The ND10 Complex Represses Lytic Human Herpesvirus 6A Replication and Promotes Silencing of the Viral Genome.
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ABSTRACT:
SUBMITTER: Sanyal A
PROVIDER: S-EPMC6115956 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:We report the application of size selection of small RNA species isolated from Jjhan cells harboring the human herpesvirus 6A genome. We ammassed >3.4million reads of sequence from three different sources: Normal Brain cell total RNA, Jjhan total RNA and HHV-6A BAC transfected Jjhan total RNA. Sequences were mapped to the HHV-6A Uganda 1102 strain genome (GenBank: X83413.1) with no less than 100% match for reads >20nt and <23nt. The resulting pool of candidates was mapped to the HHV-6A genome. Single pass 36nt sequencing of samples either with or without HHV-6a genomes present.
2015-01-09 | E-GEOD-62233 | biostudies-arrayexpress
Project description:UnlabelledHuman herpesvirus 6A (HHV-6A), a member of the betaherpesvirus family, is associated with several human diseases. Like all herpesviruses, HHV-6A establishes a lifelong, latent infection in its host. Reactivation of HHV-6A is frequent within the immunosuppressed and immunocompromised populations and results in lytic viral replication within multiple organs, often leading to severe disease. MicroRNAs (miRNAs) are key regulators of multiple cellular processes that regulate the translation of specific transcripts. miRNAs carried by herpesviruses play important roles in modulating the host cell, thereby facilitating a suitable environment for productive viral infection and/or latency. Currently, there are approximately 150 known human herpesvirus-encoded miRNAs, although an miRNA(s) encoded by HHV-6A has yet to be reported. We hypothesized that HHV-6A, like other members of the human herpesvirus family, encodes miRNAs, which function to promote viral infection. We utilized deep sequencing of small RNA species isolated from cells harboring HHV-6A to identify five novel small noncoding RNA species that originate from the viral genome, one of which has the characteristics of a viral miRNA. These RNAs are expressed during productive infection by either bacterial artificial chromosome (BAC)-derived virus in Jjhan cells or wild-type HHV-6A strain U1102 virus in HSB2 cells and are associated with the RNA induced silencing complex (RISC) machinery. Growth analyses of mutant viruses that lack each individual miRNA revealed that a viral miRNA candidate (miR-U86) targets the HHV-6A IE gene U86, thereby regulating lytic replication. The identification and biological characterization of this HHV-6A-specific miRNA is the first step to defining how the virus regulates its life cycle.ImportanceA majority of the human population is infected with human herpesvirus 6A (HHV-6A), a betaherpesvirus family member. Infections usually occur in young children, and upon resolution, the virus remains in a latent state within the host. Importantly, during times of weakened immune responses, the virus can reactivate and is correlated with significant disease states. Viruses encode many different types of factors that both undermine the host antiviral response and regulate viral replication, including small RNA species called microRNAs (miRNAs). Here we report that HHV-6A encodes at least one miRNA, which we named miR-U86. We have characterized the requirement of this viral miRNA and its impact on the viral life cycle and found that it functions to regulate a viral protein important for efficient viral replication. Our data suggest that viral miRNAs are important for HHV-6A and that they may serve as an important therapeutic target to inhibit the virus.
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Project description:We report the application of size selection of small RNA species isolated from Jjhan cells harboring the human herpesvirus 6A genome. We ammassed >3.4million reads of sequence from three different sources: Normal Brain cell total RNA, Jjhan total RNA and HHV-6A BAC transfected Jjhan total RNA. Sequences were mapped to the HHV-6A Uganda 1102 strain genome (GenBank: X83413.1) with no less than 100% match for reads >20nt and <23nt. The resulting pool of candidates was mapped to the HHV-6A genome.
2015-01-09 | GSE62233 | GEO
Project description:Human herpesvirus 6A (HHV-6A) is a common virus with a worldwide distribution that has been associated with multiple sclerosis. Whether HHV-6A can replicate in dendritic cells (DC) and how the infection might modulate the functional properties of the cell are currently not well known and need further investigations. Here, we show that a non-productive infection of HHV-6A in DC leads to the up-regulation of HLA-ABC, via autocrine IFN-? signaling, as well as the up-regulation of HLA-DR and CD86. However, HHV-6A exposure reduces IL-8 secretion by DC and their capacity to stimulate allogenic T cell proliferation. The ability to suppress DC functions important for activation of innate and adaptive immune responses might be one successful strategy by which HHV-6A avoids the induction of appropriate host defense mechanisms, and thus facilitating persistent infection.
