Mechanism of kinetin-induced death of Vicia faba ssp. minor root cortex cells.

Cell death (CD) may be induced by endogenous or exogenous factors and contributes to all the steps of plant development. This paper presents results related to the mechanism of CD regulation induced by kinetin (Kin) in the root cortex of Vicia faba ssp. minor. To explain the process, 6-(2-hydroxy-3-methylbenzylamino)purine (PI-55), adenine (Ad), 5'-amine-5'-deoxyadenosine (Ado) and N-(2-chloro-4-piridylo)-N'-phenylurea (CPPU) were applied to (i) block cytokinin receptors (CKs) and inhibit the activities of enzymes of CK metabolism, i.e., (ii) phosphoribosyltransferase, (iii) kinases, and (iv) oxidases, respectively. Moreover, ethylene glycol-bis(ß-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA), lanthanum chloride (LaCl3), ruthenium red (RRed) and cyclosporine A (CS-A) were applied to (i) chelate extracellular calcium ions (Ca2+) as well as blocks of (ii) plasma-, (iii) endoplasmic reticulum- (ER) membrane Ca2+ ion channels and (iv) mitochondria- (MIT) Ca2+ ions release by permeability transition por (PTP), respectively. The measured physiological effectiveness of these factors was the number of living and dying cortex cells estimated with orange acridine (OA) and ethidium bromide (EB), the amounts of cytosolic Ca2+ ions with chlortetracycline (CTC) staining and the intensity of chromatin and Ca2+-CTC complex fluorescence, respectively. Moreover, the role of sorafenib, an inhibitor of RAF kinase, on the vitality of cortex cells and ethylene levels as well as the activities of RAF-like kinase and MEK2 with Syntide-2 and Mek2 as substrates were studied. The results clarified the previously presented suggestion that Kin is converted to appropriate ribotides (5'-monophosphate ribonucleotides), which cooperate with the ethylene and Ca2+ ion signalling pathways to transduce the signal of kinetin-programmed cell death (Kin-PCD). Based on the present and previously published results related to Kin-PCD, the crosstalk between ethylene and MAP kinase signalling, as well as inhibitors of CK receptors and enzymes of their metabolism, is proposed.

Synergistic Effects of Melatonin and Gamma-Aminobutyric Acid on Protection of Photosynthesis System in Response to Multiple Abiotic Stressors.

GABA (gamma-aminobutyric acid) and melatonin are endogenous compounds that enhance plant responses to abiotic stresses. The response of Vicia faba to different stressors (salinity (NaCl), poly ethylene glycol (PEG), and sulfur dioxide (SO2)) was studied after priming with sole application of GABA and melatonin or their co-application (GABA + melatonin). Both melatonin and GABA and their co-application increased leaf area, number of flowers, shoot dry and fresh weight, and total biomass. Plants treated with GABA, melatonin, and GABA + melatonin developed larger stomata with wider aperture compared to the stomata of control plants. The functionality of the photosynthetic system was improved in primed plants. To investigate the photosynthetic functionality in details, the leaf samples of primed plants were exposed to different stressors, including SO2, PEG, and NaCl. The maximum quantum yield of photosystem II (PS II) was higher in the leaf samples of primed plants, while the non-photochemical quenching (NPQ) of primed plants was decreased when leaf samples were exposed to the stressors. Correlation analysis showed the association of initial PIabs with post-stress FV/FM and NPQ. Stressors attenuated the association of initial PIabs with both FV/FM and NPQ, while priming plants with GABA, melatonin, or GABA + melatonin minimized the effect of stressors by attenuating these correlations. In conclusion, priming plants with both GABA and melatonin improved growth and photosynthetic performance of Vicia faba and mitigated the effects of abiotic stressors on the photosynthetic performance.

The cropping system matters - Contrasting responses of winter faba bean (Vicia faba L.) genotypes to drought stress.

