Inhibition of Nuclear Translocation of Transcription Factor NF-κB by a Synthetic Peptide Containing a Cell Membrane-permeable Motif and Nuclear Localization Sequence

Abstract

To control agonist-induced nuclear translocation of transcription factor kappa B (NF-kappa B) in intact cells, cell-permeable synthetic peptides were devised. Their import into intact cells was dependent on a hydrophobic region selected from the signal peptide sequences and was verified by their inaccessibility to extracellular proteases and by confocal laser scanning microscopy. When a cell-permeable peptide carried a functional cargo representing the nuclear localization sequence of NF-kappa B p50, it inhibited in a concentration-dependent manner nuclear translocation of NF-kappa B in cultured endothelial and monocytic cells stimulated with lipopolysaccharide or tumor necrosis factor-alpha. Synthetic peptide analogues with deleted hydrophobic cell membrane-permeable motif or with a mutated nuclear localization sequence were inactive. Cell membrane-permeable peptides were not cytotoxic within the concentration range used in these experiments. These results suggest that cell-permeable synthetic peptides carrying a functional cargo can be applied to control signal transduction-dependent subcellular traffic of transcription factors mediating the cellular responses to different agonists. Moreover, this approach can be used to study other intracellular processes involving proteins with functionally distinct domains.

Toll-like receptor 4-mediated nuclear factor-κB activation in spinal cord contributes to chronic morphine-induced analgesic tolerance and hyperalgesia in rats

“…SN50 is a cellpermeable synthetic peptide that is known to inhibit NF-κB activation directly by blocking the translocation of the active NF-κB complex into the nucleus [41] . Our results showed that NF-κB inhibition prevented the development of chronic morphine-induced analgesic tolerance, alleviated morphine withdrawal-induced allodynia and hyperalgesia, and partially reversed the established analgesic tolerance.…”

Inhibitors of NF-κB signaling: 785 and counting

“…One approach to block this step has used cellpermeable peptides that contain the nuclear localizing sequence of p50. These peptides are thought to inhibit nuclear translocation of p50-containing dimers by saturating the nuclear import machinery responsible for the uptake of NF-kB dimers containing p50 (Lin et al, 1995;Torgerson et al, 1998;Letoha et al, 2005). However, one of the more commonly used peptides of this type, SN-50, has been reported to block nuclear translocation of a number of non-NF-kB transcription factors as well (Torgerson et al, 1998).…”

Activation of NF-kB mediates ICAM-1 induction in respiratory cells exposed to an adenovirus-derived vector

“…The peptide SN50M has mutations in two of the 10 residues within the NLS of SN50 and is used as control for specificity of inhibition by SN50. 19 The effect of these inhibitors was first tested on ICAM-1 mRNA expression, which was quantitated by single-tube competitive RT-PCR as described. 2 We utilized the model system of the alveolar type II-derived A549 cells exposed to Ad.CFTR for 24 h. Figure 2a shows that Ad.CFTR up-regulated ICAM-1 mRNA levels from two-to four-fold at doses ranging from 0.1 to 50 MOI.…”