Development Of Frost Tolerance Research Articles

Relationship between WCS120 Protein Family Accumulation and Frost Tolerance in Wheat Cultivars Grown under Different Temperature Treatments

Frost tolerance (FT) is generally acquired after exposure of plants to low, but non-freezing temperatures, where it is associated with the accumulation of COR proteins. The aim of the study was to reveal the effect of different temperature treatments (25, 17, 9 and 4 °C) on accumulation of cold-regulated dehydrins, dry weight content, and the development of FT in five wheat cultivars of different frost-tolerances in detail. The levels of cold-regulated dehydrins, WCS120 proteins in wheat were determined by immunoblot analysis, probed with an anti-dehydrin antibody. The lower the growth temperature: the higher the level of frost tolerance, dry weight content, and dehydrin accumulation, in all cultivars. There was a significant correlation between the level of induced FT and the accumulation of WCS120 proteins in cultivars grown at lower temperatures (9 and 4 °C). Moreover, the highly frost-tolerant wheat cultivars (as opposed to the lower-tolerant) accumulated higher levels of WCS120 proteins at 17 °C, a temperature at which it was not possible to differentiate between them via a frost test. Here, we demonstrated the possibility to distinguish differently frost-tolerant cultivars grown at different temperatures by the accumulation of different members of WCS120 family.

Frost tolerance and metabolite changes of rye (Secale cereale) during the cold hardening and overwintering

Successful winter survival of the plants needs a programmed capacity to activate appropriate mechanisms in development of frost tolerance (FT). Cold hardening is a complicated operation facilitating the overwintering and is associated by extensive changes in metabolome. The purpose of the present investigation is to simplify the association between metabolites changes and FT in winter rye varieties (Puma and Anatolien) acclimated under field situation in NW Iran, regions with long winters. Plants were sampled on five dates (Oct. 31, Nov. 29, Dec. 17, Jan. 4, and Feb. 1) as different cold hardening periods. FT, as measured by LT50 (the temperature at 50% lethality) significantly expanded during the autumn and winter seasons. Cold adaptation persuaded accumulation of sucrose and sorbitol in both varieties; however, the accumulation was more profound in Puma. The content of proline, aspartic acid, glutamic acid, tyrosine, and glycine betaine linearly expanded in the Anatolien variety during the overwintering. However, content of glycine betaine, proline, and aspartic acid in Puma was considerably higher than Anatolien. Hydrogen peroxide reached to its greatest quantities at November and December in Anatolien and Puma, respectively. These levels significantly reduced up to the next months. Glutathione and ascorbic acid contents, as important antioxidants, noticeably expanded during overwintering. There was a significant positive association between FT, total protein content, free amino acid, soluble sugars, and glycine betaine. These data suggest that the development of FT during the cold adaptation and overwintering is partly resulted from metabolome changes and such modifications are strongly linked to the genetic potential.

Photoprotective mechanisms in photosystem II of Ephedra monosperma during development of frost tolerance

Dissipation of light energy absorbed by photosystem II (PSII) in assimilating shoots of an ever� green shrub Ephedra monosperma was investigated during its transition from the vegetative to frosttolerant state under natural conditions of Central Yakutia. The dynamics of modulated chlorophyll fluorescence and carotenoid content was analyzed during seasonal decrease in ambient temperature. The seasonal cooling was accompanied by a stepwise decrease in photochemical activity of PSII (Fv/Fm = (Fm - F0)/Fm). The decrease in Fv/Fm occurred from the beginning of September to the end of October, when the temperature was lowered from 10 to -8°C. During winter period the residual activity of PSII was retained at about 30% of the summer values. The seasonal decrease in temperature was accompanied by a significant stimulation of pHindepen� dent dissipative processes in reaction centers and antenna of PSII. The increase in energy losses was paral� leled by a proportional increase in zeaxanthin content on the background of decreasing content of violaxan� thin and β�carotene as possible zeaxanthin precursors. At the same time, inhibition of lightinduced non� photochemical quenching in the PSII antenna was observed. The results suggest that principal photoprotec� tive mechanisms during seasonal lowering of temperature are: (1) inactivation of PSII and dissipation of exci� tation energy in PSII reaction centers and (2) zeaxanthinmediated ener gy dissipation in the antenna com� plexes. The first mechanism seems to prevail at early stages of seasonal cooling, whereas both mechanisms are recruited from the onset of sustained freezing temperatures.

