The p14 fusion-associated small transmembrane (FAST) protein effects membrane fusion from a subset of membrane microdomains.
The reovirus fusion-associated small transmembrane (FAST) proteins are a unique family of viral membrane fusion proteins. These nonstructural viral proteins induce efficient cell-cell rather than virus-cell membrane fusion. We analyzed the lipid environment in which the reptilian reovirus p14 FAST protein resides to determine the influence of the cell membrane on the fusion activity of the FAST proteins. Topographical mapping of the surface of fusogenic p14-containing liposomes by atomic force microscopy under aqueous conditions revealed that p14 resides almost exclusively in thickened membrane microdomains. In transfected cells, p14 was found in both Lubrol WX- and Triton X-100-resistant membrane complexes. Cholesterol depletion of donor cell membranes led to preferential disruption of p14 association with Lubrol WX (but not Triton X-100)-resistant membranes and decreased cell-cell fusion activity, both of which were reversed upon subsequent cholesterol repletion. Furthermore, co-patching analysis by fluorescence microscopy indicated that p14 did not co-localize with classical lipid-anchored raft markers. These data suggest that the p14 FAST protein associates with heterogeneous membrane microdomains, a distinct subset of which is defined by cholesterol-dependent Lubrol WX resistance and which may be more relevant to the membrane fusion process.
Corcoran
Jennifer A
JA
Departmentnof Microbiology, Dalhousie University, Halifax, Nova Scotia B3H 1X5, Canada.
Salsman
Jayme
J
de Antueno
Roberto
R
Touhami
Ahmed
A
Jericho
Manfred H
MH
Clancy
Eileen K
EK
Duncan
Roy
R
D000818:Animals; D002462:Cell Membrane / Q000378:metabolism; D002784:Cholesterol / Q000737:chemistry / Q000378:metabolism; D003902:Detergents / Q000494:pharmacology; D005347:Fibroblasts / Q000378:metabolism; D008055:Lipids / Q000737:chemistry; D021962:Membrane Microdomains / Q000378:metabolism; D017830:Octoxynol / Q000494:pharmacology; D030102:Orthoreovirus, Mammalian / Q000378:metabolism*; D010957:Plasmids / Q000378:metabolism; D011092:Polyethylene Glycols / Q000494:pharmacology; D017434:Protein Structure, Tertiary; D011784:Quail; D014162:Transfection; D014760:Viral Fusion Proteins / Q000737:chemistry / Q000502:physiology*
D003902:Detergents; D008055:Lipids; D014760:Viral Fusion Proteins; C017474:Lubrol WX; D011092:Polyethylene Glycols; D017830:Octoxynol; D002784:Cholesterol