Girdling induces oxidative damage and triggers enzymatic and non-enzymatic antioxidative defences in Citrus leaves

The effects of girdling on oxidative damage, antioxidant enzyme activity, antioxidant metabolites and proline (Pro) were studied in leaves arising from different shoot types of potted 2-year-old ‘Loretina’ mandarin (Citrus reticulata Blanco) trees during the spring flush period. Girdling increased malonyldialdehyde (MDA) and basal chlorophyll (Chl) a fluorescence (F_o) in young leaves 30 days after girdling but not in the mature leaves (ML) suggesting a disruption of photosynthetic apparatus and oxidative damage in young leaves. This phenomenon was accompanied by increasing levels of Pro. Paralleling these changes, an increase of all antioxidant enzyme activities occurred in leaves from vegetative (VG) and multiflowered leafy shoots (MLY) of girdled trees. Similarly, in ML of girdled trees, ascorbate peroxidase (APX), catalase (CAT) and glutathione reductase (GR) activity also increased. However, dehydroascorbate reductase (DHAR) activity decreased and superoxide dismutase (SOD) activity remained unchanged. Total leaf carbohydrate content and starch also increased as a result of girdling in all shoot types. Whilst soluble sugars increased markedly in young leaves, they increased only slightly in ML. In conclusion, this study provides evidence that girdling gives rise to oxidative damage in Citrus during carbohydrate accumulation, triggering enzymatic and non-enzymatic defence mechanisms.     

Girdling increases survival and growth of emerald ash borer larvae on Manchurian ash

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Stem girdling influences concentrations of endogenous cytokinins and abscisic acid in relation to leaf senescence in cotton

Many studies have shown that root–shoot imbalance influences vegetative growth and development of cotton (Gossypium hirsutum L.), but few have examined changes in leaf senescence and endogenous hormones due to stem girdling. The objective of this study was to determine the correlation between some endogenous phytohormones, particularly cytokinins and abscisic acid (ABA), and leaf senescence following stem girdling. Field-grown cotton plants were girdled on the main stem 5 days after squaring (DAS), while the non-girdled plants served as control. Plant biomass, seed cotton yield, main-stem leaf photosynthetic (Pn) rate, chlorophyll (Chl) and malondialdehyde (MDA) concentrations, as well as levels of cytokinins and ABA in main-stem leaves and xylem sap were determined after girdling or at harvest. Main-stem girdling decreased the dry root weight and root/shoot ratio from 5 to 70 days after girdling (DAG) and reduced seed cotton yield at harvest. Main-stem leaf Pn and Chl concentration in girdled plants were significantly lower than in control plants. Much higher levels of MDA were observed in main-stem leaves from 5 to 70 DAG, suggesting that stem girdling accelerated leaf senescence. Girdled plants contained less trans-zeatin and its riboside (t-Z + t-ZR), dihydrozeatin and its riboside (DHZ + DHZR), and isopentenyladenine and its riboside (iP + iPA), but more ABA than control plants in both main-stem leaves and xylem sap. These results suggested that main-stem girdling accelerated leaf senescence due to reduced levels of cytokinin and/or increased ABA. Cytokinin and ABA are involved in leaf senescence following main-stem girdling.     

Net CO(2) Assimilation and Carbohydrate Partitioning of Grapevine Leaves in Response to Trunk Girdling and Gibberellic Acid Application

Leaf net CO₂ assimilation rate (A), stomatal conductance (g_s), carboxylation efficiency, and foliar nonstructural carbohydrates were measured on mature, field-grown Vitis vinifera L. (cv Thompson Seedless) vines that had been trunk girdled, sprayed with gibberellic acid, or both, shortly after anthesis. Girdling reduced A, g_s, and carboxylation efficiency when measured 2 weeks after imposition of the treatments. Diurnal measurements indicated that A of girdled vines was less than that of control vines between 1000 and 1800 hours. Gibberellic acid mitigated the depressing effect of girdling on g_s during the same diurnal measurements. The concentrations of foliar carbohydrates were greatest for the girdled vines, followed by the combination treatment and were lowest for the control vines. Foliar carbohydrates were greater for girdled vines 4 weeks after the treatments were imposed, however, by this time there was no significant difference in A between the control and girdled vines. Two and 4 weeks after the experiment was initiated root carbohydrate concentrations were less for the girdled vines when compared to the control vines. The data indicate that the reduction in A of girdled grapevines is not associated with the accumulation of leaf nonstructural carbohydrates following the girdling treatment.     

