RESUMO
MAIN CONCLUSION: Ambient concentrations of atmospheric nitrogen dioxide (NO2) inhibit the binding of PIF4 to promoter regions of auxin pathway genes to suppress hypocotyl elongation in Arabidopsis. Ambient concentrations (10-50 ppb) of atmospheric nitrogen dioxide (NO2) positively regulate plant growth to the extent that organ size and shoot biomass can nearly double in various species, including Arabidopsis thaliana (Arabidopsis). However, the precise molecular mechanism underlying NO2-mediated processes in plants, and the involvement of specific molecules in these processes, remain unknown. We measured hypocotyl elongation and the transcript levels of PIF4, encoding a bHLH transcription factor, and its target genes in wild-type (WT) and various pif mutants grown in the presence or absence of 50 ppb NO2. Chromatin immunoprecipitation assays were performed to quantify binding of PIF4 to the promoter regions of its target genes. NO2 suppressed hypocotyl elongation in WT plants, but not in the pifq or pif4 mutants. NO2 suppressed the expression of target genes of PIF4, but did not affect the transcript level of the PIF4 gene itself or the level of PIF4 protein. NO2 inhibited the binding of PIF4 to the promoter regions of two of its target genes, SAUR46 and SAUR67. In conclusion, NO2 inhibits the binding of PIF4 to the promoter regions of genes involved in the auxin pathway to suppress hypocotyl elongation in Arabidopsis. Consequently, PIF4 emerges as a pivotal participant in this regulatory process. This study has further clarified the intricate regulatory mechanisms governing plant responses to environmental pollutants, thereby advancing our understanding of how plants adapt to changing atmospheric conditions.
Assuntos
Proteínas de Arabidopsis , Arabidopsis , Fatores de Transcrição Hélice-Alça-Hélice Básicos , Regulação da Expressão Gênica de Plantas , Hipocótilo , Dióxido de Nitrogênio , Arabidopsis/genética , Arabidopsis/crescimento & desenvolvimento , Arabidopsis/metabolismo , Hipocótilo/crescimento & desenvolvimento , Hipocótilo/genética , Hipocótilo/efeitos dos fármacos , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Dióxido de Nitrogênio/farmacologia , Dióxido de Nitrogênio/metabolismo , Regiões Promotoras Genéticas/genética , Ácidos Indolacéticos/metabolismo , MutaçãoRESUMO
Although reactive oxygen species (ROS) are known to have harmful effects in organisms, recent studies have demonstrated expression of ROS synthases at various parts of the organisms and the controlled ROS generation, suggesting possible involvement of ROS signaling in physiological events of individuals. However, physiological roles of ROS in the CNS, including functional roles in higher brain functions or neuronal activity-dependent ROS production, remain to be elucidated. Here, we demonstrated involvement of ROS - 8-NO2-cGMP signaling in motor learning and synaptic plasticity in the cerebellum. In the presence of inhibitors of ROS signal or ROS synthases, cerebellar motor learning was impaired, and the stimulus inducing long-term depression (LTD), cellular basis for the motor learning, failed to induce LTD but induced long-term potentiation (LTP)-like change at cerebellar synapses. Furthermore, ROS was produced by LTD-inducing stimulus in enzyme-dependent manner, and excess administration of the antioxidant vitamin E impaired cerebellar motor learning, suggesting beneficial roles of endogenous ROS in the learning. As a downstream signal, involvement of 8-NO2-cGMP in motor learning and cerebellar LTD were also revealed. These findings indicate that ROS - 8-NO2-cGMP signal is activated by neuronal activity and is essential for cerebellum-dependent motor learning and synaptic plasticity, demonstrating involvement of the signal in physiological function of brain systems.
Assuntos
GMP Cíclico/análogos & derivados , Plasticidade Neuronal , Dióxido de Nitrogênio , Humanos , Espécies Reativas de Oxigênio/metabolismo , Dióxido de Nitrogênio/metabolismo , Plasticidade Neuronal/fisiologia , Cerebelo/metabolismo , Memória de Longo PrazoRESUMO
NH3-N and NO2-N always co-exist in the aquatic environment, but there is not a clear opinion on their joint toxicities to the molluscs. Presently, clams Ruditapes philippinarum were challenged by environmental concentrations of NH3-N and NO2-N, singly or in combination, and analyzed by metabolomics approaches, enzyme assays and transmission electron microscope (TEM) observation. Results showed that some same KEGG pathways with different enriched-metabolites were detected in the three exposed groups within one day, and completely different profiles of metabolites were found in the rest of the exposure period. The combined exposure induced heavier and more lasting toxicities to the clams compared with their single exposure. ACP activity and the number of secondary lysosomes were significantly increased after the combined exposure. The present study shed light on the joint-toxicity mechanism of NH3-N and NO2-N, and provided fundamental data for the toxicity research on inorganic nitrogen.
