CARD8 is an inflammasome sensor for HIV-1 protease activity

HIV-1 has high mutation rates and exists as mutant swarms within the host. Rapid evolution of HIV-1 allows the virus to outpace the host immune system, leading to viral persistence. Approaches to targeting immutable components are needed to clear HIV-1 infection. Here, we report that the caspase recruitment domain–containing protein 8 (CARD8) inflammasome senses HIV-1 protease activity. HIV-1 can evade CARD8 sensing because its protease remains inactive in infected cells before viral budding. Premature intracellular activation of the viral protease triggered CARD8 inflammasome–mediated pyroptosis of HIV-1–infected cells. This strategy led to the clearance of latent HIV-1 in patient CD4+ T cells after viral reactivation. Thus, our study identifies CARD8 as an inflammasome sensor of HIV-1, which holds promise as a strategy for the clearance of persistent HIV-1 infection.

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CARD8 inflammasome mediates pyroptosis of HIV-1-infected cells by sensing viral protease activity

10.1101/2020.09.25.308734 ◽

2020 ◽

Author(s):

Qiankun Wang ◽

Hongbo Gao ◽

Kolin M. Clark ◽

Pengfei Tang ◽

Gray H. Harlan ◽

...

Keyword(s):

Rapid Evolution ◽

Mutation Rates ◽

Host Immune System ◽

Virus Reactivation ◽

Viral Protease ◽

Viral Budding ◽

Infected Cells ◽

Latent Hiv ◽

Hiv 1 ◽

Recognition Receptor

AbstractHIV-1 has high mutation rates and exists as mutant swarms in the host. Rapid evolution of HIV-1 allows the virus to outpace host immune system, leading to viral persistence. Novel approaches to target immutable components are needed to clear HIV-1 infection. Here we report a pattern-recognition receptor CARD8 that senses enzymatic activity of the HIV-1 protease, which is indispensable for the virus. All subtypes of HIV-1 can be sensed by CARD8 despite substantial viral diversity. HIV-1 evades CARD8 sensing because the viral protease remains inactive in infected cells prior to viral budding. Induction of premature intracellular activation of the viral protease triggers CARD8 inflammasome-mediated pyroptosis of HIV-1-infected cells. This strategy leads to clearance of latent HIV-1 in patient CD4+ T cells after virus reactivation. Taken together, our study identifies CARD8 as an inflammasome sensor of HIV-1 that holds promise as a strategy for clearance of persistent HIV-1 infection.

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Glycolysis downregulation is a hallmark of HIV-1 latency and sensitizes infected cells to oxidative stress

10.1101/2020.12.30.424810 ◽

2020 ◽

Author(s):

Iart Luca Shytaj ◽

Francesco Andrea Procopio ◽

Mohammad Tarek ◽

Irene Carlon-Andres ◽

Hsin-Yao Tang ◽

...

Keyword(s):

Pentose Phosphate ◽

Viral Reactivation ◽

People Living With Hiv ◽

Cellular Models ◽

Downstream Effectors ◽

Latently Infected ◽

Infected Cells ◽

Latent Hiv ◽

Living With Hiv ◽

Hiv 1

AbstractHIV-1 infects lymphoid and myeloid cells, which can harbor a latent proviral reservoir responsible for maintaining lifelong infection. Glycolytic metabolism has been identified as a determinant of susceptibility to HIV-1 infection, but its role in the development and maintenance of HIV-1 latency has not been elucidated. By combining transcriptomic, proteomic and metabolomic analysis, we here show that transition to latent HIV-1 infection downregulates glycolysis, while viral reactivation by conventional stimuli reverts this effect. Decreased glycolytic output in latently infected cells is associated with downregulation of NAD+/NADH. Consequently, infected cells rely on the parallel pentose phosphate pathway and its main product, the antioxidant NADPH, fueling antioxidant pathways maintaining HIV-1 latency. Of note, blocking NADPH downstream effectors, thioredoxin and glutathione, favors HIV-1 reactivation from latency in lymphoid and myeloid cellular models. This provides a “shock and kill effect” decreasing proviral DNA in cells from people-living-with-HIV/AIDS. Overall, our data show that downmodulation of glycolysis is a metabolic signature of HIV-1 latency that can be exploited to target latently infected cells with eradication strategies.

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Excision of Latent HIV-1 from Infected Cells In Vivo: An Important Step Forward

Molecular Therapy ◽

10.1016/j.ymthe.2017.04.009 ◽

2017 ◽

Vol 25 (5) ◽

pp. 1062-1064 ◽

Cited By ~ 1

Author(s):

Harshana S. De Silva Feelixge ◽

Keith R. Jerome

Keyword(s):

Infected Cells ◽

Latent Hiv ◽

Hiv 1

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The bromodomain and extraterminal domain inhibitor bromosporine synergistically reactivates latent HIV-1 in latently infected cells

Oncotarget ◽

10.18632/oncotarget.21585 ◽

2017 ◽

Vol 8 (55) ◽

pp. 94104-94116 ◽

Cited By ~ 5

Author(s):

Hanyu Pan ◽

Panpan Lu ◽

Yinzhong Shen ◽

Yanan Wang ◽

Zhengtao Jiang ◽

...