Net CO(2) Assimilation Rate of Grapevine Leaves in Response to Trunk Girdling and Gibberellic Acid Application

Net CO₂ assimilation rate (A), stomatal conductance (g_s), and weight per unit leaf area (W) were determined on Thompson Seedless grapevines grown in the field. Treatments included fruit set applications of gibberellic acid (40 milligrams gibberellic acid (GA₃) per liter) to vines, shoots and clusters, alone and in combination with trunk girdling. Leaf A and g_s were measured prior to and 3, 6, and 13 days after fruit set. Weight per unit leaf area was determined on leaves collected subsequent to gas exchange measurements. Leaf A of girdled vines was reduced approximately 30% when compared to the control 13 days after treatment. The reduction in A due to girdling was not as great when vines were sprayed with GA₃. GA₃ sprays alone had no significant effect on A. Stomatal conductance was reduced by girdling 13 days after treatment. Weight per unit leaf area was 17% greater for trunk girdled vines when compared to the controls. Results indicate GA₃ affected net CO₂ assimilation rate only on girdled vines, a treatment which increased weight per unit leaf area.     

Birds girdling activity on exotic tree species as a form of adaptive behavior?

Four tree species in the Kostelec n. ?. l. arboretum (Czech Republic) have been repeatedly damaged by Dendrocopos medius. The unique aspect of this otherwise common behavior called girdling consists in regularly visiting the same trees every spring, although there are more than 1.200 tree species within the arboretum. We monitored transpiration, leaf phenology and the chemical composition of the xylem sap of girdled and nongirdled trees. Spectral analysis revealed slightly higher amounts of sugars, especially saccharose, in Cladrastis Raf. as the most regularly girdled tree among other conditions, comparing girdled to non-girdled trees. Higher transpiration rates were not confirmed in connection with girdling—quite the opposite—Cladrastis Raf. as the most highly favored tree for girdling showed the lowest transpiration rates (in average 6 kg water per day within spring months) compared to other non-girdled trees. We presume that the birds do not choose a particular tree on the basis of any visible or chemical traits but they examine many trees within their territory. Afterwards they probably remember the position of trees whose xylem sap starts to flow early in the spring compared to other trees, as their transpiration stream is enriched with sweet organic substances that represent an advantage for the forthcoming nesting period.     

Regulation of photosynthesis by end-product accumulation in leaves of plants storing starch, sucrose, and hexose sugars

