Activation of caspase-like proteases and induction of apoptosis by isopentenyladenosine in tobacco BY-2 cells.
The effects of isopentenyladenosine (iPA) on tobacco (Nicotiana tabacum L.) BY-2 cells were examined. The number of BY-2 cells decreased in a time- and concentration-dependent manner after being exposed to micromolar concentrations of iPA. This decrease was mainly due to a loss of cell viability, since no substantial changes in cell cycle progression were revealed by flow-cytometric analysis. Dying cells exhibited the typical morphological and biochemical hallmarks of apoptosis, including cell shrinkage, chromatin condensation, and degradation of nuclear DNA to nucleosomal size fragments. Caspase-1-like and caspase-3-like proteases also became activated, the former being dominant. Inhibitor-sensitivity studies revealed that although synthetic caspase inhibitors failed to prevent cell death they markedly reduced cell death in tobacco BY-2 cells, Nu-benzyloxycarbonyl-Val-Ala-Asp-fluoromethylketone, a specific inhibitor for caspase-1, being the most effective. Our results indicate that caspase-like proteases, and particularly caspase-1-like protease, might be critically implicated in iPA-induced apoptosis of BY-2 cells. Finally, the outcome of inhibiting adenosine kinase by 4-amino-3-iodo-1(beta- D-ribofuranosyl)pyrazolo[3,4-d]-pyrimidine revealed that intracellular phosphorylation of iPA is required for its cytotoxicity to develop.
Mlejnek
Petr
P
Department of Botany and Plant Physiology, Mendel University of Agriculture and Forestry Brno, Zemedelská 1, 613 00 Brno, Czech Republic. mlejnek@mendelu.cz
Procházka
Stanislav
S
D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects*; D020170:Caspase 1 / Q000378:metabolism; D053148:Caspase 3; D061945:Caspase Inhibitors; D020169:Caspases / Q000378:metabolism*; D002478:Cells, Cultured; D003583:Cytokinins / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D010450:Endopeptidases / Q000378:metabolism; D004789:Enzyme Activation / Q000187:drug effects; D007541:Isopentenyladenosine / Q000494:pharmacology*; D009842:Oligopeptides / Q000494:pharmacology; D010937:Plant Growth Regulators / Q000494:pharmacology; D014026:Nicotiana / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism
D000590:Amino Acid Chloromethyl Ketones; D061945:Caspase Inhibitors; D003583:Cytokinins; D009842:Oligopeptides; D010937:Plant Growth Regulators; C110772:benzoylcarbonyl-aspartyl-glutamyl-valyl-aspartyl-fluoromethyl ketone; C096713:benzyloxycarbonylvalyl-alanyl-aspartyl fluoromethyl ketone; D007541:Isopentenyladenosine; D010450:Endopeptidases; D053148:Caspase 3; D020169:Caspases; D020170:Caspase 1