Chilling tolerance of cucumber (Cucumis sativus) seedling radicles is affected by radicle length, seedling vigor, and induced osmotic- and heat-shock proteins

Cucumber seedling radicles decrease in chilling tolerance as they increase in length or decrease in vigor. The protein content of the apical 5 mm of the radicle decreased with decreases in chilling tolerance ( R ² = 0.92). This general reduction in protein content was reflected in a decrease of six dehydrin-like proteins with apparent molecular weights of 13.0, 15.0, 16.8, 23.0, 26.8, and 33.5 kDa. The disappearance of naturally occurring dehydrin-like proteins in cucumber seedling radicles as they elongate or lose vigor was correlated with a loss of chilling tolerance. Exposure to an osmotic (0.6 M mannitol) or heat (2 min at 45°C) stress enhanced chilling tolerance. The osmotic-shock treatment induced both chilling tolerance and the appearance or strengthening of dehydrin-like proteins previously present in radicles. The heat-shock treatment also induced high levels of chilling tolerance and protein(s) that reacted with a 23 and 70 kDa antibody. However, these heat-shock protein (HSPs) did not cross react with the probe for dehydrin-like proteins. When organized into high, medium, and low chilling tolerance groups, radicle that were chilling tolerant contained either the 13.0 and 16.8 kDa dehydrin-like proteins, or the 15.0 and 23.0 kDa dehydrin-like proteins, or the 23 or 70 kDa HSP.     

Reduced chilling tolerance in elongating cucumber seedling radicles is related to their reduced antioxidant enzyme and DPPH-radical scavenging activity

Cucumber seedling radicles become more chilling sensitive as they elongate. Chilling seedlings with radicles 20 mm long for 48 h at 2.5°C inhibited subsequent growth by 36%, while it reduced the growth of 70 mm-long radicles by 63%. Although the growth rate of non-chilled cucumber radicles at 25°C is constant from 20 to 80 mm, tissue viability [i.e. reduction of TTC (2,3,5-triphenyltetrazolium chloride) to formazan] and DPPH ( α,α -diphenyl- β -picrylhydrazyl) radical scavenging activity of apical tissue declines as radicles elongate from 20 to 80 mm in length. TTC reduction, DPPH-radical scavenging activity and protein content of apical tissue were higher in 20 than in 70 mm radicles immediately after chilling and after an additional 48 h of growth at 25°C. Catalase (CAT; EC 1.11.1.6) and ascorbate peroxidase (APX; EC 1.11.1.11) activity was higher in the apical tissue of 20 than in 70 mm radicles before chilling. Immediately after chilling and after an additional 48 h at 25°C, superoxide dismutase (SOD; EC 1.15.1.1), glutathione reductase (GR; EC 1.6.4.2), and guaiacol peroxidase (GPX; EC 1.11.1.7) activity increased more rapidly in 70 mm radicles than in 20 mm radicles (SOD, GR, and GPX activity in 70 mm radicles was 1.5-, 1.9- and 8.6-fold higher, respectively, than in 20 mm radicles). However, APX and CAT activity in 20 mm radicles were always higher than in 70 mm radicles. Growth after chilling enhanced the activity of all antioxidant enzymes compared to that found in non-chilled tissue; however, CAT activity in 70 mm radicles did not recover to levels found in non-chilled tissue. Higher levels of CAT, APX and DPPH-radical scavenging activity are correlated with higher chilling tolerance of 20 mm-long cucumber radicles compared to 70 mm-long radicles.     

Effect of chilling on antioxidant enzymes and DPPH-radical scavenging activity of high- and low-vigour cucumber seedling radicles

The chilling tolerance of cucumber seedling radicles was influenced by their relative levels of vigour. Radicles of high‐vigour seedlings grew to 20 mm in length in 36 h at 25 °C, whereas it took 60 h for low‐vigour seedling radicles to reach that length. Chilling at 2·5 °C for 48 h inhibited the subsequent growth of high‐ and low‐vigour seedlings by 39 and 68%, respectively. The 2,3,5‐triphenyltetrazolium chloride (TTC) viability index, and α,α‐diphenyl‐β‐picrylhydrazyl (DPPH)‐radical scavenging activity were higher in high than low‐vigour radicles. Higher ascorbate peroxidase (APX) and catalase (CAT) enzyme activity, DPPH‐radical scavenging activity, and recovery of CAT activity after chilling in high‐vigour radicles corresponded with their higher level of chilling tolerance in comparison with low‐vigour radicles. In contrast, elevated levels of superoxide dismutase, glutathione reductase and guaiacol peroxidase appear to be correlated with chilling injury since they only showed substantial increases in activity in the more chilling‐­sensitive low‐vigour radicles after chilling. Manipulation of APX, CAT, and/or DPPH activity could produce plants with superior and persistent chilling tolerance.     

