Role of calcium signaling in cadmium stress mitigation by indol-3-acetic acid and gibberellin in chickpea seedlings.

Given the adverse impacts of heavy metals on plant development and physiological processes, the present research investigated the protective role of indole-3-acetic acid (IAA) and gibberellic acid (GA3) against cadmium (Cd)-induced injury in chickpea seedlings. Therefore, seeds germinated for 6 days in a medium containing 200 µM Cd alone or combined with 10 µM GA3 or 10 µM IAA. Both GA3 and IAA mitigated Cd-imposed growth delays in roots and shoots (80% and 50% increase in root and shoot length, respectively). This beneficial effect was accompanied by a significant reduction in Cd2+ accumulation in both roots (74% for IAA and 38% for GA3) and shoots (68% and 35%, respectively). Furthermore, these phytohormones restored the cellular redox state by reducing the activity of NADPH oxidase and downregulating the transcription level of RbohF and RbohD genes. Likewise, hydrogen peroxide contents were reduced by GA3 and IAA supply. Additionally, GA3 and IAA countered the Cd-induced reduction in total phenols, flavonoids, and reducing sugars in both roots and shoots. The exogenous effectors enhanced the activities of catalase, ascorbate peroxidase, and thioredoxin, as well as the corresponding gene expressions. Interestingly, adding GA3 and IAA to the Cd-contaminated germination media corrected the level of calcium (Ca2+) ion within seedling tissues. This effect coincided with the upregulation of key genes associated with stress sensing and signal transduction, including auxin-binding protein (ABP19a), mitogen-activated protein kinase (MAPK2), calcium-dependent protein kinase (CDPK1), and calmodulin (CaM). Overall, the current results suggest that GA3 and IAA sustain the Ca2+ signaling pathway, resulting in metal phytotoxicity relief. Amendment of agricultural soils contaminated with heavy metals with GA3 or IAA could represent an effective practice to improve crop yield.

Seed pretreatment with melatonin confers cadmium tolerance to chickpea seedlings through cellular redox homeostasis and antioxidant gene expression improvement.

Several phytoremediation strategies have been undertaken to alleviate cadmium (Cd)-mediated injury to crop yield resulting from agricultural land pollution. In the present study, the potentially beneficial effect of melatonin (Me) was appraised. Therefore, chickpea (Cicer arietinum L.) seeds were imbibed for 12 H in distilled water or Me (10 µM) solution. Then, the seeds germinated in the presence or the absence of 200 µM CdCl2 for 6 days. Seedlings obtained from Me-pretreated seeds exhibited enhanced growth traits, reflected by fresh biomass and length increase. This beneficial effect was associated with a decreased Cd accumulation in seedling tissues (by 46 and 89% in roots and shoots, respectively). Besides, Me efficiently protected the cell membrane integrity of Cd-subjected seedlings. This protective effect was manifested by the decreased lipoxygenase activity and the subsequently reduced accumulation of 4-hydroxy-2-nonenal. Melatonin counteracted the Cd-mediated stimulation of the pro-oxidant NADPH-oxidase (90 and 45% decrease compared to non-pretreated Cd-stressed roots and shoots, respectively) and NADH-oxidase activities (almost 40% decrease compared to non-pretreated roots and shoots), preventing, thus, hydrogen peroxide overaccumulation (50 and 35% lesser than non-pretreated roots and shoots, respectively). Furthermore, Me enhanced the cellular content of pyridine nicotinamide reduced forms [NAD(P)H] and their redox state. This effect was associated with the Me-mediated stimulation of the glucose-6-phosphate dehydrogenase (G6PDH) and malate dehydrogenase activities, concomitantly with the inhibition of NAD(P)H-consuming activities. These effects were accompanied by the up-regulation of G6PDH gene expression (45% increase in roots) and the down-regulation of the respiratory burst oxidase homolog protein F (RBOHF) gene expression (53% decrease in roots and shoots). Likewise, Me induced an increased activity and gene transcription of the Asada-Halliwell cycle, namely ascorbate peroxidase, monodehydroascorbate reductase, dehydroascorbate reductase, and glutathione reductase, concomitantly with a reduction of the glutathione peroxidase activity. This modulating effect led to the restoration of the redox homeostasis of the ascorbate and the glutathione pools. Overall, current results attest that seed pretreatment with Me is effective in Cd stress relief and can be a beneficial crop-protective approach.

