Kaposi Sarcoma-associated Herpesvirus Latency-associated Nuclear Antigen Inhibits Interferon (IFN) β Expression by Competing with IFN Regulatory Factor-3 for Binding to IFNB Promoter (original) (raw)
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Journal of Biological Chemistry, 2010
Host cells respond to viral infections by synthesizing and producing antiviral molecules such as type I interferons (IFN). The Kaposi sarcoma-associated herpesvirus (KSHV) encodes multiple proteins expressed during the lytic replication cycle that alter the antiviral response of the host. Considering that in Kaposi sarcoma lesions and primary effusion lymphoma cells KSHV is latent in the vast majority of cells, we were interested in determining whether latently expressed viral proteins have the ability to modulate IFN synthesis. The latency-associated nuclear antigen (LANA-1) is a large nuclear protein that plays a role in the establishment and maintenance of latent KSHV episome in the nucleus of infected cells. LANA-1 is also described to modulate the cellular transcription. Here, we report that LANA-1 inhibits IFN- transcription and synthesis by competing with the binding of interferon regulatory factor-3 (IRF3) to the IFNB promoter. Using mutants of LANA-1, we have identified the central acidic repeated region as the domain essential for interfering with the binding of IRF3 to the positive regulatory domains I-III of the IFNB promoter. In addition, the nuclear localization of LANA-1 proved essential for IFN- inhibition. Thus, LANA-1 interferes with the formation of IFN- enhanceosome by competing with the fixation of IRF3 and by inhibiting the expression of the CREB-binding protein. The ability of LANA-1 to inhibit IFNB gene expression highlights a new role for this protein in cellular gene modulation and immune evasion strategies. Kaposi sarcoma-associated herpesvirus (KSHV), 3 also called human herpesvirus 8, is an oncogenic virus associated with the
Journal of Virology, 2001
Kaposi's sarcoma-associated herpesvirus (KSHV), also called human herpesvirus 8 (HHV-8), is the likely etiological agent of Kaposi's sarcoma and primary effusion lymphoma. Common to these malignancies is that tumor cells are latently infected with KSHV. Viral gene expression is limited to a few genes, one of which is the latency-associated nuclear antigen (LANA), the product of ORF73. Examination of the primary sequence of LANA reveals some structural features reminiscent of transcription factors, leading us to hypothesize that LANA may regulate viral and cellular transcription during latency. In reporter gene-based transient transfection assays, we found that LANA can have either positive or negative effects on gene expression.
Journal of Biological Chemistry, 2004
Kaposi's sarcoma-associated herpesvirus (KSHV) or human herpesvirus 8 has been established as the etiological agent of Kaposi's sarcoma and certain AIDS-associated lymphomas. KSHV establishes latent infection in these tumors, invariably expressing high levels of the viral latency-associated nuclear antigen (LANA) protein. LANA is necessary and sufficient to maintain the KSHV episome. It also modulates viral and cellular transcription and has been implicated directly in oncogenesis because of its ability to bind to the p53 and pRb tumor suppressor proteins. Previously, we identified the LANA promoter (LANAp) and showed that it was positively regulated by LANA itself. Here, we present a detailed mutational analysis and define cis-acting elements and trans-acting factors for the core LANAp. We found that a downstream promoter element, TATA box, and GC box/Sp1 site at -29 are all individually required for activity. This architecture places LANAp into the small and unusual group of eukaryotic promoters that contain both the downstream promoter element and TATA element but lack a defined initiation site. Furthermore, we demonstrate that LANA regulates its own promoter via its C-terminal domain and does bind to a defined site within the core promoter.