In the present study, leaves of different plant species were girdled by the hot wax collar method to prevent export of assimilates. Photosynthetic activity of girdled and control leaves was evaluated 3 to 7 days later by two methods: (a) carbon exchange rate (CER) of attached leaves was determined under ambient CO₂ concentrations using a closed gas system, and (b) maximum photosynthetic capacity (A_max) was determined under 3% CO₂ with a leaf disc O₂ electrode. Starch, hexoses, and sucrose were determined enzymically. Typical starch storers like soybean (Glycine max L.) (up to 87.5 milligrams of starch per square decimeter in girdled leaves), cotton (Gossypium hirsutum L.), and cucumber (Cucumis sativus L.) responded to 7 days of girdling by increased (80-100%) stomatal resistance (r_s) and decreased A_max (>50%). On the other hand, spinach (Spinacia oleracea L.), a typical sucrose storer (up to 160 milligrams of sucrose per square decimeter in girdled leaves), showed only a slight reduction in CER and almost no change in A_max. Intermediate plants like tomato (Lycopersicon esculentum Mill.), sunflower (Helianthus annuus L.), broad bean (Vicia faba L.), bean (Phaseolus vulgaris L.), and pea (Pisum sativum L.), which upon girdling store both starch and sucrose, responded to the girdle by a considerable reduction in CER but only moderate inhibition of A_max, indicating that the observed reduction in CER was primarily a stomatal response. Both the wild-type tobacco (Nicotiana sylvestris) (which upon girdling stored starch and hexoses) and the starchless mutant (which stored only hexoses, up to 90 milligrams per square decimeter) showed 90 to 100% inhibition of CER and approximately 50% inhibition of A_max. In general, excised leaves (6 days) behaved like girdled leaves of the respective species, showing 50% reduction of A_max in wild-type and starchless N. sylvestris but only slight decline of A_max in spinach. The results of the present study demonstrate the possibility of the occurrence of end-product inhibition of photosynthesis in a large number of crop plants. The long-term inhibition of photosynthesis in girdled leaves is not confined to stomatal responses since the A_max declined up to 50%. The inhibition of A_max by girdling was strongest in starch storers, but starch itself cannot be directly responsible, because the starchless mutant of N. sylvestris was also strongly inhibited. Similarly, the inhibition cannot be attributed to hexose sugars either, because soybean, cotton, and cucumber are among the plants most strongly inhibited although they do not maintain a large hexose pool. Spinach, a sucrose storer, showed the least inhibition in both girdled and excised leaf systems, which indicates that sucrose is probably not directly responsible for the end-product inhibition of photosynthesis. The occurrence of strong end-product inhibition appears to be correlated with high acid-invertase activity in fully expanded leaves. The inhibition may be related to the nature of soluble sugar metabolism in the extrachloroplastic compartment and may be caused by a metabolite that has different rates of accumulation and turnover in sucrose storers and other plants.     

Evidence of a Role for Abscisic Acid in Mediating Stomatal Closure Induced by Obstructing Translocation from Leaves of Pearl Millet (Pennisetum americanum [L.] Leeke)

Application of a heat girdle near the base of the lamina ofthe fifth, fully expanded leaf of young pearl millet (Pennisetumamericanum [L.] Leeke) plants resulted in a decrease in solutepotential, an increase in leaf dry matter content, and a declinein stomatal conductance and in the rate of CO₂ assimilation.Total water potential was largely unaffected by girdling whileturgor potential increased as a consequence of the decreasein solute potential. Abscisic acid (ABA) content of the leaf increased 5 to 6-foldwithin 1 h of girdling, then declined equally rapidly beforeincreasing again at a slower rate. The decline in conductance was correlated with both the decreasein solute potential and the increase in ABA. To determine whichof these factors could be controlling conductance, girdled leaveswere exposed either to 14 h of continuous light or to a similarperiod of darkness followed by a brief light treatment to allowstomata to open. Girdling reduced conductance equally followingdarkness or light but solute accumulation occurred only in thelight. ABA accumulated in girdled leaves in both darkness andlight. Simultaneous measurements of conductance and CO₂ assimilationshowed that intercellular CO₂ concentration did not increasefollowing girdling. It was concluded that the decrease in conductancein millet leaves after girdling was most probably mediated bythe increase in ABA content. Key words: Leaf girdling, Solute accumulation, Stomatal conductance, Abscisic acid; Pennisetum americanum     

Girdling affects carbohydrate-related gene expression in leaves,bark and roots of alternate-bearing citrus trees

Effects of girdling on carbohydrate status and carbohydrate-related gene expression in citrus trees were investigated. Alternate-bearing 'Murcott' (a Citrus reticulata hybrid of unknown origin) trees were girdled during autumn (25 Sep. 2001) and examined 10 weeks later. Girdling brought about carbohydrate (soluble sugar and starch) accumulation in leaves and shoot bark above the girdle, in trees during their fruitless, 'off' year. Trees during their heavy fruit load, 'on' year did not accumulate carbohydrates above the girdle due to the high demand for carbohydrates by the developing fruit. Girdling caused a strong decline in soluble sugar and starch concentrations in organs below the girdle (roots), in both 'on' and 'off' trees. Expression of STPH-L and STPH-H (two isoforms of starch phosphorylase), Agps (ADP-glucose pyrophosphorylase, small subunit), AATP (plastidic ADP/ATP transporter), PGM-C (phosphoglucomutase) and CitSuS1 (sucrose synthase), all of which are associated with starch accumulation, was studied. It was found that gene expression is related to starch accumulation in all 'off' tree organs. RNA levels of all the genes examined were high in leaves and bark that accumulated high concentrations of starch, and low in roots with declining starch concentrations. It may be hypothesized that changes in specific sugars signal the up- and down-regulation of genes involved in starch synthesis.     

