scholarly journals Genetic Organization and Hypoxic Activation of the Kaposi's Sarcoma-Associated Herpesvirus ORF34-37 Gene Cluster

2006 ◽  
Vol 80 (14) ◽  
pp. 7037-7051 ◽  
Author(s):  
Muzammel Haque ◽  
Victoria Wang ◽  
David A. Davis ◽  
Zhi-Ming Zheng ◽  
Robert Yarchoan
Keyword(s):  
Gene Cluster ◽  
Kaposi's Sarcoma ◽  
Primary Effusion Lymphoma ◽  
Critical Role ◽  
Kaposi’S Sarcoma ◽  
Lytic Replication ◽  
Site Directed Mutagenesis ◽  
Mobility Shift ◽  
Kaposi's Sarcoma Associated Herpesvirus ◽  
Kaposi’S Sarcoma Associated Herpesvirus

ABSTRACT Kaposi's sarcoma-associated herpesvirus (KSHV) is the etiologic agent for Kaposi's sarcoma (KS) and primary effusion lymphoma (PEL). We previously reported that hypoxia activates KSHV lytic replication and that the promoter for open reading frame 34 (ORF34) contains a functional hypoxia-responsive element (HRE). ORF34 is part of a cluster of lytic genes (ORF34-37) that includes ORF36, a phosphotransferase, and ORF37, a shutoff exonuclease. Rapid amplification of cDNA ends analysis revealed that they share a common polyadenylation signal but have two start sites. Two transcripts were identified, one 3.4 kb encoding ORF35-37, and the other 4.2 kb encoding ORF34 and also having coding potential for ORF35-37. Exposure of PEL cell lines to hypoxia induced messages of lengths consistent with those of these transcripts. Reporter assays with Hep3B cells showed activation of both transcripts by hypoxia. The ORF34-37 promoter region has six consensus HREs. Sequential deletion, site-directed mutagenesis experiments, and Northern blot analysis of RNA produced by constructs indicated that the second HRE (HRE-2) plays a critical role in the hypoxic activation of both RNA transcripts. The ORF35-37 transcript was upregulated by cotransfected hypoxia-inducible factor (HIF). Electrophoretic mobility shift assays demonstrated that HRE-2 and ancillary sequences bind and compete for HIF with hypoxic Hep3B nuclear extract. The activation of this gene cluster by hypoxia may have implications for the pathogenesis of PEL and KS. Moreover, the activation of ORF36 by hypoxia might be exploited to develop targeted therapy for PEL, which arises in a hypoxic environment (pleural effusions).

scholarly journals Lytic Replication-Defective Kaposi's Sarcoma-Associated Herpesvirus: Potential Role in Infection and Malignant Transformation

2004 ◽  
Vol 78 (20) ◽  
pp. 11108-11120 ◽  
Author(s):  
Jian-Hong Deng ◽  
Yan-Jin Zhang ◽  
Xin-Ping Wang ◽  
Shou-Jiang Gao
Keyword(s):  
Malignant Transformation ◽  
Cell Lines ◽  
Kaposi's Sarcoma ◽  
Potential Role ◽  
Primary Effusion Lymphoma ◽  
Kaposi’S Sarcoma ◽  
Lytic Replication ◽  
Screening Methods ◽  
Kaposi's Sarcoma Associated Herpesvirus ◽  
Kaposi’S Sarcoma Associated Herpesvirus

ABSTRACT Defective viruses often have pivotal roles in virus-induced diseases. Although Kaposi's sarcoma-associated herpesvirus (KSHV) is etiologically associated with Kaposi's sarcoma (KS) and primary effusion lymphoma (PEL), defective KSHV has not been reported. Using differential genetic screening methods, we show that defective KSHV is present in KS tumors and PEL cell lines. To investigate the role of defective viruses in KSHV-induced pathogenesis, we isolated and characterized a lytic replication-defective KSHV, KV-1, containing an 82-kb genomic deletion of solely lytic genes. Cells harboring KV-1 escaped G0/G1 apoptosis induced by spontaneous lytic replication occurred in cells infected with regular KSHV but maintained efficient latent replication. Consequently, KV-1-infected cells had phenotypes of enhanced cell proliferation and transformation potentials. Importantly, KV-1 was packaged as infectious virions by using regular KSHV as helpers, and KV-1-like variants were detected in cultures of two of five KSHV cell lines and 1 of 18 KS tumors. These results point to a potential role for defective viruses in the regulation of KSHV infection and malignant transformation.

