Effect of exogenous abscisic acid on cold acclimation in two Magnolia species

In northern China, freezing injury is observed frequently in the rare species Magnolia wufengensis but not in the more common species Magnolia denudata. To investigate the role of the phytohormone abscisic acid (ABA) on frost tolerance in these two species, exogenous ABA was applied to the seedlings and then physiological and biochemical responses were measured during cold acclimation. Shoot growth cessation was stimulated by ABA in M. wufengensis but not in M. denudata. Abscisic acid inhibited shoot growth in M. wufengensis but not in M. denudata. Treatment with ABA stimulated leaf senescence in both species, and this effect was greater in M. denudata. For both species, ABA-treated plants exhibited bud dormancy sooner and had an increased tolerance to freezing, decreased water content and increased accumulation of proline, glucose, and fructose in shoots. These effects were generally greater for M. denudata. Freezing tolerance was significantly correlated with content of water, proline, glucose, and fructose for both species, but freezing tolerance was significantly correlated with raffinose content only in M. wufengensis. We conclude that exogenous ABA could increase cold acclimation and improve cold hardiness of both Magnolia species, although M. denudata was more responsive to ABA than M. wufengensis, which might result from a greater dehydration and accumulation of proline and certain soluble sugars.     

Involvement of abscisic Acid in potato cold acclimation

Upon exposure to 2°C day/night (D/N), leaves of Solanum commersonii (Sc) began acclimating on the 4th day from a −5°C (killing temperature) hardy level to −12°C by the 15th day. Leaves of S. tuberosum L. (St) cv `Red Pontiac' typically failed to acclimate and were always killed at −3°C. Leaves of control (20/15°C, D/N) and treated plants (2°C, D/N) of St showed similar levels of free abscisic acid (ABA) during a 15-day sampling period. In treated Sc plants, however, free ABA contents increased 3-fold on the 4th day and then declined to their initial level thereafter. The increase was not observed in leaves of Sc control plants.

Treated St plants showed a slightly higher content of leaf soluble protein than controls. In Sc, leaves of controls maintained relatively constant soluble proteins, but leaves of treated plants showed a distinct increase. This significant increase was initiated on the 4th day, peaked on the 5th day, and remained at a high level throughout the 15-day sampling period.

Exogenously applied ABA induced frost hardiness in leaves of Sc plants whether plants were grown under a 20°C or 2°C temperature regime. When cycloheximide was added to the medium of stem-cultured plants at the beginning of 2°C acclimation, or at the beginning of the ABA treatment in the 20°C regime, it completely inhibited the development of frost hardiness. However, when cycloheximide was added to plants on the 5th day during 2°C acclimation, the induction of frost hardiness was not inhibited. The role of ABA in triggering protein synthesis needed to induce frost hardiness is discussed.

     

Potato cold hardiness development and abscisic acid. I. Conjugated abscisic acid is not the source of the increase in free abscisic acid during potato (Solanum commersonii) cold acclimation

Free and conjugated abscisic acid (ABA) levels in stem-cultured plantlets of potato ( Solanum commersonii Dun, PI 458317) during cold acclimation were measured. The levels of free and conjugated ABA were measured by an enzyme immunoassay (EIA) with rabbit anti-ABA-serum. The use of immunoglobulin G fraction purified from rabbit antiserum and the methylated form of ABA resulted in an improved measuring range (0.01 to 10 pmol ABA) and precision (slope of logit-log plot, −1.35) of EIA, compared to the use of antiserum and free ABA. Estimates of the EIA were consistent with those resulting from a commercial EIA. Under a 4/2°C (day/night) temperature regime, the potato plantlets increased cold hardiness from −5°C (warm-grown control) to −10°C by the 7th day. During the same period, there were two transitory increases in free ABA, the first one three-fold from 1.5 to 5.3 nmol (g dry weight)⁻¹ on the 2nd day and the second one five-fold from 1.5 to 7.6 nmol (g dry weight)⁻¹ on the 6th day. Each increase in ABA concentration was followed by an increase in cold hardiness. There was no significant change in conjugated ABA content (4.2±0.6 nmol [g dry weight]⁻¹) throughout the cold acclimation period. The lack of an interrelationship between levels of free and conjugated ABA suggested that the transitory increase in free ABA during cold acclimation was not a result of the conversion of conjugated ABA. The increase in free ABA due to biosynthesis of ABA during potato cold acclimation is discussed.     

Cold acclimation and abscisic acid induced alterations in carbohydrate content in calli of wheat genotypes differing in frost tolerance

The effect of cold and abscisic acid (ABA) treatment on soluble carbohydrate content was compared in callus cultures of wheat genotypes differing in frost tolerance. The effect of 5A chromosome substituted from the frost tolerant to the sensitive on cold-induced carbohydrate accumulation was also determined. Following cold hardening, the increase in sucrose and fructan level in calli of tolerant varieties was significantly higher than those of the sensitive ones. In 5A substitution line higher sucrose and fructan content was detected than in recipient . Tendentiously, cold stress caused higher degree of changes in carbohydrate content than the exogenously applied ABA did. Comparing the accumulation pattern of the components of WSC measured in vitro to the previously published in vivo results it can be concluded that in the case of sucrose and fructans it was similar, while for the reducing sugars it was different. The regulatory role of chromosome 5A either in the development of freezing tolerance or carbohydrate accumulation was confirmed in dedifferentiated calli, as well.     

