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Review
. 2018 Jun 8;430(12):1685-1695.
doi: 10.1016/j.jmb.2018.04.015. Epub 2018 Apr 18.

Monomeric Intermediates Formed by Vesiculovirus Glycoprotein during Its Low-pH-induced Structural Transition

Abbas Abou-Hamdan  1 , Laura Belot  1 , Aurélie Albertini  1 , Yves Gaudin  2
Affiliations
Review

Monomeric Intermediates Formed by Vesiculovirus Glycoprotein during Its Low-pH-induced Structural Transition

Abbas Abou-Hamdan et al. J Mol Biol. .

Abstract
in English, French

Unlabelled Image

Vesiculoviruses enter cells by membrane fusion, driven by a large, low-pH-induced, conformational change in the fusion glycoprotein (G) that involves transition from a trimeric pre-fusion to a trimeric post-fusion state. G is the model of class III fusion glycoproteins which also includes the fusion glycoproteins of herpesviruses (gB) and baculoviruses (gp64). Class III fusion proteins combine features of the previously characterized class I and class II fusion proteins. In this review, we first present and discuss the data that indicate that the Vesiculovirus G structural transition proceeds through monomeric intermediates. Then, we focus on a recently determined crystal structure of the Chandipura virus G ectodomain that contained two monomeric intermediate conformations of the glycoprotein, revealing the chronological order of the structural changes in the protein and offering a detailed pathway for the conformational change, in agreement with electron microscopy data. In the crystal, the intermediates were associated through their fusion domain in an antiparallel manner to form an intermolecular β-sheet. Mutagenesis indicated that this interface is functionally relevant. All those structural data challenge the current model proposed for viral membrane fusion. Therefore, we wonder if this mode of operating is specific to Vesiculovirus G and discuss data indicating that class II fusion glycoproteins are monomeric when they interact with the target membrane but also crystal structures suggesting the existence of non-trimeric intermediates for influenza hemagglutinin which is the prototype of class I fusion proteins.

Keywords: Influenza; Vesiculovirus; conformational change; glycoprotein; membrane fusion.

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Figures

Unlabelled Image
Graphical abstract
Fig. 1
Fig. 1
Structure of vesiculovirus glycoprotein G. (a) Overall structures of the trimeric pre- and post-fusion forms of VSV glycoprotein. Left part: Ribbon diagram of VSV Gth pre-fusion trimer (PDB code: 5I2S). Right part: Ribbon diagram of VSV Gth post-fusion trimer (PDB code: 5I2M). (b) From left to right: Overall structures of VSV Gth pre-fusion protomer, CHAV Gth EI (PDB code: 5MDM), CHAV Gth LI (PDB code: 5MDM) and CHAV Gth post-fusion protomer (PDB code: 4D6W). The major difference between the pre-fusion protomer and the EI is the movement of R5 (arrowhead 1). Then, the refolding of the hinge regions R2 and R3 between PHD and FD (arrowhead 2) together with the partial elongation of the central helix implying R4 allows for the transition to the LI state. At this stage, G is already in a hairpin conformation. The formation of the last amino-terminal turn of the central helix (arrowhead 3) and the refolding of R5 into the lateral helix (arrowhead 4) concomitantly with the final trimerization form the six-helix bundle organization and achieve the structural transition. All the structures are aligned on their TrD. The histidines of the cluster H60, H162 and H407 in the pre-fusion protomer are depicted in green sticks. A close-up view of the cluster is also represented. Color code: TrD is in red, PHD is in orange, FD is in yellow, segments R1 and R4 (indicated by thin gray arrows) are in blue, segments R2 and R3 are in green, and segment R5 and C-terminal segment (cter) are in pink. The residues in the fusion loops at the tip of the FD are in red sticks. The C-terminal segments which are linked to the transmembrane domain are depicted in dashed lines. The position of the membrane relative to the protein is indicated by a black line.
Fig. 2
Fig. 2
Non-trimeric association of monomeric intermediate states of G. (a and b) Two views of the tetrameric assembly in the CHAV Gth crystal (PDB code: 5MDM) rotated by 90°. Protomers in the EI conformation are in blue. Protomers in the LI conformation are in green. The residues in the fusion loops at the tip of the FD are in red sticks. In panel B, the C-terminal segments which are linked to the transmembrane domain are depicted in dashed lines. (c) Close-up view of the antiparallel β-sheet formed between EI (in blue) and LI (in green) FDs in the crystal asymetric unit. Key residues which cluster at the interface are represented in gray sticks. The residues in the fusion loops at the tip of the FD are in red sticks. (d) Localization of residues K76, H80, Q112, D121, E123 in the FD in the pre-fusion protomer (left part) and trimeric post-fusion (right part) conformations of VSV G.
Fig. 3
Fig. 3
A model for VSV membrane fusion. (a) At pH 7.5 in the absence of a target membrane, at the viral surface, there is an equilibrium between G pre-fusion trimer and flexible monomers , mostly in an EI-like conformation having different orientations, thanks to the flexibility of the R5 segment . (b) Lowering the pH below 7 progressively shifts the equilibrium toward more elongated monomers in an LI-like conformation , . Some post-fusion trimers are already present on the lateral side of the virus. The flat base is a favorable site for the association of monomeric intermediates into flat antiparallel dimers, which insert their fusion loops in the target membrane. (c) Formation of the fusion pore. The number of oligomers involved at this stage and their exact organization around the fusion pore is not known. For convenience, on the scheme, the fusion pore has been placed at the center of the flat base. However, note that the membrane is strongly constrained at the edge of the base, a feature that might favor pore formation at this location. (d) The formation of the helical network of spikes in their post-fusion conformation on the lateral side of the virion drives the enlargement of the initial fusion pore leading to complete membrane merger. What happens to the dimers that were involved in the initiation of the fusion process is unknown. The simplest explanation is that the dimeric association is only transient and that the dimer dissociates upon protomers refolding to LI and final retrimerization.
Fig. 4
Fig. 4
Monomeric HA structures. (a) Structure of HA pre-fusion state (H5 subtype, PDB code 2IBX[78]). Left: Ribbon diagram of HA pre-fusion trimer. The HA1 subunits are colored in a shade of yellow. The HA2 subunits are colored in a shade of blue. Right: Ribbon diagram of HA protomer. (b) First row: ribbon diagrams of HA2 structure (H1 subtype) as in the pre-fusion protomer and in two monomeric HA structures , . Second row: ribbon diagrams of HA2 structure (H3 subtype) as in the pre-fusion protomer, in two monomeric HA0 structures and as in its trimeric post-fusion conformation . Third row: ribbon diagrams of HA2 structure (H5 subtype) as in the pre-fusion protomer and in a monomeric HA structures . The residues are colored by their secondary structure in the pre-fusion protomer. Residues corresponding to helix A are in yellow, to helix B in blue and to the loop located between helices A and B in red. In several monomers, the part of the chain corresponding to the loop between helices A and B is not visible in the crystal. All the structures are aligned on helix A of the prefusion protomer. The corresponding PDB codes are indicated above each structure.
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