Significance of antioxidants and electron sinks for the cold-hardening-induced resistance of winter rye leaves to photo-oxidative stress

The contents of ascorbate and glutathione and the activities of superoxide dismutase and glutathione reductase were increased to levels as high as those in cold-hardened leaves (CHL) by incubating non-hardened leaves (NHL) of winter rye (Secale cereale L.) with the precursor substrates L -galactonic acid-γ-lactone and 2-oxothiazolidine-4-carboxylate. Reduced glutathione was rapidly depleted from NHL after application of D , L -buthionine sulfoximine, an inhibitor of its biosynthesis. In spite of greatly divergent antioxidant contents the rates of photo-inactivation of photosystem II (PSII) and catalase observed in the presence of translation inhibitors did not differ greatly. The paraquat-induced catalase inactivation and chlorophyll degradation in light were reduced in NHL with increased antioxidant levels. Paraquat-induced photo-inactivation of PSII was, however, not mitigated. The CHL had a higher capacity to prevent paraquat-induced oxidation of ascorbate and glutathione than NHL with increased antioxidant contents. Increased antioxidant contents did not establish resistance to low temperature-induced photo-inactivation of PSII and catalase in NHL. The resistance of CHL to low temperature-induced photo-inactivation of PSII and catalase required repair at low temperature and active carbon assimilation but was only little affected when photorespiration was suppressed by phosphinothricin. Protection of PSII depended also on non-photochemical quenching of excitation energy.     

Resistance to photoinhibition of photosystem II and catalase and antioxidative protection in high mountain plants

In leaves of three alpine high mountain plants, Homogyne alpina, Ranunculus glacialis and Soldanella alpina, both photosystem II (PSII) and the enzyme catalase appeared to he highly resistant to photoinactivation under natural field conditions. While the Dl protein of PSII and catalase have a rapid turnover in light and require continuous new protein synthesis in non-adapted plants, little apparent photoinactivation of PSII or catalase was induced in the alpine plants by translation inhibitors or at low temperature, suggesting that turnover of the Dl protein and catalase was slow in these leaves. In vitro PSII was rapidly inactivated in light in isolated thylakoids from H. alpina and R. glacialis. In isolated intact chloroplasts from R. glacialis, photoinactivation of PSII was slower than in thylakoids. Partially purified catalase from R. glacialis and S. alpina was as sensitive to photoinactivation in vitro as catalases from other sources. Catalase from H. alpina had, however, a 10-fold higher stability in light. The levels of xanthophyll cycle carotenoids, of the antioxidants ascorbate and glulathione, and of the activities of catalase, superoxide dismutase and glutathione reductase were very high in S. alpina, intermediate in H. alpina, but very low in R. glacialis. However, isolated chloroplasts from all three alpine species contained much higher concentrations of ascorbate and glutathione than chloroplasts from lowland plants.     

Rapid photosynthetic adaptation to heat stress triggered in potato leaves by moderately elevated temperatures

Photosystem II (PSII) is considered to be one of the most thermolabile aspects of photosynthesis. In vivo measurements of chlorophyll fluorescence and photosynthetic oxygen evolution in 25°C-grown potato leaves (cv. Haig) indicated that the threshold temperature T_c above which PSII denatures was indeed rather low–about 38°C–with temperatures higher than Tc causing a rapid and irreversible loss of PSII activity. The present study demonstrates the existence of adaptive processes which rapidly adjust the in vivo thermal stability of PSII in response to temperature increase. Transfer of potato leaves from 25°C to temperatures slightly lower than T_c (between 30 and 35°C) was observed to cause an upward shift of the Tc value without any appreciable loss of PSII activity. This increase in PSII thermotolerance was substantial (around +5°C in the Haig cultivar), rapid (with a half-time of ～20 min) and slowly reversible at 25°C (>24h). As a consequence, high temperatures (e.g. 40°C) which caused a complete and irreversible inhibition of the PSII function had very little effect in 35°C-treated leaves, thus suggesting that the above-described PSII changes could be of prime importance for the plant's behaviour in the field. Accordingly, the rise in Tc at 35°C was much larger (+8°C) in Sahel, a stress-resistant potato variety, than in the heat-sensitive Haig cultivar.     

