Indirect evidence implicates a virus in the etiology of Multiple Sclerosis (MS) and more direct evidence shows that viruses can cause other human demyelinating diseases. Coronavirus, mouse hepatitis virus (MHV), strain A59 infection of weanling mice is a good animal model system for the study of virus induced demyelination. After the initial acute encephalitis, virus becomes impossible to detect during the chronic stages of disease while viral nucleic acids may be detected in white matter by in situ hybridization. Furthermore, MHV-A59 causes a persistent, productive, but non-lytic infection in primary glial cells in culture. The long term goal of this project is to determine the molecular basis for MHV-A59 persistence in glial cells in culture and in the mouse central nervous system (CNS) and to determine the relationship between viral persistence and chronic demyelination. In this proposal we plan to study the infection of cultures enriched in either astrocytes, oligodendrocyte or neurons with MHV-A59 and its attenuated fusion deficient variants. 1) We will determine why wild type MHV-A59 induces only a minimal amount of cell fusion in glial cells while nearly 100% of fibroblasts infected with this virus undergo fusion. For this question we will focus on synthesis and processing of the viral S glycoprotein in glial cells. 2) We will compare both acute and persistent infection with wild type and fusion deficient variants in cultures enriched for oligodendrocytes and astrocytes. We will examine mutations in viral surface glycoprotein genes that evolve during persistent infection.. We will clone viruses from enriched oligodendrocyte cultures and compare these to the fusion deficient viruses we have isolated and characterized from persistently infected mixed glial cultures. 3) We will infect cultures of neuronal origin with both wild type and variant viruses as well as the more virulent MHV strain, JHM. We will determine whether these viruses cause acute and/or persistent infections in neuronal cells. If these cells are infected, we will determine whether cell fusion occurs and we will characterize viruses isolated from such infections with respect to tropism and the ability to induce fusion.
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