scholarly journals ORF73 of Herpesvirus Saimiri, a Viral Homolog of Kaposi's Sarcoma-Associated Herpesvirus, Modulates the Two Cellular Tumor Suppressor Proteins p53 and pRb

2004 ◽  
Vol 78 (19) ◽  
pp. 10336-10347 ◽  
Author(s):  
Sumit Borah ◽  
Subhash C. Verma ◽  
Erle S. Robertson
Keyword(s):  
Cell Cycle ◽  
Kaposi's Sarcoma ◽  
Kaposi’S Sarcoma ◽  
Herpesvirus Saimiri ◽  
Nuclear Antigen ◽  
Kaposi's Sarcoma Associated Herpesvirus ◽  
Kaposi’S Sarcoma Associated Herpesvirus ◽  
P53 Inactivation

ABSTRACT All known DNA tumor viruses are known to target and inactivate two main cell cycle regulatory proteins, retinoblastoma protein (pRb) and p53. Inactivation of pRb promotes host cell cycle progression into S phase, and inactivation of p53 promotes cell immortalization. The DNA tumor virus Kaposi's sarcoma associated herpesvirus (KSHV)-encoded latency-associated nuclear antigen (LANA) was shown to target and inactivate pRb as well as p53. In this report we provide evidence that these functions are conserved in the homologous protein encoded by the related gammaherpesvirus herpesvirus saimiri (HVS). ORF73, the HVS homologue of LANA, is shown to bind both p53 and pRb in vitro and in vivo, to colocalize with p53 in human T cells infected with HVS, and in cells overexpressing both ORF73 and p53, as well as to adversely influence pRB/E2F and p53 transcriptional regulation. The C terminus of LANA, the region most highly conserved in ORF73, is shown to be responsible for both pRb and p53 interactions, supporting the hypothesis that these functions are conserved in both homologues. Finally, the region of p53 targeted by LANA (and ORF73) maps to the domain required for tetramerization. However, preliminary cross-linking studies do not detect disruption of p53 tetramerization by either LANA or HVS-encoded ORF73, suggesting that p53 inactivation may be by a mechanism independent of tetramer disruption.

scholarly journals Kaposi's Sarcoma-Associated Herpesvirus Latency-Associated Nuclear Antigen Interacts with Bromodomain Protein Brd4 on Host Mitotic Chromosomes

2006 ◽  
Vol 80 (18) ◽  
pp. 8909-8919 ◽  
Author(s):  
Jianxin You ◽  
Viswanathan Srinivasan ◽  
Gerald V. Denis ◽  
William J. Harrington ◽  
Mary E. Ballestas ◽  
...  
Keyword(s):  
Kaposi's Sarcoma ◽  
Kaposi’S Sarcoma ◽  
Host Cells ◽  
Nuclear Antigen ◽  
Mitotic Chromosomes ◽  
Kaposi's Sarcoma Associated Herpesvirus ◽  
Kaposi’S Sarcoma Associated Herpesvirus ◽  
Chromosome Binding

