Inhibition of NF-kappaB by a TAT-NEMO-binding domain peptide accelerates constitutive apoptosis and abrogates LPS-delayed neutrophil apoptosis.
Delivery of biologically active peptides into human polymorphonuclear neutrophils (PMNs) has implications for studying cellular functions and may be therapeutically relevant. The transcription factor nuclear factor-kappaB (NF-kappaB) regulates the expression of multiple genes controlling inflammation, proliferation, and cell survival. PMNs play a crucial role in first-line defense. Targeting NF-kappaB in these cells may promote apoptosis and therefore facilitate resolution of inflammation. We used an 11-amino acid sequence NEMO-binding domain (NBD) that selectively inhibits the IKKgamma (NEMO)/IKKbeta interaction, preventing NF-kappaB activation. An HIV-TAT sequence served as a highly effective transducing shuttle. We show that lipopolysaccharide (LPS), granulocyte-macrophage colony-stimulating factor (GM-CSF), and dexamethasone (DEX) significantly reduced apoptosis after 20 hours. LPS, but not GM-CSF or DEX, activated NF-kappaB as shown by IkappaBalpha degradation, NF-kappaB DNA binding, and transcriptional activity. The TAT-NBD blocked LPS-induced NF-kappaB activation and NF-kappaB-dependent gene expression. TAT-NBD accelerated constitutive PMN apoptosis dose dependently and abrogated LPS-delayed apoptosis. These results provide a proof of principle for peptide delivery by TAT-derived protein transduction domains to specifically inhibit NF-kappaB activity in PMNs. This strategy may help in controlling various cellular functions even in short-lived, transfection-resistant primary human cells.
Choi
Mira
M
Division of Nephrology, Franz Volhard Clinic, Medical Faculty of the Charité, Humboldt Univertsity of Berlin, Wiltbergstrasse 50, 13122 Berlin, Germany.
Rolle
Susanne
S
Wellner
Maren
M
Cardoso
M Cristina
MC
Scheidereit
Claus
C
Luft
Friedrich C
FC
Kettritz
Ralph
R
D017209:Apoptosis / Q000187:drug effects* / Q000276:immunology; D001665:Binding Sites / Q000502:physiology; D003907:Dexamethasone / Q000494:pharmacology; D015696:Gene Products, tat / Q000737:chemistry; D005938:Glucocorticoids / Q000494:pharmacology; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000494:pharmacology; D006801:Humans; D051550:I-kappa B Kinase; D008070:Lipopolysaccharides / Q000494:pharmacology*; D016328:NF-kappa B / Q000037:antagonists & inhibitors*; D009504:Neutrophils / Q000166:cytology* / Q000187:drug effects; D010446:Peptide Fragments / Q000494:pharmacology*; D017346:Protein Serine-Threonine Kinases / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology
D015696:Gene Products, tat; D005938:Glucocorticoids; D008070:Lipopolysaccharides; D016328:NF-kappa B; D010446:Peptide Fragments; D003907:Dexamethasone; D016178:Granulocyte-Macrophage Colony-Stimulating Factor; D017346:Protein Serine-Threonine Kinases; C496559:CHUK protein, human; D051550:I-kappa B Kinase; C496560:IKBKB protein, human; C496565:IKBKE protein, human