The TMEV are ideally suited viruses for studying molecular pathogenesis. First, the TMEV as picornaviruses are small, nonenveloped viruses which store their genetic information on one piece of single-strand, message- sense RNA, and thus have a relatively simple genome for microbes. Second, these naturally occurring enteric pathogens of mice can be divided into two neurovirulence groups. Not only do members of the two groups produce different CNS diseases in mice, they also can be distinguished by a number of phenotypic markers in vitro. Third, the nucleotide sequences of the complete genomes of three TMEV, the highly virulent GDVII virus and the less virulent BeAn and DA viruses have been determined revealing finite sequence differences between the members of the two neurovirulence groups. And finally, my lab has constructed a three-dimensional model of the coat proteins of one TMEV, BeAn virus, on the atomic coordinates of the closely related Mengovirus, providing valuable information about structural features of the Theiler's virion prior to crystallographic analysis. In this proposal we plan to determine whether amino acids in the putative viral receptor attachment site are involved in binding of TMEV to its cellular receptor (pit). This will be accomplished by oligonucleotide- directed mutation of selected amino acids of the 32 surface residues that comprise the pit. We also plan to determine if deletion of the two major VP1 loops, or parts of these loops, between beta strands C and D will result in viable BeAn virus deletion mutants that are susceptible to the Sterling Research Group WIN antiviral agents. Two WIN compounds have been shown to be effective in vivo against several of the human enteroviruses and human rhinoviruses but not against the cardioviruses. The two VP1 loops in cardioviruses appear to block drug entrance to the pore in the bottom of the pit and to the hydrophobic pocket in VP1. Having a viable BeAn deletion mutant that is susceptible the WIN antiviral drugs would permit treatment of mice with TMEV-induced demyelinating disease. Being able to eradicate TMEV CNS persistence would be a use tool for studying the immune effector mechanism of demyelination in this system.
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