The specificity of gene activation was revealed by RNA-seq in the HEK293T cells transfected with the dCasMINI-VPR or dhpCasMINI-VPR system targeting IL1RN [RNA-seq]
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ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE270586 | GEO | 2024/06/29
REPOSITORIES: GEO
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Project description:The specificity of gene activation was revealed by RNA-seq in the HEK293T cells transfected with the dCasMINI-VPR or dhpCasMINI-VPR system targeting IL1RN [RNA-seq]
| PRJNA1127525 | ENA
Project description:Three 293 cell-derivative cell line were constructed using commercially available Invitrogen Flp-In TREx 293 cells. The cell lines express wild type HIV-1 Vpr and two mutant Vprs, F72A/R73A and R80A. The mutant Vprs are defective in the Vpr cell cycle arrest phenotype. The cell lines were characterized for their ability to express Vpr at the RNA and protein levels and functionally for the ability of the inducibly expressed Vprs to arrest cell cycle progression at G2/M. To study whether certain host cell genes showed alterations in expression following induced expression of Vpr, we conducted a large scale host cell gene expression profiling study using microarray technologies. To determine whether the differential expression of certain genes was associated with Vpr-induced cell cycle arrest, we compared the gene expression pattern of wild type Vpr expressing cells with the pattern of Vpr mutant F72A/R73A- or R80A-Vpr expressing cells. Doxycycline-induced and uninduced and Flp-In TREx 293/Vpr cells were collected 0, 1, 2, 4, 6, 8, 12, 16 and 24 hours post induction (hpi). Doxycyclineinduced and uninduced Flp-In TREx 293/Blank cells were also collected as a reference for following experiments. Total RNA was isolated using RNAeasy Midi Kit (QIAGEN), according to the manufacturer's protocol. Human long oligonucleotide microarrays were obtained from the National Cancer Institute Microarray Facility, Advanced Technology Center (Gaithersburg, MD). The microarrays (Hs Operon V2) contained 22,434 oligonucleotide (60-70 mer) spots on a glass slide. 25 µg of total RNAs isolated from doxycycline-induced Flp-In TREx 293 stably transfected blank- and FLAG tagged-Vpr cells were reverse transcribed using an oligo (dT) primer and aminoallyl-dNTP (Amersham Pharmacia). Unincorporated dNTPs were removed with QIAquick PCR purification kit (QIAGEN). Purified cDNA was processed for coupling reaction with Cy 5 or Cy 3-ester dye, respectively. Keywords: other
2006-02-10 | GSE2296 | GEO
Project description:We analyzed the transcriptomic profiles Pepck-Vpr mouse kidney by single-nuclear RNA-seq.
2024-06-12 | GSE223777 | GEO
Project description:Lentiviral accessory genes enhance replication through diverse mechanisms. HIV-1 accessory protein Vpr modulates the host DNA damage response (DDR) at multiple steps through DNA damage, cell cycle arrest, the degradation of host proteins, and both the activation and repression of DDR signaling. Vpr also alters host and viral transcription; however, the connection between Vpr-mediated DDR modulation and transcriptional activation remains unclear. Here, we determined the cellular consequences of Vpr-induced DNA damage using Vpr mutants that allow us to separate the ability of Vpr to induce DNA damage from cell cycle arrest and other DDR phenotypes including host protein degradation and repression of DDR. RNA-sequencing of cells expressing Vpr or Vpr mutants identified that Vpr alters cellular transcription through mechanisms both dependent and independent of cell cycle arrest. In tissue-cultured U2OS cells and primary human monocyte-derived macrophages (MDMs), Vpr-induced DNA damage activates the ATM-NEMO pathway and alters cellular transcription via NF-κB/RelA signaling. HIV-1 infection of primary MDMs validated Vpr-dependent NF-κB transcriptional activation during infection. Both virion delivered and de novo expressed Vpr induced DNA damage and activated ATM-NEMO dependent NF-κB transcription, suggesting that engagement of the DDR and transcriptional reprogramming can occur during early and late stages of viral replication. Together, our data identifies a mechanism by which Vpr activates NF-κB through DNA damage and the ATM-NEMO pathway, which occur independent of cell cycle arrest. We propose this is essential to overcoming restrictive environments, such as in macrophages, to enhance viral transcription and replication.
