocytes with IR, a well-defined stress signal, led to increase of HIV-1 transcription, as evidenced by the presence of RNA polymerase II and reduction of HDAC1 and methyl transferase SUV39H1 on the HIV-1 promoter. This correlated with the increased GFP signal and elevated level of intracellular HIV-1 RNA in the IR-treated quiescent CD4{sup +} T cells infected with GFP-encoding HIV-1. Exposition of latently HIV-1infected monocytes treated with PKC agonist bryostatin 1 to IR enhanced transcription activation effect of this latency-reversing agent. Increased HIV-1 replication after IR correlated with higher cell death: the level of phosphorylated Ser46 in p53, responsible for apoptosis induction, was markedly higher in the HIV-1 infected cells following IR treatment. Exposure of HIV-1 infected humanized mice with undetectable viral RNA level to IR resulted in a significant increase of HIV-1 RNA in plasma, lung and brain tissues. Collectively, these data point to the use of low to moderate dose of IR alone or in combination with HIV-1 transcription activators as a potential application for the “Shock and Kill” strategy for latently HIV-1 infected cells. - Highlights: • X-ray irradiation

  7. Hyperthermia stimulates HIV-1 replication.

    Directory of Open Access Journals (Sweden)

    Ferdinand Roesch

    Full Text Available HIV-infected individuals may experience fever episodes. Fever is an elevation of the body temperature accompanied by inflammation. It is usually beneficial for the host through enhancement of immunological defenses. In cultures, transient non-physiological heat shock (42-45°C and Heat Shock Proteins (HSPs modulate HIV-1 replication, through poorly defined mechanisms. The effect of physiological hyperthermia (38-40°C on HIV-1 infection has not been extensively investigated. Here, we show that culturing primary CD4+ T lymphocytes and cell lines at a fever-like temperature (39.5°C increased the efficiency of HIV-1 replication by 2 to 7 fold. Hyperthermia did not facilitate viral entry nor reverse transcription, but increased Tat transactivation of the LTR viral promoter. Hyperthermia also boosted HIV-1 reactivation in a model of latently-infected cells. By imaging HIV-1 transcription, we further show that Hsp90 co-localized with actively transcribing provirus, and this phenomenon was enhanced at 39.5°C. The Hsp90 inhibitor 17-AAG abrogated the increase of HIV-1 replication in hyperthermic cells. Altogether, our results indicate that fever may directly stimulate HIV-1 replication, in a process involving Hsp90 and facilitation of Tat-mediated LTR activity.

  8. Transcriptionally active LTR retrotransposons in Eucalyptus genus are differentially expressed and insertionally polymorphic.

    Science.gov (United States)

    Marcon, Helena Sanches; Domingues, Douglas Silva; Silva, Juliana Costa; Borges, Rafael Junqueira; Matioli, Fábio Filippi; Fontes, Marcos Roberto de Mattos; Marino, Celso Luis

    2015-08-14

    In Eucalyptus genus, studies on genome composition and transposable elements (TEs) are particularly scarce. Nearly half of the recently released Eucalyptus grandis genome is composed by retrotransposons and this data provides an important opportunity to understand TE dynamics in Eucalyptus genome and transcriptome. We characterized nine families of transcriptionally active LTR retrotransposons from Copia and Gypsy superfamilies in Eucalyptus grandis genome and we depicted genomic distribution and copy number in two Eucalyptus species. We also evaluated genomic polymorphism and transcriptional profile in three organs of five Eucalyptus species. We observed contrasting genomic and transcriptional behavior in the same family among different species. RLC_egMax_1 was the most prevalent family and RLC_egAngela_1 was the family with the lowest copy number. Most families of both superfamilies have their insertions occurring Eucalyptus species. Using EST analysis and qRT-PCRs, we observed transcriptional activity in several tissues and in all evaluated species. In some families, osmotic stress increases transcript values. Our strategy was successful in isolating transcriptionally active retrotransposons in Eucalyptus, and each family has a particular genomic and transcriptional pattern. Overall, our results show that retrotransposon activity have differentially affected genome and transcriptome among Eucalyptus species.

  9. Reactivation of Latent HIV-1 Expression by Engineered TALE Transcription Factors.

    Science.gov (United States)

    Perdigão, Pedro; Gaj, Thomas; Santa-Marta, Mariana; Barbas, Carlos F; Goncalves, Joao

    2016-01-01

    The presence of replication-competent HIV-1 -which resides mainly in resting CD4+ T cells--is a major hurdle to its eradication. While pharmacological approaches have been useful for inducing the expression of this latent population of virus, they have been unable to purge HIV-1 from all its reservoirs. Additionally, many of these strategies have been associated with adverse effects, underscoring the need for alternative approaches capable of reactivating viral expression. Here we show that engineered transcriptional modulators based on customizable transcription activator-like effector (TALE) proteins can induce gene expression from the HIV-1 long terminal repeat promoter, and that combinations of TALE transcription factors can synergistically reactivate latent viral expression in cell line models of HIV-1 latency. We further show that complementing TALE transcription factors with Vorinostat, a histone deacetylase inhibitor, enhances HIV-1 expression in latency models. Collectively, these findings demonstrate that TALE transcription factors are a potentially effective alternative to current pharmacological routes for reactivating latent virus and that combining synthetic transcriptional activators with histone deacetylase inhibitors could lead to the development of improved therapies for latent HIV-1 infection.

  10. Reactivation of Latent HIV-1 Expression by Engineered TALE Transcription Factors.

    Directory of Open Access Journals (Sweden)

    Pedro Perdigão

    Full Text Available The presence of replication-competent HIV-1 -which resides mainly in resting CD4+ T cells--is a major hurdle to its eradication. While pharmacological approaches have been useful for inducing the expression of this latent population of virus, they have been unable to purge HIV-1 from all its reservoirs. Additionally, many of these strategies have been associated with adverse effects, underscoring the need for alternative approaches capable of reactivating viral expression. Here we show that engineered transcriptional modulators based on customizable transcription activator-like effector (TALE proteins can induce gene expression from the HIV-1 long terminal repeat promoter, and that combinations of TALE transcription factors can synergistically reactivate latent viral expression in cell line models of HIV-1 latency. We further show that complementing TALE transcription factors with Vorinostat, a histone deacetylase inhibitor, enhances HIV-1 expression in latency models. Collectively, these findings demonstrate that TALE transcription factors are a potentially effective alternative to current pharmacological routes for reactivating latent virus and that combining synthetic transcriptional activators with histone deacetylase inhibitors could lead to the development of improved therapies for latent HIV-1 infection.

  11. Curaxin CBL0100 Blocks HIV-1 Replication and Reactivation through Inhibition of Viral Transcriptional Elongation

    Directory of Open Access Journals (Sweden)

    Maxime J. Jean

    2017-10-01

    Full Text Available Despite combination antiretroviral therapy (cART, acquired immunodeficiency syndrome (AIDS, predominantly caused by the human immunodeficiency virus type 1 (HIV-1, remains incurable. The barrier to a cure lies in the virus' ability to establish a latent infection in HIV/AIDS patients. Unsurprisingly, efforts for a sterilizing cure have focused on the “shock and kill” strategy using latency-reversing agents (LRAs to complement cART in order to eliminate these latent reservoirs. However, this method faces numerous challenges. Recently, the “block and lock” strategy has been proposed. It aims to reinforce a deep state of latency and prevent sporadic reactivation (“blip” of HIV-1 using latency-promoting agents (LPAs for a functional cure. Our studies of curaxin 100 (CBL0100, a small-molecule targeting the facilitates chromatin transcription (FACT complex, show that it blocks both HIV-1 replication and reactivation in in vitro and ex vivo models of HIV-1. Mechanistic investigation elucidated that CBL0100 preferentially targets HIV-1 transcriptional elongation and decreases the occupancy of RNA Polymerase II (Pol II and FACT at the HIV-1 promoter region. In conclusion, CBL0100 is a newly identified inhibitor of HIV-1 transcription that can be used as an LPA in the “block and lock” cure strategy.

  12. Strand transfer and elongation of HIV-1 reverse transcription is facilitated by cell factors in vitro.

    Directory of Open Access Journals (Sweden)

    David Warrilow

    Full Text Available Recent work suggests a role for multiple host factors in facilitating HIV-1 reverse transcription. Previously, we identified a cellular activity which increases the efficiency of HIV-1 reverse transcription in vitro. Here, we describe aspects of the activity which shed light on its function. The cellular factor did not affect synthesis of strong-stop DNA but did improve downstream DNA synthesis. The stimulatory activity was isolated by gel filtration in a single fraction of the exclusion volume. Velocity-gradient purified HIV-1, which was free of detectable RNase activity, showed poor reverse transcription efficiency but was strongly stimulated by partially purified cell proteins. Hence, the cell factor(s did not inactivate an RNase activity that might degrade the viral genomic RNA and block completion of reverse transcription. Instead, the cell factor(s enhanced first strand transfer and synthesis of late reverse transcription suggesting it stabilized the reverse transcription complex. The factor did not affect lysis of HIV-1 by Triton X-100 in the endogenous reverse transcription (ERT system, and ERT reactions with HIV-1 containing capsid mutations, which varied the biochemical stability of viral core structures and impeded reverse transcription in cells, showed no difference in the ability to be stimulated by the cell factor(s suggesting a lack of involvement of the capsid in the in vitro assay. In addition, reverse transcription products were found to be resistant to exogenous DNase I activity when the active fraction was present in the ERT assay. These results indicate that the cell factor(s may improve reverse transcription by facilitating DNA strand transfer and DNA synthesis. It also had a protective function for the reverse transcription products, but it is unclear if this is related to improved DNA synthesis.

  13. LTR real-time PCR for HIV-1 DNA quantitation in blood cells for early diagnosis in infants born to seropositive mothers treated in HAART area (ANRS CO 01).

    Science.gov (United States)

    Avettand-Fènoël, Véronique; Chaix, Marie-Laure; Blanche, Stéphane; Burgard, Marianne; Floch, Corinne; Toure, Kadidia; Allemon, Marie-Christine; Warszawski, Josiane; Rouzioux, Christine

    2009-02-01

    HIV-1 diagnosis in babies born to seropositive mothers is one of the challenges of HIV epidemics in children. A simple, rapid protocol was developed for quantifying HIV-1 DNA in whole blood samples and was used in the ANRS French pediatric cohort in conditions of prevention of mother-to-child transmission. A quantitative HIV-1 DNA protocol (LTR real-time PCR) requiring small blood volumes was developed. First, analytical reproducibility was evaluated on 172 samples. Results obtained on blood cell pellets and Ficoll-Hypaque separated mononuclear cells were compared in 48 adult HIV-1 samples. Second, the protocol was applied to HIV-1 diagnosis in infants in parallel with plasma HIV-RNA quantitation. This prospective study was performed in children born between May 2005 and April 2007 included in the ANRS cohort. The assay showed good reproducibility. The 95% detection cut-off value was 6 copies/PCR, that is, 40 copies/10(6) leukocytes. HIV-DNA levels in whole blood were highly correlated with those obtained after Ficoll-Hypaque separation (r = 0.900, P mothers have received HAART. (c) 2008 Wiley-Liss, Inc.

  14. Identification of a methylated oligoribonucleotide as a potent inhibitor of HIV-1 reverse transcription complex.

    Science.gov (United States)

    Grigorov, Boyan; Bocquin, Anne; Gabus, Caroline; Avilov, Sergey; Mély, Yves; Agopian, Audrey; Divita, Gilles; Gottikh, Marina; Witvrouw, Myriam; Darlix, Jean-Luc

    2011-07-01

    Upon HIV-1 infection of a target cell, the viral reverse transcriptase (RT) copies the genomic RNA to synthesize the viral DNA. The genomic RNA is within the incoming HIV-1 core where it is coated by molecules of nucleocapsid (NC) protein that chaperones the reverse transcription process. Indeed, the RT chaperoning properties of NC extend from the initiation of cDNA synthesis to completion of the viral DNA. New and effective drugs against HIV-1 continue to be required, which prompted us to search for compounds aimed at inhibiting NC protein. Here, we report that the NC chaperoning activity is extensively inhibited in vitro by small methylated oligoribonucleotides (mODN). These mODNs were delivered intracellularly using a cell-penetrating-peptide and found to impede HIV-1 replication in primary human cells at nanomolar concentrations. Extensive analysis showed that viral cDNA synthesis was severely impaired by mODNs. Partially resistant viruses with mutations in NC and RT emerged after months of passaging in cell culture. A HIV-1 molecular clone (NL4.3) bearing these mutations was found to replicate at high concentrations of mODN, albeit with a reduced fitness. Small, methylated ODNs such as mODN-11 appear to be a new type of highly potent inhibitor of HIV-1.

  15. RNA glycosidase and other agents target Tat to inhibit HIV-1 transcription.

    Science.gov (United States)

    Harrich, David; Jin, Hongping

    2018-03-20

    The HIV-1 tat gene encodes a small 86-104 amino acid protein depending on the HIV-1 strain. Tat is essential for HIV-1 replication through interactions with numerous cellular transcription factors. The interaction between Tat and P-TEFb, which is a cellular protein complex composed of cyclin T1 and CDK9, delivers P-TEFb to the newly transcribed viral mRNAs where phosphorylation of RNA polymerase II by CDK9 leads to highly efficient mRNA transcription. It has long been recognized that Tat is a potential anti-HIV-1 target and possibly a viral Achilles' heel. However, specifically targeting Tat without affecting normal host cell functions has been challenging. Means to inactivate Tat have been reported that includes small compounds, transdominant negative Tat proteins, and by plant-derived antivirals. Investigations of these agents have reported encouraging outcomes that inform and may hopefully affect strategies for a functional HIV-1 cure. © 2018 The Author(s). Published by Portland Press Limited on behalf of the Biochemical Society.

  16. Interaction between HIV-1 Tat and DNA-PKcs modulates HIV transcription and class switch recombination.

    Science.gov (United States)

    Zhang, Shi-Meng; Zhang, He; Yang, Tian-Yi; Ying, Tian-Yi; Yang, Pei-Xiang; Liu, Xiao-Dan; Tang, Sheng-Jian; Zhou, Ping-Kun

    2014-01-01

    HIV-1 tat targets a variety of host cell proteins to facilitate viral transcription and disrupts host cellular immunity by inducing lymphocyte apoptosis, but whether it influences humoral immunity remains unclear. Previously, our group demonstrated that tat depresses expression of DNA-PKcs, a critical component of the non-homologous end joining pathway (NHEJ) of DNA double-strand breaks repair, immunoglobulin class switch recombination (CSR) and V(D)J recombination, and sensitizes cells to ionizing radiation. In this study, we demonstrated that HIV-1 Tat down-regulates DNA-PKcs expression by directly binding to the core promoter sequence. In addition, Tat interacts with and activates the kinase activity of DNA-PKcs in a dose-dependent and DNA independent manner. Furthermore, Tat inhibits class switch recombination (CSR) at low concentrations (≤ 4 µg/ml) and stimulates CSR at high concentrations (≥ 8 µg/ml). On the other hand, low protein level and high kinase activity of DNA-PKcs promotes HIV-1 transcription, while high protein level and low kinase activity inhibit HIV-1 transcription. Co-immunoprecipitation results revealed that DNA-PKcs forms a large complex comprised of Cyclin T1, CDK9 and Tat via direct interacting with CDK9 and Tat but not Cyclin T1. Taken together, our results provide new clues that Tat regulates host humoral immunity via both transcriptional depression and kinase activation of DNA-PKcs. We also raise the possibility that inhibitors and interventions directed towards DNA-PKcs may inhibit HIV-1 transcription in AIDS patients.

  17. Development of 5' LTR DNA methylation of latent HIV-1 provirus in cell line models and in long-term-infected individuals

    Czech Academy of Sciences Publication Activity Database

    Trejbalová, K.; Kovářová, D.; Blažková, J.; Machala, L.; Jilich, D.; Weber, Jan; Kučerová, D.; Vencálek, O.; Hirsch, Ivan; Hejnar, J.

    2016-01-01

    Roč. 8, Feb 19 (2016), č. článku 19. ISSN 1868-7083 Institutional support: RVO:61388963 Keywords : HIV-1 * latent reservoir * DNA methylation * chromatin conformation * latent HIV-1 provirus reactivation * HIV-1-infected individuals Subject RIV: EE - Microbiology, Virology Impact factor: 4.987, year: 2016 http://clinicalepigeneticsjournal.biomedcentral.com/articles/10.1186/s13148-016-0185-6

  18. Development of 5 ' LTR DNA methylation of latent HIV-1 provirus in cell line models and in long-term-infected individuals

    Czech Academy of Sciences Publication Activity Database

    Trejbalová, Kateřina; Kovářová, Denisa; Blažková, Jana; Machala, L.; Jilich, D.; Weber, J.; Kučerová, Dana; Vencálek, O.; Hirsch, Ivan; Hejnar, Jiří

    2016-01-01

    Roč. 8, zima (2016), č. článku 19. ISSN 1868-7083 R&D Projects: GA ČR GAP304/12/1736 Institutional support: RVO:68378050 Keywords : HIV-1 * latent reservoir * DNA methylation * chromatin conformation * latent HIV-1 provirus reactivation * HIV-1-infected individuals Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 4.987, year: 2016

  19. Eliminating HIV-1 Packaging Sequences from Lentiviral Vector Proviruses Enhances Safety and Expedites Gene Transfer for Gene Therapy.

    Science.gov (United States)

    Vink, Conrad A; Counsell, John R; Perocheau, Dany P; Karda, Rajvinder; Buckley, Suzanne M K; Brugman, Martijn H; Galla, Melanie; Schambach, Axel; McKay, Tristan R; Waddington, Simon N; Howe, Steven J

    2017-08-02

    Lentiviral vector genomic RNA requires sequences that partially overlap wild-type HIV-1 gag and env genes for packaging into vector particles. These HIV-1 packaging sequences constitute 19.6% of the wild-type HIV-1 genome and contain functional cis elements that potentially compromise clinical safety. Here, we describe the development of a novel lentiviral vector (LTR1) with a unique genomic structure designed to prevent transfer of HIV-1 packaging sequences to patient cells, thus reducing the total HIV-1 content to just 4.8% of the wild-type genome. This has been achieved by reconfiguring the vector to mediate reverse-transcription with a single strand transfer, instead of the usual two, and in which HIV-1 packaging sequences are not copied. We show that LTR1 vectors offer improved safety in their resistance to remobilization in HIV-1 particles and reduced frequency of splicing into human genes. Following intravenous luciferase vector administration to neonatal mice, LTR1 sustained a higher level of liver transgene expression than an equivalent dose of a standard lentivirus. LTR1 vectors produce reverse-transcription products earlier and start to express transgenes significantly quicker than standard lentiviruses after transduction. Finally, we show that LTR1 is an effective lentiviral gene therapy vector as demonstrated by correction of a mouse hemophilia B model. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

  20. SUN2 Modulates HIV-1 Infection and Latency through Association with Lamin A/C To Maintain the Repressive Chromatin.

    Science.gov (United States)

    Sun, Wei-Wei; Jiao, Shi; Sun, Li; Zhou, Zhaocai; Jin, Xia; Wang, Jian-Hua

    2018-05-01

    The postintegrational latency of HIV-1 is characterized by reversible silencing of long terminal repeat (LTR)-driven transcription of the HIV genome. It is known that the formation of repressive chromatin at the 5'-LTR of HIV-1 proviral DNA impedes viral transcription by blocking the recruitment of positive transcription factors. How the repressive chromatin is formed and modulated during HIV-1 infection remains elusive. Elucidation of which chromatin reassembly factor mediates the reorganization of chromatin is likely to facilitate the understanding of the host's modulation of HIV-1 transcription and latency. Here we revealed that "Sad1 and UNC84 domain containing 2" (SUN2), an inner nuclear membrane protein, maintained the repressive chromatin and inhibited HIV LTR-driven transcription of proviral DNA through an association with lamin A/C. Specifically, lamin A/C tethered SUN2 to the nucleosomes 1 and 2 of the HIV-1 5'-LTR to block the initiation and elongation of HIV-1 transcription. SUN2 knockdown converted chromatin to an active form and thus enhanced the phosphorylation of RNA polymerase II and its recruitment to the 5'-LTR HIV-1 proviral DNA, leading to reactivation of HIV-1 from latency. Conversely, the exogenous factors such as tumor necrosis factor alpha (TNF-α) induced reactivation, and the replication of HIV-1 led to the disassociation between SUN2 and lamin A/C, suggesting that disruption of the association between SUN2 and lamin A/C to convert the repressive chromatin to the active form might be a prerequisite for the initiation of HIV-1 transcription and replication. Together, our findings indicate that SUN2 is a novel chromatin reassembly factor that helps to maintain chromatin in a repressive state and consequently inhibits HIV-1 transcription. IMPORTANCE Despite the successful use of scores of antiretroviral drugs, HIV latency poses a major impediment to virus eradication. Elucidation of the mechanism of latency facilitates the discovery of new

  1. Human-Phosphate-Binding-Protein inhibits HIV-1 gene transcription and replication

    Directory of Open Access Journals (Sweden)

    Candolfi Ermanno

    2011-07-01

    Full Text Available Abstract The Human Phosphate-Binding protein (HPBP is a serendipitously discovered lipoprotein that binds phosphate with high affinity. HPBP belongs to the DING protein family, involved in various biological processes like cell cycle regulation. We report that HPBP inhibits HIV-1 gene transcription and replication in T cell line, primary peripherical blood lymphocytes and primary macrophages. We show that HPBP is efficient in naïve and HIV-1 AZT-resistant strains. Our results revealed HPBP as a new and potent anti HIV molecule that inhibits transcription of the virus, which has not yet been targeted by HAART and therefore opens new strategies in the treatment of HIV infection.

  2. Controlling cellular P-TEFb activity by the HIV-1 transcriptional transactivator Tat.

    Directory of Open Access Journals (Sweden)

    Lisa Muniz

    Full Text Available The human immunodeficiency virus 1 (HIV-1 transcriptional transactivator (Tat is essential for synthesis of full-length transcripts from the integrated viral genome by RNA polymerase II (Pol II. Tat recruits the host positive transcription elongation factor b (P-TEFb to the HIV-1 promoter through binding to the transactivator RNA (TAR at the 5'-end of the nascent HIV transcript. P-TEFb is a general Pol II transcription factor; its cellular activity is controlled by the 7SK small nuclear RNA (snRNA and the HEXIM1 protein, which sequester P-TEFb into transcriptionally inactive 7SK/HEXIM/P-TEFb snRNP. Besides targeting P-TEFb to HIV transcription, Tat also increases the nuclear level of active P-TEFb through promoting its dissociation from the 7SK/HEXIM/P-TEFb RNP by an unclear mechanism. In this study, by using in vitro and in vivo RNA-protein binding assays, we demonstrate that HIV-1 Tat binds with high specificity and efficiency to an evolutionarily highly conserved stem-bulge-stem motif of the 5'-hairpin of human 7SK snRNA. The newly discovered Tat-binding motif of 7SK is structurally and functionally indistinguishable from the extensively characterized Tat-binding site of HIV TAR and importantly, it is imbedded in the HEXIM-binding elements of 7SK snRNA. We show that Tat efficiently replaces HEXIM1 on the 7SK snRNA in vivo and therefore, it promotes the disassembly of the 7SK/HEXIM/P-TEFb negative transcriptional regulatory snRNP to augment the nuclear level of active P-TEFb. This is the first demonstration that HIV-1 specifically targets an important cellular regulatory RNA, most probably to promote viral transcription and replication. Demonstration that the human 7SK snRNA carries a TAR RNA-like Tat-binding element that is essential for the normal transcriptional regulatory function of 7SK questions the viability of HIV therapeutic approaches based on small drugs blocking the Tat-binding site of HIV TAR.

  3. A real-time view of the TAR:Tat:P-TEFb complex at HIV-1 transcription sites

    Directory of Open Access Journals (Sweden)

    Knezevich Anna

    2007-05-01

    Full Text Available Abstract HIV-1 transcription is tightly regulated: silent in long-term latency and highly active in acutely-infected cells. Transcription is activated by the viral protein Tat, which recruits the elongation factor P-TEFb by binding the TAR sequence present in nascent HIV-1 RNAs. In this study, we analyzed the dynamic of the TAR:Tat:P-TEFb complex in living cells, by performing FRAP experiments at HIV-1 transcription sites. Our results indicate that a large fraction of Tat present at these sites is recruited by Cyclin T1. We found that in the presence of Tat, Cdk9 remained bound to nascent HIV-1 RNAs for 71s. In contrast, when transcription was activated by PMA/ionomycin, in the absence of Tat, Cdk9 turned-over rapidly and resided on the HIV-1 promoter for only 11s. Thus, the mechanism of trans-activation determines the residency time of P-TEFb at the HIV-1 gene, possibly explaining why Tat is such a potent transcriptional activator. In addition, we observed that Tat occupied HIV-1 transcription sites for 55s, suggesting that the TAR:Tat:P-TEFb complex dissociates from the polymerase following transcription initiation, and undergoes subsequent cycles of association/dissociation.

  4. Transcriptional Reprogramming during Effector-to-Memory Transition Renders CD4+ T Cells Permissive for Latent HIV-1 Infection.

    Science.gov (United States)

    Shan, Liang; Deng, Kai; Gao, Hongbo; Xing, Sifei; Capoferri, Adam A; Durand, Christine M; Rabi, S Alireza; Laird, Gregory M; Kim, Michelle; Hosmane, Nina N; Yang, Hung-Chih; Zhang, Hao; Margolick, Joseph B; Li, Linghua; Cai, Weiping; Ke, Ruian; Flavell, Richard A; Siliciano, Janet D; Siliciano, Robert F

    2017-10-17

    The latent reservoir for HIV-1 in resting memory CD4 + T cells is the major barrier to curing HIV-1 infection. Studies of HIV-1 latency have focused on regulation of viral gene expression in cells in which latent infection is established. However, it remains unclear how infection initially becomes latent. Here we described a unique set of properties of CD4 + T cells undergoing effector-to-memory transition including temporary upregulation of CCR5 expression and rapid downregulation of cellular gene transcription. These cells allowed completion of steps in the HIV-1 life cycle through integration but suppressed HIV-1 gene transcription, thus allowing the establishment of latency. CD4 + T cells in this stage were substantially more permissive for HIV-1 latent infection than other CD4 + T cells. Establishment of latent HIV-1 infection in CD4 + T could be inhibited by viral-specific CD8 + T cells, a result with implications for elimination of latent HIV-1 infection by T cell-based vaccines. Copyright © 2017. Published by Elsevier Inc.

  5. Transcriptional profiling of primary endometrial epithelial cells following acute HIV-1 exposure reveals gene signatures related to innate immunity.

    Science.gov (United States)

    Zahoor, Muhammad Atif; Woods, Matthew William; Dizzell, Sara; Nazli, Aisha; Mueller, Kristen M; Nguyen, Philip V; Verschoor, Chris P; Kaushic, Charu

    2018-04-01

    Genital epithelial cells (GECs) line the mucosal surface of the female genital tract (FGT) and are the first cells that interface with both commensal microbiota and sexually transmitted pathogens. Despite the protective barrier formed by GECs, the FGT is a major site of HIV-1 infection. This highlights the importance of studying the interaction of HIV-1 and GECs. Using microarray analysis, we characterized the transcriptional profile of primary endometrial GECs grown in the presence or absence of physiological levels of E2 (10 -9  mol/L) or P4 (10 -7  mol/L) following acute exposure to HIV-1 for 6 hours. Acute exposure of primary endometrial GECs to HIV-1 resulted in the expression of genes related to inflammation, plasminogen activation, adhesion and diapedesis and interferon response. Interestingly, exposure to HIV-1 in the presence of E2 and P4 resulted in differential transcriptional profiles, suggesting that the response of primary endometrial GECs to HIV-1 exposure is modulated by female sex hormones. The gene expression signature of endometrial GECs indicates that the response of these cells may be key to determining host susceptibility to HIV-1 and that sex hormones modulate these interactions. This study allows us to explore possible mechanisms that explain the hormone-mediated fluctuation of HIV-1 susceptibility in women. © 2018 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  6. The HIV-1 transcriptional activator Tat has potent nucleic acid chaperoning activities in vitro.

    Science.gov (United States)

    Kuciak, Monika; Gabus, Caroline; Ivanyi-Nagy, Roland; Semrad, Katharina; Storchak, Roman; Chaloin, Olivier; Muller, Sylviane; Mély, Yves; Darlix, Jean-Luc

    2008-06-01

    The human immunodeficiency virus type 1 (HIV-1) is a primate lentivirus that causes the acquired immunodeficiency syndrome (AIDS). In addition to the virion structural proteins and enzyme precursors, that are Gag, Env and Pol, HIV-1 encodes several regulatory proteins, notably a small nuclear transcriptional activator named Tat. The Tat protein is absolutely required for virus replication since it controls proviral DNA transcription to generate the full-length viral mRNA. Tat can also regulate mRNA capping and splicing and was recently found to interfere with the cellular mi- and siRNA machinery. Because of its extensive interplay with nucleic acids, and its basic and disordered nature we speculated that Tat had nucleic acid-chaperoning properties. This prompted us to examine in vitro the nucleic acid-chaperoning activities of Tat and Tat peptides made by chemical synthesis. Here we report that Tat has potent nucleic acid-chaperoning activities according to the standard DNA annealing, DNA and RNA strand exchange, RNA ribozyme cleavage and trans-splicing assays. The active Tat(44-61) peptide identified here corresponds to the smallest known sequence with DNA/RNA chaperoning properties.

  7. Structure and possible function of a G-quadruplex in the long terminal repeat of the proviral HIV-1 genome.

    Science.gov (United States)

    De Nicola, Beatrice; Lech, Christopher J; Heddi, Brahim; Regmi, Sagar; Frasson, Ilaria; Perrone, Rosalba; Richter, Sara N; Phan, Anh Tuân

    2016-07-27

    The long terminal repeat (LTR) of the proviral human immunodeficiency virus (HIV)-1 genome is integral to virus transcription and host cell infection. The guanine-rich U3 region within the LTR promoter, previously shown to form G-quadruplex structures, represents an attractive target to inhibit HIV transcription and replication. In this work, we report the structure of a biologically relevant G-quadruplex within the LTR promoter region of HIV-1. The guanine-rich sequence designated LTR-IV forms a well-defined structure in physiological cationic solution. The nuclear magnetic resonance (NMR) structure of this sequence reveals a parallel-stranded G-quadruplex containing a single-nucleotide thymine bulge, which participates in a conserved stacking interaction with a neighboring single-nucleotide adenine loop. Transcription analysis in a HIV-1 replication competent cell indicates that the LTR-IV region may act as a modulator of G-quadruplex formation in the LTR promoter. Consequently, the LTR-IV G-quadruplex structure presented within this work could represent a valuable target for the design of HIV therapeutics. © The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.

  8. Identification and functional analysis of a second RBF-2 binding site within the HIV-1 promoter

    International Nuclear Information System (INIS)

    Dahabieh, Matthew S.; Ooms, Marcel; Malcolm, Tom; Simon, Viviana; Sadowski, Ivan

    2011-01-01

    Transcription from the HIV-1 long terminal repeat (LTR) is mediated by numerous host transcription factors. In this study we characterized an E-box motif (RBE1) within the core promoter that was previously implicated in both transcriptional activation and repression. We show that RBE1 is a binding site for the RBF-2 transcription factor complex (USF1, USF2, and TFII-I), previously shown to bind an upstream viral element, RBE3. The RBE1 and RBE3 elements formed complexes of identical mobility and protein constituents in gel shift assays, both with Jurkat T-cell nuclear extracts and recombinant USF/TFII-I. Furthermore, both elements are regulators of HIV-1 expression; mutations in LTR-luciferase reporters and in HIV-1 molecular clones resulted in decreased transcription, virion production, and proviral expression in infected cells. Collectively, our data indicate that RBE1 is a bona fide RBF-2 binding site and that the RBE1 and RBE3 elements are necessary for mediating proper transcription from the HIV-1 LTR.

  9. Histone acetyltransferase (HAT) activity of p300 modulates human T lymphotropic virus type 1 p30II-mediated repression of LTR transcriptional activity

    International Nuclear Information System (INIS)

    Michael, Bindhu; Nair, Amrithraj M.; Datta, Antara; Hiraragi, Hajime; Ratner, Lee; Lairmore, Michael D.

    2006-01-01

    Human T-lymphotropic virus type-1 (HTLV-1) is a deltaretrovirus that causes adult T cell leukemia/lymphoma, and is implicated in a variety of lymphocyte-mediated inflammatory disorders. HTLV-1 provirus has regulatory and accessory genes in four pX open reading frames. HTLV-1 pX ORF-II encodes two proteins, p13 II and p30 II , which are incompletely defined in virus replication or pathogenesis. We have demonstrated that pX ORF-II mutations block virus replication in vivo and that ORF-II encoded p30 II , a nuclear-localizing protein that binds with CREB-binding protein (CBP)/p300, represses CREB and Tax responsive element (TRE)-mediated transcription. Herein, we have identified p30 II motifs important for p300 binding and in regulating TRE-mediated transcription in the absence and presence of HTLV-1 provirus. Within amino acids 100-179 of p30 II , a region important for repression of LTR-mediated transcription, we identified a single lysine residue at amino acid 106 (K3) that significantly modulates the ability of p30 II to repress TRE-mediated transcription. Exogenous p300, in a dose-responsive manner, reverses p30 II -dependent repression of TRE-mediated transcription, in the absence or presence of the provirus, In contrast to wild type p300, p300 HAT mutants (defective in histone acetyltransferase activity) only partially rescued p30 II -mediated LTR repression. Deacetylation by histone deacetylase-1 (HDAC-1) enhanced p30 II -mediated LTR repression, while inhibition of deacetylation by trichostatin A decreases p30 II -mediated LTR repression. Collectively, our data indicate that HTLV-1 p30 II modulates viral gene expression in a cooperative manner with p300-mediated acetylation

  10. UV-induced transcription from the human immunodeficiency virus type 1 (HIV-1) long terminal repeat and UV-induced secretion of an extracellular factor that induces HIV-1 transcription in nonirradiated cells

    International Nuclear Information System (INIS)

    Stein, B.; Kraemer, M.R.; Rahmsdorf, H.J.; Ponta, H.; Herrlich, P.

    1989-01-01

    UV irradiation, but not visible sunlight, induces the transcription of human immunodeficiency virus type 1 (HIV-1). Chimeric constructs carrying all or parts of the HIV-1 long terminal repeat linked to an indicator gene were transfected into HeLa cells or murine and human T-cell lines, and their response to irradiation was tested. The cis-acting element conferring UV responsiveness is identical to the sequence binding transcription factor NF kappa B. UV irradiation enhances NF kappa B binding activity as assayed by gel retardation experiments. Interestingly, the requirement for UV irradiation can be replaced by cocultivation of transfected cells with UV-irradiated nontransfected (HIV-1-negative) cells. A UV-induced extracellular protein factor is detected in the culture medium conditioned by UV-treated cells. The factor is produced upon UV irradiation by several murine and human cell lines, including HeLa, Molt-4, and Jurkat, and acts on several cells. These data suggest that the UV response of keratinocytes in human skin can be magnified and spread to deeper layers that are more shielded, including the Langerhans cells, and that this indirect UV response may contribute to the activation of HIV-1 in humans

  11. Selective histonedeacetylase inhibitor M344 intervenes in HIV-1 latency through increasing histone acetylation and activation of NF-kappaB.

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    Hao Ying

    Full Text Available Histone deacetylase (HDAC inhibitors present an exciting new approach to activate HIV production from latently infected cells to potentially enhance elimination of these cells and achieve a cure. M344, a novel HDAC inhibitor, shows robust activity in a variety of cancer cells and relatively low toxicity compared to trichostatin A (TSA. However, little is known about the effects and action mechanism of M344 in inducing HIV expression in latently infected cells.Using the Jurkat T cell model of HIV latency, we demonstrate that M344 effectively reactivates HIV-1 gene expression in latently infected cells. Moreover, M344-mediated activation of the latent HIV LTR can be strongly inhibited by a NF-κB inhibitor aspirin. We further show that M344 acts by increasing the acetylation of histone H3 and histone H4 at the nucleosome 1 (nuc-1 site of the HIV-1 long terminal repeat (LTR and by inducing NF-κB p65 nuclear translocation and direct RelA DNA binding at the nuc-1 region of the HIV-1 LTR. We also found that M344 synergized with prostratin to activate the HIV-1 LTR promoter in latently infected cells.These results suggest the potential of M344 in anti-latency therapies and an important role for histone modifications and NF-κB transcription factors in regulating HIV-1 LTR gene expression.

  12. On the role of the second coding exon of the HIV-1 Tat protein in virus replication and MHC class I downregulation

    NARCIS (Netherlands)

    Verhoef, K.; Bauer, M.; Meyerhans, A.; Berkhout, B.

    1998-01-01

    Tat is an essential protein of human immunodeficiency virus type 1 (HIV-1) and activates transcription from the viral long terminal repeat (LTR) promoter. The tat gene is composed of two coding exons of which the first, corresponding to the N-terminal 72 amino acid residues, has been reported to be

  13. Human Immunodeficiency Virus-Type 1 LTR DNA contains an intrinsic gene producing antisense RNA and protein products

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    Hsiao Chiu-Bin

    2006-11-01

    Full Text Available Abstract Background While viruses have long been shown to capitalize on their limited genomic size by utilizing both strands of DNA or complementary DNA/RNA intermediates to code for viral proteins, it has been assumed that human retroviruses have all their major proteins translated only from the plus or sense strand of RNA, despite their requirement for a dsDNA proviral intermediate. Several studies, however, have suggested the presence of antisense transcription for both HIV-1 and HTLV-1. More recently an antisense transcript responsible for the HTLV-1 bZIP factor (HBZ protein has been described. In this study we investigated the possibility of an antisense gene contained within the human immunodeficiency virus type 1 (HIV-1 long terminal repeat (LTR. Results Inspection of published sequences revealed a potential transcription initiator element (INR situated downstream of, and in reverse orientation to, the usual HIV-1 promoter and transcription start site. This antisense initiator (HIVaINR suggested the possibility of an antisense gene responsible for RNA and protein production. We show that antisense transcripts are generated, in vitro and in vivo, originating from the TAR DNA of the HIV-1 LTR. To test the possibility that protein(s could be translated from this novel HIV-1 antisense RNA, recombinant HIV antisense gene-FLAG vectors were designed. Recombinant protein(s were produced and isolated utilizing carboxy-terminal FLAG epitope (DYKDDDDK sequences. In addition, affinity-purified antisera to an internal peptide derived from the HIV antisense protein (HAP sequences identified HAPs from HIV+ human peripheral blood lymphocytes. Conclusion HIV-1 contains an antisense gene in the U3-R regions of the LTR responsible for both an antisense RNA transcript and proteins. This antisense transcript has tremendous potential for intrinsic RNA regulation because of its overlap with the beginning of all HIV-1 sense RNA transcripts by 25 nucleotides. The

  14. Single Nucleotide Polymorphism in Gene Encoding Transcription Factor Prep1 Is Associated with HIV-1-Associated Dementia

    Science.gov (United States)

    van Manen, Daniëlle; Bunnik, Evelien M.; van Sighem, Ard I.; Sieberer, Margit; Boeser-Nunnink, Brigitte; de Wolf, Frank; Schuitemaker, Hanneke; Portegies, Peter; Kootstra, Neeltje A.; van 't Wout, Angélique B.

    2012-01-01

    Background Infection with HIV-1 may result in severe cognitive and motor impairment, referred to as HIV-1-associated dementia (HAD). While its prevalence has dropped significantly in the era of combination antiretroviral therapy, milder neurocognitive disorders persist with a high prevalence. To identify additional therapeutic targets for treating HIV-associated neurocognitive disorders, several candidate gene polymorphisms have been evaluated, but few have been replicated across multiple studies. Methods We here tested 7 candidate gene polymorphisms for association with HAD in a case-control study consisting of 86 HAD cases and 246 non-HAD AIDS patients as controls. Since infected monocytes and macrophages are thought to play an important role in the infection of the brain, 5 recently identified single nucleotide polymorphisms (SNPs) affecting HIV-1 replication in macrophages in vitro were also tested. Results The CCR5 wt/Δ32 genotype was only associated with HAD in individuals who developed AIDS prior to 1991, in agreement with the observed fading effect of this genotype on viral load set point. A significant difference in genotype distribution among all cases and controls irrespective of year of AIDS diagnosis was found only for a SNP in candidate gene PREP1 (p = 1.2×10−5). Prep1 has recently been identified as a transcription factor preferentially binding the −2,518 G allele in the promoter of the gene encoding MCP-1, a protein with a well established role in the etiology of HAD. Conclusion These results support previous findings suggesting an important role for MCP-1 in the onset of HIV-1-associated neurocognitive disorders. PMID:22347417

  15. Single nucleotide polymorphism in gene encoding transcription factor Prep1 is associated with HIV-1-associated dementia.

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    Sebastiaan M Bol

    Full Text Available BACKGROUND: Infection with HIV-1 may result in severe cognitive and motor impairment, referred to as HIV-1-associated dementia (HAD. While its prevalence has dropped significantly in the era of combination antiretroviral therapy, milder neurocognitive disorders persist with a high prevalence. To identify additional therapeutic targets for treating HIV-associated neurocognitive disorders, several candidate gene polymorphisms have been evaluated, but few have been replicated across multiple studies. METHODS: We here tested 7 candidate gene polymorphisms for association with HAD in a case-control study consisting of 86 HAD cases and 246 non-HAD AIDS patients as controls. Since infected monocytes and macrophages are thought to play an important role in the infection of the brain, 5 recently identified single nucleotide polymorphisms (SNPs affecting HIV-1 replication in macrophages in vitro were also tested. RESULTS: The CCR5 wt/Δ32 genotype was only associated with HAD in individuals who developed AIDS prior to 1991, in agreement with the observed fading effect of this genotype on viral load set point. A significant difference in genotype distribution among all cases and controls irrespective of year of AIDS diagnosis was found only for a SNP in candidate gene PREP1 (p = 1.2 × 10(-5. Prep1 has recently been identified as a transcription factor preferentially binding the -2,518 G allele in the promoter of the gene encoding MCP-1, a protein with a well established role in the etiology of HAD. CONCLUSION: These results support previous findings suggesting an important role for MCP-1 in the onset of HIV-1-associated neurocognitive disorders.

  16. Requirements for DNA strand transfer during reverse transcription in mutant HIV-1 virions

    NARCIS (Netherlands)

    Berkhout, B.; van Wamel, J.; Klaver, B.

    1995-01-01

    Retroviruses convert their RNA genome into a DNA form by means of reverse transcription. According to the current model of reverse transcription, two strand transfer reactions are needed to synthesize a full-length DNA genome. Because reverse transcription is initiated close to the 5' end of the RNA

  17. Differential expression and interaction of host factors augment HIV-1 gene expression in neonatal mononuclear cells

    International Nuclear Information System (INIS)

    Sundaravaradan, Vasudha; Mehta, Roshni; Harris, David T.; Zack, Jerome A.; Ahmad, Nafees

    2010-01-01

    We have previously shown a higher level of HIV-1 replication and gene expression in neonatal (cord) blood mononuclear cells (CBMC) compared with adult blood cells (PBMC), which could be due to differential expression of host factors. We performed the gene expression profile of CBMC and PBMC and found that 8013 genes were expressed at higher levels in CBMC than PBMC and 8028 genes in PBMC than CBMC, including 1181 and 1414 genes upregulated after HIV-1 infection in CBMC and PBMC, respectively. Several transcription factors (NF-κB, E2F, HAT-1, TFIIE, Cdk9, Cyclin T1), signal transducers (STAT3, STAT5A) and cytokines (IL-1β, IL-6, IL-10) were upregulated in CBMC than PBMC, which are known to influence HIV-1 replication. In addition, a repressor of HIV-1 transcription, YY1, was down regulated in CBMC than PBMC and several matrix metalloproteinase (MMP-7, -12, -14) were significantly upregulated in HIV-1 infected CBMC than PBMC. Furthermore, we show that CBMC nuclear extracts interacted with a higher extent to HIV-1 LTR cis-acting sequences, including NF-κB, NFAT, AP1 and NF-IL6 compared with PBMC nuclear extracts and retroviral based short hairpin RNA (shRNA) for STAT3 and IL-6 down regulated their own and HIV-1 gene expression, signifying that these factors influenced differential HIV-1 gene expression in CBMC than PBMC.

  18. HIV-1 Promoter Single Nucleotide Polymorphisms Are Associated with Clinical Disease Severity.

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    Michael R Nonnemacher

    Full Text Available The large majority of human immunodeficiency virus type 1 (HIV-1 markers of disease progression/severity previously identified have been associated with alterations in host genetic and immune responses, with few studies focused on viral genetic markers correlate with changes in disease severity. This study presents a cross-sectional/longitudinal study of HIV-1 single nucleotide polymorphisms (SNPs contained within the viral promoter or long terminal repeat (LTR in patients within the Drexel Medicine CNS AIDS Research and Eradication Study (CARES Cohort. HIV-1 LTR SNPs were found to associate with the classical clinical disease parameters CD4+ T-cell count and log viral load. They were found in both defined and undefined transcription factor binding sites of the LTR. A novel SNP identified at position 108 in a known COUP (chicken ovalbumin upstream promoter/AP1 transcription factor binding site was significantly correlated with binding phenotypes that are potentially the underlying cause of the associated clinical outcome (increase in viral load and decrease in CD4+ T-cell count.

  19. NFAT5 regulates HIV-1 in primary monocytes via a highly conserved long terminal repeat site.

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    Shahin Ranjbar

    2006-12-01

    Full Text Available To replicate, HIV-1 capitalizes on endogenous cellular activation pathways resulting in recruitment of key host transcription factors to its viral enhancer. RNA interference has been a powerful tool for blocking key checkpoints in HIV-1 entry into cells. Here we apply RNA interference to HIV-1 transcription in primary macrophages, a major reservoir of the virus, and specifically target the transcription factor NFAT5 (nuclear factor of activated T cells 5, which is the most evolutionarily divergent NFAT protein. By molecularly cloning and sequencing isolates from multiple viral subtypes, and performing DNase I footprinting, electrophoretic mobility shift, and promoter mutagenesis transfection assays, we demonstrate that NFAT5 functionally interacts with a specific enhancer binding site conserved in HIV-1, HIV-2, and multiple simian immunodeficiency viruses. Using small interfering RNA to ablate expression of endogenous NFAT5 protein, we show that the replication of three major HIV-1 viral subtypes (B, C, and E is dependent upon NFAT5 in human primary differentiated macrophages. Our results define a novel host factor-viral enhancer interaction that reveals a new regulatory role for NFAT5 and defines a functional DNA motif conserved across HIV-1 subtypes and representative simian immunodeficiency viruses. Inhibition of the NFAT5-LTR interaction may thus present a novel therapeutic target to suppress HIV-1 replication and progression of AIDS.

  20. HIV-1 reverse transcription initiation: a potential target for novel antivirals?

    NARCIS (Netherlands)

    Abbink, Truus E. M.; Berkhout, Ben

    2008-01-01

    Reverse transcription is an essential step in the retroviral life cycle, as it converts the genomic RNA into DNA. In this review, we describe recent developments concerning the initiation step of this complex, multi-step reaction. During initiation of reverse transcription, a cellular tRNA primer is

  1. A Novel Toll-Like Receptor 9 Agonist, MGN1703, Enhances HIV-1 Transcription and NK Cell-Mediated Inhibition of HIV-1-Infected Autologous CD4+ T Cells.

    Science.gov (United States)

    Offersen, Rasmus; Nissen, Sara Konstantin; Rasmussen, Thomas A; Østergaard, Lars; Denton, Paul W; Søgaard, Ole Schmeltz; Tolstrup, Martin

    2016-05-01

    Toll-like receptor (TLR) agonists are potent enhancers of innate antiviral immunity and may also reverse HIV-1 latency. Therefore, TLR agonists have a potential role in the context of a "shock-and-kill" approach to eradicate HIV-1. Our extensive preclinical evaluation suggests that a novel TLR9 agonist, MGN1703, may indeed perform both functions in an HIV-1 eradication trial. Peripheral blood mononuclear cells (PBMCs) from aviremic HIV-1-infected donors on antiretroviral therapy (ART) that were incubated with MGN1703 ex vivo exhibited increased secretion of interferon alpha (IFN-α) (P= 0.005) and CXCL10 (P= 0.0005) in culture supernatants. Within the incubated PBMC pool, there were higher proportions of CD69-positive CD56(dim)CD16(+)NK cells (P= 0.001) as well as higher proportions of CD107a-positive (P= 0.002) and IFN-γ-producing (P= 0.038) NK cells. Incubation with MGN1703 also increased the proportions of CD69-expressing CD4(+)and CD8(+)T cells. Furthermore, CD4(+)T cells within the pool of MGN1703-incubated PBMCs showed enhanced levels of unspliced HIV-1 RNA (P= 0.036). Importantly, MGN1703 increased the capacity of NK cells to inhibit virus spread within a culture of autologous CD4(+)T cells assessed by using an HIV-1 p24 enzyme-linked immunosorbent assay (ELISA) (P= 0.03). In conclusion, we show that MGN1703 induced strong antiviral innate immune responses, enhanced HIV-1 transcription, and boosted NK cell-mediated suppression of HIV-1 infection in autologous CD4(+)T cells. These findings support clinical testing of MGN1703 in HIV-1 eradication trials. We demonstrate that MGN1703 (a TLR9 agonist currently undergoing phase 3 clinical testing for the treatment of metastatic colorectal cancer) induces potent antiviral responses in immune effector cells from HIV-1-infected individuals on suppressive antiretroviral therapy. The significantly improved safety and tolerability profiles of MGN1703 versus TLR9 agonists of the CpG-oligodeoxynucleotide (CpG-ODN) family

  2. HIV-1 in the RNA world: Transcription regulation, miRNAs and antiviral RNAs

    NARCIS (Netherlands)

    Harwig, A.

    2015-01-01

    All organisms, from bacteria to human, use three biological molecules that each serve critical functions in the expression of genes in the cell. These are deoxyribonucleic acid (DNA), ribonucleic acid (RNA) and proteins. RNA is synthesized from DNA in a process called transcription. RNA differs from

  3. Structural features in the HIV-1 repeat region facilitate strand transfer during reverse transcription

    NARCIS (Netherlands)

    Berkhout, B.; Vastenhouw, N. L.; Klasens, B. I.; Huthoff, H.

    2001-01-01

    Two obligatory DNA strand transfers take place during reverse transcription of a retroviral RNA genome. The first strand transfer is facilitated by terminal repeat (R) elements in the viral genome. This strand-transfer reaction depends on base pairing between the cDNA of the 5'R and the 3'R. There

  4. The RNA binding protein HuR does not interact directly with HIV-1 reverse transcriptase and does not affect reverse transcription in vitro

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    Gronenborn Angela M

    2010-05-01

    Full Text Available Abstract Background Lemay et al recently reported that the RNA binding protein HuR directly interacts with the ribonuclease H (RNase H domain of HIV-1 reverse transcriptase (RT and influences the efficiency of viral reverse transcription (Lemay et al., 2008, Retrovirology 5:47. HuR is a member of the embryonic lethal abnormal vision protein family and contains 3 RNA recognition motifs (RRMs that bind AU-rich elements (AREs. To define the structural determinants of the HuR-RT interaction and to elucidate the mechanism(s by which HuR influences HIV-1 reverse transcription activity in vitro, we cloned and purified full-length HuR as well as three additional protein constructs that contained the N-terminal and internal RRMs, the internal and C-terminal RRMs, or the C-terminal RRM only. Results All four HuR proteins were purified and characterized by biophysical methods. They are well structured and exist as monomers in solution. No direct protein-protein interaction between HuR and HIV-1 RT was detected using NMR titrations with 15N labeled HuR variants or the 15N labeled RNase H domain of HIV-1 RT. Furthermore, HuR did not significantly affect the kinetics of HIV-1 reverse transcription in vitro, even on RNA templates that contain AREs. Conclusions Our results suggest that HuR does not impact HIV-1 replication through a direct protein-protein interaction with the viral RT.

  5. NF-kappaB and p53 are the dominant apoptosis-inducing transcription factors elicited by the HIV-1 envelope.

    Science.gov (United States)

    Perfettini, Jean-Luc; Roumier, Thomas; Castedo, Maria; Larochette, Nathanael; Boya, Patricia; Raynal, Brigitte; Lazar, Vladimir; Ciccosanti, Fabiola; Nardacci, Roberta; Penninger, Josef; Piacentini, Mauro; Kroemer, Guido

    2004-03-01

    The coculture of cells expressing the HIV-1 envelope glycoprotein complex (Env) with cells expressing CD4 results into cell fusion, deregulated mitosis, and subsequent cell death. Here, we show that NF-kappaB, p53, and AP1 are activated in Env-elicited apoptosis. The nuclear factor kappaB (NF-kappaB) super repressor had an antimitotic and antiapoptotic effect and prevented the Env-elicited phosphorylation of p53 on serine 15 and 46, as well as the activation of AP1. Transfection with dominant-negative p53 abolished apoptosis and AP1 activation. Signs of NF-kappaB and p53 activation were also detected in lymph node biopsies from HIV-1-infected individuals. Microarrays revealed that most (85%) of the transcriptional effects of HIV-1 Env were blocked by the p53 inhibitor pifithrin-alpha. Macroarrays led to the identification of several Env-elicited, p53-dependent proapoptotic transcripts, in particular Puma, a proapoptotic "BH3-only" protein from the Bcl-2 family known to activate Bax/Bak. Down modulation of Puma by antisense oligonucleotides, as well as RNA interference of Bax and Bak, prevented Env-induced apoptosis. HIV-1-infected primary lymphoblasts up-regulated Puma in vitro. Moreover, circulating CD4+ lymphocytes from untreated, HIV-1-infected donors contained enhanced amounts of Puma protein, and these elevated Puma levels dropped upon antiretroviral therapy. Altogether, these data indicate that NF-kappaB and p53 cooperate as the dominant proapoptotic transcription factors participating in HIV-1 infection.

  6. Transcriptional regulation of HIV-1 host factor COMMD1 by the Sp family.

    Science.gov (United States)

    Kudo, Eriko; Taura, Manabu; Suico, Mary Ann; Goto, Hiroki; Kai, Hirofumi; Okada, Seiji

    2018-04-01

    Copper metabolism Murr1 domain containing 1 (COMMD1) has multiple functions in the regulation of protein stability at the plasma membrane and in the cytoplasm. However, the regulation of COMMD1 transcriptional has remained to be elucidated. In the present study, the 5'‑flanking region (‑1,192/+83 bp) of the human COMMD1 gene was cloned. It was observed that the COMMD1 promoter region contains GC‑rich region that has 7 putative Sp1‑binding sites via in silico analysis. The proximal promoter region at ‑289/+83 bp was required for COMMD1 basal promoter activity by deletion constructs of COMMD1 promoter. Moreover, Sp1 inhibitor, mithramycin A, suppressed basal COMMD1 promoter activity. The Sp1‑binding site (‑11/‑1 bp) in the proximal promoter region was a critical site for COMMD1 gene regulation by Sp1 and Sp3. Sp1 upregulated COMMD1 promoter activity, whereas Sp3 suppressed it. Endogenous Sp1 and Sp3 bound to the proximal promoter region of COMMD1. Taken together, Sp1 constitutively regulates the basal expression of the COMMD1 gene in human epithelial cell lines.

  7. N-terminally truncated POM121C inhibits HIV-1 replication.

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    Hideki Saito

    Full Text Available Recent studies have identified host cell factors that regulate early stages of HIV-1 infection including viral cDNA synthesis and orientation of the HIV-1 capsid (CA core toward the nuclear envelope, but it remains unclear how viral DNA is imported through the nuclear pore and guided to the host chromosomal DNA. Here, we demonstrate that N-terminally truncated POM121C, a component of the nuclear pore complex, blocks HIV-1 infection. This truncated protein is predominantly localized in the cytoplasm, does not bind to CA, does not affect viral cDNA synthesis, reduces the formation of 2-LTR and diminished the amount of integrated proviral DNA. Studies with an HIV-1-murine leukemia virus (MLV chimeric virus carrying the MLV-derived Gag revealed that Gag is a determinant of this inhibition. Intriguingly, mutational studies have revealed that the blockade by N-terminally-truncated POM121C is closely linked to its binding to importin-β/karyopherin subunit beta 1 (KPNB1. These results indicate that N-terminally-truncated POM121C inhibits HIV-1 infection after completion of reverse transcription and before integration, and suggest an important role for KPNB1 in HIV-1 replication.

  8. Knockdown of MAP4 and DNAL1 produces a post-fusion and pre-nuclear translocation impairment in HIV-1 replication

    International Nuclear Information System (INIS)

    Gallo, Daniel E.; Hope, Thomas J.

    2012-01-01

    DNAL1 and MAP4 are both microtubule-associated proteins. These proteins were identified as HIV-1 dependency factors in a screen with wild-type HIV-1. In this study we demonstrate that knockdown using DNAL1 and MAP4 siRNAs and shRNAs inhibits HIV-1 infection regardless of envelope. Using a fusion assay, we show that DNAL1 and MAP4 do not impact fusion. By assaying for late reverse transcripts and 2-LTR circles, we show that DNAL1 and MAP4 inhibit both by approximately 50%. These results demonstrate that DNAL1 and MAP4 impact reverse transcription but not nuclear translocation. DNAL1 and MAP4 knockdown cells do not display cytoskeletal defects. Together these experiments indicate that DNAL1 and MAP4 may exert their functions in the HIV life cycle at reverse transcription, prior to nuclear translocation.

  9. Regulatory elements involved in tax-mediated transactivation of the HTLV-I LTR.

    Science.gov (United States)

    Seeler, J S; Muchardt, C; Podar, M; Gaynor, R B

    1993-10-01

    HTLV-I is the etiologic agent of adult T-cell leukemia. In this study, we investigated the regulatory elements and cellular transcription factors which function in modulating HTLV-I gene expression in response to the viral transactivator protein, tax. Transfection experiments into Jurkat cells of a variety of site-directed mutants in the HTLV-1 LTR indicated that each of the three motifs A, B, and C within the 21-bp repeats, the binding sites for the Ets family of proteins, and the TATA box all influenced the degree of tax-mediated activation. Tax is also able to activate gene expression of other viral and cellular promoters. Tax activation of the IL-2 receptor and the HIV-1 LTR is mediated through NF-kappa B motifs. Interestingly, sequences in the 21-bp repeat B and C motifs contain significant homology with NF-kappa B regulatory elements. We demonstrated that an NF-kappa B binding protein, PRDII-BF1, but not the rel protein, bound to the B and C motifs in the 21-bp repeat. PRDII-BF1 was also able to stimulate activation of HTLV-I gene expression by tax. The role of the Ets proteins on modulating tax activation was also studied. Ets 1 but not Ets 2 was capable of increasing the degree of tax activation of the HTLV-I LTR. These results suggest that tax activates gene expression by either direct or indirect interaction with several cellular transcription factors that bind to the HTLV-I LTR.

  10. Regulatory CD4 T cells inhibit HIV-1 expression of other CD4 T cell subsets via interactions with cell surface regulatory proteins.

    Science.gov (United States)

    Zhang, Mingce; Robinson, Tanya O; Duverger, Alexandra; Kutsch, Olaf; Heath, Sonya L; Cron, Randy Q

    2018-03-01

    During chronic HIV-1 infection, regulatory CD4 T cells (Tregs) frequently represent the largest subpopulation of CD4 T cell subsets, implying relative resistant to HIV-1. When HIV-1 infection of CD4 T cells was explored in vitro and ex vivo from patient samples, Tregs possessed lower levels of HIV-1 DNA and RNA in comparison with conventional effector and memory CD4 T cells. Moreover, Tregs suppressed HIV-1 expression in other CD4 T cells in an in vitro co-culture system. This suppression was mediated in part via multiple inhibitory surface proteins expressed on Tregs. Antibody blockade of CTLA-4, PD-1, and GARP on Tregs resulted in increased HIV-1 DNA integration and mRNA expression in neighboring CD4 T cells. Moreover, antibody blockade of Tregs inhibitory proteins resulted in increased HIV-1 LTR transcription in co-cultured CD4 T cells. Thus, Tregs inhibit HIV-1 infection of other CD4 T cell subsets via interactions with inhibitory cell surface proteins. Copyright © 2018 Elsevier Inc. All rights reserved.

  11. Trans-activation of the 5' to 3' viral DNA strand transfer by nucleocapsid protein during reverse transcription of HIV1 RNA.

    Science.gov (United States)

    Darlix, J L; Vincent, A; Gabus, C; de Rocquigny, H; Roques, B

    1993-08-01

    Two DNA strand transfer reactions take place during reverse transcription of the retroviral genome. The first transfer, that of the minus-strand strong stop DNA from the 5' end of the viral RNA to the 3' end, has been studied in vitro with two RNAs mimicking the 5' and 3' regions of the HIV1 genome and with nucleocapsid protein, NCp7, and reverse transcriptase. The results show that NCp7 strongly activates the 5' to 3' DNA strand transfer during reverse transcription while a basic peptide resembling NCp7 is inactive. Activation of the first transfer by several NCp7 derived peptides and the influence of the terminal redundancies (R) present at the 5' and 3' ends of HIV1 RNA were also examined. The first transfer is optimal in the presence of intact NCp7 and necessitates R on both the 5' and 3' RNAs. Sequencing of full length viral DNA products reveals approximately 40% misincorporations at the first nucleotide beyond the transfer point. If such base misincorporations occur during proviral DNA synthesis with possible homologous recombinations it may well contribute to the high level of genetic variability of HIV.

  12. Yeast tRNAPhe expressed in human cells can be selected by HIV-1 for use as a reverse transcription primer

    International Nuclear Information System (INIS)

    Kelly, Nathan J.; Morrow, Casey D.

    2003-01-01

    All naturally occurring human immune deficiency viruses (HIV-1) select and use tRNA Lys,3 as the primer for reverse transcription. Studies to elucidate the mechanism of tRNA selection from the intracellular milieu have been hampered due to the difficulties in manipulating the endogenous levels of tRNA Lys,3 . We have previously described a mutant HIV-1 with a primer binding site (PBS) complementary to yeast tRNA Phe (psHIV-Phe) that relies on transfection of yeast tRNA Phe for infectivity. To more accurately recapitulate the selection process, a cDNA was designed for the intracellular expression of the yeast tRNA Phe . Increasing amounts of the plasmid encoding tRNA Phe resulted in a corresponding increase in levels of yeast tRNA Phe in the cell. The yeast tRNA Phe isolated from cells transfected with the cDNA for yeast tRNA Phe , or in the cell lines expressing yeast tRNA Phe , were aminoacylated, indicating that the expressed yeast tRNA Phe was incorporated into tRNA biogenesis pathways and translation. Increasing the cytoplasmic levels of tRNA Phe resulted in increased encapsidation of tRNA Phe in viruses with a PBS complementary to tRNA Phe (psHIV-Phe) or tRNA Lys,3 (wild-type HIV-1). Production of infectious psHIV-Phe was dependent on the amount of cotransfected tRNA Phe cDNA. Increasing amounts of plasmids encoding yeast tRNA Phe produced an increase of infectious psHIV-Phe that plateaued at a level lower than that from the transfection of the wild-type genome, which uses tRNA Lys,3 as the primer for reverse transcription. Cell lines were generated that expressed yeast tRNA Phe at levels approximately 0.1% of that for tRNA Lys,3 . Even with this reduced level of yeast tRNA Phe , the cell lines complemented psHIV-Phe over background levels. The results of these studies demonstrate that intracellular levels of primer tRNA can have a direct effect on HIV-1 infectivity and further support the role for PBS-tRNA complementarity in the primer selection process

  13. LTR retrotransposons in fungi.

    Directory of Open Access Journals (Sweden)

    Anna Muszewska

    Full Text Available Transposable elements with long terminal direct repeats (LTR TEs are one of the best studied groups of mobile elements. They are ubiquitous elements present in almost all eukaryotic genomes. Their number and state of conservation can be a highlight of genome dynamics. We searched all published fungal genomes for LTR-containing retrotransposons, including both complete, functional elements and remnant copies. We identified a total of over 66,000 elements, all of which belong to the Ty1/Copia or Ty3/Gypsy superfamilies. Most of the detected Gypsy elements represent Chromoviridae, i.e. they carry a chromodomain in the pol ORF. We analyzed our data from a genome-ecology perspective, looking at the abundance of various types of LTR TEs in individual genomes and at the highest-copy element from each genome. The TE content is very variable among the analyzed genomes. Some genomes are very scarce in LTR TEs (8000 elements. The data shows that transposon expansions in fungi usually involve an increase both in the copy number of individual elements and in the number of element types. The majority of the highest-copy TEs from all genomes are Ty3/Gypsy transposons. Phylogenetic analysis of these elements suggests that TE expansions have appeared independently of each other, in distant genomes and at different taxonomical levels. We also analyzed the evolutionary relationships between protein domains encoded by the transposon pol ORF and we found that the protease is the fastest evolving domain whereas reverse transcriptase and RNase H evolve much slower and in correlation with each other.

  14. LEDGF/p75 Deficiency Increases Deletions at the HIV-1 cDNA Ends.

    Science.gov (United States)

    Bueno, Murilo T D; Reyes, Daniel; Llano, Manuel

    2017-09-15

    Processing of unintegrated linear HIV-1 cDNA by the host DNA repair system results in its degradation and/or circularization. As a consequence, deficient viral cDNA integration generally leads to an increase in the levels of HIV-1 cDNA circles containing one or two long terminal repeats (LTRs). Intriguingly, impaired HIV-1 integration in LEDGF/p75-deficient cells does not result in a correspondent increase in viral cDNA circles. We postulate that increased degradation of unintegrated linear viral cDNA in cells lacking the lens epithelium-derived growth factor (LEDGF/p75) account for this inconsistency. To evaluate this hypothesis, we characterized the nucleotide sequence spanning 2-LTR junctions isolated from LEDGF/p75-deficient and control cells. LEDGF/p75 deficiency resulted in a significant increase in the frequency of 2-LTRs harboring large deletions. Of note, these deletions were dependent on the 3' processing activity of integrase and were not originated by aberrant reverse transcription. Our findings suggest a novel role of LEDGF/p75 in protecting the unintegrated 3' processed linear HIV-1 cDNA from exonucleolytic degradation.

  15. Tat-dependent repression of human immunodeficiency virus type 1 long terminal repeat promoter activity by fusion of cellular transcription factors

    International Nuclear Information System (INIS)

    Zhao Cunyou; Chen Yali; Park, Jiyoung; Kim, Jae Bum; Tang Hong

    2004-01-01

    Transcription initiation from HIV-1 long terminal repeat (LTR) promoter requires the virally encoded transactivator, Tat, and several cellular co-factors to accomplish the Tat-dependent processive transcription elongation. Individual cellular transcription activators, LBP-1b and Oct-1, on the other hand, have been shown to inhibit LTR promoter activities probably via competitive binding against TFIID to the TATA-box in LTR promoter. To explore the genetic interference strategies against the viral replication, we took advantage of the existence of the bipartite DNA binding domains and the repression domains of LBP-1b and Oct-1 factors to generate a chimeric transcription repressor. Our results indicated that the fusion protein of LBP-1b and Oct-1 exhibited higher DNA binding affinity to the viral promoter than the individual factors, and little interference with the host cell gene expression due to its anticipated rare cognate DNA sites in the host cell genome. Moreover, the chimera exerted increased Tat-dependent repression of transcription initiation at the LTR promoter both in vitro and in vivo compared to LBP-1b, Oct-1 or combination of LBP-1b and Oct-1. These results might provide the lead in generating a therapeutic reagent useful to suppress HIV-1 replication

  16. Keeping your armour intact: how HIV-1 evades detection by the innate immune system: HIV-1 capsid controls detection of reverse transcription products by the cytosolic DNA sensor cGAS.

    Science.gov (United States)

    Maelfait, Jonathan; Seiradake, Elena; Rehwinkel, Jan

    2014-07-01

    HIV-1 infects dendritic cells (DCs) without triggering an effective innate antiviral immune response. As a consequence, the induction of adaptive immune responses controlling virus spread is limited. In a recent issue of Immunity, Lahaye and colleagues show that intricate interactions of HIV capsid with the cellular cofactor cyclophilin A (CypA) control infection and innate immune activation in DCs. Manipulation of HIV-1 capsid to increase its affinity for CypA results in reduced virus infectivity and facilitates access of the cytosolic DNA sensor cGAS to reverse transcribed DNA. This in turn induces a strong host response. Here, we discuss these findings in the context of recent developments in innate immunity and consider the implications for disease control and vaccine design. © 2014 The Authors. Bioessays published by WILEY Periodicals, Inc.

  17. HIV-1 Tat affects the programming and functionality of human CD8⁺ T cells by modulating the expression of T-box transcription factors.

    Science.gov (United States)

    Sforza, Fabio; Nicoli, Francesco; Gallerani, Eleonora; Finessi, Valentina; Reali, Eva; Cafaro, Aurelio; Caputo, Antonella; Ensoli, Barbara; Gavioli, Riccardo

    2014-07-31

    HIV infection is characterized by several immune dysfunctions of both CD8⁺ and CD4⁺ T cells as hyperactivation, impairment of functionality and expansion of memory T cells. CD8⁺ T-cell dysfunctions have been associated with increased expression of T-bet, Eomesdermin and pro-inflammatory cytokines, and with down-regulation of CD127. The HIV-1 trans-activator of transcription (Tat) protein, which is released by infected cells and detected in tissues of HIV-positive individuals, is known to contribute to the dysregulation of CD4⁺ T cells; however, its effects on CD8⁺ T cells have not been investigated. Thus, in this study, we sought to address whether Tat may affect CD8⁺ T-cell functionality and programming. CD8⁺ T cells were activated by T-cell receptor engagement in the presence or absence of Tat. Cytokine production, killing capacity, surface phenotype and expression of transcription factors important for T-cell programming were evaluated. Tat favors the secretion of interleukin-2, interferon-γ and granzyme B in CD8⁺ T cells. Behind this functional modulation we observed that Tat increases the expression of T-bet, Eomesdermin, Blimp-1, Bcl-6 and Bcl-2 in activated but not in unstimulated CD8⁺ T lymphocytes. This effect is associated with the down-regulation of CD127 and the up-regulation of CD27. Tat deeply alters the programming and functionality of CD8⁺ T lymphocytes.

  18. [Non-LTR retrotransposons: LINEs and SINEs in plant genome].

