Cellular delivery of a double-stranded oligonucleotide
Open Access
- 1 August 2004
- journal article
- research article
- Published by Springer Nature in Gene Therapy
- Vol. 11 (16), 1264-1272
- https://doi.org/10.1038/sj.gt.3302291
Abstract
The activation of nuclear factor κB (NFκB) is a key event in immune and inflammatory responses. In this study, a cell-penetrating transport peptide, transportan (TP) or its shorter analogue TP 10, was used to facilitate the cellular uptake of an NFκB decoy. Peptide nucleic acid (PNA) hexamer or nonamer was linked to the transport peptide by a disulfide bond. NFκB decoy oligonucleotide consisted of a double-stranded consensus sequence corresponding to the κB site localized in the IL-6 gene promoter, 5′-GGGACTTTCCC-3′, with a single-stranded protruding 3′-terminal sequence complementary to the PNA sequence was hybridized to the transport peptide–PNA construct. The ability of the transport peptide–PNA–NFκB decoy complex to block the effect of interleukin (IL)-1β-induced NFκB activation and IL-6 gene expression was analyzed by electrophoretic mobility shift assay and reverse transcriptase-polymerase chain reaction in rat Rinm5F insulinoma cells. Preincubation with transport peptide–PNA–NFκB decoy (1 μM, 1 h) blocked IL-1β-induced NFκB-binding activity and significantly reduced the IL-6 mRNA expression. The same concentration of NFκB decoy in the absence of transport peptide–PNA had no effect even after longer incubations. Our results showed that binding of the oligonucleotide NFκB decoy to the nonamer PNA sequence resulted in a stable complex that was efficiently translocated across the plasma membrane.Keywords
This publication has 62 references indexed in Scilit:
- Recent advances torwards understanding redox mechanisms in the activation of nuclear factor κbFree Radical Biology & Medicine, 2000
- Activators and target genes of Rel/NF-κB transcription factorsOncogene, 1999
- MyD88, an Adapter Protein Involved in Interleukin-1 SignalingJournal of Biological Chemistry, 1998
- Signal transduction through NF-κBImmunology Today, 1998
- IκB Kinase-β: NF-κB Activation and Complex Formation with IκB Kinase-α and NIKScience, 1997
- Identification and Characterization of an IκB KinaseCell, 1997
- NF-kB: a crucial transcription factor for glial and neuronal cell functionTrends in Neurosciences, 1997
- THE NF-κB AND IκB PROTEINS: New Discoveries and InsightsAnnual Review of Immunology, 1996
- Function and Activation of NF-kappaB in the Immune SystemAnnual Review of Immunology, 1994
- IκB: a Specific Inhibitor of the NF-κB Transcription FactorScience, 1988