Cytosolic and Nuclear Delivery of Oligonucleotides Mediated by an Amphiphilic Anionic Peptide
- 1 August 1997
- journal article
- Published by Mary Ann Liebert Inc in Antisense and Nucleic Acid Drug Development
- Vol. 7 (4), 335-343
- https://doi.org/10.1089/oli.1.1997.7.335
Abstract
Antisense oligonucleotides (ODN) were easily introduced into the cytosol of mammalian cells on permeabilization of the plasma membrane by an amphiphilic anionic peptide. The E5CA peptide (GLFEAIAEFIEGGWEGLIEGCA) is an E5 peptide analog derived from the N-terminal segment of the HA2 subunit of influenza virus hemagglutinin. This peptide undergoes a conformational change when the pH shifts from neutral to around 6.0, inducing a transient permeabilization of the plasma membrane. In the presence of the E5CA peptide at pH close to 6.0, fluoresceinylated ODN were rapidly taken up by cells and diffused into the nucleus. The uptake of ODN was dependent on the E5CA peptide concentration and on the duration of the incubation at low pH, as shown by confocal microscopy and flow cytometry analyses. This procedure is suitable for loading adherent cells as well as nonadherent cells with single-stranded or double-stranded ODN. Under optimal conditions, a high percentage of cells were nuclei loaded, and the viability was not affected. This method makes use of a well-defined chemical product without the requirement of any special equipment. It will be useful to study the interactions of single-stranded or double-stranded ODN used as antisense, antigenes, or decoys.Keywords
This publication has 33 references indexed in Scilit:
- Altered mRNA Splicing and Inhibition of Human E-selectin Expression by an Antisense Oligonucleotide in Human Umbilical Vein Endothelial CellsPublished by Elsevier ,1996
- Dilation of the influenza hemagglutinin fusion pore revealed by the kinetics of individual cell-cell fusion events.The Journal of cell biology, 1996
- Membrane fusion mediated by the influenza virus hemagglutinin requires the concerted action of at least three hemagglutinin trimers.The Journal of cell biology, 1996
- A Simple Method for Delivering Morpholino Antisense Oligos into the Cytoplasm of CellsAntisense and Nucleic Acid Drug Development, 1996
- Transport of oligonucleotides across natural and model membranesBiochimica et Biophysica Acta (BBA) - Reviews on Biomembranes, 1994
- Cationic Lipids Improve Antisense Oligonucleotide Uptake and Prevent Degradation in Cultured Cells and in Human SerumBiochemical and Biophysical Research Communications, 1993
- pH-Dependent membrane fusion and vesiculation of phospholipid large unilamellar vesicles induced by amphiphilic anionic and cationic peptidesBiochemistry, 1992
- Conformation of membrane fusion-active 20-residue peptides with or without lipid bilayers. Implication of .alpha.-helix formation for membrane fusionBiochemistry, 1990
- Uptake of neoglycoproteins via membrane lectin(s) of L1210 cells evidenced by quantitative flow cytofluorometry and drug targetingBiology of the Cell, 1984
- Weak bases and ionophores rapidly and reversibly raise the pH of endocytic vesicles in cultured mouse fibroblasts.The Journal of cell biology, 1982