Plant Cold Acclimation Research Articles

Development of freezing tolerance in different altitudinal ecotypes of Salix paraplesia

Salix paraplesia was used as an experimental model to investigate the effect of short day photoperiod (SD) and low temperature (LT) on development of freezing tolerance and on endogenous abscisic acid (ABA) contents. We characterized differences in SD and LT-induced cold acclimation in three ecotypes from different altitudes. The results demonstrated that cold acclimation could be triggered by exposing the plants to SD or LT alone, and that a combination of the different treatments had an additive effect on freezing tolerance in all ecotypes studied. However, the high altitudinal ecotype was more responsive to SD and LT than the low altitudinal ecotype. Development of freezing tolerance induced by SD and LT was accompanied by changes in ABA contents which were ecotype-dependent. Although the stem had higher initial freezing tolerance, the leaves developed freezing tolerance more quickly than the stem and thus leaves may provide an interesting experimental system for physiological and molecular studies of cold acclimation in woody plants.

On the induction of cold acclimation in carrots ( Daucus carota L.) and its influence on storage performance

We investigated the role of cold acclimation in carrot plants with respect to its influence on the storage performance of the harvested taproots. The induction of cold acclimation was followed in plants cultivated in a growth chamber under strict climate control and in taproots harvested from two separate field cultivations where the plants had been exposed to the natural variations in climate. Under controlled growth conditions, levels of antifreeze protein (AFP) mRNA were used as a marker for cold acclimation in carrot taproot tissue. Expression of this gene was induced by cold in discs excised from harvested taproots and this induction was clearly affected by the growth temperature of the plants from which the taproots were taken. These in vitro data were consistent with those from field-grown plants. In the cell wall of taproots harvested in year 2000, where the intact plants had frequently been exposed to temperatures below 6 °C, a 36 kDa AFP accumulated to higher levels during storage than in the taproots harvested from plants grown in year 2001, where cultivation temperatures had rarely dropped below 6 °C. The taproots from 2001 exhibited poor storage performance as shown by an earlier increase in relative electrolyte leakage and decrease in dry matter compared to taproots harvested in 2000. The capacity of the AFP to accumulate during storage was consistent with a high storage performance.

Phylogenetic analyses in cornus substantiate ancestry of xylem supercooling freezing behavior and reveal lineage of desiccation related proteins.

The response of woody plant tissues to freezing temperature has evolved into two distinct behaviors: an avoidance strategy, in which intracellular water supercools, and a freeze-tolerance strategy, where cells tolerate the loss of water to extracellular ice. Although both strategies involve extracellular ice formation, supercooling cells are thought to resist freeze-induced dehydration. Dehydrin proteins, which accumulate during cold acclimation in numerous herbaceous and woody plants, have been speculated to provide, among other things, protection from desiccative extracellular ice formation. Here we use Cornus as a model system to provide the first phylogenetic characterization of xylem freezing behavior and dehydrin-like proteins. Our data suggest that both freezing behavior and the accumulation of dehydrin-like proteins in Cornus are lineage related; supercooling and nonaccumulation of dehydrin-like proteins are ancestral within the genus. The nonsupercooling strategy evolved within the blue- or white-fruited subgroup where representative species exhibit high levels of freeze tolerance. Within the blue- or white-fruited lineage, a single origin of dehydrin-like proteins was documented and displayed a trend for size increase in molecular mass. Phylogenetic analyses revealed that an early divergent group of red-fruited supercooling dogwoods lack a similar protein. Dehydrin-like proteins were limited to neither nonsupercooling species nor to those that possess extreme freeze tolerance.

Low temperature sensing in silver birch (Betula pendula Roth) ecotypes

Non-freezing low temperature (LT) is the main factor controlling cold acclimation and dormancy release of woody plants in the cold and temperate regions. Through evolution many tree species have developed different ecotypes, which are closely adapted to the local climatic conditions. In order to elucidate the physiological basis of climatic adaptation involved in LT sensing in the northern tree species, we investigated responses of the northern and southern ecotypes of silver birch (Betula pendula Roth) to a range of decreasing temperatures. Development of freezing tolerance, determined by the electrolyte leakage, and bud dormancy release were monitored. A range of decreasing temperatures triggered cold acclimation and development of freezing tolerance in actively growing plants, and enhanced freezing tolerance and released bud dormancy in the dormant plants in both ecotypes tested. However, there were significant ecotypic differences in LT-induced cold acclimation and bud dormancy release. The northern ecotype was more responsive to a range of decreasing temperatures than the southern ecotype, resulting in more rapid cold acclimation and development of freezing tolerance in actively growing plants, and earlier enhancement of freezing tolerance and bud dormancy release in the dormant plants. Development of freezing tolerance and dormancy release induced by LT was accompanied by changes in ABA levels. Alterations in ABA levels paralleled with development of freezing tolerance and preceded bud dormancy release in both ecotypes tested, but these alterations in ABA levels were ecotype-dependent, the northern ecotype reacting more strongly to LT.