Changes in Photosynthesis and Fruit Characteristics in Olive in Response to Assimilate Availability

The source level in the olive cultivar Leccino was varied by girdling at different stages of fruit growth. Afterwards, the effects on gas exchange, fruit growth, and ripening and blooming were studied. Girdling during fruit growth did not significantly influence net photosynthetic rate (P _N) except in the last phase of fruit growth when the P _N was reduced. In the girdled branch, P _N began to decrease at the onset of starch accumulation because fruit growth ceased. In mid-November stomatal conductance (g _s) and transpiration rate (E) were also reduced by girdling, whereas sub-stomatal CO₂ concentration (C _i) increased in leaves from the girdled branches. The total chlorophyll content (Chl) tended to decrease in parallel with the reduced P _N. Girdling did not substantially influence the leaf and shoot water contents. The large availability of assimilates seems to cause an earlier fruit ripening. In general, girdling increased fruit dry mass. Healing before the time when the majority of pulp growth occurs reduced the effect of girdling. June girdling increased the pit dry mass. Girdling at the beginning of August and September, compared to the control, increased the pulp dry mass, but the pit dry mass did not differ with respect to the control. The percentage of oil in the fruit, on a dry mass basis, increased with August and September girdlings, but the percentage of oil in the pulp did not change. Girdling reduced shoot growth, but the internode length was unchanged. Girdling slightly stimulated differentiation of flower buds.     

Carbohydrate supply and n(2) fixation in soybean : the effect of varied daylength and stem girdling

When arrival of shoot supplied carbohydrate to the nodulated root system of soybean was interrupted by stem girdling, stem chilling, or leaf removal, nodule carbohydrate pools were utilized, and a marked decline in the rates of CO₂ and H₂ evolution was observed within approximately 30 minutes of treatment. Nodule excision studies demonstrated that the decline in nodulated root respiration was associated with nodule rather than root metabolism, since within 3.5 hours of treatment, nodules respired at less than 10% of the initial rates. Apparently, a continuous supply of carbohydrate from the shoot is required to support nodule, but not root, function. Depletion of nodular carbohydrate pools was sufficient to account for the (diminishing) nodule respiration of girdled plants. Of starch and soluble sugar pools within the whole plant, only leaf starch exhibited a diurnal variation which was sufficient to account for the respiratory carbon loss of nodules over an 8 hour night. Under 16 hour nights, or in continuous dark, first the leaf starch pools were depleted, and then nodule starch reserves declined concomitant with a decrease in the rates of CO₂ and H₂ evolution from the nodules. Nodule soluble sugar levels were maintained in dark treated plants but declined in girdled plants. The depletion of starch in root nodules is an indicator of carbohydrate limitation of nodule function.     

环剥对红松(Pinus koraiensis)韧皮部和木质部碳水化合物的影响

树干环剥可以阻碍韧皮部光合产物的运输并进一步影响光合产物的分配。长时期内,环剥能够导致环痕上部可溶性糖和淀粉的积累,但对于短期内如何影响碳水化合物在木质部和韧皮部内的运输模式所知甚少。以38年生红松(Pinus koraiensis Sieb.etZucc.)为研究材料,分别对环剥上部、下部每隔1~2d采样,区分木质部和韧皮部(树皮)进行可溶性糖和淀粉含量及树干糖呼吸消耗速率测定,确定环剥后的日变化和周变化,并对木质部可溶性糖、淀粉含量与韧皮部中相应指标进行相关关系的回归分析。结果发现:(1)环剥后4周内,在环剥痕上、下部间木质部可溶性糖和淀粉含量,韧皮部中淀粉含量均不存在显著差异(p>0.05),而韧皮部内可溶性糖含量,环剥后第2周出现显著差异,从第4周出现环剥上部显著高于下部的碳水化合物积累现象(p<0.05);(2)环剥阻隔了韧皮部可溶性糖的纵向运输,但是并不影响木质部的纵向运输,而且环剥并没有影响木质部和韧皮部之间的糖和淀粉的相关关系;(3)环剥第1周内环剥上部和下部呼吸消耗速率差异不显著,第2周环剥上部显著高于环剥下部,从第3周开始环剥下部呼吸消耗速率显著下降。推断认为,在环剥处理的4周内,环剥上部冠层新形成的碳水化合物很大一部分均被呼吸消耗掉,导致环剥上部较环剥下部可溶性糖稍有增加;红松胸高直径以下部分所储藏的碳水化合物足以保障2周内红松树干呼吸。     

