title: GPI-anchored influenza hemagglutinin induces hemifusion to both red blood cell and planar bilayer membranes.
abstract: Under fusogenic conditions, fluorescent dye redistributed from the outer monolayer leaflet of red blood cells (RBCs) to cells expressing glycophosphatidylinositol-anchored influenza virus hemagglutinin (GPI-HA) without transfer of aqueous dye. This suggests that hemifusion, but not full fusion, occurred (Kemble, G. W., T. Danieli, and J. M. White. 1994. Cell. 76:383-391). We extended the evidence for hemifusion by labeling the inner monolayer leaflets of RBCs with FM4-64 and observing that these inner leaflets did not become continuous with GPI-HA-expressing cells. The region of hemifusion-separated aqueous contents, the hemifusion diaphragm, appeared to be extended and was long-lived. But when RBCs hemifused to GPI-HA-expressing cells were osmotically swollen, some diaphragms were disrupted, and spread of both inner leaflet and aqueous dyes was observed. This was characteristic of full fusion: inner leaflet and aqueous probes spread to cells expressing wild-type HA (wt-HA). By simultaneous video fluorescence microscopy and time-resolved electrical admittance measurements, we rigorously demonstrated that GPI-HA-expressing cells hemifuse to planar bilayer membranes: lipid continuity was established without formation of fusion pores. The hemifusion area became large. In contrast, for cells expressing wt-HA, before lipid dye spread, fusion pores were always observed, establishing that full fusion occurred. We present an elastic coupling model in which the ectodomain of wt-HA induces hemifusion and the transmembrane domain, absent in the GPI-HA-expressing cells, mediates full fusion.
authors.affiliation: Department of Molecular Biophysics and Physiology, Rush Medical College, Chicago, Illinois 60612, USA.
authors.forename: G B
authors.initials: GB
authors.lastname: Melikyan
authors.forename: J M
authors.initials: JM
authors.lastname: White
authors.forename: F S
authors.initials: FS
authors.lastname: Cohen
mesh_terms: D000818:Animals; D016466:CHO Cells / Q000502:physiology; D002459:Cell Fusion / Q000502:physiology; D002462:Cell Membrane / Q000502:physiology*; D004396:Coloring Agents / Q000493:pharmacokinetics; D006224:Cricetinae; D004594:Electrophysiology; D004912:Erythrocytes / Q000502:physiology*; D004996:Ethidium; D017261:Glycosylphosphatidylinositols / Q000502:physiology*; D006389:Hemagglutinins, Viral / Q000502:physiology*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008051:Lipid Bilayers; D050356:Lipid Metabolism; D009975:Orthomyxoviridae / Q000737:chemistry; D009995:Osmosis / Q000502:physiology; D018272:Porins / Q000378:metabolism; D014760:Viral Fusion Proteins / Q000502:physiology
chemical_list: D004396:Coloring Agents; D017261:Glycosylphosphatidylinositols; D006389:Hemagglutinins, Viral; D008051:Lipid Bilayers; D018272:Porins; D014760:Viral Fusion Proteins; D004996:Ethidium