Molecular biological studies on the neurovirulence of murine coronavirus

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A highly neurotropic murine coronavirus JHM strain has been studied as a model of viral agent to cause acute and subacute encephalomyelitis with demyelination. As a viral factor involving in the neurovirulence. the spike(S)protein comprising the projections on the virion surface is considered to be deeply involved. To study the mechanisms by which JHM strain exhibits the neurovirulence, we have done the molecular biological analysis of S protein of variant JHM viruses showing different neurovirulence. We first of all have isolated a highly neurovirulent variant, cl-2, for rats. This variant has been shown to have a larger S protein as compared with other variants with avirulence. We have isolated the cDNA from mRNA3 encoding the larger S protein and sequence analysis has identified that the larger S protein has 141 amino acid clusters which is missing in a small S protein of avirulent viruses. The 141 amino acid cluster may be implicated with high neurovirulence, although the precise role of the amino acids is yet unclear. We further studied on the larger S protein by expressing in mammalian cells with recombinant vaceinia viruses. The S protein expressed showed no difference in molecular size, immunogenicity or antigenicity as compared with the authentic S protein. Furthermore, the expressed S protein exhibited the fusionability in various cultured cells. These results suggest that the S protein expressed by recombinant vaceinia virus is appropriate for the study of biological activities of the S protein. Experiments are now in progress to see the neurovirulence of JHMV using the recombinant vaccinia viruses with large S protein cDNA.