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| Status |
Public on Aug 17, 2018 |
| Title |
A Virus-Packageable CRISPR Screen Identifies Host Factors Mediating Interferon Inhibition of HIV |
| Organisms |
Homo sapiens; Human immunodeficiency virus |
| Experiment type |
Other
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| Summary |
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.
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| Overall design |
A CRISPR screen targeting Interferon-Stimulated Genes in which enrichment of sgRNA sequences is compared in viral RNA to the library representation in genomic DNA. 2 biological replicates were performed in 3 indpendent cell lines (one wild type THP-1 and two independent ZAP Knockout THP-1 clonal cell lines). Cells were infected following treatment overnight with IFN and viral supernatants and cell pellets collected 3 days post-infection. sgRNA representation in viral RNA is compared to genomic DNA to determine which sgRNAs are over- or under-represented in the viral population.
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| Contributor(s) |
Ohainle MO, Pendergast L, Vermeire J, Roesch F, Humes D, Basom R, Delrow JJ, Overbaugh J, Emerman M |
| Citation(s) |
30520725 |
| Submission date |
Aug 16, 2018 |
| Last update date |
Mar 27, 2019 |
| Contact name |
Molly Ohainle |
| E-mail(s) |
mohainle@fredhutch.org
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| Organization name |
Fred Hutchinson Cancer Research Center
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| Street address |
1100 Fairview Ave N, C2-023
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| City |
Seattle |
| State/province |
WA |
| ZIP/Postal code |
98109 |
| Country |
USA |
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| Platforms (2) |
| GPL16791 |
Illumina HiSeq 2500 (Homo sapiens) |
| GPL25457 |
Illumina HiSeq 2500 (Human immunodeficiency virus) |
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| Samples (18)
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| Relations |
| BioProject |
PRJNA486290 |
| SRA |
SRP158114 |
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