The overall aim of the proposed research program is the identify the functions of the proteins encoded by reovirus.
Six specific aims are proposed. The first is to isolate the native forms of all reovirus proteins from cells infected with a powerful mammalian expression vector into which the reovirus genome segments have been cloned. The isolated proteins will be examined for enzymic activities and ability to bind to RNA and other proteins, both viral and cellular. Second, transient expression vectors will be used to examine the effect of reovirus proteins on DNA replication, transcription and protein synthesis. Genetically engineered genome segments will be used to identify functional domains. The intracellular location of reovirus proteins into which reovirus RNA species have been lipofected singly and in recombinant vaccinia virus strains containing the various genome segments. Third, techniques will be developed to introduce genetically engineered genome segments into reovirus genomes so as to permit study of the effect of alterations in functional domains on viral phenotype. Fourth, temperature-sensitive mutants and anti-sense technology will be used to determine at what stage inability of individual viral proteins to function arrests virus multiplication, thereby providing further clues to viral protein function. Fifth, selected SORFs in reovirus serotype 1, 2 and 3 cell receptors will be identified in gene libraries and sequenced in order to identify the nature of the receptors.
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