Environmental regulation and physiological basis of freezing tolerance in woody plants

Cold acclimation of plants is a complex process involving a number of biochemical and physiological changes. The ability to cold acclimate is under genetic control. The development of freezing tolerance in woody plants is generally triggered by non-freezing low temperatures but can also be induced by mild drought or exogenous abscisic acid, as well as by short photoperiod. In nature, the extreme freezing tolerance of woody plants is achieved during sequential stages of cold acclimation the first of which is initiated by short photoperiods and non-freezing low temperatures, and the second by freezing temperatures. Although recent breakthroughs have increased our knowledge on the physiological molecular basis of freezing tolerance in herbaceous species, which acclimate primarily in response to non-freezing low temperatures, very little is known about cold acclimation of woody plants. This article attempts to review our current understanding of the physiological aspects that underline cold acclimation in woody plants.

Low temperature sensing in silver birch ( Betula pendula Roth) ecotypes

Non-freezing low temperature (LT) is the main factor controlling cold acclimation and dormancy release of woody plants in the cold and temperate regions. Through evolution many tree species have developed different ecotypes, which are closely adapted to the local climatic conditions. In order to elucidate the physiological basis of climatic adaptation involved in LT sensing in the northern tree species, we investigated responses of the northern and southern ecotypes of silver birch ( Betula pendula Roth) to a range of decreasing temperatures. Development of freezing tolerance, determined by the electrolyte leakage, and bud dormancy release were monitored. A range of decreasing temperatures triggered cold acclimation and development of freezing tolerance in actively growing plants, and enhanced freezing tolerance and released bud dormancy in the dormant plants in both ecotypes tested. However, there were significant ecotypic differences in LT-induced cold acclimation and bud dormancy release. The northern ecotype was more responsive to a range of decreasing temperatures than the southern ecotype, resulting in more rapid cold acclimation and development of freezing tolerance in actively growing plants, and earlier enhancement of freezing tolerance and bud dormancy release in the dormant plants. Development of freezing tolerance and dormancy release induced by LT was accompanied by changes in ABA levels. Alterations in ABA levels paralleled with development of freezing tolerance and preceded bud dormancy release in both ecotypes tested, but these alterations in ABA levels were ecotype-dependent, the northern ecotype reacting more strongly to LT.

Overexpression of multiple dehydrin genes enhances tolerance to freezing stress in Arabidopsis.

To elucidate the contribution of dehydrins (DHNs) to freezing stress tolerance in Arabidopsis, transgenic plants overexpressing multiple DHN genes were generated. Chimeric double constructs for expression of RAB18 and COR47 (pTP9) or LTI29 and LTI30 (pTP10) were made by fusing the coding sequences of the respective DHN genes to the cauliflower mosaic virus 35S promoter. Overexpression of the chimeric genes in Arabidopsis resulted in accumulation of the corresponding dehydrins to levels similar or higher than in cold-acclimated wild-type plants. Transgenic plants exhibited lower LT50 values and improved survival when exposed to freezing stress compared to the control plants. Post-embedding immuno electron microscopy of high-pressure frozen, freeze-substituted samples revealed partial intracellular translocation from cytosol to the vicinity of the membranes of the acidic dehydrin LTI29 during cold acclimation in transgenic plants. This study provides evidence that dehydrins contribute to freezing stress tolerance in plants and suggests that this could be partly due to their protective effect on membranes.

ABA and gibberellin-like substances during prehardening, cold acclimation, de- and reacclimation of oilseed rape

Previously published results showed that high relative reduction state of PSII (PSII excitation pressure) during both early seedling growth (prehardening) as well as cold deacclimation caused significant changes in growth pattern. The differences in elongation growth rate were related to the cold acclimation of photosynthetic apparatus and to frost resistance. To study changes in the hormonal balance connected with alterations in elongation growth rate observed during prehardening and deacclimation under different PSII excitation pressure (modulated by day-temperatures), endogenous concentration of ABA, GA3 and GA-like substances (GAs) were analysed. Analyses were also performed during cold acclimation and reacclimation of plants characterized by different elongation growth rate triggered by prehardening or deacclimation under different day-temperatures. Growth under high PSII excitation pressure (prehardening) resulted in a significant increase in ABA and a considerable decrease in GAs contents. On the other hand, different ABA content played almost no role in controlling growth rate during cold deacclimation and subsequent reacclimation, when the induction of elongation growth was connected with the changes in concentration of GAs including GA3. The possible role of ABA and GAs in controlling prehardening, cold acclimation and deacclimation is discussed.

