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Polyamines are critical metabolites involved in various cellular processes and often dysregulated in cancers. Kaposi's sarcoma-associated Herpesvirus (KSHV), a defined human oncogenic virus, leads to profound alterations of host metabolic landscape to favor development of KSHV-associated malignancies. In our studies, we identified that polyamine biosynthesis and eIF5A hypusination are dynamically regulated by KSHV infection through modulation of key enzymes (ODC1 and DHPS) of these pathways. During KSHV latency, ODC1 and DHPS are upregulated along with increase of hypusinated eIF5A (hyp-eIF5A), while hyp-eIF5A is further induced along with reduction of ODC1 and intracellular polyamines during KSHV lytic reactivation. In return these metabolic pathways are required for both KSHV lytic reactivation and de novo infection. Further analysis unraveled that synthesis of critical KSHV latent and lytic proteins (LANA, RTA) depends on hypusinated-eIF5A. We also demonstrated that KSHV infection can be efficiently and specifically suppressed by inhibitors targeting these pathways. Collectively, our results illustrated that the dynamic and profound interaction of a DNA tumor virus (KSHV) with host polyamine biosynthesis and eIF5A hypusination pathways promote viral propagation, thus defining new therapeutic targets to treat KSHV-associated malignancies. Author summary: Understanding virus-host interactions is crucial to develop and improve therapies. Kaposi's sarcoma associated Herpesvirus (KSHV) is a human gamma-herpesvirus which deeply modulates the host metabolism and is associated with various cancers of endothelial and lymphoid origin. Polyamines are critical metabolites often dysregulated in cancers. In this study we demonstrated KSHV dynamically modulates polyamine metabolism to favor eIF5A hypusination and translation of critical KSHV latent and lytic proteins (LANA, RTA). Consequently, we found KSHV lytic switch from latency and de novo infection were dependent on polyamines and hypusination and pharmacological inhibition efficiently and specifically restricted KSHV infection. Our study provides new insights into KSHV alteration of the host metabolism and describe new therapeutic targets to treat KSHV-associated malignancies. [ABSTRACT FROM AUTHOR]

EBV-associated gastric cancer (EBVaGC) is characterized by high frequency of DNA methylation. In this study, we investigated how epigenetic alteration of host genome contributes to pathogenesis of EBVaGC through the analysis of transcriptomic and epigenomic datasets from NIH TCGA (The Cancer Genome Atlas) consortium. We identified that immune related genes (IRGs) is a group of host genes preferentially silenced in EBV-positive gastric cancers through DNA hypermethylation. Further functional characterizations of selected IRGs reveal their novel antiviral activity against not only EBV but also KSHV. In particular, we showed that metallothionein-1 (MT1) and homeobox A (HOXA) gene clusters are down-regulated via EBV-driven DNA hypermethylation. Several MT1 isoforms suppress EBV lytic replication and release of progeny virions as well as KSHV lytic reactivation, suggesting functional redundancy of these genes. In addition, single HOXA10 isoform exerts antiviral activity against both EBV and KSHV. We also confirmed the antiviral effect of other dysregulated IRGs, such as IRAK2 and MAL, in scenario of EBV and KSHV lytic reactivation. Collectively, our results demonstrated that epigenetic silencing of IRGs is a viral strategy to escape immune surveillance and promote viral propagation, which is overall beneficial to viral oncogenesis of human gamma-herpesviruses (EBV and KSHV), considering that these IRGs possess antiviral activities against these oncoviruses. Author summary: Epstein-Barr virus (EBV), one of the human gamma-herpesviruses, is a well-defined viral agent that strongly associates with malignancies of lymphoid and epithelial origin. EBV-associated gastric carcinoma (EBVaGC) is the most common malignancy caused by EBV infection. In this paper, we identified that beyond tumor suppressor genes, immune related genes (IRGs) are another group of host genes that are extensively DNA hypermethylated and epigenetically silenced in EBVaGC comparing to non-EBV GC. For certain IRGs, such as metallothionein-1 (MT1) and homeobox A (HOXA) genes, the whole gene clusters undergo DNA hypermethylation in EBVaGC, while it also occurs for isolated individual IRGs, such as IRAK2 and MAL. Considering the critical role of innate immunity in antiviral control, we expected that silencing of IRGs would benefit EBV viral replication and propagation. Indeed, our further investigation of several IRGs (MT1G, HOXA10, IRAK2, and MAL) confirmed their antiviral activities against EBV lytic replication. Some of them also suppress lytic replication of another human gamma-herpesvirus, Kaposi's sarcoma-associated herpesvirus (KSHV), indicating their broad antiviral spectrum. Collectively, our results demonstrated that epigenetic silencing of antiviral IRGs is an efficient viral strategy utilized by oncoviruses, such as EBV, to escape immune surveillance and promote viral propagation, which is overall beneficial to viral oncogenesis. [ABSTRACT FROM AUTHOR]

