Hemagglutinin-neuraminidase enhances F protein-mediated membrane fusion of reconstituted Sendai virus envelopes with cells.
Reconstituted Sendai virus envelopes containing both the fusion (F) protein and the hemagglutinin-neuraminidase (HN) (F,HN-virosomes) or only the F protein (F-virosomes) were prepared by solubilization of the intact virus with Triton X-100 followed by its removal by using SM2 Bio-Beads. Viral envelopes containing HN whose disulfide bonds were irreversibly reduced (HNred) were also prepared by treating the envelopes with dithiothreitol followed by dialysis (F,HNred-virosomes). Both F-virosomes and F,HNred-virosomes induced hemolysis of erythrocytes in the presence of wheat germ agglutinin, but the rates and extents were markedly lower than those for hemolysis induced by F,HN-virosomes. Using an assay based on the relief of self-quenching of a lipid probe incorporated in the Sendai virus envelopes, we demonstrate the fusion of both F,HN-virosomes and F-virosomes with cultured HepG2 cells containing the asialoglycoprotein receptor, which binds to a terminal galactose moiety of F. By desialylating the HepG2 cells, the entry mediated by HN-terminal sialic acid receptor interactions was bypassed. We show that both F-virosomes and F,HN-virosomes fuse with desialylated HepG2 cells, although the rate was two- to threefold higher if HN was included in the viral envelope. We also observed enhancement of fusion rates when both F and HN envelope proteins were attached to their specific receptors.
Department of Biochemistry, University of Delhi, New Delhi, India.
S
S
Bagai
A
A
Puri
R
R
Blumenthal
D P
DP
Sarkar
D000818:Animals; D037263:Asialoglycoprotein Receptor; D002642:Chick Embryo; D004220:Disulfides / Q000378:metabolism; D006461:Hemolysis; D006801:Humans; D008561:Membrane Fusion; D015335:Molecular Probes; D019158:N-Acetylneuraminic Acid; D010084:Oxidation-Reduction; D010222:Parainfluenza Virus 1, Human / Q000378:metabolism*; D011971:Receptors, Immunologic; D012794:Sialic Acids; D013050:Spectrometry, Fluorescence; D014407:Tumor Cells, Cultured; D014759:Viral Envelope Proteins / Q000378:metabolism*; D014760:Viral Fusion Proteins / Q000378:metabolism*; D014909:Wheat Germ Agglutinins
D037263:Asialoglycoprotein Receptor; D004220:Disulfides; D015335:Molecular Probes; D011971:Receptors, Immunologic; D012794:Sialic Acids; D014759:Viral Envelope Proteins; D014760:Viral Fusion Proteins; D014909:Wheat Germ Agglutinins; D019158:N-Acetylneuraminic Acid