Effect of abscisic acid on chilling injury of zucchini squash

The endogenous levels of abscisic acid (ABA) in zucchini squash were increased by temperature conditioning at 10°C for 2 days. This temperature conditioning treatment reduced the severity of chilling injury in the squash during subsequent storage at 2.5°C. The ABA levels remained higher in treated squash than in untreated samples throughout the storage. Direct treatments of squash with ABA at 0.5 and 1.0 mM before storage at 2.5°C increased ABA levels in the tissue and were also effective in reducing chilling injury.     

Changes in the levels of abscisic acid and its metabolites resulting from chilling of tomato fruits

The 6,6,6-[²H]-analogues of abscisic acid (ABA), phaseic (PA) and dihydrophaseic (DPA) acids were used in GC-MS-SIM determination of free and total alkali hydrolyzable ABA, PA and DPA in the pericarp of tomato (Lycopersicon esculentum L. cv. Pik Red) fruit. Determinations were made on breaker-stage fruit stored 1, 2, 3 or 4 weeks at 2.5°C or at 10°C, and after subsequent ripening for 1 week in darkness at 20°C. Two-fold increases in levels of ABA occurred after storage at low temperatures with a slightly but significantly greater increase in ABA level occurring with 2.5°C storage. These increases in ABA levels were not associated with the appearance of damage symptoms that occurred with storage at the chilling temperature (2.5°C). Differences in ABA metabolism were found resulting from storage at the two temperatures, 2.5 or 10°C. Significantly greater DPA levels were found after 10°C storage than after 2.5°C storage (2 weeks). Levels of ABA ester-conjugates increased with 20°C ripening only after 10°C storage while free ABA levels decreased after both storage temperature conditions. Levels of DPA conjugates also increased only after 20°C ripening following 10°C storage. A longer period of storage resulted in decreases of free DPA levels after 10°C storage but increased DPA levels were found after 2.5°C storage.Abbreviations ABA abscisic acid - PA phaseic acid - DPA dihydrophaseic acid - GC-MS-SIM gas chromatography-mass spectrometry-selected ion monitoring - HPLC high pressure liquid chromatography - fw. fresh weight author for correspondence     

Relationship between ABA and chilling injury in mangosteen fruit treated with spermine

Theeffects of spermine on abscisic acid (ABA), hardening, and browning in storedmangosteen fruit were investigated. The hardening and browning, which areassociated with chilling injury (CI), were observed in only the skin of fruitstored at 7 °C. However, the hardening of skin was notaccompanied by moisture loss. The spermine treatment decreased the browning andhardening of the skin and extended storage time. Carbon dioxide(CO₂)production from stored fruit gradually increased with d in storage(DIS). The increase of CO₂ may be associated with the moisture lossbecause these levels coincided. ABA concentrations in the skin were highest infruit stored at 7 °C, followed by spermine treatment at 7°C, and the lowest at 13 °C. That is, thespermine treatment inhibited the increase of ABA in the skin of stored fruit.ABA concentrations in the skin may be associated with the degree of CI becausetheir fluctuations coincided. ABA metabolism in fruit stored at 7°C or 13 °C was also examined. The PA-DPApathway may not be the primary pathway of ABA metabolism because theconcentrations of PA and DPA were very low compared with those of ABA. ABAconcentrations in the aril were not significantly different between 7°C and 13 °C. This may be related to the lackof CI observed in the aril. ABA metabolism was different at each temperature.The decrease of ABA and the increase of DPA correlated at 13°C, however this correlation was not observed at 7°C. ABA metabolism may be influenced by temperature.     

Effects of Chilling Temperatures on Ethylene Binding by Banana Fruit

Banana fruit are highly susceptible to chilling injury during low temperature storage. Experiments were conducted to compare ethylene binding during storage at chilling (3 and 8 °C) versus optimum (13 °C) temperatures. The skins of fruit stored at 3 and 8 °C gradually darkened as storage duration increased. This chilling effect was reflected in increasing membrane permeability as shown by increased relative electrolyte leakage from skin tissue. In contrast, banana fruit stored for 8 days at 13 °C showed no chilling injury symptoms. Exposure of banana fruit to the ethylene binding inhibitor 1-methylcyclopropene (1 l l^-1 1-MCP) prevented ripening. However, this treatment also enhanced the chilling injury accelerated the occurrence of chilling injury-associated increased membrane permeability. ¹⁴C-ethylene release assay showed that ethylene binding by banana fruit stored at low temperature decreased with reduced storage temperature and/or prolonged storage time. Fruit exposed to 1-MCP for 12 h and then stored at 3 or 8 °C exhibited lower ethylene binding than those stored at 13 °C. Thus, chilling injury of banana fruit stored at low temperature is associated with a decrease in ethylene binding. The ability of tissue to respond to ethylene is evidently reduced, thereby resulting in failure to ripen.     

