Repression of Bax Gene Expression by the HTLV-I Tax Protein: Implications for Suppression of Apoptosis in Virally Infected Cells

Abstract Background Vertebrate hosts limit the availability of iron to microbial pathogens in order to nutritionally starve the invaders. The impact of iron deficiency induced by the iron chelator deferoxamine mesylate (DFO) was investigated in Atlantic salmon SHK-1 cells infected with the facultative intracellular bacterium Piscirickettsia salmonis. Results Effects of the DFO treatment and P. salmonis on SHK-1 cells were gaged by assessing cytopathic effects, bacterial load and activity, and gene expression profiles of eight immune biomarkers at 4- and 7-days post infection (dpi) in the control group, groups receiving single treatments (DFO or P. salmonis) and their combination. The chelator appears to be well-tolerated by host cells, while it had a negative impact on the number of bacterial cells and associated cytotoxicity. DFO alone had minor effects on gene expression of SHK-1 cells, including an early activation of IL-1β at 4 dpi. In contrast to few moderate changes induced by single treatments (either infection or chelator), most genes had highest upregulation in the infected groups receiving DFO. The mildest induction of hepcidin-1 (antimicrobial peptide precursor and regulator of iron homeostasis) was observed in cells exposed to DFO alone, followed by P. salmonis infected cells while the addition of DFO to infected cells further increased the mRNA abundance of this gene. Transcripts encoding TNF-α (immune signaling) and iNOS (immune effector) showed sustained increase at both time points in this group while cathelicidin-1 (immune effector) and IL-8 (immune signaling) were upregulated at 7 dpi. The stimulation of protective gene responses seen in infected cultures supplemented with DFO coincided with the reduction of bacterial load and activity (judged by the expression of P. salmonis 16S rRNA), and damage to cultured host cells. Conclusion The absence of immune gene activation under normal iron conditions suggests modulation of host responses by P. salmonis. The negative effect of iron deficiency on bacteria likely allowed host cells to respond in a more protective manner to the infection, further decreasing its progression. Presented findings encourage in vivo exploration of iron chelators as a promising strategy against piscirickettsiosis.

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Targeting the latent human cytomegalovirus reservoir for T-cell-mediated killing with virus-specific nanobodies

Nature Communications ◽

10.1038/s41467-021-24608-5 ◽

2021 ◽

Vol 12 (1) ◽

Author(s):

Timo W. M. De Groof ◽

Elizabeth G. Elder ◽

Eleanor Y. Lim ◽

Raimond Heukers ◽

Nick D. Bergkamp ◽

...

Keyword(s):

Gene Expression ◽

Human Cytomegalovirus ◽

Early Gene ◽

Latent Reservoir ◽

Solid Organ ◽

Cell Transplant ◽

Viral Receptor ◽

Transplant Patients ◽

Latently Infected ◽

Infected Cells

AbstractLatent human cytomegalovirus (HCMV) infection is characterized by limited gene expression, making latent HCMV infections refractory to current treatments targeting viral replication. However, reactivation of latent HCMV in immunosuppressed solid organ and stem cell transplant patients often results in morbidity. Here, we report the killing of latently infected cells via a virus-specific nanobody (VUN100bv) that partially inhibits signaling of the viral receptor US28. VUN100bv reactivates immediate early gene expression in latently infected cells without inducing virus production. This allows recognition and killing of latently infected monocytes by autologous cytotoxic T lymphocytes from HCMV-seropositive individuals, which could serve as a therapy to reduce the HCMV latent reservoir of transplant patients.

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The Pro-Inflammatory Chemokines CXCL9, CXCL10 and CXCL11 Are Upregulated Following SARS-CoV-2 Infection in an AKT-Dependent Manner

Viruses ◽

10.3390/v13061062 ◽

2021 ◽

Vol 13 (6) ◽

pp. 1062

Author(s):

Victoria Callahan ◽

Seth Hawks ◽

Matthew A. Crawford ◽

Caitlin W. Lehman ◽

Holly A. Morrison ◽

...

