Ricardo A. Azevedo

Making the life of heavy metal-stressed plants a little easier

The contamination of soils and water with metals has created a major environmental problem, leading to considerable losses in plant productivity and hazardous health effects. Exposure to toxic metals can intensify the production of reactive oxygen species (ROS), which are continuously produced in both unstressed and stressed plants cells. Some of the ROS species are highly toxic and must be detoxified by cellular stress responses, if the plant is to survive and grow. The aim of this review is to assess the mode of action and role of antioxidants in protecting plants from stress caused by the presence of heavy metals in the environment.

Antioxidant enzymes responses to cadmium in radish tissues

To investigate the antioxidant responses of radish (Raphanus sativus L.) to cadmium (Cd) treatment, seedlings of a tolerant variety were grown in increasing concentrations of CdCl(2), ranging from 0.25-1 mM, for up to 72 h in a hydroponic system. Analysis of Cd uptake indicated that most of the Cd accumulated in the roots, but some was also translocated and accumulated in the leaves, especially at the higher concentrations of Cd used in the experiments. Roots and leaves were analysed for catalase, glutathione reductase and superoxide dismutase activities. Catalase and glutathione reductase activities increased considerably in the roots and leaves after 24 h exposure to the metal, indicating a direct correlation with Cd accumulation. The analysis of native PAGE enzyme activity staining, revealed several superoxide dismutase isoenzymes in leaves, with the two predominant isoenzymes exhibiting increases in activity in response to Cd treatment. The results suggest that in radish, the activity of antioxidant enzymes responds to Cd treatment. The main response may be via the activation of the ascorbate-glutathione cycle for the removal of hydrogen peroxide, or to ensure the availability of glutathione for the synthesis of Cd-binding proteins.

The biosynthesis and metabolism of the aspartate derived amino acids in higher plants

The essential amino acids lysine, threonine, methionine and isoleucine are synthesised in higher plants via a common pathway starting with aspartate. The regulation of the pathway is discussed in detail, and the properties of the key enzymes described. Recent data obtained from studies of regulation at the gene level and information derived from mutant and transgenic plants are also discussed. The herbicide target enzyme acetohydroxyacid synthase involved in the synthesis of the branched chain amino acids is reviewed.

Activity of antioxidant enzymes in response to cadmium in Crotalaria juncea

The effects of the heavy metal Cadmium (Cd) on the growth and the activities of the antioxidant enzymes, catalase (CAT, EC 1.11.1.6), superoxide dismutase (SOD, EC 1.15.1.1) and glutathione reductase (GR, EC 1.6.4.2) have been investigated in Crotalaria juncea seedlings. Concentrations above 0.2 mM CdCl2 were shown to inhibit strongly the growth of roots and shoots. Cd was shown to accumulate to very high concentrations in the roots, whilst in the leaves, the maximum concentration obtained following treatment with 2 mM CdCl2, was only 6% of that determined in the roots. Although CAT activity did not exhibit any major variation in the roots following CdCl2 treatment, 2 mM CdCl2 induced a 6-fold increase in activity in the leaves when compared to the untreated control. Non-denaturing PAGE gels stained for SOD activity revealed four isoenzymes, two Mn-SOD and two Cu/Zn-SOD. The results observed for SOD were different of those observed for CAT activity, since in both, leaves and roots, no significant changes in the total activity or of the four isoenzymes were observed following the treatment with CdCl2. GR activity exhibited a similar pattern of that of CAT activity. The concentration of 2 mM CdCl2 induced a small increase in activity in the roots after 48 h of exposure, whereas in leaves a 7-fold increase in GR activity was detected after 48 hr exposure to 2 mM CdCl2. The results suggest that in C. juncea the reactive oxygen species (ROS) induced by Cd, are metabolised by CAT in the peroxisomes. In the case of GR activity, the increase observed in the leaves suggest that GR is also playing a role in the detoxification of Cd-induced ROS possibly via the glutathione-ascorbate cycle.

The aspartic acid metabolic pathway, an exciting and essential pathway in plants.

Summary.Aspartate is the common precursor of the essential amino acids lysine, threonine, methionine and isoleucine in higher plants. In addition, aspartate may also be converted to asparagine, in a potentially competing reaction. The latest information on the properties of the enzymes involved in the pathways and the genes that encode them is described. An understanding of the overall regulatory control of the flux through the pathways is undisputedly of great interest, since the nutritive value of all cereal and legume crops is reduced due to low concentrations of at least one of the aspartate-derived amino acids. We have reviewed the recent literature and discussed in this paper possible methods by which the concentrations of the limiting amino acids may be increased in the seeds.

Effects of Cadmium on Antioxidant Enzyme Activities in Sugar Cane

Sugar cane (Saccharum officinarum L. cv. Copersucar SP80-3280) seedlings were grown in nutrient solution with varying concentrations (0, 2 and 5 mM) of cadmium chloride for 96 h. Leaves were analysed for catalase (CAT), glutathione reductase (GR) and superoxide dismutase (SOD) activities. Although a clear effect of CdCl2 on plant growth was observed, the activity of SOD was not altered significantly. However, the CAT activity decreased as the concentration of CdCl2 increased. GR exhibits a significant increase in activity at 2 and 5 mM CdCl2. CAT and SOD isoenzymes were further characterised by analysis in non-denaturing PAGE. Activity staining for SOD revealed up to seven isoenzymes in untreated control and 2 mM CdCl2 treated plants, corresponding to Cu/Zn-SOD isoenzymes. At 5 mM CdCl2, only six Cu/Zn-SOD isoenzymes were observed. No Fe-SOD and Mn-SOD isoenzymes were detected. For CAT, one band of activity was observed.

