Modulation of Cell Signaling Pathways by Kaposi's Sarcoma ...

INTRODUCTION

Kaposi’s sarcoma-associated herpesvirus(KSHV), also known as human herpesvirus 8,was originally isolated from Kaposi’s sarcomalesions using representational differenceanalysis (1). KSHV has been linked to thedevelopment of Kaposi’s sarcoma (2–4) andlymphoproliferative disorders such as primary

effusion lymphoma (PEL) and multicentricCastleman’s disease (MCD) (5,6). KSHV hasbeen classified as a gammaherpesvirus, basedon genomic organization and sequence infor-mation. The gammaherpesviruses are lym-photropic, although most are also capable ofreplicating in epithelial or fibroblast cells tosome degree. There are two classes of gamma-herpesviruses: γ-1 (lymphocryptoviruses) and γ-2 (rhadinoviruses) based on their biologicalproperties and genomic organization. Anotherhuman virus, Epstein-Barr virus (EBV or humanherpesvirus 4) is a γ-1 herpesvirus, whereas theγ-2 subgroup includes KSHV, rhesus monkey

Modulation of Cell Signaling Pathways by Kaposi’sSarcoma-Associated Herpesvirus (KSHV/HHV-8)

Blossom Damania*,1

Lineberger Comprehensive Cancer Center, CB #7295, University of North Carolina, Chapel Hill, NC

Abstract

Kaposi’s sarcoma-associated herpesvirus (KSHV), also known as human herpesvirus 8, hasbeen associated with the development of Kaposi’s sarcoma, pleural effusion lymphoma, and mul-ticentric Castleman’s disease. KSHV is a double-stranded DNA virus that has been classified as agammaherpesvirus. The viral genome is approx 160 kb long and encodes for several genes that areinvolved in cell signaling pathways. These include genes that are unique to the virus as well asviral homologues of cellular genes. The latter are likely to have been usurped from the hostgenome and include both virokines and viral receptor proteins. This article reviews how theseKSHV proteins modulate cellular signal transduction pathways.

Index Entries: Kaposi’s sarcoma-associated herpesvirus (KSHV); human herpesvirus (HHV-8);herpesviruses; signal transduction; cytokines.

*Author to whom all correspondence and reprintrequests should be addressed. E-mail: [email protected].

1Article received 07/01/03; accepted 09/02/03.

REVIEW ARTICLE

© Copyright 2004 by Humana Press Inc.All rights of any nature whatsoever reserved.1085-9195/04/40/305–322/$25.00

Cell Biochemistry and Biophysics 305 Volume 40, 2004

rhadinovirus (RRV), herpesvirus saimiri (HVS),and murine herpesvirus-68 (7–10).

Herpesviruses have latent and lytic phasesto their life cycle. A hallmark of all her-pesviruses is their ability to establish a life-long latent infection in the host. During latentinfection, viral gene expression is highlyattenuated and the viral genome remains sta-bly associated with the cell. In the lytic phase,viral gene expression and DNA replicationensue, leading to the production of progenyvirions and eventual lysis of the cell.Pathogenesis caused by these viruses is usu-ally seen in the context of host immunosup-pression or cross-species transmission. EBV,KSHV, and HVS are among the most widelystudied gammaherpesviruses to date. Allthree viruses have been shown to be associ-ated with a wide variety of cancers. HVS andEBV have also been shown to transform lym-phoid cells in culture and to induce lympho-proliferative diseases in the natural orexperimental host. The striking correlationbetween gammaherpesviruses and diseaseinduction in primates enables a study of thecontributions of individual herpes viral genesto cell growth transformation.

Since 1994, KSHV DNA sequences havebeen widely identified in KS tumors from HIV-positive and HIV-negative patients (1,11,12).In addition, KSHV has also been consistentlyfound in AIDS-associated lymphoproliferativediseases such as PELs or body cavity-basedlymphomas (BCBLs) and lymphoblastic vari-ants of MCD (5,13–16). Although it remainscontroversial, KSHV may also be associatedwith multiple myeloma (17–19). BCBLs werefirst identified in AIDS patients and were laterfound to have a high incidence of EBV andKSHV coinfection, although some lymphomaswere only positive for KSHV infection (5,20).BCBLs are thought to be monoclonal B cells inorigin and lack many B-lymphocyte antigenssuch as CD19, CD20, and lymphocyte homingand adhesion markers (21,22). MCD is anatypical lymphoproliferative disorder thatincludes hyperplasia, lymphadenopathy, andsplenomegaly. Both HIV-infected and HIV-

