title: Translocation of 5' mRNA cap analogue--peptide conjugates across the membranes of giant unilamellar vesicles.
abstract: Cell-penetrating peptides (CPPs) have been extensively studied because of their ability to deliver various cargo molecules, which are often potential therapeutic agents. However, in most cases, the exact entry mechanism of CPPs is still unknown. In this study, we focused our attention on the membrane permeability sequence (MPS) peptide (AAVALLPAVLLALLAK) conjugated to analogues of a 5' mRNA cap. This unique RNA structure plays a pivotal role in eukaryotic gene expression and has a large therapeutic application potential. We validated the translocation abilities of conjugates across the membranes of giant unilamellar vesicles (GUVs) composed of POPC lipids by application of fluorescence microscopy. Translocation of the MPS peptide itself was observed in contrast to peptide conjugates containing mono- and dinucleotide cap analogues, indicating that even for such small cargos, passive translocation does not occur. However, membrane permeability was observed in the case of conjugated mononucleotides. Fluorescence lifetime microscopy (FLIM) of the C6-NBD-phospholipid revealed changes in lipid packing induced by a penetrating peptide. Our results support the usefulness of artificial membrane systems applied to elucidate membrane crossing mechanisms.
authors.affiliation: Institute of Physics, Polish Academy of Sciences, Al. Lotników 32/45, 02-668 Warsaw, Poland. Electronic address: remiwo@ifpan.edu.pl.
authors.forename: Remigiusz
authors.initials: R
authors.lastname: Worch
authors.affiliation: Faculty of Chemistry, University of Warsaw, Pasteura 1, 02-093 Warsaw, Poland.
authors.forename: Karolina
authors.initials: K
authors.lastname: Piecyk
authors.affiliation: Institute of Physics, Polish Academy of Sciences, Al. Lotników 32/45, 02-668 Warsaw, Poland.
authors.forename: Anna Brygida
authors.initials: AB
authors.lastname: Kolasa
authors.affiliation: Faculty of Chemistry, University of Warsaw, Pasteura 1, 02-093 Warsaw, Poland; Department of Biochemistry, Second Faculty of Medicine, Medical University of Warsaw, Zwirki and Wigury 101, 02-089 Warsaw, Poland. Electronic address: marzena@chem.uw.edu.pl.
authors.forename: Marzena
authors.initials: M
authors.lastname: Jankowska-Anyszka
mesh_terms: D057846:Cell-Penetrating Peptides / Q000737:chemistry*; D008856:Microscopy, Fluorescence; D010713:Phosphatidylcholines / Q000737:chemistry*; D012315:RNA Caps / Q000737:chemistry*; D053835:Unilamellar Liposomes / Q000737:chemistry*
chemical_list: D057846:Cell-Penetrating Peptides; D010713:Phosphatidylcholines; D012315:RNA Caps; D053835:Unilamellar Liposomes; C028694:1-palmitoyl-2-oleoylphosphatidylcholine
keywords: Cap analogue; Cell penetrating peptides; Fluorescence microscopy; Membrane transport; Peptide-oligonucleotide conjugate; Synthetic lipid bilayer