title: Production of reactive oxygen species, alteration of cytosolic ascorbate peroxidase, and impairment of mitochondrial metabolism are early events in heat shock-induced programmed cell death in tobacco Bright-Yellow 2 cells.
abstract: To gain some insight into the mechanisms by which plant cells die as a result of abiotic stress, we exposed tobacco (Nicotiana tabacum) Bright-Yellow 2 cells to heat shock and investigated cell survival as a function of time after heat shock induction. Heat treatment at 55 degrees C triggered processes leading to programmed cell death (PCD) that was complete after 72 h. In the early phase, cells undergoing PCD showed an immediate burst in hydrogen peroxide (H2O2) and superoxide (O2*-) anion production. Consistently, death was prevented by the antioxidants ascorbate (ASC) and superoxide dismutase (SOD). Actinomycin D and cycloheximide, inhibitors of transcription and translation, respectively, also prevented cell death, but with a lower efficiency. Induction of PCD resulted in gradual oxidation of endogenous ASC; this was accompanied by a decrease in both the amount and the specific activity of the cytosolic ASC peroxidase (cAPX). A reduction in cAPX gene expression was also found in the late PCD phase. Moreover, changes of cAPX kinetic properties were found in PCD cells. Production of ROS in PCD cells was accompanied by early inhibition of glucose (Glc) oxidation, with a strong impairment of mitochondrial function as shown by an increase in cellular NAD(P)H fluorescence, and by failure of mitochondria isolated from cells undergoing PCD to generate membrane potential and to oxidize succinate in a manner controlled by ADP. Thus, we propose that in the early phase of tobacco Bright-Yellow 2 cell PCD, ROS production occurs, perhaps because of damage of the cell antioxidant system, with impairment of the mitochondrial oxidative phosphorylation.
authors.affiliation: Istituto di Biomembrane e Bioenergetica, Consiglio Nazionale delle Ricerche, Via Amendola 165/A, I-70126 Bari, Italy.
authors.forename: Rosa Anna
authors.initials: RA
authors.lastname: Vacca
authors.forename: Maria Concetta
authors.initials: MC
authors.lastname: de Pinto
authors.forename: Daniela
authors.initials: D
authors.lastname: Valenti
authors.forename: Salvatore
authors.initials: S
authors.lastname: Passarella
authors.forename: Ersilia
authors.initials: E
authors.lastname: Marra
authors.forename: Laura
authors.initials: L
authors.lastname: De Gara
mesh_terms: D017209:Apoptosis / Q000187:drug effects; D060387:Ascorbate Peroxidases; D001205:Ascorbic Acid / Q000378:metabolism; D001483:Base Sequence; D002460:Cell Line; D003513:Cycloheximide / Q000494:pharmacology; D003600:Cytosol / Q000201:enzymology; D018744:DNA, Plant / Q000235:genetics; D003609:Dactinomycin / Q000494:pharmacology; D004734:Energy Metabolism; D016166:Free Radical Scavengers / Q000494:pharmacology; D018869:Heat-Shock Response; D007700:Kinetics; D008928:Mitochondria / Q000378:metabolism; D010544:Peroxidases / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D018749:RNA, Plant / Q000235:genetics / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism; D014026:Nicotiana / Q000166:cytology* / Q000235:genetics / Q000378:metabolism*
chemical_list: D018744:DNA, Plant; D016166:Free Radical Scavengers; D012333:RNA, Messenger; D018749:RNA, Plant; D017382:Reactive Oxygen Species; D003609:Dactinomycin; D003513:Cycloheximide; D010544:Peroxidases; D060387:Ascorbate Peroxidases; D001205:Ascorbic Acid