Publications by authors named "Bożena Szal"

22 Publications

  • Page 1 of 1

Spatiotemporal auxin distribution in Arabidopsis tissues is regulated by anabolic and catabolic reactions under long-term ammonium stress.

BMC Plant Biol 2021 Dec 18;21(1):602. Epub 2021 Dec 18.

Institute of Plant Bioenergetics, Faculty of Biology, University of Warsaw, I. Miecznikowa 01, 02-096, Warsaw, Poland.

Background: The plant hormone auxin is a major coordinator of plant growth and development in response to diverse environmental signals, including nutritional conditions. Sole ammonium (NH) nutrition is one of the unique growth-suppressing conditions for plants. Therefore, the quest to understand NH-mediated developmental defects led us to analyze auxin metabolism.

Results: Indole-3-acetic acid (IAA), the most predominant natural auxin, accumulates in the leaves and roots of mature Arabidopsis thaliana plants grown on NH, but not in the root tips. We found changes at the expressional level in reactions leading to IAA biosynthesis and deactivation in different tissues. Finally, NH nutrition would facilitate the formation of inactive oxidized IAA as the final product.

Conclusions: NH-mediated accelerated auxin turnover rates implicate transient and local IAA peaks. A noticeable auxin pattern in tissues correlates with the developmental adaptations of the short and highly branched root system of NH-grown plants. Therefore, the spatiotemporal distribution of auxin might be a root-shaping signal specific to adjust to NH-stress conditions.
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http://dx.doi.org/10.1186/s12870-021-03385-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8684078PMC
December 2021

Markers for Mitochondrial ROS Status.

Methods Mol Biol 2022 ;2363:199-213

Department of Plant Bioenergetics, Institute of Experimental Plant Biology and Biotechnology, Faculty of Biology, University of Warsaw, Warsaw, Poland.

Mitochondria actively participate in oxygenic metabolism and are one of the major sources of reactive oxygen species (ROS) production in plant cells. However, instead of measuring ROS concentrations in organelles it is more worthwhile to observe active ROS generation or downstream oxidation products, because the steady state level of ROS is easily buffered. Here, we describe how to measure the in vitro production of superoxide anion radicals (O·) by mitochondria and the release of O· into the cytosol. A method to determine glutathione, which is the most abundant mitochondrial low-mass antioxidant, is presented since changes in the redox state of glutathione can be indicative of the oxidative action of ROS. The identification of oxidative damage to mitochondrial components is the ultimate symptom that ROS homeostasis is not under control. We present how to determine the extent of oxidation of membrane lipids and the carbonylation of mitochondrial proteins. In summary, oxidative stress symptoms have to be analyzed at different levels, including ROS production, scavenging capacity, and signs of destruction, which only together can be considered markers of mitochondrial ROS status.
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http://dx.doi.org/10.1007/978-1-0716-1653-6_15DOI Listing
January 2022

Quantification of Methylglyoxal Levels in Cowpea Leaves in Response to Cowpea Aphid Infestation.

Bio Protoc 2020 Oct 20;10(20):e3795. Epub 2020 Oct 20.

Graduate program in Biochemistry and Molecular Biology, University of California, Riverside, USA.

Aphids are a serious pest of crops across the world. Aphids feed by inserting their flexible hypodermal needlelike mouthparts, or stylets, into their host plant tissues. They navigate their way to the phloem where they feed on its sap causing little mechanical damage to the plant. Additionally, while feeding, aphids secrete proteinaceous effectors in their saliva to alter plant metabolism and disrupt plant defenses to gain an advantage over the plant. Even with these arsenals to overcome plant responses, plants have evolved ways to detect and counter with defense responses to curtail aphid infestation. One of such response of cowpea to cowpea aphid infestation, is accumulation of the metabolite methylglyoxal. Methylglyoxal is an α,β-dicarbonyl ketoaldehyde that is toxic at high concentrations. Methylglyoxal levels increase modestly after exposure to a number of different abiotic and biotic stresses and has been shown to act as an emerging defense signaling molecule at low levels. Here we describe a protocol to measure methylglyoxal in cowpea leaves after cowpea aphid infestation, by utilizing a perchloric acid extraction process. The extracted supernatant was neutralized with potassium carbonate, and methylglyoxal was quantified through its reaction with N-acetyl-L-cysteine to form N-α-acetyl-S-(1-hydroxy-2-oxo-prop-1-yl)cysteine, a product that is quantified spectrophotometrically.
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http://dx.doi.org/10.21769/BioProtoc.3795DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7842737PMC
October 2020

Mitochondrial NAD(P)H oxidation pathways and nitrate/ammonium redox balancing in plants.

