title: Epstein-Barr virus nuclear antigen 1 interacts with regulator of chromosome condensation 1 dynamically throughout the cell cycle.
abstract: The Epstein-Barr virus (EBV) nuclear antigen 1 (EBNA1) is a sequence-specific DNA-binding protein that plays an essential role in viral episome replication and segregation, by recruiting the cellular complex of DNA replication onto the origin (oriP) and by tethering the viral DNA onto the mitotic chromosomes. Whereas the mechanisms of viral DNA replication are well documented, those involved in tethering EBNA1 to the cellular chromatin are far from being understood. Here, we have identified regulator of chromosome condensation 1 (RCC1) as a novel cellular partner for EBNA1. RCC1 is the major nuclear guanine nucleotide exchange factor for the small GTPase Ran enzyme. RCC1, associated with chromatin, is involved in the formation of RanGTP gradients critical for nucleo-cytoplasmic transport, mitotic spindle formation and nuclear envelope reassembly following mitosis. Using several approaches, we have demonstrated a direct interaction between these two proteins and found that the EBNA1 domains responsible for EBNA1 tethering to the mitotic chromosomes are also involved in the interaction with RCC1. The use of an EBNA1 peptide array confirmed the interaction of RCC1 with these regions and also the importance of the N-terminal region of RCC1 in this interaction. Finally, using confocal microscopy and Förster resonance energy transfer analysis to follow the dynamics of interaction between the two proteins throughout the cell cycle, we have demonstrated that EBNA1 and RCC1 closely associate on the chromosomes during metaphase, suggesting an essential role for the interaction during this phase, perhaps in tethering EBNA1 to mitotic chromosomes.
authors.affiliation: Université Lyon 1, Centre International de Recherche en Infectiologie, Lyon 69364, France.|CNRS, UMR5308, Lyon 69364, France.|CIRI, International Center for Infectiology Research, Oncogenic Herpesviruses Team, Université de Lyon, Lyon 69364, France.|Ecole Normale Supérieure de Lyon, Lyon 69364, France.|INSERM, U1111, Lyon 69364, France.
authors.forename: Thibaut
authors.initials: T
authors.lastname: Deschamps
authors.affiliation: Ecole Normale Supérieure de Lyon, Lyon 69364, France.|CNRS, UMR5308, Lyon 69364, France.|Université Lyon 1, Centre International de Recherche en Infectiologie, Lyon 69364, France.|Present address: Section of Virology, Department of Medicine, Imperial College London, St Mary's Campus, London, UK.|CIRI, International Center for Infectiology Research, Oncogenic Herpesviruses Team, Université de Lyon, Lyon 69364, France.|INSERM, U1111, Lyon 69364, France.
authors.forename: Quentin
authors.initials: Q
authors.lastname: Bazot
authors.affiliation: Present address: University of Oxford, Ludwig Institute for Cancer Research, Oxford, UK.|College of Medical, Veterinary and Life Sciences, University of Glasgow, Glasgow G12 8QQ, UK.
authors.forename: Derek M
authors.initials: DM
authors.lastname: Leske
authors.affiliation: College of Medical, Veterinary and Life Sciences, University of Glasgow, Glasgow G12 8QQ, UK.
authors.forename: Ruth
authors.initials: R
authors.lastname: MacLeod
authors.affiliation: Present address: University Joseph Fourier, Pathogenesis and Lentiviral Vaccination Laboratory, Grenoble, France.|INSERM, U1111, Lyon 69364, France.|Université Lyon 1, Centre International de Recherche en Infectiologie, Lyon 69364, France.|Ecole Normale Supérieure de Lyon, Lyon 69364, France.|CIRI, International Center for Infectiology Research, Oncogenic Herpesviruses Team, Université de Lyon, Lyon 69364, France.|CNRS, UMR5308, Lyon 69364, France.
authors.forename: Dimitri
authors.initials: D
authors.lastname: Mompelat
authors.affiliation: CIRI, International Center for Infectiology Research, Cell Biology of Viral Infections Team, Université de Lyon, Lyon 69364, France.|INSERM, U1111, Lyon 69364, France.|Present address: Cell Biology Lab, University of Mons, Mons, Belgium.|Université Lyon 1, Centre International de Recherche en Infectiologie, Lyon 69364, France.
authors.forename: Lionel
authors.initials: L
authors.lastname: Tafforeau
authors.affiliation: Université Lyon 1, Centre International de Recherche en Infectiologie, Lyon 69364, France.|Ecole Normale Supérieure de Lyon, Lyon 69364, France.|CNRS, UMR5308, Lyon 69364, France.|INSERM, U1111, Lyon 69364, France.|CIRI, International Center for Infectiology Research, Cell Biology of Viral Infections Team, Université de Lyon, Lyon 69364, France.
authors.forename: Vincent
authors.initials: V
authors.lastname: Lotteau
authors.affiliation: UPMC Université Paris 6, Inserm, Centre d'Immunologie et des Maladies Infectieuses (Cimi-Paris), UMR 1135, ERL CNRS 8255, F-75013 Paris, France.
authors.forename: Vincent
authors.initials: V
authors.lastname: Maréchal
authors.affiliation: College of Medical, Veterinary and Life Sciences, University of Glasgow, Glasgow G12 8QQ, UK.
authors.forename: George S
authors.initials: GS
authors.lastname: Baillie
authors.affiliation: CNRS, UMR5308, Lyon 69364, France.|INSERM, U1111, Lyon 69364, France.|Université Lyon 1, Centre International de Recherche en Infectiologie, Lyon 69364, France.|CIRI, International Center for Infectiology Research, Oncogenic Herpesviruses Team, Université de Lyon, Lyon 69364, France.|Ecole Normale Supérieure de Lyon, Lyon 69364, France.
authors.forename: Henri
authors.initials: H
authors.lastname: Gruffat
authors.affiliation: College of Medical, Veterinary and Life Sciences, University of Glasgow, Glasgow G12 8QQ, UK.
authors.forename: Joanna B
authors.initials: JB
authors.lastname: Wilson
authors.affiliation: INSERM, U1111, Lyon 69364, France.|Université Lyon 1, Centre International de Recherche en Infectiologie, Lyon 69364, France.|CIRI, International Center for Infectiology Research, Oncogenic Herpesviruses Team, Université de Lyon, Lyon 69364, France.|Ecole Normale Supérieure de Lyon, Lyon 69364, France.|CNRS, UMR5308, Lyon 69364, France.
authors.forename: Evelyne
authors.initials: E
authors.lastname: Manet
mesh_terms: D020816:Amino Acid Motifs; D018797:Cell Cycle Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002843:Chromatin / Q000378:metabolism; D002877:Chromosomes, Human / Q000378:metabolism; D019309:Epstein-Barr Virus Nuclear Antigens / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D031541:Fluorescence Resonance Energy Transfer; D020662:Guanine Nucleotide Exchange Factors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D057809:HEK293 Cells; D006367:HeLa Cells; D006801:Humans; D008677:Metaphase; D018613:Microscopy, Confocal; D008938:Mitosis; D009687:Nuclear Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D040081:Protein Array Analysis; D054730:Protein Interaction Domains and Motifs; D025941:Protein Interaction Mapping; D008941:Spindle Apparatus / Q000378:metabolism
chemical_list: D018797:Cell Cycle Proteins; D002843:Chromatin; D019309:Epstein-Barr Virus Nuclear Antigens; D020662:Guanine Nucleotide Exchange Factors; D009687:Nuclear Proteins; C062521:RCC1 protein, human; C417029:EBV-encoded nuclear antigen 1