VIROLOGY 227, 271–280 (1997) ARTICLE NO. VY968316

MINIREVIEW

View metadata, citation and similar papers at core.ac.uk brought to you by CORE DNA Tumor and Src Family Tyrosine Kinases, an Intimate Relationship provided by Elsevier - Publisher Connector

ANJA S. MESSERSCHMITT, NICOLAS DUNANT, and KURT BALLMER-HOFER1

Friedrich Miescher-Institute, P.O. Box 2543, CH-4002 Basel, Switzerland Received August 28, 1996; returned to author for revision October 7, 1996; accepted October 24, 1996

INTRODUCTION Weiss, 1995; Erpel and Courtneidge, 1995). Members of the Src family of tyrosine kinases are membrane-associ- Replication of viruses depends on a plethora of ated and act as transducers of signals emanating from highly specific interactions with the host resulting plasma membrane receptors, like, e.g., the PDGF recep- in the subversion of multiple cellular signal transduc- tor (Kypta et al., 1990) or the B and T cell receptors tion pathways. Cell cycle progression, intracellular (reviewed in Rudd et al., 1993). The activity of Src kinases protein transport, and protein and nucleotide metabo- is regulated by intramolecular interaction between a car- lism are among the many cellular processes deregu- boxy-terminal phosphorylated tyrosine residue and the lated by viral proteins in order to establish a cellular SH2 (Src homology 2) domain located in the amino-termi- environment favorable for replication. Viruses nal half of the molecule (Liu et al., 1993) (Fig. 1). When the also modulate a host’s immune response to escape SH2 domain is engaged in binding the carboxy-terminal immunosurveillance. Many of the pathways controlling phosphotyrosine, the kinase is inactive. The SH3 (Src cell behavior are tightly regulated by cellular tyrosine kinases. One particular family of these enzymes, Src homology 3) domain immediately adjacent to the SH2 family kinases, are involved in processing signals ema- domain is capable of binding proline-rich peptides and nating from the plasma membrane. Two families of seems to further stabilize the closed conformation of Src- DNA viruses, polyoma- and herpesviruses, encode related kinases perhaps upon binding to the catalytic or proteins targeted at tyrosine kinases. Middle-T anti- SH1 (Src homology 1) domain (Abrams and Zhao, 1995; gens expressed by mouse and polyomavirus Seidel-Dugan et al., 1992). Alternatively, the SH3, to- associate with and activate Src family tyrosine kinases, gether with the SH2 domain and the tail, might be in- while two members of the herpes family of DNA vi- volved in dimerization of Src kinases. This model was ruses, Epstein–Barr virus (EBV) and herpesvirus saim- suggested by a crystal structure of the covalently linked iri (HVS), encode proteins, LMP2A and Tip, respec- SH3 and SH2 domains of Lck complexed with the tail tively, that associate with cellular tyrosine kinases of peptide (Eck et al., 1994). Src-related tyrosine kinases are the Src and Syk/Zap family. All four viral proteins tightly activated by upstream signaling molecules that appear bind these kinases resulting in altered enzymatic activ- to weaken the intramolecular interaction between the ity and changes in downstream signaling. In this re- regulatory tyrosine residue and the SH2 domain. Dephos- view we focus on the mechanisms involved in the de- phorylation of this tyrosine residue by cellular phospha- regulation of tyrosine kinases by viral proteins and tases might stabilize the open, active conformation. Acti- discuss their possible function in virus replication and vation results in the phosphorylation of substrates down- spreading and in virus-induced tumorigenesis. stream in the signaling cascade, triggering interactions with a variety of cellular proteins. VIRAL PROTEINS ASSOCIATING WITH CELLULAR Polyoma- as well as some herpesviruses encode pro- TYROSINE KINASES teins capable of deregulating Src-related tyrosine ki- nases. Four viral proteins have been identified so far: Tyrosine kinases play a pivotal role in cell signaling mouse polyomavirus middle-T antigen (mouse middle-T) regulating cell growth, differentiation, and morphology associates with three Src-related kinases, c-Src (Bolen (reviewed in Taylor and Shalloway, 1996; Howe and et al., 1984; Courtneidge and Smith, 1983, 1984), c-Yes (Kornbluth et al., 1987), and Fyn (Cheng et al., 1988a; 1 To whom correspondence and reprint requests should be ad- Kypta et al., 1988; Horak et al., 1989), resulting in the dressed. Fax: 61 697 3976. E-mail: [email protected]. activation of the former two, while hamster polyomavirus

0042-6822/97 $25.00 271 Copyright ᭧ 1997 by Academic Press All rights of reproduction in any form reserved.

AID VY 8316 / 6a22$$$281 12-31-96 09:48:41 vira AP: Virology I Y81 6a22$$8316 / 8316 VY AID 23-60:84 iaA:Virology AP: vira 09:48:41 12-31-96 272