Mitochondrion 2020 07 30;53:158-165. Epub 2020 May 30.

University of Warsaw, Faculty of Biology, Institute of Experimental Plant Biology and Biotechnology, Ilii Miecznikowa 1, 02-096 Warsaw, Poland.

Plant mitochondrial oxidative phosphorylation is characterised by alternative electron transport pathways with different energetic efficiencies, allowing turnover of cellular redox compounds like NAD(P)H. These electron transport chain pathways are profoundly affected by soil nitrogen availability, most commonly as oxidized nitrate (NO) and/or reduced ammonium (NH). The bioenergetic strategies involved in assimilating different N sources can alter redox homeostasis and antioxidant systems in different cellular compartments, including the mitochondria and the cell wall. Conversely, changes in mitochondrial redox systems can affect plant responses to N. This review explores the integration between N assimilation, mitochondrial redox metabolism, and apoplast metabolism.
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http://dx.doi.org/10.1016/j.mito.2020.05.010DOI Listing
July 2020

Efficient Photosynthetic Functioning of Through Electron Dissipation in Chloroplasts and Electron Export to Mitochondria Under Ammonium Nutrition.

Front Plant Sci 2020 26;11:103. Epub 2020 Feb 26.

Institute of Experimental Plant Biology and Biotechnology, Faculty of Biology, University of Warsaw, Warsaw, Poland.

An improvement in photosynthetic rate promotes the growth of crop plants. The sink-regulation of photosynthesis is crucial in optimizing nitrogen fixation and integrating it with carbon balance. Studies on these processes are essential in understanding growth inhibition in plants with ammonium ( ) syndrome. Hence, we sought to investigate the effects of using nitrogen sources with different states of reduction (during assimilation of versus ) on the photosynthetic performance of . Our results demonstrated that photosynthetic functioning during long-term nutrition was not disturbed and that no indication of photoinhibition of PSII was detected, revealing the robustness of the photosynthetic apparatus during stressful conditions. Based on our findings, we propose multiple strategies to sustain photosynthetic activity during limited reductant utilization for assimilation. One mechanism to prevent chloroplast electron transport chain overreduction during nutrition is for cyclic electron flow together with plastid terminal oxidase activity. Moreover, redox state in chloroplasts was optimized by a dedicated type II NAD(P)H dehydrogenase. In order to reduce the amount of energy that reaches the photosynthetic reaction centers and to facilitate photosynthetic protection during nutrition, non-photochemical quenching (NPQ) and ample xanthophyll cycle pigments efficiently dissipate excess excitation. Additionally, high redox load may be dissipated in other metabolic reactions outside of chloroplasts due to the direct export of nucleotides through the malate/oxaloacetate valve. Mitochondrial alternative pathways can downstream support the overreduction of chloroplasts. This mechanism correlated with the improved growth of with the overexpression of the alternative oxidase 1a (AOX1a) during nutrition. Most remarkably, our findings demonstrated the capacity of chloroplasts to tolerate syndrome instead of providing redox poise to the cells.
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http://dx.doi.org/10.3389/fpls.2020.00103DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7054346PMC
February 2020

Nitrogen Source Dependent Changes in Central Sugar Metabolism Maintain Cell Wall Assembly in Mitochondrial Complex I-Defective and Secondarily Affect Programmed Cell Death.

Int J Mol Sci 2018 Jul 28;19(8). Epub 2018 Jul 28.

Institute of Experimental Plant Biology and Biotechnology, Faculty of Biology, University of Warsaw, I. Miecznikowa 1, 02-096 Warsaw, Poland.

For optimal plant growth, carbon and nitrogen availability needs to be tightly coordinated. Mitochondrial perturbations related to a defect in complex I in the () mutant, carrying a point mutation in the 8-kD Fe-S subunit of NDUFS4 protein, alter aspects of fundamental carbon metabolism, which is manifested as stunted growth. During nitrate nutrition, plants showed a dominant sugar flux toward nitrogen assimilation and energy production, whereas cellulose integration in the cell wall was restricted. However, when cultured on NH₄⁺ as the sole nitrogen source, which typically induces developmental disorders in plants (i.e., the ammonium toxicity syndrome), showed improved growth as compared to NO₃ nourishing. Higher energy availability in plants was correlated with restored cell wall assembly during NH₄⁺ growth. To determine the relationship between mitochondrial complex I disassembly and cell wall-related processes, aspects of cell wall integrity and sugar and reactive oxygen species signaling were analyzed in plants. The responses of plants to NH₄⁺ treatment were consistent with the inhibition of a form of programmed cell death. Resistance of plants to NH₄⁺ toxicity coincided with an absence of necrotic lesion in plant leaves.
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http://dx.doi.org/10.3390/ijms19082206DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6121878PMC
July 2018

Enhanced Formation of Methylglyoxal-Derived Advanced Glycation End Products in Under Ammonium Nutrition.