The spatiotemporal dynamics of Hepatitis C Virus (HCV) RNA localisation are poorly understood. To address this we engineered HCV genomes harbouring MS2 bacteriophage RNA stem-loops within the 3′-untranslated region to allow tracking of HCV RNA via specific interaction with a MS2-Coat-mCherry fusion protein. Despite the impact of these insertions on viral fitness, live imaging revealed that replication of tagged-HCV genomes induced specific redistribution of the mCherry-tagged-MS2-Coat protein to motile and static foci. Further analysis showed that HCV RNA was associated with NS5A in both static and motile structures while a subset of motile NS5A structures was devoid of HCV RNA. Further investigation of viral RNA traffic with respect to lipid droplets (LDs) revealed HCV RNA-positive structures in close association with LDs. These studies provide new insights into the dynamics of HCV RNA traffic with NS5A and LDs and provide a platform for future investigations of HCV replication and assembly. [ABSTRACT FROM AUTHOR]

Hepatitis C virus (HCV) NS5A is essential for viral genome replication within cytoplasmic replication complexes and virus assembly at the lipid droplet (LD) surface, although its definitive functions are poorly understood. We developed approaches to investigate NS5A dynamics during a productive infection. We report here that NS5A motility and efficient HCV RNA replication require the microtubule network and the cytoplasmic motor dynein and demonstrate that both motile and relatively static NS5A-positive foci are enriched with host factors VAP-A and Rab5A. Pulse-chase imaging revealed that newly synthesized NS5A foci are small and distinct from aged foci, while further studies using a unique dual fluorescently tagged infectious HCV chimera showed a relatively stable association of NS5A foci with core-capped LDs. These results reveal new details about the dynamics and maturation of NS5A and the nature of potential sites of convergence of HCV replication and assembly pathways. [ABSTRACT FROM AUTHOR]

Both Kaposi's sarcoma-associated herpesvirus (KSHV) and Epstein-Barr virus (EBV) establish the persistent, life-long infection primarily at the latent status, and associate with certain types of tumors, such as B cell lymphomas, especially in immuno-compromised individuals including people living with HIV (PLWH). Lytic reactivation of these viruses can be employed to kill tumor cells harboring latently infected viral episomes through the viral cytopathic effects and the subsequent antiviral immune responses. In this study, we identified that polo-like kinase 1 (PLK1) is induced by KSHV de novo infection as well as lytic switch from KSHV latency. We further demonstrated that PLK1 depletion or inhibition facilitates KSHV reactivation and promotes cell death of KSHV-infected lymphoma cells. Mechanistically, PLK1 regulates Myc that is critical to both maintenance of KSHV latency and support of cell survival, and preferentially affects the level of H3K27me3 inactive mark both globally and at certain loci of KSHV viral episomes. Furthremore, we recognized that PLK1 inhibition synergizes with STAT3 inhibition to efficiently induce KSHV reactivation. We also confirmed that PLK1 depletion or inhibition yields the similar effect on EBV lytic reactivation and cell death of EBV-infected lymphoma cells. Lastly, we noticed that PLK1 in B cells is elevated in the context of HIV infection and caused by HIV Nef protein to favor KSHV/EBV latency. Author summary: KSHV is a large double-stranded DNA virus belonging to human gamma-herpesviruses and capable of establishing the long-term persistent infection, which causes cancers under immunocompromised states, such as acquired immunodeficiency syndrome (AIDS). There are currently still no effective therapies to specifically treat KSHV-associated tumors, nor eliminate its persistent infection. In this study, we illustrated that PLK1 is upregulated by KSHV, which in return suppresses KSHV lytic infection. We further demonstrated that PLK1 is a novel host target that can be inhibited pharmacologically to benefit the viral oncolysis for promotion of KSHV lytic reactivation and elimination of KSHV-positive lymphoma cells, particularly for people living with HIV (PLWH). PLK1 inhibitors also exerted the similar effect on EBV that is the other member of human gamma-herpesvirus. [ABSTRACT FROM AUTHOR]

