Characterization of the nonstructural protein NS80 of grass carp reovirus

Nonstructural proteins of members of the family Reoviridae are believed to play significant roles in the virus replication cycle. Phylogenetic analyses indicate that the nonstructural protein NS80 of grass carp reovirus, encoded by a gene of Segment 4 (S4), is a primary determinant that is related to the formation of viroplasmic inclusion bodies (VIB), where viral replication and assembly are thought to occur. To understand the role of the NS80 protein in viral replication, an initial investigation of NS80 gene expression in both infected and transfected cells was conducted. Transmission electron microscopy results indicate that replication and assembly of GCRV occur within VIB-like structures in the perinuclear region of the cell cytoplasm. Furthermore, expression of the S4 gene in infected cells was detected with an NS80-specific antibody by western blot and immunofluorescence. Moreover, globular VIB-like structures were observed when expressing GFP-derived full-length NS80 (pEGFP-C1/NS80) and recombinants containing the C-terminal conserved region (pEGFP-C1/NS80₃₃₅₋₇₂₄) in transfected Vero. No such structures were detected in cells transfected with an N-terminal recombinant (pEGFP-C1/NS80₁₋₃₃₄), suggesting that the NS80 C-terminal conserved region may be involved in the formation of inclusion structures. These data provide a foundation for further functional studies of NS80 related to viral inclusion formation in viral replication.