The Importance of Nitric Oxide as the Molecular Basis of the Hydrogen Gas Fumigation-Induced Alleviation of Cd Stress on Ganoderma lucidum

Whether or not hydrogen gas (H2) can reduce cadmium (Cd) toxicity in Ganoderma lucidum has remained largely unknown. Here, we report that Cd-induced growth inhibition in G. lucidum was significantly alleviated by H2 fumigation or hydrogen-rich water (HRW), evaluated by lower oxidative damage and Cd accumulation. Moreover, the amelioration effects of H2 fumigation were better than of HRW in an optimum concentration of H2 under our experimental conditions. Further results showed that H2-alleviated growth inhibition in G. lucidum was accompanied by increased nitric oxide (NO) level and nitrate reductase (NR) activity under Cd stress. On the other hand, the mitigation effects were reversed after removing endogenous NO with its scavenger cPTIO or inhibiting H2-induced NR activity with sodium tungstate. The role of NO in H2-alleviated growth inhibition under Cd stress was proved to be achieved through a restoration of redox balance, an increase in cysteine and proline contents, and a reduction in Cd accumulation. In summary, these results clearly revealed that NR-dependent NO might be involved in the H2-alleviated Cd toxicity in G. lucidum through rebuilding redox homeostasis, increasing cysteine and proline levels, and reducing Cd accumulation. These findings may open a new window for H2 application in Cd-stressed economically important fungi.

Persulfidation of Nitrate Reductase 2 Is Involved in L-Cysteine Desulfhydrase-Regulated Rice Drought Tolerance

Hydrogen sulfide (H2S) is an important signaling molecule that regulates diverse cellular signaling pathways through persulfidation. Our previous study revealed that H2S is involved in the improvement of rice drought tolerance. However, the corresponding enzymatic sources of H2S and its regulatory mechanism in response to drought stress are not clear. Here, we cloned and characterized a putative L-cysteine desulfhydrase (LCD) gene in rice, which encodes a protein possessing H2S-producing activity and was named OsLCD1. Overexpression of OsLCD1 results in enhanced H2S production, persulfidation of total soluble protein, and confers rice drought tolerance. Further, we found that nitrate reductase (NR) activity was decreased under drought stress, and the inhibition of NR activity was controlled by endogenous H2S production. Persulfidation of NIA2, an NR isoform responsible for the main NR activity, led to a decrease in total NR activity in rice. Furthermore, drought stress-triggered inhibition of NR activity and persulfidation of NIA2 was intensified in the OsLCD1 overexpression line. Phenotypical and molecular analysis revealed that mutation of NIA2 enhanced rice drought tolerance by activating the expression of genes encoding antioxidant enzymes and ABA-responsive genes. Taken together, our results showed the role of OsLCD1 in modulating H2S production and provided insight into H2S-regulated persulfidation of NIA2 in the control of rice drought stress.

Nitroreductase Activites in Giardia lamblia: ORF 17150 Encodes a Quinone Reductase with Nitroreductase Activity

The intestinal diplomonadid Giardia lamblia is a causative agent of persistent diarrhea. Current treatments are based on nitro drugs, especially metronidazole. Nitro compounds are activated by reduction, yielding toxic intermediates. The enzymatic systems responsible for this activation are not completely understood. By fractionating cell free crude extracts by size exclusion chromatography followed by mass spectrometry, enzymes with nitroreductase (NR) activities are identified. The protein encoded by ORF 17150 found in two pools with NR activities is overexpressed and characterized. In pools of fractions with main NR activities, previously-known NRs are identified, as well as a previously uncharacterized protein encoded by ORF 17150. Recombinant protein 17150 is a flavoprotein with NADPH-dependent quinone reductase and NR activities. Besides a set of previously identified NRs, we have identified a novel enzyme with NR activity.

