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Nitric oxide signaling, metabolism and toxicity in nitrogen-fixing symbiosis.
Berger, A, Boscari, A, Frendo, P, Brouquisse, R
Journal of experimental botany. 2019;(17):4505-4520
Abstract
Interactions between legumes and rhizobia lead to the establishment of a symbiotic relationship characterized by the formation of a new organ, the nodule, which facilitates the fixation of atmospheric nitrogen (N2) by nitrogenase through the creation of a hypoxic environment. Significant amounts of nitric oxide (NO) accumulate at different stages of nodule development, suggesting that NO performs specific signaling and/or metabolic functions during symbiosis. NO, which regulates nodule gene expression, accumulates to high levels in hypoxic nodules. NO accumulation is considered to assist energy metabolism within the hypoxic environment of the nodule via a phytoglobin-NO-mediated respiration process. NO is a potent inhibitor of the activity of nitrogenase and other plant and bacterial enzymes, acting as a developmental signal in the induction of nodule senescence. Hence, key questions concern the relative importance of the signaling and metabolic functions of NO versus its toxic action and how NO levels are regulated to be compatible with nitrogen fixation functions. This review analyses these paradoxical roles of NO at various stages of symbiosis, and highlights the role of plant phytoglobins and bacterial hemoproteins in the control of NO accumulation.
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Phytoglobins in the nuclei, cytoplasm and chloroplasts modulate nitric oxide signaling and interact with abscisic acid.
Rubio, MC, Calvo-Begueria, L, Díaz-Mendoza, M, Elhiti, M, Moore, M, Matamoros, MA, James, EK, Díaz, I, Pérez-Rontomé, C, Villar, I, et al
The Plant journal : for cell and molecular biology. 2019;(1):38-54
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Abstract
Symbiotic hemoglobins provide O2 to N2 -fixing bacteria within legume nodules, but the functions of non-symbiotic hemoglobins or phytoglobins (Glbs) are much less defined. Immunolabeling combined with confocal microscopy of the Glbs tagged at the C-terminus with green fluorescent protein was used to determine their subcellular localizations in Arabidopsis and Lotus japonicus. Recombinant proteins were used to examine nitric oxide (NO) scavenging in vitro and transgenic plants to show S-nitrosylation and other in vivo interactions with NO and abscisic acid (ABA) responses. We found that Glbs occur in the nuclei, chloroplasts and amyloplasts of both model plants, and also in the cytoplasm of Arabidopsis cells. The proteins show similar NO dioxygenase activities in vitro, are nitrosylated in Cys residues in vivo, and scavenge NO in the stomatal cells. The Cys/Ser mutation does not affect NO dioxygenase activity, and S-nitrosylation does not significantly consume NO. We demonstrate an interaction between Glbs and ABA on several grounds: Glb1 and Glb2 scavenge NO produced in stomatal guard cells following ABA supply; plants overexpressing Glb1 show higher constitutive expression of the ABA responsive genes Responsive to ABA (RAB18), Responsive to Dehydration (RD29A) and Highly ABA-Induced 2 (HAI2), and are more tolerant to dehydration; and ABA strongly upregulates class 1 Glbs. We conclude that Glbs modulate NO and interact with ABA in crucial physiological processes such as the plant's response to dessication.
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Cyclooxygenase (COX) Inhibition by Acetyl Salicylic Acid (ASA) Enhances Antitumor Effects of Nitric Oxide in Glioblastoma In Vitro.
Guenzle, J, Garrelfs, NWC, Goeldner, JM, Weyerbrock, A
Molecular neurobiology. 2019;(9):6046-6055
Abstract
Glioblastoma multiforme (GBM) is the most aggressive brain tumor with a high recurrence rate and a median survival of about 16 months even with multimodal therapy. Novel experimental strategies against malignant gliomas include cyclooxygenase (COX) inhibition and nitric oxide (NO)-based therapies. Therapeutic doses of NO can be delivered to tumor cells by NO donors such as JS-K (O2-(2,4-dinitrophenyl)1-[(4-ethoxycarbonyl)piperazin-1-yl]diazen-1-ium-1,2-diolate) which releases NO upon enzymatic activation by glutathione S-transferase. COX-2 is frequently overexpressed in tumors and increases tumor invasiveness and angiogenesis. In this study, we show that pretreatment with acetyl salicylic acid (ASA) enhanced the cytotoxic antitumor effect of NO in vitro. Combination of low doses of JS-K and ASA revealed a dose-dependent synergistic increase of necrotic cell death under circumvention of classical apoptosis and alteration of the metabolic calcium level. These findings provide an opportunity to improve currently used therapeutic strategies in the treatment of gliomas with a well-established remedy.
