The role of salicylic acid in response of two rice cultivars to chilling stress

Two rice (Oryza sativa L.) cultivars differing in chilling sensitivity, Changbaijiu (chilling-tolerant) and Zhongjian (chilling-sensitive) were pre-treated with 0.5, 1.0 and 2.0 mM salicylic acid (SA) for 24 h before chilling at 5°C for 1 d. Chilling induced SA accumulation, particularly conjugated SA in both leaves and roots of the two rice cultivars. After SA administration, SA accumulated in the roots of both cultivars at a concentration-dependent manner, whereas only a slight increase was observed in their leaves. Conjugated SA accounted for most of the increase. The beneficial effect of SA treatment on protecting rice seedlings from chilling injury was not observed at any concentration in either cultivar. Pre-treatment with SA even decreased their chilling tolerance confirmed by increased electrolyte leakage and lipid peroxidation. Further, most of the activities of antioxidant enzymes decreased or remained unchanged in leaves and roots of SA pre-treated seedlings after chilling. These results implied that down-regulation of antioxidant defence might be involved in the reduction of chilling tolerance in SA-pre-treated plants.     

Leaf photosynthetic parameters of two maize (Zea mays) cultivars in response to various patterns of chilling temperatures and photon flux densities

Chilling‐induced photosynthetic impairment was examined in leaves of maize (Zea mays L.) seedlings of two cultivars, one adapted to western Europe and one adapted to Mexican highlands. Three experiments were performed in a controlled environment. The effects of chilling night temperatures, of chilling at high light intensity and of variable chilling day temperatures on photosynthetic parameters, were evaluated. Chilling in the dark period resulted in stomatal limitation of net photosynthesis. Chilling at moderate to high light intensities caused chilling‐dependent photoinhibition of CO₂ uptake. Photobleached maize leaves did not resume normal photosynthetic function. Maize cv. Batan 8686 from the highlands of Mexico was less susceptible to photosynthetic damage than maize cv. Bastion adapted for cultivation in W. Europe, when exposed to chilling night temperatures, or to mild chilling photoinhibitory conditions.     

Low temperature acclimation to chilling tolerance in rice roots

Rice seedlings (Oryza sativa L.) were subjected to low temperature pretreatment (LT-PT; 10°C) for various length of time (1, 2, 4, 6, 12, 18, 24 h) followed by a 48-h chilling temperature stress (2°C). Chilling temperature tolerance of rice roots was improved with increasing duration of LT-PT, but LT-PT longer than 12 h gave no additional improvement. Alcohol dehydrogenase activity and ethanol concentration in the roots were increased with increasing duration of LT-PT up to 12 h. Chilling tolerance was also improved by exogenously applied ethanol. These results suggest that LT-PT may increase chilling tolerance in rice roots owing to ethanol accumulation in the roots and LT-PT acclimation to chilling temperature may occur within 12 h.     

Effect of paclobutrazol and chilling on leaf membrane lipids in cucumber seedlings

The effects of paclobutrazol on the leaf membrane lipid composition of seedlings of cucumber ( Cucumis sativus L. cv. Victory) subjected to chilling temperatures were assessed. At a non-injurious temperature (12.5°C), there was no difference in the polar lipid fatty acid composition or in the glycolipid, phospholipid or free sterol content of leaves from treated vs untreated seedlings, regardless of whether paclobutrazol was administered 1 or 7 days prior to analysis. In the latter case (7 days pretreatment), there were clear effects of the bioregulator on plant growth and morphology as well as on leaf chlorophyll content. At an injurious chilling temperature (5°C), desaturation of leaf polar lipid fatty acids was markedly reduced in both treated and untreated seedlings. Chilling at 5°C resulted in losses of fresh weight and membrane lipids in leaves of both groups of plants. These losses were either reversible or irreversible, depending upon the duration of chilling and of pretreatment with paclobutrazol. Seedlings pretreated with 10 μg ml⁻¹ paclobutrazol generally sustained less chilling injury than untreated controls, as judged by the extent of wilting, necrosis and desiccation. This correlated with reduced losses of leaf fresh Weight and membrane lipids.     

