Effects of cold-hardening on compatible solutes and antioxidant enzyme activities related to freezing tolerance in Ammopiptanthus mongolicus seedlings

Cold acclimation is associated with many metabolic changes that lead to an increase of freezing tolerance. In order to investigate the biochemical process of cold acclimation in Ammopiptanthus mongolicus, seedlings were acclimated at 2℃ under 16-h photoperiod （150 μmol·m^-2·s^-1 photosynthetically active radiation） for 14 d. Freezing tolerance in seedlings increased after 14 d of cold-hardening. Contents of protein, proline and solute carbohydrate in cotyledon increased after cold acclimation. Patterns of isozymes of superoxide dismutase （SOD）, peroxidase, catalase and polyphenol oxidase （PPO） were investigated. The activities of SOD, peroxidase and PPO in cold acclimated plants were increased during cold-hardening. We deduced that compatible solutes and antioxidant enzymes play important roles in development of freezing tolerance during cold acclimation in this evergreen woody plant.     

低温锻炼与CaCl2处理对毛白杨幼苗可溶性总蛋白、CaM及抗冻性的影响

低温锻炼提高了毛白杨幼苗叶片和枝条中钙调素 (CaM)和可溶性总蛋白含量 ,同时也提高了幼苗的抗冻性 .CaCl2 处理对上述低温锻炼的作用有加强的效果 ,体现在明显提高了CaM和可溶性总蛋白含量以及幼苗抗冻性 ;钙离子螯合剂EGTA、钙离子通道阻断剂LaCl3 或钙调素拮抗剂CPZ处理则减弱了低温锻炼的效应 ,表现出CaM和可溶性总蛋白含量以及幼苗抗冻性明显降低 .进一步研究发现 ,CaCl2 处理的低温锻炼不仅能减轻低温胁迫引起的CaM和可溶性总蛋白含量的下降程度 ,而且有利于幼苗在恢复过程中CaM和可溶性总蛋白水平的迅速回升 .这表明Ca2 + CaM可能参与了CaM和可溶性总蛋白的合成及抗冻性的诱导     

在低温锻炼中毛白杨幼苗可溶性总蛋白、RNA、DNA、RNase及抗冻性的变化

低温锻炼不仅提高了毛白杨幼苗存活率和抗冻性以及RNA和可溶性总蛋白的含量 ,降低了RNase活性 ,而且能减轻低温胁迫引起的RNA和可溶性总蛋白含量的下降程度和RNase活性的提高 ,有利于幼苗在恢复过程中RNA和可溶性总蛋白水平的迅速回升以及RNase活性的降低 .进一步研究发现 ,DNA含量无论在低温锻炼中还是在低温胁迫下或是在随后的恢复生长期均保持相对稳定 ;低温锻炼所引起的RNA含量的增加 ,与RNase活性的降低呈明显的负相关 ,与可溶性总蛋白含量的增加以及幼苗抗冻性的提高成正相关 .这表明低温锻炼可能抑制RNase活性 ,有效地促进RNA含量的增加 ,而RNA可能参与了可溶性总蛋白的合成及抗冻性的诱导     

Influence of ozone on cold acclimation in sugar maple seedlings

During summer 1994, sugar maple (Acer saccharum Marsh.) seedlings were grown in open-top chambers supplied with air containing near ambient ozone concentration (control, low O(3)) or three times the ambient ozone concentration (high O(3)). The rate of CO(2) assimilation was significantly reduced by chronic exposure to a high concentration of ozone during the summer. During fall, seedlings were removed from the open-top chambers and acclimated to cold under natural conditions. In both species during cold acclimation, the starch concentration decreased, whereas the sucrose concentration increased. There was no treatment effect on the freezing tolerance of roots, even though roots in the high-O(3) treatment accumulated higher concentrations of the cryoprotective oligosaccharides raffinose and stachyose than control roots. Cold acclimation occurred earlier and stachyose concentration of stems was higher in high-O(3)-treated seedlings than in low-O(3)-treated seedlings. Cold acclimation was associated with an earlier accumulation of ABA in the xylem sap of high-O(3)-treated seedlings compared with low-O(3)-treated seedlings.     