    Science.gov (United States)

    Cheng, Xu-Dong; Ling, Hong-Qing

    2006-06-01

    Retrotransposons are one of the drivers of genome evolution. They include LTR (long terminal repeat) retrotransposons, which widespread in Eukaryotagenomes, show structural similarity to retroviruses. Non-LTR retrotransposons were first discovered in animal genomes and then identified as ubiquitous components of nuclear genomes in many species across the plant kingdom. They constitute a large fraction of the repetitive DNA. Non-LTR retrotransposons are divided into LINEs (long interspersed nuclear elements) and SINEs (short interspersed nuclear elements). Transposition of non-LTR retrotransposons is rarely observed in plants indicating that most of them are inactive and/or under regulation of the host genome. Transposition is poorly understood, but experimental evidence from other genetic systems shows that LINEs are able to transpose autonomously while non-autonomous SINEs depend on the reverse transcription machinery of other retrotransposons. Phylogenic analysis shows LINEs are probably the most ancient class of retrotransposons in plant genomes, while the origin of SINEs is unknown. This review sums up the above data and wants to show readers a clear picture of non-LTR retrotransposons.

  19. HIV-1 transcripts use IRES-initiation under conditions where Cap-dependent translation is restricted by poliovirus 2A protease.

    Directory of Open Access Journals (Sweden)

    Raquel Amorim

    Full Text Available The 30 different species of mRNAs synthesized during the HIV-1 replication cycle are all capped and polyadenilated. Internal ribosome entry sites have been recognized in the 5' untranslated region of some mRNA species of HIV-1, which would contribute to an alternative mechanism of initiation of mRNA translation. However, the Cap-dependent translation is assumed to be the main mechanism driving the initiation of HIV-1 protein synthesis. In this work, we describe a cell system in which lower to higher levels of transient expression of the poliovirus 2A protease strongly inhibited cellular Cap-dependent translation with no toxic effect to the cells during a 72-hour time frame. In this system, the synthesis of HIV-1 proteins was inhibited in a temporal dose-dependent way. Higher levels of 2A protease expression severely inhibited HIV-1 protein synthesis during the first 24 hours of infection consequently inhibiting viral production and infectivity. Intermediate to lower levels of 2A Protease expression caused the inhibition of viral protein synthesis only during the first 48 hours of viral replication. After this period both protein synthesis and viral release were recovered to the control levels. However, the infectivity of viral progeny was still partially inhibited. These results indicate that two mechanisms of mRNA translation initiation contribute to the synthesis of HIV-1 proteins; during the first 24-48 hours of viral replication HIV-1 protein synthesis is strongly dependent on Cap-initiation, while at later time points IRES-driven translation initiation is sufficient to produce high amounts of viral particles.

  20. HIV-1 envelope glycoprotein

    Science.gov (United States)

    Caulfield, Michael; Cupo, Albert; Dean, Hansi; Hoffenberg, Simon; King, C. Richter; Klasse, P. J.; Marozsan, Andre; Moore, John P.; Sanders, Rogier W.; Ward, Andrew; Wilson, Ian; Julien, Jean-Philippe

    2017-08-22

    The present application relates to novel HIV-1 envelope glycoproteins, which may be utilized as HIV-1 vaccine immunogens, and antigens for crystallization, electron microscopy and other biophysical, biochemical and immunological studies for the identification of broad neutralizing antibodies. The present invention encompasses the preparation and purification of immunogenic compositions, which are formulated into the vaccines of the present invention.

  1. Accurate episomal HIV 2-LTR circles quantification using optimized DNA isolation and droplet digital PCR.

    Science.gov (United States)

    Malatinkova, Eva; Kiselinova, Maja; Bonczkowski, Pawel; Trypsteen, Wim; Messiaen, Peter; Vermeire, Jolien; Verhasselt, Bruno; Vervisch, Karen; Vandekerckhove, Linos; De Spiegelaere, Ward

    2014-01-01

    In HIV-infected patients on combination antiretroviral therapy (cART), the detection of episomal HIV 2-LTR circles is a potential marker for ongoing viral replication. Quantification of 2-LTR circles is based on quantitative PCR or more recently on digital PCR assessment, but is hampered due to its low abundance. Sample pre-PCR processing is a critical step for 2-LTR circles quantification, which has not yet been sufficiently evaluated in patient derived samples. We compared two sample processing procedures to more accurately quantify 2-LTR circles using droplet digital PCR (ddPCR). Episomal HIV 2-LTR circles were either isolated by genomic DNA isolation or by a modified plasmid DNA isolation, to separate the small episomal circular DNA from chromosomal DNA. This was performed in a dilution series of HIV-infected cells and HIV-1 infected patient derived samples (n=59). Samples for the plasmid DNA isolation method were spiked with an internal control plasmid. Genomic DNA isolation enables robust 2-LTR circles quantification. However, in the lower ranges of detection, PCR inhibition caused by high genomic DNA load substantially limits the amount of sample input and this impacts sensitivity and accuracy. Moreover, total genomic DNA isolation resulted in a lower recovery of 2-LTR templates per isolate, further reducing its sensitivity. The modified plasmid DNA isolation with a spiked reference for normalization was more accurate in these low ranges compared to genomic DNA isolation. A linear correlation of both methods was observed in the dilution series (R2=0.974) and in the patient derived samples with 2-LTR numbers above 10 copies per million peripheral blood mononuclear cells (PBMCs), (R2=0.671). Furthermore, Bland-Altman analysis revealed an average agreement between the methods within the 27 samples in which 2-LTR circles were detectable with both methods (bias: 0.3875±1.2657 log10). 2-LTR circles quantification in HIV-infected patients proved to be more

  2. Characterization of EIAV LTR variability and compartmentalization in various reservoir tissues of long-term inapparent carrier ponies

    International Nuclear Information System (INIS)

    Reis, Jenner K.P.; Craigo, Jodi K.; Cook, Sheila J.; Issel, Charles J.; Montelaro, Ronald C.

    2003-01-01

    Dynamic genomic variation resulting in changes in envelope antigenicity has been established as a fundamental mechanism of persistence by equine infectious anemia virus (EIAV), as observed with other lentiviruses, including HIV-1. In addition to the reported changes in envelope sequences, however, certain studies indicate the viral LTR as a second variable EIAV gene, with the enhancer region being designated as hypervariable. These observations have lead to the suggestion that LTR variation may alter viral replication properties to optimize to the microenvironment of particular tissue reservoirs. To test this hypothesis directly, we examined the population of LTR quasispecies contained in various tissues of two inapparent carrier ponies experimentally infected with a reference EIAV biological clone for 18 months. The results of these studies demonstrated that the EIAV LTR is in fact highly conserved with respect to the infecting LTR species after 1.5 years of persistent infection and regardless of the tissue reservoir. Thus, these comprehensive analyses demonstrate for the first time that the EIAV LTR is highly conserved during long-term persistent infection and that the observed variations in viral LTR are associated more with in vitro adaptation to replication in cultured cells rather than in vivo replication in natural target cells

  3. Human cyclin T1 expression ameliorates a T-cell-specific transcriptional limitation for HIV in transgenic rats, but is not sufficient for a spreading infection of prototypic R5 HIV-1 strains ex vivo

    Directory of Open Access Journals (Sweden)

    Littman Dan R

    2009-01-01

    Full Text Available Abstract Background Cells derived from native rodents have limits at distinct steps of HIV replication. Rat primary CD4 T-cells, but not macrophages, display a profound transcriptional deficit that is ameliorated by transient trans-complementation with the human Tat-interacting protein Cyclin T1 (hCycT1. Results Here, we generated transgenic rats that selectively express hCycT1 in CD4 T-cells and macrophages. hCycT1 expression in rat T-cells boosted early HIV gene expression to levels approaching those in infected primary human T-cells. hCycT1 expression was necessary, but not sufficient, to enhance HIV transcription in T-cells from individual transgenic animals, indicating that endogenous cellular factors are critical co-regulators of HIV gene expression in rats. T-cells from hCD4/hCCR5/hCycT1-transgenic rats did not support productive infection of prototypic wild-type R5 HIV-1 strains ex vivo, suggesting one or more significant limitation in the late phase of the replication cycle in this primary rodent cell type. Remarkably, we identify a replication-competent HIV-1 GFP reporter strain (R7/3 YU-2 Env that displays characteristics of a spreading, primarily cell-to-cell-mediated infection in primary T-cells from hCD4/hCCR5-transgenic rats. Moreover, the replication of this recombinant HIV-1 strain was significantly enhanced by hCycT1 transgenesis. The viral determinants of this so far unique replicative ability are currently unknown. Conclusion Thus, hCycT1 expression is beneficial to de novo HIV infection in a transgenic rat model, but additional genetic manipulations of the host or virus are required to achieve full permissivity.

  4. NF-κB and p53 Are the Dominant Apoptosis-inducing Transcription Factors Elicited by the HIV-1 Envelope

    OpenAIRE

    Perfettini, Jean-Luc; Roumier, Thomas; Castedo, Maria; Larochette, Nathanael; Boya, Patricia; Raynal, Brigitte; Lazar, Vladimir; Ciccosanti, Fabiola; Nardacci, Roberta; Penninger, Josef; Piacentini, Mauro; Kroemer, Guido

    2004-01-01

    The coculture of cells expressing the HIV-1 envelope glycoprotein complex (Env) with cells expressing CD4 results into cell fusion, deregulated mitosis, and subsequent cell death. Here, we show that NF-κB, p53, and AP1 are activated in Env-elicited apoptosis. The nuclear factor κB (NF-κB) super repressor had an antimitotic and antiapoptotic effect and prevented the Env-elicited phosphorylation of p53 on serine 15 and 46, as well as the activation of AP1. Transfection with dominant-negative p5...

  5. SUN1 Regulates HIV-1 Nuclear Import in a Manner Dependent on the Interaction between the Viral Capsid and Cellular Cyclophilin A.

    Science.gov (United States)

    Luo, Xinlong; Yang, Wei; Gao, Guangxia

    2018-07-01

    Human immunodeficiency virus type 1 (HIV-1) can infect nondividing cells via passing through the nuclear pore complex. The nuclear membrane-imbedded protein SUN2 was recently reported to be involved in the nuclear import of HIV-1. Whether SUN1, which shares many functional similarities with SUN2, is involved in this process remained to be explored. Here we report that overexpression of SUN1 specifically inhibited infection by HIV-1 but not that by simian immunodeficiency virus (SIV) or murine leukemia virus (MLV). Overexpression of SUN1 did not affect reverse transcription but led to reduced accumulation of the 2-long-terminal-repeat (2-LTR) circular DNA and integrated viral DNA, suggesting a block in the process of nuclear import. HIV-1 CA was mapped as a determinant for viral sensitivity to SUN1. Treatment of SUN1-expressing cells with cyclosporine (CsA) significantly reduced the sensitivity of the virus to SUN1, and an HIV-1 mutant containing CA-G89A, which does not interact with cyclophilin A (CypA), was resistant to SUN1 overexpression. Downregulation of endogenous SUN1 inhibited the nuclear entry of the wild-type virus but not that of the G89A mutant. These results indicate that SUN1 participates in the HIV-1 nuclear entry process in a manner dependent on the interaction of CA with CypA. IMPORTANCE HIV-1 infects both dividing and nondividing cells. The viral preintegration complex (PIC) can enter the nucleus through the nuclear pore complex. It has been well known that the viral protein CA plays an important role in determining the pathways by which the PIC enters the nucleus. In addition, the interaction between CA and the cellular protein CypA has been reported to be important in the selection of nuclear entry pathways, though the underlying mechanisms are not very clear. Here we show that both SUN1 overexpression and downregulation inhibited HIV-1 nuclear entry. CA played an important role in determining the sensitivity of the virus to SUN1: the regulatory

  6. Nup153 and Nup98 bind the HIV-1 core and contribute to the early steps of HIV-1 replication

    International Nuclear Information System (INIS)

    Di Nunzio, Francesca; Fricke, Thomas; Miccio, Annarita; Valle-Casuso, Jose Carlos; Perez, Patricio; Souque, Philippe; Rizzi, Ermanno; Severgnini, Marco; Mavilio, Fulvio; Charneau, Pierre; Diaz-Griffero, Felipe

    2013-01-01

    The early steps of HIV-1 replication involve the entry of HIV-1 into the nucleus, which is characterized by viral interactions with nuclear pore components. HIV-1 developed an evolutionary strategy to usurp the nuclear pore machinery and chromatin in order to integrate and efficiently express viral genes. In the current work, we studied the role of nucleoporins 153 and 98 (Nup153 and Nup98) in infection of human Jurkat lymphocytes by HIV-1. We showed that Nup153-depleted cells exhibited a defect in nuclear import, while depletion of Nup 98 caused a slight defect in HIV integration. To explore the biochemical viral determinants for the requirement of Nup153 and Nup98 during HIV-1 infection, we tested the ability of these nucleoporins to interact with HIV-1 cores. Our findings showed that both nucleoporins bind HIV-1 cores suggesting that this interaction is important for HIV-1 nuclear import and/or integration. Distribution analysis of integration sites in Nup153-depleted cells revealed a reduced tendency of HIV-1 to integrate in intragenic sites, which in part could account for the large infectivity defect observed in Nup153-depleted cells. Our work strongly supports a role for Nup153 in HIV-1 nuclear import and integration. - Highlights: ► We studied the role of Nup98 and Nup153 in HIV-1 infection. ► Nup98 binds the HIV-1 core and is involved in HIV-1 integration. ► Nup153 binds the HIV-1 core and is involved in HIV-1 nuclear import. ► Depletion of Nup153 decreased the integration of HIV-1 in transcriptionally active sites

  7. Nup153 and Nup98 bind the HIV-1 core and contribute to the early steps of HIV-1 replication

    Energy Technology Data Exchange (ETDEWEB)

    Di Nunzio, Francesca, E-mail: francesca.di-nunzio@pasteur.fr [Molecular Virology and Vaccinology unit, CNRS URA 3015, Department of Virology, Institut Pasteur, 25-28 rue du Dr. Roux, 75015 Paris (France); Fricke, Thomas [Department of Microbiology and Immunology, Albert Einstein College of Medicine Bronx, NY 10461 (United States); Miccio, Annarita [University of Modena e Reggio Emilia, Centro di Medicina Rigenerativa, Modena (Italy); Valle-Casuso, Jose Carlos; Perez, Patricio [Department of Microbiology and Immunology, Albert Einstein College of Medicine Bronx, NY 10461 (United States); Souque, Philippe [Molecular Virology and Vaccinology unit, CNRS URA 3015, Department of Virology, Institut Pasteur, 25-28 rue du Dr. Roux, 75015 Paris (France); Rizzi, Ermanno; Severgnini, Marco [Institute of Biomedical Technologies, CNR, Milano (Italy); Mavilio, Fulvio [University of Modena e Reggio Emilia, Centro di Medicina Rigenerativa, Modena (Italy); Genethon, Evry (France); Charneau, Pierre [Molecular Virology and Vaccinology unit, CNRS URA 3015, Department of Virology, Institut Pasteur, 25-28 rue du Dr. Roux, 75015 Paris (France); Diaz-Griffero, Felipe, E-mail: felipe.diaz-griffero@einstein.yu.edu [Department of Microbiology and Immunology, Albert Einstein College of Medicine Bronx, NY 10461 (United States)

    2013-05-25

    The early steps of HIV-1 replication involve the entry of HIV-1 into the nucleus, which is characterized by viral interactions with nuclear pore components. HIV-1 developed an evolutionary strategy to usurp the nuclear pore machinery and chromatin in order to integrate and efficiently express viral genes. In the current work, we studied the role of nucleoporins 153 and 98 (Nup153 and Nup98) in infection of human Jurkat lymphocytes by HIV-1. We showed that Nup153-depleted cells exhibited a defect in nuclear import, while depletion of Nup 98 caused a slight defect in HIV integration. To explore the biochemical viral determinants for the requirement of Nup153 and Nup98 during HIV-1 infection, we tested the ability of these nucleoporins to interact with HIV-1 cores. Our findings showed that both nucleoporins bind HIV-1 cores suggesting that this interaction is important for HIV-1 nuclear import and/or integration. Distribution analysis of integration sites in Nup153-depleted cells revealed a reduced tendency of HIV-1 to integrate in intragenic sites, which in part could account for the large infectivity defect observed in Nup153-depleted cells. Our work strongly supports a role for Nup153 in HIV-1 nuclear import and integration. - Highlights: ► We studied the role of Nup98 and Nup153 in HIV-1 infection. ► Nup98 binds the HIV-1 core and is involved in HIV-1 integration. ► Nup153 binds the HIV-1 core and is involved in HIV-1 nuclear import. ► Depletion of Nup153 decreased the integration of HIV-1 in transcriptionally active sites.

  8. Novel 3′-Processing Integrase Activity Assay by Real-Time PCR for Screening and Identification of HIV-1 Integrase Inhibitors

    Directory of Open Access Journals (Sweden)

    Supachai Sakkhachornphop

    2015-01-01

    Full Text Available The 3′-end processing (3′P of each viral long terminal repeat (LTR during human immunodeficiency virus type-1 (HIV-1 integration is a vital step in the HIV life cycle. Blocking the 3′P using 3′P inhibitor has recently become an attractive strategy for HIV-1 therapeutic intervention. Recently, we have developed a novel real-time PCR based assay for the detection of 3′P activity in vitro. The methodology usually involves biotinylated HIV-1 LTR, HIV-1 integrase (IN, and specific primers and probe. In this novel assay, we designed the HIV-1 LTR substrate based on a sequence with a homology to HIV-1 LTR labeled at its 3′ end with biotin on the sense strand. Two nucleotides at the 3′ end were subsequently removed by IN activity. Only two nucleotides labeled biotin were captured on an avidin-coated tube; therefore, inhibiting the binding of primers and probe results in late signals in the real-time PCR. This novel assay has successfully detected both the 3′P activity of HIV-1 IN and the anti-IN activity by Raltegravir and sodium azide agent. This real-time PCR assay has been shown to be effective and inexpensive for a high-throughput screening of novel IN inhibitors.

  9. Involvement of histone methyltransferase GLP in HIV-1 latency through catalysis of H3K9 dimethylation

    International Nuclear Information System (INIS)

    Ding, Donglin; Qu, Xiying; Li, Lin; Zhou, Xin; Liu, Sijie; Lin, Shiguan; Wang, Pengfei; Liu, Shaohui; Kong, Chuijin; Wang, Xiaohui; Liu, Lin; Zhu, Huanzhang

    2013-01-01

    Understanding the mechanism of HIV-1 latency is crucial to eradication of the viral reservoir in HIV-1-infected individuals. However, the role of histone methyltransferase (HMT) G9a-like protein (GLP) in HIV-1 latency is still unclear. In the present work, we established four clonal cell lines containing HIV-1 vector. We found that the integration sites of most clonal cell lines favored active gene regions. However, we also observed hypomethylation of CpG of HIV 5′LTR in all four clonal cell lines. Additionally, 5′-deoxy-5′-methylthioadenosine (MTA), a broad-spectrum histone methyltransferase inhibitor, was used to examine the role of histone methylation in HIV-1 latency. MTA was found to decrease the level of H3K9 dimethylation, causing reactivation of latent HIV-1 in C11 cells. GLP knockdown by small interfering RNA clearly induced HIV-1 LTR expression. Results suggest that GLP may play a significant role in the maintenance of HIV-1 latency by catalyzing dimethylation of H3K9. - Highlights: ► We have established an in vitro model of HIV-1 latency. ► The integration sites of most clonal cell lines favor in active gene regions. ► Hypomethylation occurs in CpG islands of HIV 5′LTR in all four clonal cell lines. ► MTA can reactivate latent HIV-1 by decreasing the level of H3K9 me2 in C11 cells. ► HMT GLP may play a significant role in the maintenance of HIV-1 latency

  10. HIV-1 vaccines

    Science.gov (United States)

    Excler, Jean-Louis; Robb, Merlin L; Kim, Jerome H

    2014-01-01

    The development of a safe and effective preventive HIV-1 vaccine remains a public health priority. Despite scientific difficulties and disappointing results, HIV-1 vaccine clinical development has, for the first time, established proof-of-concept efficacy against HIV-1 acquisition and identified vaccine-associated immune correlates of risk. The correlate of risk analysis showed that IgG antibodies against the gp120 V2 loop correlated with decreased risk of HIV infection, while Env-specific IgA directly correlated with increased risk. The development of vaccine strategies such as improved envelope proteins formulated with potent adjuvants and DNA and vectors expressing mosaics, or conserved sequences, capable of eliciting greater breadth and depth of potentially relevant immune responses including neutralizing and non-neutralizing antibodies, CD4+ and CD8+ cell-mediated immune responses, mucosal immune responses, and immunological memory, is now proceeding quickly. Additional human efficacy trials combined with other prevention modalities along with sustained funding and international collaboration remain key to bring an HIV-1 vaccine to licensure. PMID:24637946

  11. Reactivation of latent HIV-1 by new semi-synthetic ingenol esters

    Energy Technology Data Exchange (ETDEWEB)

    Pandeló José, Diego [Departamento de Genética, Instituto de Biologia, Universidade Federal do Rio de Janeiro, Rio de Janeiro 21941-902 (Brazil); Bartholomeeusen, Koen [Department of Medicine, Microbiology and Immunology, University of California at San Francisco, San Francisco, CA 94143-0703 (United States); Delveccio da Cunha, Rodrigo; Abreu, Celina Monteiro [Departamento de Genética, Instituto de Biologia, Universidade Federal do Rio de Janeiro, Rio de Janeiro 21941-902 (Brazil); Glinski, Jan [PlantaAnalytica LLC, Danbury, CT 06810 (United States); Barbizan Ferreira da Costa, Thais; Bacchi Rabay, Ana Flávia Mello; Pianowski Filho, Luiz Francisco [Kyolab Laboratories, Valinhos, São Paulo 13273-105 (Brazil); Dudycz, Lech W. [Lex Company Research Laboratories, Shirley 01464, MA (United States); Ranga, Udaykumar [Molecular Biology and Genetics Unit, Jawaharlal Nehru Centre for Advanced Scientific Research, Bengaluru 560064 (India); Peterlin, Boris Matija [Department of Medicine, Microbiology and Immunology, University of California at San Francisco, San Francisco, CA 94143-0703 (United States); Pianowski, Luiz Francisco [Kyolab Laboratories, Valinhos, São Paulo 13273-105 (Brazil); Tanuri, Amilcar [Departamento de Genética, Instituto de Biologia, Universidade Federal do Rio de Janeiro, Rio de Janeiro 21941-902 (Brazil); Aguiar, Renato Santana, E-mail: santana@biologia.ufrj.br [Departamento de Genética, Instituto de Biologia, Universidade Federal do Rio de Janeiro, Rio de Janeiro 21941-902 (Brazil)

    2014-08-15

    The ability of HIV to establish long-lived latent infection is mainly due to transcriptional silencing of viral genome in resting memory T lymphocytes. Here, we show that new semi-synthetic ingenol esters reactivate latent HIV reservoirs. Amongst the tested compounds, 3-caproyl-ingenol (ING B) was more potent in reactivating latent HIV than known activators such as SAHA, ingenol 3,20-dibenzoate, TNF-α, PMA and HMBA. ING B activated PKC isoforms followed by NF-κB nuclear translocation. As virus reactivation is dependent on intact NF-κB binding sites in the LTR promoter region ING B, we have shown that. ING B was able to reactivate virus transcription in primary HIV-infected resting cells up to 12 fold and up to 25 fold in combination with SAHA. Additionally, ING B promoted up-regulation of P-TEFb subunits CDK9/Cyclin T1. The role of ING B on promoting both transcription initiation and elongation makes this compound a strong candidate for an anti-HIV latency drug combined with suppressive HAART. - Highlights: • 3-caproyl-ingenol (ING B) reactivates latent HIV better than SAHA, ingenol 3,20-dibenzoate, TNF-α, PMA and HMBA. • ING B promotes PKC activation and NF-kB translocation to the nucleus. • ING B activates virus transcription of B and non-B subtypes of HIV-1. • ING B activates HIV transcription in infected primary resting CD4+ T cells. • ING B induces higher levels of P-TEFb components in human primary cells.

  12. The effects of multiple UV exposures on HIV-LTR (long terminal repeat) expression

    International Nuclear Information System (INIS)

    Schreck, S.; Milton, J.; Panozzo, J.; Libertin, C.R.; Woloschak, G.E.; Loyola Univ., Maywood, IL

    1995-01-01

    Previous studies have shown that cellular stress agents such as UV radiation induce transcription from the long terminal repeat (LTR) of the human immunodeficiency virus (HIV). Using HeLa cells stably transfected with the HIV-LTR sequence, which transcriptionally drives the chloramphenicol acetyl transferase (CAT) reporter gene, we examined the effects of multiple exposures to UVC (254 nm) on HIV-LTR-CAT expression. Low doses (≤ 5 J m -2 ) had no effect on CAT expression, but up to 29-fold induction was observed with 10 J m -2 when cells were harvested 48 h after completion of the exposure. Little difference was noted in induction levels when cells were exposed to one 25 J m -2 dose, viable cells were harvested at 24 h, 48 h or 72 h, and cell lysates were assayed for CAT expression. Two sequential 12.5 J m -2 exposures, given 24 h apart, resulted in an additive effect on CAT expression; these two exposures produced CAT activity equivalent to that induced following a single 25 J m -2 dose. Our data suggest that HIV-LTR requires a specific threshold UV dose in order to elicit induction; a maximal induction dose is also evident; exposures higher than this maximal dose contribute no more to HIV-LTR induction in viable cells. (author)

  13. Insertion of a solo LTR retrotransposon associates with spur mutations in 'Red Delicious' apple (Malus × domestica).

    Science.gov (United States)

    Han, Mengxue; Sun, Qibao; Zhou, Junyong; Qiu, Huarong; Guo, Jing; Lu, Lijuan; Mu, Wenlei; Sun, Jun

    2017-09-01

    Insertion of a solo LTR, which possesses strong bidirectional, stem-specific promoter activities, is associated with the evolution of a dwarfing apple spur mutation. Spur mutations in apple scions revolutionized global apple production. Since long terminal repeat (LTR) retrotransposons are tightly related to natural mutations, inter-retrotransposon-amplified polymorphism technique and genome walking were used to find sequences in the apple genome based on these LTRs. In 'Red Delicious' spur mutants, a novel, 2190-bp insertion was identified as a spur-specific, solo LTR (sLTR) located at the 1038th nucleotide of another sLTR, which was 1536 bp in length. This insertion-within-an-insertion was localized within a preexisting Gypsy-50 retrotransposon at position 3,762,767 on chromosome 4. The analysis of transcriptional activity of the two sLTRs (the 2190- and 1536-bp inserts) indicated that the 2190-bp sLTR is a promoter, capable of bidirectional transcription. GUS expression in the 2190-bp-sense and 2190-bp-antisense transgenic lines was prominent in stems. In contrast, no promoter activity from either the sense or the antisense strand of the 1536-bp sLTR was detected. From ~150 kb of DNA on each side of the 2190 bp, sLTR insertion site, corresponding to 300 kb of the 'Golden Delicious' genome, 23 genes were predicted. Ten genes had predicted functions that could affect shoot development. This first report, of a sLTR insertion associated with the evolution of apple spur mutation, will facilitate apple breeding, cloning of spur-related genes, and discovery of mechanisms behind dwarf habit.

  14. Smartphone-Imaged HIV-1 Reverse-Transcription Loop-Mediated Isothermal Amplification (RT-LAMP on a Chip from Whole Blood

    Directory of Open Access Journals (Sweden)

    Gregory L. Damhorst

    2015-09-01

    Full Text Available Viral load measurements are an essential tool for the long-term clinical care of human immunodeficiency virus (HIV-positive individuals. The gold standards in viral load instrumentation, however, are still too limited by their size, cost, and sophisticated operation for these measurements to be ubiquitous in remote settings with poor healthcare infrastructure, including parts of the world that are disproportionately affected by HIV infection. The challenge of developing a point-of-care platform capable of making viral load more accessible has been frequently approached but no solution has yet emerged that meets the practical requirements of low cost, portability, and ease-of-use. In this paper, we perform reverse-transcription loop-mediated isothermal amplification (RT-LAMP on minimally processed HIV-spiked whole blood samples with a microfluidic and silicon microchip platform, and perform fluorescence measurements with a consumer smartphone. Our integrated assay shows amplification from as few as three viruses in a ~ 60 nL RT-LAMP droplet, corresponding to a whole blood concentration of 670 viruses per μL of whole blood. The technology contains greater power in a digital RT-LAMP approach that could be scaled up for the determination of viral load from a finger prick of blood in the clinical care of HIV-positive individuals. We demonstrate that all aspects of this viral load approach, from a drop of blood to imaging the RT-LAMP reaction, are compatible with lab-on-a-chip components and mobile instrumentation.

  15. Genetic architecture of HIV-1 genes circulating in north India & their functional implications.

    Science.gov (United States)

    Neogi, Ujjwal; Sood, Vikas; Ronsard, Larence; Singh, Jyotsna; Lata, Sneh; Ramachandran, V G; Das, S; Wanchu, Ajay; Banerjea, Akhil C

    2011-12-01

    This review presents data on genetic and functional analysis of some of the HIV-1 genes derived from HIV-1 infected individuals from north India (Delhi, Punjab and Chandigarh). We found evidence of novel B/C recombinants in HIV-1 LTR region showing relatedness to China/Myanmar with 3 copies of Nfκb sites; B/C/D mosaic genomes for HIV-1 Vpr and novel B/C Tat. We reported appearance of a complex recombinant form CRF_02AG of HIV-1 envelope sequences which is predominantly found in Central/Western Africa. Also one Indian HIV-1 envelope subtype C sequence suggested exclusive CXCR4 co-receptor usage. This extensive recombination, which is observed in about 10 per cent HIV-1 infected individuals in the Vpr genes, resulted in remarkably altered functions when compared with prototype subtype B Vpr. The Vpu C was found to be more potent in causing apoptosis when compared with Vpu B when analyzed for subG1 DNA content. The functional implications of these changes as well as in other genes of HIV-1 are discussed in detail with possible implications for subtype-specific pathogenesis highlighted.

  16. The HIV-1 Rev/RRE system is required for HIV-1 5' UTR cis elements to augment encapsidation of heterologous RNA into HIV-1 viral particles

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    Ma Hong

    2011-06-01

    Full Text Available Abstract Background The process of HIV-1 genomic RNA (gRNA encapsidation is governed by a number of viral encoded components, most notably the Gag protein and gRNA cis elements in the canonical packaging signal (ψ. Also implicated in encapsidation are cis determinants in the R, U5, and PBS (primer binding site from the 5' untranslated region (UTR. Although conventionally associated with nuclear export of HIV-1 RNA, there is a burgeoning role for the Rev/RRE in the encapsidation process. Pleiotropic effects exhibited by these cis and trans viral components may confound the ability to examine their independent, and combined, impact on encapsidation of RNA into HIV-1 viral particles in their innate viral context. We systematically reconstructed the HIV-1 packaging system in the context of a heterologous murine leukemia virus (MLV vector RNA to elucidate a mechanism in which the Rev/RRE system is central to achieving efficient and specific encapsidation into HIV-1 viral particles. Results We show for the first time that the Rev/RRE system can augment RNA encapsidation independent of all cis elements from the 5' UTR (R, U5, PBS, and ψ. Incorporation of all the 5' UTR cis elements did not enhance RNA encapsidation in the absence of the Rev/RRE system. In fact, we demonstrate that the Rev/RRE system is required for specific and efficient encapsidation commonly associated with the canonical packaging signal. The mechanism of Rev/RRE-mediated encapsidation is not a general phenomenon, since the combination of the Rev/RRE system and 5' UTR cis elements did not enhance encapsidation into MLV-derived viral particles. Lastly, we show that heterologous MLV RNAs conform to transduction properties commonly associated with HIV-1 viral particles, including in vivo transduction of non-dividing cells (i.e. mouse neurons; however, the cDNA forms are episomes predominantly in the 1-LTR circle form. Conclusions Premised on encapsidation of a heterologous RNA into

  17. Measuring replication competent HIV-1: advances and challenges in defining the latent reservoir

    OpenAIRE

    Wang, Zheng; Simonetti, Francesco R.; Siliciano, Robert F.; Laird, Gregory M.

    2018-01-01

    Antiretroviral therapy cannot cure HIV-1 infection due to the persistence of a small number of latently infected cells harboring replication-competent proviruses. Measuring persistent HIV-1 is challenging, as it consists of a mosaic population of defective and intact proviruses that can shift from a state of latency to active HIV-1 transcription. Due to this complexity, most of the current assays detect multiple categories of persistent HIV-1, leading to an overestimate of the true size of th...

  18. Stable assembly of HIV-1 export complexes occurs cotranscriptionally

    DEFF Research Database (Denmark)

    Nawroth, Isabel; Mueller, Florian; Basyuk, Eugenia

    2014-01-01

    The HIV-1 Rev protein mediates export of unspliced and singly spliced viral transcripts by binding to the Rev response element (RRE) and recruiting the cellular export factor CRM1. Here, we investigated the recruitment of Rev to the transcription sites of HIV-1 reporters that splice either post......- or cotranscriptionally. In both cases, we observed that Rev localized to the transcription sites of the reporters and recruited CRM1. Rev and CRM1 remained at the reporter transcription sites when cells were treated with the splicing inhibitor Spliceostatin A (SSA), showing that the proteins associate with RNA prior...... to or during early spliceosome assembly. Fluorescence recovery after photobleaching (FRAP) revealed that Rev and CRM1 have similar kinetics as the HIV-1 RNA, indicating that Rev, CRM1, and RRE-containing RNAs are released from the site of transcription in one single export complex. These results suggest...

  19. The Depsipeptide Romidepsin Reverses HIV-1 Latency In Vivo.

    Directory of Open Access Journals (Sweden)

    Ole S Søgaard

    2015-09-01

    Full Text Available Pharmacologically-induced activation of replication competent proviruses from latency in the presence of antiretroviral treatment (ART has been proposed as a step towards curing HIV-1 infection. However, until now, approaches to reverse HIV-1 latency in humans have yielded mixed results. Here, we report a proof-of-concept phase Ib/IIa trial where 6 aviremic HIV-1 infected adults received intravenous 5 mg/m2 romidepsin (Celgene once weekly for 3 weeks while maintaining ART. Lymphocyte histone H3 acetylation, a cellular measure of the pharmacodynamic response to romidepsin, increased rapidly (maximum fold range: 3.7–7.7 relative to baseline within the first hours following each romidepsin administration. Concurrently, HIV-1 transcription quantified as copies of cell-associated un-spliced HIV-1 RNA increased significantly from baseline during treatment (range of fold-increase: 2.4–5.0; p = 0.03. Plasma HIV-1 RNA increased from <20 copies/mL at baseline to readily quantifiable levels at multiple post-infusion time-points in 5 of 6 patients (range 46–103 copies/mL following the second infusion, p = 0.04. Importantly, romidepsin did not decrease the number of HIV-specific T cells or inhibit T cell cytokine production. Adverse events (all grade 1–2 were consistent with the known side effects of romidepsin. In conclusion, romidepsin safely induced HIV-1 transcription resulting in plasma HIV-1 RNA that was readily detected with standard commercial assays demonstrating that significant reversal of HIV-1 latency in vivo is possible without blunting T cell-mediated immune responses. These finding have major implications for future trials aiming to eradicate the HIV-1 reservoir.clinicaltrials.gov NTC02092116.

  20. Excision of HIV-1 proviral DNA by recombinant cell permeable tre-recombinase.

    Directory of Open Access Journals (Sweden)

    Lakshmikanth Mariyanna

    Full Text Available Over the previous years, comprehensive studies on antiretroviral drugs resulted in the successful introduction of highly active antiretroviral therapy (HAART into clinical practice for treatment of HIV/AIDS. However, there is still need for new therapeutic approaches, since HAART cannot eradicate HIV-1 from the infected organism and, unfortunately, can be associated with long-term toxicity and the development of drug resistance. In contrast, novel gene therapy strategies may have the potential to reverse the infection by eradicating HIV-1. For example, expression of long terminal repeat (LTR-specific recombinase (Tre-recombinase has been shown to result in chromosomal excision of proviral DNA and, in consequence, in the eradication of HIV-1 from infected cell cultures. However, the delivery of Tre-recombinase currently depends on the genetic manipulation of target cells, a process that is complicating such therapeutic approaches and, thus, might be undesirable in a clinical setting. In this report we demonstrate that E.coli expressed Tre-recombinases, tagged either with the protein transduction domain (PTD from the HIV-1 Tat trans-activator or the translocation motif (TLM of the Hepatitis B virus PreS2 protein, were able to translocate efficiently into cells and showed significant recombination activity on HIV-1 LTR sequences. Tre activity was observed using episomal and stable integrated reporter constructs in transfected HeLa cells. Furthermore, the TLM-tagged enzyme was able to excise the full-length proviral DNA from chromosomal integration sites of HIV-1-infected HeLa and CEM-SS cells. The presented data confirm Tre-recombinase activity on integrated HIV-1 and provide the basis for the non-genetic transient application of engineered recombinases, which may be a valuable component of future HIV eradication strategies.

  1. Extracellular histones identified in crocodile blood inhibit in-vitro HIV-1 infection.

    Science.gov (United States)

    Kozlowski, Hannah N; Lai, Eric T L; Havugimana, Pierre C; White, Carl; Emili, Andrew; Sakac, Darinka; Binnington, Beth; Neschadim, Anton; McCarthy, Stephen D S; Branch, Donald R

    2016-08-24

    It has been reported that crocodile blood contains potent antibacterial and antiviral properties. However, its effects on HIV-1 infection remain unknown. We obtained blood from saltwater crocodiles to examine whether serum or plasma could inhibit HIV-1 infection. We purified plasma fractions then used liquid chromatography-mass spectrometry to identify the inhibitory protein factor(s). We then analyzed the ability of recombinant proteins to recapitulate HIV-1 inhibition and determine their mechanism of action. Crocodylus porosus plasma was tested for inhibition of Jurkat T-cell HIV-1 infection. Inhibitor(s) were purified by reverse-phase chromatography then identified by protein liquid chromatography-mass spectrometry. Anti-HIV-1 activity of purified plasma or recombinant proteins were measured by p24 enzyme-linked immunosorbent assay and luciferase readouts, and mechanism of action was determined by measuring HIV-1 RNA, cDNA and transcription (using 1G5 cells). Crocodile plasma contains potent inhibitors of HIV-1IIIB infection, which were identified as histones. Recombinant human histones H1 and H2A significantly reduced HIV-1JR-FL infection (IC50 of 0.79 and 0.45 μmol/l, respectively), whereas H4 enhanced JR-FL luciferase activity. The inhibitory effects of crocodile plasma, recombinant H1 or recombinant H2A on HIV-1 infection were during or post-viral transcription. Circulating histones in crocodile blood, possibly released by neutrophil extracellular traps, are significant inhibitors of HIV-1 infection in-vitro. Extracellular recombinant histones have different effects on HIV-1 transcription and protein expression and are downregulated in HIV-1 patients. Circulating histones may be a novel resistance factor during HIV-1 infection, and peptide versions should be explored as future HIV-1 therapeutics that modulate viral transcription.

  2. Integrated and Total HIV-1 DNA Predict Ex Vivo Viral Outgrowth.

    Directory of Open Access Journals (Sweden)

    Maja Kiselinova

    2016-03-01

    Full Text Available The persistence of a reservoir of latently infected CD4 T cells remains one of the major obstacles to cure HIV. Numerous strategies are being explored to eliminate this reservoir. To translate these efforts into clinical trials, there is a strong need for validated biomarkers that can monitor the reservoir over time in vivo. A comprehensive study was designed to evaluate and compare potential HIV-1 reservoir biomarkers. A cohort of 25 patients, treated with suppressive antiretroviral therapy was sampled at three time points, with median of 2.5 years (IQR: 2.4-2.6 between time point 1 and 2; and median of 31 days (IQR: 28-36 between time point 2 and 3. Patients were median of 6 years (IQR: 3-12 on ART, and plasma viral load (<50 copies/ml was suppressed for median of 4 years (IQR: 2-8. Total HIV-1 DNA, unspliced (us and multiply spliced HIV-1 RNA, and 2LTR circles were quantified by digital PCR in peripheral blood, at 3 time points. At the second time point, a viral outgrowth assay (VOA was performed, and integrated HIV-1 DNA and relative mRNA expression levels of HIV-1 restriction factors were quantified. No significant change was found for long- and short-term dynamics of all HIV-1 markers tested in peripheral blood. Integrated HIV-1 DNA was associated with total HIV-1 DNA (p<0.001, R² = 0.85, us HIV-1 RNA (p = 0.029, R² = 0.40, and VOA (p = 0.041, R2 = 0.44. Replication-competent virus was detected in 80% of patients by the VOA and it correlated with total HIV-1 DNA (p = 0.039, R² = 0.54. The mean quantification difference between Alu-PCR and VOA was 2.88 log10, and 2.23 log10 between total HIV-1 DNA and VOA. The levels of usHIV-1 RNA were inversely correlated with mRNA levels of several HIV-1 restriction factors (TRIM5α, SAMHD1, MX2, SLFN11, pSIP1. Our study reveals important correlations between the viral outgrowth and total and integrated HIV-1 DNA measures, suggesting that the total pool of HIV-1 DNA may predict the size of the

  3. Analysis of plant LTR-retrotransposons at the fine-scale family level reveals individual molecular patterns

    Directory of Open Access Journals (Sweden)

    Domingues Douglas S

    2012-04-01

    Full Text Available Abstract Background Sugarcane is an important crop worldwide for sugar production and increasingly, as a renewable energy source. Modern cultivars have polyploid, large complex genomes, with highly unequal contributions from ancestral genomes. Long Terminal Repeat retrotransposons (LTR-RTs are the single largest components of most plant genomes and can substantially impact the genome in many ways. It is therefore crucial to understand their contribution to the genome and transcriptome, however a detailed study of LTR-RTs in sugarcane has not been previously carried out. Results Sixty complete LTR-RT elements were classified into 35 families within four Copia and three Gypsy lineages. Structurally, within lineages elements were similar, between lineages there were large size differences. FISH analysis resulted in the expected pattern of Gypsy/heterochromatin, Copia/euchromatin, but in two lineages there was localized clustering on some chromosomes. Analysis of related ESTs and RT-PCR showed transcriptional variation between tissues and families. Four distinct patterns were observed in sRNA mapping, the most unusual of which was that of Ale1, with very large numbers of 24nt sRNAs in the coding region. The results presented support the conclusion that distinct small RNA-regulated pathways in sugarcane target the lineages of LTR-RT elements. Conclusions Individual LTR-RT sugarcane families have distinct structures, and transcriptional and regulatory signatures. Our results indicate that in sugarcane individual LTR-RT families have distinct behaviors and can potentially impact the genome in diverse ways. For instance, these transposable elements may affect nearby genes by generating a diverse set of small RNA's that trigger gene silencing mechanisms. There is also some evidence that ancestral genomes contribute significantly different element numbers from particular LTR-RT lineages to the modern sugarcane cultivar genome.

  4. Potent inhibition of HIV-1 replication by a Tat mutant.

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    Luke W Meredith

    Full Text Available Herein we describe a mutant of the two-exon HIV-1 Tat protein, termed Nullbasic, that potently inhibits multiple steps of the HIV-1 replication cycle. Nullbasic was created by replacing the entire arginine-rich basic domain of wild type Tat with glycine/alanine residues. Like similarly mutated one-exon Tat mutants, Nullbasic exhibited transdominant negative effects on Tat-dependent transactivation. However, unlike previously reported mutants, we discovered that Nullbasic also strongly suppressed the expression of unspliced and singly-spliced viral mRNA, an activity likely caused by redistribution and thus functional inhibition of HIV-1 Rev. Furthermore, HIV-1 virion particles produced by cells expressing Nullbasic had severely reduced infectivity, a defect attributable to a reduced ability of the virions to undergo reverse transcription. Combination of these inhibitory effects on transactivation, Rev-dependent mRNA transport and reverse transcription meant that permissive cells constitutively expressing Nullbasic were highly resistant to a spreading infection by HIV-1. Nullbasic and its activities thus provide potential insights into the development of potent antiviral therapeutics that target multiple stages of HIV-1 infection.

  5. Anti-HIV-1 activity of anionic polymers: a comparative study of candidate microbicides

    Directory of Open Access Journals (Sweden)

    Li Yun-Yao

    2002-11-01

    Full Text Available Abstract Background Cellulose acetate phthalate (CAP in soluble form blocks coreceptor binding sites on the virus envelope glycoprotein gp120 and elicits gp41 six-helix bundle formation, processes involved in virus inactivation. CAP is not soluble at pH Methods Enzyme linked immunosorbent assays (ELISA were used to (1 study HIV-1 IIIB and BaL binding to micronized CAP; (2 detect virus disintegration; and (3 measure gp41 six-helix bundle formation. Cells containing integrated HIV-1 LTR linked to the β-gal gene and expressing CD4 and coreceptors CXCR4 or CCR5 were used to measure virus infectivity. Results 1 HIV-1 IIIB and BaL, respectively, effectively bound to micronized CAP. 2 The interaction between HIV-1 and micronized CAP led to: (a gp41 six-helix bundle formation; (b virus disintegration and shedding of envelope glycoproteins; and (c rapid loss of infectivity. Polymers other than CAP, except Carbomer 974P, elicited gp41 six-helix bundle formation in HIV-1 IIIB but only poly(napthalene sulfonate, in addition to CAP, had this effect on HIV-1 BaL. These polymers differed with respect to their virucidal activities, the differences being more pronounced for HIV-1 BaL. Conclusions Micronized CAP is the only candidate topical microbicide with the capacity to remove rapidly by adsorption from physiological fluids HIV-1 of both the X4 and R5 biotypes and is likely to prevent virus contact with target cells. The interaction between micronized CAP and HIV-1 leads to rapid virus inactivation. Among other anionic polymers, cellulose sulfate, BufferGel and aryl sulfonates appear most effective in this respect.

  6. Measuring replication competent HIV-1: advances and challenges in defining the latent reservoir.

    Science.gov (United States)

    Wang, Zheng; Simonetti, Francesco R; Siliciano, Robert F; Laird, Gregory M

    2018-02-13

    Antiretroviral therapy cannot cure HIV-1 infection due to the persistence of a small number of latently infected cells harboring replication-competent proviruses. Measuring persistent HIV-1 is challenging, as it consists of a mosaic population of defective and intact proviruses that can shift from a state of latency to active HIV-1 transcription. Due to this complexity, most of the current assays detect multiple categories of persistent HIV-1, leading to an overestimate of the true size of the latent reservoir. Here, we review the development of the viral outgrowth assay, the gold-standard quantification of replication-competent proviruses, and discuss the insights provided by full-length HIV-1 genome sequencing methods, which allowed us to unravel the composition of the proviral landscape. In this review, we provide a dissection of what defines HIV-1 persistence and we examine the unmet needs to measure the efficacy of interventions aimed at eliminating the HIV-1 reservoir.

  7. Cellular and molecular mechanisms of HIV-1 integration targeting.

    Science.gov (United States)

    Engelman, Alan N; Singh, Parmit K

    2018-07-01

    Integration is central to HIV-1 replication and helps mold the reservoir of cells that persists in AIDS patients. HIV-1 interacts with specific cellular factors to target integration to interior regions of transcriptionally active genes within gene-dense regions of chromatin. The viral capsid interacts with several proteins that are additionally implicated in virus nuclear import, including cleavage and polyadenylation specificity factor 6, to suppress integration into heterochromatin. The viral integrase protein interacts with transcriptional co-activator lens epithelium-derived growth factor p75 to principally position integration within gene bodies. The integrase additionally senses target DNA distortion and nucleotide sequence to help fine-tune the specific phosphodiester bonds that are cleaved at integration sites. Research into virus-host interactions that underlie HIV-1 integration targeting has aided the development of a novel class of integrase inhibitors and may help to improve the safety of viral-based gene therapy vectors.

  8. HIV-1 nef suppression by virally encoded microRNA

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    Brisibe Ebiamadon

    2004-12-01

    Full Text Available Abstract Background MicroRNAs (miRNAs are 21~25-nucleotides (nt long and interact with mRNAs to trigger either translational repression or RNA cleavage through RNA interference (RNAi, depending on the degree of complementarity with the target mRNAs. Our recent study has shown that HIV-1 nef dsRNA from AIDS patients who are long-term non-progressors (LTNPs inhibited the transcription of HIV-1. Results Here, we show the possibility that nef-derived miRNAs are produced in HIV-1 persistently infected cells. Furthermore, nef short hairpin RNA (shRNA that corresponded to a predicted nef miRNA (~25 nt, miR-N367 can block HIV-1 Nef expression in vitro and the suppression by shRNA/miR-N367 would be related with low viremia in an LTNP (15-2-2. In the 15-2-2 model mice, the weight loss, which may be rendered by nef was also inhibited by shRNA/miR-N367 corresponding to suppression of nef expression in vivo. Conclusions These data suggest that nef/U3 miRNAs produced in HIV-1-infected cells may suppress both Nef function and HIV-1 virulence through the RNAi pathway.

  9. DBR1 siRNA inhibition of HIV-1 replication

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    Naidu Yathi

    2005-10-01

    Full Text Available Abstract Background HIV-1 and all retroviruses are related to retroelements of simpler organisms such as the yeast Ty elements. Recent work has suggested that the yeast retroelement Ty1 replicates via an unexpected RNA lariat intermediate in cDNA synthesis. The putative genomic RNA lariat intermediate is formed by a 2'-5' phosphodiester bond, like that found in pre-mRNA intron lariats and it facilitates the minus-strand template switch during cDNA synthesis. We hypothesized that HIV-1 might also form a genomic RNA lariat and therefore that siRNA-mediated inhibition of expression of the human RNA lariat de-branching enzyme (DBR1 expression would specifically inhibit HIV-1 replication. Results We designed three short interfering RNA (siRNA molecules targeting DBR1, which were capable of reducing DBR1 mRNA expression by 80% and did not significantly affect cell viability. We assessed HIV-1 replication in the presence of DBR1 siRNA and found that DBR1 knockdown led to decreases in viral cDNA and protein production. These effects could be reversed by cotransfection of a DBR1 cDNA indicating that the inhibition of HIV-1 replication was a specific effect of DBR1 underexpression. Conclusion These data suggest that DBR1 function may be needed to debranch a putative HIV-1 genomic RNA lariat prior to completion of reverse transcription.

  10. Enrichment of intersubtype HIV-1 recombinants in a dual infection system using HIV-1 strain-specific siRNAs

    Science.gov (United States)

    2011-01-01

    Background Intersubtype HIV-1 recombinants in the form of unique or stable circulating recombinants forms (CRFs) are responsible for over 20% of infections in the worldwide epidemic. Mechanisms controlling the generation, selection, and transmission of these intersubtype HIV-1 recombinants still require further investigation. All intersubtype HIV-1 recombinants are generated and evolve from initial dual infections, but are difficult to identify in the human population. In vitro studies provide the most practical system to study mechanisms, but the recombination rates are usually very low in dual infections with primary HIV-1 isolates. This study describes the use of HIV-1 isolate-specific siRNAs to enrich intersubtype HIV-1 recombinants and inhibit the parental HIV-1 isolates from a dual infection. Results Following a dual infection with subtype A and D primary HIV-1 isolates and two rounds of siRNA treatment, nearly 100% of replicative virus was resistant to a siRNA specific for an upstream target sequence in the subtype A envelope (env) gene as well as a siRNA specific for a downstream target sequence in the subtype D env gene. Only 20% (10/50) of the replicating virus had nucleotide substitutions in the siRNA-target sequence whereas the remaining 78% (39/50) harbored a recombination breakpoint that removed both siRNA target sequences, and rendered the intersubtype D/A recombinant virus resistant to the dual siRNA treatment. Since siRNAs target the newly transcribed HIV-1 mRNA, the siRNAs only enrich intersubtype env recombinants and do not influence the recombination process during reverse transcription. Using this system, a strong bias is selected for recombination breakpoints in the C2 region, whereas other HIV-1 env regions, most notably the hypervariable regions, were nearly devoid of intersubtype recombination breakpoints. Sequence conservation plays an important role in selecting for recombination breakpoints, but the lack of breakpoints in many conserved

  11. Differentially-Expressed Pseudogenes in HIV-1 Infection

    Directory of Open Access Journals (Sweden)

    Aditi Gupta

    2015-09-01

    Full Text Available Not all pseudogenes are transcriptionally silent as previously thought. Pseudogene transcripts, although not translated, contribute to the non-coding RNA pool of the cell that regulates the expression of other genes. Pseudogene transcripts can also directly compete with the parent gene transcripts for mRNA stability and other cell factors, modulating their expression levels. Tissue-specific and cancer-specific differential expression of these “functional” pseudogenes has been reported. To ascertain potential pseudogene:gene interactions in HIV-1 infection, we analyzed transcriptomes from infected and uninfected T-cells and found that 21 pseudogenes are differentially expressed in HIV-1 infection. This is interesting because parent genes of one-third of these differentially-expressed pseudogenes are implicated in HIV-1 life cycle, and parent genes of half of these pseudogenes are involved in different viral infections. Our bioinformatics analysis identifies candidate pseudogene:gene interactions that may be of significance in HIV-1 infection. Experimental validation of these interactions would establish that retroviruses exploit this newly-discovered layer of host gene expression regulation for their own benefit.

  12. Differentially-Expressed Pseudogenes in HIV-1 Infection.

    Science.gov (United States)

    Gupta, Aditi; Brown, C Titus; Zheng, Yong-Hui; Adami, Christoph

    2015-09-29

    Not all pseudogenes are transcriptionally silent as previously thought. Pseudogene transcripts, although not translated, contribute to the non-coding RNA pool of the cell that regulates the expression of other genes. Pseudogene transcripts can also directly compete with the parent gene transcripts for mRNA stability and other cell factors, modulating their expression levels. Tissue-specific and cancer-specific differential expression of these "functional" pseudogenes has been reported. To ascertain potential pseudogene:gene interactions in HIV-1 infection, we analyzed transcriptomes from infected and uninfected T-cells and found that 21 pseudogenes are differentially expressed in HIV-1 infection. This is interesting because parent genes of one-third of these differentially-expressed pseudogenes are implicated in HIV-1 life cycle, and parent genes of half of these pseudogenes are involved in different viral infections. Our bioinformatics analysis identifies candidate pseudogene:gene interactions that may be of significance in HIV-1 infection. Experimental validation of these interactions would establish that retroviruses exploit this newly-discovered layer of host gene expression regulation for their own benefit.

  13. Chemical Composition of Essential Oils from Thymus vulgaris, Cymbopogon citratus, and Rosmarinus officinalis, and Their Effects on the HIV-1 Tat Protein Function.

    Science.gov (United States)

    Feriotto, Giordana; Marchetti, Nicola; Costa, Valentina; Beninati, Simone; Tagliati, Federico; Mischiati, Carlo

    2018-02-01

    New drugs would be beneficial to fight resistant HIV strains, in particular those capable of interfering with essential viral functions other than those targeted by highly active antiretroviral therapy drugs. Despite the central role played by Tat protein in HIV transcription, a search for vegetable extracts able to hamper this important viral function was never carried out. In this work, we evaluated the chemical composition and possible interference of essential oil from Thymus vulgaris, Cananga odorata, Cymbopogon citratus, and Rosmarinus officinalis with the Tat/TAR-RNA interaction and with Tat-induced HIV-1 LTR transcription. GC/MS Analysis demonstrated the biodiversity of herbal species translated into essential oils composed of different blends of terpenes. In all of them, 4 - 6 constituents represent from 81.63% to 95.19% of the total terpenes. Essential oils of Thymus vulgaris, Cymbopogon citratus, and Rosmarinus officinalis were active in interfering with Tat functions, encouraging further studies to identify single terpenes responsible for the antiviral activity. In view of the quite different composition of these essential oils, we concluded that their interference on Tat function depends on specific terpene or a characteristic blend. © 2018 Wiley-VHCA AG, Zurich, Switzerland.

  14. Hairpin-induced tRNA-mediated (HITME) recombination in HIV-1

    NARCIS (Netherlands)

    Konstantinova, Pavlina; de Haan, Peter; Das, Atze T.; Berkhout, Ben

    2006-01-01

    Recombination due to template switching during reverse transcription is a major source of genetic variability in retroviruses. In the present study we forced a recombination event in human immunodeficiency virus type 1 (HIV-1) by electroporation of T cells with DNA from a molecular HIV-1 clone that

  15. Zinc finger nuclease: a new approach for excising HIV-1 proviral DNA from infected human T cells.

    Science.gov (United States)

    Qu, Xiying; Wang, Pengfei; Ding, Donglin; Wang, Xiaohui; Zhang, Gongmin; Zhou, Xin; Liu, Lin; Zhu, Xiaoli; Zeng, Hanxian; Zhu, Huanzhang

    2014-09-01

    A major reason that Acquired Immune Deficiency Syndrome (AIDS) cannot be completely cured is the human immunodeficiency virus 1 (HIV-1) provirus integrated into the human genome. Though existing therapies can inhibit replication of HIV-1, they cannot eradicate it. A molecular therapy gains popularity due to its specifically targeting to HIV-1 infected cells and effectively removing the HIV-1, regardless of viral genes being active or dormant. Now, we propose a new method which can excellently delete the HIV provirus from the infected human T cell genome. First, we designed zinc-finger nucleases (ZFNs) that target a sequence within the long terminal repeat (LTR) U3 region that is highly conserved in whole clade. Then, we screened out one pair of ZFN and named it as ZFN-U3. We discovered that ZFN-U3 can exactly target and eliminate the full-length HIV-1 proviral DNA after the infected human cell lines treated with it, and the frequency of its excision was about 30 % without cytotoxicity. These results prove that ZFN-U3 can efficiently excise integrated HIV-1 from the human genome in infected cells. This method to delete full length HIV-1 in human genome can therefore provide a novel approach to cure HIV-infected individuals in the future.

  16. HIV-1 RNAs are Not Part of the Argonaute 2 Associated RNA Interference Pathway in Macrophages.

    Directory of Open Access Journals (Sweden)

    Valentina Vongrad

    Full Text Available MiRNAs and other small noncoding RNAs (sncRNAs are key players in post-transcriptional gene regulation. HIV-1 derived small noncoding RNAs (sncRNAs have been described in HIV-1 infected cells, but their biological functions still remain to be elucidated. Here, we approached the question whether viral sncRNAs may play a role in the RNA interference (RNAi pathway or whether viral mRNAs are targeted by cellular miRNAs in human monocyte derived macrophages (MDM.The incorporation of viral sncRNAs and/or their target RNAs into RNA-induced silencing complex was investigated using photoactivatable ribonucleoside-induced cross-linking and immunoprecipitation (PAR-CLIP as well as high-throughput sequencing of RNA isolated by cross-linking immunoprecipitation (HITS-CLIP, which capture Argonaute2-bound miRNAs and their target RNAs. HIV-1 infected monocyte-derived macrophages (MDM were chosen as target cells, as they have previously been shown to express HIV-1 sncRNAs. In addition, we applied small RNA deep sequencing to study differential cellular miRNA expression in HIV-1 infected versus non-infected MDMs.PAR-CLIP and HITS-CLIP data demonstrated the absence of HIV-1 RNAs in Ago2-RISC, although the presence of a multitude of HIV-1 sncRNAs in HIV-1 infected MDMs was confirmed by small RNA sequencing. Small RNA sequencing revealed that 1.4% of all sncRNAs were of HIV-1 origin. However, neither HIV-1 derived sncRNAs nor putative HIV-1 target sequences incorporated into Ago2-RISC were identified suggesting that HIV-1 sncRNAs are not involved in the canonical RNAi pathway nor is HIV-1 targeted by this pathway in HIV-1 infected macrophages.

  17. HIV-1 and the macrophage

    NARCIS (Netherlands)

    Bol, Sebastiaan M.; Cobos-Jimenez, Viviana; Kootstra, Neeltje A.; van 't Wout, Angelique B.

    2011-01-01

    Macrophages and CD4(+) T cells are natural target cells for HIV-1, and both cell types contribute to the establishment of the viral reservoir that is responsible for continuous residual virus replication during antiretroviral therapy and viral load rebound upon treatment interruption. Scientific

  18. Developing strategies for HIV-1 eradication

    Science.gov (United States)

    Durand, Christine M.; Blankson, Joel N.; Siliciano, Robert F.

    2014-01-01

    Highly active antiretroviral therapy (HAART) suppresses HIV-1 replication, transforming the outlook for infected patients. However, reservoirs of replication-competent forms of the virus persist during HAART, and when treatment is stopped, high rates of HIV-1 replication return. Recent insights into HIV-1 latency, as well as a report that HIV-1 infection was eradicated in one individual, have renewed interest in finding a cure for HIV-1 infection. Strategies for HIV-1 eradication include gene therapy and hematopoietic stem cell transplantation, stimulating host immunity to control HIV-1 replication, and targeting latent HIV-1 in resting memory CD4+ T cells. Future efforts should aim to provide better understanding of how to reconstitute the CD4+ T cell compartment with genetically engineered cells, exert immune control over HIV-1 replication, and identify and eliminate all viral reservoirs. PMID:22867874

  19. LTR-retrotransposons-based molecular markers in cultivated ...

    African Journals Online (AJOL)

    GRACE

    2006-07-03

    Jul 3, 2006 ... LTR-retrotransposons represent a standard component of the Gossypium Genome (Zaki and Abdel Ghany,. 2003). The analysis of the molecular existence and distribution of ancient and active LTR-retrotransposons, therefore, provides a comprehensive evaluation of the evolutionary history of Gossypium.

  20. Identifying HIV-1 dual infections

    Directory of Open Access Journals (Sweden)

    Cornelissen Marion

    2007-09-01

    Full Text Available Abstract Transmission of human immunodeficiency virus (HIV is no exception to the phenomenon that a second, productive infection with another strain of the same virus is feasible. Experiments with RNA viruses have suggested that both coinfections (simultaneous infection with two strains of a virus and superinfections (second infection after a specific immune response to the first infecting strain has developed can result in increased fitness of the viral population. Concerns about dual infections with HIV are increasing. First, the frequent detection of superinfections seems to indicate that it will be difficult to develop a prophylactic vaccine. Second, HIV-1 superinfections have been associated with accelerated disease progression, although this is not true for all persons. In fact, superinfections have even been detected in persons controlling their HIV infections without antiretroviral therapy. Third, dual infections can give rise to recombinant viruses, which are increasingly found in the HIV-1 epidemic. Recombinants could have increased fitness over the parental strains, as in vitro models suggest, and could exhibit increased pathogenicity. Multiple drug resistant (MDR strains could recombine to produce a pan-resistant, transmittable virus. We will describe in this review what is presently known about super- and re-infection among ambient viral infections, as well as the first cases of HIV-1 superinfection, including HIV-1 triple infections. The clinical implications, the impact of the immune system, and the effect of anti-retroviral therapy will be covered, as will as the timing of HIV superinfection. The methods used to detect HIV-1 dual infections will be discussed in detail. To increase the likelihood of detecting a dual HIV-1 infection, pre-selection of patients can be done by serotyping, heteroduplex mobility assays (HMA, counting the degenerate base codes in the HIV-1 genotyping sequence, or surveying unexpected increases in the

  1. A novel function for spumaretrovirus integrase: an early requirement for integrase-mediated cleavage of 2 LTR circles

    Directory of Open Access Journals (Sweden)

    Mouscadet Jean-François

    2005-05-01

    Full Text Available Abstract Retroviral integration is central to viral persistence and pathogenesis, cancer as well as host genome evolution. However, it is unclear why integration appears essential for retrovirus production, especially given the abundance and transcriptional potential of non-integrated viral genomes. The involvement of retroviral endonuclease, also called integrase (IN, in replication steps apart from integration has been proposed, but is usually considered to be accessory. We observe here that integration of a retrovirus from the spumavirus family depends mainly on the quantity of viral DNA produced. Moreover, we found that IN directly participates to linear DNA production from 2-LTR circles by specifically cleaving the conserved palindromic sequence found at LTR-LTR junctions. These results challenge the prevailing view that integrase essential function is to catalyze retroviral DNA integration. Integrase activity upstream of this step, by controlling linear DNA production, is sufficient to explain the absolute requirement for this enzyme. The novel role of IN over 2-LTR circle junctions accounts for the pleiotropic effects observed in cells infected with IN mutants. It may explain why 1 2-LTR circles accumulate in vivo in mutants carrying a defective IN while their linear and integrated DNA pools decrease; 2 why both LTRs are processed in a concerted manner. It also resolves the original puzzle concerning the integration of spumaretroviruses. More generally, it suggests to reassess 2-LTR circles as functional intermediates in the retrovirus cycle and to reconsider the idea that formation of the integrated provirus is an essential step of retrovirus production.

  2. Curcumin derivatives as HIV-1 protease inhibitors

    Energy Technology Data Exchange (ETDEWEB)

    Sui, Z.; Li, J.; Craik, C.S.; Ortiz de Montellano, P.R. [Univ. of California, San Francisco, CA (United States)

    1993-12-31

    Curcumin, a non-toxic natural compound from Curcuma longa, has been found to be an HIV-1 protease inhibitor. Some of its derivatives were synthesized and their inhibitory activity against the HIV-1 protease was tested. Curcumin analogues containing boron enhanced the inhibitory activity. At least of the the synthesized compounds irreversibly inhibits the HIV-1 protease.

  3. An application of LTR design in fault detection

    DEFF Research Database (Denmark)

    Niemann, Hans Henrik

    1998-01-01

    The fault detection and isolation (FDI) problem is considered in this paper. The FDI problem is formulated as a filter design problem, where the faults in the system is estimated and the disturbance acting on the system is rejected. It turns out that the filter design problem can be considered...... as a standard Loop Transfer Recovery (LTR) design problem. As a consequence of the connection between LTR and FDI design, it is shown in an example how the LQG/LTR design method for full order and a proportional-integral observer can be applied with advantages in connection with FDI....

  4. A parametric LTR solution for discrete-time systems

    DEFF Research Database (Denmark)

    Niemann, Hans Henrik; Jannerup, Ole Erik

    1989-01-01

    A parametric LTR (loop transfer recovery) solution for discrete-time compensators incorporating filtering observers which achieve exact recovery is presented for both minimum- and non-minimum-phase systems. First the recovery error, which defines the difference between the target loop transfer...... and the full loop transfer function, is manipulated into a general form involving the target loop transfer matrix and the fundamental recovery matrix. A parametric LTR solution based on the recovery matrix is developed. It is shown that the LQR/LTR (linear quadratic Gaussian/loop transfer recovery) solution...

  5. Epigenetic regulation of HIV-1 latency: focus on polycomb group (PcG) proteins.

    Science.gov (United States)

    Khan, Sheraz; Iqbal, Mazhar; Tariq, Muhammad; Baig, Shahid M; Abbas, Wasim

    2018-01-01

    HIV-1 latency allows the virus to persist until reactivation, in a transcriptionally silent form in its cellular reservoirs despite the presence of effective cART. Such viral persistence represents a major barrier to HIV eradication since treatment interruption leads to rebound plasma viremia. Polycomb group (PcG) proteins have recently got a considerable attention in regulating HIV-1 post-integration latency as they are involved in the repression of proviral gene expression through the methylation of histones. This epigenetic regulation plays an important role in the establishment and maintenance of HIV-1 latency. In fact, PcG proteins act in complexes and modulate the epigenetic signatures of integrated HIV-1 promoter. Key role played by PcG proteins in the molecular control of HIV-1 latency has led to hypothesize that PcG proteins may represent a valuable target for future HIV-1 therapy in purging HIV-1 reservoirs. In this regard, various small molecules have been synthesized or explored to specifically block the epigenetic activity of PcG. In this review, we will highlight the possible therapeutic approaches to achieve either a functional or sterilizing cure of HIV-1 infection with special focus on histone methylation by PcG proteins together with current and novel pharmacological approaches to reactivate HIV-1 from latency that could ultimately lead towards a better clearance of viral latent reservoirs.

  6. A European multicientre study on the comparison of HIV-1 viral loads between VERIS HIV-1 Assay and Roche COBAS® TAQMAN® HIV-1 test, Abbott RealTime HIV-1 Assay, and Siemens VERSANT HIV-1 Assay.

    Science.gov (United States)

    Braun, Patrick; Delgado, Rafael; Drago, Monica; Fanti, Diana; Fleury, Hervé; Hofmann, Jörg; Izopet, Jacques; Kühn, Sebastian; Lombardi, Alessandra; Mancon, Alessandro; Marcos, Mª Angeles; Mileto, Davide; Sauné, Karine; O'Shea, Siobhan; Pérez-Rivilla, Alfredo; Ramble, John; Trimoulet, Pascale; Vila, Jordi; Whittaker, Duncan; Artus, Alain; Rhodes, Daniel

    2017-07-01

    Viral load monitoring is essential for patients under treatment for HIV. Beckman Coulter has developed the VERIS HIV-1 Assay for use on the novel, automated DxN VERIS Molecular Diagnostics System. ¥ OBJECTIVES: Evaluation of the clinical performance of the new quantitative VERIS HIV-1 Assay at multiple EU laboratories. Method comparison with the VERIS HIV-1 Assay was performed with 415 specimens at 5 sites tested with COBAS ® AmpliPrep/COBAS ® TaqMan ® HIV-1 Test, v2.0, 169 specimens at 3 sites tested with RealTime HIV-1 Assay, and 202 specimens from 2 sites tested with VERSANT HIV-1 Assay. Patient monitoring sample results from 4 sites were also compared. Bland-Altman analysis showed the average bias between VERIS HIV-1 Assay and COBAS HIV-1 Test, RealTime HIV-1 Assay, and VERSANT HIV-1 Assay to be 0.28, 0.39, and 0.61 log 10 cp/mL, respectively. Bias at low end levels below 1000cp/mL showed predicted bias to be <0.3 log 10 cp/mL for VERIS HIV-1 Assay versus COBAS HIV-1 Test and RealTime HIV-1 Assay, and <0.5 log 10 cp/mL versus VERSANT HIV-1 Assay. Analysis on 174 specimens tested with the 0.175mL volume VERIS HIV-1 Assay and COBAS HIV-1 Test showed average bias of 0.39 log 10 cp/mL. Patient monitoring results using VERIS HIV-1 Assay demonstrated similar viral load trends over time to all comparators. The VERIS HIV-1 Assay for use on the DxN VERIS System demonstrated comparable clinical performance to COBAS ® HIV-1 Test, RealTime HIV-1 Assay, and VERSANT HIV-1 Assay. Copyright © 2017 Elsevier B.V. All rights reserved.

  7. A Theory of LTR Junk-food Consumption

    OpenAIRE

    Levy, Amnon

    2003-01-01

    LTR junk-food consumption balances the marginal satisfaction with the marginal deterioration of health. An LTR person discounts the instantaneous marginal satisfaction from junk-food consumption by its implications for his survival probability. His change rate of health evaluation is increased (decreased) by junk-food consumption when health is better (worse) than a critical level. The moderating direct effects of age and relative price on junk-food consumption may be amplified, or dimmed, by...