uninfected individuals develop MCD, andthere is a high rate of KSHV infection in thelymph nodes of HIV patients with MCD(6,16,23). MCD is also principally or exclu-sively of B-cell origin. Different from theselymphomas, the KS lesion comprises a mixedcell phenotype. One unusual cell consistentlypresent is a spindle-shaped cell of endothelialorigin. Spindle cells in the KS lesion containKSHV genetic information. There is a highlevel of cytokine and chemokine expressionwithin KS lesions and a dependence on thesecytokines and chemokines for the mainte-nance of the lesion (1,24). In addition, KSHVhas been shown to immortalize primaryhuman endothelial cells to long-term prolifer-ation and survival (25). A herpesvirus namedRRV was isolated, that is related to, but dis-tinct from KSHV (8,9,26). Homologues ofKSHV from other macaque species have alsobeen identified (27–29). Complete DNAsequence analysis of RRV shows that it is veryclosely related to KSHV (8,9). The transcrip-tional program of RRV has been shown to beextremely similar to that of KSHV (30), whichsupports its role as a suitable KSHV model.

The KSHV viral genome is comprised of anapprox 140-kb long unique region flanked bymultiple terminal repeat sequences with thetotal genomic size being approx 160–170 kb.KSHV has at least 80 open-reading frames(ORFs) that encode for proteins greater than100 amino acids (31). Some of the viral genesencoded by KSHV are common to all her-pesviruses and another set of genes is uniqueto this virus, whereas a third subset of KSHV-encoded genes are homologous of cellulargenes. It is thought that the latter are likely tohave been usurped from the host genome.Viral homologs of cellular genes fall into dif-ferent classes such as cell-cycle regulatorygenes (e.g., v-cyclin), antiapoptotic genes (e.g.,v-Bcl2), immune modulatory genes (e.g., vIRF(32)), and genes involved in signaling and cellproliferation (e.g., viral G protein-coupledreceptor, viral interleukin [vIL]-6). Genesunique to KSHV are designated by the letter“K.” Many of these unique genes are also

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involved in the aforementioned cellular path-ways. Here we review KSHV genes that havebeen shown to modulate signaling pathways.These include both viral homologs of cellulargenes and viral genes unique to KSHV.

KSHV K1 AND K15: SIGNALINGPROTEINS LOCATED AT DISTAL ENDSOF THE VIRAL GENOME

KSHV K1

A striking feature of the gammaher-pesviruses KSHV, RRV, EBV, and HVS is thatthey all contain a distinct ORF at the left end oftheir respective genomes, each of which hascharacteristic transforming ability. These ORFscode for the K1 protein of KSHV, the R1 proteinof RRV, the latent membrane protein-1 of EBV,and Saimiri transformation protein (STP) ofHVS (33–35). All of these proteins are capableof transforming cells and activating cellularsignaling pathways. This functional conserva-tion exists in the complete absence of anysequence homology because KSHV K1 andRRV R1 can substitute for STP in the context ofHVS (36,37).

The first ORF of the KSHV genome encodesfor the K1 protein, which is a 46-kd transmem-brane glycoprotein that is predicted to have asignal peptide sequence at the amino terminus,an extracellular domain, transmembranedomain, and a short cytoplasmic tail at the car-boxyl terminus (37,38). Sequence analysis hasdemonstrated that the extracellular domain ofthe K1 protein is extremely variable, showing asmuch as 40% divergence at the amino acid level(39,40). This is likely a result of its proximity tothe terminal repeats of the viral genome, a regionof high mutagenicity arising from the fact thatthese repetitive sequences undergo homologousrecombination during the viral life cycle.Although the extracellular domain of K1 isextremely variable, the cytoplasmic tail is rela-tively well-conserved (37,39–42). This carboxy-terminal cytoplasmic tail contains a functionalimmunoreceptor tyrosine-based activation motif(ITAM) (43,44). The K1 ITAM can transduce sig-