2024-08-23 | GSE253779 | GEO
Project description:Viruses target host proteins for degradation to enhance their replication and transmission, and identifying these targets has provided key insights into the host-pathogen interaction1-3. Here, we use complementary unbiased mass spectrometry-based approaches to dissect the widespread proteomeic remodelling seen in HIV-1 infected T-cells. Remarkably, the HIV accessory protein Vpr is both necessary and sufficient to cause the vast majority of these changes. Protein regulation requires recruitment of the DCAF1/DDB1/CUL4 E3 ubiquitin ligase complex, and pulsed-Stable Isotope Labelling with Amino Acids in Cell Culture (SILAC) and immunoprecipitation-mass spectrometry (IP-MS) identified at least 38 cellular proteins directly targeted for degradation by Vpr. Whilst other HIV-1 accessory proteins downregulate a small number of specific host factors, Vpr depletes multiple protein targets, causing systems-level changes to the cellular proteome. A subset of the novel cellular targets identified in this study are depleted by Vpr variants from across HIV-1/SIVcpz and other primate lentiviral lineages, confirming their biological importance in vivo.
2019-05-01 | PXD013221 | Pride
Project description:HIV-1 Vpr protein is a multifunctional protein which perturbs human transcriptome and interacts with a number of cellular proteins. In this study, we have attempted to explore the efffects of Vpr on human transcriptome and have identified several genes which are involved in innate immune respone and cell signaling pathways. We used the microarray analysis to elucidate the differnetail expression pattern of differnet genes in human macrophages infected with HIV-1 Vpr. As result we found that HIV-1 Vpr protein leads to the induction of various interferon stimualted genes (ISGs) and chemokines in human macrophages. Human monocytes-derived macrophages (MDMs) were isolated from peripheral blood mononuclear cells (PBMCs) from two healthy donors and were infected with recombinant adenoviruses either expressing HIV-1 Vpr or ZsGreen1 as a control. At 48 hours post-infection, RNA was isolated and subjected to microarray analysis.
2014-04-09 | E-GEOD-56591 | biostudies-arrayexpress
Project description:HIV-1 Vpr protein is a multifunctional protein which perturbs human transcriptome and interacts with a number of cellular proteins. In this study, we have attempted to explore the efffects of Vpr on human transcriptome and have identified several genes which are involved in innate immune responses. We used the microarray analysis to elucidate the differnetail expression pattern of differnet genes in human dendritic cells infected with HIV-1 Vpr. As result we found that HIV-1 Vpr protein leads to the induction of various interferon stimualted genes (ISGs) in human monocyte derived dendritic cells. Human monocytes-derived dendritic cells (MDDCs) were isolated from peripheral blood mononuclear cells (PBMCs) from two healthy donors and were infected with recombinant adenoviruses either expressing HIV-1 Vpr or ZsGreen1 as a control. At 48 hours post-infection, RNA was isolated and subjected to microarray analysis.
2014-09-06 | E-GEOD-61171 | biostudies-arrayexpress
Project description:HIV-1 Vpr protein is a multifunctional protein which perturbs human transcriptome and interacts with a number of cellular proteins. In this study, we have attempted to explore the efffects of Vpr on human transcriptome and have identified several genes which are involved in innate immune respone and cell signaling pathways. We used the microarray analysis to elucidate the differnetail expression pattern of differnet genes in human macrophages infected with HIV-1 Vpr. As result we found that HIV-1 Vpr protein leads to the induction of various interferon stimualted genes (ISGs) and chemokines in human macrophages.
2014-04-09 | GSE56591 | GEO
Project description:HIV-1 Vpr protein is a multifunctional protein which perturbs human transcriptome and interacts with a number of cellular proteins. In this study, we have attempted to explore the efffects of Vpr on human transcriptome and have identified several genes which are involved in innate immune responses. We used the microarray analysis to elucidate the differnetail expression pattern of differnet genes in human dendritic cells infected with HIV-1 Vpr. As result we found that HIV-1 Vpr protein leads to the induction of various interferon stimualted genes (ISGs) in human monocyte derived dendritic cells.
2014-09-06 | GSE61171 | GEO
Project description:The accessory viral protein R (Vpr) is encoded by all primate lentiviruses. Vpr counteracts DNA repair pathways, modulates viral immune sensing, and induces cell-cycle arrest in cell culture. However, its impact in vivo is controversial. Here, we show that deletion of vpr is associated with delayed viral replication kinetics, rapid innate immune activation, development and maintenance of strong B cell responses, and generation of higher avidity p27 Gag antibodies in rhesus macaques. All wild-type SIVmac239 infected animals maintained high viral loads and five of six developed fatal immunodeficiency during ~80 weeks of follow-up. Lack of Vpr was associated with better preservation of CD4+ T cells, lower viral loads and an attenuated clinical course of infection in most animals. Our results demonstrate that Vpr contributes to efficient viral immune evasion and the full pathogenic potential of SIVmac in vivo. Inhibition of Vpr may improve humoral immune control of viral replication.
2023-12-06 | GSE229310 | GEO