Polyoma virus will continue to serve as a model system to study the synthesis and processing of mammalian RNA molecules. The applicant plans to perform a number of studies designed to understand the switch from mostly early-strand mRNAs before the initiation of viral DNA synthesis to mostly late-strand mRNAs afterwards. The regulation of both early and late strand RNA levels is not at the promoter level, and is post-transcriptional. Late-strand RNA levels are regulated in an unexpected way-they appear to positively influence their own accumulation during infection. He will determine what sequences are critical for the activation of late gene expression after DNA replication initiation. The regulation of early-strand gene expression by a novel mechanism (nuclear antisense RNA from the late strand) that is likely to be generally used by cells to regulate the expression of many of their own genes. This virus provides a powerful genetic and biochemical system in which to study antisense regulation and to learn how to exploit this knowledge to regulate the expression of other genes, both viral and cellular. He has shown that, in the presence of late-strand antisense molecules, many nuclear early-strand RNAs are modified by the enzyme double strand RNA adenosine deaminase (ADAR1). This appears to be a major aspect of the downregulation of early gene expression at late times in the viral life cycle. Edited RNAs are retained in the nucleus. He will use a variety of approaches to learn more about the nuclear retention of inosine-containing RNAs, and will investigate the role of ADAR1 in polyoma infection, and the influence of infection on ADAR1 expression, activity, and localization.
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