Rapid membrane fusion of individual virus particles with supported lipid bilayers.
Many enveloped viruses employ low-pH-triggered membrane fusion during cell penetration. Solution-based in vitro assays in which viruses fuse with liposomes have provided much of our current biochemical understanding of low-pH-triggered viral membrane fusion. Here, we extend this in vitro approach by introducing a fluorescence assay using single particle tracking to observe lipid mixing between individual virus particles (influenza or Sindbis) and supported lipid bilayers. Our single-particle experiments reproduce many of the observations of the solution assays. The single-particle approach naturally separates the processes of membrane binding and membrane fusion and therefore allows measurement of details that are not available in the bulk assays. We find that the dynamics of lipid mixing during individual Sindbis fusion events is faster than 30 ms. Although neither virus binds membranes at neutral pH, under acidic conditions, the delay between membrane binding and lipid mixing is less than half a second for nearly all virus-membrane combinations. The delay between binding and lipid mixing lengthened only for Sindbis virus at the lowest pH in a cholesterol-dependent manner, highlighting the complex interaction between lipids, virus proteins, and buffer conditions in membrane fusion.
Wessels
Laura
L
Department of Physics, North Carolina State University, Raleigh, North Carolina 27695, USA.
Elting
Mary Williard
MW
Scimeca
Dominic
D
Weninger
Keith
K
D000818:Animals; D055592:Biophysical Phenomena; D001703:Biophysics; D002642:Chick Embryo; D006224:Cricetinae; D005456:Fluorescent Dyes; D006863:Hydrogen-Ion Concentration; D066298:In Vitro Techniques; D053122:Influenza A Virus, H3N2 Subtype / Q000502:physiology; D008051:Lipid Bilayers / Q000737:chemistry*; D008561:Membrane Fusion / Q000502:physiology*; D008954:Models, Biological; D012235:Rhodamines; D012845:Sindbis Virus / Q000502:physiology; D014771:Virion / Q000502:physiology*
D005456:Fluorescent Dyes; D008051:Lipid Bilayers; D012235:Rhodamines; C044542:octadecyl Rhodamine B chloride