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| Status |
Public on Nov 16, 2019 |
| Title |
HIV-CRISPR screen of HIV-1 CA-P90A and CA-N74D mutant viruses with the PIKAHIV library |
| Organisms |
Homo sapiens; Human immunodeficiency virus |
| Experiment type |
Other
|
| Summary |
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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| Overall design |
An HIV-CRISPR screen targeting Interferon-Stimulated Genes in which enrichment of 20bp sgRNA sequences is compared in viral RNA to the library representation of the sgRNA sequences in genomic DNA. 2 biological replicates were performed in ZAP Knockout THP-1 clonal cell lines for all screens, except the HIV-1 CA-P90A mutant screen with IFN and the HIV-1 CA-N74D mutant screen without IFN in which only a single replicate for the viral RNA was performed. 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 M, Emerman M |
| Citation(s) |
32282853 |
| NIH grant(s) |
| Grant ID |
Grant title |
Affiliation |
Name |
| R01 AI147877 |
Discovery and Characterization of Capsid-Targeting Lentiviral Restriction Factors |
FRED HUTCHINSON CANCER RESEARCH CENTER |
Molly OhAinle |
| R01 AI030927 |
HIV Host-Cell Interactions |
FRED HUTCHINSON CANCER RESEARCH CENTER |
Michael Emerman |
| P30 AI027757 |
University of Washington/Fred Hutch Center for AIDS Research |
UNIVERSITY OF WASHINGTON |
Jared Baeten |
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| Submission date |
Nov 15, 2019 |
| Last update date |
May 07, 2020 |
| Contact name |
Molly Ohainle |
| E-mail(s) |
mohainle@fredhutch.org
|
| Organization name |
Fred Hutchinson Cancer Research Center
|
| Street address |
1100 Fairview Ave N, C2-023
|
| 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 (12)
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| Relations |
| BioProject |
PRJNA589859 |
| SRA |
SRP230201 |
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