Front Plant Sci 2018 24;9:667. Epub 2018 May 24.

Institute of Experimental Plant Biology and Biotechnology, Faculty of Biology, University of Warsaw, Warsaw, Poland.

Nitrate (NO) and ammonium (NH) are prevalent nitrogen (N) sources for plants. Although NH should be the preferred form of N from the energetic point of view, ammonium nutrition often exhibits adverse effects on plant physiological functions and induces an important growth-limiting stress referred as ammonium syndrome. The effective incorporation of NH into amino acid structures requires high activity of the mitochondrial tricarboxylic acid cycle and the glycolytic pathway. An unavoidable consequence of glycolytic metabolism is the production of methylglyoxal (MG), which is very toxic and inhibits cell growth in all types of organisms. Here, we aimed to investigate MG metabolism in plants grown on NH as a sole N source. We found that changes in activities of glycolytic enzymes enhanced MG production and that markedly elevated MG levels superseded the detoxification capability of the glyoxalase pathway. Consequently, the excessive accumulation of MG was directly involved in the induction of dicarbonyl stress by introducing MG-derived advanced glycation end products (MAGEs) to proteins. The severe damage to proteins was not within the repair capacity of proteolytic enzymes. Collectively, our results suggest the impact of MG (mediated by MAGEs formation in proteins) in the contribution to NH toxicity symptoms in .
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http://dx.doi.org/10.3389/fpls.2018.00667DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5976750PMC
May 2018

Suppression of External NADPH Dehydrogenase-NDB1 in Confers Improved Tolerance to Ammonium Toxicity via Efficient Glutathione/Redox Metabolism.

Int J Mol Sci 2018 May 9;19(5). Epub 2018 May 9.

Institute of Experimental Plant Biology and Biotechnology, Faculty of Biology, University of Warsaw, I. Miecznikowa 1, 02-096 Warsaw, Poland.

Environmental stresses, including ammonium (NH₄⁺) nourishment, can damage key mitochondrial components through the production of surplus reactive oxygen species (ROS) in the mitochondrial electron transport chain. However, alternative electron pathways are significant for efficient reductant dissipation in mitochondria during ammonium nutrition. The aim of this study was to define the role of external NADPH-dehydrogenase (NDB1) during oxidative metabolism of NH₄⁺-fed plants. Most plant species grown with NH₄⁺ as the sole nitrogen source experience a condition known as “ammonium toxicity syndrome”. Surprisingly, transgenic plants suppressing were more resistant to NH₄⁺ treatment. The knock-down line was characterized by milder oxidative stress symptoms in plant tissues when supplied with NH₄⁺. Mitochondrial ROS accumulation, in particular, was attenuated in the knock-down plants during NH₄⁺ treatment. Enhanced antioxidant defense, primarily concerning the glutathione pool, may prevent ROS accumulation in NH₄⁺-grown -suppressing plants. We found that induction of glutathione peroxidase-like enzymes and peroxiredoxins in the -surpressing line contributed to lower ammonium-toxicity stress. The major conclusion of this study was that NDB1 suppression in plants confers tolerance to changes in redox homeostasis that occur in response to prolonged ammonium nutrition, causing cross tolerance among plants.
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http://dx.doi.org/10.3390/ijms19051412DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5983774PMC
May 2018

Extra-Cellular But Extra-Ordinarily Important for Cells: Apoplastic Reactive Oxygen Species Metabolism.

Front Plant Sci 2017 22;8:1353. Epub 2017 Aug 22.

Institute of Experimental Plant Biology and Biotechnology, Faculty of Biology, University of WarsawWarsaw, Poland.

