RNA PROTEIN INTERACTIONS IN POLIOVIRUS REPLICATION

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DESCRIPTION (Adapted from Applicant's Summary): The long-term goal of this project is to understand the mechanism of poliovirus replication and its interaction with the host-cell. Specifically, the applicant is interested in studying initiation of RNA synthesis and its relationship to translation. The focus is on dissecting the structure and function of a ribonucleoprotein complex (RNP-B) that appears to participate in the initiation of positive-strand RNA synthesis. The RNA component of this ribonucleoprotein complex folds into a cloverleaf-like structure and binds the viral protein 3CD and a ribosome-associated factor (p 36/PCBP). The identity of the host-cell factor has recently been determined, but the function of the ribonucleoprotein complex has yet to be defined. Dr. Andino has developed a novel system in which to study poliovirus replication. Microinjection of poliovirus RNA into Xenopus laevis oocytes initiates a complete and authentic viral replication cycle, which yields a high level of infectious viruses, but only if polioviral RNA is co-infected with factors present in HeLa cells. Two HeLa cell factors are required for viral replication in oocytes, one involved in initiation of translation, and the other in RNA synthesis. Thus, microinjection in oocytes can be used essentially as an in vitro system in which to identify and further analyze the function of viral and cellular factors, and to biochemically dissect the mechanism of initiation of poliovirus RNA synthesis. The specific aims of this proposal are to determine the structure of the NRP-B complex, to study its function in oocytes and human cells, and to characterize human factors require for poliovirus replication in oocytes.