ABSTRACT The latency-associated nuclear antigen (LANA) of Kaposi's sarcoma-associated herpesvirus (KSHV) is required for viral episome maintenance in host cells during latent infection. Two regions of the protein have been implicated in tethering LANA/viral episomes to the host mitotic chromosomes, and LANA chromosome-binding sites are subjects of high interest. Because previous studies had identified bromodomain protein Brd4 as the mitotic chromosome anchor for the bovine papillomavirus E2 protein, which tethers the viral episomes to host mitotic chromosomes (J. You, J. L. Croyle, A. Nishimura, K. Ozato, and P. M. Howley, Cell 117:349-360, 2004, and J. You, M. R. Schweiger, and P. M. Howley, J. Virol. 79:14956-14961, 2005), we examined whether KSHV LANA interacts with Brd4. We found that LANA binds Brd4 in vivo and in vitro and that the binding is mediated by a direct protein-protein interaction between the ET (extraterminal) domain of Brd4 and a carboxyl-terminal region of LANA previously implicated in chromosome binding. Brd4 associates with mitotic chromosomes throughout mitosis and demonstrates a strong colocalization with LANA and the KSHV episomes on host mitotic chromosomes. Although another bromodomain protein, RING3/Brd2, binds to LANA in a similar fashion in vitro, it is largely excluded from the mitotic chromosomes in KSHV-uninfected cells and is partially recruited to the chromosomes in KSHV-infected cells. These data identify Brd4 as an interacting protein for the carboxyl terminus of LANA on mitotic chromosomes and suggest distinct functional roles for the two bromodomain proteins RING3/Brd2 and Brd4 in LANA binding. Additionally, because Brd4 has recently been shown to have a role in transcription, we examined whether Brd4 can regulate the CDK2 promoter, which can be transactivated by LANA.

scholarly journals Early Events in Kaposi's Sarcoma-Associated Herpesvirus Infection of Target Cells

2009 ◽  
Vol 84 (5) ◽  
pp. 2188-2199 ◽  
Author(s):  
Bala Chandran
Keyword(s):  
Kaposi's Sarcoma ◽  
Kaposi’S Sarcoma ◽  
Target Cells ◽  
Mode Of Entry ◽  
Successful Infection ◽  
Kaposi's Sarcoma Associated Herpesvirus ◽  
Kaposi’S Sarcoma Associated Herpesvirus ◽  
Cell Signaling Pathways

ABSTRACT Kaposi's sarcoma-associated herpesvirus (KSHV), the most recently identified member of the herpesvirus family, infects a variety of target cells in vitro and in vivo. This minireview surveys current information on the early events of KSHV infection, including virus-receptor interactions, involved envelope glycoproteins, mode of entry, intracellular trafficking, and initial viral and host gene expression programs. We describe data supporting the hypothesis that KSHV manipulates preexisting host cell signaling pathways to allow successful infection. The various signaling events triggered by infection, and their potential roles in the different stages of infection and disease pathogenesis, are summarized.

scholarly journals CAK-independent Activation of CDK6 by a Viral Cyclin

2001 ◽  
Vol 12 (12) ◽  
pp. 3987-3999 ◽  
Author(s):  
Philipp Kaldis ◽  
Päivi M. Ojala ◽  
Lily Tong ◽  
Tomi P. Mäkelä ◽  
Mark J. Solomon
Keyword(s):  
Conformational Changes ◽  
Kaposi's Sarcoma ◽  
Cell Cycle Progression ◽  
Kaposi’S Sarcoma ◽  
Binding Assays ◽  
Kaposi's Sarcoma Associated Herpesvirus ◽  
Induced Apoptosis ◽  
Kaposi’S Sarcoma Associated Herpesvirus

In normal cells, activation of cyclin-dependent kinases (cdks) requires binding to a cyclin and phosphorylation by the cdk-activating kinase (CAK). The Kaposi's sarcoma-associated herpesvirus encodes a protein with similarity to D-type cyclins. This KSHV-cyclin activates CDK6, alters its substrate specificity, and renders CDK6 insensitive to inhibition by the cdk inhibitor p16INK4a. Here we investigate the regulation of the CDK6/KSHV-cyclin kinase with the use of purified proteins and a cell-based assay. We find that KSHV-cyclin can activate CDK6 independent of phosphorylation by CAK in vitro. In addition, CAK phosphorylation decreased the p16INK4asensitivity of CDK6/KSHV-cyclin complexes. In cells, expression of CDK6 or to a lesser degree of a nonphosphorylatable CDK6T177Atogether with KSHV-cyclin induced apoptosis, indicating that CDK6 activation by KSHV-cyclin can proceed in the absence of phosphorylation by CAK in vivo. Coexpression of p16 partially protected cells from cell death. p16 and KSHV-cyclin can form a ternary complex with CDK6 that can be detected by binding assays as well as by conformational changes in CDK6. The Kaposi's sarcoma-associated herpesvirus has adopted a clever strategy to render cell cycle progression independent of mitogenic signals, cdk inhibition, or phosphorylation by CAK.

