Cholesterol enriched lipid rafts are key determinants for entry and post-entry control of porcine rotavirus infection

Authors: DOU, XIUJING - Northeast Agricultural University; LI, XUNLIANG - Northeast Agricultural University; LI, YANG - Northeast Agricultural University; Zarlenga, Dante; REN, XIAOFENG - Northeast Agricultural University; DONG, NA - Northeast Agricultural University; LI, GUANGXING - Northeast Agricultural University

Submitted to: Virology Journal
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 12/1/2018
Publication Date: 12/1/2018
Citation: Dou, X., Li, X., Li, Y., Zarlenga, D.S., Ren, X., Dong, N., Li, G. 2018. Cholesterol enriched lipid rafts are key determinants for entry and post-entry control of porcine rotavirus infection. Virology Journal. 14(1):45. doi: 10.1186/s12917-018-1366-7
DOI: https://doi.org/10.1186/s12917-018-1366-7

Interpretive Summary: Porcine rotavirus (PRV) is the most important cause of severe diarrhea in newborn piglets, and results in significant economic losses to pig industry. To date, efficient drugs for preventing PRV infection have not been identified. It remains unclear as to how non-enveloped viruses which lack an outer lipid membrane infect host cells, but it is generally believed that membrane fusion is not involved in virus entry. In recent years, research has exposed a crucial role for lipid rafts (microdomains within plasma membranes that are high in cholesterol) in virus function and potentially in cell entry. In this work, we explored the importance of cellular cholesterol during PRV infection by demonstrating that viral protein and mRNA levels were reduced significantly following drug treatment to deplete cellular cholesterol. We further showed that viral protein and mRNA levels were restored by subsequently treating the cells with exogenous cholesterol. Our data co-localized the PRV predominantly to lipid rafts. Inasmuch as cholesterol was indispensable for both PRV entry and the post-entry process, lipid rafts could be a target for development of anti-PRV strategies. This research can be used for general research and by the pharmaceutical community to help develop alternative methods to treat PRV.

Technical Abstract: Lipid 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. When cellular cholesterol was depleted by Methyl-ß-cyclodextrin (MßCD), 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 MßCD 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. We conclude from these studies that membrane-based cholesterol and in particular that localized to lipid drafts, 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.