Keyword(s):

Latently Infected ◽

Infected Cells ◽

Latent Hiv ◽

Hiv 1

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Human retroviral antisense mRNAs are retained in the nuclei of infected cells for viral persistence

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.2014783118 ◽

2021 ◽

Vol 118 (17) ◽

pp. e2014783118

Author(s):

Guangyong Ma ◽

Jun-ichirou Yasunaga ◽

Kazuya Shimura ◽

Keiko Takemoto ◽

Miho Watanabe ◽

...

Keyword(s):

Viral Persistence ◽

Protein Translation ◽

Regulatory Function ◽

Antisense Gene ◽

Cell Leukemia Virus Type ◽

Antisense Mrna ◽

Human T Cell ◽

Infected Cells ◽

Hiv 1

Human retroviruses, including human T cell leukemia virus type 1 (HTLV-1) and HIV type 1 (HIV-1), encode an antisense gene in the negative strand of the provirus. Besides coding for proteins, the messenger RNAs (mRNAs) of retroviral antisense genes have also been found to regulate transcription directly. Thus, it has been proposed that retroviruses likely localize their antisense mRNAs to the nucleus in order to regulate nuclear events; however, this opposes the coding function of retroviral antisense mRNAs that requires a cytoplasmic localization for protein translation. Here, we provide direct evidence that retroviral antisense mRNAs are localized predominantly in the nuclei of infected cells. The retroviral 3′ LTR induces inefficient polyadenylation and nuclear retention of antisense mRNA. We further reveal that retroviral antisense RNAs retained in the nucleus associate with chromatin and have transcriptional regulatory function. While HTLV-1 antisense mRNA is recruited to the promoter of C-C chemokine receptor type 4 (CCR4) and enhances transcription from it to support the proliferation of HTLV-1–infected cells, HIV-1 antisense mRNA is recruited to the viral LTR and inhibits sense mRNA expression to maintain the latency of HIV-1 infection. In summary, retroviral antisense mRNAs are retained in nucleus, act like long noncoding RNAs instead of mRNAs, and contribute to viral persistence.

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A new small-molecule compound, Q308, silences latent HIV-1 provirus by suppressing Tat- and FACT-mediated transcription

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00470-21 ◽

2021 ◽

Author(s):

Chen-liang Zhou ◽

Yi-fan Huang ◽

Yi-bin Li ◽

Tai-zhen Liang ◽

Teng-yi Zheng ◽

...

Keyword(s):

Small Molecule ◽

Difficult Problem ◽

Viral Reservoir ◽

Functional Cure ◽

Latently Infected ◽

Small Molecule Compound ◽

Infected Cells ◽

Latent Hiv ◽

Anti Hiv ◽

Hiv 1

Eliminating the latent HIV reservoir remains a difficult problem for creating an HIV functional cure or achieving remission. The “block-and-lock” strategy aims to steadily suppress transcription of the viral reservoir and lock the HIV promoter in deep latency using latency-promoting agents (LPAs). However, to date, most of the investigated LPA candidates are not available for clinical trials, and some of them exhibit immune-related adverse reactions. The discovery and development of new, active, and safe LPA candidates for an HIV cure are necessary to eliminate residual HIV-1 viremia through the “block-and-lock” strategy. In this study, we demonstrated that a new small-molecule compound, Q308, silenced the HIV-1 provirus by inhibiting Tat-mediated gene transcription and selectively downregulating the expression levels of the facilitated chromatin transcription (FACT) complex. Strikingly, Q308 induced the preferential apoptosis in HIV-1 latently infected cells, indicating that Q308 may reduce the size of the viral reservoir and thus further prevent viral rebound. These findings highlight that Q308 is a novel and safe anti-HIV-1 inhibitor candidate for a functional cure.

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A CRISPR/Cas9 screen identifies the histone demethylase MINA53 as a novel HIV-1 latency-promoting gene (LPG)

Nucleic Acids Research ◽

10.1093/nar/gkz493 ◽

2019 ◽

Vol 47 (14) ◽

pp. 7333-7347 ◽

Cited By ~ 8

Author(s):

Huachao Huang ◽

Weili Kong ◽

Maxime Jean ◽

Guillaume Fiches ◽

Dawei Zhou ◽

...