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Plant peroxisomes at the crossroad of NO and H2 O2 metabolism.
Corpas, FJ, Del Río, LA, Palma, JM
Journal of integrative plant biology. 2019;(7):803-816
Abstract
Plant peroxisomes are subcellular compartments involved in many biochemical pathways during the life cycle of a plant but also in the mechanism of response against adverse environmental conditions. These organelles have an active nitro-oxidative metabolism under physiological conditions but this could be exacerbated under stress situations. Furthermore, peroxisomes have the capacity to proliferate and also undergo biochemical adaptations depending on the surrounding cellular status. An important characteristic of peroxisomes is that they have a dynamic metabolism of reactive nitrogen and oxygen species (RNS and ROS) which generates two key molecules, nitric oxide (NO) and hydrogen peroxide (H2 O2 ). These molecules can exert signaling functions by means of post-translational modifications that affect the functionality of target molecules like proteins, peptides or fatty acids. This review provides an overview of the endogenous metabolism of ROS and RNS in peroxisomes with special emphasis on polyamine and uric acid metabolism as well as the possibility that these organelles could be a source of signal molecules involved in the functional interconnection with other subcellular compartments.
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Effect of fenofibrate on serum nitric oxide levels in patients with hypertriglyceridemia.
Esenboga, K, Çiçek, ÖF, Oktay, AA, Ayral, PA, Gürlek, A
Advances in clinical and experimental medicine : official organ Wroclaw Medical University. 2019;(7):931-936
Abstract
BACKGROUND Fenofibrate, a peroxisome proliferator-activated receptor-α (PPARα) agonist, is used to treat patients with hypercholesterolemia and hypertriglyceridemia in order to reduce the risk of development of the atherosclerotic cardiovascular disease. However, it exerts pleiotropic effects beyond correcting atherogenic dyslipidemia to treat hypercholesterolemia. OBJECTIVES The aim of this study was to investigate the potential effects of fenofibrate on endothelial function by analyzing the serum nitric oxide (NO) levels in patients with hypertriglyceridemia. MATERIAL AND METHODS Lipid profiles and serum NO levels were assessed in 56 healthy adults aged 29 to 84 years, before and after 12 weeks of fenofibrate (250 mg/d; n = 30) or placebo (n = 26). Appropriate dietary suggestions for hypertriglyceridemia were made for all patients. This study was randomized, double-blind and placebo-controlled in design. RESULTS Total cholesterol, low-density lipoprotein (LDL), very low-density lipoprotein (VLDL) and triglyceride levels significantly decreased; high-density lipoprotein (HDL) and NO levels significantly increased after 12 weeks of fenofibrate therapy. We observed a statistically significant correlation between the increase in serum NO levels and decrease in serum triglyceride levels (r = -0.42, p = 0.02) in the fenofibrate group. CONCLUSIONS The positive effect of short-term fenofibrate treatments on vascular endothelial functions in patients with hypertriglyceridemia has been demonstrated by increasing the serum NO levels. Agents such as fenofibrate targeting PPARα-associated signaling pathways show promise as an alternative treatment of vascular dysfunction related to advanced age and hyperlipidemia.
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Transcriptional Regulation Contributes to Prioritized Detoxification of Hydrogen Peroxide over Nitric Oxide.