High temperature pulses decrease indirect chilling injury and elevate ATP levels in the flesh fly, Sarcophaga crassipalpis

Indirect chilling injury commonly occurs during long-term exposure to low temperature in many organisms including insects. A previous study revealed increased rates of survival and reduced cold injury in flesh flies, Sarcophaga crassipalpis, that experienced an intermittent pulse of high temperature during a low-temperature regiment. We extended these studies by determining survival rates and ATP levels for flies that had undergone continuous long-term exposure at 0 °C versus those experiencing a 24-h warming pulse of either 15 or 20 °C. Survival among flies that had undergone a warming pulse was significantly greater than for flies that were maintained continuously at 0 °C. Furthermore, ATP levels of flies that had experienced a warming pulse were significantly higher than those of flies maintained at 0 °C. These data suggest that brief warming pulses during long-term cold storage allow regeneration of energy reserves that promote survival and reduce indirect chilling injury.     

Genotype-specific differences in chilling tolerance of maize in relation to chilling-induced changes in water status and abscisic acid accumulation

Four inbred maize lines differing in chilling tolerance were used to study changes in water status and abscisic acid (ABA) levels before, during and after a chilling period. Seedlings were raised in fertilized soil at 24/22°C (day/night), 70% relative humidity. and a 12-h photoperiod with 200 μmol m⁻² s⁻¹ from fluorescent tubes. At an age of 2 weeks the plants were conditioned at 14/12°C for 4 days and then chilled for 5 days at 5/3°C. The other conditions (relative humidity, quantum flux, photoperiod) were unchanged. After the chilling period the plants were transferred to the original conditions for recovery. The third leaves were used to study changes in leaf necrosis, ion efflux, transpiration, water status and ABA accumulation. Pronounced differences in chilling tolerance between the 4 lines as estimated by necrotic leaf areas, ion efflux and whole plant survival were observed. Conditioning significantly increased tolerance against chilling at 5/3°C in all genotypes. The genotypes with low chilling tolerance had lower water and osmotic potentials than the more tolerant genotypes during a chilling period at 5/3°C. These differences were related to higher transpiration rates and lower diffusive resistance values of the more susceptible lines. During chilling stress at 5/3°C ABA levels were quadrupled. Only a small rise was measurable during conditioning at 14/12°C. However, conditioning enhanced the rise of ABA during subsequent chilling. ABA accumulation in the two lines with a higher chilling tolerance was triggered at a higher leaf water potential and reached higher levels than in the less tolerant lines. We conclude that chilling tolerance in maize is related to the ability for fast and pronounced formation of ABA as a protective agent against chilling injury.     

Chilling, oxidative stress and antioxidant responses in shoot cultures of rice

Chilling of shoot cultures from Oryza sativa L. cv. Taipei 309, to 4 °C leads to conditions of oxidative stress. Tissue H₂O₂ was observed to increase more than fourfold by 8 d of chilling, and levels of reduced glutathione, which normally rise in growing shoot cultures at 25 °C, were considerably repressed in chilled cultures. Whilst the activity of ascorbate peroxidase in chilled shoots remained similar to the activities in control cultures at 25 °C, the most notable effects of chilling to 4 °C were the very significant loss of catalase and glutathione reductase activity. Although prior exposure of shoot cultures to abscisic acid (ABA) at 25 °C increased levels of catalase activity, such increased levels were not sustained when the pre-treated cultures were placed at 4 °C. Moreover such pre-treatment with ABA did not increase the subsequent ability of shoot cultures to grow at 4 °C.Abbreviations GSH reduced glutathione - GSSG oxidised glutathione - ABA cis-abscisic acid This work is supported by a grant from the Biotechnology and Biological Sciences Research Council.     

