Cold deacclimation mechanisms and reacclimation potential in flower buds of blackcurrant (Ribes nigrum)

As a consequence of global climate change, cold acclimation and deacclimation cycles are becoming increasingly frequent during winter in temperate regions. However, little is known about plant deacclimation and in particular reacclimation mechanisms, although deacclimation resistance and the ability to reacclimate may have wide‐ranging consequences regarding plant productivity in a changing climate. Here, we report time‐dependent responses of freezing tolerance, respiration rates, metabolite contents (high‐resolution magic angle spinning NMR) and fatty acid levels (gas chromatography) in flower buds of two ecodormant Ribes nigrum cultivars exposed to three different deacclimation temperatures followed by a reacclimation treatment at 4°C. The data reveal that despite differences in the progression of deacclimation, the capacity of blackcurrant flower buds to reharden in late winter is virtually non‐existing, implying that increasingly irregular temperature patterns is critical for blackcurrant fruit yield. The early phase of deacclimation is associated with a transient increase in respiration and decreasing contents of amino acids, tricarboxylic acid (TCA) cycle intermediates and sugars, indicating an increased need for carbon sources and respiratory energy production for the activation of growth. Decreasing sugar levels may additionally cause loss of freezing tolerance. Deacclimation also involves desaturation of membrane lipids, which likely also contributes to decreased freezing tolerance but may also reflect biosynthesis of signaling molecules stimulating growth and floral organ differentiation. These data provide new insights into the under‐researched deacclimation mechanisms and the ability of blackcurrant to reacclimate following different advancements of deacclimation and contribute to our understanding of plant responses to increasingly irregular temperature patterns.     

Changes in carbohydrates, ABA and bark proteins during seasonal cold acclimation and deacclimation in Hydrangea species differing in cold hardiness

Cold injury is frequently seen in the commercially important shrub Hydrangea macrophylla but not in Hydrangea paniculata. Cold acclimation and deacclimation and associated physiological adaptations were investigated from late September 2006 to early May 2007 in stems of field-grown H. macrophylla ssp. macrophylla (Thunb.) Ser. cv. Blaumeise and H. paniculata Sieb. cv. Kyushu. Acclimation and deacclimation appeared approximately synchronized in the two species, but they differed significantly in levels of mid-winter cold hardiness, rates of acclimation and deacclimation and physiological traits conferring tolerance to freezing conditions. Accumulation patterns of sucrose and raffinose in stems paralleled fluctuations in cold hardiness in both species, but H. macrophylla additionally accumulated glucose and fructose during winter, indicating species-specific differences in carbohydrate metabolism. Protein profiles differed between H. macrophylla and H. paniculata, but distinct seasonal patterns associated with winter acclimation were observed in both species. In H. paniculata concurrent increases in xylem sap abscisic acid (ABA) concentrations ([ABA](xylem)) and freezing tolerance suggests an involvement of ABA in cold acclimation. In contrast, ABA from the root system was seemingly not involved in cold acclimation in H. macrophylla, suggesting that species-specific differences in cold hardiness may be related to differences in [ABA](xylem). In both species a significant increase in stem freezing tolerance appeared long after growth ceased, suggesting that cold acclimation is more regulated by temperature than by photoperiod.     

Changes in Sugar Content during Cold Acclimation and Deacclimation of Cabbage Seedlings

The relationship between freezing tolerance and sugar contentin cabbage seedlings was investigated. Seedlings exposed tonon-freezing low temperature (5 °C) acquired freezing tolerancedown to -6 °C. The degree of freezing tolerance increasedwith duration of exposure to low temperature (up to 10 d). Sucrose,glucose, fructose and myo -inositol were detected as solublesugars in cabbage leaves, and all soluble sugars, except formyo -inositol, and starch increased gradually during cold acclimationsuch that their levels were positively correlated with the degreeof freezing tolerance. The induced freezing tolerance was attributednot to ontogenetic changes but to cold acclimation. However,the induced freezing tolerance was lost after only 1 d of deacclimationat control temperatures, and this change was associated witha large reduction in sugar content. These results reveal that the sugar content of cabbage leavesis positively correlated with freezing tolerance. Brassica oleracea L.; cabbage; cold acclimation; deacclimation; freezing tolerance; sugars     

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.     