FIG. 1. Schematic representation of the various mechanisms employed by viral proteins associating with Src family kinases. All proteins seem to associate with Src kinases in the open conformation which usually represents the activated kinase. However, in the case of LMP2A, downregulation of the kinase has been observed, while the middle antigens and Tip activate Src kinases or maintain theses enzymes in an activated state. ‘‘Y’’ indicates tyrosine residues. Src KINASE TARGETING BY DNA TUMOR VIRUSES 273 middle-T (hamster middle-T) associates with and acti- dle-T associates with one of the SH2 domains of the vates only Fyn (Courtneidge et al., 1991). Lymphocytes regulatory 85-kDa subunit of phosphatidylinositol 3-ki- persistently infected with Epstein–Barr virus (EBV) ex- nase (PI 3-kinase) through phosphorylated tyrosine 315 press a , LMP2A, that associates with Src leading to the activation of the catalytic p110 subunit, family kinases, mainly Fyn and Lyn, and with the tyrosine which gives rise to the production of 3؅ phosphorylated kinase Syk (Miller et al., 1995; Burkhardt et al., 1992; phosphatidylinositides (Whitman et al., 1985). Phospholi- Longnecker et al., 1991). Finally, herpesvirus saimiri sub- pase C-g1 (PLC-g1) binds phosphorylated tyrosine 322 groups 484 and 488 contain open reading frames, ORF2 via an SH2 domain, presumably resulting in stimulation and ORF1, respectively, which give rise to highly related of enzyme activity and the generation of diacylglycerol proteins associating with Lck, a Src family kinase pre- and inositol triphosphate (Su et al., 1995). The adapter dominantly expressed in lymphocytes (Lund et al., 1996; protein SHC, which contains an SH2 domain, a PTB do- Biesinger et al., 1995). The HVS-C488-encoded polypep- main, and a collagen-homologous region and activates tide is also called Tip, for ‘‘tyrosine kinase interacting the Ras/MAP kinase pathway, associates with tyrosine protein’’ (Biesinger et al., 1995). 250 in the context NPTY via its PTB domain (Campbell Interestingly, neither adeno- nor papillomaviruses, et al., 1994; Dilworth et al., 1994). Hamster middle-T con- both distantly related to polyomaviruses, seem to ex- tains two residues, tyrosines 298 and 330, that are puta- press proteins capable of directly associating with and tive binding sites for PI 3-kinase and PLC-g1, respec- thereby altering the function of Src-related kinases. How- tively (Courtneidge et al., 1991; Brizuela et al., 1995), yet ever, both viruses express proteins, adeno E3 and papil- lacks a SHC binding site (Table 1). loma E5, respectively, targeted at tyrosine kinase growth Both middle-T’s contain several proline-rich se- factor receptors, like the EGF and PDGF receptors quences that might bind signaling molecules carrying (Straight et al., 1993; Kuivinen et al., 1993; Goldstein et SH3 domains, but experimental evidence in support of al., 1992; Carlin et al., 1989; Hoffman et al., 1992). These this idea is still missing. Mouse middle-T also associates receptors have been shown to associate with and acti- with members of the 14-3-3 family of proteins, which vate Src-related kinases like c-Src and Fyn upon stimula- are suggested to act as scaffolding proteins during the tion by their ligands (reviewed in Courtneidge et al., formation of protein complexes (Pallas et al., 1994). Fi- 1993). E3 and E5 might therefore indirectly influence the nally, phosphatase 2A (PP2A) has been shown to bind activity of Src-related kinases complexed with tyrosine the amino-terminal domain of mouse and hamster mid- kinase growth factor receptors. dle-T which is shared with small-T, resulting in altered in vitro substrate specificity of this enzyme (Cayla et al., THE MIDDLE-T ANTIGENS OF POLYOMAVIRUSES 1993; Ruediger et al., 1994; Yang et al., 1991; Scheidt- mann et al., 1991; Campbell et al., 1995; Pallas et al., Upon alternative splicing of the primary early tran- 1990) (Table 1). Association of PP2A with mouse middle- script, polyomaviruses produce three mRNA’s encoding T and small-T has been studied in detail and the epitopes the tumor antigens, large-, middle-, and small-T (Tooze, involved are well defined (Campbell et al., 1995; Mungre 1980). Mouse middle-T and hamster middle-T are 55% et al., 1994; Ruediger et al., 1992). identical within the amino-terminal domain which is shared by large- and small-T antigens. Except for the fact MECHANISM OF ASSOCIATION OF MOUSE AND that both proteins have a carboxy-terminal membrane HAMSTER MIDDLE-T WITH Src-RELATED KINASES anchor, very little homology is detected in the middle-T- specific unique domains. Expression of mouse middle-T The mechanisms responsible for association of mouse results in a 5- to 10-fold increase in overall c-Src (Bolen and hamster middle-T with Src-related tyrosine kinases et al., 1984; Courtneidge and Smith, 1984) or c-Yes (Korn- are distinct. Earlier work gave conflicting results sug- bluth et al., 1987) kinase activity, while Fyn is not acti- gesting either the carboxy-terminal (Cheng et al., 1988b) vated (Horak et al., 1989). However, no concomitant in- or amino-terminal domains of c-Src to be responsible for crease in overall tyrosine phosphorylation of cellular pro- association with mouse middle-T (Louie et al., 1988). We teins is observed in middle-T-transformed cultured cells. have shown recently that mouse middle-T associates Hamster middle-T, on the other hand, exclusively associ- with Src kinase mutants lacking all domains amino-termi- ates with Fyn which is activated about 2-fold (Court- nal to the catalytic (SH1) domain (Dunant et al., 1996). neidge et al., 1991). When c-Src was truncated after residue 518, mouse mid- Both middle-T’s contain a series of tyrosine residues dle-T was not phosphorylated anymore in vitro (Cheng in their unique domains that are phosphorylated upon et al., 1988b), yet defective complexes could be detected association with Src-related tyrosine kinases and are in metabolically labeled cells (Dunant et al., unpub- required for binding cellular signaling molecules via SH2 lished). This suggests that proper positioning of mouse (reviewed in Pawson, 1994) or PTB (reviewed in van der middle-T relative to the catalytic domain of c-Src is dic- Geer and Pawson, 1995) domains (Table 1). Mouse mid- tated by the c-Src tail.

AID VY 8316 / 6a22$$$281 12-31-96 09:48:41 vira AP: Virology 274 MESSERSCHMITT, DUNANT, AND BALLMER-HOFER

TABLE 1 Synopsis of Viral Proteins Associating with Src-Related and Syk/Zap Family Kinases

Additional signaling Viral protein Kinases bound Binding mechanism for tyrosine kinases proteins bound

Mouse middle-T c-Src, c-Yes, Fyn Association with SH1 domain PP2A, PI 3-K, SHC, PLC-g1, 14-3-3 Hamster middle-T Fyn Association with SH2 domain via pYEEI motif PP2A, PI 3-K, PLC-g1

LMP2A Syk, Zap Association with SH2 domain via pYXXLN7pYXXL motif PI 3-K, PLC-g2, Vav Lyn Association with SH2 domain via pYEEA motif Tip Lck Association with SH3 domain via proline-rich SH3B domain; CSKH domain also required

Note. In the case of LMP2A, association with tyrosine kinases is well established, while binding to signaling molecules is speculative and has not been shown experimentally so far.

