ABSTRACT
Aluminum (Al) causes programmed cell death (PCD) in plants. Our previous studies have confirmed that nitric oxide (NO) inhibits Al-induced PCD in the root tips of peanut. However, the mechanism by which NO inhibits Al-induced PCD is unclear. Here the effects of NO on mitochondrial reactive oxygen species (ROS), malondialdehyde (MDA), activities of superoxide dismutase (SOD) and ascorbate peroxidase (APX), expression of alternative oxidase (AhAOX) and cytochrome oxidase (AhCOX) were investigated in peanut (Arachis hypogaea L.) root tips treated with Al. The results showed that Al stress induced rapid accumulation of H2O2 and MDA and increased the ratio of SOD/APX. The up-regulation of AhAOX and AhCOX expressions was not enough to inhibit PCD occurrence. Sodium nitroprusside (SNP, a NO donor) decreased the ratio of SOD/APX and eliminated excess H2O2 and MDA, thereby inhibiting Al-induced PCD in the root tips of peanut. The expression of AhAOX and AhCOX was significantly enhanced in Al-induced PCD treated with SNP. But cPTIO (a NO specific scavenger) supply had the opposite effect. Taken together, these results suggested that lipid peroxidation induced by higher levels of H2O2 was an important cause of Al-induced PCD. NO-mediated inhibition of Al-induced PCD was related to a significant elimination of H2O2 accumulation by decreasing the ratio of SOD/APX and up-regulating the expression of AhAOX and AhCOX.
Subject(s)
Aluminum/toxicity , Antioxidants/metabolism , Apoptosis/drug effects , Arachis/metabolism , Nitric Oxide/metabolism , Ascorbate Peroxidases/metabolism , Cluster Analysis , Down-Regulation/drug effects , Electron Transport Complex IV/metabolism , Hydrogen Peroxide/metabolism , Lipid Peroxidation/drug effects , Malondialdehyde/metabolism , Mitochondria/drug effects , Mitochondria/metabolism , Nitroprusside/pharmacology , Plant Roots/metabolism , Reactive Oxygen Species/metabolism , Up-Regulation/drug effectsABSTRACT
Mitochondria play a crucial role in programmed cell death (PCD) in plants. In most cases of mitochondria-dependent PCD, cytochrome c (Cyt c) released from mitochondria due to the opening of mitochondrial permeability transition pore (MPTP) and the activation of caspase-like proteases. Here we describe the analytic methods of mitochondrial markers of PCD including mitochondria isolation, mitochondrial membrane permeability, mitochondrial inner membrane potential, Cytc release, ATP, and mitochondrial reactive oxygen species (ROS).
Subject(s)
Apoptosis , Biomarkers , Mitochondria/genetics , Mitochondria/metabolism , Adenosine Triphosphate/metabolism , Arachis/genetics , Arachis/metabolism , Cell Membrane Permeability , Cytochromes c/metabolism , Hydrogen Peroxide/metabolism , Membrane Potential, Mitochondrial , Plant Roots/genetics , Plant Roots/metabolism , Reactive Oxygen Species/metabolismABSTRACT
Aluminum (Al) stress alters nitric oxide (NO) and induces programmed cell death (PCD) in plants. Recent study has shown that NO inhibits Al-induced PCD. However, the mechanism of NO inhibiting Al-induced PCD has not been revealed yet. Here, we investigated the behavior of mitochondria during Al-induced PCD suppressed by NO in peanut. Seedlings of peanut was grown hydroponically in a controllable growth room. The mitochondrial physiological parameters were determined spectrophotometrically. The expression of AhANT and AhHsp70 was determined by quantitative RT-PCR. Al-induced cell death rapidly in peanut root tips is mitochondria-dependent PCD. There was a significantly negative relationship between PCD and mitochondrial NO/H2O2 level. Compared with Al treatment alone, the addition of NO donor sodium nitroprusside (SNP) increased the ratio of NO/H2O2, down-regulated AhANT expression and inhibited the opening of mitochondrial permeability transition pore (MPTP), up-regulated AhHsp70 expression and increased mitochondrial inner membrane potential (ΔΨm), reduced cytochrome c (Cyt c) release from mitochondria and caspase 3-like protease activity, while the effect of NO specific scavenger cPTIO supplement was opposite. NO suppresses Al-induce PCD in peanut root tips by improving mitochondrial physiological properties.
Subject(s)
Aluminum/pharmacology , Arachis/cytology , Arachis/drug effects , Mitochondria/drug effects , Nitric Oxide/metabolism , Plant Roots/cytology , Plant Roots/drug effects , Arachis/metabolism , Cell Death/drug effects , Mitochondria/metabolism , Plant Roots/metabolismABSTRACT
It had been reported that Aluminum (Al) stress altered nitric oxide (NO) concentration and induced programmed cell death (PCD) in plants. However, the relationship between NO and PCD occurrence under Al stress is unclear. The results showed that cell death induced by Al was significant negative correlation with the inhibition of Al on root elongation growth in peanut. AlCl3 at 100µmolL-1 induced DNA ladder, chromatin condensation, typical apoptotic chromatin condensation staining with DAPI, apoptosis related gene Hrs203j expression and caspase3-like protease activation in peanut root tip cells, and showed that Al-induced cell death in peanut root tip cells was a typical PCD. Exogenous NO donor sodium nitroprusside (SNP) at 200µmolL-1 inhibited Al-induced PCD occurrence, but NO specific scavenger cPTIO aggravated PCD production. It suggests that NO is a negative regulator of Al-induced PCD in peanut root tips.
Subject(s)
Aluminum/toxicity , Apoptosis/drug effects , Arachis/drug effects , Nitric Oxide Donors/pharmacology , Nitric Oxide/physiology , Nitroprusside/pharmacology , Plant Roots/drug effects , Adaptation, Physiological , Apoptosis/genetics , Arachis/genetics , Arachis/physiology , Caspase 3/metabolism , Chromatin/metabolism , Enzyme Activation , Genes, Plant , Plant Roots/cytology , Plant Roots/growth & developmentABSTRACT
BACKGROUND: Although many studies suggested that aluminum (Al) induced programmed cell death (PCD) in plants, the mechanism of Al-induced PCD and its effects in Al tolerance is limited. This study was to investigate the mechanism and type of Al induced PCD and the relationship between PCD and Al tolerance. RESULTS: In this study, two genotypes of peanut 99-1507 (Al tolerant) and ZH2 (Al sensitive) were used to investigate Al-induced PCD. Peanut root growth inhibition induced by AlCl3 was concentration and time-dependent in two peanut varieties. AlCl3 at 100 µM could induce rapidly peanut root tip PCD involved in DNA cleavage, typical apoptotic chromatin condensation staining with DAPI, apoptosis related gene Hrs203j expression and cytochrome C (Cyt c) release from mitochondria to cytosol. Caspase3-like protease was activated by Al; it was higher in ZH2 than in 99-1507. Al increased the opening of mitochondrial permeability transition pore (MPTP), decreased inner membrane potential (ΔΨm) of mitochondria. Compared with the control, Al stress increased O2â¢- and H2O2 production in mitochondria. Reactive oxygen species (ROS) burst was produced at Al treatment for 4 h. CONCLUSIONS: Al-induced PCD is earlier and faster in Al-sensitive peanut cultivar than in Al-tolerant cultivar. There is a negative relationship between PCD and Al resistance. Mitochondria- dependence PCD was induced by Al and ROS was involved in this process. The mechanism can be explained by the model of acceleration of senescence under Al stress.