Reactive oxygen species (ROS), by their very nature, are highly reactive, and it is no surprise that they can cause damage to organic molecules. In cells, ROS are produced as byproducts of many metabolic reactions, but plants are prepared for this ROS output. Even though extracellular ROS generation constitutes only a minor part of a cell's total ROS level, this fraction is of extraordinary importance. In an active apoplastic ROS burst, it is mainly the respiratory burst oxidases and peroxidases that are engaged, and defects of these enzymes can affect plant development and stress responses. It must be highlighted that there are also other less well-known enzymatic or non-enzymatic ROS sources. There is a need for ROS detoxification in the apoplast, and almost all cellular antioxidants are present in this space, but the activity of antioxidant enzymes and the concentration of low-mass antioxidants is very low. The low antioxidant efficiency in the apoplast allows ROS to accumulate easily, which is a condition for ROS signaling. Therefore, the apoplastic ROS/antioxidant homeostasis is actively engaged in the reception and reaction to many biotic and abiotic stresses.
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http://dx.doi.org/10.3389/fpls.2017.01353DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5572287PMC
August 2017

Altered Cell Wall Plasticity Can Restrict Plant Growth under Ammonium Nutrition.

Front Plant Sci 2017 10;8:1344. Epub 2017 Aug 10.

Institute of Experimental Plant Biology and Biotechnology, Faculty of Biology, University of WarsawWarsaw, Poland.

Plants mainly utilize inorganic forms of nitrogen (N), such as nitrate (NO) and ammonium (NH). However, the composition of the N source is important, because excess of NH promotes morphological disorders. Plants cultured on NH as the sole N source exhibit serious growth inhibition, commonly referred to as "ammonium toxicity syndrome." NH-mediated suppression of growth may be attributable to both repression of cell elongation and reduction of cell division. The precondition for cell enlargement is the expansion of the cell wall, which requires the loosening of the cell wall polymers. Therefore, to understand how NH nutrition may trigger growth retardation in plants, properties of their cell walls were analyzed. We found that using NH as the sole N source has smaller cells with relatively thicker cell walls. Moreover, cellulose, which is the main load-bearing polysaccharide revealed a denser assembly of microfibrils. Consequently, the leaf blade tissue showed elevated tensile strength and indicated higher cell wall stiffness. These changes might be related to changes in polysaccharide and ion content of cell walls. Further, NH toxicity was associated with altered activities of cell wall modifying proteins. The lower activity and/or expression of pectin hydrolyzing enzymes and expansins might limit cell wall expansion. Additionally, the higher activity of cell wall peroxidases can lead to higher cross-linking of cell wall polymers. Overall, the NH-mediated inhibition of growth is related to a more rigid cell wall structure, which limits expansion of cells. The changes in cell wall composition were also indicated by decreased expression of , a receptor-like kinase involved in the control of cell wall extension.
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http://dx.doi.org/10.3389/fpls.2017.01344DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5554365PMC
August 2017

[Alternative oxidase - never ending story].

Postepy Biochem 2016;62(2):138-148

Department of Plant Anatomy and Cytology, Institute of Experimental Plant Biology and Biotechnology, Faculty of Biology, Warsaw University, 1 Miecznikowa St., 02-096 Warsaw, Poland.

Investigations of plant cyanide resistant respiration lead to the discovery in mitochondrial respiratory chain of the second terminal oxidase, alternative oxidase (AOX). AOX transfers electrons from reduced ubiquinone to oxygen omitting two coupling places thus lowering energetic efficiency of respiration. The presence of AOX was shown in all plants and also in some fungi, mollusca and protista. In termogenic plants the activity of AOX is connected with heat production. In other organisms AOX activity is important for maintaining metabolic homeostasis (carbon metabolism, cell redox state and energy demand) and ROS homeostasis. In this article structure of plant AOX protein and the regulation on molecular levels was described. Possible role of AOX as stress marker was pointed and the possibility of using AOX in human gene therapy was discussed.
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December 2017

Short-term ammonium supply induces cellular defence to prevent oxidative stress in Arabidopsis leaves.

Physiol Plant 2017 May 14;160(1):65-83. Epub 2017 Feb 14.

Institute of Experimental Plant Biology and Biotechnology, Faculty of Biology, University of Warsaw, Warsaw, 02-096, Poland.

