Characterization of leucine aminopeptidase (LAP) activity in sweet pepper fruits during ripening and its inhibition by nitration and reducing events.

KEY MESSAGE: Pepper fruits contain two leucine aminopeptidase (LAP) genes which are differentially modulated during ripening and by nitric oxide. The LAP activity increases during ripening but is negatively modulated by nitration. Leucine aminopeptidase (LAP) is an essential metalloenzyme that cleaves N-terminal leucine residues from proteins but also metabolizes dipeptides and tripeptides. LAPs play a fundamental role in cell protein turnover and participate in physiological processes such as defense mechanisms against biotic and abiotic stresses, but little is known about their involvement in fruit physiology. This study aims to identify and characterize genes encoding LAP and evaluate their role during the ripening of pepper (Capsicum annuum L.) fruits and under a nitric oxide (NO)-enriched environment. Using a data-mining approach of the pepper plant genome and fruit transcriptome (RNA-seq), two LAP genes, designated CaLAP1 and CaLAP2, were identified. The time course expression analysis of these genes during different fruit ripening stages showed that whereas CaLAP1 decreased, CaLAP2 was upregulated. However, under an exogenous NO treatment of fruits, both genes were downregulated. On the contrary, it was shown that during fruit ripening LAP activity increased by 81%. An in vitro assay of the LAP activity in the presence of different modulating compounds including peroxynitrite (ONOO-), NO donors (S-nitrosoglutathione and nitrosocyteine), reducing agents such as reduced glutathione (GSH), L-cysteine (L-Cys), and cyanide triggered a differential response. Thus, peroxynitrite and reducing compounds provoked around 50% inhibition of the LAP activity in green immature fruits, whereas cyanide upregulated it 1.5 folds. To our knowledge, this is the first characterization of LAP in pepper fruits as well as of its regulation by diverse modulating compounds. Based on the capacity of LAP to metabolize dipeptides and tripeptides, it could be hypothesized that the LAP might be involved in the GSH recycling during the ripening process.

Aquaporins: a vital nexus in H2O2-gasotransmitter signaling.

Land plants have evolved with a complex mechanism of water uptake facilitated by the activity of aquaporins under normal and challenging environments. However, we lack a clear understanding of its interactions with reactive oxygen species, particularly hydrogen peroxide (H2O2) and the gasotransmitters nitric oxide (NO) and hydrogen sulfide (H2S), under oxidative stress. Here, we assess the crosstalk of aquaporin function, H2O2 homeostasis, and NO-H2S signaling in plants and provide a computational prediction of cysteine-based oxidative post-translational modifications (oxiPTMs) in plant aquaporins. We propose that aquaporin activity could be regulated by three major oxiPTMs, S-nitrosation, S-sulfenylation, and persulfidation, mediated by NO, H2O2, and H2S, respectively. Therefore, aquaporins might be key players in the gasotransmitter-mediated long-distance oxidative stress signals in plant cells.

In Silico RNAseq and Biochemical Analyses of Glucose-6-Phosphate Dehydrogenase (G6PDH) from Sweet Pepper Fruits: Involvement of Nitric Oxide (NO) in Ripening and Modulation.

