Nitric oxide production is involved in maintaining energy state in Alfalfa (Medicago sativa L.) nodulated roots under both salinity and flooding.

MAIN CONCLUSION: In Medicago sativa nodulated roots, NR-dependent NO production is involved in maintaining energy state, presumably through phytoglobin NO respiration, under both salinity and hypoxia stress. The response to low and average salinity stress and to a 5 day-long flooding period was analyzed in M. sativa nodulated roots. The two treatments result in a decrease in the biological nitrogen fixation capacity and the energy state (evaluated by the ATP/ADP ratio), and conversely in an increase nitric oxide (NO) production. Under salinity and hypoxia treatments, the use of either sodium tungstate, an inhibitor of nitrate reductase (NR), or carboxy-PTIO, a NO scavenger, results in a decrease in NO production and ATP/ADP ratio, meaning that NR-dependent NO production participates to the maintenance of the nodulated roots energy state.

The role of nitrate reductase in brassinosteroid-induced endogenous nitric oxide generation to improve cadmium stress tolerance of pepper plants by upregulating the ascorbate-glutathione cycle.

A study was performed to assess if nitrate reductase (NR) participated in brassinosteroid (BR)-induced cadmium (Cd) stress tolerance primarily by accelerating the ascorbate-glutathione (AsA-GSH) cycle. Prior to initiating Cd stress (CdS), the pepper plants were sprayed with 0.5 µM 24-epibrassinolide (EBR) every other day for 10 days. Thereafter the seedlings were subjected to control or CdS (0.1 mM CdCl2) for four weeks. Cadmium stress decreased the plant growth related attributes, water relations as well as the activities of monodehydroascorbate reductase (MDHAR) and dehydroascorbate reductase (DHAR), but enhanced proline content, leaf Cd2+ content, oxidative stress-related traits, activities of ascorbate peroxidase (APX) and glutathione reductase (GR), and the activities of antioxidant defence system-related enzymes as well as NR activity and endogenous nitric oxide content. EBR reduced leaf Cd2+ content and oxidative stress-related parameters, enhanced plant growth, regulated water relations, and led to further increases in proline content, AsA-GSH cycle-related enzymes' activities, antioxidant defence system-related enzymes as well as NR activity and endogenous nitric oxide content. The EBR and the inhibitor of NR (tungstate) reversed the positive effects of EBR by reducing NO content, showing that NR could be a potential contributor of EBR-induced generation of NO which plays an effective role in tolerance to CdS in pepper plants by accelerating the AsA-GSH cycle and antioxidant enzymes.

Protective Role of Nitrate/Nitrite Reductase System during Transient Global Cerebral Ischemia.

The effects of a non-selective blocker of NO synthases LNNA in a dose of 25 mg/kg and nitrates KNO3, NaNO3, Mg(NO3)2, and Ca(NO3)2 in doses of 5 and 50 mg/kg were studied on the model of experimental ischemic stroke induced by bilateral occlusion of the common carotid arteries. Wistar rats were randomized into 40 treatment (n=960) and 8 control (n=192) groups. Treatment group rats received injection of either nitrate, or LNNA, or a combination LNNA+nitrate. All substances were administered intraperitoneally 1 h prior to brain ischemia or 5 sec after occlusion of the common carotid arteries. Control rats received the same volume of 0.9% NaCl at the same terms. The non-selective inhibitor of NO synthases LNNA administered against the background of brain ischemia increased neurological deficit and animal mortality. Nitrate/nitrite reductase system had protective properties depending on the cation of nitrate and concentration of a nitrate during inhibition of NO synthase system under conditions of brain ischemia and hypoxia.

An Unsaturated Quinolone N-Oxide of Pseudomonas aeruginosa Modulates Growth and Virulence of Staphylococcus aureus.

