Project description:The goal of this CRISPR-based screen (HIV-CRISPR) is to identify HIV-1 dependency factors by evaluating multiple pathways simultaneously. Here are Illumina sequencing data and counts files from HIV-CRISPR screens using a guide RNA library targeting the whole genome (TKOv3), a custom guide RNA library targeting human epigenome genes (HuEpi), a custom guide RNA library targeting interferon-stimulated genes (PIKA), and a custom guide RNA library of genes containing of a subset of each of the aforementioned libraries designed to target human dependency factors (HIVDEP). The HIV-CRISPR screens described here were performed in clonal ZAP knockout Jurkat cell lines as ZAP inhibition of the HIV-CRISPR vector has been previously described (PMID: 30520725).
2023-01-26 | GSE223326 | GEO
Project description:HIV-CRISPR screens with the CD4-ISG library
Project description:The goal of this CRISPR-based screen (Latency HIV-CRISPR) is to identify HIV-1 latency factors by evaluating multiple pathways simultaneously. Here are Illumina sequencing data and counts files from a Latency HIV-CRISPR screen using a custom guide RNA library targeting human epigenome genes (HuEpi). The Latency HIV-CRISPR screens described here were performed in clonal ZAP knockout J-Lat 10.6 (PMID: 12682019) or J-Lat 5A8 (PMID: 24204950) cells lines as ZAP inhibition of the HIV-CRISPR vector has been previously described (PMID: 30520725). The screens were performed in the presence or absence of AZD5582, a SMAC mimetic and latency reversal agent, in order to identify factors that are dependent and independent of this transcriptional activator.
Project description:During the course of HIV infection, host ubiquitination pathways are both attempting to fight off infection and are being hijacked by the virus in an attempt to optimally infect our cells. In order to more fully understand the role ubiquitination plays in HIV infection, we first defined the set of ubiquitin ligases expressed in primary CD4+ T cells using mass spectrometry. Out of the approximately 380 total ligases, we identified 116 that were expressed, and proceeded to delete them individually in primary CD4+ cells using an arrayed CRISPR/Cas9 approach.
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.
Project description:This is data for the evaluation of a new way of counting sgRNAs in CRISPR screens using padlock probes and UMIs. It is compared to the typical PCR-based approach. In particular, a dropout screen was performed in MiaPaCa-2 cells using the Human Kinome CRISPR pooled library (Addgene #75314)
Project description:4T1 mouse tumor cells were screened with a CRISPR library concurrently in vivo in both SCID and immunocompetent mice. Screens were also carried out in vitro.
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.
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.
Project description:All bulk CRISPR based screens CD2 and B2M CRISPRi tiling screens (primary human CD8 T cells), IL2RA CRISPRa tiling screens (Jurkats), CRISPRi/a TF screens (primary human CD8 T cells), and CRISPR TFome KO (primary human T cells)