Effect of acetaldehyde, arsenite, ethanol, and heat shock on protein synthesis and chilling sensitivity of cucumber radicles

Exposure to a chilling temperature of 2.5°C for 96 h inhibited the subsequent growth of cucumber seedling radicles at 25°C by 92%. Exposing seedling with 5 ± 1 mm long radicles to acetaldehyde vapour (275 µl l⁻¹) or to an aqueous ethanol solution (0.6  M ) for 2 h, or to 45°C for 10 min before chilling, increased chilling tolerance so that the chilling treatment reduced growth by only 47, 39 or 36%, respectively. All of these effective treatments induced the synthesis of a number of proteins, and suppressed de novo protein synthesis (i.e. the incorporation of [³⁵S]-methionine) by about 70%. In contrast, treatment for 2 h with an aqueous arsenite solution (100 µ M ) had no effect on chilling sensitivity or the incorporation of [³⁵S]-methionine, yet it induced the synthesis of a complement of proteins that were similar to that induced by the effective heat-shock treatment. A unique protein or set of proteins may be responsible for heat-shock-induced chilling tolerance, but none was detected. The ability of various abiotic stresses to suppress protein synthesis may be more important in increasing tolerance to chilling injury than their ability to induce the synthesis of specific proteins.     

Effect of chilling, heat shock, and vigor on the growth of cucumber (Cucumis sativus) radicles

Chilling at 2.5°C reduced the subsequent growth of cucumber ( Cucumis sativus L.) radicles at 25°C. The reduction in radicle growth was linear for 1–3 days of chilling at ≈10% per day of treatment, but then it increased in a non-linear pattern until subsequent radicle growth was all but eliminated by 6 days of chilling. A heat shock of 40°C for 4–12 min increased chilling tolerance such that 4 days of chilling caused only a 36% decrease in radicle growth, compared to 66% for seedlings not heat shocked. Heat shocks were only able to protect that part of radicle growth that was in excess of the linear decrease in radicle growth projected from 0–3 days. There appear to be two effects of chilling on radicle growth. The first inhibition of subsequent growth was linear and was not affected by heat shocks. The second inhibition was much more severe; it appeared after 3 days of chilling and could be prevented by heat shock. Seeds classified with different levels of vigor (i.e., different initial rates of growth) did not respond significantly different to chilling stresses following heat-shock treatments.     

Relationship between Proline and Abscisic Acid in the Induction of Chilling Tolerance in Maize Suspension-Cultured Cells

Both proline and abscisic acid (ABA) induce chilling tolerance in chilling-sensitive plants. However, the relationship between proline and ABA in the induction of chilling tolerance is unclear. We compared the time course of the increase in chilling tolerance induced by proline and ABA, and the time course of the uptake of both into the cultured cells of maize (Zea mays L. cv Black Mexican Sweet) at 28[deg]C. The plateau of proline-induced chilling tolerance preceded by 12 h the plateau of ABA-induced chilling tolerance. The uptake of exogenous ABA into the cells reached a plateau in 1 h, whereas the uptake of exogenous proline gradually increased throughout the 24-h culture period. Although the proline content in ABA-treated cells was 2-fold higher than in untreated cells at the end of the 24-h ABA treatment at 28[deg]C, the correlation between the endogenous free proline content and the chilling tolerance in the ABA-treated cells was insignificant. Isobutyric acid treatment, which resulted in a larger accumulation of proline in the cells than ABA treatment, did not increase chilling tolerance. The induction of chilling tolerance by proline and ABA appeared to be additive. Cycloheximide inhibited ABA-induced chilling tolerance, but it did not inhibit proline-induced chilling tolerance. Newly synthesized proteins accumulate in ABA-treated cells at 28[deg]C while the chilling tolerance is developing (Z. Xin and P.H. Li [1993] Plant Physiol 101: 277-284), but none of these proteins were observed in the proline-treated cells. Results suggest that proline and ABA induce chilling tolerance in maize cultured cells by different mechanisms.     