Pepper (Capsicum annuum L.) fruit is a horticultural product consumed worldwide which has great nutritional and economic relevance. Besides the phenotypical changes that pepper fruit undergo during ripening, there are many associated modifications at transcriptomic, proteomic, biochemical, and metabolic levels. Nitric oxide (NO) is a recognized signal molecule that can exert regulatory functions in diverse plant processes including fruit ripening, but the relevance of NADPH as a fingerprinting of the crop physiology including ripening has also been proposed. Glucose-6-phosphate dehydrogenase (G6PDH) is the first and rate-limiting enzyme of the oxidative phase of the pentose phosphate pathway (oxiPPP) with the capacity to generate NADPH. Thus far, the available information on G6PDH and other NADPH-generating enzymatic systems in pepper plants, and their expression during the ripening of sweet pepper fruit, is very scarce. Therefore, an analysis at the transcriptomic, molecular and functional levels of the G6PDH system has been accomplished in this work for the first time. Based on a data-mining approach to the pepper genome and fruit transcriptome (RNA-seq), four G6PDH genes were identified in pepper plants and designated CaG6PDH1 to CaG6PDH4, with all of them also being expressed in fruits. While CaG6PDH1 encodes a cytosolic isozyme, the other genes code for plastid isozymes. The time-course expression analysis of these CaG6PDH genes during different fruit ripening stages, including green immature (G), breaking point (BP), and red ripe (R), showed that they were differentially modulated. Thus, while CaG6PDH2 and CaG6PDH4 were upregulated at ripening, CaG6PDH1 was downregulated, and CaG6PDH3 was slightly affected. Exogenous treatment of fruits with NO gas triggered the downregulation of CaG6PDH2, whereas the other genes were positively regulated. In-gel analysis using non-denaturing PAGE of a 50-75% ammonium-sulfate-enriched protein fraction from pepper fruits allowed for identifying two isozymes designated CaG6PDH I and CaG6PDH II, according to their electrophoretic mobility. In order to test the potential modulation of such pepper G6PDH isozymes, in vitro analyses of green pepper fruit samples in the presence of different compounds including NO donors (S-nitrosoglutathione and nitrosocysteine), peroxynitrite (ONOO-), a hydrogen sulfide (H2S) donor (NaHS, sodium hydrosulfide), and reducing agents such as reduced glutathione (GSH) and L-cysteine (L-Cys) were assayed. While peroxynitrite and the reducing compounds provoked a partial inhibition of one or both isoenzymes, NaHS exerted 100% inhibition of the two CaG6PDHs. Taken together these data provide the first data on the modulation of CaG6PDHs at gene and activity levels which occur in pepper fruit during ripening and after NO post-harvest treatment. As a consequence, this phenomenon may influence the NADPH availability for the redox homeostasis of the fruit and balance its active nitro-oxidative metabolism throughout the ripening process.

Pepper Fruit Extracts Show Anti-Proliferative Activity against Tumor Cells Altering Their NADPH-Generating Dehydrogenase and Catalase Profiles.

Cancer is considered one of the main causes of human death worldwide, being characterized by an alteration of the oxidative metabolism. Many natural compounds from plant origin with anti-tumor attributes have been described. Among them, capsaicin, which is the molecule responsible for the pungency in hot pepper fruits, has been reported to show antioxidant, anti-inflammatory, and analgesic activities, as well as anti-proliferative properties against cancer. Thus, in this work, the potential anti-proliferative activity of pepper (Capsicum annuum L.) fruits from diverse varieties with different capsaicin contents (California < Piquillo < Padrón < Alegría riojana) against several tumor cell lines (lung, melanoma, hepatoma, colon, breast, pancreas, and prostate) has been investigated. The results showed that the capsaicin content in pepper fruits did not correspond with their anti-proliferative activity against tumor cell lines. By contrast, the greatest activity was promoted by the pepper tissues which contained the lowest capsaicin amount. This indicates that other compounds different from capsaicin have this anti-tumor potentiality in pepper fruits. Based on this, green fruits from the Alegría riojana variety, which has negligible capsaicin levels, was used to study the effect on the oxidative and redox metabolism of tumor cell lines from liver (Hep-G2) and pancreas (MIA PaCa-2). Different parameters from both lines treated with crude pepper fruit extracts were determined including protein nitration and protein S-nitrosation (two post-translational modifications (PTMs) promoted by nitric oxide), the antioxidant capacity, as well as the activity of the antioxidant enzymes superoxide dismutase (SOD), catalase, and glutathione peroxidase (GPX), among others. In addition, the activity of the NADPH-generating enzymes glucose-6-phosphate dehydrogenase (G6PDH), 6-phosphogluconate dehydrogenase (6PGDH), and NADP-isocitrate dehydrogenase (NADP-ICDH) was followed. Our data revealed that the treatment of both cell lines with pepper fruit extracts altered their antioxidant capacity, enhanced their catalase activity, and considerably reduced the activity of the NADPH-generating enzymes. As a consequence, less H2O2 and NADPH seem to be available to cells, thus avoiding cell proliferation and possibly triggering cell death in both cell lines.

