scholarly journals The chromatin landscape at the HIV-1 provirus integration site determines viral expression

2020 ◽  
Vol 48 (14) ◽  
pp. 7801-7817 ◽  
Author(s):  
Gerlinde Vansant ◽  
Heng-Chang Chen ◽  
Eduard Zorita ◽  
Katerina Trejbalová ◽  
Dalibor Miklík ◽  
...  
Keyword(s):  
Integration Site ◽  
Treatment Interruption ◽  
Latent Reservoir ◽  
Rna Expression ◽  
Direct Link ◽  
Viral Expression ◽  
Active Transcription ◽  
Provirus Integration Site ◽  
Provirus Integration ◽  
Hiv 1

Abstract HIV-1 persists lifelong in memory cells of the immune system as latent provirus that rebounds upon treatment interruption. Therefore, the latent reservoir is the main target for an HIV cure. Here, we studied the direct link between integration site and transcription using LEDGINs and Barcoded HIV-ensembles (B-HIVE). LEDGINs are antivirals that inhibit the interaction between HIV-1 integrase and the chromatin-tethering factor LEDGF/p75. They were used as a tool to retarget integration, while the effect on HIV expression was measured with B-HIVE. B-HIVE tracks insert-specific HIV expression by tagging a unique barcode in the HIV genome. We confirmed that LEDGINs retarget integration out of gene-dense and actively transcribed regions. The distance to H3K36me3, the marker recognized by LEDGF/p75, clearly increased. LEDGIN treatment reduced viral RNA expression and increased the proportion of silent provirus. Finally, silent proviruses obtained after LEDGIN treatment were located further away from epigenetic marks associated with active transcription. Interestingly, proximity to enhancers stimulated transcription irrespective of LEDGIN treatment, while the distance to H3K36me3 only changed after treatment with LEDGINs. The fact that proximity to these markers are associated with RNA expression support the direct link between provirus integration site and viral expression.

scholarly journals Key Players in HIV-1 Transcriptional Regulation: Targets for a Functional Cure

Viruses ◽  
2020 ◽  
Vol 12 (5) ◽  
pp. 529 ◽  
Author(s):  
Luisa Mori ◽  
Susana T. Valente
Keyword(s):  
Integration Site ◽  
Great Part ◽  
Treatment Interruption ◽  
Host Genome ◽  
Viral Reservoir ◽  
Latent Reservoir ◽  
Functional Cure ◽  
Transcript Elongation ◽  
Rna Transcript ◽  
Hiv 1

HIV-1 establishes a life-long infection when proviral DNA integrates into the host genome. The provirus can then either actively transcribe RNA or enter a latent state, without viral production. The switch between these two states is governed in great part by the viral protein, Tat, which promotes RNA transcript elongation. Latency is also influenced by the availability of host transcription factors, integration site, and the surrounding chromatin environment. The latent reservoir is established in the first few days of infection and serves as the source of viral rebound upon treatment interruption. Despite effective suppression of HIV-1 replication by antiretroviral therapy (ART), to below the detection limit, ART is ineffective at reducing the latent reservoir size. Elimination of this reservoir has become a major goal of the HIV-1 cure field. However, aside from the ideal total HIV-1 eradication from the host genome, an HIV-1 remission or functional cure is probably more realistic. The “block-and-lock” approach aims at the transcriptional silencing of the viral reservoir, to render suppressed HIV-1 promoters extremely difficult to reactivate from latency. There are unfortunately no clinically available HIV-1 specific transcriptional inhibitors. Understanding the mechanisms that regulate latency is expected to provide novel targets to be explored in cure approaches.

scholarly journals Autologous IgG antibodies block outgrowth of a substantial but variable fraction of viruses in the latent reservoir for HIV-1

2020 ◽  
Vol 117 (50) ◽  
pp. 32066-32077
Author(s):  
Lynn N. Bertagnolli ◽  
Joseph Varriale ◽  
Sarah Sweet ◽  
Jacqueline Brockhurst ◽  
Francesco R. Simonetti ◽  
...  
Keyword(s):  
T Cell Activation ◽  
Neutralizing Antibodies ◽  
Cell Activation ◽  
Viral Evolution ◽  
Treatment Interruption ◽  
Latent Reservoir ◽  
Env Protein ◽  
Autologous Antibodies ◽  
Hiv 1