复合外源物质对玉米幼苗抗寒性的影响

该研究采用不同浓度的水杨酸(SA)、脱落酸(ABA)、磷酸二氢钾(KH2PO4)以及抗坏血酸(VC)进行正交组合获得不同浓度组合的复合外源物质,同时以蒸馏水处理作为对照,将其喷施于玉米幼苗后进行低温胁迫处理,在胁迫结束后使幼苗恢复生长,并测定叶片相关生长指标及生理生化指标的变化。结果表明:(1)与对照组相比,9种复合外源物质处理均可显著提高玉米幼苗的相对生长速率、干物质积累速率及脯氨酸含量,降低相对电导率,总体上提高了玉米幼苗的素质。(2)在4℃低温胁迫下,9种复合外源物质处理下幼苗相对生长速率、干物质积累速率、根系活力、脯氨酸含量、可溶性蛋白含量及可溶性糖含量,均显著高于对照,并减缓了丙二醛的积累和相应的膜脂过氧化,降低了相对电导率,提高了玉米幼苗的耐冷性。低温胁迫条件下,水杨酸、脱落酸、磷酸二氢钾和抗坏血酸四种物质组成的复合物可以提高玉米幼苗的耐冷性,其中以SA0.14 g·L~(-1)+ABA 0.015 g·L~(-1)+KH_2PO_4 3.0 g·L~(-1)+V_C 3.0 g·L~(-1)的复合物效果最好。该研究结果为新型复合抗寒剂的应用推广提供了技术支持。     

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.     

Metabolic dysfunction and unabated respiration precede the loss of membrane integrity during dehydration of germinating radicles

This study shows that dehydration induces imbalanced metabolism before loss of membrane integrity in desiccation-sensitive germinated radicles. Using a photoacoustic detection system, responses of CO(2) emission and fermentation to drying were analyzed non-invasively in desiccation-tolerant and -intolerant radicles of cucumber (Cucumis sativa) and pea (Pisum sativum). Survival after drying and a membrane integrity assay showed that desiccation tolerance was present during early imbibition and lost in germinated radicles. However, tolerance could be re-induced in germinated cucumber radicles by incubation in polyethylene glycol before drying. Tolerant and polyethylene glycol (PEG)-induced tolerant radicles exhibited a much-reduced CO(2) production before dehydration compared with desiccation-sensitive radicles. This difference was maintained during dehydration. In desiccation-sensitive tissues, dehydration induced an increase in the emission of acetaldehyde and ethanol that peaked well before the loss of membrane integrity. Acetaldehyde emission from sensitive radicles was significantly reduced when dehydration occurred in 50% O(2) instead of air. Acetaldehyde/ethanol were not detected in dehydrating tolerant radicles of either species or in polyethylene glycol-induced tolerant cucumber radicles. Thus, a balance between down-regulation of metabolism during drying and O(2) availability appears to be associated with desiccation tolerance. Using Fourier transform infrared spectroscopy, acetaldehyde was found to disturb the phase behavior of phospholipid vesicles, suggesting that the products resulting from imbalanced metabolism in seeds may aggravate membrane damage induced by dehydration.     

Activity of enzymatic antioxidant defense systems in chilled and heat shocked cucumber seedling radicles

Chilling whole cucumber seedlings that had 10‐mm long radicles for 4 days at 2.5°C significantly inhibited subsequent radicle growth both by increasing the time it took the seedlings to recover from chilling and attain a linear rate of radicle growth, and by decreasing the subsequent rate of linear growth. Exposing cucumber seedlings to 45°C for up to 20 min had no effect on subsequent radicle growth, while longer exposures produced reductions in growth. A heat shock at 45°C for 10 min induced the optimal protection to 4 days of chilling at 2.5°C by reducing chilling inhibition from 60 to 42%. Two hours after being chilled, heat shocked or heat shocked and then chilled, there was no difference in protein content of the apical 1 cm of the seedling radicle among these treatments and the non‐heat shocked, non‐chilled control. Two days after treatment, the protein content was still similar in tissue that had been heat shocked or heat shocked and chilled, while it was significantly reduced in tissue that had been chilled. In general, 2 h after treatment, the activity of the 5 antioxidant enzymes examined in this study [superoxide dismutase (SOD; EC 1.15.1.1), catalase (CAT; EC 1.11.1.6), ascorbate peroxidase (APX; EC 1.11.1.11), guaiacol peroxidase (GPX; EC 1.11.1.7) and glutathione reductase (GR; EC 1.6.4.2)] were reduced by chilling and unaffected or increased by heat shock. When heat shock was followed by chilling, there was a consistent effect of the heat shock treatment on preventing the loss of enzyme activity following chilling. This protective effect of the heat shock treatment was even more pronounced after 2 days of recovery at 25°C for SOD, CAT and APX. In contrast, the activity of GR and GPX was substantially higher in chilled tissue than in tissue that had been heat shocked before being chilled. Elevated levels of GR and GPX therefore appear to be correlated with the development of chilling injury, while elevated levels of SOD, CAT and APX appear to be correlated with the development of heat shock‐induced chilling tolerance.     

