scholarly journals The antiviral factor APOBEC3G improves CTL recognition of cultured HIV-infected T cells

2009 ◽  
Vol 207 (1) ◽  
pp. 39-49 ◽  
Author(s):  
Nicoletta Casartelli ◽  
Florence Guivel-Benhassine ◽  
Romain Bouziat ◽  
Samantha Brandler ◽  
Olivier Schwartz ◽  
...  
Keyword(s):  
Cytidine Deaminase ◽  
Adaptive Immune System ◽  
Antiviral Effect ◽  
Inhibitory Effect ◽  
Hiv Replication ◽  
Viral Infectivity Factor ◽  
Cellular Immune ◽  
Vif Protein ◽  
The Impact ◽  
Hiv 1

The cytidine deaminase APOBEC3G (A3G) enzyme exerts an intrinsic anti–human immunodeficiency virus (HIV) defense by introducing lethal G-to-A hypermutations in the viral genome. The HIV-1 viral infectivity factor (Vif) protein triggers degradation of A3G and counteracts this antiviral effect. The impact of A3G on the adaptive cellular immune response has not been characterized. We examined whether A3G-edited defective viruses, which are known to express truncated or misfolded viral proteins, activate HIV-1–specific (HS) CD8+ cytotoxic T lymphocytes (CTLs). To this end, we compared the immunogenicity of cells infected with wild-type or Vif-deleted viruses in the presence or absence of the cytidine deaminase. The inhibitory effect of A3G on HIV replication was associated with a strong activation of cocultivated HS-CTLs. CTL activation was particularly marked with Vif-deleted HIV and with viruses harboring A3G. Enzymatically inactive A3G mutants failed to enhance CTL activation. We also engineered proviruses bearing premature stop codons in their genome as scars of A3G editing. These viruses were not infectious but potently activated HS-CTLs. Therefore, the pool of defective viruses generated by A3G represents an underestimated source of viral antigens. Our results reveal a novel function for A3G, acting not only as an intrinsic antiviral factor but also as an inducer of the adaptive immune system.

scholarly journals Towards Inhibition of Vif-APOBEC3G Interaction: Which Protein to Target?

2010 ◽  
Vol 2010 ◽  
pp. 1-10 ◽  
Author(s):  
Iris Cadima-Couto ◽  
Joao Goncalves
Keyword(s):  
Antiviral Activity ◽  
Protein Binding ◽  
Mechanism Of Action ◽  
Cellular Immunity ◽  
Viral Infectivity ◽  
Binding Partners ◽  
Viral Infectivity Factor ◽  
Vif Protein ◽  
Hiv 1 ◽  
Antiretroviral Activity

APOBEC proteins appeared in the cellular battle against HIV-1 as part of intrinsic cellular immunity. The antiretroviral activity of some of these proteins is overtaken by the action of HIV-1 Viral Infectivity Factor (Vif) protein. Since the discovery of APOBEC3G (A3G) as an antiviral factor, many advances have been made to understand its mechanism of action in the cell and how Vif acts in order to counteract its activity. The mainstream concept is that Vif overcomes the innate antiviral activity of A3G by direct protein binding and promoting its degradation via the cellular ubiquitin/proteasomal pathway. Vif may also inhibit A3G through mechanisms independent of proteasomal degradation. Binding of Vif to A3G is essential for its degradation since disruption of this interaction is predicted to stimulate intracellular antiviral immunity. In this paper we will discuss the different binding partners between both proteins as one of the major challenges for the development of new antiviral drugs.

scholarly journals PF74 Inhibits HIV-1 Integration by Altering the Composition of the Preintegration Complex

2018 ◽  
Vol 93 (6) ◽  
Author(s):  
Muthukumar Balasubramaniam ◽  
Jing Zhou ◽  
Amma Addai ◽  
Phillip Martinez ◽  
Jui Pandhare ◽  
...  
Keyword(s):  
Reverse Transcription ◽  
Integrase Inhibitor ◽  
Antiviral Effect ◽  
Inhibitory Effect ◽  
Clear Understanding ◽  
Nuclear Entry ◽  
Preintegration Complex ◽  
Gradient Based ◽  
Hiv 1

