Zhiguang Zhao

Effects of silicon on defense of wheat against oxidative stress under drought at different developmental stages

The effects of silicon application before sowing on the drought-induced oxidative stress and antioxidant defense in wheat (Triticum aestivum L.) were investigated. Drought stress was applied by withholding watering till sampling at booting or filling stage. Application of Si increased the water potential of drought-stressed plants at filling stage, whereas it did not at booting stage. The superoxide dismutase (SOD) activity was inhibited and peroxidase (POD) activity was enhanced by drought at booting stage, and no differences were observed due to the Si treatment. At filling stage, however, application of Si increased the SOD activity and decreased the POD activity of drought-stressed plants. The catalase (CAT) activity was slightly increased by drought only in the absence of Si and at booting stage. The activity of glutathione reductase (GR) was not greatly influenced. Application of Si did not change the contents of H2O2, total soluble protein and protein carbonyl of drought-stressed plants at booting stage, whereas at filling stage, it decreased the content of H2O2 and protein carbonyl and increased the content of total soluble protein. The content of thiobarbituric acid reactive substances (TBARS) and the activities of acid phospholipase (AP) and lipoxygenase (LOX) in drought-stressed plants were also decreased by application of Si at both stages.

Evidence for the involvement of nitric oxide and reactive oxygen species in osmotic stress tolerance of wheat seedlings: Inverse correlation between leaf abscisic acid accumulation and leaf water loss

Nitric oxide (NO) and reactive oxygen species (ROS) play important roles in both abscisic acid (ABA) signaling and stress-induced ABA accumulation. However, little is known about their physiological roles in the whole plant. In this study, the effects of NO and ROS on leaf water control and the roles of ABA were determined using wheat (Triticum aestivum L.) seedlings. As compared with the control, osmotic stress reduced leaf water loss (LWL) while it increased leaf ABA content. The effects of osmotic stress on LWL and ABA contents were partially reversed by NO scavengers or NO synthase (NOS) inhibitors. Furthermore, sodium nitroprusside (SNP) at concentrations between 0.01 and 10 mM all reduced LWL efficiently and induced ABA accumulation in a dose-dependent manner. When ABA synthesis was inhibited by fluridone or actidione, the effects of SNP on LWL were partially reversed. These results suggest that NO is involved in leaf water maintenance of wheat seedlings under osmotic stress, and one of the possible mechanisms is by stimulating ABA synthesis. The ROS scavengers used in our experiments had no effects on either LWL or ABA accumulation induced by osmotic stress. However, all ROS induced LWL reduction and ABA accumulation significantly. Hydrogen peroxide had the same effects as SNP on LWL and induced ABA accumulation in a dose-dependent manner but had a maximal effect at 1 mM. Fluridone reversed the effects of H2O2 on both LWL reduction and ABA accumulation, while actidione had no effect. These results suggest that ROS are also involved in leaf water maintenance of wheat seedlings by stimulating ABA biosynthesis, but with a different mechanism to that of NO. The ABA-independent mechanism in NO/ROS regulation of leaf water balance is discussed, in relation to our results.

Deep-sequencing transcriptome analysis of chilling tolerance mechanisms of a subnival alpine plant, Chorispora bungeana