  8. Convergent evolution of ribonuclease h in LTR retrotransposons and retroviruses.

    Science.gov (United States)

    Ustyantsev, Kirill; Novikova, Olga; Blinov, Alexander; Smyshlyaev, Georgy

    2015-05-01

    Ty3/Gypsy long terminals repeat (LTR) retrotransposons are structurally and phylogenetically close to retroviruses. Two notable structural differences between these groups of genetic elements are 1) the presence in retroviruses of an additional envelope gene, env, which mediates infection, and 2) a specific dual ribonuclease H (RNH) domain encoded by the retroviral pol gene. However, similar to retroviruses, many Ty3/Gypsy LTR retrotransposons harbor additional env-like genes, promoting concepts of the infective mode of these retrotransposons. Here, we provide a further line of evidence of similarity between retroviruses and some Ty3/Gypsy LTR retrotransposons. We identify that, together with their additional genes, plant Ty3/Gypsy LTR retrotransposons of the Tat group have a second RNH, as do retroviruses. Most importantly, we show that the resulting dual RNHs of Tat LTR retrotransposons and retroviruses emerged independently, providing strong evidence for their convergent evolution. The convergent resemblance of Tat LTR retrotransposons and retroviruses may indicate similar selection pressures acting on these diverse groups of elements and reveal potential evolutionary constraints on their structure. We speculate that dual RNH is required to accelerate retrotransposon evolution through increased rates of strand transfer events and subsequent recombination events. © The Author 2015. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

  9. Nucleolar protein trafficking in response to HIV-1 Tat: rewiring the nucleolus.

    Science.gov (United States)

    Jarboui, Mohamed Ali; Bidoia, Carlo; Woods, Elena; Roe, Barbara; Wynne, Kieran; Elia, Giuliano; Hall, William W; Gautier, Virginie W

    2012-01-01

    The trans-activator Tat protein is a viral regulatory protein essential for HIV-1 replication. Tat trafficks to the nucleoplasm and the nucleolus. The nucleolus, a highly dynamic and structured membrane-less sub-nuclear compartment, is the site of rRNA and ribosome biogenesis and is involved in numerous cellular functions including transcriptional regulation, cell cycle control and viral infection. Importantly, transient nucleolar trafficking of both Tat and HIV-1 viral transcripts are critical in HIV-1 replication, however, the role(s) of the nucleolus in HIV-1 replication remains unclear. To better understand how the interaction of Tat with the nucleolar machinery contributes to HIV-1 pathogenesis, we investigated the quantitative changes in the composition of the nucleolar proteome of Jurkat T-cells stably expressing HIV-1 Tat fused to a TAP tag. Using an organellar proteomic approach based on mass spectrometry, coupled with Stable Isotope Labelling in Cell culture (SILAC), we quantified 520 proteins, including 49 proteins showing significant changes in abundance in Jurkat T-cell nucleolus upon Tat expression. Numerous proteins exhibiting a fold change were well characterised Tat interactors and/or known to be critical for HIV-1 replication. This suggests that the spatial control and subcellular compartimentaliation of these cellular cofactors by Tat provide an additional layer of control for regulating cellular machinery involved in HIV-1 pathogenesis. Pathway analysis and network reconstruction revealed that Tat expression specifically resulted in the nucleolar enrichment of proteins collectively participating in ribosomal biogenesis, protein homeostasis, metabolic pathways including glycolytic, pentose phosphate, nucleotides and amino acids biosynthetic pathways, stress response, T-cell signaling pathways and genome integrity. We present here the first differential profiling of the nucleolar proteome of T-cells expressing HIV-1 Tat. We discuss how these

  10. APOBEC3G inhibits HIV-1 RNA elongation by inactivating the viral trans-activation response element.

    Science.gov (United States)

    Nowarski, Roni; Prabhu, Ponnandy; Kenig, Edan; Smith, Yoav; Britan-Rosich, Elena; Kotler, Moshe

    2014-07-29

    Deamination of cytidine residues in viral DNA is a major mechanism by which APOBEC3G (A3G) inhibits vif-deficient human immunodeficiency virus type 1 (HIV-1) replication. dC-to-dU transition following RNase-H activity leads to viral cDNA degradation, production of non-functional proteins, formation of undesired stop codons and decreased viral protein synthesis. Here, we demonstrate that A3G provides an additional layer of defense against HIV-1 infection dependent on inhibition of proviral transcription. HIV-1 transcription elongation is regulated by the trans-activation response (TAR) element, a short stem-loop RNA structure required for elongation factors binding. Vif-deficient HIV-1-infected cells accumulate short viral transcripts and produce lower amounts of full-length HIV-1 transcripts due to A3G deamination of the TAR apical loop cytidine, highlighting the requirement for TAR loop integrity in HIV-1 transcription. We further show that free single-stranded DNA (ssDNA) termini are not essential for A3G activity and a gap of CCC motif blocked with juxtaposed DNA or RNA on either or 3'+5' ends is sufficient for A3G deamination. These results identify A3G as an efficient mutator and that deamination of (-)SSDNA results in an early block of HIV-1 transcription. Copyright © 2014 Elsevier Ltd. All rights reserved.

  11. Modulation of chromatin structure by the FACT histone chaperone complex regulates HIV-1 integration.

    Science.gov (United States)

    Matysiak, Julien; Lesbats, Paul; Mauro, Eric; Lapaillerie, Delphine; Dupuy, Jean-William; Lopez, Angelica P; Benleulmi, Mohamed Salah; Calmels, Christina; Andreola, Marie-Line; Ruff, Marc; Llano, Manuel; Delelis, Olivier; Lavigne, Marc; Parissi, Vincent

    2017-07-28

    Insertion of retroviral genome DNA occurs in the chromatin of the host cell. This step is modulated by chromatin structure as nucleosomes compaction was shown to prevent HIV-1 integration and chromatin remodeling has been reported to affect integration efficiency. LEDGF/p75-mediated targeting of the integration complex toward RNA polymerase II (polII) transcribed regions ensures optimal access to dynamic regions that are suitable for integration. Consequently, we have investigated the involvement of polII-associated factors in the regulation of HIV-1 integration. Using a pull down approach coupled with mass spectrometry, we have selected the FACT (FAcilitates Chromatin Transcription) complex as a new potential cofactor of HIV-1 integration. FACT is a histone chaperone complex associated with the polII transcription machinery and recently shown to bind LEDGF/p75. We report here that a tripartite complex can be formed between HIV-1 integrase, LEDGF/p75 and FACT in vitro and in cells. Biochemical analyzes show that FACT-dependent nucleosome disassembly promotes HIV-1 integration into chromatinized templates, and generates highly favored nucleosomal structures in vitro. This effect was found to be amplified by LEDGF/p75. Promotion of this FACT-mediated chromatin remodeling in cells both increases chromatin accessibility and stimulates HIV-1 infectivity and integration. Altogether, our data indicate that FACT regulates HIV-1 integration by inducing local nucleosomes dissociation that modulates the functional association between the incoming intasome and the targeted nucleosome.

  12. In vitro nuclear interactome of the HIV-1 Tat protein.

    LENUS (Irish Health Repository)

    Gautier, Virginie W

    2009-01-01

    BACKGROUND: One facet of the complexity underlying the biology of HIV-1 resides not only in its limited number of viral proteins, but in the extensive repertoire of cellular proteins they interact with and their higher-order assembly. HIV-1 encodes the regulatory protein Tat (86-101aa), which is essential for HIV-1 replication and primarily orchestrates HIV-1 provirus transcriptional regulation. Previous studies have demonstrated that Tat function is highly dependent on specific interactions with a range of cellular proteins. However they can only partially account for the intricate molecular mechanisms underlying the dynamics of proviral gene expression. To obtain a comprehensive nuclear interaction map of Tat in T-cells, we have designed a proteomic strategy based on affinity chromatography coupled with mass spectrometry. RESULTS: Our approach resulted in the identification of a total of 183 candidates as Tat nuclear partners, 90% of which have not been previously characterised. Subsequently we applied in silico analysis, to validate and characterise our dataset which revealed that the Tat nuclear interactome exhibits unique signature(s). First, motif composition analysis highlighted that our dataset is enriched for domains mediating protein, RNA and DNA interactions, and helicase and ATPase activities. Secondly, functional classification and network reconstruction clearly depicted Tat as a polyvalent protein adaptor and positioned Tat at the nexus of a densely interconnected interaction network involved in a range of biological processes which included gene expression regulation, RNA biogenesis, chromatin structure, chromosome organisation, DNA replication and nuclear architecture. CONCLUSION: We have completed the in vitro Tat nuclear interactome and have highlighted its modular network properties and particularly those involved in the coordination of gene expression by Tat. Ultimately, the highly specialised set of molecular interactions identified will

  13. Inhibition of HIV Replication by Cyclic and Hairpin PNAs Targeting the HIV-1 TAR RNA Loop

    Science.gov (United States)

    Upert, Gregory; Di Giorgio, Audrey; Upadhyay, Alok; Manvar, Dinesh; Pandey, Nootan; Pandey, Virendra N.; Patino, Nadia

    2012-01-01

    Human immunodeficiency virus-1 (HIV-1) replication and gene expression entails specific interaction of the viral protein Tat with its transactivation responsive element (TAR), to form a highly stable stem-bulge-loop structure. Previously, we described triphenylphosphonium (TPP) cation-based vectors that efficiently deliver nucleotide analogs (PNAs) into the cytoplasm of cells. In particular, we showed that the TPP conjugate of a linear 16-mer PNA targeting the apical stem-loop region of TAR impedes Tat-mediated transactivation of the HIV-1 LTR in vitro and also in cell culture systems. In this communication, we conjugated TPP to cyclic and hairpin PNAs targeting the loop region of HIV-1 TAR and evaluated their antiviral efficacy in a cell culture system. We found that TPP-cyclic PNAs containing only 8 residues, showed higher antiviral potency compared to hairpin PNAs of 12 or 16 residues. We further noted that the TPP-conjugates of the 8-mer cyclic PNA as well as the 16-mer linear PNA displayed similar antiviral efficacy. However, cyclic PNAs were shown to be highly specific to their target sequences. This communication emphasizes on the importance of small constrained cyclic PNAs over both linear and hairpin structures for targeting biologically relevant RNA hairpins. PMID:23029603

  14. Inhibition of HIV Replication by Cyclic and Hairpin PNAs Targeting the HIV-1 TAR RNA Loop

    Directory of Open Access Journals (Sweden)

    Gregory Upert

    2012-01-01

    Full Text Available Human immunodeficiency virus-1 (HIV-1 replication and gene expression entails specific interaction of the viral protein Tat with its transactivation responsive element (TAR, to form a highly stable stem-bulge-loop structure. Previously, we described triphenylphosphonium (TPP cation-based vectors that efficiently deliver nucleotide analogs (PNAs into the cytoplasm of cells. In particular, we showed that the TPP conjugate of a linear 16-mer PNA targeting the apical stem-loop region of TAR impedes Tat-mediated transactivation of the HIV-1 LTR in vitro and also in cell culture systems. In this communication, we conjugated TPP to cyclic and hairpin PNAs targeting the loop region of HIV-1 TAR and evaluated their antiviral efficacy in a cell culture system. We found that TPP-cyclic PNAs containing only 8 residues, showed higher antiviral potency compared to hairpin PNAs of 12 or 16 residues. We further noted that the TPP-conjugates of the 8-mer cyclic PNA as well as the 16-mer linear PNA displayed similar antiviral efficacy. However, cyclic PNAs were shown to be highly specific to their target sequences. This communication emphasizes on the importance of small constrained cyclic PNAs over both linear and hairpin structures for targeting biologically relevant RNA hairpins.

  15. HIV-1 Latency in Monocytes/Macrophages

    Directory of Open Access Journals (Sweden)

    Amit Kumar

    2014-04-01

    Full Text Available Human immunodeficiency virus type 1 (HIV-1 targets CD4+ T cells and cells of the monocyte/macrophage lineage. HIV pathogenesis is characterized by the depletion of T lymphocytes and by the presence of a population of cells in which latency has been established called the HIV-1 reservoir. Highly active antiretroviral therapy (HAART has significantly improved the life of HIV-1 infected patients. However, complete eradication of HIV-1 from infected individuals is not possible without targeting latent sources of infection. HIV-1 establishes latent infection in resting CD4+ T cells and findings indicate that latency can also be established in the cells of monocyte/macrophage lineage. Monocyte/macrophage lineage includes among others, monocytes, macrophages and brain resident macrophages. These cells are relatively more resistant to apoptosis induced by HIV-1, thus are important stable hideouts of the virus. Much effort has been made in the direction of eliminating HIV-1 resting CD4+ T-cell reservoirs. However, it is impossible to achieve a cure for HIV-1 without considering these neglected latent reservoirs, the cells of monocyte/macrophage lineage. In this review we will describe our current understanding of the mechanism of latency in monocyte/macrophage lineage and how such cells can be specifically eliminated from the infected host.

  16. Analytical Performances of Human Immunodeficiency Virus Type 1 RNA-Based Amplix® Real-Time PCR Platform for HIV-1 RNA Quantification

    Directory of Open Access Journals (Sweden)

    Christian Diamant Mossoro-Kpinde

    2016-01-01

    Full Text Available Objectives. We evaluated the performances of Amplix real-time PCR platform developed by Biosynex (Strasbourg, France, combining automated station extraction (Amplix station 16 Dx and real-time PCR (Amplix NG, for quantifying plasma HIV-1 RNA by lyophilized HIV-1 RNA-based Amplix reagents targeting gag and LTR, using samples from HIV-1-infected adults from Central African Republic. Results. Amplix real-time PCR assay showed low limit of detection (28 copies/mL, across wide dynamic range (1.4–10 log copies/mL, 100% sensitivity and 99% specificity, high reproducibility, and accuracy with mean bias < 5%. The assay showed excellent correlations and concordance of 95.3% with the reference HIV-1 RNA load assay (Roche, with mean absolute bias of +0.097 log copies/mL by Bland-Altman analysis. The assay was able to detect and quantify the most prevalent HIV-1 subtype strains and the majority of non-B subtypes, CRFs of HIV-1 group M, and HIV-1 groups N and O circulating in Central Africa. The Amplix assay showed 100% sensitivity and 99.6% specificity to diagnose virological failure in clinical samples from antiretroviral drug-experienced patients. Conclusions. The HIV-1 RNA-based Amplix real-time PCR platform constitutes sensitive and reliable system for clinical monitoring of HIV-1 RNA load in HIV-1-infected children and adults, particularly adapted to intermediate laboratory facilities in sub-Saharan Africa.

  17. HIV-1 Tat regulates the expression of the dcw operon and stimulates the proliferation of bacteria.

    Science.gov (United States)

    Wei, Jinsong; Zhang, Yumin; Knapp, Pamela E; Zhao, Tianyong

    2016-01-01

    Infections of pathogenic bacteria are very common in acquired immunodeficiency syndrome (AIDS) patients. However, the biological effects of HIV-1 Tat on bacteria are incompletely understood. In this study, HIV-1 Tat was expressed in Escherichia coli and Pseudomonas aeruginosa (PA01) to investigate its biological effects on bacteria. Bacterial cells expressing either HIV-1 Tat1-86 (Tat1-86) or HIV-1 Tat1-72 (Tat1-72) grow significantly faster than those with either only an empty vector or an unrelated control (GFP or Rluc). Supplementation of purified HIV-1 Tat1-86 or Tat1-101 protein into bacterial culture medium stimulated the growth of both E. coli and PA01. The expression profile of certain cell division-associated genes, such as those in the division cell wall (dcw) operon (ftsA, ftsQ, ftsW and ftsZ), yafO and zipA, was altered in HIV-1 Tat1-86 expressing E. coli BL21(DE3). Furthermore, the expression of firefly luciferase (Fluc) reporter gene, when engineered for control by the dcw promoter and terminator, was enhanced by HIV-1 Tat in E. coli, confirming that HIV-1 Tat transcriptionally regulates the expression of the dcw operon. The finding that HIV-1 Tat stimulates bacterial growth whether it is produced intracellularly or applied extracellularly may have relevance for HIV patients who are highly susceptible to opportunistic bacterial infections. Contents category: Viruses -Retroviruses. The GenBank accession number for the sequence of HIV-1 Tat1-86 is AF324439.1. Copyright © 2015 Elsevier Ltd. All rights reserved.

  18. Regulation of FeLV-945 by c-Myb binding and CBP recruitment to the LTR

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    Finstad Samantha L

    2004-09-01

    Full Text Available Abstract Background Feline leukemia virus (FeLV induces degenerative, proliferative and malignant hematologic disorders in its natural host, the domestic cat. FeLV-945 is a viral variant identified as predominant in a cohort of naturally infected animals. FeLV-945 contains a unique sequence motif in the long terminal repeat (LTR comprised of a single copy of transcriptional enhancer followed by a 21-bp sequence triplicated in tandem. The LTR is precisely conserved among independent cases of multicentric lymphoma, myeloproliferative disease and anemia in animals from the cohort. The 21-bp triplication was previously shown to act as a transcriptional enhancer preferentially in hematopoietic cells and to confer a replicative advantage. The objective of the present study was to examine the molecular mechanism by which the 21-bp triplication exerts its influence and the selective advantage responsible for its precise conservation. Results Potential binding sites for the transcription factor, c-Myb, were identified across the repeat junctions of the 21-bp triplication. Such sites would not occur in the absence of the repeat; thus, a requirement for c-Myb binding to the repeat junctions of the triplication would exert a selective pressure to conserve its sequence precisely. Electrophoretic mobility shift assays demonstrated specific binding of c-Myb to the 21-bp triplication. Reporter gene assays showed that the triplication-containing LTR is responsive to c-Myb, and that responsiveness requires the presence of both c-Myb binding sites. Results further indicated that c-Myb in complex with the 21-bp triplication recruits the transcriptional co-activator, CBP, a regulator of normal hematopoiesis. FeLV-945 replication was shown to be positively regulated by CBP in a manner dependent on the presence of the 21-bp triplication. Conclusion Binding sites for c-Myb across the repeat junctions of the 21-bp triplication may account for its precise conservation in

  19. Cytoplasmic Dynein Promotes HIV-1 Uncoating

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    Paulina Pawlica

    2014-11-01

    Full Text Available Retroviral capsid (CA cores undergo uncoating during their retrograde transport (toward the nucleus, and/or after reaching the nuclear membrane. However, whether HIV-1 CA core uncoating is dependent upon its transport is not understood. There is some evidence that HIV-1 cores retrograde transport involves cytoplasmic dynein complexes translocating on microtubules. Here we investigate the role of dynein-dependent transport in HIV-1 uncoating. To interfere with dynein function, we depleted dynein heavy chain (DHC using RNA interference, and we over-expressed p50/dynamitin. In immunofluorescence microscopy experiments, DHC depletion caused an accumulation of CA foci in HIV-1 infected cells. Using a biochemical assay to monitor HIV-1 CA core disassembly in infected cells, we observed an increase in amounts of intact (pelletable CA cores upon DHC depletion or p50 over-expression. Results from these two complementary assays suggest that inhibiting dynein-mediated transport interferes with HIV-1 uncoating in infected cells, indicating the existence of a functional link between HIV-1 transport and uncoating.

  20. Natural Plant Alkaloid (Emetine Inhibits HIV-1 Replication by Interfering with Reverse Transcriptase Activity

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    Ana Luiza Chaves Valadão

    2015-06-01

    Full Text Available Ipecac alkaloids are secondary metabolites produced in the medicinal plant Psychotria ipecacuanha. Emetine is the main alkaloid of ipecac and one of the active compounds in syrup of Ipecac with emetic property. Here we evaluated emetine’s potential as an antiviral agent against Human Immunodeficiency Virus. We performed in vitro Reverse Transcriptase (RT Assay and Natural Endogenous Reverse Transcriptase Activity Assay (NERT to evaluate HIV RT inhibition. Emetine molecular docking on HIV-1 RT was also analyzed. Phenotypic assays were performed in non-lymphocytic and in Peripheral Blood Mononuclear Cells (PBMC with HIV-1 wild-type and HIV-harboring RT-resistant mutation to Nucleoside Reverse Transcriptase Inhibitors (M184V. Our results showed that HIV-1 RT was blocked in the presence of emetine in both models: in vitro reactions with isolated HIV-1 RT and intravirion, measured by NERT. Emetine revealed a strong potential of inhibiting HIV-1 replication in both cellular models, reaching 80% of reduction in HIV-1 infection, with low cytotoxic effect. Emetine also blocked HIV-1 infection of RT M184V mutant. These results suggest that emetine is able to penetrate in intact HIV particles, and bind and block reverse transcription reaction, suggesting that it can be used as anti-HIV microbicide. Taken together, our findings provide additional pharmacological information on the potential therapeutic effects of emetine.

  1. Reactivation of HIV-1 from Latency by an Ingenol Derivative from Euphorbia Kansui.

    Science.gov (United States)

    Wang, Pengfei; Lu, Panpan; Qu, Xiying; Shen, Yinzhong; Zeng, Hanxian; Zhu, Xiaoli; Zhu, Yuqi; Li, Xian; Wu, Hao; Xu, Jianqing; Lu, Hongzhou; Ma, Zhongjun; Zhu, Huanzhang

    2017-08-25

    Cells harboring latent HIV-1 pose a major obstacle to eradication of the virus. The 'shock and kill' strategy has been broadly explored to purge the latent reservoir; however, none of the current latency-reversing agents (LRAs) can safely and effectively activate the latent virus in patients. In this study, we report an ingenol derivative called EK-16A, isolated from the traditional Chinese medicinal herb Euphorbia kansui, which displays great potential in reactivating latent HIV-1. A comparison of the doses used to measure the potency indicated EK-16A to be 200-fold more potent than prostratin in reactivating HIV-1 from latently infected cell lines. EK-16A also outperformed prostratin in ex vivo studies on cells from HIV-1-infected individuals, while maintaining minimal cytotoxicity effects on cell viability and T cell activation. Furthermore, EK-16A exhibited synergy with other LRAs in reactivating latent HIV-1. Mechanistic studies indicated EK-16A to be a PKCγ activator, which promoted both HIV-1 transcription initiation by NF-κB and elongation by P-TEFb signal pathways. Further investigations aimed to add this compound to the therapeutic arsenal for HIV-1 eradication are in the pipeline.

  2. Evolutionary characterization of Ty3/gypsy-like LTR retrotransposons in the parasitic cestode Echinococcus granulosus.

    Science.gov (United States)

    Bae, Young-An

    2016-11-01

    Cyclophyllidean cestodes including Echinococcus granulosus have a smaller genome and show characteristics such as loss of the gut, a segmented body plan, and accelerated growth rate in hosts compared with other tissue-invading helminths. In an effort to address the molecular mechanism relevant to genome shrinkage, the evolutionary status of long-terminal-repeat (LTR) retrotransposons, which are known as the most potent genomic modulators, was investigated in the E. granulosus draft genome. A majority of the E. granulosus LTR retrotransposons were classified into a novel characteristic clade, named Saci-2, of the Ty3/gypsy family, while the remaining elements belonged to the CsRn1 clade of identical family. Their nucleotide sequences were heavily corrupted by frequent base substitutions and segmental losses. The ceased mobile activity of the major retrotransposons and the following intrinsic DNA loss in their inactive progenies might have contributed to decrease in genome size. Apart from the degenerate copies, a gag gene originating from a CsRn1-like element exhibited substantial evidences suggesting its domestication including a preserved coding profile and transcriptional activity, the presence of syntenic orthologues in cestodes, and selective pressure acting on the gene. To my knowledge, the endogenized gag gene is reported for the first time in invertebrates, though its biological function remains elusive.

  3. Immune defence against HIV-1 infection in HIV-1-exposed seronegative persons.

    Science.gov (United States)

    Schmechel, S C; Russell, N; Hladik, F; Lang, J; Wilson, A; Ha, R; Desbien, A; McElrath, M J

    2001-11-01

    Rare individuals who are repeatedly exposed to HIV-1 through unprotected sexual contact fail to acquire HIV-1 infection. These persons represent a unique study population to evaluate mechanisms by which HIV-1 replication is either prevented or controlled. We followed longitudinally a group of healthy HIV-1 seronegative persons each reporting repeated high-risk sexual activities with their HIV-1-infected partner at enrollment. The volunteers were primarily (90%) male homosexuals, maintaining high risk activities with their known infected partner (45%) or multiple other partners (61%). We evaluated the quantity and specificity of HIV-1-specific T cells in 31 exposed seronegatives (ES) using a IFN-gamma ELISPOT assay to enumerate T cells recognizing epitopes within HIV-1 Env, Gag, Pol and Nef. PBMC from only three of the 31 volunteers demonstrated ex vivo HIV-1-specific IFN-gamma secretion, in contrast to nearly 30% exhibiting cytolytic responses in previous studies. These findings suggest that if T cell responses in ES are induced by HIV-1 exposure, the frequency is at low levels in most of them, and below the level of detection using the ELISPOT assay. Alternative approaches to improve the sensitivity of detection may include use of dendritic cells as antigen-presenting cells in the ex vivo assay and more careful definition of the risk behavior and extent of HIV-1 exposure in conjunction with the evaluation of T cell responses.

  4. Full Length Research Paper LTR-retrotransposons-based molecular ...

    African Journals Online (AJOL)

    LTR-retrotransposons possess unique properties that make them appropriate for investigating relationships between closely related species and populations. The aim of the current study was to employ Ty1-copia group retrotransposons as molecular markers in cultivated Egyptian cottons, G. barbadense L. Restriction site ...

  5. HIV-1 replication in cell lines harboring INI1/hSNF5 mutations.

    Science.gov (United States)

    Sorin, Masha; Yung, Eric; Wu, Xuhong; Kalpana, Ganjam V

    2006-08-31

    INI1/hSNF5 is a cellular protein that directly interacts with HIV-1 integrase (IN). It is specifically incorporated into HIV-1 virions. A dominant negative mutant derived from INI1 inhibits HIV-1 replication. Recent studies indicate that INI1 is associated with pre-integration and reverse transcription complexes that are formed upon viral entry into the target cells. INI1 also is a tumor suppressor, biallelically deleted/mutated in malignant rhabdoid tumors. We have utilized cell lines derived from the rhabdoid tumors, MON and STA-WT1, that harbor either null or truncating mutations of INI1 respectively, to assess the effect of INI1 on HIV-1 replication. We found that while HIV-1 virions produced in 293T cells efficiently transduced MON and STA-WT1 cells, HIV-1 particle production was severely reduced in both of these cells. Reintroduction of INI1 into MON and STA-WT1 significantly enhanced the particle production in both cell lines. HIV-1 particles produced in MON cells were reduced for infectivity, while those produced in STA-WT1 were not. Further analysis indicated the presence of INI1 in those virions produced from STA-WT1 but not from those produced from MON cells. HIV-1 produced in MON cells were defective for synthesis of early and late reverse transcription products in the target cells. Furthermore, virions produced in MON cells were defective for exogenous reverse transcriptase activity carried out using exogenous template, primer and substrate. Our results suggest that INI1-deficient cells exhibit reduced particle production that can be partly enhanced by re-introduction of INI1. Infectivity of HIV-1 produced in some but not all INI1 defective cells, is affected and this defect may correlate to the lack of INI1 and/or some other proteins in these virions. The block in early events of virion produced from MON cells appears to be at the stage of reverse transcription. These studies suggest that presence of INI1 or some other host factor in virions and

  6. HIV-1 replication in cell lines harboring INI1/hSNF5 mutations

    Directory of Open Access Journals (Sweden)

    Wu Xuhong

    2006-08-01

    Full Text Available Abstract Background INI1/hSNF5 is a cellular protein that directly interacts with HIV-1 integrase (IN. It is specifically incorporated into HIV-1 virions. A dominant negative mutant derived from INI1 inhibits HIV-1 replication. Recent studies indicate that INI1 is associated with pre-integration and reverse transcription complexes that are formed upon viral entry into the target cells. INI1 also is a tumor suppressor, biallelically deleted/mutated in malignant rhabdoid tumors. We have utilized cell lines derived from the rhabdoid tumors, MON and STA-WT1, that harbor either null or truncating mutations of INI1 respectively, to assess the effect of INI1 on HIV-1 replication. Results We found that while HIV-1 virions produced in 293T cells efficiently transduced MON and STA-WT1 cells, HIV-1 particle production was severely reduced in both of these cells. Reintroduction of INI1 into MON and STA-WT1 significantly enhanced the particle production in both cell lines. HIV-1 particles produced in MON cells were reduced for infectivity, while those produced in STA-WT1 were not. Further analysis indicated the presence of INI1 in those virions produced from STA-WT1 but not from those produced from MON cells. HIV-1 produced in MON cells were defective for synthesis of early and late reverse transcription products in the target cells. Furthermore, virions produced in MON cells were defective for exogenous reverse transcriptase activity carried out using exogenous template, primer and substrate. Conclusion Our results suggest that INI1-deficient cells exhibit reduced particle production that can be partly enhanced by re-introduction of INI1. Infectivity of HIV-1 produced in some but not all INI1 defective cells, is affected and this defect may correlate to the lack of INI1 and/or some other proteins in these virions. The block in early events of virion produced from MON cells appears to be at the stage of reverse transcription. These studies suggest that

  7. Infection and depletion of CD4+ group-1 innate lymphoid cells by HIV-1 via type-I interferon pathway.

    Directory of Open Access Journals (Sweden)

    Juanjuan Zhao

    2018-01-01

    Full Text Available Innate lymphoid cells (ILCs are severely depleted during chronic HIV-1 infection by unclear mechanisms. We report here that human ILC1s comprising of CD4+ and CD4- subpopulations were present in various human lymphoid organs but with different transcription programs and functions. Importantly, CD4+ ILC1s expressed HIV-1 co-receptors and were productively infected by HIV-1 in vitro and in vivo. Furthermore, chronic HIV-1 infection activated and depleted both CD4+ and CD4- ILC1s, and impaired their cytokine production activity. Highly active antiretroviral (HAART therapy in HIV-1 patients efficiently rescued the ILC1 numbers and reduced their activation, but failed to restore their functionality. We also found that blocking type-I interferon (IFN-I signaling during HIV-1 infection in vivo in humanized mice prevented HIV-1 induced depletion or apoptosis of ILC1 cells. Therefore, we have identified the CD4+ ILC1 cells as a new target population for HIV-1 infection, and revealed that IFN-I contributes to the depletion of ILC1s during HIV-1 infection.

  8. Persistence of attenuated HIV-1 rev alleles in an epidemiologically linked cohort of long-term survivors infected with nef-deleted virus

    Directory of Open Access Journals (Sweden)

    Wesselingh Steven L

    2007-07-01

    Full Text Available Abstract Background The Sydney blood bank cohort (SBBC of long-term survivors consists of multiple individuals infected with nef-deleted, attenuated strains of human immunodeficiency virus type 1 (HIV-1. Although the cohort members have experienced differing clinical courses and now comprise slow progressors (SP as well as long-term nonprogressors (LTNP, longitudinal analysis of nef/long-terminal repeat (LTR sequences demonstrated convergent nef/LTR sequence evolution in SBBC SP and LTNP. Thus, the in vivo pathogenicity of attenuated HIV-1 strains harboured by SBBC members is dictated by factors other than nef/LTR. Therefore, to determine whether defects in other viral genes contribute to attenuation of these HIV-1 strains, we characterized dominant HIV-1 rev alleles that persisted in 4 SBBC subjects; C18, C64, C98 and D36. Results The ability of Rev derived from D36 and C64 to bind the Rev responsive element (RRE in RNA binding assays was reduced by approximately 90% compared to Rev derived from HIV-1NL4-3, C18 or C98. D36 Rev also had a 50–60% reduction in ability to express Rev-dependent reporter constructs in mammalian cells. In contrast, C64 Rev had only marginally decreased Rev function despite attenuated RRE binding. In D36 and C64, attenuated RRE binding was associated with rare amino acid changes at 3 highly conserved residues; Gln to Pro at position 74 immediately N-terminal to the Rev activation domain, and Val to Leu and Ser to Pro at positions 104 and 106 at the Rev C-terminus, respectively. In D36, reduced Rev function was mapped to an unusual 13 amino acid extension at the Rev C-terminus. Conclusion These findings provide new genetic and mechanistic insights important for Rev function, and suggest that Rev function, not Rev/RRE binding may be rate limiting for HIV-1 replication. In addition, attenuated rev alleles may contribute to viral attenuation and long-term survival of HIV-1 infection in a subset of SBBC members.

  9. Chimeric peptide-mediated siRNA transduction to inhibit HIV-1 infection.

    Science.gov (United States)

    Bivalkar-Mehla, Shalmali; Mehla, Rajeev; Chauhan, Ashok

    2017-04-01

    Persistent human immunodeficiency virus 1 (HIV-1) infection provokes immune activation and depletes CD4 +  lymphocytes, leading to acquired immunodeficiency syndrome. Uninterrupted administration of combination antiretroviral therapy (cART) in HIV-infected patients suppresses viral replication to below the detectable level and partially restores the immune system. However, cART-unresponsive residual HIV-1 infection and elusive transcriptionally silent but reactivatable viral reservoirs maintain a permanent viral DNA blue print. The virus rebounds within a few weeks after interruption of suppressive therapy. Adjunct gene therapy to control viral replication by ribonucleic acid interference (RNAi) is a post-transcriptional gene silencing strategy that could suppress residual HIV-1 burden and overcome viral resistance. Small interfering ribonucleic acids (siRNAs) are efficient transcriptional inhibitors, but need delivery systems to reach inside target cells. We investigated the potential of chimeric peptide (FP-PTD) to deliver specific siRNAs to HIV-1-susceptible and permissive cells. Chimeric FP-PTD peptide was designed with an RNA binding domain (PTD) to bind siRNA and a cell fusion peptide domain (FP) to enter cells. FP-PTD-siRNA complex entered and inhibited HIV-1 replication in susceptible cells, and could be a candidate for in vivo testing.

  10. HIV-1 transmission linkage in an HIV-1 prevention clinical trial

    Energy Technology Data Exchange (ETDEWEB)

    Leitner, Thomas [Los Alamos National Laboratory; Campbell, Mary S [UNIV OF WASHINGTON; Mullins, James I [UNIV OF WASHINGTON; Hughes, James P [UNIV OF WASHINGTON; Wong, Kim G [UNIV OF WASHINGTON; Raugi, Dana N [UNIV OF WASHINGTON; Scrensen, Stefanie [UNIV OF WASHINGTON

    2009-01-01

    HIV-1 sequencing has been used extensively in epidemiologic and forensic studies to investigate patterns of HIV-1 transmission. However, the criteria for establishing genetic linkage between HIV-1 strains in HIV-1 prevention trials have not been formalized. The Partners in Prevention HSV/HIV Transmission Study (ClinicaITrials.gov NCT00194519) enrolled 3408 HIV-1 serodiscordant heterosexual African couples to determine the efficacy of genital herpes suppression with acyclovir in reducing HIV-1 transmission. The trial analysis required laboratory confirmation of HIV-1 linkage between enrolled partners in couples in which seroconversion occurred. Here we describe the process and results from HIV-1 sequencing studies used to perform transmission linkage determination in this clinical trial. Consensus Sanger sequencing of env (C2-V3-C3) and gag (p17-p24) genes was performed on plasma HIV-1 RNA from both partners within 3 months of seroconversion; env single molecule or pyrosequencing was also performed in some cases. For linkage, we required monophyletic clustering between HIV-1 sequences in the transmitting and seroconverting partners, and developed a Bayesian algorithm using genetic distances to evaluate the posterior probability of linkage of participants sequences. Adjudicators classified transmissions as linked, unlinked, or indeterminate. Among 151 seroconversion events, we found 108 (71.5%) linked, 40 (26.5%) unlinked, and 3 (2.0%) to have indeterminate transmissions. Nine (8.3%) were linked by consensus gag sequencing only and 8 (7.4%) required deep sequencing of env. In this first use of HIV-1 sequencing to establish endpoints in a large clinical trial, more than one-fourth of transmissions were unlinked to the enrolled partner, illustrating the relevance of these methods in the design of future HIV-1 prevention trials in serodiscordant couples. A hierarchy of sequencing techniques, analysis methods, and expert adjudication contributed to the linkage

  11. Nuclear trafficking of the HIV-1 pre-integration complex depends on the ADAM10 intracellular domain

    International Nuclear Information System (INIS)

    Endsley, Mark A.; Somasunderam, Anoma D.; Li, Guangyu; Oezguen, Numan; Thiviyanathan, Varatharasa; Murray, James L.; Rubin, Donald H.; Hodge, Thomas W.

    2014-01-01

    Previously, we showed that ADAM10 is necessary for HIV-1 replication in primary human macrophages and immortalized cell lines. Silencing ADAM10 expression interrupted the HIV-1 life cycle prior to nuclear translocation of viral cDNA. Furthermore, our data indicated that HIV-1 replication depends on the expression of ADAM15 and γ-secretase, which proteolytically processes ADAM10. Silencing ADAM15 or γ-secretase expression inhibits HIV-1 replication between reverse transcription and nuclear entry. Here, we show that ADAM10 expression also supports replication in CD4 + T lymphocytes. The intracellular domain (ICD) of ADAM10 associates with the HIV-1 pre-integration complex (PIC) in the cytoplasm and immunoprecipitates and co-localizes with HIV-1 integrase, a key component of PIC. Taken together, our data support a model whereby ADAM15/γ-secretase processing of ADAM10 releases the ICD, which then incorporates into HIV-1 PIC to facilitate nuclear trafficking. Thus, these studies suggest ADAM10 as a novel therapeutic target for inhibiting HIV-1 prior to nuclear entry. - Highlights: • Nuclear trafficking of the HIV-1 pre-integration complex depends on ADAM10. • ADAM10 associates with HIV-1 integrase in the pre-integration complex. • HIV-1 replication depends on the expression of ADAM15 and γ-secretase. • Silencing ADAM15 or γ-secretase expression inhibits nuclear import of viral cDNA. • ADAM10 is important for HIV-1 replication in human macrophages and CD4 + T lymphocytes

  12. Nuclear trafficking of the HIV-1 pre-integration complex depends on the ADAM10 intracellular domain

    Energy Technology Data Exchange (ETDEWEB)

    Endsley, Mark A., E-mail: maendsle@utmb.edu [Department Internal Medicine, Division of Infectious Diseases, University of Texas Medical Branch, 301 University Blvd, Galveston, TX 77555 (United States); Somasunderam, Anoma D., E-mail: asomasun@utmb.edu [Department Internal Medicine, Division of Infectious Diseases, University of Texas Medical Branch, 301 University Blvd, Galveston, TX 77555 (United States); Li, Guangyu, E-mail: LIG001@mail.etsu.edu [Department of Internal Medicine, Quillen College of Medicine, East Tennessee State University, Johnson City, TN 37614 (United States); Oezguen, Numan, E-mail: numan.oezguen@bcm.edu [Department of Pathology and Immunology, Microbiome Center, Texas Children' s Hospital, Houston, TX 77030 (United States); Thiviyanathan, Varatharasa, E-mail: Varatharasa.Thiviyanathan@uth.tmc.edu [Institute of Molecular Medicine, University of Texas Health Science Center, Houston, TX 77030 (United States); Murray, James L., E-mail: jmurray100@yahoo.com [GeneTAG Technology, Inc., 3155 Northwoods Place, Norcross, GA 30071 (United States); Rubin, Donald H., E-mail: don.h.rubin@vanderbilt.edu [Research Medicine, VA Tennessee Valley Healthcare System, 1310 24th Ave. South, Nashville, TN 37212 (United States); Departments of Medicine, Pathology, Microbiology and Immunology, Vanderbilt University School of Medicine, 1161 21st Ave South, Nashville, TN 37232 (United States); Hodge, Thomas W., E-mail: twhodge3@gmail.com [Pre-clinical and Antiviral Research, Tamir Biotechnology, Inc., 12625 High Bluff Dr., Suite 113, San Diego, CA 92130 (United States); and others

    2014-04-15

    Previously, we showed that ADAM10 is necessary for HIV-1 replication in primary human macrophages and immortalized cell lines. Silencing ADAM10 expression interrupted the HIV-1 life cycle prior to nuclear translocation of viral cDNA. Furthermore, our data indicated that HIV-1 replication depends on the expression of ADAM15 and γ-secretase, which proteolytically processes ADAM10. Silencing ADAM15 or γ-secretase expression inhibits HIV-1 replication between reverse transcription and nuclear entry. Here, we show that ADAM10 expression also supports replication in CD4{sup +} T lymphocytes. The intracellular domain (ICD) of ADAM10 associates with the HIV-1 pre-integration complex (PIC) in the cytoplasm and immunoprecipitates and co-localizes with HIV-1 integrase, a key component of PIC. Taken together, our data support a model whereby ADAM15/γ-secretase processing of ADAM10 releases the ICD, which then incorporates into HIV-1 PIC to facilitate nuclear trafficking. Thus, these studies suggest ADAM10 as a novel therapeutic target for inhibiting HIV-1 prior to nuclear entry. - Highlights: • Nuclear trafficking of the HIV-1 pre-integration complex depends on ADAM10. • ADAM10 associates with HIV-1 integrase in the pre-integration complex. • HIV-1 replication depends on the expression of ADAM15 and γ-secretase. • Silencing ADAM15 or γ-secretase expression inhibits nuclear import of viral cDNA. • ADAM10 is important for HIV-1 replication in human macrophages and CD4{sup +} T lymphocytes.

  13. The effect of CD4 receptor downregulation and its downstream signaling molecules on HIV-1 latency

    International Nuclear Information System (INIS)

    Kim, Kyung-Chang; Kim, Hyeon Guk; Roh, Tae-Young; Park, Jihwan; Jung, Kyung-Min; Lee, Joo-Shil; Choi, Sang-Yun; Kim, Sung Soon; Choi, Byeong-Sun

    2011-01-01

    Research highlights: → CD4 receptors were downregulated on the surface of HIV-1 latently infected cells. → CD4 downstream signaling molecules were suppressed in HIV-1 latently infected cells. → HIV-1 progeny can be reactivated by induction of T-cell activation signal molecules. → H3K4me3 and H3K9ac were highly enriched in CD4 downstream signaling molecules. → HIV-1 latency can be maintained by the reduction of downstream signaling molecules. -- Abstract: HIV-1 can establish a latent infection in memory CD4 + T cells to evade the host immune response. CD4 molecules can act not only as the HIV-1 receptor for entry but also as the trigger in an intracellular signaling cascade for T-cell activation and proliferation via protein tyrosine kinases. Novel chronic HIV-1-infected A3.01-derived (NCHA) cells were used to examine the involvement of CD4 downstream signaling in HIV-1 latency. CD4 receptors in NCHA cells were dramatically downregulated on its surface but were slightly decreased in whole-cell lysates. The expression levels of CD4 downstream signaling molecules, including P56 Lck , ZAP-70, LAT, and c-Jun, were sharply decreased in NCHA cells. The lowered histone modifications of H3K4me3 and H3K9ac correlated with the downregulation of P56 Lck , ZAP-70, and LAT in NCHA cells. AP-1 binding activity was also reduced in NCHA cells. LAT and c-Jun suppressed in NCHA cells were highly induced after PMA treatment. In epigenetic analysis, other signal transduction molecules which are associated with active and/or latent HIV-1 infection showed normal states in HIV-1 latently infected cells compared to A3.01 cells. In conclusion, we demonstrated that the HIV-1 latent state is sustained by the reduction of downstream signaling molecules via the downregulation of CD4 and the attenuated activity of transcription factor as AP-1. The HIV-1 latency model via T-cell deactivation may provide some clues for the development of the new antireservoir therapy.

  14. The effect of CD4 receptor downregulation and its downstream signaling molecules on HIV-1 latency

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Kyung-Chang [National Institute of Health, Chungbuk (Korea, Republic of); School of Life Science and Biotechnology, Korea University, Seoul (Korea, Republic of); Kim, Hyeon Guk [National Institute of Health, Chungbuk (Korea, Republic of); Roh, Tae-Young [Division of Molecular and Life Science, Pohang University of Science and Technology, Pohang, Gyeongbuk (Korea, Republic of); Division of Integrative Biosciences and Biotechnology, Pohang University of Science and Technology, Pohang, Gyeongbuk (Korea, Republic of); Park, Jihwan [Division of Molecular and Life Science, Pohang University of Science and Technology, Pohang, Gyeongbuk (Korea, Republic of); Jung, Kyung-Min; Lee, Joo-Shil [National Institute of Health, Chungbuk (Korea, Republic of); Choi, Sang-Yun [School of Life Science and Biotechnology, Korea University, Seoul (Korea, Republic of); Kim, Sung Soon [National Institute of Health, Chungbuk (Korea, Republic of); Choi, Byeong-Sun, E-mail: byeongsun@korea.kr [National Institute of Health, Chungbuk (Korea, Republic of)

    2011-01-14

    Research highlights: {yields} CD4 receptors were downregulated on the surface of HIV-1 latently infected cells. {yields} CD4 downstream signaling molecules were suppressed in HIV-1 latently infected cells. {yields} HIV-1 progeny can be reactivated by induction of T-cell activation signal molecules. {yields} H3K4me3 and H3K9ac were highly enriched in CD4 downstream signaling molecules. {yields} HIV-1 latency can be maintained by the reduction of downstream signaling molecules. -- Abstract: HIV-1 can establish a latent infection in memory CD4 + T cells to evade the host immune response. CD4 molecules can act not only as the HIV-1 receptor for entry but also as the trigger in an intracellular signaling cascade for T-cell activation and proliferation via protein tyrosine kinases. Novel chronic HIV-1-infected A3.01-derived (NCHA) cells were used to examine the involvement of CD4 downstream signaling in HIV-1 latency. CD4 receptors in NCHA cells were dramatically downregulated on its surface but were slightly decreased in whole-cell lysates. The expression levels of CD4 downstream signaling molecules, including P56{sup Lck}, ZAP-70, LAT, and c-Jun, were sharply decreased in NCHA cells. The lowered histone modifications of H3K4me3 and H3K9ac correlated with the downregulation of P56{sup Lck}, ZAP-70, and LAT in NCHA cells. AP-1 binding activity was also reduced in NCHA cells. LAT and c-Jun suppressed in NCHA cells were highly induced after PMA treatment. In epigenetic analysis, other signal transduction molecules which are associated with active and/or latent HIV-1 infection showed normal states in HIV-1 latently infected cells compared to A3.01 cells. In conclusion, we demonstrated that the HIV-1 latent state is sustained by the reduction of downstream signaling molecules via the downregulation of CD4 and the attenuated activity of transcription factor as AP-1. The HIV-1 latency model via T-cell deactivation may provide some clues for the development of the new

  15. Engineering HIV-1-resistant T-cells from short-hairpin RNA-expressing hematopoietic stem/progenitor cells in humanized BLT mice.

    Directory of Open Access Journals (Sweden)

    Gene-Errol E Ringpis

    Full Text Available Down-regulation of the HIV-1 coreceptor CCR5 holds significant potential for long-term protection against HIV-1 in patients. Using the humanized bone marrow/liver/thymus (hu-BLT mouse model which allows investigation of human hematopoietic stem/progenitor cell (HSPC transplant and immune system reconstitution as well as HIV-1 infection, we previously demonstrated stable inhibition of CCR5 expression in systemic lymphoid tissues via transplantation of HSPCs genetically modified by lentiviral vector transduction to express short hairpin RNA (shRNA. However, CCR5 down-regulation will not be effective against existing CXCR4-tropic HIV-1 and emergence of resistant viral strains. As such, combination approaches targeting additional steps in the virus lifecycle are required. We screened a panel of previously published shRNAs targeting highly conserved regions and identified a potent shRNA targeting the R-region of the HIV-1 long terminal repeat (LTR. Here, we report that human CD4(+ T-cells derived from transplanted HSPC engineered to co-express shRNAs targeting CCR5 and HIV-1 LTR are resistant to CCR5- and CXCR4- tropic HIV-1-mediated depletion in vivo. Transduction with the combination vector suppressed CXCR4- and CCR5- tropic viral replication in cell lines and peripheral blood mononuclear cells in vitro. No obvious cytotoxicity or interferon response was observed. Transplantation of combination vector-transduced HSPC into hu-BLT mice resulted in efficient engraftment and subsequent stable gene marking and CCR5 down-regulation in human CD4(+ T-cells within peripheral blood and systemic lymphoid tissues, including gut-associated lymphoid tissue, a major site of robust viral replication, for over twelve weeks. CXCR4- and CCR5- tropic HIV-1 infection was effectively inhibited in hu-BLT mouse spleen-derived human CD4(+ T-cells ex vivo. Furthermore, levels of gene-marked CD4(+ T-cells in peripheral blood increased despite systemic infection with either

  16. Semen Bacterial Concentrations and HIV-1 RNA Shedding Among HIV-1–Seropositive Kenyan Men

    Science.gov (United States)

    Srinivasan, Sujatha; Huang, Dandi; Ko, Daisy L.; Sanders, Eduard J.; Peshu, Norbert M.; Krieger, John N.; Muller, Charles H.; Coombs, Robert W.; Fredricks, David N.; Graham, Susan M.

    2017-01-01

    Introduction: HIV-1 is transmitted through semen from men to their sexual partners. Genital infections can increase HIV-1 RNA shedding in semen, but shedding also occurs in the absence of typical pathogens. We hypothesized that higher bacterial concentrations in semen would be associated with higher HIV-1 RNA levels. Methods: We analyzed semen samples from 42 HIV-1–seropositive Kenyan men using quantitative polymerase chain reaction (PCR) to assess bacterial concentrations and real-time PCR to measure HIV-1 RNA levels. Generalized estimation equations were used to evaluate associations between these 2 measures. Broad-range 16S rRNA gene PCR with pyrosequencing was performed on a subset of 13 samples to assess bacterial community composition. Results: Bacteria were detected in 96.6% of 88 samples by quantitative PCR. Semen bacterial concentration and HIV-1 RNA levels were correlated 0.30 (P = 0.01). The association between bacterial concentration and HIV-1 RNA detection was not significant after adjustment for antiretroviral therapy (ART) (adjusted odds ratio: 1.27, 95% CI: 0.84 to 1.91). Factors associated with semen bacterial concentration included insertive anal sex (adjusted beta 0.92, 95% CI: 0.12 to 1.73) and ART use (adjusted beta: −0.77, 95% CI: −1.50 to 0.04). Among 13 samples with pyrosequencing data, Corynebacterium spp., Staphylococcus spp., and Streptococcus spp. were most frequently detected. Conclusion: Most of these HIV-1–infected men had bacteria in their semen. ART use was associated with undetectable semen HIV-1 RNA and lower semen bacterial concentrations, whereas insertive anal sex was associated with higher bacterial concentrations. Additional studies evaluating the relationship between semen bacteria, inflammation, mucosal immunity, and HIV-1 shedding are needed to understand implications for HIV-1 transmission. PMID:27861240

  17. Semen Bacterial Concentrations and HIV-1 RNA Shedding Among HIV-1-Seropositive Kenyan Men.

    Science.gov (United States)

    Korhonen, Christine J; Srinivasan, Sujatha; Huang, Dandi; Ko, Daisy L; Sanders, Eduard J; Peshu, Norbert M; Krieger, John N; Muller, Charles H; Coombs, Robert W; Fredricks, David N; Graham, Susan M

    2017-03-01

    HIV-1 is transmitted through semen from men to their sexual partners. Genital infections can increase HIV-1 RNA shedding in semen, but shedding also occurs in the absence of typical pathogens. We hypothesized that higher bacterial concentrations in semen would be associated with higher HIV-1 RNA levels. We analyzed semen samples from 42 HIV-1-seropositive Kenyan men using quantitative polymerase chain reaction (PCR) to assess bacterial concentrations and real-time PCR to measure HIV-1 RNA levels. Generalized estimation equations were used to evaluate associations between these 2 measures. Broad-range 16S rRNA gene PCR with pyrosequencing was performed on a subset of 13 samples to assess bacterial community composition. Bacteria were detected in 96.6% of 88 samples by quantitative PCR. Semen bacterial concentration and HIV-1 RNA levels were correlated 0.30 (P = 0.01). The association between bacterial concentration and HIV-1 RNA detection was not significant after adjustment for antiretroviral therapy (ART) (adjusted odds ratio: 1.27, 95% CI: 0.84 to 1.91). Factors associated with semen bacterial concentration included insertive anal sex (adjusted beta 0.92, 95% CI: 0.12 to 1.73) and ART use (adjusted beta: -0.77, 95% CI: -1.50 to 0.04). Among 13 samples with pyrosequencing data, Corynebacterium spp., Staphylococcus spp., and Streptococcus spp. were most frequently detected. Most of these HIV-1-infected men had bacteria in their semen. ART use was associated with undetectable semen HIV-1 RNA and lower semen bacterial concentrations, whereas insertive anal sex was associated with higher bacterial concentrations. Additional studies evaluating the relationship between semen bacteria, inflammation, mucosal immunity, and HIV-1 shedding are needed to understand implications for HIV-1 transmission.

  18. Construction of Nef-positive doxycycline-dependent HIV-1 variants using bicistronic expression elements

    Energy Technology Data Exchange (ETDEWEB)

    Velden, Yme U. van der; Kleibeuker, Wendy; Harwig, Alex; Klaver, Bep; Siteur-van Rijnstra, Esther; Frankin, Esmay; Berkhout, Ben; Das, Atze T., E-mail: a.t.das@amc.uva.nl

    2016-01-15

    Conditionally replicating HIV-1 variants that can be switched on and off at will are attractive tools for HIV research. We previously developed a genetically modified HIV-1 variant that replicates exclusively when doxycycline (dox) is administered. The nef gene in this HIV-rtTA variant was replaced with the gene encoding the dox-dependent rtTA transcriptional activator. Because loss of Nef expression compromises virus replication in primary cells and precludes studies on Nef function, we tested different approaches to restore Nef production in HIV-rtTA. Strategies that involved translation via an EMCV or synthetic internal ribosome entry site (IRES) failed because these elements were incompatible with efficient virus replication. Fusion protein approaches with the FMDV 2A peptide and human ubiquitin were successful and resulted in genetically-stable Nef-expressing HIV-rtTA strains that replicate more efficiently in primary T-cells and human immune system (HIS) mice than Nef-deficient variants, thus confirming the positive effect of Nef on in vivo virus replication. - Highlights: • Different approaches to encode additional proteins in the HIV-1 genome were tested. • IRES translation elements are incompatible with efficient HIV-1 replication. • Ubiquitin and 2A fusion protein approaches allow efficient HIV-1 replication. • Doxycycline-controlled HIV-1 variants that encode all viral proteins were developed. • Nef stimulates HIV-rtTA replication in primary cells and human immune system mice.

  19. μ-opioid modulation of HIV-1 coreceptor expressionand HIV-1 replication

    International Nuclear Information System (INIS)

    Steele, Amber D.; Henderson, Earl E.; Rogers, Thomas J.

    2003-01-01

    A substantial proportion of HIV-1-infected individuals are intravenous drug users (IVDUs) who abuse opiates. Opioids induce a number of immunomodulatory effects that may directly influence HIV-1 disease progression. In the present report, we have investigated the effect of opioids on the expression of the major HIV-1 coreceptors CXCR4 and CCR5. For these studies we have focused on opiates which are ligands for the μ-opioid receptor. Our results show that DAMGO, a selective μ-opioid agonist, increases CXCR4 and CCR5 expression in both CD3 + lymphoblasts and CD14 + monocytes three- to fivefold. Furthermore, DAMGO-induced elevation of HIV-1 coreceptor expression translates into enhanced replication of both X4 and R5 viral strains of HIV-1. We have confirmed the role of the μ-opioid receptor based on the ability of a μ-opioid receptor-selective antagonist to block the effects of DAMGO. We have also found that morphine enhances CXCR4 and CCR5 expression and subsequently increases both X4 and R5 HIV-1 infection. We suggest that the capacity of μ-opioids to increase HIV-1 coreceptor expression and replication may promote viral binding, trafficking of HIV-1-infected cells, and enhanced disease progression

  20. Influenza vaccination of HIV-1-positive and HIV-1-negative former intravenous drug users.

    Science.gov (United States)

    Amendola, A; Boschini, A; Colzani, D; Anselmi, G; Oltolina, A; Zucconi, R; Begnini, M; Besana, S; Tanzi, E; Zanetti, A R

    2001-12-01

    The immunogenicity of an anti-influenza vaccine was assessed in 409 former intravenous drug user volunteers and its effect on the levels of HIV-1 RNA, proviral DNA and on CD4+ lymphocyte counts in a subset HIV-1-positive subjects was measured. HIV-1-positive individuals (n = 72) were divided into three groups on the basis of their CD4+ lymphocyte counts, while the 337 HIV-1-negative participants were allocated into group four. Haemagglutination inhibiting (HI) responses varied from 45.8 to 70% in the HIV-1-positive subjects and were significantly higher in group four (80.7% responses to the H1N1 strain, 81.6% to the H3N2 strain, and 83% to the B strain). The percentage of subjects with HI protective antibody titres (> or = 1:40) increased significantly after vaccination, especially in HIV-1 uninfected subjects. Immunization caused no significant changes in CD4+ counts and in neither plasma HIV-1 RNA nor proviral DNA levels. Therefore, vaccination against influenza may benefit persons infected by HIV-1. Copyright 2001 Wiley-Liss, Inc.

  1. Molecular control of HIV-1 postintegration latency: implications for the development of new therapeutic strategies

    Directory of Open Access Journals (Sweden)

    Van Lint Carine

    2009-12-01

    Full Text Available Abstract The persistence of HIV-1 latent reservoirs represents a major barrier to virus eradication in infected patients under HAART since interruption of the treatment inevitably leads to a rebound of plasma viremia. Latency establishes early after infection notably (but not only in resting memory CD4+ T cells and involves numerous host and viral trans-acting proteins, as well as processes such as transcriptional interference, RNA silencing, epigenetic modifications and chromatin organization. In order to eliminate latent reservoirs, new strategies are envisaged and consist of reactivating HIV-1 transcription in latently-infected cells, while maintaining HAART in order to prevent de novo infection. The difficulty lies in the fact that a single residual latently-infected cell can in theory rekindle the infection. Here, we review our current understanding of the molecular mechanisms involved in the establishment and maintenance of HIV-1 latency and in the transcriptional reactivation from latency. We highlight the potential of new therapeutic strategies based on this understanding of latency. Combinations of various compounds used simultaneously allow for the targeting of transcriptional repression at multiple levels and can facilitate the escape from latency and the clearance of viral reservoirs. We describe the current advantages and limitations of immune T-cell activators, inducers of the NF-κB signaling pathway, and inhibitors of deacetylases and histone- and DNA- methyltransferases, used alone or in combinations. While a solution will not be achieved by tomorrow, the battle against HIV-1 latent reservoirs is well- underway.

  2. Acyclovir and Transmission of HIV-1 from Persons Infected with HIV-1 and HSV-2

    Science.gov (United States)

    Celum, Connie; Wald, Anna; Lingappa, Jairam R.; Magaret, Amalia S.; Wang, Richard S.; Mugo, Nelly; Mujugira, Andrew; Baeten, Jared M.; Mullins, James I.; Hughes, James P.; Bukusi, Elizabeth A.; Cohen, Craig R.; Katabira, Elly; Ronald, Allan; Kiarie, James; Farquhar, Carey; Stewart, Grace John; Makhema, Joseph; Essex, Myron; Were, Edwin; Fife, Kenneth H.; de Bruyn, Guy; Gray, Glenda E.; McIntyre, James A.; Manongi, Rachel; Kapiga, Saidi; Coetzee, David; Allen, Susan; Inambao, Mubiana; Kayitenkore, Kayitesi; Karita, Etienne; Kanweka, William; Delany, Sinead; Rees, Helen; Vwalika, Bellington; Stevens, Wendy; Campbell, Mary S.; Thomas, Katherine K.; Coombs, Robert W.; Morrow, Rhoda; Whittington, William L.H.; McElrath, M. Juliana; Barnes, Linda; Ridzon, Renee; Corey, Lawrence

    2010-01-01

    BACKGROUND Most persons who are infected with human immunodeficiency virus type 1 (HIV-1) are also infected with herpes simplex virus type 2 (HSV-2), which is frequently reactivated and is associated with increased plasma and genital levels of HIV-1. Therapy to suppress HSV-2 reduces the frequency of reactivation of HSV-2 as well as HIV-1 levels, suggesting that suppression of HSV-2 may reduce the risk of transmission of HIV-1. METHODS We conducted a randomized, placebo-controlled trial of suppressive therapy for HSV-2 (acyclovir at a dose of 400 mg orally twice daily) in couples in which only one of the partners was seropositive for HIV-1 (CD4 count, ≥250 cells per cubic millimeter) and that partner was also infected with HSV-2 and was not taking antiretroviral therapy at the time of enrollment. The primary end point was transmission of HIV-1 to the partner who was not initially infected with HIV-1; linkage of transmissions was assessed by means of genetic sequencing of viruses. RESULTS A total of 3408 couples were enrolled at 14 sites in Africa. Of the partners who were infected with HIV-1, 68% were women, and the baseline median CD4 count was 462 cells per cubic millimeter. Of 132 HIV-1 seroconversions that occurred after randomization (an incidence of 2.7 per 100 person-years), 84 were linked within couples by viral sequencing: 41 in the acyclovir group and 43 in the placebo group (hazard ratio with acyclovir, 0.92, 95% confidence interval [CI], 0.60 to 1.41; P = 0.69). Suppression with acyclovir reduced the mean plasma concentration of HIV-1 by 0.25 log10 copies per milliliter (95% CI, 0.22 to 0.29; P<0.001) and the occurrence of HSV-2–positive genital ulcers by 73% (risk ratio, 0.27; 95% CI, 0.20 to 0.36; P<0.001). A total of 92% of the partners infected with HIV-1 and 84% of the partners not infected with HIV-1 remained in the study for 24 months. The level of adherence to the dispensed study drug was 96%. No serious adverse events related to acyclovir

  3. Astrocytes sustain long-term productive HIV-1 infection without establishment of reactivable viral latency.

    Science.gov (United States)

    Barat, Corinne; Proust, Alizé; Deshiere, Alexandre; Leboeuf, Mathieu; Drouin, Jean; Tremblay, Michel J

    2018-02-21

    The "shock and kill" HIV-1 cure strategy proposes eradication of stable cellular reservoirs by clinical treatment with latency-reversing agents (LRAs). Although resting CD4 + T cells latently infected with HIV-1 constitute the main reservoir that is targeted by these approaches, their consequences on other reservoirs such as the central nervous system are still unknown and should be taken into consideration. We performed experiments aimed at defining the possible role of astrocytes in HIV-1 persistence in the brain and the effect of LRA treatments on this viral sanctuary. We first demonstrate that the diminished HIV-1 production in a proliferating astrocyte culture is due to a reduced proliferative capacity of virus-infected cells compared with uninfected astrocytes. In contrast, infection of non-proliferating astrocytes led to a robust HIV-1 infection that was sustained for over 60 days. To identify astrocytes latently infected with HIV-1, we designed a new dual-color reporter virus called NL4.3 eGFP-IRES-Crimson that is fully infectious and encodes for all viral proteins. Although we detected a small fraction of astrocytes carrying silent HIV-1 proviruses, we did not observe any reactivation using various LRAs and even strong inducers such as tumor necrosis factor, thus suggesting that these proviruses were either not transcriptionally competent or in a state of deep latency. Our findings imply that astrocytes might not constitute a latent reservoir per se but that relentless virus production by this brain cell population could contribute to the neurological disorders seen in HIV-1-infected persons subjected to combination antiretroviral therapy. © 2018 Wiley Periodicals, Inc.

  4. Nuclear retention of multiply spliced HIV-1 RNA in resting CD4+ T cells.

    Directory of Open Access Journals (Sweden)

    Kara G Lassen

    2006-07-01

    Full Text Available HIV-1 latency in resting CD4+ T cells represents a major barrier to virus eradication in patients on highly active antiretroviral therapy (HAART. We describe here a novel post-transcriptional block in HIV-1 gene expression in resting CD4+ T cells from patients on HAART. This block involves the aberrant localization of multiply spliced (MS HIV-1 RNAs encoding the critical positive regulators Tat and Rev. Although these RNAs had no previously described export defect, we show that they exhibit strict nuclear localization in resting CD4+ T cells from patients on HAART. Overexpression of the transcriptional activator Tat from non-HIV vectors allowed virus production in these cells. Thus, the nuclear retention of MS HIV-1 RNA interrupts a positive feedback loop and contributes to the non-productive nature of infection of resting CD4+ T cells. To define the mechanism of nuclear retention, proteomic analysis was used to identify proteins that bind MS HIV-1 RNA. Polypyrimidine tract binding protein (PTB was identified as an HIV-1 RNA-binding protein differentially expressed in resting and activated CD4+ T cells. Overexpression of PTB in resting CD4+ T cells from patients on HAART allowed cytoplasmic accumulation of HIV-1 RNAs. PTB overexpression also induced virus production by resting CD4+ T cells. Virus culture experiments showed that overexpression of PTB in resting CD4+ T cells from patients on HAART allowed release of replication-competent virus, while preserving a resting cellular phenotype. Whether through effects on RNA export or another mechanism, the ability of PTB to reverse latency without inducing cellular activation is a result with therapeutic implications.

  5. Ex vivo analysis identifies effective HIV-1 latency–reversing drug combinations

    Science.gov (United States)

    Laird, Gregory M.; Bullen, C. Korin; Rosenbloom, Daniel I.S.; Martin, Alyssa R.; Hill, Alison L.; Durand, Christine M.; Siliciano, Janet D.; Siliciano, Robert F.

    2015-01-01

    Reversal of HIV-1 latency by small molecules is a potential cure strategy. This approach will likely require effective drug combinations to achieve high levels of latency reversal. Using resting CD4+ T cells (rCD4s) from infected individuals, we developed an experimental and theoretical framework to identify effective latency-reversing agent (LRA) combinations. Utilizing ex vivo assays for intracellular HIV-1 mRNA and virion production, we compared 2-drug combinations of leading candidate LRAs and identified multiple combinations that effectively reverse latency. We showed that protein kinase C agonists in combination with bromodomain inhibitor JQ1 or histone deacetylase inhibitors robustly induce HIV-1 transcription and virus production when directly compared with maximum reactivation by T cell activation. Using the Bliss independence model to quantitate combined drug effects, we demonstrated that these combinations synergize to induce HIV-1 transcription. This robust latency reversal occurred without release of proinflammatory cytokines by rCD4s. To extend the clinical utility of our findings, we applied a mathematical model that estimates in vivo changes in plasma HIV-1 RNA from ex vivo measurements of virus production. Our study reconciles diverse findings from previous studies, establishes a quantitative experimental approach to evaluate combinatorial LRA efficacy, and presents a model to predict in vivo responses to LRAs. PMID:25822022

  6. HIV-1 Reservoir Association with Immune Activation

    Directory of Open Access Journals (Sweden)

    Alejandro Vallejo

    2015-09-01

    Full Text Available In this issue of EBioMedicine, Ruggiero and colleagues describe immune activation biomarkers associated with the size of the HIV reservoir in a carefully designed cross-sectional study. The cohort consists of a homogeneous sample of HIV-1-infected patients with long-term plasma HIV-1 RNA suppression under antiretroviral treatment (ART. It is crucial to explore the potential utility of biomarkers that are easier (less labor intensive, less expensive to measure than integrated HIV DNA load, in order to quickly and accurately quantify cellular reservoirs of HIV.

  7. MAS NMR of HIV-1 protein assemblies

    Science.gov (United States)

    Suiter, Christopher L.; Quinn, Caitlin M.; Lu, Manman; Hou, Guangjin; Zhang, Huilan; Polenova, Tatyana

    2015-04-01

    The negative global impact of the AIDS pandemic is well known. In this perspective article, the utility of magic angle spinning (MAS) NMR spectroscopy to answer pressing questions related to the structure and dynamics of HIV-1 protein assemblies is examined. In recent years, MAS NMR has undergone major technological developments enabling studies of large viral assemblies. We discuss some of these evolving methods and technologies and provide a perspective on the current state of MAS NMR as applied to the investigations into structure and dynamics of HIV-1 assemblies of CA capsid protein and of Gag maturation intermediates.

  8. Imperatorin inhibits HIV-1 replication through an Sp1-dependent pathway.

    Science.gov (United States)

    Sancho, Rocío; Márquez, Nieves; Gómez-Gonzalo, Marta; Calzado, Marco A; Bettoni, Giorgio; Coiras, Maria Teresa; Alcamí, José; López-Cabrera, Manuel; Appendino, Giovanni; Muñoz, Eduardo

    2004-09-03

    Coumarins and structurally related compounds have been recently shown to present anti-human immunodeficiency virus, type 1 (HIV-1) activity. Among them, the dietary furanocoumarin imperatorin is present in citrus fruits, in culinary herbs, and in some medicinal plants. In this study we report that imperatorin inhibits either vesicular stomatitis virus-pseudotyped or gp160-enveloped recombinant HIV-1 infection in several T cell lines and in HeLa cells. These recombinant viruses express luciferase as a marker of viral replication. Imperatorin did not inhibit the reverse transcription nor the integration steps in the viral cell cycle. Using several 5' long terminal repeat-HIV-1 constructs where critical response elements were either deleted or mutated, we found that the transcription factor Sp1 is critical for the inhibitory activity of imperatorin induced by both phorbol 12-myristate 13-acetate and HIV-1 Tat. Moreover in transient transfections imperatorin specifically inhibited phorbol 12-myristate 13-acetate-induced transcriptional activity of the Gal4-Sp1 fusion protein. Since Sp1 is also implicated in cell cycle progression we further studied the effect of imperatorin on cyclin D1 gene transcription and protein expression and in HeLa cell cycle progression. We found that imperatorin strongly inhibited cyclin D1 expression and arrested the cells at the G(1) phase of the cell cycle. These results highlight the potential of Sp1 transcription factor as a target for natural anti-HIV-1 compounds such as furanocoumarins that might have a potential therapeutic role in the management of AIDS.

