Influenza hemagglutinin-mediated membrane fusion does not involve inverted phase lipid intermediates

Intermediate lipid structures such as inverted micelles and interlamellar attachments, which can form near liquid crystalline lamellar (L alpha) to inverted hexagonal (HII) phase boundaries, are thought to play a role in membrane fusion. To investigate whether these structures are also involved in influenza hemagglutinin-mediated membrane fusion, measurement of fusion under conditions where such structures could not form was attempted. It was found that the fusion of influenza virus with liposomal membranes containing phosphatidylcholine and gangliosides, which cannot form HII phases, was only slightly slower than fusion with liposomes that also contained the HII competent phosphatidylethanolamine. Furthermore, the virus fused efficiently with liposomes consisting either of pure saturated phosphatidylcholines or phosphatidylcholine/ganglioside mixtures, even when the liposomal membranes were in the gel (L beta') phase and thus far from L alpha/HII transitions. Isolated hemagglutinin, reconstituted into dimyristoylphosphatidylcholine membranes, induced fusion with liposomes composed of dimyristoylphosphatidylcholine and gangliosides at temperatures below the L beta' to L alpha phase transition temperature of dimyristoylphosphatidylcholine. This latter finding excluded the possibility that the viral lipids alone could have formed inverted phase intermediates, thus enabling them to fuse with liposomes that do not contain lipids capable of forming inverted phases. Therefore, it is concluded that structures resembling intermediates in L alpha/HII transitions are most likely not involved in influenza hemagglutinin-mediated fusion.