The influence of nitrogen supply on antioxidant enzymes in plant roots

Plants of Zea mays L., Hordeum vulgare L. and Arabidopsis thaliana (L.) Heynh. were grown at different concentrations of nitrogen, as NH4NO3, and the antioxidant enzyme activities and quantities in the roots, were studied. Maize plants were grown at 1 and 10 mM N for 32 d after germination and their roots were analysed by native PAGE for the isoenzymic profiles of glutathione reductase (GR; EC 1.6.4.2), catalase (CAT; EC 1.11.1.6) and superoxide dismutase (SOD; EC 1.15.1.1) and by spectrophotometric assays for total activity of CAT and GR. Barley and A. thaliana plants were grown at five N concentrations (0.25, 0.5, 1, 10 and 20 mM) for 27 and 26 d, respectively, and the isoenzymic profile of the three enzymes were analysed in their roots. The number of GR isoforms in the three plant species was increased at high N supply compared with low N. In addition, the CAT and GR activities detected in the roots were increased in plants grown at higher N. The increase in activity and number of bands of GR at high N in the roots of all three plants studied was not accompanied by a change in activity or the number of SOD isoforms. The possibility that the application of high N promotes the formation of reactive oxygen species by stimulating an increased rate of growth is discussed.

Nanoparticles applied to plant science: a review.

The present review addresses certain important aspects regarding nanoparticles and the environment, with an emphasis on plant science. The production and characterization of nanoparticles is the focus of this review, providing an idea of the range and the consolidation of these aspects in the literature, with modifications on the routes of synthesis and the application of the analytical techniques for characterization of the nanoparticles (NPs). Additionally, aspects related to the interaction between the NPs and plants, their toxicities, and the phytoremediation process, among others, are also discussed. Future trends are also presented, supplying evidence for certain possibilities regarding new research involving nanoparticles and plants.

CHANGES IN ANTIOXIDANT ENZYME ACTIVITIES IN SOYBEAN UNDER CADMIUM STRESS

Soybean (Glycine max L.) seedlings were grown in a hydroponic system containing nutrient solution with varying concentrations (0, 0.01, 0.5 mM) of cadmium (Cd) for a 48 h period. Roots and leaves were analyzed for catalase, glutathione reductase, and superoxide dismutase activities. Although a clear effect of Cd on plant growth was observed, the activities of catalase and superoxide dismutase were not altered significantly in the roots or leaves at all concentrations tested. Glutathione reductase did not exhibit any variation in activity in leaves, however, in roots an increase in activity was observed at 0.5 mM Cd. Catalase and superoxide dismutase isoenzymes were further characterized by analysis in non-denaturing PAGE. Activity staining for superoxide dismutase (SOD) revealed seven isoenzymes in leaves and eight isoenzymes in roots, corresponding to Mn-SOD and Cu/Zn-SOD. No Fe-SOD isoenzymes were observed. For catalase, one band of activity was observed in leaves and roots. Although no significant variation was observed in leaf tissue, in roots the 0.5 mM Cd treatment exhibited a slight increase in catalase activity. The results suggest that in soybean, glutathione reductase is stimulated by Cd in roots, to produce reduced glutathione to be used in the synthesis of phytochelatins or possibly as a response to the generation of reactive oxygen species induced by Cd.

Zn uptake, physiological response and stress attenuation in mycorrhizal jack bean growing in soil with increasing Zn concentrations

The influence of arbuscular mycorrhizal fungi (AMF) inoculation on Canavalia ensiformis growth, nutrient and Zn uptake, and on some physiological parameters in response to increasing soil Zn concentrations was studied. Treatments were applied in seven replicates in a 2 x 4 factorial design, consisting of the inoculation or not with the AMF Glomus etunicatum, and the addition of Zn to soil at the concentrations of 0, 100, 300 and 900 mg kg(-1). AMF inoculation enhanced the accumulation of Zn in tissues and promoted biomass yields and root nodulation. Mycorrhizal plants exhibited relative tolerance to Zn up to 300 mg kg(-1) without exhibiting visual symptoms of toxicity, in contrast to non-mycorrhizal plants which exhibited a significant growth reduction at the same soil Zn concentration. The highest concentration of Zn added to soil was highly toxic to the plants. Leaves of plants grown in high Zn concentration exhibited a Zn-induced proline accumulation and also an increase in soluble amino acid contents; however proline contents were lower in mycorrhizal jack beans. Plants in association or not with the AMF exhibited marked differences in the foliar soluble amino acid profile and composition in response to Zn addition to soil. In general, Zn induced oxidative stress which could be verified by increased lipid peroxidation rates and changes in catalase, ascorbate peroxidase, glutathione reductase and superoxide dismutase activities. In summary, G. etunicatum was able to maintain an efficient symbiosis with jack bean plants in moderately contaminated Zn-soils, improving plant performance under those conditions, which is likely to be due to a combination of physiological and nutritional changes caused by the intimate relation between fungus and plant. The enhanced Zn uptake by AMF inoculated jack bean plants might be of interest for phytoremediation purposes.