Adolfsen, KJ, Chou, WK, Brynildsen, MP
Journal of bacteriology. 2019;(14)
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Abstract
Hydrogen peroxide (H2O2) and nitric oxide (NO·) are toxic metabolites that immune cells use to attack pathogens. These antimicrobials can be present at the same time in phagosomes, and it remains unclear how bacteria deal with these insults when simultaneously present. Here, using Escherichia coli, we observed that simultaneous exposure to H2O2 and NO· leads to prioritized detoxification, where enzymatic removal of NO· is impeded until H2O2 has been eliminated. This phenomenon is reminiscent of carbon catabolite repression (CCR), where preferred carbon sources are catabolized prior to less desirable substrates; however, H2O2 and NO· are toxic, growth-inhibitory compounds rather than growth-promoting nutrients. To understand how NO· detoxification is delayed by H2O2 whereas H2O2 detoxification proceeds unimpeded, we confirmed that the effect depended on Hmp, which is the main NO· detoxification enzyme, and used an approach that integrated computational modeling and experimentation to delineate and test potential mechanisms. Plausible interactions included H2O2-dependent inhibition of hmp transcription and translation, direct inhibition of Hmp catalysis, and competition for reducing equivalents between Hmp and H2O2-degrading enzymes. Experiments illustrated that Hmp catalysis and NAD(P)H supply were not impaired by H2O2, whereas hmp transcription and translation were diminished. A dependence of this phenomenon on transcriptional regulation parallels CCR, and we found it to involve the transcriptional repressor NsrR. Collectively, these data suggest that bacterial regulation of growth inhibitor detoxification has similarities to the regulation of growth substrate consumption, which could have ramifications for infectious disease, bioremediation, and biocatalysis from inhibitor-containing feedstocks.IMPORTANCE Bacteria can be exposed to H2O2 and NO· concurrently within phagosomes. In such multistress situations, bacteria could have evolved to simultaneously degrade both toxic metabolites or preferentially detoxify one over the other. Here, we found that simultaneous exposure to H2O2 and NO· leads to prioritized detoxification, where detoxification of NO· is hampered until H2O2 has been eliminated. This phenomenon resembles CCR, where bacteria consume one substrate over others in carbon source mixtures. Further experimentation revealed a central role for transcriptional regulation in the prioritization of H2O2 over NO·, which is also important to CCR. This study suggests that regulatory scenarios observed in bacterial consumption of growth-promoting compound mixtures can be conserved in bacterial detoxification of toxic metabolite mixtures.
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Effects of acute nitric oxide precursor intake on peripheral and central fatigue during knee extensions in healthy men.
Le Roux-Mallouf, T, Laurent, J, Besset, D, Marillier, M, Larribaut, J, Belaidi, E, Corne, C, Doutreleau, S, Verges, S
Experimental physiology. 2019;(7):1100-1114
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Abstract
NEW FINDINGS What is the central question of this study? What is the effect of acute NO precursor intake on vascular function, muscle and cerebral oxygenation and peripheral and central neuromuscular fatigue during knee-extension exercise? What is the main finding and its importance? Acute NO precursor ingestion increases the plasma concentrations of NO precursors (nitrate, arginine and citrulline) and enhances post-ischaemic vasodilatation, but has no significant effect on muscle and cerebral oxygenation, peripheral and central mechanisms of neuromuscular fatigue and, consequently, does not improve exercise performance. ABSTRACT Nitric oxide (NO) plays an important role in matching blood flow to oxygen demand in the brain and contracting muscles during exercise. Previous studies have shown that increasing NO bioavailability can improve muscle function. The aim of this study was to assess the effect of acute NO precursor intake on muscle and cerebral oxygenation and on peripheral and central neuromuscular fatigue during exercise. In four experimental sessions, 15 healthy men performed a thigh ischaemia-reperfusion test followed by submaximal isometric knee extensions (5 s on-4 s off; 45% of maximal voluntary contraction) until task failure. In each session, subjects drank a nitrate-rich beetroot juice containing 520 mg nitrate (N), N and citrulline (6 g; N+C), N and arginine (6 g; N+A) or a placebo (PLA). Prefrontal cortex and quadriceps near-infrared spectroscopy parameters were monitored continuously. Transcranial magnetic stimulation and femoral nerve electrical stimulation were used to assess central and peripheral determinants of fatigue. The post-ischaemic increase in thigh blood total haemoglobin concentration was larger in N (10.1 ± 3.7 mmol) and N+C (10.9 ± 3.3 mmol) compared with PLA (8.2 ± 2.7 mmol; P < 0.05). Nitric oxide precursors had no significant effect on muscle and cerebral oxygenation or on peripheral and central mechanisms of neuromuscular fatigue during exercise. The total number of knee extensions did not differ between sessions (N, 71.9 ± 33.2; N+A, 73.3 ± 39.4; N+C, 74.6 ± 34.0; PLA, 71.8 ± 39.9; P > 0.05). In contrast to the post-ischaemic hyperaemic response, NO bioavailability in healthy subjects might not be the limiting factor for tissue perfusion and oxygenation during submaximal knee extensions to task failure.
