Reoviruses provide a well-established experimental model for studies of viral neuropathogenesis. Different reovirus serotypes elicit distinct patterns of disease within the central nervous system (CNS) of newborn mice. Following primary infection in the intestine, type 1 reoviruses spread hematogenously and infect ependymal cells, producing hydrocephalus. In contrast, type 3 reoviruses spread through nerves and infect neurons, causing encephalitis. The capacity of the reovirus attachment protein, sigma1, to bind either ependymal cells or neurons is a major determinant of these different patterns of CNS disease. In addition, sigma1 plays an important role in the capacity of reovirus to infect intestinal tissue and disseminate to the CNS. The sigma1 protein of type 3 reovirus is hypothesized to mediate cell-attachment by two receptor-binding domains, one of which binds sialic acid. The objective of this proposal is to define the molecular determinants of sigma1 that mediate the pathogenesis of reovirus-induced CNS injury. In Specific Aim 1, sequences in sigma1 that confer viral tropism for neural tissues will be identified using both chimeric sigma1 proteins constructed from different reovirus serotypes and synthetic sigma1 oligopeptides. Mutant sigma1 proteins and synthetic sigma1 oligopeptides will be tested for their capacity to bind reovirus receptors in primary cultures of ependymal cells and neurons. In Specific Aim 2, the role of 01 binding to sialylated receptors in reovirus-induced neurologic injury will be determined using type 3 reovirus mutants that vary in their capacity to bind sialic acid. These mutants will be tested for properties of growth, dissemination, and tropism in newborn mice. In Specific Aim 3, the mechanism of sigma1 -mediated differences in reovirus infectivity in the intestine will be investigated by determining the molecular basis for susceptibility of sigma1 to cleavage by intestinal proteases. Reovirus reassortant genetics will be used to determine whether sigma1 cleavage susceptibility is linked to poor growth in the intestine and failure to spread to the CNS. These studies will yield a precise understanding of reovirus cell-attachment at the molecular level and provide novel insights into general mechanisms by which viruses select cellular targets and produce neurologic disease.