Keyword(s):

Gene Expression ◽

Rna Virus ◽

Pulmonary Complications ◽

Lung Epithelial Cells ◽

Dependent Manner ◽

Akt Inhibitor ◽

Inflammatory Chemokines ◽

Inflammatory State ◽

Infected Cells ◽

Ifn Γ

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a highly transmissible RNA virus that is the causative agent of the Coronavirus disease 2019 (COVID-19) pandemic. Patients with severe COVID-19 may develop acute lung injury (ALI) or acute respiratory distress syndrome (ARDS) and require mechanical ventilation. Key features of SARS-CoV-2 induced pulmonary complications include an overexpression of pro-inflammatory chemokines and cytokines that contribute to a ‘cytokine storm.’ In the current study an inflammatory state in Calu-3 human lung epithelial cells was characterized in which significantly elevated transcripts of the immunostimulatory chemokines CXCL9, CXCL10, and CXCL11 were present. Additionally, an increase in gene expression of the cytokines IL-6, TNFα, and IFN-γ was observed. The transcription of CXCL9, CXCL10, IL-6, and IFN-γ was also induced in the lungs of human transgenic angiotensin converting enzyme 2 (ACE2) mice infected with SARS-CoV-2. To elucidate cell signaling pathways responsible for chemokine upregulation in SARS-CoV-2 infected cells, small molecule inhibitors targeting key signaling kinases were used. The induction of CXCL9, CXCL10, and CXCL11 gene expression in response to SARS-CoV-2 infection was markedly reduced by treatment with the AKT inhibitor GSK690693. Samples from COVID-19 positive individuals also displayed marked increases in CXCL9, CXCL10, and CXCL11 transcripts as well as transcripts in the AKT pathway. The current study elucidates potential pathway specific targets for reducing the induction of chemokines that may be contributing to SARS-CoV-2 pathogenesis via hyperinflammation.

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Human T-lymphotropic Virus Type 1-infected Cells Secrete Exosomes That Contain Tax Protein

Journal of Biological Chemistry ◽

10.1074/jbc.m114.549659 ◽

2014 ◽

Vol 289 (32) ◽

pp. 22284-22305 ◽

Cited By ~ 86

Author(s):

Elizabeth Jaworski ◽

Aarthi Narayanan ◽

Rachel Van Duyne ◽

Shabana Shabbeer-Meyering ◽

Sergey Iordanskiy ◽

...

Keyword(s):

Virus Type ◽

T Lymphotropic Virus ◽

Tax Protein ◽

Infected Cells

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Identification of RISC-Associated Adenoviral MicroRNAs, a Subset of Their Direct Targets, and Global Changes in the Targetome upon Lytic Adenovirus 5 Infection

Journal of Virology ◽

10.1128/jvi.02336-14 ◽

2014 ◽

Vol 89 (3) ◽

pp. 1608-1627 ◽

Cited By ~ 20

Author(s):

Florian Bellutti ◽

Maximilian Kauer ◽

Doris Kneidinger ◽

Thomas Lion ◽

Reinhard Klein

Keyword(s):

Gene Expression ◽

Relative Abundance ◽

A549 Cells ◽

Global Changes ◽

Cellular Mirnas ◽

Total Rna ◽

Cellular Mrnas ◽

Comprehensive Picture ◽

Infected Cells ◽

Adenovirus 5

ABSTRACTAdenoviruses encode a set of highly abundant microRNAs (mivaRNAs), which are generated by Dicer-mediated cleavage of the larger noncoding virus-associated RNAs (VA RNAs) I and II. We performed deep RNA sequencing to thoroughly investigate the relative abundance of individual single strands of mivaRNA isoforms in human A549 cells lytically infected with human adenovirus 5 (Ad5) at physiologically relevant multiplicities of infection (MOIs). In addition, we investigated their relative abundance in the endogenous RNA-induced silencing complexes (RISCs). The occupation of endogenous RISCs by mivaRNAs turned out to be pronounced but not as dominant as previously inferred from experiments with AGO2-overexpressing cells infected at high MOIs. In parallel, levels of RISC-incorporated mRNAs were investigated as well. Analysis of mRNAs enriched in RISCs in Ad5-infected cells revealed that only mRNAs with complementarity to the seed sequences of mivaRNAs derived from VA RNAI but not VA RNAII were overrepresented among them, indicating that only mivaRNAs derived from VA RNAI are likely to contribute substantially to the posttranscriptional downregulation of host gene expression. Furthermore, to generate a comprehensive picture of the entire transcriptome/targetome in lytically infected cells, we determined changes in cellular miRNA levels in both total RNA and RISC RNA as well, and bioinformatical analysis of mRNAs of total RNA/RISC fractions revealed a general, genome-wide trend toward detargeting of cellular mRNAs upon infection. Lastly, we identified the direct targets of both single strands of a VA RNAI-derived mivaRNA that constituted one of the two most abundant isoforms in RISCs of lytically infected A549 cells.IMPORTANCEViral and cellular miRNAs have been recognized as important players in virus-host interactions. This work provides the currently most comprehensive picture of the entire mRNA/miRNA transcriptome and of the complete RISC targetome during lytic adenovirus infection and thus represents the basis for a deeper understanding of the interplay between the virus and the cellular RNA interference machinery. Our data suggest that, at least in the model system that was employed, lytic infection by Ad5 is accompanied by a measurable global net detargeting effect on cellular mRNAs, and analysis of RISC-associated viral small RNAs revealed that the VA RNAs are the only source of virus-encoded miRNAs. Moreover, this work allows to assess the power of individual viral miRNAs to regulate cellular gene expression and provides a list of proven and putative direct targets of these miRNAs, which is of importance, given the fact that information about validated targets of adenovirus-encoded miRNAs is scarce.