The pathogen Pseudomonas aeruginosa produces over 50 different quinolones, 16 of which belong to the class of 2-alkyl-4-quinolone N-oxides (AQNOs) with various chain lengths and degrees of saturation. We present the first synthesis of a previously proposed unsaturated compound that is confirmed to be present in culture extracts of P. aeruginosa, and its structure is shown to be trans-Δ1 -2-(non-1-enyl)-4-quinolone N-oxide. This compound is the most active agent against S.  aureus, including MRSA strains, by more than one order of magnitude whereas its cis isomer is inactive. At lower concentrations, the compound induces small-colony variants of S. aureus, reduces the virulence by inhibiting hemolysis, and inhibits nitrate reductase activity under anaerobic conditions. These studies suggest that this unsaturated AQNO is one of the major agents that are used by P. aeruginosa to modulate competing bacterial species.

Nitric oxide is required for hydrogen gas-induced adventitious root formation in cucumber.

Hydrogen gas (H2) is involved in plant development and stress responses. Cucumber explants were used to study whether nitric oxide (NO) is involved in H2-induced adventitious root development. The results revealed that 50% and 100% hydrogen-rich water (HRW) apparently promoted the development of adventitious root in cucumber. While, the responses of HRW-induced adventitious rooting were blocked by a specific NO scavenger 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide potassium salt (cPTIO), NO synthase (NOS) enzyme inhibitor N(G)-nitro-l-arginine methylester hydrochloride (l-NAME) and nitrate reductase (NR) inhibitor NaN3. HRW also increased NO content and NOS and NR activity both in a dose- and time-dependent fashion. Moreover, molecular evidence showed that HRW up-regulated NR genes expression in explants. The results indicate the importance of NOS and NR enzymes, which might be responsible for NO production in explants during H2-induced root organogenesis. Additionally, peroxidase (POD) and indoleacetic acid oxidase (IAAO) activity was significantly decreased in the explants treated with HRW, while HRW treatment significantly increased polyphenol oxidase (PPO) activity. In addition, cPTIO, l-NAME and NaN3 inhibited the actions of HRW on the activity of these enzymes. Together, NO may be involved in H2-induced adventitious rooting, and NO may be acting downstream in plant H2 signaling cascade.

Remodeling of intermediate metabolism in the diatom Phaeodactylum tricornutum under nitrogen stress.

Diatoms are unicellular algae that accumulate significant amounts of triacylglycerols as storage lipids when their growth is limited by nutrients. Using biochemical, physiological, bioinformatics, and reverse genetic approaches, we analyzed how the flux of carbon into lipids is influenced by nitrogen stress in a model diatom, Phaeodactylum tricornutum. Our results reveal that the accumulation of lipids is a consequence of remodeling of intermediate metabolism, especially reactions in the tricarboxylic acid and the urea cycles. Specifically, approximately one-half of the cellular proteins are cannibalized; whereas the nitrogen is scavenged by the urea and glutamine synthetase/glutamine 2-oxoglutarate aminotransferase pathways and redirected to the de novo synthesis of nitrogen assimilation machinery, simultaneously, the photobiological flux of carbon and reductants is used to synthesize lipids. To further examine how nitrogen stress triggers the remodeling process, we knocked down the gene encoding for nitrate reductase, a key enzyme required for the assimilation of nitrate. The strain exhibits 40-50% of the mRNA copy numbers, protein content, and enzymatic activity of the wild type, concomitant with a 43% increase in cellular lipid content. We suggest a negative feedback sensor that couples photosynthetic carbon fixation to lipid biosynthesis and is regulated by the nitrogen assimilation pathway. This metabolic feedback enables diatoms to rapidly respond to fluctuations in environmental nitrogen availability.

SIGNAL MEDIATORS AT INDUCTION OF HEAT RESISTANCE OF WHEAT PLANTLETS BY SHORT-TERM HEATING.