Alteration of Gene Expression Associated with Abscisic Acid-Induced Chilling Tolerance in Maize Suspension-Cultured Cells

ABA induces chilling tolerance in maize (Zea mays L., cv Black Mexican Sweet) suspension-cultured cells at 28[deg] C when ABA was added to the culture medium at least 6 h prior to chilling (4[deg] C), and this induction can be inhibited by blocking protein synthesis with cycloheximide treatment (Z. Xin, P.H. Li [1992] Plant Physiol 99: 707-711). De novo synthesis of proteins and changes in poly(A+) RNAs were investigated during the ABA induction of chilling tolerance at 28[deg] C as well as during chilling exposure. At 28[deg] C, ABA increased the net synthesis of 11 proteins. Five of these proteins, whose net synthesis was also increased by chilling (4[deg] C), were called group I ABA-induced proteins; the remaining six proteins, whose net synthesis was not altered by chilling, were called group II ABA-induced proteins. Chilling suppressed the net synthesis of three proteins. ABA treatment prior to chilling did not alleviate this suppression. ABA applied at the inception of chilling induced neither chilling tolerance nor accumulation of any of the group II proteins; however, once the group II proteins appeared, they were continually synthesized even in a chilling regimen. ABA induced seven in vitro translation products at 28[deg] C. Three of these products could also be induced by chilling; the remaining four were induced by ABA only at 28[deg] C. These results suggest that ABA-induced alteration of protein synthesis at 28[deg] C is associated with an increased chilling tolerance in maize suspension-cultured cells.     

Fourier transform infrared spectra of zucchini squash stored at chilling or non-chilling temperatures

Mention of a trademark, proprietary product, or vendor does not constitute a guarantee or warranty of the product by the U.S. Department of Agriculture and does not imply its approval to the exclusion of other products or vendors that might be suitable.Fourier transform infrared spectroscopy (FTIR) was used to investigate spectral changes in the epidermis of zucchini squash resulting from low temperature storage-chilling (5°C) or non-chilling (15°C). Increased levels of fluid (water) were found in the tissue after 2 days of 5°C storage. This increase was reversed to harvest levels when chilled squash were warmed to room temperature for 1 day. After 3 days at 5°C and 1 day at room temperature, a further increase in fluid levels was found in the epidermis. Squash chilled for 3 days were apparently beyond recovery as indicated by spectral changes, although visual symptoms of chilling injury were not apparent until another 3 days of exposure to 5°C. Spectra of epidermis tissues from squash stored at 15°C indicated a pattern of increased non-reversible fluid accumulation when storage was prolonged. These results suggest that FTIR spectroscopy may be a rapid way to detect changes in chilled tissues before the eventual appearance of visible symptoms.     

Abscisic acid relationships in the chill-related dormancy mechanism in apple seeds

Abscisic acid (ABA) levels in seeds from three cultivars of apple (Malus domestica Borkh.) which have substantially different chilling requirements were investigated by gas chromatography mass-spectrometry selected ion monitoring (GCMS-SIM) during stratification. The ABA content of dormant unchilled seeds was similar in the three cultivars, suggesting no relationship between the chilling requirement of those seeds and their ABA status. That chilling is not related to ABA changes during stratification was confirmed by warm (20°C) and cold (5°C) stratification experiments. ABA content dropped rapidly and nearly identically under both temperature regimes, but only cold stratification promoted germination. The decline in ABA during stratification was due in large part to leaching from the seed coat and nucellar membrane; the ABA content of the embryo remained nearly constant. The radicle in intact seeds stratified at 5°C began growing 20–30 days after the ABA in the seed coat and nucellar membrane had nearly disappeared. Radicle growth did not occur in unchilled seeds, even though ABA had leached from them as well. It is possible that the leaching of ABA from the seed allows certain promotive forces to develop, but if so, these can develop only at chilling temperatures. Studies were also conducted on 2-trans ABA relationships to apple seed dormancy, but no association was evident.Report No. 12, Department of Fruit and Vegetable Science, Cornell University.     