Plants can assimilate nitrogen from soil pools of both ammonium and nitrate, and the relative levels of these two nitrogen sources are highly variable in soil. Long-term ammonium nutrition is known to cause damage to Arabidopsis that has been linked to mitochondrial oxidative stress. Using hydroponic cultures, we analysed the consequences of rapid shifts between nitrate and ammonium nutrition. This did not induce growth retardation, showing that Arabidopsis can compensate for the changes in redox metabolism associated with the variations in nitrogen redox status. During the first 3 h of ammonium treatment, we observed distinct transient shifts in reactive oxygen species (ROS), low-mass antioxidants, ROS-scavenging enzymes, and mitochondrial alternative electron transport pathways, indicating rapid but temporally separated changes in chloroplastic, mitochondrial and cytosolic ROS metabolism. The fast induction of antioxidant defences significantly lowered intracellular H O levels, and thus protected Arabidopsis leaves from oxidative stress. On the other hand elevated extracellular ROS production in response to ammonium supply may be involved in signalling. The response pattern displays an intricate plasticity of Arabidopsis redox metabolism to minimise stress in responses to nutrient changes.
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http://dx.doi.org/10.1111/ppl.12538DOI Listing
May 2017

Dissecting the Metabolic Role of Mitochondria during Developmental Leaf Senescence.

Plant Physiol 2016 12 15;172(4):2132-2153. Epub 2016 Oct 15.

Department of Plant Physiology, Umeå Plant Science Centre, Umeå University, S-90187 Umea, Sweden (D.C., S.R.L., B.B., A.Z., D.L., P.G., O.K.);

The functions of mitochondria during leaf senescence, a type of programmed cell death aimed at the massive retrieval of nutrients from the senescing organ to the rest of the plant, remain elusive. Here, combining experimental and analytical approaches, we showed that mitochondrial integrity in Arabidopsis (Arabidopsis thaliana) is conserved until the latest stages of leaf senescence, while their number drops by 30%. Adenylate phosphorylation state assays and mitochondrial respiratory measurements indicated that the leaf energy status also is maintained during this time period. Furthermore, after establishing a curated list of genes coding for products targeted to mitochondria, we analyzed in isolation their transcript profiles, focusing on several key mitochondrial functions, such as the tricarboxylic acid cycle, mitochondrial electron transfer chain, iron-sulfur cluster biosynthesis, transporters, as well as catabolic pathways. In tandem with a metabolomic approach, our data indicated that mitochondrial metabolism was reorganized to support the selective catabolism of both amino acids and fatty acids. Such adjustments would ensure the replenishment of α-ketoglutarate and glutamate, which provide the carbon backbones for nitrogen remobilization. Glutamate, being the substrate of the strongly up-regulated cytosolic glutamine synthase, is likely to become a metabolically limiting factor in the latest stages of developmental leaf senescence. Finally, an evolutionary age analysis revealed that, while branched-chain amino acid and proline catabolism are very old mitochondrial functions particularly enriched at the latest stages of leaf senescence, auxin metabolism appears to be rather newly acquired. In summation, our work shows that, during developmental leaf senescence, mitochondria orchestrate catabolic processes by becoming increasingly central energy and metabolic hubs.
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http://dx.doi.org/10.1104/pp.16.01463DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5129728PMC
December 2016

In comparison with nitrate nutrition, ammonium nutrition increases growth of the frostbite1 Arabidopsis mutant.

Plant Cell Environ 2015 Jan 13;38(1):224-37. Epub 2014 Aug 13.

Institute of Experimental Plant Biology and Biotechnology, Faculty of Biology, University of Warsaw, I. Miecznikowa 1, 02-096, Warsaw, Poland.

Ammonium nutrition inhibits the growth of many plant species, including Arabidopsis thaliana. The toxicity of ammonium is associated with changes in the cellular redox state. The cellular oxidant/antioxidant balance is controlled by mitochondrial electron transport chain. In this study, we analysed the redox metabolism of frostbite1 (fro1) plants, which lack mitochondrial respiratory chain complex I. Surprisingly, the growth of fro1 plants increased under ammonium nutrition. Ammonium nutrition increased the reduction level of pyridine nucleotides in the leaves of wild-type plants, but not in the leaves of fro1 mutant plants. The observed higher activities of type II NADH dehydrogenases and cytochrome c oxidase in the mitochondrial electron transport chain may improve the energy metabolism of fro1 plants grown on ammonium. Additionally, the observed changes in reactive oxygen species (ROS) metabolism in the apoplast may be important for determining the growth of fro1 under ammonium nutrition. Moreover, bioinformatic analyses showed that the gene expression changes in fro1 plants significantly overlap with the changes previously observed in plants with a modified apoplastic pH. Overall, the results suggest a pronounced connection between the mitochondrial redox system and the apoplastic pH and ROS levels, which may modify cell wall plasticity and influence growth.
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http://dx.doi.org/10.1111/pce.12404DOI Listing
January 2015

Long-term ammonium nutrition of Arabidopsis increases the extrachloroplastic NAD(P)H/NAD(P)(+) ratio and mitochondrial reactive oxygen species level in leaves but does not impair photosynthetic capacity.

