Abstract
BackgroundLipid rafts are major structural components in plasma membranes that play critical roles in many biological processes including virus infection. However, few reports have described the relationship between lipid rafts and porcine rotavirus (PRV) infection. In this study, we investigated whether or not the locally high concentrations (3–5 fold) of cholesterol present in lipid rafts are required for PRV infection, and further examined which stages of the infection process are most affected.ResultsWhen cellular cholesterol was depleted by methyl-β-cyclodextrin (MβCD), PRV infectivity significantly declined in a dose-dependent manner. This inhibition was partially reversed upon reintroduction of cholesterol into the system. This was corroborated by the co-localization of PRV with a recombinant, GPI-anchored green fluorescent protein, which functioned as a marker for membranous regions high in cholesterol and indicative of lipid rafts. Changes in virus titer and western blot analyses indicated that depletion of cellular cholesterol with MβCD had no apparent effect on PRV adsorption; however, depletion of cholesterol significantly restricted entry and post-entry of PRV into the cell. Both points of inhibition were restored to near normal levels by the addition of exogenous cholesterol.ConclusionsWe conclude from these studies that membrane-based cholesterol and in particular that localized to lipid rafts, is an indispensable biomolecule for PRV infection, and that cholesterol-based control of the infection process takes place during entry and immediately post-entry into the cell.
Highlights
Lipid rafts are major structural components in plasma membranes that play critical roles in many biological processes including virus infection
Effects of cholesterol depletion on Porcine rotavirus (PRV) infection To determine whether the cholesterol component of lipid rafts plays a role in PRV infection, the cholesterol level in cellular membranes of MA104 cells for PRV and BHK-21 cells for Vesicular stomatitis virus (VSV), respectively, were depleted by treatment with MβCD before infecting with PRV or VSV
After 24 h, VP7 polyclonal antibody for PRV binding exhibited a dose-dependent decrease in MA104 cells pretreated with increasing concentrations of MβCD (Fig. 1a)
Summary
Lipid rafts are major structural components in plasma membranes that play critical roles in many biological processes including virus infection. Few reports have described the relationship between lipid rafts and porcine rotavirus (PRV) infection. PRV, which belongs to the genus Rotavirus in the family Reoviridae, is a non-enveloped virus formed by three concentric layers of proteins [1]. It remains unclear as to how non-enveloped viruses which lack an outer lipid membrane infect host cells. Studies with simian virus 40 (SV40) [5] and poliovirus [6] have suggested that non-enveloped viruses are transported to specific domains within the plasma membrane using a vesicular transport mechanism, followed by replication in large inclusions in
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