Polyamine content and chilling susceptibility are affected by seasonal changes in temperature and by conditioning temperature in cold-stored 'Fortune' mandarin fruit

‘Fortune’ mandarins are prone to develop pitting and necrosis upon exposure to low temperatures. We have examined the effect of field temperature during fruit maturation and the effect of conditioning temperatures (from 2 to 37°C) prior to cold storage on the content of polyamines (PAs) and on chilling susceptibility in order to understand the role of PAs in maturation and chilling tolerance of this citrus cultivar. Chilling susceptibility and the content of PAs were more affected by seasonal changes in field temperature than by the stage of fruit maturity. The highest putrescine (Put) and spermidine (Spd) content was found in fruits exposed to the lowest field temperatures. These fruits were in turn more susceptible to develop chilling injury (CI) after storage at 2°C. Spermine (Spm), however, decreased in attached fruit with time of exposure to temperatures below 12°C. Temperature pretreatments for 3 days above 20°C of fruits detached from the tree reduced CI, the more so the higher the conditioning temperature. Put and Spd increased with temperature conditioning in detached fruits, differing from the response of fruits attached to the tree. No direct relationship between induced levels of these PAs and the tolerance to CI was found. Levels of Put and Spd increased at temperatures (22, 30 and 37°C) which increased the tolerance and also at temperatures (6 and 12°C) which accelerated the appearance of chilling symptoms. In contrast, a significant increase in Spm levels was only found after conditioning at 30 or 37°C. After cold storage a general decline in PA levels occurred in all temperature‐conditioned mandarins. In most cases no significant differences among fruit exposed to effective and non‐effective pretreatments were observed. PA content increased again after transferring cold‐stored fruits to 20°C, whereas the CI index was barely affected. In conclusion, PA changes in the flavedo of ‘Fortune’ mandarins appear to be related to variations in temperature rather than to stage of maturity or tolerance to chilling.     

Chilling-induced ethylene production by beans and peas

The effects of chilling on ethylene production by leaf discs and whole plants of bean (chilling-sensitive) and pea (chilling-tolerant) were studied. When pea or bean leaf discs were excised and incubated at 25°C, transient increases in ethylene production and 1-aminocyclopropane-1-carboxylic acid (ACC) accumulation were observed. Both pea and bean discs kept at 5°C evolved little ethylene, but levels of ACC increased in pea discs and not in bean discs. When discs of either species were chilled at 5°C immediately after excision and then transferred to 25°C 9 h later, increases in their ACC levels and ethylene production rates were observed. Discs were also incubated at 25°C for 12 h to allow excision-induced ethylene production to subside and then chilled at 5°C. Nine hours later, these discs were transferred to 25°C, and an increase in ethylene production was observed. These data indicate that chilling suppresses excision-induced ethylene production and enhances the production of ethylene after transfer to 25°C. Chilling of whole plants resulted in increased production of ethylene and ACC in the chilling-sensitive bean but not in the chilling-tolerant pea. Treatment of bean plants with the ethylene antagonists silver thiosulfate, norbornadiene, or aminooxyacetic acid, or of pea plants with ethylene, did not affect the appearance of chilling injury symptoms, indicating that ethylene does not induce injury symptoms and may not have an adaptive role in chilling stress.     

Effect of temperature conditioning on chilling injury of cucumber cotyledons: possible role of abscisic Acid and heat shock proteins

Endogenous abscisic acid levels and induced heat shock proteins were measured in tissue exposed for 6 hours to temperatures that reduced their subsequent chilling sensitivity. One-centimeter discs excised from fully expanded cotyledons of 11-day-old seedlings of cucumber (Cucumis sativus L., cv Poinsett 76) were exposed to 12.5 or 37°C for 6 hours followed by 4 days at 2.5 or 12.5°C. Ion leakage, a qualitative indicator of chilling injury, increased after 2 to 3 day exposure to 2.5°C, but not to 12.5°C, a nonchilling temperature. Exposure to 37°C before chilling significantly reduced the rate of ion leakage by about 60% compared to tissue exposed to 12.5°C before chilling, but slightly increased leakage compared to tissue exposed to 12.5 or 37°C and held at the nonchilling temperature of 12.5°C. There was no relationship between abscisic acid content following exposure to 12.5 or 37°C and chilling tolerance. Five heat shock proteins, with apparent molecular mass of 25, 38, 50, 70, and 80 kilodaltons, were induced by exposure to 37 or 42°C for 6 hours, and their appearance coincided with increased chilling resistance. Heat shock treatments reduced the synthesis of three proteins with apparent molecular mass of 14, 17, and 43 kilodaltons. Induction of heat shock proteins could be a possible cause of reduced chilling injury in tissue exposed to 37 or 42°C.     

