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.

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.

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.

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.

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.