title: Penetratin and related cell-penetrating cationic peptides can translocate across lipid bilayers in the presence of a transbilayer potential.
abstract: Fluorescent-labeled derivatives of the Antennapedia-derived cell-penetating peptide penetratin, and of the simpler but similarly charged peptides R(6)GC-NH(2) and K(6)GC-NH(2), are shown to be able to translocate into large unilamellar lipid vesicles in the presence of a transbilayer potential (inside negative). Vesicles with diverse lipid compositions, and combining physiological proportions of neutral and anionic lipids, are able to support substantial potential-dependent uptake of all three cationic peptides. The efficiency of peptide uptake under these conditions is strongly modulated by the vesicle lipid composition, in a manner that suggests that more than one mechanism of peptide uptake may operate in different systems. Remarkably, peptide uptake is accompanied by only minor perturbations of the overall barrier function of the lipid bilayer, as assessed by assays of vesicle leakiness under the same conditions. Fluorescence microscopy of living CV-1 and HeLa cells incubated with the labeled peptides shows that the peptides accumulate in peripheral vesicular structures at early times of incubation, consistent with an initial endosomal localization as recently reported, but gradually accumulate in the cytoplasm and nucleus during more extended incubations (several hours). Our findings indicate that these relatively hydrophilic, polybasic cell-penetrating peptides can translocate through lipid bilayers by a potential- and composition-dependent pathway that causes only minimal perturbation to the overall integrity and barrier function of the bilayer.
authors.affiliation: Department of Biochemistry, McGill University, Montréal, Québec, Canada H3G 1Y6.
authors.forename: Donato
authors.initials: D
authors.lastname: Terrone
authors.forename: Stephane Leung Wai
authors.initials: SL
authors.lastname: Sang
authors.forename: Liya
authors.initials: L
authors.lastname: Roudaia
authors.forename: John R
authors.initials: JR
authors.lastname: Silvius
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D051943:Antennapedia Homeodomain Protein; D002352:Carrier Proteins / Q000737:chemistry* / Q000378:metabolism; D002460:Cell Line; D002463:Cell Membrane Permeability / Q000502:physiology*; D057846:Cell-Penetrating Peptides; D002522:Chlorocebus aethiops; D029721:Drosophila Proteins / Q000737:chemistry* / Q000378:metabolism; D006367:HeLa Cells; D018398:Homeodomain Proteins / Q000737:chemistry* / Q000378:metabolism; D006801:Humans; D008051:Lipid Bilayers / Q000737:chemistry* / Q000378:metabolism; D008564:Membrane Potentials; D008969:Molecular Sequence Data; D009687:Nuclear Proteins; D010446:Peptide Fragments / Q000737:chemistry* / Q000378:metabolism; D010713:Phosphatidylcholines / Q000737:chemistry; D010714:Phosphatidylethanolamines / Q000737:chemistry; D010715:Phosphatidylglycerols / Q000737:chemistry; D021381:Protein Transport; D013997:Time Factors; D014157:Transcription Factors
chemical_list: C410262:1,2-dioleoyl-glycero-3-phosphatidyl ethanolamine; D051943:Antennapedia Homeodomain Protein; D002352:Carrier Proteins; D057846:Cell-Penetrating Peptides; D029721:Drosophila Proteins; D018398:Homeodomain Proteins; D008051:Lipid Bilayers; D009687:Nuclear Proteins; D010446:Peptide Fragments; D010713:Phosphatidylcholines; D010714:Phosphatidylethanolamines; D010715:Phosphatidylglycerols; D014157:Transcription Factors; C051388:1,2-dioleoyl-sn-glycero-3-phosphoglycerol; C414409:penetratin; C017251:1,2-oleoylphosphatidylcholine; C028694:1-palmitoyl-2-oleoylphosphatidylcholine