Potassium is a trigger for conformational change in the fusion spike of an enveloped RNA virus

Abstract

Many enveloped viruses enter cells through the endocytic network, from which they must subsequently escape through fusion of viral and endosomal membranes. This membrane fusion is mediated by virus-encoded spikes that respond to the dynamic endosomal environment, which triggers conformational changes in the spikes that initiate the fusion process. Several fusion triggers have been identified and include pH, membrane composition, and endosome-resident proteins, and these cues dictate when and where viral fusion occurs. We recently reported that infection with an enveloped bunyavirus requires elevated potassium ion concentrations [K+], controlled by cellular K+ channels, that are encountered during viral transit through maturing endosomes. Here we reveal the molecular basis for the K+ requirement of bunyaviruses through the first direct visualization of a member of the Nairoviridae family, namely Hazara virus (HAZV), using cryo-EM. Using cryo-electron tomography, we observed HAZV spike glycoproteins within infectious HAZV particles exposed to both high and low [K+], which showed that exposure to K+ alone results in dramatic changes to the ultrastructural architecture of the virion surface. In low [K+], the spikes adopted a compact conformation arranged in locally ordered arrays, whereas, following exposure to high [K+], the spikes became extended, and spike–membrane interactions were observed. Viruses exposed to high [K+] also displayed enhanced infectivity, thus identifying K+ as a newly defined trigger that helps promote viral infection. Finally, we confirmed that K+ channel blockers are inhibitory to HAZV infection, highlighting the potential of K+ channels as anti-bunyavirus targets.

Novel ionophores active against La Crosse virus identified through rapid antiviral screening

“…We thus hypothesized that valinomycin’s antiviral activity was due to its effect on the cell. The role of potassium in bunyavirus infection has been well-documented, and bunyavirus entry is potassium dependent 21,22 . To test if valinomycin was affecting the cell rather than the virus, we treated Huh7 cells with 2 µM valinomycin and, immediately before infection, we washed away the drug.…”

Structures of enveloped virions determined by cryogenic electron microscopy and tomography

“…These patches have been proposed to create membrane curvature and contribute to the budding of hantaviruses (Huiskonen et al, 2010). Interestingly, nairovirus HAZV GP has also been observed to arrange in ordered tetrameric patches (Punch et al, 2018). Analogously, locally ordered patches of GP trimers have been observed also in BUNV, an orthobunyavirus, suggesting that such local order may be a generic driver of budding in pleomorphic bunyavirus virions (Bowden et al, 2013).…”

“…Nairoviruses are another example of a pleomorphic bunyavirus displaying an ordered lattice of glycoproteins on their envelope. The first structural insights into this lattice came from STA of HAZV, revealing a tetrameric array of spikes on the virion envelope (Punch et al, 2018).…”

“…Other factors such as the concentration of potassium ions (K + ) has also been shown to be crucial for fusion events. An STA study of HAZV GP has demonstrated that the Gc fusion protein can be triggered by high K + concentration into an extended conformation and subsequently embedded into the target membrane of co-purified vesicles (Punch et al, 2018).…”

“…(C) A virion with a nonicosahedrally symmetric, but locally ordered outer protein shell covering most of the lipid bilayer and lacking a matrix layer. Example structures include members of order Bunyavirales such as Tula virus (TULV) (Hantaviridae) (Huiskonen et al, 2010), Hazara virus (HAZV) (Nairoviridae) (Punch et al, 2018) and Bunyamwera virus (BUNV) (Orthobunyaviridae) (Bowden et al, 2013). (D) A virion with an icosahedral inner capsid and a nonicosahedrally symmetric, but locally ordered outer protein shell.…”