Assuntos
Bivalves , Poluentes Químicos da Água , Animais , Nitritos/toxicidade , Nitritos/metabolismo , Amônia/toxicidade , Amônia/metabolismo , Dióxido de Nitrogênio/metabolismo , Bivalves/metabolismo , Estresse Oxidativo , Poluentes Químicos da Água/toxicidade , Poluentes Químicos da Água/metabolismoRESUMO
BACKGROUND: Skeletal muscles are postulated to be a potent regulator of systemic nitric oxide homeostasis. In this study, we aimed to evaluate the impact of physical training on the heart and skeletal muscle nitric oxide bioavailability (judged on the basis of intramuscular nitrite and nitrate) in rats. METHODS AND RESULTS: Rats were trained on a treadmill for 8 weeks, performing mainly endurance running sessions with some sprinting runs. Muscle nitrite (NO2-) and nitrate (NO3-) concentrations were measured using a high-performance liquid chromatography-based method, while amino acids, pyruvate, lactate, and reduced and oxidized glutathione were determined using a liquid chromatography coupled with tandem mass spectrometry technique. The content of muscle nitrite reductases (electron transport chain proteins, myoglobin, and xanthine oxidase) was assessed by western immunoblotting. We found that 8 weeks of endurance training decreased basal NO2- in the locomotory muscles and in the heart, without changes in the basal NO3-. In the slow-twitch oxidative soleus muscle, the decrease in NO2- was already present after the first week of training, and the content of nitrite reductases remained unchanged throughout the entire period of training, except for the electron transport chain protein content, which increased no sooner than after 8 weeks of training. CONCLUSIONS: Muscle NO2- level, opposed to NO3-, decreases in the time course of training. This effect is rapid and already visible in the slow-oxidative soleus after the first week of training. The underlying mechanisms of training-induced muscle NO2- decrease may involve an increase in the oxidative stress, as well as metabolite changes related to an increased muscle anaerobic glycolytic activity contributing to (1) direct chemical reduction of NO2- or (2) activation of muscle nitrite reductases.
Assuntos
Nitratos , Condicionamento Físico Animal , Ratos , Animais , Nitratos/metabolismo , Nitritos , Óxido Nítrico/metabolismo , Dióxido de Nitrogênio/metabolismo , Músculo Esquelético/metabolismo , Exercício Físico , Nitrito Redutases/metabolismoRESUMO
Nitro-conjugated linoleic acid (NO2-CLA) has been observed to manifest salutary signaling responses, including anti-inflammatory and antioxidant properties. Here, the authors have explored the influence and underlying mechanisms of NO2-CLA on the proinflammatory reaction of murine macrophages that were challenged with lipopolysaccharide (LPS) derived from Prevotella intermedia, a putative periodontopathic bacterium. Treatment of LPS-activated RAW264.7 cells with NO2-CLA notably dampened the secretion of iNOS-derived NO, IL-1ß and IL-6 as well as their gene expressions and significantly enhanced the markers for M2 macrophage polarization. NO2-CLA promoted the HO-1 expression in cells challenged with LPS, and tin protoporphyrin IX, an HO-1 inhibitor, significantly reversed the NO2-CLA-mediated attenuation of NO secretion, but not IL-1ß or IL-6. We found that cells treated with NO2-CLA significantly increased mRNA expression of PPAR-γ compared to control cells, and NO2-CLA significantly reverted the decrease in PPAR-γ mRNA caused by LPS. Nonetheless, antagonists to PPAR-γ were unable to reverse the NO2-CLA-mediated suppression of inflammatory mediators. In addition, NO2-CLA did not alter the p38 and JNK activation elicited by LPS. Both NF-κB reporter activity and IκB-α degradation caused by LPS were notably diminished by NO2-CLA. NO2-CLA was observed to interrupt the nuclear translocation and DNA binding of p50 subunits caused by LPS with no obvious alterations in p65 subunits. Further, NO2-CLA attenuated the phosphorylation of STAT1/3 elicited in response to LPS. We propose that NO2-CLA could be considered as a possible strategy for the therapy of periodontal disease, although additional researches are certainly required to confirm this.
Assuntos
Ácidos Linoleicos Conjugados , Lipopolissacarídeos , Animais , Camundongos , Lipopolissacarídeos/farmacologia , Prevotella intermedia/química , Interleucina-6/metabolismo , Ácidos Linoleicos Conjugados/farmacologia , Ácidos Linoleicos Conjugados/metabolismo , Dióxido de Nitrogênio/metabolismo , Dióxido de Nitrogênio/farmacologia , Receptores Ativados por Proliferador de Peroxissomo/metabolismo , Receptores Ativados por Proliferador de Peroxissomo/farmacologia , Macrófagos , RNA Mensageiro/metabolismoRESUMO
Nitrite (NO2-) interacts with hemoglobin (Hb) in various ways to regulate blood flow. During hypoxic vasodilation, nitrite is reduced by deoxyHb to yield nitric oxide (NO). While NO, a hydrophobic gas, could freely diffuse across the cell membrane, how the reactant nitrite anion could permeate through the red blood cell (RBC) membrane remains unclear. We hypothesized that Cl-/HCO3- anion exchanger-1 (AE1; band 3) abundantly embedded in the RBC membrane could transport NO2-, as HCO3- and NO2- exhibit similar hydrated radii. Here, we monitored NO/N2O3 generated from NO2- inside human RBCs by DAF-FM fluorophore. NO2-, not NO3-, increased intraerythrocytic DAF-FM fluorescence. To test the involvement of AE1-mediated transport in intraerythrocytic NO/N2O3 production from nitrite, we lowered Cl- or HCO3- in the RBC-incubating buffer by 20 % and indeed observed slower rise of the DAF-FM fluorescence. Anti-extracellular AE1, but not anti-intracellular AE1 antibodies, reduced the rates of NO formation from nitrite. The AE1 blocker DIDS similarly reduced the rates of NO/N2O3 production from nitrite in a dose-dependent fashion, confirming that nitrite entered RBCs through AE1. Nitrite inside the RBCs reacted with both deoxyHb and oxyHb, as evidenced by 6.1 % decrease in deoxyHb, 14.7 % decrease in oxyHb, and 20.7 % increase in methemoglobin (metHb). Lowering Cl- in the milieu equally delayed metHb production from nitrite-oxyHb and nitrite-deoxyHb reactions. Thus, AE1-mediated NO2- transport facilitates NO2--Hb reactions inside the red cells, supporting NOx metabolism in circulation.