Exogenous application of molybdenum affects the expression of CBF14 and the development of frost tolerance in wheat

Wheat is able to cold acclimate in response to low temperatures and thereby increase its frost tolerance and the extent of this acclimation is greater in winter genotypes compared to spring genotypes. Such up-regulation of frost tolerance is controlled by Cbf transcription factors. Molybdenum (Mo) application has been shown to enhance frost tolerance of wheat and this study aimed to investigate the effect of Mo on the development of frost tolerance in winter and spring wheat. Results showed that Mo treatment increased the expression of Cbf14 in wheat under non-acclimating condition but did not alter frost tolerance. However, when Mo was applied in conjunction with exposure of plants to low temperature, Mo increased the expression of Cbf14 and enhanced frost tolerance in both spring and winter genotypes but the effect was more pronounced in the winter genotype. It was concluded that the application of Mo could be useful in situations where enhanced frost resistance is required. Further studies are proposed to elucidate the effect of exogenous of applications of Mo on frost resistance in spring and winter wheat at different growth stages.

The development of frost tolerance and DHN5 protein accumulation in barley ( Hordeum vulgare) doubled haploid lines derived from Atlas 68×Igri cross during cold acclimation

The dynamics of a long-term cold acclimation (CA) was studied in spring barley cultivar Atlas 68, winter barley cultivar Igri and a set of doubled haploid (DH) lines derived from an Atlas 68×Igri cross. The aim was to evaluate the effect of plant development on the ability to induce frost tolerance (FT) and to accumulate dehydrin 5 (DHN5) during CA. The plant developmental stage was evaluated by phenological development of the shoot apex and by determination of days to heading after a certain period of CA. FT was determined by direct frost tests. Plant winter survival was also determined. DHN5 was evaluated by densitometric analysis of protein gel blots. Cold led to the induction of increased FT and to the accumulation of DHN5 in both spring and winter lines. However, with the progression of CA, differences between the growth habits occurred as the winter lines were able to maintain increased FT and DHN5 levels for a significantly longer period of time than the spring lines. After vegetative/reproductive transition, a significant decrease in DHN5 accumulation was found in all lines; however, a discrepancy between the acquired FT level and DHN5 accumulation in vernalized winter barley plants was found. A correlation between DHN5 accumulation and plant winter survival was found when the studied lines were differentiated according to their developmental stage and DHN5 level. Possible explanations for these phenomena are provided.

The relationship between vernalization-and photoperiodically-regulated genes and the development of frost tolerance in wheat and barley

The review summarizes the level of current knowledge of impacts of vernalization and photoperiod on the induction and maintenance of frost tolerance (FrT) in wheat and barley. The phenomenon of vernalization is briefly described and the major vernalization (VRN) loci are characterised. Vernalization requirement and the three major growth habits of Triticeae (facultative, winter and spring) are defined on the basis of the two-locus VRN-2/VRN-1 epistatic model. Major photoperiodically regulated genes, which influence the transition to flowering, are characterised and their interactions with VRN genes are briefly discussed. The phenomenon of induction of FrT during the process of cold acclimation (CA) is described and the major cold-induced Cor/Lea genes are listed. Important regulatory mechanisms, i.e., CBF pathway, controlling the expression of Cor/Lea genes under cold, are discussed. The major loci affecting the development of FrT in Triticeae, the Fr loci, are characterised. In conclusion, current progress in this research field is summarized and new questions arising in the area are formulated.

Expression of dehydrin 5 during the development of frost tolerance in barley ( Hordeum vulgare)

The Dhn5 gene is the major cold-inducible dehydrin gene in barley. This study deals with the relationship between Dhn5 gene expression and its protein product accumulation, and the development of frost tolerance (FT) upon cold acclimation (CA) in 10 barley cultivars of different growth habits and geographical origins. The activation of Dhn5 gene expression was determined by quantitative reverse transcription-polymerase chain reaction (qRT-PCR), the accumulation of DHN5 protein was evaluated by protein gel blot analysis using a specific anti-dehydrin antibody, and the acquired level of FT was determined by a direct frost test. During the first 2 weeks of CA, there was a rapid increase in Dhn5 gene expression, DHN5 protein accumulation and FT in all cultivars examined. After 2 weeks of CA, differences in DHN5 accumulation and in FT measured as lethal temperature (LT(50)) were observed between the cultivars belonging to different growth habits. Specifically, intermediate (I) and winter (W) cultivars showed a higher level of DHN5 accumulation and FT than the spring (S) cultivars, which exhibited a lower level of accumulated DHN5 and FT. (Intermediate cultivars do not have vernalization requirement, but they are able to induce a relatively high level of FT upon CA.) In contrast, no differences between the cultivars belonging to different growth habits in Dhn5 mRNA accumulation were found. After 3 weeks of CA, the differences in accumulated DHN5 and FT between the individual growth habits became evident due to different developmental regulation of FT. The amount of accumulated DHN5 corresponded well with the level of FT of individual cultivars. We conclude that the amount of accumulated DHN5 after a certain period of CA differed according to the growth habits of cultivars and can be used as a marker for determination of FT in barley.