杜仲剥皮再生对生长的影响

杜仲（Ｅｕｃｏｍｍｉａ ｕｌｍｏｉｄｅｓ Ｌｌｉｖ．）剥皮后每年７月测定剥皮株和照株的径向与高度生长量,每年秋季观测这些树的落叶情况,春季观测其萌芽情况,连续观察４年。同是对剥皮３年后在原剥皮部位作第二次剥皮的植株作了同样的３年观测。结果表明,剥皮当年树的落叶提前、发芽推迟,高度和树粗的生长明显减慢,剥皮后第二年其各方面的生长状况与对照株接近,第三年剥皮株的生长与对照株相似,但剥皮株的木材年生长量     

Differential controls by climate and substrate over the heterotrophic and rhizospheric components of soil respiration

The partitioning of soil respiration rates into the component processes of rhizospheric respiration (because of live roots and those microorganisms that subsist on root exudations) and heterotrophic respiration (because of decomposer microorganisms that subsist on the oxidation of soil organic matter) is difficult to accomplish through experimental observation. In order to minimize disturbance to the soil and maximize preservation of the natural relationships among roots, rhizospheric microorganisms, and decomposers, we conducted a girdling experiment in a subalpine forest dominated by lodgepole pine trees. In two separate years, we girdled trees in small forest plots (5–7 m in diameter) and trenched around the plots to sever invading roots in order to experimentally stop the transport of photosynthate from needles to roots, and eliminate rhizospheric respiration. Soil respiration rates in plots with trees girdled over 1 year prior to measurement were higher than those in plots with trees girdled 2–3 months prior to measurement. These results suggest that any stimulation of respiration because of the experimental artifact of fine root death and addition of labile carbon to the pool of decomposer substrates is slow, and occurs beyond the first growing season after girdling. Compared with control plots with nongirdled trees, soil respiration rates in plots with girdled trees were reduced by 31–44% at the mid‐summer respiratory maximum. An extreme drought during one of the 2 years used for observations caused greater reductions in the heterotrophic component of soil respiration compared with the rhizospheric component. In control plots, we observed a pulse in K₂SO₄‐extractable carbon during the spring snowmelt period, which was absent in plots with girdled trees. In control plots, soil microbial biomass increased from spring to summer, coincident with a seasonal increase in the rhizospheric component of soil respiration. In plots with girdled trees, the seasonal increase in microbial biomass was lower than in control plots. These results suggest that the observed seasonal increase in rhizospheric respiration rate in control plots was because of an increase in rhizospheric microbial biomass following ‘soil priming’ by a spring‐time pulse in dissolved organic carbon. Winter‐time, beneath‐snow microbial biomass was relatively high in control plots. Soil sucrose concentrations were approximately eight times higher during winter than during spring or summer, possibly being derived from the mechanical damage of shallow roots that use sucrose as protection against low‐temperature extremes. The winter‐time sucrose pulse was not observed in plots with girdled trees. The results of this study demonstrate that (1) the rhizospheric component of soil respiration rate at this site is significant in magnitude, (2) the heterotrophic component of soil respiration rate is more susceptible to seasonal drought than the rhizospheric component, and (3) the trees in this ecosystem exert a major control over soil carbon dynamics by ‘priming’ the soil with sugar exudates during the late‐spring snowmelt period and releasing high concentrations of sucrose to the soil during winter.     