Assuntos
Óxido Nítrico , Nitritos , Humanos , Nitritos/metabolismo , Óxido Nítrico/metabolismo , Dióxido de Nitrogênio/metabolismo , Hemoglobinas/química , Eritrócitos/metabolismo , Metemoglobina , Proteína 1 de Troca de Ânion do Eritrócito/metabolismo , Membrana Eritrocítica/metabolismoRESUMO
Nitrate (NO3-)-reducing Fe(II) oxidation (NRFO) is prevalent in anoxic environments. However, it is uncertain in which step(s) the biological Fe(II) oxidation is coupled with denitrification during NRFO. In this study, a heterotrophic NRFO bacterium, Diaphorobacter caeni LI3T, was isolated from paddy soil and used to investigate the transformation of Fe(II) and nitrogen as well as nitrogen isotopic fractionation (δ15N-N2O) during NRFO. Fe(II) oxidation was observed in the Cell+NO3- +Fe(II), Cell+NO2- + Fe(II), and NO2- + Fe(II) treatments, resulting in precipitation of amorphous Fe(III) minerals and lepidocrocite on the surface and in the periplasm of cells. The presence of Fe(II) slightly accelerated microbial NO3- reduction in the Cell+NO3- + Fe(II) treatment relative to the Cell+NO3- treatment, but slowed down the NO2- reduction in the Cell+NO2- + Fe(II) treatment relative to the Cell+NO2- treatment likely due to cell encrustation that blocking microbial NO2- reduction in the periplasm. The δ15N-N2O results in the Cell+NO3- + Fe(II) treatment were close to those in the Cell+NO3- and Cell+NO2- treatments, indicating that the accumulative N2O is primarily of biological origin during NRFO. The genome analysis found a complete set of denitrification and oxidative phosphorylation genes in strain LI3T, the metabolic pathways of which were closely related with cyc2 and cytc as indicated by protein-protein interactions network analysis. It is proposed that Fe(II) oxidation is catalyzed by the outer membrane protein Cyc2, with the resulting electrons being transferred to the nitrite reductase NirS via CytC in the periplasm, and the CytC can also accept electrons from the oxidative phosphorylation in the cytoplasmic membrane. Overall, our findings provide new insights into the potential pathways of biological Fe(II) oxidation coupled with nitrate reduction in heterotrophic NRFO bacteria.
Assuntos
Compostos Férricos , Nitratos , Nitratos/metabolismo , Compostos Férricos/metabolismo , Nitritos/metabolismo , Isótopos de Nitrogênio , Dióxido de Nitrogênio/metabolismo , Oxirredução , Bactérias/metabolismo , Nitrogênio/metabolismo , Compostos Ferrosos/metabolismo , DesnitrificaçãoRESUMO
Denitrification is a form of anaerobic respiration wherein nitrate (NO3-) is sequentially reduced via nitrite (NO2-), nitric oxide, and nitrous oxide (N2O) to dinitrogen gas (N2) by four reductase enzymes. Partial denitrifying bacteria possess only one or some of these four reductases and use them as independent respiratory modules. However, it is unclear if partial denitrifiers sense and respond to denitrification intermediates outside of their reductase repertoire. Here, we tested the denitrifying capabilities of two purple nonsulfur bacteria, Rhodopseudomonas palustris CGA0092 and Rhodobacter capsulatus SB1003. Each had denitrifying capabilities that matched their genome annotation; CGA0092 reduced NO2- to N2, and SB1003 reduced N2O to N2. For each bacterium, N2O reduction could be used both for electron balance during growth on electron-rich organic compounds in light and for energy transformation via respiration in darkness. However, N2O reduction required supplementation with a denitrification intermediate, including those for which there was no associated denitrification enzyme. For CGA0092, NO3- served as a stable, non-catalyzable molecule that was sufficient to activate N2O reduction. Using a ß-galactosidase reporter, we found that NO3- acted, at least in part, by stimulating N2O reductase gene expression. In SB1003, NO2- but not NO3- activated N2O reduction, but NO2- was slowly removed, likely by a promiscuous enzyme activity. Our findings reveal that partial denitrifiers can still be subject to regulation by denitrification intermediates that they cannot use.IMPORTANCEDenitrification is a form of microbial respiration wherein nitrate is converted via several nitrogen oxide intermediates into harmless dinitrogen gas. Partial denitrifying bacteria, which individually have some but not all denitrifying enzymes, can achieve complete denitrification as a community by cross-feeding nitrogen oxide intermediates. However, the last intermediate, nitrous oxide (N2O), is a potent greenhouse gas that often escapes, motivating efforts to understand and improve the efficiency of denitrification. Here, we found that at least some partial denitrifying N2O reducers can sense and respond to nitrogen oxide intermediates that they cannot otherwise use. The regulatory effects of nitrogen oxides on partial denitrifiers are thus an important consideration in understanding and applying denitrifying bacterial communities to combat greenhouse gas emissions.