Clue of zinc oxide and copper oxide nanoparticles in the remediation of cadmium toxicity in Phaseolus vulgaris L. via the modulation of antioxidant and redox systems.

The present study represents new evidence of the successful use of metal oxide nanoparticles in the remediation of heavy metals. Zinc oxide nanoparticles (ZnO NP) and copper oxide nanoparticles (CuO NP) were assessed to alleviate cadmium (Cd) toxicity in Phaseolus vulgaris L. seedlings and plants. Monitoring physiological and metabolic parameters allowed to elucidate Cd mechanism and process whereby it exerts phytotoxic effects on bean. The response of P. vulgaris seedlings is NP dose-dependent (10 mg/L, 50 mg/L, 100 mg/L, and 200 mg/L). Similarly, applied concentrations triggered a differential response of growing plants in terms of length and biomass. Our physiological data allowed to select 100 mg/L as the most appropriate concentration to apply, in order to avoid any risk of phytotoxicity. The regulatory mechanisms by which ZnO NP and CuO NP act are for the first time compared in the embryonic axes of bean seedlings under Cd stress. Both NP were able to reduce the hypergeneration of hydrogen peroxide (H2O2). They also acted via enhancing ROS scavenging enzymatic capacity, and activity of antioxidant enzymes CAT, APX, GPOX, GPX, and GR, and inhibited the activity of ROS producing enzymes such as GOX and NOX. Another mechanistic effect of NP consisted of the modulation of redox enzymes Trx, NTR, Fd, and FNR evolved in cellular homeostasis and maintaining reduced status in cells. Taken together, ZnO NP triggered more significant metabolic regulations allowing to mitigate the oxidative damage caused by Cd.

Cu and CuO Nanoparticles Affected the Germination and the Growth of Barley (Hordeum vulgare L.) Seedling.

The application of Cu and CuO nanofertilizers in horticulture has been a promising strategy to promote plants' growth. In our study, increasing concentrations (10, 25, 50, 100, 250, 500, 1000, 2000 mg/L) were assessed for their inhibitory or stimulatory effects on barley (Hordeum vulgare L.) seedlings. Our results showed an enhancement of seed germination parameters (FGP, t50, cumulative germination rate, AUC), and seedling growth parameters (roots and shoots' lengths, fresh biomasses and dry biomasses) by the low concentrations of Cu NPs and CuO NPs, while concentrations above 500 mg/L displayed inhibiting effects. CuO NPs treatment showed a significant similitude with CuSO4, which confirms that CuO NPs act mainly via released Cu ions. However, Cu NPs exhibited a different behavior since the nanosized particles contribute together with Cu ions in barley response to Cu NPs. This provides an overall picture of the way these nanoparticles may behave in plant systems.

Assessment of the risks of copper- and zinc oxide-based nanoparticles used in Vigna radiata L. culture on food quality, human nutrition and health.

The present article aims to assess the phytotoxic effects of copper and zinc oxide nanoparticles (Cu NPs, ZnO NPs) on mung bean (Vigna radiata L.) and their possible risk on food quality and safety. We also study the molecular mechanisms underlying the toxicity of nanosized Cu and ZnO. Seeds of mung bean were germinated under increasing concentrations of Cu NPs and ZnO NPs (10, 100, 1000, 2000 mg/L). We analyzed levels of free amino acids, total soluble sugars, minerals, polyphenols and antioxidant capacity. Our results showed that depending on the concentrations used of Cu NPs and ZnO NPs, the physiology of seed germination and embryo growth were modified. Both free metal ions and nanoparticles themselves may impact plant cellular and physiological processes. At 10 mg/L, an improvement of the nutritive properties, in terms of content in free amino acids, total soluble sugars, essential minerals, antioxidant polyphenols and flavonoids, was shown. However, higher concentrations (100-2000 mg/L) caused an alteration in the nutritional balance, which was revealed by the decrease in contents and quality of phenolic compounds, macronutrients (Na, Mg, Ca) and micronutrients (Cu, Fe, Mn, Zn, K). The overall effects of Cu and ZnO nanoparticles seem to interfere with the bioavailability of mineral and organic nutrients and alter the beneficial properties of the antioxidant phytochemicals, mineral compounds, phenolic acids and flavonoids. This may result in a potential hazard to human food and health, at some critical doses of nanofertilizers. This study may contribute in the guidelines to the safe use of nanofertilizers or nanosafety, for more health benefit and less potential risks.