Intercropping of legumes and cereals provides many ecological advantages and contributes to a sustainable agriculture. These agricultural systems face ongoing shifts in precipitation patterns and seasonal drought. Although the effect of drought stress on legumes has been frequently studied, knowledge about water deficits influencing legumes under different cropping systems is still limited. Therefore, we investigated the impact of water deficit and re-irrigation on two winter faba bean genotypes (S_004 and S_062) and winter wheat (var. Genius) in pure and intercropped stands under greenhouse conditions. Various physiological and biochemical parameters, such as canopy surface temperature, leaf relative water content and proline content, were collected at three time points (beginning of water deficit, end of water deficit, after re-irrigation). In addition, water use efficiency (WUE) was analyzed at the end of the experiment. The overall drought stress tolerance was determined as conceptual analysis of all measured parameters. Water deficit significantly affected WUE, surface temperature and proline content of both winter faba bean genotypes. Interestingly, intercropping with wheat resulted in an overall high drought tolerance of genotype S_004, while genotype S_062 had a high drought tolerance in pure stands. Under water deficit, pure stands of S_062 substantially increased WUE by 30.5%. Intercropping of genotype S_004 increased the dry matter per plant by 31.7% compared to pure stands under water deficit. Contrary, intercropping of genotype S_062 did not improve the dry matter production. Our findings indicate that genotype S_004 benefits from resource complementarity in intercropping systems with wheat, whereas S_062 is better suitable for pure stands due to competitive effects. Thus, our study highlights that the drought tolerance of winter faba bean genotypes depends on the cropping system, leading to a demand for drought-adapted cultivars specifically selected for intercropping.

Stomatal morphology and physiology explain varied sensitivity to abscisic acid across vascular plant lineages.

Abscisic acid (ABA) can induce rapid stomatal closure in seed plants, but the action of this hormone on the stomata of fern and lycophyte species remains equivocal. Here, ABA-induced stomatal closure, signaling components, guard cell K+ and Ca2+ fluxes, vacuolar and actin cytoskeleton dynamics, and the permeability coefficient of guard cell protoplasts (Pf) were analyzed in species spanning the diversity of vascular land plants including 11 seed plants, 6 ferns, and 1 lycophyte. We found that all 11 seed plants exhibited ABA-induced stomatal closure, but the fern and lycophyte species did not. ABA-induced hydrogen peroxide elevation was observed in all species, but the signaling pathway downstream of nitric oxide production, including ion channel activation, was only observed in seed plants. In the angiosperm faba bean (Vicia faba), ABA application caused large vacuolar compartments to disaggregate, actin filaments to disintegrate into short fragments and Pf to increase. None of these changes was observed in the guard cells of the fern Matteuccia struthiopteris and lycophyte Selaginella moellendorffii treated with ABA, but a hypertonic osmotic solution did induce stomatal closure in fern and the lycophyte. Our results suggest that there is a major difference in the regulation of stomata between the fern and lycophyte plants and the seed plants. Importantly, these findings have uncovered the physiological and biophysical mechanisms that may have been responsible for the evolution of a stomatal response to ABA in the earliest seed plants.

Kinetics of DNA Repair in Vicia faba Meristem Regeneration Following Replication Stress.

The astonishing survival abilities of Vicia faba, one the earliest domesticated plants, are associated, among other things, to the highly effective replication stress response system which ensures smooth cell division and proper preservation of genomic information. The most crucial pathway here seems to be the ataxia telangiectasia-mutated kinase (ATM)/ataxia telangiectasia and Rad3-related kinase (ATR)-dependent replication stress response mechanism, also present in humans. In this article, we attempted to take an in-depth look at the dynamics of regeneration from the effects of replication inhibition and cell cycle checkpoint overriding causing premature chromosome condensation (PCC) in terms of DNA damage repair and changes in replication dynamics. We were able to distinguish a unique behavior of replication factors at the very start of the regeneration process in the PCC-induced cells. We extended the experiment and decided to profile the changes in replication on the level of a single replication cluster of heterochromatin (both alone and with regard to its position in the nucleus), including the mathematical profiling of the size, activity and shape. The results obtained during these experiments led us to the conclusion that even "chaotic" events are dealt with in a proper degree of order.