Assuntos
Gases de Efeito Estufa , Óxido Nitroso , Óxido Nitroso/metabolismo , Desnitrificação , Nitratos/metabolismo , Gases de Efeito Estufa/metabolismo , Dióxido de Nitrogênio/metabolismo , Dióxido de Nitrogênio/farmacologia , Bactérias/genética , Óxido Nítrico/metabolismo , Oxirredutases/metabolismoRESUMO
Current research has described improving multisystem disease and organ function through dietary nitrate (DN) supplementation. They have provided some evidence that these floras with nitrate (NO3-) reductase are mediators of the underlying mechanism. Symbiotic bacteria with nitrate reductase activity (NRA) are found in the human digestive tract, including the mouth, esophagus and gastrointestinal tract (GT). Nitrate in food can be converted to nitrite under the tongue or in the stomach by these symbiotic bacteria. Then, nitrite is transformed to nitric oxide (NO) by non-enzymatic synthesis. NO is currently recognized as a potent bioactive agent with biological activities, such as vasodilation, regulation of cardiomyocyte function, neurotransmission, suppression of platelet agglutination, and prevention of vascular smooth muscle cell proliferation. NO also can be produced through the conventional L-arginine-NO synthase (L-NOS) pathway, whereas endogenous NO production by L-arginine is inhibited under hypoxia-ischemia or disease conditions. In contrast, exogenous NO3-/NO2-/NO activity is enhanced and becomes a practical supplemental pathway for NO in the body, playing an essential role in various physiological activities. Moreover, many diseases (such as metabolic or geriatric diseases) are primarily associated with disorders of endogenous NO synthesis, and NO generation from the exogenous NO3-/NO2-/NO route can partially alleviate the disease progression. The imbalance of NO in the body may be one of the potential mechanisms of disease development. Therefore, the impact of these floras with nitrate reductase on host systemic health through exogenous NO3-/NO2-/NO pathway production of NO or direct regulation of floras ecological balance is essential (e.g., regulation of body homeostasis, amelioration of diseases, etc.). This review summarizes the bacteria with nitrate reductase in humans, emphasizing the relationship between the metabolic processes of this microflora and host systemic health and disease. The potential effects of nitrate reduction bacteria on human health and disease were also highlighted in disease models from different human systems, including digestive, cardiovascular, endocrine, nervous, respiratory, and urinary systems, providing innovative ideas for future disease diagnosis and treatment based on nitrate reduction bacteria.
Assuntos
Nitratos , Nitritos , Humanos , Idoso , Nitratos/farmacologia , Nitratos/metabolismo , Nitritos/metabolismo , Óxido Nítrico/metabolismo , Dióxido de Nitrogênio/metabolismo , Bactérias/metabolismo , Nitrato Redutases/metabolismo , Arginina/metabolismoRESUMO
Lactoperoxidase (LPO) is a heme containing mammalian enzyme which uses hydrogen peroxide (H2O2) to catalyze the conversion of substrates into oxidized products. LPO is found in body fluids and tissues such as milk, saliva, tears, mucosa and other body secretions. The previous structural studies have shown that LPO converts substrates, thiocyanate (SCN-) and iodide (I-) ions into oxidized products, hypothiocyanite (OSCN-) and hypoiodite (IO-) ions respectively. We report here a new structure of the complex of LPO with an oxidized product, nitrite (NO2-). This product was generated from NO using the two step reaction of LPO by adding hydrogen peroxide (H2O2) in the solution of LPO in 0.1 M phosphate buffer at pH 6.8 as the first step. In the second step, NO gas was added to the above mixture. This was crystallized using 20% (w/v) PEG-3350 and 0.2 M ammonium iodide at pH 6.8. The structure determination showed the presence of NO2- ion in the distal heme cavity of the substrate binding site of LPO. The structure also showed that the propionate group which is linked to pyrrole ring D of the heme moiety was disordered. Similarly, the side chain of Asp108, which is covalently linked to heme moiety, was also split into two components. As a result of these changes, the conformation of the side chain of Arg255 was altered allowing it to form new interactions with the disordered carboxylic group of propionate moiety. These structural changes are indicative of an intermediate state in the catalytic reaction pathway of LPO.