Plant Cell Environ 2013 Nov 6;36(11):2034-45. Epub 2013 May 6.

Faculty of Biology, Institute of Experimental Plant Biology and Biotechnology, University of Warsaw, I. Miecznikowa 1, 02-096, Warsaw, Poland.

Ammonium nutrition has been suggested to be associated with alterations in the oxidation-reduction state of leaf cells. Herein, we show that ammonium nutrition in Arabidopsis thaliana increases leaf NAD(P)H/NAD(P)(+) ratio, reactive oxygen species content and accumulation of biomolecules oxidized by free radicals. We used the method of rapid fractionation of protoplasts to analyse which cellular compartments were over-reduced under ammonium supply and revealed that observed changes in NAD(P)H/NAD(P)(+) ratio involved only the extrachloroplastic fraction. We also showed that ammonium nutrition changes mitochondrial electron transport chain activity, increasing mitochondrial reactive oxygen species production. Our results indicate that the functional impairment associated with ammonium nutrition is mainly associated with redox reactions outside the chloroplast.
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http://dx.doi.org/10.1111/pce.12113DOI Listing
November 2013

The role of mitochondria in leaf nitrogen metabolism.

Plant Cell Environ 2012 Oct 6;35(10):1756-68. Epub 2012 Jul 6.

Institute of Experimental Plant Biology and Biotechnology, Faculty of Biology, University of Warsaw, Warsaw, Poland.

For optimal plant growth and development, cellular nitrogen (N) metabolism must be closely coordinated with other metabolic pathways, and mitochondria are thought to play a central role in this process. Recent studies using genetically modified plants have provided insight into the role of mitochondria in N metabolism. Mitochondrial metabolism is linked with N assimilation by amino acid, carbon (C) and redox metabolism. Mitochondria are not only an important source of C skeletons for N incorporation, they also produce other necessary metabolites and energy used in N remobilization processes. Nitric oxide of mitochondrial origin regulates respiration and influences primary N metabolism. Here, we discuss the changes in mitochondrial metabolism during ammonium or nitrate nutrition and under low N conditions. We also describe the involvement of mitochondria in the redistribution of N during senescence. The aim of this review was to demonstrate the role of mitochondria as an integration point of N cellular metabolism.
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http://dx.doi.org/10.1111/j.1365-3040.2012.02559.xDOI Listing
October 2012

Oxidation-reduction and reactive oxygen species homeostasis in mutant plants with respiratory chain complex I dysfunction.

Plant Cell Environ 2012 Feb 21;35(2):296-307. Epub 2011 Apr 21.

Department of Plant Bioenergetics, Institute of Experimental Plant Biology and Biotechnology, University of Warsaw, Miecznikowa 1, 02-096 Warsaw, Poland.

Mutations in a mitochondrial or nuclear gene encoding respiratory chain complex I subunits lead to decreased or a total absence of complex I activity. Plant mutants with altered or lost complex I activity adapt their respiratory metabolism by inducing alternative pathways of the respiratory chain and changing energy metabolism. Apparently, complex I is a crucial component of the oxidation-reduction (redox) regulatory system in photosynthetic cells, and alternative NAD(P)H dehydrogenases of the mitochondrial electron transport chain (mtETC) cannot fully compensate for its impairment. In most cases, dysfunction of complex I is associated with lowered or unchanged hydrogen peroxide (H(2)O(2)) concentrations, but increased superoxide (O(2)(-)) levels. Higher production of reactive oxygen species (ROS) by mitochondria in the mosaic (MSC16) cucumber mutant may be related to retrograde signalling. Different effects of complex I dysfunction on H(2)O(2) and O(2)(-) levels in described mutants might result from diverse regulation of processes involved in H(2)O(2) and O(2)(-) production. Often, dysfunction of complex I did not lead to oxidative stress, but increased the capacity of the antioxidative system and enhanced stress tolerance. The new cellular homeostasis in mutants with dysfunction of complex I allows growth and development, reflecting the plasticity of plant metabolism.
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http://dx.doi.org/10.1111/j.1365-3040.2011.02314.xDOI Listing
February 2012

Influence of mitochondrial genome rearrangement on cucumber leaf carbon and nitrogen metabolism.

Planta 2010 Nov 10;232(6):1371-82. Epub 2010 Sep 10.