Arsenic-induced oxidative stress in the common bean legume,Phaseolus vulgaris L. seedlings and its amelioration by exogenous nitric oxide

The adverse effects of arsenic (As) toxicity on seedling growth, root and shoot anatomy, chlorophyll and carotenoid contents, root oxidizability (RO), antioxidant enzyme activities, H₂O₂ content, lipid peroxidation and electrolyte leakage (EL%) in common bean (Phaseolus vulgaris L.) were investigated. The role of exogenous nitric oxide (NO) in amelioration of As-induced inhibitory effect was also evaluated using sodium nitroprusside (100 μM SNP) as NO donor and 2-(4-carboxy-2-phenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (200 μM PTIO) as NO scavenger in different combinations with 50 μM As. As-induced growth inhibition was associated with marked anomalies in anatomical features, reduction in pigment composition, increased RO and severe perturbations in antioxidant enzyme activities. While activity of superoxide dismutase and catalase increased, levels of ascorbate peroxidase, dehydroascorbate reductase and glutathione reductase decreased significantly and guaiacol peroxidase remained normal. The over-accumulation of H₂O₂ content along with high level of lipid peroxidation and electrolyte leakage indicates As-induced oxidative damage in P. vulgaris seedlings with more pronounced effect on the roots than the shoots. Exogenous addition of NO significantly reversed the As-induced oxidative stress, maintaining H₂O₂ in a certain level through balanced alterations of antioxidant enzyme activities. The role of NO in the process of amelioration has ultimately been manifested by significant reduction of membrane damage and improvement of growth performance in plants grown on As + SNP media. Onset of oxidative stress was more severe after addition of PTIO, which confirms the protective role of NO against As-induced oxidative damage in P. vulgaris seedlings.     

Tree-girdling to separate root and heterotrophic respiration in two Eucalyptus stands in Brazil

The release of carbon as CO₂ from belowground processes accounts for about 70% of total ecosystem respiration. Insights about factors controlling soil CO₂ efflux are constrained by the challenge of apportioning sources of CO₂ between autotrophic tree roots (and mycorrhizal fungi) and heterotrophic microorganisms. In some temperate conifer forests, the reduction in soil CO₂ efflux after girdling (phloem removal) has been used to separate these sources. Girdling stops the flow of carbohydrates to the belowground portion of the ecosystem, which should slow respiration by roots and mycorrhizae while heterotrophic respiration should remain constant or be enhanced by the decomposition of newly dead roots. Therefore, the reduction in CO₂ efflux after girdling should be a conservative estimate of the belowground flux of C from trees. We tested this approach in two tropical Eucalyptus plantations. Tree canopies remained intact for more than 3 months after girdling, showing no reduction in light interception. The reduction in soil CO₂ efflux averaged 16–24% for the 3-month period after girdling. The reduction in CO₂ efflux was similar for plots with one half of the trees girdled and those with all of the trees girdled. Girdling did not reduce live fine root biomass for at least 5 months after treatment, indicating that large reserves of carbohydrates in the root systems of Eucalyptus trees maintained the roots and root respiration. Our results suggest that the girdling approach is unlikely to provide useful insights into the contribution of tree roots and heterotrophs to soil CO₂ efflux in this type of forest ecosystem.     

THE ROLE OF RESERVES IN LEAVES,BRANCHES, STEMS,AND ROOTS ON SHOOT GROWTH OF RED PINE

By defoliation, girdling, and various combinations of these before the growing season began, the role of foods in old needles, branches, main stems, and roots on shoot growth of 8-yr-old red pine (Pinus resinosa) trees was studied in northern Wisconsin. Defoliation and girdling, alone or in combination, reduced shoot growth but defoliation reduced shoot dry weight more than it reduced shoot elongation. Reductions in shoot growth due to treatment were in the following decreasing order: branch girdling + needle removal > girdling at stem base + needle removal > needle removal > branch girdling > base girdling. The old needles were the major source of food for shoot growth and accounted for four-fifths or more of all shoot growth. The combined reserves in the branches, main stem, and roots accounted for less than 15% of shoot growth. The contribution of reserves from tissues other than old leaves was in the following order: branches > main stem > roots. Defoliation weakened apical dominance relations. In defoliated trees, many secondary axes elongated more than the terminal leader, and secondary axes in lower whorls often grew more than those in upper ones. The data suggest an important role of nutritional factors in correlative growth inhibition.     