Assuntos
Lactoperoxidase , Nitritos , Animais , Lactoperoxidase/química , Nitritos/metabolismo , Óxido Nítrico/metabolismo , Peróxido de Hidrogênio/metabolismo , Dióxido de Nitrogênio/metabolismo , Propionatos , Mamíferos/metabolismo , Heme/químicaRESUMO
An important role of nitric oxide (NO) as either a free intermediate in the NH3 oxidation pathway or a potential oxidant for NH3 or NH2OH has been proposed for ammonia-oxidizing bacteria (AOB) and archaea (AOA), respectively. However, tracing NO metabolism at low concentrations remains notoriously difficult. Here, we use electrochemical sensors and the mild NO scavenger 2-phenyl-4,4,5,5-tetramethylimidazoline-1-oxyl 3-oxide (PTIO) to trace apparent NO concentration and determine production rates at low micromolar concentrations in the model AOB strain Nitrosomonas europaea. In agreement with previous studies, we found that PTIO does not affect NH3 oxidation instantaneously in both Nitrosospira briensis and Nitrosomonas europaea, unlike inhibitors for ammonia oxidation such as allylthiourea and acetylene, although it effectively scavenged NO from the cell suspensions. Quantitative analysis showed that NO production by N. europaea amounted to 3.15% to 6.23% of NO2- production, whereas N. europaea grown under O2 limitation produced NO equivalent to up to 40% of NO2- production at high substrate concentrations. In addition, we found that PTIO addition to N. europaea grown under O2 limitation abolished N2O production. These results indicate different turnover rates of NO during NH3 oxidation under O2-replete and O2-limited growth conditions in AOB. The results suggest that NO may not be a free intermediate or remain tightly bound to iron centers of enzymes during hydroxylamine oxidation and that only NH3 saturation and adaptation to O2 limitation may lead to significant dissociation of NO from hydroxylamine dehydrogenase. IMPORTANCE Ammonia oxidation by chemolithoautotrophic ammonia-oxidizing bacteria (AOB) is thought to contribute significantly to global nitrous oxide (N2O) emissions and leaching of oxidized nitrogen, particularly through their activity in nitrogen (N)-fertilized agricultural production systems. Although substantial efforts have been made to characterize the N metabolism in AOB, recent findings suggest that nitric oxide (NO) may play an important mechanistic role as a free intermediate of hydroxylamine oxidation in AOB, further implying that besides hydroxylamine dehydrogenase (HAO), additional enzymes may be required to complete the ammonia oxidation pathway. However, the NO spin trap PTIO was found to not inhibit ammonia oxidation in AOB. This study provides a combination of physiological and spectroscopic evidence that PTIO indeed scavenges only free NO in AOB and that significant amounts of free NO are produced only during incomplete hydroxylamine oxidation or nitrifier denitrification under O2-limited growth conditions.
Assuntos
Betaproteobacteria , Óxido Nítrico , Óxido Nítrico/metabolismo , Amônia/metabolismo , Hidroxilamina/química , Hidroxilamina/metabolismo , Dióxido de Nitrogênio/metabolismo , Oxirredução , Óxido Nitroso/metabolismo , Archaea/metabolismo , Betaproteobacteria/metabolismo , Nitrogênio/metabolismo , Hidroxilaminas/metabolismo , NitrificaçãoRESUMO
Prochlorococcus is an abundant photosynthetic bacterium in the open ocean, where nitrogen (N) often limits phytoplankton growth. In the low-light-adapted LLI clade of Prochlorococcus, nearly all cells can assimilate nitrite (NO2-), with a subset capable of assimilating nitrate (NO3-). LLI cells are maximally abundant near the primary NO2- maximum layer, an oceanographic feature that may, in part, be due to incomplete assimilatory NO3- reduction and subsequent NO2- release by phytoplankton. We hypothesized that some Prochlorococcus exhibit incomplete assimilatory NO3- reduction and examined NO2- accumulation in cultures of three Prochlorococcus strains (MIT0915, MIT0917, and SB) and two Synechococcus strains (WH8102 and WH7803). Only MIT0917 and SB accumulated external NO2- during growth on NO3-. Approximately 20-30% of the NO3- transported into the cell by MIT0917 was released as NO2-, with the rest assimilated into biomass. We further observed that co-cultures using NO3- as the sole N source could be established for MIT0917 and Prochlorococcus strain MIT1214 that can assimilate NO2- but not NO3-. In these co-cultures, the NO2- released by MIT0917 is efficiently consumed by its partner strain, MIT1214. Our findings highlight the potential for emergent metabolic partnerships that are mediated by the production and consumption of N cycle intermediates within Prochlorococcus populations. IMPORTANCE Earth's biogeochemical cycles are substantially driven by microorganisms and their interactions. Given that N often limits marine photosynthesis, we investigated the potential for N cross-feeding within populations of Prochlorococcus, the numerically dominant photosynthetic cell in the subtropical open ocean. In laboratory cultures, some Prochlorococcus cells release extracellular NO2- during growth on NO3-. In the wild, Prochlorococcus populations are composed of multiple functional types, including those that cannot use NO3- but can still assimilate NO2-. We show that metabolic dependencies arise when Prochlorococcus strains with complementary NO2- production and consumption phenotypes are grown together on NO3-. These findings demonstrate the potential for emergent metabolic partnerships, possibly modulating ocean nutrient gradients, that are mediated by cross-feeding of N cycle intermediates.
