Rapid membrane fusion of individual virus particles with supported lipid bilayers
Laura Wessels 1, Mary Williard Elting 1, Dominic Scimeca 1 and Keith Weninger 1*
1 North Carolina State University
* To whom correspondence should be addressed. E-mail: keith_weninger{at}ncsu.edu.
Submitted on September 14, 2006
Revised on November 6, 2006
Accepted on 8 March 2007
 |
Abstract |
|---|
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 milliseconds. 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.
Key Words:
Sindbis virus, cholesterol, influenza, lipid mixing, single molecule, single particle tracking