Zinc induced regulation of PCR1 gene for cadmium stress resistance in rice roots.

In this study, the interaction between zinc (Zn) and cadmium (Cd) was investigated in rice roots to evaluate how Zn can protect the plants from Cd stress. Rice seedlings were treated with Cd (100 µM) and Zn (100 µM) in different combinations (Cd alone, Zn alone, Zn+ Cd, Zn+ Cd+ L-NAME, Zn+ Cd+ L-NAME+ SNP). Rice roots treated with only Zn also displayed similar toxic effects, however when combined with Cd exhibited improved growth. Treating the plant with Zn along with Cd distinctly reduced Cd concentration in roots while increasing its own accumulation due to modulation in expression of Zinc-Regulated Transporter (ZRT)-/IRT-Like Protein (OsZIP1) and Plant Cadmium Resistance1 (OsPCR1). Cd reduced plant biomass, cell viability, pigments, photosynthesis and causing oxidative stress due to inhibition in ascorbate-glutathione cycle. L-NAME (NG-nitro L-arginine methyl ester), prominently suppressed the beneficial impacts of Zn against Cd stress, whereas the presence of a NO donor, sodium nitroprusside (SNP), significantly reversed this effect of L-NAME. Collectively, results point that NO signalling is essential for Zn- mediated cross-tolerance against Cd stress via by modulating uptake of Cd and Zn and expression of OsZIP1 and OsPCR1, and ROS homeostasis due to fine tuning of ascorbate-glutathione cycle which finally lessened oxidative stress in rice roots. The results of this study can be utilized to develop new varieties of rice through genetic modifications which will be of great significance for maintaining crop productivity in Cd-contaminated areas throughout the world.

Soybean (Glycine max L.) Lipoxygenase 1 (LOX 1) Is Modulated by Nitric Oxide and Hydrogen Sulfide: An In Vitro Approach.

Hydrogen sulfide (H2S) and nitric oxide (NO) are two relevant signal molecules that can affect protein function throughout post-translational modifications (PTMs) such as persulfidation, S-nitrosation, metal-nitrosylation, and nitration. Lipoxygenases (LOXs) are a group of non-heme iron enzymes involved in a wide range of plant physiological functions including seed germination, plant growth and development, and fruit ripening and senescence. Likewise, LOXs are also involved in the mechanisms of response to diverse environmental stresses. Using purified soybean (Glycine max L.) lipoxygenase type 1 (LOX 1) and nitrosocysteine (CysNO) and sodium hydrosulfide (NaHS) as NO and H2S donors, respectively, the present study reveals that both compounds negatively affect LOX activity, suggesting that S-nitrosation and persulfidation are involved. Mass spectrometric analysis of nitrated soybean LOX 1 using a peroxynitrite (ONOO-) donor enabled us to identify that, among the thirty-five tyrosine residues present in this enzyme, only Y214 was exclusively nitrated by ONOO-. The nitration of Y214 seems to affect its interaction with W500, a residue involved in the substrate binding site. The analysis of the structure 3PZW demonstrates the existence of several tunnels that directly communicate the surface of the protein with different internal cysteines, thus making feasible their potential persulfidation, especially C429 and C127. On the other hand, the CysNO molecule, which is hydrophilic and bulkier than H2S, can somehow be accommodated throughout the tunnel until it reaches C127, thus facilitating its nitrosation. Overall, a large number of potential persulfidation targets and the ease by which H2S can reach them through the diffuse tunneling network could be behind their efficient inhibition.