Impact of salicylic acid on the antioxidant enzyme system and hydrogen peroxide production in Cucumis sativus under chilling stress

Salicylic acid (SA) is a naturally produced compound and has been implicated to play important roles in defense of plants against diverse biotic and abiotic stresses. To understand how SA functions in the tolerance of cucumber (Cucumis sativus) to chilling stress, endogenous SA levels in two different cultivars with opposite chilling responsiveness were quantified. Membrane integrity, including malondialdehyde (MDA) content and leakage of electrolyte, was also examined in SA-pretreated cucumber plants under chilling conditions. In addition, activities of the two antioxidant enzymes peroxidase (POD) and catalase (CAT) were quantified, and hydrogen peroxide (H2O2) production was investigated histochemically in SA-treated leaves under chilling temperature. Chilling stress resulted in greater induction of SA levels in the chilling-tolerant cultivar Changchun mici in both leaves and seeds compared to the chilling-sensitive one Beijing jietou, while the former one contained higher levels of SA than the latter one in the seeds under normal conditions. Pretreatment with SA diminished the increased electrolyte leakage and MDA content caused by chilling in the leaves of both cultivars, while much less MDA and electrolyte leakage were produced in Changchun mici compared to Beijing jietou. Moreover, exogenous application of SA increased significantly the POD and CAT activities and soluble protein content. Most importantly, exogenous SA treatment could eliminate the accumulation of H202 in leaves and cotyledons of both cultivars caused by chilling stress. The data clearly demonstrated that the chilling-tolerant cultivar displays a higher SA level than the chilling-sensitive one, and that exogenous SA can enhance the chilling tolerance ability, which might be achieved through modulating the antioxidant system in cucumber.     

Endogenous salicylic acid accumulation is required for chilling tolerance in cucumber (Cucumis sativus L.) seedlings

Salicylic acid (SA) is an important plant hormone, and its exogenous application can induce tolerance to multiple environmental stresses in plants. In this study, we examine the potential involvement of endogenous SA in response to chilling in cucumber (Cucumis sativus L.) seedlings. A low temperature of 8 °C induces a moderate increase in endogenous SA levels. Chilling stimulates the enzymatic activities and the expression of genes for phenylalanine ammonia-lyase (PAL) and benzoic acid-2-hydroxylase rather than isochorismate synthase. This indicates that the PAL enzymatic pathway contributes to chilling-induced SA production. Cucumber seedlings pretreated with SA biosynthesis inhibitors accumulate less endogenous SA and suffer more from chilling damage. The expression of cold-responsive genes is also repressed by SA inhibitors. The reduction in stress tolerance and in gene expression can be restored by the exogenous application of SA, confirming the critical roles of SA in chilling responses in cucumber seedlings. Furthermore, the inhibition of SA biosynthesis under chilling stress results in a prolonged and enhanced hydrogen peroxide (H₂O₂) accumulation. The application of exogenous SA and the chemical scavenger of H₂O₂ reduces the excess H₂O₂ and alleviates chilling injury. In contrast, the protective effects of SA are negated by foliar spraying with high concentrations of H₂O₂ and an inhibitor of the antioxidant enzyme. These results suggest that endogenous SA is required in response to chilling stress in cucumber seedlings, by modulating the expression of cold-responsive genes and the precise induction of cellular H₂O₂ levels.     

Oxygen-mediated cold-acclimation in cucumber (Cucumis sativus) seedlings

Cold acclimation of etiolated cucumber seedlings, consisting of cooling at 12°C for 48 h followed by a warming period at 25°C, led to tolerance to subsequent chilling at 2°C. Tolerance, as evidenced by freedom from chilling injury and continued growth, developed during the warming period in a time-course manner for 12 h but decreased with prolonged warming. A similar increase and subsequent decrease was also observed in the content of palmitic, linoleic and linolenic acids in total lipid fraction from cucumber hypocotyl tissue. During the warming period supra-ambient oxygen stimulated, whereas subambient oxygen inhibited, the increase in fatty acid content as well as development of chilling tolerance. A strong correlation between oxygen-mediated changes in fatty acid content and associated development of cold tolerance suggests that both these processes are interrelated. Cold acclimation, but not cold stress, led to an increase followed by a decrease in CO₂ evolution suggesting that a respiratory upsurge is yet another feature of cold acclimation in cucumbers.     