The interaction between c-Src and mouse middle-T is bind the SH2 domains of Src-related kinases, as deter- not stoichiometric. Ten to 15% of mouse middle-T are mined in a phosphopeptide library screen (Songyang et engaged in binding wt c-Src and only 10% of c-Src are al., 1993). Based on the finding that a Fyn mutant lacking bound to mouse middle-T (Bolen et al., 1987; Cheng et the SH2 domain did not bind hamster middle-T, we mu- al., 1990). However, stoichiometric complexes are formed tated tyrosine 324 to phenylalanine. The mutant protein with activated Src kinase mutants lacking either the SH2 showed threefold reduced association with Fyn (Dunant or the SH3 domain (Dunant et al., 1996). These mutants et al., in press). Residual association with Fyn is most are believed to assume an open conformation resulting likely due to phosphotyrosine-independent binding. in increased association with mouse middle-T. The Moreover, the same study showed that this mutation mechanism by which mouse middle-T activates Src ki- abolished the oncogenicity of hamster middle-T. There- nases might therefore be to stabilize activated molecules fore, phosphorylation of hamster middle-T at tyrosine present in the open conformation (Fig. 1). We hypothesize 324, most likely by Fyn itself, is essential to form a func- that intermolecular interaction between mouse middle-T tional complex with Fyn. This is in contrast to mouse and c-Src or c-Yes prevents an intramolecular interaction middle-T, where none of the tyrosine residues is required between the regulatory phosphotyrosine and the SH2 for c-Src binding. It has also been shown that even cata- domain in these enzymes, resulting in constitutive activa- lytically inactive c-Src associates with mouse middle-T tion of mouse middle-T-associated c-Src and c-Yes. This (Cheng et al., 1990). is in agreement with the earlier finding that c-Src associ- The question remains why mouse middle-T binds c- ated with mouse middle-T is dephosphorylated at tyro- Src, c-Yes, and Fyn, while hamster middle-T associates sine 527 (Cartwright et al., 1986; Courtneidge, 1985). only with Fyn. The most likely concept is that mouse It has been more difficult to define epitope(s) present middle-T associates with epitopes well conserved in the in mouse middle-T that are required for c-Src binding. A SH1 domain of these kinases, but not in other Src family comprehensive mutational analysis suggests that even members like Lck or Lyn. In the case of hamster middle- minor changes in the amino-terminal domain of mouse T, it is likely that other regions, besides the YEEI motif, middle-T, which is also responsible for PP2A binding, are responsible for determining specificity of the interac- block association with c-Src (Cheng et al., 1989; Mark- tion with Fyn since this motif has been shown to tightly land and Smith, 1987; Campbell et al., 1995). Recent evi- bind the SH2 domain of several Src family members dence suggests that PP2A association with mouse mid- (Songyang et al., 1993). dle-T is essential for c-Src binding and indicates that structural determinants present in PP2A might take part LMP2 AND Tip OF HERPESVIRUSES in the formation of a ternary complex (Glenn and Eckhart, 1995). An alternative interpretation of these data is that Two classes of herpesviruses, a human virus, EBV, the phosphorylation state of mouse middle-T is tightly and HVS endemic in a new world monkey, have been regulated by PP2A and determines its ability to associate shown to target Src kinases by LMP2 and Tip, respec- with c-Src. An epitope required to bind c-Src, but not tively. Both viruses are tumorigenic. EBV is associated PP2A, has been located between amino acids 203 and with Burkitt’s lymphoma while in the case of HVS, tumors 240 of mouse middle-T using T antigen-specific mono- arise when the virus infects different highly related mon- clonal antibodies (Dilworth and Horner, 1993). key species. In contrast to mouse middle-T, hamster middle-T car- LMP2 exists in two splice variants, LMP2A and LMP2B, ries a stretch of amino acids, YEEI (tyrosine in position which are both membrane-associated and differ in their 324), corresponding to the optimal sequence known to amino-terminal domains. LMP2A associates with Src-re-

AID VY 8316 / 6a22$$$281 12-31-96 09:48:41 vira AP: Virology Src KINASE TARGETING BY DNA TUMOR VIRUSES 275 lated kinases (Longnecker et al., 1991; Burkhardt et al., and (Scherneck and Feunteun, 1990; Tooze, 1992) and a 72-kDa tyrosine kinase which is probably 1980). Polyomavirus large-T antigen is essential for viral Syk (Miller et al., 1994a, 1995). These interactions might DNA replication and capable of immortalizing cells (re- be mediated by tyrosine peptide sequences present in viewed in Pipas, 1992). The middle-T antigens of mouse the amino-terminal domain of LMP2A that have also been and hamster polyomaviruses show low sequence homol- found in cellular signaling molecules. A motif pYEEA ogy in their unique domains, although these viruses are known to bind Src family SH2 domains in vitro (Songyang highly homologous in structure, clearly setting et al., 1993) might recruit Fyn or Lyn to LMP2A. Syk/Zap them apart from the related monkey and human viruses tyrosine kinases might associate with LMP2A through that all lack the middle-T reading frame (Delmas et al., a sequence pYXXLN7pYXXL that is known to mediate 1985; Tooze, 1980). The unique domain of middle-T is association of these kinases with the phosphorylated T encoded in an area of the genome that is poorly con- cell receptor z chain (Weiss, 1993; Beaufils et al., 1993). served among the SV40 (SV40, BK, JC) and the polyoma The fact that LMP2A can block B lymphocyte signaling group of papovaviruses (Tooze, 1980). In SV40-like vi- through the surface immunoglobulin (sIg) receptor (Miller ruses, this region encodes the large-T epitope capable et al., 1994a,b) is probably the consequence of seques- of binding p53, the ‘‘guardian of the genome’’ (reviewed tering both Lyn and Syk into complexes with LMP2A in Lane and Benchimol, 1990; Pipas, 1992). This results that act as dominant negative constituents in the B cell in inactivation of the cell cycle surveillance function of receptor signaling cascade (Miller et al., 1995). The p53 and might prevent virus-induced apoptosis. Mouse pYXXLN7pYXXL peptide has also been shown to be func- and hamster polyomaviruses express large-T antigens tional in a chimeric protein consisting of the extracellular that do not bind p53, and one might therefore speculate and transmembrane domain of the CD8 receptor and the that polyomaviruses inactivate or subvert the function of cytoplasmic domain of the bovine leukemia virus enve- p53 or some of its downstream targets through middle- lope protein gp30 (Beaufils et al., 1993). The chimeric T antigen. In this case, though, a completely different protein elicited early and late events required for lympho- mechanism than employed by SV40 large-T must be envi- cyte activation and has been shown to give rise to B cell sioned. It will be interesting to study whether middle- proliferation. To initiate association with tyrosine ki- T-induced pathways, in particular the Ras/MAP kinase nases, LMP2A must be phosphorylated at the specific cascade, and the functions altered in SV40-transformed tyrosine residues mentioned above. It is not yet clear cells as a consequence of sequestering p53 into large- how this is brought about, but phosphorylation by Src T complexes converge on any of the known cell cycle kinases, e.g., Lck, upon stimulation through ancillary sub- regulatory proteins, like, e.g., cyclin-dependent kinase units of the B cell receptor complex seems to be a likely inhibitors, cyclins, or cdk’s (reviewed in Eick and Hermek- explanation. ing, 1994; Cox and Lane, 1995). Tip of HVS strain C488 is composed of a series of The middle-T antigens of polyomaviruses have been peptide motifs that are all required for binding to Lck: a shown to be required for virus-induced oncogenesis and 38-amino-acid peptide called the CSKH domain which cell transformation in vitro. Transgenic animals express- is related to a sequence present close to the carboxy- ing mouse middle-T show endotheliomas, while hamster terminus of the kinase domain of all Src-related kinases, middle-T causes mainly skin epitheliomas and a proline-rich motif called SH3B, and a spacer linking lymphoma (reviewed in Kiefer et al., 1994b). This is a these domains (Jung et al., 1995a) (Fig. 2). The SH3 do- consequence of the activation of a variety of cellular main of Lck is sufficient for Tip binding (Jung et al., 1995b) signaling proteins regulating cell growth. Phosphoryla- and none of the other Src-related kinases seems to asso- tion of middle-T by Src-related kinases at specific tyro- ciate with Tip (Wiese et al., 1996). A hydrophobic se- sine residues is essential to assemble these cellular quence at the carboxy-terminus of Tip probably localizes proteins into an active signaling complex (reviewed in this protein to cellular membranes but seems not to be Dilworth, 1995). The ability of mouse middle-T to induce required for interaction with Lck. Membrane-bound Tip tumors has also been analyzed in knockout mice might interfere with lymphocyte-specific early signaling lacking c-Src, c-Yes, or Fyn (Thomas et al., 1993; Kiefer events localized at the plasma membrane. et al., 1994b). These studies showed that expression of only two of the kinases known to associate with mouse WHAT IS THE ROLE OF MIDDLE-T IN THE middle-T is sufficient for endothelioma formation. In con- POLYOMAVIRUS LIFE CYCLE AND IN trast to c-src and fyn knockout animals, c-yes-deficient VIRUS-MEDIATED ONCOGENICITY? mice showed a reduced number of tumors and a longer latency period (Kiefer et al., 1994a), suggesting that the Polyomaviruses were originally discovered as agents contribution of c-Yes in tumor formation is only partially causing tumors in mice and . The mouse virus compensated by other Src family kinases. Hamster mid- usually induces tumors derived from mesenchymal cells, dle-T was still able to transform endothelial cells from while the hamster virus causes epitheliomas, lymphoma, fyn knockout mice (Kiefer et al., 1994b). This suggests