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Use of a Beet necrotic yellow vein virus RNA-5-derived replicon as a new tool for gene expression

Journal of General Virology ◽

10.1099/vir.0.80720-0 ◽

2005 ◽

Vol 86 (2) ◽

pp. 463-467 ◽

Cited By ~ 19

Author(s):

Laure Schmidlin ◽

Didier Link ◽

Jérôme Mutterer ◽

Hubert Guilley ◽

David Gilmer

Keyword(s):

Gene Expression ◽

Recombinant Proteins ◽

Expression System ◽

Green Fluorescence Protein ◽

Yellow Vein ◽

Expression Levels ◽

Virus Rna ◽

New Gene ◽

Infected Cells ◽

Fluorescence Protein

A new gene-expression system based on RNA-5 of Beet necrotic yellow vein virus (BNYVV) was constructed to allow the expression of recombinant proteins in virally infected cells. Replication and expression levels of the RNA-5-based replicon containing the green fluorescence protein (GFP) gene were compared with those obtained with the well-characterized RNA-3-derived replicon (Rep-3). When RNA-3 and/or RNA-4 BNYVV RNAs were added to the inoculum, the expression levels of RNA-5-encoded GFP were considerably reduced. To a lesser extent, RNA-3-derived GFP expression was also affected by the presence of RNA-4 and -5. Both RNA-3- and RNA-5-derived molecules were able to express proteins within the same infected cells. Together with Rep-3, the RNA-5-derived replicon thus provides a new tool for the co-expression of different recombinant proteins. In Beta macrocarpa, Rep-5-GFP was able to move in systemic tissues in the presence of RNA-3 and thus provides a new expression system that is not restricted to the inoculated leaves.

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HIV-1 Infection Transcriptomics: Meta-Analysis of CD4+ T Cells Gene Expression Profiles

Viruses ◽

10.3390/v13020244 ◽

2021 ◽

Vol 13 (2) ◽

pp. 244 ◽

Cited By ~ 1

Author(s):

Antonio Victor Campos Coelho ◽

Rossella Gratton ◽

João Paulo Britto de Melo ◽

José Leandro Andrade-Santos ◽

Rafael Lima Guimarães ◽

...

Keyword(s):

Gene Expression ◽

T Cells ◽

Differentially Expressed Genes ◽

Cd4 T Cells ◽

Expression Profiles ◽

Meta Analysis ◽

Differentially Expressed ◽

Rna Seq ◽

Infected Cells ◽

Hiv 1

HIV-1 infection elicits a complex dynamic of the expression various host genes. High throughput sequencing added an expressive amount of information regarding HIV-1 infections and pathogenesis. RNA sequencing (RNA-Seq) is currently the tool of choice to investigate gene expression in a several range of experimental setting. This study aims at performing a meta-analysis of RNA-Seq expression profiles in samples of HIV-1 infected CD4+ T cells compared to uninfected cells to assess consistently differentially expressed genes in the context of HIV-1 infection. We selected two studies (22 samples: 15 experimentally infected and 7 mock-infected). We found 208 differentially expressed genes in infected cells when compared to uninfected/mock-infected cells. This result had moderate overlap when compared to previous studies of HIV-1 infection transcriptomics, but we identified 64 genes already known to interact with HIV-1 according to the HIV-1 Human Interaction Database. A gene ontology (GO) analysis revealed enrichment of several pathways involved in immune response, cell adhesion, cell migration, inflammation, apoptosis, Wnt, Notch and ERK/MAPK signaling.

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The gammaherpesvirus 68 viral cyclin facilitates reactivation by promoting latent gene expression

10.1101/761700 ◽

2019 ◽

Author(s):

Brian F Niemeyer ◽

Joy E Gibson ◽

Jennifer N Berger ◽

Lauren M Oko ◽

Eva Medina ◽

...

