Activation of phospholipase activity during Semliki Forest virus infection

Infection of animal cells by a number of cytolytic viruses leads to increased membrane permeability. Thus, Semliki Forest virus (SFV) infection of susceptible cells modifies the permeability of the membrane for a number of cations and metabolites (Muñoz et al. (1985), Virology 146, 203-212). The molecular basis of this modification of the cell membrane has not been investigated in detail. We report that during the infection of HeLa cells with SFV, or BHK cells with vesicular stomatitis virus, there is a significant increase in the release of choline and arachidonic acid into the culture medium, suggesting that both phospholipases (PLases) C and A2 become activated during infection. Both choline and phosphorylcholine are released into the medium as expected when PLase C is activated. Cells prelabeled with arachidonic acid release a significant amount of radioactivity from the third hour postinfection. Most of this radioactivity is present in the medium of SFV-infected cells in the form of free fatty acid, suggesting that phospholipid hydrolysis has occurred; no intact phospholipids are detected in the culture medium. Finally, the action of several inhibitors of PLases, such as zinc and cadmium ions, chloroquine, chlorpromazine, amantadine, and dansylcadaverine were assayed. Our findings indicate that the release of choline or arachidonic acid is potently blocked by some of these lipase inhibitors. Following infection by SFV HeLa cells become susceptible to the inhibition of protein synthesis by hygromycin B due to increased uptake of this antibiotic. Entry of hygromycin B was prevented by zinc ions or chloroquine, suggesting that the increase in membrane permeability in SFV-infected cells may be mediated in part by lipase activation.