scholarly journals Kaposi's Sarcoma-Associated Herpesvirus LANA2 Is a B-Cell-Specific Latent Viral Protein That Inhibits p53

2001 ◽  
Vol 75 (1) ◽  
pp. 429-438 ◽  
Author(s):  
Carmen Rivas ◽  
Ai-En Thlick ◽  
Carlo Parravicini ◽  
Patrick S. Moore ◽  
Yuan Chang
Keyword(s):  
Kaposi's Sarcoma ◽  
Primary Effusion Lymphoma ◽  
Blot Hybridization ◽  
Human Herpesvirus ◽  
Kaposi’S Sarcoma ◽  
Nuclear Antigen ◽  
P53 Overexpression ◽  
Kaposi's Sarcoma Associated Herpesvirus ◽  
Kaposi’S Sarcoma Associated Herpesvirus

ABSTRACT Kaposi's sarcoma-associated herpesvirus (KSHV), or human herpesvirus 8, is associated with three proliferative diseases ranging from viral cytokine-induced hyperplasia to monoclonal neoplasia: multicentric Castleman's disease (CD), Kaposi's sarcoma (KS), and primary effusion lymphoma (PEL). Here we report a new latency-associated 1,704-bp KSHV spliced gene belonging to a cluster of KSHV sequences having homology to the interferon regulatory factor (IRF) family of transcription factors. ORFK10.5 encodes a protein, latency-associated nuclear antigen 2 (LANA2), which is expressed in KSHV-infected hematopoietic tissues, including PEL and CD but not KS lesions. LANA2 is abundantly expressed in the nuclei of cultured KSHV-infected B cells. Transcription of K10.5 in PEL cell cultures is not inhibited by DNA polymerase inhibitors nor significantly induced by phorbol ester treatment. Unlike LANA1, LANA2 does not elicit a serologic response from patients with KS, PEL, or CD as measured by Western blot hybridization. Both KSHV vIRF1 (ORFK9) and LANA2 (ORFK10.5) appear to have arisen through gene duplication of a captured cellular IRF gene. LANA2 is a potent inhibitor of p53-induced transcription in reporter assays. LANA2 antagonizes apoptosis due to p53 overexpression in p53-null SAOS-2 cells and apoptosis due to doxorubicin treatment of wild-type p53 U2OS cells. While LANA2 specifically interacts with amino acids 290 to 393 of p53 in glutathione S-transferase pull-down assays, we were unable to demonstrate LANA2-p53 interaction in vivo by immunoprecipitation. These findings show that KSHV has tissue-specific latent gene expression programs and identify a new latent protein which may contribute to KSHV tumorigenesis in hematopoietic tissues via p53 inhibition.

scholarly journals Expression in a Recombinant Murid Herpesvirus 4 Reveals the In Vivo Transforming Potential of the K1 Open Reading Frame of Kaposi's Sarcoma-Associated Herpesvirus

2004 ◽  
Vol 78 (16) ◽  
pp. 8878-8884 ◽  
Author(s):  
Jill Douglas ◽  
Bernadette Dutia ◽  
Susan Rhind ◽  
James P. Stewart ◽  
Simon J. Talbot
Keyword(s):  
Kaposi's Sarcoma ◽  
Fluorescent Protein ◽  
Salivary Gland Tumor ◽  
Kaposi’S Sarcoma ◽  
Common Marmoset ◽  
Kaposi's Sarcoma Associated Herpesvirus ◽  
Herpesvirus 4 ◽  
Kaposi’S Sarcoma Associated Herpesvirus

