scholarly journals The Transcriptional Repressor K-RBP Modulates RTA-Mediated Transactivation and Lytic Replication of Kaposi's Sarcoma-Associated Herpesvirus

2007 ◽  
Vol 81 (12) ◽  
pp. 6294-6306 ◽  
Author(s):  
Zhilong Yang ◽  
Charles Wood
Keyword(s):  
Zinc Finger ◽  
Kaposi's Sarcoma ◽  
Zinc Finger Protein ◽  
Mutational Analysis ◽  
Binding Protein ◽  
Transcriptional Repressor ◽  
Human Herpesvirus ◽  
Kaposi’S Sarcoma ◽  
Lytic Replication ◽  
Transcription Activator

ABSTRACT The replication and transcription activator (RTA) protein of Kaposi's sarcoma (KS)-associated herpesvirus (KSHV)/human herpesvirus 8 functions as the key regulator to induce KSHV lytic replication from latency through activation of the lytic cascade of KSHV. Elucidation of the host factors involved in RTA-mediated transcriptional activation is pivotal for understanding the transition between viral latency and lytic replication. KSHV-RTA binding protein (K-RBP) was previously isolated as a cellular RTA binding protein of unknown function. Sequence analysis showed that K-RBP contains a Kruppel-associated box (KRAB) at the N terminus and 12 adjacent zinc finger motifs. In similarity to other KRAB-containing zinc finger proteins, K-RBP is a transcriptional repressor. Mutational analysis revealed that the KRAB domain is responsible for the transcriptional suppression activity of this protein and that the repression is histone deacetylase independent. K-RBP was found to repress RTA-mediated transactivation and interact with TIF1β (transcription intermediary factor 1β), a common corepressor of KRAB-containing protein, to synergize with K-RBP in repression. Overexpression and knockdown experiment results suggest that K-RBP is a suppressor of RTA-mediated KSHV reactivation. Our findings suggest that the KRAB-containing zinc finger protein K-RBP can suppress RTA-mediated transactivation and KSHV lytic replication and that KSHV utilizes this protein as a regulator to maintain a balance between latency and lytic replication.

scholarly journals Cellular Corepressor TLE2 Inhibits Replication-and-Transcription- Activator-Mediated Transactivation and Lytic Reactivation of Kaposi's Sarcoma-Associated Herpesvirus

2009 ◽  
Vol 84 (4) ◽  
pp. 2047-2062 ◽  
Author(s):  
Zhiheng He ◽  
Yunhua Liu ◽  
Deguang Liang ◽  
Zhuo Wang ◽  
Erle S. Robertson ◽  
...  
Keyword(s):  
Kaposi's Sarcoma ◽  
Binding Protein ◽  
Kaposi’S Sarcoma ◽  
Lytic Replication ◽  
Host Protein ◽  
Transcription Activator ◽  
Rich Domain ◽  
Lytic Reactivation ◽  
Kaposi's Sarcoma Associated Herpesvirus ◽  
Kaposi’S Sarcoma Associated Herpesvirus

ABSTRACT Replication and transcription activator (RTA) encoded by open reading frame 50 (ORF50) of Kaposi's sarcoma-associated herpesvirus (KSHV) is essential and sufficient to initiate lytic reactivation. RTA activates its target genes through direct binding with high affinity to its responsive elements or by interaction with cellular factors, such as RBP-Jκ, Ap-1, C/EBP-α, and Oct-1. In this study, we identified transducin-like enhancer of split 2 (TLE2) as a novel RTA binding protein by using yeast two-hybrid screening of a human spleen cDNA library. The interaction between TLE2 and RTA was confirmed by glutathione S-transferase (GST) binding and coimmunoprecipitation assays. Immunofluorescence analysis showed that TLE2 and RTA were colocalized in the same nuclear compartment in KSHV-infected cells. This interaction recruited TLE2 to RTA bound to its recognition sites on DNA and repressed RTA auto-activation and transactivation activity. Moreover, TLE2 also inhibited the induction of lytic replication and virion production driven by RTA. We further showed that the Q (Gln-rich), SP (Ser-Pro-rich), and WDR (Trp-Asp repeat) domains of TLE2 and the Pro-rich domain of RTA were essential for this interaction. RBP-Jκ has been shown previously to bind to the same Pro-rich domain of RTA, and this binding can be subject to competition by TLE2. In addition, TLE2 can form a complex with RTA to access the cognate DNA sequence of the RTA-responsive element at different promoters. Intriguingly, the transcription level of TLE2 could be upregulated by RTA during the lytic reactivation process. In conclusion, we identified a new RTA binding protein, TLE2, and demonstrated that TLE2 inhibited replication and transactivation mediated by RTA. This provides another potentially important mechanism for maintenance of KSHV viral latency through interaction with a host protein.

