Copper-mediated oxidative burst in Nicotiana tabacum L. cv. Bright Yellow 2 cell suspension cultures

In cell suspension cultures of Nicotiana tabacum L. cv. Bright Yellow 2 (BY-2) a rapid and concentration-dependent accumulation of H2O2 is induced by excess concentrations of copper (up to 100 muM). This specific and early response towards copper stress was shown to be extracellular. Addition of 300 U of catalase per ml decreased the level of H2O2. Superoxide dismutase (5 U/ml) induced an increase in H2O2 production by 22.2%. This indicates that at least part of the H2O2 is produced by dismutation of superoxide. Pretreatment of the cell cultures with the NAD(P)H oxidase inhibitors diphenylene iodonium (2 and 10 muM) and quinacrine (1 and 5 mM) prevented the generation of H2O2 under copper stress for 90%. The influence of the pH on the H2O2 production revealed the possible involvement of cell-wall-dependent peroxidases in the generation of reactive oxygen species after copper stress.