Induction Of Cold Tolerance Research Articles

Effect of cold stress on polyamine metabolism and antioxidant responses in chickpea

Metabolic and genomic characteristics of polyamines (PAs) may be associated with the induction of cold tolerance (CT) responses in plants. Characteristics of PAs encoding genes in chickpea (Cicer arietinum L.) and their function under cold stress (CS) are currently unknown. In this study, the potential role of PAs along with the antioxidative defense systems were assessed in two chickpea genotypes (Sel96th11439, cold-tolerant and ILC533, cold-sensitive) under CS conditions. Six days after exposure to CS, the leaf H2O2 content and electrolyte leakage index increased in the sensitive genotype by 47.7 and 59 %, respectively, while these values decreased or remained unchanged, respectively, in the tolerant genotype. In tolerant genotype, the enhanced activity of superoxide dismutase (SOD) (by 50 %) was accompanied by unchanged activities of ascorbate peroxidase (APX), guaiacol peroxidase (GPX) and catalase (CAT) as well as the accumulation of glutathione (GSH) (by 43 %) on the sixth day of CS. Higher levels of putrescine (Put) (322 %), spermidine (Spd) (45 %), spermine (Spm) (69 %) and the highest ratio of Put/(Spd + Spm) were observed in tolerant genotype compared to the sensitive one on the sixth day of CS. Gamma-aminobutyric acid (GABA) accumulation was 74 % higher in tolerant genotype compared to the sensitive one on the sixth day of CS. During CS, the activity of diamine oxidase (DAO) and polyamine oxidase (PAO) increased in tolerant (by 3.02- and 2.46-fold) and sensitive (by 2.51- and 2.8-fold) genotypes, respectively, in comparison with the respective non-stressed plants (normal conditions). The highest activity of DAO and PAO in the tolerant genotype was accompanied by PAs decomposition and a peak in GABA content on the sixth day of CS. The analysis of chickpea genome revealed the presence of five PAs biosynthetic genes, their chromosomal locations, and cis-regulatory elements. A significant increase in transcript levels of arginine decarboxylase (ADC) (24.26- and 7.96-fold), spermidine synthase 1 (SPDS1) (3.03- and 1.53-fold), SPDS2 (5.5- and 1.62-fold) and spermine synthase (SPMS) (3.92- and 1.65-fold) genes was detected in tolerant and sensitive genotypes, respectively, whereas the expression of ornithine decarboxylase (ODC) genes decreased significantly under CS conditions in both genotypes. Leaf chlorophyll and carotenoid contents exhibited declining trends in the sensitive genotype, while these photosynthetic pigments were stable in the tolerant genotype due to the superior performance of defensive processes under CS conditions. Overall, these results suggested the specific roles of putative PAs genes and PAs metabolism in development of effective CT responses in chickpea.

The bZIP gene family in watermelon: genome-wide identification and expression analysis under cold stress and root-knot nematode infection.

The basic leucine zipper (bZIP) family transcription factors play crucial roles in regulating plant development and stress response. In this study, we identified 62 ClabZIP genes from watermelon genome, which were unevenly distributed across the 11 chromosomes. These ClabZIP proteins could be classified into 13 groups based on the phylogenetic relationships, and members in the same group showed similar compositions of conserved motifs and gene structures. Transcriptome analysis revealed that a number of ClabZIP genes have important roles in the melatonin (MT) induction of cold tolerance. In addition, some ClabZIP genes were induced or repressed under red light (RL) or root-knot nematode infection according to the transcriptome data, and the expression patterns of several ClabZIP genes were further verified by quantitative real-time PCR, revealing their possible roles in RL induction of watermelon defense against nematode infection. Our results provide new insights into the functions of different ClabZIP genes in watermelon and their roles in response to cold stress and nematode infection.

Transcriptomic Insights into Phenological Development and Cold Tolerance of Wheat Grown in the Field.

Cold acclimation and winter survival in cereal species is determined by complicated environmentally regulated gene expression. However, studies investigating these complex cold responses are mostly conducted in controlled environments that only consider the responses to single environmental variables. In this study, we have comprehensively profiled global transcriptional responses in crowns of field-grown spring and winter wheat (Triticum aestivum) genotypes and their near-isogenic lines with the VRN-A1 alleles swapped. This in-depth analysis revealed multiple signaling, interactive pathways that influence cold tolerance and phenological development to optimize plant growth and development in preparation for a wide range of over-winter stresses. Investigation of genetic differences at the VRN-A1 locus revealed that a vernalization requirement maintained a higher level of cold response pathways while VRN-A1 genetically promoted floral development. Our results also demonstrated the influence of genetic background on the expression of cold and flowering pathways. The link between delayed shoot apex development and the induction of cold tolerance was reflected by the gradual up-regulation of abscisic acid-dependent and C-REPEAT-BINDING FACTOR pathways. This was accompanied by the down-regulation of key genes involved in meristem development as the autumn progressed. The chromosome location of differentially expressed genes between the winter and spring wheat genetic backgrounds showed a striking pattern of biased gene expression on chromosomes 6A and 6D, indicating a transcriptional regulation at the genome level. This finding adds to the complexity of the genetic cascades and gene interactions that determine the evolutionary patterns of both phenological development and cold tolerance traits in wheat.