scholarly journals Transcriptional Downregulation of ORF50/Rta by Methotrexate Inhibits the Switch of Kaposi's Sarcoma-Associated Herpesvirus/Human Herpesvirus 8 from Latency to Lytic Replication

2002 ◽  
Vol 76 (10) ◽  
pp. 5208-5219 ◽  
Author(s):  
Francesca Curreli ◽  
Francesca Cerimele ◽  
Sumitra Muralidhar ◽  
Leonard J. Rosenthal ◽  
Ethel Cesarman ◽  
...  
Keyword(s):  
Kaposi's Sarcoma ◽  
Cultured Cells ◽  
Primary Effusion Lymphoma ◽  
Human Herpesvirus ◽  
Kaposi’S Sarcoma ◽  
Lytic Replication ◽  
Early Gene ◽  
Transcription Activator ◽  
Kaposi's Sarcoma Associated Herpesvirus ◽  
Kaposi’S Sarcoma Associated Herpesvirus

ABSTRACT Kaposi's sarcoma-associated herpesvirus (KSHV) encodes a cellular dihydrofolate reductase (DHFR) homologue. Methotrexate (MTX), a potent anti-inflammatory agent, inhibits cellular DHFR activity. We investigated the effect of noncytotoxic doses of MTX on latency and lytic KSHV replication in two KSHV-infected primary effusion lymphoma cell lines (BC-3 and BC-1) and in MTX-resistant BC-3 cells (MTX-R-BC-3 cells). Treatment with MTX completely prevented tetradecanoyl phorbol acetate-induced viral DNA replication and strongly decreased viral lytic transcript levels, even in MTX-resistant cells. However, the same treatment had no effect on transcription of cellular genes and KSHV latent genes. One of the lytic transcripts inhibited by MTX, ORF50/Rta (open reading frame), is an immediate-early gene encoding a replication-transcription activator required for expression of other viral lytic genes. Therefore, transcription of genes downstream of ORF50/Rta was inhibited, including those encoding the viral G-protein-coupled receptor (GPCR), viral interleukin-6, and K12/kaposin, which have been shown to be transforming in vitro and oncogenic in mice. Resistance to MTX has been documented in cultured cells and also in patients treated with this drug. However, MTX showed an inhibitory activity even in MTX-R-BC-3 cells. Two currently available antiherpesvirus drugs, cidofovir and foscarnet, had no effect on the transcription of these viral oncogenes and ORF50/Rta. MTX is the first example of a compound shown to downregulate the expression of ORF50/Rta and therefore prevent viral transforming gene transcription. Given that the expression of these genes may be important for tumor development, MTX could play a role in the future management of KSHV-associated malignancies.

scholarly journals Kaposi's Sarcoma-Associated Herpesvirus ori-Lyt-Dependent DNA Replication: DualRole of Replication and Transcription Activator

2006 ◽  
Vol 80 (24) ◽  
pp. 12171-12186 ◽  
Author(s):  
Yan Wang ◽  
Qiyi Tang ◽  
Gerd G. Maul ◽  
Yan Yuan
Keyword(s):  
Dna Replication ◽  
Kaposi's Sarcoma ◽  
Kaposi’S Sarcoma ◽  
Lytic Replication ◽  
Transcription Activator ◽  
Binding Motifs ◽  
Viral Propagation ◽  
Kaposi's Sarcoma Associated Herpesvirus ◽  
Kaposi’S Sarcoma Associated Herpesvirus ◽  
Lytic Dna Replication

