title: Acidic residues in the membrane-proximal stalk region of vaccinia virus protein B5 are required for glycosaminoglycan-mediated disruption of the extracellular enveloped virus outer membrane.
abstract: The extracellular enveloped virus (EEV) form of vaccinia virus (VACV) is surrounded by two lipid envelopes. This presents a topological problem for virus entry into cells, because a classical fusion event would only release a virion surrounded by a single envelope into the cell. Recently, we described a mechanism in which the EEV outer membrane is disrupted following interaction with glycosaminoglycans (GAGs) on the cell surface and thus allowing fusion of the inner membrane with the plasma membrane and penetration of a naked core into the cytosol. Here we show that both the B5 and A34 viral glycoproteins are required for this process. A34 is required to recruit B5 into the EEV membrane and B5 acts as a molecular switch to control EEV membrane rupture upon exposure to GAGs. Analysis of VACV strains expressing mutated B5 proteins demonstrated that the acidic stalk region between the transmembrane anchor sequence and the fourth short consensus repeat of B5 are critical for GAG-induced membrane rupture. Furthermore, the interaction between B5 and A34 can be disrupted by the addition of polyanions (GAGs) and polycations, but only the former induce membrane rupture. Based on these data we propose a revised model for EEV entry.
authors.affiliation: Department of Virology, Faculty of Medicine, Imperial College London, St Mary's Campus, Norfolk Place, London W2 1PG, UK.
authors.forename: Kim L
authors.initials: KL
authors.lastname: Roberts
authors.affiliation: Department of Virology, Faculty of Medicine, Imperial College London, St Mary's Campus, Norfolk Place, London W2 1PG, UK.
authors.forename: Adrien
authors.initials: A
authors.lastname: Breiman
authors.affiliation: Department of Virology, Faculty of Medicine, Imperial College London, St Mary's Campus, Norfolk Place, London W2 1PG, UK.
authors.forename: Gemma C
authors.initials: GC
authors.lastname: Carter
authors.affiliation: Department of Virology, Faculty of Medicine, Imperial College London, St Mary's Campus, Norfolk Place, London W2 1PG, UK.
authors.forename: Helen A
authors.initials: HA
authors.lastname: Ewles
authors.affiliation: Department of Virology, Faculty of Medicine, Imperial College London, St Mary's Campus, Norfolk Place, London W2 1PG, UK.
authors.forename: Michael
authors.initials: M
authors.lastname: Hollinshead
authors.affiliation: Department of Virology, Faculty of Medicine, Imperial College London, St Mary's Campus, Norfolk Place, London W2 1PG, UK.
authors.forename: Mansun
authors.initials: M
authors.lastname: Law
authors.affiliation: Department of Virology, Faculty of Medicine, Imperial College London, St Mary's Campus, Norfolk Place, London W2 1PG, UK.
authors.forename: Geoffrey L
authors.initials: GL
authors.lastname: Smith
mesh_terms: D000818:Animals; D002460:Cell Line; D006025:Glycosaminoglycans / Q000378:metabolism*; D011073:Polyamines / Q000378:metabolism; D000071228:Polyelectrolytes; D011108:Polymers / Q000378:metabolism; D054730:Protein Interaction Domains and Motifs; D014616:Vaccinia virus / Q000502:physiology*; D014763:Viral Matrix Proteins / Q000235:genetics / Q000378:metabolism*; D053586:Virus Internalization
chemical_list: D006025:Glycosaminoglycans; D011073:Polyamines; D000071228:Polyelectrolytes; D011108:Polymers; D014763:Viral Matrix Proteins; C534336:B5 protein, vaccinia virus