Changes in activity of antioxidative enzymes in wheat (Triticum aestivum) seedlings under cold acclimation

Activated oxygen species such as superoxide radicals, singlet oxygen, hydrogen peroxide and hydroxyl radicals can be produced in plants exposed to low, non-freezing, non-injurious temperatures. To prevent or alleviate oxidative injury, plants have evolved several mechanisms which include scavenging by natural antioxidants and enzymatic antioxidant systems such as superoxide dismutases, catalase and peroxidases. Although overproduction of hydrogen peroxide and increased tolerance to oxidative stress can be induced in wheat by low-temperature treatments, data concerning changes in the enzymatic antioxidant systems are almost absent. With the aim to provide this information, antioxidant enzyme (superoxide dismutases, catalase and peroxidases) activities were analysed in leaves and roots of Triticum aestivum cvs Brasilia (frost resistant in field) and Eridano (less frost resistant in field) seedlings grown at day/night temperatures of 24/22°C (control treatment) and 12/5°C (low-temperature treatment). Our data showed that superoxide dismutase activities were unaffected by low-temperature treatment both in leaves and roots. Catalase activity in leaves and roots was decreased in 12/5°C-grown seedlings, but Brasilia maintained higher catalase activity than Eridano. Differences were also observed in guaiacol peroxidase activities between control and acclimated seedlings: Higher guaiacol peroxidase activities were found in the leaves of 12/5°C-grown seedlings while in roots these activities were lower. Moreover, Brasilia guaiacol peroxidase activities were higher than Eridano. Superoxide dismutase and peroxidase zymogram analyses showed that synthesis of new isoforms was not induced by low-temperature treatment. Changes in the activities of antioxidant enzymes induced by cold acclimation support the hypothesis that a frost-resistant wheat cultivar, in comparison with a less frost-resistant one, maintains a better defence against activated oxygen species during low-temperature treatment.     

Temperature-sensitivity of D1 protein metabolism in isolated Zea mays chloroplasts

The effects of low temperature on the synthesis and stability of the 32 kDa D1 protein of photosystem II were investigated in chloroplasts isolated from maize (Zea mays cv. LG11) leaves. The synthesis of D1 by intact chloroplasts in vitro was strongly dependent on temperature; the Q₁₀ for the initial rate of incorporation of [³⁵S]-methionine into D1 was ca. 2.6 over the range 13–25°C. The synthesis of other thylakoid polypeptides exhibited a similar temperature dependence, whilst synthesis of stromal proteins was considerably less temperature-dependent, with the exception of two polypeptides of ca. 56 and 59.5 kDa. The stability of newly-synthesized D1 in the thylakoid membranes was dependent both on the temperature at which the plants were grown and on the temperature during the pulse-labelling period when the protein was synthesized. In chloroplasts isolated from maize leaves grown at 25°C, D1 that was synthesized and assembled at 25 °C in vitro was rapidly degraded during the chase period. At lower chase temperatures the protein was more stable. When chloroplasts from 25°C-grown leaves were pulse-labelled at 13°C, the stability of D1 was markedly enhanced at all temperatures during the chase period. This effect was even more pronounced in chloroplasts isolated from plants grown at 14°C. The implications of these results are discussed with regard to the ability of maize to recover from photoinhibitory damage at low temperatures.     

Chloroplast thylakoid protein changes induced by low growth temperature in maize revealed by immunocytology

Tissue-specific effects of low growth temperature on maize chloroplast thylakoid protein accumulation were analysed using immunocytology. Sections of leaves from plants grown at 25 and 14°C were probed with antibodies to specific chloroplast thylakoid proteins from the four major protein multisubunit complexes of the thylakoid membrane followed by fluorescein-conjugated goat anti-rabbit antibodies. At a normal growth temperature of 25°C, the 32 kDa D1 protein of the photosystem II reaction centre and the 33 kDa protein of the extrinsic oxygen-evolving complex of photosystem II are both accumulated to a greater degree in the mesophyll than in the bundle sheath chloroplasts. In contrast, subunit II of photosystem I, cytochrome f and the α- and β-subunits of ATP synthetase are predominant in the bundle sheath thylakoids at 25°C. A striking difference between the 25°C-grown and the 14°C-grown leaf tissue was the presence in the latter of (20–30%) cells whose chloroplasts apparently completely lack several of the thylakoid proteins. In plants grown at 14°C, the accumulation of the 33 kDa protein of the extrinsic oxygen-evolving complex of photosystem II was apparently unchanged, but other thylakoid proteins showed a significant reduction. The uneven distribution of proteins between the bundle sheath and mesophyll chloroplasts observed at 25°C was also maintained at 14°C. Reduction in the fluorescence at 14°C was manifested either as an overall reduction in the diffuse fluorescence across the chloroplast profiles or less frequently as a reduction to small discrete bodies of intense fluorescence. The significance of these results to low-temperature-induced reduction in the photosynthetic productivity of maize is discussed.     