BackgroundThe plant tolerance mechanisms to low temperature have been studied extensively in the model plant Arabidopsis at the transcriptional level. However, few studies were carried out in plants with strong inherited cold tolerance. Chorispora bungeana is a subnival alpine plant possessing strong cold tolerance mechanisms. To get a deeper insight into its cold tolerance mechanisms, the transcriptome profiles of chilling-treated C. bungeana seedlings were analyzed by Illumina deep-sequencing and compared with Arabidopsis.ResultsTwo cDNA libraries constructed from mRNAs of control and chilling-treated seedlings were sequenced by Illumina technology. A total of 54,870 unigenes were obtained by de novo assembly, and 3,484 chilling up-regulated and 4,571 down-regulated unigenes were identified. The expressions of 18 out of top 20 up-regulated unigenes were confirmed by qPCR analysis. Functional network analysis of the up-regulated genes revealed some common biological processes, including cold responses, and molecular functions in C. bungeana and Arabidopsis responding to chilling. Karrikins were found as new plant growth regulators involved in chilling responses of C. bungeana and Arabidopsis. However, genes involved in cold acclimation were enriched in chilling up-regulated genes in Arabidopsis but not in C. bungeana. In addition, although transcription activations were stimulated in both C. bungeana and Arabidopsis, no CBF putative ortholog was up-regulated in C. bungeana while CBF2 and CBF3 were chilling up-regulated in Arabidopsis. On the other hand, up-regulated genes related to protein phosphorylation and auto-ubiquitination processes were over-represented in C. bungeana but not in Arabidopsis.ConclusionsWe conducted the first deep-sequencing transcriptome profiling and chilling stress regulatory network analysis of C. bungeana, a subnival alpine plant with inherited cold tolerance. Comparative transcriptome analysis suggests that cold acclimation is not a major chilling tolerance mechanism of C. bungeana. Activation of protein phosphorylation and ubiquitination may confer chilling tolerance to C. bungeana in a more rapid and flexible way than cold acclimation. Such differences may have contributed to the differences in cold tolerance between C. bungeana and Arabidopsis. The results presented in this paper will be informative for gene discovery and the molecular mechanisms related to plant cold tolerance.

Abscisic acid is involved in brassinosteroids-induced chilling tolerance in the suspension cultured cells from Chorispora bungeana

The objective of this study was to investigate whether abscisic acid (ABA), a second messenger in chilling stress responses, is involved in brassinosteroids (BRs)-induced chilling tolerance in suspension cultured cells from Chorispora bungeana. The suspension cells were treated with 24-epibrassinolide (EBR), ABA, ABA biosynthesis inhibitor fluridone (Flu) and EBR in combination with Flu. Their effects on chilling tolerance, reactive oxygen species (ROS) levels and antioxidant defense system were analyzed. The results showed that EBR treatment markedly alleviated the decrease of cell viability and the increases of ion leakage and lipid peroxidation induced by chilling stress, suggesting that application of EBR could improve the chilling tolerance of C. bungeana suspension cultures. In addition, similar results were observed when exogenous ABA was applied. Treatment with Flu alone and in combination with EBR significantly suppressed cell viability and increased ion leakage and lipid peroxidation under low temperature conditions, indicating that the inhibition of ABA biosynthesis could decrease the chilling tolerance of C. bungeana suspension cultures and the EBR-enhanced chilling tolerance. Further analyses showed that EBR and ABA enhanced antioxidant defense and slowed down the accumulation of ROS caused by chilling. However, Flu application differentially blocked these protective effects of EBR. Moreover, EBR was able to mimic the effect of ABA by markedly increasing ABA content in the suspension cells under chilling conditions, whereas the EBR-induced ABA accumulation was inhibited by the addition of Flu. Taken together, these results demonstrate that EBR may confer chilling tolerance to C. bungeana suspension cultured cells by enhancing the antioxidant defense system, which is partially mediated by ABA, resulting in preventing the overproduction of ROS to alleviate oxidative injury induced by chilling.

Brassinosteroids regulate pectin methylesterase activity and AtPME41 expression in Arabidopsis under chilling stress