Silicon nanoparticles (SiNPs) restore photosynthesis and essential oil content by upgrading enzymatic antioxidant metabolism in lemongrass (Cymbopogon flexuosus) under salt stress.

Lemongrass (Cymbopogon flexuosus) has great relevance considering the substantial commercial potential of its essential oil. Nevertheless, the increasing soil salinity poses an imminent threat to lemongrass cultivation given its moderate salt-sensitivity. For this, we used silicon nanoparticles (SiNPs) to stimulate salt tolerance in lemongrass considering SiNPs special relevance to stress settings. Five foliar sprays of SiNPs 150 mg L-1 were applied weekly to NaCl 160 and 240 mM-stressed plants. The data indicated that SiNPs minimised oxidative stress markers (lipid peroxidation, H2O2 content) while triggering a general activation of growth, photosynthetic performance, enzymatic antioxidant system including superoxide dismutase (SOD), catalase (CAT), and peroxidase (POD), and osmolyte proline (PRO). SiNPs amplified stomatal conductance and photosynthetic CO2 assimilation rate by about 24% and 21% in NaCl 160 mM-stressed plants. Associated benefits contributed to pronounced plant phenotype over their stressed counterparts, as we found. Foliar SiNPs sprays assuaged plant height by 30% and 64%, dry weight by 31% and 59%, and leaf area by 31% and 50% under NaCl 160 and 240 mM concentrations, respectively. SiNPs relieved enzymatic antioxidants (SOD, CAT, POD) and osmolyte (PRO) in lemongrass plants stressed with NaCl 160 mM (9%, 11%, 9%, and 12%, respectively) and NaCl 240 mM (13%, 18%, 15%, and 23%, respectively). The same treatment supported the oil biosynthesis improving essential oil content by 22% and 44% during 160 and 240 mM salt stress, respectively. We found SiNPs can completely overcome NaCl 160 mM stress while significantly palliating NaCl 240 mM stress. Thus, we propose that SiNPs can be a useful biotechnological tool to palliate salinity stress in lemongrass and related crops.

H2S-Generating Cytosolic L-Cysteine Desulfhydrase and Mitochondrial D-Cysteine Desulfhydrase from Sweet Pepper (Capsicum annuum L.) Are Regulated During Fruit Ripening and by Nitric Oxide.

Aims: Pepper fruit is a horticultural product worldwide consumed that has great nutritional and economic relevance. Besides the phenotypical changes that undergo pepper fruit during ripening, there are many associated modifications at transcriptomic, proteomic, biochemical, and metabolic levels. Nitric oxide (NO) and hydrogen sulfide (H2S) are recognized signal molecules that can exert regulatory functions in diverse plant processes. This study aims at analyzing the interrelationship between NO and H2S during fruit ripening. Results: Our data indicate that the H2S-generating cytosolic L-cysteine desulfhydrase (LCD) and the mitochondrial D-cysteine desulfhydrase (DCD) activities are downregulated during ripening but this effect was reverted after NO treatment of fruits. Innovation and Conclusion: Using as a model the non-climacteric pepper fruits at different ripening stages and under an NO-enriched atmosphere, the activity of the H2S-generating LCD and DCD was analyzed. LCD and DCD activities were downregulated during ripening, but this effect was reverted after NO treatment of fruits. The analysis of LCD activity by non-denaturing polyacrylamide gel electrophoresis (PAGE) allowed identifying three isozymes designated CaLCD I to CaLCD III, which were differentially modulated by NO and strictly dependent on pyridoxal 5'-phosphate (PLP). In vitro analyses of green fruit samples in the presence of different compounds including NO donors, peroxynitrite (ONOO-), and reducing agents such as reduced glutathione (GSH) and L-cysteine (L-Cys) triggered an almost 100% inhibition of CaLCD II and CaLCD III. This redox adaptation process of both enzymes could be cataloged as a hormesis phenomenon. The protein tyrosine (Tyr) nitration (an NO-promoted post-translational modification) of the recombinant LCD was corroborated by immunoblot and by mass spectrometry (MS) analyses. Among the 11 Tyr residues present in this enzyme, MS of the recombinant LCD enabled us to identify that Tyr82 and Tyr254 were nitrated by ONOO-, this occurring near the active center on the enzyme, where His237 and Lys260 together with the cofactor PLP are involved. These data support the relationship between NO and H2S during pepper fruit ripening, since LCD and DCD are regulated by NO during this physiological event, and this could also be extrapolated to other plant species.