Thermal acclimation and photoacclimation of photosynthesis in the brown alga Laminaria saccharina

Thermal acclimation and photoacclimation of photosynthesis were compared in Laminaria saccharina sporophytes grown at temperatures of 5 and 17 °C and irradiances of 15 and 150μmol photons m^?2 s^?1. When measured at a standard temperature (17°C), rates of light-saturated photosynthesis (P_max) were higher in 5 °C-grown algae (c. 3.0 μmol O₂ m^?2 s^?1) than in 17 °C-grown algae (c. 0.9 μmol O₂ m_-2 s_-1). Concentrations of Rubisco were also 3-fold higher (per unit protein) in 5 °C-grown algae than in algae grown at 17 °C. Light-limited photosynthesis responded similarly to high temperature and low light Photon yields (α) were higher in algae grown at high temperature (regardless of light), and at 5 °C in low light, than in algae grown at 5 °C in high light Differences in a were correlated with light absorption; both groups of 17 °C algae and 5 °C low-light algae absorbed c. 75% of incident light, whereas 5 °C high-light algae absorbed c. 55%. Increased absorption was correlated with increases in pigment content PSII reaction centre densities and the fucoxanthin-Chl ale protein complex (FCP). Changes in a were also attributed, in part, to changes in the maximum photon yield of photosynthesis (0_max). PSI reaction centre densities were unaffected by growth temperature, but the areal concentration of PSI in low-light-grown algae was twice that of high-light-grown algae (c. 160.0 versus 80.0 nmol m^?2). We suggest that complex metabolic regulation allows L, saccharina to optimize photosynthesis over the wide range of temperatures and light levels encountered in nature.     

Acclimation by suboptimal growth temperature diminishes photooxidative damage in maize leaves

Leaves of Zea mays L. seedlings which developed at optimal (25°C) or suboptimal (15°C) temperature were exposed to high irradiance (1000 μmol m^?2 s^?1) and a severe chilling temperature (5°C) for up to 24 h to investigate their ability to withstand photooxidative stress. During this stress, the degradation of the endogenous antioxidants ascorbate, glutathione and α-tocopherol was delayed and less pronounced in 15°C leaves. Similarly, the decline in chlorophyll a, chlorophyll b, β-carotene and lutein was slower throughout the stress period. Faster development and a higher level of non-photochemical quenching (NPQ) of chlorophyll fluorescence, related to a stronger de-poxidation of the larger xanthophyll cycle pool in 15°C leaves, could act as a defence mechanism to reduce the formation of reactive oxygen species during severe chilling. Furthermore, plants grown at suboptimal temperature exhibited a higher amount of the antioxidants glutathione and α-tocopherol. The higher α-tocopherol content in leaves (double based on leaf area; 4-fold higher based on chlorophyll content) which developed at suboptimal temperature may play an especially important role in the stabilization of the thylakoid membrane and thus prevent lipid peroxidation.     

Carotenoid composition in Zea mays developed at sub-optimal temperature and different light intensities