ABSTRACT Murid herpesvirus 4 (commonly called MHV-68) is closely related to Kaposi's sarcoma-associated herpesvirus (KSHV) and provides an excellent model system for investigating gammaherpesvirus-associated pathogenesis. MHV-76 is a naturally occurring deletion mutant of MHV-68 that lacks 9,538 bp of the left end of the unique portion of the genome encoding nonessential pathogenesis-related genes. The KSHV K1 protein has been shown to transform rodent fibroblasts in vitro and common marmoset T lymphocytes in vivo. Using homologous recombination techniques, we successfully generated recombinants of MHV-76 that encode green fluorescent protein (MHV76-GFP) and KSHV K1 (MHV76-K1). The replication of MHV76-GFP and MHV76-K1 in cell culture was identical to that of MHV-76. However, infection of BALB/c mice via the intranasal route revealed that MHV76-K1 replicated to a 10-fold higher titer than MHV76-GFP in the lungs at day 5 postinfection (p.i.). We observed type 2 pneumocyte proliferation in areas of consolidation and interstitial inflammation of mice infected with MHV76-K1 at day 10 p.i. MHV76-K1 established a 2- to 3-fold higher latent viral load than MHV76-GFP in the spleens of infected mice on days 10 and 14 p.i., although this was 10-fold lower than that established by wild-type MHV-76. A salivary gland tumor was present in one of four mice infected with MHV76-K1, as well as an increased inflammatory response in the lungs at day 120 p.i. compared with that of mice infected with MHV-76 and MHV76-GFP.

scholarly journals Exploring the DNA Binding Interactions of the Kaposi's Sarcoma-Associated Herpesvirus Lytic Switch Protein by Selective Amplification of Bound Sequences In Vitro

2006 ◽  
Vol 80 (6) ◽  
pp. 2958-2967 ◽  
Author(s):  
Joseph Ziegelbauer ◽  
Adam Grundhoff ◽  
Don Ganem
Keyword(s):  
Protein Interactions ◽  
Kaposi's Sarcoma ◽  
Genomic Library ◽  
Consensus Sequence ◽  
Kaposi’S Sarcoma ◽  
Genomic Libraries ◽  
Kaposi's Sarcoma Associated Herpesvirus ◽  
Kaposi’S Sarcoma Associated Herpesvirus

ABSTRACT The lytic switch protein RTA of Kaposi's sarcoma-associated herpesvirus (KSHV) can be targeted to DNA by either direct sequence-specific recognition or via protein-protein interactions with host transcription factors. We have searched for sequences capable of direct RTA binding by screening synthetic oligonucleotide pools and KSHV genomic libraries for RTA-interacting elements, using repeated cycles of in vitro binding followed by amplification of the bound sequences. Multiple low-affinity sequences were recovered from the random pools, with generation of only a weak consensus sequence. The genomic library, by contrast, yielded many biologically relevant fragments, most of which could be shown to interact with RTA in vitro and some of which likely play important regulatory roles in vivo. Surprisingly, the most highly selected fragment came from the promoter of a late gene (gB) and contained at least two direct RTA binding sites, as well as one RBP-Jκ binding site. This raises the possibility that some late KSHV genes may also be subject to direct RTA regulation, though indirect models are not excluded.

scholarly journals Kaposi's Sarcoma-Associated Herpesvirus Encodes a Viral Deubiquitinase

2009 ◽  
Vol 83 (19) ◽  
pp. 10224-10233 ◽  
Author(s):  
Carlos M. González ◽  
Ling Wang ◽  
Blossom Damania
Keyword(s):  
Kaposi's Sarcoma ◽  
Kaposi’S Sarcoma ◽  
Lytic Replication ◽  
Cell Fractionation ◽  
Reading Frame ◽  
Latently Infected ◽  
Kaposi's Sarcoma Associated Herpesvirus ◽  
Kaposi’S Sarcoma Associated Herpesvirus