Acclimatisation of guard cell metabolism to long-term salinity.

Stomatal movements are enabled by changes in guard cell turgor facilitated via transient accumulation of inorganic and organic ions imported from the apoplast or biosynthesized within guard cells. Under salinity, excess salt ions accumulate within plant tissues resulting in osmotic and ionic stress. To elucidate whether (a) Na+ and Cl- concentrations increase in guard cells in response to long-term NaCl exposure and how (b) guard cell metabolism acclimates to the anticipated stress, we profiled the ions and primary metabolites of leaves, the apoplast and isolated guard cells at darkness and during light, that is, closed and fully opened stomata. In contrast to leaves, the primary metabolism of guard cell preparations remained predominantly unaffected by increased salt ion concentrations. Orchestrated reductions of stomatal aperture and guard cell osmolyte synthesis were found, but unlike in leaves, no increases of stress responsive metabolites or compatible solutes occurred. Diverging regulation of guard cell metabolism might be a prerequisite to facilitate the constant adjustment of turgor that affects aperture. Moreover, the photoperiod-dependent sucrose accumulation in the apoplast and guard cells changed to a permanently replete condition under NaCl, indicating that stress-related photosynthate accumulation in leaves contributes to the permanent closing response of stomata under stress.

Potential risks of nicotine on the germination, growth, and nutritional properties of broad bean.

This study evaluated the allelopathy, uptake and accumulation, and potential agricultural and food safety risks of nicotine in broad bean (Vicia faba L.) during seed germination and seedling growth. Nicotine stress has an allelopathic inhibitory effect on seeds and a hormesis effect on germinated seeds and seedlings, which has an enhancement effect (<50 mg kg-1) and an inhibition effect (>100 mg kg-1) on the germinated seeds and an enhancement effect (<100 mg kg-1) and an inhibition effect (>200 mg kg-1) on the seedlings. Exogenous nicotine can be absorbed by broad bean roots from nicotine-contaminated soil and accumulated in the main organs of the seedlings, especially the leaves, which exceeded the maximum residue level (0.03 mg kg-1 DW) at 50 mg kg-1. Moreover, nicotine resulted in a bitter taste in the edible broad bean leaves, disrupting the balance of basic nutritional properties, decreasing sucrose, and increasing bitter substances such as choline and procyanidin. These results demonstrated that residual nicotine in the soil not only poses potential risks to sustainable agricultural development but also a food safety risk for consumers. The present study provides insight into the potential risks of nicotine in agroecosystems.

Investigating the underlying mechanism of cadmium-induced plant adaptive response to genotoxic stress.

Plants as sessile organisms have developed some unique strategies to withstand environmental stress and adaptive response (AR) is one of them. In the present study Cadmium (Cd)-induced AR was evaluated to ameliorate the genotoxicity of a known chemical mutagen ethyl methanesulphonate (EMS) based on cytotoxicity, genotoxicity and oxidative stress in two model plant systems Allium cepa L. and Vicia faba L. Priming the plants with cadmium chloride (CdCl2, 25 and 50 µM) reduced the genotoxicity of EMS (0.25 mM). Cd-induced AR was evident by the magnitude of adaptive response (MAR) values calculated for cytotoxicity, genotoxicity and biochemical parameters. In addition the involvement of some major metabolic pathways and epigenetic modifications in AR was investigated. Metabolic blockers of protein kinase cascades, DNA repair, oxidative stress and de novo translation interfered with the adaptive response implying their role in AR whereas, inhibitors involved in post-replication repair and autophagy were ineffective implicating that they probably have no role in the AR studied. Moreover to find the role of DNA methylation in AR, methylation-sensitive comet assay was carried out. Simultaneously 5-methyl- 2'-deoxycytidine (5mdC) levels were quantified by HPLC (high performance liquid chromatography). AR was eliminated in cells treated with a demethylating agent, 5-aza- 2'deoxycytidine (AZA). Results implied a contribution of DNA hypermethylation. To the best of our knowledge this is a first report correlating DNA methylation to Cd-induced adaptive response in plants undergoing genotoxic stress.