Institute of Experimental Plant Biology, University of Warsaw, Miecznikowa 1, 02-096, Warsaw, Poland.

The MSC16 cucumber (Cucumis sativus L.) mitochondrial mutant was used to study the effect of mitochondrial dysfunction and disturbed subcellular redox state on leaf day/night carbon and nitrogen metabolism. We have shown that the mitochondrial dysfunction in MSC16 plants had no effect on photosynthetic CO(2) assimilation, but the concentration of soluble carbohydrates and starch was higher in leaves of MSC16 plants. Impaired mitochondrial respiratory chain activity was associated with the perturbation of mitochondrial TCA cycle manifested, e.g., by lowered decarboxylation rate. Mitochondrial dysfunction in MSC16 plants had different influence on leaf cell metabolism under dark or light conditions. In the dark, when the main mitochondrial function is the energy production, the altered activity of TCA cycle in mutated plants was connected with the accumulation of pyruvate and TCA cycle intermediates (citrate and 2-OG). In the light, when TCA activity is needed for synthesis of carbon skeletons required as the acceptors for NH(4) (+) assimilation, the concentration of pyruvate and TCA intermediates was tightly coupled with nitrate metabolism. Enhanced incorporation of ammonium group into amino acids structures in mutated plants has resulted in decreased concentration of organic acids and accumulation of Glu.
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http://dx.doi.org/10.1007/s00425-010-1261-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2957574PMC
November 2010

Chilling stress and mitochondrial genome rearrangement in the MSC16 cucumber mutant affect the alternative oxidase and antioxidant defense system to a similar extent.

Physiol Plant 2009 Dec 21;137(4):435-45. Epub 2009 May 21.

Faculty of Biology, Institute of Experimental Plant Biology, University of Warsaw, Miecznikowa 1, 02-096 Warsaw, Poland.

The mosaic MSC16 cucumber (Cucumis sativus L.) mutant, which houses a rearranged mitochondrial genome, has altered respiratory chain activity, with a dysfunctional Complex I, increased external NADH dehydrogenases (ND(ex)) activity, and a higher alternative oxidase (AOX) capacity and AOX protein level. In the present study, changes in oxidative defense metabolism resulting from the respiratory chain dysfunction in the MSC16 mutant were compared with those induced by chilling. Chilling increased the enzymatic and non-enzymatic antioxidant defense systems in the wild-type (WT) but not in MSC16, which displays elevated antioxidant defenses as a result of the mitochondrial mutation. The high AOX capacity and protein level in MSC16 were unchanged as a result of chilling, whereas chilling increased these parameters in WT leaves. In mitochondria isolated from WT plants, superoxide was produced to a similar extent in the matrix and the intermembrane space, but in MSC16 mitochondria superoxide was produced largely within the intermembrane space. Mitochondria isolated from both genotypes after chilling showed increased superoxide production within the intermembrane space. Cytochemical detection revealed an increased abundance of H2O2 in the mitochondrial membrane in mesophyll cells of MSC16 leaves. The mitochondrial mutation also resulted in changes in the antioxidative defense system, including AOX, which were similar to those observed following chilling. The results presented here support the hypothesis that AOX is an effective marker of the cellular reprogramming resulting from stress. Moreover, we propose a role for reactive oxygen species (ROS) generated within the mitochondria in signal transduction.
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http://dx.doi.org/10.1111/j.1399-3054.2009.01255.xDOI Listing
December 2009

Effect of mitochondrial genome rearrangement on respiratory activity, photosynthesis, photorespiration and energy status of MSC16 cucumber (Cucumis sativus) mutant.

Physiol Plant 2007 Dec;131(4):527-41

Institute of Experimental Plant Biology, University of Warsaw, Miecznikowa 1, PL-02-096 Warsaw, Poland.