Nitric oxide donor, sodium nitroprusside modulates hydrogen sulfide metabolism and cysteine homeostasis to aid the alleviation of chromium toxicity in maize seedlings (Zea mays L.).

The current research aimed to assess the protective role of nitric oxide (NO) against chromium (Cr) toxicity in maize seedlings. Chromium (200 µM) lowered osmotic potential in epicotyls and mostly in radicles (by 38% and 63%, respectively) as compared to the control. Sodium nitroprusside (SNP, NO donor) restored seedling biomass (+90% for both organs) and water potential, whereas application of Nω-nitro-L-arginine methylester (L-NAME, a NOS inhibitor) increased sensitivity to Cr. SNP suppressed Cr-triggered proline accumulation by inhibiting Δ1-pyrroline-5-carboxylate synthetase activity and stimulating proline dehydrogenase activity, leading to glutamate over-accumulation (~30% for both organs). Cr stimulated cysteine metabolism and this was further enhanced by SNP which stimulated serine acetyl-transferase and O-acetylserine (thiol) lyase activities. This was followed by an increase in endogenous hydrogen sulfide (H2S) generation by up-regulating L-cysteine desulfhydrase (+205%), D-cysteine desulfhydrase (+150%) and cyanoalanine synthase (+65%) activities in radicles compared to Cr-treatments plants. These positive effects were reduced in L-NAME compared to control. Combined Cr+SNP affected the levels of compounds involved in glutathione metabolism (γ-glutamyl-cysteinyl, γ-glutamyl-cysteinyl-clycine, γ-cysteinyl-glycine, and glycine.). All together, our findings indicate that NO and elicited cellular H2S act synergistically to alleviate Cr stress in maize seedlings by influencing a metabolic interplay between cysteine, proline, and glutathione.

Exogenous nitric oxide alleviates manganese toxicity in bean plants by modulating photosynthesis in relation to leaf lipid composition.

Nitric oxide (NO) is a signaling molecule controlling several steps of plant development and defense process under stress conditions. NO-induced alleviation of manganese (Mn) toxicity was investigated on bean plants submitted for 28 days to 500 µM MnCl2. Manganese excess decreased plant dry weight and elongation and increased levels of reactive oxygen species and lipid peroxidation leading to up-regulation of superoxide dismutase, catalase, and ascorbate peroxidase activities. The inhibitory effects of Mn on plant growth were associated to reduction of light-saturated carbon assimilation (Amax), stomatal conductance (gs), and transpiration (E). By contrast, Mn induced significant increase in the apparent quantum yield (ɸ) and light compensation point (LCP). Interestingly, intracellular CO2 (Ci) remains stable under Mn stress. Concomitantly, leaf membrane lipids have drastically reduced under high Mn concentration. After Mn exposition, leaf fatty acids exhibited a significant loss of linolenic acid, accompanied by an accumulation of palmitoleic, stearic, and linoleic acids leading to alteration of lipid desaturation. NO supply reversed Mn toxicity as evidenced by enhancement of growth biomass and recovery of Amax, E, ɸ, and LCP. Similarly, NO addition has positive effects on leaf lipid content and composition leading to restoration of lipid unsaturation. The modulation of fatty acid composition can be a way to reduce leaf membrane damages and maintain optimal photosynthesis and plant growth. Despite the absence of enough evidences in how NO is involved in lipid and photosynthesis recovery under Mn stress conditions, it is assumed that NO beneficial effects are attributable to NO/Mn cross-talk.

Calcium and ethylene glycol tetraacetic acid mitigate toxicity and alteration of gene expression associated with cadmium stress in chickpea (Cicer arietinum L.) shoots.