nals to induce nuclear factor of activated T-cells,nuclear factor-κB (NF-κB), calcium mobilization,and tyrosine phosphorylation, events that areindicative of lymphocyte activation (43–45).However, unlike other ITAM-based signal trans-duction events, which require a ligand-receptorinteraction, K1 signaling occurs constitutively(Fig. 1A) and K1 is thought to oligomerizethrough disulfide bonding of its extracellulardomain (37,43,44). It is controversial as towhether ITAM-dependent signaling by K1 alsoplays a role in viral lytic reactivation or sup-pression (46,47). K1 is transcribed during theKSHV lytic cycle but may also be expressed atlow levels in latent cells (48). K1 has also beenshown to upregulate vascular endothelialgrowth factor (VEG) and matrix metallopro-teinase 9 (MMP-9) in epithelial and endothelialcells (147).

The K1 protein has been shown to inducephosphorylation of several cellular signal trans-duction proteins, including vav, p85, syk, andAkt kinase (43,44,49). These adaptor moleculesare normally involved in B-cell receptor (BCR)-mediated signal transduction. Samaniego et al.found that cells expressing K1 showedincreased NF-κB–dependent promoter activity(45). NF-κB is responsible for activating a num-ber of inflammatory response genes. K1 maytherefore activate uninfected endothelial cells ina paracrine manner through the activation ofNF-κB–dependent promoters and secretion ofinflammatory cytokines (45).

In addition to activating signaling pathways,K1 has been shown to transform rodent fibrob-lasts (37). It can also functionally substitute forSTP in HVS for the immortalization of com-mon marmoset T-lymphocytes to interleukin(IL)-2–independent growth and for the induc-tion of lymphomas in common marmosets (37).RRV R1, the simian homolog of KSHV K1, alsohas similar transforming and signaling activi-ties as KSHV K1 (36,50).

Of 13 transgenic mice expressing K1 underthe transcriptional control of the simian virus40 promoter, only 2 animals developedtumors after 14 mo. One K1 transgenic mousedeveloped a spindle-cell sarcomatoid tumor

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and a second animal developed malignantplasmablastic lymphoma (51). B-lymphocytesfrom these K1-transgenic mice showed consti-tutive activation of NF-κB and Oct-2 transcrip-tion factors. These B-lymphocytes also showedincreased phosphorylation of Lyn, a Src familytyrosine kinase, and enhanced Lyn activity(51). Thus it appears that the KSHV K1 proteincan induce a cell signaling cascade, which maylikely be the mechanism by which it is able toinduce morphological changes and foci forma-tion in vitro.

KSHV K15

At the opposite end of the genome from K1lies the KSHV K15 gene. It is located in thesame genomic position as the EBV-latentmembrane protein-2A gene (39,52,53) andabuts the right hand terminal repeats.

Although K15 isolates exhibit an extremelycomplex splicing pattern, all isoforms consistof 4 to 12 transmembrane spanning domainsand a short stretch of cytoplasmic domain(Fig. 1B) (39,52,53). K15 is weakly expressed inlatently infected BCBLs, and the level of itsexpression was significantly increased by 12-O-tetradecanoylphorbol-13-acetate (TPA) stim-ulation (52,53). K15 from different KSHVisolates exhibit dramatic sequence variation,showing as much as 60–70% divergence at theamino acid level (39). However, analogous toK1, the cytoplasmic effector domain of K15,which contains signaling motifs, is conservedin most isolates (39). These include SH2 andSH3 binding motifs and a YASIL sequence(39,53,54). The cytoplasmic domain of K15 isconstitutively tyrosine phosphorylated, andthe tyrosine residue within the putative SH2binding motif is indeed a major site of phos-

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Fig. 1. The Kaposi’s sarcoma-associated herpesvirus (KSHV) K1 and K15 signaling proteins. (A) Aschematic diagram of the KSHV K1 protein. The K1 protein has an extracellular domain, transmem-brane domain, and a cytoplasmic tail. The cytoplasmic tail contains an immunoreceptor tyrosine-basedactivation motif (ITAM). K1 signaling induces activity of NF-κB and NFAT and results in B-lympho-cyte activation. (B) The KSHV K15 protein. K15 is a 12-transmembrane domain protein and contains acytoplasmic tail that is capable of associating with major B-cell receptor (BCR)-associated kinases andinhibiting BCR signaling activity. The K15 C-terminus can also interact with TNF receptor-associatedfactors 1, 2, and 3.

