The level of jasmonic acid in Arabidopsis thaliana and Phaseolus coccineus plants under heavy metal stress

The effect of heavy metal stress as a potent abiotic elicitor for triggering an accumulation of jasmonic acid (JA) was investigated. Copper and cadmium in in vivo conditions induced accumulation of jasmonates in mature leaves of Arabidopsis thaliana and in young and oldest Phaseolus coccineus plants. The dynamics of jasmonate accumulation showed a biphasic character in both plants. In the first phase, after 7 (A. thaliana) or 14 h (P. coccineus) of exposure to Cu or Cd, a rapid increase of JA level occurred, followed by a rapid decrease observed during 7 successive hours. In the next phase, a repeated but slow increase of JA content occurred. The heavy metal stress induced in particular a more stable (3R,7R) form of jasmonates. These results indicate that JA is connected with the mechanism of toxic action of both heavy metals in plants, differentially reacting to exogenous JA and possessing variable dynamics depending on the plants studied as well as their growth stage.     

NOX5 is expressed at the plasma membrane and generates superoxide in response to protein kinase C activation

NOX5 is a ROS-generating NADPH oxidase which contains an N-terminal EF-hand region and can be activated by cytosolic Ca(2+) elevations. However the C-terminal region of NOX5 also contains putative phosphorylation sites. In this study we used HEK cells stably expressing NOX5 to analyze the size and subcellular localization of the NOX5 protein, its mechanisms of activation, and the characteristics of the ROS released. We demonstrate that NOX5 can be activated both by the protein kinase C activating phorbol esther PMA and by the Ca(2+) ionophore ionomycin. The PMA- but not the ionomycin-dependent activation can be inhibited by protein kinase C inhibitors. NOX5 activity is inhibited by submicromolar concentrations of diphenyl iodonium (DPI), but not by apocynin. Western blot analysis showed a lower ( approximately 70 kDa) than expected (82 kDa) molecular mass. Two arguments suggest that NOX5 is at least partially expressed on the plasma membrane: (i) the membrane-impermeant superoxide was readily detected by extracellular probes, and (ii) immunofluorescent labeling of NOX5 detected a fraction of the NOX5 protein at the plasma membrane. In summary, we demonstrate that NOX5 can be found intracellularly and at the cell surface. We also describe that it can be activated through protein kinase C, in addition to its Ca(2+) activation.     

Toxicity in leaves of rice exposed to cadmium is due to hydrogen peroxide accumulation

The production of H₂O₂ in detached rice leaves of Taichung Native 1 (TN1) caused by CdCl₂ was investigated. CdCl₂ treatment resulted in H₂O₂ production in detached rice leaves. Diphenyleneiodonium chloride (DPI) and imidazole (IMD), inhibitors of NADPH oxidase (NOX), prevented CdCl₂-induced H₂O₂ production, suggesting that NOX is a H₂O₂-genearating enzyme in CdCl₂-treated detached rice leaves. Phosphatidylinositol 3-kinase inhibitors wortmanin (WM) or LY294002 (LY) inhibited CdCl₂-inducted H₂O₂ production in detached rice leaves. Exogenous H₂O₂ reversed the inhibitory effect of WM or LY, suggesting that phosphatidylinositol 3-phosphate is required for Cd-induced H₂O₂ production in detached rice leaves. Nitric oxide donor sodium nitroprusside (SNP) was also effective in reducing CdCl₂-inducing accumulation of H₂O₂ in detached rice leaves. Cd toxicity was judged by the decrease in chlorophyll content. The results indicated that DPI, IMD, WM, LY, and SNP were able to reduce Cd-induced toxicity of detached rice leaves. Twelve-day-old TN1 and Tainung 67 (TNG67) rice seedlings were treated with or without CdCl₂. In terms of Cd toxicity (leaf chlorosis), it was observed that rice seedlings of cultivar TN1 are Cd-sensitive and those of cultivar TNG67 are Cd-tolerant. On treatment with CdCl₂, H₂O₂ accumulated in the leaves of TN1 seedlings but not in the leaves of TNG67. Prior exposure of TN1 seedlings to 45^oC for 3 h resulted in a reduction of H₂O₂ accumulation, as well as Cd tolerance of TN1 seedlings treated with CdCl₂. The results strongly suggest that Cd toxicity of detached leaves and leaves attached to rice seedlings are due to H₂O₂ accumulation.     

