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The seeds of soft winter wheat for long-term storage are treated with low-temperature plasma of a surface discharge according to an indirect method, i.e., without contact between a layer of immobile seeds and a surface discharge in a strip electrode system with a distance of 10 mm between strips. The seeds are treated with a surface discharge for 60 s in two modes of feeding the electrodes with a sinusoidal voltage (16 kHz/1.5 kV and 4.4 kHz/2.4 kV). The biometric parameters of seedlings on the third and seventh days, the resistance of cold hardened seedlings to low temperatures, and the degree of seed infection are measured. It is found that seed treatment does not provide an increase in the quality characteristics of germination and freezing tolerance and also does not reduce the seed infection under the condition of a high mycobiotic load. [ABSTRACT FROM AUTHOR]

Abstract Background In the agricultural areas of Qinghai-Tibet Plateau, temperature varies widely from day to night during the growing season, which makes the extreme temperature become one of the limiting factors of crop yield. Turnip (Brassica rapa var. rapa) is a traditional crop of Tibet grown in the Tibet Plateau, but its molecular and metabolic mechanisms of freezing tolerance are unclear. Results Here, based on the changes in transcriptional and metabolic levels of Tibetan turnip under freezing treatment, the expression of the arginine decarboxylase gene BrrADC2.2 exhibited an accumulative pattern in accordance with putrescine content. Moreover, we demonstrated that BrrICE1.1 (Inducer of CBF Expression 1) could directly bind to the BrrADC2.2 promoter, activating BrrADC2.2 to promote the accumulation of putrescine, which was verified by RNAi and overexpression analyses for both BrrADC2.2 and BrrICE1.1 using transgenic hair root. The function of putrescine in turnip was further analyzed by exogenous application putrescine and its inhibitor DL-α-(Difluoromethyl) arginine (DFMA) under freezing tolerance. In addition, the BrrICE1.1 was found to be involved in the ICE1-CBF pathway to increase the freezing stress of turnip. Conclusions BrrICE1.1 could bind the promoter of BrrADC2.2 or CBFs to participate in freezing tolerance of turnip by transcriptomics and targeted metabolomics analyses. This study revealed the regulatory network of the freezing tolerance process in turnip and increased our understanding of the plateau crops response to extreme environments in Tibet.

Abstract Background Sheepgrass (Leymus chinensis (Trin.) Tzvel) is a perennial forage grass that can survive extreme freezing winters (− 47.5 °C) in China. In this study, we isolated an unknown function MYB transcription factor gene, LcMYB4, from sheepgrass. However, the function of LcMYB4 and its homologous genes has not been studied in other plants. Results The expression of the LcMYB4 gene was upregulated in response to cold induction, and the LcMYB4 fusion protein was localized in the nucleus, with transcriptional activation activity. Biological function analysis showed that compared with WT plants, LcMYB4-overexpressing Arabidopsis presented significantly increased chilling and freezing tolerance as evidenced by increased germination rate, survival rate, and seed setting rate under conditions of low temperature stress. Furthermore, LcMYB4-overexpressing plants showed increased soluble sugar content, leaf chlorophyll content and superoxide dismutase activity but decreased malondialdehyde (MDA) under chilling stress. Moreover, the expression of the CBF1, KIN1, KIN2 and RCI2A genes were significantly upregulated in transgenic plants with chilling treatment. These results suggest that LcMYB4 overexpression increased the soluble sugar content and cold-inducible gene expression and alleviated oxidative damage and membrane damage, resulting in enhanced cold resistance in transgenic plants. Interestingly, our results showed that the LcMYB4 protein interacts with fructose-1,6-bisphosphate aldolase protein1 (LcFBA1) and that the expression of the LcFBA1 gene was also upregulated during cold induction in sheepgrass, similar to LcMYB4. Conclusion Our findings suggest that LcMYB4 encodes MYB transcription factor that plays a positive regulatory role in cold stress.