The content and composition of pigments were examined in the third leaf of Zea mays L. plants grown under controlled environment at near-optimal temperature (24°C) or sub-optimal temperature (14°C) at a light intensity of either 200 or 600 μmol m^?2 s^?1. Compared to leaves grown at 24°C, leaves grown at 14°C showed a large reduction in the chlorophyll (Chl) content, a marked decrease in the Chl a/b ratio, and a large increase in the ratio of total carotenoids/Chl a+b. Leaves grown at 14°C showed a much lower content of β-carotene than leaves grown at 24°C, while the content of the carotenoids of the xanthophyll cycle (violaxanthin [V] + antheraxanthin [A] + zeaxanthin [Z]) was markedly higher in the former leaves as compared to the latter leaves; neoxanthin and lutein were affected by the growth temperature to a much lesser extent. The xanthophylls/β-carotene ratio was about three times higher in leaves grown at 14°C as compared to leaves grown at 24°C. On a chlorophyll basis, the two types of leaves hardly differed in their level of β-carotene, while the levels of the xanthophylls (including lutein and neoxanthin) were higher in 14°C-grown leaves as compared to 24°C-grown leaves. In leaves grown at 14°C, 40 and 56% of the V+A+Z pool was in the form of zeaxanthin at low light intensity and high light intensity, respectively. Only trace amounts of zeaxanthin, if any, were present in leaves grown at 24°C. The changes in the pigment composition induced by growth at sub-optimal temperature were more pronounced at a light intensity of 600 as compared to 200 μmol m^?2 s^?1. In the given range, the light intensity slightly affected the composition of pigments in leaves grown at 24°C. The physiological significance of the modifications to the pigment composition induced by growth at sub-optimal temperature is discussed.     

The effect of acute high light and low temperature stresses on the ascorbate–glutathione cycle and superoxide dismutase activity in two Dunaliella salina strains

Dunaliella species accumulate carotenoids and their role in protection against photooxidative stress has been investigated extensively. By contrast, the role of other antioxidants in this alga, has received less attention. Therefore, the components of the ascorbate–glutathione cycle, along with superoxide dismutase (E.C. 1.15.1.1) and peroxidase (E.C. 1.11.1.11) activity were compared in two strains of Dunaliella salina. Strain IR‐1 had two‐fold higher chlorophyll and β‐carotene concentration than Gh‐U. IR‐1 had around four‐fold higher superoxide dismutase, ascorbate peroxidase and pyrogallol peroxidase activities than Gh‐U on a protein basis. Ascorbate and glutathione concentrations and redox state did not differ between strains and there was little difference in the activity of ascorbate–glutathione cycle enzymes (monodehydroascorbate reductase [E.C. 1.6.5.4], dehydroascorbate reductase [E.C. 1.8.5.1] and glutathione reductase [E.C. 1.8.1.7]). The response of these antioxidants to high light and low temperature was assessed by transferring cells from normal growth conditions (28°C, photon flux density of 100 μmol m^?2 s^?1)to 28°C/1200 μmol m^?2 s^?1; 13°C/100 μmol m^?2 s^?1; 13°C/1200 μmol m^?2 s^?1 and 28°C/100 μmol m^?2 s^?1 for 24 h. Low temperature and combined high light‐low temperature decreased chlorophyll and β‐carotene in both strains indicating that these treatments cause photooxidative stress. High light, low temperature and combined high light‐low temperature treatments increased the total ascorbate pool by 10–50% and the total glutathione pool by 20–100% with no consistent effect on their redox state. Activities of ascorbate–glutathione cycle enzymes were not greatly affected but all the treatments increased superoxide dismutase activity. It is concluded that D. salina can partially adjust to photooxidative conditions by increasing superoxide dismutase activity, ascorbate and glutathione.     

The relationship between respiration and temperature in leaves of the arctic plant Saxifraga cernua

Abstract Saxifraga cernua, a perennial herb distributed throughout the arctic and subarctic regions, shows high levels of dark respiration. The amount of respiration exhibited by leaves and whole plants at any temperature is influenced by the pretreatment temperature. Plants grown at 10°C typically show higher dark respiration rates than plants grown at 20°C. The levels of alternative-pathway respiration (or cyanide-insensitive respiration) in leaves of S. cernua grown at high and low temperatures were assessed by treating leaf discs with 0.25 mol m^?3 salicylhydroxamic acid during measurements of oxygen consumption. Alternative pathway respiration accounted for up to 75% of the total respiration. Tissues from 20°C-grown plants yielded a Q₁₀ of 3.37 for normal respiration, and of 0.97 for alternative-pathway respiration. Tissues from 10°C-grown plants yielded a Q₁₀ of 2.55 for normal respiration, and of 0.79 for alternative-pathway respiration. The alternative pathway does not appear to be as temperature sensitive as the normal cytochrome pathway. A simple energy model was used to predict the temperature gain expected from these high rates of alternative-pathway respiration. The model shows that less than 0.02°C can be gained by leaves experiencing these high respiration rates.     

