Permeability changes resulting from virus-cell fusion: temperature-dependence of the contributing processes.

Abstract

A new assay for membrane fusion, using the fluorescent probe pyrene-sulphonyl-phosphatidyl ethanolamine, has been developed. Fusion between the envelope of Sendai virus and human erythrocytes or Lettre cells has a Q10 of approximately 4 at 37 degrees C, increasing to approximately 7 at 7 degrees C; there is no lag to onset of fusion. Viral neuraminidase has a Q10 of 2.3 between 37 degrees C and 4 degrees C. Its action limits the extent of fusion by causing the elution of virus; this effect is particularly marked at low temperature because of the difference in Q10 for fusion and neuraminidase. The temperature-dependence of the initiation of permeability changes following the removal of inhibitory amounts of Ca2+ is approximately 2; thus membrane fusion is the principal temperature-sensitive step during the permeabilization of cells by Sendai virus. A recovery process, by which cells become insensitive to the removal of Ca2+ and which therefore limits the extent of permeabilization, has a Q10 of 7.4 between 37 degrees C and 21 degrees C. It is concluded that the lag to onset of permeability changes is not due to a lag in virus-cell membrane fusion, but to the gradual acquisition of a threshold level of membrane damage; the extent of permeabilization depends on the rate of fusion relative to the rates of neuraminidase and recovery.