Association of polyamines in governing the chilling sensitivity of maize genotypes

Chilling stress is an important constraint of global production of maize. This study was undertaken to compare the chilling responses of different maize seedling tissues and to analyze changes in polyamines as a result of chilling stress. Reponses to chilling were characterized in two maize (Zea mays L.) inbred lines, ‘HuangC’ and ‘Mo17’, that putatively differ in chilling sensitivity. Seedlings were exposed to low temperature (5°C) and chilling injury was estimated by electrical conductivity (EC), malonaldehyde (MDA) concentration, and by changes in putrescine (Put), spermidine (Spd) and spermine (Spm) concentrations in root, mesocotyl, and coleoptile tissues. Membrane permeability (as measured by EC), MDA concentrations and Put concentrations in the three tissue of maize seedlings increased after chilling stress, except for the Put concentration in roots. Spd and Spm concentrations in the three tissues of seedlings decreased after chilling stress. The EC for cold stressed tissues were lower in HuangC than Mo17. Also, the EC of coleoptile tissues were lower than for mesocotyl in both inbred lines. We suggest that mesocotyl tissue can be used to evaluate cold tolerance in maize. Stepwise regression analyses showed that chilling injury in roots was generally correlated with Spd concentration while in the mesocotyl injury was mainly correlated with Put and Spd concentrations. Spermidine and Spm concentrations in the coleoptile were correlated with chilling injury. Characteristics changes of polyamines in chill-tolerant maize seedling combined with regression analysis are a reliable method for evaluating chill tolerance in maize lines.     

Reversibility of chilling injury to corn seedlings

Seedlings of corn (Zea mays) were tested for recovery from chilling injury incurred at 0.3 ± 0.3 C. At 0.3 C visual leaf injury appeared in 36 hours, whereas stem and root injuries appeared later. Appearance of leaf injury was preceded by a rise in O₂ uptake and a lessened effect of 2,4-dinitrophenol on O₂ uptake by leaf segments and was accompanied by increased ion leakage from the leaves. These effects were reversible, in that transfer of seedlings to 21 C after 36 hours at 0.3 C produced a return of O₂ uptake, 2,4-dinitrophenol stimulation, and ion leakage to the levels of unchilled leaves, as well as a disappearance of leaf symptoms, within 72 hours. For most seedlings, transfer to 21 C after 48 to 60 hours at 0.3 C reversed the chilling effects on O₂ uptake, 2,4-dinitrophenol stimulation, and injury symptoms but not on ion leakage within 108 hours. However, some seedlings collapsed during 48 to 60 hours of chilling, and these never recovered. Transfer to 21 C after 72 hours at 0.3 C did not produce recovery from any symptom of chilling injury examined, and these seedlings soon died. No growth occurred at 0.3 C, but growth began soon after transfer to 21 C. Seedlings chilled 24 or 36 hours grew at reduced rates during the first 72 hours at 21 C, but within 96 hours at 21 C were growing at the same rate as nonchilled seedlings. These results demonstrate considerable capacity of growing plants to recover from short chilling treatments even though significant physiological changes occurred at low temperatures.     

Chilling injury and recovery in detached and attached leaves measured by chlorophyll fluorescence