ABSTRACT Lytic replication of Kaposi's sarcoma-associated herpesvirus (KSHV) is essential for viral propagation and pathogenicity. In Kaposi's sarcoma lesions, constant lytic replication plays a role in sustaining the population of latently infected cells that otherwise are quickly lost by segregation of latent viral episomes as spindle cells divide. Lytic DNA replication initiates from an origin (ori-Lyt) and requires trans-acting elements. Two functional ori-Lyts have been identified in the KSHV genome. Some cis-acting and trans-acting elements for ori-Lyt-dependent DNA replication have been found. Among these, K8 binding sites, a cluster of C/EBP binding motifs, and a replication and transcription activator (RTA) responsive element (RRE) are crucial cis-acting elements. Binding of K8 and RTA proteins to these motifs in ori-Lyt DNA was demonstrated to be absolutely essential for DNA replication. In the present study, functional roles of RTA in ori-Lyt-dependent DNA replication have been investigated. Two distinct functions of RTA were revealed. First, RTA activates an ori-Lyt promoter and initiates transcription across GC-rich tandem repeats. This RTA-mediated transcription is indispensable for DNA replication. Second, RTA is a component of the replication compartment, where RTA interacts with prereplication complexes composed of at least six core machinery proteins and K8. The prereplication complexes are recruited to ori-Lyt DNA through RTA, which interacts with the RRE, as well as K8, which binds to a cluster of C/EBP binding motifs with the aid of C/EBP α. The revelation of these two functions of RTA, together with its role in initiation of a transcriptional cascade that leads to transcription of all viral lytic genes, shows that RTA is a critical initiator and regulator of KSHV lytic DNA replication and viral propagation.

scholarly journals Activation of Kaposi's Sarcoma-Associated Herpesvirus (Human Herpesvirus 8) Lytic Replication by Human Cytomegalovirus

2001 ◽  
Vol 75 (3) ◽  
pp. 1378-1386 ◽  
Author(s):  
Jeffrey Vieira ◽  
Patricia O'Hearn ◽  
Louise Kimball ◽  
Bala Chandran ◽  
Lawrence Corey
Keyword(s):  
Human Cytomegalovirus ◽  
Kaposi's Sarcoma ◽  
Human Fibroblasts ◽  
Human Herpesvirus ◽  
Kaposi’S Sarcoma ◽  
Lytic Replication ◽  
Lytic Cycle ◽  
Nuclear Antigen ◽  
Kaposi's Sarcoma Associated Herpesvirus ◽  
Kaposi’S Sarcoma Associated Herpesvirus

ABSTRACT The majority of Kaposi's sarcoma-associated herpesvirus (KSHV)-infected cells identified in vivo contain latent KSHV, with lytic replication in only a few percent of cells, as is the case for the cells of Kaposi's sarcoma (KS) lesions. Factors that influence KSHV latent or lytic replication are not well defined. Because persons with KS are often immunosuppressed and susceptible to many infectious agents, including human cytomegalovirus (HCMV), we have investigated the potential for HCMV to influence the replication of KSHV. Important to this work was the construction of a recombinant KSHV, rKSHV.152, expressing the green fluorescent protein (GFP) andneo (conferring resistance to G418). The expression of GFP was a marker of KSHV infection in cells of both epithelial and endothelial origin. The rKSHV.152 virus was used to establish cells, including human fibroblasts (HF), containing only latent KSHV, as demonstrated by latency-associated nuclear antigen expression and Gardella gel analysis. HCMV infection of KSHV latently infected HF activated KSHV lytic replication with the production of infectious KSHV. Dual-color immunofluorescence detected both the KSHV lytic open reading frame 59 protein and the HCMV glycoprotein B in coinfected cells, and UV-inactivated HCMV did not activate the production of infectious KSHV-GFP. In addition, HCMV coinfection increased the production of KSHV from endothelial cells and activated lytic cycle gene expression in keratinocytes. These data demonstrate that HCMV can activate KSHV lytic replication and suggest that HCMV could influence KSHV pathogenesis.

scholarly journals Kaposi's Sarcoma-Associated Herpesvirus (Human Herpesvirus 8) Contains Hypoxia Response Elements: Relevance to Lytic Induction by Hypoxia

2003 ◽  
Vol 77 (12) ◽  
pp. 6761-6768 ◽  
Author(s):  
Muzammel Haque ◽  
David A. Davis ◽  
Victoria Wang ◽  
Isabelle Widmer ◽  
Robert Yarchoan
Keyword(s):  
Promoter Region ◽  
Kaposi's Sarcoma ◽  
Human Herpesvirus ◽  
Kaposi’S Sarcoma ◽  
Lytic Replication ◽  
Human Herpesvirus 8 ◽  
Viral Gene ◽  
Hypoxia Response ◽  
Herpesvirus 8 ◽  
Response Elements