Pectin methylesterases (PMEs) are important cell wall enzymes that may play important roles in plant chilling/freezing tolerance. We investigated the possible roles of brassinosteroids (BRs) in regulation of PMEs under chilling stress. Chilling stress or 24-epibrassinolide (eBL) treatments induced significant increases in PME activity in wild type (Col-0) seedlings of Arabidopsis. The chilling-stress-induced increases in PME activity were also found in bzr1-D mutant, a BZR1 stabilized mutant with a constitutively active BR signaling pathway, but not in bri1-116, a BR insensitive null allele of the BR receptor BRI1. The results suggest that the regulation of PME activity in Arabidopsis under chilling stress depends on the BR signaling pathway. Furthermore, we showed that the effect of chilling stress on PME activity was impaired in pme41, a knockout mutant of AtPME41. Semi-quantitative RT-PCR results showed that expression of AtPME41 was induced by chilling stress in wild type plants but not in the bri1-116 mutant. The expression of AtPME41 increased in bzr1-D and eBL treated wild type seedlings, but decreased in bri1-116 seedlings. Furthermore, ion leakage induced by low temperature were dramatically increased in both bri1-116 and pme41, while lipid peroxidation was increased in bri1-116 only. The results suggest that BRs may modulate total PME activity in Arabidopsis under chilling stress by regulating AtPME41 expression. Regulation of PME activity may serve as one of the mechanisms that BR participates in chilling tolerance of plants.

Drought‐Stimulated Activity of Plasma Membrane Nicotinamide Adenine Dinucleotide Phosphate Oxidase and Its Catalytic Properties in Rice

The activity of plasma membrane (PM) nicotinamide adenine dinucleotide phosphate (NADPH) oxidase and its catalytic properties in rice was investigated under drought stress conditions. Drought stress led to decreased leaf relative water content (RWC) and, as a result of drought-induced oxidative stress, the activities of antioxidant enzymes increased significantly. More interestingly, the intensity of applied water stress was correlated with increased production of H2O2 and O2 (-) and elevated activity of PM NADPH oxidase, a key enzyme of reactive oxygen species generation in plants. Histochemical analyses also revealed increased H2O2 and O2 (-) production in drought-stressed leaves. Application of diphenylene iodonium (DPI), an inhibitor of PM NADPH oxidase, did not alleviate drought-induced production of H2O2 and O2 (-). Catalysis experiments indicated that the rice PM NADPH oxidase was partially flavin-dependent. The pH and temperature optima for this enzyme were 9.8 and 40 degrees C, respectively. In addition, drought stress enhanced the activity under alkaline pH and high temperature conditions. These results suggest that a complex regulatory mechanism, associated with the NADPH oxidase-H2O2 system, is involved in the response of rice to drought stress.

Molecular cloning and characterization of a γ-glutamylcysteine synthetase gene from Chorispora bungeana

Chorispora bungeana Fisch. and C.A. Mey (C. bungeana) is a rare alpine subnival plant species that is highly tolerant of freezing temperatures. Glutathione (GSH) is a major player in various metabolic processes involved in plant growth and development and stress responses. A recent study has found that the inhibition of GSH synthesis decreases the chilling tolerance of C. bungeana callus (J. Wu et al.: Cryobiology 57:9–17, 2008). We have isolated and characterized a full-length cDNA encoding γ-glutamylcysteine synthetase (CbγECS), the key enzyme of GSH synthesis, from the leaves of C. bungeana, with the aim of furthering our understanding of the role of GSH at the molecular level. CbγECS was found to encode a 524-amino acid protein with moderate to high nucleotide sequence similar to previously reported plant γ-ECS genes. CbγECS transcripts were detected in the calluses, roots and leaves of C. bungeana, but there was no tissue-specific expression. The transcripts of CbγECS accumulated both rapidly and at high levels when C. bungeana was treated with chilling (4°C), freezing (−4°C), several other environmental stresses (such as heat, salinity, osmotic and heavy metals), abscisic acid and hydrogen peroxide. These results suggest that CbγECS may participate in the cellular responses against multiple environmental stimuli in C. bungeana.

Inhibition of glutathione synthesis decreases chilling tolerance in Chorispora bungeana callus.