In the aim to estimate the protective role of calcium (Ca) and ethylene glycol tetraacetic acid (EGTA) against cadmium (Cd)-induced damage, chickpea (Cicer arietinum L.) seeds were exposed to 200 µM Cd stress for 6 days or 3 days then subjected to co-treatment of the metal with either 100 mM CaCl2 or 100 µM EGTA for 3 additional days. The addition of Ca and EGTA improved seedling growth. This protecting effect was correlated to the alleviation of the metal-induced oxidative stress, exemplified by the reduction of hydrogen peroxide (H2O2) contents. Besides, Ca and EGTA stimulated thioredoxin (Trx) and thioredoxin reductase (NTR) activities (2.75- and 1.75-fold increase when compared to Cd-stressed, respectively) protecting, thereby, protein -SH groups from the Cd-mediated oxidation, and modulated ferredoxin (Fdx) activity to a control level. Moreover, Ca and EGTA reinstated the glutathione redox steady state, mainly via preserving a high level of glutathione reduced form (GSH). This effect coincided with the maintaining of the Cd-stimulated glutathione reductase (GR) activity and the decline of glutathione peroxidase (GPX, 43% lower than Cd-stressed shoots) activity. Ca and EGTA counteracted the inhibitory effect of Cd on the activity and gene expression of Cu/Zn-superoxide dismutase (Cu/Zn-SOD) isoenzyme and modulated the activities of catalase (CAT) and ascorbate peroxidase (APX). Overall, our results provided evidence that Ca and EGTA supplement could be a promising approach in the remediation of Cd-contaminated environment.

Nitric oxide and hydrogen sulfide protect plasma membrane integrity and mitigate chromium-induced methylglyoxal toxicity in maize seedlings.

The present study aims to analyse the potential crosstalk between nitric oxide (NO) and hydrogen sulfide (H2S) in triggering resilience of maize (Zea mays L.) seedlings to hexavalent chromium (Cr VI). Exogenous application of 500 µM sodium nitroprusside (SNP, as a NO donor) or sodium hydrosulfide (NaHS, as a H2S donor) to 9-day-old maize seedlings, countered a Cr (200 µM) -elicited reduction in embryonic axis biomass. Cr caused cellular membrane injury by enhancing the levels of superoxide and hydroxyl radicals as well as methylglyoxal, and 4-hydroxy-2-nonenal. The application of SNP or NaHS considerably improved the endogenous NO and H2S pool, decreased oxidative stress and lipid peroxidation by suppressing lipoxygenase activity and improving some antioxidant enzymes activities in radicles and epicotyls. Radicles were more affected than epicotyls by Cr-stress with enhanced electrolyte leakage and decreased proton extrusion as indicated by lesser H+-ATPase activity. H2S appeared to mitigate Cr toxicity through up-regulated H+-ATPase and glyoxalase pathways and by maintaining optimal GSH levels as downstream effects of ROS and MG suppression. Hence, H2S-mediated the regeneration of GSH pool is associated with the attenuation of MG toxicity by enhancing S-lactoglutathione and D-lactate production. Taken together, our results indicate complementary roles for H2S and GSH to strengthen membrane integrity against Cr stress in maize seedlings.

Exogenous application of hydrogen sulfide reduces chromium toxicity in maize seedlings by suppressing NADPH oxidase activities and methylglyoxal accumulation.

Chromium (Cr) represents an important source of metallic stress in plants. Working with maize (Zea mays) seedlings, we characterize the suppressive effects of exogenously applied NaHS (a hydrogen sulfide; [H2S] donor) on the toxic effects of Cr (VI). Heavy metal treatment reduced radicle and epicotyl lengths and fresh weights in seedlings at 6 and 9 days following germination. The negative Cr (200 µM) effect was countered by application with NaHS (500 µM) but this countering was reduced with the co-application of the H2S generation inhibitor hydroxylamine (HA) or the H2S scavenger hypotaurine (HT). The Cr-elicited H2O2 production was suppressed by NaHS and also by an inhibitor of the reactive oxygen species (ROS) generating NADPH oxidase (NOX). These effects were correlated with relative changes in carbomyl (-CO) and thiol (-SH) groups. Nitric oxide (NO) production increased by NaHS application with associated increase in S-nitrosoglutathione (GSNO) level, but low S-nitrosoglutathione reductase (GSNOR) activities indicating an elevated S-nitrosylation. Assessment of the role of the ascorbate-glutathione antioxidant cycle indicated that whilst ascorbate played at a best minor role, glutathione was more prominent. Methylglyoxal (MG) production was increased by Cr but reduced by NaHS through a mechanism which could be based on glutathione-S-transferase (GST) detoxification. Taken together data suggest that H2S acts to counter Cr effect in maize by NOX suppression, mostly likely by the well-characterised S-nitrosylation mechanism, as well as a reduction of MG accumulation.