Keyword(s):

Gene Expression ◽

Critical Role ◽

Productive Infection ◽

Viral Gene ◽

Cellular Distribution ◽

Viral Cyclin ◽

Latently Infected ◽

Infected Cells ◽

Virally Encoded ◽

The Impact

AbstractGammaherpesviruses establish life-long infections within their host and have been shown to be the causative agents of devastating malignancies. Chronic infection within the host is mediated through cycles of transcriptionally quiescent stages of latency with periods of reactivation into more active lytic and productive infection. The mechanisms that regulate reactivation from latency remain poorly understood. Previously, we defined a critical role for the viral cyclin in promoting reactivation from latency. Disruption of the viral cyclin had no impact on the frequency of cells containing viral genome during latency, yet it remains unclear whether the viral cyclin influences latently infected cells in a qualitative manner. To define the impact of the viral cyclin on latent gene expression, we utilized a viral cyclin deficient variant expressing a LANA-beta-lactamase fusion protein (LANA::βla), to enumerate both the cellular distribution and frequency of latent gene expression. Disruption of the viral cyclin did not affect the cellular distribution of latently infected cells, but did result in a significant decrease in the frequency of cells that expressed LANA::βla across multiple tissues and in both immunocompetent and immunodeficient hosts. Strikingly, whereas the cyclin-deficient virus had a reactivation defect in bulk culture, sort purified cyclin-deficient LANA::βla expressing cells were fully capable of reactivation. These data emphasize that the γHV68 latent reservoir is comprised of at least two distinct stages of infection characterized by differential latent gene expression, and that a primary function of the viral cyclin is to promote latent gene expression within infected cells in vivo.AUTHOR SUMMARYGammaherpesviruses are ubiquitous viruses with oncogenic potential that establish latency for the life of the host. These viruses can emerge from latency through reactivation, a process that is controlled by the immune system. Control of viral latency and reactivation is thought to be critical to prevent γHV-associated disease. This study focuses on a virally-encoded cyclin that is required for reactivation from latency. By characterizing how the viral cyclin influences latent infection in pure cell populations, we find that the viral cyclin has a vital role in promoting viral gene expression during latency. This work provides new insight into the function of a virally encoded cyclin in promoting reactivation from latency.

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Tax induces the recruitment of NF-kB to unintegrated HIV-1 DNA to rescue viral gene expression and replication

10.1101/2021.02.16.431555 ◽

2021 ◽

Author(s):

Ishak D. Irwan ◽

Bryan R. Cullen

Keyword(s):

Gene Expression ◽

T Cells ◽

Lentiviral Vectors ◽

Expression Vectors ◽

Viral Gene ◽

Epigenetic Marks ◽

Tax Protein ◽

Human T Cell ◽

Activation Of Transcription ◽

Hiv 1

AbstractWe have previously reported that the normally essential step of integration of the HIV-1 proviral DNA intermediate into the host cell genome becomes dispensable in T cells that express the Human T cell leukemia virus 1 (HTLV-1) Tax protein. The rescue of integrase (IN) deficient HIV-1 replication by Tax results from the strong activation of transcription from the long terminal repeat (LTR) promoter on episomal HIV-1 DNA, an effect that is closely correlated with the recruitment of activating epigenetic marks, such as H3Ac, and depletion of repressive epigenetic marks, such as H3K9me3, from chromatinized unintegrated proviruses. In addition, activation of transcription from unintegrated HIV-1 DNA coincides with the recruitment of NF-kB to the two NF-kB binding sites found in the HIV-1 LTR enhancer. Here we report that the recruitment of NF-kB to unintegrated viral DNA precedes, and is a prerequisite for, Tax-induced changes in epigenetic marks, so that an IN-HIV-1 mutant lacking both LTR NF-kB sites is entirely non-responsive to Tax and fails to undergo the epigenetic changes listed above. We also report that heterologous promoters introduced into IN-HIV-1-based vectors are transcriptionally active even in the absence of Tax. Finally, we failed to reproduce a recent report arguing that heterologous promoters introduced into IN-vectors based on HIV-1 are more active if the HIV-1 promoter and enhancer, located in the LTR U3 region, are deleted, in a so-called self inactivating or SIN lentivector design.ImportanceIntegrase-deficient expression vectors based on HIV-1 are becoming increasingly popular as tools for gene therapy in vivo due to their inability to cause insertional mutagenesis. However, many IN-lentiviral vectors are able to achieve only low levels of gene expression and methods to increase this low level have not been extensively explored. Here we analyze how the HTLV-1 Tax protein is able to rescue the replication of IN-HIV-1 in T cells and describe IN-lentiviral vectors that are able to express a heterologous gene effectively.

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