  9. Viral linkage in HIV-1 seroconverters and their partners in an HIV-1 prevention clinical trial.

    Directory of Open Access Journals (Sweden)

    Mary S Campbell

    2011-03-01

    Full Text Available Characterization of viruses in HIV-1 transmission pairs will help identify biological determinants of infectiousness and evaluate candidate interventions to reduce transmission. Although HIV-1 sequencing is frequently used to substantiate linkage between newly HIV-1 infected individuals and their sexual partners in epidemiologic and forensic studies, viral sequencing is seldom applied in HIV-1 prevention trials. The Partners in Prevention HSV/HIV Transmission Study (ClinicalTrials.gov #NCT00194519 was a prospective randomized placebo-controlled trial that enrolled serodiscordant heterosexual couples to determine the efficacy of genital herpes suppression in reducing HIV-1 transmission; as part of the study analysis, HIV-1 sequences were examined for genetic linkage between seroconverters and their enrolled partners.We obtained partial consensus HIV-1 env and gag sequences from blood plasma for 151 transmission pairs and performed deep sequencing of env in some cases. We analyzed sequences with phylogenetic techniques and developed a Bayesian algorithm to evaluate the probability of linkage. For linkage, we required monophyletic clustering between enrolled partners' sequences and a Bayesian posterior probability of ≥ 50%. Adjudicators classified each seroconversion, finding 108 (71.5% linked, 40 (26.5% unlinked, and 3 (2.0% indeterminate transmissions, with linkage determined by consensus env sequencing in 91 (84%. Male seroconverters had a higher frequency of unlinked transmissions than female seroconverters. The likelihood of transmission from the enrolled partner was related to time on study, with increasing numbers of unlinked transmissions occurring after longer observation periods. Finally, baseline viral load was found to be significantly higher among linked transmitters.In this first use of HIV-1 sequencing to establish endpoints in a large clinical trial, more than one-fourth of transmissions were unlinked to the enrolled partner

  10. Molecular mechanisms of HIV-1 associated neurodegeneration

    Indian Academy of Sciences (India)

    Since identification of the human immunodeficiency virus-1 (HIV-1), numerous studies suggest a link between neurological impairments, in particular dementia, with acquired immunodeficiency syndrome (AIDS) with alarming occurrence worldwide. Approximately, 60% of HIV-infected people show some form of neurological ...

  11. Epidemiology of HIV-1 and emerging problems

    NARCIS (Netherlands)

    Lukashov, V. V.; de Ronde, A.; de Jong, J. J.; Goudsmit, J.

    2000-01-01

    Broad use of antiretroviral drugs is becoming a factor that is important to consider for understanding the HIV-1 epidemiology. Since 1993, we observe that a proportion of new infections within major risk groups in Amsterdam is caused by azidothymidine (AZT)-resistant viruses. After the introduction

  12. Interferon-α Subtypes in an Ex Vivo Model of Acute HIV-1 Infection: Expression, Potency and Effector Mechanisms.

    Directory of Open Access Journals (Sweden)

    Michael S Harper

    Full Text Available HIV-1 is transmitted primarily across mucosal surfaces and rapidly spreads within the intestinal mucosa during acute infection. The type I interferons (IFNs likely serve as a first line of defense, but the relative expression and antiviral properties of the 12 IFNα subtypes against HIV-1 infection of mucosal tissues remain unknown. Here, we evaluated the expression of all IFNα subtypes in HIV-1-exposed plasmacytoid dendritic cells by next-generation sequencing. We then determined the relative antiviral potency of each IFNα subtype ex vivo using the human intestinal Lamina Propria Aggregate Culture model. IFNα subtype transcripts from the centromeric half of the IFNA gene complex were highly expressed in pDCs following HIV-1 exposure. There was an inverse relationship between IFNA subtype expression and potency. IFNα8, IFNα6 and IFNα14 were the most potent in restricting HIV-1 infection. IFNα2, the clinically-approved subtype, and IFNα1 were both highly expressed but exhibited relatively weak antiviral activity. The relative potencies correlated with binding affinity to the type I IFN receptor and the induction levels of HIV-1 restriction factors Mx2 and Tetherin/BST-2 but not APOBEC3G, F and D. However, despite the lack of APOBEC3 transcriptional induction, the higher relative potency of IFNα8 and IFNα14 correlated with stronger inhibition of virion infectivity, which is linked to deaminase-independent APOBEC3 restriction activity. By contrast, both potent (IFNα8 and weak (IFNα1 subtypes significantly induced HIV-1 GG-to-AG hypermutation. The results unravel non-redundant functions of the IFNα subtypes against HIV-1 infection, with strong implications for HIV-1 mucosal immunity, viral evolution and IFNα-based functional cure strategies.

  13. Plectin regulates the signaling and trafficking of the HIV-1 co-receptor CXCR4 and plays a role in HIV-1 infection

    International Nuclear Information System (INIS)

    Ding Yun; Zhang Li; Goodwin, J. Shawn; Wang Ziqing; Liu Bingdong; Zhang Jingwu; Fan Guohuang

    2008-01-01

    The CXC chemokine CXCL12 and its cognate receptor CXCR4 play an important role in inflammation, human immunodeficiency virus (HIV) infection and cancer metastasis. The signal transduction and intracellular trafficking of CXCR4 are involved in these functions, but the underlying mechanisms remain incompletely understood. In the present study, we demonstrated that the CXCR4 formed a complex with the cytolinker protein plectin in a ligand-dependent manner in HEK293 cells stably expressing CXCR4. The glutathione-S-transferase (GST)-CXCR4 C-terminal fusion proteins co-precipitated with the full-length and the N-terminal fragments of plectin isoform 1 but not with the N-terminal deletion mutants of plectin isoform 1, thereby suggesting an interaction between the N-terminus of plectin and the C-terminus of CXCR4. This interaction was confirmed by confocal microscopic reconstructions showing co-distribution of these two proteins in the internal vesicles after ligand-induced internalization of CXCR4 in HEK293 cells stably expressing CXCR4. Knockdown of plectin with RNA interference (RNAi) significantly inhibited ligand-dependent CXCR4 internalization and attenuated CXCR4-mediated intracellular calcium mobilization and activation of extracellular signal regulated kinase 1/2 (ERK1/2). CXCL12-induced chemotaxis of HEK293 cells stably expressing CXCR4 and of Jurkat T cells was inhibited by the plectin RNAi. Moreover, CXCR4 tropic HIV-1 infection in MAGI (HeLa-CD4-LTR-Gal) cells was inhibited by the RNAi of plectin. Thus, plectin appears to interact with CXCR4 and plays an important role in CXCR4 signaling and trafficking and HIV-1 infection

  14. Inhibition of HIV-1 by curcumin A, a novel curcumin analog

    Science.gov (United States)

    Kumari, Namita; Kulkarni, Amol A; Lin, Xionghao; McLean, Charlee; Ammosova, Tatiana; Ivanov, Andrey; Hipolito, Maria; Nekhai, Sergei; Nwulia, Evaristus

    2015-01-01

    Despite the remarkable success of combination antiretroviral therapy at curtailing HIV progression, emergence of drug-resistant viruses, chronic low-grade inflammation, and adverse effects of combination antiretroviral therapy treatments, including metabolic disorders collectively present the impetus for development of newer and safer antiretroviral drugs. Curcumin, a phytochemical compound, was previously reported to have some in vitro anti-HIV and anti-inflammatory activities, but poor bioavailability has limited its clinical utility. To circumvent the bioavailability problem, we derivatized curcumin to sustain retro-aldol decomposition at physiological pH. The lead compound derived, curcumin A, showed increased stability, especially in murine serum where it was stable for up to 25 hours, as compared to curcumin that only had a half-life of 10 hours. Both curcumin and curcumin A showed similar inhibition of one round of HIV-1 infection in cultured lymphoblastoid (also called CEM) T cells (IC50=0.7 μM). But in primary peripheral blood mononuclear cells, curcumin A inhibited HIV-1 more potently (IC50=2 μM) compared to curcumin (IC50=12 μM). Analysis of specific steps of HIV-1 replication showed that curcumin A inhibited HIV-1 reverse transcription, but had no effect on HIV-1 long terminal repeat basal or Tat-induced transcription, or NF-κB-driven transcription at low concentrations that affected reverse transcription. Finally, we showed curcumin A induced expression of HO-1 and decreased cell cycle progression of T cells. Our findings thus indicate that altering the core structure of curcumin could yield more stable compounds with potent antiretroviral and anti-inflammatory activities. PMID:26366056

  15. Safety and immunogenicity of HIV-1 Tat toxoid in immunocompromised HIV-1-infected patients.

    Science.gov (United States)

    Gringeri, A; Santagostino, E; Muça-Perja, M; Mannucci, P M; Zagury, J F; Bizzini, B; Lachgar, A; Carcagno, M; Rappaport, J; Criscuolo, M; Blattner, W; Burny, A; Gallo, R C; Zagury, D

    1998-01-01

    To antagonize the deleterious effects of the HIV-1 toxin extracellular Tat on uninfected immune cells, we developed a new strategy of anti-HIV-1 vaccine using an inactivated but immunogenic Tat (Tat toxoid). Tat toxoid has been assayed for safety and immunogenicity in seropositive patients. The phase I vaccine clinical trial testing Tat toxoid preparation in Seppic Isa 51 oil adjuvant was performed on 14 HIV-1-infected asymptomatic although biologically immunocompromised individuals (500-200 CD4+ cells/mm3). Following as many as 8 injections, no clinical defects were observed. All patients exhibited an antibody (Ab) response to Tat, and some had cell-mediated immunity (CMI) as evaluated by skin test in vivo and T-cell proliferation in vitro. These results provide initial evidence of safety and potency of Tat toxoid vaccination in HIV-1-infected individuals.

  16. Suppression of APOBEC3-mediated restriction of HIV-1 by Vif

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    Yuqing eFeng

    2014-08-01

    Full Text Available The APOBEC3 restriction factors are a family of deoxycytidine deaminases that are able to suppress replication of viruses with a single-stranded DNA intermediate by inducing mutagenesis and functional inactivation of the virus. Of the seven human APOBEC3 enzymes, only APOBEC3-D, -F, -G, and -H appear relevant to restriction of HIV-1 in CD4+ T cells and will be the focus of this review. The restriction of HIV-1 occurs most potently in the absence of HIV-1 Vif that induces polyubiquitination and degradation of APOBEC3 enzymes through the proteasome pathway. To restrict HIV-1, APOBEC3 enzymes must be encapsidated into budding virions. Upon infection of the target cell during reverse transcription of the HIV-1 RNA into (-DNA APOBEC3 enzymes deaminate cytosines to forms uracils in single-stranded (- DNA regions. Upon replication of the (-DNA to (+DNA, the HIV-1 reverse transcriptase incorporates adenines opposite the uracils thereby inducing C/G to T/A mutations that can functionally inactivate HIV-1. APOBEC3G is the most studied APOBEC3 enzyme and it is known that Vif attempts to thwart APOBEC3 function not only by inducing its proteasomal degradation but by several degradation-independent mechanisms such as inhibiting APOBEC3G virion encapsidation, mRNA translation, and for those APOBEC3G molecules that still become virion encapsidated, Vif can inhibit APOBEC3G mutagenic activity. Although most Vif variants can induce efficient degradation of APOBEC3-D, -F, and -G, there appears to be differential sensitivity to Vif-mediated degradation for APOBEC3H. This review examines APOBEC3-mediated HIV restriction mechanisms, how Vif acts as a substrate receptor for a Cullin5 ubiquitin ligase complex to induce degradation of APOBEC3s, and the determinants and functional consequences of the APOBEC3 and Vif interaction from a biological and biochemical perspective.

  17. Picomolar dichotomous activity of gnidimacrin against HIV-1.

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    Li Huang

    Full Text Available Highly active antiretroviral therapy (HAART has offered a promising approach for controlling HIV-1 replication in infected individuals. However, with HARRT, HIV-1 is suppressed rather than eradicated due to persistence of HIV-1 in latent viral reservoirs. Thus, purging the virus from latent reservoirs is an important strategy toward eradicating HIV-1 infection. In this study, we discovered that the daphnane diterpene gnidimacrin, which was previously reported to have potent anti-cancer cell activity, activated HIV-1 replication and killed persistently-infected cells at picomolar concentrations. In addition to its potential to purge HIV-1 from latently infected cells, gnidimacrin potently inhibited a panel of HIV-1 R5 virus infection of peripheral blood mononuclear cells (PBMCs at an average concentration lower than 10 pM. In contrast, gnidimacrin only partially inhibited HIV-1 ×4 virus infection of PBMCs. The strong anti-HIV-1 R5 virus activity of gnidimacrin was correlated with its effect on down-regulation of the HIV-1 coreceptor CCR5. The anti-R5 virus activity of gnidimacrin was completely abrogated by a selective protein kinase C beta inhibitor enzastaurin, which suggests that protein kinase C beta plays a key role in the potent anti-HIV-1 activity of gnidimacrin in PBMCs. In summary, these results suggest that gnidimacrin could activate latent HIV-1, specifically kill HIV-1 persistently infected cells, and inhibit R5 viruses at picomolar concentrations.

  18. Demonstration of a novel HIV-1 restriction phenotype from a human T cell line.

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    Yanxing Han

    2008-07-01

    Full Text Available Although retroviruses may invade host cells, a productive infection can be established only after the virus counteracts inhibition from different types of host restriction factors. Fv1, APOBEC3G/F, TRIM5alpha, ZAP, and CD317 inhibit the replication of different retroviruses by interfering with viral uncoating, reverse transcription, nuclear import, RNA stability, and release. In humans, although APOBEC3G/3F and CD317 block HIV-1 replication, their antiviral activities are neutralized by viral proteins Vif and Vpu. So far, no human gene has been found to effectively block wild type HIV-1 replication under natural condition. Thus, identification of such a gene product would be of great medical importance for the development of HIV therapies.In this study, we discovered a new type of host restriction against the wild type HIV-1 from a CD4/CXCR4 double-positive human T cell line. We identified a CEM-derived cell line (CEM.NKR that is highly resistant to productive HIV-1 infection. Viral production was reduced by at least 1000-fold when compared to the other permissive human T cell lines such as H9, A3.01, and CEM-T4. Importantly, this resistance was evident at extremely high multiplicity of infection. Further analyses demonstrated that HIV-1 could finish the first round of replication in CEM.NKR cells, but the released virions were poorly infectious. These virions could enter the target cells, but failed to initiate reverse transcription. Notably, this restriction phenotype was also present in CEM.NKR and 293T heterokaryons.These results clearly indicate that CEM.NKR cells express a HIV inhibitory gene(s. Further characterization of this novel gene product(s will reveal a new antiretroviral mechanism that directly inactivates wild type HIV-1.

  19. Interactions between the HIV-1 Unspliced mRNA and Host mRNA Decay Machineries

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    Daniela Toro-Ascuy

    2016-11-01

    Full Text Available The human immunodeficiency virus type-1 (HIV-1 unspliced transcript is used both as mRNA for the synthesis of structural proteins and as the packaged genome. Given the presence of retained introns and instability AU-rich sequences, this viral transcript is normally retained and degraded in the nucleus of host cells unless the viral protein REV is present. As such, the stability of the HIV-1 unspliced mRNA must be particularly controlled in the nucleus and the cytoplasm in order to ensure proper levels of this viral mRNA for translation and viral particle formation. During its journey, the HIV-1 unspliced mRNA assembles into highly specific messenger ribonucleoproteins (mRNPs containing many different host proteins, amongst which are well-known regulators of cytoplasmic mRNA decay pathways such as up-frameshift suppressor 1 homolog (UPF1, Staufen double-stranded RNA binding protein 1/2 (STAU1/2, or components of miRNA-induced silencing complex (miRISC and processing bodies (PBs. More recently, the HIV-1 unspliced mRNA was shown to contain N6-methyladenosine (m6A, allowing the recruitment of YTH N6-methyladenosine RNA binding protein 2 (YTHDF2, an m6A reader host protein involved in mRNA decay. Interestingly, these host proteins involved in mRNA decay were shown to play positive roles in viral gene expression and viral particle assembly, suggesting that HIV-1 interacts with mRNA decay components to successfully accomplish viral replication. This review summarizes the state of the art in terms of the interactions between HIV-1 unspliced mRNA and components of different host mRNA decay machineries.

  20. Synthetic AAV/CRISPR vectors for blocking HIV-1 expression in persistently infected astrocytes.

    Science.gov (United States)

    Kunze, Christine; Börner, Kathleen; Kienle, Eike; Orschmann, Tanja; Rusha, Ejona; Schneider, Martha; Radivojkov-Blagojevic, Milena; Drukker, Micha; Desbordes, Sabrina; Grimm, Dirk; Brack-Werner, Ruth

    2018-02-01

    Astrocytes, the most abundant cells in the mammalian brain, perform key functions and are involved in several neurodegenerative diseases. The human immunodeficiency virus (HIV) can persist in astrocytes, contributing to the HIV burden and neurological dysfunctions in infected individuals. While a comprehensive approach to HIV cure must include the targeting of HIV-1 in astrocytes, dedicated tools for this purpose are still lacking. Here we report a novel Adeno-associated virus-based vector (AAV9P1) with a synthetic surface peptide for transduction of astrocytes. Analysis of AAV9P1 transduction efficiencies with single brain cell populations, including primary human brain cells, as well as human brain organoids demonstrated that AAV9P1 targeted terminally differentiated human astrocytes much more efficiently than neurons. We then investigated whether AAV9P1 can be used to deliver HIV-inhibitory genes to astrocytes. To this end we generated AAV9P1 vectors containing genes for HIV-1 proviral editing by CRISPR/Cas9. Latently HIV-1 infected astrocytes transduced with these vectors showed significantly diminished reactivation of proviruses, compared with untransduced cultures. Sequence analysis identified mutations/deletions in key HIV-1 transcriptional control regions. We conclude that AAV9P1 is a promising tool for gene delivery to astrocytes and may facilitate inactivation/destruction of persisting HIV-1 proviruses in astrocyte reservoirs. © 2017 Wiley Periodicals, Inc.

  1. The global transmission network of HIV-1.

    Science.gov (United States)

    Wertheim, Joel O; Leigh Brown, Andrew J; Hepler, N Lance; Mehta, Sanjay R; Richman, Douglas D; Smith, Davey M; Kosakovsky Pond, Sergei L

    2014-01-15

    Human immunodeficiency virus type 1 (HIV-1) is pandemic, but its contemporary global transmission network has not been characterized. A better understanding of the properties and dynamics of this network is essential for surveillance, prevention, and eventual eradication of HIV. Here, we apply a simple and computationally efficient network-based approach to all publicly available HIV polymerase sequences in the global database, revealing a contemporary picture of the spread of HIV-1 within and between countries. This approach automatically recovered well-characterized transmission clusters and extended other clusters thought to be contained within a single country across international borders. In addition, previously undescribed transmission clusters were discovered. Together, these clusters represent all known modes of HIV transmission. The extent of international linkage revealed by our comprehensive approach demonstrates the need to consider the global diversity of HIV, even when describing local epidemics. Finally, the speed of this method allows for near-real-time surveillance of the pandemic's progression.

  2. Human vaginal fluid contains exosomes that have an inhibitory effect on an early step of the HIV-1 life cycle.

    Science.gov (United States)

    Smith, Johanna A; Daniel, Rene

    2016-11-13

    Vaginal transmission is crucial to the spread of HIV-1 around the world. It is not yet clear what type (s) of innate defenses against HIV-1 infection are present in the vagina. Here, we aimed to determine whether human vaginal fluid contains exosomes that may possess anti-HIV-1 activity. The exosomal fraction was isolated from samples of vaginal fluids. The presence of exosomes was confirmed by flow cytometry and western blotting. The newly discovered exosomes were tested for their ability to block early steps of HIV-1 infection in vitro using established cell culture systems and real time PCR-based methods. Vaginal fluid contains exosomes expressing CD9, CD63, and CD81 exosomal markers. The exosomal fraction of the fluid-reduced transmission of HIV-1 vectors by 60%, the efficiency of reverse transcription step by 58.4%, and the efficiency of integration by 47%. Exosomes had no effect on the entry of HIV-1 vectors. Human vaginal fluid exosomes are newly discovered female innate defenses that may protect women against HIV-1 infection.

  3. Morphogenesis of the infectious HIV-1 virion

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    Jun-Ichi eSakuragi

    2011-12-01

    Full Text Available The virion of HIV-1 is spherical and viral glycoprotein spikes (gp120, gp41 protrude from its envelope. The characteristic cone-shaped core exists within the virion, caging the ribonucleoprotein (RNP complex, which is comprised of viral RNA, nucleocapsid (NC and viral enzymes. The HIV-1 virion is budded and released from the infected cell as an immature donut-shaped particle. During or immediately after release, viral protease (PR is activated and subsequently processes the viral structural protein Gag. Through this maturation process, virions acquire infectivity, but its mechanism and transition of morphology largely remain unclear. Recent technological advances in experimental devices and techniques have made it possible to closely dissect the viral production site on the cell, the exterior – or even the interior – of an individual virion, and many new aspects on virion morphology and maturation. In this manuscript, I review the morphogenesis of HIV-1 virions. I focus on several studies, including some of our recent findings, which examined virion formation and/or maturation processes. The story of novel compound, which inhibits virion maturation, and the importance of maturation research are also discussed.

  4. HIV-1 Eradication Strategies: Design and Assessment

    Science.gov (United States)

    Siliciano, Robert F.

    2014-01-01

    Purpose of review Recent developments have generated renewed interest in the possibility of curing HIV-1 infection. This review describes some of the practical challenges that will need to be overcome if curative strategies are to be successful. Recent findings The latent reservoir for HIV-1 in resting memory CD4+ T cells is the major barrier to curing the infection. The most widely discussed approach to curing the infection involves finding agents that reverse latency in resting CD4+ T cells, with the assumption that the cells will then die from viral cytopathic effects or be lysed by host cytolytic T lymphocytes (CTL). A major challenge is the development of in vitro models that can be used to explore mechanisms and identify latency reversing agents (LRA). Although several models have been developed, including primary cell models, none of them may fully capture the quiescent state of the cells that harbor latent HIV-1 in vivo. An additional problem is that LRA that do not cause T cell activation may not lead to the death of infected cells. Finally, measuring the effects of LRAs in vivo is complicated by the lack of correlation between different assays for the latent reservoir. Summary Progress on these practical issues is essential to finding a cure. PMID:23698561

  5. Potent Inhibition of HIV-1 Replication in Resting CD4 T Cells by Resveratrol and Pterostilbene.

    Science.gov (United States)

    Chan, Chi N; Trinité, Benjamin; Levy, David N

    2017-09-01

    HIV-1 infection of resting CD4 T cells plays a crucial and numerically dominant role during virus transmission at mucosal sites and during subsequent acute replication and T cell depletion. Resveratrol and pterostilbene are plant stilbenoids associated with several health-promoting benefits. Resveratrol has been shown to inhibit the replication of several viruses, including herpes simplex viruses 1 and 2, papillomaviruses, severe acute respiratory syndrome virus, and influenza virus. Alone, resveratrol does not inhibit HIV-1 infection of activated T cells, but it does synergize with nucleoside reverse transcriptase inhibitors in these cells to inhibit reverse transcription. Here, we demonstrate that resveratrol and pterostilbene completely block HIV-1 infection at a low micromolar dose in resting CD4 T cells, primarily at the reverse transcription step. The anti-HIV effect was fully reversed by exogenous deoxynucleosides and Vpx, an HIV-1 and simian immunodeficiency virus protein that increases deoxynucleoside triphosphate (dNTP) levels. These findings are consistent with the reported ability of resveratrol to inhibit ribonucleotide reductase and to lower dNTP levels in cells. This study supports the potential use of resveratrol, pterostilbene, or related compounds as adjuvants in anti-HIV preexposure prophylaxis (PrEP) formulations. Copyright © 2017 American Society for Microbiology.

  6. Requirements for capsid-binding and an effector function in TRIMCyp-mediated restriction of HIV-1

    International Nuclear Information System (INIS)

    Diaz-Griffero, Felipe; Vandegraaff, Nick; Li Yuan; McGee-Estrada, Kathleen; Stremlau, Matthew; Welikala, Sohanya; Si Zhihai; Engelman, Alan; Sodroski, Joseph

    2006-01-01

    In owl monkeys, a retrotransposition event replaced the gene encoding the retroviral restriction factor TRIM5α with one encoding TRIMCyp, a fusion between the RING, B-box 2 and coiled-coil domains of TRIM5 and cyclophilin A. TRIMCyp restricts human immunodeficiency virus (HIV-1) infection by a mechanism dependent on the interaction of the cyclophilin A moiety and the HIV-1 capsid protein. Here, we show that infection by retroviruses other than HIV-1 can be restricted by TRIMCyp, providing an explanation for the evolutionary retention of the TRIMCyp gene in owl monkey lineages. The TRIMCyp-mediated block to HIV-1 infection occurs before the earliest step of reverse transcription. TRIMCyp-mediated restriction involves at least two functions: (1) capsid binding, which occurs most efficiently for trimeric TRIMCyp proteins that retain the coiled-coil and cyclophilin A domains, and (2) an effector function that depends upon the B-box 2 domain

  7. Recruitment of a SAP18-HDAC1 complex into HIV-1 virions and its requirement for viral replication.

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    Masha Sorin

    2009-06-01

    Full Text Available HIV-1 integrase (IN is a virally encoded protein required for integration of viral cDNA into host chromosomes. INI1/hSNF5 is a component of the SWI/SNF complex that interacts with HIV-1 IN, is selectively incorporated into HIV-1 (but not other retroviral virions, and modulates multiple steps, including particle production and infectivity. To gain further insight into the role of INI1 in HIV-1 replication, we screened for INI1-interacting proteins using the yeast two-hybrid system. We found that SAP18 (Sin3a associated protein 18 kD, a component of the Sin3a-HDAC1 complex, directly binds to INI1 in yeast, in vitro and in vivo. Interestingly, we found that IN also binds to SAP18 in vitro and in vivo. SAP18 and components of a Sin3A-HDAC1 complex were specifically incorporated into HIV-1 (but not SIV and HTLV-1 virions in an HIV-1 IN-dependent manner. Using a fluorescence-based assay, we found that HIV-1 (but not SIV virion preparations harbour significant deacetylase activity, indicating the specific recruitment of catalytically active HDAC into the virions. To determine the requirement of virion-associated HDAC1 to HIV-1 replication, an inactive, transdominant negative mutant of HDAC1 (HDAC1(H141A was utilized. Incorporation of HDAC1(H141A decreased the virion-associated histone deacetylase activity. Furthermore, incorporation of HDAC1(H141A decreased the infectivity of HIV-1 (but not SIV virions. The block in infectivity due to virion-associated HDAC1(H141A occurred specifically at the early reverse transcription stage, while entry of the virions was unaffected. RNA-interference mediated knock-down of HDAC1 in producer cells resulted in decreased virion-associated HDAC1 activity and a reduction in infectivity of these virions. These studies indicate that HIV-1 IN and INI1/hSNF5 bind SAP18 and selectively recruit components of Sin3a-HDAC1 complex into HIV-1 virions. Furthermore, HIV-1 virion-associated HDAC1 is required for efficient early post

  8. Human endogenous retrovirus K Gag coassembles with HIV-1 Gag and reduces the release efficiency and infectivity of HIV-1.

    Science.gov (United States)

    Monde, Kazuaki; Contreras-Galindo, Rafael; Kaplan, Mark H; Markovitz, David M; Ono, Akira

    2012-10-01

    Human endogenous retroviruses (HERVs), which are remnants of ancestral retroviruses integrated into the human genome, are defective in viral replication. Because activation of HERV-K and coexpression of this virus with HIV-1 have been observed during HIV-1 infection, it is conceivable that HERV-K could affect HIV-1 replication, either by competition or by cooperation, in cells expressing both viruses. In this study, we found that the release efficiency of HIV-1 Gag was 3-fold reduced upon overexpression of HERV-K(CON) Gag. In addition, we observed that in cells expressing Gag proteins of both viruses, HERV-K(CON) Gag colocalized with HIV-1 Gag at the plasma membrane. Furthermore, HERV-K(CON) Gag was found to coassemble with HIV-1 Gag, as demonstrated by (i) processing of HERV-K(CON) Gag by HIV-1 protease in virions, (ii) coimmunoprecipitation of virion-associated HERV-K(CON) Gag with HIV-1 Gag, and (iii) rescue of a late-domain-defective HERV-K(CON) Gag by wild-type (WT) HIV-1 Gag. Myristylation-deficient HERV-K(CON) Gag localized to nuclei, suggesting cryptic nuclear trafficking of HERV-K Gag. Notably, unlike WT HERV-K(CON) Gag, HIV-1 Gag failed to rescue myristylation-deficient HERV-K(CON) Gag to the plasma membrane. Efficient colocalization and coassembly of HIV-1 Gag and HERV-K Gag also required nucleocapsid (NC). These results provide evidence that HIV-1 Gag heteromultimerizes with HERV-K Gag at the plasma membrane, presumably through NC-RNA interaction. Intriguingly, HERV-K Gag overexpression reduced not only HIV-1 release efficiency but also HIV-1 infectivity in a myristylation- and NC-dependent manner. Altogether, these results indicate that Gag proteins of endogenous retroviruses can coassemble with HIV-1 Gag and modulate the late phase of HIV-1 replication.

  9. The RNA helicase DDX1 is involved in restricted HIV-1 Rev function in human astrocytes

    International Nuclear Information System (INIS)

    Fang Jianhua; Acheampong, Edward; Dave, Rajnish; Wang Fengxiang; Mukhtar, Muhammad; Pomerantz, Roger J.

    2005-01-01

    Productive infection by human immunodeficiency virus type I (HIV-1) in the central nervous system (CNS) involves mainly macrophages and microglial cells. A frequency of less than 10% of human astrocytes is estimated to be infectable with HIV-1. Nonetheless, this relatively low percentage of infected astrocytes, but associated with a large total number of astrocytic cells in the CNS, makes human astrocytes a critical part in the analyses of potential HIV-1 reservoirs in vivo. Investigations in astrocytic cell lines and primary human fetal astrocytes revealed that limited HIV-1 replication in these cells resulted from low-level viral entry, transcription, viral protein processing, and virion maturation. Of note, a low ratio of unspliced versus spliced HIV-1-specific RNA was also investigated, as Rev appeared to act aberrantly in astrocytes, via loss of nuclear and/or nucleolar localization and diminished Rev-mediated function. Host cellular machinery enabling Rev function has become critical for elucidation of diminished Rev activity, especially for those factors leading to RNA metabolism. We have recently identified a DEAD-box protein, DDX1, as a Rev cellular co-factor and now have explored its potential importance in astrocytes. Cells were infected with HIV-1 pseudotyped with envelope glycoproteins of amphotropic murine leukemia viruses (MLV). Semi-quantitative reverse transcriptase-polymerase chain reactions (RT-PCR) for unspliced, singly-spliced, and multiply-spliced RNA clearly showed a lower ratio of unspliced/singly-spliced over multiply-spliced HIV-1-specific RNA in human astrocytes as compared to Rev-permissive, non-glial control cells. As well, the cellular localization of Rev in astrocytes was cytoplasmically dominant as compared to that of Rev-permissive, non-glial controls. This endogenous level of DDX1 expression in astrocytes was demonstrated directly to lead to a shift of Rev sub-cellular distribution dominance from nuclear and/or nucleolar to

  10. HIV-1 phylogenetic analysis shows HIV-1 transits through the meninges to brain and peripheral tissues.

    Science.gov (United States)

    Lamers, Susanna L; Gray, Rebecca R; Salemi, Marco; Huysentruyt, Leanne C; McGrath, Michael S

    2011-01-01

    Brain infection by the human immunodeficiency virus type 1 (HIV-1) has been investigated in many reports with a variety of conclusions concerning the time of entry and degree of viral compartmentalization. To address these diverse findings, we sequenced HIV-1 gp120 clones from a wide range of brain, peripheral and meningeal tissues from five patients who died from several HIV-1 associated disease pathologies. High-resolution phylogenetic analysis confirmed previous studies that showed a significant degree of compartmentalization in brain and peripheral tissue subpopulations. Some intermixing between the HIV-1 subpopulations was evident, especially in patients that died from pathologies other than HIV-associated dementia. Interestingly, the major tissue harboring virus from both the brain and peripheral tissues was the meninges. These results show that (1) HIV-1 is clearly capable of migrating out of the brain, (2) the meninges are the most likely primary transport tissues, and (3) infected brain macrophages comprise an important HIV reservoir during highly active antiretroviral therapy. Copyright © 2010 Elsevier B.V. All rights reserved.

  11. Crosstalk between HDAC6 and Nox2-based NADPH oxidase mediates HIV-1 Tat-induced pro-inflammatory responses in astrocytes

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    Gi Soo Youn

    2017-08-01

    Full Text Available Histone deacetylase 6 (HDAC6 likely is important in inflammatory diseases. However, how HDAC6 exerts its effect on inflammatory processes remains unclear. HIV-1 transactivator of transcription (Tat activates NADPH oxidase resulting in generation of reactive oxygen species (ROS, leading to extensive neuro-inflammation in the central nervous system. We investigated the correlation of HDAC6 and NADPH oxidase in HIV-1 Tat-stimulated astrocytes. HDAC6 knockdown attenuated HIV-1 Tat-induced ROS generation and NADPH oxidase activation. HDAC6 knockdown suppressed HIV-1 Tat-induced expression of NADPH oxidase subunits, such as Nox2, p47phox, and p22phox. Specific inhibition of HDAC6 using tubastatin A suppressed HIV-1 Tat-induced ROS generation and activation of NADPH oxidase. N-acetyl cysteine, diphenyl iodonium, and apocynin suppressed HIV-1 Tat-induced expression of HDAC6 and the pro-inflammatory chemokines CCL2, CXCL8, and CXCL10. Nox2 knockdown attenuated HIV-1 Tat-induced HDAC6 expression and subsequent expression of chemokines. The collective results point to the potential crosstalk between HDAC6 and NADPH oxidase, which could be a combined therapeutic target for relief of HIV-1 Tat-mediated neuro-inflammation. Keywords: HIV-1 Tat, HDAC6, NADPH oxidase, ROS, Inflammation, Astrocytes

  12. Cyclophilin A potentiates TRIM5α inhibition of HIV-1 nuclear import without promoting TRIM5α binding to the viral capsid.

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    Mallori Burse

    Full Text Available The host immunophilin cyclophilin A (CypA binds to the capsid protein (CA of HIV-1 and regulates its infectivity. Depending on the target cell type, CypA can either promote or inhibit HIV-1 infection. The ability of CypA to promote HIV-1 infection has been extensively studied and linked to several steps in early replication including uncoating, reverse transcription and nuclear import. By contrast, the mechanism by which CypA inhibits infection is less well understood. We investigated the mechanism by which CypA potentiates restriction of HIV-1 by the tripartite motif-containing protein 5 (TRIM5α. Depletion of TRIM5α in the African green monkey cell line Vero, resulted in a loss of inhibition of infection by CypA, demonstrating that inhibition by CypA is mediated by TRIM5α. Complementary genetic and biochemical assays failed to demonstrate an ability of CypA to promote binding of TRIM5α to the viral capsid. TRIM5α inhibits HIV-1 reverse transcription in a proteasome-dependent manner; however, we observed that inhibition of proteasome activity did not reduce the ability of CypA to inhibit infection, suggesting that CypA acts at a step after reverse transcription. Accordingly, we observed a CypA-dependent reduction in the accumulation of nuclear HIV-1 DNA, indicating that CypA specifically promotes TRIM5α inhibition of HIV-1 nuclear import. We also observed that the ability of CypA to inhibit HIV-1 infection is abolished by amino acid substitutions within the conserved CPSF6-binding surface in CA. Our results indicate that CypA inhibits HIV-1 infection in Vero cells not by promoting TRIM5α binding to the capsid but by blocking nuclear import of the HIV-1 preintegration complex.

  13. A Real Time PCR Platform for the Simultaneous Quantification of Total and Extrachromosomal HIV DNA Forms in Blood of HIV-1 Infected Patients

    Science.gov (United States)

    Canovari, Benedetta; Scotti, Maddalena; Acetoso, Marcello; Valentini, Massimo; Petrelli, Enzo; Magnani, Mauro

    2014-01-01

    Background The quantitative measurement of various HIV-1 DNA forms including total, unintegrated and integrated provirus play an increasingly important role in HIV-1 infection monitoring and treatment-related research. We report the development and validation of a SYBR Green real time PCR (TotUFsys platform) for the simultaneous quantification of total and extrachromosomal HIV-1 DNA forms in patients. This innovative technique makes it possible to obtain both measurements in a single PCR run starting from frozen blood employing the same primers and standard curve. Moreover, due to identical amplification efficiency, it allows indirect estimation of integrated level. To specifically detect 2-LTR a qPCR method was also developed. Methodology/Findings Primers used for total HIV-1 DNA quantification spanning a highly conserved region were selected and found to detect all HIV-1 clades of group M and the unintegrated forms of the same. A total of 195 samples from HIV-1 patients in a wide range of clinical conditions were analyzed with a 100% success rate, even in patients with suppressed plasma viremia, regardless of CD4+ or therapy. No significant correlation was observed between the two current prognostic markers, CD4+ and plasma viremia, while a moderate or high inverse correlation was found between CD4+ and total HIV DNA, with strong values for unintegrated HIV DNA. Conclusions/Significance Taken together, the results support the use of HIV DNA as another tool, in addition to traditional assays, which can be used to estimate the state of viral infection, the risk of disease progression and to monitor the effects of ART. The TotUFsys platform allowed us to obtain a final result, expressed as the total and unintegrated HIV DNA copy number per microgram of DNA or 104 CD4+, for 12 patients within two working days. PMID:25364909

  14. Selection of reliable reference genes for gene expression studies in Trichoderma afroharzianum LTR-2 under oxalic acid stress.

    Science.gov (United States)

    Lyu, Yuping; Wu, Xiaoqing; Ren, He; Zhou, Fangyuan; Zhou, Hongzi; Zhang, Xinjian; Yang, Hetong

    2017-10-01

    An appropriate reference gene is required to get reliable results from gene expression analysis by quantitative real-time reverse transcription PCR (qRT-PCR). In order to identify stable and reliable reference genes in Trichoderma afroharzianum under oxalic acid (OA) stress, six commonly used housekeeping genes, i.e., elongation factor 1, ubiquitin, ubiquitin-conjugating enzyme, glyceraldehyde-3-phosphate dehydrogenase, α-tubulin, actin, from the effective biocontrol isolate T. afroharzianum strain LTR-2 were tested for their expression during growth in liquid culture amended with OA. Four in silico programs (comparative ΔCt, NormFinder, geNorm and BestKeeper) were used to evaluate the expression stabilities of six candidate reference genes. The elongation factor 1 gene EF-1 was identified as the most stably expressed reference gene, and was used as the normalizer to quantify the expression level of the oxalate decarboxylase coding gene OXDC in T. afroharzianum strain LTR-2 under OA stress. The result showed that the expression of OXDC was significantly up-regulated as expected. This study provides an effective method to quantify expression changes of target genes in T. afroharzianum under OA stress. Copyright © 2017 Elsevier B.V. All rights reserved.

  15. Increased Risk of HIV-1 Transmission in Pregnancy: A Prospective Study among African HIV-1 Serodiscordant Couples

    Science.gov (United States)

    MUGO, Nelly R.; HEFFRON, Renee; DONNELL, Deborah; WALD, Anna; WERE, Edwin O.; REES, Helen; CELUM, Connie; KIARIE, James N.; COHEN, Craig R.; KAYINTEKORE, Kayitesi; BAETEN, Jared M.

    2011-01-01

    Background Physiologic and behavioral changes during pregnancy may alter HIV-1 susceptibility and infectiousness. Prospective studies exploring pregnancy and HIV-1 acquisition risk in women have found inconsistent results. No study has explored the effect of pregnancy on HIV-1 transmission risk from HIV-1 infected women to male partners. Methods In a prospective study of African HIV-1 serodiscordant couples, we evaluated the relationship between pregnancy and the risk of 1) HIV-1 acquisition among women and 2) HIV-1 transmission from women to men. Results 3321 HIV-1 serodiscordant couples were enrolled, 1085 (32.7%) with HIV-1 susceptible female partners and 2236 (67.3%) with susceptible male partners. HIV-1 incidence in women was 7.35 versus 3.01 per 100 person-years during pregnant and non-pregnant periods (hazard ratio [HR] 2.34, 95% confidence interval [CI] 1.33–4.09). This effect was attenuated and not statistically significant after adjusting for sexual behavior and other confounding factors (adjusted HR 1.71, 95% CI 0.93–3.12). HIV-1 incidence in male partners of infected women was 3.46 versus 1.58 per 100 person-years when their partners were pregnant versus not pregnant (HR 2.31, 95% CI 1.22–4.39). This effect was not attenuated in adjusted analysis (adjusted HR 2.47, 95% CI 1.26–4.85). Conclusions HIV-1 risk increased two-fold during pregnancy. Elevated risk of HIV-1 acquisition in pregnant women appeared in part to be explained by behavioral and other factors. This is the first study to show pregnancy increased the risk of female-to-male HIV-1 transmission, which may reflect biological changes of pregnancy that could increase HIV-1 infectiousness. PMID:21785321

  16. Human CNS cultures exposed to HIV-1 gp120 reproduce dendritic injuries of HIV-1-associated dementia

    Directory of Open Access Journals (Sweden)

    Hammond Robert R

    2004-05-01

    Full Text Available Abstract HIV-1-associated dementia remains a common subacute to chronic central nervous system degeneration in adult and pediatric HIV-1 infected populations. A number of viral and host factors have been implicated including the HIV-1 120 kDa envelope glycoprotein (gp120. In human post-mortem studies using confocal scanning laser microscopy for microtubule-associated protein 2 and synaptophysin, neuronal dendritic pathology correlated with dementia. In the present study, primary human CNS cultures exposed to HIV-1 gp120 at 4 weeks in vitro suffered gliosis and dendritic damage analogous to that described in association with HIV-1-associated dementia.

  17. HIV-1 genetic diversity and its distribution characteristics among newly diagnosed HIV-1 individuals in Hebei province, China.

    Science.gov (United States)

    Lu, Xinli; Zhao, Cuiying; Wang, Wei; Nie, Chenxi; Zhang, Yuqi; Zhao, Hongru; Chen, Suliang; Cui, Ze

    2016-01-01

    Since the first HIV-1 case in 1989, Hebei province has presented a clearly rising trend of HIV-1 prevalence, and HIV-1 genetic diversity has become the vital barrier to HIV prevention and control in this area. To obtain detailed information of HIV-1 spread in different populations and in different areas of Hebei, a cross-sectional HIV-1 molecular epidemiological investigation was performed across the province. Blood samples of 154 newly diagnosed HIV-1 individuals were collected from ten prefectures in Hebei using stratified sampling. Partial gag and env genes were amplified and sequenced. HIV-1 genotypes were identified by phylogenetic tree analyses. Among the 139 subjects genotyped, six HIV-1 subtypes were identified successfully, including subtype B (41.0 %), CRF01_AE (40.3 %), CRF07_BC (11.5 %), CRF08_BC (4.3 %), unique recombinant forms (URFs) (1.4 %) and subtype C (1.4 %). Subtype B was identified as the most frequent subtype. Two URF recombination patterns were the same as CRF01_AE/B. HIV-1 genotype distribution showed a significant statistical difference in different demographic characteristics, such as source (P  0.05). The differences in HIV-1 genotype distribution were closely associated with transmission routes. Particularly, all six subtype strains were found in heterosexuals, showing that HIV-1 has spread from the high-risk populations to the general populations in Hebei, China. In addition, CRF01_AE instead of subtype B has become the major strain of HIV-1 infection among homosexuals. Our study revealed HIV-1 evolution and genotype distribution by investigating newly diagnosed HIV-1 individuals in Hebei, China. This study provides important information to enhance the strategic plan for HIV prevention and control in China.

  18. Methamphetamine inhibits HIV-1 replication in CD4+ T cells by modulating anti-HIV-1 miRNA expression.

    Science.gov (United States)

    Mantri, Chinmay K; Mantri, Jyoti V; Pandhare, Jui; Dash, Chandravanu

    2014-01-01

    Methamphetamine is the second most frequently used illicit drug in the United States. Methamphetamine abuse is associated with increased risk of HIV-1 acquisition, higher viral loads, and enhanced HIV-1 pathogenesis. Although a direct link between methamphetamine abuse and HIV-1 pathogenesis remains to be established in patients, methamphetamine has been shown to increase HIV-1 replication in macrophages, dendritic cells, and cells of HIV transgenic mice. Intriguingly, the effects of methamphetamine on HIV-1 replication in human CD4(+) T cells that serve as the primary targets of infection in vivo are not clearly understood. Therefore, we examined HIV-1 replication in primary CD4(+) T cells in the presence of methamphetamine in a dose-dependent manner. Our results demonstrate that methamphetamine had a minimal effect on HIV-1 replication at concentrations of 1 to 50 μmol/L. However, at concentrations >100 μmol/L, it inhibited HIV-1 replication in a dose-dependent manner. We also discovered that methamphetamine up-regulated the cellular anti-HIV-1 microRNAs (miR-125b, miR-150, and miR-28-5p) in CD4(+) T cells. Knockdown experiments illustrated that up-regulation of the anti-HIV miRNAs inhibited HIV-1 replication. These results are contrary to the paradigm that methamphetamine accentuates HIV-1 pathogenesis by increasing HIV-1 replication. Therefore, our findings underline the complex interaction between drug use and HIV-1 and necessitate comprehensive understanding of the effects of methamphetamine on HIV-1 pathogenesis. Copyright © 2014 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

  19. Cyclophilin B enhances HIV-1 infection

    Energy Technology Data Exchange (ETDEWEB)

    DeBoer, Jason; Madson, Christian J. [Department of Medical Microbiology and Immunology, Creighton University, Omaha, NE (United States); Belshan, Michael, E-mail: michaelbelshan@creighton.edu [Department of Medical Microbiology and Immunology, Creighton University, Omaha, NE (United States); The Nebraska Center for Virology, University of Nebraska, Lincoln, NE (United States)

    2016-02-15

    Cyclophilin B (CypB) is a member of the immunophilin family and intracellular chaperone. It predominantly localizes to the ER, but also contains a nuclear localization signal and is secreted from cells. CypB has been shown to interact with the Gag protein of human immunodeficiency type 1 (HIV-1). Several proteomic and genetic studies identified it as a potential factor involved in HIV replication. Herein, we show that over-expression of CypB enhances HIV infection by increasing nuclear import of viral DNA. This enhancement was unaffected by cyclosporine treatment and requires the N-terminus of the protein. The N-terminus contains an ER leader sequence, putative nuclear localization signal, and is required for secretion. Deletion of the N-terminus resulted in mislocalization from the ER and suppression of HIV infection. Passive transfer experiments showed that secreted CypB did not impact HIV infection. Combined, these experiments show that intracellular CypB modulates a pathway of HIV nuclear import. - Highlights: • CypB has been identified in several proteomic studies of HIV-1 infection. • CypB expression is upregulated in activated and infected T-cells. • Over-expression of CypB enhances HIV nuclear import and infection. • The N-terminus of CypB is necessary for these effects.

  20. Rational development of radiopharmaceuticals for HIV-1

    International Nuclear Information System (INIS)

    Lau, Chuen-Yen; Maldarelli, Frank; Eckelman, William C.; Neumann, Ronald D.

    2014-01-01

    The global battle against HIV-1 would benefit from a sensitive and specific radiopharmaceutical to localize HIV-infected cells. Ideally, this probe would be able to identify latently infected host cells containing replication competent HIV sequences. Clinical and research applications would include assessment of reservoirs, informing clinical management by facilitating assessment of burden of infection in different compartments, monitoring disease progression and monitoring response to therapy. A “rational” development approach could facilitate efficient identification of an appropriate targeted radiopharmaceutical. Rational development starts with understanding characteristics of the disease that can be effectively targeted and then engineering radiopharmaceuticals to hone in on an appropriate target, which in the case of HIV-1 (HIV) might be an HIV-specific product on or in the host cell, a differentially expressed gene product, an integrated DNA sequence specific enzymatic activity, part of the inflammatory response, or a combination of these. This is different from the current approach that starts with a radiopharmaceutical for a target associated with a disease, mostly from autopsy studies, without a strong rationale for the potential to impact patient care. At present, no targeted therapies are available for HIV latency, although a number of approaches are under study. Here we discuss requirements for a radiopharmaceutical useful in strategies targeting persistently infected cells. The radiopharmaceutical for HIV should be developed based on HIV biology, studied in an animal model and then in humans, and ultimately used in clinical and research settings

  1. Cyclophilin B enhances HIV-1 infection

    International Nuclear Information System (INIS)

    DeBoer, Jason; Madson, Christian J.; Belshan, Michael

    2016-01-01

    Cyclophilin B (CypB) is a member of the immunophilin family and intracellular chaperone. It predominantly localizes to the ER, but also contains a nuclear localization signal and is secreted from cells. CypB has been shown to interact with the Gag protein of human immunodeficiency type 1 (HIV-1). Several proteomic and genetic studies identified it as a potential factor involved in HIV replication. Herein, we show that over-expression of CypB enhances HIV infection by increasing nuclear import of viral DNA. This enhancement was unaffected by cyclosporine treatment and requires the N-terminus of the protein. The N-terminus contains an ER leader sequence, putative nuclear localization signal, and is required for secretion. Deletion of the N-terminus resulted in mislocalization from the ER and suppression of HIV infection. Passive transfer experiments showed that secreted CypB did not impact HIV infection. Combined, these experiments show that intracellular CypB modulates a pathway of HIV nuclear import. - Highlights: • CypB has been identified in several proteomic studies of HIV-1 infection. • CypB expression is upregulated in activated and infected T-cells. • Over-expression of CypB enhances HIV nuclear import and infection. • The N-terminus of CypB is necessary for these effects.

  2. HIV-1 proteins dysregulate motivational processes and dopamine circuitry.

    Science.gov (United States)

    Bertrand, Sarah J; Mactutus, Charles F; Harrod, Steven B; Moran, Landhing M; Booze, Rosemarie M

    2018-05-18

    Motivational alterations, such as apathy, in HIV-1+ individuals are associated with decreased performance on tasks involving frontal-subcortical circuitry. We used the HIV-1 transgenic (Tg) rat to assess effect of long-term HIV-1 protein exposure on motivated behavior using sucrose (1-30%, w/v) and cocaine (0.01-1.0 mg/kg/infusion) maintained responding with fixed-ratio (FR) and progressive-ratio (PR) schedules of reinforcement. For sucrose-reinforced responding, HIV-1 Tg rats displayed no change in EC 50 relative to controls, suggesting no change in sucrose reinforcement but had a downward shifted concentration-response curves, suggesting a decrease in response vigor. Cocaine-maintained responding was attenuated in HIV-1 Tg rats (FR1 0.33 mg/kg/infusion and PR 1.0 mg/kg/infusion). Dose-response tests (PR) revealed that HIV-1 Tg animals responded significantly less than F344 control rats and failed to earn significantly more infusions of cocaine as the unit dose increased. When choosing between cocaine and sucrose, control rats initially chose sucrose but with time shifted to a cocaine preference. In contrast, HIV-1 disrupted choice behaviors. DAT function was altered in the striatum of HIV-1 Tg rats; however, prior cocaine self-administration produced a unique effect on dopamine homeostasis in the HIV-1 Tg striatum. These findings of altered goal directed behaviors may determine neurobiological mechanisms of apathy in HIV-1+ patients.

  3. The HIV-1 Tat protein modulates CD4 expression in human T cells through the induction of miR-222.

    Science.gov (United States)

    Orecchini, Elisa; Doria, Margherita; Michienzi, Alessandro; Giuliani, Erica; Vassena, Lia; Ciafrè, Silvia Anna; Farace, Maria Giulia; Galardi, Silvia

    2014-01-01

    Several cellular microRNAs show substantial changes in expression during HIV-1 infection and their active role in the viral life cycle is progressively emerging. In the present study, we found that HIV-1 infection of Jurkat T cells significantly induces the expression of miR-222. We show that this induction depends on HIV-1 Tat protein, which is able to increase the transcriptional activity of NFkB on miR-222 promoter. Moreover, we demonstrate that miR-222 directly targets CD4, a key receptor for HIV-1, thus reducing its expression. We propose that Tat, by inducing miR-222 expression, complements the CD4 downregulation activity exerted by other viral proteins (i.e., Nef, Vpu, and Env), and we suggest that this represents a novel mechanism through which HIV-1 efficiently represses CD4 expression in infected cells.

  4. The HIV-1 Tat protein modulates CD4 expression in human T cells through the induction of miR-222

    Science.gov (United States)

    Orecchini, Elisa; Doria, Margherita; Michienzi, Alessandro; Giuliani, Erica; Vassena, Lia; Ciafrè, Silvia Anna; Farace, Maria Giulia; Galardi, Silvia

    2014-01-01

    Several cellular microRNAs show substantial changes in expression during HIV-1 infection and their active role in the viral life cycle is progressively emerging. In the present study, we found that HIV-1 infection of Jurkat T cells significantly induces the expression of miR-222. We show that this induction depends on HIV-1 Tat protein, which is able to increase the transcriptional activity of NFkB on miR-222 promoter. Moreover, we demonstrate that miR-222 directly targets CD4, a key receptor for HIV-1, thus reducing its expression. We propose that Tat, by inducing miR-222 expression, complements the CD4 downregulation activity exerted by other viral proteins (i.e., Nef, Vpu, and Env), and we suggest that this represents a novel mechanism through which HIV-1 efficiently represses CD4 expression in infected cells. PMID:24717285

  5. An Analysis Of Pole/zero Cancellation In LTR-based Feedback Design

    DEFF Research Database (Denmark)

    Niemann, Hans Henrik; Jannerup, Ole Erik

    1990-01-01

    The pole/zero cancellation in LTR-based feedback design will be analyzed for both full-order as well as minimal-order observers. The asymptotic behaviour of the sensitivity function from the LTR-procedure are given in explicit expressions in the case when a zero is not cancelled by an equivalent...... pole. It will be shown that the non-minimum phase case is included as a special case. The results are not based on any specific LTR-method....

  6. Antibodyomics: bioinformatics technologies for understanding B-cell immunity to HIV-1.

    Science.gov (United States)

    Kwong, Peter D; Chuang, Gwo-Yu; DeKosky, Brandon J; Gindin, Tatyana; Georgiev, Ivelin S; Lemmin, Thomas; Schramm, Chaim A; Sheng, Zizhang; Soto, Cinque; Yang, An-Suei; Mascola, John R; Shapiro, Lawrence

    2017-01-01

    Numerous antibodies have been identified from HIV-1-infected donors that neutralize diverse strains of HIV-1. These antibodies may provide the basis for a B cell-mediated HIV-1 vaccine. However, it has been unclear how to elicit similar antibodies by vaccination. To address this issue, we have undertaken an informatics-based approach to understand the genetic and immunologic processes controlling the development of HIV-1-neutralizing antibodies. As DNA sequencing comprises the fastest growing database of biological information, we focused on incorporating next-generation sequencing of B-cell transcripts to determine the origin, maturation pathway, and prevalence of broadly neutralizing antibody lineages (Antibodyomics1, 2, 4, and 6). We also incorporated large-scale robotic analyses of serum neutralization to identify and quantify neutralizing antibodies in donor cohorts (Antibodyomics3). Statistical analyses furnish another layer of insight (Antibodyomics5), with physical characteristics of antibodies and their targets through molecular dynamics simulations (Antibodyomics7) and free energy perturbation analyses (Antibodyomics8) providing information-rich output. Functional interrogation of individual antibodies (Antibodyomics9) and synthetic antibody libraries (Antibodyomics10) also yields multi-dimensional data by which to understand and improve antibodies. Antibodyomics, described here, thus comprise resolution-enhancing tools, which collectively embody an information-driven discovery engine aimed toward the development of effective B cell-based vaccines. © 2017 The Authors. Immunological Reviews published by John Wiley & Sons Ltd.

  7. HIV-1 matrix dependent membrane targeting is regulated by Gag mRNA trafficking.

    Directory of Open Access Journals (Sweden)

    Jing Jin

    Full Text Available Retroviral Gag polyproteins are necessary and sufficient for virus budding. Productive HIV-1 Gag assembly takes place at the plasma membrane. However, little is known about the mechanisms by which thousands of Gag molecules are targeted to the plasma membrane. Using a bimolecular fluorescence complementation (BiFC assay, we recently reported that the cellular sites and efficiency of HIV-1 Gag assembly depend on the precise pathway of Gag mRNA export from the nucleus, known to be mediated by Rev. Here we describe an assembly deficiency in human cells for HIV Gag whose expression depends on hepatitis B virus (HBV post-transcriptional regulatory element (PRE mediated-mRNA nuclear export. PRE-dependent HIV Gag expressed well in human cells, but assembled with slower kinetics, accumulated intracellularly, and failed to associate with a lipid raft compartment where the wild-type Rev-dependent HIV-1 Gag efficiently assembles. Surprisingly, assembly and budding of PRE-dependent HIV Gag in human cells could be rescued in trans by co-expression of Rev-dependent Gag that provides correct membrane targeting signals, or in cis by replacing HIV matrix (MA with other membrane targeting domains. Taken together, our results demonstrate deficient membrane targeting of PRE-dependent HIV-1 Gag and suggest that HIV MA function is regulated by the trafficking pathway of the encoding mRNA.

  8. HIV-1 subtype C unproductively infects human cardiomyocytes in vitro and induces apoptosis mitigated by an anti-Gp120 aptamer.

    Science.gov (United States)

    Lopes de Campos, Walter R; Chirwa, Nthato; London, Grace; Rotherham, Lia S; Morris, Lynn; Mayosi, Bongani M; Khati, Makobetsa

    2014-01-01

    HIV-associated cardiomyopathy (HIVCM) is of clinical concern in developing countries because of a high HIV-1 prevalence, especially subtype C, and limited access to highly active antiretroviral therapy (HAART). For these reasons, we investigated the direct and indirect effects of HIV-1 subtype C infection of cultured human cardiomyocytes and the mechanisms leading to cardiomyocytes damage; as well as a way to mitigate the damage. We evaluated a novel approach to mitigate HIVCM using a previously reported gp120 binding and HIV-1 neutralizing aptamer called UCLA1. We established a cell-based model of HIVCM by infecting human cardiomyocytes with cell-free HIV-1 or co-culturing human cardiomyocytes with HIV-infected monocyte derived macrophages (MDM). We discovered that HIV-1 subtype C unproductively (i.e. its life cycle is arrested after reverse transcription) infects cardiomyocytes. Furthermore, we found that HIV-1 initiates apoptosis of cardiomyocytes through caspase-9 activation, preferentially via the intrinsic or mitochondrial initiated pathway. CXCR4 receptor-using viruses were stronger inducers of apoptosis than CCR5 utilizing variants. Importantly, we discovered that HIV-1 induced apoptosis of cardiomyocytes was mitigated by UCLA1. However, UCLA1 had no protective effective on cardiomyocytes when apoptosis was triggered by HIV-infected MDM. When HIV-1 was treated with UCLA1 prior to infection of MDM, it failed to induce apoptosis of cardiomyocytes. These data suggest that HIV-1 causes a mitochondrial initiated apoptotic cascade, which signal through caspase-9, whereas HIV-1 infected MDM causes apoptosis predominantly via the death-receptor pathway, mediated by caspase-8. Furthermore the data suggest that UCLA1 protects cardiomyocytes from caspase-mediated apoptosis, directly by binding to HIV-1 and indirectly by preventing infection of MDM.

  9. Upregulation of Glucose Uptake and Hexokinase Activity of Primary Human CD4+ T Cells in Response to Infection with HIV-1

    Directory of Open Access Journals (Sweden)

    Maia Kavanagh Williamson

    2018-03-01

    Full Text Available Infection of primary CD4+ T cells with HIV-1 coincides with an increase in glycolysis. We investigated the expression of glucose transporters (GLUT and glycolytic enzymes in human CD4+ T cells in response to infection with HIV-1. We demonstrate the co-expression of GLUT1, GLUT3, GLUT4, and GLUT6 in human CD4+ T cells after activation, and their concerted overexpression in HIV-1 infected cells. The investigation of glycolytic enzymes demonstrated activation-dependent expression of hexokinases HK1 and HK2 in human CD4+ T cells, and a highly significant increase in cellular hexokinase enzyme activity in response to infection with HIV-1. HIV-1 infected CD4+ T cells showed a marked increase in expression of HK1, as well as the functionally related voltage-dependent anion channel (VDAC protein, but not HK2. The elevation of GLUT, HK1, and VDAC expression in HIV-1 infected cells mirrored replication kinetics and was dependent on virus replication, as evidenced by the use of reverse transcription inhibitors. Finally, we demonstrated that the upregulation of HK1 in HIV-1 infected CD4+ T cells is independent of the viral accessory proteins Vpu, Vif, Nef, and Vpr. Though these data are consistent with HIV-1 dependency on CD4+ T cell glucose metabolism, a cellular response mechanism to infection cannot be ruled out.

  10. BI-2 destabilizes HIV-1 cores during infection and Prevents Binding of CPSF6 to the HIV-1 Capsid.

    Science.gov (United States)

    Fricke, Thomas; Buffone, Cindy; Opp, Silvana; Valle-Casuso, Jose; Diaz-Griffero, Felipe

    2014-12-11

    The recently discovered small-molecule BI-2 potently blocks HIV-1 infection. BI-2 binds to the N-terminal domain of HIV-1 capsid. BI-2 utilizes the same capsid pocket used by the small molecule PF74. Although both drugs bind to the same pocket, it has been proposed that BI-2 uses a different mechanism to block HIV-1 infection when compared to PF74. This work demonstrates that BI-2 destabilizes the HIV-1 core during infection, and prevents the binding of the cellular factor CPSF6 to the HIV-1 core. Overall this short-form paper suggests that BI-2 is using a similar mechanism to the one used by PF74 to block HIV-1 infection.

  11. LTR retrotransposon landscape in Medicago truncatula: more rapid removal than in rice

    Directory of Open Access Journals (Sweden)

    Liu Jin-Song

    2008-08-01

    Full Text Available Abstract Background Long terminal repeat retrotransposons (LTR elements are ubiquitous Eukaryotic TEs that transpose through RNA intermediates. Accounting for significant proportion of many plant genomes, LTR elements have been well established as one of the major forces underlying the evolution of plant genome size, structure and function. The accessibility of more than 40% of genomic sequences of the model legume Medicago truncatula (Mt has made the comprehensive study of its LTR elements possible. Results We use a newly developed tool LTR_FINDER to identify LTR retrotransposons in the Mt genome and detect 526 full-length elements as well as a great number of copies related to them. These elements constitute about 9.6% of currently available genomic sequences. They are classified into 85 families of which 64 are reported for the first time. The majority of the LTR retrotransposons belong to either Copia or Gypsy superfamily and the others are categorized as TRIMs or LARDs by their length. We find that the copy-number of Copia-like families is 3 times more than that of Gypsy-like ones but the latter contribute more to the genome. The analysis of PBS and protein-coding domain structure of the LTR families reveals that they tend to use only 4–5 types of tRNAs and many families have quite conservative ORFs besides known TE domains. For several important families, we describe in detail their abundance, conservation, insertion time and structure. We investigate the amplification-deletion pattern of the elements and find that the detectable full-length elements are relatively young and most of them were inserted within the last 0.52 MY. We also estimate that more than ten million bp of the Mt genomic sequences have been removed by the deletion of LTR elements and the removal of the full-length structures in Mt has been more rapid than in rice. Conclusion This report is the first comprehensive description and analysis of LTR retrotransposons in the

  12. Opening of the TAR hairpin in the HIV-1 genome causes aberrant RNA dimerization and packaging

    Directory of Open Access Journals (Sweden)

    Das Atze T

    2012-07-01

    Full Text Available Abstract Background The TAR hairpin is present at both the 5′ and 3′ end of the HIV-1 RNA genome. The 5′ element binds the viral Tat protein and is essential for Tat-mediated activation of transcription. We recently observed that complete TAR deletion is allowed in the context of an HIV-1 variant that does not depend on this Tat-TAR axis for transcription. Mutations that open the 5′ stem-loop structure did however affect the leader RNA conformation and resulted in a severe replication defect. In this study, we set out to analyze which step of the HIV-1 replication cycle is affected by this conformational change of the leader RNA. Results We demonstrate that opening the 5′ TAR structure through a deletion in either side of the stem region caused aberrant dimerization and reduced packaging of the unspliced viral RNA genome. In contrast, truncation of the TAR hairpin through deletions in both sides of the stem did not affect RNA dimer formation and packaging. Conclusions These results demonstrate that, although the TAR hairpin is not essential for RNA dimerization and packaging, mutations in TAR can significantly affect these processes through misfolding of the relevant RNA signals.

  13. Multifaceted counter-APOBEC3G mechanisms employed by HIV-1 Vif.

    Science.gov (United States)

    Britan-Rosich, Elena; Nowarski, Roni; Kotler, Moshe

    2011-07-29

    In the absence of human immunodeficiency virus type 1 (HIV-1) Vif protein, the host antiviral deaminase apolipoprotein B mRNA-editing enzyme-catalytic polypeptide-like 3G (A3G) restricts the production of infectious HIV-1 by deamination of dC residues in the negative single-stranded DNA produced by reverse transcription. The Vif protein averts the lethal threat of deamination by precluding the packaging of A3G into assembling virions by mediating proteasomal degradation of A3G. In spite of this robust Vif activity, residual A3G molecules that escape degradation and incorporate into newly assembled virions are potentially deleterious to the virus. We hypothesized that virion-associated Vif inhibits A3G enzymatic activity and therefore prevents lethal mutagenesis of the newly synthesized viral DNA. Here, we show that (i) Vif-proficient HIV-1 particles released from H9 cells contain A3G with lower specific activity compared with Δvif-virus-associated A3G, (ii) encapsidated HIV-1 Vif inhibits the deamination activity of recombinant A3G, and (iii) purified HIV-1 Vif protein and the Vif-derived peptide Vif25-39 inhibit A3G activity in vitro at nanomolar concentrations in an uncompetitive manner. Our results manifest the potentiality of Vif to control the deamination threat in virions or in the pre-integration complexes following entry to target cells. Hence, virion-associated Vif could serve as a last line of defense, protecting the virus against A3G antiviral activity. Copyright © 2011 Elsevier Ltd. All rights reserved.

  14. Striking HIV-1 Entry by Targeting HIV-1 gp41. But, Where Should We Target?

    Directory of Open Access Journals (Sweden)

    Cátia Teixeira

    Full Text Available HIV-1 gp41 facilitates the viral fusion through a conformational switch involving the association of three C-terminal helices along the conserved hydrophobic grooves of three N-terminal helices coiled-coil. The control of these structural rearrangements is thought to be central to HIV-1 entry and, therefore, different strategies of intervention are being developed. Herewith, we describe a procedure to simulate the folding of an HIV-1 gp41 simplified model. This procedure is based on the construction of plausible conformational pathways, which describe protein transition between non-fusogenic and fusogenic conformations. The calculation of the paths started with 100 molecular dynamics simulations of the non-fusogenic conformation, which were found to converge to different intermediate states. Those presenting defined criteria were selected for separate targeted molecular dynamics simulations, subjected to a force constant imposing a movement towards the gp41 fusogenic conformation. Despite significant diversity, a preferred sequence of events emerged when the simulations were analyzed in terms of the formation, breakage and evolution of the contacts. We pointed out 29 residues as the most relevant for the movement of gp41; also, 2696 possible interactions were reduced to only 48 major interactions, which reveals the efficiency of the method. The analysis of the evolution of the main interactions lead to the detection of four main behaviors for those contacts: stable, increasing, decreasing and repulsive interactions. Altogether, these results suggest a specific small cavity of the HIV-1 gp41 hydrophobic groove as the preferred target to small molecules.

  15. The anti-HIV-1 effect of scutellarin

    International Nuclear Information System (INIS)

    Zhang Gaohong; Wang Qian; Chen Jijun; Zhang Xuemei; Tam, S.-C.; Zheng Yongtang

    2005-01-01

    Scutellarin was purified from the plant Erigeron breviscapus (Vant.) Hand.-Mazz. The activity against 3 strains of human immunodeficiency virus (HIV) was determined in vitro in this study. These were laboratory-derived virus (HIV-1 IIIB ), drug-resistant virus (HIV-1 74V ), and low-passage clinical isolated virus (HIV-1 KM018 ). From syncytia inhibition study, the EC 50 of scutellarin against HIV-1 IIIB direct infection in C8166 cells was 26 μM with a therapeutic index of 36. When the mode of infection changed from acute infection to cell-to-cell infection, this compound became even more potent and the EC 50 reduced to 15 μM. This suggested that cell fusion might be affected by this compound. By comparing the inhibitory effects on p24 antigen, scutellarin was also found to be active against HIV-1 74V (EC 50 253 μM) and HIV-1 KM018 (EC 50 136 μM) infection with significant difference in potency. The mechanism of its action was also explored in this study. At a concentration of 433 μM, scutellarin inhibited 48% of the cell free recombinant HIV-1 RT activity. It also caused 82% inhibition of HIV-1 particle attachment and 45% inhibition of fusion at the concentrations of 54 μM. In summary, scutellarin was found to inhibit several strains of HIV-1 replication with different potencies. It appeared to inhibit HIV-1 RT activity, HIV-1 particle attachment and cell fusion. These are essential activities for viral transmission and replication

  16. Cloning, Expression and Purification of the Recombinant HIV-1 Tat-Nef Fusion Protein in Prokaryotic Expression System

    OpenAIRE

    Somayeh Kadkhodayan; Shiva Irani; Seyed Mehdi Sadat; Fatemeh Fotouhi; Azam Bolhassani

    2016-01-01

    Abstract Background: Nef is one of the HIV-1 critical proteins, because it is essential for viral replication and AIDS disease progression and induction of immune response against it can partially inhibit viral infection. Moreover, a domain of the HIV-1 Trans-Activator of Transcription (Tat, 48-60 aa) could act as a cell penetrating peptide (CPP). In current study, cloning and expression of Tat-Nef fusion protein was performed in E. coli for the first time. The protein expression was confi...

  17. The origin and emergence of an HIV-1 epidemic:

    DEFF Research Database (Denmark)

    Bruhn, Christian Anders Wathne; Audelin, Anne M.; Helleberg, Marie

    2014-01-01

    To describe, at patient-level detail, the determining events and factors involved in the development of a country's HIV-1 epidemic.......To describe, at patient-level detail, the determining events and factors involved in the development of a country's HIV-1 epidemic....

  18. Schistosomiasis and HIV-1 infection in rural Zimbabwe

    DEFF Research Database (Denmark)

    Kallestrup, Per; Zinyama, Rutendo; Gomo, Exnevia

    2005-01-01

    Stunted development and reduced fecundity of Schistosoma parasites in immunodeficient mice and the impaired ability of human immunodeficiency virus 1 (HIV-1)-infected humans to excrete schistosome eggs have been described. This study explores the effect that HIV-1-associated immunodeficiency has...

  19. HIV-1 Nef control of cell signalling molecules: multiple strategies

    Indian Academy of Sciences (India)

    HIV-1 has at its disposal numerous proteins encoded by its genome which provide the required arsenal to establish and maintain infection in its host for a considerable number of years. One of the most important and enigmatic of these proteins is Nef. The Nef protein of HIV-1 plays a fundamental role in the virus life cycle.

