Peptide-Mediated Disruption of NFB/NRF Interaction Inhibits IL-8 Gene Activation by IL-1 or Helicobacter pylori

Authors:
Bartels M, Schweda AT, Dreikhausen U, Frank R, Resch K, Beil W, Nourbakhsh M
In:
Source: J Immunol
Publication Date: (2007)
Issue: 179(11): 7605-13
Research Area:
Cancer Research/Cell Biology
Immunotherapy / Hematology
Cells used in publication:
HeLa
Species: human
Tissue Origin: cervix
AGS
Species: human
Tissue Origin: stomach
Platform:
Nucleofector™ I/II/2b
Experiment
For each nucleofection 10^6 AGS or HeLa cells were harvested and resuspended in 100 µl nucleofector solution with 20 µg of synthetic fluorescin labeled peptides. The cell-peptide mixtures were nucleofected and cells were immediately transferred into 3 ml medium with 10% FCS for 3 h at 37°C. Immediately following nucleofection, HeLa cells were treated with IL-1-beta (10 ng/ml) and AGS cells were infected with 50 CFU of a cagA-positive H. pylori (HP87) per cell for following 3 h. Approximately 93 % of AGS and HeLa cells nucleofected with fluorescin-labeled peptide displayed positive signals, determined by FACS, compared with nonlabeled peptide.
Abstract
Selective inhibition of proinflammatory chemokines such as IL-8 is an important approach to combat inflammatory and infection diseases. Previous studies suggested that interaction of transcription factors NFkappaB repressing factor (NRF) and NFkappaB play a crucial role in activation of IL-8 gene expression. In a search for a specific inhibitor of IL-8 expression, we applied tandem affinity purification to investigate interaction of NRF and NFkappaB p65 in cells. We identified a synthetic peptide corresponding to aa 223-238 of NRF interfering with binding of endogenous p65 to NRF. Furthermore, nucleofection experiments were established to introduce this inhibitory peptide into the nucleus of IL-1 stimulated human cervical and Helicobacter pylori infected gastric epithelial cells. Our data clearly show that the specific peptide disturbing NRF/NFkappaB interaction is able to significantly decrease endogenous IL-8 gene transcription in response to IL-1 or Helicobacter pylori infection. Thus, our study provides novel insights into NRF and NFkappaB interaction in vivo and may facilitate the design of new anti-IL-8 drugs based on novel strategies.