ABSTRACTThe HIV-1 capsid protein (CA) facilitates reverse transcription and nuclear entry of the virus. However, CA’s role in post-nuclear entry steps remains speculative. We describe a direct link between CA and integration by employing the capsid inhibitor PF74 as a probe coupled with the biochemical analysis of HIV-1 preintegration complexes (PICs) isolated from acutely infected cells. At a low micromolar concentration, PF74 potently inhibited HIV-1 infection without affecting reverse transcription. Surprisingly, PF74 markedly reduced proviral integration owing to inhibition of nuclear entry and/or integration. However, a 2-fold reduction in nuclear entry by PF74 did not quantitatively correlate with the level of antiviral activity. Titration of PF74 against the integrase inhibitor raltegravir showed an additive antiviral effect that is dependent on a block at the post-nuclear entry step. PF74’s inhibitory effect was not due to the formation of defective viral DNA ends or a delay in integration, suggesting that the compound inhibits PIC-associated integration activity. Unexpectedly, PICs recovered from cells infected in the presence of PF74 exhibited elevated integration activity. PF74’s effect on PIC activity is CA specific since the compound did not increase the integration activity of PICs of a PF74-resistant HIV-1 CA mutant. Sucrose gradient-based fractionation studies revealed that PICs assembled in the presence of PF74 contained lower levels of CA, suggesting a negative association between CA and PIC-associated integration activity. Finally, the addition of a CA-specific antibody or PF74 inhibited PIC-associated integration activity. Collectively, our results demonstrate that PF74’s targeting of PIC-associated CA results in impaired HIV-1 integration.IMPORTANCEAntiretroviral therapy (ART) that uses various combinations of small molecule inhibitors has been highly effective in controlling HIV. However, the drugs used in the ART regimen are expensive, cause side effects, and face viral resistance. The HIV-1 CA plays critical roles in the virus life cycle and is an attractive therapeutic target. While currently there is no CA-based therapy, highly potent CA-specific inhibitors are being developed as a new class of antivirals. Efforts to develop a CA-targeted therapy can be aided through a clear understanding of the role of CA in HIV-1 infection. CA is well established to coordinate reverse transcription and nuclear entry of the virus. However, the role of CA in post-nuclear entry steps of HIV-1 infection is poorly understood. We show that a CA-specific drug PF74 inhibits HIV-1 integration revealing a novel role of this multifunctional viral protein in a post-nuclear entry step of HIV-1 infection.

scholarly journals The Structural Interface between HIV-1 Vif and Human APOBEC3H

2016 ◽  
Vol 91 (5) ◽  
Author(s):  
Marcel Ooms ◽  
Michael Letko ◽  
Viviana Simon
Keyword(s):  
Binding Site ◽  
Drug Targets ◽  
Cytidine Deaminase ◽  
Interaction Model ◽  
Proteasomal Degradation ◽  
Large Set ◽  
Structure Model ◽  
Contact Points ◽  
Vif Protein ◽  
Hiv 1

ABSTRACT Human APOBEC3H (A3H) is a cytidine deaminase that inhibits HIV-1 replication. To evade this restriction, the HIV-1 Vif protein binds A3H and mediates its proteasomal degradation. To date, little information on the Vif-A3H interface has been available. To decipher how both proteins interact, we first mapped the Vif-binding site on A3H by functionally testing a large set of A3H mutants in single-cycle infectivity and replication assays. Our data show that the two A3H α-helixes α3 and α4 represent the Vif-binding site of A3H. We next used viral adaptation and a set of Vif mutants to identify novel, reciprocal Vif variants that rescued viral infectivity in the presence of two Vif-resistant A3H mutants. These A3H-Vif interaction points were used to generate the first A3H-Vif structure model, which revealed that the A3H helixes α3 and α4 interact with the Vif β-sheet (β2-β5). This model is in good agreement with previously reported Vif and A3H amino acids important for interaction. Based on the predicted A3H-Vif interface, we tested additional points of contact, which validated our model. Moreover, these experiments showed that the A3H and A3G binding sites on HIV-1 Vif are largely distinct, with both host proteins interacting with Vif β-strand 2. Taken together, this virus-host interface model explains previously reported data and will help to identify novel drug targets to combat HIV-1 infection. IMPORTANCE HIV-1 needs to overcome several intracellular restriction factors in order to replicate efficiently. The human APOBEC3 locus encodes seven proteins, of which A3D, A3F, A3G, and A3H restrict HIV-1. HIV encodes the Vif protein, which binds to the APOBEC3 proteins and leads to their proteasomal degradation. No HIV-1 Vif-APOBEC3 costructure exists to date despite extensive research. We and others previously generated HIV-1 Vif costructure models with A3G and A3F by mapping specific contact points between both proteins. Here, we applied a similar approach to HIV-1 Vif and A3H and successfully generated a Vif-A3H interaction model. Importantly, we find that the HIV-1 Vif-A3H interface is distinct from the Vif-A3G and Vif-A3F interfaces, with a small Vif region being important for recognition of both A3G and A3H. Our Vif-A3H structure model informs on how both proteins interact and could guide toward approaches to block the Vif-A3H interface to target HIV replication.