  20. Molecular Mechanisms in Activation of Latent HIV-1

    NARCIS (Netherlands)

    H. Rafati (Haleh)

    2014-01-01

    markdownabstract__Abstract__ Finding a cure for the human immunodeficiency virus type 1 (HIV-1) is extremely challenging. Development of highly active anti-retroviral therapy (HAART), transformed HIV-1 infection from an acute syndrome into chronic disease. Although using HAART results in

  1. Vaginalmycosis and HIV-1 infection in Kaduna, Nigeria. | Eni ...

    African Journals Online (AJOL)

    ... mycosis in HIV-1positive women and managed accordingly. Proper management of these two conditions will improve reproductive health of women in Nigeria. Keywords: Vaginal mycosis, Genital candidiasis, Reproductive health: Candida albicans: HIV-1 infection. Journal of Biomedical Investigation Vol. 3 (1) 2005: pp.

  2. Antiviral Therapy by HIV-1 Broadly Neutralizing and Inhibitory Antibodies

    Directory of Open Access Journals (Sweden)

    Zhiqing Zhang

    2016-11-01

    Full Text Available Human immunodeficiency virus type 1 (HIV-1 infection causes acquired immune deficiency syndrome (AIDS, a global epidemic for more than three decades. HIV-1 replication is primarily controlled through antiretroviral therapy (ART but this treatment does not cure HIV-1 infection. Furthermore, there is increasing viral resistance to ART, and side effects associated with long-term therapy. Consequently, there is a need of alternative candidates for HIV-1 prevention and therapy. Recent advances have discovered multiple broadly neutralizing antibodies against HIV-1. In this review, we describe the key epitopes on the HIV-1 Env protein and the reciprocal broadly neutralizing antibodies, and discuss the ongoing clinical trials of broadly neutralizing and inhibitory antibody therapy as well as antibody combinations, bispecific antibodies, and methods that improve therapeutic efficacy by combining broadly neutralizing antibodies (bNAbs with latency reversing agents. Compared with ART, HIV-1 therapeutics that incorporate these broadly neutralizing and inhibitory antibodies offer the advantage of decreasing virus load and clearing infected cells, which is a promising prospect in HIV-1 prevention and treatment.

  3. Cold denaturation of the HIV-1 protease monomer

    DEFF Research Database (Denmark)

    Rösner, Heike Ilona; Caldarini, Martina; Prestel, Andreas

    2017-01-01

    The HIV-1-protease is a complex protein which in its active form adopts a homodimer dominated by -sheet structures. We have discovered a cold-denatured state of the monomeric subunit of HIV-1-protease which is populated above 0ºC and therefore directly accessible to various spectroscopic approac...

  4. Raltegravir with optimized background therapy for resistant HIV-1 infection

    DEFF Research Database (Denmark)

    Steigbigel, Roy T; Cooper, David A; Kumar, Princy N

    2008-01-01

    BACKGROUND: Raltegravir (MK-0518) is an inhibitor of human immunodeficiency virus type 1 (HIV-1) integrase active against HIV-1 susceptible or resistant to older antiretroviral drugs. METHODS: We conducted two identical trials in different geographic regions to evaluate the safety and efficacy of...

  5. HIV-1 protease inhibitory substances from Cassia garrettiana

    Directory of Open Access Journals (Sweden)

    Jindaporn Puripattanvong

    2007-01-01

    Full Text Available Cassia garrettiana Craib, a Thai medicinal plant locally known as Samae-sarn, was investigated for its active constituents against HIV-1 protease (HIV-1 PR. Bioassay-guided fractionation of the heart woodof this plant led to the isolation of a stilbene derivative (1, piceatannol and an anthraquinone derivative (2, chrysophanol. Piceatannol exhibited appreciable inhibitory effect against HIV-1 PR with an IC50 value of25.4 μg/ml, whereas that of chrysophanol was 73.5 μg/ml. In addition, other two stilbenoids together with three anthraquinone derivatives were also investigated for their anti-HIV-1 PR activities. The resultindicated that resveratrol possessed anti-HIV-1 PR activity with an IC50 value of 85.0 μg/ml, whereas other stilbenoid (oxyresveratrol and anthraquinone derivatives (emodin, aloe-emodin, rhein were inactive (IC50 > 100 μg/ml.

  6. SP-D impedes transfer of HIV-1 from multi-layered vaginal epithelium with a distinct gene signature

    Directory of Open Access Journals (Sweden)

    Hrishikesh Pandit

    2017-12-01

    Full Text Available Surfactant Protein (SP D is a member of the collectin family of soluble pattern recognition receptors. We have previously shown that a recombinant fragment of SP-D (rhSP-D inhibits gp120-CD4 interaction and HIV-1 entry in target cells. To potentiate its prophylactic use as a vaginal microbicide, we determined ex vivo efficacy using organotypic human vaginal-ectocervical epithelia (VEC-100 that closely resemble the native tissues of origin. VEC-100, stratified human vaginal-ectocervical tissues grown on membrane inserts were treated with rhSP-D followed by a challenge with HIV-1 to assess the transfer of HIV-1 through the VEC-100 tissues to PBMCs in the basal submucosal compartment. Treated VEC tissues were subjected to mRNA Illumina microarray analysis. Levels of transcripts encoding for immune mediators, adhesion and tight junction proteins were also evaluated. Effect of rhSP-D on viability, NFκB activation, cytokine secretion and bacterial colonization of cervical vaginal epithelial cells was determined. rhSP-D significantly inhibited HIV-1 transfer from the multi-layered epithelial tissues to the basal PBMCs as compared to HIV-1 alone. Global gene expression profile of HIV-1 challenged VEC-100 tissues revealed differential regulation of genes and pathways majorly involved in inflammation, cell survival and transcription factors. Levels of Guanylate-binding proteins (GBPs and interferon-inducible proteins were significantly upregulated suggesting an interferon host defense response. rhSP-D showed an inhibition in the levels of GBPs and rescued the cell adhesion molecules such as Claudin 2, 3, 4, 5 and Occludin, known to be down regulated by HIV-1 in primary vaginal cells. Importantly, rhSP-D conditioned VEC tissue supernatants did not enhance susceptibility of target cells to HIV-1. rhSP-D treated vaginal epithelial cells did not show any significant alteration in viability, NFκB activation and levels of immune mediators like IL-1RA, Elafin

  7. Interferon-β induced in female genital epithelium by HIV-1 glycoprotein 120 via Toll-like-receptor 2 pathway acts to protect the mucosal barrier.

    Science.gov (United States)

    Nazli, Aisha; Dizzell, Sara; Zahoor, Muhammad Atif; Ferreira, Victor H; Kafka, Jessica; Woods, Matthew William; Ouellet, Michel; Ashkar, Ali A; Tremblay, Michel J; Bowdish, Dawn Me; Kaushic, Charu

    2018-03-19

    More than 40% of HIV infections occur via female reproductive tract (FRT) through heterosexual transmission. Epithelial cells that line the female genital mucosa are the first line of defense against HIV-1 and other sexually transmitted pathogens. These sentient cells recognize and respond to external stimuli by induction of a range of carefully balanced innate immune responses. Previously, we have shown that in response to HIV-1 gp120, the genital epithelial cells (GECs) from upper reproductive tract induce an inflammatory response that may facilitate HIV-1 translocation and infection. In this study, we report that the endometrial and endocervical GECs simultaneously induce biologically active interferon-β (IFNβ) antiviral responses following exposure to HIV-1 that act to protect the epithelial tight junction barrier. The innate antiviral response was directly induced by HIV-1 envelope glycoprotein gp120 and addition of gp120 neutralizing antibody inhibited IFNβ production. Interferon-β was induced by gp120 in upper GECs through Toll-like receptor 2 signaling and required presence of heparan sulfate on epithelial cell surface. The induction of IFNβ was dependent upon activation of transcription factor IRF3 (interferon regulatory factor 3). The IFNβ was biologically active, had a protective effect on epithelial tight junction barrier and was able to inhibit HIV-1 infection in TZM-bl indicator cells and HIV-1 replication in T cells. This is the first report that recognition of HIV-1 by upper GECs leads to induction of innate antiviral pathways. This could explain the overall low infectivity of HIV-1 in the FRT and could be exploited for HIV-1 prophylaxis.Cellular and Molecular Immunology advance online publication, 19 March 2018; doi:10.1038/cmi.2017.168.

  8. Comparison of glycerolisation with cryopreservation methods on HIV-1 inactivation

    International Nuclear Information System (INIS)

    Van Baare, J.; Pagnon, J.; Cameron, P.; Vardaxis, N.; Middlekoop, E.; Crowe, S.

    1999-01-01

    Cryopreservation and glycerolisation are two successful long-term preservation methods for human cadaveric donor skin, which is used in the treatment of bum patients. High concentrations of glycerol has been shown to be antibacterial and virucidal. Because fear of possible transmission of HIV-1 following allograft transplantation, this study was undertaken to investigate whether HIV can be effectively eliminated from skin explants. HIV-1 Ba-L, which has been shown to infect monocytes in skin explants and also dendritic cells, was. For the experiments we used cell-free virus, exogenously HIV infected peripheral blood mononuclear cells (PBMCs) and exogenously HIV infected cadaver split skin. Different concentrations of glycerol at various temperatures and the glycerolisation procedure as used by the Euro Skin Bank were used to determine the effects on HIV-1 Ba-L infectivity. For the cryopreservation technique we used 10% DMSO and a controlled rate freezer. HIV-1 Ba-L transfer was determined by adding uninfected PBMCs to the infected material and reverse transcriptase was measured. Cell-free HIV-1 Ba-L was not inactivated by 50% glycerol but was effectively inactivated within 30 minutes by 70% and 85% glycerol at 4 degree C, room temperature and 37 degree C. In contrast, cell-free HIV-1 Ba-L was not inactivated by cryopreservation. Most importantly, we have shown that HIV-1 Ba-L present in split skin is inactivated by incubating skin in 70% glycerol for three hours at 37-C. HIV in exogenously infected skin was not inactivated by cryopreservation. High concentrations of glycerol effectively inactivates free HIV-1 Ba-L and intracellular HIV-1 Ba-L. Also the current glycerolisation procedure carried out by the Euro Skin Bank effectively inactivates infectious virus. However, the cryopreservation technique did not show any reduction in HIV-1 Ba-L infectivity

  9. Sexually transmitted infections among HIV-1-discordant couples.

    Directory of Open Access Journals (Sweden)

    Brandon L Guthrie

    2009-12-01

    Full Text Available More new HIV-1 infections occur within stable HIV-1-discordant couples than in any other group in Africa, and sexually transmitted infections (STIs may increase transmission risk among discordant couples, accounting for a large proportion of new HIV-1 infections. Understanding correlates of STIs among discordant couples will aid in optimizing interventions to prevent HIV-1 transmission in these couples.HIV-1-discordant couples in which HIV-1-infected partners were HSV-2-seropositive were tested for syphilis, chlamydia, gonorrhea, and trichomoniasis, and HIV-1-uninfected partners were tested for HSV-2. We assessed sociodemographic, behavioral, and biological correlates of a current STI.Of 416 couples enrolled, 16% were affected by a treatable STI, and among these both partners were infected in 17% of couples. A treatable STI was found in 46 (11% females and 30 (7% males. The most prevalent infections were trichomoniasis (5.9% and syphilis (2.6%. Participants were 5.9-fold more likely to have an STI if their partner had an STI (P<0.01, and STIs were more common among those reporting any unprotected sex (OR = 2.43; P<0.01 and those with low education (OR = 3.00; P<0.01. Among HIV-1-uninfected participants with an HSV-2-seropositive partner, females were significantly more likely to be HSV-2-seropositive than males (78% versus 50%, P<0.01.Treatable STIs were common among HIV-1-discordant couples and the majority of couples affected by an STI were discordant for the STI, with relatively high HSV-2 discordance. Awareness of STI correlates and treatment of both partners may reduce HIV-1 transmission.ClinicalTrials.gov NCT00194519.

  10. Cyclophilin B enhances HIV-1 infection.

    Science.gov (United States)

    DeBoer, Jason; Madson, Christian J; Belshan, Michael

    2016-02-01

    Cyclophilin B (CypB) is a member of the immunophilin family and intracellular chaperone. It predominantly localizes to the ER, but also contains a nuclear localization signal and is secreted from cells. CypB has been shown to interact with the Gag protein of human immunodeficiency type 1 (HIV-1). Several proteomic and genetic studies identified it as a potential factor involved in HIV replication. Herein, we show that over-expression of CypB enhances HIV infection by increasing nuclear import of viral DNA. This enhancement was unaffected by cyclosporine treatment and requires the N-terminus of the protein. The N-terminus contains an ER leader sequence, putative nuclear localization signal, and is required for secretion. Deletion of the N-terminus resulted in mislocalization from the ER and suppression of HIV infection. Passive transfer experiments showed that secreted CypB did not impact HIV infection. Combined, these experiments show that intracellular CypB modulates a pathway of HIV nuclear import. Copyright © 2015 Elsevier Inc. All rights reserved.

  11. Changes in the topology of gene expression networks by human immunodeficiency virus type 1 (HIV-1) integration in macrophages.

    Science.gov (United States)

    Soto-Girón, María Juliana; García-Vallejo, Felipe

    2012-01-01

    One key step of human immunodeficiency virus type 1 (HIV-1) infection is the integration of its viral cDNA. This process is mediated through complex networks of host-virus interactions that alter several normal cell functions of the host. To study the complexity of disturbances in cell gene expression networks by HIV-1 integration, we constructed a network of human macrophage genes located close to chromatin regions rich in proviruses. To perform the network analysis, we selected 28 genes previously identified as the target of cDNA integration and their transcriptional profiles were obtained from GEO Profiles (NCBI). A total of 2770 interactions among the 28 genes located around the HIV-1 proviruses in human macrophages formed a highly dense main network connected to five sub-networks. The overall network was significantly enriched by genes associated with signal transduction, cellular communication and regulatory processes. To simulate the effects of HIV-1 integration in infected macrophages, five genes with the most number of interaction in the normal network were turned off by putting in zero the correspondent expression values. The HIV-1 infected network showed changes in its topology and alteration in the macrophage functions reflected in a re-programming of biosynthetic and general metabolic process. Understanding the complex virus-host interactions that occur during HIV-1 integration, may provided valuable genomic information to develop new antiviral treatments focusing on the management of some specific gene expression networks associated with viral integration. This is the first gene network which describes the human macrophages genes interactions related with HIV-1 integration. Copyright © 2011 Elsevier B.V. All rights reserved.

  12. Cis elements and trans-acting factors involved in the RNA dimerization of the human immunodeficiency virus HIV-1.

    Science.gov (United States)

    Darlix, J L; Gabus, C; Nugeyre, M T; Clavel, F; Barré-Sinoussi, F

    1990-12-05

    The retroviral genome consists of two identical RNA molecules joined at their 5' ends by the Dimer Linkage Structure (DLS). To study the mechanism of dimerization and the DLS of HIV-1 RNA, large amounts of bona fide HIV-1 RNA and of mutants have been synthesized in vitro. We report that HIV-1 RNA forms dimeric molecules and that viral nucleocapsid (NC) protein NCp15 greatly activates dimerization. Deletion mutagenesis in the RNA 5' 1333 nucleotides indicated that a small domain of 100 nucleotides, located between positions 311 to 415 from the 5' end, is necessary and sufficient to promote HIV-1 RNA dimerization. This dimerization domain encompasses an encapsidation element located between the 5' splice donor site and initiator AUG of gag and shows little sequence variations in different strains of HIV-1. Furthermore, cross-linking analysis of the interactions between NC and HIV-1 RNA (311 to 415) locates a major contact site in the encapsidation element of HIV-1 RNA. The genomic RNA dimer is tightly associated with nucleocapsid protein molecules in avian and murine retroviruses, and this ribonucleoprotein structure is believed to be the template for reverse transcription. Genomic RNA-protein interactions have been analyzed in human immunodeficiency virus (HIV) virions and results showed that NC protein molecules are tightly bound to the genomic RNA dimer. Since retroviral RNA dimerization and packaging appear to be under the control of the same cis element, the encapsidation sequences, and trans-acting factor, the NC protein, they are probably related events in the course of virion assembly.

  13. Development of an epitope-based HIV-1 vaccine strategy from HIV-1 lipopeptide to dendritic-based vaccines.

    Science.gov (United States)

    Surenaud, Mathieu; Lacabaratz, Christine; Zurawski, Gérard; Lévy, Yves; Lelièvre, Jean-Daniel

    2017-10-01

    Development of a safe, effective and globally affordable Human Immunodeficiency Virus strain 1 (HIV-1) vaccine offers the best hope for future control of the HIV-1 pandemic. However, with the exception of the recent RV144 trial, which elicited a modest level of protection against infection, no vaccine candidate has shown efficacy in preventing HIV-1 infection or in controlling virus replication in humans. There is also a great need for a successful immunotherapeutic vaccine since combination antiretroviral therapy (cART) does not eliminate the reservoir of HIV-infected cells. But to date, no vaccine candidate has proven to significantly alter the natural history of an individual with HIV-1 infection. Areas covered: For over 25 years, the ANRS (France Recherche Nord&Sud Sida-HIV hépatites) has been committed to an original program combining basic science and clinical research developing an epitope-based vaccine strategy to induce a multiepitopic cellular response against HIV-1. This review describes the evolution of concepts, based on strategies using HIV-1 lipopeptides towards the use of dendritic cell (DC) manipulation. Expert commentary: Understanding the crucial role of DCs in immune responses allowed moving from the non-specific administration of HIV-1 sequences with lipopeptides to DC-based vaccines. These DC-targeting strategies should improve HIV-1 vaccine efficacy.

  14. HIV-1-Specific IgA Monoclonal Antibodies from an HIV-1 Vaccinee Mediate Galactosylceramide Blocking and Phagocytosis

    Science.gov (United States)

    2018-01-01

    ABSTRACT Vaccine-elicited humoral immune responses comprise an array of antibody forms and specificities, with only a fraction contributing to protective host immunity. Elucidation of antibody effector functions responsible for protective immunity against human immunodeficiency virus type 1 (HIV-1) acquisition is a major goal for the HIV-1 vaccine field. Immunoglobulin A (IgA) is an important part of the host defense against pathogens; however, little is known about the role of vaccine-elicited IgA and its capacity to mediate antiviral functions. To identify the antiviral functions of HIV-1-specific IgA elicited by vaccination, we cloned HIV-1 envelope-specific IgA monoclonal antibodies (MAbs) by memory B cell cultures from peripheral blood mononuclear cells from an RV144 vaccinee and produced two IgA clonal cell lines (HG129 and HG130) producing native, nonrecombinant IgA MAbs. The HG129 and HG130 MAbs mediated phagocytosis by monocytes, and HG129 blocked HIV-1 Env glycoprotein binding to galactosylceramide, an alternative HIV-1 receptor. These findings elucidate potential antiviral functions of vaccine-elicited HIV-1 envelope-specific IgA that may act to block HIV-1 acquisition at the portal of entry by preventing HIV-1 binding to galactosylceramide and mediating antibody Fc receptor-mediated virion phagocytosis. Furthermore, these findings highlight the complex and diverse interactions of vaccine-elicited IgA with pathogens that depend on IgA fine specificity and form (e.g., multimeric or monomeric) in the systemic circulation and mucosal compartments. IMPORTANCE Host-pathogen interactions in vivo involve numerous immune mechanisms that can lead to pathogen clearance. Understanding the nature of antiviral immune mechanisms can inform the design of efficacious HIV-1 vaccine strategies. Evidence suggests that both neutralizing and nonneutralizing antibodies can mediate some protection against HIV in animal models. Although numerous studies have characterized the

  15. Trans-dissemination of exosomes from HIV-1-infected cells fosters both HIV-1 trans-infection in resting CD4+ T lymphocytes and reactivation of the HIV-1 reservoir.

    Science.gov (United States)

    Chiozzini, Chiara; Arenaccio, Claudia; Olivetta, Eleonora; Anticoli, Simona; Manfredi, Francesco; Ferrantelli, Flavia; d'Ettorre, Gabriella; Schietroma, Ivan; Andreotti, Mauro; Federico, Maurizio

    2017-09-01

    Intact HIV-1 and exosomes can be internalized by dendritic cells (DCs) through a common pathway leading to their transmission to CD4 + T lymphocytes by means of mechanisms defined as trans-infection and trans-dissemination, respectively. We previously reported that exosomes from HIV-1-infected cells activate both uninfected quiescent CD4 + T lymphocytes, which become permissive to HIV-1, and latently infected cells, with release of HIV-1 particles. However, nothing is known about the effects of trans-dissemination of exosomes produced by HIV-1-infected cells on uninfected or latently HIV-1-infected CD4 + T lymphocytes. Here, we report that trans-dissemination of exosomes from HIV-1-infected cells induces cell activation in resting CD4 + T lymphocytes, which appears stronger with mature than immature DCs. Using purified preparations of both HIV-1 and exosomes, we observed that mDC-mediated trans-dissemination of exosomes from HIV-1-infected cells to resting CD4 + T lymphocytes induces efficient trans-infection and HIV-1 expression in target cells. Most relevant, when both mDCs and CD4 + T lymphocytes were isolated from combination anti-retroviral therapy (ART)-treated HIV-1-infected patients, trans-dissemination of exosomes from HIV-1-infected cells led to HIV-1 reactivation from the viral reservoir. In sum, our data suggest a role of exosome trans-dissemination in both HIV-1 spread in the infected host and reactivation of the HIV-1 reservoir.

  16. HMBA Enhances Prostratin-Induced Activation of Latent HIV-1 via Suppressing the Expression of Negative Feedback Regulator A20/TNFAIP3 in NF-κB Signaling

    Directory of Open Access Journals (Sweden)

    Duchu Chen

    2016-01-01

    Full Text Available In the past decade, much emphasis has been put on the transcriptional activation of HIV-1, which is proposed as a promised strategy for eradicating latent HIV-1 provirus. Two drugs, prostratin and hexamethylene bisacetamide (HMBA, have shown potent effects as inducers for releasing HIV-1 latency when used alone or in combination, although their cellular target(s are currently not well understood, especially under drug combination. Here, we have shown that HMBA and prostratin synergistically release HIV-1 latency via different mechanisms. While prostratin strongly stimulates HMBA-induced HIV-1 transcription via improved P-TEFb activation, HMBA is capable of boosting NF-κB-dependent transcription initiation by suppressing prostratin-induced expression of the deubiquitinase A20, a negative feedback regulator in the NF-κB signaling pathway. In addition, HMBA was able to increase prostratin-induced phosphorylation and degradation of NF-κB inhibitor IκBα, thereby enhancing and prolonging prostratin-induced nuclear translocation of NF-κB, a prerequisite for stimulation of transcription initiation. Thus, by blocking the negative feedback circuit, HMBA functions as a signaling enhancer of the NF-κB signaling pathway.

  17. HIV-1 molecular epidemiology among newly diagnosed HIV-1 individuals in Hebei, a low HIV prevalence province in China.

    Directory of Open Access Journals (Sweden)

    Xinli Lu

    Full Text Available New human immunodeficiency virus type 1 (HIV-1 diagnoses are increasing rapidly in Hebei. The aim of this study presents the most extensive HIV-1 molecular epidemiology investigation in Hebei province in China thus far. We have carried out the most extensive systematic cross-sectional study based on newly diagnosed HIV-1 positive individuals in 2013, and characterized the molecular epidemiology of HIV-1 based on full length gag-partial pol gene sequences in the whole of Hebei. Nine HIV-1 genotypes based on full length gag-partial pol gene sequence were identified among 610 newly diagnosed naïve individuals. The four main genotypes were circulating recombinant form (CRF01_AE (53.4%, CRF07_BC (23.4%, subtype B (15.9%, and unique recombinant forms URFs (4.9%. Within 1 year, three new genotypes (subtype A1, CRF55_01B, CRF65_cpx, unknown before in Hebei, were first found among men who have sex with men (MSM. All nine genotypes were identified in the sexually contracted HIV-1 population. Among 30 URFs, six recombinant patterns were revealed, including CRF01_AE/BC (40.0%, CRF01_AE/B (23.3%, B/C (16.7%, CRF01_AE/C (13.3%, CRF01_AE/B/A2 (3.3% and CRF01_AE/BC/A2 (3.3%, plus two potential CRFs. This study elucidated the complicated characteristics of HIV-1 molecular epidemiology in a low HIV-1 prevalence northern province of China and revealed the high level of HIV-1 genetic diversity. All nine HIV-1 genotypes circulating in Hebei have spread out of their initial risk groups into the general population through sexual contact, especially through MSM. This highlights the urgency of HIV prevention and control in China.

  18. HIV-1 molecular epidemiology among newly diagnosed HIV-1 individuals in Hebei, a low HIV prevalence province in China.

    Science.gov (United States)

    Lu, Xinli; Kang, Xianjiang; Liu, Yongjian; Cui, Ze; Guo, Wei; Zhao, Cuiying; Li, Yan; Chen, Suliang; Li, Jingyun; Zhang, Yuqi; Zhao, Hongru

    2017-01-01

    New human immunodeficiency virus type 1 (HIV-1) diagnoses are increasing rapidly in Hebei. The aim of this study presents the most extensive HIV-1 molecular epidemiology investigation in Hebei province in China thus far. We have carried out the most extensive systematic cross-sectional study based on newly diagnosed HIV-1 positive individuals in 2013, and characterized the molecular epidemiology of HIV-1 based on full length gag-partial pol gene sequences in the whole of Hebei. Nine HIV-1 genotypes based on full length gag-partial pol gene sequence were identified among 610 newly diagnosed naïve individuals. The four main genotypes were circulating recombinant form (CRF)01_AE (53.4%), CRF07_BC (23.4%), subtype B (15.9%), and unique recombinant forms URFs (4.9%). Within 1 year, three new genotypes (subtype A1, CRF55_01B, CRF65_cpx), unknown before in Hebei, were first found among men who have sex with men (MSM). All nine genotypes were identified in the sexually contracted HIV-1 population. Among 30 URFs, six recombinant patterns were revealed, including CRF01_AE/BC (40.0%), CRF01_AE/B (23.3%), B/C (16.7%), CRF01_AE/C (13.3%), CRF01_AE/B/A2 (3.3%) and CRF01_AE/BC/A2 (3.3%), plus two potential CRFs. This study elucidated the complicated characteristics of HIV-1 molecular epidemiology in a low HIV-1 prevalence northern province of China and revealed the high level of HIV-1 genetic diversity. All nine HIV-1 genotypes circulating in Hebei have spread out of their initial risk groups into the general population through sexual contact, especially through MSM. This highlights the urgency of HIV prevention and control in China.

  19. Anti-HIV-1 activity of flavonoid myricetin on HIV-1 infection in a dual-chamber in vitro model.

    Directory of Open Access Journals (Sweden)

    Silvana Pasetto

    Full Text Available HIV infection by sexual transmission remains an enormous global health concern. More than 1 million new infections among women occur annually. Microbicides represent a promising prevention strategy that women can easily control. Among emerging therapies, natural small molecules such as flavonoids are an important source of new active substances. In this study we report the in vitro cytotoxicity and anti-HIV-1 and microbicide activity of the following flavonoids: Myricetin, Quercetin and Pinocembrin. Cytotoxicity tests were conducted on TZM-bl, HeLa, PBMC, and H9 cell cultures using 0.01-100 µM concentrations. Myricetin presented the lowest toxic effect, with Quercetin and Pinocembrin relatively more toxic. The anti-HIV-1 activity was tested with TZM-bl cell plus HIV-1 BaL (R5 tropic, H9 and PBMC cells plus HIV-1 MN (X4 tropic, and the dual tropic (X4R5 HIV-1 89.6. All flavonoids showed anti-HIV activity, although Myricetin was more effective than Quercetin or Pinocembrin. In TZM-bl cells, Myricetin inhibited ≥90% of HIV-1 BaL infection. The results were confirmed by quantification of HIV-1 p24 antigen in supernatant from H9 and PBMC cells following flavonoid treatment. In H9 and PBMC cells infected by HIV-1 MN and HIV-1 89.6, Myricetin showed more than 80% anti-HIV activity. Quercetin and Pinocembrin presented modest anti-HIV activity in all experiments. Myricetin activity was tested against HIV-RT and inhibited the enzyme by 49%. Microbicide activities were evaluated using a dual-chamber female genital tract model. In the in vitro microbicide activity model, Myricetin showed promising results against different strains of HIV-1 while also showing insignificant cytotoxic effects. Further studies of Myricetin should be performed to identify its molecular targets in order to provide a solid biological foundation for translational research.

  20. Molecular Basis for Drug Resistance in HIV-1 Protease

    Directory of Open Access Journals (Sweden)

    Celia A. Schiffer

    2010-11-01

    Full Text Available HIV-1 protease is one of the major antiviral targets in the treatment of patients infected with HIV-1. The nine FDA approved HIV-1 protease inhibitors were developed with extensive use of structure-based drug design, thus the atomic details of how the inhibitors bind are well characterized. From this structural understanding the molecular basis for drug resistance in HIV-1 protease can be elucidated. Selected mutations in response to therapy and diversity between clades in HIV-1 protease have altered the shape of the active site, potentially altered the dynamics and even altered the sequence of the cleavage sites in the Gag polyprotein. All of these interdependent changes act in synergy to confer drug resistance while simultaneously maintaining the fitness of the virus. New strategies, such as incorporation of the substrate envelope constraint to design robust inhibitors that incorporate details of HIV-1 protease’s function and decrease the probability of drug resistance, are necessary to continue to effectively target this key protein in HIV-1 life cycle.

  1. Correlates of HIV-1 genital shedding in Tanzanian women.

    Directory of Open Access Journals (Sweden)

    Clare Tanton

    2011-03-01

    Full Text Available Understanding the correlates of HIV shedding is important to inform strategies to reduce HIV infectiousness. We examined correlates of genital HIV-1 RNA in women who were seropositive for both herpes simplex virus (HSV-2 and HIV-1 and who were enrolled in a randomised controlled trial of HSV suppressive therapy (aciclovir 400 mg b.i.d vs. placebo in Tanzania.Samples, including a cervico-vaginal lavage, were collected and tested for genital HIV-1 and HSV and reproductive tract infections (RTIs at randomisation and 6, 12 and 24 months follow-up. Data from all women at randomisation and women in the placebo arm during follow-up were analysed using generalised estimating equations to determine the correlates of cervico-vaginal HIV-1 RNA detection and load.Cervico-vaginal HIV-1 RNA was detected at 52.0% of 971 visits among 482 women, and was independently associated with plasma viral load, presence of genital ulcers, pregnancy, bloody cervical or vaginal discharge, abnormal vaginal discharge, cervical ectopy, Neisseria gonorrhoeae, Chlamydia trachomatis, Trichomonas vaginalis, an intermediate bacterial vaginosis score and HSV DNA detection. Similar factors were associated with genital HIV-1 RNA load.RTIs were associated with increased presence and quantity of genital HIV-1 RNA in this population. These results highlight the importance of integrating effective RTI treatment into HIV care services.

  2. CCR5 Gene Disruption via Lentiviral Vectors Expressing Cas9 and Single Guided RNA Renders Cells Resistant to HIV-1 Infection

    Science.gov (United States)

    Liu, Jingjing; Zhang, Di; Kimata, Jason T.; Zhou, Paul

    2014-01-01

    CCR5, a coreceptor for HIV-1 entry, is a major target for drug and genetic intervention against HIV-1. Genetic intervention strategies have knocked down CCR5 expression levels by shRNA or disrupted the CCR5 gene using zinc finger nucleases (ZFN) or Transcription activator-like effector nuclease (TALEN). In the present study, we silenced CCR5 via CRISPR associated protein 9 (Cas9) and single guided RNAs (sgRNAs). We constructed lentiviral vectors expressing Cas9 and CCR5 sgRNAs. We show that a single round transduction of lentiviral vectors expressing Cas9 and CCR5 sgRNAs into HIV-1 susceptible human CD4+ cells yields high frequencies of CCR5 gene disruption. CCR5 gene-disrupted cells are not only resistant to R5-tropic HIV-1, including transmitted/founder (T/F) HIV-1 isolates, but also have selective advantage over CCR5 gene-undisrupted cells during R5-tropic HIV-1 infection. Importantly, using T7 endonuclease I assay we did not detect genome mutations at potential off-target sites that are highly homologous to these CCR5 sgRNAs in stably transduced cells even at 84 days post transduction. Thus we conclude that silencing of CCR5 via Cas9 and CCR5-specific sgRNAs could be a viable alternative strategy for engineering resistance against HIV-1. PMID:25541967

  3. Novel host restriction factors implicated in HIV-1 replication.

    Science.gov (United States)

    Ghimire, Dibya; Rai, Madhu; Gaur, Ritu

    2018-04-01

    Human immunodeficiency virus-1 (HIV-1) is known to interact with multiple host cellular proteins during its replication in the target cell. While many of these host cellular proteins facilitate viral replication, a number of them are reported to inhibit HIV-1 replication at various stages of its life cycle. These host cellular proteins, which are known as restriction factors, constitute an integral part of the host's first line of defence against the viral pathogen. Since the discovery of apolipoprotein B mRNA-editing enzyme 3G (APOBEC3G) as an HIV-1 restriction factor, several human proteins have been identified that exhibit anti-HIV-1 restriction. While each restriction factor employs a distinct mechanism of inhibition, the HIV-1 virus has equally evolved complex counter strategies to neutralize their inhibitory effect. APOBEC3G, tetherin, sterile alpha motif and histidine-aspartate domain 1 (SAMHD1), and trim-5α are some of the best known HIV-1 restriction factors that have been studied in great detail. Recently, six novel restriction factors were discovered that exhibit significant antiviral activity: endoplasmic reticulum α1,2-mannosidase I (ERManI), translocator protein (TSPO), guanylate-binding protein 5 (GBP5), serine incorporator (SERINC3/5) and zinc-finger antiviral protein (ZAP). The focus of this review is to discuss the antiviral mechanism of action of these six restriction factors and provide insights into the probable counter-evasion strategies employed by the HIV-1 virus. The recent discovery of new restriction factors substantiates the complex host-pathogen interactions occurring during HIV-1 pathogenesis and makes it imperative that further investigations are conducted to elucidate the molecular basis of HIV-1 replication.

  4. HIV-1 Replication and the Cellular Eukaryotic Translation Apparatus

    Directory of Open Access Journals (Sweden)

    Santiago Guerrero

    2015-01-01

    Full Text Available Eukaryotic translation is a complex process composed of three main steps: initiation, elongation, and termination. During infections by RNA- and DNA-viruses, the eukaryotic translation machinery is used to assure optimal viral protein synthesis. Human immunodeficiency virus type I (HIV-1 uses several non-canonical pathways to translate its own proteins, such as leaky scanning, frameshifting, shunt, and cap-independent mechanisms. Moreover, HIV-1 modulates the host translation machinery by targeting key translation factors and overcomes different cellular obstacles that affect protein translation. In this review, we describe how HIV-1 proteins target several components of the eukaryotic translation machinery, which consequently improves viral translation and replication.

  5. HIV-1 Non-Nucleoside Reverse Transcriptase Inhibitors

    DEFF Research Database (Denmark)

    Vanangamudi, Murugesan; Poongavanam, Vasanthanathan; Namasivayam, Vigneshwaran

    2017-01-01

    BACKGROUND: Design of inhibitors for HIV-1 reverse transcriptase inhibition (HIV-1 RT) is one of the successful chemotherapies for the treatment of HIV infection. Among the inhibitors available for HIV-1 RT, non-nucleoside reverse transcriptase inhibitors (NNRTIs) have shown to be very promising......: The conformation dependent-alignment based (CoMFA and CoMSIA) methods have been proven very successful ligand based strategy in the drug design. Here, CoMFA and CoMSIA studies reported for structurally distinct NNRTIs including thiazolobenzimidazole, dipyridodiazepinone, 1,1,3-trioxo [1,2,4]-thiadiazine...

  6. Novel Latency Reversal Agents for HIV-1 Cure.

    Science.gov (United States)

    Spivak, Adam M; Planelles, Vicente

    2018-01-29

    Antiretroviral therapy (ART) has rendered HIV-1 infection a treatable illness; however, ART is not curative owing to the persistence of replication-competent, latent proviruses in long-lived resting T cells. Strategies that target these latently infected cells and allow immune recognition and clearance of this reservoir will be necessary to eradicate HIV-1 in infected individuals. This review describes current pharmacologic approaches to reactivate the latent reservoir so that infected cells can be recognized and targeted, with the ultimate goal of achieving an HIV-1 cure.

  7. Towards an HIV-1 cure: measuring the latent reservoir.

    Science.gov (United States)

    Bruner, Katherine M; Hosmane, Nina N; Siliciano, Robert F

    2015-04-01

    The latent reservoir (LR) of HIV-1 in resting memory CD4(+) T cells serves as a major barrier to curing HIV-1 infection. While many PCR- and culture-based assays have been used to measure the size of the LR, correlation between results of different assays is poor and recent studies indicate that no available assay provides an accurate measurement of reservoir size. The discrepancies between assays are a hurdle to clinical trials that aim to measure the efficacy of HIV-1 eradication strategies. Here we describe the advantages and disadvantages of various approaches to measuring the LR. Copyright © 2015 Elsevier Ltd. All rights reserved.

  8. Towards an HIV-1 cure: measuring the latent reservoir

    Science.gov (United States)

    Bruner, Katherine M.; Hosmane, Nina N.; Siliciano, Robert F.

    2015-01-01

    The latent reservoir of HIV-1 in resting memory CD4+ T cells serves as a major barrier to curing HIV-1 infection. While many PCR- and culture-based assays have been used to measure the size of the latent reservoir, correlation between results of different assays is poor and recent studies indicate that no available assay provides an accurate measurement of reservoir size. The discrepancies between assays are a hurdle to clinical trials that aim to measure the efficacy of HIV-1 eradication strategies. Here we describe the advantages and disadvantages of various approaches to measure the latent reservoir. PMID:25747663

  9. Sensitive non-radioactive detection of HIV-1

    DEFF Research Database (Denmark)

    Teglbjærg, Lars Stubbe; Nielsen, C; Hansen, J E

    1992-01-01

    This report describes the use of the polymerase chain reaction (PCR) for the non-radioactive detection of HIV-1 proviral genomic sequences in HIV-1 infected cells. We have developed a sensitive assay, using three different sets of nested primers and our results show that this method is superior...... to standard PCR for the detection of HIV-1 DNA. The assay described features the use of a simple and inexpensive sample preparation technique and a non-radioactive hybridization procedure for confirmation of results. To test the suitability of the assay for clinical purposes, we tested cell samples from 76...

  10. The cross-talk of HIV-1 Tat and methamphetamine in HIV-associated neurocognitive disorders

    Directory of Open Access Journals (Sweden)

    Susana T Valente

    2015-10-01

    Full Text Available Antiretroviral therapy (ART has dramatically improved the lives of HIV-1 infected individuals. Nonetheless, HIV associated neurocognitive disorders (HAND, which range from undetectable neurocognitive impairments to severe dementia, still affect approximately 50% of the infected population, hampering their quality of life.The persistence of HAND is promoted by several factors, including longer life expectancies, the residual levels of virus in the central nervous system and the continued presence of HIV-1 regulatory proteins such as the transactivator of transcription (Tat in the brain. Tat is a secreted viral protein that crosses the blood brain barrier into the central nervous system, where it has the ability to directly act on neurons and non-neuronal cells alike. These actions result in the release of soluble factors involved in inflammation, oxidative stress and excitotoxicity, ultimately resulting in neuronal damage. The percentage of methamphetamine abusers is high among the HIV-1-positive population compared to the general population. On the other hand, methamphetamine abuse is correlated with increased viral replication, enhanced Tat-mediated neurotoxicity and neurocognitive impairments. Although several strategies have been investigated to reduce HAND and methamphetamine use, no clinically approved treatment is currently available. Here, we review the latest findings of the effects of Tat and methamphetamine in HAND and discuss a few promising potential therapeutic developments.

  11. Compounds producing an effective combinatorial regimen for disruption of HIV-1 latency.

    Science.gov (United States)

    Hashemi, Pargol; Barreto, Kris; Bernhard, Wendy; Lomness, Adam; Honson, Nicolette; Pfeifer, Tom A; Harrigan, P Richard; Sadowski, Ivan

    2018-02-01

    Highly active antiretroviral therapy (HAART) has improved the outlook for the HIV epidemic, but does not provide a cure. The proposed "shock-and-kill" strategy is directed at inducing latent HIV reservoirs, which may then be purged via boosted immune response or targeting infected cells. We describe five novel compounds that are capable of reversing HIV latency without affecting the general T-cell activation state. The new compounds exhibit synergy for reactivation of latent provirus with other latency-reversing agents (LRAs), in particular ingenol-3-angelate/PEP005. One compound, designated PH02, was efficient at reactivating viral transcription in several cell lines bearing reporter HIV-1 at different integration sites. Furthermore, it was capable of reversing latency in resting CD4 + T lymphocytes from latently infected aviremic patient cells on HAART, while producing minimal cellular toxicity. The combination of PH02 and PEP005 produces a strong synergistic effect for reactivation, as demonstrated through a quantitative viral outgrowth assay (qVOA), on CD4 + T lymphocytes from HIV-1-infected individuals. We propose that the PH02/PEP005 combination may represent an effective novel treatment for abrogating persistent HIV-1 infection. © 2017 The Authors. Published under the terms of the CC BY 4.0 license.

  12. A Rough Set-Based Model of HIV-1 Reverse Transcriptase Resistome

    Directory of Open Access Journals (Sweden)

    Marcin Kierczak

    2009-10-01

    Full Text Available Reverse transcriptase (RT is a viral enzyme crucial for HIV-1 replication. Currently, 12 drugs are targeted against the RT. The low fidelity of the RT-mediated transcription leads to the quick accumulation of drug-resistance mutations. The sequence-resistance relationship remains only partially understood. Using publicly available data collected from over 15 years of HIV proteome research, we have created a general and predictive rule-based model of HIV-1 resistance to eight RT inhibitors. Our rough set-based model considers changes in the physicochemical properties of a mutated sequence as compared to the wild-type strain. Thanks to the application of the Monte Carlo feature selection method, the model takes into account only the properties that significantly contribute to the resistance phenomenon. The obtained results show that drug-resistance is determined in more complex way than believed. We confirmed the importance of many resistance-associated sites, found some sites to be less relevant than formerly postulated and— more importantly—identified several previously neglected sites as potentially relevant. By mapping some of the newly discovered sites on the 3D structure of the RT, we were able to suggest possible molecular-mechanisms of drug-resistance. Importantly, our model has the ability to generalize predictions to the previously unseen cases. The study is an example of how computational biology methods can increase our understanding of the HIV-1 resistome.

  13. BET bromodomain inhibition as a novel strategy for reactivation of HIV-1.

    Science.gov (United States)

    Banerjee, Camellia; Archin, Nancie; Michaels, Daniel; Belkina, Anna C; Denis, Gerald V; Bradner, James; Sebastiani, Paola; Margolis, David M; Montano, Monty

    2012-12-01

    The persistence of latent HIV-1 remains a major challenge in therapeutic efforts to eradicate infection. We report the capacity for HIV reactivation by a selective small molecule inhibitor of BET family bromodomains, JQ1, a promising therapeutic agent with antioncogenic properties. JQ1 reactivated HIV transcription in models of latent T cell infection and latent monocyte infection. We also tested the effect of exposure to JQ1 to allow recovery of replication-competent HIV from pools of resting CD4(+) T cells isolated from HIV-infected, ART-treated patients. In one of three patients, JQ1 allowed recovery of virus at a frequency above unstimulated conditions. JQ1 potently suppressed T cell proliferation with minimal cytotoxic effect. Transcriptional profiling of T cells with JQ1 showed potent down-regulation of T cell activation genes, including CD3, CD28, and CXCR4, similar to HDAC inhibitors, but JQ1 also showed potent up-regulation of chromatin modification genes, including SIRT1, HDAC6, and multiple lysine demethylases (KDMs). Thus, JQ1 reactivates HIV-1 while suppressing T cell activation genes and up-regulating histone modification genes predicted to favor increased Tat activity. Thus, JQ1 may be useful in studies of potentially novel mechanisms for transcriptional control as well as in translational efforts to identify therapeutic molecules to achieve viral eradication.

  14. Seroprevalence and molecular epidemiology of HTLV-1 isolates from HIV-1 co-infected women in Feira de Santana, Bahia, Brazil.

    Science.gov (United States)

    de Almeida Rego, Filipe Ferreira; Mota-Miranda, Aline; de Souza Santos, Edson; Galvão-Castro, Bernardo; Alcantara, Luiz Carlos

    2010-12-01

    HTLV-1/HIV-1 co-infection is associated with severe clinical manifestations, marked immunodeficiency, and opportunistic pathogenic infections, as well as risk behavior. Salvador, the capital of the State of Bahia, Brazil, has the highest HTLV-1 prevalence (1.74%) found in Brazil. Few studies exist which describe this co-infection found in Salvador and its surrounding areas, much less investigate how these viruses circulate or assess the relationship between them. To describe the epidemiological and molecular features of HTLV in HIV co-infected women. To investigate the prevalence of HTLV/HIV co-infection in surrounding areas, as well as the molecular epidemiology of HTLV, a cross sectional study was carried out involving 107 women infected with HIV-1 from the STD/HIV/AIDS Reference Center located in the neighboring City of Feira de Santana. Patient samples were submitted to ELISA, and HTLV infection was confirmed using Western Blot and Polymerase Chain Reaction (PCR). Phylogenetic analysis using Neighbor-Joining (NJ) and Maximum Likelihood (ML) was performed on HTLV LTR sequences in order to gain further insights about molecular epidemiology and the origins of this virus in Bahia. Four out of five reactive samples were confirmed to be infected with HTLV-1, and one with HTLV-2. The seroprevalence of HTLV among HIV-1 co-infected women was 4.7%. Phylogenetic analysis of the LTR region from four HTLV-1 sequences showed that all isolates were clustered into the main Latin American group within the Transcontinental subgroup of the Cosmopolitan subtype. The HTLV-2 sequence was classified as the HTLV-2c subtype. It was also observed that four HTLV/HIV-1 co-infected women exhibited risk behavior with two having parenteral exposure, while another two were sex workers. This article describes the characteristics of co-infected patients. This co-infection is known to be severe and further studies should be conducted to confirm the suggestion that HTLV-1 is spreading from

  15. Structural Study of a New HIV-1 Entry Inhibitor and Interaction with the HIV-1 Fusion Peptide in Dodecylphosphocholine Micelles.

    Science.gov (United States)

    Pérez, Yolanda; Gómara, Maria José; Yuste, Eloísa; Gómez-Gutierrez, Patricia; Pérez, Juan Jesús; Haro, Isabel

    2017-08-25

    Previous studies support the hypothesis that the envelope GB virus C (GBV-C) E1 protein interferes the HIV-1 entry and that a peptide, derived from the region 139-156 of this protein, has been defined as a novel HIV-1 entry inhibitor. In this work, we firstly focus on the characterization of the structural features of this peptide, which are determinant for its anti-HIV-1 activity and secondly, on the study of its interaction with the proposed viral target (i.e., the HIV-1 fusion peptide). We report the structure of the peptide determined by NMR spectroscopy in dodecylphosphocholine (DPC) micelles solved by using restrained molecular dynamics calculations. The acquisition of different NMR experiments in DPC micelles (i.e., peptide-peptide titration, diffusion NMR spectroscopy, and addition of paramagnetic relaxation agents) allows a proposal of an inhibition mechanism. We conclude that a 18-mer peptide from the non-pathogenic E1 GBV-C protein, with a helix-turn-helix structure inhibits HIV-1 by binding to the HIV-1 fusion peptide at the membrane level, thereby interfering with those domains in the HIV-1, which are critical for stabilizing the six-helix bundle formation in a membranous environment. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

  16. HIV transcription is induced with some forms of cell killing

    International Nuclear Information System (INIS)

    Woloschak, G.E.; Schreck, S.; Chang-Liu, C.-M.; Libertin, C.R.

    1996-01-01

    Using HeLa cells stably transfected with an HIV-LTR-CAT construct', we demonstrated a peak in CAT induction that occurs in viable (but not necessarily cell-division-competent) cells 24 h following exposure to some cell-killing agents. Γ rays were the only cell-killing agent which did not induce HIV transcription; this can be attributed to the fact that γ-ray-induced apoptotic death requires function p53, which is missing in HeLa cells. For all other agents, HIV-LTR induction was dose-dependent and correlated with the amount of cell killing that occurred in the culture

  17. Nanoparticles containing siRNA to silence CD4 and CCR5 reduce expression of these receptors and inhibit HIV-1 infection in human female reproductive tract tissue explants

    Directory of Open Access Journals (Sweden)

    Susan K. Eszterhas

    2011-09-01

    Full Text Available Human Immunodeficiency Virus-type 1 (HIV- 1 binds to CD4 and CCR5 receptors on target cells in the human female reproductive tract. We sought to determine whether reducing levels of messenger RNA (mRNA transcripts that encode these receptors in female reproductive tract cells could protect mucosal tissue explants from HIV- 1 infection. Explants prepared from the endometrium, endocervix, and ectocervix of hysterectomy tissues from HIV-1 sero-negative women were exposed to nanoparticles containing CD4- and CCR5-specific short-interfering RNA (siRNA sequences. Explants were then exposed two days later to HIV-1, and HIV-1 reverse transcripts were measured five days post-infection. Explants treated with nanoparticles containing CD4- and CCR5-specific siRNA showed reduced levels of CD4 and CCR5 transcripts, and significantly lower levels of HIV-1 reverse transcripts compared to those treated with an irrelevant siRNA. In female reproductive tract explants and in peripheral blood cell cultures, siRNA transfection induced the secretion of IFN-alpha (IFN-α, a potent antiviral cytokine. In female mice, murine-specific Cd4-siRNA nanoparticles instilled within the uterus significantly reduced murine Cd4 transcripts by day 3. Our findings demonstrate that siRNA nanoparticles reduce expression of HIV-1 infectivity receptors in human female reproductive tract tissues and also inhibit HIV-1 infection. Murine studies demonstrate that nanoparticles can penetrate the reproductive tract tissues in vivo and silence gene expression. The induction of IFN-α after siRNA transfection can potentially contribute to the antiviral effect. These findings support the therapeutic development of nanoparticles to deliver siRNA molecules to silence host cell receptors in the female reproductive tract as a novel microbicide to inhibit mucosal HIV-1 transmission.

  18. Isolation and characterization of a replication-competent molecular clone of an HIV-1 circulating recombinant form (CRF33_01B.

    Directory of Open Access Journals (Sweden)

    Kok Keng Tee

    Full Text Available A growing number of emerging HIV-1 recombinants classified as circulating recombinant forms (CRFs have been identified in Southeast Asia in recent years, establishing a molecular diversity of increasing complexity in the region. Here, we constructed a replication-competent HIV-1 clone for CRF33_01B (designated p05MYKL045.1, a newly identified recombinant comprised of CRF01_AE and subtype B. p05MYKL045.1 was reconstituted by cloning of the near full-length HIV-1 sequence from a newly-diagnosed individual presumably infected heterosexually in Kuala Lumpur, Malaysia. The chimeric clone, which contains the 5' LTR (long terminal repeat region of p93JP-NH1 (a previously isolated CRF01_AE infectious clone, showed robust viral replication in the human peripheral blood mononuclear cells. This clone demonstrated robust viral propagation and profound syncytium formation in CD4+, CXCR4-expressing human glioma NP-2 cells, indicating that p05MYKL045.1 is a CXCR4-using virus. Viral propagation, however, was not detected in various human T cell lines including MT-2, M8166, Sup-T1, H9, Jurkat, Molt-4 and PM1. p05MYKL045.1 appears to proliferate only in restricted host range, suggesting that unknown viral and/or cellular host factors may play a role in viral infectivity and replication in human T cell lines. Availability of a CRF33_01B molecular clone will be useful in facilitating the development of vaccine candidates that match the HIV-1 strains circulating in Southeast Asia.

  19. Characterization of two candidate genes, NCoA3 and IRF8, potentially involved in the control of HIV-1 latency

    Directory of Open Access Journals (Sweden)

    Gumez Audrey

    2005-11-01

    Full Text Available Abstract Background The persistence of latent HIV-1 reservoirs is the principal barrier preventing the eradication of HIV-1 infection in patients by current antiretroviral therapy. It is thus crucial to understand the molecular mechanisms involved in the establishment, maintenance and reactivation of HIV-1 latency. Since chromatin remodeling has been implicated in the transcriptional reactivation of the HIV-1 promoter, we assessed the role of the histone deacetylase inhibitor sodium butyrate (NaB on two HIV-1 latently infected cell lines (U1 and ACH-2 gene expression. Results Analysis of microarrays data led us to select two candidate genes: NCoA3 (Nuclear Receptor Coactivator 3, a nuclear receptor coactivator and IRF8 (Interferon Regulatory Factor 8, an interferon regulatory factor. NCoA3 gene expression is upregulated following NaB treatment of latently infected cells whereas IRF8 gene expression is strongly downregulated in the promonocytic cell line following NaB treatment. Their differential expressions were confirmed at the transcriptional and translational levels. Moreover, NCoA3 gene expression was also upregulated after treatment of U1 and ACH-2 cells with phorbol myristyl acetate (PMA but not trichostatin A (TSA and after treatment with NaB of two others HIV-1 latently infected cell lines (OM10.1 and J1.1. IRF8 gene is only expressed in U1 cells and was also downregulated after treatment with PMA or TSA. Functional analyses confirmed that NCoA3 synergizes with Tat to enhance HIV-1 promoter transcription and that IRF8 represses the IRF1-mediated activation through the HIV-1 promoter Interferon-stimulated response element (ISRE. Conclusion These results led us to postulate that NCoA3 could be involved in the transcriptional reactivation of the HIV-1 promoter from latency and that IRF8 may contribute to the maintenance of the latent state in the promonocytic cell line. Implication of these factors in the maintenance or reactivation of the

  20. Hepatitis B Virus Vaccination in HIV-1-Infected Young Adults: A Tool to Reduce the Size of HIV-1 Reservoirs?

    Science.gov (United States)

    Bekele, Yonas; Graham, Rebecka Lantto; Soeria-Atmadja, Sandra; Nasi, Aikaterini; Zazzi, Maurizio; Vicenti, Ilaria; Naver, Lars; Nilsson, Anna; Chiodi, Francesca

    2017-01-01

    During anti-retroviral therapy (ART) HIV-1 persists in cellular reservoirs, mostly represented by CD4+ memory T cells. Several approaches are currently being undertaken to develop a cure for HIV-1 infection through elimination (or reduction) of these reservoirs. Few studies have so far been conducted to assess the possibility of reducing the size of HIV-1 reservoirs through vaccination in virologically controlled HIV-1-infected children. We recently conducted a vaccination study with a combined hepatitis A virus (HAV) and hepatitis B virus (HBV) vaccine in 22 HIV-1-infected children. We assessed the size of the virus reservoir, measured as total HIV-1 DNA copies in blood cells, pre- and postvaccination. In addition, we investigated by immunostaining whether the frequencies of CD4+ and CD8+ T cells and parameters of immune activation and proliferation on these cells were modulated by vaccination. At 1 month from the last vaccination dose, we found that 20 out of 22 children mounted a serological response to HBV; a majority of children had antibodies against HAV at baseline. The number of HIV-1 DNA copies in blood at 1 month postvaccination was reduced in comparison to baseline although this reduction was not statistically significant. A significant reduction of HIV-1 DNA copies in blood following vaccination was found in 12 children. The frequencies of CD4+ (naïve, effector memory) and CD8+ (central memory) T-cell subpopulations changed following vaccinations and a reduction in the activation and proliferation pattern of these cells was also noticed. Multivariate linear regression analysis revealed that the frequency of CD8+ effector memory T cells prior to vaccination was strongly predictive of the reduction of HIV-1 DNA copies in blood following vaccination of the 22 HIV-1-infected children. The results of this study suggest a beneficial effect of vaccination to reduce the size of virus reservoir in HIV-1-infected children receiving ART. A reduced frequency of

  1. Hepatitis B Virus Vaccination in HIV-1-Infected Young Adults: A Tool to Reduce the Size of HIV-1 Reservoirs?

    Directory of Open Access Journals (Sweden)

    Yonas Bekele

    2018-01-01

    Full Text Available During anti-retroviral therapy (ART HIV-1 persists in cellular reservoirs, mostly represented by CD4+ memory T cells. Several approaches are currently being undertaken to develop a cure for HIV-1 infection through elimination (or reduction of these reservoirs. Few studies have so far been conducted to assess the possibility of reducing the size of HIV-1 reservoirs through vaccination in virologically controlled HIV-1-infected children. We recently conducted a vaccination study with a combined hepatitis A virus (HAV and hepatitis B virus (HBV vaccine in 22 HIV-1-infected children. We assessed the size of the virus reservoir, measured as total HIV-1 DNA copies in blood cells, pre- and postvaccination. In addition, we investigated by immunostaining whether the frequencies of CD4+ and CD8+ T cells and parameters of immune activation and proliferation on these cells were modulated by vaccination. At 1 month from the last vaccination dose, we found that 20 out of 22 children mounted a serological response to HBV; a majority of children had antibodies against HAV at baseline. The number of HIV-1 DNA copies in blood at 1 month postvaccination was reduced in comparison to baseline although this reduction was not statistically significant. A significant reduction of HIV-1 DNA copies in blood following vaccination was found in 12 children. The frequencies of CD4+ (naïve, effector memory and CD8+ (central memory T-cell subpopulations changed following vaccinations and a reduction in the activation and proliferation pattern of these cells was also noticed. Multivariate linear regression analysis revealed that the frequency of CD8+ effector memory T cells prior to vaccination was strongly predictive of the reduction of HIV-1 DNA copies in blood following vaccination of the 22 HIV-1-infected children. The results of this study suggest a beneficial effect of vaccination to reduce the size of virus reservoir in HIV-1-infected children receiving ART. A reduced

  2. Hepatitis B Virus Vaccination in HIV-1-Infected Young Adults: A Tool to Reduce the Size of HIV-1 Reservoirs?

    Science.gov (United States)

    Bekele, Yonas; Graham, Rebecka Lantto; Soeria-Atmadja, Sandra; Nasi, Aikaterini; Zazzi, Maurizio; Vicenti, Ilaria; Naver, Lars; Nilsson, Anna; Chiodi, Francesca

    2018-01-01

    During anti-retroviral therapy (ART) HIV-1 persists in cellular reservoirs, mostly represented by CD4+ memory T cells. Several approaches are currently being undertaken to develop a cure for HIV-1 infection through elimination (or reduction) of these reservoirs. Few studies have so far been conducted to assess the possibility of reducing the size of HIV-1 reservoirs through vaccination in virologically controlled HIV-1-infected children. We recently conducted a vaccination study with a combined hepatitis A virus (HAV) and hepatitis B virus (HBV) vaccine in 22 HIV-1-infected children. We assessed the size of the virus reservoir, measured as total HIV-1 DNA copies in blood cells, pre- and postvaccination. In addition, we investigated by immunostaining whether the frequencies of CD4+ and CD8+ T cells and parameters of immune activation and proliferation on these cells were modulated by vaccination. At 1 month from the last vaccination dose, we found that 20 out of 22 children mounted a serological response to HBV; a majority of children had antibodies against HAV at baseline. The number of HIV-1 DNA copies in blood at 1 month postvaccination was reduced in comparison to baseline although this reduction was not statistically significant. A significant reduction of HIV-1 DNA copies in blood following vaccination was found in 12 children. The frequencies of CD4+ (naïve, effector memory) and CD8+ (central memory) T-cell subpopulations changed following vaccinations and a reduction in the activation and proliferation pattern of these cells was also noticed. Multivariate linear regression analysis revealed that the frequency of CD8+ effector memory T cells prior to vaccination was strongly predictive of the reduction of HIV-1 DNA copies in blood following vaccination of the 22 HIV-1-infected children. The results of this study suggest a beneficial effect of vaccination to reduce the size of virus reservoir in HIV-1-infected children receiving ART. A reduced frequency of

  3. Therapeutic strategies to fight HIV-1 latency: progress and challenges

    CSIR Research Space (South Africa)

    Manoto, Sello L

    2017-10-01

    Full Text Available —1112, 2017 Therapeutic strategies to fight HIV-1 latency: progress and challenges Sello Lebohang Manoto, Lebogang Thobakgale, Rudzani Malabi, Charles Maphanga, Saturnin Ombinda-Lemboumba, Patience Mthunzi-Kufa Abstract: The life...

  4. Distribution of HIV-1 resistance-conferring polymorphic alleles SDF ...

    Indian Academy of Sciences (India)

    Unknown

    involved with delay in disease progression. ... proteins in 525 healthy individuals without any history of HIV-1 infection from 11 diverse populations of ... in three populations (Yamani, Pathan and Kamma), all in low frequencies (i.e. 1% to 3%).

  5. Intragenic HIV-1 env sequences that enhance gag expression

    International Nuclear Information System (INIS)

    Suptawiwat, Ornpreya; Sutthent, Ruengpung; Lee, T.-H.; Auewarakul, Prasert

    2003-01-01

    Expression of HIV-1 genes is regulated at multiple levels including the complex RNA splicing and transport mechanisms. Multiple cis-acting elements involved in these regulations have been previously identified in various regions of HIV-1 genome. Here we show that another cis-acting element was present in HIV-1 env region. This element enhanced the expression of Gag when inserted together with Rev response element (RRE) into a truncated HIV-1 genome in the presence of Rev. The enhancing activity was mapped to a 263-bp fragment in the gp41 region downstream to RRE. RNA analysis showed that it might function by promoting RNA stability and Rev-dependent RNA export. The enhancement was specific to Rev-dependent expression, since it did not enhance Gag expression driven by Sam68, a cellular protein that has been shown to be able to substitute for Rev in RNA export function

  6. The latest evidence for possible HIV-1 curative strategies.

    Science.gov (United States)

    Pham, Hanh Thi; Mesplède, Thibault

    2018-01-01

    Human immunodeficiency virus type 1 (HIV-1) infection remains a major health issue worldwide. In developed countries, antiretroviral therapy has extended its reach from treatment of people living with HIV-1 to post-exposure prophylaxis, treatment as prevention, and, more recently, pre-exposure prophylaxis. These healthcare strategies offer the epidemiological tools to curve the epidemic in rich settings and will be concomitantly implemented in developing countries. One of the remaining challenges is to identify an efficacious curative strategy. This review manuscript will focus on some of the current curative strategies aiming at providing a sterilizing or functional cure to HIV-1-positive individuals. These include the following: early treatment initiation in post-treatment controllers as a long-term HIV-1 remission strategy, latency reversal, gene editing with or without stem cell transplantation, and antibodies against either the viral envelope protein or the host integrin α4β7.

  7. Towards HIV-1 remission: potential roles for broadly neutralizing antibodies.

    Science.gov (United States)

    Halper-Stromberg, Ariel; Nussenzweig, Michel C

    2016-02-01

    Current antiretroviral drug therapies do not cure HIV-1 because they do not eliminate a pool of long-lived cells harboring immunologically silent but replication-competent proviruses - termed the latent reservoir. Eliminating this reservoir and stimulating the immune response to control infection in the absence of therapy remain important but unsolved goals of HIV-1 cure research. Recently discovered broadly neutralizing antibodies (bNAbs) exhibit remarkable breadth and potency in their ability to neutralize HIV-1 in vitro, and recent studies have demonstrated new therapeutic applications for passively administered bNAbs in vivo. This Review discusses the roles bNAbs might play in HIV-1 treatment regimens, including prevention, therapy, and cure.

  8. Human immunodeficiency virus type-1 (HIV-1) genetic diversity and ...

    African Journals Online (AJOL)

    PROGMANAGER

    2013-04-24

    Apr 24, 2013 ... objective of this study was to determine the genetic diversity of HIV-1 and the prevalence of antiretroviral (ARV) ... individuals in resource limited settings. Key words: ... management of HIV infection even as antiretroviral (ARV).

  9. Purinergic Receptors: Key Mediators of HIV-1 infection and inflammation

    Directory of Open Access Journals (Sweden)

    Talia H Swartz

    2015-11-01

    Full Text Available Human immunodeficiency virus (HIV-1 causes a chronic infection that afflicts more than 38 million individuals worldwide. While the infection can be suppressed with potent anti-retroviral therapies, individuals infected with HIV have elevated levels of inflammation as indicated by increased T cell activation, soluble biomarkers, and associated morbidity and mortality. A single mechanism linking HIV pathogenesis to this inflammation has yet to be identified. Purinergic receptors are known to mediate inflammation and have been shown to be required for HIV-1 infection at the level of HIV-1 membrane fusion. Here we review the literature on the role of purinergic receptors in HIV-1 infection and associated inflammation and describe a role for these receptors as potential therapeutic targets.

  10. [Molecular epidemiology and transmission of HIV-1 infection in Zhejiang province, 2015].

    Science.gov (United States)

    Yang, J Z; Chen, W J; Zhang, W J; He, L; Zhang, J F; Pan, X H

    2017-11-10

    Objective: To understand the distribution of HIV-1 subtype diversity and its transmission characteristics in Zhejiang province. Methods: A total of 302 newly diagnosed HIV-1 positive patients were selected through stratified random sampling in Zhejiang in 2015. HIV-1 pol genes were sequenced successfully with reverse transcription PCR/nested PCR and phylogenetic analysis was conducted for 276 patients. Then a molecular epidemiologic study was performed combined with field epidemiological investigation. Results: Of 276 sequence samples analyzed, 122 CRF07_BC strains (44.2%), 103 CRF01_AE strains (37.3%), 17 CRF08_BC strains (6.1%), 9 B strains (3.2%), 6 CRF55_01B strains (2.2%), 5 C strains (1.8%), 1 CRF59_01B strain (0.4%), 1 CRF67_01B strain (0.4%), 1 A1 strain (0.4%), and 11 URFs strains (4.0%) were identified. Phylogenetic analysis revealed 16 clusters with only 15.1% (34/225) sequences involved among CRF07_BC and CRF01_AE strains. The clustered cases in MSM were higher than that in populations with other transmission routes. And clusters existed between the populations with different transmission routes. Conclusion: The major strains of HIV-1 in Zhejiang are CRF07_BC and CRF01_AE. The HIV subtypes showed more complexity in Zhejiang. It is necessary to strengthen the surveillance for HIV subtypes, carry out classified management and conduct effective prevention and control in the population at high risk.

  11. Ex vivo response to histone deacetylase (HDAC inhibitors of the HIV long terminal repeat (LTR derived from HIV-infected patients on antiretroviral therapy.

    Directory of Open Access Journals (Sweden)

    Hao K Lu

    Full Text Available Histone deacetylase inhibitors (HDACi can induce human immunodeficiency virus (HIV transcription from the HIV long terminal repeat (LTR. However, ex vivo and in vivo responses to HDACi are variable and the activity of HDACi in cells other than T-cells have not been well characterised. Here, we developed a novel assay to determine the activity of HDACi on patient-derived HIV LTRs in different cell types. HIV LTRs from integrated virus were amplified using triple-nested Alu-PCR from total memory CD4+ T-cells (CD45RO+ isolated from HIV-infected patients prior to and following suppressive antiretroviral therapy. NL4-3 or patient-derived HIV LTRs were cloned into the chromatin forming episomal vector pCEP4, and the effect of HDACi investigated in the astrocyte and epithelial cell lines SVG and HeLa, respectively. There were no significant differences in the sequence of the HIV LTRs isolated from CD4+ T-cells prior to and after 18 months of combination antiretroviral therapy (cART. We found that in both cell lines, the HDACi panobinostat, trichostatin A, vorinostat and entinostat activated patient-derived HIV LTRs to similar levels seen with NL4-3 and all patient derived isolates had similar sensitivity to maximum HDACi stimulation. We observed a marked difference in the maximum fold induction of luciferase by HDACi in HeLa and SVG, suggesting that the effect of HDACi may be influenced by the cellular environment. Finally, we observed significant synergy in activation of the LTR with vorinostat and the viral protein Tat. Together, our results suggest that the LTR sequence of integrated virus is not a major determinant of a functional response to an HDACi.

  12. Determination of HIV-1 co-receptor usage.

    Science.gov (United States)

    Cavarelli, Mariangela; Scarlatti, Gabriella

    2014-01-01

    Human immunodeficiency virus type I (HIV-1) infects target cells through interaction with the CD4 molecule and chemokine receptors, mainly the β-chemokine receptor 5 (CCR5) and the α-chemokine receptor 4 (CXCR4). Viral isolates can be phenotypically classified based on the co-receptor they utilize to infect target cells. In this chapter, methods to determine the co-receptor usage of HIV-1 variants are described.

  13. Increased T cell trafficking as adjunct therapy for HIV-1

    OpenAIRE

    Fryer, HR; Wolinsky, SM; McLean, AR

    2018-01-01

    Although antiretroviral drug therapy suppresses human immunodeficiency virus-type 1 (HIV-1) to undetectable levels in the blood of treated individuals, reservoirs of replication competent HIV-1 endure. Upon cessation of antiretroviral therapy, the reservoir usually allows outgrowth of virus and approaches to targeting the reservoir have had limited success. Ongoing cycles of viral replication in regions with low drug penetration contribute to this persistence. Here, we use a mathematical mode...

  14. Low levels of LTR retrotransposon deletion by ectopic recombination in the gigantic genomes of salamanders.

    Science.gov (United States)

    Frahry, Matthew Blake; Sun, Cheng; Chong, Rebecca A; Mueller, Rachel Lockridge

    2015-02-01

    Across the tree of life, species vary dramatically in nuclear genome size. Mutations that add or remove sequences from genomes-insertions or deletions, or indels-are the ultimate source of this variation. Differences in the tempo and mode of insertion and deletion across taxa have been proposed to contribute to evolutionary diversity in genome size. Among vertebrates, most of the largest genomes are found within the salamanders, an amphibian clade with genome sizes ranging from ~14 to ~120 Gb. Salamander genomes have been shown to experience slower rates of DNA loss through small (i.e., genomes. However, no studies have addressed DNA loss from salamander genomes resulting from larger deletions. Here, we focus on one type of large deletion-ectopic-recombination-mediated removal of LTR retrotransposon sequences. In ectopic recombination, double-strand breaks are repaired using a "wrong" (i.e., ectopic, or non-allelic) template sequence-typically another locus of similar sequence. When breaks occur within the LTR portions of LTR retrotransposons, ectopic-recombination-mediated repair can produce deletions that remove the internal transposon sequence and the equivalent of one of the two LTR sequences. These deletions leave a signature in the genome-a solo LTR sequence. We compared levels of solo LTRs in the genomes of four salamander species with levels present in five vertebrates with smaller genomes. Our results demonstrate that salamanders have low levels of solo LTRs, suggesting that ectopic-recombination-mediated deletion of LTR retrotransposons occurs more slowly than in other vertebrates with smaller genomes.

