Susana M. Gallego

Cadmium toxicity in plants

Heavy metals are important environmental pollutants and their toxicity is a problem of increasing significance for ecological, evolutionary, nutritional, and environmental reasons. Plants posses homeostatic cellular mechanisms to regulate the concentration of metal ions inside the cell to minimize the potential damage that could result from the exposure to nonessential metal ions. This paper summarizes present knowledge in the field of higher plant responses to cadmium, an important environmental pollutant. Knowledge concerning metal toxicity, including mechanisms of cadmium homeostasis, uptake, transport and accumulation are evaluated. The role of the cell wall, the plasma membrane and the mycorrhizas, as the main barriers against cadmium entrance to the cell, as well as some aspects related to phytochelatin-based sequestration and compartmentalization processes are also reviewed. Cadmium-induced oxidative stress was also considered as one of the most studied topics of cadmium toxicity.

Effect of heavy metal ion excess on sunflower leaves: evidence for involvement of oxidative stress

Abstract The relationship between heavy metal ion toxicity and oxidative stress in plant cells was studied. Leaf segments from 14 day old sunflower seedlings were incubated in solutions containing 0.5 mM Fe(II), Cu(II) or Cd(II) ions for 12 h in the light. Treatment with metal ions studied produced a decrease in chlorophyll and GSH contents as well as increases in lipid peroxidation and lipoxygenase activity. Free radical scavengers, such as sodium benzoate and mannitol, prevented the decrease in chlorophyll and GSH content and the lipid peroxidation and lipoxygenase increases. While Fe(II) and Cd(II) ions caused a decrease in superoxide dismutase activity, Cu(II) ions raised its level. However, all three metal ions caused decreases in other antioxidant enzymes (catalase, ascorbate peroxidase, glutathione reductase and dehydroascorbate reductase). Free radical scavengers protected these enzymes against inactivation. No effect of these scavengers was observed on superoxide dismutase activity. These results indicate that excess Fe(II), Cu(II) or Cd(II) ions produce oxidative damage in plant leaves.

Relationship between antioxidant defence systems and salt tolerance in Solanum tuberosum

A relationship between the antioxidant defence system and salt tolerance in two clones of potato (Solanum tuberosum L.) differing in salt sensitivity was studied. The antioxidant defence system of the sensitive clone responded differently to 100 and 150 mM NaCl. At 100 mM NaCl, growth, dehydroascorbate reductase and catalase activities remained unaltered, but chlorophyll and reduced glutathione content decreased (23% and 35%, respectively), while ascorbate content and superoxide dismutase activity were increased 34% and 63%, with respect to the control (0 mM NaCl). The superoxide dismutase increment was higher under 150 mM NaCl treatment, while a general decrease (except for dehydroascorbate reductase and catalase activities) in all the antioxidant parameters studied was observed in the sensitive clone. Reduced glutathione and ascorbate, the main antioxidant soluble defences, and all antioxidant enzymes (except catalase) were significantly elevated in the tolerant clone compared to the sensitive one when both were grown in the absence of NaCl. Under 100 and 150 mM NaCl treatments, no changes in the antioxidant stress parameters were detected in the tolerant clone. These results suggest a relationship between salt tolerance and the antioxidant defence system in the two clones.

Behaviour of antioxidant defense system in the adaptive response to salt stress in Helianthus annuus L. cells

A relationship between the antioxidant defense system and salt tolerance in two types of sunflower calli differing in salt sensitivity was studied. No reduction in growth occurred in the NaCl-salt-adapted cell line (T) when grown on 175 mM NaCl but growth of the salt-stressed cell line (S) was reduced by 83%. Lipid peroxidation and protein oxidation increased during acute stress of salt stressed cells at 14 and 28 d of the experiment, while salt-adapted calli (T) remained similar to non-shocked (C) values. The antioxidant defense system of callus adapted to growth under NaCl responded differently to 175 mM of salt compared with the corresponding controls under shock treatment. Salt-adapted and salt-stressed calli showed a similar pattern in GSH content at day 14 but at day 28 in S calli, GSH content was increased 100% over the non-shocked calli, while T calli returned to the initial values. In the salt-stressed calli, a general decrease in all the antioxidant enzymes studied (except for glutathione reductase and dehydroascorbate reductase activities) was observed at day 28. Except for catalase, the antioxidant enzymes were elevated constitutively in adapted calli as compared to stressed cells, when both were grown in the absence of NaCl (time 0), and remained unaltered until 28 d after the beginning of the experiment. These results suggest the involvement of an enzymatic antioxidant defense system in the adaptive response to salt stress in Helianthus annuus L. cells.