First approach to pod dehiscence in faba bean: genetic and histological analyses.

Pod dehiscence causes important yield losses in cultivated crops and therefore has been a key trait strongly selected against in crop domestication. In spite of the growing knowledge on the genetic basis of dehiscence in different crops, no information is available so far for faba bean. Here we conduct the first comprehensive study for faba bean pod dehiscence by combining, linkage mapping, comparative genomics, QTL analysis and histological examination of mature pods. Mapping of dehiscence-related genes revealed conservation of syntenic blocks among different legumes. Three QTLs were identified in faba bean chromosomes II, IV and VI, although none of them was stable across years. Histological analysis supports the convergent phenotypic evolution previously reported in cereals and related legume species but revealed a more complex pattern in faba bean. Contrary to common bean and soybean, the faba bean dehiscence zone appears to show functional equivalence to that described in crucifers. The lignified wall fiber layer, which is absent in the paucijuga primitive line Vf27, or less lignified and vacuolated in other dehiscent lines, appears to act as the major force triggering pod dehiscence in this species. While our findings, provide new insight into the mechanisms underlying faba bean dehiscence, full understanding of the molecular bases will require further studies combining precise phenotyping with genomic analysis.

Earthworm Eisenia andrei modulates oxidative stress in bean plants Vicia faba irrigated with treated wastewater.

With respect to reducing the pressure on freshwater resources, treated wastewater (TWW) irrigation represents a sustainable alternative in agriculture. Due to their low quality and variable composition, TWW could entail harmful consequences for living organisms in terrestrial ecosystems. This study aims to evaluate how earthworm (Eisenia andrei) can modulate oxidative stress in bean plants (Vicia faba) that are irrigated over a course of 60 days with two doses of TWW (50 and 100%) in addition to a control condition (0%) irrigated with distilled water. This is achieved by measuring glutathione-S-transferase (GST) activity and malondialdehyde accumulation (MDA) in plants. Furthermore, catalase (CAT), GST, MDA, and acetylcholinesterase (AChE) activities of the earthworms are also assessed. Our results show that growth and physiological parameters are modified when applying TWW irrigation. Moreover, oxidative stress apprehended by GST activity and MDA accumulation is exacerbated in V. faba plants after exposure to increased TWW doses. Similarly, TWW irrigation enhances oxidative stress parameters in earthworms with a crucial decrease in AChE activity. In addition, the presence of earthworms increases growth and physiological parameters; it also results in a significant reduction in GST activity and MDA rate in V. faba plants. Our results provide new insights into the impact of TWW irrigation on soil organisms and the importance of earthworms in the reduction of oxidative stress in plants.

Transcriptome profile analysis of two Vicia faba cultivars with contrasting salinity tolerance during seed germination.

Faba bean (Vicia faba L.) is an important food legume crop. Salinity soils severely constrain the production of faba bean, however, the seed germination of faba bean, which is a vital plant growth stage, is sensitive to salinity. Planting improved varieties of faba bean, which exhibit salt tolerance in seed germination stage, is an optimal strategy for faba bean product. To investigate the genes dynamics during the seed germination stage under salinity, RNA-seq method was used to investigate genome-wide transcription profiles of two faba bean varieties with contrast salt-tolerance during the seed germination. A total of 4,486 differentially expressed genes (DEGs) were identified among the comparison of salt-tolerant variety Y134 and salt-sensitive variety Y078 treated with salinity or not. Of these, 1,410 candidate DEGs were identified as salt-stress response genes. Furthermore, 623 DEGs were identified as variety-specific response gene during seed germination at 16 h or 24 h with salt treatment. Based on the pathway enrichment according to the Kyoto Encyclopedia of Genes and Genomes database (KEGG), these DEGs involving in cell wall loosening (e.g., xyloglucan endotransglucosylase/hydrolase, chitinase, and expansin), hormone metabolism (e.g., LEA genes, genes associated with ABA or ethylene signal pathway), chromatin remodeling (e.g., chromatin structure proteins, LHP1), small interfering RNA pathway, etc., were significantly up-regulated in salt-tolerance variety with salt treatment, indicating that they play critical roles in regulation of seed germination. The results indicated that a clearer mechanism of gene regulation that regulates the seed germination responding to salinity in faba bean. These findings are helpful to increase the understanding of the salt tolerance mechanism of crops during seed germination, and provide valuable genetic resource for the breeding of salt-tolerant faba bean varieties in future.