TSAO-T Analogues Bearing Amino Acids at Position N-3 of Thymine: Synthesis and Anti-Human Immunodeficiency Virus Activity

2000 ◽  
Vol 11 (1) ◽  
pp. 61-69 ◽  
Author(s):  
C Chamorro ◽  
E De Clercq ◽  
J Balzarini ◽  
M-J Camarasa ◽  
A San-Félix
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Inhibitory Effect ◽  
Acid Moiety ◽  
Hiv Replication ◽  
Virus Activity ◽  
Immunodeficiency Virus ◽  
Amino Acid Moiety ◽  
Anti Hiv ◽  
Hiv 1

Novel analogues of the anti-HIV-1 lead compound [1-[2‘,5’-bis- O-( tert-butyldimethylsilyl)-β-D-ribofuranosyl]thymine]-3‘-spiro-5’-(4“-amino-1”,2“-oxathiole-2‘,2’-dioxide) (TSAO-T) bearing different amino acids at position N-3 of thymine were prepared and evaluated as inhibitors of HIV replication. The synthesis of the target compounds was accomplished by coupling of the appropriate TSAO intermediate with a conveniently protected (L) amino acid in the presence of BOP and triethylamine, followed by depro-tection of the amino acid moiety. Several TSAO derivatives, bearing at N-3 position of the thymine base an L-amino acid retaining the free carboxylic acid, acquired activity against HIV-2, in addition to their inhibitory effect on HIV-1.

scholarly journals The Impact of HIV-1 Genetic Diversity on CRISPR-Cas9 Antiviral Activity and Viral Escape

Viruses ◽  
2019 ◽  
Vol 11 (3) ◽  
pp. 255 ◽  
Author(s):  
Gilles Darcis ◽  
Caroline Binda ◽  
Bep Klaver ◽  
Elena Herrera-Carrillo ◽  
Ben Berkhout ◽  
...  
Keyword(s):  
Cleavage Site ◽  
Inhibitory Effect ◽  
Viral Escape ◽  
Target Sequence ◽  
Guide Rna ◽  
Gradual Loss ◽  
Minor Sequence ◽  
Viral Target ◽  
The Impact ◽  
Hiv 1

The clustered regularly interspaced short palindromic repeats (CRISPR)-Cas9 system is widely explored for sequence-specific attack on HIV-1 proviral DNA. We recently identified dual-guide RNA (dual-gRNA) combinations that can block HIV-1 replication permanently in infected cell cultures and prevent viral escape. Although the gRNAs were designed to target highly conserved viral sequences, their efficacy may be challenged by high genetic variation in the HIV-1 genome. We therefore evaluated the breadth of these dual-gRNA combinations against distinct HIV-1 isolates, including several subtypes. Replication of nearly all virus isolates could be prevented by at least one gRNA combination, which caused inactivation of the proviral genomes and the gradual loss of replication-competent virus over time. The dual-gRNA efficacy was not affected by most single nucleotide (nt) mismatches between gRNA and the viral target. However, 1-nt mismatches at the Cas9 cleavage site and two mismatches anywhere in the viral target sequence significantly reduced the inhibitory effect. Accordingly, sequence analysis of viruses upon breakthrough replication revealed the acquisition of escape mutations in perfectly matching and most 1-nt mismatching targets, but not in targets with a mismatch at the Cas9 cleavage site or with two mismatches. These results demonstrate that combinatorial CRISPR-Cas9 treatment can cure T cells infected by distinct HIV-1 isolates, but even minor sequence variation in conserved viral target sites can affect the efficacy of this strategy. Successful cure attempts against isolates with divergent target sequences may therefore require adaptation of the gRNAs.

scholarly journals Mechanism of Enhanced HIV Restriction by Virion Coencapsidated Cytidine Deaminases APOBEC3F and APOBEC3G

