A Virus-Packageable CRISPR Screen Identifies Host Factors Mediating Interferon Inhibition of HIV
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA486290 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR7702340.fastq.gz | Fastqsanger.gz | |||
| SRR7702341.fastq.gz | Fastqsanger.gz | |||
| SRR7702342.fastq.gz | Fastqsanger.gz | |||
| SRR7702343.fastq.gz | Fastqsanger.gz | |||
| SRR7702344.fastq.gz | Fastqsanger.gz |
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Project description:Interferon (IFN) inhibits HIV replication by inducing an array of antiviral effectors. Here we describe a novel CRISPR knockout screening approach to identify the ensemble of these HIV restriction factors. We assembled a CRISPR sgRNA library specific for Interferon Stimulated Genes (ISGs) into a modified lentiviral vector that allows for packaging of sgRNA-encoding genomes in trans into budding HIV-1 particles. We observed that knockout of Zinc Antiviral Protein (ZAP) improved the performance of the screen due to ZAP-mediated inhibition of the vector. We identify a small panel of IFN-induced HIV restriction factors, including MxB, IFITM1, Tetherin/BST2 and TRIM5 which together explain the inhibitory effects of IFN on the HIV-1 LAI strain in THP-1 cells. Further, we identify novel HIV dependency factors, including SEC62 and TLR2. The ability of IFN-induced restriction factors to inhibit an HIV strain to replicate in human cells suggests that these human restriction factors are incompletely antagonized.
2018-08-17 | GSE118631 | GEO
Project description:Interferon (IFN) inhibits HIV replication by inducing antiviral effectors. To comprehensively identify IFN-induced HIV restriction factors, we assembled a CRISPR sgRNA library of Interferon Stimulated Genes (ISGs) into a modified lentiviral vector that allows for packaging of sgRNA-encoding genomes in trans into budding HIV-1 particles. We observed that knockout of Zinc Antiviral Protein (ZAP) improved the performance of the screen due to ZAP-mediated inhibition of the vector. A small panel of IFN-induced HIV restriction factors, including MxB, IFITM1, Tetherin/BST2 and TRIM5alpha together explain the inhibitory effects of IFN on the CXCR4-tropic HIV-1 strain, HIV-1LAI, in THP-1 cells. A second screen with a CCR5-tropic primary strain, HIV-1Q23.BG505, described an overlapping, but non-identical, panel of restriction factors. Further, this screen also identifies HIV dependency factors. The ability of IFN-induced restriction factors to inhibit HIV strains to replicate in human cells suggests that these human restriction factors are incompletely antagonized.Editorial noteThis article has been through an editorial process in which the authors decide how to respond to the issues raised during peer review. The Reviewing Editor's assessment is that all the issues have been addressed (see decision letter).
| S-EPMC6286125 | biostudies-literature
Project description:To identify novel host factors as putative targets to reverse HIV-1 latency, we performed an insertional mutagenesis genetic screen in a latently HIV-1-infected pseudohaploid KBM7 cell line (Hap-Lat). Following mutagenesis, insertions were mapped to the genome, and bioinformatic analysis resulted in the identification of 69 candidate host genes involved in maintaining HIV-1 latency.
2021-11-22 | GSE189314 | GEO
Project description:Type I interferon plays a critical role in the control of viral infections, including HIV-1. Interferon induces a number of restriction factors that block HIV-1 entry, replication and release from the host cell. Currently, systemic treatment of HIV-1 infection with interferon has little efficacy in the clinic due to side effects including fatigue and flu-like symptoms. However, understanding the role of interferon in HIV-1 restriction, and developing molecular tools to generate type I interferons locally, provide an opportunity to inhibit HIV-1 replication while avoiding the side effects associated with systemic administration. Here, we tested a constitutively active inducer of high levels of interferon beta (dLMP1-MAVS). Supernatant from cell transfected with dLMP1-MAVS inhibited HIV-1 replication in both culture cells and primary human CD4+ T cells. CD4+ T cells upregulated a number of known HIV-1 restriction factors, including Viperin, Tetherin, MxB, and ISG56 in response to dLMP1-MAVS. In addition, dLMP1-MAVS activated human dendritic and acted as a molecular adjuvant in a mouse HIV-1 vaccine model. Our study generates new insights into the role of type I interferon in HIV-1 restriction, and provides a novel strategy to induce both type I interferon and anti-HIV-1 immune responses at sites of ongoing viral replication.
