Both α2,3- and α2,6-Linked Sialic Acids on O-Linked Glycoproteins Act as Functional Receptors for Porcine Sapovirus

Abstract

Sapovirus, a member of the Caliciviridae family, is an important cause of acute gastroenteritis in humans and pigs. Currently, the porcine sapovirus (PSaV) Cowden strain remains the only cultivable member of the Sapovirus genus. While some caliciviruses are known to utilize carbohydrate receptors for entry and infection, a functional receptor for sapovirus is unknown. To characterize the functional receptor of the Cowden strain of PSaV, we undertook a comprehensive series of protein-ligand biochemical assays in mock and PSaV-infected cell culture and/or piglet intestinal tissue sections. PSaV revealed neither hemagglutination activity with red blood cells from any species nor binding activity to synthetic histo-blood group antigens, indicating that PSaV does not use histo-blood group antigens as receptors. Attachment and infection of PSaV were markedly blocked by sialic acid and Vibrio cholerae neuraminidase (NA), suggesting a role for α2,3-linked, α2,6-linked or α2,8-linked sialic acid in virus attachment. However, viral attachment and infection were only partially inhibited by treatment of cells with sialidase S (SS) or Maackia amurensis lectin (MAL), both specific for α2,3-linked sialic acid, or Sambucus nigra lectin (SNL), specific for α2,6-linked sialic acid. These results indicated that PSaV recognizes both α2,3- and α2,6-linked sialic acids for viral attachment and infection. Treatment of cells with proteases or with benzyl 4-O-β-D-galactopyranosyl-β-D-glucopyranoside (benzylGalNAc), which inhibits O-linked glycosylation, also reduced virus binding and infection, whereas inhibition of glycolipd synthesis or N-linked glycosylation had no such effect on virus binding or infection. These data suggest PSaV binds to cellular receptors that consist of α2,3- and α2,6-linked sialic acids on glycoproteins attached via O-linked glycosylation.

Comprehensive Review of Human Sapoviruses

“…Susceptibility to human sapoviruses is not associated with histo-blood group antigen (HBGA) phenotypes (214). In vitro data also support no binding of sapovirus to HBGAs (257,258). This differs from the case for the prototype norovirus (Norwalk virus): certain HBGA phenotypes (e.g., nonsecretor) of an individual are clearly related to resistance to virus infection (259,260).…”

“…Other different genogroups/genotypes of norovirus strains can also bind to HBGAs (113,(261)(262)(263) but lack a clear relatedness between the HBGA phenotypes and resistance to infection (264)(265)(266). Sialic acids have recently been reported as binding factors for porcine sapovirus (258).…”

Mechanism of Cell Culture Adaptation of an Enteric Calicivirus, the Porcine Sapovirus Cowden Strain

“…Since the P2 region of caliciviruses is the most protruding part of VP1 and is highly variable, it has been considered responsible for binding to host receptors (52). A recent study indicated that the ␣2,3-and ␣2,6-linked sialic acids on O-linked glycoproteins are receptors on LLC-PK cells for the PoSaV Cowden strain (53). In our study, based on their locations in the structural model, amino acid position 178 was located in the S domain, which may influence VP1 oligomerization, virion assembly, and stability.…”

Role of histo-blood group antigens in primate enteric calicivirus infections

“…Recent in vitro experiments with synthetic glycoconjugates with relevance as CV receptors have revealed at least three distinct patterns of virus recognition [17] . While human NoV, ReCV/TV and bovine NoV all utilize HBGAs; other CVs such as feline CV and porcine sapovirus utilize sialic acid via N-and O-linked glycoproteins, respectively; and murine NoV uses sialic acid in a strain-dependent manner [17,18] .…”