Abstract Background Alfalfa is a high-quality forage cultivated widely in northern China. Recently, the failure of alfalfa plants to survive the winter has caused substantial economic losses. Water management has attracted considerable attention as a method for the potential improvement of winter survival. The aim of this study was to determine whether and how changes in the water regime affect the freezing tolerance of alfalfa. Results The alfalfa variety WL353LH was cultivated under water regimes of 80 and 25% of water-holding capacity, and all the plants were subjected to low temperatures at 4/0 °C (light/dark) and then − 2/− 6 °C (light/dark). The semi-lethal temperatures were lower for water-stressed than well-watered alfalfa. The pool sizes of total soluble sugars, total amino acids, and proline changed substantially under water-deficit and low-temperature conditions. Metabolomics analyses revealed 72 subclasses of differential metabolites, among which lipid and lipid-like molecules (e.g., fatty acids, unsaturated fatty acids, and glycerophospholipids) and amino acids, peptides, and analogues (e.g., proline betaine) were upregulated under water-deficit conditions. Some carbohydrates (e.g., D-maltose and raffinose) and flavonoids were also upregulated at low temperatures. Finally, Kyoto Encyclopedia of Genes and Genomes analyses revealed 18 significantly enriched pathways involved in the biosynthesis and metabolism of carbohydrates, unsaturated fatty acids, amino acids, and glycerophospholipids. Conclusions Water deficit significantly enhanced the alfalfa’ freezing tolerance, and this was correlated with increased soluble sugar, amino acid, and lipid and lipid-like molecule contents. These substances are involved in osmotic regulation, cryoprotection, and the synthesis, fluidity, and stability of the cellular membrane. Our study provides a reference for improving alfalfa’ winter survival through water management.

Low temperature is an important factor limiting plant growth. Most cultivars of Vitis vinifera L. are sensitive to low temperatures and are at risk of freezing injury or even plant death during winter. In this study, we analyzed the transcriptome of branches of dormant cv. Cabernet Sauvignon exposed to several low-temperature conditions to identify differentially expressed genes and determine their function based on Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG)enrichment analyses. Our results indicated that exposure to subzero low temperatures resulted in damage to plant cell membranes and extravasation of intracellular electrolytes, and that this damage increased with decreasing temperature or increasing duration. The number of differential genes increased as the duration of stress increased, but most of the common differentially expressed genes reached their highest expression at 6 h of stress, indicating that 6 h may be a turning point for vines to tolerate extreme low temperatures. Several pathways play key roles in the response of Cabernet Sauvignon to low-temperature injury, namely: (1) the role of calcium/calmodulin-mediated signaling; (2) carbohydrate metabolism, including the hydrolysis of cell wall pectin and cellulose, decomposition of sucrose, synthesis of raffinose, and inhibition of glycolytic processes; (3) the synthesis of unsaturated fatty acids and metabolism of linolenic acid; and (4) the synthesis of secondary metabolites, especially flavonoids. In addition, pathogenesis-related protein may also play a role in plant cold resistance, but the mechanism is not yet clear. This study reveals possible pathways for the freezing response and leads to new insights into the molecular basis of the tolerance to low temperature in grapevine.

Low-temperatures pose extreme challenges to crops causing significant economical impacts. Frosts are responsible for more than 30% of weather-related insured crop losses in some temperate climate jurisdictions, but are particularly devastating for small holdings and communities reliant on a bountiful harvest. Low-temperatures are also frequently accompanied by other abiotic and biotic stresses, including pathogen attacks. Some pathogens have sub-zero temperature optima, while others leverage low-temperatures to promote freezing at high sub-zero temperatures by way of ice-nucleating proteins in order to access intracellular nutrients. To survive low-temperatures and the attendant risks, various plant species have evolved complex and intricate signaling networks, molecular mechanisms, and physiological changes, in addition to symbiotic relationships with microbiota. Enhancing low-temperature survival and pathogen-induced freezing tolerance in cold susceptible, agriculturally significant crops is an attractive area of research with immense translatable value to all aspects of society. This area of research will be particularly important in our near future as climate change increases the unpredictability of frosts, particularly in the spring and autumn. Against this backdrop, the world population continues to grow while arable land remains finite and wealth inequality exacerbates food poverty. In this review, we examine plant (i) low-temperature stress, (ii) cold acclimation responses, particularly in crops (iii) antifreeze proteins, and (iv) frost-associated pathogens. Lastly, we suggest integrated approaches to improve crop frost tolerance.