Functions of NO and H2S Signal Molecules Against Plant Abiotic Stress.

Nitric oxide (NO) and hydrogen sulfide (H2S) are two recognized signal molecules in higher plants involved in a wide range of physiological processes and the mechanisms of response against adverse environmental conditions. These molecules can interact to provide an adequate response to palliate the negative impact exerted by stressful conditions, particularly by regulating key components of the metabolism of reactive oxygen species (ROS) to avoid their overproduction and further oxidative damage which, finally, affects cellular functioning. NO and H2S can exert the regulation over the function of susceptible proteins by posttranslational modifications (PTMs) including nitration, S-nitrosation, and persulfidation but also through the regulation of gene expression by the induction of specific transcription factors which modulate the expression of genes encoding proteins related to stress resistance. This chapter encompasses a wide perspective of the signaling and functional relationships between NO and H2S to modulate the overproduction of reactive oxygen species, particularly under abiotic stress conditions.

Analysis of Plant L-Cysteine Desulfhydrase (LCD) Isozymes by Non-denaturing Polyacrylamide Gel Electrophoresis.

Hydrogen sulfide (H2S) is a signaling molecule that achieves different regulatory functions in animal and plant cells. The cytosolic enzyme L-cysteine desulfhydrase (LCD; EC 4.4.1.28) catalyzes the conversion of cysteine (L-Cys) to pyruvate and ammonium with the concomitant generation of H2S, this enzyme being considered one of the main sources of H2S in higher plants. Using non-denaturing polyacrylamide gel electrophoresis (PAGE) in combination with a specific assay for LCD activity, the present protocol allows identifying diverse LCD isozymes present in different organs (roots, shoots, leaves, and fruits) and plant species including pea, garlic, Arabidopsis, and pepper.

Lipoxygenase (LOX) in Sweet and Hot Pepper (Capsicum annuum L.) Fruits during Ripening and under an Enriched Nitric Oxide (NO) Gas Atmosphere.

Lipoxygenases (LOXs) catalyze the insertion of molecular oxygen into polyunsaturated fatty acids (PUFA) such as linoleic and linolenic acids, being the first step in the biosynthesis of a large group of biologically active fatty acid (FA)-derived metabolites collectively named oxylipins. LOXs are involved in multiple functions such as the biosynthesis of jasmonic acid (JA) and volatile molecules related to the aroma and flavor production of plant tissues, among others. Using sweet pepper (Capsicum annuum L.) plants as a model, LOX activity was assayed by non-denaturing polyacrylamide gel electrophoresis (PAGE) and specific in-gel activity staining. Thus, we identified a total of seven LOX isozymes (I to VII) distributed among the main plant organs (roots, stems, leaves, and fruits). Furthermore, we studied the FA profile and the LOX isozyme pattern in pepper fruits including a sweet variety (Melchor) and three autochthonous Spanish varieties that have different pungency levels (Piquillo, Padrón, and Alegría riojana). It was observed that the number of LOX isozymes increased as the capsaicin content increased in the fruits. On the other hand, a total of eight CaLOX genes were identified in sweet pepper fruits, and their expression was differentially regulated during ripening and by the treatment with nitric oxide (NO) gas. Finally, a deeper analysis of the LOX IV isoenzyme activity in the presence of nitrosocysteine (CysNO, a NO donor) suggests a regulatory mechanism via S-nitrosation. In summary, our data indicate that the different LOX isozymes are differentially regulated by the capsaicin content, fruit ripening, and NO.