2016 ◽  
Vol 91 (3) ◽  
Author(s):  
Anjuman Ara ◽  
Robin P. Love ◽  
Tyson B. Follack ◽  
Khawaja A. Ahmed ◽  
Madison B. Adolph ◽  
...  
Keyword(s):  
Size Exclusion ◽  
Minus Strand ◽  
Cytidine Deaminases ◽  
Proviral Dna ◽  
Exclusion Chromatography ◽  
Viral Infectivity Factor ◽  
Biochemical Analyses ◽  
Vif Protein ◽  
Hiv 1

ABSTRACT The APOBEC3 (A3) enzymes, A3G and A3F, are coordinately expressed in CD4+ T cells and can become coencapsidated into HIV-1 virions, primarily in the absence of the viral infectivity factor (Vif). A3F and A3G are deoxycytidine deaminases that inhibit HIV-1 replication by inducing guanine-to-adenine hypermutation through deamination of cytosine to form uracil in minus-strand DNA. The effect of the simultaneous presence of both A3G and A3F on HIV-1 restriction ability is not clear. Here, we used a single-cycle infectivity assay and biochemical analyses to determine if coencapsidated A3G and A3F differ in their restriction capacity from A3G or A3F alone. Proviral DNA sequencing demonstrated that compared to each A3 enzyme alone, A3G and A3F, when combined, had a coordinate effect on hypermutation. Using size exclusion chromatography, rotational anisotropy, and in vitro deamination assays, we demonstrate that A3F promotes A3G deamination activity by forming an A3F/G hetero-oligomer in the absence of RNA which is more efficient at deaminating cytosines. Further, A3F caused the accumulation of shorter reverse transcripts due to decreasing reverse transcriptase efficiency, which would leave single-stranded minus-strand DNA exposed for longer periods of time, enabling more deamination events to occur. Although A3G and A3F are known to function alongside each other, these data provide evidence for an A3F/G hetero-oligomeric A3 with unique properties compared to each individual counterpart. IMPORTANCE The APOBEC3 enzymes APOBEC3F and APOBEC3G act as a barrier to HIV-1 replication in the absence of the HIV-1 Vif protein. After APOBEC3 enzymes are encapsidated into virions, they deaminate cytosines in minus-strand DNA, which forms promutagenic uracils that induce transition mutations or proviral DNA degradation. Even in the presence of Vif, footprints of APOBEC3-catalyzed deaminations are found, demonstrating that APOBEC3s still have discernible activity against HIV-1 in infected individuals. We undertook a study to better understand the activity of coexpressed APOBEC3F and APOBEC3G. The data demonstrate that an APOBEC3F/APOBEC3G hetero-oligomer can form that has unique properties compared to each APOBEC3 alone. This hetero-oligomer has increased efficiency of virus hypermutation, raising the idea that we still may not fully realize the antiviral mechanisms of endogenous APOBEC3 enzymes. Hetero-oligomerization may be a mechanism to increase their antiviral activity in the presence of Vif.

scholarly journals C–C Chemokines Released by Lipopolysaccharide (LPS)-stimulated Human Macrophages Suppress HIV-1 Infection in Both Macrophages and T Cells

1997 ◽  
Vol 185 (5) ◽  
pp. 805-816 ◽  
Author(s):  
Alessia Verani ◽  
Gabriella Scarlatti ◽  
Manola Comar ◽  
Eleonora Tresoldi ◽  
Simona Polo ◽  
...  
Keyword(s):  
T Cells ◽  
Inhibitory Effect ◽  
Hiv Replication ◽  
Soluble Factors ◽  
Human Macrophages ◽  
Immunodeficiency Virus ◽  
Releasing Factors ◽  
Hiv 1 ◽  
Human Immunodeficiency Virus 1

Human immunodeficiency virus-1 (HIV-1) expression in monocyte-derived macrophages (MDM) infected in vitro is known to be inhibited by lipopolysaccharide (LPS). However, the mechanisms are incompletely understood. We show here that HIV-1 suppression is mediated by soluble factors released by MDM stimulated with physiologically significant concentrations of LPS. LPS-conditioned supernatants from MDM inhibited HIV-1 replication in both MDM and T cells. Depletion of C–C chemokines (RANTES, MIP-1α, and MIP-1β) neutralized the ability of LPS-conditioned supernatants to inhibit HIV-1 replication in MDM. A combination of recombinant C–C chemokines blocked HIV-1 infection as effectively as LPS. Here, we report an inhibitory effect of C–C chemokines on HIV replication in primary macrophages. Our results raise the possibility that monocytes may play a dual role in HIV infection: while representing a reservoir for the virus, they may contribute to the containment of the infection by releasing factors that suppress HIV replication not only in monocytes but also in T lymphocytes.