The effect of LAB 173711 and ethephon on time of flowering and cold hardiness of peach flower buds

Peach flowers are often killed during bloom by spring frosts. LAB 173711, a compound with abscisic (ABA)-like activity, and ethephon delayed flowering in peach trees. In greenhouse experiments, LAB 173711, at concentrations of 10^–3–10^–2 M, was most effective in delaying bloom when applied after a 5°C cold storage period, rather than before the dormancy breaking treatment. In contrast, ethephon delayed bloom most effectively when applied before 5°C cold storage; ethephon caused flower bud abscission when treatments were made after the chilling requirement had been satisfied. In field experiments, ethephon delayed flowering by 6–7 days, which reduced bud injury after a spring frost during bloom. No flower bud injury was found on ethephon-treated trees after temperatures of –4.3°C; whereas without ethephon 25% of the flower buds were frost damaged. LAB 173711 delayed the time to 50% bloom by 2–3 days. However, this was not long enough to avoid low-temperature injury to the flower buds.     

Temperature effects on embryonic abscisic acid levels during development of wheat grain dormancy

Abscisic acid (ABA) levels were determined in both the embryo and remaining grain remnant during development of wheat caryopses under temperature conditions which produced either high or low levels of dormancy in mature grain. Higher levels of grain dormancy were produced in grain from plants grown at 15°C as compared to 25°C. In grain grown at 15°C, embryonic ABA levels steadily increased during development, reaching a maximum at stage IV, just before grain desiccation. At 25°C, ABA levels were very high at the earliest stages of embryonic development, but dropped rapidly during maturation. Only small cultivar differences in ABA levels were observed during development at either temperature. In general, higher levels of dormancy in mature grain correlated with prolonged elevation of ABA levels during grain maturation.Contribution from USDA-ARS and the College of Agriculture and Home Economics Research Center, Washington State University, scientific paper no. 8901-13.Mention of a specific product name by the United States Department of Agriculture does not constitute an endorsement and does not imply a recommendation over other suitable products.     

Kinetics of dormancy release and the high temperature germination response in Aesculus hippocastanum seeds

The kinetics of primary dormancy loss were investigated in seeds of horse chestnut (Aesculus hippocastanum L.) harvested in four different years. Freshly collected seeds from 1991 held for up to 1 year at temperatures between 2C and 42C exhibited two peaks in germination (radicle growth), representing a low temperature (2-8°C) and a high temperature response (31-36°C). Germination at 36°C generally occurred within 1 month of sowing, but was never fully expressed in the seedlots investigated. At low temperatures (2-8°C), germination started after around 4 months. Generally, very low levels of termination were observed at intermediate temperatures (11-26°C). Stratification at 6°C prior to germination at warmer temperatures increased the proportion of seeds that germinated, and the rate of germination for all seedlots. Within a harvest, germination percentage (on a probit scale) increased linearly with stratification time and this relationship was independent of germination temperature (16-26°C). However, inter-seasonal differences in the increases in germination capacity following chilling were observed, varying from 0.044 to 0.07 probits d^-1 of chilling at 6°C. Increased sensitivity to chilling was associated with warmer temperatures during the period of seed filling. The estimated base temperature for germination, Tb, for newly harvested seeds varied slightly between collection years but was close to 25°C. For all seedlots, Tb decreased by 1°C every 6 d of chilling at 6°C. This systematic reduction in Tb with chilling ultimately facilitated germination at 6°C after dormancy release.     

