title: Chelerythrin activates caspase-8, downregulates FLIP long and short, and overcomes resistance to tumour necrosis factor-related apoptosis-inducing ligand in KG1a cells.
abstract: KG1a cells (CD34+/38-) express FAS and TRAIL (tumour-necrosis-factor-related apoptosis-inducing ligand) receptors but are resistant to FAS-ligand and TRAIL/APO2-L (apoptosis antigen-2 ligand)-induced apoptosis. KG1a cells are sensitized to FAS-induced apoptosis by chelerythrin, an inhibitor of protein kinase C (PKC). As cytoplasmatic adaptor molecules of FAS, e.g. FLIP [Fas-associated death domain protein (FADD)-like interleukin 1 beta-converting enzyme [FLICE (caspase-8)-inhibitory protein]], also modulate TRAIL signals, we determined whether chelerythrin affected TRAIL-mediated apoptosis. Chelerythrin by itself induced apoptosis in KG1a cells, and apoptosis was associated with activation of caspase-8. While TRAIL alone failed to activate caspase-8 or induce apoptosis, the addition of TRAIL to chelerythrin-treated cells significantly enhanced cleavage of caspase-8 and apoptosis. Chelerythrin-pretreated KG1a cells showed decreased phosphorylation of protein kinase C (PKC)-zeta and downregulation of both FLIP long and FLIP short proteins. Downregulation of FLIP and induction of apoptosis were partially abrogated by pretreatment with the specific caspase-8 inhibitor, Z-IETD-FMK. The decrease in FLIP protein expression induced by chelerythrin was accompanied by a progressive increase in mRNA levels of both FLIP long and FLIP short. CD34+ precursors from normal human marrow were also sensitive to chelerythrin but, in contrast to KG1a cells, were not sensitized to TRAIL-mediated apoptosis. Thus, resistance to TRAIL-induced apoptosis in leukaemic KG1a cells but not in normal CD34+ precursors was overcome in the presence of chelerythrin. The mechanism appeared to involve inhibition of PKC. Central targets were FLIP long and FLIP short, and their interactions with caspase-8. Whether such a pathway can be exploited to selectively target leukaemic progenitor cells remains to be determined.
authors.affiliation: Fred Hutchinson Cancer Research Center, School of Medicine, Seattle, WA, USA. platzbecker@mk1.med.tu-dresden.de
authors.forename: Uwe
authors.initials: U
authors.lastname: Platzbecker
authors.forename: Jessica L
authors.initials: JL
authors.lastname: Ward
authors.forename: H Joachim
authors.initials: HJ
authors.lastname: Deeg
mesh_terms: D000470:Alkaloids; D017209:Apoptosis; D051017:Apoptosis Regulatory Proteins; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D053119:Benzophenanthridines; D015153:Blotting, Western; D053181:Caspase 8; D053453:Caspase 9; D020169:Caspases / Q000378:metabolism*; D002460:Cell Line; D004789:Enzyme Activation; D018922:HL-60 Cells; D006801:Humans; D019169:Jurkat Cells; D015473:Leukemia, Promyelocytic, Acute / Q000188:drug therapy* / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000378:metabolism*; D010617:Phenanthridines / Q000494:pharmacology*; D016133:Polymerase Chain Reaction; D011233:Precipitin Tests; D011493:Protein Kinase C / Q000037:antagonists & inhibitors*; D012333:RNA, Messenger / Q000032:analysis; D053221:TNF-Related Apoptosis-Inducing Ligand; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism*
chemical_list: D000470:Alkaloids; D051017:Apoptosis Regulatory Proteins; D001426:Bacterial Proteins; D053119:Benzophenanthridines; D008562:Membrane Glycoproteins; D010617:Phenanthridines; D012333:RNA, Messenger; D053221:TNF-Related Apoptosis-Inducing Ligand; C505653:TNFSF10 protein, human; D014409:Tumor Necrosis Factor-alpha; C074890:FliP protein, Bacillus; C016299:chelerythrine; D011493:Protein Kinase C; C505530:CASP8 protein, human; C506433:CASP9 protein, human; D053181:Caspase 8; D053453:Caspase 9; D020169:Caspases