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Multiple Local Contact Sites are Induced by GPI‐Linked Influenza Hemagglutinin During Hemifusion and Flickering Pore Formation
Author(s) -
Frolov Vadim A.,
Cho MyoungSoon,
Bronk Peter,
Reese Thomas S.,
Zimmerberg Joshua
Publication year - 2000
Publication title -
traffic
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.677
H-Index - 130
eISSN - 1600-0854
pISSN - 1398-9219
DOI - 10.1034/j.1600-0854.2000.010806.x
Subject(s) - ectodomain , lipid bilayer fusion , biophysics , membrane , fusion , hemagglutinin (influenza) , lipid bilayer , transmembrane protein , electron microscope , cell membrane , biology , chemistry , biochemistry , optics , linguistics , philosophy , receptor , physics , gene
Membrane fusion intermediates induced by the glycosylphosphatidylinositol‐linked ectodomain of influenza hemagglutinin (GPI‐HA) were investigated by rapid freeze, freeze‐substitution, thin section electron microscopy, and with simultaneous recordings of whole‐cell admittance and fluorescence. Upon triggering, the previously separated membranes developed numerous hourglass shaped points of membrane contact (∼10–130 nm waist) when viewed by electron microscopy. Stereo pairs showed close membrane contact at peaks of complementary protrusions, arising from each membrane. With HA, there were fewer contacts, but wide fusion pores. Physiological measurements showed fast lipid dye mixing between cells after acidification, and either fusion pore formation or the lack thereof (true hemifusion). For the earliest pores, a similar conductance distribution and frequency of flickering pores were detected for both HA and GPI‐HA. For GPI‐HA, lipid mixing was detected prior to, during, or after pore opening, whereas for HA, lipid mixing is seen only after pore opening. Our findings are consistent with a pathway wherein conformational changes in the ectodomain of HA pull membranes towards each other to form a contact site, then hemifusion and pore formation initiate in a small percentage of these contact sites. Finally, the transmembrane domain of HA is needed to complete membrane fusion for macromolecular content mixing.