AID VY 8316 / 6a22$$$281 12-31-96 09:48:41 vira AP: Virology 276 MESSERSCHMITT, DUNANT, AND BALLMER-HOFER

FIG. 2. Domain structure of viral proteins associating with Src family kinases. Epitope assigned to PP2A binding site of the middle-T’s is tentative and derived from mutants made in SV40 small-T and mouse middle-T (Campbell et al., 1995; Mungre et al., 1994). Dark gray boxes indicate transmembrane domains. that another, not yet identified tyrosine kinase might com- portant to note that peptides derived from polyomavirus pensate for the lack of Fyn or that kinase activity in the T antigens act as tumor-specific transplantation antigens hamster middle-T complex is dispensable for transforma- (TSTA) and play a role in eliciting an immune response tion of this cell type. to polyomavirus-induced tumor cells (Reinholdsson Ljun- Studies performed with mouse polyomavirus in tissue ggren et al., 1992; reviewed in Dalianis, 1990). culture cells showed that mouse middle-T is also re- A role for mouse middle-T in activating the Ras/MAP quired for virus replication (Garcea et al., 1989). Mutants kinase signaling cascade has been proposed in virus- of mouse middle-T that are only capable of binding PP2A mediated S phase induction (Urich et al., 1995; Srinivas were sufficient to support virus growth in . et al., 1994). Immediate early induced through the In vivo, however, these mutants were defective in virus MAP kinase pathway are also expressed upon binding replication and unable to spread to secondary sites of of virus to cell surface receptors (Zullo et al., replication (Freund et al., 1992). This study also showed 1987). Virus mutants expressing a mouse middle-T un- that tumor formation in animals requires an intact mouse able to bind Src kinases and to activate the MAP kinase middle-T and correlates with the ability of the virus to cascade in the absence of growth factors grow like wild maintain a persistent infection. In this context it is im- type in NIH 3T3 cells in serum-containing medium (Gar-

AID VY 8316 / 6a22$$$281 12-31-96 09:48:41 vira AP: Virology Src KINASE TARGETING BY DNA TUMOR VIRUSES 277 cea et al., 1989). These middle-T mutants that still associ- the kinase from phosphorylating cellular targets. Binding ate with PP2A might cooperate with small-T in preventing of Tip to the SH3 domain might also block the interaction downregulation of growth factor-induced MAP kinase ac- of Lck with specific cellular targets. One of the biological tivity as reported for SV40 small-T (Frost et al., 1994; consequences of Tip-induced Lck inactivation in lympho- Sontag et al., 1993). The mitogenic function of mouse cytes is downregulation of surface antigens like CD2 and middle-T might therefore be required when a virus repli- CD4 (Jung et al., 1995b). This is reminiscent of the effects cates in cells grown in low serum or in low multiplicity observed in Lck knockout mice (Molina et al., 1992), sug- infections. gesting that active signaling through Lck is required for The function of mouse middle-T during virus replica- expression of T cell-specific surface antigens. A recent tion is, at least in part, to increase the efficiency of virus study shows that Tip can activate Lck in some T cells encapsidation late in the replicative cycle (Garcea et al., as well as in in vitro phosphorylation assays (Wiese et 1985, 1989; Garcea and Benjamin, 1983). To initiate en- al., 1996). This might be the consequence of differences capsidation, the major coat protein, VP1, is phosphory- in the experimental system used by these authors. lated at canonical MAP kinase phosphorylation sites, A different virus strain, HVS 484-77, expresses a pro- suggesting that MAP kinase activated by mouse middle- tein highly related to Tip. Expression of this protein has T might phosphorylate VP1 in virus-infected cells (Li and been associated with IL-2-independent growth of virus- Garcea, 1994). infected T lymphocytes (Lund et al., 1996). Since Lck has been shown to mediate signaling downstream from the WHAT IS THE ROLE OF LMP2A AND Tip IN IL-2 receptor (Hatakeyama et al., 1991), it is tempting to HERPESVIRUS PROPAGATION AND speculate that Tip of HVS 484-77 functions in activating ONCOGENICITY? this pathway in a ligand-independent way. This might confer HVS-infected and immortalized cells with a factor- In vivo LMP2A is expressed in latently infected B lym- independent growth phenotype. phocytes and nasopharyngeal carcinoma cells (Qu and Rowe, 1992; Busson et al., 1992; Brooks et al., 1992). This EVOLUTION OF VIRAL PROTEINS TARGETING led to the suggestion that LMP2A plays a crucial role in CELLULAR TYROSINE KINASES establishing virus persistence in EBV-infected patients. Considering the various strategies that polyoma- and Lytic reactivation of EBV has also been shown to be herpesviruses use to target cellular tyrosine kinases blocked in LMP2A-expressing lymphocytes in cell culture raises the question of how the middle-T antigens, (Miller et al., 1994a,b). As described above, LMP2A mod- LMP2A, and Tip evolved. A direct relationship with ances- ulates cell signaling downstream of the sIg cell surface tral cellular genes has not been found so far, yet many receptor in persistently infected B cells blocking the ex- of the motifs carried by these proteins are also present pression of both cellular and viral genes that are essen- in cellular proteins involved in cell signaling. Herpesvirus tial for virus replication. LMP2A might thereby allow af- reading frames encoding LMP2A and Tip show no appar- flicted B lymphocytes to escape from immunosurveil- ent sequence homology, although they are located at lance. Epstein–Barr virus also transforms cells in vitro similar positions in their respective viral (Al- and causes malignant growth in rare cases. Both LMP’s brecht et al., 1992). Similarly, the reading frames coding and a set of nuclear proteins, EBNA’s, might be involved for the middle-T antigens of mouse and hamster poly- in this process (reviewed in Kieff, 1996; Klein, 1994). How- omavirus map to the same domain in the respective viral ever, contrary to the polyomavirus middle-T antigens that genomes yet are poorly conserved in the second, middle- are highly oncogenic, LMP2A does not transform cells T-specific exon. Therefore, targeting of tyrosine kinases in culture as demonstrated using EBV mutants lacking a by LMP2A, Tip, and the middle-T antigens is most likely functional LMP2A reading frame (Longnecker et al., 1992, the product of independent evolutionary processes. Se- 1993a,b). quences encoding viral protein motifs capable of associ- A similar function might be postulated for Tip in her- ating with cellular tyrosine kinases might have been ac- pesvirus saimiri-infected cells. Tip seems to cooperate quired by recombination with host genes during virus with STP during virus-mediated transformation and tumor replication. Alternatively, the evolution of these motifs formation in HVS-infected cells (Bro¨ker et al., 1993; Jung might be the product of random mutations propagated et al., 1991; Biesinger et al., 1992). Tip has been shown as a consequence of increased viral fitness. to attenuate the activity of Lck in fibroblasts transformed with an activated Lck mutant and to reduce phosphoryla- CONCLUSIONS tion of substrates like calpactin I (Jung et al., 1995b). Tip also blocks signaling through the T cell receptor and Both polyoma- and herpesviruses express proteins ca- prevents in vitro phosphorylation of substrates upon CD3 pable of targeting tyrosine kinases that regulate the cellu- stimulation (Jung et al., 1995b). In HVS-infected cells, Tip lar signaling network. Similar to LMP2A in EBV-infected might be the primary substrate of Lck, thereby preventing cells, Tip and middle-T may cooperate with the immor-