scholarly journals Natural scrub typhus antibody suppresses HIV CXCR4(X4) viruses

2013 ◽  
Vol 5 (1) ◽  
pp. 8 ◽  
Author(s):  
George Watt ◽  
Pacharee Kantipong ◽  
Thierry Burnouf ◽  
Cecilia Shikuma ◽  
Sean Philpott
Keyword(s):  
Scrub Typhus ◽  
Inhibitory Effect ◽  
Cross Reactivity ◽  
Viral Population ◽  
Hiv Replication ◽  
Fusion Inhibitors ◽  
Hiv Gp120 ◽  
Hiv 1

Viral load generally rises in HIV-infected individuals with a concomitant infection, but falls markedly in some individuals with scrub typhus (ST), a common Asian rickettsial infection. ST infection appears to shift the viral population from CXCR4-using (X4) to CCR5-utilizing (R5) strains, and there is evidence of cross-reactivity between ST-specific antibodies and HIV-1. We examined the mechanism of ST suppression of HIV by measuring the effects of ST infection on X4 and R5 viruses <em>in vivo</em> and <em>in vitro</em>, and assessing the relative contributions of antibodies and chemokines to the inhibitory effect. <em>In vivo</em>, a single scrub typhus plasma infusion markedly reduced the subpopulation of HIV-1 viruses using the X4 co-receptor in all 8 recipients, and eliminated X4 viruses 6 patients. <em>In vitro</em>, the 14 ST sera tested all inhibited the replication of an X4 but not an R5 virus. This inhibitory effect was maintained if ST sera were depleted of chemokines but was lost upon removal of antibodies. Sera from ST-infected mice recognized a target that co-localized with X4 HIV gp120 in immunofluorescent experiments. These <em>in vivo </em>and <em>in vitro </em>data suggest that acute ST infection generates cross-reactive antibodies that produce potent suppression of CXCR4- but not CCR5-using HIV-1 viruses. ST suppression of HIV replication could reveal novel mechanisms that could be exploited for vaccination strategies, as well as aid in the development of fusion inhibitors and other new therapeutic regimens. This also appears to be the first instance where one pathogen is neutralized by antibody produced in response to infection by a completely unrelated organism.

scholarly journals Induction of APOBEC3 family proteins, a defensive maneuver underlying interferon-induced anti–HIV-1 activity

2006 ◽  
Vol 203 (1) ◽  
pp. 41-46 ◽  
Author(s):  
Gang Peng ◽  
Ke Jian Lei ◽  
Wenwen Jin ◽  
Teresa Greenwell-Wild ◽  
Sharon M. Wahl
Keyword(s):  
Cytidine Deaminase ◽  
Intracellular Protein ◽  
Mrna Editing ◽  
Hiv Replication ◽  
Potent Inducer ◽  
Virion Infectivity Factor ◽  
Immunodeficiency Virus ◽  
Editing Enzyme ◽  
Anti Hiv ◽  
Hiv 1

Apolipoprotein B mRNA-editing enzyme-catalytic polypeptide-like 3G (APOBEC3G), a cytidine deaminase, is a recently recognized innate intracellular protein with lethal activity against human immunodeficiency virus (HIV). Packaged into progeny virions, APOBEC3G enzymatic activity leads to HIV DNA degradation. As a counterattack, HIV virion infectivity factor (Vif) targets APOBEC3G for proteasomal proteolysis to exclude it from budding virions. Based on the ability of APOBEC3G to antagonize HIV infection, considerable interest hinges on elucidating its mechanism(s) of regulation. In this study, we provide the first evidence that an innate, endogenous host defense factor has the potential to promote APOBEC3G and rebuke the virus-mediated attempt to control its cellular host. We identify interferon (IFN)-α as a potent inducer of APOBEC3G to override HIV Vif neutralization of APOBEC3 proteins that pose a threat to efficient macrophage HIV replication. Our data provide a new dimension by which IFN-α mediates its antiviral activity and suggest a means to render the host nonpermissive for viral replication.

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