Monitoring expression profiles of Arabidopsis genes during cold acclimation and deacclimation using DNA microarrays

A comparative analysis of gene expression profiles during cold acclimation and deacclimation is necessary to elucidate the molecular mechanisms of cold stress responses in higher plants. We analyzed gene expression profiles in the process of cold acclimation and deacclimation (recovery from cold stress) using two microarray systems, the 7K RAFL cDNA microarray and the Agilent 22K oligonucleotide array. By both microarray analyses, we identified 292 genes up-regulated and 320 genes down-regulated during deacclimation, and 445 cold up-regulated genes and 341 cold down-regulated genes during cold acclimation. Many genes up-regulated during deacclimation were found to be down-regulated during cold acclimation, and vice versa. The genes up-regulated during deacclimation were classified into (1) regulatory proteins involved in further regulation of signal transduction and gene expression and (2) functional proteins involved in the recovery process from cold-stress-induced damages and plant growth. We also applied expression profiling studies to identify the key genes involved in the biosynthesis of carbohydrates and amino acids that are known to play important roles in cold acclimation. We compared genes that are regulated during deacclimation with those regulated during rehydration after dehydration to discuss the similarity and difference of each recovery process.Electronic Supplementary Material Supplementary materials are available for this article at     

Induction of freezing tolerance in spinach is associated with the synthesis of cold acclimation induced proteins

Spinach (Spinacia oleracea L. cv Bloomsdale) seedlings cultured in vitro were used to study changes in protein synthesis during cold acclimation. Seedlings grown for 3 weeks postsowing on an inorganic-nutrient-agar medium were able to increase their freezing tolerance when grown at 5°C. During cold acclimation at 5°C and deacclimation at 25°C, the kinetics of freezing tolerance induction and loss were similar to that of soil-grown plants. Freezing tolerance increased after 1 day of cold acclimation and reached a maximum within 7 days. Upon deacclimation at 25°C, freezing tolerance declined within 1 day and was largely lost by the 7th day. Leaf proteins of intact plants grown at 5 and 25°C were in vivo radiolabeled, without wounding or injury, to high specific activities with [³⁵S]methionine. Leaf proteins were radiolabeled at 0, 1, 2, 3, 4, 7, and 14 days of cold acclimation and at 1, 3, and 7 days of deacclimation. Up to 500 labeled proteins were separated by two-dimensional gel electrophoresis and visualized by fluorography. A rapid and stable change in the protein synthesis pattern was observed when seedlings were transferred to the low temperature environment. Cold-acclimated leaves contained 22 polypeptides not found in nonacclimated leaves. Exposure to 5°C induced the synthesis of three high molecular weight cold acclimation proteins (CAPs) (M_r of about 160,000, 117,000, and 85,000) and greatly increased the synthesis of a fourth high molecular weight protein (M_r 79,000). These proteins were synthesized during day 1 and throughout the 14 day exposure to 5°C. During deacclimation, the synthesis of CAPs 160, 117, and 85 was greatly reduced by the first day of exposure to 25°C. However, CAP 79 was synthesized throughout the 7 day deacclimation treatment. Thus, the induction at low temperature and termination at warm temperature of the synthesis of CAPs 160, 117, and 85 was highly correlated with the induction and loss of freezing tolerance. Cold acclimation did not result in a general posttranslational modification of leaf proteins. Most of the observed changes in the two-dimensional gel patterns could be attributed to the de novo synthesis of proteins induced by low temperature. In spinach leaf tissue, heat shock altered the pattern of protein synthesis and induced the synthesis of several heat shock proteins (HSPs). One polypeptide synthesized in cold-acclimated leaves had a molecular weight and net charge (M_r 79,000, pI 4.8) similar to that of a HSP (M_r 83,000, pI 4.8). However, heat shock did not increase the freezing tolerance, and cold acclimation did not increase heat tolerance over that of nonacclimated plants, but heat-shocked leaf tissue was more tolerant to high temperatures than nonacclimated or cold-acclimated leaf tissue. When protein extracts from heat-shocked and cold-acclimated leaves were mixed and separated in the same two-dimensional gel, the CAP and HSP were shown to be two separate polypeptides with slightly different isoelectric points and molecular weights.     

Differential remodeling of the lipidome during cold acclimation in natural accessions of Arabidopsis thaliana

Freezing injury is a major factor limiting the geographical distribution of plant species and the growth and yield of crop plants. Plants from temperate climates are able to increase their freezing tolerance during exposure to low but non‐freezing temperatures in a process termed cold acclimation. Damage to cellular membranes is the major cause of freezing injury in plants, and membrane lipid composition is strongly modified during cold acclimation. Forward and reverse genetic approaches have been used to probe the role of specific lipid‐modifying enzymes in the freezing tolerance of plants. In the present paper we describe an alternative ecological genomics approach that relies on the natural genetic variation within a species. Arabidopsis thaliana has a wide geographical range throughout the Northern Hemisphere with significant natural variation in freezing tolerance that was used for a comparative analysis of the lipidomes of 15 Arabidopsis accessions using ultra‐performance liquid chromatography coupled to Fourier‐transform mass spectrometry, allowing the detection of 180 lipid species. After 14 days of cold acclimation at 4°C the plants from most accessions had accumulated massive amounts of storage lipids, with most of the changes in long‐chain unsaturated triacylglycerides, while the total amount of membrane lipids was only slightly changed. Nevertheless, major changes in the relative amounts of different membrane lipids were also evident. The relative abundance of several lipid species was highly correlated with the freezing tolerance of the accessions, allowing the identification of possible marker lipids for plant freezing tolerance.     