Assuntos
Prochlorococcus , Synechococcus , Nitritos/metabolismo , Dióxido de Nitrogênio/metabolismo , Nitratos/metabolismo , Synechococcus/genética , FitoplânctonRESUMO
Antibiotics and heavy metals can have a negative impact on the nitrogen (N) cycle and microbial metabolism in coastal aquaculture environment. An indoor simulated culture experiment was conducted to explore how sulfadiazine and lead influence the N cycling in aquatic environment. Specifically, the experiment involved adding sulfadiazine (SDZ), lead (Pb), a combination of SDZ and Pb (SP), and a control group (CK). The fluxes and contents of ammonia nitrogen (NH4+-N), nitrate nitrogen (NO3--N) and nitrite nitrogen (NO2--N) in sediment-water interface and sediments, the abundance of N cycle function genes (amoA_AOB, hzsA, nar, nirK, nirS, norB and nosZ) and microbiota in sediments were analyzed. The results showed that the presence of SDZ and Pb inhibited the nitrification function gene and nitrifiers abundance in surface sediment, and thus leading to more accumulation of NH4+ and NO2- in overlying water. Pb exposure increased the abundances of denitrifying bacteria stimulated the first three steps of denitrification in the sediment, resulting in more removal of NO3- from the sediment, but possibly had the risk of releasing more greenhouse gas N2O. Conversely, the presence of SDZ ultimately inhibited denitrification and anammox bacterial activities in the sediment. This study revealed how heavy metal and antibiotic impair the microbial communities and N cycling function gene expression, leading to the deterioration of typical coastal aquaculture environments.
Assuntos
Desnitrificação , Metais Pesados , Antibacterianos/toxicidade , Antibacterianos/metabolismo , Água/metabolismo , Dióxido de Nitrogênio/metabolismo , Chumbo/metabolismo , Ciclo do Nitrogênio , Bactérias/metabolismo , Metais Pesados/toxicidade , Metais Pesados/metabolismo , Nitrogênio/metabolismo , Sulfadiazina/metabolismoRESUMO
This study tested the hypothesis that the increases in salivary and plasma [NO2-] after dietary NO3- supplementation would be greater when oral temperature and pH were independently elevated, and increased further when oral temperature and pH were elevated concurrently. Seven healthy males (mean ± SD, age 23 ± 4 years) ingested 70 mL of beetroot juice concentrate (BR, which provided ~6.2 mmol NO3-) during six separate laboratory visits. In a randomised crossover experimental design, salivary and plasma [NO3-] and [NO2-] were assessed at a neutral oral pH with a low (TLo-pHNorm), intermediate (TMid-pHNorm), and high (THi-pHNorm) oral temperature, and when the oral pH was increased at a low (TLo-pHHi), intermediate (TMid-pHHi), and high (THi-pHHi) oral temperature. Compared with the TMid-pHNorm condition (976 ± 388 µM), the mean salivary [NO2-] 1-3 h post BR ingestion was higher in the TMid-pHHi (1855 ± 423 µM), THi-pHNorm (1371 ± 653 µM), THi-pHHi (1792 ± 741 µM), TLo-pHNorm (1495 ± 502 µM), and TLo-pHHi (2013 ± 662 µM) conditions, with salivary [NO2-] also higher at a given oral temperature when the oral pH was increased (p < 0.05). Plasma [NO2-] was higher 3 h post BR ingestion in the TMid-pHHi, THi-pHHi, and TLo-pHHi conditions, but not the TLo-pHNorm and THi-pHNorm conditions, compared with TMid-pHNorm (p < 0.05). Therefore, despite ingesting the same NO3- dose, the increases in salivary [NO2-] varied depending on the temperature and pH of the oral cavity, while the plasma [NO2-] increased independently of oral temperature, but to a greater extent at a higher oral pH.
Assuntos
Beta vulgaris , Nitratos , Humanos , Adulto , Masculino , Adulto Jovem , Nitritos , Dióxido de Nitrogênio/metabolismo , Temperatura , Saliva/metabolismo , Concentração de Íons de Hidrogênio , Suplementos NutricionaisRESUMO
Inflammation and oxidative and nitrosative stress are involved in the pathogenesis of proliferative retinopathies (PR). In PR, a loss of balance between pro-angiogenic and anti-angiogenic factors favors the secretion of vascular endothelial growth factor (VEGF). This vascular change results in alterations in the blood-retinal barrier, with extravasation of plasma proteins such as α2-macroglobulin (α2M) and gliosis in Müller glial cells (MGCs, such as MIO-M1). It is well known that MGCs play important roles in healthy and sick retinas, including in PR. Nitro-fatty acids are electrophilic lipid mediators with anti-inflammatory and cytoprotective properties. Our aim was to investigate whether nitro-oleic acid (NO2-OA) is beneficial against oxidative stress, gliosis, and the pro-angiogenic response in MGCs. Pure synthetic NO2-OA increased HO-1 expression in a time- and concentration-dependent manner, which was abrogated by the Nrf2 inhibitor trigonelline. In response to phorbol 12-myristate 13-acetate (PMA) and lipopolysaccharide (LPS), NO2-OA prevented the ROS increase and reduced the gliosis induced by α2M. Finally, when hypoxic MGCs were incubated with NO2-OA, the increase in VEGF mRNA expression was not affected, but under hypoxia and inflammation (IL-1ß), NO2-OA significantly reduced VEGF mRNA levels. Furthermore, NO2-OA inhibited endothelial cell (BAEC) tubulogenesis. Our results highlight NO2-OA's protective effect on oxidative damage, gliosis; and the exacerbated pro-angiogenic response in MGCs.