H2S in Horticultural Plants: Endogenous Detection by an Electrochemical Sensor, Emission by a Gas Detector, and Its Correlation with L-Cysteine Desulfhydrase (LCD) Activity.

H2S has acquired great attention in plant research because it has signaling functions under physiological and stress conditions. However, the direct detection of endogenous H2S and its potential emission is still a challenge in higher plants. In order to achieve a comparative analysis of the content of H2S among different plants with agronomical and nutritional interest including pepper fruits, broccoli, ginger, and different members of the genus Allium such as garlic, leek, Welsh and purple onion, the endogenous H2S and its emission was determined using an ion-selective microelectrode and a specific gas detector, respectively. The data show that endogenous H2S content range from pmol to µmol H2S · g-1 fresh weight whereas the H2S emission of fresh-cut vegetables was only detected in the different species of the genus Allium with a maximum of 9 ppm in garlic cloves. Additionally, the activity and isozymes of the L-cysteine desulfhydrase (LCD) were analyzed, which is one of the main enzymatic sources of H2S, where the different species of the genus Allium showed the highest activities. Using non-denaturing gel electrophoresis, the data indicated the presence of up to nine different LCD isozymes from one in ginger to four in onion, leek, and broccoli. In summary, the data indicate a correlation between higher LCD activity with the endogenous H2S content and its emission in the analyzed horticultural species. Furthermore, the high content of endogenous H2S in the Allium species supports the recognized benefits for human health, which are associated with its consumption.

Thiol-based Oxidative Posttranslational Modifications (OxiPTMs) of Plant Proteins.

The thiol group of cysteine (Cys) residues, often present in the active center of the protein, is of particular importance to protein function, which is significantly determined by the redox state of a protein's environment. Our knowledge of different thiol-based oxidative posttranslational modifications (oxiPTMs), which compete for specific protein thiol groups, has increased over the last 10 years. The principal oxiPTMs include S-sulfenylation, S-glutathionylation, S-nitrosation, persulfidation, S-cyanylation and S-acylation. The role of each oxiPTM depends on the redox cellular state, which in turn depends on cellular homeostasis under either optimal or stressful conditions. Under such conditions, the metabolism of molecules such as glutathione, NADPH (reduced nicotinamide adenine dinucleotide phosphate), nitric oxide, hydrogen sulfide and hydrogen peroxide can be altered, exacerbated and, consequently, outside the cell's control. This review provides a broad overview of these oxiPTMs under physiological and unfavorable conditions, which can regulate the function of target proteins.

The Modus Operandi of Hydrogen Sulfide(H2S)-Dependent Protein Persulfidation in Higher Plants.

Protein persulfidation is a post-translational modification (PTM) mediated by hydrogen sulfide (H2S), which affects the thiol group of cysteine residues from target proteins and can have a positive, negative or zero impact on protein function. Due to advances in proteomic techniques, the number of potential protein targets identified in higher plants, which are affected by this PTM, has increased considerably. However, its precise impact on biological function needs to be evaluated at the experimental level in purified proteins in order to identify the specific cysteine(s) residue(s) affected. It also needs to be evaluated at the cellular redox level given the potential interactions among different oxidative post-translational modifications (oxiPTMs), such as S-nitrosation, glutathionylation, sulfenylation, S-cyanylation and S-acylation, which also affect thiol groups. This review aims to provide an updated and comprehensive overview of the important physiological role exerted by persulfidation in higher plants, which acts as a cellular mechanism of protein protection against irreversible oxidation.

Identification of Compounds with Potential Therapeutic Uses from Sweet Pepper (Capsicum annuum L.) Fruits and Their Modulation by Nitric Oxide (NO).

