title: RNA polymerase I-driven minigenome system for Ebola viruses.
abstract: In general, Ebola viruses are well known for their ability to cause severe hemorrhagic fever in both human and nonhuman primates. However, despite substantial sequence homology to other members of the family Filoviridae, Reston ebolavirus displays reduced pathogenicity for nonhuman primates and has never been demonstrated to cause clinical disease in humans, despite its ability to cause infection. In order to develop a tool to explore potential roles for transcription and replication in the reduced pathogenicity of Reston ebolavirus, we developed an RNA polymerase I (Pol I)-driven minigenome system. Here we demonstrate successful Reston ebolavirus minigenome rescue, including encapsidation, transcription, and replication, as well as the packaging of minigenome transcripts into progeny particles. The Pol I-driven Reston ebolavirus minigenome system provides a higher signal intensity with less background (higher signal-to-noise ratio) than a comparable T7-driven Reston ebolavirus minigenome system which was developed simultaneously. Successful Reston ebolavirus minigenome rescue was also achieved by the use of helper plasmids derived from the closely related Zaire ebolavirus or the more distantly related Lake Victoria marburgvirus. The use of heterologous helper plasmids in the Reston ebolavirus minigenome system yielded levels of reporter expression which far exceeded the level produced by the homologous helper plasmids. This comparison between minigenomes and helper plasmids from different filovirus species and genera indicates that inherent differences in the transcription and/or replication capacities of the ribonucleoprotein complexes of pathogenic and apathogenic filoviruses may exist, as these observations were confirmed in a Lake Victoria marburgvirus minigenome system.
authors.affiliation: National Laboratory for Zoonotic Diseases and Special Pathogens, National Microbiology Laboratory, Public Health Agency of Canada, Winnipeg, MB R3E 3R2, Canada.
authors.forename: Allison
authors.initials: A
authors.lastname: Groseth
authors.forename: Heinz
authors.initials: H
authors.lastname: Feldmann
authors.forename: Steven
authors.initials: S
authors.lastname: Theriault
authors.forename: Gülsah
authors.initials: G
authors.lastname: Mehmetoglu
authors.forename: Ramon
authors.initials: R
authors.lastname: Flick
mesh_terms: D019677:Artificial Gene Fusion; D015500:Chloramphenicol O-Acetyltransferase / Q000032:analysis / Q000235:genetics; D029043:Ebolavirus / Q000235:genetics* / Q000502:physiology*; D017930:Genes, Reporter; D016679:Genome, Viral; D006378:Helper Viruses / Q000235:genetics; D029024:Marburgvirus / Q000235:genetics; D012318:RNA Polymerase I / Q000378:metabolism*; D012367:RNA, Viral / Q000235:genetics / Q000502:physiology*; D014158:Transcription, Genetic; D019065:Virus Assembly / Q000502:physiology; D014779:Virus Replication / Q000502:physiology*
chemical_list: D012367:RNA, Viral; D015500:Chloramphenicol O-Acetyltransferase; D012318:RNA Polymerase I