Efficiency of the Green Synthesized Nanoparticles as New Tools in Cancer Therapy: Insights on Plant-Based Bioengineered Nanoparticles, Biophysical Properties, and Anticancer Roles.

The aim of this work is to review the current knowledge on the efficiency of plant-based synthesized nanoparticles in medical field, particularly in the prevention, diagnosis, and therapy of cancer. For this, we examine the advantages of nanotechnological tools. Besides, a particular attention was given to understand the mechanism by which plant-based bioengineered nanoparticles can interact with components of cancerous cells. Green biosynthesized nanoparticles seem to be novel tool for prognostic biomarkers for cancer diagnosis and drug delivery in tumor cells. They can act either by leading to the damage of tumor cells, or by the protection of healthy cells, via mechanisms involving the specific properties of nanoparticles themselves and the antioxidative and antitumor properties found in plants. However, special attention should be given to the choice of plant species, extracts, and the toxic dose of some phytocompounds during the biosynthesis process. An increase in metal or trace element release from metal and metal oxide biosynthesized nanoparticles can lead to greater oxidative stress, which is associated with higher risk of cancer. Hence, plant-based nanosystems should be more developed to increase their specific targeting of the cancerous cells, in order to preserve the healthy ones.

Cadmium effects on embryo growth of pea seeds during germination: Investigation of the mechanisms of interference of the heavy metal with protein mobilization-related factors.

This work aims to give more insight into mechanisms of action of cadmium (Cd) on germinating pea seeds (Pisum sativum L. var. douce province), specifically the different ways by which Cd cations may interfere with the principal factors involved during germination process, notably storage proteins mobilization, amino acids freeing and proteolytic activities. Obtained results revealed that the process of hydrolysis of main storage proteins showed a significant disruption, which resulted in the decrease of the release of free amino acids, thus imposing a lack in nitrogen supply of essential nutrients to growing embryo under Cd stress. This hypothesis was evidenced by Cd-induced changes occurring in main purified protein fractions; Albumins, Legumins and Vicilins, during their breakdown. Besides, at enzymatic level, the activities of main proteases responsible for this hydrolysis were altered. Indeed, assays using synthetic substrates and specific protease inhibitors followed by protease activity measurements demonstrated that Cd inhibited drastically the total azocaseinolytic activity (ACA) and activities of different proteolytic classes: cysteine-, aspartic-, serine- and metallo-endopeptidases (EP), leucine- and proline-aminopeptidases (LAP and PAP, respectively), and glycine-carboxypeptidases (Gly-CP). The data here presented may suggest that the vulnerability of the embryonic axes towards Cd toxicity could be explained as a result of eventual disruption of metabolic pathways that affect mobilization of reserves and availability of nutrients. In vitro studies suggest that Cd cations may act either directly on the catalytic sites of the proteolytic enzymes, which may cause their deactivation, or indirectly via the generation of oxidative stress and overproduction of free radicals that can interact with enzymes, by altering their activity and structure.

Protective role of exogenous phytohormones on redox status in pea seedlings under copper stress.

The present work aims to provide insight on the role of phytohormone application in developing efficient practical defense strategies to improve plants tolerance under heavy metal contamination. For this purpose, pea (Pisum sativum L.) seeds were germinated in an aqueous solution of 200µM CuCl2 up to the 3rd day and then continued to germinate in the presence of distilled water (stress cessation) or were subjected to following combinations: Cu+1µM IAA and Cu+1µM GA3 for 3 additional days. The results showed that copper excess induced oxidative stress in germinating seeds, which resulted in changes of the redox state of glutathione and cysteine, and proteomics revealed Cu-induced modifications of thiols (SH) and carbonyls (CO) (indicators of protein oxidation). However, application of IAA or GA3 in the germination medium after 3days of Cu exposure alleviated toxicity on seedlings, despite the persistence of Cu up to 6th day. This improving effect seems to be mediated by a cell Cu accumulation decrease and a protein reduced status recovery, since phytohormones modulate thioredoxin/ferredoxin systems in favor of protecting proteins against oxidation. In addition, an IAA and GA3 protective effect was evidenced by a cellular homeostasis amelioration resulting from the balance conservation between the regeneration and consumption processes of glutathione and cysteine reduced forms. The exogenous effectors also induced modifications of profiles of SH and CO, suggesting changes in the regulation and expression of proteins that could be involved in defense mechanism against Cu stress.