Low temperature acclimation and treatment with exogenous abscisic acid induce common polypeptides in Arabidopsis thaliana (L.) Heynh

Summary Exogenously applied abscisic acid (ABA) induced frost hardening of Arabidopsis thaliana (L.) Heynh. The freezing tolerance of A. thaliana plantlets treated with ABA (15 mg/l) at a non-acclimating temperature (20 °C) appeared to increase even more rapidly than following a low temperature (4 °C) acclimation. Analysis of in vivo-labelled soluble proteins by two-dimensional gel electrophoresis revealed several low temperature — or ABA — induced proteins, which where not produced in non-acclimated plants. A subset of these proteins was induced by both low temperature and ABA treatments, suggesting that they might be directly involved in the frost hardening process in A. thaliana.     

Potato cold hardiness development and abscisic acid. II. De novo synthesis of proteins is required for the increase in free abscisic acid during potato (Solanum commersonii) cold acclimation

During cold acclimation of potato plantlets ( Solanum commersonii Dun, PI 458317), there are two transitory increases in free ABA content corresponding to a three-fold increase on the 2nd day and a five-fold increase on the 6th day (Ryu and Li 1993). During this period, plantlets increased in cold hardiness from −5°C (killing temperature, control grown at 22/18°C, day/night) to −10°C by the 7th day of exposure to 4/2°C (day/night). This increase in free ABA was not found when cycloheximide (CHI), an inhibitor of cytoplasmic protein synthesis, was added to the culture medium 6 h before exposure to low temperatures. Plantlets treated with CHI did not acclimate to cold, maintaining a hardiness level (−5°C) similar to that of the 22/18°C-grown plantlets. When the CHI-treated plantlets were exposed to low temperatures for 3 days, transferred to CHI-free culture medium and grown at low temperatures, the plantlets showed a transitory increase in free ABA 2 days later. This increase was followed by the development of cold hardiness (−8°C). Application of CHI to the culture medium after 3 days of cold acclimation, when the first ABA peak and a partial development of cold hardiness (−8°C) had occurred, blocked the second transitory increase in free ABA and resulted in no further development of cold hardiness. These results suggest that de novo synthesis of proteins is required for these transitory increases in free ABA during cold acclimation of potato plantlets.     

Effect of abscisic acid and cold acclimation on the cytoskeletal and phosphorylated proteins in different cultivars of Triticum aestivum L

In winter wheat, the tubulin and 60 kDa-phosphorylated proteins/actin ratio is considerably higher in the roots than in the leaves. Differences in the content of the main cytoskeletal proteins were also found in the leaves of the different cultivars. It is suggested that the lower amount of the tubulin and 60 kDa-phosphorylated proteins and higher content of actin determine the greater tubulin cytoskeletal stability in the leaves and their higher frost resistance, as compared with the roots. Also, it is possible that the higher content of the tubulin and 60 kDa-phosphorylated proteins defines the lower microtubule (MT) stability in the leaves of the low frost resistant cultivar than in the leaves of the more frost resistant ones. In the roots and leaves of the low frost resistant cultivar, the low stability of the numerous tubulin structures is apparently one reason for the abscisic acid (ABA)-induced reduction of the cytoskeletal and 60 kDa-phosphorylated proteins in the cells. The cold acclimation compensated the ABA effect in the roots of the very frost resistant cultivar in the most extent. This suggests the existence of the different pathways in the increased plant cell frost resistance through the action of ABA and low temperature.     

Ultrastructural and protein changes in cell suspension cultures of peach associated with low temperature-induced cold acclimation and abscisic acid treatment

Cell suspension cultures were initiated from callus derived from xylem tissues of peach [Prunus persica (L.) Batsch]. Cold acclimation was induced (LT₅₀ of-13°C) in cell suspensions at 3°C in the dark for 10 days. Freezing tolerance returned to the level of nonacclimated cells (LT₅₀ of –4.5°C) when cold-acclimated cells were transferred to 24°C (in dark) for 3 days. Addition of 75 M abscisic acid (ABA) to the growth medium failed to induce cold acclimation after cells were cultured for 5 days at 24°C. Microvacuolation, cytoplasmic augmentation and disappearance of starch grains were observed in cells that were cold-acclimated by exposure to low temperature. Similar ultrastructural alterations were not observed in ABA-treated cells. Several qualitative and quantitative changes in proteins were noted during both cold acclimation and ABA treatment. Both the ultrastructural and protein changes observed during cold acclimation were reversed during deacclimation. The relationship of these changes to cold acclimation in peach cell-cultures is discussed.Abbreviations ABA abscisic acid - 2,4-d 2,4-dichlorophenoxyacetic acid - IBA indole-3-butyric acid - Ms Murashige & Skoog - PMSF phenylmethylsulfonyl fluoride - LT₅₀ or Freezing Tolerance temperature that resulted in 50% decrease in TTC reduction - TTC 2,3,5-triphenyltetrazolium chloride     