Assuntos
Dióxido de Nitrogênio , Fator A de Crescimento do Endotélio Vascular , Humanos , Dióxido de Nitrogênio/metabolismo , Dióxido de Nitrogênio/farmacologia , Fator A de Crescimento do Endotélio Vascular/metabolismo , Células Ependimogliais/metabolismo , Gliose/metabolismo , Estresse Oxidativo , Hipóxia/metabolismo , Inflamação/metabolismo , RNA Mensageiro/metabolismoRESUMO
In order to solve nitrogen pollution in environmental water, two heterotrophic nitrifying and aerobic denitrifying strains isolated from acid paddy soil were identified as Achromobacter sp. strain HNDS-1 and Enterobacter sp. strain HNDS-6 respectively. Strain HNDS-1 and strain HNDS-6 exhibited amazing ability to nitrogen removal. When (NH4)2SO4, KNO3, NaNO2 were used as nitrogen resource respectively, the NH4+-N, NO3--N, NO2--N removal efficiencies of strain HNDS-1 were 93.31%, 89.47%, and 100% respectively, while those of strain HNDS-6 were 82.39%, 96.92%, and 100%. And both of them could remove mixed nitrogen effectively in low C/N (C/Nâ¯=â¯5). Strain HNDS-1 could remove 76.86% NH4+-N and 75.13% NO3--N. And strain HNDS-6 can remove 65.07% NH4+-N and 78.21% NO3--N. A putative ammonia monooxygenase, nitrite reductase, nitrate reductase, assimilatory nitrate reductase, nitrate/nitrite transport protein and nitric oxide reductase of strain HNDS-1, while hydroxylamine reductase, nitrite reductase, nitrate reductase, assimilatory nitrate reductase, nitrate/nitrite transport protein, and nitric oxide reductase of strain HNDS-6 were identified by genomic analysis. DNA-SIP analysis showed that genes Nxr, narG, nirK, norB, nosZ were involved in nitrogen removal pathway, which indicates that the denitrification pathway of strain HNDS-1 and strain HNDS-6 was NO3-âNO2-âNOâN2OâN2 during NH4+-N removal process. And the nitrification pathway of strain HNDS-1 and strain HNDS-6 was NO2-âNO3-, but the nitrification pathway of NH4+â NO2- needs further studies.
Assuntos
Achromobacter , Desnitrificação , Enterobacter , Nitrificação , Achromobacter/genética , Achromobacter/metabolismo , Aerobiose/genética , Aerobiose/fisiologia , Desnitrificação/genética , Desnitrificação/fisiologia , Enterobacter/genética , Enterobacter/metabolismo , Nitratos/metabolismo , Nitrificação/genética , Nitrificação/fisiologia , Nitrito Redutases/metabolismo , Nitritos/metabolismo , Nitrogênio/metabolismo , Dióxido de Nitrogênio/metabolismoRESUMO
The second step of nitrification can be mediated by nitrite oxidizing bacteria (NOB), i.e. Nitrospira and Nitrobacter, with different characteristics in terms of the r/K theory. In this study, an activated sludge model was developed to account for competition between two groups of canonical NOB and comammox bacteria. Heterotrophic denitrification on soluble microbial products was also incorporated into the model. Four 5-week washout trials were carried out at dissolved oxygen-limited conditions for different temperatures (12 °C vs. 20 °C) and main substrates (NH4+-N vs. NO2--N). Due to the aggressive reduction of solids retention time (from 4 to 1 d), the biomass concentrations were continuously decreased and stabilized after two weeks at a level below 400 mg/L. The collected experimental data (N species, biomass concentrations, and microbiological analyses) were used for model calibration and validation. In addition to the standard predictions (N species and biomass), the newly developed model also accurately predicted two microbiological indicators, including the relative abundance of comammox bacteria as well as nitrifiers to heterotrophs ratio. Sankey diagrams revealed that the relative contributions of specific microbial groups to N conversion pathways were significantly shifted during the trial. The contribution of comammox did not exceed 5 % in the experiments with both NH4+-N and NO2--N substrates. This study contributes to a better understanding of the novel autotrophic N removal processes (e.g. deammonification) with nitrite as a central intermediate product.
Assuntos
Nitritos , Esgotos , Nitritos/metabolismo , Amônia/metabolismo , Dióxido de Nitrogênio/metabolismo , Oxirredução , Bactérias/metabolismo , Nitrificação , Reatores BiológicosRESUMO
A marine aerobic denitrifying bacterium was isolated and identified as Pseudomonas stutzeri BBW831 from the seabed silt of Beibu Gulf in China. According to the genome analysis, P. stutzeri BBW831 possessed a total of 14 genes (narG, narH, narI, narJ, napA, napB, nirB, nirD, nirS, norB, norC, norD, norQ, and nosZ) responsible for fully functional enzymes (nitrate reductase, nitrite reductase, nitric oxide reductase, and nitrous oxide reductase) involved in the complete aerobic denitrification pathway, suggesting that it had the potential for reducing nitrate to the final N2. Denitrification results showed that P. stutzeri BBW831 exhibited efficient nitrogen removal characteristics. Within 12 h, the NO3--N removal efficiency and rate reached 94.64% and 13.09 mg·L-1·h-1 under 166.10 ± 3.75 mg/L NO3--N as the sole nitrogen source, and removal efficiency of the mixed nitrogen (50.50 ± 0.55, 62.28 ± 0.74, and 64.26 ± 0.90 mg/L of initial NH4+-N, NO3--N, and NO2--N, respectively) was nearly 100%. Furthermore, a simplified strategy, by augmenting the inoculation biomass, was developed for promoting the nitrogen removal performance under high levels of NO2--N and salinity. As a result, the removal efficiency of the initial NO2--N up to approximately 130 mg/L reached 99.46% within 8 h, and the NO3--N removal efficiency achieved at 59.46% under the NaCl concentration even up to 50 g/L. The C/N ratio of 10 with organic acid salt such as trisodium citrate and sodium acetate as the carbon source was most conducive for cell growth and nitrogen removal by P. stutzeri BBW831, respectively. In conclusion, the marine P. stutzeri BBW831 contained the functional genes responsible for a complete aerobic denitrification pathway (NO3--N â NO2--N â NO â N2O â N2), and had great potential for the practical treatment of high-salinity nitrogenous mariculture wastewater.