phorylation by cellular tyrosine kinases(52,54). In addition, experiments with CD8-K15 chimeras indicate that the cytoplasmicdomain of K15 is capable of inhibiting BCRsignal transduction (52). KSHV K15 can inter-act with major B-cell receptor-associatedkinases and alter their signaling activity(52,53). This inhibition requires the putativeSH2/SH3 binding motifs present in the cyto-plasmic region of K15. The cytoplasmic tailcan also interact with tumor necrosis factorreceptor-associated factors 1, 2, and 3 (52,53).The ability of K15 to antagonize BCR signal-ing events may help prevent the virus fromundergoing reactivation from latency becauseactivation of the B cell through BCR signalingis one plausible mechanism by which thevirus could be reactivated. In addition, K15has been shown to bind an antiapoptotic pro-tein, HAX-1 (55), and hence may also play arole in cell survival of the virus-infected cell.

Virocrines: Virokines and Viral Receptors

Gammaherpesviruses encode homologs ofcellular genes that are likely to have been cap-tured from the host. Some of these genes haveextensive amino acid similarity to cellular genes,whereas others show more diversity. AlthoughEBV contains only a few homologs of cellulargenes, KSHV and HVS both contain numerousviral counterparts of cellular genes that may con-tribute to the deregulation of cellular growth.

Cytokines play a critical role in the regula-tion of immune responses and are importanttargets of virus immune evasion mechanisms.One strategy used by these gammaherpes-viruses is to encode virocrines, which com-prise virokines and viral receptor proteins(56). Virokines are viral proteins that mimiccellular cytokines and chemokines, whereasviral receptors mimic cellular receptors.

KSHV Viral G Protein-Coupled Receptor

KSHV viral G protein-coupled receptor(vGPCR) ORF 74 encodes for a receptor withhigh sequence homology to the IL-8 cellularreceptor (Fig. 2A) (57,58). It is a 7-transmem-

brane receptor protein (Fig. 2B) that is consti-tutively active and does not require ligandbinding for its activity (59), although it canbind to both the CXC and CC families ofchemokines (59–61). Geras-Raaka et al.showed that human interferon (IFN)-γ-inducible protein 10 (HuIP-10), vMIP-II, andstromal derived growth factor-1 inhibit KSHVvGPCR signaling (62–64). Through activationof the phospholipase C and phosphatidylinos-itol 3-kinase pathways, the KSHV vGPCRprotein tickles several cell signaling networksincluding the protein kinase C and proteinkinase B, Akt, and mitogen-activated proteinkinase (MAPK) pathways. This eventuallyleads to increased transcription of cellulargenes involved in cell proliferation, immortal-ization, and transformation (59–61,65–73).Indeed, expression of the KSHV vGPCR genein rat kidney cells and NIH 3T3 fibroblastsinduced cellular proliferation indicative of atransforming function for vGPCR (59,65,68).In addition, endothelial cells expressingvGPCR were further able to promote immor-talization and tumor formation by cellsexpressing KSHV latent genes, suggestive of acooperative role among viral genes in the pro-motion of Kaposi’s sarcomagenesis (66,68).Most interestingly, transformation induced byvGPCR is associated with an increased secre-tion of vascular endothelial growth factor(VEGF), which leads to the induction of anangiogenic response in cell culture and innude mice (65,74). Transgenic mice expressingvGPCR within hematopoietic cells developedangioproliferative lesions morphologicallysimilar to KS lesions (75). In addition, trans-genic mice expressing vGPCR under anendothelial-specific promoter or the SV40 pro-moter also develop angioproliferative lesionsthat resemble human KS (68,76).

KSHV vGPCR-mediated signaling has beenshown to activate two protein kinases,JNK/SAPK and p38MAPK, characteristic ofgeneral inflammatory responses (65,70). Thereare a number of autocrine and paracrine factorsthat are induced in response to vGPCR expres-sion. These include IL-1β, IL-6, IL-8, tumor



necrosis factor-α, granulocyte-macrophagecolony-stimulating factor (GMCSF), vascularendothelial growth factor (VEGF), and basicfibroblast growth factor (bFGF) (65,69,71–74,77).