Plant species are precursors of a wide variety of secondary metabolites that, besides being useful for themselves, can also be used by humans for their consumption and economic benefit. Pepper (Capsicum annuum L.) fruit is not only a common food and spice source, it also stands out for containing high amounts of antioxidants (such as vitamins C and A), polyphenols and capsaicinoids. Particular attention has been paid to capsaicin, whose anti-inflammatory, antiproliferative and analgesic activities have been reported in the literature. Due to the potential interest in pepper metabolites for human use, in this project, we carried out an investigation to identify new bioactive compounds of this crop. To achieve this, we applied a metabolomic approach, using an HPLC (high-performance liquid chromatography) separative technique coupled to metabolite identification by high resolution mass spectrometry (HRMS). After chromatographic analysis and data processing against metabolic databases, 12 differential bioactive compounds were identified in sweet pepper fruits, including quercetin and its derivatives, L-tryptophan, phytosphingosin, FAD, gingerglycolipid A, tetrahydropentoxylin, blumenol C glucoside, colnelenic acid and capsoside A. The abundance of these metabolites varied depending on the ripening stage of the fruits, either immature green or ripe red. We also studied the variation of these 12 metabolites upon treatment with exogenous nitric oxide (NO), a free radical gas involved in a good number of physiological processes in higher plants such as germination, growth, flowering, senescence, and fruit ripening, among others. Overall, it was found that the content of the analyzed metabolites depended on the ripening stage and on the presence of NO. The metabolic pattern followed by quercetin and its derivatives, as a consequence of the ripening stage and NO treatment, was also corroborated by transcriptomic analysis of genes involved in the synthesis of these compounds. This opens new research perspectives on the pepper fruit's bioactive compounds with nutraceutical potentiality, where biotechnological strategies can be applied for optimizing the level of these beneficial compounds.

Crosstalk between abscisic acid and nitric oxide under heat stress: exploring new vantage points.

Heat stress adversely affects plants growth potential. Global warming is reported to increase in the intensity, frequency, and duration of heatwaves, eventually affecting ecology, agriculture and economy. With an expected increase in average temperature by 2-3 °C over the next 30-50 years, crop production is facing a severe threat to sub-optimum growth conditions. Abscisic acid (ABA) and nitric oxide (NO) are growth regulators that are involved in the adaptation to heat stress by affecting each other and changing the adaptation process. The interaction between these molecules has been discussed in various studies in general or under stress conditions; however, regarding high temperature, their interaction has little been worked out. In the present review, the focus is shifted on the role of these molecules under heat stress emphasizing the different possible interactions between ABA and NO as both regulate stomatal closure and other molecules including hydrogen peroxide (H2O2), hydrogen sulfide (H2S), antioxidants, proline, glycine betaine, calcium (Ca2+) and heat shock protein (HSP). Exploring the crosstalk between ABA and NO with other molecules under heat stress will provide us with a comprehensive knowledge of plants mechanism of heat tolerance which could be useful to develop heat stress-resistant varieties.

Silicon nanoparticles elicit an increase in lemongrass (Cymbopogon flexuosus (Steud.) Wats) agronomic parameters with a higher essential oil yield.