Reorganization of the microtubule and actin cytoskeleton in root cells of Triticum aestivum L during low temperature and abscisic acid treatments.

Information on the effects of low temperatures and abscisic acid (ABA) on the structural organization of MTs and MFs is scarce and contradictory (Chu et al., 1993; Eun and Lee, 1997; Jiang et al., 1996; PihakaskiMaunsbach and Puhakainen, 1995; Rikin et al., 1983; Sakiyama and Shibaoka, 1990). Nothing is known about the interaction of these components on the cold acclimation of plants. Seedling roots of winter wheat cultivars differing in their frost resistance—Bezostaya 1 (low frost resistant), Mironovskaya 808 (moderately frost resistant) and Albidum 114 (highly frost resistant)—were the objects of the investigation. The plants were grown in hydroculture at an illumination of 100 W/m and with a photoperiod of 12 h. Nonacclimated plants were grown at 23 (C for 5 or 9 days. ABA (30 μM) was added to the 7-day-old plant roots and the plants were grown in ABA solution for 3 days. For cold acclimation, 8-day-old plants were transferred to 3 (C for 7 days. In treatments, in which the effect of ABA was studied together with cold acclimation, ABA was added 1 day before the beginning of acclimation. The visualization of the cytoskeleton was conducted by indirect immunofluorescent microscopy using monoclonal tubulin and actin antibodies (Baluska et al., 1997). In the cold-acclimated and ABA-treated winter wheat plants, the comparative investigation of the state (localization, orientation, structure) and the stability of actin and microtubule cytoskeleton have been done in the cells of different root zones. Cold acclimation caused the aggregation of MTs; it increased the fluorescence and stability of microfilaments (MFs) and MTs mainly in the zone of root differentiation. This could testify to the strengthening of the contacts between MTs and MFs. The stability of MTs was estimated by their response to the depolymerizing action of oryzalin. The roots were incubated in a solution of oryzalin (10 μM) for 3 h. To define the stability of MTs in cells of the differentiation zone, a quantitative approach was used. For each of the four variants (without acclimation, cold acclimation, ABA without acclimation, ABA with acclimation), 25 sections of several roots were investigated, and the number of sections, containing intact MTs in differentiating cells, was calculated. As in cold acclimation, ABA induced the formation of MT bunches, but in cells of the meristem and the elongation zones only. However, in the zone of differentiation, an increase in the stability of MT structures correlated with a decrease in the content, the degree of aggregation and the immunofluorescence of MTs and with the complete depolymerization of MFs. It was shown that the stabilizing effect of ABA on MTs depended on the degree of plant frost resistance (Fig. 1). For this reason, the ABA-induced increase of MT stability may be used as a diagnostic marker of frost resistance in different winter wheat cultivars. It was revealed that plant cold acclimation removed the effect of hormone on the structural organization of MTs and MFs in the zone of differentiation. It is suggested that such processes as depolymerization of MTs, a decrease of the number of unstable MT populations and an increase in the number of stable ones cause the slowing down of the growth of ABAtreated plants or plants at the initial stage of cold acclimation. At the end of the plant cold acclimation, the induction of plant growth is evidently determined by an increase of the number of dynamic/unstable populations of MTs and by an increase of polymerization of MFs. * Corresponding author. Tel.: +7-8432-32-51-45; fax: +7-8432-38-74-18. E-mail address: olga.olinevich@ksu.ru (L.P. Khokhlova). Cell Biology International 27 (2003) 211–212 Cell Biology International

Enhancement of cold tolerance and inhibition of lipid peroxidation by citrus dehydrin in transgenic tobacco.