In untreated HIV-1 infection, rapid viral evolution allows escape from immune responses. Viral replication can be blocked by antiretroviral therapy. However, HIV-1 persists in a latent reservoir in resting CD4+ T cells, and rebound viremia occurs following treatment interruption. The reservoir, which is maintained in part by clonal expansion, can be measured using quantitative viral outgrowth assays (QVOAs) in which latency is reversed with T cell activation to allow viral outgrowth. Recent studies have shown that viruses detected in QVOAs prior to treatment interruption often differ from rebound viruses. We hypothesized that autologous neutralizing antibodies directed at the HIV-1 envelope (Env) protein might block outgrowth of some reservoir viruses. We modified the QVOA to reflect pressure from low concentrations of autologous antibodies and showed that outgrowth of a substantial but variable fraction of reservoir viruses is blocked by autologous contemporaneous immunoglobulin G (IgG). A reduction in outgrowth of >80% was seen in 6 of 15 individuals. This effect was due to direct neutralization. We established a phylogenetic relationship between rebound viruses and viruses growing out in vitro in the presence of autologous antibodies. Some large infected cell clones detected by QVOA carried neutralization-sensitive viruses, providing a cogent explanation for differences between rebound virus and viruses detected in standard QVOAs. Measurement of the frequency of reservoir viruses capable of outgrowth in the presence of autologous IgG might allow more accurate prediction of time to viral rebound. Ultimately, therapeutic immunization targeting the subset of variants resistant to autologous IgG might contribute to a functional cure.

scholarly journals Characterization of Intact Proviruses in Blood and Lymph Node from HIV-Infected Individuals Undergoing Analytical Treatment Interruption

2019 ◽  
Vol 93 (8) ◽  
Author(s):  
Line K. Vibholm ◽  
Julio C. C. Lorenzi ◽  
Joy A. Pai ◽  
Yehuda Z. Cohen ◽  
Thiago Y. Oliveira ◽  
...  
Keyword(s):  
T Cells ◽  
Lymph Node ◽  
Treatment Interruption ◽  
Latent Reservoir ◽  
Lymphoid Tissues ◽  
Viral Rebound ◽  
Analytical Treatment ◽  
Latent Hiv ◽  
Hiv 1

ABSTRACT The role of lymphoid tissue as a potential source of HIV-1 rebound following interruption of antiretroviral therapy (ART) is uncertain. To address this issue, we compared the latent viruses obtained from CD4+ T cells in peripheral blood and lymph nodes to viruses emerging during treatment interruption. Latent viruses were characterized by sequencing near-full-length (NFL) proviral DNA and env from viral outgrowth assays (VOAs). Five HIV-1-infected individuals on ART were studied, four of whom participated in a clinical trial of a TLR9 agonist that included an analytical treatment interruption. We found that 98% of intact or replication-competent clonal sequences overlapped between blood and lymph node. In contrast, there was no overlap between 205 latent reservoir and 125 rebound sequences in the four individuals who underwent treatment interruption. However, rebound viruses could be accounted for by recombination. The data suggest that CD4+ T cells carrying latent viruses circulate between blood and lymphoid tissues in individuals on ART and support the idea that recombination may play a role in the emergence of rebound viremia. IMPORTANCE HIV-1 persists as a latent infection in CD4+ T cells that can be found in lymphoid tissues in infected individuals during ART. However, the importance of this tissue reservoir and its contribution to viral rebound upon ART interruption are not clear. In this study, we sought to compare latent HIV-1 from blood and lymph node CD4+ T cells from five HIV-1-infected individuals. Further, we analyzed the contribution of lymph node viruses to viral rebound. We observed that the frequencies of intact proviruses were the same in blood and lymph node. Moreover, expanded clones of T cells bearing identical proviruses were found in blood and lymph node. These latent reservoir sequences did not appear to be the direct origin of rebound virus. Instead, latent proviruses were found to contribute to the rebound compartment by recombination.

scholarly journals Concurrent administration of IFNα14 and cART in TKO-BLT mice enhances suppression of HIV-1 viremia but does not eliminate the latent reservoir

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Kathrin Sutter ◽  
Kerry J. Lavender ◽  
Ronald J. Messer ◽  
Marek Widera ◽  
Katie Williams ◽  
...  
Keyword(s):  
Infection Type ◽  
Treatment Interruption ◽  
Type I Interferons ◽  
Latent Reservoir ◽  
Type I ◽  
Concurrent Administration ◽  
Viral Dna ◽  
Therapy Interruption ◽  
Hiv 1

