Impacts of ozone on Plantago major: apoplastic and symplastic antioxidant status (original) (raw)
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Plant Physiology, 1997
Transgenic tobacco (Nicotiana tabacum cv Bel W3) plants were used to test the hypothesis that protection from O, injury could be conferred by overproduction of ascorbate peroxidase (APX) in the chloroplast. The 10-fold increase in soluble APX activity in the chloroplast was expected to alleviate an implied increase in oxidative potential and prevent damage caused by O,. Three different O, exposure experiments (one acute and two chronic) with two replicates each were conducted. APX activity in nontransgenic plants increased in response to chronic O, exposure. However, most responses to O, were similar between transgenic and nontransgenic plants. These included reductions in net photosynthesis and stomatal conductance, increases in ethylene emission and visible injury, and a decline in the leve1 of the small subunit of ribulose-1,5biphosphate carboxylase/oxygenase mRNA transcripts observed in response to the air pollutant in the acute and/or chronic experiments. No O,-induced effect on ribulose-1,5-biphosphate carboxylase/oxygenase quantity was observed in the chronic experiments. O, did not induce acceleration of senescence, as expected from studies with most other species; rather, the tobacco plants rapidly developed necrotic lesions. Thus, overproduction of APX in the chloroplast did not protect this cultivar of tobacco from O,.
Ascorbate in the leaf apoplast is a factor mediating ozone resistance in Plantago major
Plant Physiology and Biochemistry, 2000
The role of ascorbate in mediating ozone resistance was examined in Plantago major L. Seedlings of eleven populations which exhibited differential resistance to ozone were fumigated in controlled environment chambers with charcoal/Purafilt-filtered air (CFA) or CFA plus 15 nmol·mol -1 ozone overnight rising to a maximum between 12:00-16:00 hours of 75 nmol·mol -1 for 14 d. Measurements of ascorbate content were made on apoplastic and symplastic extracts. Populations differed in their constitutive level of ascorbate in youngest fully expanded leaves, and regression analysis revealed a significant correlation between ascorbate content in ozone-treated leaves and the ozone resistance of the populations. The relationship was stronger using apoplastic ascorbate levels than with corresponding symplastic measurements. The ascorbate content of the youngest fully expanded leaf of an ozone sensitive population was increased by foliar application of ascorbate. No significant difference in stomatal conductance was found between control and ascorbate-treated plants. Following spraying, plants were fumigated with 400 nmol·mol -1 ozone for 7 h. In control plants, ozone exposure resulted in extensive visible leaf damage (20-70 % at the end of the fumigation period) and decreased rates of CO 2 assimilation (-57 %). However, ascorbate treatment prevented the appearance of visible injury, and ameliorated the decline in photosynthesis induced by ozone (-26 %). Modelled data estimating the extent of protection afforded by apoplastic ascorbate against ozone supported the experimental observations. The results suggested that although apoplastic ascorbate plays an important role, other factors must also contribute to the mediation of ozone resistance in P. major. © 2000 Éditions scientifiques et médicales Elsevier SAS Apoplast / ascorbate / detoxification / ozone / Plantago major / resistance A, rate of CO 2 assimilation / ASC, ascorbate / AWF, apoplastic washing fluid / CFA, charcoal / Purafil t -filtered air / DHA, dehydroascorbate / g H2O , stomatal conductance to water vapour / O 3 , ozone / RLE, residual leaf extract / total ASC, ascorbate+dehydroascorbate Plant Physiol. Biochem., 2000, 38 (5), 403−411 /
Planta, 2003
Transgenic tobacco (Nicotiana tabacum L. cv. Xanthi) plants expressing cucumber ascorbate oxidase (EC.1.10.3.3) were used to examine the role of extracellular ascorbic acid in mediating tolerance to the ubiquitous air pollutant, ozone (O 3 ). Three homozygous transgenic lines, chosen on the basis of a preliminary screen of AO activity in the leaves of 29 lines, revealed up to a 380-fold increase in AO activity, with expression predominantly associated with leaf cell walls. Overexpression of AO resulted in no change in the total ascorbate content recovered in apoplast washing fluid, but the redox state of ascorbate was reduced from 30% in wild-type leaves to below the threshold for detection in transgenic plants. Levels of ascorbic acid and glutathione in the symplast were not affected by AO overexpression, but the redox state of ascorbate was reduced, while that of glutathione was increased. AO overexpressing plants exposed to 100 nmol mol -1 ozone for 7 h day -1 exhibited a substantial increase in foliar injury, and a greater pollutant-induced reduction in both the light-saturated rate of CO 2 assimilation and the maximum in vivo rate of ribulose-1,5-bisphosphate carboxylase/oxygenase carboxylation, compared with wild-type plants. Transgenic plants also exhibited a greater decline in CO 2 assimilation rate when exposed to a brief ozone episode (300 nmol mol -1 for 8 h). Stomatal conductance, hence O 3 uptake, was unaffected by AO over-expression. Our findings illustrate the important role played by ascorbate redox state and sub-cellular compartmentation in mediating the tolerance of plants to ozone-induced oxidative stress.
