Related Experiment Video
Updated: Aug 14, 2026

A Reporter Based Cellular Assay for Monitoring Splicing Efficiency
Published on: September 15, 2021
Endosome trapping limits the efficiency of splicing correction by PNA-oligolysine conjugates
Saïd Abes1, Donna Williams, Paul Prevot
1UMR 5124 CNRS, CC 086, Université Montpellier 2, Montpellier, France.
Abstract:
Splicing correction by steric-blocking oligonucleotides (ON) might lead to important clinical applications but requires efficient delivery to cell nuclei. The conjugation of short oligolysine tails has been used to deliver a correcting peptide nucleic acid (PNA) sequence in a positive readout assay in which ON hybridization to the cryptic splice site is strictly required for the expression of a luciferase reporter gene. We have investigated the mechanism of cellular uptake and the efficiency of a (Lys)(8)-PNA-Lys construction in this model system. Cell uptake is temperature-dependent and leads to sequestration of the conjugate in cytoplasmic vesicles in keeping with an endocytic mechanism of internalization. Accordingly a significant and sequence-specific splicing correction is achieved only in the presence of endosome-disrupting agents as chloroquine or 0.5 M sucrose. These endosome-disrupting agents do not affect the activity of free PNA, and do not increase (Lys)(8)-PNA-Lys uptake.
Related Concept Videos
RNA Splicing
Nonsense-mediated mRNA Decay
Usually, Upf3 binds to an Exon Junction Complex (EJC) at mRNA splice sites. If a ribosome fully translates the mRNA,...
Translesion DNA Polymerases
TLS polymerases are found in all three domains of life - archaea, bacteria, and eukaryotes. Of the different classes of TLS polymerases, members of the Y family are fitted with specialized structures that...
Proofreading
Errors During Replication are Corrected by the DNA Polymerase Enzyme
Proofreading
Regulation of Nuclear Protein Sorting
