Rapid inactivation of chloroplastic ascorbate peroxidase is responsible for oxidative modification to Rubisco in tomato (Lycopersicon esculentum) under cadmium stress.
To investigate the sensitive site of antioxidant systems in chloroplast under cadmium stress and its consequence on reactive oxygen species production and action, the sub-organellar localization of chloroplast superoxide dismutases (SOD, EC 1.15.1.1) and ascorbic peroxidase (APX, EC 1.11.1.11) isoenzymes and changes of enzymes activities under cadmium stress were investigated in tomato seedlings. Two APX isoforms, one thylakoid-bound and one stromal, were detected. Cd at 50 microM induced a moderate increase of SOD activities but a rapid inactivation of both APX isoenzymes. APX inactivation was mainly related to the decrease of ascorbate concentration, as supported by in vitro treatment of exogenous ascorbate and APX kinetic properties under Cd stress. H2O2 accumulation in chloroplast, as a consequence of APX inactivation, was associated with a 60% loss of Rubisco (EC 4.1.1.39) activity, which could be partially accounted for by a 10% loss of Rubisco content. Protein oxidation assay found that the Rubisco large subunit was the most prominent carbonylated protein; the level of carbonylated Rubisco large subunit increased fivefold after Cd exposure. Thiol groups in the Rubisco large subunit were oxidized, as indicated by non-reducing electrophoresis. Treating crude extract with H2O2 resulted in a similar pattern of protein oxidation and thiols oxidation with that observed in Cd-treated plants. Our study indicates that APXs in the chloroplast is a highly sensitive site of antioxidant systems under Cd stress, and the inactivation of APX could be mainly responsible for oxidative modification to Rubisco and subsequent decrease in its activity.
Liu
Kai-Lang
KL
College of Food Science and Nutritional Engineering, China Agricultural University, Beijing 100083, China.
Shen
Lin
L
Wang
Jia-Qi
JQ
Sheng
Ji-Ping
JP
D060387:Ascorbate Peroxidases; D001205:Ascorbic Acid / Q000378:metabolism / Q000494:pharmacology; D015415:Biomarkers / Q000378:metabolism; D002104:Cadmium / Q000633:toxicity*; D002736:Chloroplasts / Q000187:drug effects* / Q000201:enzymology*; D004789:Enzyme Activation / Q000187:drug effects; D006861:Hydrogen Peroxide / Q000378:metabolism / Q000494:pharmacology; D007150:Immunohistochemistry; D007527:Isoenzymes; D007700:Kinetics; D018551:Solanum lycopersicum / Q000187:drug effects* / Q000201:enzymology*; D010084:Oxidation-Reduction / Q000187:drug effects; D010544:Peroxidases / Q000302:isolation & purification / Q000378:metabolism*; D010940:Plant Proteins / Q000378:metabolism; D021122:Protein Subunits / Q000378:metabolism; D021381:Protein Transport / Q000187:drug effects; D012273:Ribulose-Bisphosphate Carboxylase / Q000378:metabolism*; D013438:Sulfhydryl Compounds / Q000378:metabolism; D013482:Superoxide Dismutase / Q000378:metabolism
D015415:Biomarkers; D007527:Isoenzymes; D010940:Plant Proteins; D021122:Protein Subunits; D013438:Sulfhydryl Compounds; D002104:Cadmium; D006861:Hydrogen Peroxide; D010544:Peroxidases; D060387:Ascorbate Peroxidases; D013482:Superoxide Dismutase; D012273:Ribulose-Bisphosphate Carboxylase; D001205:Ascorbic Acid