N-linked glycosylation and reticuloendotheliosis retrovirus envelope glycoprotein function

Different properties of the spleen necrosis virus (SNV) envelope glycoprotein were analyzed following biosynthesis in the presence of glycosylation inhibitors. Tunicamycin, which inhibits all asparagine N-linked glycosylation, prevented intracellular processing and translocation to the cell surface of the envelope protein. In contrast, castanospermine or deoxymannojirimycin, which block glycosidase trimming of the early high-mannose chains and subsequent complex type N-glycosylation, did not inhibit proteolytic cleavage or cellular translocation. The ability of unglycosylated and partially glycosylated envelope protein to bind the viral receptor was assayed using an infection interference assay. Tunicamycin abrogated SNV envelope glycoprotein-induced receptor interference, whereas the trimming glycosidase inhibitors had no effect on interference. Similarly, tunicamycin but not the glycosidase inhibitors reduced the titers of released virus 100-fold. We conclude that carbohydrate trimming and complex N-glycosylation are not essential for envelope glycoprotein translocation, proteolytic cleavage, receptor binding, or infectivity, whereas cotranslational high-mannose N-glycosylation is essential for all of the SNV envelope glycoprotein properties tested. Syncytia formation can be induced following transfection into D17 cells of an envelope glycoprotein expression plasmid. Unlike virus particle infectivity, cell fusion is strongly inhibited by the glycosidase inhibitors.