title: Disruption of clathrin-dependent trafficking results in the failure of grass carp reovirus cellular entry.
abstract: BACKGROUND
  Grass carp reovirus (GCRV) is responsible for viral hemorrhagic disease in cultured grass carp (Ctenopharyngon idellus). GCRV is a non-enveloped, double-stranded RNA virus in the genus Aquareovirus, of the family Reoviridae, which encodes seven structural proteins (VP1-VP7) and five nonstructural proteins (NS80, NS38, NS31, NS26, and NS16). To date, the mechanism of GCRV entry into CIK Ctenopharyngon idellus kidney (CIK) cells remains poorly understood.
  
  
  RESULTS
  Here, we present a study of the GCRV internalization mechanism in CIK cells. Our results indicated that GCRV infection was inhibited by chlorpromazine, the specific inhibitor for clathrin-mediated endocytosis. Colocalization of GCRV virions with endogenous clathrin was observed during early infection by confocal microscopy. Moreover, GCRV infection of CIK cells depended on acidification of the endosome. This was indicated by significant inhibition of viral infection following prophylactic treatment with the lysosomotropic drugs chloroquine or ammonium chloride. In addition, the disturbance of dynamin activity blocked GCRV entry, which confirmed the dynamin-dependent nature of clathrin-mediated endocytosis.
  
  
  CONCLUSIONS
  Our findings suggest that GCRV might enter CIK cells via clathrin-mediated endocytosis in a pH-dependent manner. Additionally, dynamin is critical for efficient viral entry.
authors.affiliation: National Pathogen Collection Center for Aquatic Animals, Shanghai Ocean university, Shanghai, 201306, China. haowang1000@163.com.|Key Laboratory of Aquatic Genetic Resources of Ministry of Aquaculture, Shanghai Ocean University, Shanghai, 201306, China. haowang1000@163.com.
authors.forename: Hao
authors.initials: H
authors.lastname: Wang
authors.affiliation: National Pathogen Collection Center for Aquatic Animals, Shanghai Ocean university, Shanghai, 201306, China. 373455917@qq.com.
authors.forename: Weisha
authors.initials: W
authors.lastname: Liu
authors.affiliation: National Pathogen Collection Center for Aquatic Animals, Shanghai Ocean university, Shanghai, 201306, China. yufei1011126@yeah.net.
authors.forename: Fei
authors.initials: F
authors.lastname: Yu
authors.affiliation: National Pathogen Collection Center for Aquatic Animals, Shanghai Ocean university, Shanghai, 201306, China. lqlv@shou.edu.cn.|Key Laboratory of Aquatic Genetic Resources of Ministry of Aquaculture, Shanghai Ocean University, Shanghai, 201306, China. lqlv@shou.edu.cn.
authors.forename: Liqun
authors.initials: L
authors.lastname: Lu
mesh_terms: D000818:Animals; D000998:Antiviral Agents / Q000494:pharmacology; D002460:Cell Line; D002966:Clathrin / Q000378:metabolism*; D034281:Dynamins / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005372:Filipin / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D021381:Protein Transport; D012087:Reoviridae / Q000502:physiology* / Q000648:ultrastructure; D012088:Reoviridae Infections / Q000378:metabolism / Q000821:virology; D053586:Virus Internalization
chemical_list: D000998:Antiviral Agents; D002966:Clathrin; D004791:Enzyme Inhibitors; D005372:Filipin; D034281:Dynamins