Cryopreservation of in vitro sugar beet shoot tips using the encapsulation-dehydration technique: influence of abscisic acid and cold acclimation

It has been previously shown that shoot tips of in vitro plantlets of sugar beet (Beta vulgaris L. clone SES1) can be cryopreserved using the encapsulation-dehydration technique (survival rate of 37% after freezing). This article reports the influence of abscisic acid (ABA) and cold acclimation on survival after cryopreservation. When ABA was added to the multiplication medium of the plants, the survival rate of shoot tips after cryopreservation was not increased (45%). After cold acclimation of the plants, their growth pattern differed (plants became apically dominant) and the survival rate of the shoot tips after cryopreservation clearly increased (70% survival and 50% plant regeneration after freezing). This improved protocol was successfully applied to three other clones. Received: 28 October 1996 / Revision received: 28 January 1997 / Accepted: 15 March 1997     

Effects of abscisic acid on the orientation and cold stability of cortical microtubules in epicotyl cells of the dwarf pea

Summary The effects of abscisic acid (ABA) on the orientation and cold stability of cortical microtubules (MTs) in epidermal cells of epicotyls of the dwarf pea,Pisum sativum L. cv. Little Marvel, were examined by immunofluorescence microscopy. The effect of ABA on the elongation of epicotyls and on the orientation of cortical MTs was opposite to that of gibberellin A₃ (GA₃). Treatment with ABA, which reduced the promotion of epicotyl elongation by GA₃, eliminated the GA₃-induced predominance of transverse MTs and resulted in a predominance of longitudinal MTs. The effect of ABA on the cold stability of cortical MTs was also opposite to that of GA₃. ABA increased the cold stability of MTs, while GA₃ decreased it. The predominance of longitudinal MTs brought about by ABA may have some relationship to ABA-induced inhibition of the elongation of the epicotyl. ABA may alter membrane proteins to stabilize cortical MTs and induce cold hardiness of plants.Abbreviations ABA abscisic acid - DMSO dimethylsulfoxide - FITC fluorescein isothiocyanate - GA₃ gibberellin A₃ - MT microtubule - PBS phosphate-buffered saline Dedicated to the memory of Professor Oswald Kiermayer     

Cold shock and cold acclimation proteins in the psychrotrophic bacterium Arthrobacter globiformis SI55.

The psychrotrophic bacterium Arthrobacter globiformis SI55 was grown at 4 and 25 degrees C, and the cell protein contents were analyzed by two-dimensional electrophoresis. Cells subjected to cold shocks of increasing magnitude were also analyzed. Correspondence analysis of protein appearance distinguished four groups of physiological significance. Group I contained cold shock proteins (Csps) overexpressed only after a large temperature downshift. Group II contained Csps with optimal expression after mild shocks. Group III contained proteins overexpressed after all cold shocks. These last proteins were also overexpressed in cells growing at 4 degrees C and were considered to be early cold acclimation proteins (Caps). Group IV contained proteins which were present at high concentrations only in 4 degrees C steady-state cells and appeared to be late Caps. A portion of a gene very similar to the Escherichia coli cspA gene (encoding protein CS7.4) was identified. A synthetic peptide was used to produce an antibody which detected a CS7.4-like protein (A9) by immunoblotting two-dimensional electrophoresis gels of A. globiformis SI55 total proteins. Unlike mesophilic microorganisms, this CS7.4-like protein was still produced during prolonged growth at low temperature, and it might have a particular adaptive function needed for balanced growth under harsh conditions. However, A9 was induced at high temperature by chloramphenicol, suggesting that CS7.4-like proteins have a more general role than their sole implication in cold acclimation processes.     

The expression of a rab-related gene,rab 18, is induced by abscisic acid during the cold acclimation process of Arabidopsis thaliana (L.) Heynh

Erratum

The expression of a rab-related gene, rab 18, is induced by abscisic acid during the cold acclimation process of Arabidopsis thaliana (L.) Heynh     

The expression of a rab-related gene,rab18, is induced by abscisic acid during the cold acclimation process of Arabidopsis thaliana (L.) Heynh

We have isolated a rab-related (responsive to ABA) gene, rab18 from Arabidopsis thaliana. The gene encodes a hydrophilic, glycine-rich protein (18.5 kDa), which contains the conserved serine- and lysine-rich domains characteristic of similar RAB proteins in other plant species. The rab18 mRNA accumulates in plants exposed to low temperature, water stress or exogenous ABA but not in plants subjected to heat shock. This stress-related accumulation of the rab18 mRNA is markedly decreased in the ABA-synthesis mutant aba-1, the ABA-response mutant abi-1 or in wild-type plants treated with the carotenoid synthesis inhibitor, fluridone. Exogenous ABA treatment can induce the rab18 mRNA in the aba-1 mutant but not in the abi-1 mutant. These results provide direct genetic evidence for the ABA-dependent regulation of the rab18 gene in A. thaliana.