scholarly journals Kaposi's Sarcoma-Associated Herpesvirus Protein LANA2 Disrupts PML Oncogenic Domains and Inhibits PML-Mediated Transcriptional Repression of the Survivin Gene

2009 ◽  
Vol 83 (17) ◽  
pp. 8849-8858 ◽  
Author(s):  
Laura Marcos-Villar ◽  
Fernando Lopitz-Otsoa ◽  
Pedro Gallego ◽  
Cesar Muñoz-Fontela ◽  
José González-Santamaría ◽  
...  
Keyword(s):  
Rna Interference ◽  
Viral Protein ◽  
Transcriptional Repression ◽  
Kaposi's Sarcoma ◽  
Primary Effusion Lymphoma ◽  
Kaposi’S Sarcoma ◽  
Lytic Replication ◽  
Negative Results ◽  
Kaposi's Sarcoma Associated Herpesvirus ◽  
Kaposi’S Sarcoma Associated Herpesvirus

ABSTRACT Infection by herpesviruses causes a dramatic disturbance of PML oncogenic domains (PODs) that has been suggested to be essential for viral lytic replication. Several proteins from Kaposi's sarcoma-associated herpesvirus (KSHV) have been tested as putative POD-disrupting factors with negative results. Here, we show that LANA2, a viral protein that is absolutely required for the viability and proliferation of KSHV-infected primary effusion lymphoma (PEL) cells, increases the levels of SUMO2-ubiquitin-modified PML and induces the disruption of PODs by a proteasome-mediated mechanism. In addition, we demonstrate that this disruption is largely dependent on both the integrity of a SUMO interaction motif in LANA2 and the lysine 160 from PML. Moreover, silencing of LANA2 expression in PEL cells by RNA interference led to an increase in the PML levels. Finally, we demonstrate that LANA2 relieves PML-mediated transcriptional repression of survivin, a protein that directly contributes to malignant progression of PEL. This represents the first example of inactivation of these important antiviral structures by KSHV.

scholarly journals Kaposi's Sarcoma-Associated Herpesvirus Latent and Lytic Gene Expression as Revealed by DNA Arrays

2001 ◽  
Vol 75 (2) ◽  
pp. 891-902 ◽  
Author(s):  
Richard G. Jenner ◽  
M. Mar Albà ◽  
Chris Boshoff ◽  
Paul Kellam
Keyword(s):  
Gene Expression ◽  
Kaposi's Sarcoma ◽  
Primary Effusion Lymphoma ◽  
Human Herpesvirus ◽  
Kaposi’S Sarcoma ◽  
Lytic Replication ◽  
Dna Arrays ◽  
Nylon Membrane ◽  
Kaposi's Sarcoma Associated Herpesvirus ◽  
Kaposi’S Sarcoma Associated Herpesvirus

ABSTRACT Kaposi's sarcoma-associated herpesvirus (KSHV; human herpesvirus 8) is associated with three human tumors, Kaposi's sarcoma, primary effusion lymphoma (PEL), and multicentric Castleman's disease. KSHV encodes a number of homologs of cellular proteins involved in the cell cycle, signal transduction, and modulation of the host immune response. Of the virus complement of over 85 open reading frames (ORFs), the expression of only a minority has been characterized individually. We have constructed a nylon membrane-based DNA array which allows the expression of almost every ORF of KSHV to be measured simultaneously. A PEL-derived cell line, BC-3, was used to study the expression of KSHV during latency and after the induction of lytic replication. Cluster analysis, which arranges genes according to their expression profile, revealed a correlation between expression and assigned gene function that is consistent with the known stages of the herpesvirus life cycle. Furthermore, latent and lytic genes thought to be functionally related cluster into groups. The correlation between gene expression and function also infers possible roles for KSHV genes yet to be characterized.