  15. Dendritic cells exposed to MVA-based HIV-1 vaccine induce highly functional HIV-1-specific CD8(+ T cell responses in HIV-1-infected individuals.

    Directory of Open Access Journals (Sweden)

    Núria Climent

    Full Text Available Currently, MVA virus vectors carrying HIV-1 genes are being developed as HIV-1/AIDS prophylactic/therapeutic vaccines. Nevertheless, little is known about the impact of these vectors on human dendritic cells (DC and their capacity to present HIV-1 antigens to human HIV-specific T cells. This study aimed to characterize the interaction of MVA and MVA expressing the HIV-1 genes Env-Gag-Pol-Nef of clade B (referred to as MVA-B in human monocyte-derived dendritic cells (MDDC and the subsequent processes of HIV-1 antigen presentation and activation of memory HIV-1-specific T lymphocytes. For these purposes, we performed ex vivo assays with MDDC and autologous lymphocytes from asymptomatic HIV-infected patients. Infection of MDDC with MVA-B or MVA, at the optimal dose of 0.3 PFU/MDDC, induced by itself a moderate degree of maturation of MDDC, involving secretion of cytokines and chemokines (IL1-ra, IL-7, TNF-α, IL-6, IL-12, IL-15, IL-8, MCP-1, MIP-1α, MIP-1β, RANTES, IP-10, MIG, and IFN-α. MDDC infected with MVA or MVA-B and following a period of 48 h or 72 h of maturation were able to migrate toward CCL19 or CCL21 chemokine gradients. MVA-B infection induced apoptosis of the infected cells and the resulting apoptotic bodies were engulfed by the uninfected MDDC, which cross-presented HIV-1 antigens to autologous CD8(+ T lymphocytes. MVA-B-infected MDDC co-cultured with autologous T lymphocytes induced a highly functional HIV-specific CD8(+ T cell response including proliferation, secretion of IFN-γ, IL-2, TNF-α, MIP-1β, MIP-1α, RANTES and IL-6, and strong cytotoxic activity against autologous HIV-1-infected CD4(+ T lymphocytes. These results evidence the adjuvant role of the vector itself (MVA and support the clinical development of prophylactic and therapeutic anti-HIV vaccines based on MVA-B.

  16. Structure of HIV-1 protease determined by neutron crystallography

    International Nuclear Information System (INIS)

    Adachi, Motoyasu; Kuroki, Ryota

    2009-01-01

    HIV-1 protease is an aspartic protease, and plays an essential role in replication of HIV. To develop HIV-1 protease inhibitors through structure-based drug design, it is necessary to understand the catalytic mechanism and inhibitor recognition of HIV-1 protease. We have determined the crystal structure of HIV-1 protease in complex with KNI-272 to 1.9 A resolution by neutron crystallography in combination with 1.4 A resolution X-ray diffraction data. The results show that the carbonyl group of hydroxymethylcarbonyl (HMC) in KNI-272 forms a hydrogen bonding interaction with protonated Asp 25 and the hydrogen atom from the hydroxyl group of HMC forms a hydrogen bonding interaction with the deprotonated Asp125. This is the first neutron report for HIV-1/inhibitor complex and shows directly the locations of key hydrogen atoms in catalysis and in the binding of a transition-state analog. The results confirm key aspect of the presumed catalytic mechanism of HIV-1 protease and will aid in the further development of protease inhibitors. (author)

  17. HIV-1, Methamphetamine and Astrocytes at Neuroinflammatory crossroads

    Directory of Open Access Journals (Sweden)

    Kathleen eBorgmann

    2015-10-01

    Full Text Available As a popular psychostimulant, methamphetamine (METH use leads to long-lasting, strong euphoric effects. While METH abuse is common in the general population, between 10-15% of human immunodeficiency virus-1 (HIV-1 patients report having abused METH. METH exacerbates the severity and onset of HIV-1-associated neurocognitive disorders (HAND through direct and indirect mechanisms. Repetitive METH use decreases adherence to antiretroviral drug regimens, increasing the likelihood of HIV-1 disease progression towards AIDS. METH exposure also directly affects both innate and adaptive immunity, altering lymphocyte number and activity, cytokine signaling, phagocytic function, and CNS infiltration through the blood brain barrier. Further, METH triggers the neuronal dopamine reward pathway and leads to altered neuronal activity and direct toxicity. Concurrently, METH and HIV-1 alter the neuroimmune balance and induce neuroinflammation. Neuroinflammation modulates a wide range of brain functions including neuronal signaling and activity, glial activation, viral infection, oxidative stress and excitotoxicity. Pathologically, glial activation is a hallmark of both HIV-1 and METH-associated neuroinflammation. Significant commonality exists in the neurotoxic mechanisms for both METH and HAND; however, the pathways dysregulated in astroglia during METH exposure are less clear. Thus alterations in astrocyte intracellular signaling pathways, gene expression and function during METH and HIV-1 comorbidity, neuroinflammation and HAND are carefully reviewed. Interventions targeting astrocytes in HAND and METH are presented as potential novel therapeutic approaches.

  18. Clinical presentation and opportunistic infections in HIV-1, HIV-2 and HIV-1/2 dual seropositive patients in Guinea-Bissau

    DEFF Research Database (Denmark)

    Sørensen, Allan; Jespersen, Sanne; Katzenstein, Terese L

    2016-01-01

    HIV-2 is prevalent. In this study, we aimed to characterize the clinical presentations among HIV-1, HIV-2 and HIV-1/2 dual seropositive patients. Methods: In a cross-sectional study, newly diagnosed HIV patients attending the HIV outpatient clinic at Hospital Nacional Sim~ao Mendes in Guinea......-Bissau were enrolled. Demographical and clinical data were collected and compared between HIV-1, HIV-2 and HIV-1/2 dual seropositive patients. Results: A total of 169 patients (76% HIV-1, 17% HIV-2 and 6% HIV 1/2) were included in the study between 21 March 2012 and 14 December 2012. HIV-1 seropositive...... antigen. Conclusion: HIV-1 and HIV-1/2 seropositive patients have lower CD4 cell counts than HIV-2 seropositive patients when diagnosed with HIV with only minor clinical and demographic differences among groups. Few patients were diagnosed with TB and cryptococcal disease was not found to be a major...

  19. Phylogenetic analysis of HIV-1 pol gene: first subgenomic evidence of CRF29-BF among Iranian HIV-1 patients

    Directory of Open Access Journals (Sweden)

    Kazem Baesi

    2014-09-01

    Full Text Available Objective: To identify the dominant subtype among the HIV-1 strains circulation in Iran. Methods: In this cross sectional study 100 HIV positive patients participated. HIV-1 RNA was extracted from plasma. RT nested-PCR was performed and the final products were sequenced and phylogenetically analyzed; reference sequences were downloaded from Los Alamos, aligned with Iranian pol sequences in the study and analyzed by neighbor-joining method. Results: The results of the phylogenetic analysis showed that HIV-1 subtype CRF-35AD was the dominant subtype among HIV-1 infected patients in Iran; this analysis also suggested a new circulating recombinant form that had not previously been identified in Iran: CRF-29BF. Conclusions: The impact of HIV diversity on pathogenesis, transmission and clinical management have been discussed in different studies; therefore, analyses of HIV genetic diversity is required to design effective antiretroviral strategies for different HIV subtypes.

  20. Silencing of human T-cell leukemia virus type I gene transcription by epigenetic mechanisms

    Directory of Open Access Journals (Sweden)

    Mueller Nancy

    2005-10-01

    Full Text Available Abstract Background Human T-cell leukemia virus type I (HTLV-I causes adult T-cell leukemia (ATL after a long latent period. Among accessory genes encoded by HTLV-I, the tax gene is thought to play a central role in oncogenesis. However, Tax expression is disrupted by several mechanims including genetic changes of the tax gene, deletion/hypermethylation of 5'-LTR. To clarify the role of epigenetic changes, we analyzed DNA methylation and histone modification in the whole HTLV-I provirus genome. Results The gag, pol and env genes of HTLV-I provirus were more methylated than pX region, whereas methylation of 5'-LTR was variable and 3'-LTR was not methylated at all. In ATL cell lines, complete DNA methylation of 5'-LTR was associated with transcriptional silencing of viral genes. HTLV-I provirus was more methylated in primary ATL cells than in carrier state, indicating the association with disease progression. In seroconvertors, DNA methylation was already observed in internal sequences of provirus just after seroconversion. Taken together, it is speculated that DNA methylation first occurs in the gag, pol and env regions and then extends in the 5' and 3' directions in vivo, and when 5'-LTR becomes methylated, viral transcription is silenced. Analysis of histone modification in the HTLV-I provirus showed that the methylated provirus was associated with hypoacetylation. However, the tax gene transcript could not be detected in fresh ATL cells regardless of hyperacetylated histone H3 in 5'-LTR. The transcription rapidly recovered after in vitro culture in such ATL cells. Conclusion These results showed that epigenetic changes of provirus facilitated ATL cells to evade host immune system by suppressing viral gene transcription. In addition, this study shows the presence of another reversible mechanism that suppresses the tax gene transcription without DNA methylation and hypoacetylated histone.

  1. HIV-1 infection during pregnancy and in children : significance of HIV-1 variability and the placental barrier

    OpenAIRE

    Casper, Charlotte

    2001-01-01

    With the global increase in human immunodeficiency virus 1 (HIV-1) infection in women of childbearing age, there has also been an alarming increase in the number of mother-to-child transmissions of HIV-1. Although antiretroviral therapy and Cesarian section have been demonstrated to significantly decrease the vertical transmission rate of , these interventions are not widely available in the developing world. Therefore, studies of the mechanisms of vertical transmission are ...

  2. Characterization and frequency of a newly identified HIV-1 BF1 intersubtype circulating recombinant form in São Paulo, Brazil

    Directory of Open Access Journals (Sweden)

    Neto Walter

    2010-04-01

    Full Text Available Abstract Background HIV circulating recombinant forms (CRFs play an important role in the global and regional HIV epidemics, particularly in regions where multiple subtypes are circulating. To date, several (>40 CRFs are recognized worldwide with five currently circulating in Brazil. Here, we report the characterization of near full-length genome sequences (NFLG of six phylogenetically related HIV-1 BF1 intersubtype recombinants (five from this study and one from other published sequences representing CRF46_BF1. Methods Initially, we selected 36 samples from 888 adult patients residing in São Paulo who had previously been diagnosed as being infected with subclade F1 based on pol subgenomic fragment sequencing. Proviral DNA integrated in peripheral blood mononuclear cells (PBMC was amplified from the purified genomic DNA of all 36-blood samples by five overlapping PCR fragments followed by direct sequencing. Sequence data were obtained from the five fragments that showed identical genomic structure and phylogenetic trees were constructed and compared with previously published sequences. Genuine subclade F1 sequences and any other sequences that exhibited unique mosaic structures were omitted from further analysis Results Of the 36 samples analyzed, only six sequences, inferred from the pol region as subclade F1, displayed BF1 identical mosaic genomes with a single intersubtype breakpoint identified at the nef-U3 overlap (HXB2 position 9347-9365; LTR region. Five of these isolates formed a rigid cluster in phylogentic trees from different subclade F1 fragment regions, which we can now designate as CRF46_BF1. According to our estimate, the new CRF accounts for 0.56% of the HIV-1 circulating strains in São Paulo. Comparison with previously published sequences revealed an additional five isolates that share an identical mosaic structure with those reported in our study. Despite sharing a similar recombinant structure, only one sequence appeared to

  3. Identifying potential survival strategies of HIV-1 through virus-host protein interaction networks

    Directory of Open Access Journals (Sweden)

    Boucher Charles AB

    2010-07-01

    Full Text Available Abstract Background The National Institute of Allergy and Infectious Diseases has launched the HIV-1 Human Protein Interaction Database in an effort to catalogue all published interactions between HIV-1 and human proteins. In order to systematically investigate these interactions functionally and dynamically, we have constructed an HIV-1 human protein interaction network. This network was analyzed for important proteins and processes that are specific for the HIV life-cycle. In order to expose viral strategies, network motif analysis was carried out showing reoccurring patterns in virus-host dynamics. Results Our analyses show that human proteins interacting with HIV form a densely connected and central sub-network within the total human protein interaction network. The evaluation of this sub-network for connectivity and centrality resulted in a set of proteins essential for the HIV life-cycle. Remarkably, we were able to associate proteins involved in RNA polymerase II transcription with hubs and proteasome formation with bottlenecks. Inferred network motifs show significant over-representation of positive and negative feedback patterns between virus and host. Strikingly, such patterns have never been reported in combined virus-host systems. Conclusions HIV infection results in a reprioritization of cellular processes reflected by an increase in the relative importance of transcriptional machinery and proteasome formation. We conclude that during the evolution of HIV, some patterns of interaction have been selected for resulting in a system where virus proteins preferably interact with central human proteins for direct control and with proteasomal proteins for indirect control over the cellular processes. Finally, the patterns described by network motifs illustrate how virus and host interact with one another.

  4. Phylodynamics of the HIV-1 epidemic in Cuba.

    Science.gov (United States)

    Delatorre, Edson; Bello, Gonzalo

    2013-01-01

    Previous studies have shown that the HIV-1 epidemic in Cuba displayed a complex molecular epidemiologic profile with circulation of several subtypes and circulating recombinant forms (CRF); but the evolutionary and population history of those viral variants remains unknown. HIV-1 pol sequences of the most prevalent Cuban lineages (subtypes B, C and G, CRF18_cpx, CRF19_cpx, and CRFs20/23/24_BG) isolated between 1999 and 2011 were analyzed. Maximum-likelihood analyses revealed multiple introductions of subtype B (n≥66), subtype C (n≥10), subtype G (n≥8) and CRF18_cpx (n≥2) viruses in Cuba. The bulk of HIV-1 infections in this country, however, was caused by dissemination of a few founder strains probably introduced from North America/Europe (clades B(CU-I) and B(CU-II)), east Africa (clade C(CU-I)) and central Africa (clades G(CU), CRF18(CU) and CRF19(CU)), or locally generated (clades CRFs20/23/24_BG). Bayesian-coalescent analyses show that the major HIV-1 founder strains were introduced into Cuba during 1985-1995; whereas the CRFs_BG strains emerged in the second half of the 1990s. Most HIV-1 Cuban clades appear to have experienced an initial period of fast exponential spread during the 1990s and early 2000s, followed by a more recent decline in growth rate. The median initial growth rate of HIV-1 Cuban clades ranged from 0.4 year⁻¹ to 1.6 year⁻¹. Thus, the HIV-1 epidemic in Cuba has been a result of the successful introduction of a few viral strains that began to circulate at a rather late time of the AIDS pandemic, but then were rapidly disseminated through local transmission networks.

  5. Diverse fates of uracilated HIV-1 DNA during infection of myeloid lineage cells.

    Science.gov (United States)

    Hansen, Erik C; Ransom, Monica; Hesselberth, Jay R; Hosmane, Nina N; Capoferri, Adam A; Bruner, Katherine M; Pollack, Ross A; Zhang, Hao; Drummond, Michael Bradley; Siliciano, Janet M; Siliciano, Robert; Stivers, James T

    2016-09-20

    We report that a major subpopulation of monocyte-derived macrophages (MDMs) contains high levels of dUTP, which is incorporated into HIV-1 DNA during reverse transcription (U/A pairs), resulting in pre-integration restriction and post-integration mutagenesis. After entering the nucleus, uracilated viral DNA products are degraded by the uracil base excision repair (UBER) machinery with less than 1% of the uracilated DNA successfully integrating. Although uracilated proviral DNA showed few mutations, the viral genomic RNA was highly mutated, suggesting that errors occur during transcription. Viral DNA isolated from blood monocytes and alveolar macrophages (but not T cells) of drug-suppressed HIV-infected individuals also contained abundant uracils. The presence of viral uracils in short-lived monocytes suggests their recent infection through contact with virus producing cells in a tissue reservoir. These findings reveal new elements of a viral defense mechanism involving host UBER that may be relevant to the establishment and persistence of HIV-1 infection.

  6. Diverse fates of uracilated HIV-1 DNA during infection of myeloid lineage cells

    Science.gov (United States)

    Hansen, Erik C; Ransom, Monica; Hesselberth, Jay R; Hosmane, Nina N; Capoferri, Adam A; Bruner, Katherine M; Pollack, Ross A; Zhang, Hao; Drummond, Michael Bradley; Siliciano, Janet M; Siliciano, Robert; Stivers, James T

    2016-01-01

    We report that a major subpopulation of monocyte-derived macrophages (MDMs) contains high levels of dUTP, which is incorporated into HIV-1 DNA during reverse transcription (U/A pairs), resulting in pre-integration restriction and post-integration mutagenesis. After entering the nucleus, uracilated viral DNA products are degraded by the uracil base excision repair (UBER) machinery with less than 1% of the uracilated DNA successfully integrating. Although uracilated proviral DNA showed few mutations, the viral genomic RNA was highly mutated, suggesting that errors occur during transcription. Viral DNA isolated from blood monocytes and alveolar macrophages (but not T cells) of drug-suppressed HIV-infected individuals also contained abundant uracils. The presence of viral uracils in short-lived monocytes suggests their recent infection through contact with virus producing cells in a tissue reservoir. These findings reveal new elements of a viral defense mechanism involving host UBER that may be relevant to the establishment and persistence of HIV-1 infection. DOI: http://dx.doi.org/10.7554/eLife.18447.001 PMID:27644592

  7. Deletion of the LTR enhancer/promoter has no impact on the integration profile of MLV vectors in human hematopoietic progenitors.

    Directory of Open Access Journals (Sweden)

    Arianna Moiani

    Full Text Available Moloney murine leukemia virus (MLV-derived gamma-retroviral vectors integrate preferentially near transcriptional regulatory regions in the human genome, and are associated with a significant risk of insertional gene deregulation. Self-inactivating (SIN vectors carry a deletion of the U3 enhancer and promoter in the long terminal repeat (LTR, and show reduced genotoxicity in pre-clinical assays. We report a high-definition analysis of the integration preferences of a SIN MLV vector compared to a wild-type-LTR MLV vector in the genome of CD34(+ human hematopoietic stem/progenitor cells (HSPCs. We sequenced 13,011 unique SIN-MLV integration sites and compared them to 32,574 previously generated MLV sites in human HSPCs. The SIN-MLV vector recapitulates the integration pattern observed for MLV, with the characteristic clustering of integrations around enhancer and promoter regions associated to H3K4me3 and H3K4me1 histone modifications, specialized chromatin configurations (presence of the H2A.Z histone variant and binding of RNA Pol II. SIN-MLV and MLV integration clusters and hot spots overlap in most cases and are generated at a comparable frequency, indicating that the reduced genotoxicity of SIN-MLV vectors in hematopoietic cells is not due to a modified integration profile.

  8. HIV-1 viral load measurement in venous blood and fingerprick blood using Abbott RealTime HIV-1 DBS assay.

    Science.gov (United States)

    Tang, Ning; Pahalawatta, Vihanga; Frank, Andrea; Bagley, Zowie; Viana, Raquel; Lampinen, John; Leckie, Gregor; Huang, Shihai; Abravaya, Klara; Wallis, Carole L

    2017-07-01

    HIV RNA suppression is a key indicator for monitoring success of antiretroviral therapy. From a logistical perspective, viral load (VL) testing using Dried Blood Spots (DBS) is a promising alternative to plasma based VL testing in resource-limited settings. To evaluate the analytical and clinical performance of the Abbott RealTime HIV-1 assay using a fully automated one-spot DBS sample protocol. Limit of detection (LOD), linearity, lower limit of quantitation (LLQ), upper limit of quantitation (ULQ), and precision were determined using serial dilutions of HIV-1 Virology Quality Assurance stock (VQA Rush University), or HIV-1-containing armored RNA, made in venous blood. To evaluate correlation, bias, and agreement, 497 HIV-1 positive adult clinical samples were collected from Ivory Coast, Uganda and South Africa. For each HIV-1 participant, DBS-fingerprick, DBS-venous and plasma sample results were compared. Correlation and bias values were obtained. The sensitivity and specificity were analyzed at a threshold of 1000 HIV-1 copies/mL generated using the standard plasma protocol. The Abbott HIV-1 DBS protocol had an LOD of 839 copies/mL, a linear range from 500 to 1×10 7 copies/mL, an LLQ of 839 copies/mL, a ULQ of 1×10 7 copies/mL, and an inter-assay SD of ≤0.30 log copies/mL for all tested levels within this range. With clinical samples, the correlation coefficient (r value) was 0.896 between DBS-fingerprick and plasma and 0.901 between DBS-venous and plasma, and the bias was -0.07 log copies/mL between DBS-fingerprick and plasma and -0.02 log copies/mL between DBS-venous and plasma. The sensitivity of DBS-fingerprick and DBS-venous was 93%, while the specificity of both DBS methods was 95%. The results demonstrated that the Abbott RealTime HIV-1 assay with DBS sample protocol is highly sensitive, specific and precise across a wide dynamic range and correlates well with plasma values. The Abbott RealTime HIV-1 assay with DBS sample protocol provides an

  9. Characterization of natural polymorphic sites of the HIV-1 integrase before the introduction of HIV-1 integrase inhibitors in Germany

    Science.gov (United States)

    Meixenberger, Karolin; Pouran Yousef, Kaveh; Somogyi, Sybille; Fiedler, Stefan; Bartmeyer, Barbara; von Kleist, Max; Kücherer, Claudia

    2014-01-01

    Introduction The aim of our study was to analyze the occurrence and evolution of HIV-1 integrase polymorphisms during the HIV-1 epidemic in Germany prior to the introduction of the first integrase inhibitor raltegravir in 2007. Materials and Methods Plasma samples from drug-naïve HIV-1 infected individuals newly diagnosed between 1986 and 2006 were used to determine PCR-based population sequences of the HIV-1 integrase (amino acids 1–278). The HIV-1 subtype was determined using the REGA HIV-1 subtyping tool. We calculated the frequency of amino acids at each position of the HIV-1 integrase in 337 subtype B strains for the time periods 1986–1989, 1991–1994, 1995–1998, 1999–2002, and 2003–2006. Positions were defined as polymorphic if amino acid variation was >1% in any period. Logistic regression was used to identify trends in amino acid variation over time. Resistance-associated mutations were identified according to the IAS 2013 list and the HIVdb, ANRS and GRADE algorithms. Results Overall, 56.8% (158/278) amino acid positions were polymorphic and 15.8% (25/158) of these positions exhibited a significant trend in amino acid variation over time. Proportionately, most polymorphic positions (63.3%, 31/49) were detected in the N-terminal zinc finger domain of the HIV-1 integrase. Motifs and residues essential for HIV-1 integrase activity were little polymorphic, but within the minimal non-specific DNA binding region I220-D270 up to 18.1% amino acid variation was noticed, including four positions with significant amino acid variation over time (S230, D232, D256, A265). No major resistance mutations were identified, and minor resistance mutations were rarely observed without trend over time. E157Q considered by HIVdb, ANRS, and GRADE algorithms was the most frequent resistance-associated polymorphism with an overall prevalence of 2.4%. Conclusions Detailed knowledge of the evolutionary variation of HIV-1 integrase polymorphisms is important to understand

  10. HIV-1 uncoating: connection to nuclear entry and regulation by host proteins

    Energy Technology Data Exchange (ETDEWEB)

    Ambrose, Zandrea, E-mail: zaa4@pitt.edu [Division of Infectious Diseases, Department of Medicine, University of Pittsburgh, School of Medicine, Pittsburgh, PA 15261 (United States); Aiken, Christopher [Department of Pathology, Microbiology and Immunology, Vanderbilt University, School of Medicine, Nashville, TN 37232 (United States)

    2014-04-15

    The RNA genome of human immunodeficiency virus type 1 (HIV-1) is enclosed by a capsid shell that dissociates within the cell in a multistep process known as uncoating, which influences completion of reverse transcription of the viral genome. Double-stranded viral DNA is imported into the nucleus for integration into the host genome, a hallmark of retroviral infection. Reverse transcription, nuclear entry, and integration are coordinated by a capsid uncoating process that is regulated by cellular proteins. Although uncoating is not well understood, recent studies have revealed insights into the process, particularly with respect to nuclear import pathways and protection of the viral genome from DNA sensors. Understanding uncoating will be valuable toward developing novel antiretroviral therapies for HIV-infected individuals.

  11. Identification of a non-LTR retrotransposon from the gypsy moth

    Science.gov (United States)

    K.J. Garner; J.M. Slavicek

    1999-01-01

    A family of highly repetitive elements, named LDT1, has been identified in the gypsy moth, Lymantria dispar. The complete element is 5.4 kb in length and lacks long-terminal repeats, The element contains two open reading frames with a significant amino acid sequence similarity to several non-LTR retrotransposons. The first open reading frame contains...

  12. Akv murine leukemia virus enhances bone tumorigenesis in hMT-c-fos-LTR transgenic mice

    DEFF Research Database (Denmark)

    Schmidt, Jörg; Krump-Konvalinkova, Vera; Luz, Arne

    1995-01-01

    hMt-c-fos-LTR transgenic mice (U. Rüther, D. Komitowski, F. R. Schubert, and E. F. Wagner. Oncogene 4, 861–865, 1989) developed bone sarcomas in 20% (3/15) of females at 448 ± 25 days and in 8% (1/12) of males at 523 days. After infection of newborns with Akv, an infectious retrovirus derived from...

  13. LTR retrotransposon dynamics in the evolution of the olive (Olea europaea) genome

    Czech Academy of Sciences Publication Activity Database

    Barghini, E.; Natali, L.; Giordani, T.; Cossu, R.M.; Scalabrin, S.; Cattonaro, F.; Šimková, Hana; Vrána, Jan; Doležel, Jaroslav; Morgante, M.; Cavallini, A.

    2015-01-01

    Roč. 22, č. 1 (2015), s. 91-100 ISSN 1340-2838 R&D Projects: GA ČR GBP501/12/G090; GA MŠk(CZ) LO1204 Institutional support: RVO:61389030 Keywords : LTR retrotransposons * next-generation sequencing * olive Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 5.267, year: 2015

  14. RNA helicase MOV10 functions as a co-factor of HIV-1 Rev to facilitate Rev/RRE-dependent nuclear export of viral mRNAs

    International Nuclear Information System (INIS)

    Huang, Feng; Zhang, Junsong; Zhang, Yijun; Geng, Guannan; Liang, Juanran; Li, Yingniang; Chen, Jingliang; Liu, Chao; Zhang, Hui

    2015-01-01

    Human immunodeficiency virus type 1 (HIV-1) exploits multiple host factors during its replication. The REV/RRE-dependent nuclear export of unspliced/partially spliced viral transcripts needs the assistance of host proteins. Recent studies have shown that MOV10 overexpression inhibited HIV-1 replication at various steps. However, the endogenous MOV10 was required in certain step(s) of HIV-1 replication. In this report, we found that MOV10 potently enhances the nuclear export of viral mRNAs and subsequently increases the expression of Gag protein and other late products through affecting the Rev/RRE axis. The co-immunoprecipitation analysis indicated that MOV10 interacts with Rev in an RNA-independent manner. The DEAG-box of MOV10 was required for the enhancement of Rev/RRE-dependent nuclear export and the DEAG-box mutant showed a dominant-negative activity. Our data propose that HIV-1 utilizes the anti-viral factor MOV10 to function as a co-factor of Rev and demonstrate the complicated effects of MOV10 on HIV-1 life cycle. - Highlights: • MOV10 can function as a co-factor of HIV-1 Rev. • MOV10 facilitates Rev/RRE-dependent transport of viral mRNAs. • MOV10 interacts with Rev in an RNA-independent manner. • The DEAG-box of MOV10 is required for the enhancement of Rev/RRE-dependent export.

  15. Novel HIV-1 knockdown targets identified by an enriched kinases/phosphatases shRNA library using a long-term iterative screen in Jurkat T-cells.

    Directory of Open Access Journals (Sweden)

    Sylvie Rato

    2010-02-01

    Full Text Available HIV-1 is a complex retrovirus that uses host machinery to promote its replication. Understanding cellular proteins involved in the multistep process of HIV-1 infection may result in the discovery of more adapted and effective therapeutic targets. Kinases and phosphatases are a druggable class of proteins critically involved in regulation of signal pathways of eukaryotic cells. Here, we focused on the discovery of kinases and phosphatases that are essential for HIV-1 replication but dispensable for cell viability. We performed an iterative screen in Jurkat T-cells with a short-hairpin-RNA (shRNA library highly enriched for human kinases and phosphatases. We identified 14 new proteins essential for HIV-1 replication that do not affect cell viability. These proteins are described to be involved in MAPK, JNK and ERK pathways, vesicular traffic and DNA repair. Moreover, we show that the proteins under study are important in an early step of HIV-1 infection before viral integration, whereas some of them affect viral transcription/translation. This study brings new insights for the complex interplay of HIV-1/host cell and opens new possibilities for antiviral strategies.

  16. RNA helicase MOV10 functions as a co-factor of HIV-1 Rev to facilitate Rev/RRE-dependent nuclear export of viral mRNAs

    Energy Technology Data Exchange (ETDEWEB)

    Huang, Feng; Zhang, Junsong; Zhang, Yijun; Geng, Guannan; Liang, Juanran; Li, Yingniang; Chen, Jingliang [Institute of Human Virology, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou 510080 (China); Key Laboratory of Tropical Disease Control of Ministry of Education, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou 510080 (China); Liu, Chao, E-mail: liuchao9@mail.sysu.edu.cn [Institute of Human Virology, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou 510080 (China); Key Laboratory of Tropical Disease Control of Ministry of Education, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou 510080 (China); Zhang, Hui [Institute of Human Virology, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou 510080 (China); Key Laboratory of Tropical Disease Control of Ministry of Education, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou 510080 (China)

    2015-12-15

    Human immunodeficiency virus type 1 (HIV-1) exploits multiple host factors during its replication. The REV/RRE-dependent nuclear export of unspliced/partially spliced viral transcripts needs the assistance of host proteins. Recent studies have shown that MOV10 overexpression inhibited HIV-1 replication at various steps. However, the endogenous MOV10 was required in certain step(s) of HIV-1 replication. In this report, we found that MOV10 potently enhances the nuclear export of viral mRNAs and subsequently increases the expression of Gag protein and other late products through affecting the Rev/RRE axis. The co-immunoprecipitation analysis indicated that MOV10 interacts with Rev in an RNA-independent manner. The DEAG-box of MOV10 was required for the enhancement of Rev/RRE-dependent nuclear export and the DEAG-box mutant showed a dominant-negative activity. Our data propose that HIV-1 utilizes the anti-viral factor MOV10 to function as a co-factor of Rev and demonstrate the complicated effects of MOV10 on HIV-1 life cycle. - Highlights: • MOV10 can function as a co-factor of HIV-1 Rev. • MOV10 facilitates Rev/RRE-dependent transport of viral mRNAs. • MOV10 interacts with Rev in an RNA-independent manner. • The DEAG-box of MOV10 is required for the enhancement of Rev/RRE-dependent export.

  17. Impaired production of cytokines is an independent predictor of mortality in HIV-1-infected patients

    DEFF Research Database (Denmark)

    Ostrowski, Sisse R; Gerstoft, Jan; Pedersen, Bente K

    2003-01-01

    With regard to the natural history of HIV-1 infection this study investigated whether whole-blood culture cytokine production was associated with mortality in HIV-1-infected patients.......With regard to the natural history of HIV-1 infection this study investigated whether whole-blood culture cytokine production was associated with mortality in HIV-1-infected patients....

  18. Raltegravir cerebrospinal fluid concentrations in HIV-1 infection.

    Directory of Open Access Journals (Sweden)

    Aylin Yilmaz

    2009-09-01

    Full Text Available Raltegravir is an HIV-1 integrase inhibitor currently used in treatment-experienced HIV-1-infected patients resistant to other drug classes. In order to assess its central nervous system penetration, we measured raltegravir concentrations in cerebrospinal fluid (CSF and plasma in subjects receiving antiretroviral treatment regimens containing this drug.Raltegravir concentrations were determined by liquid chromatography tandem mass spectrometry in 25 paired CSF and plasma samples from 16 HIV-1-infected individuals. The lower limit of quantitation was 2.0 ng/ml for CSF and 10 ng/ml for plasma.Twenty-four of the 25 CSF samples had detectable raltegravir concentrations with a median raltegravir concentration of 18.4 ng/ml (range, <2.0-126.0. The median plasma raltegravir concentration was 448 ng/ml (range, 37-5180. CSF raltegravir concentrations correlated with CSF:plasma albumin ratios and CSF albumin concentrations.Approximately 50% of the CSF specimens exceeded the IC(95 levels reported to inhibit HIV-1 strains without resistance to integrase inhibitors. In addition to contributing to control of systemic HIV-1 infection, raltegravir achieves local inhibitory concentrations in CSF in most, but not all, patients. Blood-brain and blood-CSF barriers likely restrict drug entry, while enhanced permeability of these barriers enhances drug entry.

  19. [HIV-1 genetic variability in non Spaniard infected children].

    Science.gov (United States)

    Piñeiro Pérez, R; Mellado Peña, M J; Holguín, A; Cilleruelo, M J; García Hortelano, M; Villota, J; Martín Fontelos, P

    2009-01-01

    The prevalence of HIV-1 non-B subtypes (HIV-NBS) is increasing in Europe, because of emigration from countries where genetic variants are endemic. Although HIV-NBS could have a different clinical evolution and could respond differently to antiretrovirals (AR) than B-subtypes, these variant's response remain undocumented. To identify HIV-1 genetic variants and to determine clinical evolution in a non-Spaniard children infected with HIV-1. Children with HIV-1 infection from endemic countries were tested for HIV-1 subtypes between 1-1-1988 and 31-12-2006. Twelve children less than 18 years old and born abroad were selected. HIV-NBS were isolated in 5 children (42%): CRF2_AG recombinant in 3 cases (Equatorial Guinea), Subtype C in one (Equatorial Guinea) and CRF13_cpx in last one (India). Because of the increasing frequency of patients with HIV-NBS and their unknown long-term evolution, all children from endemic countries should be tested for HIV subtypes. We believe new studies with more patients during longer times could reveal differences in these patient's clinical, immunological and virological evolution.

  20. Raltegravir cerebrospinal fluid concentrations in HIV-1 infection.

    Science.gov (United States)

    Yilmaz, Aylin; Gisslén, Magnus; Spudich, Serena; Lee, Evelyn; Jayewardene, Anura; Aweeka, Francesca; Price, Richard W

    2009-09-01

    Raltegravir is an HIV-1 integrase inhibitor currently used in treatment-experienced HIV-1-infected patients resistant to other drug classes. In order to assess its central nervous system penetration, we measured raltegravir concentrations in cerebrospinal fluid (CSF) and plasma in subjects receiving antiretroviral treatment regimens containing this drug. Raltegravir concentrations were determined by liquid chromatography tandem mass spectrometry in 25 paired CSF and plasma samples from 16 HIV-1-infected individuals. The lower limit of quantitation was 2.0 ng/ml for CSF and 10 ng/ml for plasma. Twenty-four of the 25 CSF samples had detectable raltegravir concentrations with a median raltegravir concentration of 18.4 ng/ml (range, <2.0-126.0). The median plasma raltegravir concentration was 448 ng/ml (range, 37-5180). CSF raltegravir concentrations correlated with CSF:plasma albumin ratios and CSF albumin concentrations. Approximately 50% of the CSF specimens exceeded the IC(95) levels reported to inhibit HIV-1 strains without resistance to integrase inhibitors. In addition to contributing to control of systemic HIV-1 infection, raltegravir achieves local inhibitory concentrations in CSF in most, but not all, patients. Blood-brain and blood-CSF barriers likely restrict drug entry, while enhanced permeability of these barriers enhances drug entry.

  1. Neuropsychological performance in patients with asymptomatic HIV-1 infection.

    Science.gov (United States)

    Martínez-Banfi, Martha; Vélez, Jorge I; Perea, M Victoria; García, Ricardo; Puentes-Rozo, Pedro J; Mebarak Chams, Moises; Ladera, Valentina

    2018-05-01

    Human immunodeficiency virus (HIV-1) infection and acquired immunodeficiency syndrome (AIDS) lead to neurocognitive disorders; however, there is still much knowledge to be gained regarding HIV-associated neurocognitive disorders. The purpose of this study was to assess the cognitive performance, instrumental activities of daily living, depression, and anxiety in patients with asymptomatic HIV-1 infections compared with seronegative participants without neurocognitive impairment. We studied a sample consisted of 60 patients with asymptomatic HIV-1 infections and 60 seronegative participants without neurocognitive impairment from the city of Barranquilla, Colombia, with a mean age of 36.07 years. A protocol of neuropsychological and psychopathological tests was applied to the participants. The group of patients with asymptomatic HIV infections significantly underperformed on tasks that assessed global cognitive screening, attention span, learning, phonemic verbal fluency, auditory-verbal comprehension, information processing speed, cognitive flexibility, and motor skills compared to the group of seronegative participants. No significant differences were found in memory, visual confrontation naming, vocabulary, inhibition, and instrumental activities of daily living. Additionally, the patients with asymptomatic HIV-1 infection had a higher anxiety index than the seronegative participants, but no significant difference was found in depression. A correlation was found between depression and anxiety. In conclusion, the patients with asymptomatic HIV-1 infection had lower cognitive performances than the seronegative participants in the cognitive functions mentioned above and more anxiety but still performed the instrumental activities of daily living.

  2. Sieve analysis in HIV-1 vaccine efficacy trials.

    Science.gov (United States)

    Edlefsen, Paul T; Gilbert, Peter B; Rolland, Morgane

    2013-09-01

    The genetic characterization of HIV-1 breakthrough infections in vaccine and placebo recipients offers new ways to assess vaccine efficacy trials. Statistical and sequence analysis methods provide opportunities to mine the mechanisms behind the effect of an HIV vaccine. The release of results from two HIV-1 vaccine efficacy trials, Step/HVTN-502 (HIV Vaccine Trials Network-502) and RV144, led to numerous studies in the last 5 years, including efforts to sequence HIV-1 breakthrough infections and compare viral characteristics between the vaccine and placebo groups. Novel genetic and statistical analysis methods uncovered features that distinguished founder viruses isolated from vaccinees from those isolated from placebo recipients, and identified HIV-1 genetic targets of vaccine-induced immune responses. Studies of HIV-1 breakthrough infections in vaccine efficacy trials can provide an independent confirmation to correlates of risk studies, as they take advantage of vaccine/placebo comparisons, whereas correlates of risk analyses are limited to vaccine recipients. Through the identification of viral determinants impacted by vaccine-mediated host immune responses, sieve analyses can shed light on potential mechanisms of vaccine protection.

  3. Antimalarial activity of HIV-1 protease inhibitor in chromone series.

    Science.gov (United States)

    Lerdsirisuk, Pradith; Maicheen, Chirattikan; Ungwitayatorn, Jiraporn

    2014-12-01

    Increasing parasite resistance to nearly all available antimalarial drugs becomes a serious problem to human health and necessitates the need to continue the search for new effective drugs. Recent studies have shown that clinically utilized HIV-1 protease (HIV-1 PR) inhibitors can inhibit the in vitro and in vivo growth of Plasmodium falciparum. In this study, a series of chromone derivatives possessing HIV-1 PR inhibitory activity has been tested for antimalarial activity against P. falciparum (K1 multi-drug resistant strain). Chromone 15, the potent HIV-1 PR inhibitor (IC50=0.65μM), was found to be the most potent antimalarial compound with IC50=0.95μM while primaquine and tafenoquine showed IC50=2.41 and 1.95μM, respectively. Molecular docking study of chromone compounds against plasmepsin II, an aspartic protease enzyme important in hemoglobin degradation, revealed that chromone 15 exhibited the higher binding affinity (binding energy=-13.24kcal/mol) than the known PM II inhibitors. Thus, HIV-1 PR inhibitor in chromone series has the potential to be a new class of antimalarial agent. Copyright © 2014 Elsevier Inc. All rights reserved.

  4. Mechanisms for Cell-to-Cell Transmission of HIV-1

    Science.gov (United States)

    Bracq, Lucie; Xie, Maorong; Benichou, Serge; Bouchet, Jérôme

    2018-01-01

    While HIV-1 infection of target cells with cell-free viral particles has been largely documented, intercellular transmission through direct cell-to-cell contact may be a predominant mode of propagation in host. To spread, HIV-1 infects cells of the immune system and takes advantage of their specific particularities and functions. Subversion of intercellular communication allows to improve HIV-1 replication through a multiplicity of intercellular structures and membrane protrusions, like tunneling nanotubes, filopodia, or lamellipodia-like structures involved in the formation of the virological synapse. Other features of immune cells, like the immunological synapse or the phagocytosis of infected cells are hijacked by HIV-1 and used as gateways to infect target cells. Finally, HIV-1 reuses its fusogenic capacity to provoke fusion between infected donor cells and target cells, and to form infected syncytia with high capacity of viral production and improved capacities of motility or survival. All these modes of cell-to-cell transfer are now considered as viral mechanisms to escape immune system and antiretroviral therapies, and could be involved in the establishment of persistent virus reservoirs in different host tissues. PMID:29515578

  5. Transcriptional and Bioinformatic Analysis Provide a Relationship between Host Response Changes to Marek’s Disease Viruses Infection and an Integrated Long Terminal Repeat

    Directory of Open Access Journals (Sweden)

    Ning eCui

    2016-04-01

    Full Text Available GX0101, Marek’s disease virus (MDV strain with a long terminal repeat (LTR insert of reticuloendotheliosis virus (REV, was isolated from CVI988/Rispens vaccinated birds showing tumors. We have constructed a LTR deleted strain GX0101∆LTR in our previous study. To compare the host responses to GX0101 and GX0101∆LTR, chicken embryo fibroblasts (CEF cells were infected with two MDV strains and a gene-chip containing chicken genome was employed to examine gene transcription changes in host cells in the present study. Of the 42 368 chicken transcripts on the chip, there were 2199 genes that differentially expressed in CEF infected with GX0101 compared to GX0101∆LTR significantly. Differentially expressed genes were distributed to 25 possible gene networks according to their intermolecular connections and were annotated to 56 pathways. The insertion of REV LTR showed the greatest influence on cancer formation and metastasis, followed with immune changes, atherosclerosis and nervous system disorders in MDV-infected CEF cells. Based on these bio functions, GX0101 infection was predicated with a greater growth and survival inhibition but lower oncogenicity in chickens than GX0101∆LTR, at least in the acute phase of infection. In summary, the insertion of REV LTR altered the expression of host genes in response to MDV infection, possibly resulting in novel phenotypic properties in chickens. Our study has provided the evidence of retroviral insertional changes of host responses to herpesvirus infection for the first time, which will promote to elucidation of the possible relationship between the LTR insertion and the observed phenotypes.

  6. [Analysis on HIV-1 genetics and threshold of drug resistance in Dehong prefecture of Yunnan province in 2013].

    Science.gov (United States)

    Ma, Yanling; Wang, Jibao; Xing, Hui; Chen, Min; Yao, Shitang; Chen, Huichao; Yang, Jin; Li, Yanling; Duan, Song; Jia, Manhong

    2015-06-01

    To study the HIV-1 genotypes and transmitted drug resistance (TDR) in Dehong prefecture of Yunnan province in 2013. Referring to the guidelines for HIV drug resistance threshold survey (HIVDR-TS), 54 plasma samples of recently reported HIV-infected individuals, aged between 16 and 25 years, were collected in Dehong prefecture from January to August 2013. Genotyping of partial pol gene was performed by using reverse transcriptional PCR. HIV-1 genotype. Prevalent levels of HIV-1 drug resistance transmission were analyzed. Forty-eight plasma samples were successfully sequenced and analyzed. Among them, 45.8% were Chinese and the rest 54.2% were all Burmese. Based on pol sequences, identified HIV genotypes included subtype C (41.7%), URF (31.3%), CRF01_AE (12.5%), CRF07_BC (10.4%), CRF08_BC (2.1%) and subtype B (2.1%), C subtype appeared dominated in Chinese while URF was dominated in Burmese. One drug resistant mutation to non-nucleoside reverse transcriptase inhibitors (NNRTIs) was detected in one sequence from Burmese. Based on the statistical method of HIVDR-TS, the prevalence of transmitted HIV-1 drug resistance was adjusted as scientific management for people living with HIV/AIDS should be strictly followed. Meanwhile, relevant surveillance, including drug resistance surveillance should also be performed among cross-border migrant population.

  7. No evidence of association between HIV-1 and malaria in populations with low HIV-1 prevalence.

    Directory of Open Access Journals (Sweden)

    Diego F Cuadros

    Full Text Available The geographic overlap between HIV-1 and malaria has generated much interest in their potential interactions. A variety of studies have evidenced a complex HIV-malaria interaction within individuals and populations that may have dramatic effects, but the causes and implications of this co-infection at the population level are still unclear. In a previous publication, we showed that the prevalence of malaria caused by the parasite Plasmodium falciparum is associated with HIV infection in eastern sub-Saharan Africa. To complement our knowledge of the HIV-malaria co-infection, the objective of this work was to assess the relationship between malaria and HIV prevalence in the western region of sub-Saharan Africa.Population-based cross-sectional data were obtained from the HIV/AIDS Demographic and Health Surveys conducted in Burkina Faso, Ghana, Guinea, Mali, Liberia and Cameroon, and the malaria atlas project. Using generalized linear mixed models, we assessed the relationship between HIV-1 and Plasmodium falciparum parasite rate (PfPR adjusting for important socio-economic and biological cofactors. We found no evidence that individuals living in areas with stable malaria transmission (PfPR>0.46 have higher odds of being HIV-positive than individuals who live in areas with PfPR≤0.46 in western sub-Saharan Africa (estimated odds ratio 1.14, 95% confidence interval 0.86-1.50. In contrast, the results suggested that PfPR was associated with being infected with HIV in Cameroon (estimated odds ratio 1.56, 95% confidence interval 1.23-2.00.Contrary to our previous research on eastern sub-Saharan Africa, this study did not identify an association between PfPR and infection with HIV in western sub-Saharan Africa, which suggests that malaria might not play an important role in the spread of HIV in populations where the HIV prevalence is low. Our work highlights the importance of understanding the epidemiologic effect of co-infection and the relevant

  8. Artificial 64-Residue HIV-1 Enhancer-Binding Peptide Is a Potent Inhibitor of Viral Replication in HIV-1-Infected Cells.

    Science.gov (United States)

    Oufir, Mouhssin; Bisset, Leslie R; Hoffmann, Stefan R K; Xue, Gongda; Klauser, Stephan; Bergamaschi, Bianca; Gervaix, Alain; Böni, Jürg; Schüpbach, Jörg; Gutte, Bernd

    2011-01-01

    An artificial HIV-1 enhancer-binding peptide was extended by nine consecutive arginine residues at the C-terminus and by the nuclear localization signal of SV40 large T antigen at the N-terminus. The resulting synthetic 64-residue peptide was found to bind to the two enhancers of the HIV-1 long terminal repeat, cross the plasma membrane and the nuclear envelope of human cells, and suppress the HIV-1 enhancer-controlled expression of a green fluorescent protein reporter gene. Moreover, HIV-1 replication is inhibited by this peptide in HIV-1-infected CEM-GFP cells as revealed by HIV-1 p24 ELISA and real-time RT-PCR of HIV-1 RNA. Rapid uptake of this intracellular stable and inhibitory peptide into the cells implies that this peptide may have the potential to attenuate HIV-1 replication in vivo.

  9. Artificial 64-Residue HIV-1 Enhancer-Binding Peptide Is a Potent Inhibitor of Viral Replication in HIV-1-Infected Cells

    Directory of Open Access Journals (Sweden)

    Mouhssin Oufir

    2011-01-01

    Full Text Available An artificial HIV-1 enhancer-binding peptide was extended by nine consecutive arginine residues at the C-terminus and by the nuclear localization signal of SV40 large T antigen at the N-terminus. The resulting synthetic 64-residue peptide was found to bind to the two enhancers of the HIV-1 long terminal repeat, cross the plasma membrane and the nuclear envelope of human cells, and suppress the HIV-1 enhancer-controlled expression of a green fluorescent protein reporter gene. Moreover, HIV-1 replication is inhibited by this peptide in HIV-1-infected CEM-GFP cells as revealed by HIV-1 p24 ELISA and real-time RT-PCR of HIV-1 RNA. Rapid uptake of this intracellular stable and inhibitory peptide into the cells implies that this peptide may have the potential to attenuate HIV-1 replication in vivo.

  10. Mining the Human Complexome Database Identifies RBM14 as an XPO1-Associated Protein Involved in HIV-1 Rev Function

    OpenAIRE

    Budhiraja, Sona; Liu, Hongbing; Couturier, Jacob; Malovannaya, Anna; Qin, Jun; Lewis, Dorothy E.; Rice, Andrew P.

    2015-01-01

    By recruiting the host protein XPO1 (CRM1), the HIV-1 Rev protein mediates the nuclear export of incompletely spliced viral transcripts. We mined data from the recently described human nuclear complexome to identify a host protein, RBM14, which associates with XPO1 and Rev and is involved in Rev function. Using a Rev-dependent p24 reporter plasmid, we found that RBM14 depletion decreased Rev activity and Rev-mediated enhancement of the cytoplasmic levels of unspliced viral transcripts. RBM14 ...

  11. Interleukin-27 is a potent inhibitor of cis HIV-1 replication in monocyte-derived dendritic cells via a type I interferon-independent pathway.

    Directory of Open Access Journals (Sweden)

    Qian Chen

    Full Text Available IL-27, a member of the IL-12 family of cytokines, plays an important and diverse role in the function of the immune system. Whilst generally recognized as an anti-inflammatory cytokine, in addition IL-27 has been found to have broad anti-viral effects. Recently, IL-27 has been shown to be a potent inhibitor of HIV-1 infection in CD4+ T cells and macrophages. The main objective of this study was to see whether IL-27 has a similar inhibitory effect on HIV-1 replication in dendritic cells (DCs. Monocytes were differentiated into immature DCs (iDCs and mature DCs (mDCs with standard techniques using a combination of GM-CSF, IL-4 and LPS. Following differentiation, iDCs were infected with HIV-1 and co-cultured in the presence or absence of IL-27. IL-27 treated DCs were shown to be highly potent inhibitors of cis HIV-1, particularly of CCR5 tropic strains. Of note, other IL-12 family members (IL-12, IL-23 and IL-35 had no effect on HIV-1 replication. Microarray studies of IL-27 treated DCs showed no up-regulation of Type I (IFN gene expression. Neutralization of the Type-I IFN receptor had no impact on the HIV inhibition. Lastly, IL-27 mediated inhibition was shown to act post-viral entry and prior to completion of reverse transcription. These results show for the first time that IL-27 is a potent inhibitor of cis HIV-1 infection in DCs by a Type I IFN independent mechanism. IL-27 has previously been reported to inhibit HIV-1 replication in CD4+ T cells and macrophages, thus taken together, this cytokine is a potent anti-HIV agent against all major cell types targeted by the HIV-1 virus and may have a therapeutic role in the future.

  12. Back to the future: revisiting HIV-1 lethal mutagenesis

    Science.gov (United States)

    Dapp, Michael J.; Patterson, Steven E.; Mansky, Louis M.

    2012-01-01

    The concept of eliminating HIV-1 infectivity by elevating the viral mutation rate was first proposed over a decade ago, even though the general concept had been conceived earlier for RNA viruses. Lethal mutagenesis was originally viewed as a novel chemotherapeutic approach for treating HIV-1 infection in which use of a viral mutagen would over multiple rounds of replication lead to the lethal accumulation of mutations, rendering the virus population non infectious – known as the slow mutation accumulation model. There have been limitations in obtaining good efficacy data with drug leads, leaving some doubt into clinical translation. More recent studies of the APOBEC3 proteins as well as new progress in the use of nucleoside analogs for inducing lethal mutagenesis have helped to refocus attention on rapid induction of HIV-1 lethal mutagenesis in a single or limited number of replication cycles leading to a rapid mutation accumulation model. PMID:23195922

  13. Increased T cell trafficking as adjunct therapy for HIV-1

    Science.gov (United States)

    Wolinsky, Steven M.; McLean, Angela R.

    2018-01-01

    Although antiretroviral drug therapy suppresses human immunodeficiency virus-type 1 (HIV-1) to undetectable levels in the blood of treated individuals, reservoirs of replication competent HIV-1 endure. Upon cessation of antiretroviral therapy, the reservoir usually allows outgrowth of virus and approaches to targeting the reservoir have had limited success. Ongoing cycles of viral replication in regions with low drug penetration contribute to this persistence. Here, we use a mathematical model to illustrate a new approach to eliminating the part of the reservoir attributable to persistent replication in drug sanctuaries. Reducing the residency time of CD4 T cells in drug sanctuaries renders ongoing replication unsustainable in those sanctuaries. We hypothesize that, in combination with antiretroviral drugs, a strategy to orchestrate CD4 T cell trafficking could contribute to a functional cure for HIV-1 infection. PMID:29499057

  14. Copy number variation of KIR genes influences HIV-1 control

    DEFF Research Database (Denmark)

    Pelak, Kimberly; Need, Anna C; Fellay, Jacques

    2011-01-01

    A genome-wide screen for large structural variants showed that a copy number variant (CNV) in the region encoding killer cell immunoglobulin-like receptors (KIR) associates with HIV-1 control as measured by plasma viral load at set point in individuals of European ancestry. This CNV encompasses...... the KIR3DL1-KIR3DS1 locus, encoding receptors that interact with specific HLA-Bw4 molecules to regulate the activation of lymphocyte subsets including natural killer (NK) cells. We quantified the number of copies of KIR3DS1 and KIR3DL1 in a large HIV-1 positive cohort, and showed that an increase in KIR3...... amounts of these activating and inhibitory KIR play a role in regulating the peripheral expansion of highly antiviral KIR3DS1+ NK cells, which may determine differences in HIV-1 control following infection....

  15. Glycosylation in HIV-1 envelope glycoprotein and its biological implications

    KAUST Repository

    Ho, Yung Shwen

    2013-08-01

    Glycosylation of HIV-1 envelope proteins (Env gp120/gp41) plays a vital role in viral evasion from the host immune response, which occurs through the masking of key neutralization epitopes and the presentation of the Env glycosylation as \\'self\\' to the host immune system. Env glycosylation is generally conserved, yet its continual evolution plays an important role in modulating viral infectivity and Env immunogenicity. Thus, it is believed that Env glycosylation, which is a vital part of the HIV-1 architecture, also controls intra- and inter-clade genetic variations. Discerning intra- and inter-clade glycosylation variations could therefore yield important information for understanding the molecular and biological differences between HIV clades and may assist in effectively designing Env-based immunogens and in clearly understanding HIV vaccines. This review provides an in-depth perspective of various aspects of Env glycosylation in the context of HIV-1 pathogenesis. © 2013 Future Medicine Ltd.

  16. Redefining the Viral Reservoirs That Prevent HIV-1 Eradication

    Science.gov (United States)

    Eisele, Evelyn; Siliciano, Robert F.

    2014-01-01

    Summary This review proposes definitions for key terms in the field of HIV-1 latency and eradication. In the context of eradication, a reservoir is a cell type that allows persistence of replication-competent HIV-1 on a time scale of years in patients on optimal antiretroviral therapy. Reservoirs act as a barrier to eradication in the patient population in whom cure attempts will likely be made. Halting viral replication is essential to eradication, and definitions and criteria for assessing whether this goal has been achieved are proposed. The cell types that may serve as reservoirs for HIV-1 are discussed. Currently, only latently infected resting CD4+ T cells fit the proposed definition of a reservoir, and more evidence is necessary to demonstrate that other cell types including hematopoietic stem cells and macrophages fit this definition. Further research is urgently required on potential reservoirs in the gut-associated lymphoid tissue and the central nervous system. PMID:22999944

  17. HLA Class I-Mediated HIV-1 Control in Vietnamese Infected with HIV-1 Subtype A/E.

    Science.gov (United States)

    Chikata, Takayuki; Tran, Giang Van; Murakoshi, Hayato; Akahoshi, Tomohiro; Qi, Ying; Naranbhai, Vivek; Kuse, Nozomi; Tamura, Yoshiko; Koyanagi, Madoka; Sakai, Sachiko; Nguyen, Dung Hoai; Nguyen, Dung Thi; Nguyen, Ha Thu; Nguyen, Trung Vu; Oka, Shinichi; Martin, Maureen P; Carrington, Mary; Sakai, Keiko; Nguyen, Kinh Van; Takiguchi, Masafumi

    2018-03-01

    HIV-1-specific cytotoxic T cells (CTLs) play an important role in the control of HIV-1 subtype B or C infection. However, the role of CTLs in HIV-1 subtype A/E infection still remains unclear. Here we investigated the association of HLA class I alleles with clinical outcomes in treatment-naive Vietnamese infected with subtype A/E virus. We found that HLA-C*12:02 was significantly associated with lower plasma viral loads (pVL) and higher CD4 counts and that the HLA-A*29:01-B*07:05-C*15:05 haplotype was significantly associated with higher pVL and lower CD4 counts than those for individuals without these respective genotypes. Nine Pol and three Nef mutations were associated with at least one HLA allele in the HLA-A*29:01-B*07:05-C*15:05 haplotype, with a strong negative correlation between the number of HLA-associated Pol mutations and CD4 count as well as a positive correlation with pVL for individuals with these HLA alleles. The results suggest that the accumulation of mutations selected by CTLs restricted by these HLA alleles affects HIV control. IMPORTANCE Most previous studies on HLA association with disease progression after HIV-1 infection have been performed on cohorts infected with HIV-1 subtypes B and C, whereas few such population-based studies have been reported for cohorts infected with the Asian subtype A/E virus. In this study, we analyzed the association of HLA class I alleles with clinical outcomes for 536 HIV-1 subtype A/E-infected Vietnamese individuals. We found that HLA-C*12:02 is protective, while the HLA haplotype HLA-A*29:01-B*07:05-C*15:05 is deleterious. The individuals with HIV-1 mutations associated with at least one of the HLA alleles in the deleterious HLA haplotype had higher plasma viral loads and lower CD4 counts than those of individuals without the mutations, suggesting that viral adaptation and escape from HLA-mediated immune control occurred. The present study identifies a protective allele and a deleterious haplotype for HIV-1

  18. Evolutionary genomics revealed interkingdom distribution of Tcn1-like chromodomain-containing Gypsy LTR retrotransposons among fungi and plants

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    Blinov Alexander

    2010-04-01

    Full Text Available Abstract Background Chromodomain-containing Gypsy LTR retrotransposons or chromoviruses are widely distributed among eukaryotes and have been found in plants, fungi and vertebrates. The previous comprehensive survey of chromoviruses from mosses (Bryophyta suggested that genomes of non-seed plants contain the clade which is closely related to the retrotransposons from fungi. The origin, distribution and evolutionary history of this clade remained unclear mainly due to the absence of information concerning the diversity and distribution of LTR retrotransposons in other groups of non-seed plants as well as in fungal genomes. Results In present study we preformed in silico analysis of chromodomain-containing LTR retrotransposons in 25 diverse fungi and a number of plant species including spikemoss Selaginella moellendorffii (Lycopodiophyta coupled with an experimental survey of chromodomain-containing Gypsy LTR retrotransposons from diverse non-seed vascular plants (lycophytes, ferns, and horsetails. Our mining of Gypsy LTR retrotransposons in genomic sequences allowed identification of numerous families which have not been described previously in fungi. Two new well-supported clades, Galahad and Mordred, as well as several other previously unknown lineages of chromodomain-containing Gypsy LTR retrotransposons were described based on the results of PCR-mediated survey of LTR retrotransposon fragments from ferns, horsetails and lycophytes. It appeared that one of the clades, namely Tcn1 clade, was present in basidiomycetes and non-seed plants including mosses (Bryophyta and lycophytes (genus Selaginella. Conclusions The interkingdom distribution is not typical for chromodomain-containing LTR retrotransposons clades which are usually very specific for a particular taxonomic group. Tcn1-like LTR retrotransposons from fungi and non-seed plants demonstrated high similarity to each other which can be explained by strong selective constraints and the

  19. Selective elimination of HIV-1-infected cells by Env-directed, HIV-1-based virus-like particles

    International Nuclear Information System (INIS)

    Peretti, Silvia; Schiavoni, Ilaria; Pugliese, Katherina; Federico, Maurizio

    2006-01-01

    We recently showed that both replicating and resting cells cultivated with ganciclovir (GCV) were killed when challenged with vesicular stomatitis virus G glycoprotein pseudotyped HIV-1-based virus-like particles (VLPs) carrying the Nef7 (i.e., an HIV-1 Nef mutant incorporating in virions at high levels)/herpes simplex virus-1 thymidine kinase (HSV-TK) fusion product. On this basis, a novel anti-HIV therapeutic approach based on Nef7/TK VLPs expressing X4 or R5 HIV cell receptor complexes has been attempted. We here report that (CD4-CXCR4) and (CD4-CCR5) Nef7-based VLPs efficiently enter cells infected by X4- or R5-tropic HIV-1 strains, respectively. Importantly, the delivery of the VLP-associated Nef7/TK led to cell death upon GCV treatment. Of interest, VLPs were effective also against non-replicating, HIV-1-infected primary human monocyte-derived macrophages. HIV-targeted VLPs represent a promising candidate for the treatment of persistently HIV-1-infected cells that are part of virus reservoirs resistant to HAART therapies

  20. HIV-1 induces DCIR expression in CD4+ T cells.

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    Alexandra A Lambert

    2010-11-01

    Full Text Available The C-type lectin receptor DCIR, which has been shown very recently to act as an attachment factor for HIV-1 in dendritic cells, is expressed predominantly on antigen-presenting cells. However, this concept was recently challenged by the discovery that DCIR can also be detected in CD4(+ T cells found in the synovial tissue from rheumatoid arthritis (RA patients. Given that RA and HIV-1 infections share common features such as a chronic inflammatory condition and polyclonal immune hyperactivation status, we hypothesized that HIV-1 could promote DCIR expression in CD4(+ T cells. We report here that HIV-1 drives DCIR expression in human primary CD4(+ T cells isolated from patients (from both aviremic/treated and viremic/treatment naive persons and cells acutely infected in vitro (seen in both virus-infected and uninfected cells. Soluble factors produced by virus-infected cells are responsible for the noticed DCIR up-regulation on uninfected cells. Infection studies with Vpr- or Nef-deleted viruses revealed that these two viral genes are not contributing to the mechanism of DCIR induction that is seen following acute infection of CD4(+ T cells with HIV-1. Moreover, we report that DCIR is linked to caspase-dependent (induced by a mitochondria-mediated generation of free radicals and -independent intrinsic apoptotic pathways (involving the death effector AIF. Finally, we demonstrate that the higher surface expression of DCIR in CD4(+ T cells is accompanied by an enhancement of virus attachment/entry, replication and transfer. This study shows for the first time that HIV-1 induces DCIR membrane expression in CD4(+ T cells, a process that might promote virus dissemination throughout the infected organism.

  1. Flail arm-like syndrome associated with HIV-1 infection

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    Nalini A

    2009-01-01

    Full Text Available During the last 20 years at least 23 cases of motor neuron disease have been reported in HIV-1 seropositive patients. In this report we describe the clinical picture of a young man with HIV-1 clade C infection and flail arm-like syndrome, who we were able to follow-up for a long period. We investigated and prospectively monitored a 34-year-old man with features of flail arm syndrome, who developed the weakness and wasting 1 year after being diagnosed with HIV-1 infection after a routine blood test. He presented in 2003 with progressive, symmetrical wasting and weakness of the proximal muscles of the upper limb of 2 years′ duration. He had severe wasting and weakness of the shoulder and arm muscles. There were no pyramidal signs. He has been on HAART for the last 4 years and the weakness or wasting has not worsened. At the last follow-up in July 2007, the patient had the same neurological deficit and no other symptoms or signs of HIV-1 infection. MRI of the spinal cord in 2007 showed characteristic T2 hyperintense signals in the central part of the spinal cord, corresponding to the central gray matter. Thus, our patient had HIV-1 clade C infection associated with a ′flail arm-like syndrome.′ The causal relationship between HIV-1 infection and amyotrophic lateral sclerosis (ALS-like syndrome is still uncertain. The syndrome usually manifests as a lower motor neuron syndrome, as was seen in our young patient. It is known that treatment with antiretroviral therapy (ART stabilizes/improves the condition. In our patient the weakness and atrophy remained stable over a period of 3.5 years after commencing HAART regimen.

  2. HIV-1 protease-substrate coevolution in nelfinavir resistance.

    Science.gov (United States)

    Kolli, Madhavi; Ozen, Ayşegül; Kurt-Yilmaz, Nese; Schiffer, Celia A

    2014-07-01

    Resistance to various human immunodeficiency virus type 1 (HIV-1) protease inhibitors (PIs) challenges the effectiveness of therapies in treating HIV-1-infected individuals and AIDS patients. The virus accumulates mutations within the protease (PR) that render the PIs less potent. Occasionally, Gag sequences also coevolve with mutations at PR cleavage sites contributing to drug resistance. In this study, we investigated the structural basis of coevolution of the p1-p6 cleavage site with the nelfinavir (NFV) resistance D30N/N88D protease mutations by determining crystal structures of wild-type and NFV-resistant HIV-1 protease in complex with p1-p6 substrate peptide variants with L449F and/or S451N. Alterations of residue 30's interaction with the substrate are compensated by the coevolving L449F and S451N cleavage site mutations. This interdependency in the PR-p1-p6 interactions enhances intermolecular contacts and reinforces the overall fit of the substrate within the substrate envelope, likely enabling coevolution to sustain substrate recognition and cleavage in the presence of PR resistance mutations. Resistance to human immunodeficiency virus type 1 (HIV-1) protease inhibitors challenges the effectiveness of therapies in treating HIV-1-infected individuals and AIDS patients. Mutations in HIV-1 protease selected under the pressure of protease inhibitors render the inhibitors less potent. Occasionally, Gag sequences also mutate and coevolve with protease, contributing to maintenance of viral fitness and to drug resistance. In this study, we investigated the structural basis of coevolution at the Gag p1-p6 cleavage site with the nelfinavir (NFV) resistance D30N/N88D protease mutations. Our structural analysis reveals the interdependency of protease-substrate interactions and how coevolution may restore substrate recognition and cleavage in the presence of protease drug resistance mutations. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

  3. Human cellular restriction factors that target HIV-1 replication

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    Jeang Kuan-Teh

    2009-09-01

    Full Text Available Abstract Recent findings have highlighted roles played by innate cellular factors in restricting intracellular viral replication. In this review, we discuss in brief the activities of apolipoprotein B mRNA-editing enzyme 3G (APOBEC3G, bone marrow stromal cell antigen 2 (BST-2, cyclophilin A, tripartite motif protein 5 alpha (Trim5α, and cellular microRNAs as examples of host restriction factors that target HIV-1. We point to countermeasures encoded by HIV-1 for moderating the potency of these cellular restriction functions.

  4. The Latent Reservoir for HIV-1: How Immunologic Memory and Clonal Expansion Contribute to HIV-1 Persistence

    Science.gov (United States)

    Murray, Alexandra J.; Kwon, Kyungyoon J.; Farber, Donna L.; Siliciano, Robert F.

    2016-01-01

    Combination antiretroviral therapy (ART) for HIV-1 infection reduces plasma virus levels to below the limit of detection of clinical assays. However, even with prolonged suppression of viral replication with ART, viremia rebounds rapidly after treatment interruption. Thus ART is not curative. The principal barrier to cure is a remarkably stable reservoir of latent HIV-1 in resting memory CD4+ T cells. Here we consider explanations for the remarkable stability of the latent reservoir. Stability does not appear to reflect replenishment from new infection events but rather normal physiologic processes that provide for immunologic memory. Of particular importance are proliferative processes that drive clonal expansion of infected cells. Recent evidence suggests that in some infected cells, proliferation is a consequence of proviral integration into host genes associated with cell growth. Efforts to cure HIV-1 infection by targeting the latent reservoir may need to consider the potential of latently infected cells to proliferate. PMID:27382129

  5. Recognition of HIV-1 peptides by host CTL is related to HIV-1 similarity to human proteins.

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    Morgane Rolland

    Full Text Available BACKGROUND: While human immunodeficiency virus type 1 (HIV-1-specific cytotoxic T lymphocytes preferentially target specific regions of the viral proteome, HIV-1 features that contribute to immune recognition are not well understood. One hypothesis is that similarities between HIV and human proteins influence the host immune response, i.e., resemblance between viral and host peptides could preclude reactivity against certain HIV epitopes. METHODOLOGY/PRINCIPAL FINDINGS: We analyzed the extent of similarity between HIV-1 and the human proteome. Proteins from the HIV-1 B consensus sequence from 2001 were dissected into overlapping k-mers, which were then probed against a non-redundant database of the human proteome in order to identify segments of high similarity. We tested the relationship between HIV-1 similarity to host encoded peptides and immune recognition in HIV-infected individuals, and found that HIV immunogenicity could be partially modulated by the sequence similarity to the host proteome. ELISpot responses to peptides spanning the entire viral proteome evaluated in 314 individuals showed a trend indicating an inverse relationship between the similarity to the host proteome and the frequency of recognition. In addition, analysis of responses by a group of 30 HIV-infected individuals against 944 overlapping peptides representing a broad range of individual HIV-1B Nef variants, affirmed that the degree of similarity to the host was significantly lower for peptides with reactive epitopes than for those that were not recognized. CONCLUSIONS/SIGNIFICANCE: Our results suggest that antigenic motifs that are scarcely represented in human proteins might represent more immunogenic CTL targets not selected against in the host. This observation could provide guidance in the design of more effective HIV immunogens, as sequences devoid of host-like features might afford superior immune reactivity.

  6. The porcine circovirus type 1 capsid gene promoter improves antigen expression and immunogenicity in a HIV-1 plasmid vaccine

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    Burger Marieta

    2011-02-01

    Full Text Available Abstract Background One of the promising avenues for development of vaccines against Human immunodeficiency virus type 1 (HIV-1 and other human pathogens is the use of plasmid-based DNA vaccines. However, relatively large doses of plasmid must be injected for a relatively weak response. We investigated whether genome elements from Porcine circovirus type 1 (PCV-1, an apathogenic small ssDNA-containing virus, had useful expression-enhancing properties that could allow dose-sparing in a plasmid vaccine. Results The linearised PCV-1 genome inserted 5' of the CMV promoter in the well-characterised HIV-1 plasmid vaccine pTHgrttnC increased expression of the polyantigen up to 2-fold, and elicited 3-fold higher CTL responses in mice at 10-fold lower doses than unmodified pTHgrttnC. The PCV-1 capsid gene promoter (Pcap alone was equally effective. Enhancing activity was traced to a putative composite host transcription factor binding site and a "Conserved Late Element" transcription-enhancing sequence previously unidentified in circoviruses. Conclusions We identified a novel PCV-1 genome-derived enhancer sequence that significantly increased antigen expression from plasmids in in vitro assays, and improved immunogenicity in mice of the HIV-1 subtype C vaccine plasmid, pTHgrttnC. This should allow significant dose sparing of, or increased responses to, this and other plasmid-based vaccines. We also report investigations of the potential of other circovirus-derived sequences to be similarly used.