Journal of Virology, 2012
LANA is essential for tethering the Kaposi's sarcoma-associated herpesvirus (KSHV) genome to metaphase chromosomes and for modulating host-cell gene expression, but the binding sites in the host-chromosome remain unknown. Here, we use LANAspecific chromatin immunoprecipitation coupled with high-throughput sequencing (ChIP-Seq) to identify LANA binding sites in the viral and host-cell genomes of a latently infected pleural effusion lymphoma cell line BCBL1. LANA bound with high occupancy to the KSHV genome terminal repeats (TR) and to a few minor binding sites in the KSHV genome, including the LANA promoter region. We identified 256 putative LANA binding site peaks with P < 0.01 and overlap in two independent ChIP-Seq experiments. We validated several of the high-occupancy binding sites by conventional ChIP assays and quantitative PCR. Candidate cellular LANA binding motifs were identified and assayed for binding to purified recombinant LANA protein in vitro but bound with low affinity compared to the viral TR binding site. More than half of the LANA binding sites (170/256) could be mapped to within 2.5 kb of a cellular gene transcript. Pathways and Gene Ontogeny (GO) analysis revealed that LANA binds to genes within the p53 and tumor necrosis factor (TNF) regulatory network. Further analysis revealed partial overlap of LANA and STAT1 binding sites in several gamma interferon (IFN-␥)-regulated genes. We show that ectopic expression of LANA can downmodulate IFN-␥-mediated activation of a subset of genes, including the TAP1 peptide transporter and proteasome subunit beta type 9 (PSMB9), both of which are required for class I antigen presentation. Our data provide a potential mechanism through which LANA may regulate several host cell pathways by direct binding to gene regulatory elements.
PLOS Pathogens
Kaposi's sarcoma-associated herpesvirus (KSHV; human herpesvirus 8) belongs to the subfamily of Gammaherpesvirinae and is the etiological agent of Kaposi's sarcoma as well as of two lymphoproliferative diseases: primary effusion lymphoma and multicentric Castleman disease. The KSHV life cycle is divided into a latent and a lytic phase and is highly regulated by viral immunomodulatory proteins which control the host antiviral immune response. Among them is a group of proteins with homology to cellular interferon regulatory factors, the viral interferon regulatory factors 1-4. The KSHV vIRFs are known as inhibitors of cellular interferon signaling and are involved in different oncogenic pathways. Here we characterized the role of the second vIRF protein, vIRF2, during the KSHV life cycle. We found the vIRF2 protein to be expressed in different KSHV positive cells with early lytic kinetics. Importantly, we observed that vIRF2 suppresses the expression of viral early lytic genes in both newly infected and reactivated persistently infected endothelial cells. This vIRF2-dependent regulation of the KSHV life cycle might involve the increased expression of cellular interferon-induced genes such as the IFIT proteins 1, 2 and 3, which antagonize the expression of early KSHV lytic proteins. Our findings suggest a model in which the viral protein vIRF2 allows KSHV to harness an IFN-dependent pathway to regulate KSHV early gene expression.
Journal of General Virology, 2003
Kaposi's sarcoma-associated herpesvirus (KSHV) uses several strategies to counteract the interferon (IFN) system. In this study, the relationship of the protein LANA2 from KSHV to the IFN-activated protein kinase (PKR) and 2-5A system was analysed. It was found that LANA2 could not abrogate apoptosis or RNA degradation mediated by the 2-5A system. However, expression of LANA2 inhibited apoptosis triggered by PKR. LANA2 also counteracted the PKR-mediated inhibition of protein synthesis and partially blocked PKR-induced phosphorylation of eIF-2a. Analysis of PKR-induced activation of caspases 3 and 9 revealed that LANA2 abrogated activation of caspase 3 but not of caspase 9. These findings show that LANA2 is able to interfere with downstream events triggered by PKR. Hence, LANA2 should be considered as a KSHV defence protein against IFN.
Journal of Virology, 2007
Upon viral infection, the major defense mounted by the host immune system is activation of the interferon (IFN)-mediated antiviral pathway that is mediated by IFN regulatory factors (IRFs). In order to complete their life cycle, viruses must modulate the host IFN-mediated immune response. Kaposi's sarcoma-associated herpesvirus (KSHV), a human tumor-inducing herpesvirus, has developed a unique mechanism for antagonizing cellular IFN-mediated antiviral activity by incorporating viral homologs of the cellular IRFs, called vIRFs. Here, we report a novel immune evasion mechanism of KSHV vIRF3 to block cellular IRF7-mediated innate immunity in response to viral infection. KSHV vIRF3 specifically interacts with either the DNA binding domain or the central IRF association domain of IRF7, and this interaction leads to the inhibition of IRF7 DNA binding activity and, therefore, suppression of alpha interferon (IFN-␣) production and IFN-mediated immunity. Remarkably, the central 40 amino acids of vIRF3, containing the double ␣ helix motifs, are sufficient not only for binding to IRF7, but also for inhibiting IRF7 DNA binding activity. Consequently, the expression of the double ␣ helix motif-containing peptide effectively suppresses IRF7-mediated IFN-␣ production. This demonstrates a remarkably efficient means of viral avoidance of host antiviral activity.