The effects of functional interplay of calcium ions, reactive oxygen species (ROS) and nitric oxide (NO) in the cells of wheat plantlets roots (Triticum aestivum L.) at the induction of their heat resistance by a short-term influence of hyperthermia (heating at the temperature of 42 degrees C during 1 minute) have been investigated. The transitional increase of NO and H2O2 content, invoked by heating, was suppressed by the treatment of plantlets with the antagonists of calcium EGTA (chelator of exocellular calcium), lanthanum chloride (blocker of calcium channels of various types) and neomycin (inhibitor of phosphatidylinositol-dependent phospholipase C). The rise of hydrogen peroxide content, caused by hardening, was partially suppressed by the action of inhibitors of nitrate reductase (sodium wolframate) and NO-synthase (N(G)-nitro-L-arginine methyl ester--L-NAME), and the increasing of nitric oxide content was suppressed by the treatment of plants with the antioxidant ionol and with the scavenger of hydrogen peroxide (dimethylthiourea). These compounds and antagonists of calcium also partially removed the effect of the rise of plantlets' heat resistance, invoked by hardening heating. The conclusion on calcium's role in the activation of enzymatic systems, generating reactive oxygen species and nitric oxide, and on the functional interplay of these signal mediators at the induction of heat resistance of plantlets by hardening heating is made.

The endogenous nitric oxide mediates selenium-induced phytotoxicity by promoting ROS generation in Brassica rapa.

Selenium (Se) is suggested as an emerging pollutant in agricultural environment because of the increasing anthropogenic release of Se, which in turn results in phytotoxicity. The most common consequence of Se-induced toxicity in plants is oxidative injury, but how Se induces reactive oxygen species (ROS) burst remains unclear. In this work, histofluorescent staining was applied to monitor the dynamics of ROS and nitric oxide (NO) in the root of Brassica rapa under Se(IV) stress. Se(IV)-induced faster accumulation of NO than ROS. Both NO and ROS accumulation were positively correlated with Se(IV)-induced inhibition of root growth. The NO accumulation was nitrate reductase (NR)- and nitric oxide synthase (NOS)-dependent while ROS accumulation was NADPH oxidase-dependent. The removal of NO by NR inhibitor, NOS inhibitor, and NO scavenger could alleviate Se(IV)-induced expression of Br_Rbohs coding for NADPH oxidase and the following ROS accumulation in roots, which further resulted in the amelioration of Se(IV)-induced oxidative injury and growth inhibition. Thus, we proposed that the endogenous NO played a toxic role in B. rapa under Se(IV) stress by triggering ROS burst. Such findings can be used to evaluate the toxic effects of Se contamination on crop plants.

Kinetics of substrate inhibition of periplasmic nitrate reductase.

Periplasmic nitrate reductase catalyzes the reduction of nitrate into nitrite using a mononuclear molybdenum cofactor that has nearly the same structure in all enzymes of the DMSO reductase family. In previous electrochemical investigations, we found that the enzyme exists in several inactive states, some of which may have been previously isolated and mistaken for catalytic intermediates. In particular, the enzyme slowly and reversibly inactivates when exposed to high concentrations of nitrate. Here, we study the kinetics of substrate inhibition and its dependence on electrode potential and substrate concentration to learn about the properties of the active and inactive forms of the enzyme. We conclude that the substrate-inhibited enzyme never significantly accumulates in the EPR-active Mo(+V) state. This conclusion is relevant to spectroscopic investigations where attempts are made to trap a Mo(+V) catalytic intermediate using high concentrations of nitrate.

The nitrate reductase inhibitor, tungsten, disrupts actin microfilaments in Zea mays L.

