title: Role of N-linked glycans on bunyamwera virus glycoproteins in intracellular trafficking, protein folding, and virus infectivity.
abstract: The membrane glycoproteins (Gn and Gc) of Bunyamwera virus (BUN, family Bunyaviridae) contain three potential sites for the attachment of N-linked glycans: one site (N60) on Gn and two (N624 and N1169) on Gc. We determined that all three sites are glycosylated. Digestion of the glycoproteins with endo-beta-N-acetylglucosaminidase H (endo H) or peptide:N-glycosidase F revealed that Gn and Gc differ significantly in their glycan status and that late in infection Gc glycans remain endo H sensitive. The roles of the N-glycans in intracellular trafficking of the glycoproteins to the Golgi, protein folding, and virus replication were investigated by mutational analysis and confocal immunofluorescence. Elimination of the glycan on Gn, by changing N60 to a Q residue, resulted in the protein misfolding and failure of both Gn and Gc proteins to traffic to the Golgi complex. We were unable to rescue a viable virus by reverse genetics from a cDNA containing the N60Q mutation. In contrast, mutant Gc proteins lacking glycans on either N624 or N1169, or both sites, were able to target to the Golgi. Gc proteins containing mutations N624Q and N1169Q acquired endo H resistance. Three viable N glycosylation-site-deficient viruses, lacking glycans on one site or both sites on Gc, were created by reverse genetics. The viability of these recombinant viruses and analysis of growth kinetics indicates that the glycans on Gc are not essential for BUN replication, but they do contribute to the efficiency of virus infection.
authors.affiliation: Division of Virology, Institute of Virology, University of Glasgow, Church St., Glasgow G11 5JR, Scotland, United Kingdom.
authors.forename: Xiaohong
authors.initials: X
authors.lastname: Shi
authors.forename: Kristina
authors.initials: K
authors.lastname: Brauburger
authors.forename: Richard M
authors.initials: RM
authors.lastname: Elliott
mesh_terms: D000818:Animals; D002042:Bunyamwera virus / Q000254:growth & development* / Q000472:pathogenicity; D002044:Bunyaviridae Infections / Q000821:virology*; D002460:Cell Line; D003593:Cytoplasm / Q000378:metabolism; D006023:Glycoproteins / Q000378:metabolism; D006031:Glycosylation; D006801:Humans; D009154:Mutation; D011134:Polysaccharides / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D017510:Protein Folding; D021381:Protein Transport; D014764:Viral Proteins / Q000378:metabolism
chemical_list: D006023:Glycoproteins; D011134:Polysaccharides; D014764:Viral Proteins