Antisense inhibition of protein phosphatase 2C accelerates cold acclimation in Arabidopsis thaliana.

Two related protein phosphatases 2C, ABI1 and AtPP2CA have been implicated as negative regulators of ABA signalling. In this study we characterized the role of AtPP2CA in cold acclimation. The pattern of expression of AtPP2CA and ABI1 was studied in different tissues and in response to abiotic stresses. The expression of both AtPP2CA and ABI1 was induced by low temperature, drought, high salt and ABA. The cold and drought-induced expression of these genes was ABA-dependent, but divergent in various ABA signalling mutants. In addition, the two PP2C genes exhibited differences in their tissue-specific expression as well as in temporal induction in response to low temperature. To elucidate the function of AtPP2CA in cold acclimation further, the corresponding gene was silenced by antisense inhibition. Transgenic antisense plants exhibited clearly accelerated development of freezing tolerance. Both exposure to low temperature and application of ABA resulted in enhanced freezing tolerance in antisense plants. These plants displayed increased sensitivity to ABA both during development of frost tolerance and during seed germination, but not in their drought responses. Furthermore, the expression of cold-and ABA-induced genes was enhanced in transgenic antisense plants. Our results suggest that AtPP2CA is a negative regulator of ABA responses during cold acclimation.

The role of inorganic phosphate in the development of freezing tolerance and the acclimatization of photosynthesis to low temperature is revealed by the pho mutants of Arabidopsis thaliana.

Low temperature inhibits sucrose synthesis, leading to a phosphate-limitation of photosynthesis. We have used the Arabidopsis pho1-2 and pho2-1 mutants with decreased and increased shoot phosphate, respectively, to investigate whether low phosphate triggers cold acclimatization of photosynthetic carbon metabolism. Wild-type Arabidopsis, pho1-2 and pho2-1 were grown at 23 degrees C and transferred to 5 degrees C to investigate acclimatization in pre-existing leaves and in new leaves developing at 5 degrees C. The development of frost tolerance and the accumulation of proline and sugars was unaltered or improved in pho1-2, and impaired in pho2-1. Sucrose phosphate synthase and cytoplasmic fructose-1,6-bisphosphatase activity and protein increase after transfer to 5 degrees C. This increase was accentuated in pho1-2 and attenuated in pho2-1. RBCS and LHCB2 transcript levels decrease in pre-formed wild-type leaves after transfer to 5 degrees C and recover in new leaves that develop at 5 degrees C. The initial decrease was attenuated in pho1-2, and accentuated in pho2-1, where the recovery in new leaves was also suppressed. Rubisco activity increased in wild-type leaves that developed at 5 degrees C. This increase was accentuated in pho1-2 and absent in pho2-1. NADP-glyceraldehyde-3-phosphate dehydrogenase, plastidic fructose-1,6-bisphosphatase and aldolase activity increase relative to phosphoglycerate kinase, transketolase and phosphoribulokinase in wild-type leaves at 5 degrees C. This shift was accentuated in pho1-2 and reversed in pho2-1. Transcript levels for COR genes increase transiently 1 day after transfer to 5 degrees C but were very low in leaves that developed at 5 degrees C in wild-type Arabidopsis, pho1-2 and pho2-1. We conclude that low phosphate plays an important role in triggering cold acclimatization of leaves, leading in particular to an increase of Rubisco expression, changes in other Calvin cycle enzymes to minimize sequestration of phosphate in metabolites, and increased expression of sucrose biosynthesis enzymes.