Effect of cadmium stress on nitrogen metabolism in nodules and roots of soybean plants

The nitrogen metabolism of soybean (Glycine max L.) nodules and roots was studied in plants subjected to two different concentrations (50 and 200 μM) of CdCl2. Nitrogenase activity was decreased in nodules treated with 200 μM Cd2+. In 50 μM Cd2+-treated plants, NH4+ content showed similar values to controls in nodules, but increased by 55% in roots. However, after treatment with 200 μM Cd2+, NH4+ levels increased in both tissues. Glutamate (Glu) and protein contents remained unaltered in nodules treated with 50 μM Cd2+, while at the higher Cd2+ concentration both were decreased. Nevertheless, polyamine content was increased at the two Cd2+ concentrations. In roots, Glu, polyamine and protein levels were significantly diminished at 50 and 200 μM CdCl2. For nitrogen-assimilation enzymes, glutamate dehydrogenase activity was moderately increased in nodules and roots following the lower Cd2+ treatment, though at the higher Cd2+ concentration root enzyme activity returned to control levels. An impressive increase in enzyme activity was found in nodules. In roots, the glutamine synthetase / glutamate synthase pathway was decreased at the two Cd2+ concentrations, though in nodules it was diminished only at 200 μM Cd2+. No changes in protease activity were found in the two tissues treated with 50μMCd2+. However, at 200 μM Cd2+, nodule and root protease activities decreased and increased, respectively. These results suggest that, in general, treatment with Cd2+ affects nitrogen assimilation and metabolism to a greater extent in soybean roots than in nodules.

Response of antioxidant defence system in soybean nodules and roots subjected to cadmium stress

The antioxidant defence system of soybean (Glycine maxL.) nodules and roots was studied in plants subjected to three different concentrations (50, 100 and µ200 M ) of CdCl 2 . Cadmium (Cd)-induced oxidative stress was determined by lipid peroxidation, which remained unaltered in nodules and roots treated with 50 µM Cd(II). No changes were observed in nodules treated with 100 M Cd(II), while a 20% increase was found in roots and 200 µM Cd(II) produced an increase of about 55% in both tissues. The soluble antioxidant defence, reduced glutath one(GSH) and the corresponding reduced/oxidised glutathione (GSH/GSSG) ratio showed different behaviour in both tissues, decreasing in roots with 100 and 200 µM Cd(II). No changes were observed in the GSH/GSSG ratio in nodules under the three Cd treatments. However, ascorbate content and ascorbate/dehydroascorbate (As/DAs) ratio were diminished in nodules and roots subjected to the three Cd concentrations. Regarding the antioxidant enzymes, it was found that, except for catalase, a general decrease in nodule enzyme activities was produced only under the 200 µM Cd(II) treatment. Nevertheless, root enzymatic antioxidant defences showed significant increments in L -ascorbate peroxidase (APOX), dehydroascorbate reductase (DHAR) and glutathione reductase (GR) activities under lower Cd treatment, while the enzyme activities decreased with the two higher concentrations of CdCl 2 . These results suggest that soybean roots were more affected than nodules by Cd treatments, although the higher Cd concentration produced oxidative stress and deleterious effects in antioxidant defence system in both tissues.

Effect of Cadmium Ions on Antioxidant Defense System in Sunflower Cotyledons

Sunflower (Helianthus annuus L.) seeds were germinated and grown in the presence of 50, 100 and 200 µM CdCl2. The lower concentration (50 µM) of Cd2 ions produced slight decrease in reduced glutathione (GSH) content and overall increase (except superoxide dismutase) in antioxidant enzyme activities, and in H2O2 concentration. Chlorophyll content, lipid peroxidation and protein oxidation were not affected under 50 µM CdCl2. GSH content was diminished under 100 and 200 µM CdCl2, and except for superoxide dismutase, which activity remained unaltered, overall decreases in the antioxidant enzyme activities (catalase, ascorbate peroxidase, dehydroascorbate peroxidase, glutathione reductase) and in guaiacol peroxidase were observed. These Cd2 concentrations caused a decrease in chlorophyll content as well as an increase in lipid peroxidation, protein oxidation and H2O2 concentration. All the observed effects were more evident with the highest concentration of cadmium chloride used.

