Peroxisomes as a source of superoxide and hydrogen peroxide in stressed plants (original) (raw)
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PLANT PHYSIOLOGY, 1988
The intraorganellar distribution of superoxide dismutase (SOD) (EC 1.15.1.1) in two types of plant peroxisomes (glyoxysomes and leaf peroxisomes) was studied by determinations of SOD latency in intact organelles and by solubilization assays with 0.2 molar KCI. Glyoxysomes were purified from watermelon (Citrullus vulgaris Schrad.) cotyledons, and their integrity, calculated on the basis of glyoxysomal marker enzymes, was about 60%. Under the same conditions, the latency of SOD activity determined in glyoxysomes was 40%. The difference between glyoxysomal intactness and SOD latency was very close to the percentage of isozyme Mn-SOD previously determined in glyoxysomes (LM Sandalio, LA Del Rio 1987 J Plant Physiol 127: 395409). In matrix and membrane fractions of glyoxysomes, SOD exhibited a solubilization pattern very similar to catalase, a typical soluble enzyme of glyoxysomes. The analysis of the distribution of individual SOD isozymes in glyoxysomal fractions treated with KCI showed that Cu,Zn-SOD II, the major SOD isozyme in glyoxysomes, was present in the soluble fraction of these organelles, whereas Mn-SOD was bound to the glyoxysomal membrane. These data in conjunction with those of latency of SOD activity in intact glyoxysomes suggest that Mn-SOD is bound to the external side of the membrane of glyoxysomes. On the other hand, in intact leaf peroxisomes where only a Mn-containing SOD is present (LM Sandalio, JM Palma, LA Del Rio 1987 Plant Sci 51: 1-8), this isozyme was found in the peroxisomal matrix. The physiological meaning of SOD localization in matrix and membrane fractions of glyoxysomes and the possibility of new roles for plant peroxisomes in cellular metabolism related to activated oxygen species is discussed.
New Phytologist, 1998
The number and type of isoforms of superoxide dismutase (SOD) and their activities were compared in mitochondria and peroxisomes isolated from cotyledons of three different oilseed seedlings. Mitochondrial and peroxisomal isoforms of SOD could be distinguished in nondenaturing polyacrylamide gels by their differential sensitivities to KCN and\or H # O # . The type of SOD was not the same for each organelle in each of the three oilseed species. For example, a single Mn-SOD was found in cotton and cucumber mitochondria, whereas four CuZn-SODs were present in mitochondria from sunflower. At least one CuZn-SOD isoform was found in the peroxisomes of all three species. Cucumber peroxisomes contained both a CuZn-SOD and a Mn-SOD, cotton peroxisomes contained a single CuZn-SOD, whilst four separate CuZn-SODs, but no Mn-SOD were found in sunflower peroxisomes. Using antibodies against CuZn-SOD from watermelon peroxisomes or from chloroplasts of Equisetum, a single polypeptide of c. 16n5 kDa was detected on immunoblots of peroxisomal fractions from the three species. Post-embedment, electron-microscopic double immunogold-labelling showed that CuZn-SOD, with malate synthase used as marker enzyme of peroxisomes, was localized in the matrix of these organelles of all three species. These results suggest that CuZn-SOD is a characteristic matrix enzyme of peroxisomes in oilseed cotyledons.
Journal of Experimental Botany, 2007
In this work the manganese superoxide dismutase (Mn-SOD) bound to peroxisomal membranes of watermelon cotyledons (Citrullus lanatus Schrad.) was purified to homogeneity and some of its molecular properties were determined. The stepwise purification procedure consisted of ammonium sulphate fractionation, batch anion-exchange chromatography, and anion-exchange and gel-filtration column chromatography using a fast protein liquid chromatography system. Peroxisomal membrane Mn-SOD (perMn-SOD; EC 1.15.1.1) was purified 5600-fold with a yield of 2.6 mg of enzyme g 21 of cotyledons, and had a specific activity of 480 U mg 21 of protein. The native molecular mass determined for perMn-SOD was 108 000 Da, and it was composed of four equal subunits of 27 kDa, which indicates that perMn-SOD is a homotetramer. Ultraviolet and visible absorption spectra of the enzyme showed a shoulder at 275 nm and two absorption maxima at 448 nm and 555 nm, respectively. By isoelectric focusing, a pI of 5.75 was determined for perMn-SOD. In immunoblot assays, purified perMn-SOD was recognized by a polyclonal antibody against Mn-SOD from pea leaves, and the peroxisomal enzyme rapidly dissociated in the presence of dithiothreitol and SDS. The potential binding of the Mn-SOD isozyme to the peroxisomal membrane was confirmed by immunoelectron microscopy analysis. The properties of perMn-SOD and the mitMn-SOD are compared and the possible function in peroxisomal membranes of the peripheral protein Mn-SOD is discussed.
