title: Epstein-Barr virus protein kinase BGLF4 targets the nucleus through interaction with nucleoporins.
abstract: BGLF4 of Epstein-Barr virus (EBV) encodes a serine/threonine protein kinase that phosphorylates multiple viral and cellular substrates to optimize the cellular environment for viral DNA replication and the nuclear egress of viral nucleocapsids. BGLF4 is expressed predominantly in the nucleus at early and late stages of virus replication, while a small portion of BGLF4 is distributed in the cytoplasm at the late stage of virus replication and packaged into the virion. Here, we analyzed systematically the functional domains crucial for nuclear localization of BGLF4 and found that both the N and C termini play important modulating roles. Analysis of amino acid substitution mutants revealed that the C terminus of BGLF4 does not contain a conventional nuclear localization signal (NLS). Additionally, deletion of the C-terminal putative helical regions at amino acids 386 to 393 and 410 to 419 diminished the nuclear translocation of BGLF4, indicating that the secondary structure of the C terminus is important for the localization of BGLF4. The green fluorescent protein-fused wild-type or C-terminal helical regions of BGLF4 associate with phenylalanine/glycine repeat-containing nucleoporins (Nups) in nuclear envelope fractionation. Both coimmunoprecipitation and in vitro pull-down assays further demonstrated that BGLF4 binds to Nup62 and Nup153. Remarkably, nuclear import assay with permeabilized HeLa cells demonstrated that BGLF4 translocated into nucleus independent of cytosolic factors. Data presented here suggest that BGLF4 employs a novel mechanism through direct interactions with nucleoporins for its nuclear targeting.
authors.affiliation: Graduate Institute and Department of Microbiology, College of Medicine, National Taiwan University, Taipei, Taiwan.
authors.forename: Chou-Wei
authors.initials: CW
authors.lastname: Chang
authors.forename: Chung-Pei
authors.initials: CP
authors.lastname: Lee
authors.forename: Yu-Hao
authors.initials: YH
authors.lastname: Huang
authors.forename: Pei-Wen
authors.initials: PW
authors.lastname: Yang
authors.forename: Jiin-Tarng
authors.initials: JT
authors.lastname: Wang
authors.forename: Mei-Ru
authors.initials: MR
authors.lastname: Chen
mesh_terms: D021581:Active Transport, Cell Nucleus / Q000235:genetics; D019943:Amino Acid Substitution; D002467:Cell Nucleus / Q000201:enzymology* / Q000235:genetics / Q000821:virology; D004261:DNA Replication / Q000502:physiology; D004279:DNA, Viral / Q000235:genetics / Q000378:metabolism; D020031:Epstein-Barr Virus Infections / Q000235:genetics / Q000378:metabolism*; D006367:HeLa Cells; D004854:Herpesvirus 4, Human / Q000502:physiology*; D006801:Humans; D008562:Membrane Glycoproteins / Q000235:genetics / Q000378:metabolism*; D020125:Mutation, Missense; D028861:Nuclear Pore Complex Proteins / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism*; D019065:Virus Assembly / Q000502:physiology*
chemical_list: D004279:DNA, Viral; D008562:Membrane Glycoproteins; C079377:NUP153 protein, human; D028861:Nuclear Pore Complex Proteins; D014764:Viral Proteins; C049312:nuclear pore protein p62; C406063:BGLF4 protein, Epstein-Barr virus; D017346:Protein Serine-Threonine Kinases