  7. Fluorescent protein-tagged Vpr dissociates from HIV-1 core after viral fusion and rapidly enters the cell nucleus.

    Science.gov (United States)

    Desai, Tanay M; Marin, Mariana; Sood, Chetan; Shi, Jiong; Nawaz, Fatima; Aiken, Christopher; Melikyan, Gregory B

    2015-10-29

    HIV-1 Vpr is recruited into virions during assembly and appears to remain associated with the viral core after the reverse transcription and uncoating steps of entry. This feature has prompted the use of fluorescently labeled Vpr to visualize viral particles and to follow trafficking of post-fusion HIV-1 cores in the cytoplasm. Here, we tracked single pseudovirus entry and fusion and observed that fluorescently tagged Vpr gradually dissociates from post-fusion viral cores over the course of several minutes and accumulates in the nucleus. Kinetics measurements showed that fluorescent Vpr released from the cores very rapidly entered the cell nucleus. More than 10,000 Vpr molecules can be delivered into the cell nucleus within 45 min of infection by HIV-1 particles pseudotyped with the avian sarcoma and leukosis virus envelope glycoprotein. The fraction of Vpr from cell-bound viruses that accumulated in the nucleus was proportional to the extent of virus-cell fusion and was fully blocked by viral fusion inhibitors. Entry of virus-derived Vpr into the nucleus occurred independently of envelope glycoproteins or target cells. Fluorescence correlation spectroscopy revealed two forms of nuclear Vpr-monomers and very large complexes, likely involving host factors. The kinetics of viral Vpr entering the nucleus after fusion was not affected by point mutations in the capsid protein that alter the stability of the viral core. The independence of Vpr shedding of capsid stability and its relatively rapid dissociation from post-fusion cores suggest that this process may precede capsid uncoating, which appears to occur on a slower time scale. Our results thus demonstrate that a bulk of fluorescently labeled Vpr incorporated into HIV-1 particles is released shortly after fusion. Future studies will address the question whether the quick and efficient nuclear delivery of Vpr derived from incoming viruses can regulate subsequent steps of HIV-1 infection.

  8. Minimal Contribution of APOBEC3-Induced G-to-A Hypermutation to HIV-1 Recombination and Genetic Variation.

    Science.gov (United States)

    Delviks-Frankenberry, Krista A; Nikolaitchik, Olga A; Burdick, Ryan C; Gorelick, Robert J; Keele, Brandon F; Hu, Wei-Shau; Pathak, Vinay K

    2016-05-01

    Although the predominant effect of host restriction APOBEC3 proteins on HIV-1 infection is to block viral replication, they might inadvertently increase retroviral genetic variation by inducing G-to-A hypermutation. Numerous studies have disagreed on the contribution of hypermutation to viral genetic diversity and evolution. Confounding factors contributing to the debate include the extent of lethal (stop codon) and sublethal hypermutation induced by different APOBEC3 proteins, the inability to distinguish between G-to-A mutations induced by APOBEC3 proteins and error-prone viral replication, the potential impact of hypermutation on the frequency of retroviral recombination, and the extent to which viral recombination occurs in vivo, which can reassort mutations in hypermutated genomes. Here, we determined the effects of hypermutation on the HIV-1 recombination rate and its contribution to genetic variation through recombination to generate progeny genomes containing portions of hypermutated genomes without lethal mutations. We found that hypermutation did not significantly affect the rate of recombination, and recombination between hypermutated and wild-type genomes only increased the viral mutation rate by 3.9 × 10-5 mutations/bp/replication cycle in heterozygous virions, which is similar to the HIV-1 mutation rate. Since copackaging of hypermutated and wild-type genomes occurs very rarely in vivo, recombination between hypermutated and wild-type genomes does not significantly contribute to the genetic variation of replicating HIV-1. We also analyzed previously reported hypermutated sequences from infected patients and determined that the frequency of sublethal mutagenesis for A3G and A3F is negligible (4 × 10-21 and1 × 10-11, respectively) and its contribution to viral mutations is far below mutations generated during error-prone reverse transcription. Taken together, we conclude that the contribution of APOBEC3-induced hypermutation to HIV-1 genetic

  9. SAMHD1 restricts HIV-1 replication and regulates interferon production in mouse myeloid cells.

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    Ruonan Zhang

    Full Text Available SAMHD1 restricts the replication of HIV-1 and other retroviruses in human myeloid and resting CD4(+ T cells and that is counteracted in SIV and HIV-2 by the Vpx accessory protein. The protein is a phosphohydrolase that lowers the concentration of deoxynucleoside triphosphates (dNTP, blocking reverse transcription of the viral RNA genome. Polymorphisms in the gene encoding SAMHD1 are associated with Aicardi-Goutières Syndrome, a neurological disorder characterized by increased type-I interferon production. SAMHD1 is conserved in mammals but its role in restricting virus replication and controlling interferon production in non-primate species is not well understood. We show that SAMHD1 is catalytically active and expressed at high levels in mouse spleen, lymph nodes, thymus and lung. siRNA knock-down of SAMHD1 in bone marrow-derived macrophages increased their susceptibility to HIV-1 infection. shRNA knock-down of SAMHD1 in the murine monocytic cell-line RAW264.7 increased its susceptibility to HIV-1 and murine leukemia virus and increased the levels of the dNTP pool. In addition, SAMHD1 knock-down in RAW264.7 cells induced the production of type-I interferon and several interferon-stimulated genes, modeling the situation in Aicardi-Goutières Syndrome. Our findings suggest that the role of SAMHD1 in restricting viruses is conserved in the mouse. The RAW264.7 cell-line serves as a useful tool to study the antiviral and innate immune response functions of SAMHD1.

  10. Identification of cell surface targets for HIV-1 therapeutics using genetic screens

    International Nuclear Information System (INIS)

    Dunn, Stephen J.; Khan, Imran H.; Chan, Ursula A.; Scearce, Robin L.; Melara, Claudia L.; Paul, Amber M.; Sharma, Vikram; Bih, Fong-Yih; Holzmayer, Tanya A.; Luciw, Paul A.; Abo, Arie

    2004-01-01

    Human immunodeficiency virus (HIV) drugs designed to interfere with obligatory utilization of certain host cell factors by virus are less likely to encounter development of resistant strains than drugs directed against viral components. Several cellular genes required for productive infection by HIV were identified by the use of genetic suppressor element (GSE) technology as potential targets for anti-HIV drug development. Fragmented cDNA libraries from various pools of human peripheral blood mononuclear cells (PBMC) were expressed in vitro in human immunodeficiency virus type 1 (HIV-1)-susceptible cell lines and subjected to genetic screens to identify GSEs that interfered with viral replication. After three rounds of selection, more than 15 000 GSEs were sequenced, and the cognate genes were identified. The GSEs that inhibited the virus were derived from a diverse set of genes including cell surface receptors, cytokines, signaling proteins, transcription factors, as well as genes with unknown function. Approximately 2.5% of the identified genes were previously shown to play a role in the HIV-1 life cycle; this finding supports the biological relevance of the assay. GSEs were derived from the following 12 cell surface proteins: CXCR4, CCR4, CCR7, CD11C, CD44, CD47, CD68, CD69, CD74, CSF3R, GABBR1, and TNFR2. Requirement of some of these genes for viral infection was also investigated by using RNA interference (RNAi) technology; accordingly, 10 genes were implicated in early events of the viral life cycle, before viral DNA synthesis. Thus, these cell surface proteins represent novel targets for the development of therapeutics against HIV-1 infection and AIDS

  11. Different patterns of HIV-1 DNA after therapy discontinuation

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    Ghinelli Florio

    2005-09-01

    Full Text Available Abstract Background By persisting in infected cells for a long period of time, proviral HIV-1 DNA can represent an alternative viral marker to RNA viral load during the follow-up of HIV-1 infected individuals. In the present study sequential blood samples of 10 patients under antiretroviral treatment from 1997 with two NRTIs, who refused to continue any antiviral regimen, were analyzed for 16 – 24 weeks to study the possible relationship between DNA and RNA viral load. Methods The amount of proviral DNA was quantified by SYBR green real-time PCR in peripheral blood mononuclear cells from a selected group of ten patients with different levels of plasmatic viremia (RNA viral load. Results Variable levels of proviral DNA were found without any significant correlation between proviral load and plasma HIV-1 RNA levels. Results obtained showed an increase or a rebound in viral DNA in most patients, suggesting that the absence of therapy reflects an increase and/or a persistence of cells containing viral DNA. Conclusion Even though plasma HIV RNA levels remain the basic parameter to monitor the intensity of viral replication, the results obtained seem to indicate that DNA levels could represent an adjunct prognostic marker in monitoring HIV-1 infected subjects.

  12. Stepping toward a Macaque Model of HIV-1 Induced AIDS

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    Jason T. Kimata

    2014-09-01

    Full Text Available HIV-1 exhibits a narrow host range, hindering the development of a robust animal model of pathogenesis. Past studies have demonstrated that the restricted host range of HIV-1 may be largely due to the inability of the virus to antagonize and evade effector molecules of the interferon response in other species. They have also guided the engineering of HIV-1 clones that can replicate in CD4 T-cells of Asian macaque species. However, while replication of these viruses in macaque hosts is persistent, it has been limited and without progression to AIDS. In a new study, Hatziioannou et al., demonstrate for the first time that adapted macaque-tropic HIV-1 can persistently replicate at high levels in pigtailed macaques (Macaca nemestrina, but only if CD8 T-cells are depleted at the time of inoculation. The infection causes rapid disease and recapitulates several aspects of AIDS in humans. Additionally, the virus undergoes genetic changes to further escape innate immunity in association with disease progression. Here, the importance of these findings is discussed, as they relate to pathogenesis and model development.

  13. Copy number variation of KIR genes influences HIV-1 control

    DEFF Research Database (Denmark)

    Pelak, Kimberly; Need, Anna C; Fellay, Jacques

    2011-01-01

    A genome-wide screen for large structural variants showed that a copy number variant (CNV) in the region encoding killer cell immunoglobulin-like receptors (KIR) associates with HIV-1 control as measured by plasma viral load at set point in individuals of European ancestry. This CNV encompasses t...

  14. Innate immune factors associated with HIV-1 transmission

    NARCIS (Netherlands)

    Pollakis, Georgios; Stax, Martijn J.; Paxton, William A.

    2011-01-01

    Relatively little is known with regards to the mechanisms of HIV-1 transmission across a mucosal surface and more specifically what effects host factors have on influencing infection and early viral dissemination. The purpose of this review is to summarize which factors of the innate immune response

  15. Recent evolutionary history of HIV-1 subtype B - Rebuttal

    NARCIS (Netherlands)

    Lukashov, V. V.; Goudsmit, J.

    2003-01-01

    The history of the HIV-1 B epidemic is the subject of a continuing debate. Did the epidemic start in the 1970s, as it was established based on the epidemiological data, or decades earlier, as it was suggested based on the analysis of nucleotide distances in the env gene? Our study [Lukashov and

  16. HIV-1 adaptation to NK cell-mediated immune pressure

    DEFF Research Database (Denmark)

    Elemans, Marjet; Boelen, Lies; Rasmussen, Michael

    2017-01-01

    The observation, by Alter et al., of the enrichment of NK cell “escape” variants in individuals carrying certain Killer-cell Immunoglobulin-like Receptor (KIR) genes is compelling evidence that natural killer (NK) cells exert selection pressure on HIV-1. Alter et al hypothesise that variant pepti...

  17. Incidence of HIV-1 infection and changes in prevalence of ...

    African Journals Online (AJOL)

    Sexual risk behaviours and RTIs may have contributed to HIV-1 transmission in this community. The data collected may help to inform the future design and evaluation of various intervention measures. Keywords: Africa, bacterial vaginosis, candidiasis, chlamydia, epidemiological synergy, gonorrhoea, incidence, sequelae

  18. Effects of human SAMHD1 polymorphisms on HIV-1 susceptibility

    International Nuclear Information System (INIS)

    White, Tommy E.; Brandariz-Nuñez, Alberto; Valle-Casuso, Jose Carlos; Knowlton, Caitlin; Kim, Baek; Sawyer, Sara L.; Diaz-Griffero, Felipe

    2014-01-01

    SAMHD1 is a human restriction factor that prevents efficient infection of macrophages, dendritic cells and resting CD4+ T cells by HIV-1. Here we explored the antiviral activity and biochemical properties of human SAMHD1 polymorphisms. Our studies focused on human SAMHD1 polymorphisms that were previously identified as evolving under positive selection for rapid amino acid replacement during primate speciation. The different human SAMHD1 polymorphisms were tested for their ability to block HIV-1, HIV-2 and equine infectious anemia virus (EIAV). All studied SAMHD1 variants block HIV-1, HIV-2 and EIAV infection when compared to wild type. We found that these variants did not lose their ability to oligomerize or to bind RNA. Furthermore, all tested variants were susceptible to degradation by Vpx, and localized to the nuclear compartment. We tested the ability of human SAMHD1 polymorphisms to decrease the dNTP cellular levels. In agreement, none of the different SAMHD1 variants lost their ability to reduce cellular levels of dNTPs. Finally, we found that none of the tested human SAMHD1 polymorphisms affected the ability of the protein to block LINE-1 retrotransposition. - Highlights: • Human SAMHD1 single-nucleotide polymorphisms block HIV-1 and HIV-2 infection. • SAMHD1 polymorphisms do not affect its ability to block LINE-1 retrotransposition. • SAMHD1 polymorphisms decrease the cellular levels of dNTPs

  19. HIV-1 isolation from infected peripheral blood mononuclear cells

    NARCIS (Netherlands)

    Dispinseri, Stefania; Saba, Elisa; Vicenzi, Elisa; Kootstra, Neeltje A.; Schuitemaker, Hanneke; Scarlatti, Gabriella

    2014-01-01

    Human immunodeficiency virus 1 (HIV-1) isolation from peripheral blood mononuclear cells (PBMCs) allows retrieval of replication-competent viral variants. In order to impose the smallest possible selective pressure on the viral isolates, isolation must be carried out in primary cultures of cells and

  20. The global spread of HIV-1 subtype B epidemic

    NARCIS (Netherlands)

    Magiorkinis, Gkikas; Angelis, Konstantinos; Mamais, Ioannis; Katzourakis, Aris; Hatzakis, Angelos; Albert, Jan; Lawyer, Glenn; Hamouda, Osamah; Struck, Daniel; Vercauteren, Jurgen; Wensing, Annemarie; Alexiev, Ivailo; Åsjö, Birgitta; Balotta, Claudia; Gomes, Perpétua; Camacho, Ricardo J.; Coughlan, Suzie; Griskevicius, Algirdas; Grossman, Zehava; Horban, Anders; Kostrikis, Leondios G.; Lepej, Snjezana J.; Liitsola, Kirsi; Linka, Marek; Nielsen, Claus; Otelea, Dan; Paredes, Roger; Poljak, Mario; Puchhammer-Stöckl, Elizabeth; Schmit, Jean Claude; Sönnerborg, Anders; Staneková, Danica; Stanojevic, Maja; Stylianou, Dora C.; Boucher, Charles A B; Nikolopoulos, Georgios; Vasylyeva, Tetyana; Friedman, Samuel R.; van de Vijver, David; Angarano, Gioacchino; Chaix, Marie Laure; de Luca, Andrea; Korn, Klaus; Loveday, Clive; Soriano, Vincent; Yerly, Sabine; Zazzi, Mauricio; Vandamme, Anne Mieke; Paraskevis, Dimitrios

    2016-01-01

    Human immunodeficiency virus type 1 (HIV-1) was discovered in the early 1980s when the virus had already established a pandemic. For at least three decades the epidemic in the Western World has been dominated by subtype B infections, as part of a sub-epidemic that traveled from Africa through Haiti

  1. Selection dramatically reduces effective population size in HIV-1 infection

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    Mittler John E

    2008-05-01

    Full Text Available Abstract Background In HIV-1 evolution, a 100–100,000 fold discrepancy between census size and effective population size (Ne has been noted. Although it is well known that selection can reduce Ne, high in vivo mutation and recombination rates complicate attempts to quantify the effects of selection on HIV-1 effective size. Results We use the inbreeding coefficient and the variance in allele frequency at a linked neutral locus to estimate the reduction in Ne due to selection in the presence of mutation and recombination. With biologically realistic mutation rates, the reduction in Ne due to selection is determined by the strength of selection, i.e., the stronger the selection, the greater the reduction. However, the dependence of Ne on selection can break down if recombination rates are very high (e.g., r ≥ 0.1. With biologically likely recombination rates, our model suggests that recurrent selective sweeps similar to those observed in vivo can reduce within-host HIV-1 effective population sizes by a factor of 300 or more. Conclusion Although other factors, such as unequal viral reproduction rates and limited migration between tissue compartments contribute to reductions in Ne, our model suggests that recurrent selection plays a significant role in reducing HIV-1 effective population sizes in vivo.

  2. Defective proviruses rapidly accumulate during acute HIV-1 infection

    Science.gov (United States)

    Bruner, Katherine M.; Murray, Alexandra J.; Pollack, Ross A.; Soliman, Mary G.; Laskey, Sarah B.; Capoferri, Adam A.; Lai, Jun; Strain, Matthew C.; Lada, Steven M.; Hoh, Rebecca; Ho, Ya-Chi; Richman, Douglas D.; Deeks, Steven G.; Siliciano, Janet D.; Siliciano, Robert F.

    2016-01-01

    Although antiretroviral therapy (ART) suppresses viral replication to clinically undetectable levels, HIV-1 persists in CD4+ T cells in a latent form not targeted by the immune system or ART1–5. This latent reservoir is a major barrier to cure. Many individuals initiate ART during chronic infection, and in this setting, most proviruses are defective6. However, the dynamics of the accumulation and persistence of defective proviruses during acute HIV-1 infection are largely unknown. Here we show that defective proviruses accumulate rapidly within the first few weeks of infection to make up over 93% of all proviruses, regardless of how early ART is initiated. Using an unbiased method to amplify near full-length proviral genomes from HIV-1 infected adults treated at different stages of infection, we demonstrate that early ART initiation limits the size of the reservoir but does not profoundly impact the proviral landscape. This analysis allows us to revise our understanding of the composition of proviral populations and estimate the true reservoir size in individuals treated early vs. late in infection. Additionally, we demonstrate that common assays for measuring the reservoir do not correlate with reservoir size. These findings reveal hurdles that must be overcome to successfully analyze future HIV-1 cure strategies. PMID:27500724

  3. Effects of human SAMHD1 polymorphisms on HIV-1 susceptibility

    Energy Technology Data Exchange (ETDEWEB)

    White, Tommy E.; Brandariz-Nuñez, Alberto; Valle-Casuso, Jose Carlos [Department of Microbiology and Immunology, Albert Einstein College of Medicine, Bronx, 1301 Morris Park – Price Center 501, New York, NY 10461 (United States); Knowlton, Caitlin; Kim, Baek [Department of Microbiology and Immunology, University of Rochester School of Medicine and Dentistry, Rochester, NY 14642 (United States); Sawyer, Sara L. [Department of Molecular Biosciences, University of Texas at Austin, Austin, TX 78712 (United States); Diaz-Griffero, Felipe, E-mail: Felipe.Diaz-Griffero@einstein.yu.edu [Department of Microbiology and Immunology, Albert Einstein College of Medicine, Bronx, 1301 Morris Park – Price Center 501, New York, NY 10461 (United States)

    2014-07-15

    SAMHD1 is a human restriction factor that prevents efficient infection of macrophages, dendritic cells and resting CD4+ T cells by HIV-1. Here we explored the antiviral activity and biochemical properties of human SAMHD1 polymorphisms. Our studies focused on human SAMHD1 polymorphisms that were previously identified as evolving under positive selection for rapid amino acid replacement during primate speciation. The different human SAMHD1 polymorphisms were tested for their ability to block HIV-1, HIV-2 and equine infectious anemia virus (EIAV). All studied SAMHD1 variants block HIV-1, HIV-2 and EIAV infection when compared to wild type. We found that these variants did not lose their ability to oligomerize or to bind RNA. Furthermore, all tested variants were susceptible to degradation by Vpx, and localized to the nuclear compartment. We tested the ability of human SAMHD1 polymorphisms to decrease the dNTP cellular levels. In agreement, none of the different SAMHD1 variants lost their ability to reduce cellular levels of dNTPs. Finally, we found that none of the tested human SAMHD1 polymorphisms affected the ability of the protein to block LINE-1 retrotransposition. - Highlights: • Human SAMHD1 single-nucleotide polymorphisms block HIV-1 and HIV-2 infection. • SAMHD1 polymorphisms do not affect its ability to block LINE-1 retrotransposition. • SAMHD1 polymorphisms decrease the cellular levels of dNTPs.

  4. Analysis of dinucleotide signatures in HIV-1 subtype B genomes

    Indian Academy of Sciences (India)

    It was also shown that the profile generated by taking all dinucleotides together ... Keywords. genome signature; DRAP; HIV-1; chaos game representation. Journal of .... be used to quantify low levels of variation as are observed within species ..... Dayton A.I., Sodroski J.G., Rosen C.A., Goh W.C. and Haseltine. W.A. 1986 ...

  5. Distribution of HIV-1 resistance-conferring polymorphic alleles SDF ...

    Indian Academy of Sciences (India)

    Polymorphic allelic variants of chemokine receptors CCR2 and CCR5, as well as of stromal-derived factor-1 SDF-1, the ligand for the chemokine receptor CXCR4, are known to have protective effects against HIV-1 infection and to be involved with delay in disease progression. We have studied the DNA polymorphisms at ...

  6. Alemtuzumab-induced elimination of HIV-1-infected immune cells.

    Science.gov (United States)

    Ruxrungtham, Kiat; Sirivichayakul, Sunee; Buranapraditkun, Supranee; Krause, Werner

    2016-01-01

    Currently, there is no drug known that is able to eradicate either HIV or HIV-infected host cells. The effectiveness of all available treatments is based on the prevention of viral replication. We investigated whether the monoclonal, CD52 receptor-targeting antibody, alemtuzumab, which is currently approved for the treatment of multiple sclerosis, is able to eliminate HIV-infected immune cells. In blood samples from healthy donors and from HIV-1-infected subjects who were either treatment-naïve or resistant to HAART, we studied whether the CD52 expression on T cells and their subsets (CD3, CD4, CD8), B cells (CD19), dendritic cells (CD123) and monocytes (CD11c) is retained in HIV-1 infection and whether alemtuzumab is able to eradicate infected cells, using four-colour flow cytometry. We found that CD52 expression on immune cells is retained in HIV-1 infection regardless of CD4 cell count, viral load and treatment status, and is amenable to alemtuzumab-induced depletion. For the first time it could be shown in vitro that HIV-1-infected immune cells can be eliminated by using the monoclonal antibody alemtuzumab.

  7. The global spread of HIV-1 subtype B epidemic

    NARCIS (Netherlands)

    G. Magiorkinis (Gkikas); K. Angelis (Konstantinos); I. Mamais (Ioannis); Katzourakis, A. (Aris); A. Hatzakis (Angelos); J. Albert (Jan); Lawyer, G. (Glenn); O. Hamouda (Osamah); D. Struck (Daniel); J. Vercauteren (Jurgen); A. Wensing (Amj); I. Alexiev (Ivailo); B. Åsjö (Birgitta); C. Balotta (Claudia); Gomes, P. (Perpétua); R.J. Camacho (Ricardo Jorge); S. Coughlan (Suzie); A. Griskevicius (Algirdas); Z. Grossman (Zehava); Horban, A. (Anders); L.G. Kostrikis (Leondios); Lepej, S.J. (Snjezana J.); K. Liitsola (Kirsi); M. Linka (Marek); C. Nielsen; D. Otelea (Dan); R. Paredes (Roger); M. Poljak (Mario); E. Puchhammer-Stöckl (Elisabeth); J.C. Schmit; A. Sonnerborg (Anders); D. Stanekova (Danica); M. Stanojevic (Maja); Stylianou, D.C. (Dora C.); C.A.B. Boucher (Charles); Nikolopoulos, G. (Georgios); Vasylyeva, T. (Tetyana); Friedman, S.R. (Samuel R.); D.A.M.C. van de Vijver (David); G. Angarano (Guiseppe); M.L. Chaix (Marie Laure); A. de Luca (Andrea); K. Korn (Klaus); Loveday, C. (Clive); V. Soriano (Virtudes); S. Yerly (Sabine); M. Zazzi; A.M. Vandamme (Anne Mieke); D. Paraskevis (Dimitrios)

    2016-01-01

    textabstractHuman immunodeficiency virus type 1 (HIV-1) was discovered in the early 1980s when the virus had already established a pandemic. For at least three decades the epidemic in the Western World has been dominated by subtype B infections, as part of a sub-epidemic that traveled from Africa

  8. Molecular epidemiology of HIV-1 infection in Europe: An overview.

    Science.gov (United States)

    Beloukas, Apostolos; Psarris, Alexandros; Giannelou, Polina; Kostaki, Evangelia; Hatzakis, Angelos; Paraskevis, Dimitrios

    2016-12-01

    Human Immunodeficiency Virus type 1 (HIV-1) is characterised by vast genetic diversity. Globally circulating HIV-1 viruses are classified into distinct phylogenetic strains (subtypes, sub-subtypes) and several recombinant forms. Here we describe the characteristics and evolution of European HIV-1 epidemic over time through a review of published literature and updated queries of existing HIV-1 sequence databases. HIV-1 in Western and Central Europe was introduced in the early-1980s in the form of subtype B, which is still the predominant clade. However, in Eastern Europe (Former Soviet Union (FSU) countries and Russia) the predominant strain, introduced into Ukraine in the mid-1990s, is subtype A (A FSU ) with transmission mostly occurring in People Who Inject Drugs (PWID). In recent years, the epidemic is evolving towards a complex tapestry with an increase in the prevalence of non-B subtypes and recombinants in Western and Central Europe. Non-B epidemics are mainly associated with immigrants, heterosexuals and females but more recently, non-B clades have also spread amongst groups where non-B strains were previously absent - non-immigrant European populations and amongst men having sex with men (MSM). In some countries, non-B clades have spread amongst the native population, for example subtype G in Portugal and subtype A in Greece, Albania and Cyprus. Romania provides a unique case where sub-subtype F1 has predominated throughout the epidemic. In contrast, HIV-1 epidemic in FSU countries remains more homogeneous with A FSU clade predominating in all countries. The differences between the evolution of the Western epidemic and the Eastern epidemic may be attributable to differences in transmission risk behaviours, lifestyle and the patterns of human mobility. The study of HIV-1 epidemic diversity provides a useful tool by which we can understand the history of the pandemic in addition to allowing us to monitor the spread and growth of the epidemic over time

  9. Similarities and differences in the nucleic acid chaperone activity of HIV-2 and HIV-1 nucleocapsid proteins in vitro.

    Science.gov (United States)

    Pachulska-Wieczorek, Katarzyna; Stefaniak, Agnieszka K; Purzycka, Katarzyna J

    2014-07-03

    The nucleocapsid domain of Gag and mature nucleocapsid protein (NC) act as nucleic acid chaperones and facilitate folding of nucleic acids at critical steps of retroviral replication cycle. The basic N-terminus of HIV-1 NC protein was shown most important for the chaperone activity. The HIV-2 NC (NCp8) and HIV-1 NC (NCp7) proteins possess two highly conserved zinc fingers, flanked by basic residues. However, the NCp8 N-terminal domain is significantly shorter and contains less positively charged residues. This study characterizes previously unknown, nucleic acid chaperone activity of the HIV-2 NC protein. We have comparatively investigated the in vitro nucleic acid chaperone properties of the HIV-2 and HIV-1 NC proteins. Using substrates derived from the HIV-1 and HIV-2 genomes, we determined the ability of both proteins to chaperone nucleic acid aggregation, annealing and strand exchange in duplex structures. Both NC proteins displayed comparable, high annealing activity of HIV-1 TAR DNA and its complementary nucleic acid. Interesting differences between the two NC proteins were discovered when longer HIV substrates, particularly those derived from the HIV-2 genome, were used in chaperone assays. In contrast to NCp7, NCp8 weakly facilitates annealing of HIV-2 TAR RNA to its complementary TAR (-) DNA. NCp8 is also unable to efficiently stimulate tRNALys3 annealing to its respective HIV-2 PBS motif. Using truncated NCp8 peptide, we demonstrated that despite the fact that the N-terminus of NCp8 differs from that of NCp7, this domain is essential for NCp8 activity. Our data demonstrate that the HIV-2 NC protein displays reduced nucleic acid chaperone activity compared to that of HIV-1 NC. We found that NCp8 activity is limited by substrate length and stability to a greater degree than that of NCp7. This is especially interesting in light of the fact that the HIV-2 5'UTR is more structured than that of HIV-1. The reduced chaperone activity observed with NCp8 may

  10. Effects of As2O3 on DNA methylation, genomic instability, and LTR retrotransposon polymorphism in Zea mays.

    Science.gov (United States)

    Erturk, Filiz Aygun; Aydin, Murat; Sigmaz, Burcu; Taspinar, M Sinan; Arslan, Esra; Agar, Guleray; Yagci, Semra

    2015-12-01

    Arsenic is a well-known toxic substance on the living organisms. However, limited efforts have been made to study its DNA methylation, genomic instability, and long terminal repeat (LTR) retrotransposon polymorphism causing properties in different crops. In the present study, effects of As2O3 (arsenic trioxide) on LTR retrotransposon polymorphism and DNA methylation as well as DNA damage in Zea mays seedlings were investigated. The results showed that all of arsenic doses caused a decreasing genomic template stability (GTS) and an increasing Random Amplified Polymorphic DNAs (RAPDs) profile changes (DNA damage). In addition, increasing DNA methylation and LTR retrotransposon polymorphism characterized a model to explain the epigenetically changes in the gene expression were also found. The results of this experiment have clearly shown that arsenic has epigenetic effect as well as its genotoxic effect. Especially, the increasing of polymorphism of some LTR retrotransposon under arsenic stress may be a part of the defense system against the stress.

  11. Enhancement of Intranasal Vaccination in Mice with Deglycosylated Chain A Ricin by LTR72, a Novel Mucosal Adjuvant

    National Research Council Canada - National Science Library

    Kende, Meir; Del Giudice, Giuseppe; Rivera, Noelia; Hewetson, John

    2006-01-01

    .... However, in the presence of 4, 2, or 1 microg of the mucosal adjuvant LTR72, a mutant of the heat-labile enterotoxin of Escherichia coli, the low antibody response and protection were substantially enhanced...

  12. Enhancement of Intranasal Vaccination in Mice with Deglycosylated Chain A Ricin by LTR72, a Novel Mucosal Adjuvant

    National Research Council Canada - National Science Library

    Kende, Meir; Del Giudice, Giuseppe; Rivera, Noelia; Hewetson, John

    2006-01-01

    .... However, in the presence of 4, 2, or 1 micro-gram of the mucosal adjuvant LTR72, a mutant of the heat-labile enterotoxin of Escherichia coli, the low antibody response and protection were substantially enhanced...

  13. Dolutegravir reshapes the genetic diversity of HIV-1 reservoirs.

    Science.gov (United States)

    Gantner, Pierre; Lee, Guinevere Q; Rey, David; Mesplede, Thibault; Partisani, Marialuisa; Cheneau, Christine; Beck-Wirth, Geneviève; Faller, Jean-Pierre; Mohseni-Zadeh, Mahsa; Martinot, Martin; Wainberg, Mark A; Fafi-Kremer, Samira

    2018-04-01

    Better understanding of the dynamics of HIV reservoirs under ART is a critical step to achieve a functional HIV cure. Our objective was to assess the genetic diversity of archived HIV-1 DNA over 48 weeks in blood cells of individuals starting treatment with a dolutegravir-based regimen. Eighty blood samples were prospectively and longitudinally collected from 20 individuals (NCT02557997) including: acutely (n = 5) and chronically (n = 5) infected treatment-naive individuals, as well as treatment-experienced individuals who switched to a dolutegravir-based regimen and were either virologically suppressed (n = 5) or had experienced treatment failure (n = 5). The integrase and V3 loop regions of HIV-1 DNA isolated from PBMCs were analysed by pyrosequencing at baseline and weeks 4, 24 and 48. HIV-1 genetic diversity was calculated using Shannon entropy. All individuals achieved or maintained viral suppression throughout the study. A low and stable genetic diversity of archived HIV quasispecies was observed in individuals starting treatment during acute infection. A dramatic reduction of the genetic diversity was observed at week 4 of treatment in the other individuals. In these patients and despite virological suppression, a recovery of the genetic diversity of the reservoirs was observed up to 48 weeks. Viral variants bearing dolutegravir resistance-associated substitutions at integrase position 50, 124, 230 or 263 were detected in five individuals (n = 5/20, 25%) from all groups except those who were ART-failing at baseline. None of these substitutions led to virological failure. These data demonstrate that the genetic diversity of the HIV-1 reservoir is reshaped following the initiation of a dolutegravir-based regimen and strongly suggest that HIV-1 can continue to replicate despite successful treatment.

  14. Prevalence of genotypic HIV-1 drug resistance in Thailand, 2002

    Directory of Open Access Journals (Sweden)

    Watitpun Chotip

    2003-03-01

    Full Text Available Abstract Background The prices of reverse transcriptase (RT inhibitors in Thailand have been reduced since December 1, 2001. It is expected that reduction in the price of these inhibitors may influence the drug resistance mutation pattern of HIV-1 among infected people. This study reports the frequency of HIV-1 genetic mutation associated with drug resistance in antiretroviral-treated patients from Thailand. Methods Genotypic resistance testing was performed on samples collected in 2002 from 88 HIV-1 infected individuals. Automated DNA sequencing was used to genotype the HIV-1 polymerase gene isolated from patients' plasma. Results Resistance to protease inhibitors, nucleoside and non-nucleoside reverse transcriptase inhibitors were found in 10 (12%, 42 (48% and 19 (21% patients, respectively. The most common drug resistance mutations in the protease gene were at codon 82 (8%, 90 (7% and 54 (6%, whereas resistant mutations at codon 215 (45%, 67 (40%, 41 (38% and 184 (27% were commonly found in the RT gene. This finding indicates that genotypic resistance to nucleoside reverse transcriptase inhibitors was prevalent in 2002. The frequency of resistant mutations corresponding to non-nucleoside reverse transcriptase inhibitors was three times higher-, while resistant mutation corresponding to protease inhibitors was two times lower than those frequencies determined in 2001. Conclusion This study shows that the frequencies of RT inhibitor resistance mutations have been increased after the reduction in the price of RT inhibitors since December 2001. We believe that this was an important factor that influenced the mutation patterns of HIV-1 protease and RT genes in Thailand.

  15. Characteristics of HIV-1 discordant couples enrolled in a trial of HSV-2 suppression to reduce HIV-1 transmission: the partners study.

    Directory of Open Access Journals (Sweden)

    Jairam R Lingappa

    Full Text Available The Partners HSV-2/HIV-1 Transmission Study (Partners Study is a phase III, placebo-controlled trial of daily acyclovir for genital herpes (HSV-2 suppression among HIV-1/HSV-2 co-infected persons to reduce HIV-1 transmission to their HIV-1 susceptible partners, which requires recruitment of HIV-1 serodiscordant heterosexual couples. We describe the baseline characteristics of this cohort.HIV-1 serodiscordant heterosexual couples, in which the HIV-1 infected partner was HSV-2 seropositive, had a CD4 count >or=250 cells/mcL and was not on antiretroviral therapy, were enrolled at 14 sites in East and Southern Africa. Demographic, behavioral, clinical and laboratory characteristics were assessed.Of the 3408 HIV-1 serodiscordant couples enrolled, 67% of the HIV-1 infected partners were women. Couples had cohabitated for a median of 5 years (range 2-9 with 28% reporting unprotected sex in the month prior to enrollment. Among HIV-1 susceptible participants, 86% of women and 59% of men were HSV-2 seropositive. Other laboratory-diagnosed sexually transmitted infections were uncommon (500 relative to <350, respectively, p<0.001.The Partners Study successfully enrolled a cohort of 3408 heterosexual HIV-1 serodiscordant couples in Africa at high risk for HIV-1 transmission. Follow-up of this cohort will evaluate the efficacy of acyclovir for HSV-2 suppression in preventing HIV-1 transmission and provide insights into biological and behavioral factors determining heterosexual HIV-1 transmission.ClinicalTrials.gov NCT00194519.

  16. The role of Vif oligomerization and RNA chaperone activity in HIV-1 replication.

    Science.gov (United States)

    Batisse, Julien; Guerrero, Santiago; Bernacchi, Serena; Sleiman, Dona; Gabus, Caroline; Darlix, Jean-Luc; Marquet, Roland; Tisné, Carine; Paillart, Jean-Christophe

    2012-11-01

    The viral infectivity factor (Vif) is essential for the productive infection and dissemination of HIV-1 in non-permissive cells that involve most natural HIV-1 target cells. Vif counteracts the packaging of two cellular cytidine deaminases named APOBEC3G (A3G) and A3F by diverse mechanisms including the recruitment of an E3 ubiquitin ligase complex and the proteasomal degradation of A3G/A3F, the inhibition of A3G mRNA translation or by a direct competition mechanism. In addition, Vif appears to be an active partner of the late steps of viral replication by participating in virus assembly and Gag processing, thus regulating the final stage of virion formation notably genomic RNA dimerization and by inhibiting the initiation of reverse transcription. Vif is a small pleiotropic protein with multiple domains, and recent studies highlighted the importance of Vif conformation and flexibility in counteracting A3G and in binding RNA. In this review, we will focus on the oligomerization and RNA chaperone properties of Vif and show that the intrinsic disordered nature of some Vif domains could play an important role in virus assembly and replication. Experimental evidence demonstrating the RNA chaperone activity of Vif will be presented. Copyright © 2012 Elsevier B.V. All rights reserved.

  17. Formation of stable and functional HIV-1 nucleoprotein complexes in vitro.

    Science.gov (United States)

    Tanchou, V; Gabus, C; Rogemond, V; Darlix, J L

    1995-10-06

    HIV genomic RNA resides within the nucleocapsid, in the interior of the virus, which serves to protect the RNA against nuclease degradation and to promote its reverse transcription. To investigate the role of nucleocapsid protein (NCp7) in the stability and replication of genomic RNA within the nucleocapsid, we used NCp7, reverse transcriptase (RT) and RNAs representing the 5' and 3' regions of the genome to reconstitute functional HIV-1 nucleocapsids. The nucleoprotein complexes generated in vitro were found to be stable, which, according to biochemical and genetic data, probably results from the tight binding of NCp7 molecules to the RNA and strong NCp7/NCp7 interactions. The nucleoprotein complexes efficiently protected viral RNA against RNase degradation and, at the same time, promoted viral DNA synthesis by RT. DNA strand transfer from the 5' to the 3' RNA template was very efficient in nucleoprotein complexes formed in the presence of both RNAs, but not when the RNAs were in separate complexes. These results indicate that the in vitro reconstituted HIV-1 nucleoprotein complexes function like virion nucleocapsids and thus provide a way to study at the molecular level this viral substructure and the synthesis of proviral DNA, and to search for new anti-HIV agents.

  18. A HIV-1 heterosexual transmission chain in Guangzhou, China: a molecular epidemiological study.

    Science.gov (United States)

    Han, Zhigang; Leung, Tommy W C; Zhao, Jinkou; Wang, Ming; Fan, Lirui; Li, Kai; Pang, Xinli; Liang, Zhenbo; Lim, Wilina W L; Xu, Huifang

    2009-09-25

    We conducted molecular analyses to confirm four clustering HIV-1 infections (Patient A, B, C & D) in Guangzhou, China. These cases were identified by epidemiological investigation and suspected to acquire the infection through a common heterosexual transmission chain. Env C2V3V4 region, gag p17/p24 junction and partial pol gene of HIV-1 genome from serum specimens of these infected cases were amplified by reverse transcription polymerase chain reaction (RT-PCR) and nucleotide sequenced. Phylogenetic analyses indicated that their viral nucleotide sequences were significantly clustered together (bootstrap value is 99%, 98% and 100% in env, gag and pol tree respectively). Evolutionary distance analysis indicated that their genetic diversities of env, gag and pol genes were significantly lower than non-clustered controls, as measured by unpaired t-test (env gene comparison: p Epidemiological results and molecular analyses consistently illustrated these four cases represented a transmission chain which dispersed in the locality through heterosexual contact involving commercial sex worker.

  19. Structure of long terminal repeats of transcriptionally active and inactive copies of Drosophila mobile dispersed genetic elements mdg3

    International Nuclear Information System (INIS)

    Dzhumagaliev, E.B.; Mazo, A.N.; Baev, A.A. Jr.; Gorelova, T.V.; Arkhipova, I.R.; Shuppe, N.G.; Il'in, Yu.V.

    1986-01-01

    The authors have determined the nucleotide sequences of long terminal repeats (LTRS) and adjacent regions in the transcribed and nontranscribed variants of the mobile dispersed gene mdg3. In its main characteristics the mdg3 is similar to other mdg. Its integration into chromosomal DNA brings about duplication of the 4 bp of the host DNA, no specificity of the mdg integration at the nucleotide level being detected. The mdg3 is flanked by a 5 bp inverted repeat. The variations in the length of the LTR in different mdg copies is mainly due to duplication of certain sequences in the U3 and R regions. mdg3 copies with a LTR length of 267 bp are the most abundant and are completely conservative in their primary structure. They are transcribed in the cells of the 67J25D culture, but not transcribed in the K/sub c/ line, where another mdg3 variant with a LTR length of 293 bp is transcriptionally active. The SI mapping of transcription initiation and termination sites has shown that in both mdg3 variants they are localized in the same LTR regions, and that the LTR itself has a characteristic U3-R-U5 structure-like retroviral LTRs. The possible factors involved in the regulation of mdg transcription are discussed

  20. German-austrian recommendations for HIV1-therapy in pregnancy and in HIV1-exposed newborn - update 2008

    Directory of Open Access Journals (Sweden)

    Buchholz Bernd

    2009-11-01

    Full Text Available Abstract German-Austrian recommendations for HIV1-therapy in pregnancy - Update 2008 Bernd Buchholz (University Medical Centre Mannheim, Pediatric Clinic, Matthias Beichert (Mannheim, Gynecology and Obstetrics Practice, Ulrich Marcus (Robert Koch Institute, Berlin, Thomas Grubert, Andrea Gingelmaier (Gynecology Clinic of the Ludwig Maximilians University of Munich, Dr. med. Annette Haberl (HIV-Department, J. W. Goethe-University Hospital, Frankfurt, Dr. med. Brigitte Schmied (Otto-Wagner Spital, Wien. In Germany during the last years about 200-250 HIV1-infected pregnant women delivered a baby each year, a number that is currently increasing. To determine the HIV-status early in pregnancy voluntary HIV-testing of all pregnant women is recommended in Germany and Austria as part of prenatal care. In those cases, where HIV1-infection was known during pregnancy, since 1995 the rate of vertical transmission of HIV1 was reduced to 1-2%. This low transmission rate has been achieved by the combination of anti-retroviral therapy of pregnant women, caesarean section scheduled before onset of labour, anti-retroviral post exposition prophylaxis in the newborn and refraining from breast-feeding by the HIV1-infected mother. To keep pace with new results in research, approval of new anti-retroviral drugs and changes in the general treatment recommendations for HIV1-infected adults, in 1998, 2001, 2003 and 2005 an interdisciplinary consensus meeting was held. Gynaecologists, infectious disease specialists, paediatricians, pharmacologists, virologists and members of the German AIDS Hilfe (NGO were participating in this conference to update the prevention strategies. A fifth update became necessary in 2008. The updating process was started in January 2008 and was terminated in September 2008. The guidelines provide new recommendations on the indication and the starting point for HIV-therapy in pregnancies without complications, drugs and drug combinations to be

  1. Reduced expression of glutamate transporter EAAT2 and impaired glutamate transport in human primary astrocytes exposed to HIV-1 or gp120

    International Nuclear Information System (INIS)

    Wang Zhuying; Pekarskaya, Olga; Bencheikh, Meryem; Chao Wei; Gelbard, Harris A.; Ghorpade, Anuja; Rothstein, Jeffrey D.; Volsky, David J.

    2003-01-01

    L-Glutamate is the major excitatory neurotransmitter in the brain. Astrocytes maintain low levels of synaptic glutamate by high-affinity uptake and defects in this function may lead to neuronal cell death by excitotoxicity. We tested the effects of HIV-1 and its envelope glycoprotein gp120 upon glutamate uptake and expression of glutamate transporters EAAT1 and EAAT2 in fetal human astrocytes in vitro. Astrocytes isolated from fetal tissues between 16 and 19 weeks of gestation expressed EAAT1 and EAAT2 RNA and proteins as detected by Northern blot analysis and immunoblotting, respectively, and the cells were capable of specific glutamate uptake. Exposure of astrocytes to HIV-1 or gp120 significantly impaired glutamate uptake by the cells, with maximum inhibition within 6 h, followed by gradual decline during 3 days of observation. HIV-1-infected cells showed a 59% reduction in V max for glutamate transport, indicating a reduction in the number of active transporter sites on the cell surface. Impaired glutamate transport after HIV-1 infection or gp120 exposure correlated with a 40-70% decline in steady-state levels of EAAT2 RNA and protein. EAAT1 RNA and protein levels were less affected. Treatment of astrocytes with tumor necrosis factor-α (TNF-α) decreased the expression of both EAAT1 and EAAT2, but neither HIV-1 nor gp120 were found to induce TNF-α production by astrocytes. These findings demonstrate that HIV-1 and gp120 induce transcriptional downmodulation of the EAAT2 transporter gene in human astrocytes and coordinately attenuate glutamate transport by the cells. Reduction of the ability of HIV-1-infected astrocytes to take up glutamate may contribute to the development of neurological disease

  2. Maturation Pathways of Cross-Reactive HIV-1 Neutralizing Antibodies

    Directory of Open Access Journals (Sweden)

    Dimiter S. Dimitrov

    2009-11-01

    Full Text Available Several human monoclonal antibodies (hmAbs and antibody fragments, including the best characterized in terms of structure-function b12 and Fab X5, exhibit relatively potent and broad HIV-1 neutralizing activity. However, the elicitation of b12 or b12-like antibodies in vivo by vaccine immunogens based on the HIV-1 envelope glycoprotein (Env has not been successful. B12 is highly divergent from the closest corresponding germline antibody while X5 is less divergent. We have hypothesized that the relatively high degree of specific somatic hypermutations may preclude binding of the HIV-1 envelope glycoprotein (Env to closest germline antibodies, and that identifying antibodies that are intermediates in the pathways to maturation could help design novel vaccine immunogens to guide the immune system for their enhanced elicitation. In support of this hypothesis we have previously found that a germline-like b12 (monovalent and bivalent scFv as an Fc fusion protein or IgG lacks measurable binding to an Env as measured by ELISA with a sensitivity in the μM range [1]; here we present evidence confirming and expanding these findings for a panel of Envs. In contrast, a germline-like scFv X5 bound Env with high (nM affinity. To begin to explore the maturation pathways of these antibodies we identified several possible b12 intermediate antibodies and tested their neutralizing activity. These intermediate antibodies neutralized only some HIV-1 isolates and with relatively weak potency. In contrast, germline-like scFv X5 neutralized a subset of the tested HIV-1 isolates with comparable efficiencies to that of the mature X5. These results could help explain the relatively high immunogenicity of the coreceptor binding site on gp120 and the abundance of CD4-induced (CD4i antibodies in HIV-1-infected patients (X5 is a CD4i antibody as well as the maturation pathway of X5. They also can help identify antigens that can bind specifically to b12 germline and

  3. Contrasting roles for TLR ligands in HIV-1 pathogenesis.

    Directory of Open Access Journals (Sweden)

    Beda Brichacek

    2010-09-01

    Full Text Available The first line of a host's response to various pathogens is triggered by their engagement of cellular pattern recognition receptors (PRRs. Binding of microbial ligands to these receptors leads to the induction of a variety of cellular factors that alter intracellular and extracellular environment and interfere directly or indirectly with the life cycle of the triggering pathogen. Such changes may also affect any coinfecting microbe. Using ligands to Toll-like receptors (TLRs 5 and 9, we examined their effect on human immunodeficiency virus (HIV-1 replication in lymphoid tissue ex vivo. We found marked differences in the outcomes of such treatment. While flagellin (TLR5 agonist treatment enhanced replication of CC chemokine receptor 5 (CCR 5-tropic and CXC chemokine receptor 4 (CXCR4-tropic HIV-1, treatment with oligodeoxynucleotide (ODN M362 (TLR9 agonist suppressed both viral variants. The differential effects of these TLR ligands on HIV-1 replication correlated with changes in production of CC chemokines CCL3, CCL4, CCL5, and of CXC chemokines CXCL10, and CXCL12 in the ligand-treated HIV-1-infected tissues. The nature and/or magnitude of these changes were dependent on the ligand as well as on the HIV-1 viral strain. Moreover, the tested ligands differed in their ability to induce cellular activation as evaluated by the expression of the cluster of differentiation markers (CD 25, CD38, CD39, CD69, CD154, and human leukocyte antigen D related (HLA-DR as well as of a cell proliferation marker, Ki67, and of CCR5. No significant effect of the ligand treatment was observed on apoptosis and cell death/loss in the treated lymphoid tissue ex vivo. Our results suggest that binding of microbial ligands to TLRs is one of the mechanisms that mediate interactions between coinfected microbes and HIV-1 in human tissues. Thus, the engagement of appropriate TLRs by microbial molecules or their mimetic might become a new strategy for HIV therapy or prevention.

  4. HIV-1 pre-mRNA commitment to Rev mediated export through PSF and Matrin 3

    International Nuclear Information System (INIS)

    Kula, Anna; Gharu, Lavina; Marcello, Alessandro

    2013-01-01

    Human immunodeficiency virus gene expression and replication are regulated at several levels. Incompletely spliced viral RNAs and full-length genomic RNA contain the RRE element and are bound by the viral trans-acting protein Rev to be transported out of the nucleus. Previously we found that the nuclear matrix protein MATR3 was a cofactor of Rev-mediated RNA export. Here we show that the pleiotropic protein PSF binds viral RNA and is associated with MATR3. PSF is involved in the maintenance of a pool of RNA available for Rev activity. However, while Rev and PSF bind the viral pre-mRNA at the site of viral transcription, MATR3 interacts at a subsequent step. We propose that PSF and MATR3 define a novel pathway for RRE-containing HIV-1 RNAs that is hijacked by the viral Rev protein.

  5. HIV-1 resistance dynamics in patients failing dolutegravir maintenance monotherapy.

    Science.gov (United States)

    Wijting, Ingeborg E A; Lungu, Cynthia; Rijnders, Bart J A; van der Ende, Marchina E; Pham, Hanh T; Mesplede, Thibault; Pas, Suzan D; Voermans, Jolanda J C; Schuurman, Rob; van de Vijver, David A M C; Boers, Patrick H M; Gruters, Rob A; Boucher, Charles A B; van Kampen, Jeroen J A

    2018-03-29

    A high genetic resistance barrier to the integrase-strand-transfer-inhibitor (INSTI) dolutegravir has been reported in vitro and in vivo. We describe the dynamics of INSTI-resistance-associated-mutations (INSTI-RAMs) and mutations in the 3'-polypurine tract (3'-PPT) in relation to virological failure (VF) observed in the randomized dolutegravir maintenance monotherapy study (DOMONO, NCT02401828). From ten patients with VF plasma samples prior to start cART and during VF were used to generate Sanger sequences of integrase, the 5' terminal bases of the 3'- LTR, and the 3'-PPT. Median HIV-RNA (IQR) at VF was 3,490 (1,440-4,990) c/mL. INSTI-RAMs were detected in 4/10 patients (S230R, R263K, N155H, E92Q+N155H) and in 4/10 patients no INSTI-RAMs were detected (2/10 patients integrase sequencing was unsuccessful). The time-to-VF ranged from 4 weeks to 72 weeks. In one patient, mutations developed in the highly conserved 3'-PPT. No changes in the terminal bases of the 3'-LTR were observed. The genetic barrier to resistance is too low to justify dolutegravir maintenance monotherapy as single INSTI-RAMs are sufficient to cause VF. The large variation in time-to-VF suggests that stochastic reactivation of a pre-existing provirus containing a single INSTI-RAM is the mechanism for failure. Changes in the 3'-PPT point to a new dolutegravir resistance mechanism in vivo.

  6. Laser irradiation reduces HIV-1 infection in TZM-bl cells

    CSIR Research Space (South Africa)

    Lugongolo, Masixole Y

    2016-10-01

    Full Text Available HIV-1 epidemic remains a major health challenge. This study explores the effects of low level laser therapy on HIV-1 infected cells. Infection is reduced by irradiation and the mechanism needs to be investigated further....

  7. HIV-1 isolation from infected peripheral blood mononuclear cells.

    Science.gov (United States)

    Dispinseri, Stefania; Saba, Elisa; Vicenzi, Elisa; Kootstra, Neeltje A; Schuitemaker, Hanneke; Scarlatti, Gabriella

    2014-01-01

    Human immunodeficiency virus 1 (HIV-1) isolation from peripheral blood mononuclear cells (PBMCs) allows retrieval of replication-competent viral variants. In order to impose the smallest possible selective pressure on the viral isolates, isolation must be carried out in primary cultures of cells and not in tumor derived cell lines. The procedure involves culture of PBMCs from an infected patient with phytohemagglutinin (PHA)-stimulated PBMC from seronegative donors, which provide susceptible target cells for HIV replication. HIV can be isolated from the bulk population of PBMCs or after cloning of the cells to obtain viral biological clones. Viral production is determined with p24 antigen (Ag) detection assays or with reverse transcriptase (RT) activity assay. Once isolated, HIV-1 can be propagated by infecting PHA-stimulated PBMCs from healthy donors. Aliquots from culture with a high production of virus are stored for later use.

  8. HIV-1 Nef in Macrophage-Mediated Disease Pathogenesis

    Science.gov (United States)

    Lamers, Susanna L.; Fogel, Gary B.; Singer, Elyse J.; Salemi, Marco; Nolan, David J.; Huysentruyt, Leanne C.; McGrath, Michael S.

    2013-01-01

    Combined anti-retroviral therapy (cART) has significantly reduced the number of AIDS-associated illnesses and changed the course of HIV-1 disease in developed countries. Despite the ability of cART to maintain high CD4+ T-cell counts, a number of macrophage-mediated diseases can still occur in HIV-infected subjects. These diseases include lymphoma, metabolic diseases, and HIV-associated neurological disorders. Within macrophages, the HIV-1 regulatory protein “Nef” can modulate surface receptors, interact with signaling pathways, and promote specific environments that contribute to each of these pathologies. Moreover, genetic variation in Nef may also guide the macrophage response. Herein, we review findings relating to the Nef–macrophage interaction and how this relationship contributes to disease pathogenesis. PMID:23215766

  9. Cytokine expression during syphilis infection in HIV-1-infected individuals

    DEFF Research Database (Denmark)

    Knudsen, Andreas; Benfield, Thomas; Kofoed, Kristian

    2009-01-01

    BACKGROUND: Little is known about cytokine responses to syphilis infection in HIV-1-infected individuals. METHODS: We retrospectively identified patients with HIV-1 and Treponema pallidum coinfection. Plasma samples from before, during, and after coinfection were analyzed for interleukin (IL)-2, IL......-4, IL-6, IL-8, IL-10, interferon (IFN)-gamma, and tumor necrosis factor (TNF)-alpha. RESULTS: Thirty-six patients were included. IL-10 levels increased significantly in patients with primary or secondary stage syphilis from a median of 12.8 pg/mL [interquartile range (IQR), 11.0-27.8] before...... infection to 46.7 pg/mL (IQR, 28.4-78.9) at the time of diagnosis (P = 0.027) and decreased to 13.0 pg/mL (IQR, 6.2-19.4) after treatment of syphilis (P syphilis in patients with primary or secondary stage syphilis (median 3.9 pg...

  10. Characteristics of HIV-1 serodiscordant couples enrolled in a clinical trial of antiretroviral pre-exposure prophylaxis for HIV-1 prevention.

    Directory of Open Access Journals (Sweden)

    Andrew Mujugira

    Full Text Available Stable heterosexual HIV-1 serodiscordant couples in Africa have high HIV-1 transmission rates and are a critical population for evaluation of new HIV-1 prevention strategies. The Partners PrEP Study is a randomized, double-blind, placebo-controlled trial of tenofovir and emtricitabine-tenofovir pre-exposure prophylaxis to decrease HIV-1 acquisition within heterosexual HIV-1 serodiscordant couples. We describe the trial design and characteristics of the study cohort.HIV-1 serodiscordant couples, in which the HIV-1 infected partner did not meet national guidelines for initiation of antiretroviral therapy, were enrolled at 9 research sites in Kenya and Uganda. The HIV-1 susceptible partner was randomized to daily oral tenofovir, emtricitabine-tenofovir, or matching placebo with monthly follow-up for 24-36 months.From July 2008 to November 2010, 7920 HIV-1 serodiscordant couples were screened and 4758 enrolled. For 62% (2966/4758 of enrolled couples, the HIV-1 susceptible partner was male. Median age was 33 years for HIV-1 susceptible and HIV-1 infected partners [IQR (28-40 and (26-39 respectively]. Most couples (98% were married, with a median duration of partnership of 7.0 years (IQR 3.0-14.0 and recent knowledge of their serodiscordant status [median 0.4 years (IQR 0.1-2.0]. During the month prior to enrollment, couples reported a median of 4 sex acts (IQR 2-8; 27% reported unprotected sex and 14% of male and 1% of female HIV-1 susceptible partners reported sex with outside partners. Among HIV-1 infected partners, the median plasma HIV-1 level was 3.94 log(10 copies/mL (IQR 3.31-4.53 and median CD4 count was 496 cells/µL (IQR 375-662; the majority (64% had WHO stage 1 HIV-1 disease.Couples at high risk of HIV-1 transmission were rapidly recruited into the Partners PrEP Study, the largest efficacy trial of oral PrEP. (ClinicalTrials.gov NCT00557245.

  11. Barriers to Antiretroviral Initiation in HIV-1-Discordant Couples

    Science.gov (United States)

    Guthrie, Brandon L.; Choi, Robert Y.; Liu, Amy Y.; Mackelprang, Romel D.; Rositch, Anne F.; Bosire, Rose; Manyara, Lucy; Gatuguta, Anne; Kiarie, James N.; Farquhar, Carey

    2011-01-01

    BACKGROUND In Kenya and much of sub-Saharan Africa, nearly half of all couples affected by HIV are discordant. Antiretroviral therapy (ART) slows disease progression in HIV-1-infected individuals, and reduces transmission to uninfected partners. We examined time to ART initiation and factors associated with delayed initiation in HIV-1-discordant couples in Nairobi. METHODS HIV-1-discordant couples were enrolled and followed quarterly for up to 2 years. Clinical staff administered questionnaires and conducted viral loads and CD4 counts. Participants with a CD4 count meeting ART criteria were referred to a nearby PEPFAR-funded treatment center. Barriers to ART initiation among participants with a CD4 count eligible for ART were assessed by Cox regression. RESULTS Of 439 HIV-1-infected participants (63.6% females and 36.4% males) 146 met CD4 count criteria for ART during follow-up. Median time from meeting CD4 criteria until ART initiation was 8.9 months, with 42.0% of eligible participants on ART by 6 months and 63.4% on ART by 1 year. The CD4 count at the time of eligibility was inversely associated with time to ART initiation (HR=0.49, p< 0.001). Compared to homeowners, those paying higher rents started ART 48% more slowly (p=0.062) and those paying lower rents started 71% more slowly (p=0.002). CONCLUSIONS Despite access to regular health care, referrals to treatment centers, and free access to ART, over a third of participants with an eligible CD4 count had not started ART within 1 year. Factors of lower socioeconomic status may slow ART initiation and targeted approaches are needed to avoid delays in treatment initiation. PMID:21826010

  12. HIV-1 Infection in adults with haematological malignancies in ...

    African Journals Online (AJOL)

    Burkett's lymphoma, Hodgkin's disease and myelodysplastic syndrome had been less frequently diagnosed. Forty-five of all cases (26.2%) had antibodies to the HIV-1 virus, predominantly in patients with Non-Hodgkin's lymphomas (p<0.001, OR=5.8, adjusted for age; CI=2.7 – 12.4). About 19.9% and 11.8% of cases with ...

  13. HIV-1 binding and neutralizing antibodies of injecting drug users

    Directory of Open Access Journals (Sweden)

    E.P. Ouverney

    2005-09-01

    Full Text Available Previous studies have demonstrated a stronger seroreactivity against some synthetic peptides responsible for inducing neutralizing antibodies in injecting drug users (IDU compared to that of individuals sexually infected with HIV-1 (S, but the effectiveness in terms of the neutralizing ability of these antibodies has not been evaluated. Our objective was to study the humoral immune response of IDU by determining the specificity of their antibodies and the presence of neutralizing antibodies. The neutralization capacity against the HIV-1 isolate MN (genotype B, the primary HIV-1 isolate 95BRRJ021 (genotype F, and the seroreactivity with peptides known to induce neutralizing antibodies, from the V2 and V3 loops of different HIV-1 subtypes, were analyzed. Seroreactivity indicates that IDU plasma are more likely to recognize a broader range of peptides than S plasma, with significantly higher titers, especially of V3 peptides. Similar neutralization frequencies of the MN isolate were observed in plasma of the IDU (16/47 and S (20/60 groups in the 1:10 dilution. The neutralization of the 95BRRJ021 isolate was more frequently observed for plasma from the S group (15/23 than from the IDU group (15/47, P = 0.0108. No correlation between neutralization and seroreactivity with the peptides tested was observed. These results suggest that an important factor responsible for the extensive and broad humoral immune response observed in IDU is their infection route. There was very little difference in neutralizing antibody response between the IDU and S groups despite their differences in seroreactivity and health status.

  14. nef gene sequence variation among HIV-1-infected African children

    Czech Academy of Sciences Publication Activity Database

    Chakraborty, R.; Reiniš, Milan; Rostron, T.; Philpott, S.; Dong, T.; D'Agostino, A.; Musoke, R.; de Silva, E.; Stumpf, M.; Weiser, B.; Burger, H.; Rowland-Jones, S.L.

    2006-01-01

    Roč. 7, č. 2 (2006), s. 75-84 ISSN 1464-2662 Grant - others:Fogarty International Center, NIH(US) 3D43TW00915; NIH(US) RO1 AI 42555 Institutional research plan: CEZ:AV0Z50520514 Keywords : HIV-1 nef gene * non-clade B * Kenya Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.674, year: 2006

  15. A Novel Leu92 Mutant of HIV-1 Reverse Transcriptase with a Selective Deficiency in Strand Transfer Causes a Loss of Viral Replication.

    Science.gov (United States)

    Herzig, Eytan; Voronin, Nickolay; Kucherenko, Nataly; Hizi, Amnon

    2015-08-01

    The process of reverse transcription (RTN) in retroviruses is essential to the viral life cycle. This key process is catalyzed exclusively by the viral reverse transcriptase (RT) that copies the viral RNA into DNA by its DNA polymerase activity, while concomitantly removing the original RNA template by its RNase H activity. During RTN, the combination between DNA synthesis and RNA hydrolysis leads to strand transfers (or template switches) that are critical for the completion of RTN. The balance between these RT-driven activities was considered to be the sole reason for strand transfers. Nevertheless, we show here that a specific mutation in HIV-1 RT (L92P) that does not affect the DNA polymerase and RNase H activities abolishes strand transfer. There is also a good correlation between this complete loss of the RT's strand transfer to the loss of the DNA clamp activity of the RT, discovered recently by us. This finding indicates a mechanistic linkage between these two functions and that they are both direct and unique functions of the RT (apart from DNA synthesis and RNA degradation). Furthermore, when the RT's L92P mutant was introduced into an infectious HIV-1 clone, it lost viral replication, due to inefficient intracellular strand transfers during RTN, thus supporting the in vitro data. As far as we know, this is the first report on RT mutants that specifically and directly impair RT-associated strand transfers. Therefore, targeting residue Leu92 may be helpful in selectively blocking this RT activity and consequently HIV-1 infectivity and pathogenesis. Reverse transcription in retroviruses is essential for the viral life cycle. This multistep process is catalyzed by viral reverse transcriptase, which copies the viral RNA into DNA by its DNA polymerase activity (while concomitantly removing the RNA template by its RNase H activity). The combination and balance between synthesis and hydrolysis lead to strand transfers that are critical for reverse transcription

  16. New approaches to design HIV-1 T-cell vaccines.

    Science.gov (United States)

    Perrin, Hélène; Canderan, Glenda; Sékaly, Rafick-Pierre; Trautmann, Lydie

    2010-09-01

    Following the evidence that T-cell responses are crucial in the control of HIV-1 infection, vaccines targeting T-cell responses were tested in recent clinical trials. However, these vaccines showed a lack of efficacy. This review attempts to define the qualitative and quantitative features that are desirable for T-cell-induced responses by vaccines. We also describe strategies that could lead to achievement of this goal. Using the yellow fever vaccine as a benchmark of an efficient vaccine, recent studies identified factors of immune protection and more importantly innate immune pathways needed for the establishment of long-term protective adaptive immunity. To prevent or control HIV-1 infection, a vaccine must induce efficient and persistent antigen-specific T cells endowed with mucosal homing capacity. Such cells should have the capability to counteract HIV-1 diversity and its rapid spread from the initial site of infection. To achieve this goal, the activation of a diversified innate immune response is critical. New systems biology approaches will provide more precise correlates of immune protection that will pave the way for new approaches in T-cell-based vaccines.

  17. The global spread of HIV-1 subtype B epidemic.

    Science.gov (United States)

    Magiorkinis, Gkikas; Angelis, Konstantinos; Mamais, Ioannis; Katzourakis, Aris; Hatzakis, Angelos; Albert, Jan; Lawyer, Glenn; Hamouda, Osamah; Struck, Daniel; Vercauteren, Jurgen; Wensing, Annemarie; Alexiev, Ivailo; Åsjö, Birgitta; Balotta, Claudia; Gomes, Perpétua; Camacho, Ricardo J; Coughlan, Suzie; Griskevicius, Algirdas; Grossman, Zehava; Horban, Anders; Kostrikis, Leondios G; Lepej, Snjezana J; Liitsola, Kirsi; Linka, Marek; Nielsen, Claus; Otelea, Dan; Paredes, Roger; Poljak, Mario; Puchhammer-Stöckl, Elizabeth; Schmit, Jean Claude; Sönnerborg, Anders; Staneková, Danica; Stanojevic, Maja; Stylianou, Dora C; Boucher, Charles A B; Nikolopoulos, Georgios; Vasylyeva, Tetyana; Friedman, Samuel R; van de Vijver, David; Angarano, Gioacchino; Chaix, Marie-Laure; de Luca, Andrea; Korn, Klaus; Loveday, Clive; Soriano, Vincent; Yerly, Sabine; Zazzi, Mauricio; Vandamme, Anne-Mieke; Paraskevis, Dimitrios

    2016-12-01

    Human immunodeficiency virus type 1 (HIV-1) was discovered in the early 1980s when the virus had already established a pandemic. For at least three decades the epidemic in the Western World has been dominated by subtype B infections, as part of a sub-epidemic that traveled from Africa through Haiti to United States. However, the pattern of the subsequent spread still remains poorly understood. Here we analyze a large dataset of globally representative HIV-1 subtype B strains to map their spread around the world over the last 50years and describe significant spread patterns. We show that subtype B travelled from North America to Western Europe in different occasions, while Central/Eastern Europe remained isolated for the most part of the early epidemic. Looking with more detail in European countries we see that the United Kingdom, France and Switzerland exchanged viral isolates with non-European countries than with European ones. The observed pattern is likely to mirror geopolitical landmarks in the post-World War II era, namely the rise and the fall of the Iron Curtain and the European colonialism. In conclusion, HIV-1 spread through specific migration routes which are consistent with geopolitical factors that affected human activities during the last 50years, such as migration, tourism and trade. Our findings support the argument that epidemic control policies should be global and incorporate political and socioeconomic factors. Copyright © 2016 The Authors. Published by Elsevier B.V. All rights reserved.

  18. HIV-1 transgenic rats develop T cell abnormalities

    International Nuclear Information System (INIS)

    Reid, William; Abdelwahab, Sayed; Sadowska, Mariola; Huso, David; Neal, Ashley; Ahearn, Aaron; Bryant, Joseph; Gallo, Robert C.; Lewis, George K.; Reitz, Marvin

    2004-01-01

    HIV-1 infection leads to impaired antigen-specific T cell proliferation, increased susceptibility of T cells to apoptosis, progressive impairment of T-helper 1 (Th1) responses, and altered maturation of HIV-1-specific memory cells. We have identified similar impairments in HIV-1 transgenic (Tg) rats. Tg rats developed an absolute reduction in CD4 + and CD8 + T cells able to produce IFN-γ following activation and an increased susceptibility of T cells to activation-induced apoptosis. CD4 + and CD8 + effector/memory (CD45RC - CD62L - ) pools were significantly smaller in Tg rats compared to non-Tg controls, although the converse was true for the naieve (CD45RC + CD62L + ) T cell pool. Our interpretation is that the HIV transgene causes defects in the development of T cell effector function and generation of specific effector/memory T cell subsets, and that activation-induced apoptosis may be an essential factor in this process

  19. Direct and dynamic detection of HIV-1 in living cells.

    Directory of Open Access Journals (Sweden)

    Jonas Helma

    Full Text Available In basic and applied HIV research, reliable detection of viral components is crucial to monitor progression of infection. While it is routine to detect structural viral proteins in vitro for diagnostic purposes, it previously remained impossible to directly and dynamically visualize HIV in living cells without genetic modification of the virus. Here, we describe a novel fluorescent biosensor to dynamically trace HIV-1 morphogenesis in living cells. We generated a camelid single domain antibody that specifically binds the HIV-1 capsid protein (CA at subnanomolar affinity and fused it to fluorescent proteins. The resulting fluorescent chromobody specifically recognizes the CA-harbouring HIV-1 Gag precursor protein in living cells and is applicable in various advanced light microscopy systems. Confocal live cell microscopy and super-resolution microscopy allowed detection and dynamic tracing of individual virion assemblies at the plasma membrane. The analysis of subcellular binding kinetics showed cytoplasmic antigen recognition and incorporation into virion assembly sites. Finally, we demonstrate the use of this new reporter in automated image analysis, providing a robust tool for cell-based HIV research.

