VIRAL SMALL RNPS--ROLES IN CELL TRANSFORMATION

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Small RNPs present in cells transformed by certain primate Herpesviruses are assembled from virus-encoded small RNAs and host protein components. The two EBERs (Epstein Barr encoded RNAS) of EBV tightly bind the cellular lupus antigen La (50 kd), while the four newly discovered HSURs (Herpes saimiri U-like RNAS) acquire 5'-m3G caps and associate with proteins common to the snRNPs responsible for splicing and other premessenger RNA processing reactions in mammalian cell nuclei. Both the EBERs and HSURs are localized in the nucleus and are among the few viral gene products expressed in transformed lymphocytes. Our long term goals are to understand how these viral small RNPs contribute to the transformed state and how t cell in turn controls their biogenesis. To achieve these goals we propose the following strategies: 1) We will ask, using in fl= assays and immunoprecipitation, whether EBERs might regulate the activity of the interferon-induced 2',5'-oligoA synthetase, since they do not have the same inhibitory effect as VAI on the activity of the interferon-induced protein kinase. We will construct EBER affinity columns to identify and subsequently characterize nuclear proteins that bind EBERS. 2) We will ask whether the high abundance of EBERs in EBV-transformed cells serves to ' maintain high levels of important cellular small RNAs (tRNAs, 5S) because of the sequestration of a large fraction of the cellular La protein. We will further probe La's function in transcription termination in vitro. 3) We will identify and characterize the transcription factors that bind to regulatory sequences upstream of the EBER genes (and the genes for HVP1 and 2 RNAs of Herpesvirus papio). Those that are also involved in transcription of cellular and viral oncogenes will be studied to ascertain co-regulation by phosphorylation, extracellular signals, protein interaction, etc. 4) The function of HSURs will be probed based on the assumption that they act in polyadenylation. Long-elusive comparable host-RNAs will be sought using HSUR probes. 5) Other primate Herpesviruses will be examined for the presence of novel small RNAs using various patient autoantibodies.