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TNFα stimulates NO release in EA.hy926 cells by activating the CaMKKβ-AMPK-eNOS pathway.
Dymkowska, D, Drabarek, B, Michalik, A, Nowak, N, Zabłocki, K
The international journal of biochemistry & cell biology. 2019;:57-67
Abstract
Previously we showed that a mild stimulation of EA.hy926 cells with tumour necrosis factor alpha (TNFα) activated mitochondrial biogenesis, probably as a mechanism preventing cell death. This was accompanied by an increased phosphorylation of eNOS and elevation of NO release. The aim of the present study was to explain the biochemical basis of this effect. Our results indicate that eNOS is the only enzyme catalysing NO generation in EA.hy926 cells, and TNFα stimulates its activity by activating AMP-activated protein kinase (AMPK). Inhibition of AMPK with Compound C prevents the TNFα-induced activatory phosphorylation of endothelial nitric oxide synthase (eNOS) at Ser1177 and reduces the NO release. AMPK is activated by phosphorylation catalysed by liver kinase B1 (LKB1) and calcium/calmodulin-dependent protein kinase kinase beta (CaMKKβ), which are phosphorylated and thereby activated in the presence of TNFα. Moreover, CaMKKβ catalyses an activatory phosphorylation of sirtuin 1, which could deacetylate and activate eNOS both directly and indirectly by an elevating the LKB1 activity. TNFα hardly increases the nuclear fraction of sirtuin 1, thus its major activity is probably attributed to the cytosolic pool. This is in line with the elevated activity of eNOS. We conclude that the increased AMPK-dependent phosphorylation of eNOS at least partially explains the stimulation of NO generation by TNFα in EA.hy926 cells.
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Ethylene-mediated nitric oxide depletion pre-adapts plants to hypoxia stress.
Hartman, S, Liu, Z, van Veen, H, Vicente, J, Reinen, E, Martopawiro, S, Zhang, H, van Dongen, N, Bosman, F, Bassel, GW, et al
Nature communications. 2019;(1):4020
Abstract
Timely perception of adverse environmental changes is critical for survival. Dynamic changes in gases are important cues for plants to sense environmental perturbations, such as submergence. In Arabidopsis thaliana, changes in oxygen and nitric oxide (NO) control the stability of ERFVII transcription factors. ERFVII proteolysis is regulated by the N-degron pathway and mediates adaptation to flooding-induced hypoxia. However, how plants detect and transduce early submergence signals remains elusive. Here we show that plants can rapidly detect submergence through passive ethylene entrapment and use this signal to pre-adapt to impending hypoxia. Ethylene can enhance ERFVII stability prior to hypoxia by increasing the NO-scavenger PHYTOGLOBIN1. This ethylene-mediated NO depletion and consequent ERFVII accumulation pre-adapts plants to survive subsequent hypoxia. Our results reveal the biological link between three gaseous signals for the regulation of flooding survival and identifies key regulatory targets for early stress perception that could be pivotal for developing flood-tolerant crops.
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Sulfur: the heart of nitric oxide-dependent redox signalling.
Umbreen, S, Lubega, J, Loake, GJ
Journal of experimental botany. 2019;(16):4279-4286
Abstract
Nitric oxide (NO), more benign than its more reactive and damaging related molecules, reactive oxygen species (ROS), is perfectly suited for duties as a redox signalling molecule. A key route for NO bioactivity is through S-nitrosation, the addition of an NO moiety to a protein Cys thiol (-SH). This redox-based, post-translational modification (PTM) can modify protein function analogous to more well established PTMs such as phosphorylation, for example by modulating enzyme activity, localization, or protein-protein interactions. At the heart of the underpinning chemistry associated with this PTM is sulfur. The emerging evidence suggests that S-nitrosation is integral to a myriad of plant biological processes embedded in both development and environmental relations. However, a role for S-nitrosation is perhaps most well established in plant-pathogen interactions.