Tungsten is a widely used inhibitor of nitrate reductase, applied to diminish the nitric oxide levels in plants. It was recently shown that tungsten also has heavy metal attributes. Since information about the toxic effects of tungsten on actin is limited, and considering that actin microfilaments are involved in the entry of tungsten inside plant cells, the effects of tungsten on them were studied in Zea mays seedlings. Treatments with sodium tungstate for 3, 6, 12 or 24 h were performed on intact seedlings and seedlings with truncated roots. Afterwards, actin microfilaments in meristematic root and leaf tissues were stained with fluorescent phalloidin, and the specimens were examined by confocal laser scanning microscopy. While the actin microfilament network was well organized in untreated seedlings, in tungstate-treated ones it was disrupted in a time-dependent manner. In protodermal root cells, the effects of tungsten were stronger as cortical microfilaments were almost completely depolymerized and the intracellular ones appeared highly bundled. Fluorescence intensity measurements confirmed the above results. In the meristematic leaf tissue of intact seedlings, no depolymerization of actin microfilaments was noticed. However, when root tips were severed prior to tungstate application, both cortical and endoplasmic actin networks of leaf cells were disrupted and bundled after 24 h of treatment. The differential response of root and leaf tissues to tungsten toxicity may be due to differential penetration and absorption, while the effects on actin microfilaments could not be attributed to the nitric oxide depletion by tungsten.

Fosfomycin and tobramycin in combination downregulate nitrate reductase genes narG and narH, resulting in increased activity against Pseudomonas aeruginosa under anaerobic conditions.

The activity of aminoglycosides, which are used to treat Pseudomonas aeruginosa respiratory infection in cystic fibrosis (CF) patients, is reduced under the anaerobic conditions that reflect the CF lung in vivo. In contrast, a 4:1 (wt/wt) combination of fosfomycin and tobramycin (F:T), which is under investigation for use in the treatment of CF lung infection, has increased activity against P. aeruginosa under anaerobic conditions. The aim of this study was to elucidate the mechanisms underlying the increased activity of F:T under anaerobic conditions. Microarray analysis was used to identify the transcriptional basis of increased F:T activity under anaerobic conditions, and key findings were confirmed by microbiological tests, including nitrate utilization assays, growth curves, and susceptibility testing. Notably, growth in subinhibitory concentrations of F:T, but not tobramycin or fosfomycin alone, significantly downregulated (P < 0.05) nitrate reductase genes narG and narH, which are essential for normal anaerobic growth of P. aeruginosa. Under anaerobic conditions, F:T significantly decreased (P < 0.001) nitrate utilization in P. aeruginosa strains PAO1, PA14, and PA14 lasR::Gm, a mutant known to exhibit increased nitrate utilization. A similar effect was observed with two clinical P. aeruginosa isolates. Growth curves indicate that nitrate reductase transposon mutants had reduced growth under anaerobic conditions, with these mutants also having increased susceptibility to F:T compared to the wild type under similar conditions. The results of this study suggest that downregulation of nitrate reductase genes resulting in reduced nitrate utilization is the mechanism underlying the increased activity of F:T under anaerobic conditions.

Endogenous nitric oxide generation in protoplast chloroplasts.

KEY MESSAGE : NO generation is studied in the protoplast chloroplasts. NO, ONOO ( - ) and ROS (O ( 2 ) ( - ) and H ( 2 ) O ( 2 ) ) are generated in chloroplasts. Nitric oxide synthase-like protein appears to be involved in NO generation. Nitric oxide stimulates chlorophyll biosynthesis and chloroplast differentiation. The present study was conducted to better understand the process of NO generation in the leaf chloroplasts and protoplasts. NO, peroxynitrite and superoxide anion were investigated in the protoplasts and isolated chloroplasts using specific dyes, confocal laser scanning and light microscopy. The level of NO was highest after protoplast isolation and subsequently decreased during culture. Suppression of NO signal in the presence of PTIO, suggests that diaminofluorescein-2 diacetate (DAF-2DA) detected NO. Detection of peroxynitrite, a reaction product of NO and superoxide anion, further suggests NO generation. Moreover, generation of NO and peroxynitrite in the chloroplasts of wild-type Arabidopsis and their absence or weak signals in the leaf-derived protoplasts of Atnoa1 mutants confirmed the reactivity of DAF-2DA and aminophenyl fluorescein to NO and peroxynitrite, respectively. Isolated chloroplasts also showed signal of NO. Suppression of NO signal in the presence of 100 µM nitric oxide synthase inhibitors [L-NNA, Nω-nitro-L-arginine and PBIT, S,S'-1,3-phenylene-bis(1,2-ethanediyl)-bis-isothiourea] revealed that nitric oxide synthase-like system is involved in NO synthesis. Suppression of NO signal in the protoplasts isolated in the presence of cycloheximide suggests de novo synthesis of NO generating protein during the process of protoplast isolation. Furthermore, the lack of inhibition of NO production by sodium tungstate (250 µM) and inhibition by L-NNA, and PBIT suggest involvement NOS-like protein, but not nitrate reductase, in NO generation in the leaf chloroplasts and protoplasts.