Ultrastructural and Extracellular Protein Changes in Cell Suspension Cultures of Populus euphratica Associated with Low Temperature-induced Cold Acclimation

Populus euphratica Olive is the only tree species that can grow in the saline land and also survive cold winters in northwest China, and it plays a very important role in stabilizing the vulnerable ecosystem there. A cell suspension culture was initiated from callus derived from plantlets of Populus euphratica. Cold acclimation was induced (LT50 of-17.5 ℃) in cell suspension at 4-5 ℃ in the dark for 30 days and the freezing tolerance increased from LT50 of-12.5 ℃ in nonacclimated cells to LT50 of-17.5 ℃ in cold-acclimated cells. Microvacuolation, cytoplasmic augmentation and accumulation of starch granules were observed in cells that were cold-acclimated by exposure to low temperatures. Several qualitative and quantitative changes in proteins were noted during cold acclimation. Antibodies to carrot extracellular (apoplastic) 36 kD antifreeze protein did not cross react on immunoelectroblots with extracellular proteins in cell suspension culture medium of Populus euphratica, indicating no common epitopes in the carrot 36 kD antifreeze protein and P euphratica extracellular proteins. The relationship of these changes to cold acclimation in Populus euphratica cell cultures was discussed.     

Characterizing the frost sensitivity of black spruce photosynthesis during cold acclimation

We used photosynthetic light response curves to measure and model the responses of two provenances of 3-year-old black spruce (Picea mariana (Mill.) BSP) seedlings to severe artificial frost treatments applied at 2-week intervals during cold acclimation. Black spruce seedlings responded to cold acclimation with long-term suppression of photosynthetic capacity (Amax) and apparent quantum-use efficiency (alpha'). Short-term reductions in both photosynthetic parameters following frost treatments were dependent on the extent of cold acclimation of the seedlings and the severity of the frost treatments. Large reductions in Amax in response to the frost treatments were observed in seedlings that had undergone little cold acclimation and these reductions were associated with an irreversible reduction in alpha'. Such seedlings recovered only partially during the subsequent 23 days, whereas seedlings in most other treatments showed complete recovery of Amax after 13 days. The impact of frost treatments on Amax and alpha' did not vary with seedling provenance. We propose an algorithm that predicts the combined effects of cold acclimation and severe freezing temperatures on the extent of the suppression of A(max) during autumn. The algorithm is based on (1) the maximum Amax observed during the growing season, (2) the accumulation of cold degree-days, based on a minimum nocturnal temperature < 5 degrees C, and (3) the severity of freezing temperatures during autumn. The parameters developed in the algorithm showed that cold acclimation of black spruce seedlings had a greater impact on the reduction of Amax in autumn than did the severe frost treatments. Mean Amax of seedlings subjected to artificial frosts showed a strong correlation with values predicted by the algorithm (r2 = 0.91).     

毛白杨抗冻性低温诱导过程中糖的效应(英文)

在低温锻炼和结合蔗糖处理的低温锻炼下 ,对毛白杨 (Populustomentosa)幼苗枝条和叶片的可溶性糖含量 ,以及幼苗存活率和抗冻性分别进行了测定 .结果表明 ,低温锻炼明显提高了幼苗枝条和叶片的可溶性糖含量 ,同时也提高了幼苗的存活率和抗冻性 .2 0 %蔗糖处理对上述低温锻炼的作用有加强的效果 .进一步分析发现 ,无论是单纯的低温锻炼还是结合蔗糖预处理的低温锻炼 ,幼苗枝条中可溶性糖含量的提高程度均较叶片明显 ,而且它们的含量增加与幼苗抗冻性的提高密切相关 .这表明低温锻炼所引起的可溶性糖积累可能参与了幼苗抗冻性的诱导     

Detection of a chitinase-like protein in the roots of Douglas-fir trees infected with Armillaria ostoyae and Phellinus weirii

