Similar Datasets
Project description:Genome-wide CRISPR screen identifies TSC1 and DEPDC5 that control HIV-1 latency via the mTOR Signaling pathway
| PRJNA432068 | ENA
Project description:Transcriptional silencing of latent HIV-1 proviruses entails complex and overlapping mechanisms and are a major barrier to in vivo elimination of HIV-1. We developed a new latency CRISPR screening strategy, called Latency HIV-CRISPR, which uses the packaging of guideRNA-encoding lentiviral vector genomes into the supernatant of budding virions as a direct readout of factors involved in the maintenance of HIV-1 latency. We developed a custom guideRNA library targeting epigenetic regulatory genes and paired the screen with and without a latency reversal agent – AZD5582, an activator of the non-canonical NFkB pathway – to examine a combination of mechanisms controlling HIV-1 latency. A component of the Nucleosome Acetyltransferase of H4 histone acetylation (NuA4 HAT) complex, ING3, acts in concert with AZD5582 to activate proviruses in J-Lat cell lines and in a primary CD4+ T cell model of HIV-1 latency. We found that the knockout of ING3 reduces acetylation of the H4 histone tail and BRD4 occupancy on the HIV-1 LTR, and the combination of ING3 knockout with the activation of non-canonical NFkB via AZD5582 act together to dramatically increase initiation and elongation of RNA Polymerase II on the HIV-1 provirus in a manner that is nearly unique among all cellular promoters.
2022-12-26 | GSE213639 | GEO
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.
2022-12-21 | GSE213995 | GEO
Project description:The latent reservoir of HIV-1 presents a major barrier to viral eradication. The mechanism of the establishment and maintenance of the latent viral reservoir is not yet fully understood, which hinders the development of effective curative strategies. In this study, we identified two inhibitory genes, TSC1 and DEPDC5, that maintained HIV-1 latency by suppressing the mTORC1 pathway. We first adapted a genome-wide CRISPR screening approach to identify host factors required for HIV latency in a T-cell-based latency model and discovered two inhibitory genes, TSC1 and DEPDC5, which are potentially involved in HIV-1 latency. Knockout of either TSC1 or DEPDC5 led to enhanced HIV-1 reactivation in both a T-cell line (C11) and a monocyte cell line (U1), and this enhancement could be antagonized by the mTORC1 inhibitor rapamycin. Further evaluation of the mechanism revealed that TSC1 suppresses AKT-mTORC1-S6 via downregulation of Rheb, whereas DEPDC5 inhibits AKT-mTORC1-S6 through RagA. Overall, both TSC1 and DEPDC5 negatively regulate the AKT-mTORC1 pathway, and thus their agonists could be used in the development of new therapeutic approaches for activating HIV-1 latency.
| S-EPMC6081400 | biostudies-literature
Project description:Perturb-seq reveals intrinsic and extrinsic biological factors that regulate gene functions in laten HIV expression.
2024-08-22 | GSE247599 | GEO
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: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.
2023-08-15 | GSE240894 | GEO
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:Resting CD4+ T cells that are both persistent and latently infected with HIV represent the most important challenge to HIV eradication. Estimates indicate the need for >70 years of continuous, fully suppressive, antiretroviral therapy (ART) to eliminate the HIV reservoir. Alternatively, induction of HIV from its latent state could accelerate the decline of the reservoir, thereby shortening the time to eradication. Previous attempts to reactivate latent HIV in preclinical animal models and in clinical trials have measured HIV induction in peripheral blood with minimal focus on tissue reservoirs and had limited effect. We found that activation of the non-canonical NF-B signaling pathway via AZD5582 results in induction of HIV- and SIV-RNA expression in the blood and tissues of ART-suppressed humanized mice and rhesus macaques. Analysis of resting CD4+ T cells from tissues after AZD55852 treatment revealed increased SIV-RNA in lymph nodes in macaques and robust induction of HIV in virtually all tissues analyzed in humanized mice including lymph nodes, thymus, bone marrow, liver, and lung. This promising new approach to latency reversal, in combination with the appropriate tools for systemic clearance of persistent HIV infection, greatly increases opportunities for HIV eradication.
2020-01-07 | GSE141546 | GEO
Project description:Despite the widespread use of anti-retroviral therapy, human immunodeficiency virus (HIV) still persists in an infected cell reservoir that harbors transcriptionally silent yet replication-competent proviruses. While significant progress has been made in understanding how the HIV reservoir is established, transcription repression mechanisms that are enforced on the integrated viral promoter have not been fully revealed. In this study, we performed a whole-genome CRISPR knockout screen in HIV infected T cells to identify host genes that potentially promote HIV latency. Of several top candidates, the KRAB-containing zinc finger protein, ZNF304, was identified as the top hit. ZNF304 silences HIV gene transcription through associating with TRIM28 and recruiting to the viral promoter heterochromatin-inducing methyltransferases, including the polycomb repression complex (PRC) and SETB1. Depletion of ZNF304 expression reduced levels of H3K9me3, H3K27me3 and H2AK119ub repressive histone marks on the HIV promoter as well as SETB1 and TRIM28, ultimately enhancing HIV gene transcription. Significantly, ZNF304 also promoted HIV latency, as its depletion delayed the entry of HIV infected cells into latency. In primary CD4+ cells, ectopic expression of ZNF304 silenced viral transcription. We conclude that by associating with TRIM28 and recruiting host transcriptional repressive complexes, SETB1 and PRC, to the HIV promoter, ZNF304 silences HIV gene transcription and promotes viral latency.
| S-EPMC7529202 | biostudies-literature