Nitric oxide enhances development of lateral roots in tomato (Solanum lycopersicum L.) under elevated carbon dioxide.

Elevated carbon dioxide (CO2) has been shown to enhance the growth and development of plants, especially of roots. Amongst them, lateral roots play an important role in nutrient uptake, and thus alleviate the nutrient limitation to plant growth under elevated CO2. This paper examined the mechanism underlying CO2 elevation-induced lateral root formation in tomato. The endogenous nitric oxide (NO) in roots was detected by the specific probe 4-amino-5-methylamino-2',7'-difluorofluorescein diacetate (DAF-FM DA). We suggest that CO2 elevation-induced NO accumulation was important for lateral root formation. Elevated CO2 significantly increased the activity of nitric oxide synthase in roots, but not nitrate reductase activity. Moreover, the pharmacological evidence showed that nitric oxide synthase rather than nitrate reductase was responsible for CO2 elevation-induced NO accumulation. Elevated CO2 enhanced the activity of nitric oxide synthase and promoted production of NO, which was involved in lateral root formation in tomato under elevated CO2.

NOS-like-mediated nitric oxide is involved in Pinus thunbergii response to the invasion of Bursaphelenchus xylophilus.

The content of NO and H(2)O(2) as well as the activities of nitric oxide synthase (NOS)-like and nitrate reductase (NR) were monitored in the needles of Pinus thunbergii infected by Bursaphelenchus xylophilus. The results showed that the content of NO increased significantly only 8 h after the invasion of B. xylophilus, while H(2)O(2) increased 12 h after invasion. NO donor SNP could promote and NO scavenger cPTIO could prevent the production of NO and H(2)O(2). The content of NO changed earlier than that of H(2)O(2). In addition, the symptoms appeared 9, 5 and 12 days, respectively, after the inoculation with B. xylophilus, SNP pre-treatment and cPTIO pre-treatment followed by B. xylophilus infection. After B. xylophilus infection, the content of NO in P. thunbergii changed fiercely more earlier than the appearance of external symptoms, which indicated that the content of NO was related with the appearance and the development of the symptoms. The treatment with L-NNA (NOS inhibitor) inhibited the content of NO significantly, whereas, Na(2)WO(4) (NR inhibitor) had no effect. The further analysis of NOS revealed that NO changed in consistent with cNOS activity. To sum up, NO, as the upstream signal molecule of H(2)O(2), was involved in the pine early response to the invasion of B. xylophilus and influenced the accumulation of the content of H(2)O(2). Moreover, NOS-like rather than NR was responsible for the endogenous NO generation, which was modulated by cNOS during the interaction between P. thunbergii and B. xylophilus. Key message NO is involved in early response of P. thunbergii to the invasion of B. xylophilus and NOS is the key enzyme responsible for NO generation in P. thunbergii.

Activities of nitrate reductase and glutamine synthetase in rice seedlings during cyanide metabolism.

