title: A novel life cycle modeling system for Ebola virus shows a genome length-dependent role of VP24 in virus infectivity.
abstract: Work with infectious Ebola viruses is restricted to biosafety level 4 (BSL4) laboratories, presenting a significant barrier for studying these viruses. Life cycle modeling systems, including minigenome systems and transcription- and replication-competent virus-like particle (trVLP) systems, allow modeling of the virus life cycle under BSL2 conditions; however, all current systems model only certain aspects of the virus life cycle, rely on plasmid-based viral protein expression, and have been used to model only single infectious cycles. We have developed a novel life cycle modeling system allowing continuous passaging of infectious trVLPs containing a tetracistronic minigenome that encodes a reporter and the viral proteins VP40, VP24, and GP1,2. This system is ideally suited for studying morphogenesis, budding, and entry, in addition to genome replication and transcription. Importantly, the specific infectivity of trVLPs in this system was ∼ 500-fold higher than that in previous systems. Using this system for functional studies of VP24, we showed that, contrary to previous reports, VP24 only very modestly inhibits genome replication and transcription when expressed in a regulated fashion, which we confirmed using infectious Ebola viruses. Interestingly, we also discovered a genome length-dependent effect of VP24 on particle infectivity, which was previously undetected due to the short length of monocistronic minigenomes and which is due at least partially to a previously unknown function of VP24 in RNA packaging. Based on our findings, we propose a model for the function of VP24 that reconciles all currently available data regarding the role of VP24 in nucleocapsid assembly as well as genome replication and transcription.
  
  
  OBJECTIVE
  Ebola viruses cause severe hemorrhagic fevers in humans, with no countermeasures currently being available, and must be studied in maximum-containment laboratories. Only a few of these laboratories exist worldwide, limiting our ability to study Ebola viruses and develop countermeasures. Here we report the development of a novel reverse genetics-based system that allows the study of Ebola viruses without maximum-containment laboratories. We used this system to investigate the Ebola virus protein VP24, showing that, contrary to previous reports, it only modestly inhibits virus genome replication and transcription but is important for packaging of genomes into virus particles, which constitutes a previously unknown function of VP24 and a potential antiviral target. We further propose a comprehensive model for the function of VP24 in nucleocapsid assembly. Importantly, on the basis of this approach, it should easily be possible to develop similar experimental systems for other viruses that are currently restricted to maximum-containment laboratories.
authors: Laboratory of Virology, Division of Intramural Research, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Hamilton, Montana, USA.
authors: Ari
authors: A
authors: Watt
authors: Institute for Virology, Philipps University Marburg, Marburg, Germany.
authors: Felicien
authors: F
authors: Moukambi
authors: Laboratory of Virology, Division of Intramural Research, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Hamilton, Montana, USA.
authors: Logan
authors: L
authors: Banadyga
authors: Laboratory of Virology, Division of Intramural Research, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Hamilton, Montana, USA Institute for Virology, Philipps University Marburg, Marburg, Germany.
authors: Allison
authors: A
authors: Groseth
authors: Laboratory of Virology, Division of Intramural Research, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Hamilton, Montana, USA.
authors: Julie
authors: J
authors: Callison
authors: Institute for Virology, Philipps University Marburg, Marburg, Germany.
authors: Astrid
authors: A
authors: Herwig
authors: Laboratory of Virology, Division of Intramural Research, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Hamilton, Montana, USA.
authors: Hideki
authors: H
authors: Ebihara
authors: Laboratory of Virology, Division of Intramural Research, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Hamilton, Montana, USA feldmannh@niaid.nih.gov thomas.hoenen@nih.gov.
authors: Heinz
authors: H
authors: Feldmann
authors: Laboratory of Virology, Division of Intramural Research, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Hamilton, Montana, USA Institute for Virology, Philipps University Marburg, Marburg, Germany feldmannh@niaid.nih.gov thomas.hoenen@nih.gov.
authors: Thomas
authors: T
authors: Hoenen
mesh_terms: D029043:Ebolavirus / Q000235:genetics / Q000254:growth & development* / Q000472:pathogenicity / Q000502:physiology*; D016679:Genome, Viral; D019142:Hemorrhagic Fever, Ebola / Q000821:virology*; D006801:Humans; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism*; D014774:Virulence; D019065:Virus Assembly; D014779:Virus Replication
chemical_list: C446938:VP24 protein, Ebola virus; D014764:Viral Proteins