Plant, Cell and Environment, 2002
Ozone exposure stimulates an oxidative burst in leaves of sensitive plants, resulting in the generation and accumulation of hydrogen peroxide (H 2 O 2 ) in tobacco and tomato, and superoxide (O 2 -• ) together with H 2 O 2 in Arabidopsis accessions. Accumulation of these reactive oxygen species (ROS) preceded the induction of cell death, and both responses co-occurred spatially in the periveinal regions of the leaves. Re-current ozone exposure of the sensitive tobacco cv. Bel W3 in closed chambers or in the field led to an enlargement of existing lesions by priming the border cells for H 2 O 2 accumulation. Open top chamber experiments with native herbaceous plants in the field showed that Malva sylvestris L. accumulates O 2 -• at those sites that later exhibit plant cell death. Blocking of ROS accumulation markedly reduced ozone-induced cell death in tomato, Arabidopsis and M. sylvestris. It is concluded that ozone triggers an in planta generation and accumulation of H 2 O 2 and/or O 2 -• depending on the species, accession and cultivar, and that both these reactive oxygen species are involved in the induction of cell death in sensitive crop and native plants.
Redox state and peroxidase system in sunflower plants exposed to ozone
Plant Science, 2000
Sunflower plants subjected to a short-term fumigation with O 3 (150 ppb for 4 h repeated for 4 days) exhibited an increase in total ascorbate content, accompanied by a marked oxidation of ascorbate, leading to a decrease in its redox state, either at intracellular or extracellular level. O 3 exposure induced a rise in free extracellular peroxidase (POD) activity, assayed by syringaldazine as electron donor, as well as in the ionically and covalently cell wall bound PODs. On the contrary, the activity of both extracellular and intracellular guaiacol-POD did not show significant changes as a consequence of the pollutant exposure. The stimulation of syringaldazine-POD activities may be related to the effect of ozone on the growth of the cells, inducing an early senescence through the activation or acceleration of lignification processes. Beside, the reduced plasticity of the cell wall may oppose an unspecific mechanical resistance against the abiotic stress induced by the ozone exposure.
The apoplastic oxidative burst in response to biotic stress in plants: a three-component system
Journal of Experimental Botany, 2002
The oxidative burst, the generation of reactive oxygen species (ROS) in response to microbial pathogen attack, is a ubiquitous early part of the resistance mechanisms of plant cells. It has also become apparent from the study of a number of plant-pathogen interactions and those modelled by elicitor treatment of cultured cells that there may be more than one mechanism operating. However, one mechanism may be dominant in any given species. NADPH oxidases have been implicated in a number of systems and have been cloned and characterized. However, the enzyme system which is the major source of ROS in French bean (Phaseolus vulgaris) cells treated with a cell wall elicitor from Colletotrichum lindemuthianum, appears to be dependent on an exocellular peroxidase. The second component, the extracellular alkalinization, occurs as a result of the Ca 2[ and proton influxes and the K [ efflux common to most elicitation systems as one of the earliest responses. The third component, the actual reductantu substrate, has remained elusive. The low molecular weight compound composition of apoplastic fluid was compared before and after elicitation. The substrate only becomes available some min after elicitation and can be extracted, so that by comparing the profiles by LC-MS it has been possible to identify possible substrates. The mechanism has proved to be complex and may involve a number of low molecular weight components. Stimulation of H 2 O 2 production was observed with saturated fatty acids such as palmitate and stearate without concomitant oxylipin production. This biochemical evidence is supported by immunolocalization studies on papillae forming at bacterial infection sites that show the peroxidase isoform present at sites of H 2 O 2 production revealed by cerium chloride staining together with the crosslinked wall proteins and callose and callose synthase. The peroxidase has been cloned and expressed in Pichia pastoris and has been shown to catalyse the oxidation reaction with the same kinetics as the purified enzyme. Furthermore, Arabidopsis plants transformed heterologously using the French bean peroxidase in antisense orientation have proved to be highly susceptible to bacterial and fungal pathogens. Thus it is possible that Arabidopsis is another species with the potential to mount an apoplastic oxidative burst and these transformed plant lines may be useful to identify the peroxidase that is responsible.
Archives of Environmental Protection, 2018
Tropospheric ozone is a harmful air pollutant which may cause oxidative stress in plant cells, leading to biochemical and physiological changes and yield reduction. The aim of the study was to examine the cumulative effect of long-term ozone stress on the activity of antioxidative enzymes in leaves of ozone sensitive and resistant tobacco cultivar growing at the sites of various ozone concentrations. The one-month experiment was conducted in the 2009, 2010 and 2011 growing seasons with different meteorological conditions and ozone concentrations. The activity of SOD, APX and GuPX was measured four times at weekly intervals. The highest tropospheric ozone concentration was recorded in 2010 along with high level of solar radiation and temperature. The enhanced ozone level caused the increase in the activity of all examined enzymes. However, at more elevated ozone level the activity of all enzymes was higher in sensitive (Bel W3) than resistant (Bel B) tobacco cultivar. On the other hand, at lower ozone level the activity of examined enzymes was rather similar in both cultivars or even higher in ozone-resistant one, which was especially valid for SOD. A positive correlation between the activity of all enzymes and ozone concentration was shown in both cultivars. The presented results show that meteorological conditions modify tropospheric ozone concentration and real plant response to this environmental pollutant. It is highly important to interpret week-by-week plant response and environmental conditions bearing in mind the cumulative ozone effect resulting from previous weeks conditions.