Citrus ( Citrus unshiu Marcov.) dehydrin in response to chilling stress was overexpressed in tobacco ( Nicotiana tabacum L.), and the cold stress tolerance of transgenics at low temperature was analyzed. The freezing at -4 degrees C for 3 h of 24 independent lines indicated that a phenotype expressing citrus dehydrin showed less electrolyte leakage than the control. Dehydrin protein content was correlated with freezing tolerance in transgenics. Dehydrin-expressing tobacco exhibited earlier germination and better seedling growth than the control at 15 degrees C. Cell fractionation experiments suggested that the protein was predominantly expressed in mitochondria and the soluble fraction. Malondialdehyde production enhanced by chilling stress was lower in tobacco plants expressing citrus dehydrin than in control phenotypes. Dehydrin protein, purified from Escherichia coli expressing citrus dehydrin cDNA, prevented peroxidation of soybean ( Glycine max L.) liposomes in vitro. The inhibitory activity of dehydrin against liposome oxidation was stronger than that of albumin, glutathione, proline, glycine betaine, and sucrose. These results suggest that dehydrin facilitates plant cold acclimation by acting as a radical-scavenging protein to protect membrane systems under cold stress.

Abscisic acid-induced freezing tolerance in the moss Physcomitrella patens is accompanied by increased expression of stress-related genes

Abscisic acid (ABA)-induced genes are implicated in the development of freezing tolerance during cold acclimation in higher plants, but their roles in lower land plants have not been determined. We examined ABA- and cold-induced changes in freezing tolerance and gene expression in the moss Physcomitrella patens. Slow equilibrium freezing to -4 degrees C of P. patens protonemata grown under normal growth conditions killed more than 90% of the cells, indicating that the protonema cells are freezing-sensitive. ABA treatment for 24 h dramatically increased the freezing tolerance of the protonemata, while cold treatment only slightly increased the freezing tolerance within the same period. We examined the expressions of fourteen Physcomitrella patens ABA-responsive genes (PPARs), isolated from ABA-treated protonemata. ABA treatment resulted in a remarkable increase in the expression of all the PPAR genes within 24 h. Several of the PPAR genes (PPAR 1 to 8, and 14) were also responsive to cold, but the response was much slower than that to ABA. Treatment with hyperosmotic concentrations of NaCl and mannitol increased freezing tolerance of protonemata and also increased the expression levels of eleven PPAR genes (PPAR2, 3, 5 to 8, and 10 to 14). These results suggest that ABA and environmental stresses positively affect the expression of common genes that participate in protection of protonema cells leading to the development of freezing tolerance.

Seasonal regulation of a 24-kDa protein from red-osier dogwood (Cornus sericea) xylem.

Previously, we showed that an apparent cell wall-plasma membrane interaction in xylem ray parenchyma differed between cold acclimated and non-acclimated red-osier dogwood (Cornus sericea L.) (Ristic and Ashworth 1994). For the present study, a calcium chloride extraction method was used to identify cell-wall-associated xylem proteins that accumulated during periods of cold acclimation. A 24-kDa protein represented the predominant protein in both total protein and CaCl2 extracts during cold acclimation of field-grown plants. Two-dimensional gel electrophoresis separated the 24-kDa protein into four basic isoforms. The most abundant and basic isoform had a high glycine content. In-gel digestion of this basic 24-kDa isoform generated three partial peptide fragments, of which one exhibited homology to the dehydrin protein family. An anti-dehydrin polyclonal antibody cross-reacted with the 24-kDa protein, providing further evidence that this protein is related to dehydrins. The 24-kDa protein began to accumulate in late August, reached a maximum in midwinter, declined during the spring months and was absent in early summer.

Characterisation of a plasma membrane-associated phospholipase A 2 activity increased in response to cold acclimation

We here demonstrate the presence of a plasma membrane-associated phospholipase A(2) (EC 3.1.1.4; PLA(2)) activity in spinach (Spinacia oleracea) leaves. The pH profile of the spinach plasma membrane PLA(2) activity revealed two peaks, one at pH 4.4 and one at pH 5.5. The activity at pH 5.5 had an absolute requirement of Ca2+, with full enzyme activity at 10mumol/L Ca2+. The Ca2+-dependent PLA(2) activity was both heat sensitive and stimulated by diacylglycerol, whereas ATP completely inhibited the activity. Thus, the spinach plasma membrane contains a Ca2+-dependent PLA(2) activity, which has not previously been characterised in plants. Cold acclimation of spinach resulted in a 2.2-fold higher plasma membrane PLA(2) activity whereas the plasma membrane phospholipase D activity remained unaffected. Taken together, our data suggest a role of PLA(2) in cold acclimation in plants. (Less)

Role of glutathione in adaptation and signalling during chilling and cold acclimation in plants.