Keyword(s):

Local Level ◽

Histone Demethylase ◽

Host Factors ◽

Epigenetic Mark ◽

Combination Antiretroviral Therapy ◽

Downstream Effectors ◽

Infected Cells ◽

Latent Hiv ◽

Hiv 1

AbstractAlthough combination antiretroviral therapy is potent to block active replication of HIV-1 in AIDS patients, HIV-1 persists as transcriptionally inactive proviruses in infected cells. These HIV-1 latent reservoirs remain a major obstacle for clearance of HIV-1. Investigation of host factors regulating HIV-1 latency is critical for developing novel antiretroviral reagents to eliminate HIV-1 latent reservoirs. From our recently accomplished CRISPR/Cas9 sgRNA screens, we identified that the histone demethylase, MINA53, is potentially a novel HIV-1 latency-promoting gene (LPG). We next validated MINA53’s function in maintenance of HIV-1 latency by depleting MINA53 using the alternative RNAi approach. We further identified that in vitro MINA53 preferentially demethylates the histone substrate, H3K36me3 and that in cells MINA53 depletion by RNAi also increases the local level of H3K36me3 at LTR. The effort to map the downstream effectors unraveled that H3K36me3 has the cross-talk with another epigenetic mark H4K16ac, mediated by KAT8 that recognizes the methylated H3K36 and acetylated H4K16. Removing the MINA53-mediated latency mechanisms could benefit the reversal of post-integrated latent HIV-1 proviruses for purging of reservoir cells. We further demonstrated that a pan jumonji histone demethylase inhibitor, JIB-04, inhibits MINA53-mediated demethylation of H3K36me3, and JIB-04 synergizes with other latency-reversing agents (LRAs) to reactivate latent HIV-1.

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Thiostrepton Reactivates Latent HIV-1 through the p-TEFb and NF-κB Pathways Mediated by Heat Shock Response

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.02328-19 ◽

2020 ◽

Vol 64 (5) ◽

Cited By ~ 2

Author(s):

Wen Peng ◽

Zhongsi Hong ◽

Xi Chen ◽

Hongbo Gao ◽

Zhuanglin Dai ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Heat Shock ◽

Viral Reactivation ◽

Immune Recognition ◽

Bryostatin 1 ◽

Functional Cure ◽

Cellular Models ◽

Latent Hiv ◽

Hiv 1

ABSTRACT Antiretroviral therapy (ART) suppresses HIV-1 replication but fails to cure the infection. The presence of an extremely stable viral latent reservoir, primarily in resting memory CD4+ T cells, remains a major obstacle to viral eradication. The “shock and kill” strategy targets these latently infected cells and boosts immune recognition and clearance, and thus, it is a promising approach for an HIV-1 functional cure. Although some latency-reversing agents (LRAs) have been reported, no apparent clinical progress has been made, so it is still vital to seek novel and effective LRAs. Here, we report that thiostrepton (TSR), a proteasome inhibitor, reactivates latent HIV-1 effectively in cellular models and in primary CD4+ T cells from ART-suppressed individuals ex vivo. TSR does not induce global T cell activation, severe cytotoxicity, or CD8+ T cell dysfunction, making it a prospective LRA candidate. We also observed a significant synergistic effect of reactivation when TSR was combined with JQ1, prostratin, or bryostatin-1. Interestingly, six TSR analogues also show reactivation abilities that are similar to or more effective than that of TSR. We further verified that TSR upregulated expression of heat shock proteins (HSPs) in CD4+ T cells, which subsequently activated positive transcriptional elongation factor b (p-TEFb) and NF-κB signals, leading to viral reactivation. In summary, we identify TSR as a novel LRA which could have important significance for applications to an HIV-1 functional cure in the future.

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Epigenetic compound screening uncovers small molecules for re-activation of latent HIV-1

10.1101/2020.08.21.262311 ◽

2020 ◽

Author(s):

Ariane Zutz ◽

Lin Chen ◽

Franziska Sippl ◽

Christian Schölz

Keyword(s):

High Throughput Screening ◽

Model Systems ◽

Reporter System ◽

Compound Screening ◽

Infected Cells ◽

Low Toxicity ◽

Immunological Destruction ◽

New Treatment ◽

Latent Hiv ◽

Hiv 1

AbstractDuring infection with the human immunodeficiency virus type 1 (HIV-1), latent reservoirs are established, which circumvent full eradication of the virus by antiretroviral therapy (ART) and are the source for viral rebound after cessation of therapy. As these reservoirs are phenotypically undistinguishable from infected cells, current strategies aim to reactivate these reservoirs, followed by pharmaceutical and immunological destruction of the cells.Here, we employed a simple and convenient cell-based reporter system, which enables sample handling under biosafety level (BSL)-1 conditions, to screen for compounds that were able to reactivate latent HIV-1. The assay showed a high dynamic signal range and reproducibility with an average Z-factor of 0.77, classifying the system as robust. The assay was used for high-throughput screening (HTS) of an epigenetic compound library in combination with titration and cell-toxicity studies and revealed several potential new latency reversing agents (LRAs). Further validation in well-known latency model systems verified earlier studies and identified two novel compounds with very high reactivation efficiency and low toxicity. Both drugs, namely N-hydroxy-4-(2-[(2-hydroxyethyl)(phenyl)amino]-2-oxoethyl)benzamide (HPOB) and 2',3'-difluoro-[1,1'-biphenyl]-4-carboxylic acid, 2-butylhydrazide (SR-4370), showed comparable performances to other already known LRAs, did not activate CD4+ T-cells or caused changes in the composition of PBMCs as shown by flow cytometry analyses.Both compounds may represent an effective new treatment possibility for revocation of latency in HIV-1 infected individuals.

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