Utilization of Sialic Acid as a Coreceptor Enhances Reovirus Attachment by Multistep Adhesion Strengthening

Abstract

Many serotype 3 reoviruses bind to two different host cell molecules, sialic acid and an unidentified protein, using discrete receptor-binding domains in viral attachment protein, 1. To determine mechanisms by which these receptor-binding events cooperate to mediate cell attachment, we generated isogenic reovirus strains that differ in the capacity to bind sialic acid. Strain SA؉, but not SA؊, bound specifically to sialic acid on a biosensor chip with nanomolar avidity. SA؉ displayed 5-fold higher avidity for HeLa cells when compared with SA؊, although both strains recognized the same proteinaceous receptor. Increased avidity of SA؉ binding was mediated by increased k on . Neuraminidase treatment to remove cell-surface sialic acid decreased the k on of SA؉ to that of SA؊. Increased k on of SA؉ enhanced an infectious attachment process, since SA؉ was 50 -100-fold more efficient than SA؊ at infecting HeLa cells in a kinetic fluorescent focus assay. Sialic acid binding was operant early during SA؉ attachment, since the capacity of soluble sialyllactose to inhibit infection decreased rapidly during the first 20 min of adsorption. These results indicate that reovirus binding to sialic acid enhances virus infection through adhesion of virus to the cell surface where access to a proteinaceous receptor is thermodynamically favored.

Cathepsin L and Cathepsin B Mediate Reovirus Disassembly in Murine Fibroblast Cells

“…Monolayers were washed twice with PBS, 0.5% Triton X-100, and infected cells were visualized by indirect immunofluorescence. Infected cells were identified by the presence of intense cytoplasmic fluorescence that was excluded from the nucleus (41). Background staining in uninfected control monolayers was not detected.…”

Crystal structure of human junctional adhesion molecule 1: Implications for reovirus binding

“…For type 3 reoviruses, this coreceptor is ␣-linked sialic acid (22), and its binding site has been mapped to a region close to the midpoint of the 1 tail (20,23). The finding that reoviruses bind to different receptors by using distinct domains within the 1 protein has led to the suggestion that reoviruses use a multiplestep adhesion-strengthening mechanism to engage the cell surface (21). In this scenario, reovirus binding to carbohydrate facilitates viral attachment through low-affinity adhesion.…”

“…and T.S.D., unpublished observations). Some reoviruses also use additional carbohydratebased coreceptors for cell attachment (20,21). For type 3 reoviruses, this coreceptor is ␣-linked sialic acid (22), and its binding site has been mapped to a region close to the midpoint of the 1 tail (20,23).…”

The Reovirus σ1 Aspartic Acid Sandwich

“…For serotype 3 reoviruses, viral attachment is a multistep process initiated by low affinity binding to sialic acid followed by high affinity binding to junctional adhesion molecule-A (JAM-A) (1,2). These steps are mediated by discrete receptorbinding domains in the attachment protein, 1 (3), a fiber-like molecule with head-and-tail morphology (4 -6).…”