Improved Corn Yields When Humic Acid Extracted from Composted Manure Is Applied to Acid Soils with Phosphorus Fertilizer

Most corn planted on tropical acidic soils suffers substantial yield loss caused by low crop phosphorus (P) uptake. Humic acid is recommended to increase crop P uptake since it is capable of competing with P to be bound to soil adsorption complexes. Humic acid extracted from composted manure (MHA) is a good alternative to humic acid because it is more reactive and has a higher complexation ability compared with leonardite. Therefore, the effectiveness of soil-applied MHA combined with P was evaluated to improve corn yields in acidic soils from the aspect of crop nutrient uptake efficiency and crop physiological performance. Two high-yielding corn cultivars that are highly accepted by Indonesian farmers and resistant to downy mildew disease were subjected to five different types of MHA and P combinations. The amount of P fertilizer was equivalent to 120 mg P2O5 kg−1 soil. The H10P1 produced the most significant result p < 0.05 in terms of total crop dry weight, grain dry weight, and corn yield. The highest efficiency for P uptake in the leaves and grains was found with H10P1 and H5P1, respectively. Although chlorophyll content was not substantially improved, the stomatal apertures 7 weeks after planting (WAP) were significantly increased p < 0.05 with H10P1 and H15P1. The photosynthetic rate and nitrate reductase (NR) activity at 10 WAP were significantly increased p < 0.05 with H5P1 and H10P1, respectively. The results clearly indicated significant increases in the efficiency of crop P uptake and physiological performance for stomatal aperture, photosynthetic rate, and NR activity can highly contribute to higher corn yields.

Nuclear Receptors and Development of Marine Invertebrates

Nuclear Receptors (NRs) are a superfamily of transcription factors specific to metazoans that have the unique ability to directly translate the message of a signaling molecule into a transcriptional response. In vertebrates, NRs are pivotal players in countless processes of both embryonic and adult physiology, with embryonic development being one of the most dynamic periods of NR activity. Accumulating evidence suggests that NR signaling is also a major regulator of development in marine invertebrates, although ligands and transactivation dynamics are not necessarily conserved with respect to vertebrates. The explosion of genome sequencing projects and the interpretation of the resulting data in a phylogenetic context allowed significant progress toward an understanding of NR superfamily evolution, both in terms of molecular activities and developmental functions. In this context, marine invertebrates have been crucial for characterizing the ancestral states of NR-ligand interactions, further strengthening the importance of these organisms in the field of evolutionary developmental biology.

Al exposure increases proline levels by different pathways in an Al-sensitive and an Al-tolerant rye genotype

Aluminium (Al) toxicity limits crop productivity, particularly at low soil pH. Proline (Pro) plays a role in protecting plants against various abiotic stresses. Using the relatively Al-tolerant cereal rye (Secale cereale L.), we evaluated Pro metabolism in roots and shoots of two genotypes differing in Al tolerance, var. RioDeva (sensitive) and var. Beira (tolerant). Most enzyme activities and metabolites of Pro biosynthesis were analysed. Al induced increases in Pro levels in each genotype, but the mechanisms were different and were also different between roots and shoots. The Al-tolerant genotype accumulated highest Pro levels and this stronger increase was ascribed to simultaneous activation of the ornithine (Orn)-biosynthetic pathway and decrease in Pro oxidation. The Orn pathway was particularly enhanced in roots. Nitrate reductase (NR) activity, N levels, and N/C ratios demonstrate that N-metabolism is less inhibited in the Al-tolerant line. The correlation between Pro changes and differences in Al-sensitivity between these two genotypes, supports a role for Pro in Al tolerance. Our results suggest that differential responses in Pro biosynthesis may be linked to N-availability. Understanding the role of Pro in differences between genotypes in stress responses, could be valuable in plant selection and breeding for Al resistance.

Different Pharmacokinetic Response to an Acute Dose of Inorganic Nitrate in Patients with Type 2 Diabetes

Aim: We aimed to compare the pharmacokinetics of nitrate (NO3) in patients with type 2 diabetes mellitus (T2DM) and healthy adults. Potential effects of salivary nitrate reductase (NR) activity on cardiometabolic responses to an acute dose of NO3 was also assessed. Methods: Nine healthy adults and nine T2DM patients were recruited to consume a NO3-rich breakfast (~410 mg NO3). Pharmacokinetics of NO3 were examined using repeated measurements of NOx (nitrate+nitrite) concentrations of serum and saliva over 8 hours and NO3 concentrations of spot and 24-h urine samples. Cardiometabolic parameters including serum levels of glucose, insulin, and triglycerides as well as blood pressures were also measured. Results: Compared to patients with T2DM, serum NOx concentration (Δ1= 16.7 vs. 4.4 µmol/L, P=0.057) of healthy subjects sharply increased within 1 hour after NO3 loading. Healthy subjects had a higher NR activity index, and higher peak salivary NO3 concentration with a lower time to peak. Diabetic patients with high- compared to low-NR values had a higher whole-body NOx exposure (103±31.4 vs. 58.9±22.1 µmol*h/L); they also showed a better glycemic response and more reduction of blood pressure following ingestion of a NO3-rich meal. Conclusion: T2DM may be associated with a different pattern of NOx pharmacokinetics (especially salivary NOx metabolism). Salivary NR activity may have a critical role in postprandial metabolism of NO3, and diabetic patients with higher NR activity may take more advantages from NO3 supplementation.

