title: Liposomal nanocontainers as models for viral infection: monitoring viral genomic RNA transfer through lipid membranes.
abstract: After uptake into target cells, many nonenveloped viruses undergo conformational changes in the low-pH environment of the endocytic compartment. This results in exposure of amphipathic viral peptides and/or hydrophobic protein domains that are inserted into and either disrupt or perforate the vesicular membranes. The viral nucleic acids thereby gain access to the cytosol and initiate replication. We here demonstrate the in vitro transfer of the single-stranded positive-sense RNA genome of human rhinovirus 2 into liposomes decorated with recombinant very-low-density lipoprotein receptor fragments. Membrane-attached virions were exposed to pH 5.4, mimicking the in vivo pH environment of late endosomes. This triggered the release of the RNA whose arrival in the liposomal lumen was detected via in situ cDNA synthesis by encapsulated reverse transcriptase. Subsequently, cDNA was PCR amplified. At a low ratio between virions and lipids, RNA transfer was positively correlated with virus concentration. However, membranes became leaky at higher virus concentrations, which resulted in decreased cDNA synthesis. In accordance with earlier in vivo data, the RNA passes through the lipid membrane without causing gross damage to vesicles at physiologically relevant virus concentrations.
authors.affiliation: Max F. Perutz Laboratories, Dept. Med. Biochemistry, Medical University of Vienna, Vienna, Austria.
authors.forename: Gerhard
authors.initials: G
authors.lastname: Bilek
authors.forename: Nena M
authors.initials: NM
authors.lastname: Matscheko
authors.forename: Angela
authors.initials: A
authors.lastname: Pickl-Herk
authors.forename: Victor U
authors.initials: VU
authors.lastname: Weiss
authors.forename: Xavier
authors.initials: X
authors.lastname: Subirats
authors.forename: Ernst
authors.initials: E
authors.lastname: Kenndler
authors.forename: Dieter
authors.initials: D
authors.lastname: Blaas
mesh_terms: D002462:Cell Membrane / Q000378:metabolism* / Q000821:virology; D016679:Genome, Viral; D006863:Hydrogen-Ion Concentration; D008081:Liposomes; D008563:Membrane Lipids / Q000378:metabolism; D046529:Microscopy, Electron, Transmission; D049329:Nanostructures; D012327:RNA Virus Infections / Q000821:virology*; D012367:RNA, Viral / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012229:Rhinovirus / Q000502:physiology*; D053586:Virus Internalization; D057646:Virus Uncoating
chemical_list: D008081:Liposomes; D008563:Membrane Lipids; D012367:RNA, Viral