A study was conducted to investigate activities of nitrate reductase (NR) and glutamine synthetase (GS) in plants during cyanide metabolism. Young rice seedlings (Oryza sativa L. cv. XZX 45) were grown in the nutrient solutions containing KNO(3) or NH(4)Cl and treated with free cyanide (KCN). Cyanide in solutions and in plant materials was analyzed to estimate the phyto-assimilation potential. Activities of NR and GS in different parts of rice seedlings were assayed in vivo. Seedlings grown on NH(4)(+) showed significantly higher relative growth rate than those on NO(3)(-) (p<0.05) in the presence of exogenous cyanide. The metabolic rates of cyanide by seedlings were all positively correlated to the concentrations supplied. A negligible difference was observed between the two treatments with nitrate and ammonium (p>0.05). Enzymatic assays showed that cyanide (≥0.97mg CN L(-1)) impaired NR activity significantly in both roots and shoots (p<0.05). The effect of cyanide on GS activity in roots was more evident at 1.93mg CN L(-1), suggesting that NR activity was more susceptible to change from cyanide application than GS activity. The results observed here suggest that the exogenous cyanide, which to a certain level has a beneficial role in plant nutrition.

Nitric oxide is involved in hemin-induced cucumber adventitious rooting process.

Hemin, a heme oxygenase-1 (HO-1) inducer, was shown to exert numerous beneficial physiological functions in animals. Our previous study suggests that HO-1/carbon monoxide (CO) acts as a novel downstream signal system in the auxin-induced adventitious rooting. The objective of this study was to test whether nitric oxide (NO) is involved in hemin-induced cucumber adventitious rooting. Applications of hemin or CO aqueous solution to auxin-depleted cucumber explant induced up-regulation of cucumber HO-1 transcripts (CsHO1), NO production, and thereafter adventitious root formation, and some above responses were blocked by the combination treatment with two nitric oxide synthase (NOS)-like enzyme inhibitors N(G)-nitro-L-arginine methylester hydrochloride and N(G)-nitro-L-arginine, a HO-1 specific inhibitor zinc protoporphyrin IX, and a specific NO scavenger 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide potassium salt. However, these blocking responses were not observed using tungstate, an inhibitor of nitrate reductase, another NO producing enzyme in plants. Furthermore, the guanylate cyclase inhibitors 1H-(1,2,4)-oxadiazole[4,3-a]quinoxalin-1-one and 6-anilino-5,8-quinolinedione reduced root development induced by hemin, whereas the cell-permeable cyclic guanosine monophosphate (cGMP) derivative 8-Br-cGMP reversed this effect. Together, our results indicated that at least in our experimental conditions, NO might operate downstream of hemin promoting adventitious root formation probably in a cGMP-dependent manner.

[Influence of environmental factors on the generation of nitric oxide in the roots of etiolated pea seedlings].

The article studies the nitric oxide (NO) levels in the roots of etiolated seedlings of garden peas (Pisum sativum L.) using the DAF-2DA fluorescent probe and fluorescent microscopy. Cross sections of roots of 100-150 microm (the site of a root which is 10-15 mm from the apex) are analyzed. It is shown that the level of NO in the roots after 24 h increased by more than a factor of 2 in the versions with NaNO2 and sodium nitroprusside. At feeding the seedlings with KNO3, a peak in the accumulation of NO in the roots (twofold increase) was observed after 30 min. Fertilizing seedlings with L-arginine (2 mM) increased the intensity of the fluorescence of the root sections by more than a factor of 2. The inoculation of seedlings of rhizobia (Rhizobium leguminosarum by. viceae) contributed to the reduction of NO on the background of the control (H20) and sodium nitroprusside and nitrogen compounds. Scavengers of NO (2-phenyl-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (PTIO), hemoglobin) and inhibitors of nitrate reductase and animal NO synthase (sodium tungstate and aminoguanidine hydrochloride) reduced the level of NO in the roots. The results are discussed in relation to the role of NO in plants under the influence of biotic and abiotic factors.

Tungstate: is it really a specific nitrate reductase inhibitor in plant nitric oxide research?