Nitrogen Assimilation in the Highly Salt- and Boron-Tolerant Ecotype Zea mays L. Amylacea

The Lluta Valley in Northern Chile is an important agricultural area affected by both salinity and boron (B) toxicity. Zea mays L. amylacea, an ecotype arisen because of the seed selection practiced in this valley, shows a high tolerance to salt and B levels. In the present study the interaction between B and salt was studied after 20 days of treatment at low (100 mM) and high salinity (430 mM NaCl), assessing changes in nitrogen metabolites and in the activity of key nitrogen-assimilating enzymes. Under non-saline conditions, the presence of excessive B favored higher nitrate and ammonium mobilization to leaves, increasing nitrate reductase (NR) activity but not glutamine synthetase (GS). Thus, the increment of nitrogen use efficiency by B application would contribute partially to maintain the biomass production in this ecotype. Positive relationships between NR activity, nitrate, and stomatal conductance were observed in leaves. The increment of major amino acids alanine and serine would indicate a photoprotective role of photorespiration under low-salinity conditions, thus the inhibition of nitrogen assimilation pathway (NR and GS activities) occurred only at high salinity. The role of cytosolic GS regarding the proline accumulation is discussed.

Relationship of nitrogen use and nitrate reductase activity by sugarcane cultivars

Intrinsic genetic potential of each cultivar of sugarcane may determine the absorption intensity and nitrogen assimilation. It is possible to occur a lower expression of the productive potential by limitations related to low nitrate reductase (NR) activity, since this enzyme is "key" of the N metabolism. The objectives were to compare cultivars of sugarcane as the level of NR activity and its relationship to productive and nutritional variables. The experiment was conducted under condition of a greenhouse, where were grown ten sugarcane cultivars in pots of 4 dm3, filled with sand and vermiculite plus nutrients as indicated for the crop. The variables evaluated were: plant height, NR activity, dry matter (shoot and roots), N content and N accumulation and N uptake and utilization efficiency, moreover, the correlation between the variables was evaluated. The results showed that NR activity varies with the cultivar and further, that NR activity in sugarcane leaves does not correlate with the N uptake efficiency, but is positively correlated with N content and negatively with N utilization efficiency. The NR activity is not a good physiological parameter to discriminate N efficient use sugarcane genotypes

Adaptation of nitrate reductase activity assay for high throughput screening of crops

The possibility of freeze drying of plant material and its grinding in a shaking bead mill to determine the activity of nitrate reductase (NR) was studied. The effectiveness of applying this approach to high throughput mass screening of crops was shown. The assay was carried out on seedlings of common wheat (Triticum aestivum) of the following cultivars: Altigo, Vassa, Grom, Doka, Soberbash, Starshina, Fisht and spring wheat Novosibirskaya 67. The crops were grown during 4-5 weeks on substrate without nitrogen and on substrate supplemented with 50 millimol / l (mM) of potassium nitrate. Nitrate reductase in plants retained its activity after lyophilization and grinding of dried leaves in a mill. The proposed protocol for NR activity assay is suitable for plant lysates with an NR activity suffi cient to form nitrite in the range of 5–120 micromoles / l (μM) in 800 μl of reaction mix (for instance, freeze-dried sample originated from 100 mg of wheat seedling leaves). Centrifugation of a plant lysate at 20,000 g almost did not change NR activity compared to 12,000 g that is achievable for most lab centrifuges. Lysates from fresh leaves contained signifi cantly more total protein than lysates from lyophilized leaves (with an equal amount of starting wet material). The difference in the nitrate-reducing activity in lysates from fresh and lyophilized leaves was not as high as the difference in protein concentration. Thus, the activity of NR calculated per g of total protein was higher in lyophilized leaves than in fresh leaves. The activity of NR was signifi cantly induced by nitrate for all cultivars. The basal and nitrate-induced NR activity varied widely between the cultivars, and the induction ranged from 2.5 fold for Novosibirskaya 67 variety and 2.7 fold for Vassa to 5.4 for Altigo and 5.7 fold for Grom.