Cholesterol dependence of varicella-zoster virion entry into target cells.
The entry of inhaled virions into airway cells is presumably the initiating step of varicella-zoster infection. In order to characterize viral entry, we studied the relative roles played by lipid rafts and clathrin-mediated transport. Virus and target cells were pretreated with agents designed to perturb selected aspects of endocytosis and membrane composition, and the effects of these perturbations on infectious focus formation were monitored. Infectivity was exquisitely sensitive to methyl-beta-cyclodextrin (M beta CD) and nystatin, which disrupt lipid rafts by removing cholesterol. These agents inhibited infection by enveloped, but not cell-associated, varicella-zoster virus (VZV) in a dose-dependent manner and exerted these effects on both target cell and viral membranes. Inhibition by M beta CD, which could be reversed by cholesterol replenishment, rapidly declined as a function of time after exposure of target cells to VZV, suggesting that an early step in viral infection requires cholesterol. No effect of cholesterol depletion, however, was seen on viral binding; moreover, there was no reduction in the surface expression or internalization of mannose 6-phosphate receptors, which are required for VZV entry. Viral entry was energy dependent and showed concentration-dependent inhibition by chlorpromazine, which, among other actions, blocks clathrin-mediated endocytosis. These data suggest that both membrane lipid composition and clathrin-mediated transport are critical for VZV entry. Lipid rafts are likely to contribute directly to viral envelope integrity and, in the host membrane, may influence endocytosis, evoke downstream signaling, and/or facilitate membrane fusion.
Hambleton
S
S
Department of Paediatric, College of Physicians and Surgeons, Columbia University, New York, NY, USA. shambleton@doctors.org.uk
Steinberg
S P
SP
Gershon
M D
MD
Gershon
A A
AA
D002478:Cells, Cultured; D002784:Cholesterol / Q000378:metabolism*; D004705:Endocytosis / Q000187:drug effects; D006493:Heparin / Q000494:pharmacology; D014645:Herpesvirus 3, Human / Q000502:physiology* / Q000648:ultrastructure; D006801:Humans; D008360:Mannosephosphates / Q000494:pharmacology; D008561:Membrane Fusion; D008854:Microscopy, Electron; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors; D014771:Virion / Q000502:physiology* / Q000648:ultrastructure
D008360:Mannosephosphates; C027693:mannose-6-phosphate; D006493:Heparin; D002784:Cholesterol; D019869:Phosphatidylinositol 3-Kinases; D011505:Protein-Tyrosine Kinases