Abstract
Sendai and influenza virions are able to fuse with mycoplasmata. Virus-Mycoplasma fusion was demonstrated by the use of fluorescently labeled intact virions and fluorescence dequenching, as well as by electron microscopy. A high degree of fusion was observed upon incubation of both virions with Mycoplasma gallisepticum or Mycoplasma capricolum. Significantly less virus-cell fusion was observed with Acholeplasma laidlawii, whose membrane contains relatively low amounts of cholesterol. The requirement of cholesterol for allowing virus-Mycoplasma fusion was also demonstrated by showing that a low degree of fusion was obtained with M. capricolum, whose cholesterol content was decreased by modifying its growth medium. Fluorescence dequenching was not observed by incubating unfusogenic virions with mycoplasmata. Sendai virions were rendered nonfusogenic by treatment with trypsin, phenylmethylsulfonyl fluoride, or dithiothreitol, whereas influenza virions were made nonfusogenic by treatment with glutaraldehyde, ammonium hydroxide, high temperatures, or incubation at low pH. Practically no fusion was observed using influenza virions bearing uncleaved hemagglutinin. Trypsinization of influenza virions bearing uncleaved hemagglutinin greatly stimulated their ability to fuse with Mycoplasma cells. Similarly to intact virus particles, also reconstituted virus envelopes, bearing the two viral glycoproteins, fused with M. capricolum. However, membrane vesicles, bearing only the viral binding (HN) or fusion (F) glycoproteins, failed to fuse with mycoplasmata. Fusion between animal enveloped virions and prokaryotic cells was thus demonstrated.
Highlights
From the Departmentof Biological Chemisto, Institute of Life Sciences, The Hebrew University of Jerusalem, 91904 Jerusalem, Israel, the $Department of Membrane and Ultrastructure Research, The Hebrew University-Hadassah Medical School. 91010 Jerusalem
The requiremenot f cholesterol for allowing virus-Mycoplasma fusion was demonstrated by showing thaat low degree of fusion was obtained with M. capricolum, whose cholesterol content was decreased by modifying its growth medium
Sendai virions were renderendonfusogenic by treatment with trypsin, phenylmethylsulfonyl fluoride, or dithiothreitol, whereas influenza virions were made nonfusogenic by treatment with glutaraldehyde, ammonium hydroxide, high temperatureso,r incubation at low pH
Summary
Citovsky, ShlomoRottemS, Ofer Nussbaum, Yehudit Laster, Rudolf RottQ, and Abraham Loyter. Similar observations have been made following the interaction between influenza virions and phospholipid vesicles (Maeda et al, 1981; Doms et al, 1985) Based on these observations, it is tempting to speculate that viral envelopes are able to fuse with any biological membrane whose lipid molecules are exposed to the viral glycoproteins, without the virions bearing uncleaved hemagglutinin. After 15 min of incubation at room temperature in the dark, the virions were washed in 20 volumes of Solution Na (100,000X g for 30 min a t 4 "C) and resuspended in the same buffer to give a protein concentration of 0.2-0.3 mg/ml Under such conditions, Rls was inserted into the viral membranes at selfquenching surface density (about 3 mol % of total viral phospholipids), and its decrease was shown to be proportional to the fluorescence dequenching (Hoekstra et aL, 1984; Citovsky et al, 1985). Protein Determination-Protein was determined by the method of Lowry et al (1951)with bovine serum albumin as a standard
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