title: NF-kappaB inhibition reveals differential mechanisms of TNF versus TRAIL-induced apoptosis upstream or at the level of caspase-8 activation independent of cIAP2.
abstract: Death ligands not only activate a death program but also regulate inflammatory signalling pathways, for example, through NF-kappaB induction. Although tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) and TNF both activate NF-kappaB in human keratinocytes, only TRAIL potently induces apoptosis. However, when induction of NF-kappaB was inhibited with a kinase dead IKK2 mutant (IKK2-KD), TNF- but not TRAIL-induced apoptosis was dramatically enhanced. Acquired susceptibility to TNF-induced apoptosis was due to increased caspase-8 activation. To investigate the mechanism of resistance of HaCaT keratinocytes to TNF-induced apoptosis, we analyzed a panel of NF-kappaB-regulated effector molecules. Interestingly, the inhibitor of apoptosis protein (IAP) family member cIAP2, but not cIAP1, X-linked inhibitor of apoptosis, TNF receptor-associated factor (TRAF)-1, or TRAF2, was downregulated in sensitive but not in resistant HaCaT keratinocytes. Surprisingly, however, stable inducible expression of cIAP2 was not sufficient to render IKK2-KD-sensitized keratinocytes resistant to TNF, and reduction of cIAP2 alone did not increase the sensitivity of HaCaT keratinocytes to TNF. In conclusion, we demonstrate that inhibition of NF-kappaB dramatically sensitizes human keratinocytes to TNF- but not to TRAIL-induced apoptosis and that this sensitization for TNF was largely independent of cIAP2. Our data thus clearly exclude the candidates proposed to date to confer TNF apoptosis resistance and suggest the function of an unanticipated effector of NF-kappaB critical for the survival of HaCaT keratinocytes upstream or at the level of caspase-8 activation.
authors.affiliation: Laboratory for Experimental Dermatology, Department of Dermatology and Venerology, Otto-von-Guericke-University Magdeburg, Magdeburg, Germany.
authors.forename: Philip
authors.initials: P
authors.lastname: Diessenbacher
authors.forename: Mike
authors.initials: M
authors.lastname: Hupe
authors.forename: Martin R
authors.initials: MR
authors.lastname: Sprick
authors.forename: Andreas
authors.initials: A
authors.lastname: Kerstan
authors.forename: Peter
authors.initials: P
authors.lastname: Geserick
authors.forename: Tobias L
authors.initials: TL
authors.lastname: Haas
authors.forename: Tina
authors.initials: T
authors.lastname: Wachter
authors.forename: Manfred
authors.initials: M
authors.lastname: Neumann
authors.forename: Henning
authors.initials: H
authors.lastname: Walczak
authors.forename: John
authors.initials: J
authors.lastname: Silke
authors.forename: Martin
authors.initials: M
authors.lastname: Leverkus
mesh_terms: D017209:Apoptosis / Q000502:physiology; D000075747:Baculoviral IAP Repeat-Containing 3 Protein; D053446:CASP8 and FADD-Like Apoptosis Regulating Protein / Q000378:metabolism; D053181:Caspase 8 / Q000378:metabolism*; D002461:Cell Line, Transformed; D002470:Cell Survival / Q000502:physiology; D006801:Humans; D051550:I-kappa B Kinase / Q000235:genetics / Q000378:metabolism; D051034:Inhibitor of Apoptosis Proteins / Q000378:metabolism*; D015603:Keratinocytes / Q000166:cytology / Q000378:metabolism*; D016296:Mutagenesis; D016328:NF-kappa B / Q000037:antagonists & inhibitors / Q000378:metabolism*; D015398:Signal Transduction / Q000502:physiology; D047990:TNF Receptor-Associated Factor 1 / Q000378:metabolism; D047992:TNF Receptor-Associated Factor 2 / Q000378:metabolism; D053221:TNF-Related Apoptosis-Inducing Ligand / Q000378:metabolism*; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism*; D044767:Ubiquitin-Protein Ligases
chemical_list: D053446:CASP8 and FADD-Like Apoptosis Regulating Protein; C506386:CFLAR protein, human; D051034:Inhibitor of Apoptosis Proteins; D016328:NF-kappa B; D047990:TNF Receptor-Associated Factor 1; D047992:TNF Receptor-Associated Factor 2; D053221:TNF-Related Apoptosis-Inducing Ligand; C505653:TNFSF10 protein, human; D014409:Tumor Necrosis Factor-alpha; C494832:BIRC3 protein, human; D000075747:Baculoviral IAP Repeat-Containing 3 Protein; D044767:Ubiquitin-Protein Ligases; D051550:I-kappa B Kinase; D053181:Caspase 8