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Kaposi's sarcoma-associated herpesvirus noncoding polyadenylated nuclear RNA interacts with virus- and host cell-encoded proteins and suppresses expression of genes involved in immune modulation.

ABSTRACT: During lytic infection, Kaposi's sarcoma-associated herpesvirus (KSHV) expresses a polyadenylated nuclear RNA (PAN RNA). This noncoding RNA (ncRNA) is localized to the nucleus and is the most abundant viral RNA during lytic infection; however, to date, the role of PAN RNA in the virus life cycle is unknown. Many examples exist where ncRNAs have a defined key regulatory function controlling gene expression by various mechanisms. Our goal for this study was to identify putative binding partners for PAN RNA in an effort to elucidate a possible function for the transcript in KSHV infection. We employed an in vitro affinity protocol where PAN RNA was used as bait for factors present in BCBL-1 cell nuclear extract to show that PAN RNA interacts with several virus- and host cell-encoded factors, including histones H1 and H2A, mitochondrial and cellular single-stranded binding proteins (SSBPs), and interferon regulatory factor 4 (IRF4). RNA chromatin immunoprecipitation (ChIP) assays confirmed that PAN RNA interacted with these factors in the infected cell environment. A luciferase reporter assay showed that PAN RNA expression interfered with the ability of IRF4/PU.1 to activate the interleukin-4 (IL-4) promoter, strongly suggesting a role for PAN RNA in immune modulation. Since the proteomic screen and functional data suggested a role in immune responses, we investigated if constitutive PAN RNA expression could affect other genes involved in immune responses. PAN RNA expression decreased expression of gamma interferon, interleukin-18, alpha interferon 16, and RNase L. These data strongly suggest that PAN RNA interacts with viral and cellular proteins and can function as an immune modulator.

SUBMITTER: Rossetto CC

PROVIDER: S-EPMC3233155 | biostudies-literature | 2011 Dec

REPOSITORIES: biostudies-literature

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Kaposi's sarcoma-associated herpesvirus noncoding polyadenylated nuclear RNA interacts with virus- and host cell-encoded proteins and suppresses expression of genes involved in immune modulation.

Rossetto Cyprian C CC Pari Gregory S GS

Journal of virology 20110928 24

During lytic infection, Kaposi's sarcoma-associated herpesvirus (KSHV) expresses a polyadenylated nuclear RNA (PAN RNA). This noncoding RNA (ncRNA) is localized to the nucleus and is the most abundant viral RNA during lytic infection; however, to date, the role of PAN RNA in the virus life cycle is unknown. Many examples exist where ncRNAs have a defined key regulatory function controlling gene expression by various mechanisms. Our goal for this study was to identify putative binding partners fo ...[more]

PMID: 21957289

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Project description:Kaposi's sarcoma-associated herpesvirus (KSHV) is an oncogenic human gammaherpesvirus and the causative agent of Kaposi's sarcoma (KS), primary effusion lymphoma (PEL), and multicentric Castleman's disease (MCD). During reactivation, viral genes are expressed in a temporal manner. These lytic genes encode transactivators, core replication proteins, or structural proteins. During reactivation, other viral factors that are required for lytic replication are expressed. The most abundant viral transcript is the long noncoding RNA (lncRNA) known as polyadenylated nuclear (PAN) RNA. lncRNAs have diverse functions, including the regulation of gene expression and the immune response. PAN possesses two main cis-acting elements, the Mta response element (MRE) and the expression and nuclear retention element (ENE). While PAN has been demonstrated to be required for efficient viral replication, the function of these elements within PAN remains unclear. Our goal was to determine if the ENE of PAN is required in the context of infection. A KSHV bacmid containing a deletion of the 79-nucleotide (nt) ENE in PAN was generated to assess the effects of the ENE during viral replication. Our studies demonstrated that the ENE is not required for viral DNA synthesis, lytic gene expression, or the production of infectious virus. Although the ENE is not required for viral replication, we found that the ENE functions to retain PAN in the nucleus, and the absence of the ENE results in an increased accumulation of PAN in the cytoplasm. Furthermore, open reading frame 59 (ORF59), LANA, ORF57, H1.4, and H2A still retain the ability to bind to PAN in the absence of the ENE. Together, our data highlight how the ENE affects the nuclear retention of PAN but ultimately does not play an essential role during lytic replication. Our data suggest that PAN may have other functional domains apart from the ENE. IMPORTANCE KSHV is an oncogenic herpesvirus that establishes latency and exhibits episodes of reactivation. KSHV disease pathologies are most often associated with the lytic replication of the virus. PAN RNA is the most abundant viral transcript during the reactivation of KSHV and is required for viral replication. Deletion and knockdown of PAN resulted in defects in viral replication and reduced virion production in the absence of PAN RNA. To better understand how the cis elements within PAN may contribute to its function, we investigated if the ENE of PAN was necessary for viral replication. Although the ENE had previously been extensively studied with both biochemical and in vitro approaches, this is the first study to demonstrate the role of the ENE in the context of infection and that the ENE of PAN is not required for the lytic replication of KSHV.

Dataset Information

Kaposi's sarcoma-associated herpesvirus noncoding polyadenylated nuclear RNA interacts with virus- and host cell-encoded proteins and suppresses expression of genes involved in immune modulation.

Publications

Kaposi's sarcoma-associated herpesvirus noncoding polyadenylated nuclear RNA interacts with virus- and host cell-encoded proteins and suppresses expression of genes involved in immune modulation.

Similar Datasets

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