Chromosomal responses of blowfly Lucilia cuprina to heat and heavy metal stress

Chromosomal responses to heat and heavy metal shocks were studied in the pupal trichogen polytene chromosomes of Lucilia cuprina. Heat shock induced seven distinct puffs on different chromosomes at the following loci: 1C2, 21A1, 21C, 23B, 24B, 42A and 95B3. Arsenate and mercury, two of the most common toxic environmental chemical pollutants also, induced almost the same set of puffs except 20B2, which appeared to be induced by metals only and 95B3, which was not induced by arsenate. The findings suggest that a common set of gene loci encoding the heat shock proteins is responsive to these diverse environmental stresses. This revised version was published online in July 2006 with corrections to the Cover Date.     

Role of plant respiratory burst oxidase homologs in stress responses

Plant respiratory burst oxidase homologs (Rbohs), which are also named nicotinamide adenine dinucleotide phosphate (NADPH) oxidases (NOXs), are the homologs of mammalian phagocyte gp91^phox. As a unique among other reactive oxygen species (ROS) production mechanisms in plants, NADPH oxidases can integrate different signal transduction pathways, such as calcium, protein phosphorylation catalysed by protein kinases, nitric oxide, and lipid messengers. Coupling with genetic studies, the ability of plant NADPH oxidases to integrate different signal transduction pathways with ROS production demonstrates their involvement in many important biological processes in cells, such as morphogenesis and development, and stress responses. Here, we focus on several current studies concerning the role of plant NADPH oxidases in stress responses.     

ERF转录因子及其在烟草抗逆性改良中的应用

转录因子调控的基因表达是植物逆境应答反应的关键环节。乙烯反应因子(ERF:ethylene responsive factor)在植物抗逆反应中发挥重要的调控作用,并成为近年来植物分子生物学领域的研究前沿。本文综述目前ERF及其对烟草抗逆遗传改良的相关研究进展。     

The generation of H2O2 in the xylem of Zinnia elegans is mediated by an NADPH-oxidase-like enzyme

The nature of the enzymatic system responsible for the generation of H₂O₂ in the lignifying xylem of Zinnia elegans (L.) was studied using the starch/KI method for monitoring H₂O₂ production and the nitroblue tetrazolium method for monitoring superoxide production. The results showed that lignifying xylem tissues are able to accumulate H₂O₂ and to sustain H₂O₂ production. Hydrogen peroxide production in the xylem of Z. elegans was sensitive to pyridine, imidazole, quinacrine and diphenylene iodonium, which are inhibitors of phagocytic plasma-membrane NADPH oxidase. The sensitivity of H₂O₂ production to the inhibitor of phospholipase C, neomycin, and to the inhibitor of protein kinase, staurosporine, and its reversion by the inhibitor of protein phosphatases, cantharidin, pointed to the analogies existing between the mechanism of H₂O₂ production in lignifying xylem and the oxidative burst observed during the hypersensitive plant cell response. A further support for the participation of an NADPH-oxidase-like activity in H₂O₂ production in lignifying xylem was obtained from the observation that areas of H₂O₂ production were superimposed on areas producing superoxide anion, the suspected product of NADPH oxidase, although attempts to demonstrate the existence of superoxide dismutase activity in intercellular washing fluid from Z. elegans were unsuccessful. Even so, the levels of NADPH-oxidase-like activity in microsomal fractions, and of peroxidase in intercellular washing fluids, are consistent with a role for NADPH oxidase in the delivery of H₂O₂ which may be further used by xylem peroxidases for the synthesis of lignins. This hypothesis was further confirmed through a direct histochemical probe based on the H₂O₂-dependent oxidation of tetramethylbenzidine by xylem cell wall peroxidases. These results are the first evidence for the existence of an NADPH oxidase responsible for supplying H₂O₂ to peroxidase in the lignifying xylem of Z. elegans. Received: 6 February 1998 / Accepted: 14 August 1998     

