Recent studies appear to associate natural SV40 strains with human brain tumors of early childhood. The overall goals are to understand the pathways that contribute to the formation of a DNA tumor virus, SV40, and to understand how tumor antigens play a role in morphogenesis. The work proposed here concerns defining individual protein domains of SV40 structural proteins that determine the individual steps in morphogenesis and relating these domains to their biological activities. We will use molecular biological, genetic, biochemical and immunological approaches with cultured host cells to define a number of determinants that function in SV40 morphogenesis. A basic understanding of the functional determinants is important to understand which mediates infection and which prevents the spread of the DNA tumor viruses. In addition, success in identifying the component and specific protein domains of viral proteins, which are responsible for effective nuclear targeting of SV40 DNA, may lead to development of a general and effective means to exogenously introduce DNA into mammalian cells. Our specific goals are: (l) Development and evaluation of methods that enable the measurement of gene activity at a single cells level, and characterization of assembly intermediates in the nucleus. (2) Structure-function analysis of Vp2 and Vp3 proteins. (3) Structure-function analysis of Vp1. (4) We plan to identity the viral protein and protein domain important for nuclear targeting of infectious virion particles. We also plan to study sub-virion assembly in the cytoplasm. (5) For those protein domains with defined functions, complementation tests will be performed with a newly constructed non-overlap SV40 variant, in which genes for Vp2 and Vp3 are separated from Vp1 gene, to identity each of the individual steps involved in morphogenesis. (6) Study of the role of viral structural proteins in nuclear targeting of viral DNA.
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