Patch clamp studies of single cell-fusion events mediated by a viral fusion protein.
To enter cells, viruses must fuse their envelope with a host cell membrane. Fusion is mediated by specific, membrane-spanning fusion proteins, of which the influenza virus haemagglutinins (HA) are the best characterized. Several HAs have been sequenced, and the crystal structure of the major part of one HA is known. The conditions for fusion and some of the rearrangements in the HA that accompany fusion are well understood, but it remains unclear how HA causes bilayers to fuse. We have observed, in real time, unitary cell-fusion events caused by HA. Fibroblasts expressing HA were induced to fuse with red blood cells by a rapid drop in pH. Fusion was monitored by fluorescence microscopy, and by measuring the membrane conductance and capacitance of the fibroblast. The earliest event observed was the sudden opening of an aqueous pore connecting the cytoplasms of the fusing cells. Initially, the pore conductance often fluctuated between zero and approximately 600 pS, as if the pore were opening and closing repeatedly. Later, it increased over tens of seconds, as if the pore dilated. We suggest that, as in exocytosis, HA-mediated membrane fusion begins with the formation of a narrow pore. Based on the conductance, we estimate the initial diameter of the pore to be no more than twice that of a gap junction channel.
Spruce
A E
AE
Department of Physiology and Biophysics, University of Washington, Seattle 98195.
Iwata
A
A
White
J M
JM
Almers
W
W
D000818:Animals; D002462:Cell Membrane / Q000502:physiology*; D004553:Electric Conductivity; D004594:Electrophysiology; D006389:Hemagglutinins, Viral / Q000502:physiology*; D006863:Hydrogen-Ion Concentration; D066298:In Vitro Techniques; D008561:Membrane Fusion; D008564:Membrane Potentials; D051379:Mice; D009975:Orthomyxoviridae; D014760:Viral Fusion Proteins / Q000502:physiology*
D006389:Hemagglutinins, Viral; D014760:Viral Fusion Proteins