The transgenerational effects of solar short-UV radiation differed in two accessions of Vicia faba L. from contrasting UV environments.

BACKGROUND AND AIMS: UVB radiation can rapidly induce gene regulation leading to cumulative changes for plant physiology and morphology. We hypothesized that a transgenerational effect of chronic exposure to solar short UV modulates the offspring's responses to UVB and blue light, and that the transgenerational effect is genotype dependent. METHODS: We established a factorial experiment combining two Vicia faba L. accessions, two parental UV treatments (full sunlight and exclusion of short UV, 290-350 nm), and four offspring light treatments from the factorial combination of UVB and blue light. The accessions were Aurora from southern Sweden, and ILB938 from Andean region of Colombia and Ecuador. KEY RESULTS: The transgenerational effect influenced morphological responses to blue light differently in the two accessions. In Aurora, when UVB was absent, blue light increased shoot dry mass only in plants whose parents were protected from short UV. In ILB938, blue light increased leaf area and shoot dry mass more in plants whose parents were exposed to short UV than those that were not. Moreover, when the offspring was exposed to UVB, the transgenerational effect decreased in ILB938 and disappeared in Aurora. For flavonoids, the transgenerational effect was detected only in Aurora: parental exposure to short UV was associated with a greater induction of total quercetin in response to UVB. Transcript abundance was higher in Aurora than in ILB938 for both CHALCONE SYNTHASE (99-fold) and DON-GLUCOSYLTRANSFERASE 1 (19-fold). CONCLUSIONS: The results supported both hypotheses. Solar short UV had transgenerational effects on progeny responses to blue and UVB radiation, and they differed between the accessions. These transgenerational effects could be adaptive by acclimation of slow and cumulative morphological change, and by early build-up of UV protection through flavonoid accumulation on UVB exposure. The differences between the two accessions aligned with their adaptation to contrasting UV environments.

Using advanced vibrational molecular spectroscopy (ATR-Ft/IRS) to study heating process induced changes on protein molecular structure of biodegradation residues in cool-climate adapted faba bean seeds: Relationship with rumen and intestinal protein digestion in ruminant systems.