The overexpression of an alternative oxidase gene triggers ozone sensitivity in tobacco plants
Plant, Cell & Environment, 2007
The alternative oxidase (AOX) of plant mitochondria transfers electrons from the ubiquinione pool to oxygen without energy conservation and prevents the formation of reactive oxygen species (ROS) when the ubiquinone pool is over-reduced. Thus, AOX may be involved in plant acclimation to a number of oxidative stresses. To test this hypothesis, we exposed wild-type (WT) Xanthi tobacco plants as well as Xanthi plants transformed with the Bright Yellow tobacco AOX1a cDNA with enhanced (SN21 and SN29), and decreased (SN10) AOX capacity to an acute ozone (O3) fumigation. As a result of 5 h of O3 exposition (250 nL L -1 ), SN21 and SN29 plants surprisingly showed localized leaf damage, whereas SN10, similarly to WT plants, was undamaged. In keeping with this observation, WT and SN21 plants differed in their response to O3 for the expression profiles of catalase 1 (CAT1), catalase 2 (CAT2), glutathione peroxidase (GPX) and ascorbate peroxidase (APX) genes, and for the activity of these antioxidant enzymes, which were induced in WT. Concomitantly, although ozone induced H2O2 accumulation in WT and in all transgenic lines, only in transgenics with high AOX capacity the H2O2 level in the post-fumigation period was high. The alternative pathway of WT plants was strongly stimulated by O3, whereas in SN21 plants, the respiratory capacity was always high across the treatment. The present results show that, far from exerting a protective role, the overexpression of AOX triggers an increased O3 sensitivity in tobacco plants. We hypothesize that the AOX overexpression results in a decrease of mitochondrial ROS level that in turn alters the defensive mitochondrial to nucleus signalling pathway that activates ROS scavenging systems.
Plant, Cell and Environment, 2001
The role that the constituents of the ascorbate-glutathione cycle play in the mechanism of contrasting ozone sensitivities was examined in mature and old tobacco leaves after acute ozone-fumigation (150 p.p.b., 5 h). Levels of the enzyme activities associated with the detoxifying system were lower in ozone-sensitive Bel W3 control plants than in unfumigated ozone-tolerant Bel B plants. In particular, the endogenous activities of ascorbate peroxidase (APX) and glutathione reductase (GR), and the metabolites ascorbic acid (AA) and reduced glutathione (GSH) were more abundant in Bel B than Bel W3 control plants. These results suggest that the higher tolerance of Bel B to O 3 is associated with a greater initial content of the antioxidant enzymes or metabolites. Only in the mature leaves of the ozone-tolerant Bel B cv. did fumigation trigger activation of APX and, weakly, of dehydroascorbate reductase (DHAR). The activity of these enzymes was significantly lower after ozone treatment in both mature and old leaves of Bel W3 than in control plants. Fumigation had little effect on the ascorbate content. Its main effects on the glutathione pool were that it boosted the oxidized form and lowered the reduced form, particularly in mature Bel W3 leaves. Extractable GR activity remained unchanged in both Bel B and Bel W3 immediately after fumigation, but increased slightly 24 h later, particularly in mature leaves of Bel W3. Exposure to O 3 caused a sharp decline in chloroplastic GR mRNA levels in both cultivars. However, as Western blot analysis failed to detect any major changes in GR protein content at this time, the protein must be highly stable. There is therefore a good correlation between tolerance to O 3 and high endogenous levels of antioxidant metabolites such as AA and GSH in tobacco. In addition, the degree of inducibility of the system discriminates the two cultivars investigated.
Plant, Cell and Environment, 2005
Apoplastic reactive oxygen intermediates, which are formed during the exposure of a higher plant to ozone (O 3 ), have been proposed to be detoxified by apoplastic ascorbate (ASC). An investigation to determine whether the differential sensitivity of two white clover clones ( Trifolium repens L. cv Regal) to O 3 was related with their levels of ASC, glutathione derivatives or with the total antioxidative capacity. In contrast to what might be expected, the sensitive clone of white clover (NC-S) constitutively showed a 72% higher concentration of apoplastic ASC compared to the O 3 -tolerant clone (NC-R). Furthermore, NC-S also showed a higher redox status of apoplastic ASC. These results indicate that higher ASC levels in the apoplast of NC-S are not sufficient to induce a higher O 3 tolerance. The redox status, but not the absolute concentration of homoglutathione in the symplast was found to be constitutively higher in NC-R than in NC-S. It is not clear, however, whether homoglutathione is a direct cause of the differential O 3 detoxification capacity of both clones. Total antioxidative capacity measurements ruled out the contribution of other low-molecular antioxidants to the relative tolerance of NC-R. It was concluded that elevated apoplastic ASC levels can not always be sufficient to render a plant O 3 tolerant.