scholarly journals DNA Binding and Modulation of Gene Expression by the Latency-Associated Nuclear Antigen of Kaposi's Sarcoma-Associated Herpesvirus

2001 ◽  
Vol 75 (17) ◽  
pp. 7882-7892 ◽  
Author(s):  
Alexander C. Garber ◽  
Marla A. Shu ◽  
Jianhong Hu ◽  
Rolf Renne
Keyword(s):  
Gene Expression ◽  
Dna Binding ◽  
Kaposi's Sarcoma ◽  
Primary Effusion Lymphoma ◽  
Kaposi’S Sarcoma ◽  
Nuclear Antigen ◽  
Viral Gene ◽  
Mobility Shift ◽  
Kaposi's Sarcoma Associated Herpesvirus ◽  
Kaposi’S Sarcoma Associated Herpesvirus

ABSTRACT Kaposi's sarcoma-associated herpesvirus (KSHV) is associated with Kaposi's sarcoma, primary effusion lymphoma, and multicentric Castleman's disease. The latency-associated nuclear antigen (LANA) is highly expressed in these malignancies and has been shown to play an important role in episomal maintenance, presumably by binding to a putative oriP. In addition, LANA modulates cellular and viral gene expression and interacts with the cellular tumor suppressors p53 and retinoblastoma suppressor protein. Many of these features are reminiscent of Epstein-Barr virus nuclear antigens (EBNAs), a family of six proteins expressed during latency. EBNA-1 is required for episome maintenance, binds to oriP, and strongly activates transcription from two promoters, including its own. We have previously shown that LANA can transactivate its own promoter and therefore asked whether LANA, like EBNA-1, activates transcription by direct binding to DNA. By using recombinant LANA expressed from vaccinia virus vectors for electrophoretic mobility shift assays, we found that LANA does not bind to its own promoter. In contrast, LANA binds specifically to sequences containing an imperfect 20-bp palindrome in the terminal repeat (TR) of KSHV. We further show that the C-terminal domain of LANA is sufficient for site-specific DNA binding. Unlike EBNA-1, which activates transcription through binding of oriP, we found that LANA inhibits transcription from a single TR binding site. A multimerized TR as found in the viral genome results in strong transcriptional suppression when linked to a heterologous promoter. These data suggest that LANA, although fulfilling functions similar to those of EBNA-1, does so by very different mechanisms.

scholarly journals Kaposi's Sarcoma-Associated Herpesvirus Latent Protein LANA Interacts with HIF-1α To Upregulate RTA Expression during Hypoxia: Latency Control under Low Oxygen Conditions

2006 ◽  
Vol 80 (16) ◽  
pp. 7965-7975 ◽  
Author(s):  
Qiliang Cai ◽  
Ke Lan ◽  
Subhash C. Verma ◽  
Huaxin Si ◽  
Doug Lin ◽  
...  
Keyword(s):  
Kaposi's Sarcoma ◽  
Primary Effusion Lymphoma ◽  
Kaposi’S Sarcoma ◽  
Lytic Replication ◽  
Nuclear Antigen ◽  
Regulatory Sequences ◽  
Physical Interaction ◽  
Low Oxygen ◽  
Kaposi's Sarcoma Associated Herpesvirus ◽  
Kaposi’S Sarcoma Associated Herpesvirus

ABSTRACT Hypoxia can induce lytic replication of Kaposi's sarcoma-associated herpesvirus (KSHV) in primary effusion lymphoma (PEL) cells. However, the molecular mechanism of lytic reactivation of KSHV by hypoxia remains unclear. Here we show that the latency-associated nuclear antigen (LANA), which plays a crucial role in modulating viral and cellular gene expression, directly associated with a low oxygen responder, hypoxia-inducible factor-1α (HIF-1α). LANA enhanced not only the transcriptional activities of HIF-1α but also its mRNA level. Coimmunoprecipitation and immunofluorescence studies documented a physical interaction between LANA and HIF-1α in transiently transfected 293T cells as well as in PEL cell lines during hypoxia. Through sequence analysis, several putative hypoxia response elements (HRE-1 to -6) were identified in the essential lytic gene Rta promoter. Reporter assays showed that HRE-2 (−1130 to −1123) and HRE-5 and HRE-6 (+234 to +241 and +812 to +820, respectively, within the intron sequence) were necessary and sufficient for the LANA-mediated HIF-1α response. Electrophoretic mobility shift assays showed HIF-1α-dependent binding of a LANA protein complex specifically to the HRE-2, -5, and -6 motifs within the promoter regulatory sequences. This study demonstrates that hypoxia-induced KSHV lytic replication is mediated at least in part through cooperation of HIF-1α with LANA bound to the HRE motifs of the Rta promoter.

