title: Simultaneous Visualization of Parental and Progeny Viruses by a Capsid-Specific HaloTag Labeling Strategy.
abstract: Real-time, long-term, single-particle tracking (SPT) provides us an opportunity to explore the fate of individual viruses toward understanding the mechanisms underlying virus infection, which in turn could lead to the development of therapeutics against viral diseases. However, the research focusing on the virus assembly and egress by SPT remains a challenge because established labeling strategies could neither specifically label progeny viruses nor make them distinguishable from the parental viruses. Herein, we have established a temporally controllable capsid-specific HaloTag labeling strategy based on reverse genetic technology. VP26, the smallest pseudorabies virus (PrV) capsid protein, was fused with HaloTag protein and labeled with the HaloTag ligand during virus replication. The labeled replication-competent recombinant PrV harvested from medium can be applied directly in SPT experiments without further modification. Thus, virus infectivity, which is critical for the visualization and analysis of viral motion, is retained to the largest extent. Moreover, progeny viruses can be distinguished from parental viruses using diverse HaloTag ligands. Consequently, the entire course of virus infection and replication can be visualized continuously, including virus attachment and capsid entry, transportation of capsids to the nucleus along microtubules, docking of capsids on the nucleus, endonuclear assembly of progeny capsids, and the egress of progeny viruses. In combination with SPT, the established strategy represents a versatile means to reveal the mechanisms and dynamic global picture of the life cycle of a virus.
authors.affiliation: Key Laboratory of Analytical Chemistry for Biology and Medicine (Ministry of Education), College of Chemistry and Molecular Sciences, State Key Laboratory of Virology, The Institute for Advanced Studies, and Wuhan Institute of Biotechnology, Wuhan University , Wuhan 430072, P.R. China.
authors.forename: An-An
authors.initials: AA
authors.lastname: Liu
authors.affiliation: State Key Laboratory of Virology, Wuhan Institute of Virology, Chinese Academy of Sciences , Wuhan 430071, P.R. China.
authors.forename: Zhenfeng
authors.initials: Z
authors.lastname: Zhang
authors.affiliation: Key Laboratory of Analytical Chemistry for Biology and Medicine (Ministry of Education), College of Chemistry and Molecular Sciences, State Key Laboratory of Virology, The Institute for Advanced Studies, and Wuhan Institute of Biotechnology, Wuhan University , Wuhan 430072, P.R. China.
authors.forename: En-Ze
authors.initials: EZ
authors.lastname: Sun
authors.affiliation: State Key Laboratory of Virology, Wuhan Institute of Virology, Chinese Academy of Sciences , Wuhan 430071, P.R. China.
authors.forename: Zhenhua
authors.initials: Z
authors.lastname: Zheng
authors.affiliation: Key Laboratory of Analytical Chemistry for Biology and Medicine (Ministry of Education), College of Chemistry and Molecular Sciences, State Key Laboratory of Virology, The Institute for Advanced Studies, and Wuhan Institute of Biotechnology, Wuhan University , Wuhan 430072, P.R. China.
authors.forename: Zhi-Ling
authors.initials: ZL
authors.lastname: Zhang
authors.affiliation: State Key Laboratory of Virology, Wuhan Institute of Virology, Chinese Academy of Sciences , Wuhan 430071, P.R. China.
authors.forename: Qinxue
authors.initials: Q
authors.lastname: Hu
authors.affiliation: Key Laboratory of Special Pathogens and Biosafety, Center for Emerging Infectious Diseases, Wuhan Institute of Virology, Chinese Academy of Sciences , Wuhan 430071, P.R. China.
authors.forename: Hanzhong
authors.initials: H
authors.lastname: Wang
authors.affiliation: Key Laboratory of Analytical Chemistry for Biology and Medicine (Ministry of Education), College of Chemistry and Molecular Sciences, State Key Laboratory of Virology, The Institute for Advanced Studies, and Wuhan Institute of Biotechnology, Wuhan University , Wuhan 430072, P.R. China.
authors.forename: Dai-Wen
authors.initials: DW
authors.lastname: Pang
mesh_terms: D000818:Animals; D002213:Capsid / Q000378:metabolism / Q000648:ultrastructure*; D036022:Capsid Proteins / Q000235:genetics / Q000378:metabolism; D002460:Cell Line; D002467:Cell Nucleus / Q000648:ultrastructure / Q000821:virology; D002522:Chlorocebus aethiops; D006224:Cricetinae; D005456:Fluorescent Dyes / Q000737:chemistry; D015870:Gene Expression; D011558:Herpesvirus 1, Suid / Q000502:physiology / Q000648:ultrastructure*; D051379:Mice; D008856:Microscopy, Fluorescence; D008870:Microtubules / Q000648:ultrastructure / Q000821:virology; D009930:Organic Chemicals / Q000737:chemistry; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D059386:Reverse Genetics; D013194:Staining and Labeling / Q000379:methods*; D014709:Vero Cells; D014771:Virion / Q000378:metabolism / Q000648:ultrastructure*; D019065:Virus Assembly / Q000502:physiology; D057074:Virus Release / Q000502:physiology; D014779:Virus Replication / Q000502:physiology*
chemical_list: D036022:Capsid Proteins; D005456:Fluorescent Dyes; D009930:Organic Chemicals; D011993:Recombinant Fusion Proteins; C461159:SYTO 13
keywords: HaloTag; capsid; labeling; tracking; virus