Variation in freezing tolerance,water content and carbohydrate metabolism of floral buds during deacclimation of contrasting blackcurrant cultivars

As a result of climate change, temperature patterns are expected to become increasingly irregular with longer and more frequent episodes of unseasonable warm spells during the winter season. Warm spells may promote premature loss of freezing tolerance and bud burst in woody perennials, thereby increasing the risk of tissue damage by subsequent frosts. This study investigated the variation in kinetics of deacclimation and bud break and associated changes in carbohydrate metabolism and water status in floral buds of six blackcurrant (Ribes nigrum) cultivars in response to a simulated warm spell (16/11 °C day/night). In three of the cultivars, the rate of deacclimation showed an almost logarithmic course, whereas the other three cultivars exhibited greater deacclimation resistance and a sigmoid deacclimation pattern. The timing and rate of bud development, and their relationship with deacclimation varied greatly amongst cultivars, indicating genotypic variation in time-dependent responses of freezing tolerance and bud break to warm temperatures. In all six cultivars, deacclimation and growth resumption were strongly associated with rehydration. In contrast, changes in carbohydrate metabolism were mostly associated with deacclimation. Evaluation of phenological responses of the same cultivars under field conditions showed that cultivars which were fast flushing in response to an experimental warm spell also exhibited early bud break under natural conditions, indicating that cultivar differences in phenological responses are consistent under different temperature conditions.     

Changes in glutathione content during cold acclimation in Cornus sericea and Citrus sinensis

Glutathione content was evaluated in relation to freezing tolerance in red osier dogwood stems and Valencia orange leaves. Exposure of dogwood and citrus to cold-acclimating conditions in controlled environments led to increases in reduced glutathione (GSH) content which were correlated with freezing tolerance. GSH did not accumulate in field-grown dogwood stems during cold acclimation in fall, but did increase in content prior to deacclimation in late winter. Further studies showed that accumulation of GSH in dogwood at low temperatures is dependent on adequate levels of sulfate in the soil. In citrus, modulation of GSH content by infiltration of leaf tissue with various compounds including GSH did not alter freezing tolerance. Root treatment with N,N-diallyl-2,2-dichloroacetamide (R-25788) increased leaf GSH content, but not hardiness. Evidence presented indicates that glutathione accumulates in plant tissues exposed to low temperatures, but that GSH accumulation is not associated with freezing tolerance.     

Comparison of freezing tolerance in cultured plant cells and their respective protoplasts

The possibility that the plant cell wall influences the severity of freezing injury was examined by comparing the freeze stress response of intact cells and protoplasts from four different suspension cultures. In no case did the intact cells suffer more injury than the respective wall-less protoplasts, showing that mechanical strain imposed by the cell wall during freeze-thaw stress is not a major determinant of injury. For three of the four species studied, cells from which the wall was removed showed significantly greater freezing injury, indicating that the plant cell wall may have a protective role. Other researchers have suggested that cell wall rigidity may minimize freezing injury by slowing freeze-induced loss of cell water. We found that decreased enzyme digestibility (perhaps indicating greater rigidity) of cell walls accompanied cold acclimation in various tissues. These results provide impetus to research which will characterize low-temperature-induced cell wall modification in cold acclimating tissues.     

Differential degradation of extraplastidic and plastidic lipids during freezing and post-freezing recovery in Arabidopsis thaliana

