GLYCOSYLATION WITHIN AN ANTIGENIC SITE ON THE HN GLYCOPROTEIN OF NEWCASTLE-DISEASE VIRUS INTERFERES WITH ITS ROLE IN THE PROMOTION OF MEMBRANE-FUSION

The binding of monoclonal antibodies to antigenic site 3 on the hemagg lutinin-neuraminidase (HN) glycoprotein of Newcastle disease virus neu tralizes viral infectivity and prevents syncytium formation by a mecha nism other than the prevention of viral attachment. The virus can esca pe neutralization by these antibodies by the addition of an N-glycan a t a site introduced by a D287N mutation in HN. The variant has signifi cantly reduced ability to induce fusion from within, the mode of fusio n promoted by the viral glycoproteins deposited on the cell surface la te in infection. Conversely, and unlike the parent virus, the variant has acquired the ability to promote fusion from without, the mode of f usion directly mediated by input virions at high multiplicity. This fi nding is consistent with different roles for the HN protein in virion- cell and cell-cell fusion. D287N-mutated HN with its additional N-glyc an shows a markedly reduced ability, compared to wild-type HN, to comp lement the viral fusion protein in the promotion of fusion in a BHK ce ll transient expression system. This confirms that the addition of an N-glycan in HN antigenic site 3 and the deficiency in syncytium format ion are causally related. Moreover, no alteration in cell surface expr ession, hemadsorption, or neuraminidase activity was detected in the m utated protein. This monoclonal antibody-selected mutation suggests th at a fusion-related function, secondary to receptor recognition, may b e defined by the globular head of the HN spike: However, D287C or D287 S-mutated HN is as effective as the wild-type protein in the promotion of fusion in the coexpression system. This suggests that the diminish ed fusogenicity of the D287N-mutated protein is probably due to a more global effect of glycosylation in site 3 rather than an alteration at the amino acid level. (C) 1994 Academic Press, Inc.