Background and Aims: As winter and spring temperatures continue to increase, the timing of flowering and leaf out is advancing in many seasonally cold regions. This advancement could put plants that flower early in the spring at risk of decreased reproduction in years when there are late freeze events. Unfortunately, relatively little is known about floral freezing tolerance in forest communities. In this study, we examined the impact of freezing temperatures on the flowers of woody plants in a region where there is rapid winter warming in North America., Methods: We subjected the flowers of twenty-five woody species to a hard (-5ºC) and a light freeze (0ºC). We assessed tissue damage using electrolyte leakage. In a subset of species, we also examined the impact of a hard freeze on pollen tube growth. To determine if the vulnerability of flowers to freezing damage relates to flowering time and to examine the responsiveness of flowering time to spring temperature, we recorded the date of first flower for our study species for three years., Key Results and Conclusions: Across species, we found that floral freezing tolerance was strongly tied to flowering time with the highest freezing tolerance occurring in plants that bloomed earlier in the year. We hypothesize that these early blooming species are unlikely to be impacted by a false spring. Instead, the most vulnerable species to a false spring should be those that bloom later in the season. The flowering time in these species is also more sensitive to temperature, putting them at a great risk of experiencing a false spring. Ultimately, floral damage in one year will not have a large impact on species fitness, but if false springs become more frequent, there could be long-term impacts on reproduction of vulnerable species., (© The Author(s) 2024. Published by Oxford University Press on behalf of the Annals of Botany Company. All rights reserved. For commercial re-use, please contact reprints@oup.com for reprints and translation rights for reprints. All other permissions can be obtained through our RightsLink service via the Permissions link on the article page on our site—for further information please contact journals.permissions@oup.com.)

Freezing stress can seriously affect plant growth and development, but the mechanisms of these effects and plant responses to freezing stress require further exploration. Here, we identified a NAM, ATAF1/2, and CUC2 (NAC)-family transcription factor (TF), NAC056, that can promote freezing tolerance in Arabidopsis. NAC056 mRNA levels are strongly induced by freezing stress in roots, and the nac056 mutant exhibits compromised freezing tolerance. NAC056 acts positively in response to freezing by directly promoting key C-repeat-binding factor (CBF) pathway genes. Interestingly, we found that CBF1 regulates nitrate assimilation by regulating the nitrate reductase gene NIA1 in plants; therefore, NAC056-CBF1-NIA1 form a regulatory module for the assimilation of nitrate and the growth of roots under freezing stress. In addition, 35S::NAC056 transgenic plants show enhanced freezing tolerance, which is partially reversed in the cbfs triple mutant. Thus, NAC056 confers freezing tolerance through the CBF pathway, mediating plant responses to balance growth and freezing stress tolerance., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Proanthocyanidins (PAs) are important metabolites that enhance freezing tolerance of plants. Actinidia arguta, especially freezing-tolerant germplasms, accumulate abundant PAs in dormant shoots and thereby enhance freezing tolerance, but the underlying mechanism is unknown. In this study, we used two A. arguta with contrasting cold-resistant phenotypes, KL and RB, to explore the mechanisms in response to cold tolerance. We determined that a leucoanthocyanidin reductase gene (AaLAR1) was more highly expressed in freezing-tolerant KL than in freezing-sensitive RB. Moreover, overexpressing AaLAR1 in kiwifruit promoted PAs biosynthesis and enhanced cold tolerance. The AaLAR1 promoters of various A. arguta germplasms differ due to the presence of a 60-bp deletion in cold-tolerant genotypes that forms a functional binding site for MYC-type transcription factor. Yeast one-hybrid and two-hybrid, dual-luciferase reporter, bimolecular fluorescence complementation and coimmunoprecipitation assays indicated that the AaMYC2a binds to the MYC-core cis-element in the AaLAR1 promoter with the assistance of AaMYB5a, thereby promoting PAs accumulation in the shoots of cold-tolerant kiwifruit. We conclude that the variation in the AaLAR1 promoter and the AaMYC2a-AaMYB5a-AaLAR1 module shape freezing tolerance in A. arguta. The identification of a key structural variation in the AaLAR1 promoter offers a new target for resistance breeding of kiwifruit., (© 2024 John Wiley & Sons Ltd.)