scholarly journals Kaposi’s Sarcoma-Associated Herpesvirus ORF7 Is Essential for Virus Production

2021 ◽  
Vol 9 (6) ◽  
pp. 1169
Author(s):  
Yuki Iwaisako ◽  
Tadashi Watanabe ◽  
Mizuki Hanajiri ◽  
Yuichi Sekine ◽  
Masahiro Fujimuro
Keyword(s):  
Dna Cleavage ◽  
Kaposi's Sarcoma ◽  
Stop Codon ◽  
Primary Effusion Lymphoma ◽  
Virus Production ◽  
Kaposi’S Sarcoma ◽  
Lytic Replication ◽  
Capsid Formation ◽  
Kaposi's Sarcoma Associated Herpesvirus ◽  
Kaposi’S Sarcoma Associated Herpesvirus

Kaposi’s sarcoma-associated herpesvirus (KSHV) causes Kaposi’s sarcoma, primary effusion lymphoma (PEL), and multicentric Castleman disease. Although capsid formation and maturation in the alpha-herpesvirus herpes simplex virus 1 are well understood, these processes in KSHV remain unknown. The KSHV ORF7, encoding the viral terminase (DNA cleavage and packaging protein), is thought to contribute to capsid formation; however, functional information is lacking. Here, we investigated the role of ORF7 during KSHV lytic replication by generating two types of ORF7 knock-out (KO) mutants (frameshift-induced and stop codon-induced ORF7 deficiency), KSHV BAC16, and its revertants. The results revealed that both ORF7-KO KSHVs showed significantly reduced viral production but there was no effect on lytic gene expression and viral genome replication. Complementation assays showed virus production from cells harboring ORF7-KO KSHV could be recovered by ORF7 overexpression. Additionally, exogenously expressed ORF7 partially induced nuclear relocalization of the other terminase components, ORF29 and ORF67.5. ORF7 interacted with both ORF29 and ORF67.5, whereas ORF29 and ORF67.5 failed to interact with each other, suggesting that ORF7 functions as a hub molecule in the KSHV terminase complex for interactions between ORF29 and ORF67.5. These findings indicate that ORF7 plays a key role in viral replication, as a component of terminase.

scholarly journals FoxO1 Suppresses Kaposi’s Sarcoma-Associated Herpesvirus Lytic Replication and Controls Viral Latency

2018 ◽  
Vol 93 (3) ◽  
Author(s):  
Ruoyun Gao ◽  
Tingting Li ◽  
Brandon Tan ◽  
Suzane Ramos da Silva ◽  
Jae U. Jung ◽  
...  
Keyword(s):  
Kaposi's Sarcoma ◽  
Critical Role ◽  
Kaposi’S Sarcoma ◽  
Oxygen Free Radical ◽  
Free Radical Scavenger ◽  
Lytic Replication ◽  
Radical Scavenger ◽  
Intracellular Ros ◽  
Kaposi's Sarcoma Associated Herpesvirus ◽  
Kaposi’S Sarcoma Associated Herpesvirus

