title: Scrapie infection of prion protein-deficient cell line upon ectopic expression of mutant prion proteins.
abstract: Expression of the cellular prion protein (PrP(C)) is crucial for susceptibility to prions. In vivo, ectopic expression of PrP(C) restores susceptibility to prions and transgenic mice that express heterologous PrP on a PrP knock-out background have been used extensively to study the role of PrP alterations for prion transmission and species barriers. Here we report that prion protein knock-out cells can be rendered permissive to scrapie infection by the ectopic expression of PrP. The system was used to study the influence of sheep PrP-specific residues in mouse PrP on the infection process with mouse adapted scrapie. These studies reveal several critical residues previously not associated with species barriers and demonstrate that amino acid residue alterations at positions known to have an impact on the susceptibility of sheep to sheep scrapie also drastically influence PrP(Sc) formation by mouse-adapted scrapie strain 22L. Furthermore, our data suggest that amino acid polymorphisms located on the outer surfaces of helix 2 and 3 drastically impact conversion efficiency. In conclusion, this system allows for the fast generation of mutant PrP(Sc) that is entirely composed of transgenic PrP and is, thus, ideally suited for testing if artificial PrP molecules can affect prion replication. Transmission of infectivity generated in HpL3-4 cells expressing altered PrP molecules to mice could also help to unravel the potential influence of mutant PrP(Sc) on host cell tropism and strain characteristics in vivo.
authors.affiliation: Institute of Virology, Technical University of Munich, Troger Strasse 30, 81675 Munich, Germany.
authors.forename: Elke
authors.initials: E
authors.lastname: Maas
authors.forename: Markus
authors.initials: M
authors.lastname: Geissen
authors.forename: Martin H
authors.initials: MH
authors.lastname: Groschup
authors.forename: Romina
authors.initials: R
authors.lastname: Rost
authors.forename: Takashi
authors.initials: T
authors.lastname: Onodera
authors.forename: Hermann
authors.initials: H
authors.lastname: Schätzl
authors.forename: Ina M
authors.initials: IM
authors.lastname: Vorberg
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D018929:Cell Culture Techniques / Q000379:methods*; D002460:Cell Line; D003470:Culture Media / Q000378:metabolism; D000939:Epitopes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D006624:Hippocampus / Q000166:cytology; D051379:Mice; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D018620:PrPSc Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D012190:Retroviridae / Q000235:genetics; D012608:Scrapie / Q000473:pathology / Q000503:physiopathology* / Q000635:transmission; D012756:Sheep; D014161:Transduction, Genetic
chemical_list: D003470:Culture Media; D000939:Epitopes; D018620:PrPSc Proteins