Smillie, R. M., Nott, R., Hetherington, S. E. and Öyustt, G. 1987. Chilling injury and recovery in detached and attached leaves measured by chlorophyll fluorescence Chilling injury was compared in detached and attached leaves chilled at 0 or 0.5°C by measuring the decrease in induced chlorophyll fluorescence in vivo. The fluorescence parameter measured was F_R, the maximal rate of rise of induced chlorophyll fluorescence emission after irradiating dark-adapted leaves. The plants used were bean, Phaseolus vulgaris L. cv. Pioneer, and maize, Zea mays L. cvs hybrid GH 390 and Northern Belle. Leaves were detached and placed on wet paper and covered with thin polyethylene film to prevent water loss during chilling. Leaves left attached on plants were treated similarly. When chilled in this way at 100% relative humidity, the chilling-induced decrease in F_R was the same in detached and attached leaves. For the attached leaves, the same result was obtained whether just a single leaf was chilled or the whole plant. Expression of chilling injury was greatest in fully turgid leaves and comparisons can be invalid unless the water status of the detached and attached leaves are the same. Problems arising from diurnal fluctuations in water potential of plants grown in a glasshouse were circumvented by placing leaves on the wet filter paper under polyethylene film prior to chilling, which allowed high water potentials to be regained, or mist sprays in the glasshouse were employed. Determinations of the time course for changes in F_R of maize (cv. Northern Belle) during chilling at 0°C showed that F_R decreased exponentially, at the same rate (time to 50% decrease in F_R was 9.3 h) in detached and attached leaves. Chilling injury was largely reversible for the first 20 h of chilling stress as both detached and attached leaves recovered their pre-chilling values of F_R after a further 20 h at 20°C in darkness. Leaves chilled for 48 h showed partial recovery, while those chilled for 72 h did not recover. Recovery was impeded by light. Inability to recover from chilling as indicated by measurements of F_R was paralleled by the incidence of visible symptoms of injury. It is concluded that detached and attached leaves behave similarly during chilling and short-term recovery, provided a similarity in treatments is rigorously maintained.     

Effects of chilling stress on the accumulation of soluble sugars and their key enzymes in <Emphasis Type="Italic">Jatropha curcas</Emphasis> seedlings

As osmolytes and signaling molecules, soluble sugars participate in the response and adaptation of plants to environmental stresses. In the present study, we measured the effect of chilling (12 °C) stress on the contents of eight soluble sugars in the leaves, cotyledons, stems, and roots of Jatropha curcas seedlings, as well as on the activities of eight rate-limiting enzymes that are critical to the metabolism of those soluble sugars. Chilling stress promoted both starch hydrolysis and soluble sugar accumulation. The soluble sugar contents of the leaves and cotyledons were affected more than that of the stems and roots. Meanwhile, the activities of the corresponding metabolic enzymes (e.g., β-amylase, uridine diphosphate glucose phosphorylase, and sucrose phosphate synthase) also increased in some organs. The gradual increase of soluble neutral alkaline invertase activity in the four studied organs suggested that sucrose catabolic production, such as glucose and fructose, was especially important in determining resistance to chilling stress and hexose signal transduction pathway. In addition, the substantial accumulation of raffinose family oligosaccharides and increase in corresponding metabolic enzyme activity suggested that galactinol and raffinose play an important role in determining the chilling resistance of J. curcas. Together, these findings establish a foundation for determining the relationship between the chilling resistance and soluble sugar accumulation of J. curcas and for investigating the mechanisms underlying sugar signaling transduction and stress responses.     

Frost resistance and water status of winter rape leaves as affected by differential shoot/root temperature

Twenty day-old winter rape ( Brassica napus L. var. oleifera L. cv. Jantar) seedlings, grown in nutrient solution, were exposed to different shoot/root temperature (i. e. 20/20, 20/3, 3/20 and 3/3°C) for 2 or 4 weeks. Chilling treatments modified markedly the pattern of plant growth as indicated by changes in dry matter accumulation in individual plant parts (leaves, hypocotyls, roots) and decreased leaf specific area. Growth of roots was less sensitive to low temperature than that of shoots. This was reflected by a decrease in shoot/root biomass ratio. Chilling treatments increased freezing resistance, decreased water content and water potential and modified reducing sugar, soluble protein and phospholipid contents in the leaves. A biphasic character of tissue responses to chilling temperature was observed, the most remarkable changes being registered during the first 7 or 14 days of the treatment. Effects of root or shoot exposure to chilling temperature on ice nucleation temperature, LT₅₀, water potential, accumulation of sugars and phospholipids in leaves were additive. All the observations point to the important role of the root system in plant acclimation to cold. Its impact on water status of leaves is emphasized and some mechanisms of root involvement in acclimation processes are proposed.     

