Functional analysis of the Bunyamwera orthobunyavirus Gc glycoprotein

Abstract

The virion glycoproteins Gn and Gc of Bunyamwera orthobunyavirus (family Bunyaviridae) are encoded by the M RNA genome segment and have roles in both viral attachment and membrane fusion. To investigate further the structure and function of the Gc protein in viral replication, we generated 12 mutants that contain truncations from the N terminus. The effects of these deletions were analysed with regard to Golgi targeting, low pH-dependent membrane fusion, infectious virus-like particle (VLP) formation and virus infectivity. Our results show that the N-terminal half (453 residues) of the Gc ectodomain (909 residues in total) is dispensable for Golgi trafficking and cell fusion. However, deletions in this region resulted in a significant reduction in VLP formation. Four mutant viruses that contained N-terminal deletions in their Gc proteins were rescued, and found to be attenuated to different degrees in BHK-21 cells. Taken together, our data indicate that the N-terminal half of the Gc ectodomain is dispensable for replication in cell culture, whereas the C-terminal half is required to mediate cell fusion. A model for the domain structure of the Gc ectodomain is proposed.

Analysis of the humoral immune response against the envelope glycoprotein Gc of Schmallenberg virus reveals a domain located at the amino terminus targeted by mAbs with neutralizing activity

“…Mutants of an orthobunyavirus that lacked the domain analogue to SBV Gc Amino still replicated in cell culture (Pollitt et al, 2006), and further analysis of the BUNV Gc function (Shi et al, 2009) corroborated these results. Besides that, recombinant BUNVs have been generated that lacked almost half of the Gc domain and tolerated the insertion of fluorescent proteins at this position (Shi et al, 2010).…”

“…Hence, the SBV Gc protein seems to mature to a certain extent without Gn. Alternatively, the amino terminus of SBV Gc could support a folding in which the fusion peptide is masked, and in its absence the protein might undergo conformational changes that lead to aggregation, as was hypothesized in a study where N-terminal residues of the BUNV Gc were removed (Shi et al, 2009). …”

“…One explanation might be the absence of Gn in these constructs, as it plays a role by translocating Gc to the Golgi complex and assists the correct folding of Gc (Shi et al, 2004(Shi et al, , 2009). However, the Gc protein expressed without Gn (Fig.…”

“…Besides that, recombinant BUNVs have been generated that lacked almost half of the Gc domain and tolerated the insertion of fluorescent proteins at this position (Shi et al, 2010). Nevertheless, certain mutations resulted in viruses with an attenuated phenotype and reduced virus-like particle formation (Shi et al, 2009).…”

Attenuation of Bunyamwera Orthobunyavirus Replication by Targeted Mutagenesis of Genomic Untranslated Regions and Creation of Viable Viruses with Minimal Genome Segments

“…ical region for viral replication (23). It was on the L segment, coding for the polymerase protein, that sequencing revealed the generation of most mutations.…”