AID VY 8316 / 6a22$$$281 12-31-96 09:48:41 vira AP: Virology 278 MESSERSCHMITT, DUNANT, AND BALLMER-HOFER talizing genes of HVS and polyomaviruses, STP, and cells: Coexpression of EBNA1, LMP1, and LMP2 transcripts. J. Virol. large-T, respectively, to maintain cell viability and allow 66, 2689–2697. virus-infected cells to escape immunosurveillance. This Burkhardt, A. L., Bolen, J. B., Kieff, E., and Longnecker, R. (1992). An Epstein–Barr virus transformation-associated membrane protein in- might turn out to be essential in establishing virus persis- teracts with src family tyrosine kinases. J. Virol. 66, 5161–5167. tence and ensure efficient spreading within a host spe- Busson, P., McCoy, R., Sadler, R., Gilligan, K., Tursz, T., and Raab-Traub, cies rather than increasing short-term efficiency of virus N. (1992). Consistent transcription of the Epstein–Barr virus LMP2 replication. Oncogenicity of DNA viruses probably gene in nasopharyngeal carcinoma. J. Virol. 66, 3257–3262. evolved fortuitously as a consequence of altered cell Campbell, K. S., Ogris, E., Burke, B., Su, W., Auger, K. R., Druker, B. J., Schaffhausen, B. S., Roberts, T. M., and Pallas, D. C. (1994). Polyoma signaling to support the viral replicative cycle since there interacts with SHC protein via the NPTY (Asn- is hardly a benefit derived from a virus’ ability to induce Pro-Thr-Tyr) motif in middle tumor antigen. Proc. Natl. Acad. Sci. USA tumors, as no live virus is usually released from tumor 91, 6344–6348. cells. Campbell, K. S., Auger, K. R., Hemmings, B. A., Roberts, T. M., and Pal- las, D. C. (1995). Identification of regions in polyomavirus middle T and small t antigens important for association with protein phospha- ACKNOWLEDGMENTS tase 2A. J. Virol. 69, 3721–3728. Carlin, C. R., Tollefson, A. E., Brady, H. A., Hoffman, B. L., and Wold, We thank Dr. R. Chiquet-Ehrismann and Dr. S. Kaech, FMI Basel, W. S. M. (1989). Epidermal growth factor receptor is down-regulated CH, for critically reading the manuscript and Dr. B. Biesinger, Erlangen, by a 10,400 MW protein encoded by the E3 region of adenovirus. FDR, and Dr. J.-P. Jost, FMI Basel, for helpful comments during the Cell 57, 135–144. preparation of this review. We also acknowledge the excellent secre- Cartwright, C. A., Kaplan, P. L., Cooper, J. A., Hunter, T., and Eckhart, tarial work done by E. Laschinger, FMI Basel, CH. N. Dunant was W. (1986). Altered sites of tyrosine phosphorylation in pp60c-src asso- supported by ‘‘Stipendienfonds der Basler Chemischen Industrie’’ and ciated with polyomavirus middle tumor antigen. Mol. Cell. Biol. 6, A. Messerschmitt by ‘‘Krebsliga beider Basel.’’ 1562–1570. Cayla, X., Ballmer-Hofer, K., Merlevede, W., and Goris, J. (1993). Phos- REFERENCES phatase 2A associated with polyomavirus small-T or middle-T anti- gen is an okadaic acid-sensitive tyrosyl phosphatase. Eur. J. Bio- Abrams, C. S., and Zhao, W. (1995). SH3 domains specifically regulate chem. 214, 281–286. kinase activity of expressed Src family proteins. J. Biol. Chem. 270, Cheng, S. H., Harvey, R., Espino, P. C., Semba, K., Yamamoto, T., Toyo- 333–339. shima, K., and Smith, A. E. (1988a). Peptide antibodies to the human Albrecht, J.-C., Nicholas, J., Biller, D., Cameron, K. R., Biesinger, B., c-fyn gene product demonstrate pp59c-fyn is capable of complex for- Newman, C., Wittmann, S., Craxton, M. A., Coleman, H., Fleckenstein, mation with the middle-T antigen of polyomavirus. EMBO J. 7, 3845– B., and Honess, R. W. (1992). Primary structure of the Herpesvirus 3855. saimiri genome. J. Virol. 66, 5047–5058. Cheng, S. H., Piwnica-Worms, H., Harvey, R. W., Roberts, T. M., and Beaufils, P., Choquet, D., Mamoun, R. Z., and Malissen, B. (1993). The Smith, A. E. (1988b). The carboxy terminus of pp60c-src is a regulatory (YXXL/I)2 signalling motif found in the cytoplasmic segments of the domain and is involved in complex formation with the middle-T anti- bovine leukaemia virus envelope protein and Epstein–Barr virus la- gen of polyomavirus. Mol. Cell. Biol. 8, 1736–1747. tent membrane protein 2A can elicit early and late lymphocyte activa- Cheng, S. H., Harvey, R., Piwnica-Worms, H., Espino, P. C., Roberts, tion events. EMBO J. 12, 5105–5112. T. M., and Smith, A. E. (1989). Mechanism of activation of complexed Biesinger, B., Mu¨ller-Fleckenstein, I., Simmer, B., Lang, G., Wittmann, pp60c-src the middle T antigen of polyomavirus. Curr. Top. Microbiol. S., Platzer, E., Desrosiers, R. C., and Fleckenstein, B. (1992). Stable Immunol. 144, 109–120. growth transformation of human T lymphocytes by Herpesvirus saim- Cheng, S. H., Espino, P. C., Marshall, J., Harvey, R., and Smith, A. E. iri. Proc. Natl. Acad. Sci. USA 89, 3116–3119. (1990). Stoichiometry of cellular and viral components in the poly- Biesinger, B., Tsygankov, A. Y., Fickenscher, H., Emmrich, F., Fleck- omavirus middle-T antigen–tyrosine kinase complex. Mol. Cell. Biol. enstein, B., Bolen, J. B., and Broker, B. M. (1995). The product of the 10, 5569–5574. Herpesvirus saimiri open reading frame 1 (tip) interacts with T cell- Courtneidge, S. A. (1985). Activation of the pp60c-src kinase by middle specific kinase p56lck in transformed cells. J. Biol. Chem. 270, 4729– T antigen binding or by dephosphorylation. EMBO J. 4, 1471–1477. 4734. Courtneidge, S. A., Goutebroze, L., Cartwright, A., Heber, A., Scherneck, Bolen, J. B., Thiele, C. J., Israel, M. A., Yonemoto, W., Lipsich, L. A., and S., and Feunteun, J. (1991). Identification and characterization of the Brugge, J. S. (1984). Enhancement of cellular src gene product asso- hamster polyomavirus middle T antigen. J. Virol. 65, 3301–3308. ciated tyrosyl kinase activity following polyoma virus infection and transformation. Cell 38, 767–777. Courtneidge, S. A., Fumagalli, S., Koegl, M., Superti Furga, G., and Bolen, J. B., Deseau, V., O’Shaughnessy, J., and Amini, S. (1987). Analy- Twamley, S. G. M. (1993). The Src family of protein tyrosine kinases: sis of middle tumor antigen and pp60c-src interactions in polyomavi- Regulation and functions. Development, 57–64. rus-transformed rat cells. J. Virol. 61, 3299–3305. Courtneidge, S. A., and Smith, A. E. (1983). Polyoma virus transforming Brizuela, L., Ulug, E. T., Jones, M. A., and Courtneidge, S. A. (1995). protein associates with the product of the c-src cellular gene. Nature Induction of interleukin-2 transcription by the hamster polyomavirus 303, 435–439. middle T antigen: A role for Fyn in T cell signal transduction. Eur. J. Courtneidge, S. A., and Smith, A. E. (1984). The complex of polyoma c-src Immunol. 25, 385–393. virus middle-T antigen and pp60 . EMBO J. 3, 585–591. Bro¨ker, B. M., Tsygankov, A. Y., Muller Fleckenstein, I., Guse, A. H., Cox, L. S., and Lane, D. P. (1995). Tumour suppressors, kinases and Chitaev, N. A., Biesinger, B., Fleckenstein, B., and Emmrich, F. (1993). clamps: How p53 regulates the cell cycle in response to DNA dam- Immortalization of human T cell clones by Herpesvirus saimiri. Signal age. BioEssays 17, 501–508. transduction analysis reveals functional CD3, CD4, and IL-2 recep- Dalianis, T. (1990). Studies on the polyoma virus tumor-specific trans- tors. J. Immunol. 151, 1184–1192. plantation antigen (TSTA). Adv. Cancer Res. 55, 57–85. Brooks, L., Yao, Q. Y., Rickinson, A. B., and Young, P. G. (1992). Epstein– Delmas, V., Bastien, C., Scherneck, S., and Feunteun, J. (1985). A new Barr virus latent gene transcription in nasopharyngeal carcinoma member of the polyomavirus family: The hamster papovavirus com-