Nitrate reductase (NR) is an enzymatic source of nitric oxide (NO) in plants, and it needs Mo for the Mo-cofactor to be activated. Because NR-deficient mutants are not always available in some species, a cheap and simple pharmacological application of tungstate, which substitutes for Mo in the Mo-cofactor as a competitive antagonist, is widely used as a NR inhibitor in plant NO research. However, evidence indicates that tungstate not only inactivates NR but also inhibits other molybdate-dependent enzymes in plants. In addition, a number of investigations have shown that tungstate also inhibits root growth, affects cortical microtubule formation, and induces programmed cell death (PCD) in plants, just like other heavy metals do. Therefore, tungstate has been shown to exert many other effects that are not connected with the inhibition of NR activity. The origin and mechanism of using tungstate as a NR inhibitor in plants is reviewed here and the progress regarding tungstate toxicity to plants and the possible problems involved in using tungstate as a NR inhibitor in plant NO research are analysed. In summary, the use of tungstate as a NR inhibitor in plant NO research must be treated with caution, keeping in mind that it is not completely specific. It is necessary to search for more NR-deficient mutants and new, specific NR inhibitors. A combination of pharmacological and biochemical analysis with a genetic approach will be necessary in order to investigate the roles of NO in plants.

NO synthase-generated NO acts downstream of auxin in regulating Fe-deficiency-induced root branching that enhances Fe-deficiency tolerance in tomato plants.

In response to Fe-deficiency, various dicots increase their root branching which contributes to the enhancement of ferric-chelate reductase activity. Whether this Fe-deficiency-induced response eventually enhances the ability of the plant to tolerate Fe-deficiency or not is still unclear and evidence is also scarce about the signals triggering it. In this study, it was found that the SPAD-chlorophyll meter values of newly developed leaves of four tomato (Solanum lycocarpum) lines, namely line227/1 and Roza and their two reciprocal F(1) hybrid lines, were positively correlated with their root branching under Fe-deficient conditions. It indicates that Fe-deficiency-induced root branching is critical for plant tolerance to Fe-deficiency. In another tomato line, Micro-Tom, the increased root branching in Fe-deficient plants was accompanied by the elevation of endogenous auxin and nitric oxide (NO) levels, and was suppressed either by the auxin transport inhibitors NPA and TIBA or the NO scavenger cPTIO. On the other hand, root branching in Fe-sufficient plants was induced either by the auxin analogues NAA and 2,4-D or the NO donors NONOate or SNP. Further, in Fe-deficient plants, NONOate restored the NPA-terminated root branching, but NAA did not affect the cPTIO-terminated root branching. Fe-deficiency-induced root branching was inhibited by the NO-synthase (NOS) inhibitor L-NAME, but was not affected by the nitrate reductase (NR) inhibitor NH(4)(+), tungstate or glycine. Taking all of these findings together, a novel function and signalling pathway of Fe-deficiency-induced root branching is presented where NOS-generated rather than NR-generated NO acts downstream of auxin in regulating this Fe-deficiency-induced response, which enhances the plant tolerance to Fe-deficiency.

Zinc and copper induced changes in physiological characteristics of Vigna mungo (L.).

The effect of deleterious concentration of zinc and copper provided either individually or in combination in the nutrient media was investigated in order to assess the effect of metal interaction in Vigna mungo (L.). Both metals showed negative effect and led to a marked decrease in seed germination (20%), seedling growth (91.7%) and nitrate reductase activity (85.7%) with the increase in metal concentrations. The present study also emphasizes on the response of catalase and peroxidase enzyme under zinc and copper stress. Both antioxidant enzymes exhibited an increasing trend under different treatment conditions but it was reverse at highly toxic metal concentration. The results showed active involvement of peroxidase enzyme in regulating oxidative stress rather than catalase enzyme, as the specific activity of peroxidase enzyme got increased by 8.94% under the combined metals stress whereas catalase activity got declined by 60.97% in comparison to control due to excessive stress. The combined effect of copper and zinc metal was more pronounced in comparison to their individual effects.