Influence of cytokinins on the methyl jasmonate-promoted senescence in Helianthus annuus cotyledons

Cotyledons of seven-day-old sunflower seedlings were pretreated withcytokinins prior to exposure to methyl jasmonate (MeJA). The rate of senescencewas then followed by measuring chlorophyll loss, electrolyte leakage, ethyleneproduction, and 1-aminocyclopropane-1-carboxylase (ACC) oxidase activity. MeJApromoted all these senescence parameters and the MeJA effects were partiallyblocked by cytokinin pretreatment. 8-azaguanine, which has been reported tohaveanticytokinin activity, blocked the ability of benzyl adenine (BA) to reversethe effect of MeJA on senescence. MeJA also increased SOD and catalaseactivity,decreased protein content. However, while the cytokinin BA more than overcamethe MeJA effect on protein content and SOD activity it did not antagonize theeffect of MeJA on catalase.     

Wound response in passion fruit (<Emphasis Type="Italic">Passiflora</Emphasis> f. <Emphasis Type="Italic">edulis flavicarpa</Emphasis>) plants: gene characterization of a novel chloroplast-targeted allene oxide synthase up-regulated by mechanical injury and methyl jasmonate

The induction of a chloroplast-localized 13-lipoxygenase (13-LOX) in passion fruit leaves in response to methyl jasmonate (MeJa) was previously reported. Since allene oxide synthase (AOS) is a key cytochrome P450 enzyme in the oxylipin pathway leading to AOS-derived jasmonates, the results above led in turn to an investigation of AOS in our model plant. Spectrophotometric assays showed that 24 h exposure of MeJa caused a high increase in 13-hydroperoxy linolenic acid (13-HPOT) metabolizing activity in leaf tissue. Western analysis using polyclonal antibodies against tomato AOS strongly indicate that, at least a part of the 13-HPOT metabolizing capacity can be attributed to AOS activity. We cloned the cDNA from a novel AOS encoding gene from passion fruit, named PfAOS. The 1,512 bp open reading frame of the AOS–cDNA codes a putative protein of 504 amino acid residues containing a chloroplast target sequence. Database comparisons of the deduced amino acid sequence showed highest similarity with dicot AOSs. Immunocytochemistry analysis showed the compartmentalization of AOS in chloroplasts of MeJa treated leaves, corroborating the predicted subcellular localization. Northern analysis showed that AOS gene expression is induced in leaf tissue in response to mechanical wounding and exposure to MeJa. In addition, such treatments caused an increase in papain inhibitor(s) in leaf tissue. Taken together, these results indicate that PfAOS may play an important role in systemic wound response against chewing insect attack. Furthermore, it can be useful as a tool for understanding the regulation of jasmonates biosynthesis in passion fruit.     

Chemical activity and biological effect of sludge-borne heavy metals and inorganic metal salts added to soils

Summary A greenhouse experiment was conducted in order to evaluate the chemical activity and the uptake by Italian ryegrass (Lolium perenne cv. S24) of Zn, Cu, Cd and Ni added to a sandy and a heavy clay soil in two different forms: as inorganic salts and sludge-borne.The chemical activity of heavy metals as evaluated with different extractants was higher for the inorganic salt treatment and for the sandy soil, indicating that the chemical form of the metal and soil characteristics largely affect their extractability.The different chemical activity was also reflected in plant uptake. For all metals the degree of plant accumulation decreased in the following order: sandy soil-salt sandy soil-sludge> clay soil-salt>clay soil-sludge.These findings indicate that caution must be used when using results of inorganic salt treatments and different soil types to evaluate plant uptake of heavy metals from sludge amended soils.     

