Discrimination between sialic acid-containing receptors and pseudoreceptors regulates polyomavirus spread in the mouse

Abstract

Variations in the polyomavirus major capsid protein VP1 underlie important biological differences between highly pathogenic large-plaque and relatively nonpathogenic small-plaque strains. These polymorphisms constitute major determinants of virus spread in mice and also dictate previously recognized strain differences in sialyloligosaccharide binding. X-ray crystallographic studies have shown that these determinants affect binding to the sialic acids. Here we report results of further experiments designed to test the importance of specific contacts between VP1 and the carbohydrate moieties of the receptor. With minor exceptions, substitutions at positions predicted from crystallography to be important in binding the terminal alpha-2,3-linked sialic acid or the penultimate sugar (galactose) destroyed the ability of the virus to replicate in cell culture. Substitutions that prevented binding to a branched disialyloligosaccharide were found to result in viruses that were both viable in culture and tumorigenic in the mouse. Conversely, substitutions that allowed recognition and binding of the branched carbohydrate chain inhibited spread in the mouse, though the viruses remained viable in culture. Mice of five different inbred strains, all highly susceptible to large-plaque virus, showed resistance to the spread of polyomavirus strains bearing the VP1 type which binds the branched-chain receptor. We suggest that glycoproteins bearing the appropriate O-linked branched sialyloligosaccharide chains are effective pseudoreceptors in the host and that they block the spread of potentially tumorigenic or virulent virus strains.

FIG. 1

Interactions between polyomavirus VP1 and sialyloligosaccharide receptors, as seen in their crystal structures (–34). The oligosaccharide receptor is shown as a ball-and-stick model, with the straight-chain receptor being represented with light shading and the branching α-2,6-linked sialic acid being represented with darker shading. Hydrogen bonds between VP1 residues and the receptor are shown as broken lines; hydrophobic interactions are represented with arrows. The molecular surface of a polyomavirus VP1 pentamer complexed with the branched oligosaccharide is shown in the inset. The figure was prepared with RIBBONS (M. Carson, University of Alabama at Birmingham) and GRASP (B. Honig and A. Nicholls, Columbia University).

FIG. 2

Whole-mouse-section hybridization. Newborn C3H/BiDa mice were inoculated with 2 × 10⁵ to 5 × 10⁵ PFU of viruses of the indicated strains and sacrificed at 7 days. See Materials and Methods.

FIG. 3

Whole-mouse-section hybridization with different mouse strains. Newborn mice of the indicated strains were inoculated with 2 × 10⁵ to 5 × 10⁵ PFU of wild-type PTA or A2, each encoding E91 (top in each pair), or PTA-E91G (bottom in each pair) and sacrificed at 10 to 12 days. See Materials and Methods.

FIG. 4

Whole-mouse-section hybridization. Newborn C3H/BiDa mice were inoculated with the indicated virus strains. Shown in the top row (left to right) are results with RA (5 × 10⁶ to 10 × 10⁶ PFU after 7 days), RA-G91E (2 × 10⁵ to 5 × 10⁵ PFU after 7 days), and RA-G91E V296A (2 × 10⁵ to 5 × 10⁵ PFU after 7 days). Shown in the bottom row (left to right) are results with RA (2 × 10⁶ to 5 × 10⁶ PFU after 12 days) and RA-V296A (2 × 10⁶ to 5 × 10⁶ PFU after 12 days). See Results.