Finally, the cellular environment in whichvGPCR signals appears to play a significantrole in which downstream genes are activated.Two CC chemokines were very highly upregu-

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Fig. 2. The Kaposi’s sarcoma-associated herpesvirus (KSHV) G-protein coupled receptor. (A) AClustal W alignment of the KSHV viral G-protein coupled receptor (vGPCR) and the human inter-leukin-8α receptor. Identical amino acids are depicted by “*”, “:” represents conserved substitu-tions, and “.” represents semiconserved substitutions. The NCBI accession numbers for vGPCR andinterleukin-8α are AF148805 and AAH28221, respectively. (B) A schematic diagram of the vGPCRprotein with seven transmembrane spanning domains.

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lated in B-lymphocytes by vGPCR signaling,whereas GROα and IL-6 were induced byvGPCR in endothelial cells (78). In PEL cells,vGPCR activated the ERK-2 and p38 signalingpathways and activated AP-1, NF-κB, cAMPresponse element-binding protein (CREB), andnuclear factor of activated T-cell transcriptionfactors (79).

KSHV vIL-6

Cellular IL-6 has been implicated in many B-lymphocyte-associated malignancies in whichit has been found to stimulate the growth oflymphomas, myelomas, and leukemias (80). AKSHV-encoded vIL-6 shows homology to cel-

lular IL-6 (Fig. 3A). vIL-6 has been shown to besecreted from BCBLs and can support prolifer-ation of IL-6-dependent mouse myeloma celllines (80–82). Viral IL-6, in contrast to cellularIL-6, protects KSHV-infected PELs as well asheterologous cells from the antiviral affects ofIFNα, which has been shown to downregulatesurface levels of the IL-6 receptor, gp80 (83).In addition, Chatterjee et al. showed that IFNαcan activate transcription of vIL-6 through theIFN-stimulated response elements in the vIL-6promoter (83). Despite their similarity insequence and function, cellular IL-6 and vIL-6display differences in receptor usage.Although cellular IL-6 absolutely requires boththe IL-6Rα and the gp130 subunits, vIL-6



Fig. 3. The Kaposi’s sarcoma-associated herpesvirus (KSHV) viral interleukin (vIL)-6 cytokine.(A) A Clustal W alignment of KSHV vIL-6 and human IL-6 (hIL-6). Identical amino acids aredepicted by “*”, “:” represents conserved substitutions and “.” represents semiconserved substitu-tions. “_” represents missing amino acids. The NCBI accession numbers for vIL-6 and hIL-6 areNC_003409 and NP_000591, respectively. (B) A schematic diagram of vIL6 interacting with gp130and stimulating B-cell proliferation. There is no requirement for the gp80α receptor subunit.

appears to require only gp130 (Fig. 3B) (84–92).In addition, vIL-6 has been shown to activateJak1, STAT1, and STAT3 phosphorylation inhepatoma cells (89,91). This viral cytokine hasalso been shown to induce secretion of humanIL-6 and to support IL-6-dependent cell lines(93,94). In the context of KSHV disease, vIL-6supports the growth of PELs and is also highlyexpressed in MCD, where it appears to con-tribute to progression of this disease (95). KSlesions express vIL-6 in the minor fraction oflytic cells (96–98). Furthermore, vIL-6 has beenshown to promote hematopoiesis and angio-genesis in athymic mice (99,100). Thus vIL-6 isa multifunctional cytokine that potentiallycontributes to KSHV-associated disease pro-gression by continuously stimulating IL-6receptor signaling pathways and preventingapoptosis of virus-infected cells.

KSHV Viral Macrophage InflammatoryProteins

KSHV encodes several viral macrophageinflammatory proteins (vMIPs). KSHV ORFsK4 (vMIP-1), K4.1 (v-MIP-III) and K6 (vMIP-II)encode chemokines showing homology to cel-lular CC chemokines such as MIP-1α andRANTES.