Different effects of chilling on respiration in leaves and roots of cucumber (Cucumis sativus)

The effects of chilling on respiration (SHAM-resistant, cytochrome pathway and KCN-resistant, alternative pathway), temperature sensitivity, relative electrolyte conductivity, and degrees of oxidative stress (H(2)O(2) and malonaldehyde (MDA) contents) were separately examined in leaves and roots of cucumber (Cucumis sativus L.). After chilling at 8 degrees C for 4 days, both total respiration and KCN-resistant respiration in roots increased at different measurement temperatures. In contrast, SHAM-resistant respiration remained unchanged. In comparison, chilling significantly decreased the total respiration in leaves and this decrease was mostly due to a decrease in SHAM-resistant respiration. Chilling apparently decreased the sensitivity of KCN-resistant respiration to changes of temperature. The reduction levels of ubiquinone pool (UQr/UQt) increased both in chilled leaves and roots whilst pyruvate content increased only in chilled roots, but not in chilled leaves. Furthermore increases of H(2)O(2) and MDA contents were much greater in leaves than in roots. The same trend was also observed for ion leakage from tissues. Taken together, the results suggested that the higher chilling tolerance of roots was associated with their high total respiration and KCN-resistant respiration.     

Galactolipase activity but not the level of high-melting-point phosphatidylglycerol is related to chilling tolerance in differentially sensitive Zea mays inbred lines

Galactolipase activity, the level of high-melting-point phosphatidylglycerol (HMP-PG) as well as degradation of lipids during chilling and rewarming were studied in seedlings of maize inbred lines with different chilling responses. In aged chloroplasts of chilling-sensitive (CS) lines, galactolipase activity was considerably higher than that determined in aged chloroplasts isolated from chilling-tolerant (CT) ones. Chilling of seedlings at 5 °C for 6 days induced neither loss of chlorophyll content nor visible changes in the leaves, while a slight decline in total acyl lipid content by about 15.5% and 12.5% in CS and CT lines, respectively, was observed. Among total acyl lipids, only monogalactosyldiacylglycerol (MGDG) levels were decreased significantly upon chilling. Following return to the original growth conditions for 4 days, visible chilling injury in seedlings as well as essential differences in the decrease in total acyl lipids by about 53% and 20% in CS and CT lines, respectively, were found. These changes were accompanied by more extensive degradation of MGDG, digalactosyldiacylglycerol and phosphatidylglycerol in CS than in CT lines. As the levels of HMP-PG in fresh leaves were the same in all four lines of maize, it seems that galactolipase activity and not the level of HMP-PG is related to chilling response in maize. Received: 4 July 1997 / Revision received: 18 September 1997 / Accepted: 3 March 1998     

Heat shock-mediated APX gene expression and protection against chilling injury in rice seedlings

Rice (Oryza sativa L.) seedlings, when kept at 42 degrees C for 24 h before being kept at 5 degrees C for 7 d, did not develop chilling injury. Chilling resistance was enhanced in parallel with the period of heat-treatment. The level of APX activity was higher in heated seedlings whereas CAT activity was decreased by heat stress. There was no significant difference in SOD activity between heated and unheated seedlings. The elevated activity of APX was sustained after 7 d of chilling. The cytosolic APX gene expression in response to high and low temperature was analysed with an APXa gene probe. APXa mRNA levels increased within 1 h after seedlings were exposed to 42 degrees C. Elevated APXa mRNA levels could also be detected after 24 h of heating. The APXa mRNA level in preheated seedlings was still higher than unheated seedlings under cold stress. The promoter of the APXa gene was cloned from rice genomic DNA by TAIL-PCR, and characterized by DNA sequencing. The promoter had a minimal heat shock factor-binding motif, 5'-nGAAnnTTCn-3', located in the 81 bp upstream of the TATA box. Heat shock induction of the APXa gene could be a possible cause of reduced chilling injury in rice seedlings.     