Metabolism of oxygen radicals in peroxisomes and cellular implications
Free Radical Biology and Medicine, 1992
Peroxisomes are subcellular respiratory organelles which contain catalase and HaO2-producing ravin oxidases as basic enzymatic constituents. These organelles have an essentially oxidative type of metabolism and have the potential to carry out different important metabolic pathways. In recent years the presence of different types ofsuperoxide dismutase (SOD) have been demonstrated in peroxisomes from several plant species, and more recently the occurrence of SOD has been extended to peroxisomes from human and transformed yeast cells. A copper, zinc-containing SOD from plant peroxisomes has been purified and partially characterized. The production of hydroxyl and superoxide radicals has been studied in peroxisomes. There are two sites of O~ production in peroxisomes: (1) in the matrix, the generating system being xanthine oxidase; and (2) in peroxisomal membranes, dependent on reduced nicotinamide adenine dinucleotide (NADH), and the electron transport components of the peroxisomal membrane are possibly responsible. The generation of oxygen radicals in peroxisomes could have important effects on cellular metabolism. Diverse cellular implications of oxyradical metabolism in peroxisomes are discussed in relation to phenomena such as cell injury, peroxisomal genetic diseases, peroxisome proliferation and oxidative stress, metal and salt stress, catabolism of nucleic acids, senescence, and plant pathogenic processes. her dissertation dealt with the localization of SOD in specialized plant peroxisomes and the superoxide production in peroxisomes. She did postdoctoral work (1988)(1989) in the Department of Biology at the Washington University, Saint Louis, and she is currently Reprinted with permission from Sandalio, L. M.: Palma, J. M.; del Rio, L. A. Localization of manganese superoxide dismutase in peroxisomes isolated from Pisurn sativum L. Plant Sci 51:4: 1987.
Plant Peroxisomes, Reactive Oxygen Metabolism and Nitric Oxide
Iubmb Life, 2003
In plant cells, as in most eukaryotic organisms, peroxisomes are probably the major sites of intracellular H 2 0 2 production, as a result of their essentially oxidative type of metabolism. Like mitochondria and chloroplasts, peroxisomes also produce superoxide radicals (02'-) and there are, at least, two sites of superoxide generation: one in the organelle matrix, the generating system being xanthine oxidase, and another site in the peroxisomal membranes dependent on NAD(P)H. In peroxisomal membranes, three integral polypeptides (PMPs) with molecular masses of 18, 29, and 32 kDa have been shown to generate 0 2 . -radicals. Besides catalase, several antioxidative systems have been demonstrated in plant peroxisomes, including different superoxide dismutases, the four enzymes of the ascorbate-glutathione cycle plus ascorbate and glutathione, and three NADP-dependent dehydrogenases. A CuZn-SOD and two Mn-SODS have been purified and characterized from different types of plant peroxisomes. The presence of the enzyme nitric oxide synthase (NOS) and its reaction product, nitric oxide (NO'), has been recently demonstrated in plant peroxisomes. Different experimental evidence has suggested that peroxisomes have a ROS-mediated cellular function in leaf senescence and in stress situations induced by xenobiotics and heavy metals. Peroxisomes oxygen species; SOD, superoxide dismutase; XDH, xanthine dehydrogenase; XOD, xanthine oxidase. could also have a role in plant cells as a source of signal molecules like NO', 02'radicals, H202, and possibly S-nitrosoglutathione (GSNO). It seems reasonable to think that a signal moleculeproducing function similar to that postulated for plant peroxisomes could also be performed by human, animal and yeast peroxisomes, where research on oxy radicals, antioxidants and nitric oxide is less advanced than in plant peroxisomes. IUBMB Life, 55: [71][72][73][74][75][76][77][78][79][80][81]2003
Localization of manganese superoxide dismutase in peroxisomes isolated from Pisum sativum L
Plant Science
The presence of the metalloenzyme superoxide dismutase (SOD) in peroxlsomes from leaves of Pisum sativum L. was studied. Plant organelles were isolated by differential and Percoll density-gradient centrifugation. Purified intact peroxisomes contained a superoxide dlsmutase isozyme which was identified, on the basis of polyacrylamide-gel analysis and KCNand H202-sensitivity, as a Mn-containing SOD (Mn-SOD). By determination of latency of superoxide dismutase activity in intact peroxlsomes, Mn-SOD was demonstrated to be located in the interior of these oxidative organelles. In terms of specific activity, the peroxisomal Mn superoxide dismutase represents at least 50% of the whole SOD activity occurring in mitochondria. The presence of a Mn-SOD in peroxisomes strongly suggests the generation in these oxidative organelles of superoxide free radicals (02"-), the substrate of the enzyme, as well as new activated oxygen-related functions for peroxisomes in cellular metabolism.