  20. Innate immune reconstitution with suppression of HIV-1.

    Science.gov (United States)

    Scully, Eileen P; Lockhart, Ainsley; Garcia-Beltran, Wilfredo; Palmer, Christine D; Musante, Chelsey; Rosenberg, Eric; Allen, Todd M; Chang, J Judy; Bosch, Ronald J; Altfeld, Marcus

    2016-03-17

    Progressive HIV-1 infection leads to both profound immune suppression and pathologic inflammation in the majority of infected individuals. While adaptive immune dysfunction, as evidenced by CD4 + T cell depletion and exhaustion, has been extensively studied, less is known about the functional capacity of innate immune cell populations in the context of HIV-1 infection. Given the broad susceptibility to opportunistic infections and the dysregulated inflammation observed in progressive disease, we hypothesized that there would be significant changes in the innate cellular responses. Using a cohort of patients with multiple samplings before and after antiretroviral therapy (ART) initiation, we demonstrated increased responses to innate immune stimuli following viral suppression, as measured by the production of inflammatory cytokines. Plasma viral load itself had the strongest association with this change in innate functional capacity. We further identified epigenetic modifications in the TNFA promoter locus in monocytes that are associated with viremia, suggesting a molecular mechanism for the observed changes in innate immune function following initiation of ART. These data indicate that suppression of HIV-1 viremia is associated with changes in innate cellular function that may in part determine the restoration of protective immune responses.

  1. Oligodendrocyte Injury and Pathogenesis of HIV-1-Associated Neurocognitive Disorders

    Directory of Open Access Journals (Sweden)

    Han Liu

    2016-07-01

    Full Text Available Oligodendrocytes wrap neuronal axons to form myelin, an insulating sheath which is essential for nervous impulse conduction along axons. Axonal myelination is highly regulated by neuronal and astrocytic signals and the maintenance of myelin sheaths is a very complex process. Oligodendrocyte damage can cause axonal demyelination and neuronal injury, leading to neurological disorders. Demyelination in the cerebrum may produce cognitive impairment in a variety of neurological disorders, including human immunodeficiency virus type one (HIV-1-associated neurocognitive disorders (HAND. Although the combined antiretroviral therapy has markedly reduced the incidence of HIV-1-associated dementia, a severe form of HAND, milder forms of HAND remain prevalent even when the peripheral viral load is well controlled. HAND manifests as a subcortical dementia with damage in the brain white matter (e.g., corpus callosum, which consists of myelinated axonal fibers. How HIV-1 brain infection causes myelin injury and resultant white matter damage is an interesting area of current HIV research. In this review, we tentatively address recent progress on oligodendrocyte dysregulation and HAND pathogenesis.

  2. Multimerized CHR-derived peptides as HIV-1 fusion inhibitors.

    Science.gov (United States)

    Nomura, Wataru; Hashimoto, Chie; Suzuki, Takaharu; Ohashi, Nami; Fujino, Masayuki; Murakami, Tsutomu; Yamamoto, Naoki; Tamamura, Hirokazu

    2013-08-01

    To date, several HIV-1 fusion inhibitors based on the carboxy-terminal leucine/isoleucine heptad repeat (CHR) region of an HIV-1 envelope protein gp41 have been discovered. We have shown that a synthetic peptide mimetic of a trimer form of the CHR-derived peptide C34 has potent inhibitory activity against the HIV-1 fusion mechanism, compared to a monomer C34 peptide. The present study revealed that a dimeric form of C34 is evidently structurally critical for fusion inhibitors, and that the activity of multimerized CHR-derived peptides in fusion inhibition is affected by the properties of the unit peptides C34, SC34EK, and T20. The fluorescence-based study suggested that the N36-interactive sites of the C34 trimer, including hydrophobic residues, are exposed outside the trimer and that trimerization of C34 caused a remarkable increase in fusion inhibitory activity. The present results could be useful in the design of fusion inhibitors against viral infections which proceed via membrane fusion with host cells. Copyright © 2013 The Authors. Published by Elsevier Ltd.. All rights reserved.

  3. Positron emission tomography in patients suffering from HIV-1 infection

    Energy Technology Data Exchange (ETDEWEB)

    Sathekge, Mike [University Hospital of Pretoria, Department of Nuclear Medicine, Pretoria (South Africa); Goethals, Ingeborg; Wiele, Christophe van de [University Hospital Ghent, Department of Nuclear Medicine, Ghent (Belgium); Maes, Alex [AZ Groening, Department of Nuclear Medicine, Kortrijk (Belgium)

    2009-07-15

    This paper reviews currently available PET studies performed either to improve our understanding of the pathogenesis of HIV-1 infection or to assess the value of PET imaging in the clinical decision making of patients infected with HIV-1 presenting with AIDS-related opportunistic infections and malignancies. FDG PET has shown that HIV-1 infection progresses by distinct anatomical steps, with involvement of the upper torso preceding involvement of the lower part of the torso, and that the degree of FDG uptake relates to viral load. The former finding suggests that lymphoid tissues are engaged in a predictable sequence and that diffusible mediators of activation might be important targets for vaccine or therapeutic intervention strategies. In lipodystrophic HIV-infected patients, limited available data support the hypothesis that stavudine-related lipodystrophy is associated with increased glucose uptake by adipose tissue as a result of the metabolic stress of adipose tissue in response to highly active antiretroviral treatment (HAART). Finally, in early AIDS-related dementia complex (ADC), striatal hypermetabolism is observed, whereas progressive ADC is characterized by a decrease in subcortical and cortical metabolism. In the clinical setting, PET has been shown to allow the differentiation of AIDS-related opportunistic infections and malignancies, and to allow monitoring of side effects of HAART. However, in patients suffering from HIV infection and presenting with extracerebral lymphoma or other human malignancies, knowledge of viraemia is essential when interpreting FDG PET imaging. (orig.)

  4. Positron emission tomography in patients suffering from HIV-1 infection

    International Nuclear Information System (INIS)

    Sathekge, Mike; Goethals, Ingeborg; Wiele, Christophe van de; Maes, Alex

    2009-01-01

    This paper reviews currently available PET studies performed either to improve our understanding of the pathogenesis of HIV-1 infection or to assess the value of PET imaging in the clinical decision making of patients infected with HIV-1 presenting with AIDS-related opportunistic infections and malignancies. FDG PET has shown that HIV-1 infection progresses by distinct anatomical steps, with involvement of the upper torso preceding involvement of the lower part of the torso, and that the degree of FDG uptake relates to viral load. The former finding suggests that lymphoid tissues are engaged in a predictable sequence and that diffusible mediators of activation might be important targets for vaccine or therapeutic intervention strategies. In lipodystrophic HIV-infected patients, limited available data support the hypothesis that stavudine-related lipodystrophy is associated with increased glucose uptake by adipose tissue as a result of the metabolic stress of adipose tissue in response to highly active antiretroviral treatment (HAART). Finally, in early AIDS-related dementia complex (ADC), striatal hypermetabolism is observed, whereas progressive ADC is characterized by a decrease in subcortical and cortical metabolism. In the clinical setting, PET has been shown to allow the differentiation of AIDS-related opportunistic infections and malignancies, and to allow monitoring of side effects of HAART. However, in patients suffering from HIV infection and presenting with extracerebral lymphoma or other human malignancies, knowledge of viraemia is essential when interpreting FDG PET imaging. (orig.)

  5. Binding kinetics of aptamers to gp120 derived from HIV-1 subtype C

    CSIR Research Space (South Africa)

    Millroy, L

    2011-02-01

    Full Text Available aptamers with specific and strong affinity to the HIV-1 envelope glycoprotein gp120 and act as novel HIV-1 entry inhibitor drugs or as targeted drug delivery systems to HIV-1 infected cells. Prior to any downstream applications, novel gp120 aptamers need...

  6. Blocking type I interferon signaling enhances T cell recovery and reduces HIV-1 reservoirs.

    Science.gov (United States)

    Cheng, Liang; Ma, Jianping; Li, Jingyun; Li, Dan; Li, Guangming; Li, Feng; Zhang, Qing; Yu, Haisheng; Yasui, Fumihiko; Ye, Chaobaihui; Tsao, Li-Chung; Hu, Zhiyuan; Su, Lishan; Zhang, Liguo

    2017-01-03

    Despite the efficient suppression of HIV-1 replication that can be achieved with combined antiretroviral therapy (cART), low levels of type I interferon (IFN-I) signaling persist in some individuals. This sustained signaling may impede immune recovery and foster viral persistence. Here we report studies using a monoclonal antibody to block IFN-α/β receptor (IFNAR) signaling in humanized mice (hu-mice) that were persistently infected with HIV-1. We discovered that effective cART restored the number of human immune cells in HIV-1-infected hu-mice but did not rescue their immune hyperactivation and dysfunction. IFNAR blockade fully reversed HIV-1-induced immune hyperactivation and rescued anti-HIV-1 immune responses in T cells from HIV-1-infected hu-mice. Finally, we found that IFNAR blockade in the presence of cART reduced the size of HIV-1 reservoirs in lymphoid tissues and delayed HIV-1 rebound after cART cessation in the HIV-1-infected hu-mice. We conclude that low levels of IFN-I signaling contribute to HIV-1-associated immune dysfunction and foster HIV-1 persistence in cART-treated hosts. Our results suggest that blocking IFNAR may provide a potential strategy to enhance immune recovery and reduce HIV-1 reservoirs in individuals with sustained elevations in IFN-I signaling during suppressive cART.

  7. HIV-1 seroprevalence and subtypes in police recruits from Afar regional state, Ethiopia

    NARCIS (Netherlands)

    Zewde, Ayele; Bahiru, Seifu; Sanders, Eduard; Tilahun, Tesfaye; Beyene, Asfaw; Alebachew, Mengiste; Schaap, Ab; Wolday, Dawit; Rinke de Wit, Tobias F.

    2002-01-01

    Surveillance for HIV-1 prevalence and subtypes in Afar Region, Ethiopia was performed among police recruits in the year 2000, by unlinked anonymous testing. Of 408 samples tested, 26 (6.4%) appeared positive for HIV-1 antibodies. There was a trend for higher HIV-1 seroprevalence in women (9.5%,

  8. Creatine protects against mitochondrial dysfunction associated with HIV-1 Tat-induced neuronal injury

    Science.gov (United States)

    Stevens, Patrick R.; Gawryluk, Jeremy W.; Hui, Liang; Chen, Xuesong; Geiger, Jonathan D.

    2015-01-01

    HIV-1 infected individuals are living longer but experiencing a prevalence rate of over 50% for HIV-1 associated neurocognitive disorders (HAND) for which no effective treatment is available. Viral and cellular factors secreted by HIV-1 infected cells leads to neuronal injury and HIV-1 Tat continues to be implicated in the pathogenesis of HAND. Here we tested the hypothesis that creatine protected against HIV-1 Tat-induced neuronal injury by preventing mitochondrial bioenergetic crisis and/or redox catastrophe. Creatine blocked HIV-1 Tat1-72-induced increases in neuron cell death and synaptic area loss. Creatine protected against HIV-1 Tat-induced decreases in ATP. Creatine and creatine plus HIV-1 Tat increased cellular levels of creatine, and creatine plus HIV-1 Tat further decreased ratios of phosphocreatine to creatine observed with creatine or HIV-1 Tat treatments alone. Additionally, creatine protected against HIV-1 Tat-induced mitochondrial hypopolarization and HIV-1 Tat-induced mitochondrial permeability transition pore opening. Thus, creatine may be a useful adjunctive therapy against HAND. PMID:25613139

  9. Histone deacetylase inhibitors for purging HIV-1 from the latent reservoir.

    NARCIS (Netherlands)

    Matalon, S.; Rasmussen, T.A.; Dinarello, C.A.

    2011-01-01

    A reservoir of latently infected memory CD4(+) T cells is believed to be the source of HIV-1 reemergence after discontinuation of antiretroviral therapy. HIV-1 eradication may depend on depletion of this reservoir. Integrated HIV-1 is inaccessible for expression, in part because of histone

  10. Effect of HIV-1 infection on malaria treatment outcome in Ugandan ...

    African Journals Online (AJOL)

    Background: Malaria and HIV-1 infection cause significant morbidity and mortality in sub-Saharan Africa. HIV-1 increases risk for malaria with the risk increasing as immunity declines.The effect of HIV-1 infection on antimalarial treatment outcome is still inconclusive. Objective: To compare antimalarial treatment outcome ...

  11. The yeast Ty3 retrotransposon contains a 5'-3' bipartite primer-binding site and encodes nucleocapsid protein NCp9 functionally homologous to HIV-1 NCp7.

    Science.gov (United States)

    Gabus, C; Ficheux, D; Rau, M; Keith, G; Sandmeyer, S; Darlix, J L

    1998-08-17

    Retroviruses, including HIV-1 and the distantly related yeast retroelement Ty3, all encode a nucleoprotein required for virion structure and replication. During an in vitro comparison of HIV-1 and Ty3 nucleoprotein function in RNA dimerization and cDNA synthesis, we discovered a bipartite primer-binding site (PBS) for Ty3 composed of sequences located at opposite ends of the genome. Ty3 cDNA synthesis requires the 3' PBS for primer tRNAiMet annealing to the genomic RNA, and the 5' PBS, in cis or in trans, as the reverse transcription start site. Ty3 RNA alone is unable to dimerize, but formation of dimeric tRNAiMet bound to the PBS was found to direct dimerization of Ty3 RNA-tRNAiMet. Interestingly, HIV-1 nucleocapsid protein NCp7 and Ty3 NCp9 were interchangeable using HIV-1 and Ty3 RNA template-primer systems. Our findings impact on the understanding of non-canonical reverse transcription as well as on the use of Ty3 systems to screen for anti-NCp7 drugs.

  12. Cerebrospinal fluid HIV-1 RNA levels in asymptomatic patients with early stage chronic HIV-1 infection: support for the hypothesis of local virus replication.

    Science.gov (United States)

    García, F; Niebla, G; Romeu, J; Vidal, C; Plana, M; Ortega, M; Ruiz, L; Gallart, T; Clotet, B; Miró, J M; Pumarola, T; Gatell, J M

    1999-08-20

    To assess HIV-1 RNA levels in cerebrospinal fluid (CSF) and their potential correlation with plasma viral load and central nervous system (CNS) HIV-1 infection markers in stable asymptomatic patients with a CD4 T cell count >500x10(6) cells/l. Consecutive patients screened for two trials were eligible for lumbar puncture assessment. At day 0, simultaneous samples of CSF and plasma were obtained and levels of total proteins, albumin, IgG, antibodies against HIV-1 p24 antigen, HIV-1 RNA (using the polymerase chain technique) and white cells were measured. The integrity of the blood-brain barrier was preserved (albumin index > or =7) in 59 out of 70 patients (84%). Intrathecal production of antibodies against HIV-1 p24 antigen was demonstrated in 55 out of 70 individuals (78%). Viral load in CSF was significantly lower than plasma values (3.13+/-0.95 versus 4.53+/-0.53, P = 0.0001). HIV-1 RNA was not detected in CSF in only three of the 70 patients (4%). Overall, there was a significant correlation between plasma and CSF HIV-1 RNA levels (r = 0.43, P = 0.0001); however, in 29 patients (41%) there were significant differences (>1.5 log10 copies/ml) between the viral loads in plasma and CSF. In the multivariate analysis, a high level of protein and white cells in CSF, but not the HIV-1 RNA plasma level, were factors independently associated with a higher level of HIV-1 RNA in CSF (P = 0.0001). HIV-1 RNA can be detected almost always in CSF of asymptomatic patients in early stages of HIV-1 infection including those with a preserved integrity of the blood-brain barrier. The important discrepancies between plasma and CSF viral load, and the independent association between CSF abnormalities and CSF viral load, support the hypothesis of local production of HIV-1.

  13. Mother-to-Child HIV-1 Transmission Events Are Differentially Impacted by Breast Milk and Its Components from HIV-1-Infected Women.

    Directory of Open Access Journals (Sweden)

    Ruizhong Shen

    Full Text Available Breast milk is a vehicle of infection and source of protection in post-natal mother-to-child HIV-1 transmission (MTCT. Understanding the mechanism by which breast milk limits vertical transmission will provide critical insight into the design of preventive and therapeutic approaches to interrupt HIV-1 mucosal transmission. However, characterization of the inhibitory activity of breast milk in human intestinal mucosa, the portal of entry in postnatal MTCT, has been constrained by the limited availability of primary mucosal target cells and tissues to recapitulate mucosal transmission ex vivo. Here, we characterized the impact of skimmed breast milk, breast milk antibodies (Igs and non-Ig components from HIV-1-infected Ugandan women on the major events of HIV-1 mucosal transmission using primary human intestinal cells and tissues. HIV-1-specific IgG antibodies and non-Ig components in breast milk inhibited the uptake of Ugandan HIV-1 isolates by primary human intestinal epithelial cells, viral replication in and transport of HIV-1- bearing dendritic cells through the human intestinal mucosa. Breast milk HIV-1-specific IgG and IgA, as well as innate factors, blocked the uptake and transport of HIV-1 through intestinal mucosa. Thus, breast milk components have distinct and complementary effects in reducing HIV-1 uptake, transport through and replication in the intestinal mucosa and, therefore, likely contribute to preventing postnatal HIV-1 transmission. Our data suggests that a successful preventive or therapeutic approach would require multiple immune factors acting at multiple steps in the HIV-1 mucosal transmission process.

  14. Rapid Antiretroviral Therapy Initiation for Women in an HIV-1 Prevention Clinical Trial Experiencing Primary HIV-1 Infection during Pregnancy or Breastfeeding.

    Science.gov (United States)

    Morrison, Susan; John-Stewart, Grace; Egessa, John J; Mubezi, Sezi; Kusemererwa, Sylvia; Bii, Dennis K; Bulya, Nulu; Mugume, Francis; Campbell, James D; Wangisi, Jonathan; Bukusi, Elizabeth A; Celum, Connie; Baeten, Jared M

    2015-01-01

    During an HIV-1 prevention clinical trial in East Africa, we observed 16 cases of primary HIV-1 infection in women coincident with pregnancy or breastfeeding. Nine of eleven pregnant women initiated rapid combination antiretroviral therapy (ART), despite having CD4 counts exceeding national criteria for ART initiation; breastfeeding women initiated ART or replacement feeding. Rapid ART initiation during primary HIV-1 infection during pregnancy and breastfeeding is feasible in this setting.

  15. Rapid Antiretroviral Therapy Initiation for Women in an HIV-1 Prevention Clinical Trial Experiencing Primary HIV-1 Infection during Pregnancy or Breastfeeding.

    Directory of Open Access Journals (Sweden)

    Susan Morrison

    Full Text Available During an HIV-1 prevention clinical trial in East Africa, we observed 16 cases of primary HIV-1 infection in women coincident with pregnancy or breastfeeding. Nine of eleven pregnant women initiated rapid combination antiretroviral therapy (ART, despite having CD4 counts exceeding national criteria for ART initiation; breastfeeding women initiated ART or replacement feeding. Rapid ART initiation during primary HIV-1 infection during pregnancy and breastfeeding is feasible in this setting.

  16. Interferon-α regulates glutaminase 1 promoter through STAT1 phosphorylation: relevance to HIV-1 associated neurocognitive disorders.

    Directory of Open Access Journals (Sweden)

    Lixia Zhao

    Full Text Available HIV-1 associated neurocognitive disorders (HAND develop during progressive HIV-1 infection and affect up to 50% of infected individuals. Activated microglia and macrophages are critical cell populations that are involved in the pathogenesis of HAND, which is specifically related to the production and release of various soluble neurotoxic factors including glutamate. In the central nervous system (CNS, glutamate is typically derived from glutamine by mitochondrial enzyme glutaminase. Our previous study has shown that glutaminase is upregulated in HIV-1 infected monocyte-derived-macrophages (MDM and microglia. However, how HIV-1 leads to glutaminase upregulation, or how glutaminase expression is regulated in general, remains unclear. In this study, using a dual-luciferase reporter assay system, we demonstrated that interferon (IFN α specifically activated the glutaminase 1 (GLS1 promoter. Furthermore, IFN-α treatment increased signal transducer and activator of transcription 1 (STAT1 phosphorylation and glutaminase mRNA and protein levels. IFN-α stimulation of GLS1 promoter activity correlated to STAT1 phosphorylation and was reduced by fludarabine, a chemical that inhibits STAT1 phosphorylation. Interestingly, STAT1 was found to directly bind to the GLS1 promoter in MDM, an effect that was dependent on STAT1 phosphorylation and significantly enhanced by IFN-α treatment. More importantly, HIV-1 infection increased STAT1 phosphorylation and STAT1 binding to the GLS1 promoter, which was associated with increased glutamate levels. The clinical relevance of these findings was further corroborated with investigation of post-mortem brain tissues. The glutaminase C (GAC, one isoform of GLS1 mRNA levels in HIV associated-dementia (HAD individuals correlate with STAT1 (p<0.01, IFN-α (p<0.05 and IFN-β (p<0.01. Together, these data indicate that both HIV-1 infection and IFN-α treatment increase glutaminase expression through STAT1 phosphorylation and

  17. W-curve alignments for HIV-1 genomic comparisons.

    Directory of Open Access Journals (Sweden)

    Douglas J Cork

    2010-06-01

    Full Text Available The W-curve was originally developed as a graphical visualization technique for viewing DNA and RNA sequences. Its ability to render features of DNA also makes it suitable for computational studies. Its main advantage in this area is utilizing a single-pass algorithm for comparing the sequences. Avoiding recursion during sequence alignments offers advantages for speed and in-process resources. The graphical technique also allows for multiple models of comparison to be used depending on the nucleotide patterns embedded in similar whole genomic sequences. The W-curve approach allows us to compare large numbers of samples quickly.We are currently tuning the algorithm to accommodate quirks specific to HIV-1 genomic sequences so that it can be used to aid in diagnostic and vaccine efforts. Tracking the molecular evolution of the virus has been greatly hampered by gap associated problems predominantly embedded within the envelope gene of the virus. Gaps and hypermutation of the virus slow conventional string based alignments of the whole genome. This paper describes the W-curve algorithm itself, and how we have adapted it for comparison of similar HIV-1 genomes. A treebuilding method is developed with the W-curve that utilizes a novel Cylindrical Coordinate distance method and gap analysis method. HIV-1 C2-V5 env sequence regions from a Mother/Infant cohort study are used in the comparison.The output distance matrix and neighbor results produced by the W-curve are functionally equivalent to those from Clustal for C2-V5 sequences in the mother/infant pairs infected with CRF01_AE.Significant potential exists for utilizing this method in place of conventional string based alignment of HIV-1 genomes, such as Clustal X. With W-curve heuristic alignment, it may be possible to obtain clinically useful results in a short time-short enough to affect clinical choices for acute treatment. A description of the W-curve generation process, including a comparison

  18. W-curve alignments for HIV-1 genomic comparisons.

    Science.gov (United States)

    Cork, Douglas J; Lembark, Steven; Tovanabutra, Sodsai; Robb, Merlin L; Kim, Jerome H

    2010-06-01

    The W-curve was originally developed as a graphical visualization technique for viewing DNA and RNA sequences. Its ability to render features of DNA also makes it suitable for computational studies. Its main advantage in this area is utilizing a single-pass algorithm for comparing the sequences. Avoiding recursion during sequence alignments offers advantages for speed and in-process resources. The graphical technique also allows for multiple models of comparison to be used depending on the nucleotide patterns embedded in similar whole genomic sequences. The W-curve approach allows us to compare large numbers of samples quickly. We are currently tuning the algorithm to accommodate quirks specific to HIV-1 genomic sequences so that it can be used to aid in diagnostic and vaccine efforts. Tracking the molecular evolution of the virus has been greatly hampered by gap associated problems predominantly embedded within the envelope gene of the virus. Gaps and hypermutation of the virus slow conventional string based alignments of the whole genome. This paper describes the W-curve algorithm itself, and how we have adapted it for comparison of similar HIV-1 genomes. A treebuilding method is developed with the W-curve that utilizes a novel Cylindrical Coordinate distance method and gap analysis method. HIV-1 C2-V5 env sequence regions from a Mother/Infant cohort study are used in the comparison. The output distance matrix and neighbor results produced by the W-curve are functionally equivalent to those from Clustal for C2-V5 sequences in the mother/infant pairs infected with CRF01_AE. Significant potential exists for utilizing this method in place of conventional string based alignment of HIV-1 genomes, such as Clustal X. With W-curve heuristic alignment, it may be possible to obtain clinically useful results in a short time-short enough to affect clinical choices for acute treatment. A description of the W-curve generation process, including a comparison technique of

  19. Structural and dynamic characterization of the upper part of the HIV-1 cTAR DNA hairpin

    OpenAIRE

    Zargarian, Loussin?; Kanevsky, Igor; Bazzi, Ali; Boynard, Jonathan; Chaminade, Fran?oise; Foss?, Philippe; Mauffret, Olivier

    2009-01-01

    First strand transfer is essential for HIV-1 reverse transcription. During this step, the TAR RNA hairpin anneals to the cTAR DNA hairpin; this annealing reaction is promoted by the nucleocapsid protein and involves an initial loop?loop interaction between the apical loops of TAR and cTAR. Using NMR and probing methods, we investigated the structural and dynamic properties of the top half of the cTAR DNA (mini-cTAR). We show that the upper stem located between the apical and the internal loop...

  20. HIV-1 Infection of Primary CD4+ T Cells Regulates the Expression of Specific Human Endogenous Retrovirus HERV-K (HML-2) Elements.

    Science.gov (United States)

    Young, George R; Terry, Sandra N; Manganaro, Lara; Cuesta-Dominguez, Alvaro; Deikus, Gintaras; Bernal-Rubio, Dabeiba; Campisi, Laura; Fernandez-Sesma, Ana; Sebra, Robert; Simon, Viviana; Mulder, Lubbertus C F

    2018-01-01

    Endogenous retroviruses (ERVs) occupy extensive regions of the human genome. Although many of these retroviral elements have lost their ability to replicate, those whose insertion took place more recently, such as the HML-2 group of HERV-K elements, still retain intact open reading frames and the capacity to produce certain viral RNA and/or proteins. Transcription of these ERVs is, however, tightly regulated by dedicated epigenetic control mechanisms. Nonetheless, it has been reported that some pathological states, such as viral infections and certain cancers, coincide with ERV expression, suggesting that transcriptional reawakening is possible. HML-2 elements are reportedly induced during HIV-1 infection, but the conserved nature of these elements has, until recently, rendered their expression profiling problematic. Here, we provide comprehensive HERV-K HML-2 expression profiles specific for productively HIV-1-infected primary human CD4 + T cells. We combined enrichment of HIV-1 infected cells using a reporter virus expressing a surface reporter for gentle and efficient purification with long-read single-molecule real-time sequencing. We show that three HML-2 proviruses-6q25.1, 8q24.3, and 19q13.42-are upregulated on average between 3- and 5-fold in HIV-1-infected CD4 + T cells. One provirus, HML-2 12q24.33, in contrast, was repressed in the presence of active HIV replication. In conclusion, this report identifies the HERV-K HML-2 loci whose expression profiles differ upon HIV-1 infection in primary human CD4 + T cells. These data will help pave the way for further studies on the influence of endogenous retroviruses on HIV-1 replication. IMPORTANCE Endogenous retroviruses inhabit big portions of our genome. Moreover, although they are mainly inert, some of the evolutionarily younger members maintain the ability to express both RNA and proteins. We have developed an approach using long-read single-molecule real-time (SMRT) sequencing that produces long reads that

  1. Inhibition of Non Canonical HIV-1 Tat Secretion Through the Cellular Na+,K+-ATPase Blocks HIV-1 Infection

    Directory of Open Access Journals (Sweden)

    Silvia Agostini

    2017-07-01

    Full Text Available Besides its essential role in the activation of HIV-1 gene expression, the viral Tat protein has the unusual property of trafficking in and out of cells. In contrast to Tat internalization, the mechanism involved in extracellular Tat release has so far remained elusive. Here we show that Tat secretion occurs through a Golgi-independent pathway requiring binding of Tat with three short, non-consecutive intracytoplasmic loops at the C-terminus of the cellular Na+,K+-ATPase pump alpha subunit. Ouabain, a pump inhibitor, blocked this interaction and prevented Tat secretion; virions produced in the presence of this drug were less infectious, consistent the capacity of virion-associated Tat to increase HIV-1 infectivity. Treatment of CD4+ T-cells with short peptides corresponding to the Tat-binding regions of the pump alpha subunit impaired extracellular Tat release and blocked HIV-1 replication. Thus, non canonical, extracellular Tat secretion is essential for viral infectivity.

  2. Increased Risk of Female HIV-1 Acquisition Throughout Pregnancy and Postpartum: A Prospective Per-coital Act Analysis Among Women with HIV-1 Infected Partners.

    Science.gov (United States)

    Thomson, Kerry A; Hughes, James; Baeten, Jared M; John-Stewart, Grace; Celum, Connie; Cohen, Craig R; Ngure, Kenneth; Kiarie, James; Mugo, Nelly; Heffron, Renee

    2018-03-05

    Understanding the absolute and relative risk of HIV-1 acquisition during pregnancy and postpartum can inform HIV-1 prevention strategies for women. We used a complementary log-log model and data from 2,751 HIV-1 serodiscordant couples to compare the probability of women's HIV-1 acquisition risk per sex act during early pregnancy, late pregnancy, postpartum, and non-pregnant periods. At total of 686 pregnancies were identified and 82 incident HIV-1 infections occurred. After adjustment for condom use, age, PrEP use, and HIV-1 viral load, the per act probability of HIV-1 acquisition was higher in late pregnancy (aRR 2.82, p=0.01) and postpartum (aRR 3.97, p=0.01) compared to non-pregnant periods. The HIV-1 acquisition probability per condomless sex act for a 25 year old woman not taking PrEP with an HIV-1 infected male partner with viral load of 10,000 copies/ml was 0.0011 (95% CI: 0.005, 0.0019), 0.0022 (95% CI: 0.0004, 0.0093), 0.0030 (95% CI: 0.0007, 0.0108), and 0.0042 (95% CI: 0.0007, 0.0177) in the non-pregnant, early pregnant, late pregnant, and postpartum periods, respectively. The HIV-1 acquisition probability per condomless sex act steadily increased through pregnancy and was highest during the postpartum period, suggesting that biological changes during pregnancy and postpartum increase female HIV-1 susceptibility.

  3. Genome-wide analysis of LTR-retrotransposons in oil palm.

    Science.gov (United States)

    Beulé, Thierry; Agbessi, Mawussé Dt; Dussert, Stephane; Jaligot, Estelle; Guyot, Romain

    2015-10-15

    The oil palm (Elaeis guineensis Jacq.) is a major cultivated crop and the world's largest source of edible vegetable oil. The genus Elaeis comprises two species E. guineensis, the commercial African oil palm and E. oleifera, which is used in oil palm genetic breeding. The recent publication of both the African oil palm genome assembly and the first draft sequence of its Latin American relative now allows us to tackle the challenge of understanding the genome composition, structure and evolution of these palm genomes through the annotation of their repeated sequences. In this study, we identified, annotated and compared Transposable Elements (TE) from the African and Latin American oil palms. In a first step, Transposable Element databases were built through de novo detection in both genome sequences then the TE content of both genomes was estimated. Then putative full-length retrotransposons with Long Terminal Repeats (LTRs) were further identified in the E. guineensis genome for characterization of their structural diversity, copy number and chromosomal distribution. Finally, their relative expression in several tissues was determined through in silico analysis of publicly available transcriptome data. Our results reveal a congruence in the transpositional history of LTR retrotransposons between E. oleifera and E. guineensis, especially the Sto-4 family. Also, we have identified and described 583 full-length LTR-retrotransposons in the Elaeis guineensis genome. Our work shows that these elements are most likely no longer mobile and that no recent insertion event has occurred. Moreover, the analysis of chromosomal distribution suggests a preferential insertion of Copia elements in gene-rich regions, whereas Gypsy elements appear to be evenly distributed throughout the genome. Considering the high proportion of LTR retrotransposon in the oil palm genome, our work will contribute to a greater understanding of their impact on genome organization and evolution

  4. Gene expression profiling of the host response to HIV-1 B, C, or A/E infection in monocyte-derived dendritic cells

    International Nuclear Information System (INIS)

    Solis, Mayra; Wilkinson, Peter; Romieu, Raphaelle; Hernandez, Eduardo; Wainberg, Mark A.; Hiscott, John

    2006-01-01

    Dendritic cells (DC) are among the first targets of human immunodeficiency virus type-1 (HIV-1) infection and in turn play a crucial role in viral transmission to T cells and in the regulation of the immune response. The major group of HIV-1 has diversified genetically based on variation in env sequences and comprise at least 11 subtypes. Because little is known about the host response elicited against different HIV-1 clade isolates in vivo, we sought to use gene expression profiling to identify genes regulated by HIV-1 subtypes B, C, and A/E upon de novo infection of primary immature monocyte-derived DC (iMDDCs). A total of 3700 immune-related genes were subjected to a significance analysis of microarrays (SAM); 656 genes were selected as significant and were further divided into 8 functional categories. Regardless of the time of infection, 20% of the genes affected by HIV-1 were involved in signal transduction, followed by 14% of the genes identified as transcription-related genes, and 7% were classified as playing a role in cell proliferation and cell cycle. Furthermore, 7% of the genes were immune response genes. By 72 h postinfection, genes upregulated by subtype B included the inhibitor of the matrix metalloproteinase TIMP2 and the heat shock protein 40 homolog (Hsp40) DNAJB1, whereas the IFN inducible gene STAT1, the MAPK1/ERK2 kinase regulator ST5, and the chemokine CXCL3 and SHC1 genes were induced by subtypes C and A/E. These analyses distinguish a temporally regulated host response to de novo HIV-1 infection in primary dendritic cells

  5. Maraviroc is associated with latent HIV-1 reactivation through NF-κB activation in resting CD4+ T cells from HIV-Infected Individuals on Suppressive Antiretroviral Therapy.

    Science.gov (United States)

    Madrid-Elena, Nadia; García-Bermejo, María Laura; Serrano-Villar, Sergio; Díaz-de Santiago, Alberto; Sastre, Beatriz; Gutiérrez, Carolina; Dronda, Fernando; Coronel Díaz, María; Domínguez, Ester; López-Huertas, María Rosa; Hernández-Novoa, Beatriz; Moreno, Santiago

    2018-02-14

    Maraviroc is a CCR5 antagonist used in the treatment of HIV-1 infection. We and others have suggested that maraviroc could reactivate latent HIV-1. To test the latency reversing potential of maraviroc and the mechanisms involved, we performed a phase-II, single-center, open-label study in which maraviroc was administered for 10 days to 20 HIV-1-infected individuals on suppressive antiretroviral therapy (Eudra CT: 2012-003215-66). All patients completed full maraviroc dosing and follow up. The primary endpoint was to study whether maraviroc may reactivate HIV-1 latency, eliciting signalling pathways involved in the viral reactivation. An increase in HIV-1 transcription in resting CD4 + T-cells, estimated by HIV-1 unspliced RNA, was observed. Moreover, activation of the NF-κB transcription factor was observed in these cells. In contrast, AP-1 and NFAT activity was not detected. To elucidate the mechanism of NF-κB activation by maraviroc, we have evaluated in HeLa P4 C5 cells, which stably express CCR5, if maraviroc could be acting as a partial CCR5-agonist, with no other mechanisms or pathways involved. Our results show that maraviroc can induce NF-κB activity and NF-κB target genes expression by CCR5 binding, since the use of TAK779, a CCR5 inhibitor, blocked NF-κB activation and functionality. Taken together, we show that maraviroc may have a role in the activation of latent virus transcription through the activation of NF-κB as a result of binding CCR5. Our results strongly support a novel use of maraviroc as a potential latency reversal agent in HIV-1-infected patients. IMPORTANCE HIV-1 persistence in a small pool of long-lived latently infected resting CD4 + T-cells is a major barrier to viral eradication in HIV-1-infected patients on antiretroviral therapy. A potential strategy to cure HIV-1-infection is the use of latency reversing agents to eliminate the reservoirs established in resting CD4 + T-cells. As no drug has been shown to be completely

  6. Epidemiological trends of HIV-1 infection in blood donors from Catalonia, Spain (2005-2014).

    Science.gov (United States)

    Bes, Marta; Piron, Maria; Casamitjana, Natàlia; Gregori, Josep; Esteban, Juan Ignacio; Ribera, Esteban; Quer, Josep; Puig, Lluís; Sauleda, Sílvia

    2017-09-01

    Human immunodeficiency virus 1 (HIV-1) subtype B is predominant in Spain. However, the recent arrival of immigrant populations has increased the prevalence of non-B subtypes and circulating recombinant forms. The objective of this study was to determine the prevalence of HIV-1 subtypes and transmitted drug-resistance mutations in blood donors from the Catalonian region (northeastern Spain). HIV-1-positive blood donors identified in Catalonia from 2005 to 2014 were included. Demographic variables and risk factors for HIV-1 acquisition were recorded. HIV-1 subtyping was carried out by HIV-1 DNA polymerase region sequencing, and phylogenetic analyses were performed using the neighbor-joining method. During the study period, 2.8 million blood donations were screened, and 214 HIV-1-positive donors were identified, yielding an overall prevalence of 7.7 per 100,000 donations (89% men; mean age, 34 ± 10 years). Most HIV-1-positive donors were native to Spain (81%), and 61% were regular blood donors. When risk factors were known, 62% reportedly were men who had sex with men. HIV-1 subtyping was possible in 176 HIV-1-positive individuals: 143 (81%) had HIV-1 subtype B, and 33 (19%) had non-B subtypes. Most HIV-1 non-B subtypes were circulating recombinant forms (n = 20; 61%). Factors associated with HIV-1 subtype B were male sex (p = 0.007) and men who had sex with men (p HIV-1-positive blood donors in Catalonia. Continuous local epidemiological surveillance is required to implement optimal prevention strategies for controlling transfusion-transmitted HIV and to improve health policies regarding HIV infection. © 2017 AABB.

  7. Interdependence of Inhibitor Recognition in HIV-1 Protease.

    Science.gov (United States)

    Paulsen, Janet L; Leidner, Florian; Ragland, Debra A; Kurt Yilmaz, Nese; Schiffer, Celia A

    2017-05-09

    Molecular recognition is a highly interdependent process. Subsite couplings within the active site of proteases are most often revealed through conditional amino acid preferences in substrate recognition. However, the potential effect of these couplings on inhibition and thus inhibitor design is largely unexplored. The present study examines the interdependency of subsites in HIV-1 protease using a focused library of protease inhibitors, to aid in future inhibitor design. Previously a series of darunavir (DRV) analogs was designed to systematically probe the S1' and S2' subsites. Co-crystal structures of these analogs with HIV-1 protease provide the ideal opportunity to probe subsite interdependency. All-atom molecular dynamics simulations starting from these structures were performed and systematically analyzed in terms of atomic fluctuations, intermolecular interactions, and water structure. These analyses reveal that the S1' subsite highly influences other subsites: the extension of the hydrophobic P1' moiety results in 1) reduced van der Waals contacts in the P2' subsite, 2) more variability in the hydrogen bond frequencies with catalytic residues and the flap water, and 3) changes in the occupancy of conserved water sites both proximal and distal to the active site. In addition, one of the monomers in this homodimeric enzyme has atomic fluctuations more highly correlated with DRV than the other monomer. These relationships intricately link the HIV-1 protease subsites and are critical to understanding molecular recognition and inhibitor binding. More broadly, the interdependency of subsite recognition within an active site requires consideration in the selection of chemical moieties in drug design; this strategy is in contrast to what is traditionally done with independent optimization of chemical moieties of an inhibitor.

  8. HIV-1 subtype A gag variability and epitope evolution.

    Science.gov (United States)

    Abidi, Syed Hani; Kalish, Marcia L; Abbas, Farhat; Rowland-Jones, Sarah; Ali, Syed

    2014-01-01

    The aim of this study was to examine the course of time-dependent evolution of HIV-1 subtype A on a global level, especially with respect to the dynamics of immunogenic HIV gag epitopes. We used a total of 1,893 HIV-1 subtype A gag sequences representing a timeline from 1985 through 2010, and 19 different countries in Africa, Europe and Asia. The phylogenetic relationship of subtype A gag and its epidemic dynamics was analysed through a Maximum Likelihood tree and Bayesian Skyline plot, genomic variability was measured in terms of G → A substitutions and Shannon entropy, and the time-dependent evolution of HIV subtype A gag epitopes was examined. Finally, to confirm observations on globally reported HIV subtype A sequences, we analysed the gag epitope data from our Kenyan, Pakistani, and Afghan cohorts, where both cohort-specific gene epitope variability and HLA restriction profiles of gag epitopes were examined. The most recent common ancestor of the HIV subtype A epidemic was estimated to be 1956 ± 1. A period of exponential growth began about 1980 and lasted for approximately 7 years, stabilized for 15 years, declined for 2-3 years, then stabilized again from about 2004. During the course of evolution, a gradual increase in genomic variability was observed that peaked in 2005-2010. We observed that the number of point mutations and novel epitopes in gag also peaked concurrently during 2005-2010. It appears that as the HIV subtype A epidemic spread globally, changing population immunogenetic pressures may have played a role in steering immune-evolution of this subtype in new directions. This trend is apparent in the genomic variability and epitope diversity of HIV-1 subtype A gag sequences.

  9. HIV-1 subtype A gag variability and epitope evolution.

    Directory of Open Access Journals (Sweden)

    Syed Hani Abidi

    Full Text Available OBJECTIVE: The aim of this study was to examine the course of time-dependent evolution of HIV-1 subtype A on a global level, especially with respect to the dynamics of immunogenic HIV gag epitopes. METHODS: We used a total of 1,893 HIV-1 subtype A gag sequences representing a timeline from 1985 through 2010, and 19 different countries in Africa, Europe and Asia. The phylogenetic relationship of subtype A gag and its epidemic dynamics was analysed through a Maximum Likelihood tree and Bayesian Skyline plot, genomic variability was measured in terms of G → A substitutions and Shannon entropy, and the time-dependent evolution of HIV subtype A gag epitopes was examined. Finally, to confirm observations on globally reported HIV subtype A sequences, we analysed the gag epitope data from our Kenyan, Pakistani, and Afghan cohorts, where both cohort-specific gene epitope variability and HLA restriction profiles of gag epitopes were examined. RESULTS: The most recent common ancestor of the HIV subtype A epidemic was estimated to be 1956 ± 1. A period of exponential growth began about 1980 and lasted for approximately 7 years, stabilized for 15 years, declined for 2-3 years, then stabilized again from about 2004. During the course of evolution, a gradual increase in genomic variability was observed that peaked in 2005-2010. We observed that the number of point mutations and novel epitopes in gag also peaked concurrently during 2005-2010. CONCLUSION: It appears that as the HIV subtype A epidemic spread globally, changing population immunogenetic pressures may have played a role in steering immune-evolution of this subtype in new directions. This trend is apparent in the genomic variability and epitope diversity of HIV-1 subtype A gag sequences.

  10. Predicting Bevirimat resistance of HIV-1 from genotype

    Directory of Open Access Journals (Sweden)

    Hoffmann Daniel

    2010-01-01

    Full Text Available Abstract Background Maturation inhibitors are a new class of antiretroviral drugs. Bevirimat (BVM was the first substance in this class of inhibitors entering clinical trials. While the inhibitory function of BVM is well established, the molecular mechanisms of action and resistance are not well understood. It is known that mutations in the regions CS p24/p2 and p2 can cause phenotypic resistance to BVM. We have investigated a set of p24/p2 sequences of HIV-1 of known phenotypic resistance to BVM to test whether BVM resistance can be predicted from sequence, and to identify possible molecular mechanisms of BVM resistance in HIV-1. Results We used artificial neural networks and random forests with different descriptors for the prediction of BVM resistance. Random forests with hydrophobicity as descriptor performed best and classified the sequences with an area under the Receiver Operating Characteristics (ROC curve of 0.93 ± 0.001. For the collected data we find that p2 sequence positions 369 to 376 have the highest impact on resistance, with positions 370 and 372 being particularly important. These findings are in partial agreement with other recent studies. Apart from the complex machine learning models we derived a number of simple rules that predict BVM resistance from sequence with surprising accuracy. According to computational predictions based on the data set used, cleavage sites are usually not shifted by resistance mutations. However, we found that resistance mutations could shorten and weaken the α-helix in p2, which hints at a possible resistance mechanism. Conclusions We found that BVM resistance of HIV-1 can be predicted well from the sequence of the p2 peptide, which may prove useful for personalized therapy if maturation inhibitors reach clinical practice. Results of secondary structure analysis are compatible with a possible route to BVM resistance in which mutations weaken a six-helix bundle discovered in recent experiments

  11. Assessment of phylogenetic sensitivity for reconstructing HIV-1 epidemiological relationships.

    Science.gov (United States)

    Beloukas, Apostolos; Magiorkinis, Emmanouil; Magiorkinis, Gkikas; Zavitsanou, Asimina; Karamitros, Timokratis; Hatzakis, Angelos; Paraskevis, Dimitrios

    2012-06-01

    Phylogenetic analysis has been extensively used as a tool for the reconstruction of epidemiological relations for research or for forensic purposes. It was our objective to assess the sensitivity of different phylogenetic methods and various phylogenetic programs to reconstruct epidemiological links among HIV-1 infected patients that is the probability to reveal a true transmission relationship. Multiple datasets (90) were prepared consisting of HIV-1 sequences in protease (PR) and partial reverse transcriptase (RT) sampled from patients with documented epidemiological relationship (target population), and from unrelated individuals (control population) belonging to the same HIV-1 subtype as the target population. Each dataset varied regarding the number, the geographic origin and the transmission risk groups of the sequences among the control population. Phylogenetic trees were inferred by neighbor-joining (NJ), maximum likelihood heuristics (hML) and Bayesian methods. All clusters of sequences belonging to the target population were correctly reconstructed by NJ and Bayesian methods receiving high bootstrap and posterior probability (PP) support, respectively. On the other hand, TreePuzzle failed to reconstruct or provide significant support for several clusters; high puzzling step support was associated with the inclusion of control sequences from the same geographic area as the target population. In contrary, all clusters were correctly reconstructed by hML as implemented in PhyML 3.0 receiving high bootstrap support. We report that under the conditions of our study, hML using PhyML, NJ and Bayesian methods were the most sensitive for the reconstruction of epidemiological links mostly from sexually infected individuals. Copyright © 2012 Elsevier B.V. All rights reserved.

  12. Virus-producing cells determine the host protein profiles of HIV-1 virion cores

    Science.gov (United States)

    2012-01-01

    Background Upon HIV entry into target cells, viral cores are released and rearranged into reverse transcription complexes (RTCs), which support reverse transcription and also protect and transport viral cDNA to the site of integration. RTCs are composed of viral and cellular proteins that originate from both target and producer cells, the latter entering the target cell within the viral core. However, the proteome of HIV-1 viral cores in the context of the type of producer cells has not yet been characterized. Results We examined the proteomic profiles of the cores purified from HIV-1 NL4-3 virions assembled in Sup-T1 cells (T lymphocytes), PMA and vitamin D3 activated THP1 (model of macrophages, mMΦ), and non-activated THP1 cells (model of monocytes, mMN) and assessed potential involvement of identified proteins in the early stages of infection using gene ontology information and data from genome-wide screens on proteins important for HIV-1 replication. We identified 202 cellular proteins incorporated in the viral cores (T cells: 125, mMΦ: 110, mMN: 90) with the overlap between these sets limited to 42 proteins. The groups of RNA binding (29), DNA binding (17), cytoskeleton (15), cytoskeleton regulation (21), chaperone (18), vesicular trafficking-associated (12) and ubiquitin-proteasome pathway-associated proteins (9) were most numerous. Cores of the virions from SupT1 cells contained twice as many RNA binding proteins as cores of THP1-derived virus, whereas cores of virions from mMΦ and mMN were enriched in components of cytoskeleton and vesicular transport machinery, most probably due to differences in virion assembly pathways between these cells. Spectra of chaperones, cytoskeletal proteins and ubiquitin-proteasome pathway components were similar between viral cores from different cell types, whereas DNA-binding and especially RNA-binding proteins were highly diverse. Western blot analysis showed that within the group of overlapping proteins, the level of

  13. Optical delivery of ARV drugs into HIV-1 permissive cells

    CSIR Research Space (South Africa)

    Khanyile, T

    2013-10-01

    Full Text Available - “Ongoing declines in AIDS incidence and deaths in developed nations, primarily due to widespread use of HAART” (Roger J Pomerantz and David L Horn) AIDS-related deaths, 1995–2011 (World Health Organisation, 2012) Sub-Saharan Africa region: 23....5 mil living with HIV-1 N o . o f p e o p le in m ill io n s 0 500 000 1 000 000 1 500 000 2 000 000 2 500 000 3 000 000 1995 1996 1997 1998 1999 2000 2001 2002 2003 2004 2005 2006 2007 2008 2009 2010 2011 Without antiretroviral therapy...

  14. Prediction of the Secondary Structure of HIV-1 gp120

    DEFF Research Database (Denmark)

    Hansen, Jan; Lund, Ole; Nielsen, Jens O.

    1996-01-01

    Fourier transform infrared spectroscopy. The predicted secondary structure of gp120 compared well with data from NMR analysis of synthetic peptides from the V3 loop and the C4 region. As a first step towards modeling the tertiary structure of gp120, the predicted secondary structure may guide the design......The secondary structure of HIV-1 gp120 was predicted using multiple alignment and a combination of two independent methods based on neural network and nearest-neighbor algorithms. The methods agreed on the secondary structure for 80% of the residues in BH10 gp120. Six helices were predicted in HIV...

  15. Progress and perspectives on HIV-1 microbicide development

    CSIR Research Space (South Africa)

    Alexandre, KB

    2016-10-01

    Full Text Available of in- fected women compared to men. This can be explained by women socio-economic dependence on men, domestic violence, high in- cidence of rape, cultural norms and women inability to negotiate safe sex practices, such as the use of condom... this latter found a 50% increase in the risk of con- tracting HIV-1 in the study group (Statement on results of pro- ducts containing N-9 from the CDC, 2000). The risk was even higher in womenwho reported a reduced use of condom. This was believed to be due...

  16. Cocaine modulates HIV-1 integration in primary CD4+ T cells: implications in HIV-1 pathogenesis in drug-abusing patients

    Science.gov (United States)

    Addai, Amma B.; Pandhare, Jui; Paromov, Victor; Mantri, Chinmay K.; Pratap, Siddharth; Dash, Chandravanu

    2015-01-01

    Epidemiologic studies suggest that cocaine abuse worsens HIV-1 disease progression. Increased viral load has been suggested to play a key role for the accelerated HIV disease among cocaine-abusing patients. The goal of this study was to investigate whether cocaine enhances proviral DNA integration as a mechanism to increase viral load. We infected CD4+ T cells that are the primary targets of HIV-1 in vivo and treated the cells with physiologically relevant concentrations of cocaine (1 µM–100 µM). Proviral DNA integration in the host genome was measured by nested qPCR. Our results illustrated that cocaine from 1 µM through 50 µM increased HIV-1 integration in CD4+ T cells in a dose-dependent manner. As integration can be modulated by several early postentry steps of HIV-1 infection, we examined the direct effects of cocaine on viral integration by in vitro integration assays by use of HIV-1 PICs. Our data illustrated that cocaine directly increases viral DNA integration. Furthermore, our MS analysis showed that cocaine is able to enter CD4+ T cells and localize to the nucleus-. In summary, our data provide strong evidence that cocaine can increase HIV-1 integration in CD4+ T cells. Therefore, we hypothesize that increased HIV-1 integration is a novel mechanism by which cocaine enhances viral load and worsens disease progression in drug-abusing HIV-1 patients. PMID:25691383

  17. Clustering patterns of cytotoxic T-lymphocyte epitopes in human immunodeficiency virus type 1 (HIV-1) proteins reveal imprints of immune evasion on HIV-1 global variation

    DEFF Research Database (Denmark)

    Yusim, K.; Kesmir, Can; Gaschen, B.

    2002-01-01

    The human cytotoxic T-lymphocyte (CTL) response to human immunodeficiency virus type 1 (HIV-1) has been intensely studied, and hundreds of CTL epitopes have been experimentally defined, published, and compiled in the HIV Molecular Immunology Database. Maps of CTL epitopes on HIV-1 protein sequenc...

  18. Rare HIV-1 Subtype J Genomes and a New H/U/CRF02_AG Recombinant Genome Suggests an Ancient Origin of HIV-1 in Angola.

    Science.gov (United States)

    Bártolo, Inês; Calado, Rita; Borrego, Pedro; Leitner, Thomas; Taveira, Nuno

    2016-08-01

    Angola has an extremely diverse HIV-1 epidemic fueled in part by the frequent interchange of people with the Democratic Republic of Congo (DRC) and Republic of Congo (RC). Characterization of HIV-1 strains circulating in Angola should help to better understand the origin of HIV-1 subtypes and recombinant forms and their transmission dynamics. In this study we characterize the first near full-length HIV-1 genomic sequences from HIV-1 infected individuals from Angola. Samples were obtained in 1993 from three HIV-1 infected patients living in Cabinda, Angola. Near full-length genomic sequences were obtained from virus isolates. Maximum likelihood phylogenetic tree inference and analyses of potential recombination patterns were performed to evaluate the sequence classifications and origins. Phylogenetic and recombination analyses revealed that one virus was a pure subtype J, another mostly subtype J with a small uncertain region, and the final virus was classified as a H/U/CRF02_AG recombinant. Consistent with their epidemiological data, the subtype J sequences were more closely related to each other than to other J sequences previously published. Based on the env gene, taxa from Angola occur throughout the global subtype J phylogeny. HIV-1 subtypes J and H are present in Angola at low levels since at least 1993. Low transmission efficiency and/or high recombination potential may explain their limited epidemic success in Angola and worldwide. The high diversity of rare subtypes in Angola suggests that Angola was part of the early establishment of the HIV-1 pandemic.

  19. Effects of nucleic acid local structure and magnesium ions on minus-strand transfer mediated by the nucleic acid chaperone activity of HIV-1 nucleocapsid protein

    OpenAIRE

    Wu, Tiyun; Heilman-Miller, Susan L.; Levin, Judith G.

    2007-01-01

    HIV-1 nucleocapsid protein (NC) is a nucleic acid chaperone, which is required for highly specific and efficient reverse transcription. Here, we demonstrate that local structure of acceptor RNA at a potential nucleation site, rather than overall thermodynamic stability, is a critical determinant for the minus-strand transfer step (annealing of acceptor RNA to (−) strong-stop DNA followed by reverse transcriptase (RT)-catalyzed DNA extension). In our system, destabilization of a stem-loop stru...

  20. Intermolecular masking of the HIV-1 Rev NLS by the cellular protein HIC: novel insights into the regulation of Rev nuclear import.

    LENUS (Irish Health Repository)

    Gu, Lili

    2011-01-01

    The HIV-1 regulatory protein Rev, which is essential for viral replication, mediates the nuclear export of unspliced viral transcripts. Rev nuclear function requires active nucleocytoplasmic shuttling, and Rev nuclear import is mediated by the recognition of its Nuclear Localisation Signal (NLS) by multiple import factors, which include transportin and importin β. However, it remains unclear which nuclear import pathway(s) predominate in vivo, and the cellular environment that modulates Rev nucleocytoplasmic shuttling remains to be characterised.

  1. Trade-off between synergy and efficacy in combinations of HIV-1 latency-reversing agents.

    Science.gov (United States)

    Gupta, Vipul; Dixit, Narendra M

    2018-02-01

    Eradicating HIV-1 infection is difficult because of the reservoir of latently infected cells that gets established soon after infection, remains hidden from antiretroviral drugs and host immune responses, and retains the capacity to reignite infection following the cessation of treatment. Drugs called latency-reversing agents (LRAs) are being developed to reactivate latently infected cells and render them susceptible to viral cytopathicity or immune killing. Whereas individual LRAs have failed to induce adequate reactivation, pairs of LRAs have been identified recently that act synergistically and hugely increase reactivation levels compared to individual LRAs. The maximum synergy achievable with LRA pairs is of clinical importance, as it would allow latency-reversal with minimal drug exposure. Here, we employed stochastic simulations of HIV-1 transcription and translation in latently infected cells to estimate this maximum synergy. We incorporated the predominant mechanisms of action of the two most promising classes of LRAs, namely, protein kinase C agonists and histone deacetylase inhibitors, and quantified the activity of individual LRAs in the two classes by mapping our simulations to corresponding in vitro experiments. Without any adjustable parameters, our simulations then quantitatively captured experimental observations of latency-reversal when the LRAs were used in pairs. Performing simulations representing a wide range of drug concentrations, we estimated the maximum synergy achievable with these LRA pairs. Importantly, we found with all the LRA pairs we considered that concentrations yielding the maximum synergy did not yield the maximum latency-reversal. Increasing concentrations to increase latency-reversal compromised synergy, unravelling a trade-off between synergy and efficacy in LRA combinations. The maximum synergy realizable with LRA pairs would thus be restricted by the desired level of latency-reversal, a constrained optimum we elucidated with

  2. Field evaluation of an open and polyvalent universal HIV-1/SIVcpz/SIVgor quantitative RT-PCR assay for HIV-1 viral load monitoring in comparison to Abbott RealTime HIV-1 in Cameroon.

    Science.gov (United States)

    Guichet, Emilande; Aghokeng, Avelin; Eymard-Duvernay, Sabrina; Vidal, Nicole; Ayouba, Ahidjo; Mpoudi Ngole, Eitel; Delaporte, Eric; Ciaffi, Laura; Peeters, Martine

    2016-11-01

    With the increasing demand of HIV viral load (VL) tests in resource-limited countries (RLCs) there is a need for assays at affordable cost and able to quantify all known HIV-1 variants. VLs obtained with a recently developed open and polyvalent universal HIV-1/SIVcpz/SIVgor RT-qPCR were compared to Abbott RealTime HIV-1 assay in Cameroon. On 474 plasma samples, characterized by a wide range of VLs and a broad HIV-1 group M genetic diversity, 97.5% concordance was observed when using the lower detection limit of each assay. When using the threshold of 3.00 log 10 copies/mL, according to WHO guidelines to define virological failure (VF) in RLCs, the concordance was 94.7%, 360/474 versus 339/474 patients were identified with VF with the new assay and Abbott RealTime HIV-1, respectively. Higher VLs were measured with the new assay, +0.47 log 10 copies/mL (95% CI; 0.42-0.52) as shown with Bland-Altman analysis. Eleven samples from patients on VF with drug resistance were not detected by Abbott RealTime HIV-1 versus two only with the new assay. Overall, our study showed that the new assay can be easily implemented in a laboratory in RLCs with VL experience and showed good performance on a wide diversity of HIV-1 group M variants. Copyright © 2016 Elsevier B.V. All rights reserved.

  3. Endogenous MCM7 microRNA cluster as a novel platform to multiplex small interfering and nucleolar RNAs for combinational HIV-1 gene therapy.

    Science.gov (United States)

    Chung, Janet; Zhang, Jane; Li, Haitang; Ouellet, Dominique L; DiGiusto, David L; Rossi, John J

    2012-11-01

    Combinational therapy with small RNA inhibitory agents against multiple viral targets allows efficient inhibition of viral production by controlling gene expression at critical time points. Here we explore combinations of different classes of therapeutic anti-HIV-1 RNAs expressed from within the context of an intronic MCM7 (minichromosome maintenance complex component-7) platform that naturally harbors 3 microRNAs (miRNAs). We replaced the endogenous miRNAs with anti-HIV small RNAs, including small interfering RNAs (siRNAs) targeting HIV-1 tat and rev messages that function to induce post-transcriptional gene silencing by the RNA interference pathway, a nucleolar-localizing RNA ribozyme that targets the conserved U5 region of HIV-1 transcripts for degradation, and finally nucleolar trans-activation response (TAR) and Rev-binding element (RBE) RNA decoys designed to sequester HIV-1 Tat and Rev proteins inside the nucleolus. We demonstrate the versatility of the MCM7 platform in expressing and efficiently processing the siRNAs as miRNA mimics along with nucleolar small RNAs. Furthermore, three of the combinatorial constructs tested potently suppressed viral replication during a 1-month HIV challenge, with greater than 5-log inhibition compared with untransduced, HIV-1-infected CEM T lymphocytes. One of the most effective constructs contains an anti-HIV siRNA combined with a nucleolar-localizing U5 ribozyme and TAR decoy. This represents the first efficacious example of combining Drosha-processed siRNAs with small nucleolar ribonucleoprotein (snoRNP)-processed nucleolar RNA chimeras from a single intron platform for effective inhibition of viral replication. Moreover, we demonstrated enrichment/selection for cells expressing levels of the antiviral RNAs that provide optimal inhibition under the selective pressure of HIV. The combinations of si/snoRNAs represent a new paradigm for combinatorial RNA-based gene therapy applications.

  4. Multi-step inhibition explains HIV-1 protease inhibitor pharmacodynamics and resistance

    Science.gov (United States)

    Rabi, S. Alireza; Laird, Gregory M.; Durand, Christine M.; Laskey, Sarah; Shan, Liang; Bailey, Justin R.; Chioma, Stanley; Moore, Richard D.; Siliciano, Robert F.

    2013-01-01

    HIV-1 protease inhibitors (PIs) are among the most effective antiretroviral drugs. They are characterized by highly cooperative dose-response curves that are not explained by current pharmacodynamic theory. An unresolved problem affecting the clinical use of PIs is that patients who fail PI-containing regimens often have virus that lacks protease mutations, in apparent violation of fundamental evolutionary theory. Here, we show that these unresolved issues can be explained through analysis of the effects of PIs on distinct steps in the viral life cycle. We found that PIs do not affect virion release from infected cells but block entry, reverse transcription, and post–reverse transcription steps. The overall dose-response curves could be reconstructed by combining the curves for each step using the Bliss independence principle, showing that independent inhibition of multiple distinct steps in the life cycle generates the highly cooperative dose-response curves that make these drugs uniquely effective. Approximately half of the inhibitory potential of PIs is manifest at the entry step, likely reflecting interactions between the uncleaved Gag and the cytoplasmic tail (CT) of the Env protein. Sequence changes in the CT alone, which are ignored in current clinical tests for PI resistance, conferred PI resistance, providing an explanation for PI failure without resistance. PMID:23979165

  5. Calcitonin Gene-Related Peptide Induces HIV-1 Proteasomal Degradation in Mucosal Langerhans Cells.

    Science.gov (United States)

    Bomsel, Morgane; Ganor, Yonatan

    2017-12-01

    The neuroimmune dialogue between peripheral neurons and Langerhans cells (LCs) within mucosal epithelia protects against incoming pathogens. LCs rapidly internalize human immunodeficiency virus type 1 (HIV-1) upon its sexual transmission and then trans -infect CD4 + T cells. We recently found that the neuropeptide calcitonin gene-related peptide (CGRP), secreted mucosally from peripheral neurons, inhibits LC-mediated HIV-1 trans -infection. In this study, we investigated the mechanism of CGRP-induced inhibition, focusing on HIV-1 degradation in LCs and its interplay with trans -infection. We first show that HIV-1 degradation occurs in endolysosomes in untreated LCs, and functionally blocking such degradation with lysosomotropic agents results in increased trans -infection. We demonstrate that CGRP acts via its cognate receptor and at a viral postentry step to induce faster HIV-1 degradation, but without affecting the kinetics of endolysosomal degradation. We reveal that unexpectedly, CGRP shifts HIV-1 degradation from endolysosomes toward the proteasome, providing the first evidence for functional HIV-1 proteasomal degradation in LCs. Such efficient proteasomal degradation significantly inhibits the first phase of trans -infection, and proteasomal, but not endolysosomal, inhibitors abrogate CGRP-induced inhibition. Together, our results establish that CGRP controls the HIV-1 degradation mode in LCs. The presence of endogenous CGRP within innervated mucosal tissues, especially during the sexual response, to which CGRP contributes, suggests that HIV-1 proteasomal degradation predominates in vivo Hence, proteasomal, rather than endolysosomal, HIV-1 degradation in LCs should be enhanced clinically to effectively restrict HIV-1 trans -infection. IMPORTANCE During sexual transmission, HIV-1 is internalized and degraded in LCs, the resident antigen-presenting cells in mucosal epithelia. Yet during trans -infection, infectious virions escaping degradation are transferred

  6. Appreciating HIV-1 diversity: subtypic differences in ENV

    Energy Technology Data Exchange (ETDEWEB)

    Gnanakaran, S [Los Alamos National Laboratory; Shen, Tongye [Los Alamos National Laboratory; Lynch, Rebecca M [NON LANL; Derdeyn, Cynthia A [NON LANL

    2008-01-01

    Human immunodeficiency virus type 1 (HIV-1) group M is responsible for the current AIDS pandemic and exhibits exceedingly high levels of viral genetic diversity around the world, necessitating categorization of viruses into distinct lineages, or subtypes. These subtypes can differ by around 35% in the envelope (Env) glycoproteins of the virus, which are displayed on the surface of the virion and are targets for both neutralizing antibody and cell-mediated immune responses. This diversity reflects the remarkable ability of the virus to adapt to selective pressures, the bulk of which is applied by the host immune response, and represents a serious obstacle for developing an effective vaccine with broad coverage. Thus, it is important to understand the underlying biological consequences of inter-subtype diversity. Recent studies have revealed that the HIV-1 subtypes exhibit phenotypic differences that result from subtle differences in Env structure, particularly within the highly immunogenic V3 domain, which participates directly in viral entry. This review will therefore explore current research that describes subtypic differences in Env at the genetic and phenotypic level, focusing in particular on V3, and highlighting recent discoveries about the unique features of subtype C Env, which is the most prevalent subtype globally.

  7. Developing a Dynamic Pharmacophore Model for HIV-1 Integrase

    International Nuclear Information System (INIS)

    Carlson, Heather A.; Masukawa, Keven M.; Rubins, Kathleen; Bushman, Frederic; Jorgensen, William L.; Lins, Roberto; Briggs, James; Mccammon, Andy

    2000-01-01

    We present the first receptor-based pharmacophore model for HIV-1 integrase. The development of ''dynamic'' pharmacophore models is a new method that accounts for the inherent flexibility of the active site and aims to reduce the entropic penalties associated with binding a ligand. Furthermore, this new drug discovery method overcomes the limitation of an incomplete crystal structure of the target protein. A molecular dynamics (MD) simulation describes the flexibility of the uncomplexed protein. Many conformational models of the protein are saved from the MD simulations and used in a series of multi-unit search for interacting conformers (MUSIC) simulations. MUSIC is a multiple-copy minimization method, available in the BOSS program; it is used to determine binding regions for probe molecules containing functional groups that complement the active site. All protein conformations from the MD are overlaid, and conserved binding regions for the probe molecules are identified. Those conserved binding regions define the dynamic pharmacophore model. Here, the dynamic model is compared to known inhibitors of the integrase as well as a three-point, ligand-based pharmacophore model from the literature. Also, a ''static'' pharmacophore model was determined in the standard fashion, using a single crystal structure. Inhibitors thought to bind in the active site of HIV-1 integrase fit the dynamic model but not the static model. Finally, we have identified a set of compounds from the Available Chemicals Directory that fit the dynamic pharmacophore model, and experimental testing of the compounds has confirmed several new inhibitors

  8. Large-scale functional purification of recombinant HIV-1 capsid.

    Directory of Open Access Journals (Sweden)

    Magdeleine Hung

    Full Text Available During human immunodeficiency virus type-1 (HIV-1 virion maturation, capsid proteins undergo a major rearrangement to form a conical core that protects the viral nucleoprotein complexes. Mutations in the capsid sequence that alter the stability of the capsid core are deleterious to viral infectivity and replication. Recently, capsid assembly has become an attractive target for the development of a new generation of anti-retroviral agents. Drug screening efforts and subsequent structural and mechanistic studies require gram quantities of active, homogeneous and pure protein. Conventional means of laboratory purification of Escherichia coli expressed recombinant capsid protein rely on column chromatography steps that are not amenable to large-scale production. Here we present a function-based purification of wild-type and quadruple mutant capsid proteins, which relies on the inherent propensity of capsid protein to polymerize and depolymerize. This method does not require the packing of sizable chromatography columns and can generate double-digit gram quantities of functionally and biochemically well-behaved proteins with greater than 98% purity. We have used the purified capsid protein to characterize two known assembly inhibitors in our in-house developed polymerization assay and to measure their binding affinities. Our capsid purification procedure provides a robust method for purifying large quantities of a key protein in the HIV-1 life cycle, facilitating identification of the next generation anti-HIV agents.

  9. HIV-1 infection: the role of the gastrointestinal tract.

    Science.gov (United States)

    Cavarelli, Mariangela; Scarlatti, Gabriella

    2014-06-01

    The intestinal mucosa has an important role as portal of entry during mother-to-child transmission of HIV-1 and during sexual transmission. Tissue morphology and integrity, as well as distribution of relevant cell types within the mucosa, spanning from the oropharynx to the rectum, can greatly influence viral infection, replication, presentation, and persistence. The relative contribution to transmission by cell-associated or cell-free virus is still not defined for the different routes of transmission. Although the main target cells for HIV-1 replication are the CD4+ T lymphocytes, which are rapidly depleted both in the periphery and in the mucosal tissues, dendritic cells, Langerhans' cells, and macrophages are players in each of these processes. The predominant cells involved may differ according to the tract of the gut and the route of transmission. The microenvironment of the intestinal mucosa, including mucus, antibodies, or chemo-cytokines, can as well influence infection and replication of the virus: their role is still under investigation. The understanding of these processes may help in developing efficient prevention strategies. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  10. Cellular Antiviral Factors that Target Particle Infectivity of HIV-1.

    Science.gov (United States)

    Goffinet, Christine

    2016-01-01

    In the past decade, the identification and characterization of antiviral genes with the ability to interfere with virus replication has established cell-intrinsic innate immunity as a third line of antiviral defense in addition to adaptive and classical innate immunity. Understanding how cellular factors have evolved to inhibit HIV-1 reveals particularly vulnerable points of the viral replication cycle. Many, but not all, antiviral proteins share type I interferon-upregulated expression and sensitivity to viral counteraction or evasion measures. Whereas well-established restriction factors interfere with early post-entry steps and release of HIV-1, recent research has revealed a diverse set of proteins that reduce the infectious quality of released