低温处理葡萄根系对叶片PSII活性的影响

以美乐葡萄(Vitis vinifera cv. ‘Merlot’)幼苗为试材, 对叶片进行霜冻胁迫的同时控制土壤降温过程, 造成根系冷胁迫(2°C)和冻胁迫(0°C)。测定霜冻胁迫后和恢复期间叶片的快速叶绿素荧光参数, 并分析低温胁迫不同根系对叶片霜冻害程度的影响。结果表明, 根系在不同低温胁迫下会影响叶片对霜冻的反应, 根系冻胁迫造成叶片严重的霜冻伤害, 光系统II (PSII)反应中心活性难以得到恢复; 根系冷胁迫能避免叶片严重的低温伤害, 低温胁迫后PSII的活性也能很快恢复。     

Patterns of Assimilate Production and Translocation in Muskmelon (Cucumis melo L.) : II. Low Temperature Effects

Muskmelon (Cucumis melo L.) plants were exposed to a 10°C chilling treatment for 72 hours, which induced leaf chilling injury symptoms (wilting, appearance of water-soaked areas, necrosis). Chilling caused an accumulation of starch, sucrose, hexoses (glucose and fructose), and certain amino acids (glutamate, aspartate, and citrulline) in source leaf tissues, but no accumulation of stachyose or other galactosyl-oligosaccharides occurred. Chilling also caused a general increase in sugar (stachyose, raffinose, sucrose) and amino acid content of the phloem sap, although rates of phloem transport were apparently reduced. Pretreatment of the leaves with a 20-milligram per liter abscisic acid (ABA) spray before chilling prevented the appearance of chilling injury symptoms. ABA pretreatment had little or no affect on sugar accumulation in leaf tissues but greatly reduced or eliminated the chilling-induced amino acid accumulation. Higher levels of aspartate and particularly of arginine were found in phloem saps from ABA-pretreated plants. The data indicate that changes in leaf metabolism caused by environmental stresses such as chilling may change the composition of cucurbit phloem sap. This raises the possibility that some of the deleterious effects of stress on sink tissues may, in part, be due to alterations in the nature of the assimilate supply.     

Effects of seed hydropriming in presence of exogenous proline on chilling injury limitation in <Emphasis Type="Italic">Vigna radiata</Emphasis> L. seedlings

Three-day-old seedlings (t ₀ stage) of Vigna radiata (L.) Wilczek obtained from seeds hydroprimed (H) and hydroprimed with proline (HPro) were examined. H and HPro slightly improved mung bean seed germination and seedlings growth at 5°C. The best growth was observed in the seedlings obtain from HPro5 (5 mM) seeds in comparison with the seedlings obtained from the control-non-primed seeds and H seeds. Exposure of mung bean seedlings grown from non-primed seeds to chilling for 4 days induced chilling injury: membrane lipid peroxidation, decrease in endogenous proline level and inhibition of growth of roots and hypocotyls. The seedlings obtain from HPro seeds grew better during the time of chilling and after rewarming at 25°C. The possible role of HPro in chilling injury limitation is discussed.     

Chilling injury and changes in adenosine triphosphate of cotton seedlings

Young Gossypium hirsutum L. seedlings chilled at 5° showed a continual decrease in ATP concentration with time of chilling. Chilled plants returned to optimum conditions were able to restore the initial ATP concentration when chilled only 1 day, but not when chilled 2 days. The decrease in ATP with chilling was prevented by hardening the seedlings at 15° for 2 days (14-hr-day-length) immediately before chilling. The ATP level of hardened plants was higher than of unhardened plants. When hardened plants were chilled at 5°, the ATP level increased in the leaves but decreased in the roots.     

A comparative study of low-temperature-induced ultrastructural alterations of three species with differing chilling sensitivities

Abstract. The effects of light and water stress upon chilling injury of chloroplasts have been assessed by electron microscopy in seedlings of three species known to differ in their chilling susceptibility. Chilling injury to chloroplasts was first manifested by distortion and swelling of thylakoids, reduction in starch granule size, and the formation of small vesicles of the envelope, called the peripheral reticulum. More prolonged treatment produced accumulations of lipid droplets, increased staining of the stroma, disintegration of the envelope, and mixing with cytoplasmic contents. Cotton, a notably chilling-sensitive plant, and bush bean, a somewhat less sensitive plant, showed damage within 6 h when exposed to both light and water stress at chilling temperatures (5°C). Even collard, a chilling-resistant species, exhibited signs of chilling injury to chloroplasts after 6 h when exposed to both light and water stress but the plastids remained intact throughout the 48 h of treatment. Comparable chilling injury does not occur in cotton until around 72 h if the plants are exposed to water stress or light separately. Bush bean was affected less by separate treatments of light and water stress. The least chilling injury occurred in all three species when they were kept in the dark at a high humidity.     

