Activation of Stat3 Transcription Factor by Herpesvirus Saimiri STP-A Oncoprotein

Signal transducer and activator of transcription 3 (Stat3) protein belongs to a seven-member family of latent cytoplasmic transcription factors that contribute to signal transduction initiated by cytokines, hormones, and growth factors (13). Stat3 proteins control fundamental cellular processes, including survival, proliferation, and differentiation. Upon stimulation with interferons, interleukin-6, granulocyte-macrophage colony-stimulating factor, epidermal growth factor, or platelet-derived growth factor, Stat3 protein becomes activated by phosphorylation on a single tyrosine (Y705) and dimerizes through a reciprocal interaction between the SH2 domain of one monomer and the phosphorylated tyrosine of the other (29). The dimers accumulate in the nucleus, recognize specific DNA elements, and activate transcription. Given the critical roles of Stat3 protein, it is hypothesized that inappropriate or aberrant activation of Stat3 contributes to cellular transformation and, in particular, leukemogenesis (6, 19, 20). In fact, the constitutive activation of Stat3 has been detected in many cancer cells and tissues, including those of multiple myeloma, leukemia, lymphoma, breast, lung, and head and neck cancer (6, 19, 20). Furthermore, it has been shown that Stat3 exerts its growth-deregulating activity by activating the expression of cellular genes that are involved in cell cycle progression such as fos, cyclin D1, myc, and pim-1 and by activating antiapoptotic processes such as Bcl-2 and Bcl-XL (3, 5, 7, 17, 35). Thus, activation of the Stat3 signal transduction pathway is likely a common strategy used by various viruses to transform a normal cell to a cancerous cell (8, 38). In fact, viral Src has been shown to interact with Stat3 and to activate Stat3 to induce cell growth transformation (4, 34, 37). Herpesvirus saimiri (HVS) belongs to the gamma subfamily of herpesviruses (Gammaherpesvirinae). HVS naturally infects the squirrel monkey (Saimiri sciureus), a common South American primate, but with no apparent disease association. However, HVS infection of marmosets, owl monkeys, and other species of New World primates results in rapidly progressing fulminant lymphomas, lymphosarcomas, and leukemias of T-cell origin (18, 26). HVS can be further subclassified into three subgroups (subgroups A, B, and C) on the basis of the extent of DNA sequence divergence at the left end of coding DNA (32). Subgroups A and C are highly oncogenic and are able to immortalize common marmoset T lymphocytes to interleukin 2-independent growth in vitro (16, 36). Subgroup-C strains are further capable of immortalizing human, rabbit, and rhesus monkey lymphocytes into continuously proliferating T-cell lines (1, 2). Mutational analyses have demonstrated that the leftmost open reading frame in the coding sequence of subgroup A strain 11 is not required for viral replication but is required for immortalization of common marmoset T lymphocytes in vitro and for lymphoma induction in vivo (14, 15). This open reading frame is termed STP-A11, for saimiri transformation-associated protein of subgroup A strain 11 (33). At a position and an orientation equivalent to those of the STP-A11 reading frame, the highly oncogenic HVS subgroup C strain 488 contains a distantly related reading frame termed STP-C488 (2, 28). Despite limited sequence similarity, STP-A11 and STP-C488 seem to be organized similarly in terms of the presence and localization of basic structural elements (2, 28). Both proteins are predicted to have a highly acidic amino terminus and collagen-like repeats in the central region. The primary amino acid sequence of STP-A11 has 9 repeats of a collagen-like motif (Gly-X-Y, where X and/or Y is proline), and in STP-C488 this motif is directly repeated 18 times (2, 28). The STP-A11 and STP-C488 proteins also contain a hydrophobic stretch at their carboxyl termini sufficient for a membrane-spanning domain (27). Both STPs have transforming and tumor-inducing activities independent of the rest of the herpesvirus genome (28). Specifically, both can transform rodent fibroblast cells, resulting in apparent loss of contact inhibition, formation of foci, growth at reduced serum concentrations, and formation of invasive tumors in nude mice (28). To understand the structural and functional properties of STP-A, we analyzed the primary amino acid sequences of six different subgroup-A isolates (31). This analysis revealed that STP-A contains interesting structural and functional elements, including the 60PVQES64 binding motif for tumor necrosis factor (TNF) receptor-associated factors (TRAFs) and the 115YAEV118 SH2 binding motif for Src family kinases. Indeed, biochemical analysis has demonstrated that STP-A is capable of interacting with TRAF and Src kinase through the 60PVQES64 and 115YAEV118 motifs, respectively. While the role of TRAF interaction has not been well characterized, Src interaction has been shown to induce the tyrosine phosphorylation of STP-A11 as well as of other cellular proteins (30, 31). In this report, we further demonstrate that STP-A11 interacts with Stat3 independently of TRAF and Src, and that Src kinase associated with STP-A11 phosphorylates Stat3, resulting in its nuclear localization and transcriptional activation. Consequently, the constitutive activation of Stat3 induced by STP-A11 leads to cell survival and proliferation upon serum deprivation. Thus, the STP-A11 oncoprotein targets multiple cellular signaling molecules to elicit cell growth transformation, which ultimately contributes to T-cell transformation induced by HVS subgroup-A strains.