ABSTRACT Kaposi's sarcoma (KS)-associated herpesvirus (KSHV), also known as human herpesvirus 8, is an etiologic agent of KS, primary effusion lymphoma (PEL), and multicentric Castleman's disease. We recently demonstrated that hypoxia can induce lytic replication of KSHV in PEL cell lines. Hypoxia induces the accumulation of hypoxia-inducible factors (HIF), and we hypothesized that the KSHV genome may respond to hypoxia through functional hypoxia response elements (HREs). Here, we demonstrate the presence of at least two promoters within the KSHV genome that are activated by hypoxia or hypoxia mimics. One is in the promoter region of the gene for Rta, the main lytic switch gene, and the other is within the promoter region of ORF34, a lytic gene of unknown function. The ORF34 promoter contains three putative consensus HREs oriented in the direction of the gene. Dissection and site-directed mutagenesis studies confirmed that one of the HREs of the ORF34 promoter is functional. Under conditions of hypoxia, the ORF34 promoter was strongly upregulated by HIF-1α and HIF-2α. By contrast, the promoter of the gene for Rta appeared to be preferentially upregulated by HIF-2α. Reverse transcription-PCR analysis revealed that specific messages for ORF34 and ORF50 are upregulated in BCBL-1 cells exposed to hypoxia. An HIF-1 binding and competition assay demonstrated that the HRE sequence from the ORF34 promoter can compete for HIF-1α binding to an erythropoietin HRE oligonucleotide while a mutant sequence cannot. Thus, we demonstrated that a viral gene can be activated by hypoxia through activation of a functional viral HRE. To our knowledge, this is the first example of a functional HRE in a viral promoter.

scholarly journals Principal Role of TRAP/Mediator and SWI/SNF Complexes in Kaposi's Sarcoma-Associated Herpesvirus RTA-Mediated Lytic Reactivation

2003 ◽  
Vol 23 (6) ◽  
pp. 2055-2067 ◽  
Author(s):  
Yousang Gwack ◽  
Hwa Jin Baek ◽  
Hiroyuki Nakamura ◽  
Sun Hwa Lee ◽  
Michael Meisterernst ◽  
...  
Keyword(s):  
Gene Expression ◽  
Kaposi's Sarcoma ◽  
Viral Gene Expression ◽  
Kaposi’S Sarcoma ◽  
Lytic Replication ◽  
Viral Gene ◽  
Transcription Activator ◽  
Lytic Reactivation ◽  
Kaposi's Sarcoma Associated Herpesvirus ◽  
Kaposi’S Sarcoma Associated Herpesvirus

ABSTRACT An important step in the herpesvirus life cycle is the switch from latency to lytic reactivation. The RTA transcription activator of Kaposi's sarcoma-associated herpesvirus (KSHV) acts as a molecular switch for lytic reactivation. Here we demonstrate that KSHV RTA recruits CBP, the SWI/SNF chromatin remodeling complex, and the TRAP/Mediator coactivator into viral promoters through interactions with a short acidic sequence in the carboxyl region and that this recruitment is essential for RTA-dependent viral gene expression. The Brg1 subunit of SWI/SNF and the TRAP230 subunit of TRAP/Mediator were shown to interact directly with RTA. Consequently, genetic ablation of these interactions abolished KSHV lytic replication. These results demonstrate that the recruitment of CBP, SWI/SNF, and TRAP/Mediator complexes by RTA is the principal mechanism to direct well-controlled viral gene expression and thereby viral lytic reactivation.

scholarly journals Human Herpesvirus 8 Infects and Replicates in Langerhans Cells and Interstitial Dermal Dendritic Cells and Impairs Their Function

2017 ◽  
Vol 91 (20) ◽  
Author(s):  
Giovanna Rappocciolo ◽  
Mariel Jais ◽  
Paolo A. Piazza ◽  
Diana C. DeLucia ◽  
Frank J. Jenkins ◽  
...  
Keyword(s):  
Dendritic Cells ◽  
Langerhans Cells ◽  
Kaposi's Sarcoma ◽  
Human Herpesvirus ◽  
Kaposi’S Sarcoma ◽  
Lytic Replication ◽  
Productive Infection ◽  
Receptor Protein ◽  
Human Herpesvirus 8 ◽  
Herpesvirus 8

