Abstract
Viral infection is an intricate process that requires the concerted action of both viral and host cell components. Entry of viruses into cells is initiated by interactions between viral proteins and their cell surface receptors. Despite recent progress, the molecular mechanisms underlying the multistep reovirus entry process are poorly understood. Using atomic force microscopy, we investigated how the reovirus σ1 attachment protein binds to both α-linked sialic acid (α-SA) and JAM-A cell-surface receptors. We discovered that initial σ1 binding to α-SA favors a strong multivalent anchorage to JAM-A. The enhanced JAM-A binding by virions following α-SA engagement is comparable to JAM-A binding by infectious subvirion particles (ISVPs) in the absence of α-SA. Since ISVPs have an extended σ1 conformer, this finding suggests that α-SA binding triggers a conformational change in σ1. These results provide new insights into the function of viral attachment proteins in the initiation of infection and open new avenues for the use of reoviruses as oncolytic agents.
Highlights
Viral infection is an intricate process that requires the concerted action of both viral and host cell components
We discovered that initial σ1 binding to α-linked sialic acid (α-SA) acts as a trigger that enhances the overall avidity of σ1 for junctional adhesion molecule A (JAM-A), binding to which is a critical step in viral entry
In terms of number of bonds established between reovirus virions and JAM-A, our analysis reveals that after binding to α-SA, the reovirus-JAM-A binding potential is increased to a level similar to that of infectious subvirion particles (ISVPs)-JAM-A interaction (Fig. 6f, g)
Summary
Number of established bonds a complex between the sialylated glycan and the glycan-binding site in the serotype 3 σ1 tail domain (Fig. 1b). T3SA+ virions incubated with free Neu5Ac and ISVPs have a much higher avidity for JAM-A, reaching a very high affinity (KD ∼pM range) (Fig. 8a) These observations are consistent with our AFM data showing a binding potential of T3SA+ virions incubated with α-SA compounds comparable to that of ISVPs. The dynamics of reovirus binding to the surface of living cells was evaluated by SPT using high-speed confocal microscopy. Injection of free α-SA reduced T3SA+ diffusion on the cell surface, presumably by the capacity to mediate multivalent interactions with JAM-A, and significantly increased the number of particles bound to CHOJAM-A cells (Fig. 8f) This effect was not observed for the non-SA-binding strain T3SA− (Fig. 8f, Supplementary Fig. 9). These observations strengthen our conclusion that σ1 binding to α-SA induces a conformational change in σ1 that leads to an increase in the multivalent attachment of the virus to cell-surface receptors
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