Effects of Cold Acclimation and Salicylic Acid on Changes in ACC and MACC Contents in Maize during Chilling

The effect of 0.5 mM salicylic acid (SA) pretreatment and of growing at hardening temperatures on chilling-induced changes in 1-aminocyclopropane-1-carboxylic acid (ACC) and malonyl 1-aminocyclopropane-1-carboxylic acid (MACC) was investigated in young maize (Zea mays L.) plants grown in hydroponic solution at 22/20 °C. Chilling at 5 °C caused an increase in ACC content;however, this increase was less pronounced in plants cold acclimated at 13/11 °C 4 d before the chilling treatment, and in those which were pretreated with SA for 1 d before the cold stress. Changes in MACC at low temperature showed no correlation with chilling tolerance in maize.     

Accumulation of gentisic acid as associated with systemic infections but not with the hypersensitive response in plant-pathogen interactions

In the present work we have studied the accumulation of gentisic acid (2,5-dihydroxybenzoic acid, a metabolic derivative of salicylic acid, SA) in the plant-pathogen systems, Cucumis sativus and Gynura aurantiaca, infected with either prunus necrotic ringspot virus (PNRSV) or the exocortis viroid (CEVd), respectively. Both pathogens produced systemic infections and accumulated large amounts of the intermediary signal molecule gentisic acid as ascertained by electrospray ionization mass spectrometry (ESI-MS) coupled on line with high performance liquid chromatography (HPLC). The compound was found mostly in a conjugated (β-glucoside) form. Gentisic acid has also been found to accumulate (although at lower levels) in cucumber inoculated with low doses of Pseudomonas syringae pv. tomato, producing a nonnecrotic reaction. In contrast, when cucumber was inoculated with high doses of this pathogen, a hypersensitive reaction occurred, but no gentisic-acid signal was induced. This is consistent with our results supporting the idea that gentisic-acid signaling may be restricted to nonnecrotizing reactions of the host plant (Bellés et al. in Mol Plant-Microbe Interact 12:227–235, 1999). In cucumber and Gynura plants, the activity of gentisic acid as inducing signal was different to that of SA, thus confirming the data found for tomato. Exogenously supplied gentisic acid was able to induce peroxidase activity in both Gynura and cucumber plants in a similar way as SA or pathogens. However, gentisic-acid treatments strongly induced polyphenol oxidase activity in cucumber, whereas pathogen infection or SA treatment resulted in a lower induction of this enzyme. Nevertheless, gentisic acid did not induce other defensive proteins which are induced by SA in these plants. This indicates that gentisic acid could act as an additional signal to SA for the activation of plant defenses in cucumber and Gynura plants.     

Participation of H2O2 in Enhancement of Cold Chilling by Salicylic Acid in Banana Seedlings

The possible physiological mechanism of enhancement of cold tolerance by salicylic acid (SA) in banana seedlings (Musa acuminata cv. Williams 8188) was explored. Measurements of leakage electrolyte after 2 d of recovery at 30/22 ℃ (day/night) following 3 d of cold stress at 7 ℃ showed that pretreatment with hydroponic solution containing SA 0.3－0.9 mmol/L as foliar spray under normal growth conditions (30/22 ℃) could significantly enhance cold tolerance of banana plants. The highest enhancing effect of SA occurred at 0.5 mmol/L and it showed the lowest leakage rate of electrolyte or smaller leaf wilting area after 2 d of recovery at normal temperature from 3 d of 7 ℃ or 5 ℃ cold stress. Higher concentrations (≥2.5 mmol/L) of SA, however, caused more electrolyte leakage, indicating that they aggravated chilling damage. Enhanced cold tolerance by SA could be related to H2O2 metabolism. Compared with water-treated seedlings (control), SA 0.5 mmol/L treatment inhibited activities of catalase (CAT) and ascorbate peroxidase (APX), increased peroxidase (POX) activity, but did not affect the activity of superoxide dismutase (SOD) under normal growth conditions, and these changes might lead to an accumulation of H2O2, whereas SA pretreatment enhanced the activities of CAT and APX, and reduced the increase in productions of H2O2 and thiobarbituric acid-reaction substances (TBARS) during subsequent 7 ℃ cold stress and recovery periods. Exogenous H2O2 treatments (1.5－2.5 mmol/L) also increased cold tolerance of banana seedlings. Furthermore, pretreatment of banana seedlings with dimethylthiourea (a trap for H2O2) significantly inhibited cold tolerance induced by SA. These results suggested that endogenous H2O2 may be required for SA-enhanced cold tolerance. The significance of the interaction of SA, H2O2 and H2O2-metabolizing enzymes during cold stress has been discussed.     

