Membrane rupture as an alternative membrane assembly pathway
Final Report Abstract
The aim of the application was to understand the molecular mechanism of membrane assembly of large DNA viruses of the nucleo-cytoplasmic large DNA virus (NCLDV) family. Rather than budding these viruses acquire their membrane by the rupture of cellular membranes; the open intermediates are then used to build the viral membrane. Within the frame of this proposal we could show, using advanced electron microscopy techniques, that all members of the NCLDVs we studied use this same unconventional mechanism suggesting they share a common molecular machinery. For one of its members, vaccinia virus (VACV), we subsequently identified a single viral protein required for membrane rupture and thus for assembly. We have started to clone this protein in order to express and purify it and analyse its structure by cryo-EM. Within the frame of this proposal we also showed that the membrane of purified virions is enriched in certain lipids and speculate that these could also be involved in the unusual membrane biogenesis. We have started a new series of experiments, funded by a joint ANR/DFG grant to test this hypothesis by expressing the VACV protein in liposomes of defined lipid composition.
Publications
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(2017) Vaccinia virus A11 is required for membrane rupture and viral membrane assembly. Cellular microbiology 19 (10)
Suarez, Cristina; Hoppe, Simone; Pénard, Esthel; Walther, Paul; Krijnse-Locker, Jacomine
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(2013). Open membranes are the precursors for assembly of large DNA viruses. Cell Microbiol 15, 1883-1895
Suarez, C., Welsch, S., Chlanda, P., Hagen, W., Hoppe, S., Kolovou, A., Pagnier, I., Raoult, D., and Krijnse Locker, J.
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(2013). Poxvirus membrane biogenesis: rupture not disruption. Cell Microbiol 15, 190-199
Krijnse Locker, J., Chlanda, P., Sachsenheimer, T., and Brugger, B.
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(2015). African swine fever virus assembles a single membrane derived from rupture of the endoplasmic reticulum. Cell Microbiol 17, 1683-1698
Suarez, C., Andres, G., Kolovou, A., Hoppe, S., Salas, M.L., Walther, P., and Krijnse Locker, J.
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(2017). The sleeping beauty kissed awake: new methods in electron microscopy to study cellular membranes. Biochem J 474, 1041-1053
Chlanda, P., and Krijnse Locker, J.
