Structural and Functional Analysis of Murine Polyomavirus Capsid Proteins Establish the Determinants of Ligand Recognition and Pathogenicity

Abstract

Murine polyomavirus (MuPyV) causes tumors of various origins in newborn mice and hamsters. Infection is initiated by attachment of the virus to ganglioside receptors at the cell surface. Single amino acid exchanges in the receptor-binding pocket of the major capsid protein VP1 are known to drastically alter tumorigenicity and spread in closely related MuPyV strains. The virus represents a rare example of differential receptor recognition directly influencing viral pathogenicity, although the factors underlying these differences remain unclear. We performed structural and functional analyses of three MuPyV strains with strikingly different pathogenicities: the low-tumorigenicity strain RA, the high-pathogenicity strain PTA, and the rapidly growing, lethal laboratory isolate strain LID. Using ganglioside deficient mouse embryo fibroblasts, we show that addition of specific gangliosides restores infectability for all strains, and we uncover a complex relationship between virus attachment and infection. We identify a new infectious ganglioside receptor that carries an additional linear [α-2,8]-linked sialic acid. Crystal structures of all three strains complexed with representative oligosaccharides from the three main pathways of ganglioside biosynthesis provide the molecular basis of receptor recognition. All strains bind to a range of sialylated glycans featuring the central [α-2,3]-linked sialic acid present in the established receptors GD1a and GT1b, but the presence of additional sialic acids modulates binding. An extra [α-2,8]-linked sialic acid engages a protein pocket that is conserved among the three strains, while another, [α-2,6]-linked branching sialic acid lies near the strain-defining amino acids but can be accommodated by all strains. By comparing electron density of the oligosaccharides within the binding pockets at various concentrations, we show that the [α-2,8]-linked sialic acid increases the strength of binding. Moreover, the amino acid exchanges have subtle effects on their affinity for the validated receptor GD1a. Our results indicate that both receptor specificity and affinity influence MuPyV pathogenesis.

Murine Polyomavirus Cell Surface Receptors Activate Distinct Signaling Pathways Required for Infection

“…Finally, to determine whether MuPyV ganglioside receptors activate specific signaling pathways versus non-MuPyV ganglioside receptors, we compared virus signal activation after supplementation with different gangliosides. The gangliosides GD1a and GT1a are known receptors for MuPyV, whereas GM1 is a receptor for SV40 PyV (13, 31). Virus addition to GD1a- and GT1a-supplemented ganglioside −/− MEFs resulted in increased ERK phosphorylation compared to the DMSO control (Fig.…”

Type I Interferons Regulate the Magnitude and Functionality of Mouse Polyomavirus-Specific CD8 T Cells in a Virus Strain-Dependent Manner

“…The single-amino-acid E-to-G exchange at position 91 in VP1 causes MPyV virions to bind not only to terminally unbranched sialic acid true cellular receptors but also to branched-chain sialic acid "pseudoreceptors," resulting in attenuation of viral spread and tumorigenicity (6,9). We validated that the E-to-G substitution at VP1 position 91 dampened viral replication in vivo and that it did so in adult B6 mice, which are highly resistant to MPyV tumorigenesis (53).…”

“…In SP strains, VP1 capsids with G-91 interact with branched (␣-2,6)-linked sialyloligosaccharides, which may act as pseudoreceptors by binding cell surface glycoproteins that divert virions into noninfectious pathways (8). An E at this position in VP1 leads to electrostatic repulsion of the (␣-2,6)-linked sialic acids, thereby preventing binding of such branched structures by LP strains; however, binding to gangliosides with sialic acid (␣-2,3)-linked to galactose is retained for virion uptake into an infectious pathway (9,10). Interestingly, MPyVs isolated from feral mice have exclusively E-91 VP1s, an unexpected finding given that such LP viruses are potentially more oncogenic than G-91 SP viruses (11).…”

Sialoglycovirology of Lectins: Sialyl Glycan Binding of Enveloped and Non-enveloped Viruses

“…SV40, mPy, and BKPyV (either caveolin-dependent or caveolin-independent entry) share a common binding for ganglio-series gangliosides [166,[170][171][172] with differential binding preferences. SV40 prefers the branched α2-3Sia GM1 ganglioside [170], mPyV mainly attaches α2-3Sia on GD1a/GT1b/GT1a [173,174], and BKPyV and MCPyV commonly prefer to bind to α2-8Sia b-series gangliosides including GD3/GD2/GD1b/GT1b for BKPyV [171] and GT1b in cooperative binding with a glycosaminoglycan (GAG) for MCPyV [172]. Comparison of the crystal structures of BKPyV VP1-GD3 and SV40 VP1-GM1 ( Fig.…”