The effects of changes in mitochondrial DNA in cucumber (Cucumis sativus L.) mosaic mutant (MSC16) on respiration, photosynthesis and photorespiration were analyzed under non-stressed conditions. Decreased respiratory capacity of complex I in MSC16 mitochondria was indicated by lower respiration rates of intact mitochondria with malate and by rotenone-inhibited NADH or malate oxidation in the presence of alamethicin. Moreover, blue native PAGE indicated decreased intensity of protein bands of respiratory chain complex I in MSC16 leaves. Concerning the redox state, complex I impairment could be compensated to some extent by increased external NADH dehydrogenases (ND(ex)NADH) and alternative oxidase (AOX) capacity, the latter presenting differential expression in the light and in the dark. Although MSC16 mitochondria have a higher AOX protein level and an increased capacity, the AOX activity measured in the dark conditions by oxygen discrimination technique is similar to that in wild-type (WT) plants. Photosynthesis induction by light followed different patterns in WT and MSC16, suggesting changes in feedback chloroplast DeltapH caused by different adenylate levels. At steady-state, net photosynthesis was only slightly impaired in MSC16 mutants, while photorespiration rate (PR) was significantly increased. This was the result of large decreases in both stomatal and mesophyll conductance to CO2, which resulted in a lower CO2 concentration in the chloroplasts. The observed changes on CO2 diffusion caused by mitochondrial mutations open a whole new view of interaction between organelle metabolism and whole tissue physiology. The sum of all the described changes in photosynthetic and respiratory metabolism resulted in a lower ATP availability and a slower plant growth.
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http://dx.doi.org/10.1111/j.1399-3054.2007.00984.xDOI Listing
December 2007

Changes in energy status of leaf cells as a consequence of mitochondrial genome rearrangement.

Planta 2008 Feb 30;227(3):697-706. Epub 2007 Oct 30.

Institute of Experimental Plant Biology, University of Warsaw, Miecznikowa 1, 02-096 Warsaw, Poland.

The MSC16 cucumber (Cucumis sativus L.) mutant with lower activity of mitochondrial Complex I was used to study the influence of mitochondrial metabolism on whole cell energy and redox state. Mutant plants had lower content of adenylates and NADP(H) whereas the NAD(H) pool was similar as in wild type. Subcellular compartmentation of adenylates and pyridine nucleotides were studied using the method of rapid fractionation of protoplasts. The data obtained demonstrate that dysfunction of mitochondrial respiratory chain decreased the chloroplastic ATP pool. No differences in NAD(H) pools in subcellular fractions of mutated plants were observed; however, the cytosolic fraction was highly reduced whereas the mitochondrial fraction was more oxidized in MSC16, as compared to WTc. The NADP(H) pool in MSC16 protoplasts was greatly decreased and the chloroplastic NADP(H) pool was more reduced, whereas the extrachloroplastic pool was much more oxidized, than in WTc protoplast. Changes in nucleotides distribution in cucumber MSC16 mutant were compared to changes found in tobacco (Nicotiana sylvestris) CMS II mitochondrial mutant. In contrast to MSC16 cucumber, the content of adenylates in tobacco mutant was much higher than in tobacco wild type. The differences were more pronounced in leaf tissue collected after darkness than in the middle of the photoperiod. Results obtained after tobacco protoplast fractionating showed that the increase in CMS II adenylate content was mainly due to a higher level in extrachloroplast fraction. Both mutations have a negative effect on plant growth through perturbation of chloroplast/mitochondrial interactions.
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http://dx.doi.org/10.1007/s00425-007-0652-6DOI Listing
February 2008

Factors affecting determination of superoxide anion generated by mitochondria from barley roots after anaerobiosis.

J Plant Physiol 2004 Dec;161(12):1339-46

Institute of Experimental Plant Biology, University of Warsaw, Miecznikowa 1, PL-02-096 Warsaw, Poland.

During the post-hypoxic period, symptoms of oxidative stress and activation of enzymatic and non-enzymatic antioxidant systems were observed in several plant tissues. In the roots, mitochondrial respiratory chain is the main source of ROS. Superoxide anion radical is formed in the mitochondrial electron-transport chain at the level of Complexes I and III. The purpose of this work was to estimate superoxide anion production by the mitochondria isolated after a period of hypoxic treatment. Seedlings of barley (Hordeum vulgare L.) were grown on a nutrient medium flushed for 5d with air (control) or nitrogen (hypoxia) and then transferred for 24h to aerated medium (post-hypoxia). Production of superoxide anion by the mitochondria was measured by SOD-inhibitable oxidation of adrenaline to adrenochrome with NADH as a respiratory substrate. Hypoxic treatment increased mitochondrial activity but decreased mitochondrial superoxide anion appearance outside the mitochondrial membrane as compared to the mitochondria isolated from the roots continuously grown on aerated medium. The result of lower superoxide anion determination is attributed to increased antioxidants concentration during hypoxia. This was confirmed by inhibition of O2- production by exogenous GSH and stimulation by addition of 1-chloro-2,4-dinitrobenzene (CDNB), which depleted endogenous mitochondrial GSH.
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http://dx.doi.org/10.1016/j.jplph.2004.03.005DOI Listing
December 2004
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