Fusion activity of the influenza virus hemagglutinin does not require a transbilayer pH gradient

Following reports suggesting that membrane fusion mediated by the influenza virus hemagglutinin might be dependent on a pH gradient across a putative target membrane, we have designed experiments in which this issue could be addressed directly. Accordingly, we have prepared two populations of liposomes, both simulating the plasma membrane of target cells, but with the pH of the internal aqueous medium buffered either at pH 7.4 (physiological cytosol pH) or pH 5.0 (endosomal pH at which influenza virus displays maximal fusion activity). Monitoring fusion as the relief in self-quenching of the fluorescent probe octadecylrhodamine B chloride we have found that the internal pH of the target liposomes did not influence membrane merging as mediated by the influenza virus hemagglutinin, thus demonstrating that a transmembrane pH gradient is not required for the fusion process to take place.