|
Iron is an important component of variety proteins in the cells of higher plants. For example, it participates in photosynthesis and respiration. On the other side, it is well known that excess iron was able to lead to oxidative stress, which may destroy the structure of lipid, protein and DNA, affecting plant growth. Heme oxygenase (HO) is one of the enzymes responsible for maintaining stability, and exhibiting cytoprotection roles. Meanwhile, reduced glutathione (GSH) is the major intracellular anti-oxidant substances, which could alleviate the oxidative damage of plants. Thus, this study was to explore the inter-relationship between HO and GSH under iron stressed condition, and its possible antioxidant mechanism.The results demonstrated that iron stress resulted in the increased iron and lipid peroxidation in a dose-dependence manner in detached rice leaves. When treated with 0,50, 100, and 500μmol L-1 Fe-citrate (Fe3+), HO-1 gene expression and its protein expression were increased along with the increase of iron concentrations. For example,500μmol L-1 Fe3+ treatment for 9 h led to the significant induction of lipid peroxidation and HO activity (P<0.05). However, high concentration of Fe3+(1000μmol L-1) not only blocked the HO activity, but also down-regulated the HO-1 gene expression and its protein expression, showing the lower level in comparison with those of the control sample (Con). In the following experiment,500μmol L-1 Fe3+ was selected.Further study also found that the GSH contents were negatively correlated with the concentrations of Fe3+ applied. Interestingly, the inducible effects of Fe3+ on HO activity, HO-1 gene expression, and its protein expression were alleviated by the pretreatment of GSH. Furthermore, the effect of GSH was significantly blocked by GSH synthesis inhibitor buthionine-sulfoxine (BSO). Further experiments showed that adding of HO-1 inducer Hemin pretreatment for 3 h could not only reverse the decreased GSH content, but also block the increased HO activity, HO-1 transcripts and its protein expression in detached rice leaves under Fe3+-stressed condition, thus resulting in the enhancement of antioxidant capacity.In addition, the changes of antioxidant enzymes activities showed that Fe3+ stress might lead to the increase of ascorbate peroxidase (APX), and superoxide dismutase (SOD) activities in detached rice leaves. Furthermore, Fe3+-induced increase of APX activity was significantly reversed when pretreated with Hemin. However, glutathione reductase (GR), and glutathione peroxidase (GPX) activities were increased. Whereas, glutathione S-transferase (GST) activity was down-regulated. Above results were further confirmed by the detection of hydrogen peroxide (H2O2) in detached rice leaves by staining in vivo.Taken together, combined pharmacological with molecular approaches, the effects of Fe3+ stress on detached rice leaves illustrated that there might existing the inter-relationship between HO and GSH, both of which were participating in the tolerance against iron toxicity in detached rice leaves.
|