Protein was extracted from root bark of 11- and 25-year-old interior Douglas-fir (Pseudotsuga menziesii (Mirb.) Franco) trees that were naturally infected with Armillaria ostoyae (Romagnesi) Herink. The proteins were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Root bark tissue adjacent to infected areas had a significantly higher protein concentration than healthy tissue (P < 0.05), whereas the protein concentration of infected tissue was consistently lower (P < 0.05) than that of healthy tissue. The SDS-PAGE profiles of healthy, infected, and adjacent-to-infected root bark tissues revealed significant differences in concentrations of a 29.3-kDa protein. The N-terminal amino acid sequence of the 29.3-kDa protein displayed significant homology (P = 0.013) to a basic endochitinase. Use of a polyclonal antibody raised against the 29.3-kDa putative endochitinase-like protein (ECP) indicated differences in the quantities of ECP in healthy roots compared with roots infected with A. ostoyae in 11- and 25-year-old interior Douglas-fir trees. The antibody was also used to screen for the presence of the 29.3-kDa protein in roots of 24-year-old coastal Douglas-fir (Pseudotsuga menziesii var. menziesii) trees that were artificially inoculated with and colonized by Phellinus weirii (Murr.) Gilbn. The amount of ECP was elevated in root bark of coastal Douglas-fir in response to P. weirii infection, although in lower quantities relative to those found in the A. ostoyae-interior Douglas-fir pathosystem. The sequence homology of the ECP with a basic chitinase, together with its increased synthesis in response to two fungal pathogens, indicate a possible role for this protein in the defense of Douglas-fir against fungal pathogens.     

Different responses of northern and southern ecotypes of Betula pendula to exogenous ABA application

We investigated responses of northern and southern ecotypes of silver birch (Betula pendula Roth) to exogenous abscisic acid (ABA) under controlled environmental conditions to determine the role of ABA in cold acclimation and dormancy development. Abscisic acid was sprayed on the leaves and changes in freezing tolerance, determined by the electrolyte leakage test, and bud dormancy were monitored. Applied ABA induced cold acclimation but had no effect on growth cessation in seedlings grown in long day conditions (LD, 24-h photoperiod at 18 degrees C). It enhanced freezing tolerance and accelerated growth cessation in seedlings grown in short day conditions (SD, 12-h photoperiod at 18 degrees C), and slightly enhanced freezing tolerance in seedlings grown at low temperature (LT, 24-h photoperiod at 4 degrees C) in both ecotypes. There were distinct ecotypic differences in ABA-induced cold acclimation and dormancy development. The northern ecotype was more responsive to applied ABA than the southern ecotype, resulting in more rapid development of freezing tolerance in all treatments, and earlier dormancy development in SD. When plants were grown in a photoperiod just above the critical photoperiod for the ecotype (defined as the longest photoperiod that induces growth cessation), applied ABA caused growth cessation and dormancy development. Compared with ABA-treated seedlings grown in SD, dormancy development was delayed in ABA-treated seedlings exposed to a near-critical photoperiod, but even in this treatment dormancy developed faster in the northern ecotype than in the southern ecotype.     

低温锻炼对冷胁迫下油茶幼苗光合速率与抗氧化酶活性的影响

以油茶品种"杂优闽1"为试验材料,以未经低温锻炼的油茶幼苗为对照,研究了油茶幼苗在(10±0.5)℃低温锻炼、(2±0.5)℃冷胁迫和常温恢复期间的光合速率和抗氧化酶活性的变化。结果表明:低温锻炼3 d可明显提高油茶幼苗的抗寒性;冷胁迫3 d,经低温锻炼后的油茶幼苗叶片净光合速率和相对含水量明显高于对照;且其在常温下的恢复也较对照迅速。在低温锻炼和冷胁迫期间,油茶幼苗叶片脯氨酸和可溶性糖含量明显增加,且丙二醛(MDA)含量明显低于对照;经低温锻炼的油茶幼苗叶片超氧化物歧化酶(SOD)、过氧化氢酶(CAT)和抗坏血酸过氧化物酶(APX)活性则均明显高于未经低温锻炼的油茶幼苗,说明渗透调节物质含量增加和保护酶活性增强在低温锻炼诱导油茶幼苗的抗寒性上发挥着重要的作用。     