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2023-01-26 | GSE223326 | GEO
Project description:To identify host factors that restrict HIV replication, we conducted a CRISPR activation screen in a susceptible T cell line using a high-complexity, genome-wide sgRNA library. Our results identified host factors that conferred protection from HIV infection.
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Project description:HIV is able to outpace the innate immune response, including the response mediated by interferon (IFN), to establish a productive infection. However, monocyte derived macrophages (MDMs) may be protected from HIV infection by treatment with type I IFN before virus exposure. The ability of HIV to modulate the type I IFN-mediated innate immune response when it encounters a cell that has already been exposed to IFN was investigated. To investigate the presence of HIV on an established IFN response, MDMs were subjected to four different conditions: (1) IFN-treated only, (2) IFN-treated followed by HIV infection, (3) HIV infected only, and (4) a mock-treated and mock-infected control. Microarray gene expression analysis was performed on a total of 24 samples derived from the 4 conditions assessed at 3 time points (1, 4 and 8 days following treatment/infection) for both IFN-α2 or -ω. Initially, ISGs were identified as those that were upregulated greater than 2-fold by IFN alone (condition 1) at both Days 4 and 8. Then, the IFN-treated condition was compared to the IFN-treated followed by HIV-infection condition in order to identify those ISGs that were downregulated at least 1.5-fold by the presence of HIV at both days. Assuming that it would be counterproductive for HIV infection by itself to induce the expression of ISGs with putative anti-HIV effects, those ISGs that were upregulated greater than 2-fold in the HIV control were removed. Finally, ISGs that passed these filters and were concordant with both IFN-treatments (IFN-α2 and -ω) were identified and corresponded to the following 8 ISGs: AXL receptor tyrosine kinase (AXL), interferon-alpha inducible protein 27 (IFI27), interferon-induced protein 44 (IFI44), interferon-induced protein 44-like (IFI44L), ISG15, OAS1, OAS3 and XIAP associated factor 1 (XAF1). It should be noted that the IFN-α2 and -ω microarray experiments were performed in different batches but batch effects were not corrected since genes were identified by the filtering approach just described within each batch.
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Project description:Perturb-seq reveals intrinsic and extrinsic biological factors that regulate gene functions in laten HIV expression.
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Project description:We sought to explore the hypothesis that host factors required for HIV-1 replication also play a role in latency reversal. Using a CRISPR gene library of putative HIV dependency factors, we performed a screen to identify genes required for latency reactivation. We identified several HIV- 1 dependency factors that play a key role in HIV-1 latency reactivation including ELL, UBE2M, TBL1XR1, HDAC3, AMBRA1, and ALYREF. Knockout of Cyclin T1 (CCNT1), a component of the P-TEFb complex important for transcription elongation, was the top hit in the screen and had the largest effect on HIV latency reversal with a wide variety of latency reversal agents. Moreover, CCNT1 knockout prevents latency reactivation in a primary CD4+ T cell model of HIV latency without affecting activation of these cells. RNA sequencing data showed that CCNT1 regulates HIV-1 proviral genes to a larger extent than any other host gene and had no significant effects on RNA transcripts in primary T cells after activation. We conclude that CCNT1 function is redundant in T cells but is absolutely required for HIV latency reversal.
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Project description:Illumina sequencing data used in HIV-CRISPR screen with an ISG-specific sgRNA library (PIKAHIV) to find genes that block infection by the N74D and P90A HIV-1 capsid mutant viruses in THP-1 monocytic cells. For more information on the library and approach see Ohainle et al. eLife 2018 (PMID: 30520725). All screens performed here were done in a clonal ZAP-KO cell line (ZAP may inhibit the HIV-CRISPR vector used in the screen). Here we screen the Cyclophilin A-binding deficient mutant P90A and the CPSF6-binding deficient mutant N74D together with a wild type HIV-1 virus. The N74D screen was performed both with IFN treatment and without. These two HIV-1 capsid mutant viruses are hypersensitive to the effects of IFN.
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