ABSTRACT Kaposi’s sarcoma-associated herpesvirus (KSHV) has latent and lytic replication phases, both of which contribute to the development of KSHV-induced malignancies. Among the numerous factors identified to regulate the KSHV life cycle, oxidative stress, caused by imbalanced clearing and production of reactive oxygen species (ROS), has been shown to robustly disrupt KSHV latency and induce viral lytic replication. In this study, we identified an important role of the antioxidant defense factor forkhead box protein O1 (FoxO1) in the KSHV life cycle. Either chemical inhibition of the FoxO1 function or knockdown of FoxO1 expression led to an increase in the intracellular ROS level that was subsequently sufficient to disrupt KSHV latency and induce viral lytic reactivation. On the other hand, treatment with N-acetyl-l-cysteine (NAC), an oxygen free radical scavenger, led to a reduction in the FoxO1 inhibition-induced ROS level and, ultimately, the attenuation of KSHV lytic reactivation. These findings reveal that FoxO1 plays a critical role in keeping KSHV latency in check by maintaining the intracellular redox balance. IMPORTANCE Kaposi’s sarcoma-associated herpesvirus (KSHV) is associated with several cancers, including Kaposi’s sarcoma (KS). Both the KSHV latent and lytic replication phases are important for the development of KS. Identification of factors regulating the KSHV latent phase-to-lytic phase switch can provide insights into the pathogenesis of KSHV-induced malignancies. In this study, we show that the antioxidant defense factor forkhead box protein O1 (FoxO1) maintains KSHV latency by suppressing viral lytic replication. Inhibition of FoxO1 disrupts KSHV latency and induces viral lytic replication by increasing the intracellular ROS level. Significantly, treatment with an oxygen free radical scavenger, N-acetyl-l-cysteine (NAC), attenuated the FoxO1 inhibition-induced intracellular ROS level and KSHV lytic replication. Our works reveal a critical role of FoxO1 in suppressing KSHV lytic replication, which could be targeted for antiviral therapy.

scholarly journals Antiviral Activity of (+)-Rutamarin against Kaposi's Sarcoma-Associated Herpesvirus by Inhibition of the Catalytic Activity of Human Topoisomerase II

2013 ◽  
Vol 58 (1) ◽  
pp. 563-573 ◽  
Author(s):  
Bo Xu ◽  
Ling Wang ◽  
Lorenzo González-Molleda ◽  
Yan Wang ◽  
Jun Xu ◽  
...  
Keyword(s):  
Kaposi's Sarcoma ◽  
Topoisomerase Ii ◽  
Primary Effusion Lymphoma ◽  
Kaposi’S Sarcoma ◽  
Binding Mode ◽  
Lytic Replication ◽  
Associated Diseases ◽  
Catalytic Inhibitor ◽  
Kaposi's Sarcoma Associated Herpesvirus ◽  
Kaposi’S Sarcoma Associated Herpesvirus

ABSTRACTKaposi's sarcoma-associated herpesvirus (KSHV) is an etiological agent of several AIDS-associated malignancies, including Kaposi's sarcoma (KS), primary effusion lymphoma (PEL), and multicentric Castleman's disease (MCD). Its lytic replication cycle has been proven to be critical for the pathogenesis of KSHV-associated diseases. In KS lesions, lytic viral replication, production of virion particles, and reinfection of endothelial cells are essential to sustain the population of infected cells that otherwise would be quickly lost as spindle cells divide. Thus, antivirals that block KSHV replication could be a strategy in the treatment of KSHV-associated diseases. However, there is no effective anti-KSHV drug currently available. Our previous work showed that human topoisomerase II (Topo II) is indispensable for KSHV lytic replication and is suggested to be an effective target for antiviral drugs. Here, we report the discovery and characterization of a novel catalytic inhibitor of human Topo IIα, namely, (+)-rutamarin. The binding mode of (+)-rutamarin to the ATPase domain of human Topo IIα was established by docking and validated by molecular dynamics (MD) simulations. More importantly, (+)-rutamarin efficiently inhibits KSHV lytic DNA replication in BCBL-1 cells with a half-maximal inhibitory concentration (IC50) of 1.12 μM and blocks virion production with a half-maximal antiviral effective concentration (EC50) of 1.62 μM. It possesses low cytotoxicity, as indicated by the selectivity index (SI) of 84.14. This study demonstrated great potential for (+)-rutamarin to become an effective drug for treatment of human diseases associated with KSHV infection.

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