ABSTRACT The predominant types of dendritic cells (DC) in the skin and mucosa are Langerhans cells (LC) and interstitial dermal DC (iDDC). LC and iDDC process cutaneous antigens and migrate out of the skin and mucosa to the draining lymph nodes to present antigens to T and B cells. Because of the strategic location of LC and iDDC and the ability of these cells to capture and process pathogens, we hypothesized that they could be infected with human herpesvirus 8 (HHV-8) (Kaposi's sarcoma [KS]-associated herpesvirus) and have an important role in the development of KS. We have previously shown that HHV-8 enters monocyte-derived dendritic cells (MDDC) through DC-SIGN, resulting in nonproductive infection. Here we show that LC and iDDC generated from pluripotent cord blood CD34+ cell precursors support productive infection with HHV-8. Anti-DC-SIGN monoclonal antibody (MAb) inhibited HHV-8 infection of iDDC, as shown by low expression levels of viral proteins and DNA. In contrast, blocking of both langerin and the receptor protein tyrosine kinase ephrin A2 was required to inhibit HHV-8 infection of LC. Infection with HHV-8 did not alter the cell surface expression of langerin on LC but downregulated the expression of DC-SIGN on iDDC, as we previously reported for MDDC. HHV-8-infected LC and iDDC had a reduced ability to stimulate allogeneic CD4+ T cells in the mixed-lymphocyte reaction. These results indicate that HHV-8 can target both LC and iDDC for productive infection via different receptors and alter their function, supporting their potential role in HHV-8 pathogenesis and KS. IMPORTANCE Here we show that HHV-8, a DNA tumor virus that causes Kaposi's sarcoma, infects three types of dendritic cells: monocyte-derived dendritic cells, Langerhans cells, and interstitial dermal dendritic cells. We show that different receptors are used by this virus to infect these cells. DC-SIGN is a major receptor for infection of both monocyte-derived dendritic cells and interstitial dermal dendritic cells, yet the virus fully replicates only in the latter. HHV-8 uses langerin and the ephrin A2 receptor to infect Langerhans cells, which support full HHV-8 lytic replication. This infection of Langerhans cells and interstitial dermal dendritic cells results in an impaired ability to stimulate CD4+ helper T cell responses. Taken together, our data show that HHV-8 utilizes alternate receptors to differentially infect and replicate in these tissue-resident DC and support the hypothesis that these cells play an important role in HHV-8 infection and pathogenesis.

scholarly journals Kaposi's Sarcoma-Associated Herpesvirus Reduces Cellular Myeloid Differentiation Primary-Response Gene 88 (MyD88) Expression via Modulation of Its RNA

2015 ◽  
Vol 90 (1) ◽  
pp. 180-188 ◽  
Author(s):  
Amy Lingel ◽  
Erica Ehlers ◽  
Qianli Wang ◽  
Mingxia Cao ◽  
Charles Wood ◽  
...  
Keyword(s):  
Kaposi's Sarcoma ◽  
Rna Synthesis ◽  
Kaposi’S Sarcoma ◽  
Primary Response ◽  
Lytic Replication ◽  
Myeloid Differentiation ◽  
Toll Like Receptor ◽  
Transcription Activator ◽  
Response Gene ◽  
Primary Response Gene

ABSTRACTKaposi's sarcoma (KS)-associated herpesvirus (KSHV) is a human gammaherpesvirus associated with several human malignancies. The replication and transcription activator (RTA) is necessary and sufficient for the switch from KSHV latency to lytic replication. Interleukin 1 (IL-1) is a major mediator for inflammation and plays an important role in both innate and adaptive immunity. Myeloid differentiation primary response gene 88 (MyD88) is an essential adaptor molecule for IL-1 as well as most Toll-like receptor signaling. In this study, we identified a novel mechanism by which KSHV interferes with host inflammation and immunity. KSHV RTA specifically reduces the steady-state protein levels of MyD88, and physiological levels of MyD88 are downregulated during KSHV lytic replication when RTA is expressed. The N-terminal region of RTA is required for the reduction of MyD88. Additional studies demonstrated that RTA targets MyD88 expression at the RNA level, inhibits RNA synthesis of MyD88, and may bind MyD88 RNA. Finally, RTA inhibits IL-1-mediated activation of NF-κB. Because IL-1 is abundant in the KS microenvironment and inhibits KSHV replication, this work may expand our understanding of how KSHV evades host inflammation and immunity for its survivalin vivo.IMPORTANCEMyD88 is an important molecule for IL-1-mediated inflammation and Toll-like receptor (TLR) signaling. This work shows that KSHV inhibits MyD88 expression through a novel mechanism. KSHV RTA may bind to MyD88 RNA, suppresses RNA synthesis of MyD88, and inhibits IL-1-mediated signaling. This work may expand our understanding of how KSHV evades host inflammation and immunity.

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