(20) The Functional Importance of Reovirus σ1 Attachment Protein

Non-enveloped virus entry into cells is a poorly understood process. Reovirus, a non-enveloped virus with a segmented, double-stranded (ds) RNA genome is an oncolytic virus and a useful model for studies of non-enveloped... [ view full abstract ]

Non-enveloped virus entry into cells is a poorly understood process. Reovirus, a non-enveloped virus with a segmented, double-stranded (ds) RNA genome is an oncolytic virus and a useful model for studies of non-enveloped virus entry.  The reovirus genome is composed of ten segments of dsRNA, which encode 12 proteins.  The S1 genome segment encodes viral attachment protein σ1, which forms a trimeric molecule containing tail, body, and head domains.  During infection, viral attachment protein σ1 engages sialylated glycans and junctional adhesion molecule-A, triggering clathrin-dependent endocytosis.  Within the endocytic compartment, virions are proteolytically converted to infectious subvirion particles (ISVPs). Further disassembly permits release of transcriptionally active reovirus cores into the cytosol through a mechanism that is incompletely understood.  Electron microscopy has revealed a distinct conformational change in σ1 from a compact form on virions to an extended form on ISVPs, inspiring the hypothesis that σ1 extension in the endosome is imperative for reovirus escape into the cytosol.  To explore the importance of σ1 conformational mobility, we introduced cysteine pairs into structurally adjacent sites in the σ1 head and body domains using reverse genetics. We are also engineering cysteine residues in the σ1 tail. These cysteines are anticipated to create disulfide bonds that will crosslink σ1 and may impair its endocytic extension.  We are characterizing the replication, attachment, and entry phenotypes of the engineered viruses. These studies will enhance an understanding of non-enveloped virus entry mechanisms and may provide information useful for modification of σ1 to enable more efficient targeting of cancer cells.