Developmental instability in cabbage aphid (Brevicoryne brassicae) populations exposed to heavy metal accumulated host plants

Random deviations from the perfect symmetry of normally bilaterally symmetrical characters exist during individual development as a result of various environmental conditions. Fluctuating asymmetry (FA) is often used as a measurement of developmental instability, and within-environmental variation (CV_e) is also considered as an indicator of developmental deviations. These two parameters may indicate the quality of the environmental habitat of organisms. For herbivore insects, such as aphids, any change in their host plants conditions is important and directly affects their development. The presented investigation revealed that both Lead (Pb) and Copper (Cu) accumulation in different host plants resulted in a significant amount of deviations from bilateral symmetry in cabbage aphid (Brevicoryne brassicae). Cabbage aphid populations showed higher FA and CV_e on heavy metal accumulated cabbage and radish than on non-contaminated cabbage and radish plants. However, the pattern between developmental instability and fitness measurements was inconsistent. Thus, this study does not support the hypothesis that asymmetry is a valuable bioindicator of developmental instability.     

Managing heavy metal toxicity stress in plants: Biological and biotechnological tools

The maintenance of ion homeostasis in plant cells is a fundamental physiological requirement for sustainable plant growth, development and production. Plants exposed to high concentrations of heavy metals must respond in order to avoid the deleterious effects of heavy metal toxicity at the structural, physiological and molecular levels. Plant strategies for coping with heavy metal toxicity are genotype-specific and, at least to some extent, modulated by environmental conditions. There is considerable interest in the mechanisms underpinning plant metal tolerance, a complex process that enables plants to survive metal ion stress and adapt to maintain growth and development without exhibiting symptoms of toxicity. This review briefly summarizes some recent cell biological, molecular and proteomic findings concerning the responses of plant roots to heavy metal ions in the rhizosphere, metal ion-induced reactions at the cell wall-plasma membrane interface, and various aspects of heavy metal ion uptake and transport in plants via membrane transporters. The molecular and genetic approaches that are discussed are analyzed in the context of their potential practical applications in biotechnological approaches for engineering increased heavy metal tolerance in crops and other useful plants.     

Physiological responses of four herbaceous plants to aluminum stress in South China

Different plants have physiological responses under Al stress, but there is no systematic study to examine physiological responses of herbaceous plants under Al stress. The aim of this study is to investigate the effect of Al on physiological characteristics of four herbaceous plants, which distributed in red soil area in South China, and to analyze the differences in physiological responses to Al stress between the four herbaceous plants. Four herbaceous plants (Pharbitis nil, Cassia occidentlis, Echinochloa colonum and Aeschynomene indica) were used, and the seed germination percentage, the contents of chlorophyll, proline, and malondialdehyde (MDA), membrane permeability (MP), soluble sugar, and activities of peroxides (POD) and catalase (CAT) in leaves under five Al³⁺ treatments (0, 80, 400, 2 000, and 10 000 mg/L) were assayed with the sand culture method. The results showed remarkable effects of Al³⁺ on physiological characteristics of these four herbaceous plants. The seeds of all the four species could not germinate at 10 000 mg/L, and the growth of all plants were retarded under the 2 000 mg/L Al³⁺ treatment. Compared with the control, 2 000 mg/L Al³⁺ significantly (P < 0.05) reduced the contents of chlorophyll a and chlorophyll a + b, and increased the contents of MDA and MP. The content of proline increased very significantly (P < 0.01) and activities of POD and CAT were depressed. The contents of MDA and MP in leaves of P. nil and A. indica decreased, and the activities of POD and CAT in leaves of the two plants increased under 80 mg/L and 400 mg/L. However, the changes in C. occidentlis leaves were opposite to those of the above two plants. The changes in leaves of E. colonum were similar to those of P. nil and A. indica at 80 mg/L, but were opposite to those at 400 mg/L Al³⁺. It is suggested that plants with higher activities of POD and CAT, more contents of chlorophyll and proline, and lower contents of MDA and MP consequently improve the tolerance to Al stress under low and middle Al treatments. __________ Translated from Acta Phytoecologica Sinica, 2005, 29(4): 644–651 [译自: 植物生态学报, 2005, 29(4): 644–651]     

Physiological responses of four herbaceous plants to aluminum stress in South China