Assuntos
Pseudomonas stutzeri , Desnitrificação , Nitratos , Nitrogênio/metabolismo , Dióxido de Nitrogênio/metabolismo , Pseudomonas stutzeri/genética , Pseudomonas stutzeri/metabolismoRESUMO
Phosphopantetheinyl transferases (PPTases) play important roles in activating apo-acyl carrier proteins (apo-ACPs) and apo-peptidyl carrier proteins (apo-PCPs) in both primary and secondary metabolism. PPTases catalyze the posttranslational modifications of those carrier proteins by covalent attachment of the 4'-phosphopantetheine group to a conserved serine residue. The protein-protein interactions between a PPTase and a cognate acyl or peptidyl carrier protein have important regulatory functions in microbial biosynthesis, but the molecular mechanism underlying their specific recognition remains elusive. In this study, we identified a new rishirilide biosynthetic gene cluster with a rare in-cluster PPTase from Streptomyces xanthophaeus no2. The function of this Sfp-type PPTase, SxrX, in rishirilide production was confirmed using genetic mutagenesis and biochemical characterization. We applied molecular modeling and site-directed mutagenesis to identify key residues mediating the protein-protein interaction between SxrX and its cognate ACP. In addition, six natural products were isolated from wild-type S. xanthophaeus no2 and the ΔsxrX mutant, including rishirilide A and lupinacidin A, that exhibited antimicrobial and anticancer activities, respectively. SxrX is the first Sfp-type PPTase identified from an aromatic polyketide biosynthetic gene cluster and shown to be responsible for high-level production of rishirilide derivatives. IMPORTANCE Genome mining has been a vital means for natural product drug discovery in the postgenomic era. The rishirilide-type polyketides have attracted attention due to their potent bioactivity, but the poor robustness of production hosts has limited further research and development. This study not only identifies a hyperproducer of rishirilides but also reveals a rare, in-cluster PPTase SxrX that plays an important role in boosting rishirilide biosynthesis. Experimental and computational investigations revealed new insights on the protein-protein interaction between SxrX and its cognate ACP with wide implications for understanding polyketide biosynthesis.
Assuntos
Proteínas de Bactérias , Dióxido de Nitrogênio , Dióxido de Nitrogênio/metabolismo , Proteínas de Bactérias/metabolismo , Proteínas de Transporte/metabolismoRESUMO
Nitric oxide (NO) stimulates mitochondrial biogenesis in skeletal muscle. However, NO metabolism is disrupted in individuals with type 2 diabetes mellitus (T2DM) potentially contributing to their decreased cardiorespiratory fitness (i.e., VO2max) and skeletal muscle oxidative capacity. We used a randomized, double-blind, placebo-controlled, 8-week trial with beetroot juice containing nitrate (NO3−) and nitrite (NO2−) (250 mg and 20 mg/day) to test potential benefits on VO2max and skeletal muscle oxidative capacity in T2DM. T2DM (N = 36, Age = 59 ± 9 years; BMI = 31.9 ± 5.0 kg/m2) and age- and BMI-matched non-diabetic controls (N = 15, Age = 60 ± 9 years; BMI = 29.5 ± 4.6 kg/m2) were studied. Mitochondrial respiratory capacity was assessed in muscle biopsies from a subgroup of T2DM and controls (N = 19 and N = 10, respectively). At baseline, T2DM had higher plasma NO3− (100%; p < 0.001) and lower plasma NO2− levels (−46.8%; p < 0.0001) than controls. VO2max was lower in T2DM (−26.4%; p < 0.001), as was maximal carbohydrate- and fatty acid-supported oxygen consumption in permeabilized muscle fibers (−26.1% and −25.5%, respectively; p < 0.05). NO3−/NO2− supplementation increased VO2max (5.3%; p < 0.01). Further, circulating NO2−, but not NO3−, positively correlated with VO2max after supplementation (R2= 0.40; p < 0.05). Within the NO3−/NO2− group, 42% of subjects presented improvements in both carbohydrate- and fatty acid-supported oxygen consumption in skeletal muscle (vs. 0% in placebo; p < 0.05). VO2max improvements in these individuals tended to be larger than in the rest of the NO3−/NO2− group (1.21 ± 0.51 mL/(kg*min) vs. 0.31 ± 0.10 mL/(kg*min); p = 0.09). NO3−/NO2− supplementation increases VO2max in T2DM individuals and improvements in skeletal muscle oxidative capacity appear to occur in those with more pronounced increases in VO2max.