Cholesterol depletion affects infectivity and stability of pseudorabies virus.

Statins, such as lovastatin, inhibit the cellular cholesterol biosynthesis. Addition of lovastatin to SK cells and subsequent infection with the alphaherpesvirus pseudorabies virus (PRV) did not affect the intracellular production of viral structural proteins but reduced virus titers. Addition of methyl-beta-cyclodextrin to deplete cholesterol from the viral envelope also resulted in reduced virus titers. Addition of exogenous cholesterol restored virus titers in both experimental assays. Further analysis showed that reducing cholesterol levels reduced both the infectivity of newly produced infectious virus and their stability, as assessed by determining virus titers immediately after treatment or upon storage of the virus at room temperature or frozen. This is the first report to demonstrate that cholesterol is involved in the stability of infectious alphaherpesvirus, and that treatment of host cells with statins reduces alphaherpesvirus titers. Hence, cholesterol is important for pseudorabies virus infectivity and stability.

Plasma membrane cholesterol is required for efficient pseudorabies virus entry.

Alphaherpesviruses comprise closely related viruses of man and animal, including herpes simplex virus, varicella-zoster virus and pseudorabies virus (PRV). Here, using methyl-beta-cyclodextrin and fluorescently tagged PRV, we directly show that depletion of cholesterol from the plasma membrane of host cells significantly reduces PRV entry. Cholesterol depletion did not reduce PRV attachment, but stalled virus particles at the plasma membrane before penetration of the cell. Cholesterol depletion results in destabilization of lipid raft microdomains in the plasma membrane, which have been shown before to be involved in efficient entry of different viruses. A significant fraction of PRV virions appears to localize juxtaposed to GM1, a lipid raft marker, during entry. Together, these data indicate that cholesterol and possibly cholesterol-rich lipid rafts may be important during PRV entry.

Tyrosine phosphorylation and lipid raft association of pseudorabies virus glycoprotein E during antibody-mediated capping.

In specific cell types infected with the alphaherpesviruses herpes simplex virus and pseudorabies virus (PRV), addition of virus-specific antibodies results in redistribution of cell-surface-anchored viral proteins. This redistribution is triggered by the viral protein gE and consists of the directional movement of the antibody-antigen complexes to one pole of the cell. This viral capping process has been associated with increased antibody-resistant virus spread and strongly resembles immunoreceptor capping, a process that is crucial in activation of different immune cells (e.g. capping of Fcgamma-receptors, B and T cell receptors). Here, we report that the PRV gE-mediated viral capping process results in increased Src kinase-mediated tyrosine phosphorylation of the cytoplasmic domain of gE and that a fraction of gE associates with lipid rafts, all very reminiscent of immunoreceptor capping. These results provide evidence that gE-mediated capping is a viral mimicry of immunoreceptor capping.