Epstein-Barr virus (EBV) SM protein is an essential nuclear protein produced during the lytic cycle of EBV replication. and co-opts the function of cellular SRp20 IRAK3 in alternative splicing. SM protein (EB2, Mta, and BMLF1) is a nuclear phosphoprotein synthesized by Epstein-Barr virus (EBV) during the early stage of lytic replication (for a review, see reference 38). SM has multiple functions in enhancing EBV gene expression posttranscriptionally, binds target gene mRNA, enhances nuclear mRNA export and stability, and modulates cellular and EBV RNA splicing (2, 9, 17, 18, 23, 31, 35, 39, 40). SM is essential for EBV replication and EBV recombinants with insertional deletion of the SM gene are defective for virus production (12). SM is required for the efficient accumulation of ca. Sorafenib ic50 60% of EBV lytic transcripts (13). SM is required for efficient expression of both EBV DNA primase (BSLF1) and EBV DNA polymerase (BALF5) mRNAs, leading to severely impaired lytic EBV DNA replication in the absence of SM (13). SM also directly enhances accumulation of specific past due gene mRNAs furthermore to allowing DNA replication (13). This mix of results on DNA replication and past due gene mRNAs qualified prospects to a worldwide deficiency of past due gene manifestation in the lack of SM. We lately proven that SM works alternatively splicing element and modulates mobile splicing (40). The consequences of SM on sponsor mobile gene manifestation during lytic EBV replication stay to be completely characterized. When indicated in EBV-negative cells inducibly, SM includes a broadly inhibitory influence on mobile Sorafenib ic50 mRNA accumulation (30). Nevertheless, SM causes several cellular transcripts to accumulate at higher levels (30). These transcripts include STAT1 and several interferon-stimulated genes. The STAT1 protein is an integral mediator of both type I (alpha/beta interferon [IFN-/]) and type II (IFN-) IFN signal transduction pathways (for a review, see reference 7). STAT1 is expressed as two isoforms, STAT1 and STAT1. STAT1 mRNA is generated by cleavage and polyadenylation at an alternative site in the last intron of the STAT1 pre-mRNA, leading to production of a protein which lacks the transactivating domain encoded in the last exon of the STAT1 gene (see Fig. 5A). STAT1 homodimers are not capable of activating GAS sequences, and STAT1 may therefore act as a dominant-negative repressor of STAT1 (3, 27, 41). Consistent with a role for STAT1 as an antagonist of STAT1, the ratio of STAT1 and – isoforms has been shown to affect cellular apoptosis and resistance to viral infection (1, 26). Interestingly, SM disproportionately increases the relative amounts of STAT1 mRNA. Further investigation of previous findings that SM changed the ratio of two functionally distinct STAT1 isoforms generated by alternative processing (30) led to the finding that SM directed splicing of STAT1 to an alternative 5 splice site with high efficiency and specificity (40). This activity was based on preferential binding of SM to specific regions of the pre-mRNA, indicating that SM may function in a manner similar to cellular splicing factors. Although SM does bind to RNA directly (14, 29), it does not possess arginine-serine (RS) repeats typically found in cellular SR proteins that act as alternative splicing factors (11). We report here the interaction of SM with SRp20, a cellular SR protein, and its role in modulation of splicing by SM. MATERIALS AND METHODS Cell lines and transfections. 293 is a cell line derived from human embryonic kidney cells (10). 293T and HeLa cells were maintained in Dulbecco modified Eagle medium containing 10% fetal calf serum supplemented with Glutamax (Invitrogen). HeLa cell transfections were performed with Lipofectamine Plus (Invitrogen) in six-well plates using 1 g of DNA per transfection, according to the manufacturer’s protocols. 293T cells transfections were performed with TransIT293 reagent according to the manufacturer’s protocols (Mirus). Cells Sorafenib ic50 were harvested 48 h after transfection. P3HR1/ZHT cell line was derived from P3HR-1 (28) by transfection with a plasmid expressing a tamoxifen-inducible EBV Zta activator of lytic gene expression (pCDNA3-ZHT), followed by selection in.