AID VY 8316 / 6a22$$$281 12-31-96 09:48:41 vira AP: Virology Src KINASE TARGETING BY DNA TUMOR VIRUSES 279

plete nucleotide sequence and transformation properties. EMBO J. siers, R. C. (1995b). Downregulation of Lck-mediated signal transduc- 4, 1279–1286. tion by tip of herpesvirus saimiri. J. Virol. 69, 7814–7822. Dilworth, S. M., Brewster, C. E., Jones, M. D., Lanfrancone, L., Pelicci, Kiefer, F., Anhauser, I., Soriano, P., Aguzzi, A., Courtneidge, S. A., and G., and Pelicci, P. G. (1994). Transformation by polyoma virus middle Wagner, E. F. (1994a). Endothelial cell transformation by polyomavi- T-antigen involves the binding and tyrosine phosphorylation of Shc. rus middle T antigen in mice lacking Src-related kinases. Curr. Biol. Nature 367, 87–90. 4, 100–109. Dilworth, S. M. (1995). Polyoma virus middle T antigen: Meddler or Kiefer, F., Courtneidge, S. A., and Wagner, E. F. (1994b). Oncogenic mimic? Trends Microbiol. 3, 31–35. properties of the middle T antigens of polyomaviruses. Adv. Cancer Dilworth, S. M., and Horner, V. P. (1993). Novel monoclonal antibodies Res. 64, 125–157. that differentiate between the binding of pp60c-src or protein phospha- Kieff, E. (1996). In ‘‘Virology’’ (B. N. Fields, D. M. Knipe, and P. M. Howley, tase 2A by polyomavirus middle T antigen. J. Virol. 67, 2235–2244. Eds.), 3rd ed., pp. 2343–2446. Dunant, N. M., Senften, M., and Ballmer-Hofer, K. (1996). Polyomavirus Klein, G. (1994). Epstein–Barr virus strategy in normal and neoplastic middle-T antigen associates with the kinase domain of Src-related B cells. Cell 77, 791–793. tyrosine kinases. J. Virol. 70, 1323–1330. Kornbluth, S., Sudol, M., and Hanafusa, H. (1987). Association of the Eck, M. J., Atwell, S. K., Shoelson, S. E., and Harrison, S. C. (1994). Struc- polyomavirus middle-T antigen with c-yes protein. Nature 325, 171– ture of the regulatory domains of the Src-family tyrosine kinase Lck. 173. Nature 368, 764–769. Kuivinen, E., Hoffman, B. L., Hoffman, P. A., and Carlin, C. R. (1993). Eick, D., and Hermeking, H. (1994). Viruses as pacemakers in the evolu- Structurally related Class I and Class II receptor protein tyrosine tion of defence mechanisms against cancer. Curr. Biol. 4, 100–109. kinases are down-regulated by the same E3 protein coded for by Erpel, T., and Courtneidge, S. A. (1995). Src family protein tyrosine ki- human Group C adenoviruses. J. Cell Biol. 120, 1271–1279. nases and cellular signal transduction pathways. Curr. Opin. Cell Kypta, R. M., Hemming, A., and Courtneidge, S. A. (1988). Identification Biol. 7, 176–182. and characterization of p59fyn (a src-like protein tyrosine kinase) in Freund, R., Sotnikov, A., Bronson, R. T., and Benjamin, T. L. (1992). Poly- normal and polyoma virus transformed cells. EMBO J. 7, 3837–3844. oma virus middle T is essential for virus replication and persistence Kypta, R. M., Goldberg, Y., Ulug, E. T., and Courtneidge, S. A. (1990). as well as for tumor induction in mice. Virology 191, 716–723. Association between the PDGF receptor and members of the src Frost, J. A., Alberts, A. S., Sontag, E., Guan, K., Mumby, M. C., and Fera- family of tyrosine kinases. Cell 62, 481–492. misco, J. R. (1994). Simian virus 40 small t antigen cooperates with Lane, D. P., and Benchimol, S. (1990). p53: Oncogene or anti-oncogene? mitogen-activated kinases to stimulate AP-1 activity. Mol. Cell. Biol. Genes Dev. 4, 1–8. 14, 6244–6252. Li, M., and Garcea, R. L. (1994). Identification of the threonine phosphor- Garcea, R. L., Ballmer-Hofer, K., and Benjamin, T. L. (1985). Virion as- ylation sites on the polyomavirus major protein VP1: Relation- sembly defect of polyomavirus hr-t mutants: Underphosphorylation ship to the activity of middle T antigen. J. Virol. 