Acidic pH enhancement of the fusion of Newcastle disease virus with cultured cells.
Fusion of the lentogenic strain "Clone 30" of Newcastle disease virus (NDV) with the cell line COS-7 has been studied. Fusion was monitored using the octadecylrhodamine B chloride dequenching assay [Hoekstra, D., de Boer, T., Klappe, K. and Wilschut, J. (1984). Biochemistry 23, 5675-5681]. In the present work, fusion of NDV with COS-7 cells was found to occur in a time- and temperature-dependent fashion. Significant dequenching of the probe occurred at temperatures higher than 28 degrees C. A 20-fold excess of unlabeled virus inhibited fusion by about 53% compared with the control, whereas 62% inhibition of fusion was obtained after digestion of viral glycoproteins with trypsin. The data are discussed in terms of the nonfusion transfer of the probe. In addition, preincubation of cells with 50 mM ammonium chloride or 0.1% sodium azide prevented NDV from fusing with COS-7 cells by about 30% in comparison with the control. The cytopathic effect of NDV infection in cell culture in the presence of ammonium chloride was reduced compared with control. Moreover, viral preincubation at pH 5 yielded a mild inhibition of fusogenic activity. Our results suggest that NDV may use the endocytic pathway as a complementary way of entering cells by direct fusion with the plasma membrane.
Departamento de Bioquímica y Biología Molecular, Universidad de Salamanca, Edificio Departamental Lab. 109, Salamanca, 37007, Spain.
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San Román
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Villar
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Muñoz-Barroso
D000643:Ammonium Chloride / Q000494:pharmacology; D000818:Animals; D001667:Binding, Competitive; D019556:COS Cells; D002522:Chlorocebus aethiops; D003588:Cytopathogenic Effect, Viral / Q000187:drug effects; D004705:Endocytosis / Q000187:drug effects; D005456:Fluorescent Dyes / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008561:Membrane Fusion / Q000187:drug effects; D009522:Newcastle disease virus / Q000187:drug effects / Q000378:metabolism / Q000502:physiology*; D010446:Peptide Fragments / Q000378:metabolism / Q000494:pharmacology; D012235:Rhodamines / Q000378:metabolism; D019810:Sodium Azide / Q000494:pharmacology; D013696:Temperature; D014357:Trypsin / Q000378:metabolism; D014361:Trypsin Inhibitors / Q000494:pharmacology; D014709:Vero Cells; D014760:Viral Fusion Proteins / Q000378:metabolism
D005456:Fluorescent Dyes; D010446:Peptide Fragments; D012235:Rhodamines; D014361:Trypsin Inhibitors; D014760:Viral Fusion Proteins; D000643:Ammonium Chloride; D019810:Sodium Azide; D014357:Trypsin; C044542:octadecyl Rhodamine B chloride