The Effect Of Cd And Cd+High Temperature Stress To The Growth And The Antioxidant System Of The Double Antioxidase Transgenic Rice

Rice is one of the most important crops in the world. It feeds about a half of the population.Both heavy metal pollution and heat stresses have an enormous impact on rice growth and productivity. Therefore, using genetic engineering means to improve rice resistance to heavy metal and heat stresses, and increase rice yield is significant.The rice (Oryza sativa L. cv Zhonghua No.11) and its transgenics co-expressing both GST (glutathione-S-transferase) and CAT1 (catalase) was used as the material of this experiment. The 6-week-old rice seedlings were treated with Cd (NO3)2 and Cd+high temperature (38℃) stresses, respectively, for 0-3 days, and the physiological characters were determined. The experimental results showed that Cd and high temperature poison to the rice seedlings became worse and worse with the prolongation of treatment time, but the height of seedlings, the length and quantity of roots, the relative water content, chlorophyll content, soluble sugar content, net photosynthetic rate, the vigor of root system and so on were all higher in transgenics than in non-transgenics. Whereas less MDA and H2O2 generated in transgenic rice compared to the notransgenics.A comparative analysis on variation of the antioxidant system, in particular the ascorbate-glutathione cycle and some physiological parameters, was performed in transgenic rice (Oryza sativa L. cv Zhonghua No.11) coexpressing catalase (CAT, EC 1.11.1.6) and glutathione-S-transferase (GST, EC 2.5.1.18) and nontransgenic rice under cadmium (Cd) stress condition. Significant different modifications, either at the level or/and the trend, of the antioxidant system particularly the overall ascorbate-glutathione cycle in transgenics from those of nontransgenics were found upon Cd exposure. Enhancement of CAT activity was obtained in transgenics, but inhibition occurred in nontransgenics after 2 days of Cd stress, in comparison with the control. Monodehydroascorbate reductase (MDAR, EC 1.6.5.4) showed marked stimulation after 2 days of Cd treatment in transgenics compared with nontransgenics and the control. Activities of the dehydroascorbate reductase (DHAR, EC 1.8.5.1) and superoxide dismutase (SOD, EC 1.15.1.1) in transgenics were much higher than those in nontransgenics after 3 days of Cd exposure. Furthermore, the levels of ascorbate peroxidase (APX, EC 1.11.1.11), glutathione reductase (GR, EC 1.6.4.2) activity, glutathione (GSH) content and ascorbate (ASA)/(ASA+ dehydroascorbate, DHA) ratio were all higher in transgenics than those in nontransgenics, despite these components changed in the similar manner in all stressed plants.The combination of Cd and heat stress resulted in more reduced growth and increased malon dialdehyde (MDA) and H2O2 in both transgenics and nontransgenics, compared to those of Cd alone. Distinctly different modifications, either at the level or/and the trend, of the antioxidant system particularly the overall ascorbate-glutathione cycle in transgenics treated with combined Cd and heat stress from those of Cd alone were found. One striking feature was that dramatically enhancement of CAT and glutathione reductase (GR, EC.1.6.4.2) activities were obtained upon the combined stress in transgenics, in contrast, both of them inhibited by single Cd. Another marked characteristic observed in transgenics was that the monodehydroascorbate reductase (MDAR, EC.1.6.5.4), superoxide dismutase (SOD, EC.1.15.1.1) and GST showed marked stimulation after the combined stress treatment, compared to those of Cd alone and the control, despite these enzymes changed in the similar manner in both stress conditions. Importantly, in transgenics the ascorbate (ASA)/(ASA+dehydroascorbate, DHA) ratio was also stimulated by the combination of Cd and heat stress, compared to that of single Cd. Furthermore the levels of CAT, SOD, GR, GST, MDAR and ASA/(ASA+DHA) ratio were all much higher in transgenics than in nontransgenics under the combined stress condition, regardless their change trend. These results evidenced that the stress combination was a more complex trait, and posed greater toxicity to plants compared to the individual Cd, but not simply the sum of these two different stresses. Moreover, the responses of antioxidant system to Cd and to the combined Cd and heat stress application followed very different patterns in the transgenics and nontransgenics.In general, the experimental data showed that co-expression of the GST and CAT1 in transgenic rice could increase Cd and combined Cd and high temperature stress tolerance, which provide a theoretical basis for rice breeding resistance to heavy metal and heat stresses. Taken together, our data showed that the single Cd and the combined Cd and heat stresses induced alterations of the overall ascorbate-glutathione cycle were more in transgenics than in nontransgenics, which might be due to the coexpression of CAT and GST influencing the levels of signal molecules such as reactive oxygen species (ROS) and GSH as well as the ASA redox state. The less oxidative damage caused by Cd and the combined Cd and heat stresses in the transformants might not only come from the coexpression of CAT and GST, but also from the cooperation of the function-related components, at least in ASA-GSH cycle, of the antioxidant network.