Glutathione is an important component of the ascorbate-glutathione cycle, which is involved in the regulation of hydrogen peroxide (H2O2) concentrations in plants. During chilling and cold acclimation, i.e. exposure to temperatures between 0 and 15 degrees C, H2O2 may accumulate. Excess electrons from the photosynthetic and respiratory electron transport chains can be used for the reduction of oxygen, thus producing superoxide radicals (O2.-); these are subsequently transformed to H2O2 via superoxide dismutase (SOD; EC 1.15.1.1). During the removal of excess H2O2, reduced glutathione (GSH) is converted to its oxidised form (GSSG), and GSH is regenerated by the activity of NADPH-dependent glutathione reductase (GR; EC 1.6.4.2). At low non-freezing temperatures, high GSH content and GR activity were detected in several plant species, indicating a possible contribution to chilling tolerance and cold acclimation. Changes in H2O2 concentration and GSH/GSSG ratio alter the redox state of the cells and may activate special defence mechanisms through a redox signalling chain. The finding that several defence genes have antioxidant responsive elements or GSSG binding sites in their regulatory regions supports the idea that redox signalling is involved in regulating gene expression in response to low temperature.

Ectopic expression of ABI3 gene enhances freezing tolerance in response to abscisic acid and low temperature in Arabidopsis thaliana

SummaryThe plant hormone abscisic acid (ABA) regulates several physiological and developmental processes in plants, including stress adaptation and seed maturation. ABA‐mediated processes appear to be central in plant cold acclimation and expression of cold acclimation‐related genes. Ectopic expression of ABI3 encoding a seed‐specific transcriptional activator confers on Arabidopsis vegetative tissues the ability to accumulate seed‐specific transcripts in response to ABA, and also influences some ABA‐mediated vegetative responses. In the present study we characterized the effect of ectopic expression of ABI3 on cold acclimation and development of freezing tolerance in Arabidopsis. We first determined the effect of ABI3 on ABA‐induced expression of cold acclimation‐related genes. Expression of ABI3 increased the ABA‐induced accumulation of transcripts for several ABA/cold/drought‐responsive genes such as RAB18 and LTI78. Enhanced expression of these genes was evident even after transient application of ABA, and the enhanced expression was correlated with increased freezing tolerance in ABI3 transgenic plants. Ectopic expression of ABI3 also appeared to modulate low temperature‐induced freezing tolerance. The ABI3 transgenic plants acclimated faster than the wild‐type plants, and the maximum tolerance obtained was significantly higher. These data showed that lower levels of ABA were needed to trigger the expression of the genes and to maintain the freezing‐tolerant state in the ABI3 transgenic plants, and indicate that ectopic expression of ABI3 leads to enhanced responsiveness to ABA. The ectopic expression of ABI3 could provide a new strategy for engineering plant stress tolerance.

Ectopic expression of ABI3 gene enhances freezing tolerance in response to abscisic acid and low temperature in Arabidopsis thaliana.

The plant hormone abscisic acid (ABA) regulates several physiological and developmental processes in plants, including stress adaptation and seed maturation. ABA-mediated processes appear to be central in plant cold acclimation and expression of cold acclimation-related genes. Ectopic expression of ABI3 encoding a seed-specific transcriptional activator confers on Arabidopsis vegetative tissues the ability to accumulate seed-specific transcripts in response to ABA, and also influences some ABA-mediated vegetative responses. In the present study we characterized the effect of ectopic expression of ABI3 on cold acclimation and development of freezing tolerance in Arabidopsis. We first determined the effect of ABI3 on ABA-induced expression of cold acclimation-related genes. Expression of ABI3 increased the ABA-induced accumulation of transcripts for several ABA/cold/drought-responsive genes such as RAB18 and LTI78. Enhanced expression of these genes was evident even after transient application of ABA, and the enhanced expression was correlated with increased freezing tolerance in ABI3 transgenic plants. Ectopic expression of ABI3 also appeared to modulate low temperature-induced freezing tolerance. The ABI3 transgenic plants acclimated faster than the wild-type plants, and the maximum tolerance obtained was significantly higher. These data showed that lower levels of ABA were needed to trigger the expression of the genes and to maintain the freezing-tolerant state in the ABI3 transgenic plants, and indicate that ectopic expression of ABI3 leads to enhanced responsiveness to ABA. The ectopic expression of ABI3 could provide a new strategy for engineering plant stress tolerance.