Changes in membrane lipid composition play important roles in plant adaptation to and survival after freezing. Plant response to cold and freezing involves three distinct phases: cold acclimation, freezing, and post-freezing recovery. Considerable progress has been made toward understanding lipid changes during cold acclimation and freezing, but little is known about lipid alteration during post-freezing recovery. We previously showed that phospholipase D (PLD) is involved in lipid hydrolysis and Arabidopsis thaliana freezing tolerance. This study was undertaken to determine how lipid species change during post-freezing recovery and to determine the effect of two PLDs, PLDalpha1 and PLDdelta, on lipid changes during post-freezing recovery. During post-freezing recovery, hydrolysis of plastidic lipids, monogalactosyldiacylglycerol and plastidic phosphatidylglycerol, is the most prominent change. In contrast, during freezing, hydrolysis of extraplastidic phospholipids, phosphatidylcholine and phosphatidylethanolamine, occurs. Suppression of PLDalpha1 decreased phospholipid hydrolysis and phosphatidic acid production in both the freezing and post-freezing phases, whereas ablation of PLDdelta increased lipid hydrolysis and phosphatidic acid production during post-freezing recovery. Thus, distinctly different changes in lipid hydrolysis occur in freezing and post-freezing recovery. The presence of PLDalpha1 correlates with phospholipid hydrolysis in both freezing and post-freezing phases, whereas the presence of PLDdelta correlates with reduced lipid hydrolysis during post-freezing recovery. These data suggest a negative role for PLDalpha1 and a positive role for PLDdelta in freezing tolerance.     

植物抗寒及其基因表达研究进展

植物经过逐渐降低的温度从而提高抗寒能力 ,这个过程被人们称为低温驯化。植物低温驯化过程是一个复杂的生理、生化和能量代谢变化过程 ,这些变化主要包括膜系统的稳定性、可溶性蛋白的积累和小分子渗透物质 ,比如脯氨酸、糖等 ,这些变化中的一些是植物抗寒必需的 ,而另外一些变化不是必需的。主要对冷害和低温生理生化变化、低温诱导表达基因的功能和作用、低温驯化的调节机制及其信号转导方面进行了综述。通过差别筛选 c DNA文库的方法已经鉴定了许多低温诱导表达、进而提高植物抗寒能力的基因 ,其中有脱水素、COR基因和 CBF1转录因子等。低温信号的感受、转导和调节表达是低温驯化的关键环节 ,低温信号的转导过程与干旱胁迫之间具有一定的交叉 ,这为利用 ABA等来提高植物抗寒能力成为可能 ,相信不久的将来人们可以通过提高植物抗寒能力从而增加经济产量成为现实。     

Freezing tolerance, cold acclimation and oxidative stress in potato. Paraquat tolerance is related to acclimation but is a poor indicator of freezing tolerance

Potato is a species commonly cultivated in temperate areas where the growing season may be interrupted by frosts, resulting in loss of yield. Cultivated potato, Solanum tuberosum, is freezing sensitive, but it has several freezing-tolerant wild potato relatives, one of which is S. commersonii. Our study was aimed to resolve the relationship between enhanced freezing tolerance, acclimation capacity and capacity to tolerate active oxygen species. To be able to characterize freezing tolerant ideotypes, a potato population (S1), which segregates in freezing tolerance, acclimation capacity and capacity to tolerate superoxide radicals, was produced by selfing a somatic hybrid between a freezing-tolerant Solanum commersonii (LT₅₀=-4.6°C) and -sensitive S. tuberosum (LT₅₀=-3.0°C). The distribution of non-acclimated freezing tolerance (NA-freezing tolerance) of the S1 population varied between the parental lines and we were able to identify genotypes, having significantly high or low NA-freezing tolerance. When a population of 25 genotypes was tested both for NA-freezing and paraquat (PQ) tolerance, no correlation was found between these two traits (R = 0.02). However, the most NA-freezing tolerant genotypes were also among the most PQ tolerant plants. Simultaneously, one of the NA-freezing sensitive genotypes (2022) (LT₅₀=-3.0°C) was observed to be PQ tolerant. These conflicting results may reflect a significant, but not obligatory, role of superoxide scavenging mechanisms in the NA-freezing tolerance of S. commersonii. The freezing tolerance after cold acclimation (CA-freezing tolerance) and the acclimation capacity (AC) was measured after acclimation for 7 days at 4/2°C. Lack of correlation between NA-freezing tolerance and AC (R =-0.05) in the S1 population points to independent genetic control of NA-freezing tolerance and AC in Solanum sp. Increased freezing tolerance after cold acclimation was clearly related to PQ tolerance of all S1 genotypes, especially those having good acclimation capacity. The rapid loss of improved PQ tolerance under deacclimation conditions confirmed the close relationship between the process of cold acclimation and enhanced PQ tolerance. Here, we report an increased PQ tolerance in cold-acclimated plants compared to non-acclimated controls. However, we concluded that high PQ tolerance is not a good indicator of actual freezing tolerance and should not be used as a selectable marker for the identification of a freezing-tolerant genotype.     

Effects of MPK3 and MPK6 kinases on the chloroplast architecture and function induced by cold acclimation in Arabidopsis

Photosynthesis Research - Exposure to low, non-freezing temperatures develops freezing tolerance in many plant species. Such process is called cold acclimation. Molecular changes undergone during...     

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.