Hemadsorption and fusion inhibition activities of hemagglutinin analyzed by vaccinia virus mutants.
Vaccinia virus IHD-J strain induces hemagglutinin (HA) on the surface membrane of infected cells and does not elicit cell-cell fusion (F-). We isolated 21 independent hemadsorption-negative (HAD-) mutant viruses from IHD-J and five HAD+ revertants from one of these mutants. Of the 21 mutants, 19 that synthesized either no or little HA at the cell surface caused cell-cell fusion (F+), whereas none of the five revertants that synthesized HA at the cell surface induced cell-cell fusion. Furthermore, anti-HA monoclonal antibody B2D10 induced extensive polykaryocytosis of IHD-J-infected cells and suppressed the ability of the IHD-J-infected cell extract to inhibit the polykaryocytosis induced by IHD-W. The other 2 of the 21 HAD- mutants, B1 and A2, which induced HAs at the cell surface, showed F- and F+ phenotype, respectively. The HA molecule of mutant B1 had a single amino acid substitution of Lys for Glu-121 in its extracellular domain, whereas that of mutant A2 had a single substitution mutation of Tyr for Cys-103. We conclude that the vaccinia HA is a fusion inhibition protein, that the active sites for the two activities reside separately in its extracellular domain, and that cysteine-103 is important in forming the proper tertiary structure of the protein to exert both activities.
Seki
M
M
Institute for Virus Research, Kyoto University, Japan.
Oie
M
M
Ichihashi
Y
Y
Shida
H
H
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D002459:Cell Fusion; D005455:Fluorescent Antibody Technique; D006367:HeLa Cells; D006382:Hemadsorption; D006389:Hemagglutinins, Viral / Q000096:biosynthesis / Q000235:genetics / Q000502:physiology*; D006801:Humans; D015151:Immunoblotting; D009154:Mutation; D010641:Phenotype; D011233:Precipitin Tests; D014616:Vaccinia virus / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D014709:Vero Cells
D000911:Antibodies, Monoclonal; D006389:Hemagglutinins, Viral