The objective of this study was to evaluate the effects of heating process on protein molecular structure from ruminal degradation residues in cool-climate adapted faba bean seeds in relation to crude protein (CP), in situ degradation kinetics, rumen protein degradation and intestinal protein digestion parameters in dairy cows. Seeds of six faba bean varieties with low (Snowbird, Snowdrop, 219_16) and normal tannin (Fatima, 346_10, SSNS_1) were collected from three different locations, and were heated 3 min by microwave irradiation (MI, dry heating) or heated 1 h by steam pressure toasting (SP, moist heating) or kept raw as a control. Heat treated samples were used for rumen incubating 24, 12, 8, 4, 2, 0 h(s) in two replicate runs and then residues from 12 h of rumen degradation were used for three steps in vitro technique for determining intestinal protein digestion. Attenuated total reflectance Fourier transforms infrared spectroscopy (ATR-Ft/IRS) was used for analyzing protien molecular structure of residual faba bean seeds. The results showed that SP increased the intensities of amide I, amide II, α-helix and ß-sheet but decreased amide I to amide II height and area ratio, α-helix to ß-sheet height ratio from 12 and 24 h of ruminal degradation, and MI decreased all the intensities of amide I, amide II, α-helix and ß-sheet and ratios except amide I to amide II area ratio of residues from 24 h of ruminal degradation. Additionally, the intensities of amide I, amide II, α-helix and ß-sheet had a unique pattern of increasing first and then decreasing with the increasing ruminal digestion time for SP treatment, while amide I to amide II height and area ratio, α-helix to ß-sheet height ratio were declining. For the MI groups, this pattern was not observed and the intensities were rather consistent across the digestion process. Rumen protein degradation parameters including rumen bypass crude protein (BCP) or rumen undegradable protein (RUP) and rumen degradable protein (RDP) closely correlated with protein molecular structure of to peak heights, areas and ratios. Regression equations based on residual protein molecular structure presented a good estimation power for soluble fraction (S, R2 = 0.79), degradable fraction (D, R2 = 0.805), BCP (R2 = 0.941), RUP (R2 = 0.941) and RDP (R2 = 0.811). Overall, heat-induced changes in rumen residual protein molecular structures were related to CP, in situ degradation kinetics, rumen protein degradation and rumen protein digestion parameters.

Yield benefits of additional pollination to faba bean vary with cultivar, scale, yield parameter and experimental method.

The benefits of insect pollination to crop yield are used to justify management decisions across agricultural landscapes but current methods for assessing these benefits may underestimate the importance of context. We quantify how the effects of simulated insect pollination vary between five faba bean cultivars, and to what extent this changes between years, scales, yield parameters, and experimental methods. We do this by measuring responses to standardised hand pollination treatments in controlled experiments in flight cages and in the field. Pollination treatments generally improved yield, but in some cases yield was lower with additional pollination. Pollination dependence varied with cultivar, ranging from 58% (loss in yield mass per plant without pollination) in one cultivar, to a lower yield with pollination in another (-51%). Pollination dependence also varied between flight cage and field experiments (-10 to 37% in the same cultivar and year), year (4 to 33%; same cultivar and yield parameter), and yield parameter (-4 to 46%; same cultivar and year). This variability highlights that to be robust, assessments of pollination benefits need to focus upon marketable crop outputs at a whole-plant or larger scale while including and accounting for the effects of different years, sites, methodologies and cultivars.

Is nitric oxide a critical key factor in ABA-induced stomatal closure?

The role of nitric oxide (NO) in abscisic acid (ABA)-induced stomatal closure is a matter of debate. We conducted experiments in Vicia faba leaves using NO gas and sodium nitroprusside (SNP), a NO-donor compound, and compared their effects to those of ABA. In epidermal strips, stomatal closure was induced by ABA but not by NO, casting doubt on the role of NO in ABA-mediated stomatal closure. Leaf discs and intact leaves showed a dual dose response to NO: stomatal aperture widened at low dosage and narrowed at high dosage. Overcoming stomatal resistance by means of high CO2 concentration ([CO2]) restored photosynthesis in ABA-treated leaf discs but not in those exposed to NO. NO inhibited photosynthesis immediately, causing an instantaneous increase in intercellular [CO2] (Ci), followed by stomatal closure. However, lowering Ci by using low ambient [CO2] showed that it was not the main factor in NO-induced stomatal closure. In intact leaves, the rate of stomatal closure in response to NO was about one order of magnitude less than after ABA application. Because of the different kinetics of photosynthesis and stomatal closure that were observed, we conclude that NO is not likely to be the key factor in ABA-induced rapid stomatal closure, but that it fine-tunes stomatal aperture via different pathways.

Enzymes contributing to the hydrogen peroxide signal dynamics that regulate wall labyrinth formation in transfer cells.

