Mechanism of membrane fusion induced by vesicular stomatitis virus G protein

Abstract

The glycoproteins (G proteins) of vesicular stomatitis virus (VSV) and related rhabdoviruses (e.g., rabies virus) mediate both cell attachment and membrane fusion. The reversibility of their fusogenic conformational transitions differentiates them from many other low-pH-induced viral fusion proteins. We report single-virion fusion experiments, using methods developed in previous publications to probe fusion of influenza and West Nile viruses. We show that a three-stage model fits VSV single-particle fusion kinetics: (i) reversible, pH-dependent, G-protein conformational change from the known prefusion conformation to an extended, monomeric intermediate; (ii) reversible trimerization and clustering of the G-protein fusion loops, leading to an extended intermediate that inserts the fusion loops into the target-cell membrane; and (iii) folding back of a cluster of extended trimers into their postfusion conformations, bringing together the viral and cellular membranes. From simulations of the kinetic data, we conclude that the critical number of G-protein trimers required to overcome membrane resistance is 3 to 5, within a contact zone between the virus and the target membrane of 30 to 50 trimers. This sequence of conformational events is similar to those shown to describe fusion by influenza virus hemagglutinin (a "class I" fusogen) and West Nile virus envelope protein ("class II"). Our study of VSV now extends this description to "class III" viral fusion proteins, showing that reversibility of the low-pHinduced transition and architectural differences in the fusion proteins themselves do not change the basic mechanism by which they catalyze membrane fusion. membrane fusion protein | virus entry | enveloped virus

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

“…The question, which then arises, is whether the working of the vesiculovirus machinery is different from that of other viruses. This is all the more pertinent because (i) all the experimental data suggest that the membrane fusion pathway (i.e., the lipidic intermediates which are formed) is very similar for all the enveloped viruses studied so far regardless of the organization of their fusion machinery [52][53][54], and (ii) fusion kinetics of individual virions for influenza virus (class I), West Nile virus (class II), and VSV could all be fit to models based on very similar sequence of conformational events [13,55,56]. Therefore, in this last part, we will discuss data obtained on other viral fusion glycoproteins from both class I and class II and focus on some features reminiscent of what is observed for vesiculovirus G.…”

“…Fusion is catalyzed by a low-pHinduced large structural transition from a pre-to a post-fusion conformation which are both trimeric [10]. Remarkably, for rhabdoviruses, the structural transition is reversible [11][12][13][14], and in fact, there is an equilibrium between different states of G, which is shifted toward the trimeric post-fusion conformation at low pH [5,15].…”

Single Virion Tracking Microscopy for the Study of Virus Entry Processes in Live Cells and Biomimetic Platforms

“…For West Nile virus, hemifusion has one or two rate-limiting steps, depending on the pH [138]. Vesicular stomatitis virus hemifusion also has multiple rate-limiting steps at higher pH and a single rate-limiting step at low pH [139].…”

Illuminating pathogen–host intimacy through optogenetics

“…Not least, selected geneencoded sensors that can facilitate optogenetic work in virology include GECIs (gene-encoded calcium indicators), cNMP biosensors, as well as fluorescent indicators for pH, lipids, and several other metabolites (Table 1). Among many, one assay would be to test the pH dependence of membrane fusion during the entry of influenza and stomatitis viruses, mediated by hemagglutinin and G glycoprotein, respectively [68,69]. Another conceivable application is to visualize lipid rafts as a predicted platform for the entry, assembly, and release of viral particles [70][71][72].…”