Acclimation,Hydrogen Peroxide,and Abscisic Acid Protect Mitochondria against Irreversible Chilling Injury in Maize Seedlings

Our previous results indicated that 3-d-old dark-grown chilling-sensitive maize (Zea mays L.) seedlings did not survive 7 d of 4[deg]C chilling stress, but 69% of them survived similar stress when the seedlings were either preexposed to 14[deg]C for 3 d or pretreated with 0.1 mM H2O2 for 4 h at 27[deg]C (T.K. Prasad, M.D. Anderson, B.A. Martin, C.R. Stewart [1994] Plant Cell 6: 65-74) or 1 mM abscisic acid (ABA) for 24 h at 27[deg]C (M.D. Anderson, T.K. Prasad, B.A. Martin, C.R. Stewart [1994] Plant Physiol 105: 331-339). We discovered that chilling imposed oxidative stress on the seedlings. Since H2O2 accumulated during the periods of both acclimation and nonacclimation, we concluded that H2O2 had dual effects at low temperature: (a) During acclimation, its early transient accumulation signals the induction of antioxidant enzymes such as catalase 3 and peroxidase to scavenge H2O2; and (b) at 4[deg]C in nonacclimated seedlings, it accumulates to damaging levels in the tissues because of low levels of these and perhaps other antioxidant enzymes. Three-day-old seedlings pretreated with H2O2 (a mild oxidative stress) or ABA showed induced chilling tolerance. In the present study, we investigated whether mitochondria are a target for chilling-induced oxidative stress and, if so, what differences do acclimation, H2O2, or ABA make to protect mitochondria from irreversible chilling injury. The results indicated that chilling, in general, impairs respiratory activity, the cytochrome pathway of electron transport, and ATPase activity regardless of the treatment. In pretreated seedlings, the activities of catalase 3 and peroxidase in the mitochondria increased severalfold compared with control and nonacclimated seedlings. The increases in these antioxidant enzymes imply that mitochondria are under oxidative stress and such increases could initiate a protective mechanism in the mitochondria. Mitochondrial respiration is partially cyanide resistant during chilling stress and also after the 1st d of recovery. Upon further recovery over 3 d, in contrast to nonacclimated seedlings, the mitochondria of acclimation-, H2O2-, and ABA-treated seedlings showed the following recovery features. (a) The mitochondrial respiration changed from a cyanide-resistant to a cyanide-sensitive cytochrome pathway, (b) cytochrome oxidase activity recovered to control levels, (c) the ability of mitochondria to generate ATP was regained, and (d) the antioxidant enzyme activities remained at or above control levels. Based on these results, we conclude that chilling impairs mitochondrial function and that chilling-induced oxidative stress seems to be a factor, at least in part, for causing possible irreversible damage to the mitochondrial membrance components. Acclimation, H2O2, and ABA provide a protective mechanism by inducing antioxidant enzymes to protect mitochondria from irreversible oxidative damage that is absent in nonacclimated seedlings. Therefore, we conclude that the ability of the seedlings to recover from chilling injury is, at least in part, due to the ability of the mitochondria to resume normal function.     

Chilling injury in coleus as influenced by photosynthetically active radiation, temperature and abscisic acid pretreatment. I. Morphological and physiological responses

Abstract. Coleus blumei Benth. (PI No. 354190), a green-leafed cultivar, was exposed to 5°C for 48 or 72 h after pretreatment for 48 h at two levels of photosynthetically active radiation (PAR) (8 and 320 μmol s⁻¹ m⁻²), two temperatures (13 and 20°C), and two abscisic acid (ABA) levels (0 and 200 g m⁻³ of the racemic mixture). Plants given low PAR for only 48 h prior to chilling treatment (48 or 72 h at 5°C) showed increased protection against chilling injury while those given high PAR were severely injured. The former plants were darker green, contained greater concentrations of chlorophyll- a , chlorophyll- b , total chlorophyll and anthocyanin and generally had a lower abscission rate than the latter plants. There were no differences, however, in chlorophyll- a/b ratio among plants grown at the two PAR levels, two temperatures or two ABA concentrations. Temperature and ABA pretreatment and number of hours at 5°C had no significant effect on chilling injury as measured by leaf chlorosis, but generally had a significant effect on leaf abcission, especially at 3 and 7 d after returning the plants to the greenhouse. Enclosing intact plants or excised shoots in plastic bags to maintain 100% relative humidity during 72 h chilling treatment failed to provide protection against chilling injury. These findings indicate that the protective effects of low PAR applied prior to chilling treatment may be as important or more important than that applied during chilling. They also indicate the importance of making careful measurements of PAR levels when conducting studies on chilling injury.