Exogenous proline alleviates the effects of H2O2-induced oxidative stress in wild almond species

The effect of proline on the antioxidant system in the leaves of eight species of wild almond (Prunus spp.) exposed to H₂O₂-mediated oxidative stress was studied. The levels of endogenous proline (Pro) and hydrogen peroxide, and the activities of total superoxide dismutase (SOD), catalase (CAT), ascorbate peroxidase (APX), glutathione reductase (GR), and guaiacol peroxidase (POD) were measured. The degradation of chlorophyll but not carotenoids occurred in leaves in the solution of 5 mM H₂O₂. An increase in membrane lipid peroxidation was observed in H₂O₂ treatment, as assessed by MDA level and percentage of membrane electrolyte leakage (EL). Significant increases in total SOD and CAT activities, as well as decreases in APX and POD activities, were detected in H₂O₂-treated leaves. The three SOD isoforms showed different behavior, as Mn-SOD activity was enhanced by H₂O₂, whereas Fe-SOD and Cu/Zn-SOD activities were inhibited. In addition, Pro accumulation up to 0.1 ??mol/g fr wt, accompanied by significant decreases in ascorbate and glutathione levels, was observed in H₂O₂-treated leaves. After two different treatments with 10 mM Pro + 5 mM H₂O₂, total SOD and CAT activities were similar to the levels in control plants, while POD and APX activities were higher if compared to the leaves exposed only to H₂O₂. Pro + H₂O₂ treatments also caused a strong reduction in the cellular H₂O₂ and MDA contents and EL. The results showed that Pro could have a key role in protecting against oxidative stress injury of wild almond species by decreasing membrane oxidative damage.     

Developmental changes of antioxidative systems in tobacco leaves as affected by limited sucrose export in transgenic plants expressing yeast-invertase in the apoplastic space

It is generally believed that a restricted export of carbohydrates from source leaves causes oxidative stress because of an enhanced utilisation of O₂ instead of NADP⁺ as electron acceptor in photosynthesis. To test this hypothesis, developmental changes of antioxidative systems were investigated in wild-type and transgenic tobacco (Nicotiana tabacum L.) suffering from disturbed sink-source relations by expression of yeast invertase in the apoplastic space. Young expanding leaves of the wild type contained higher activities of Superoxide dismutase (EC 1.15.1.1), ascorbate peroxidase (EC 1.11.1.11), catalase (EC 1.11.1.6), dehydroascorbate reductase (EC 1.8.5.1), glutathione reductase (EC 1.6.4.2) and a higher glutathione content than mature source leaves. The activity of monodehydroascorbate-radical reductase (EC 1.1.5.4) and the ascorbate content remained unaffected by the developmental stage in the wild type. In young expanding leaves of the transgenic plants the capacity of the antioxidative systems was similar to or higher than in corresponding leaves from the wild type. Source leaves of transgenic tobacco with an increased carbohydrate content showed a small chlorophyll loss, an increased malondialdehyde content, a selective loss of the activities of Cu/Zn-superoxide dismutase isoenzymes and a fourfold decrease in ascorbate compared with the wild type. There was no evidence that the protection from H₂O₂ was insufficient since source leaves of transgenic tobacco contained increased activities of catalase, ascorbate peroxidase, and monodehydroascorbate-radical reductase and an increased ascorbate-to-dehydroascorbate ratio compared with source leaves of the wild type. In severely chlorotic leaf sections of the transgenic plants, most components of the antioxidative system were lower than in green leaf sections, but the ascorbate-to-dehydroascorbate ratio was increased. These results suggest that carbohydrate-accumulating cells have an increased availability of reductant, which can increase the degree of reduction of the ascorbate system via glutathione-related systems or via the activity of monodehydroascorbate-radical reductase. At the same time, transgenic tobacco leaves seem to suffer from an increased oxidative stress, presumably as a result of a decreased consumption of O ₂ ^.- by Cu/Zn-superoxide dismutases in the chloroplasts. There was no evidence that carbohydrate-accumulating leaves acclimated to enhanced O ₂ ^.- production rates in the chloroplasts.