Inhibition of glutathione synthesis reduces chilling tolerance in maize

 The role of glutathione (GSH) in protecting plants from chilling injury was analyzed in seedlings of a chilling-tolerant maize (Zea mays L.) genotype using buthionine sulfoximine (BSO), a specific inhibitor of γ-glutamylcysteine (γEC) synthetase, the first enzyme of GSH synthesis. At 25 °C, 1 mM BSO significantly increased cysteine and reduced GSH content and GSH reductase (GR: EC 1.6.4.2) activity, but interestingly affected neither fresh weight nor dry weight nor relative injury. Application of BSO up to 1 mM during chilling at 5 °C reduced the fresh and dry weights of shoots and roots and increased relative injury from 10 to almost 40%. Buthionine sulfoximine also induced a decrease in GR activity of 90 and 40% in roots and shoots, respectively. Addition of GSH or γEC together with BSO to the nutrient solution protected the seedlings from the BSO effect by increasing the levels of GSH and GR activity in roots and shoots. During chilling, the level of abscisic acid increased both in controls and BSO-treated seedlings and decreased after chilling in roots and shoots of the controls and in the roots of BSO-treated seedlings, but increased in their shoots. Taken together, our results show that BSO did not reduce chilling tolerance of the maize genotype analyzed by inhibiting abscisic acid accumulation but by establishing a low level of GSH, which also induced a decrease in GR activity. Received: 9 November 1999 / Accepted: 17 February 2000     

Influence of Chilling Stress on the Intercellular Distribution of Assimilatory Sulfate Reduction and Thiols in Zea mays

Abstract: The effect of chilling on the intercellular distribution of mRNAs for enzymes of assimilatory sulfate reduction, the activity of adenosine 5'-phosphosulfate reductase (APR), and the level of glutathione was analysed in leaves and roots of maize ( Zea mays L). At 25 °C the mRNAs for APR, ATP sulfurylase, and sulfite reductase accumulated in bundle-sheath only, whereas the mRNA for O-acetylserine sulfhydrylase was also detected in mesophyll cells. Glutathione was predominantly detected in mesophyll cells; however, oxidized glutathione was equally distributed between the two cell types. Chilling at 12 °C induced oxidative stress which resulted in increased concentrations of oxidized glutathione in both cell types and a prominent increase of APR mRNA and activity in bundle-sheath cells. After chilling, mRNAs for APR and sulfite reductase, as well as low APR activity, were detected in mesophyll cells. In roots, APR mRNA and activity were at higher levels in root tips than in the mature root and were greatly increased after chilling. These results demonstrate that chilling stress affected the levels and the intercellular distribution of mRNAs for enzymes of sulfate assimilation.     

A role for young leaves in vernalization of cauliflower: I. Analysis of leaf development during curd induction

Growth analysis techniques are used to test the hypothesis that chilling induces curd (flower) initiation in the cauliflower ( Brassica oleracea Botrytis L. cv. Perfection) through inhibiting leaf growth, thereby increasing the availability of growth factors to the stem apex and enabling differentiation of the curd. Effects of chilling on leaf growth and curd induction are compared in juvenile and mature, vegetative plants. Chilling at 5°C reduced dry matter accumulation in the total leaf complement by ca 60% in juvenile plants and 40% in mature plants, compared to control plants growth at 20°C. Juvenile plants showed slower rates of leaf initiation than mature plants. Leaf initiation was retarded by chilling in both plant types with the most marked effect seen in the juvenile plants. This was consistent with dry matter availability to the stem apex limiting differentiation more severely in juvenile plants than in mature plants. The rate of dry matter accumulation in existing leaves, however, was faster in juvenile plants than in mature plants at 20°C. Plants that were juvenile during chilling produced an average of 43 leaves below the curd whereas those that were mature produced 25.
Dry matter accumulation in younger leaves was more markedly inhibited by chilling than in older leaves. Chilling also reduced the rate at which enlarging leaves became positionally more remote from the stem apex. Possible roles for such leaves in regulating apical development are considered.