Comprehensive Functional Analysis of N-Linked Glycans on Ebola Virus GP1

Lennemann, Nicholas J.; Rhein, Bethany A.; Ndungo, Esther; Chandran, Kartik; Qiu, Xiangguo; Maury, Wendy

A role for CLECs in EBOV GP-dependent entry has been firmly established by others (32, 33, 38, 39) . We have shown for the first time that the removal of N-glycans from either the glycan cap (7G) or MLD (GPm8G) dramatically decreases the utilization of DC-SIGN, L-SIGN, and LSECtin. Surprisingly, these same CLECs enhanced the transduction mediated by GP1⌬muc, despite this form of the protein sharing exactly the same pattern of NGS as GPm8G. Another surprising finding was that hMGL utilization was largely unaffected by the loss of GP1 N-glycans. This provided indirect evidence that hMGL-dependent EBOV transduction is strongly dependent on O-linked glycans in GP. However, despite the apparent utilization of O-linked glycans by hMGL, the transduction mediated by GP1⌬muc was greatly enhanced by the expression of this CLEC (32) . In combination, these two sets of results suggest that deletion of the MLD alters the species of glycans present on GP1. Such a conclusion is consistent with previous work showing that glycosylation differences exist between GP and the small, soluble form of GP (sGP), which shares the same NGS with GP1⌬muc (40). More glycan processing was found on sGP, indicated by the higher percentage of galactose residues and lower percentage of high-mannose glycans. Since the types of glycans present on proteins are largely dependent on the environment in which the protein is produced (41) , it is possible that the cytotoxicity associated with overexpression of GP (42, 43) or the large number of O-glycans present changes the glycosylation machinery in the ER/Golgi.