Redox biology response in germinating Phaseolus vulgaris seeds exposed to copper: Evidence for differential redox buffering in seedlings and cotyledon.

In agriculture, heavy metal contamination of soil interferes with processes associated with plant growth, development and productivity. Here, we describe oxidative and redox changes, and deleterious injury within cotyledons and seedlings caused by exposure of germinating (Phaseolus vulgaris L. var. soisson nain hâtif) seeds to copper (Cu). Cu induced a marked delay in seedling growth, and was associated with biochemical disturbances in terms of intracellular oxidative status, redox regulation and energy metabolism. In response to these alterations, modulation of activities of antioxidant proteins (thioredoxin and glutathione reductase, peroxiredoxin) occurred, thus preventing oxidative damage. In addition, oxidative modification of proteins was detected in both cotyledons and seedlings by one- and two-dimensional electrophoresis. These modified proteins may play roles in redox buffering. The changes in activities of redox proteins underline their fundamental roles in controlling redox homeostasis. However, observed differential redox responses in cotyledon and seedling tissues showed a major capacity of the seedlings' redox systems to protect the reduced status of protein thiols, thus suggesting quantitatively greater antioxidant protection of proteins in seedlings compared to cotyledon. To our knowledge, this is the first comprehensive redox biology investigation of the effect of Cu on seed germination.

Alleviation of Cr(VI)-induced oxidative stress in maize (Zea mays L.) seedlings by NO and H2S donors through differential organ-dependent regulation of ROS and NADPH-recycling metabolisms.

Chromium (Cr) contamination in soil is a growing concern in relation to sustainable agricultural production and food safety. Nitric oxide (NO) and, more recently, hydrogen sulfide (H2S) are considered to be new signalling molecules with biotechnological applications in the agronomical sector. Using 9-day-old maize (Zea mays) seedlings exposed to 200µM Cr(VI), the potential mitigating effects of exogenous NO and H2S on chromium-induced stress in maize seedlings were investigated in roots, cotyledons and coleoptiles. Analysis of Cr content, lipid peroxidation, antioxidant enzymes (catalase and superoxide dismutase isozymes), peroxisomal H2O2-producing glycolate oxidase and the main NADPH-regenerating system revealed that chromium causes oxidative stress, leading to a general increase in these activities in coleptiles and roots, with the latter organ being the most affected. However, cotyledons behaved in an opposite manner. Moreover, exogenous applications of NO and H2S to Cr-stressed maize seedlings triggered a significant response, involving the virtual restoration of the values for all these activities to those observed in unstressed seedlings, although their specific impact on ROS and NADPH-recycling metabolisms depends on the seedling organ involved. Taken together, the data indicate that gas transmitters, NO and H2S, which act as a defence against the negative effects of hexavalent chromium contamination, are alternative compounds with potential biotechnological applications.

Cadmium-induced changes in antioxidative systems and differentiation in roots of contrasted Medicago truncatula lines.

Defense pathways and stress responses induced under Cd stress were illustrated in roots of hydroponically grown Medicago truncatula seedlings. Actually, the ascorbate-glutathione and antioxidative system, secondary metabolism events including peroxidases, phenolic compounds, and lignification launching, and developmental modifications were described. Cd (100 µM) initially increased reactive oxygen species, enhanced antioxidative (total SOD, CAT, and PRX) and ascorbate-glutathione-related metabolism enzymes (APX and MDAR), except in A17 and TN1.11. In agreement with peroxidase enhancement, physiological measurement and in situ observation illustrated soluble phenolic compound accumulation under Cd treatment. However, lignification was restricted to recently created protoxylem elements established in the root tip area, usually constituting the elongation zone. Cell death was increased. In the absence of necrotic reactions, developmental changes including lignin deposition, increase in cellulose and pectin contents, intercellular meatus, and condensed and deformed hairs were noticed in Cd-treated roots.

Effect of cadmium pollution on mobilization of embryo reserves in seedlings of six contrasted Medicago truncatula lines.