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     

Synergistic effect of antioxidant system and osmolyte in hydrogen sulfide and salicylic acid crosstalk-induced heat tolerance in maize (Zea mays L.) seedlings

Salicylic acid (SA), is a plant hormone with multifunction that is involved in plant growth, development and the acquisition of stress tolerance. Hydrogen sulfide (H₂S) is emerging similar functions, but crosstalk between SA and H₂S in the acquisition of heat tolerance is not clear. Our recent study firstly reported that SA treatment enhanced the activity of L-cysteine desulfhydrase (L-DES), a key enzyme in H₂S biosynthesis, followed by induced endogenous H₂S accumulation, which in turn improved the heat tolerance of maize seedlings.¹ In addition, NaHS, a H₂S donor, enhanced SA-induced heat tolerance, while its biosynthesis inhibitor DL-propargylglycine (PAG) and scavenger hydroxylamine (HT) weakened SA-induced heat tolerance. Also, NaHS had no significant effect on SA accumulation and its biosynthesis enzymes phenylalanine ammonia lyase (PAL) and benzoic-acid-2-hydroxylase (BA2H) activities, as well as significant difference was not observed in NaHS-induced heat tolerance of maize seedlings by SA biosynthesis inhibitors paclobutrazol (PAC) and 2-aminoindan-2-phosph- onic acid (AIP) treatment.¹ Further study displayed that SA induced osmolytes (proline, betaine and trehalose) accumulation and enhancement in activity of antioxidant system in maize seedlings. These results showed that antioxidant system and osmolyte play a synergistic role in SA and H₂S crosstalk-induced heat tolerance of maize seedlings.     

Endoribonuclease of Vicia faba ssp. minor radicles, and enhancement of its activity by chilling stress or abscisic acid

Endoribonuclease in roots of 3-day-old seedlings of Vicia faba L. ssp, minor is a citrate activated glycoprotein of 35 kDa, with pH and temperature optima of 6.0 and 50°C. The same holds for endoribonuclease from seedlings treated with abscisic acid or exposed to a chilling stress of -3°C for 24 h, except that the temperature optimum was decreased to 40°C. The enzyme(s) preferentially hydrolysed poly (A) and poly (U). RNase activity in the radicles was enhanced by chilling stress or abscisic acid. ABA did not potentiate the effect brought about by chilling but slowed down the decrease in RNase activity in chill-stressed seedlings upon transfer to 25°C. Both factors modified the pattern of the isoelectric-points of the molecular forms of RNase.     

Induction of chilling tolerance in cucumber (Cucumis sativus) seedlings by endogenous and applied ethanol

Five-day-old etiolated cucumber ( Cucumis sativus L.) seedlings cv. Marketmore held at 2°C for 72 h developed chilling injury, resulting in desiccation and collapse of the hypocotyl tissue and eventual plant death. Hypoxia-induced accumulation of ethanol and acetaldehyde led to tolerance of subsequent chilling, as evidenced by continued hypocotyl growth and freedom from injury. Attenuated accumulation of volatiles by applied bisulfite reduced the development of hypoxia-induced chilling tolerance in seedlings. In seedlings held in normoxia cold tolerance was induced by applied ethanol vapors, whereas acetaldehyde had a marginal effect, suggesting that hypoxia-induced cold tolerance may arise from the accumulation and activity of ethanol. Cold tolerance was also induced by exposure of seedlings to volatile anesthetics including n -propanol, n -butanol, chloroform and halothane, suggesting that ethanol activity may result from fluidization of membrane lipids. This view is consistent with results which showed that ethanol activity was not associated with lipid metabolism. However, development of cold tolerance in ethanol-enriched tissues was time dependent, indicating that ethanol activity probably also entails biosynthetic event(s).