Lemongrass (Cymbopogon flexuosus (Steud.) Wats) is an aromatic grass with great industrial potential. It is cultivated for its essential oil (EO) which has great economical value due to its numerous medicinal, cosmetic and culinary applications. The present study was conducted on silicon nanoparticles (SiNPs) application to lemongrass with the objective of overall agronomic enhancements. Graded concentrations (50-200 mg L-1) of SiNPs were exogenously applied to lemongrass leaves. The physiological and biochemical analyses revealed that 150 mg L-1 SiNPs is the optimum concentration for lemongrass plants. This concentration triggered photosynthetic variables, gas exchange modules and activities of enzymes involved in EO (geraniol dehydrogenase) and nitrogen (nitrate reductase) metabolism as well as in the antioxidant system (catalase, peroxidase and superoxide dismutase). These SiNPs-induced metabolic changes altogether significantly (p ≤ 0.05) enhanced overall plant growth and yield. Moreover, SiNPs treatments assisted in palliating lipid peroxidation and H2O2 content in lemongrass leaves which added further advantage to plant metabolism. Overall, data indicates SiNPs elicit beneficial effects on lemongrass growth and yield through inducing various physiological and biochemical responses. This renders high possibility that similar objectives could be achieved with SiNPs biotechnological application on further related agronomic crops as well as in diverse industries.

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.

Appraisal of H2S metabolism in Arabidopsis thaliana: In silico analysis at the subcellular level.

Hydrogen sulfide (H2S) has become a new signal molecule in higher plants which seems to be involved in almost all physiological processes from seed germination, root and plant growth until flowering and fruit ripening. Moreover, H2S also participates in the mechanism of response against adverse environmental stresses. However, its basic biochemistry in plant cells can be considered in a nascent stage. Using the available information of the model plant Arabidopsis thaliana, the goal of the present study is to provide a broad overview of H2S metabolism and to display an in silico analysis of the 26 enzymatic components involved in the metabolism of H2S and their subcellular compartmentation (cytosol, chloroplast and mitochondrion) thus providing a wide picture of the cross-talk inside the organelles and amongst them and, consequently, to get a better understanding of the cellular and tissue implications of H2S. This information will be also relevant for other crop species, especially those whose whole genome is not yet available.

Melatonin and calcium function synergistically to promote the resilience through ROS metabolism under arsenic-induced stress.

The interplay between melatonin (Mel) and calcium (Ca2+) in enhancing tolerance to metalloid toxicity and underlying physiological and biochemical mechanisms of this relationship still remains unknown. The present study reveals that the signaling molecules Mel and/or Ca2+ enhanced tolerance of Vicia faba (cv. Tara) plant to metalloid arsenic (As) toxicity. However, a combination of Mel and Ca2+ was more efficient than alone. Plants grew with As exhibited enhanced hydrogen peroxide, superoxide anion, electrolyte leakage, lipid peroxidation together with increased reactive oxygen species (ROS) producing enzymes, such as NADPH oxidase and glycolate oxidase (GOX). On the contrary, an inhibition in chlorophyll (Chl) biosynthesis and gas exchange parameters (net photosynthetic rate, stomatal conductance, intercellular carbon dioxide concentration) was observed. Under As toxicity conditions, the application of Mel and Ca2+ synergistically suppressed the plants' program cell death features (nucleus condensation and nucleus fragmentation) in guard cells of stomata, DNA damage, and formation of ROS in guard cells, leaves and roots. Moreover, it enhanced gas exchange parameters and activity of enzymes involved in photosynthesis process (carbonic anhydrase and RuBisco), Chl biosynthesis (δ-aminolevulinic acid dehydratase), and decreased activity of Chl degrading enzyme (chlorophyllase) under As toxicity conditions. Our investigation evidently established that expression of ATP synthase, Ca2+-ATPase, Ca2+-DPKase, Hsp17.6 and Hsp40 was found maximum in the plants treated with Mel + Ca2+, resulting in higher tolerance of plants to As stress. Also, increased total soluble carbohydrates, cysteine, and Pro accumulation with increased Pro synthesizing enzyme (Δ1-pyrroline-5-carboxylate synthetase (P5CS) and decreased Pro degrading enzyme (proline dehydrogenase) in Mel + Ca2+ treated plants conferred As toxicity tolerance. The obtained results postulate strong evidence that the application of Mel along with Ca2+ enhances resilience against As toxicity by upregulating the activity of plasma membrane H+-ATPase, enzymes involved in antioxidant system, and ascorbate-glutathione pathway.