Changes in freezing tolerance, plasma membrane H+-ATPase activity and fatty acid composition in Pinus resinosa needles during cold acclimation and de-acclimation

It has previously been suggested that plasma membrane ATPase (PM H+-ATPase, EC 3.6.1.3.) is a site of incipient freezing injury because activity increases following cold acclimation and there are published data indicating that activity of PM H+-ATPase is modulated by changes in lipids associated with the enzyme. To test and extend these findings in a tree species, we analyzed PM H+-ATPase activity and the fatty acid (FA) composition of glycerolipids in purified plasma membranes (PMs) prepared by the two-phase partition method from current-year needles of adult red pine (Pinus resinosa Ait.) trees. Freezing tolerance of the needles decreased from -56 degrees C in March to -9 degrees C in May, and increased from -15 degrees C in September to -148 degrees C in January. Specific activity of vanadate-sensitive PM H+-ATPase increased more than two-fold following cold acclimation, despite a concurrent increase in protein concentration. During de-acclimation, decreases in PM H+-ATPase activity and freezing tolerance were accompanied by decreases in the proportions of oleic (18:1) and linoleic (18:2) acids and increases in the proportions of palmitic (16:0) and linolenic (18:3) acids in total glycerolipids extracted from the plasma membrane fraction. This pattern of changes in PM H+-ATPase activity and the 18:1, 18:2 and 18:3 fatty acids was reversed during cold acclimation. In the PM fractions, changes in FA unsaturation, expressed as the double bond index (1 x 18:1 + 2 x 18:2 + 3 x 18:3), were closely correlated with changes in H+-ATPase specific activity (r2 = 0.995). Changes in freezing tolerance were well correlated with DBI (r2 = 0.877) and ATPase specific activity (r2 = 0.833) in the PM fraction. Total ATPase activity in microsomal fractions also closely followed changes in freezing tolerance (r2 = 0.969). We conclude that, as in herbaceous plants, simultaneous seasonal changes in PM H+-ATPase activity and fatty acid composition occur during cold acclimation and de-acclimation in an extremely winter hardy tree species under natural conditions, lending support to the hypothesis that FA-regulated PM H+-ATPase activity is involved in the cellular response underlying cold acclimation and de-acclimation.     

Monolignol oxidation by xylem peroxidase isoforms of Norway spruce (Picea abies) and silver birch (Betula pendula)

We partially purified peroxidase isoform fractions from xylem extracts of a gymnosperm, Norway spruce (Picea abies (L.) Karst.), and an angiosperm, silver birch (Betula pendula Roth.), to determine the participation of xylem-localized peroxidases in polymerization of different types of lignin in vivo. Several peroxidase fractions varying in isoelectric point values from acidic to basic were tested for their ability to catalyze the oxidation of the monolignols coniferyl alcohol, sinapyl alcohol and p-coumaryl alcohol in vitro. All of the xylem peroxidases extracted from Norway spruce and most of those from silver birch showed the highest rate of oxidation with coniferyl alcohol in the presence of hydrogen peroxide. The exception was an acidic peroxidase fraction (pI 3.60-3.65) from silver birch that exhibited higher oxidation activity for sinapyl alcohol than for coniferyl alcohol. For the xylem enzyme fractions extracted from silver birch, the ability to oxidize the artificial phenolic substrate syringaldazine coincided with high specific activity for sinapyl alcohol. Therefore, we conclude that the acidic, neutral and basic xylem peroxidases of Norway spruce all function in the synthesis of guaiacyl-type lignin, whereas in silver birch the acidic peroxidases preferentially oxidize sinapyl subunits. The latter provides a mechanism for synthesis of guaiacyl-syringyl lignin typical of tracheid cell walls in angiosperm trees.     

