Reovirus Nonstructural Protein σNS Acts as an RNA-Stability Factor Promoting Viral Genome Replication.

Viral nonstructural proteins, which are not packaged into virions, are essential for the replication of most viruses. Reovirus, a nonenveloped, double-stranded RNA (dsRNA) virus, encodes three nonstructural proteins that are required for viral replication and dissemination in the host. The reovirus nonstructural protein sigma NS is a single-stranded RNA (ssRNA)-binding protein that must be expressed in infected cells for production of viral progeny. However, the activities of sigma NS during individual steps of the reovirus replication cycle are poorly understood. We explored the function of sigma NS by disrupting its expression during infection using cells expressing a small interfering RNA (siRNA) targeting the sigma NS-encoding S3 gene and found that sigma NS is required for viral genome replication. Using complementary biochemical assays, we determined that sigma NS forms complexes with viral and nonviral RNAs. We also discovered, using in vitro and cell-based RNA degradation experiments, that sigma NS increases the RNA half-life. Cryoelectron microscopy revealed that sigma NS and ssRNAs organize into long, filamentous structures. Collectively, our findings indicate that sigma NS functions as an RNA-binding protein that increases the viral RNA half-life. These results suggest that sigma NS forms RNA-p rote in complexes in preparation for genome replication. IMPORTANCE Following infection, viruses synthesize nonstructural proteins that mediate viral replication and promote dissemination. Viruses from the family Reoviridae encode nonstructural proteins that are required for the formation of progeny viruses. Although nonstructural proteins of different viruses in the family Reoviridae diverge in primary sequence, they are functionally homologous and appear to facilitate conserved mechanisms of dsRNA virus replication. Using in vitro and cell culture approaches, we found that the mammalian reovirus nonstructural protein sigma NS binds and stabilizes viral RNA and is required for genome synthesis. This work contributes new knowledge about basic mechanisms of dsRNA virus replication and provides a foundation for future studies to determine how viruses in the family Reoviridae assort and replicate their genomes.