Development of Frost Tolerance in Winter Wheat as Modulated by Differential Root and Shoot Temperature

Abstract: Winter wheat plants (Triticum aestivum L. cv. Urban), grown in nutrient solution, were exposed to differential shoot/root temperatures (i.e., 4/4, 4/20, 20/4 and 20/20 d̀C) for six weeks. Leaves grown at 4d̀C showed an increase in frost tolerance from ‐ 4d̀C down to ‐ 11 d̀C, irrespective of root temperature. In 4/20 d̀C plants, high root temperature decreased the rate of hardening of the leaves, but did not influence the final level of frost tolerance. In roots grown at 4d̀C frost tolerance increased from ‐ 3 d̀C down to ‐ 4 d̀C, independently of shoot temperature. An accumulation of soluble sugars in the leaves was only observed when both shoot and root were grown at 4d̀C and was not correlated with final frost tolerance achieved. However, the rate of hardening was correlated with the soluble sugar concentration. An increase in root soluble sugar concentration was exclusively observed in roots exposed to a temperature of 4d̀C, irrespective of shoot temperature. Proline concentration only increased in plant parts exposed to a temperature of 4d̀C. The present results indicate that the importance of root temperature in low‐temperature hardening of winter wheat is limited, even though exposure to differential root and shoot temperatures brought about pronounced changes in growth, soluble sugar concentration, insoluble sugar concentration and proline concentration in roots and leaves.

Development of Frost Tolerance in Winter Wheat as Modulated by Differential Root and Shoot Temperature

Abstract: Winter wheat plants (Triticum aestivum L. cv. Urban), grown in nutrient solution, were exposed to differential shoot/root temperatures (i.e., 4/4, 4/20, 20/4 and 20/20 d̀C) for six weeks. Leaves grown at 4d̀C showed an increase in frost tolerance from - 4d̀C down to - 11 d̀C, irrespective of root temperature. In 4/20 d̀C plants, high root temperature decreased the rate of hardening of the leaves, but did not influence the final level of frost tolerance. In roots grown at 4d̀C frost tolerance increased from - 3 d̀C down to - 4 d̀C, independently of shoot temperature. An accumulation of soluble sugars in the leaves was only observed when both shoot and root were grown at 4d̀C and was not correlated with final frost tolerance achieved. However, the rate of hardening was correlated with the soluble sugar concentration. An increase in root soluble sugar concentration was exclusively observed in roots exposed to a temperature of 4d̀C, irrespective of shoot temperature. Proline concentration only increased in plant parts exposed to a temperature of 4d̀C. The present results indicate that the importance of root temperature in low-temperature hardening of winter wheat is limited, even though exposure to differential root and shoot temperatures brought about pronounced changes in growth, soluble sugar concentration, insoluble sugar concentration and proline concentration in roots and leaves.

Photosynthetic depression in leaves of frost-hardened ivy is not caused by feedback inhibition via assimilate accumulation

Summary The aim of this study was to examine whether an impairment of photosynthetic activity following frost hardening is associated with an accumulation of soluble saccharides usually occurring during the development of frost tolerance. Rates of photosynthetic CO 2 uptake and O 2 evolution as well as contents of glucose, fructose and sucrose were determined in leaves of ivy plants ( Hedem helix L.) with varying source/ sink relations prior to and after hardening for 4 weeks at 5/0 °C day/night temperatures. Effects of feeding inorganic phosphate (P i ) on photosynthesis and of dehardening after transfer to 20/15 °C were also recorded. Photosynthesis was reduced by about 30 % after hardening, accompanied by a strong increase in sugar contents (sucrose 2×, glucose and fructose 10 ×). Photosynthesis and sugars tended to normalise after 4 days at 20/15 °C. Photosynthetic depression in frost-hardened leaves was similar at ambient and saturating CO 2 , irrespective of source/sink relations. Feeding the leaves with P i hardly restored their photosynthetic activity. These findings do not support the hypothesis that photosynthetic impairment during frost hardening is a result of feedback inhibition via assimilate accumulation.

Desiccation tolerance changes in winter rape leaves grown under different environmental conditions

The results of studies performed on winter rape leaves (Brassica napus L. var.oleifera L., cv. Górczański) indicated that independently of the plant cultivation conditions and of the leaf growth rate, all leaves showed significant increase in desiccation tolerance at the beginning of leaf expansion. The increase in tolerance was correlated with the formation of a central vacuole in the mesophyll cells. Therefore, the high desiccation tolerance of vacuolised cells in comparison to the non-vacuolised ones is supposed to be due to the higher ability of the former to avoid cytoplasm dehydration. Development of frost tolerance in the leaves during autumn does not seem to be causally related to desiccation tolerance changes.