Effects of changes in growth temperature on photosynthesis and carotenoid composition in Zea mays leaves

The effects of changes in growth temperature on photosynthesis and carotenoid composition were examined in Zea mays L. leaves of different age and different developmental history. The plants were first grown at sub-optimal temperature (14°C) until the full development of the third leaf. At that time, the mature third leaf and the immature fourth leaf had a low chlorophyll (Chl) content, a low Chl a/b ratio, a high carotenoid/Chl a+b ratio, a high xanthophyll/β-carotene ratio, and about 80% of the xanthophyll cycle pool (violaxanthin [V] + antheraxanthin [A] + zeaxanthin [Z]) was in the form of zeaxanthin and antheraxanthin. When the temperature was increased from 14°C to 24°C for three days, increased Chl synthesis, accompanied by an increase in the Chl a/b ratio, took place. The ratios of lutein, neoxanthin, and V+A+Z to Chl a+b decreased markedly, whereas no significant changes appeared in the β-carotene/Chl a+b ratio. Furthermore, there was a sharp decrease in the xanthophyll/β-carotene ratio and most of zeaxanthin was converted to violaxanthin in the xanthophyll cycle. The third leaf and the tip segment of the fourth leaf, both expanded at 14°C, showed little difference in their pigment contents. However, the rate of CO₂ assimilation of the tip segment of the fourth leaf was nearly twice that of the third leaf on the third day at 24°C, while the photosynthetic activity was similar in both leaves before the transfer to 24°C. During the warm period at 24°C, new leaf tissue (basal segment of the fourth leaf and part of a fifth leaf) was formed. On the third day at 24°C, the pigment content of 24°C-grown leaf tissue did not differ much from that of 14°C-grown leaf tissue with the exception that the total carotenoid content was lower in the former as compared to the latter, mainly because of a lower V+A+Z content. The rate of CO₂ assimilation of 24°C-grown leaf tissue was comparable to that of the tip segment of the fourth leaf. Regardless of which leaf tissue is considered, reducing the temperature from 24°C to 14°C for 5 days slightly affected the pigment content, but violaxanthin was largely converted to zeaxanthin and antheraxanthin in the xanthophyll cycle. The results indicate that compared to old leaf tissue of mature leaves, physiologically younger leaf tissue of immature leaves is much more able to recover from depressions in the photosynthetic activity induced by growth at sub-optimal temperature when the plants experience optimal growth temperatures, but that factors other than the pigment content must determine this capability.     

Exogenous glycine betaine and proline play a protective role in heat-stressed barley leaves (Hordeum vulgare L.): A chlorophyll a fluorescence study

Abstract

Effects of drought and exogenous glycine betaine and proline on Photosystem II (PSII) photochemistry were studied in barley leaves under heat stress induced by exposing them to 45°C for 10 min. Polyphasic fluorescence transient (OJIP) was used to evaluate PSII photochemistry in leaves treated with either glycine betaine or proline, combined or not with heat treatment. A distinct K step in the fluorescence transient OJIP appeared in control leaves, indicating an inactivation of the oxygen evolving complex (OEC). Drought stress and exogenous glycine betaine and proline modified the shape of the OJIP curve of leaves heated at 45°C and the K step was not as pronounced. Increased thermostability of PSII may be associated with the resistance of OEC and increased energy connectivity between PSII units. The thermostability of PSII was also reflected by a lower decrease in maximum quantum yield of primary photochemistry (?_Po = F _V/F _M) and performance index (PI). Exogenous application of glycine betaine or proline can play an important role in enhancing plant stress tolerance and may help reduce effects of environmental stresses.     

Effects of low temperature stress on the antioxidant system and photosynthetic apparatus of Kappaphycus alvarezii (Rhodophyta,Solieriaceae)