Melatonin alleviates low PS I-limited carbon assimilation under elevated CO2 and enhances the cold tolerance of offspring in chlorophyll b-deficient mutant wheat.

Melatonin is involved in the regulation of carbohydrate metabolism and induction of cold tolerance in plants. The objective of this study was to investigate the roles of melatonin in modulation of carbon assimilation of wild-type wheat and the Chl b-deficient mutant ANK32B in response to elevated CO2 concentration ([CO2 ]) and the transgenerational effects of application of exogenous melatonin (hereafter identified as melatonin priming) on the cold tolerance in offspring. The results showed that the melatonin priming enhanced the carbon assimilation in ANK32B under elevated [CO2 ], via boosting the activities of ATPase and sucrose synthesis and maintaining a relatively higher level of total chlorophyll concentration in leaves. In addition, melatonin priming in maternal plants at grain filling promoted the seed germination in offspring by accelerating the starch degradation and improved the cold tolerance of seedlings through activating the antioxidant enzymes and enhancing the photosynthetic electron transport efficiency. These findings suggest the important roles of melatonin in plant response to future climate change, indicating that the melatonin priming at grain filling in maternal plants could be an effective approach to improve cold tolerance of wheat offspring at seedling stage.

Effects of ethylene on photosystem II and antioxidant enzyme activity in Bermuda grass under low temperature.

The phytohormone ethylene has been reported to mediate plant response to cold stress. However, it is still debated whether the effect of ethylene on plant response to cold stress is negative or positive. The objective of the present study was to explore the role of ethylene in the cold resistance of Bermuda grass (Cynodon dactylon (L).Pers.). Under control (warm) condition, there was no obvious effect of the ethylene precursor 1-aminocyclopropane-1-carboxylic acid (ACC) or the antagonist Ag(+) of ethylene signaling on electrolyte leakage (EL) and malondialdehyde (MDA) content. Under cold stress conditions, ACC-treated plant leaves had a greater level of EL and MDA than the untreated leaves. However, the EL and MDA values were lower in the Ag(+) regime versus the untreated. In addition, after 3 days of cold treatment, ACC remarkably reduced the content of soluble protein and also altered antioxidant enzyme activity. Under control (warm) condition, there was no significant effect of ACC on the performance of photosystem II (PS II) as monitored by chlorophyll α fluorescence transients. However, under cold stress, ACC inhibited the performance of PS II. Under cold condition, ACC remarkably reduced the performance index for energy conservation from excitation to the reduction of intersystem electron acceptors (PI(ABS)), the maximum quantum yield of primary photochemistry (φP0), the quantum yield of electron transport flux from Q(A) to Q(B) (φE0), and the efficiency/probability of electron transport (ΨE0). Simultaneously, ACC increased the values of specific energy fluxes for absorption (ABS/RC) and dissipation (DI0/RC) after 3 days of cold treatment. Additionally, under cold condition, exogenous ACC altered the expressions of several related genes implicated in the induction of cold tolerance (LEA, SOD, POD-1 and CBF1, EIN3-1, and EIN3-2). The present study thus suggests that ethylene affects the cold tolerance of Bermuda grass by impacting the antioxidant system, photosystem II, as well as the CBF transcriptional regulatory cascade.

IMPROVEMENT OF COLD TOLERANCE IN 'TAHITI' LIME THROUGH HEAT TREATMENTS

The influence of heat treatments on the induction of cold tolerance in 'Tahiti' lime and their effects on ethylene biosynthesis and antioxidant enzymes were evaluated. Hot water dip (HWD) of fruit at 42°C for 15 min, delayed the onset of chilling injury (CI) symptoms for approximately 20 days at 1°C. Prestorage conditioning of fruit at 37°C for 10h, HWD at 53°C for 2 min and HWD at 42°C for 15 min reduced the CI index, as compared with untreated fruit after 15 and 30 days at 1°C (plus 3 days of shelf life at 20°C). The ACC oxidase activity increased in parallel to the CI index in conditioned and untreated fruit. In contrast, in HWD at 53°C for 2 min the ACC oxidase activity remained constant. However, the ACC oxidase activity in fruit treated with HWD at 42°C for 15 min was lower in all the assessments, remaining almost the same over the storage period. There was a great increase in catalase (CAT) activity in all the fruits following 15+3 days. Afterwards, a rapid decline in CAT activity occurred, and it was associated with increased chilling injury. On the other hand, ascorbate peroxidase (APX) activity had a linear increase during storage in fruit treated with HWD at 53°C for 2 min, while the other heat treated fruit (42°C for 15 min) showed a slight increase in APX activity after 15+3 days, remaining constant along the storage period. According to our results, the reduction in activity of ACC oxidase, caused by HWD treatments enhanced the resistance of fruit to CI development.

