title: The cooperative function of arginine residues in the Prototype Foamy Virus Gag C-terminus mediates viral and cellular RNA encapsidation.
abstract: BACKGROUND
  One unique feature of the foamy virus (FV) capsid protein Gag is the absence of Cys-His motifs, which in orthoretroviruses are irreplaceable for multitude functions including viral RNA genome recognition and packaging. Instead, FV Gag contains glycine-arginine-rich (GR) sequences at its C-terminus. In case of prototype FV (PFV) these are historically grouped in three boxes, which have been shown to play essential functions in genome reverse transcription, virion infectivity and particle morphogenesis. Additional functions for RNA packaging and Pol encapsidation were suggested, but have not been conclusively addressed.
  
  
  RESULTS
  Here we show that released wild type PFV particles, like orthoretroviruses, contain various cellular RNAs in addition to viral genome. Unlike orthoretroviruses, the content of selected cellular RNAs in capsids of PFV vector particles was not altered by viral genome encapsidation. Deletion of individual GR boxes had only minor negative effects (2 to 4-fold) on viral and cellular RNA encapsidation over a wide range of cellular Gag to viral genome ratios examined. Only the concurrent deletion of all three PFV Gag GR boxes, or the substitution of multiple arginine residues residing in the C-terminal GR box region by alanine, abolished both viral and cellular RNA encapsidation (>50 to >3,000-fold reduced), independent of the viral production system used. Consequently, those mutants also lacked detectable amounts of encapsidated Pol and were non-infectious. In contrast, particle release was reduced to a much lower extent (3 to 20-fold).
  
  
  CONCLUSIONS
  Taken together, our data provides the first identification of a full-length PFV Gag mutant devoid in genome packaging and the first report of cellular RNA encapsidation into PFV particles. Our results suggest that the cooperative action of C-terminal clustered positively charged residues, present in all FV Gag proteins, is the main viral protein determinant for viral and cellular RNA encapsidation. The viral genome independent efficiency of cellular RNA encapsidation suggests differential packaging mechanisms for both types of RNAs. Finally, this study indicates that analogous to orthoretroviruses, Gag - nucleic acid interactions are required for FV capsid assembly and efficient particle release.
authors.affiliation: Institute of Virology, Medical Faculty "Carl Gustav Carus", Technische Universität Dresden, Fetscherstr. 74, 01307, Dresden, Germany. martin.hamann@tu-dresden.de.|CRTD/DFG-Center for Regenerative Therapies Dresden - Cluster of Excellence, Technische Universität Dresden, Fetscherstr. 105, 01307, Dresden, Germany. martin.hamann@tu-dresden.de.
authors.forename: Martin V
authors.initials: MV
authors.lastname: Hamann
authors.affiliation: Institute of Virology, Medical Faculty "Carl Gustav Carus", Technische Universität Dresden, Fetscherstr. 74, 01307, Dresden, Germany. erik.mullers@ki.se.|CRTD/DFG-Center for Regenerative Therapies Dresden - Cluster of Excellence, Technische Universität Dresden, Fetscherstr. 105, 01307, Dresden, Germany. erik.mullers@ki.se.|Present address: Department of Cell and Molecular Biology, Karolinska Institutet, 171 77, Stockholm, Sweden. erik.mullers@ki.se.
authors.forename: Erik
authors.initials: E
authors.lastname: Müllers
authors.affiliation: Institute of Virology, Medical Faculty "Carl Gustav Carus", Technische Universität Dresden, Fetscherstr. 74, 01307, Dresden, Germany. juliane.reh@tu-dresden.de.|CRTD/DFG-Center for Regenerative Therapies Dresden - Cluster of Excellence, Technische Universität Dresden, Fetscherstr. 105, 01307, Dresden, Germany. juliane.reh@tu-dresden.de.
authors.forename: Juliane
authors.initials: J
authors.lastname: Reh
authors.affiliation: Institute of Virology, Medical Faculty "Carl Gustav Carus", Technische Universität Dresden, Fetscherstr. 74, 01307, Dresden, Germany. nicole.stanke@tu-dresden.de.|CRTD/DFG-Center for Regenerative Therapies Dresden - Cluster of Excellence, Technische Universität Dresden, Fetscherstr. 105, 01307, Dresden, Germany. nicole.stanke@tu-dresden.de.
authors.forename: Nicole
authors.initials: N
authors.lastname: Stanke
authors.affiliation: Univ. Grenoble Alpes, UVHCI, F-38000, Grenoble, France. greg.effantin@gmail.com.|CNRS, UVHCI, F-38000, Grenoble, France. greg.effantin@gmail.com.
authors.forename: Gregory
authors.initials: G
authors.lastname: Effantin
authors.affiliation: Univ. Grenoble Alpes, UVHCI, F-38000, Grenoble, France. weissenhorn@embl.fr.|CNRS, UVHCI, F-38000, Grenoble, France. weissenhorn@embl.fr.
authors.forename: Winfried
authors.initials: W
authors.lastname: Weissenhorn
authors.affiliation: Institute of Virology, Medical Faculty "Carl Gustav Carus", Technische Universität Dresden, Fetscherstr. 74, 01307, Dresden, Germany. dirk.lindemann@tu-dresden.de.|CRTD/DFG-Center for Regenerative Therapies Dresden - Cluster of Excellence, Technische Universität Dresden, Fetscherstr. 105, 01307, Dresden, Germany. dirk.lindemann@tu-dresden.de.
authors.forename: Dirk
authors.initials: D
authors.lastname: Lindemann
mesh_terms: D019943:Amino Acid Substitution; D001120:Arginine / Q000378:metabolism*; D002460:Cell Line; D015683:Gene Products, gag / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D020125:Mutation, Missense; D012313:RNA / Q000378:metabolism*; D017384:Sequence Deletion; D016092:Spumavirus / Q000235:genetics / Q000502:physiology*; D019065:Virus Assembly
chemical_list: D015683:Gene Products, gag; D012313:RNA; D001120:Arginine