Transfer cells are characterized by an amplified plasma membrane area supported on a wall labyrinth composed of a uniform wall layer (UWL) from which wall ingrowth (WI) papillae arise. Adaxial epidermal cells of developing Vicia faba cotyledons, when placed in culture, undergo a rapid (hours) trans-differentiation to a functional epidermal transfer cell (ETC) phenotype. The trans-differentiation event is controlled by a signalling cascade comprising auxin, ethylene, apoplasmic reactive oxygen species (apoROS), and cytosolic Ca2+. Apoplasmic hydrogen peroxide (apoH2O2) was confirmed as the apoROS regulating UWL and WI papillae formation. Informed by an ETC-specific transcriptome, a pharmacological approach identified a temporally changing cohort of H2O2 biosynthetic enzymes. The cohort contained a respiratory burst oxidase homologue, polyamine oxidase, copper amine oxidase, and a suite of class III peroxidases. Collectively these generated two consecutive bursts in apoH2O2 production. Spatial organization of biosynthetic/catabolic enzymes was deduced from responses to pharmacologically blocking their activities on the cellular and subcellular distribution of apoH2O2. The findings were consistent with catalase activity constraining the apoH2O2 signal to the outer periclinal wall of the ETCs. Strategic positioning of class III peroxidases in this outer domain shaped subcellular apoH2O2 signatures that differed during assembly of the UWL and WI papillae.

Metabolomics of red-light-induced stomatal opening in Arabidopsis thaliana: Coupling with abscisic acid and jasmonic acid metabolism.

Environmental stimuli-triggered stomatal movement is a key physiological process that regulates CO2 uptake and water loss in plants. Stomata are defined by pairs of guard cells that perceive and transduce external signals, leading to cellular volume changes and consequent stomatal aperture change. Within the visible light spectrum, red light induces stomatal opening in intact leaves. However, there has been debate regarding the extent to which red-light-induced stomatal opening arises from direct guard cell sensing of red light versus indirect responses as a result of red light influences on mesophyll photosynthesis. Here we identify conditions that result in red-light-stimulated stomatal opening in isolated epidermal peels and enlargement of protoplasts, firmly establishing a direct guard cell response to red light. We then employ metabolomics workflows utilizing gas chromatography mass spectrometry and liquid chromatography mass spectrometry for metabolite profiling and identification of Arabidopsis guard cell metabolic signatures in response to red light in the absence of the mesophyll. We quantified 223 metabolites in Arabidopsis guard cells, with 104 found to be red light responsive. These red-light-modulated metabolites participate in the tricarboxylic acid cycle, carbon balance, phytohormone biosynthesis and redox homeostasis. We next analyzed selected Arabidopsis mutants, and discovered that stomatal opening response to red light is correlated with a decrease in guard cell abscisic acid content and an increase in jasmonic acid content. The red-light-modulated guard cell metabolome reported here provides fundamental information concerning autonomous red light signaling pathways in guard cells.

Phytotoxicity of silver nanoparticles on Vicia faba: Evaluation of particle size effects on photosynthetic performance and leaf gas exchange.

Nanotechnology is an emerging field in science and engineering, which presents significant impacts on the economy, society and the environment. The nanomaterials' (NMs) production, use, and disposal is inevitably leading to their release into the environment where there are uncertainties about its fate, behaviour, and toxicity. Recent works have demonstrated that NMs can penetrate, translocate, and accumulate in plants. However, studies about the effects of the NMs on plants are still limited because most investigations are carried out in the initial stage of plant development. The present study aimed to evaluate and characterize the photochemical efficiency of photosystem II (PSII) of broad bean (Vicia faba) leaves when subjected to silver nanoparticles (AgNPs) with diameters of 20, 51, and 73 nm as well as to micrometer-size Ag particles (AgBulk). The AgNPs were characterized by transmission electron microscopy and dynamic light scattering. The analyses were performed by injecting the leaves with 100 mg L-1 aqueous solution of Ag and measuring the chlorophyll fluorescence imaging, gas exchange, thermal imaging, and reactive oxygen species (ROS) production. In addition, silver ion (Ag+) release from Ag particles was determined by dialysis. The results revealed that AgNPs induce a decrease in the photochemical efficiency of photosystem II (PSII) and an increase in the non-photochemical quenching. The data also revealed that AgNPs affected the stomatal conductance (gs) and CO2 assimilation. Further, AgNPs induced an overproduction of ROS in Vicia faba leaves. Finally, all observed effects were particle diameter-dependent, increasing with the reduction of AgNPs diameter and revealing that AgBulk caused only a small or no changes on plants. In summary, the results point out that AgNPs may negatively affect the photosynthesis process when accumulated in the leaves, and that the NPs themselves were mainly responsible since negligible Ag+ release was detected.

