title: The role of lipid rafts in the early stage of Enterovirus 71 infection.
abstract: OBJECTIVE
  Although it has been widely accepted that Enterovirus 71 (EV71) enters permissive cells via receptor-mediated endocytosis, the details of entry mechanism for EV71 still need more exploration. This study aimed to investigate the role of lipid rafts in the early stage of EV71 Infection.
  
  
  METHODS
  The effect of cholesterol depletion or addition of exogenous cholesterol was detected by immunofluorescence assays and quantitative real-time PCR. Effects of cholesterol depletion on the association of EV71 with lipid rafts were determined by flow cytometry and co-immunoprecipitation assays. Localization and internalization of EV71 and its receptor were assayed by confocal microscpoy and sucrose gradient analysis. The impact of cholesterol on the activation of phosphoinositide 3'-kinase/Akt signaling pathway during initial virus infection was analyzed by Western-blotting.
  
  
  RESULTS
  Disruption of membrane cholesterol by a pharmacological agent resulted in a significant reduction in the infectivity of EV71. The inhibitory effect could be reversed by the addition of exogenous cholesterol. Cholesterol depletion post-infection did not affect EV71 infection. While virus bound equally to cholesterol-depleted cells, EV71 particles failed to be internalized by cholesterol-depleted cells. EV71 capsid protein co-localized with cholera toxin B, a lipid-raft-dependent internalization marker.
  
  
  CONCLUSIONS
  Lipid rafts play a critical role in virus endocytosis and in the activation of PI3K/Akt signaling pathway in the early stage of EV71 infection.
authors.affiliation: Department of Microbiology, Shanghai Key Laboratory of Medical Biodefense, Second Military Medical University, Shanghai, China.
authors.forename: Yong-Zhe
authors.initials: YZ
authors.lastname: Zhu
authors.forename: Da-Ge
authors.initials: DG
authors.lastname: Wu
authors.forename: Hao
authors.initials: H
authors.lastname: Ren
authors.forename: Qing-Qiang
authors.initials: QQ
authors.lastname: Xu
authors.forename: Kui-Cheng
authors.initials: KC
authors.lastname: Zheng
authors.forename: Wei
authors.initials: W
authors.lastname: Chen
authors.forename: Sheng-Lin
authors.initials: SL
authors.lastname: Chen
authors.forename: Xi-Jing
authors.initials: XJ
authors.lastname: Qian
authors.forename: Qing-Yuan
authors.initials: QY
authors.lastname: Tao
authors.forename: Yan
authors.initials: Y
authors.lastname: Wang
authors.forename: Ping
authors.initials: P
authors.lastname: Zhao
authors.forename: Zhong-Tian
authors.initials: ZT
authors.lastname: Qi
mesh_terms: D015153:Blotting, Western; D036022:Capsid Proteins / Q000378:metabolism; D045744:Cell Line, Tumor; D002648:Child; D002772:Cholera Toxin / Q000378:metabolism; D002784:Cholesterol / Q000378:metabolism; D004705:Endocytosis / Q000187:drug effects; D029821:Enterovirus A, Human / Q000302:isolation & purification / Q000378:metabolism / Q000472:pathogenicity*; D006801:Humans; D047468:Immunoprecipitation; D008297:Male; D021962:Membrane Microdomains / Q000187:drug effects / Q000378:metabolism*; D019869:Phosphatidylinositol 3-Kinases / Q000235:genetics / Q000378:metabolism; D000081082:Phosphoinositide-3 Kinase Inhibitors; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D034622:RNA Interference; D060888:Real-Time Polymerase Chain Reaction; D015398:Signal Transduction / Q000187:drug effects; D053586:Virus Internalization / Q000187:drug effects; D047392:beta-Cyclodextrins / Q000494:pharmacology
chemical_list: D036022:Capsid Proteins; D002772:Cholera Toxin; D002784:Cholesterol; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D051057:Proto-Oncogene Proteins c-akt; D047392:beta-Cyclodextrins; C108732:methyl-beta-cyclodextrin