AbstractCombination antiretroviral therapy (cART) prevents HIV-1 replication but does not eliminate the latent reservoir and cure the infection. Type I interferons (IFN) mediate antiviral effects through different mechanisms than cART. We previously showed that IFNα14 is the most potent IFNα subtype against HIV-1 and that it can significantly reduce the HIV-1 proviral reservoir. This study sought to determine whether combining cART with IFNα14 therapy would produce greater reductions in HIV-1 viral and proviral loads than ART alone. Immunodeficient Rag2−/−γc−/−CD47−/− C57BL/6 mice were humanized by the BLT method, infected with HIV-1JR-CSF and the in vivo efficacy of cART was compared with combined cART/IFNα14 therapy. Infection was allowed to establish for 6 weeks prior to 4 weeks of treatment with oral cART either with or without IFNα14. Plasma viral RNA and splenic CD4+ T cell viral DNA levels were measured immediately after treatment and after 2 weeks of therapy interruption. Augmentation of cART with IFNα14 resulted in significantly enhanced suppression of HIV-1 plasma viremia. However, no significant reduction in total viral DNA was detectable. Furthermore, virus rebounded after treatment interruption to similar levels in both groups. Thus, augmentation of cART with IFNα14 resulted in a more pronounced reduction of HIV viremia levels over cART alone, but the effect was not potent enough to be detected at the viral DNA level or to prevent virus rebound following therapy interruption in immune system-humanized mice.

scholarly journals Characterization of intact proviruses in blood and lymph node from HIV-infected individuals undergoing analytical treatment interruption

2018 ◽  
Author(s):  
Line K. Vibholm ◽  
Julio C.C. Lorenzi ◽  
Joy A. Pai ◽  
Yehuda Z. Cohen ◽  
Thiago Y. Oliveira ◽  
...  
Keyword(s):  
T Cells ◽  
Lymph Node ◽  
Antiretroviral Therapy ◽  
Treatment Interruption ◽  
Latent Reservoir ◽  
Lymphoid Tissues ◽  
Analytical Treatment ◽  
Hiv 1

AbstractThe role of lymphoid tissue as a potential source of HIV-1 rebound following interruption of antiretroviral therapy is uncertain. To address this issue, we compared the latent viruses obtained from CD4+ T cells in peripheral blood and lymph nodes to viruses emerging during treatment interruption. Latent viruses were characterized by sequencing near full-length (NFL) proviral DNA, and env from viral outgrowth cultures (VOAs). 5 HIV-1 infected individuals on antiretroviral therapy (ART) were studied, 4 of whom participated in a clinical trial that included an analytical treatment interruption. Intact or replication competent clonal sequences from blood and lymph node overlapped. In contrast, there was no overlap between 205 latent reservoir and 125 rebound sequences in the 4 individuals who underwent treatment interruption. However, rebound viruses could be accounted for by recombination. The data suggests that CD4+ T cells carrying latent viruses circulate between blood and lymphoid tissues in individuals on ART and support the idea that recombination may play a role in the emergence of rebound viremia.

scholarly journals A point mutation in HIV-1 integrase redirects proviral integration into centromeric repeats

2021 ◽  
Author(s):  
Shelby Winans ◽  
Stephen P. Goff
Keyword(s):  
Viral Replication ◽  
Point Mutation ◽  
Integration Site ◽  
Single Point ◽  
Host Factors ◽  
Latent Reservoir ◽  
Single Point Mutation ◽  
Site Preference ◽  
Chromosomal Dna ◽  
Hiv 1

AbstractRetroviruses utilize the viral integrase (IN) protein to integrate a DNA copy of their genome into the host chromosomal DNA. HIV-1 integration sites are highly biased towards actively transcribed genes, likely mediated by binding of the IN protein to specific host factors, particularly LEDGF, located at these gene regions. We here report a dramatic redirection of integration site distribution induced by a single point mutation in HIV-1 IN. Viruses carrying the K258R IN mutation exhibit more than a 25-fold increase in integrations into centromeric alpha satellite repeat sequences, as assessed by both deep sequencing and qPCR assays. Immunoprecipitation studies identified host factors that uniquely bind to the mutant IN protein and thus may account for the novel bias for integration into centromeres. Centromeric integration events are known to be enriched in the latent reservoir of infected memory T cells, as well as in patients who control viral replication without intervention (so-called elite controllers). The K258R point mutation in HIV-1 IN reported in this study has also been found in databases of latent proviruses found in patients. The altered integration site preference induced by this mutation has uncovered a hidden feature of the establishment of viral latency and control of viral replication.

