title: Analysis of prototype foamy virus particle-host cell interaction with autofluorescent retroviral particles.
abstract: BACKGROUND
  The foamy virus (FV) replication cycle displays several unique features, which set them apart from orthoretroviruses. First, like other B/D type orthoretroviruses, FV capsids preassemble at the centrosome, but more similar to hepadnaviruses, FV budding is strictly dependent on cognate viral glycoprotein coexpression. Second, the unusually broad host range of FV is thought to be due to use of a very common entry receptor present on host cell plasma membranes, because all cell lines tested in vitro so far are permissive.
  
  
  RESULTS
  In order to take advantage of modern fluorescent microscopy techniques to study FV replication, we have created FV Gag proteins bearing a variety of protein tags and evaluated these for their ability to support various steps of FV replication. Addition of even small N-terminal HA-tags to FV Gag severely impaired FV particle release. For example, release was completely abrogated by an N-terminal autofluorescent protein (AFP) fusion, despite apparently normal intracellular capsid assembly. In contrast, C-terminal Gag-tags had only minor effects on particle assembly, egress and particle morphogenesis. The infectivity of C-terminal capsid-tagged FV vector particles was reduced up to 100-fold in comparison to wild type; however, infectivity was rescued by coexpression of wild type Gag and assembly of mixed particles. Specific dose-dependent binding of fluorescent FV particles to target cells was demonstrated in an Env-dependent manner, but not binding to target cell-extracted- or synthetic- lipids. Screening of target cells of various origins resulted in the identification of two cell lines, a human erythroid precursor- and a zebrafish- cell line, resistant to FV Env-mediated FV- and HIV-vector transduction.
  
  
  CONCLUSIONS
  We have established functional, autofluorescent foamy viral particles as a valuable new tool to study FV--host cell interactions using modern fluorescent imaging techniques. Furthermore, we succeeded for the first time in identifying two cell lines resistant to Prototype Foamy Virus Env-mediated gene transfer. Interestingly, both cell lines still displayed FV Env-dependent attachment of fluorescent retroviral particles, implying a post-binding block potentially due to lack of putative FV entry cofactors. These cell lines might ultimately lead to the identification of the currently unknown ubiquitous cellular entry receptor(s) of FVs.
authors.affiliation: Institut für Virologie, Medizinische Fakultät Carl Gustav Carus, Technische Universität Dresden, Dresden, Germany.
authors.forename: Kristin
authors.initials: K
authors.lastname: Stirnnagel
authors.forename: Daniel
authors.initials: D
authors.lastname: Lüftenegger
authors.forename: Annett
authors.initials: A
authors.lastname: Stange
authors.forename: Anka
authors.initials: A
authors.lastname: Swiersy
authors.forename: Erik
authors.initials: E
authors.lastname: Müllers
authors.forename: Juliane
authors.initials: J
authors.lastname: Reh
authors.forename: Nicole
authors.initials: N
authors.lastname: Stanke
authors.forename: Arend
authors.initials: A
authors.lastname: Grosse
authors.forename: Salvatore
authors.initials: S
authors.lastname: Chiantia
authors.forename: Heiko
authors.initials: H
authors.lastname: Keller
authors.forename: Petra
authors.initials: P
authors.lastname: Schwille
authors.forename: Helmut
authors.initials: H
authors.lastname: Hanenberg
authors.forename: Hanswalter
authors.initials: H
authors.lastname: Zentgraf
authors.forename: Dirk
authors.initials: D
authors.lastname: Lindemann
mesh_terms: D000818:Animals; D002460:Cell Line; D054884:Host-Pathogen Interactions; D006801:Humans; D008164:Luminescent Proteins / Q000235:genetics* / Q000378:metabolism; D008856:Microscopy, Fluorescence / Q000379:methods; D011995:Recombination, Genetic; D053150:Simian foamy virus / Q000235:genetics / Q000502:physiology*; D013194:Staining and Labeling / Q000379:methods; D014764:Viral Proteins / Q000235:genetics* / Q000502:physiology; D014773:Virology / Q000379:methods*; D015027:Zebrafish
chemical_list: D008164:Luminescent Proteins; D014764:Viral Proteins