Light utilization in photosynthesis after cold acclimation of plants representing two different seasonal growth strategies

Seedlings of Lodgepole pine (Pinus contorta L.) and winter wheat (Triticum aestivum L.) were cold acclimated under controlled conditions. Both temperate conifers and winter cereals have the ability to acquire high freezing tolerance during cold acclimation, and they are characterized by maintaining their green foliages during autumn and winter. Pine responds to cold acclimation by partial inhibition of photosynthesis with a low daily carbon gain, while winter wheat maintains a high activity of photosynthesis and a high daily carbon gain during cold acclimation. These different responses of photosynthesis to cold acclimation are correlated with pine reducing its need for assimilates when entering dormancy associated with termination of primary growth, while winter wheat maintains a high need for assimilates as it continues to grow and develop throughout cold acclimation. These different responses are accompanied by pine reducing its needle content of chlorophyll and developing a high and sustained ability for non-photochemical dissipation of absorbed light as heat during cold acclimation, while winter wheat show no such response. The increased capacity for non-photochemical dissipation of absorbed light energy in pine, as photosynthesis becomes inhibited by a limited sink capacity, and eventually by freezing temperatures, is viewed as an important strategy for avoiding photo-oxidative damage of the needle foliage during autumn and winter. In winter cereals, on the other hand, with its highly efficient photosynthesis at chilling temperatures through the autumn, the prevailing high yield of photochemistry can efficiently dissipate absorbed light, and no sustained non-photochemical dissipation of absorbed light is required.

So what's new in the field of plant cold acclimation? Lots!

Due to its intrinsically interesting nature and importance to agriculture, considerable effort has been directed at understanding the phenomenon of “cold acclimation,” the process whereby certain plants increase in freezing tolerance upon exposure to low temperature. By 1980, thousands of

147 A 27-kDa Rhododendron Deacclimation Protein is Related to ABA Stress Ripening/Water Deficit Stress Inducible Family of Proteins

Many reports have shown the accumulation of specific proteins associated with cold acclimation in plants. However, there is a scarcity of data on the physiological and/or biochemical changes associated with deacclimation process. This study was initiated to determine protein changes specifically associated with deacclimation in Rhododendron. Current-year leaves were collected from three Rhododendron cultivars (`Chionoides', `Grumpy Yellow', and `Vulcanís Flame'; ≈4-year-old rooted cuttings) during natural non-acclimated (June), cold-acclimated (January), and deacclimated (May) state. Leaf freezing tolerance was evaluated using controlled freezing protocol (Lim et al. 1998, J. Amer. Soc. Hort. Sci. 123:246–252). Seasonal SDS-PAGE profiles exhibited a distinct accumulation of 27 kDa protein in deacclimated and nonacclimated tissues, but this protein was essentially undetectable in cold acclimated tissues of all three cultivars. Further characterization of this polypeptide, labeled as RhDAP27 (for rhododendron deacclimation protein), revealed that it has an iso-electric point of 6.5, has a compositional bias for Glu/Gln (13.9%), His (11.4%), Gly (11%), Ala (10%), Lys (8.3%), and Asp/Asn (8.1%)—hydrophilic amino acids constitutedabout 54% of the total amino acids while 40% were nonpolar, aliphatic amino acids (Gly, Ala, Val, Leu, Ile, Pro) and only 6% were aromatic amino acids (Phe and Tyr). Micro-sequencing of the four peptides produced by partial cleavage of RhDAP27 revealed a striking homology of RhDAP27 with two proteins (from Mesembryanthemum crystallinum and Pinus taeda) that belong to the family of ABA stress ripening/water deficit stress inducible proteins.

Glycine betaine accumulation and induction of cold tolerance in strawberry ( Fragaria X ananassa Duch.) plants

Endogenous glycine betaine levels in the leaves of strawberry ( Fragaria X ananassa Duch.) plants were determined during cold acclimation and in response to exogenous abscisic acid (ABA) application. Glycine betaine levels in the leaves increased nearly two-fold after 4 weeks of cold acclimation treatment during which the cold tolerance of leaves increased from −5.8 to −17°C. Exogenous application of ABA (100 μM) to plants triggered glycine betaine accumulation in unhardened plants. It also increased cold tolerance of leaves in both unhardened and cold-hardened plants. Similar to ABA, exogenous glycine betaine was effective in inducing cold tolerance in unhardened and cold-hardening plants. Exogenous application of glycine betaine (2 mM) to unhardened plants increased the cold tolerance of leaves almost two-fold within 72 h of application. In addition, it improved freezing survival and regrowth in whole plants. The results suggest that glycine betaine is involved in inducing cold tolerance both in response to exogenous ABA and during natural cold acclimation of strawberry plants.