Marker trait association for autumn cold acclimation and growth rhythm in pinus sylvestris

The main objective of this study was to identify quantitative trait loci (QTLs) involved in the genetic control of growth rhythm and autumn cold acclimation in Pinus sylvestris L. Eighteen full‐sib families of P. sylvestris with parental origins between latitudes 62 and 67° N in Sweden were assessed for autumn cold acclimation (frost injury measured from artificial freeze testing) and growth rhythm during the first two growing seasons. One family north × south with 108 individuals was used for the construction of one linkage map for the male and one for the female parent using 286 markers from random amplification of polymorphic DNA. Marker trait associations (QTLs) for shoot elongation, growth cessation and cold acclimation were found on both maps. Some QTLs were verified in two additional north × south families. Proportions of the additive genetic variance explained by the markers (R ²/_G) indicated that genetic factors of large effect were involved in all investigated traits. Single or multiple markers accounted for between 16 and 47% of the additive variance in annual shoot elongation, between 42 and 79% of the additive variance in growth termination, and between 7 and 31% of the additive variation in autumn cold acclimation. The study confirms that there are major QTL regions on different linkage groups controlling a large part of the variation for growth rhythm and autumn cold acclimation in P. sylvestris.     

Soil water content and water relations in planted and naturally regenerated Pinus halepensis Mill. seedlings during the first year in semiarid conditions

The capacity of Aleppo pine (Pinus halpensis Mill.) seedlings to overcome the planting shock in dry conditions was assessed by firstly studying the survival and water status during the first year after planting in relation to soil water content. In spite of receiving only 177 mm rainfall during the year studied, survival of planted Aleppo pine was very high (88.5%). Soil water during summer months (after receiving 67 mm rainfall in winter and spring) was only available at 30–60 cm depth, with tension values of –1.1 and –1.3 MPa in July and August respectively; in these conditions, July predawn xylem water potential measured was –2.5 MPa, and midday potential was –3.6 MPa. According to different authors, these values don't jeopardize the survival of Aleppo pine. In addition, acclimation of outplanted seedlings to environmental conditions was followed by comparing their water relations with those of naturally regenerated seedlings on the site. Predawn and midday xylem water potential showed differences in favour of outplanted seedlings since June, indicating an adjustment to this dry site. Compared to naturally regenerated trees, nursery grown stock of the same age before field planting had much more biomass and higher N and P concentrations and contents; although shoot:root balance and Dickson quality index were not significantly different. Finally, planted seedlings acclimation level during first year was also evaluated by Transplant Stress Index, which value (–0.1278) indicated a slight planting impact.     

Variation in drought resistance, drought acclimation and water conservation in four willow cultivars used for biomass production

Growth and water-use parameters of four willow (Salix spp.) clones grown in a moderate drought regime or with ample water supply were determined to characterize their water-use efficiency, drought resistance and capacity for drought acclimation. At the end of the 10-week, outdoor pot experiment, clonal differences were observed in: (1) water-use efficiency of aboveground biomass production (WUE); (2) resistance to xylem cavitation; and (3) stomatal conductance to leaf-specific, whole-plant hydraulic conductance ratio (g(st)/K(P); an indicator of water balance). Across clones and regimes, WUE was positively correlated with the assimilation rate to stomatal conductance ratio (A/g(st)), a measure of instantaneous water-use efficiency. Both of these water-use efficiency indicators were generally higher in drought-treated trees compared with well-watered trees. However, the between-treatment differences in (shoot-based) WUE were smaller than expected, considering the differences in A/g(st) for two of the clones, possibly because plants reallocated dry mass from shoots to roots when subject to drought. Higher root hydraulic conductance to shoot hydraulic conductance ratios (K(R)/K(S)) during drought supports this hypothesis. The same clones were also the most sensitive to xylem cavitation and, accordingly, showed the strongest reduction in g(st)/K(P) in response to drought. Drought acclimation was manifested in decreased g(st), g(st)/K(P), osmotic potential and leaf area to vessel internal cross-sectional area ratio, and increased K(R), K(P) and WUE. Increased resistance to stem xylem cavitation in response to drought was observed in only one clone. It is concluded that WUE and drought resistance traits are inter-linked and that both may be enhanced by selection and breeding.     