Different plants have physiological responses under A1 stress,but there is no systematic study to examine physiological responses of herbaceous plants under Al stress.The aim of this study is to investigate the effect of Al on physiological characteristics of four herbaceous plants,which distributed in red soil area in South China,and to analyze the differences in physiological responses to Al stress between the four herbaceous plants.Four herbaceous plants(Pharbitis nil,Cassia occidentlis,Echinochloa colonum and Aeschynomene indica)were used,and the seed germination percentage,the contents of chlorophyll,proline,and malondialdehyde(MDA),membrane permeability(MP),soluble sugar,and activities of peroxides(POD)and cata lase(CAT)in leaves under five Al3+ treatments(0,80,400,2000,and 10000 mg/L)were assayed with the sand culture method.The results showed remarkable effects of Al3+ on physiological characteristics of these four herbaceous plants.The seeds of all the four species could not germinate at 10000 mg/L.and the growth of all plants were retarded under the 2000 mg/LAr3+ treatment.Compared with the control,2000 mg/L Al3+ significantly(P＜0.05)reduced the contents of chlorophyll a and chlorophyll a+b,and increased the contents of MDA and MP.The content of proline in creased very significantly(P＜0.01)and activities of POD and CAT were depressed.The contents of MDA and MP in leaves of P.nil and A.indica decreased,and the activities of POD and CAT in leaves of the two plants increased under 80mg/L and 400 mg/L.However,the changes in C.occidentlis leaves were opposite to those of the above two plants.The changes in leaves of E.colonum were similar to those of P.nil and A.indica at 80 mg/L.but were opposite to those at 400mg/LAr3+.It is suggested that plants with higher activities of POD and CAT,more contents of chlorophyll and proline,and lower contents of MDA and MP consequently improve the tolerance to Al stress under low and middle Al treatments.     

Polyamine metabolism in sunflower plants under long-term cadmium or copper stress

Summary. The effect of different doses of cadmium and copper was studied in relation to growth and polyamine (Pas) metabolism in shoots of sunflower plants. Cadmium accumulated to higher levels than copper and shoot length was reduced by 0.5 and 1 mM Cd, but only by 1 mM Cu. At 1 mM of Cd or Cu, Put content increased 270% and 160% with Cd²⁺ and Cu²⁺, respectively. Spermidine (Spd) was modified only by 1 mM Cd, while spermine (Spm) declined after seeds germinated, increasing thereafter but only with 1 mM Cd or Cu (273% over the controls for Cd and 230% for Cu at day 16). Both ADC and ODC activities were increased by 1 mM Cd, whereas 1 mM Cu enhanced ADC activity, but reduced ODC activity at every concentration used. The role of Pas as markers of Cd or Cu toxicity is discussed.     

Water status in Mesembryanthemum crystallinum under heavy metal stress

Heavy metals (HMs) are known to have negative effects on plant water status; however, the mechanisms by which plants rearrange their water relations to adapt to such conditions are poorly understood. Using the model plant Mesembryanthemum crystallinum, we studied disturbances in water status and rapid plant defence responses induced by excess copper or zinc. After a day of HM stress, reductions in root sap exudation and water deficits in leaf tissues became evident. We also observed several primary adaptive events, including a rapid decrease in the transpiration rate and progressive declines in the leaf-cell sap osmotic potential. Longer HM treatments resulted in reductions of total and relative water contents as well as proline accumulation, an increase in water retention capacity and changes in aquaporin gene expression. After 3 h of HM exposure, leaf expression of the McTIP2;2 gene, which encodes tonoplast aquaporin, was suppressed more than two-fold, thus representing one of the earliest responses to HM treatment. The expression of three additional aquaporin genes was also reduced starting at 9 h; this effect became more prominent upon longer HM exposure. These results indicate that HMs induce critical rearrangements in the water relations of M. crystallinum plants, based on the rapid suppression of transpiration flow and strong inhibition of root sap exudation. These effects then triggered an adaptive water-conserving strategy involving differential regulation of aquaporin gene expression in leaves and roots, further reductions in transpiration, and an accelerated switch to CAM photosynthesis.