The host protein viperin is an interferon stimulated gene (ISG) that is up-regulated during a number of viral infections. In this study we have shown that dengue virus type-2 (DENV-2) infection significantly induced viperin, co-incident with production of viral RNA and via a mechanism requiring retinoic acid-inducible gene I (RIG-I). Viperin did not inhibit DENV-2 entry but DENV-2 RNA and infectious virus release was inhibited in viperin expressing cells. Conversely, DENV-2 replicated to higher tires earlier in viperin shRNA expressing cells. The anti-DENV effect of viperin was mediated by residues within the C-terminal 17 amino acids of viperin and did not require the N-terminal residues, including the helix domain, leucine zipper and S-adenosylmethionine (SAM) motifs known to be involved in viperin intracellular membrane association. Viperin showed co-localisation with lipid droplet markers, and was co-localised and interacted with DENV-2 capsid (CA), NS3 and viral RNA. The ability of viperin to interact with DENV-2 NS3 was associated with its anti-viral activity, while co-localisation of viperin with lipid droplets was not. Thus, DENV-2 infection induces viperin which has anti-viral properties residing in the C-terminal region of the protein that act to restrict early DENV-2 RNA production/accumulation, potentially via interaction of viperin with DENV-2 NS3 and replication complexes. These anti-DENV-2 actions of viperin show both contrasts and similarities with other described anti-viral mechanisms of viperin action and highlight the diverse nature of this unique anti-viral host protein. Author Summary: Viperin is a virally induced host protein that has been previously shown to have antiviral activity against a variety of viruses. Here we have demonstrated that viperin is also anti-viral against the medically significant arbovirus, dengue virus. Viperin was able to inhibit dengue virus at the level of viral replication, and cell lines unable to produce normal levels of viperin grew the virus to higher titres. These anti-dengue effects of viperin were mediated by amino acid residues in its C-terminus, and did not require structural domains of the N-terminal region as has been previously shown by us and others for the related virus, hepatitis C virus. Viperin was also demonstrated to co-localise and interact with the dengue capsid protein on the surface of lipid droplets, as well as with the NS3 protein and viral RNA. Viperin's association with NS3 was further demonstrated to be involved in its anti-dengue activities. The anti-viral activities of viperin presented in this manuscript show both similarities and contrasts with other described anti-viral mechanisms for the protein and highlight the diverse nature of this unique anti-viral host protein. [ABSTRACT FROM AUTHOR]

The host protein viperin is an interferon stimulated gene (ISG) that is up-regulated during a number of viral infections. In this study we have shown that dengue virus type-2 (DENV-2) infection significantly induced viperin, co-incident with production of viral RNA and via a mechanism requiring retinoic acid-inducible gene I (RIG-I). Viperin did not inhibit DENV-2 entry but DENV-2 RNA and infectious virus release was inhibited in viperin expressing cells. Conversely, DENV-2 replicated to higher tires earlier in viperin shRNA expressing cells. The anti-DENV effect of viperin was mediated by residues within the Cterminal 17 amino acids of viperin and did not require the N-terminal residues, including the helix domain, leucine zipper and S-adenosylmethionine (SAM) motifs known to be involved in viperin intracellular membrane association. Viperin showed co-localisation with lipid droplet markers, and was co-localised and interacted with DENV-2 capsid (CA), NS3 and viral RNA. The ability of viperin to interact with DENV-2 NS3 was associated with its anti-viral activity, while co-localisation of viperin with lipid droplets was not. Thus, DENV-2 infection induces viperin which has anti-viral properties residing in the Cterminal region of the protein that act to restrict early DENV-2 RNA production/accumulation, potentially via interaction of viperin with DENV-2 NS3 and replication complexes. These anti-DENV-2 actions of viperin show both contrasts and similarities with other described anti-viral mechanisms of viperin action and highlight the diverse nature of this unique antiviral host protein. [ABSTRACT FROM AUTHOR]