scholarly journals Analysis of HIV-1 latent reservoir and rebound viruses in a clinical trial of anti-HIV-1 antibody 3BNC117

2018 ◽  
Author(s):  
Yehuda Z. Cohen ◽  
Julio C. C. Lorenzi ◽  
Lisa Krassnig ◽  
John P. Barton ◽  
Leah Burke ◽  
...  
Keyword(s):  
Clinical Trial ◽  
Antiretroviral Therapy ◽  
Dominant Species ◽  
Treatment Interruption ◽  
Latent Reservoir ◽  
Analytical Treatment ◽  
Quantitative Changes ◽  
Anti Hiv ◽  
Hiv 1 ◽  
Amino Acid Variants

AbstractA clinical trial was performed to evaluate 3BNC117, a potent anti_HIV_1 antibody, in infected individuals during suppressive antiretroviral therapy (ART) and subsequent analytical treatment interruption (ATI). The circulating reservoir was evaluated by quantitative and qualitative outgrowth assay (Q2VOA) at entry and after 6 months, prior to ATI. Although there were no significant quantitative changes in the size of the reservoir, the composition of circulating reservoir clones varied over the 6_month period before treatment interruption in a manner that did not correlate with antibody sensitivity. The neutralization profile obtained from the reservoir by Q2VOA was predictive of time to rebound after ATI, and thus of antibody efficacy. Although 3BNC117 binding site amino acid variants found in rebound viruses pre_existed in the latent reservoir, only 3 of 217 rebound viruses were identical to 868 latent viruses. Instead many of the rebound viruses appeared to be recombinants, even in individuals with resistant reservoir viruses. By incorporating the possibility of recombination, 63% of the rebound viruses could have derived from the observed latent reservoir. In conclusion, viruses emerging during ATI in individuals treated with 3BNC117 are not the dominant species found in the circulating reservoir, but instead appear to represent recombinants.SummaryIn the setting of a clinical trial evaluating the anti_HIV_1 antibody 3BNC117, Cohen et al. demonstrate that rebound viruses that emerge following interruption of antiretroviral therapy are distinct from circulating latent viruses. However, rebound viruses often appear to be recombinants between isolated latent viruses.

scholarly journals Models of SIV rebound after treatment interruption that involve multiple reactivation events

2020 ◽  
Author(s):  
Christiaan H. van Dorp ◽  
Jessica M. Conway ◽  
James B. Whitney ◽  
Dan H. Barouch ◽  
Alan S. Perelson
Keyword(s):  
Growth Rate ◽  
Viral Load ◽  
Antiretroviral Therapy ◽  
Treatment Interruption ◽  
Latent Reservoir ◽  
Viral Rebound ◽  
Viral Growth ◽  
Art Initiation ◽  
Reactivation Event ◽  
Hiv 1

AbstractIn order to assess the efficacy of novel HIV-1 treatments leading to a functional cure, the time to viral rebound is frequently used as a surrogate endpoint. The longer the time to viral rebound, the more efficacious the therapy. In support of such an approach, mathematical models serve as a connection between the size of the latent reservoir and the time to HIV-1 rebound after treatment interruption. The simplest of such models assumes that a single successful latent cell reactivation event leads to observable viremia after a period of exponential viral growth. Here we consider a generalization developed by Pinkevych et al. and Hill et al. of this simple model in which multiple reactivation events can occur, each contributing to the exponential growth of the viral load. We formalize and improve the previous derivation of the dynamics predicted by this model, and use the model to estimate relevant biological parameters from SIV rebound data. We confirm a previously described effect of very early antiretroviral therapy (ART) initiation on the rate of recrudescence and the viral load growth rate after treatment interruption. We find that every day ART initiation is delayed results in a 39% increase in the recrudescence rate, and a 11% decrease of the viral growth rate. We show that when viral rebound occurs early relative to the viral load doubling time, a model with multiple successful reactivation events fits the data better than a model with only a single successful reactivation event.Author SummaryHIV-1 persists during suppressive antiretroviral therapy (ART) due to a reservoir of latently infected cells. When ART is stopped, HIV generally rebounds within a few weeks. However, there is a small fraction of patients that do not rebound over a period of months or years. A variety of treatments are being tested for their ability to reduce the size of the latent reservoir, to induce effective immune responses against the virus, or to prevent or prolong the time to viral rebound after ART interruption. These novel treatments are typically first tested in SIV infected macaques, and the efficacy of the treatment assessed by interrupting ART and measuring the time to viral rebound. Here, we develop and test a mathematical and statistical model that describes the process of viral rebound. The model can be used for statistical inference of the efficacy of newly developed treatments. Importantly, the model takes into account that multiple recrudescence events can precede rebound. We test the model using data from early treated SIV infected macaques.