冷诱导对翅荚木一些生理生化特性的影响

采用梯度降温对翅荚木幼苗进行冷诱导,研究几项与抗低温有关的生理生化指标:膜透性、可溶性糖含量、可溶性蛋白含量和超氧化物歧化酶(SOD)活性的变化。结果表明:经过冷诱导的翅荚木幼苗的膜透性明显降低,可溶性糖和可溶性蛋白含量、SOD活性明显升高,而且湖南种源的变化幅度大于广西种源,说明湖南种源翅荚木的抗低温能力强于广西种源。     

植物抗寒性的生理生态学机制研究进展

综述植物在冷驯化过程中发生的一系列生理生化变化.环境对植物抗寒性的影响主要与光诱导、温湿度以及气候的变化有关.植物表面形成冰层会引起植物的无氧呼吸,导致植物受害;光抑制诱导活性氧的产生,从而导致植物光合系统的退化,抗寒能力下降,而短日照诱导植物休眠,有利于植物抗寒.光敏色素则被认为是启动冷驯化的光受体;植物通过冷驯化增加碳水化合物的积累及病原体相关蛋白的合成,以增强对低温病原体的抵抗能力;气候的变化使植物遭受了更大的冷伤害风险.微管最初遇冷时部分的解体可以有效诱导植物抗寒性;抗氧化酶活性增强,植物体内糖、脯氨酸、多胺等内含物含量上升.植物休眠状态中的生理变化(种子的休眠、芽的休眠)与ABA敏感性的差异有关.对植物抗寒性分子机制的研究表明:COR基因的表达对于植物抗寒性和冷驯化是十分关键的;与气候梯度有关的基因梯度的分布说明寒冷地区的树种更为抗寒;多表型性状的数量性状分析,为重要的农艺性状标记辅助选择(MAS)提供基础.对植物抗寒过程中的信号转导进行研究发现,Ca2 是低温下参与调节冷驯化应答机制中信号转导途径的重要的第二信使.未来植物抗寒领域的研究热点为信号转导和基因调节,低温抗性的遗传学和遗传应用及代谢组学,气候变化对于植物抗寒的影响等方面.     

Seasonal changes in the axial distribution of peroxidase activity in the xylem sap of beech (Fagus sylvatica L.) trees

Xylem sap was collected from trunk segments of adult beech (Fagus sylvatica L.) trees by water displacement. Peroxidase activity was analyzed in xylem saps collected in different phases of the yearly growth cycle and from different heights up the trunks (up to 14 m). The xylem saps contained two major peroxidase isozymes with acidic isoelectric points of 4.1 and 4.6, respectively. Mean peroxidase activity was low during the emergence of the new leaves and high in summer and in winter. In the cold season, peroxidase activity decreased from the stem base to the top, whereas significant gradients were not observed during the vegetative period.     

Physiological changes in red spruce seedlings during a simulated winter thaw

We evaluated net photosynthesis, respiration, leaf conductance, xylem pressure potential (XPP) and cold hardiness in red spruce (Picea rubens Sarg.) seedlings exposed to either a continuous thaw (CT) or a daytime thaw with freezing nights (FN) for 8 days during mid-winter. Physiological differences between CT and FN seedlings were evident for all measured parameters. However, the temporal expression of treatment differences varied among parameters. When compared to FN seedlings, CT seedlings had higher rates of respiration following 24 h of treatment, and a higher net photosynthetic rate, leaf conductance and XPP after 48 h of treatment. The CT seedlings were significantly less cold tolerant than the FN seedlings following 4 days of thaw, whereas FN seedlings did not deharden over the 8 days of treatment. Examination of temporal trends among thaw-associated changes in physiology suggested that, although greater carbon exchange occurred as stomatal conductance increased, the transition from negative to positive net photosynthesis was not the result of increases in conductance, but may have been associated with thaw-induced increases in XPP. Because thaw-associated changes in gas exchange and cold hardiness were offset in time, we conclude that, if changes in these processes are physiologically linked, the linkage is indirect.