Six Medicago truncatula genotypes differing in cadmium susceptibility were used to test the effect of this heavy metal on mineral, carbohydrate and amino acid supply in growing radicles. Cadmium treatment caused alteration of macronutrient (Ca and K), microelement (Fe, Zn and Cu), carbohydrate (total soluble sugars (TSS), glucose, fructose and sucrose) and free amino acid (FAAS) accumulations. These mobilization changes differed in the tested genotypes. Carbohydrates were determining to susceptible lines' growth in control condition; free amino acids enabled tolerant lines to counteract cadmium intrusion. Transcriptional changes in response to cadmium treatment were analyzed on MtMST, a gene encoding a monosaccharide transport protein. A significant down-regulation was observed in the most susceptible line TN1.11. Glucose was over-consumed in tolerant lines. Thus, glucose metabolism integrity seems essential to maintain growth under cadmium exposure. Analysis of germination medium showed solute losses at the expense of suitable mobilization to the growing embryonic axis and highlights cadmium-triggered membrane alterations. FAAS and TSS leakages were reduced in tolerant lines while monosaccharide losses were accentuated in susceptible lines. This research work gave an overview of cadmium deleterious effects on biomass mobilization and membrane integrity. Carbon metabolism is shown to be primordial to enhance early embryonic growth and nitrogen metabolism is revealed to be crucial to establish seedling growth under cadmium stress.

Effects of copper on reserve mobilization in embryo of Phaseolus vulgaris L.

The present research reports a biochemical and micro-submicroscopic analysis of copper effect on reserve mobilization during germination of Phaseolus vulgaris L. var. soisson nain hatif seeds. Dry embryonic cells are rich in protein bodies and little starch grains. In Cu-treated embryos copper inhibited 50% of albumin and globulin mobilization after 72 h imbibition. The severe alterations in treated embryo cells, observed by electron microscope, were probably the cause of the inability to utilize the amino acids freed by protein mobilization and so possibly the cause of the inhibition of P. vulgaris embryonic axis elongation.

Responses of proteolytic enzymes in embryonic axes of germinating bean seeds under copper stress.

The changes in protease activities in embryonic axes during the first days of bean (Phaseolus vulgaris L.) seed germination were investigated in response to copper stress. Synthetic substrates and specific protease inhibitors have been used to define qualitatively and quantitatively different catalytic classes, particularly endoproteases (EP), carboxypeptidases (CP) and aminopeptidases (AP), then identify which ones were affected in the presence of copper. In fact, a failure in storage proteins mobilization and a disorder of nitrogen supply at enzymatic level occurred in Cu. In fact, Cu inhibited azocaseinolytic activity (ACA) and cysteine-, aspartic-, serine-, and metallo-endopeptidases activities (Cys-EP, Asp-EP, Ser-Ep, and Met-EP, respectively). Besides, Cu affected leucine- and proline-aminopeptidases (LAP and PAP, respectively) and glycine-carboxypeptidases (Gly-CP). The proteolytic responses might also be associated with the decrease in defense capacity in the Cu-treated embryos.

Oxidative injury and antioxidant genes regulation in cadmium-exposed radicles of six contrasted Medicago truncatula genotypes.

Oxidative disorders were triggered in the presence of Cd toxicity in early seedling growth of six Medicago truncatula genotypes. Interactions between root growth inhibition, cadmium uptake, as well as the occurrence of oxidative injury suggest differential responses of the genotypes, with susceptible or tolerant accessions. ROS enhancement was observed in situ and H2O2 content was measured, that did not seem related to tolerance or susceptibility. Oxidative burst impact on cell membrane integrity was analyzed in agreement with MDA content and glucose exudation, which suggest an active role of this burst in susceptible lines. Transcriptional changes in response to cadmium treatment were analyzed on target genes involved in (1) ROS-scavenging enzymes (superoxide dismutase (SOD; EC1.15.1.1) and peroxidase (PRX; EC 1.11.1.7)), (2) reduced glutathione (γ-Glu-Cys-Gly, GSH) metabolism (glutathione-S-transferase (GST; EC: 2.5.1.18) and glutathione reductase (GR; EC 1.8.1.7)), and (3) metal-chelating metabolism (PCS). The susceptible line shows no response or non-timely gene expression patterns. This research work gave an overview of the deleterious effects and oxidative injury of cadmium stress in Medicago truncatula. Oxidative defense efficiency and gene upregulation should explain relative tolerance in tested genotypes.