68, 320–327. of major capsid protein VP1 before viral DNA encapsidation. J. Virol. Liu, X., Brodeur, S. R., Gish, G., Zhou, S., Cantley, L. C., Laudano, A. P., 54, 311–316. and Pawson, T. (1993). Regulation of c-Src tyrosine kinase activity Garcea, R. L., Talmage, D. A., Harmatz, A., Freund, R., and Benjamin, by the Src SH2 domain. Oncogene 8, 1119–1126. T. L. (1989). Separation of host range from transformation functions Longnecker, R., Druker, B., Roberts, T. M., and Kieff, E. (1991). An Ep- of the hr-t gene of polyomavirus. Virology 168, 312–319. stein–Barr virus protein associated with cell growth transformation Garcea, R. L., and Benjamin, T. L. (1983). Host range transforming gene interacts with a tyrosine kinase. J. Virol. 65, 3681–3692. of polyoma virus plays a role in virus assembly. Proc. Natl. Acad. Longnecker, R., Miller, C. L., Miao, X. Q., Marchini, A., and Kieff, E. Sci. USA 80, 3613–3617. (1992). The only domain which distinguishes Epstein–Barr virus la- Glenn, G. M., and Eckhart, W. (1995). Amino-terminal regions of poly- tent membrane protein 2A (LMP2A) from LMP2B is dispensable for omavirus middle T antigen are required for interactions with protein lymphocyte infection and growth transformation in vitro; LMP2A is phosphatase 2A. J. Virol. 69, 3729–3736. therefore nonessential. J. Virol. 66, 6461–6469. Goldstein, D. J., Andresson, T., Sparkowski, J. J., and Schlegel, R. (1992). Longnecker, R., Miller, C. L., Miao, X., Tomkinson, B., Kieff, E., and Miao, The BPV-1 E5 protein, the 16 kDa membrane pore-forming protein X. Q. (1993a). The last seven transmembrane and carboxy-terminal and the PDGF receptor exist in a complex that is dependent on cytoplasmic domains of Epstein–Barr virus latent membrane protein hydrophobic transmembrane interactions. EMBO J. 11, 4851–4859. 2 (LMP2) are dispensable for lymphocyte infection and growth trans- Hatakeyama, M., Kono, T., Kobayashi, N., Kawahara, A., Levin, S. D., formation in vitro. J. Virol. 67, 2006–2013. Perlmutter, R. M., and Taniguchi, T. (1991). Interaction of the IL-2 Longnecker, R., Miller, C. L., Tomkinson, B., Miao, X., Kieff, E., and Miao, receptor with the src-family kinase p56lck: Identification of novel inter- X. Q. (1993b). Deletion of DNA encoding the first five transmembrane molecular association. Science 252, 1523–1528. domains of Epstein–Barr virus latent membrane proteins 2A and 2B. Hoffman, P., Rajakumar, P., Hoffman, B., Heuertz, R., Wold, W. S. M., J. Virol. 67, 5068–5074. and Carlin, C. R. (1992). Evidence for intracellular down-regulation Louie, R. R., King, C. S., MacAuley, A., Marth, J. D., Perlmutter, R. M., of the epidermal growth factor (EGF) receptor during adenovirus and Cooper, J. A. (1988). p56lck protein–tyrosine kinase is cytoskele- infection by an EGF-independent mechanism. J. Virol. 66, 197–203. tal and does not bind to polyomavirus middle T antigen. J. Virol. 62, Horak, I. D., Kawakami, T., Gregory, F., Robbins, K. C., and Bolen, J. B. 4673–4679. (1989). Association of p60fyn with middle tumor antigen in murine Lund, T., Medveczky, M. M., Neame, P. J., and Medveczky, P. G. (1996). polyomavirus-transformed rat cells. J. Virol. 63, 2343–2347. A herpesvirus saimiri membrane protein required for interleukin-2 Howe, L. R., and Weiss, A. (1995). Multiple kinases mediate T-cell- independence forms a stable complex with p56lck. J. Virol. 70, 600– receptor signaling. Trends Biochem. Sci. 20, 59–64. 606. Jung, J. U., Trimble, J. J., King, N. W., Biesinger, B., Fleckenstein, B. W., Markland, W., and Smith, A. E. (1987). Mutants of polyomavirus middle- and Desrosiers, R. C. (1991). Identification of transforming genes of T antigen. Biochim. Biophys. Acta 907, 299–321. subgroup A and C strains of Herpesvirus saimiri. Proc. Natl. Acad. Miller, C. L., Lee, J. H., Kieff, E., Burkhardt, A. L., Bolen, J. B., and Long- Sci. USA 88, 7051–7055. necker, R. (1994a). Epstein–Barr virus protein LMP2A regulates reac- Jung, J. U., Lang, S. M., Friedrich, U., Jun, T., Roberts, T. M., Desrosiers, tivation from latency by negatively regulating tyrosine kinases in- R. C., and Biesinger, B. (1995a). Identification of Lck-binding elements volved in sIg-mediated signal transduction. Infect. Agents Dis. 3, in tip of herpesvirus saimiri. J. Biol. Chem. 270, 20660–20667. 128–136. Jung, J. U., Lang, S. M., Jun, T., Roberts, T. M., Veillette, A., and Desro- Miller, C. L., Lee, J. H., Kieff, E., and Longnecker, R. (1994b). An integral