The possible roles of reduced glutathione (GSH) in chilling tolerance were studied in callus generated from a representative alpine plant, Chorispora bungeana Fisch. & C.A. Mey (C. bungeana). The callus grew well under low-temperature and chilling treatment led only to slight injury, as indicated by a low level of ion leakage (IL). Malondialdehyde measurements also were not elevated, however GSH was. Exogenously application of l-buthionine-(S R)-sulfoximine (BSO), an inhibitor of gamma-glutamylcysteine synthetase (gamma-ECS), arrested the GSH accumulation induced by chilling and resulted in a significant decrease in cell growth and an increase in IL and malondialdehyde. These results implied that C. bungeana is a plant with a strong low-temperature tolerance mechanism, and the tolerance of C. bungeana may be associated with GSH accumulation. Under chilling treatment, the proportion of unsaturated fatty acid in the plasma membrane (PM) increased significantly in callus of C. bungeana mainly due to increases in C18:2 and C18:3, the membrane fluidity (indicated by DPH fluorescent polarization) however was maintained. High PM H(+)-ATPase activities were also induced by chilling. Exogenously application of BSO blocked the effects of chilling treatments on the changes of fatty acids and PM H(+)-ATPase activities, reducing the PM membrane fluidity. On the other hand, simultaneous application of GSH and BSO to callus under chilling treatments reversed the effects of BSO on the changes of fatty acids, PM fluidity and PM H(+)-ATPase activities. These results suggested that GSH induced by low-temperature treatments may confer chilling tolerance to C. bungeana, probably by increasing unsaturated fatty acid compositions and maintaining PM fluidity and high enzymatic activity.

Anatomical and Chemical Characteristics of a Rolling Leaf Mutant of Rice and Its Ecophysiological Properties

The anatomical and chemical characteristics of a rolling leaf mutant (rlm) of rice (Oryza sativa L.) and its ecophysiological properties in photosynthesis and apoplastic transport were investigated. Compared with the wild type (WT), the areas of whole vascular bundles and xylem as well as the ratios of xylem area/whole vascular bundles area and xylem area/phloem area were higher in rlm, whereas the area and the width of foliar bulliform cell were lower. The Fourier transform infrared (FTIR) microspectroscopy spectra of foliar cell walls differed greatly between rlm and WT. The rlm exhibited lower protein and polysaccharide contents of foliar cell walls. An obvious reduction of pectin content was also found in rlm by biochemical measurements. Moreover, the rate of photosynthesis was depressed while the conductance of stoma and the intercellular CO2 concentration were enhanced in rlm. The PTS fluorescence, which represents the ability of apoplastic transport, was 11% higher in rlm than in WT. These results suggest that the changes in anatomical and chemical characteristics of foliar vascular bundles, such as the reduction of proteins, pectins, and other polysaccharides of foliar cell walls, participate in the leaf rolling mutation, and consequently lead to the reduced photosynthetic dynamics and apoplastic transport ability in the mutant.

Redox system in the plasma membranes of two ecotypes of reed (Phragmites communis Trin.) leaves from different habitats

Abstract Two ecotypes of reed ( Phragmites communis Trin.), named ‘swamp reed’ and ‘heavy salt meadow reed’ according to their habitats, growing in the desert regions of northwest China were investigated for plasma membrane (PM) redox activities in their leaves. The results showed that the PM redox system in both ecotypes of reed leaves can use NADH (reduced nicotinamide adenine dinucleotide) and NADPH (reduced nicotinamide adenine dinucleotide phosphate) as the electron donors and Fe(CN) 6 3− and EDTA–Fe 3+ as the electron acceptors. Inhibitors of respiratory chain such as KCN and NaN 3 had no inhibitory effects on the PM redox activities. The rates of NADH and NADPH oxidation and Fe(CN) 6 3− reduction in heavy salt meadow reed were higher compared to those in swamp reed. Reduction rates of EDTA–Fe 3+ in both ecotypes of reed were relatively low. It is suggested that higher redox activities of PM in the heavy salt meadow reed may be responsible for its adaptation to the saline habitats, as proved by previous studies that the PM redox system was involved in intercellular [Na + ] and [K + ] regulation.