To understand the effects of low temperature stress on Kappaphycus alvarezii and the responses of antioxidant systems and photosystem II (PSII), behaviour in K. alvarezii thalli exposed to low temperatures (20°C, 17°C and 14°C) for 2 hours was evaluated. Compared with the control at 26°C, activities of some antioxidant enzymes including superoxide dismutase (SOD), peroxidase (POD), ascorbate peroxidase (APX) and the level of antioxidant substance (reduced glutathione) increased in K. alvarezii thalli when exposed to lowered temperatures (20°C, 17°C). Hydroxyl free radical (·OH) scavenging activity of K. alvarezii thalli also increased at 20°C and 17°C compared with the control. This indicated that the resistance to low temperature stress in the antioxidant system of K. alvarezii increased at lowered temperatures of 20°C and 17°C. However, at the lowest temperature (14°C), no significant increases of this algal antioxidant were observed. Under low temperature stress, the maximum quantum yield of PSII photochemistry (F_V/F_M) and PSII actual photochemical efficiency (Φ_PSII) decreased in K. alvarezii thalli, suggesting that the photosynthetic capacity declined. Components of the photosynthetic apparatus (such as the oxygen-evolving complex, light absorption antennas, reaction centres, electron acceptor sides and electron donor sides of PSII) were damaged by low temperature stress to varying degrees. In addition, it was found that low temperature stress led to decreases of both D1 protein and Rubisco LSU (Rubisco large subunit) protein levels. This work is a significant contribution towards understanding the basic mechanism involved in the resistance and the adaptation of K. alvarezii to low temperature stress conditions.     

Effect of protein modification by malondialdehyde on the interaction between the oxygen-evolving complex 33 kDa protein and photosystem II core proteins

Previously we observed that the oxygen-evolving complex 33 kDa protein (OEC33) which stabilizes the Mn cluster in photosystem II (PSII), was modified with malondialdehyde (MDA), an end-product of peroxidized polyunsaturated fatty acids, and the modification increased in heat-stressed plants (Yamauchi et al. 2008). In this study, we examined whether the modification of OEC33 with MDA affects its binding to the PSII complex and causes inactivation of the oxygen-evolving complex. Purified OEC33 and PSII membranes that had been removed of extrinsic proteins of the oxygen-evolving complex (PSII∆OEE) of spinach (Spinacia oleracea) were separately treated with MDA. The binding was diminished when both OEC33 and PSII∆OEE were modified, but when only OEC33 or PSII∆OEE was treated, the binding was not impaired. In the experiment using thylakoid membranes, release of OEC33 from PSII and corresponding loss of oxygen-evolving activity were observed when thylakoid membranes were treated with MDA at 40°C but not at 25°C. In spinach leaves treated at 40°C under light, maximal efficiency of PSII photochemistry (F _v/F _m ratio of chlorophyll fluorescence) and oxygen-evolving activity decreased. Simultaneously, MDA contents in heat-stressed leaves increased, and OEC33 and PSII core proteins including 47 and 43 kDa chlorophyll-binding proteins were modified with MDA. In contrast, these changes were to a lesser extent at 40°C in the dark. These results suggest that MDA modification of PSII proteins causes release of OEC33 from PSII and it is promoted in heat and oxidative conditions.     

Capacity for RNA synthesis in 70S ribosome-deficient plastids of heat-bleached rye leaves

In the leaves of rye seedlings (Secale cereale L.) grown at an elevated temperature of 32°C the formation of plastidic 70S ribosomes is specifically prevented. The resulting plastid ribosome-deficient leaves, which are chlorotic in light, represent a system for the identification of translation products of the 80S ribosomes among the chloroplastic proteins. Searching for the primary heat-sensitive event causing the 70S ribosome-deficiency, the thermostability of the chloroplastic capacity for RNA synthesis was investigated. The RNA polymerase activity of isolated normal chloroplasts from 22°-grown rye leaves was not inactivated in vitro at temperatures between 30° and 40°C. The ribosome-deficient plastids purified from bleached 32°-grown leaf parts contained significant RNA polymerase activity which was, however, lower than in functional chloroplasts. After application of [³H]uridine to intact leaf tissues [³H]uridine incorporation was found in ribosome-deficient plastids of 32°C-grown leaves. The amount of incorporation was similar to that in the control chloroplasts from 22°C-grown leaves. According to these results, it is unlikely that the non-permissive temperature (32°C) causes a general inactivation of the chloroplastic RNA synthesis in rye leaves.     