Sodium nitroprusside (SNP) improves tolerance to arsenic (As) toxicity in Vicia faba through the modifications of biochemical attributes, antioxidants, ascorbate-glutathione cycle and glyoxalase cycle.

The present study was conducted to evaluate the effect of arsenic (As) toxicity and the mitigating role of nitric oxide (NO) donor sodium nitroprusside (SNP) on Vicia faba. Arsenics stress decreased the growth and biomass yield, and photosynthetic pigments, but it enhanced As accumulation. Supplementation of NO enhanced the afore-mentioned parameters except As accumulation which decreased in both shoot and root. Supplementation of NO enhanced the shoot tolerance index (Shoot TI%), root tolerance index (Root TI%) but it declined the As translocation factor (TF). Application of NO alleviated the As-induced decline in net assimilation rate, stomatal conductance, transpiration and leaf relative water content. The levels of proline and glycine betaine (GB) further increased due to NO application, whereas malondialdehyde (MDA), hydrogen peroxide (H2O2), electrolyte leakage (EL) and methylglyoxal (MG) declined considerably. Activities of enzymatic antioxidants such as superoxide dismutase (SOD) and catalase (CAT) increased under As stress. Supplementation of NO up-regulated the enzymes involved in Asc-Glu cycle and glyoxalase cycle under As toxicity. Another experiment was setup to authenticate whether NO was certainly able to alleviate As toxicity. For this purpose, the NO scavenger [2-(4-carboxy-2 phenyl)-4,4,5,5-tertamethylimidazoline-1-oxyl-3-oxide (cPTIO)] was added to As and NO supplemented plants. Addition of cPTIO to NO supplemented As-treated plants showed the same effect when As alone was supplied to plants. In conclusion, addition of NO to the growth medium maintained the plant performance under As toxicity through modulation of physio-biochemical attributes, antioxidant enzymes, and the Asc-Glu and glyoxalase systems.

Genotoxicity response of Vicia faba seedlings to cadmium in soils as characterized by direct soil exposure and micronucleus test.

To overcome the drawbacks of the Vicia faba root tip micronucleus test in soil using the solution extract method, we conducted a potting experiment by direct soil exposure. Cadmium was spiked into 3 typical soils (brown soil, red soil, and black soil) to simulate environmental concentrations (0.625, 1.25, 2.5, 5, and 10 mg kg-1). Multiple Vicia faba tissues (primary root tips, secondary root tips, and leaf tips) were sampled, and mitotic index (MI), chromosome aberration frequency (CA), and micronucleus frequency (MN) were used as endpoints after a seedling period of 5 days. The results showed a response between Cd concentrations and multiple sampling tissues of Vicia faba, and the secondary root tips responded to Cd stress the most, followed by primary root tips and leaf tips. Soil physicochemical properties (e.g., pH, total phosphorus, total organic carbon, etc.) influenced the genotoxicity of Cd, and pH was the dominant factor, which resulted in the genetic toxicity response of Cd in soils in the order: red soil > brown soil > black soil. The lowest observable effect concentration (LOEC) of Cd was 1.25 mg kg-1 for both brown soil and red soil and 2.5 mg kg-1 for black soil. In view of this, we suggested that soil properties should be considered in evaluating genotoxicity risk of Cd in soil, especially with soil pH range, and the secondary root tips should be taken as suitable test tissues in the MN test due to its more sensible response feature to Cd stress in soil.