ABSTRACT Kaposi's sarcoma-associated herpesvirus (KSHV) is etiologically linked to Kaposi's sarcoma, primary effusion lymphomas, and multicentric Castleman's disease. Like other herpesviruses, KSHV can exist in either a lytic or a latent phase during its life cycle. We report that the lytic protein encoded by KSHV open reading frame 64 (Orf64) is a viral deubiquitinase (DUB) enzyme capable of deubiquitinating cellular proteins in vitro and in vivo. Orf64 DUB activity is effective against lysine 48 (K48)- and lysine 63 (K63)-linked ubiquitin chains. Thus, KSHV Orf64 is a viral DUB that does not show specificity toward K48 or K63 ubiquitin linkages. Orf64 DUB activity lies within the first 205 residues of the protein, and deubiquitination is dependent on a cysteine at position 29, since mutation of this residue ablated this activity. Cell fractionation studies revealed that the N terminus and the full-length protein localized to both the nuclear and cytoplasmic compartments. The function of Orf64 was tested by short interfering RNA (siRNA) knockdown studies on latently infected cells that were induced into lytic replication. We found that depletion of Orf64 by siRNA resulted in decreased viral lytic transcription and lytic protein expression. These experiments indicate that Orf64 plays a role in KSHV lytic replication.

scholarly journals The Kaposi's Sarcoma-Associated Herpesvirus LANA Protein Stabilizes and Activates c-Myc

2007 ◽  
Vol 81 (19) ◽  
pp. 10451-10459 ◽  
Author(s):  
Jianyong Liu ◽  
Heather J. Martin ◽  
Gangling Liao ◽  
S. Diane Hayward
Keyword(s):  
Cell Cycle ◽  
Kaposi's Sarcoma ◽  
Cell Cycle Progression ◽  
Glycogen Synthase ◽  
Primary Effusion Lymphoma ◽  
Kaposi’S Sarcoma ◽  
Nuclear Antigen ◽  
Erk Phosphorylation ◽  
Kaposi's Sarcoma Associated Herpesvirus ◽  
Kaposi’S Sarcoma Associated Herpesvirus

ABSTRACT The Kaposi's sarcoma-associated herpesvirus (KSHV) latency-associated nuclear antigen (LANA) protein is functionally pleiotropic. LANA contributes to KSHV-associated pathogenesis, in part, by increasing entry of cells into S phase through a process that is driven by LANA interaction with the serine-threonine kinase glycogen synthase kinase 3 (GSK-3) and stabilization of β-catenin. We now show that LANA affects the activity of another protein involved in cell cycle regulation, c-Myc. Sequencing of c-Myc coding sequences revealed that c-Myc in KSHV-positive primary effusion lymphoma (PEL) cell lines is wild type in the N-terminal region that regulates c-Myc protein stability. Despite this, c-Myc in PEL cells is stabilized. In LANA-expressing cells, inactivation of nuclear GSK-3 reduced phosphorylation of c-Myc at Thr58 and contributed to c-Myc stabilization by decreasing c-Myc ubiquitination. Phosphorylation of c-Myc on Ser62 also affects c-Myc stability and function. We now show that LANA increases the level of phosphorylated extracellular signal-regulated kinase 1 (ERK1) and increases ERK phosphorylation of c-Myc on Ser62. LANA also interacted with c-Myc, and c-Myc amino acids 147 to 220 were required for this interaction. LANA (L1006P) retained the ability to bind to c-Myc and activate ERK1, indicating that these events did not require LANA interaction with GSK-3. Thus, LANA stabilizes c-Myc; prevents the phosphorylation of c-Myc at Thr58, an event that promotes Myc-induced apoptosis; and independently stimulates phosphorylation of c-Myc at Ser62, an event that transcriptionally activates c-Myc. LANA-mediated manipulation of c-Myc function is likely to contribute to KSHV-associated tumorigenesis through the induction of c-Myc regulated cellular genes, as well as by the stimulation of cell cycle progression.

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