scholarly journals Antiretroviral APOBEC3 Cytidine Deaminases Alter HIV-1 Provirus Integration Site Profiles

2019 ◽  
Author(s):  
Hannah O. Ajoge ◽  
Tyler M. Renner ◽  
Kasandra Bélanger ◽  
Hinissan P. Kohio ◽  
Macon D. Coleman ◽  
...  
Keyword(s):  
Integration Site ◽  
Chromosomal Dna ◽  
Retroviral Infection ◽  
Cytidine Deaminases ◽  
Gene Coding ◽  
Integration Sites ◽  
Site Targeting ◽  
Provirus Integration Site ◽  
Hiv Integration ◽  
Hiv 1

ABSTRACTAPOBEC3 (A3) proteins are host-encoded deoxycytidine deaminases that provide an innate immune barrier to retroviral infection, notably against HIV-1. While the catalytic activity of these proteins can induce catastrophic hypermutation in proviral DNA leading to near-total restriction of infection, sublethal levels of deamination contribute to the genetic evolution of HIV-1. So far, little is known about how A3 might impact HIV-1 integrations into human chromosomal DNA. Using a deep sequencing approach, we analyzed the influence A3F and A3G on HIV-1 integration site selections. DNA editing was detected at the extremities of the long terminal repeat regions of the virus. Both catalytic active and non-catalytic A3 enzymes decreased insertions into gene coding sequences and increased integration sites into SINE elements, oncogenes and transcription-silencing non-B DNA features. Our data implicate A3 as host factors that influence HIV-1 integration site selection and promote insertions into genomic sites that are transcriptionally less active.GRAPHICAL ABSTRACTSchematic depicting the influence of APOBEC3 (A3) proteins on HIV integration site targeting.Left, in the absence of A3, HIV has a strong preference for integrating into genes. Right, both catalytic active and non-catalytic A3 mutants decrease integration into genes and increase integration into SINE elements and in transcription-silencing non-B DNA features.

scholarly journals Expanded cellular clones carrying replication-competent HIV-1 persist, wax, and wane

2018 ◽  
Vol 115 (11) ◽  
pp. E2575-E2584 ◽  
Author(s):  
Zheng Wang ◽  
Evelyn E. Gurule ◽  
Timothy P. Brennan ◽  
Jeffrey M. Gerold ◽  
Kyungyoon J. Kwon ◽  
...  
Keyword(s):  
T Cell ◽  
Cellular Proliferation ◽  
Integration Site ◽  
Virus Production ◽  
Viral Reactivation ◽  
Latent Reservoir ◽  
Major Barrier ◽  
Latently Infected ◽  
Infected Cells ◽  
Hiv 1

The latent reservoir for HIV-1 in resting CD4+ T cells is a major barrier to cure. Several lines of evidence suggest that the latent reservoir is maintained through cellular proliferation. Analysis of this proliferative process is complicated by the fact that most infected cells carry defective proviruses. Additional complications are that stimuli that drive T cell proliferation can also induce virus production from latently infected cells and productively infected cells have a short in vivo half-life. In this ex vivo study, we show that latently infected cells containing replication-competent HIV-1 can proliferate in response to T cell receptor agonists or cytokines that are known to induce homeostatic proliferation and that this can occur without virus production. Some cells that have proliferated in response to these stimuli can survive for 7 d while retaining the ability to produce virus. This finding supports the hypothesis that both antigen-driven and cytokine-induced proliferation may contribute to the stability of the latent reservoir. Sequencing of replication-competent proviruses isolated from patients at different time points confirmed the presence of expanded clones and demonstrated that while some clones harboring replication-competent virus persist longitudinally on a scale of years, others wax and wane. A similar pattern is observed in longitudinal sampling of residual viremia in patients. The observed patterns are not consistent with a continuous, cell-autonomous, proliferative process related to the HIV-1 integration site. The fact that the latent reservoir can be maintained, in part, by cellular proliferation without viral reactivation poses challenges to cure.

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