AID VY 8316 / 6a22$$$281 12-31-96 09:48:41 vira AP: Virology 280 MESSERSCHMITT, DUNANT, AND BALLMER-HOFER

membrane protein (LMP2) blocks reactivation of Epstein–Barr virus Effects of SH2 and SH3 deletions on the functional activities of wild- from latency following surface immunoglobulin crosslinking. Proc. type and transforming variants of c-Src. Mol. Cell. Biol. 12, 1835– Natl. Acad. Sci. USA 91, 772–776. 1845. Miller, C. L., Burkhardt, A. L., Lee, J. H., Stealey, B., Longnecker, R., Songyang, Z., Sheolson, S. E., Chaudhuri, M., Gish, G., Pawson, T., Bolen, J. B., and Kieff, E. (1995). Integral membrane protein 2 of Ep- Haser, W. G., King, F., Roberts, T., Ratnofsky, S., Lechleider, R. J., stein–Barr virus regulates reactivation from latency through domi- Neel, B. G., Birge, R. B., Fajardo, J. E., Chou, M. M., Hanafusa, H., nant negative effects on protein–tyrosine kinases. Immunity 2, 155– Schaffhausen, B., and Cantley, L. C. (1993). SH2 domains recognize 166. specific phosphopeptide sequences. Cell 72, 767–778. Molina, T. J., Kishihara, K., Siderovski, D. P., van Ewijk, W., Narendran, Sontag, E., Fedorov, S., Kamibayashi, C., Robbins, D., Cobb, M., and A., Timms, E., Wakeham, A., Paige, C. J., Hartmann, K. U., and Veil- Mumby, M. (1993). The interaction of SV40 with lette, A. (1992). Profound block in thymocyte development in mice protein phosphatase 2A stimulates the map kinase pathway and lacking p56lck. Nature 357, 161–164. induces cell proliferation. Cell 75, 887–897. Mungre, S., Enderle, K., Turk, B., Porras, A., Wu, Y. Q., Mumby, M. C., Srinivas, S., Scho¨nthal, A., and Eckhart, W. (1994). Polyomavirus middle- and Rundell, K. (1994). Mutations which affect the inhibition of protein sized tumor antigen modulates c-Jun phosphorylation and transcrip- phosphatase 2A by simian virus 40 small-t antigen in vitro decrease tional activity. Proc. Natl. Acad. Sci. USA 91, 10064–10068. viral transformation. J. Virol. 68, 1675–1681. Straight, S. W., Hinkle, P. M., Jewers, R. J., and McCance, D. J. (1993). Pallas, D. C., Shahrik, L. K., Martin, B. L., Jaspers, S., Miller, T. B., Brauti- The E5 oncoprotein of human papillomavirus type 16 transforms gan, D. L., and Roberts, T. M. (1990). Polyoma small and middle T fibroblasts and effects the downregulation of the epidermal growth antigens and SV40 small t antigen form stable complexes with pro- factor receptor in keratinocytes. J. Virol. 67, 4521–4532. tein phosphatase 2A. Cell 60, 167–176. Su, W., Liu, W., Schaffhausen, B. S., and Roberts, T. M. (1995). Associa- Pallas, D. C., Fu, H., Haehnel, L. C., Weller, W., Collier, R. J., and Roberts, tion of polyomavirus middle tumor antigen with phospholipase C- T. M. (1994). Association of polyomavirus middle tumor antigen with gamma 1. J. Biol. Chem. 270, 12331–12334. Taylor, S. J., and Shalloway, D. (1996). Src and the control of cell divi- 14-3-3 proteins. Science 265, 535–537. sion. BioEssays 18, 9–11. Pawson, T. (1994). SH2 and SH3 domains in signal transduction. Adv. Thomas, J. E., Aguzzi, A., Soriano, P., Wagner, E. F., and Brugge, J. S. Cancer Res. 64, 87–110. (1993). Induction of tumor formation and cell transformation by poly- Pipas, J. M. (1992). Common and unique features of T antigens encoded oma middle T antigen in the absence of Src. Oncogene 8, 2521– by the polyomavirus group. J. Virol. 66, 3979–3985. 2529. Qu, L., and Rowe, D. (1992). Epstein–Barr virus latent gene expression Tooze, J. (1980). ‘‘DNA Tumor Viruses,’’ 2nd ed. Cold Spring Harbor in uncultured peripheral blood lymphocytes. J. Virol. 66, 3715–3724. Laboratory Press, Cold Spring Harbor, NY. Reinholdsson Ljunggren, G., Ramqvist, T., Ahrlund Richter, L., and Dali- Urich, M., El Shemerly, M. Y., Besser, D., Nagamine, Y., and Ballmer- anis, T. (1992). Immunization against polyoma tumors with synthetic Hofer, K. (1995). Activation and nuclear translocation of mitogen- peptides derived from the sequences of middle- and large-T anti- activated protein kinases by polyomavirus middle-T or serum depend gens. Int. J. Cancer 50, 142–146. on phosphatidylinositol 3-kinase. J. Biol. Chem. 270, 29286–29292. Rudd, C. E., Janssen, O., Prasad, K. V., Raab, M., da Silva, A., Telfer, van der Geer, P., and Pawson, T. (1995). The PTB domain: A new protein J. C., and Yamamoto, M. (1993). src-related protein tyrosine kinases module implicated in signal transduction. Trends Biochem. Sci. 20, and their surface receptors. Biochim. Biophys. Acta 1155, 239–266. 277–280. Ruediger, R., Roeckel, D., Fait, J., Bergqvist, A., Magnusson, G., and Weiss, A. (1993). T cell antigen receptor signal transduction: A tale of Walter, G. (1992). Identification of binding sites on the regulatory A tails and cytoplasmic protein–tyrosine kinases. Cell 73, 209–212. subunit of protein phosphatase 2A for the catalytic C subunit and Whitman, M., Kaplan, D. R., Schaffhausen, B. S., Cantley, L., and Rob- for tumor antigens of simian virus 40 and polyomavirus. Mol. Cell. erts, T. M. (1985). Association of phosphatidylinositol kinase activity Biol. 12, 4872–4882. with polyoma middle-T competent for transformation. Nature 315, Ruediger, R., Hentz, M., Fait, J., Mumby, M., and Walter, G. (1994). 239–242. Molecular model of the A subunit of protein phosphatase 2A: Interac- Wiese, N., Tsygankov, A. Y., Klauenberg, U., Bolen, J. B., Fleischer, B., tion with other subunits and tumor antigens. J. Virol. 68, 123–129. and Bro¨ker, B. M. (1996). Selective activation of T cell kinase p56lck Scheidtmann, K. H., Mumby, M. C., Rundell, K., and Walter, G. (1991). by Herpesvirus saimiri protein Tip. J. Biol. Chem. 271, 847–852. Dephosphorylation of simian virus 40 large-T antigen and p53 protein Yang, S. I., Lickteig, R. L., Estes, R., Rundell, K., Walter, G., and Mumby, by protein phosphatase 2A: Inhibition by small-t antigen. Mol. Cell. M. C. (1991). Control of protein phosphatase 2A by simian virus 40 Biol. 11, 1996–2003. small-t antigen. Mol. Cell. Biol. 11, 1988–1995. Scherneck, S., and Feunteun, J. (1990). The hamster polyomavirus—A Zullo, J., Stiles, C. D., and Garcea, R. L. (1987). Regulation of c-myc and brief review of recent knowledge. Arch. Geshwulstforsch. 60, 271– c-fos mRNA levels by polyomavirus: Distinct roles for the capsid 278. protein VP1 and the viral early proteins. Proc. Natl. Acad. Sci. USA Seidel-Dugan, C., Meyer, B. E., Thomas, S. M., and Brugge, J. S. (1992). 84, 1210–1214.

AID VY 8316 / 6a22$$$281 12-31-96 09:48:41 vira AP: Virology