The Overall Objective of the proposed investigation is to elucidate the role of the interferon-inducible RNA-specific adenosine deaminase (ADAR1) in the actions that interferons (IFN) mediate on viral and host functions.
The Specific Aims of our proposed continuation study of ADAR1 and double-stranded RNA editing in IFN action are as follows: (1) To further characterize the biochemical and biophysical properties of the IFN-inducible p150 ADAR1. To determine the mechanisms by which poxvirus E3L protein and adenovirus VA1 RNA antagonize ADAR1 enzymatic activity; to characterize the functional selectivity of dsRNA binding domains of ADAR1; and to determine the structural basis and functional significance of ADAR1 protein interactions and subcellular localizations in uninfected and virus-infected cells. (2) To further delineate the structure of the 5'-flanking regions of the ADAR1 gene required for IFN-inducible and basal transcriptional activity, including the Pi promoter responsible for inducible expression of transcripts encoding the p150 ADAR1 protein isoform and the Pb and Pc promoters responsible for constitutive expression of nuclear p110 ADAR1. To define the pathways leading to inducible transcriptional activation of ADAR1 expression in IFN-treated cells and in infected mice. (3) To further characterize the effect of singular stable expression of wild type and mutant forms of ADAR1 deaminase in cultured cells on virus multiplication including VA1 mutant adenovirus, E3L mutant vaccinia virus and recombinant measles virus, to generate a targeted deletion of the mouse ADAR1 inducible genomic locus, and to determine the effect of the ADAR1 p150 disruption on mouse phenotype and viral pathogenesis. The health relatedness of the proposed research stems from the likelihood that the work may contribute to a better understanding of regulatory mechanisms operative in normal cells as well as virus-infected cells. Furthermore, elucidation of the actions of IFN at the molecular level is of importance in view of potential applications of interferon in the clinic.
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