Effects of Cold-Hardening on Chilling-Induced Photoinhibition of Photosynthesis and on Xanthophyll Cycle Pigments in Sweet Pepper

Two cultivars of Capsicum annuum L. were acclimated for 5 d at sub-optimal temperature (14 °C) and irradiance of 250 µmol m^–2 s^–1. This cold-hardening resulted in some reduction in the extent of photoinhibition during an 8 h exposure to high irradiance at 4 °C. Obvious differences were observed between non-hardened leaves (NHL) and cold-hardened leaves (CHL) in the recovery under low irradiance at room temperature. The CHL of both cultivars recovered faster than NHL, especially during the initial fast phase of recovery. Compared with NHL, the total content of carotenoids (Cars), based on chlorophyll, Chl (a+b), and the proportions of xanthophyll cycle pigments referred to total Cars increased in CHL, mainly due to an increase of violaxanthin (V) + antheraxanthin (A) + zeaxanthin (Z) content per mol Chl (a+b). Faster development and a higher non-photochemical quenching (NPQ) of Chl fluorescence, related to a stronger deepoxidation of the larger xanthophyll cycle pool in NHL, could act as a major defence mechanism to reduce the formation of reactive oxygen species during severe chilling. This is suggested by higher content of Z or Z+A in photoinhibition as well as by its rapid decline during the initial fast phase of recovery. In contrast to the chilling-sensitive cv. 0004, the chilling-tolerant cv. 1141 did more easily acclimate its photosynthetic apparatus to low temperatures.     

Combined action of antioxidant defense system and osmolytes in chilling shock-induced chilling tolerance in Jatropha curcas seedlings

Low non-freezing temperature is one of the major environmental factors affecting growth, development and geographical distribution of chilling-sensitive plants, Jatropha curcas is considered as a sustainable energy plants with great potential for biodiesel production. In this study, chilling shock at 5 °C followed by recovery at 26 °C for 4 h significantly improved survival percentage of J. curcas seedlings under chilling stress at 1 °C. In addition, chilling shock could obviously enhance the activities of antioxidant enzymes superoxide dismutase (SOD), ascorbate peroxidase (APX), catalase (CAT) and glutathione reductase (GR), and the levels of antioxidants ascorbic acid (AsA) and glutathione (GSH), as well as the contents of osmolytes proline and betaine in leaves of seedlings of J. curcas compared with the control without chilling shock. During the process of recovery, GR activity, AsA, GSH, proline and betaine contents sequentially increased, whereas SOD, APX and CAT activities gradually decreased, but they markedly maintained higher activities than those of control. Under chilling stress, activities of SOD, APX, CAT, GR and GPX, and contents of AsA, GSH, proline and betaine, as well as the ratio of the reduced antioxidants to total antioxidants [AsA/(AsA + DHA) and GSH/(GSH + GSSG)] in the shocked and non-shock seedlings all dropped, but shocked seedlings sustained significantly higher antioxidant enzyme activity, antioxidant and osmolyte contents, as well as ratio of reduced antioxidants to total antioxidants from beginning to end compared with control. These results indicated that the chilling shock followed by recovery could improve chilling tolerance of seedlings in J. curcas, and antioxidant enzymes and osmolytes play important role in the acquisition of chilling tolerance.     

Acclimation of photosynthesis, H2O2 content and antioxidants in maize (Zea mays) grown at sub-optimal temperatures

Maize plants were grown at 14, 18 and 20 °C until the fourth leaf had emerged. Leaves from plants grown at 14 and 18 °C had less chlorophyll than those grown at 20 °C. Maximal extractable ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) activity was decreased at 14 °C compared with 20 °C, but the activation state was highest at 14 °C. Growth at 14 °C increased the abundance (but not the number) of Rubisco breakdown products. Phosphoenolpyruvate carboxylase (PEPC) activity was decreased at 14 °C compared with 20 °C but no chilling-dependent effects on the abundance of the PEPC protein were observed. Maximal extractable NADP-malate dehydrogenase activity increased at 14 °C compared with 20 °C whereas the glutathione pool was similar in leaves from plants grown at both temperatures. Foliar ascorbate and hydrogen peroxide were increased at 14 °C compared with 20 °C. The foliar hydrogen peroxide content was independent of irradiance at both growth temperatures. Plants grown at 14 °C had decreased rates of CO₂ fixation together with decreased quantum efficiencies of photosystem (PS) II in the light, although there was no photo-inhibition. Growth at 14 °C decreased the abundance of the D1 protein of PSII and the PSI psaB gene product but the psaA gene product was largely unaffected by growth at low temperatures. The relationships between the photosystems and the co-ordinate regulation of electron transport and CO₂ assimilation were maintained in plants grown at 14 °C.