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

Abstract

We have monitored kinetics of fusion between cell pairs consisting of a single influenza hemaglutinin (HA)-expressing cell and a single erythrocyte (RBC) that had been labeled with both a fluorescent lipid (Dil) 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 (Δψ) that was monitored by a decrease in Dil fluorescence. This event was followed by two distinct stages of fusion pore dilation: the flux of fluorescent lipid (φ(l)) and the flux of a large aqueous fluorescent dye (φ(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 pre (FP(s)). Our data are consistent with a fusion pore comprising six HA trimers.