Heterologous Expression of Wheat VERNALIZATION 2 (TaVRN2) Gene in Arabidopsis Delays Flowering and Enhances Freezing Tolerance

The vernalization gene 2 (VRN2), is a major flowering repressor in temperate cereals that is regulated by low temperature and photoperiod. Here we show that the gene from Triticum aestivum (TaVRN2) is also regulated by salt, heat shock, dehydration, wounding and abscissic acid. Promoter analysis indicates that TaVRN2 regulatory region possesses all the specific responsive elements to these stresses. This suggests pleiotropic effects of TaVRN2 in wheat development and adaptability to the environment. To test if TaVRN2 can act as a flowering repressor in species different from the temperate cereals, the gene was ectopically expressed in the model plant Arabidopsis. Transgenic plants showed no alteration in morphology, but their flowering time was significantly delayed compared to controls plants, indicating that TaVRN2, although having no ortholog in Brassicaceae, can act as a flowering repressor in these species. To identify the possible mechanism by which TaVRN2 gene delays flowering in Arabidopsis, the expression level of several genes involved in flowering time regulation was determined. The analysis indicates that the late flowering of the 35S::TaVRN2 plants was associated with a complex pattern of expression of the major flowering control genes, FCA, FLC, FT, FVE and SOC1. This suggests that heterologous expression of TaVRN2 in Arabidopsis can delay flowering by modulating several floral inductive pathways. Furthermore, transgenic plants showed higher freezing tolerance, likely due to the accumulation of CBF2, CBF3 and the COR genes. Overall, our data suggests that TaVRN2 gene could modulate a common regulator of the two interacting pathways that regulate flowering time and the induction of cold tolerance. The results also demonstrate that TaVRN2 could be used to manipulate flowering time and improve cold tolerance in other species.

A selection procedure for identifying transgenic cells and embryos of cotton without the use of antibiotics

Transgenic cells containing inserted antibiotic resistance genes and linked genes of interest are routinely selected by exposure to antibiotics. Concerns over the widespread use of antibiotic resistance genes as selectable markers for genetic transformation have motivated researchers to find alternative selection procedures. This study describes the evaluation of an alternative protocol using temperature as the selection tool. In this method, a population of host cells is transformed with a foreign DNA construct that includes at least one gene of interest and an additional sequence encoding a protein that enhances cellular high temperature tolerance. Following transformation, the population of cells is transiently cultured under temperature conditions wherein growth of non-transformed cells is suppressed or prevented, while growth of cells containing the DNA construct continues. Thus, survival and/or significant additional growth is an indication that a cell has been successfully transformed with the DNA construct and can be subsequently recovered for further growth and development. The present study used a heat shock protein (hsp101) gene from Arabidopsis thaliana under the control of a constitutive promoter as a selectable marker; however, alternative potentially suitable genes include: other heat shock proteins; heat shock transcription factors; cold regulated proteins (COR); or protein transcription factors associated with the induction of cold tolerance.

Glycine betaine mitigates cold stress damage in Chickpea

Chickpea (Cicer arietinum L.) is sensitive to chilling temperatures (<10 °C), especially in its reproductive phase. Consequently it suffers from floral abortion, poor pod set and reduced seed size, leading to substantial yield loss. The underlying mechanisms are yet to be elucidated, which are imperative to understand how to induce cold tolerance in this crop. Glycine betaine, a cryoprotectant, has been associated with induction of cold tolerance. In the present study, chickpea plants previously growing under the warm conditions of a glasshouse (28/12 °C as average maximum and minimum temperature, respectively) were subjected to the chilling stress of the field (12-14/3-5 °C) at the (i) bud or (ii) pod-filling stages. Prior to exposure to stress, 1 mM glycine betaine was applied exogenously for 3 consecutive days. In glycine betaine-treated plants, chilling injury (assessed as electrolyte leakage) was reduced by 63% while cellular respiration increased by 69%. Application of glycine betaine at the bud stage resulted in improvement in flower functioning in terms of pollen germination (in vitro and in vivo), pollen viability, pollen tube growth, stigma receptivity and ovule viability. As a result, the floral retention, pod set and pod retention were increased by 47, 38 and 23%, respectively, over control. Treatment with glycine betaine at the pod-filling stage caused 30, 37, 46% and 9% enhancement in seed yield/plant, number of seeds/100 pods, single-seeded pods/plant and individual seed weight, respectively, while number of double-seeded pods was not affected significantly. Cold tolerance induced by glycine betaine appeared to be related to elevation of relative leaf water content, chlorophyll and sucrose, and decrease in abscisic acid as well as active oxygen species (malondialdehyde and hydrogen peroxide).