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Project description:Nuclear domain 10 (ND10) components are restriction factors that inhibit herpesviral replication. Effector proteins of different herpesviruses can antagonize this restriction by a variety of strategies, including degradation or relocalization of ND10 proteins. We investigated the interplay of Kaposi's Sarcoma-Associated Herpesvirus (KSHV) infection and cellular defense by nuclear domain 10 (ND10) components. Knock-down experiments in primary human cells show that KSHV-infection is restricted by the ND10 components PML and Sp100, but not by ATRX. After KSHV infection, ATRX is efficiently depleted and Daxx is dispersed from ND10, indicating that these two ND10 components can be antagonized by KSHV. We then identified the ORF75 tegument protein of KSHV as the viral factor that induces the disappearance of ATRX and relocalization of Daxx. ORF75 belongs to a viral protein family (viral FGARATs) that has homologous proteins in all gamma-herpesviruses. Isolated expression of ORF75 in primary cells induces a relocalization of PML and dispersal of Sp100, indicating that this viral effector protein is able to influence multiple ND10 components. Moreover, by constructing a KSHV mutant harboring a stop codon at the beginning of ORF75, we could demonstrate that ORF75 is absolutely essential for viral replication and the initiation of viral immediate-early gene expression. Using recombinant viruses either carrying Flag- or YFP-tagged variants of ORF75, we could further corroborate the role of ORF75 in the antagonization of ND10-mediated intrinsic immunity, and show that it is independent of the PML antagonist vIRF3. Members of the viral FGARAT family target different ND10 components, suggesting that the ND10 targets of viral FGARAT proteins have diversified during evolution. We assume that overcoming ND10 intrinsic defense constitutes a critical event in the replication of all herpesviruses; on the other hand, restriction of herpesviral replication by ND10 components may also promote latency as the default outcome of infection.
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Project description:The K1 gene product of Kaposi's sarcoma-associated herpesvirus (KSHV) is encoded by the first open reading frame (ORF) of the viral genome. To investigate the role of the K1 gene during the KSHV life cycle, we constructed a set of recombinant viruses that contained either wild-type (WT) K1, a deleted K1 ORF (KSHV?K1), stop codons within the K1 ORF (KSHV-K15×STOP), or a revertant K1 virus (KSHV-K1REV). We report that the recombinant viruses KSHV?K1 and KSHV-K15×STOP displayed significantly reduced lytic replication compared to WT KSHV and KSHV-K1REV upon reactivation from latency. Additionally, cells infected with the recombinant viruses KSHV?K1 and KSHV-K15×STOP also yielded smaller amounts of infectious progeny upon reactivation than did WT KSHV- and KSHV-K1REV-infected cells. Upon reactivation from latency, WT KSHV- and KSHV-K1REV-infected cells displayed activated Akt kinase, as evidenced by its phosphorylation, while cells infected with viruses deleted for K1 showed reduced phosphorylation and activation of Akt kinase. Overall, our results suggest that K1 plays an important role during the KSHV life cycle.Kaposi's sarcoma-associated herpesvirus (KSHV) is the etiological agent of three human malignancies, and KSHV K1 is a signaling protein that has been shown to be involved in cellular transformation and to activate the phosphatidylinositol 3-kinase (PI3K)/Akt/mTOR pathway. In order to investigate the role of the K1 protein in the life cycle of KSHV, we constructed recombinant viruses that were deficient for K1. We found that K1 deletion viruses displayed reduced lytic replication compared to the WT virus and also yielded smaller numbers of infectious progeny. We report that K1 plays an important role in the life cycle of KSHV.
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Project description:The stimulus-induced cAMP response element (CRE)-binding protein (CREB) family of transcription factors bind to CREs to regulate diverse cellular responses, including proliferation, survival, and differentiation. Human herpesvirus 6A (HHV-6A), which belongs to the Betaherpesvirinae subfamily, is a lymphotropic herpesvirus frequently found in patients with neuroinflammatory diseases. Previous reports implicated the importance of CREs in the HHV-6A life cycle, although the effects of the binding of transcription factors to CREs in viral replication have not been fully elucidated. In this study, we analyzed the role of the CREB family of transcription factors during HHV-6A replication. We found that HHV-6A infection enhanced phosphorylation of the CREB family members CREB1 and activating transcription factor 1 (ATF1). Knockout (KO) of CREB1 or ATF1 enhanced viral gene expression and viral replication. The increase in viral yields in supernatants from ATF1-KO cells was greater than that in supernatants from CREB1-KO cells. Transcriptome sequencing (RNA-seq) analysis showed that sensors of the innate immune system were downregulated in ATF1-KO cells, and mRNAs of beta interferon (IFN-β) and IFN-regulated genes were reduced in these cells infected with HHV-6A. IFN-β treatment of ATF1-KO cells reduced progeny viral yields significantly, suggesting that the enhancement of viral replication was caused by a reduction of IFN-β. Taken together, our results suggest that ATF1 is activated during HHV-6A infection and restricts viral replication via IFN-β induction. IMPORTANCE Human herpesvirus 6A (HHV-6A) is a ubiquitous herpesvirus implicated in Alzheimer's disease, although its role in its pathogenesis has not been confirmed. Here, we showed that the transcription factor ATF1 restricts HHV-6A replication, mediated by IFN-β induction. Our study provides new insights into the role of ATF1 in innate viral immunity and reveals the importance of IFN-β for regulation of HHV-6A replication, which possibly impairs HHV-6A pathogenesis.
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