ASSEMBLY AND APPLICATION OF RECOMBINANT RHABDOVIRUSES

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DESCRIPTION (adapted from applicant s abstract): The long term objective of the project is to understand basic aspects of enveloped virus assembly and replication. Two model viral systems, vesicular stomatitis virus (VSV), and a new minimal virus system based partially on VSV will be used. VSV is the prototype of the rhabdovirus group and is one of the simplest and best characterized enveloped viruses. Recently Dr. Rose reported development of a system in which recombinant VSVs can be recovered from a complete DNA copy of the genome, and they have also reported a stable, highly efficient gene expression vector based on VSV. Additional studies will be performed to define RNA packaging limits in VSV and to generate new vectors suitable for expression of multiple genes. Such vectors are likely to have important application in vaccine development. The recombinant VSV system will be exploited in studies on the protein domains and modifications that are critical to efficient assembly of the glycoprotein into infectious particles. Basic studies will also be performed on incorporation of foreign membrane proteins into the VSV envelope. These studies will define the signals required for protein incorporation in the presence or absence of the wild-type VSV glycoprotein (G). An understanding of the rules governing efficient glycoprotein incorporation into virus particles could permit the design of novel vaccines derived by incorporating foreign glycoproteins into VSV particles. Other studies will be directed toward deriving VSVs with new targeting specificity based on incorporation of CD4 into the viral envelope. VSV is ideal for such studies because it grows rapidly to very high titers and is easily isolated in much larger quantities than most other enveloped viruses. Cell lines expressing the L and/or P genes of VSV will also be developed to permit the growth of defective VSVs lacking these genes and expressing very large amounts of foreign genetic material. Expression of the VSV G protein from a self-replicating RNA derived from Semliki Forest virus results in production of enveloped, infectious, self-propagating virus particles that contain G protein as their single structural protein. Dr. Rose s lab will continue basic studies on these minimal virus particles to determine the role of VSV G protein in their formation, to use them in a genetic selection of revertants of G protein fusion mutants, and to determine if they display any pathogenicity in animals.