The role of vMIPs in host inflammatoryresponses has been investigated. Unlike cellularMIP-1α, vMIPs I and II bind efficiently to CCR8(101–104). vMIP-II has also been shown to elicita potent chemoattractive effect on eosinophils(101). In contrast to cellular MIP-1α andRANTES, both vMIP-I and vMIP-II are highlyangiogenic in the chorioallantoic assay (101).vMIP-1 induces the secretion of VEGF-A inPELs and blocks dexamethasone-inducedapoptosis in these cells (100). vMIP III is alsoangiogenic in the chorioallantoic assay, bindsthe CCR4 chemokine receptor, and is involvedin chemoattraction of the Th2 subset of T cells(101). An immunomodulatory role for vMIP-IIhas also been proposed in directing inflamma-tory cell recruitment away from a Th1-typeresponse toward a Th2-type response, therebyfacilitating evasion from cytotoxic reactions

(105). vMIP-I and vMIP-II have also beenshown to induce signal transduction andchemotaxis in monocytic cells and may inducethe chemotaxis of CCR5-expressing monocytes(106). Finally, HIV-1 transmission studies haveshown that similar to cellular MIP-1, vMIP-I,and -II inhibits replication of HIV-1 strainsdependent on the CCR3 and CCR5 coreceptor(80,82,101,107). Thus these multiple vMIPslikely contribute to KS pathogenesis, inflamma-tory infiltration, and angiohyperplasia and mayalso have relevance to KSHV and HIV-1 inter-actions (108,109).

KSHV Viral Interferon Response Factors

KSHV encodes for a unique series of genesnot found in HVS or EBV called viral interferonregulatory factors (vIRFs). These include vIRF-1/K9, vIRF-2/K11.1, and vIRF-3/LANA-2(80,110–114). The KSHV Orf K9 gene has highhomology to the restricted regions of cellularinterferon regulatory factors. Cellular inter-feron signaling is thought to play a significantrole in preventing cellular transformation andin inducing antiviral responses in the infectedhost. KSHV vIRFs act as viral transcription fac-tors that inhibit interferon signaling but do notdirectly bind to interferon-stimulated responseelements located in interferon-regulated pro-moters (111,114,116). Stable expression of vIRF-1 in rodent fibroblasts induced transformation,resulting in focus formation, growth on softagar, and tumor induction in nude mice(111,114,115). Recent reports have shown thatKSHV vIRF-1 interacts with the cellular tran-scriptional coactivator p300 and its homologCBP (111,117–119). This interaction induces cel-lular myc protooncogene expression andrepresses transcriptional activation of cellularinterferon gene expression, thereby downregu-lating host antiviral activity and facilitating cellgrowth transformation (111,117–119). ThevIRF-1 and vIRF-3 proteins bind p53 andinhibit its apoptotic and transcriptional func-tion (113,120). The vIRF-1 was expressed inmost KS lesions, in contrast to vIRF-3/LANA-2, which was only transcribed in KSHV-associ-

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ated lymphomas (121). This suggests that thedifferent vIRFs encoded by KSHV are impor-tant for various aspects of KSHV-associatedmalignancies depending on the tissue environ-ment of the virus.

KSHV LATENCY-ASSOCIATED GENES:LANA, vCYCLIN, vFLIP, AND KAPOSIN

KSHV Latency-Associated Nuclear Antigen

The latency-associated nuclear antigen(LANA) of KSHV is a nuclear phosphoproteinthat has been shown to be essential for epi-some maintenance and segregation of theKSHV episome by binding to the KSHV termi-nal repeat regions (122–127). Among manyother cellular proteins, LANA has been shownto bind to Rb, p53, and Ring3 (128–130). LANAhas been shown to upregulate expression of β-catenin and to stabilize it by sequestering itsinhibitor, GSK-3β. Thus LANA appears to tapinto numerous cell cycle pathways and is theguardian of the KSHV latent life cycle.

KSHV-Encoded Viral Cyclin

The KSHV-encoded viral cyclin (vCyclin) ishomologous to cellular cyclin D. vCyclin hasbeen shown to bind and activate CDK6(131,132). vCyclin has also been shown toinduce the phosphorylation of Rb, histone H1,and Bcl-2, and degrade p27 (Kip) (133–136).Inactivation of Bcl-2 may help explain whyexpression of vCyclin in most cells results inapoptosis. The viral cyclin has also been shownto deregulate growth suppressive signals byinactivating STAT3 (137). Finally, vCyclin cancooperate with p53 loss in transgenic mice toinduce B-cell lymphomas (138).

