Journal
JOURNAL OF VIROLOGY
Volume 96, Issue 13, Pages -Publisher
AMER SOC MICROBIOLOGY
DOI: 10.1128/jvi.00577-22
Keywords
CD4(+) T cells; CRISPR screen; HIV latency; HIV latent reservoir; HIV reactivation; HIV silencing factors; HUSH complex; RNA exosome; RNA processing; SLTM
Categories
Funding
- NIH [R01 AI141009, R61/R33 DA047037, R37 AI147868, R01 DA051906, R01 AI145164, UM1 DA051410, U01 DA053628, CHEETAH P50 AI150464, T32 AI055403]
- NIH REACH Martin Delaney Collaboratory [UM1 AI164565]
- NIH BEAT-HIV Martin Delaney Collaboratory [UM1 AI126620, AI164570]
- Rudolf J. Anderson fellowship
- American Foundation for AIDS Research [amfAR 110029-67-RGRL]
- Yale Gruber fellowship
- Yale College first-year summer research fellowship in the sciences and engineering
- Robert I. Jacobs Fund of the Philadelphia Foundation
- Herbert Kean, M.D., Family Professorship
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This study identified SLTM as a novel host protein that regulates HIV-1 gene expression. Knocking down SLTM can reactivate HIV-1 and induce cell death in infected cells.
Despite effective antiretroviral therapy, HIV-1 persistence in latent reservoirs remains a major obstacle to a cure. We postulate that HIV-1 silencing factors suppress HIV-1 reactivation and that inhibition of these factors will increase HIV-1 reactivation. To identify HIV-1 silencing factors, we conducted a genome-wide CRISPR inhibition (CRISPRi) screen using four CRISPRi-ready, HIV-1-d6-GFP-infected Jurkat T cell clones with distinct integration sites. We sorted cells with increased green fluorescent protein (GFP) expression and captured single guide RNAs (sgRNAs) via targeted deep sequencing. We identified 18 HIV-1 silencing factors that were significantly enriched in HIV-1-d6-GFP(high) cells. Among them, SLTM (scaffold attachment factor B-like transcription modulator) is an epigenetic and transcriptional modulator having both DNA and RNA binding capacities not previously known to affect HIV-1 transcription. Knocking down SLTM by CRISPRi significantly increased HIV-1-d6-GFP expression (by 1.9- to 4.2-fold) in three HIV-1-d6-GFP-Jurkat T cell clones. Furthermore, SLTM knockdown increased the chromatin accessibility of HIV-1 and the gene in which HIV-1 is integrated but not the housekeeping gene POLR2A. To test whether SLTM inhibition can reactivate HIV-1 and further induce cell death of HIV-1-infected cells ex vivo, we established a small interfering RNA (siRNA) knockdown method that reduced SLTM expression in CD4(+) T cells from 10 antiretroviral therapy (ART)-treated, virally suppressed, HIV-1-infected individuals ex vivo. Using limiting dilution culture, we found that SLTM knockdown significantly reduced the frequency of HIV-1-infected cells harboring inducible HIV-1 by 62.2% (0.56/10(6) versus 1.48/10(6) CD4(+) T cells [P = 0.029]). Overall, our study indicates that SLTM inhibition reactivates HIV-1 in vitro and induces cell death of HIV-1-infected cells ex vivo. Our study identified SLTM as a novel therapeutic target. IMPORTANCE HIV-1-infected cells, which can survive drug treatment and immune cell killing, prevent an HIV-1 cure. Immune recognition of infected cells requires HIV-1 protein expression; however, HIV-1 protein expression is limited in infected cells after long-term therapy. The ways in which the HIV-1 provirus is blocked from producing protein are unknown. We identified a new host protein that regulates HIV-1 gene expression. We also provided a new method of studying HIV-1-host factor interactions in cells from infected individuals. These improvements may enable future strategies to reactivate HIV-1 in infected individuals so that infected cells can be killed by immune cells, drug treatment, or the virus itself. HIV-1-infected cells, which can survive drug treatment and immune cell killing, prevent an HIV-1 cure. Immune recognition of infected cells requires HIV-1 protein expression; however, HIV-1 protein expression is limited in infected cells after long-term therapy.
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