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Nef dimerization defect abrogates HIV viremia and associated immune dysregulation in the Bone Marrow-Liver-Thymus-Spleen (BLTS) humanized mouse model.

ABSTRACT: Nef dimerization defect abrogates HIV viremia and associated immune dysregulation in the Bone Marrow-Liver-Thymus-Spleen (BLTS) humanized mouse model.

PROVIDER: PRJNA694303 | ENA |

REPOSITORIES: ENA

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Nef dimerization defect abrogates HIV viremia and associated immune dysregulation in the Bone Marrow-Liver-Thymus-Spleen (BLTS) humanized mouse model.

Project description:Loss of function mutations in the human immunodeficiency virus (HIV)-negative factor (Nef) gene are associated with reduced viremia, robust T cell immune response, and delayed acquired immunodeficiency syndrome (AIDS) progression in humans. Additionally, in vitro studies have shown that mutations in Nef's dimerization interface may play a significant role in modulating viral replication and impairing host defense. However, in vivo, mechanistic studies on the role of Nef dimerization in HIV infection are lacking. Humanized rodent models with human immune cells are robust platforms for investigating the interactions between HIV and the human immune system. We recently developed the bone marrow-liver-thymus-spleen (BLTS) humanized mouse model, which carries human immune cells, as well as primary and secondary lymphoid tissues that facilitate anti-viral immune responses. Here, we employed the BLTS-humanized mouse model to demonstrate that preventing Nef dimerization abrogates HIV viremia and the associated induction of immune dysregulation (immune activation and exhaustion). This suggests that Nef dimerization may be a therapeutic target for future HIV cure strategies, which can be explored in humanized mouse models.

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Nef transfection of Jurkat T cells

Project description:Jurkat T cell line was transfected with SIV Nef controlled by an unducible promoter system. Nef was induced by addition of 10 micromolar pronasterone A and gene expression values were determined after 24 hrs. Control Jurkat cells were untransfected and treated with 10 micromolar pronasterone A and gene expressio values were determined after 24 hrs. Keywords: SIV Nef, Jurkat, pronasterone A

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Controlled ErbB receptor dimerization

Project description:In order to better understand signaling events following receptor dimerization involving HER2, we have generated an experimental system in which ErbB dimerization can be controlled. We used gene expression microarrays to identify genes and pathways that are differentially activated by HER2 homodimers and HER2 containing heterodimers. MCF10A-HER2-FKBP-HA cells were treated with AP1510, TGFalpha or heregulin for 48 hrs prior to RNA harvest for array analysis.

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HIV Nef transfected HEK293T cell culture and HIV mock transfected HEK293T cell culture [miRNA]

Project description:Purpose: Comparison of RNA-sequencing datasets obtained from exosomes of Nef-transfected and Mock-transfected HEK293T cells Methods: Assessment of RNA content of exosomes produced by Nef-transfected HEK293T cells and and Mock-transfected HEK293T cells Results: Differences in a set of microRNAs Conclusions: Nef-transfection induces changes in the microRNA content of exosomes

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HIV Nef transfected HEK293T cell culture and HIV mock transfected HEK293T cell culture [mRNA]

Project description:Purpose: Comparison of RNA-sequencing datasets obtained from exosomes of Nef-transfected and Mock-transfected HEK293T cells. Methods: Assessment of RNA content of exosomes produced by Nef-transfected HEK293T cells and and Mock-transfected HEK293T cells. Results: Differences in a set of mRNAs. Conclusions: Nef-transfection might induces changes in the mRNA content of exosomes.

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Characterization of mammalian NEF-sp as a testis-specific nuclear 3′→5′ exoribonuclease

Project description:Ribonucleases catalyse maturation of functional RNAs or mediate degradation of cellular transcripts, activities that are critical for gene expression control. Here we identify a previously uncharacterized mammalian nuclease family member NEF-sp (LOC81691) as a testis-specific factor. Recombinant human NEF-sp demonstrates a divalent metal ion-dependent 3′→5′ exoribonuclease activity. This activity is specific to single-stranded RNA substrates and is independent of their length. Presence of a 2′-O-methyl modification at the 3′ end of the RNA substrate is inhibitory. Ectopically-expressed NEF-sp localizes to the nucleolar/nuclear compartment in human cell cultures and this is dependent on an N-terminal nuclear localization signal. Finally, mice lacking NEF-sp are viable and display normal fertility, likely indicating overlapping functions with other nucleases. Taken together, our study provides the first biochemical and genetic exploration of the role of the NEF-sp exoribonuclease in the mammalian genome.

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Controlled ErbB receptor dimerization

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Gene expression profiling of CD4+ T cell line due to presence of Nef during HIV-1 infection.

Project description:We have utilized microarray technology to compare gene expression profile in presence or absence of Nef during HIV-1 infection in T cell-line. We found several genes significantly differentially regulated due to ∆nef HIV-1 infection and WT HIV-1 infection when compared to Uninfected cells and also several genes significantly dergulated due to WT HIV-1 infection as compared to ∆nef HIV-1 infection. We redistributed genes belonging to several gene ontology into broad categories like “signaling”, “apoptosis”, “transcription” and “lipid metabolism”. We selected some lipid metabolism genes to validate the Nef directed differential expression using quantitative real-time RT-PCR. Thus the present work highlights the importance of Nef directed differential gene expression in milieu of other viral proteins present during infection.

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HIV-1 Nef changes the proteome of T cells extracellular vesicles

Project description:Extracellular vesicles (EVs) are biomolecule carriers for intercellular communication in health and disease. Nef is a human immunodeficiency virus (HIV) virulence factor that is released from cells within EVs and is present in plasma EVs of HIV-1 infected individuals. We performed a quantitative proteomic analysis to fully characterize the Nef-induced changes in protein composition of T cell-derived EVs and identify novel host targets of HIV. Several proteins with well-described roles in infection or not previously associated with HIV pathogenesis were specifically modulated by Nef in EVs. Among the downregulated proteins are the interferon-induced transmembrane 1, 2 and 3 proteins (IFITM1-3), broad-spectrum antiviral factors known to be cell-to-cell transferable by EVs. Our data establish Nef as a modulator of EVs' global protein content and as an HIV factor that antagonizes IFITMs.

2024-01-26 | PXD041028 | Pride

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