title: Visualizing hepatitis B virus with biarsenical labelling in living cells.
abstract: BACKGROUND
  Study on viruses has greatly benefited from visualization of viruses tagged with green fluorescent protein (GFP) in living cells. But GFP tag, as a large inserted fragment, is not suitable for labelling Hepatitis B virus (HBV) that is a compact virion with limited internal space.
  
  
  OBJECTIVE
  To visualize HBV in living cells, we constructed several recombinant HBV fluorescently labelled with biarsenical dye to track the behaviour of HBV in the cytoplasm of infected cells.
  
  
  METHODS
  By mutagenesis, a smaller size tetracysteine (TC) tag (C-C-P-G-C-C) that could be bound with a biarsenical fluorescent dye was genetically inserted at different cell epitopes of HBV core protein expressed in transfected cells.
  
  
  RESULTS
  Confocal microscopy and transmission electron microscopy (TEM) observations showed that TC-tagged core proteins bound with biarsenical dye could specifically fluoresce in cells and be incorporated into nucleocapsid to form fluorescent virions. The recombinant fluorescent HBV virions retained their infectivity as wild-type ones. Moreover, tracking of fluorescent HBV particles in living cells reveals microtubule-dependent motility of the intracellular particles.
  
  
  CONCLUSIONS
  To the best of our knowledge, this is the first time to generate fluorescent HBV virions with biarsenical labelling and to visualize their trafficking in living cells. The fluorescent HBV may become one highly valuable tool for further studying detailed dynamic processes of HBV life cycle and interaction of HBV with host in live-imaging approach.
authors.affiliation: Institute of Liver Diseases, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China; Department of Gastroenterology, the First Affiliated Hospital, Zhengzhou University, Zhengzhou, China.
authors.forename: Shuzhen
authors.initials: S
authors.lastname: Sun
authors.forename: Jingjun
authors.initials: J
authors.lastname: Yan
authors.forename: Chao
authors.initials: C
authors.lastname: Xia
authors.forename: Yuanyuan
authors.initials: Y
authors.lastname: Lin
authors.forename: Xiaorui
authors.initials: X
authors.lastname: Jiang
authors.forename: Haojing
authors.initials: H
authors.lastname: Liu
authors.forename: Huanping
authors.initials: H
authors.lastname: Ren
authors.forename: Junwei
authors.initials: J
authors.lastname: Yan
authors.forename: Jusheng
authors.initials: J
authors.lastname: Lin
authors.forename: Xingxing
authors.initials: X
authors.lastname: He
mesh_terms: D001152:Arsenicals / Q000378:metabolism; D015153:Blotting, Western; D003545:Cysteine / Q000235:genetics / Q000378:metabolism; D005456:Fluorescent Dyes / Q000378:metabolism; D056945:Hep G2 Cells; D006515:Hepatitis B virus / Q000302:isolation & purification / Q000648:ultrastructure*; D006801:Humans; D018613:Microscopy, Confocal; D046529:Microscopy, Electron, Transmission; D010957:Plasmids / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D013194:Staining and Labeling / Q000379:methods*; D014771:Virion / Q000235:genetics / Q000378:metabolism / Q000648:ultrastructure*
chemical_list: D001152:Arsenicals; D005456:Fluorescent Dyes; D003545:Cysteine
keywords: biarsenical dye; fluorescence; hepatitis B virus; tetracysteine; tubulin