Dilation of the influenza hemagglutinin fusion pore revealed by the kinetics of individual cell-cell fusion events

We have monitored kinetics of fusion between cell pairs consisting of a single influenza hemagglutinin (HA)-expressing cell and a single erythrocyte (RBC) that had been labeled with both a fluorescent lipid (DiI) in the membrane and a fluorescent solute (calcein) in the aqueous space. Initial fusion pore opening between the RBC and HA-expressing cell produced a change in RBC membrane potential (Delta psi) that was monitored by a decrease in DiI fluorescence. This event was followed by two distinct stages of fusion pore dilation: the flux of fluorescent lipid (phi(L)) and the flux of a large aqueous fluorescent dye (phi(S)). We have analyzed the kinetics of events that occur as a result of transitions between a fusion pore (FP) and a solute permissive fusion pore (FPS). Our data are consistent with a fusion pore comprising six HA trimers.