Cold urticaria.

Cold urticaria is one form of urticaria that may be associated with other forms of physical urticarias. Frequency is generally estimated at two or three per 100. The triggering effect of cold is found at history taking in most of the cases. The urticaria is usually superficial, and more rarely associated with deep and/or mucosal urticaria. The diagnosis is based on history taking and the ice cube test. An exhaustive search for an etiologic factor is often unfruitful, and the presence of a cryopathy should lead to a complete work-up. Therapy of cold urticaria may prove to be difficult. In patients with secondary cold urticaria, underlying disease must be treated in order to resolve the skin symptoms. H1-antihistamines can be used but the clinical responses are highly variable. Short-time treatment with low concentration corticosteroids suppresses the symptoms only partially and temporarily. In patients who do not respond to previous treatments, induction of cold tolerance may be proposed but the procedure is difficult to carry out in daily life over an extended period. Key word: cryoglobulins.

Effects of various heat treatments on the induction of cold tolerance and on the postharvest qualities of ‘Star Ruby’ grapefruit

Abstract We examined the effects of various heat treatments on the induction of cold tolerance in red grapefruit ( Citrus paradisi cv. Star Ruby) and evaluated their effects on various other postharvest quality parameters such as decay development, weight loss, peel color, and juice total soluble solids (TSS) and acid content. Various heat treatments, including prestorage conditioning regimes of 3 days at 21°C and 7 days at 16°C, a hot water dip (HWD) at 53°C for 2 min, a hot water brushing (HWB) treatment at 60°C for 30 s, and curing (3 days at 36°C) achieved 70–90% reduction in the chilling injury (CI) index, as compared with control untreated fruit, following 6 weeks of cold storage at 2°C and an additional week at 20°C. Of the various treatments that induced cold tolerance, only HWB significantly reduced postharvest decay development. Conditioning and curing, which required longer exposure periods of 3–7 days at relatively high temperatures, significantly increased fruit weight loss, enhanced peel color alteration, and increased the juice TSS/acid ratios. On the other hand, the short postharvest heat treatments, including the HWD at 53°C for 2 min and the HWB at 60°C for 30 s, did not affect fruit weight loss, color, or TSS and acidity levels. Overall, it is concluded that short postharvest heat treatments, including either HWD or HWB are preferable, since they effectively induce tolerance to cold temperatures in ‘Star Ruby’ grapefruit without impairing any other postharvest qualities. Of these treatments, the new HWB is faster and could be used to clean and disinfect the fruit, and simultaneously to enhance its CI tolerance.

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.

Induction of Tolerance to Hypothermia by Previous Heat Shock Using Human Fibroblasts in Culture

The technique of organ preservation is limited by the amount of time which organs can be hypothermically stored. A potential method to effectively extend reliable storage times involves the conditioning of cells to better withstand hypothermia by previous exposure to a less severe stress. Using human fibroblasts in culture, we have demonstrated that such an approach may be feasible. Subjecting human diploid IMR-90 fibroblasts to 5 h 42.5°C heat shock was found to improve cell survival more than 10-fold to subsequent 4°C hypothermic exposure. The prior heat shock resulted in the increased synthesis of heat shock proteins (HSPs), the absolute concentrations of which were measured by an assay which utilized sodium dodecyl sulfate–polyacrylamide gel electrophoresis and Western blotting techniques. Both the degree of cold tolerance conferred upon IMR-90 cells and the levels of HSP70 and HSP27 were dependent upon initial heat shock duration. Induced cold tolerance was found to be reversible; longer recovery times at 37°C following heat shock resulted in a loss of this cold-tolerant state as well as a disappearance of HSPs. The fact that the degree of cold tolerance and HSP concentrations showed similar trends with respect to both heat shock time at 42.5°C and subsequent recovery time at 37°C suggests that these proteins may be intimately involved in the induction of cold tolerance.

Induction of cold-tolerance in Galleria larvae by Farnesyl methyl ether

After 3 weeks of chilling at 4 °C, last-instar larvae of the wax moth G. mellonella required about 40 min at 25 °C to regain posture and locomotory activity; 65% of the larvae died subsequently. Topical application of 2 μl FME significantly reduced the "recovery time" and enabled 75% of the treated larvae to survive the cold exposure.