Proteolytic system in sunflower (Helianthus annuus L.) leaves under cadmium stress

The effect of oxidative stress induced by cadmium on growth parameters and on the balance between protein synthesis and degradation was studied in sunflower (Helianthus annuus L.) leaves. Plants were germinated for 10 days and then transferred to hydroponic medium devoid (control) or containing 100, 200 and 300μM CdCl2. Analyses were performed between days 0 and 4 of Cd-treatment. All Cd(2+) concentrations significantly reduced leaf area and, fresh and dry weight, but leaf relative water content only decreased with 200 and 300μM Cd(2+). Control and treated plants had similar soluble protein content and showed the same rate of soluble protein labeling under the assay conditions. Although protease activity increased with cadmium treatment, proteasome activity was significantly inhibited. Expression of 20S proteasome remained similar to controls in cadmium treated plants. Cadmium caused an increase in ubiquitin-conjugated proteins and carbonyl groups content of treated plants, compared to control values. Cadmium induced an increase in protease specific activity; nevertheless, this increase was not relevant enough to avoid accumulation of oxidized proteins. Oxidation of proteins is one of the most important effects of cadmium treatment. The results presented here provide evidence for the role of the proteolytic system in sunflower plants subjected to cadmium stress.

Involvement of an antioxidant defence system in the adaptive response to heavy metal ions in Helianthus annuus L. cells

A relationship between the antioxidant defence system and metal iontolerance in two types of sunflower callus differing in metal ion sensitivitywas studied. The antioxidant defence system of callus subjected to anadaptationtreatment of Cd(II), Al(III) and Cr(III) responded differently to 150μM of each metal compared with the corresponding controls undershock treatment. The GSH/GSSG ratio remained similar to control values for thethree metal-acclimated calli and in the chromium shock treatment, decreasingmoderately in the acute treatment with cadmium and aluminum. In contrast, theAs/DAs ratio was decreased in the two different treatments for the three metalsions, but the decrease was greater with acute stress. The antioxidant enzymesresponded differently according to the metal and treatment used. In chromiumadapted callus, all antioxidant enzymes increased except for glutathionereductase. However, in the shock treatment ascorbate peroxidase activity wasdiminished with each metal ion assayed. Guaiacol peroxidase was decreased bycadmium and chromium and remained similar to control values with aluminum.Glutathione reductase was only decreased by cadmium, and superoxide dismutaseand catalase activities were less increased than in tolerant cells. Theseresults suggest the involvement of an antioxidant defence system in theadaptiveresponse to heavy metal ions in Helianthus annuus L.cells.

Relationship between polyamines and paraquat toxicity in sunflower leaf discs

Polyamines have been reported as efficient antioxidantcompounds in plants. Sunflower leaf discs, treatedwith 100 µM paraquat (PQ), a well known oxidativestress inducer, showed decreased levels of putrescine(Put), spermidine (Spd) and spermine (Spm) (between33% and 80% with respect to the controls). Argininedecarboxylase (ADC) and ornithine decarboxylase (ODC)activities decreased 42% and 33% respectively. Amongthe markers of oxidative stress measured after PQtreatment, chlorophyll and glutathione content werereduced (30% and 49% respectively) andthiobarbituric acid reactive substances (TBARS)content increased (60%). Superoxide dismutase (SOD)activity declined 60% with respect to the control andlipoxygenase (LOX) increased 25% when leaf-discs weretreated with the herbicide. Pretreatment withexogenous polyamines (1 mM) reversed paraquat toxicityto different degrees according to the polyamine and/orthe tested parameter. Spermidine was able to inhibitchlorophyll loss, while Spm reverted the effect of PQon the level of TBARS almost completely and alsorestored SOD activity close to control values.Putrescine was the least effective as an oxidantprotectant. These results provide support for theargument that polyamines are effective antioxidantsthrough their ability to act as radical scavengers.