1. Site-specific glycosylation of Ebola virus glycoprotein by human polypeptide GalNAc-transferase 1 induces cell adhesion defects
- Author
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Emily J. Simon and Adam D. Linstedt
- Subjects
0301 basic medicine ,Glycosylation ,Viral protein ,Glycobiology and Extracellular Matrices ,medicine.disease_cause ,Biochemistry ,Virus ,Substrate Specificity ,03 medical and health sciences ,chemistry.chemical_compound ,Cell Adhesion ,medicine ,Humans ,Cell adhesion ,Molecular Biology ,chemistry.chemical_classification ,Binding Sites ,Ebola virus ,Chemistry ,HEK 293 cells ,Cell Biology ,Ebolavirus ,Cell biology ,HEK293 Cells ,030104 developmental biology ,Ectodomain ,N-Acetylgalactosaminyltransferases ,Glycoprotein ,Viral Fusion Proteins ,Protein Binding - Abstract
The surface glycoprotein (GP) of Ebola virus causes many of the virus's pathogenic effects, including a dramatic loss of endothelial cell adhesion associated with widespread hemorrhaging during infection. Although the GP-mediated deadhesion depends on its extracellular mucin-like domain, it is unknown whether any, or all, of this domain's densely clustered O-glycosylation sites are required. It is also unknown whether any of the 20 distinct polypeptide GalNAc-transferases (ppGalNAc-Ts) that initiate mucin-type O-glycosylation in human cells are functionally required. Here, using HEK293 cell lines lacking specific glycosylation enzymes, we demonstrate that GP requires extended O-glycans to exert its deadhesion effect. We also identified ppGalNAc-T1 as largely required for the GP-mediated adhesion defects. Despite its profound effect on GP function, the absence of ppGalNAc-T1 only modestly reduced the O-glycan mass of GP, indicating that even small changes in the bulky glycodomain can cause loss of GP function. Indeed, protein-mapping studies identified a small segment of the mucin-like domain critical for function and revealed that mutation of five glycan acceptor sites within this segment are sufficient to abrogate GP function. Together, these results argue against a mechanism of Ebola GP–induced cell detachment that depends solely on ectodomain bulkiness and identify a single host-derived glycosylation enzyme, ppGalNAc-T1, as a potential target for therapeutic intervention against Ebola virus disease.
- Published
- 2018