KSHV Viral FADD-Like Interleukin-1Converting Enzyme Inhibitory Protein

The KSHV viral FADD-like interleukin-1converting enzyme inhibitory protein (vFLIP)is homologous to cellular FLIPs and encodes

a death effector domain. vFLIP has beendemonstrated to thwart Fas-mediated apopto-sis (139), and it is postulated that vFLIP mayfunction by competing with cellular FLIPs forcaspase-8 and thereby blocking apoptosis.vFLIP has been shown to activate the NF-κBpathway by interaction with I-κB kinase(140,141). In addition to its effects on the NF-κB pathway, vFLIP also modulates theJNK/AP1 pathway and induces expression ofcellular IL-6 (142).

KSHV Kaposin/K12

Kaposin is one of the few proteins expressedby the majority of KSHV-infected latent cellsand is a highly polymorphic protein. It has beenshown to consist of several protein isoformsthat differentially express two optional 23-amino acid (aa) repeats (kaposin A, B, and C),with the largest protein product being 48kDa(143). The smallest 6 kDa isoform expressedfrom the K12 locus, kaposin A, was previouslyfound to be transforming in vitro in Rat-3fibroblasts and in nude mice (144–146).Phenotypic changes induced by the kaposin Aprotein are mediated through its direct interac-tion with cytohesin-1, a guanine nucleotideexchange factor for ADP-ribosylation factor(Arf) (ARF GTPases) and regulator of integrin-mediated cell adhesion (144). Liposome-embedded kaposin A specifically stimulatescytohesin-1-dependent guanosine 5′-triphos-phate (GTP) binding of myristoylated ARF1 invitro (144). Biochemical assays have shown thatthis is mediated through the ability of KaposinA to activates the Erk1/2 pathway (144).

CONCLUSION

Historically, DNA tumor viruses have beenessential tools in the analysis of cellular path-ways involving signal transduction, transcrip-tional regulation, and transformation. KSHVand related viruses modulate cell signalingpathways to make the environment in whichthey exist more conducive to virus survival.



Thus viral proteins interact with numerous cel-lular kinases and signaling molecules to mod-ulate the expression of proteins important forcell activation and viral progeny productionwhile counteracting proapoptotic stimuli thatcould lead to the premature lysis of the virus-infected cell.

KSHV encodes many homologs to cellularproteins that are involved in multiple path-ways such as signal transduction, immunemodulation, and cell cycle regulation (Table 1).Thus KSHV is well-equipped to survive in itshost. These different viral genes may aid thevirus to persist in different cell types, such as Blymphocytes and endothelial cells. The modu-lation of the aforementioned cellular pathwaysmay contribute to virus survival during thelatent and lytic phases of its life cycle. As aresult, the modulation of these cellular path-ways by KSHV may also be a contributing fac-tor to the onset of KSHV-associated diseasessuch as KS, MCD, and PEL.

It is curious that although encodes for awide number of transforming genes, KSHV-associated malignancy is only seen in the con-text of immune suppression. This wouldsuggest that KSHV infection is probably onlyone of many steps leading to the onset of can-cer, because tumorigenesis is thought to be amultistep process during which the contribu-tions of multiple biological events ultimately

lead to a neoplastic phenotype. Other cofactorssuch as HIV infection, iatrogenic immunosup-pression, and perhaps environmental condi-tions all greatly accelerate the onset of KS orPELs. Additionally, the precise roles that eachKSHV gene plays in this oncogenic process iscurrently being investigated; future studieswith recombinant viruses may allow for a moredetailed examination of the contributions ofeach viral gene to the neoplastic process.

ACKNOWLEDGMENTS

We thank D. Dittmer for critical reading ofthe manuscript. This work was supported byNIH/NCI grant RO1-CA096500 and grantsfrom the American Heart Association, UNC-CFAR, and V Foundation.

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Table 1Summary of KSHV Viral Proteins

Proteins ORF Cellular homolog Function

K1 K1 Unique SignalingK15 K15 Unique SignalingvGPCR 74 GPCR SignalingvIL-6 K2 IL-6 CytokinevMIPs K4, K6 MIP ChemokinevIRFs K9, K10.5, K11.1 IRF IFN responseLANA 73 Unique Episome maintenancevCyclin 72 CyclinD/E Cell cyclevFLIP 71 FLIP AntiapoptoticKaposin K12 Unique Signaling

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