Reactive oxygen species, antioxidant systems and nitric oxide in peroxisomes
Journal of Experimental Botany, 2002
Peroxisomes are subcellular organelles with an essentially oxidative type of metabolism. Like chloroplasts and mitochondria, plant peroxisomes also produce superoxide radicals (O 2 $ À ) and there are, at least, two sites of superoxide generation: one in the organelle matrix, the generating system being xanthine oxidase, and another site in the peroxisomal membranes dependent on NAD(P)H. In peroxisomal membranes, three integral polypeptides (PMPs) with molecular masses of 18, 29 and 32 kDa have been shown to generate O 2 $ À radicals. Besides catalase, several antioxidative systems have been demonstrated in plant peroxisomes, including different superoxide dismutases, the ascorbateglutathione cycle, and three NADP-dependent dehydrogenases. A CuZn-SOD and two Mn-SODs have been purified and characterized from different types of peroxisomes. The four enzymes of the ascorbate-glutathione cycle (ascorbate peroxidase, monodehydroascorbate reductase, dehydroascorbate reductase, and glutathione reductase) as well as the antioxidants glutathione and ascorbate have been found in plant peroxisomes. The recycling of NADPH from NADP q can be carried out in peroxisomes by three dehydrogenases: glucose-6phosphate dehydrogenase, 6-phosphogluconate dehydrogenase, and isocitrate dehydrogenase. In the last decade, different experimental evidence has suggested the existence of cellular functions for peroxisomes related to reactive oxygen species (ROS), but the recent demonstration of the presence of nitric oxide synthase (NOS) in plant peroxisomes implies that these organelles could also have a function in plant cells as a source of signal molecules like nitric oxide (NO $ ), superoxide radicals, hydrogen peroxide, and possibly S-nitrosoglutathione (GSNO).
Acta Physiologiae Plantarum, 2017
Peroxiredoxins (Prxs) constitute a group of thiolspecific antioxidant enzymes which are present in bacteria, yeasts, and in plant and animal cells. Although Prxs are mainly localized in the cytosol, they are also present in mitochondria, chloroplasts, and nuclei, but there is no evidence of the existence of Prxs in plant peroxisomes. Using soluble fractions (matrices) of peroxisomes purified from leaves of pea (Pisum sativum L.) plants, the immunological analysis with affinity-purified IgG against yeast Prx1 revealed the presence of an immunoreactive band of about 50 kDa. The apparent molecular mass of the peroxisomal Prx was not sensitive to oxidizing and reducing conditions what could be a mechanism of protection against the oxidative environment existing in peroxisomes. Postembedment, EM immunocytochemical analysis with affinity-purified IgG against yeast Prx1 antibodies, confirmed that this protein was present in the peroxisomal matrix, mitochondria, and chloroplasts. In pea plants grown under oxidative stress conditions, the protein level of peroxisomal Prx was differentially modulated, being slightly induced by growth of plants with 50 lM CdCl 2 , but being significantly reduced by treatment with the herbicide 2,4-dichlorophenoxyacetic acid (2,4-D). The presence in the matrix of peroxisomes of a protein immunorelated to Prx of about 50 kDa, which is in the range of molecular mass of the dimeric form of other Prxs, opens new questions on the molecular properties of Prxs, but also on their function in the metabolism of reactive oxygen and nitrogen species (ROS/RNS) in these plant cell organelles, where they could be involved in the regulation of hydrogen peroxide and/or peroxynitrite.
Free Radical Research, 1991
The effect of micronutrient stress (either deficiency or toxicity) on the expression or different superoxide dismutase isoenzymes in plants is reviewed. The induction of Fe-SOD and Mn-SOD by different metals and the potential use of the metalloentyme system SOD lor the appraisal of the micronutrient status of plants, is examined. At subcellular level, evidence for the participation of peroxisomal SOD in the molecular mechanism of plant tolerance to Cu is presented, and the activated oxygen-dependent toxicity of a xenobiotic (clofibrate) in plant peroxisomes is examined.