At high elevation or latitude, trees reach low-temperature range limits. In attempting an explanation, the range limits of individual tree species (set by freezing tolerance) and the general limit of the life-form tree (set by thermal growth constraints) need to be distinguished. The general cold edge of the fundamental niche of trees is termed the treeline, by definition, the lower edge of the alpine belt, a most important bioclimatological reference line. Trees can be absent from the treeline due to disturbances or biotic interactions. The actual local edge of tree distribution, the delineation of the realized niche, is driven by stochastic effects. Therefore, treeline theory and hypothesis testing is inevitably tied to the fundamental niche concept. Low-temperature range limits of tree species result from the interaction of phenology, freezing tolerance at flushing, and season length requirements for maturation. At the low-temperature treeline, trees become victims of their height. It is the aerodynamics of the life-form tree that impose critically low, growth-limiting air temperatures on trees, from which small-stature plants escape. Humans cannot shift the low-temperature limit of trees (the treeline), but they may have removed trees from the treeline by logging or burning. The treeline tracks seasonal low-temperature means, the tree species range limit tracks low-temperature extremes. The treeline is a most important global bioclimatic reference line against which other bioclimatic zones and the associated biomes can be defined. [ABSTRACT FROM AUTHOR]

Peat bog pools around Tamsweg (Lungau, Austria) are typical habitats of the unicellular green alga Micrasterias denticulata. By measurement of water temperature and irradiation throughout a 1-year period (2018/2019), it was intended to assess the natural environmental strain in winter. Freezing resistance of Micrasterias cells and their ability to frost harden and become tolerant to ice encasement were determined after natural hardening and exposure to a cold acclimation treatment that simulated the natural temperature decrease in autumn. Transmission electron microscopy (TEM) was performed in laboratory-cultivated cells, after artificial cold acclimation treatment and in cells collected from field. Throughout winter, the peat bog pools inhabited by Micrasterias remained unfrozen. Despite air temperature minima down to −17.3 °C, the water temperature was mostly close to +0.8 °C. The alga was unable to frost harden, and upon ice encasement, the cells showed successive frost damage. Despite an unchanged freezing stress tolerance, significant ultrastructural changes were observed in field-sampled cells and in response to the artificial cold acclimation treatment: organelles such as the endoplasmic reticulum and thylakoids of the chloroplast showed distinct membrane bloating. Still, in the field samples, the Golgi apparatus appeared in an impeccable condition, and multivesicular bodies were less frequently observed suggesting a lower overall stress strain. The observed ultrastructural changes in winter and after cold acclimation are interpreted as cytological adjustments to winter or a resting state but are not related to frost hardening as Micrasterias cells were unable to improve their freezing stress tolerance. [ABSTRACT FROM AUTHOR]

Diatoms are microalgae that thrive in a range of habitats worldwide including polar areas. Remarkably, non-marine pennate diatoms do not create any morphologically distinct dormant stages that could help them to successfully face unfavourable conditions. Their survival is probably connected with the adaptation of vegetative cells to freezing and desiccation. Here we assessed the freezing tolerance of vegetative cells and vegetative-looking resting cells of 12 freshwater strains of benthic pennate diatoms isolated from polar habitats. To test the effect of various environmental factors, the strains were exposed to −20 °C freezing in four differently treated cultures: (1) vegetative cells growing in standard conditions in standard WC medium and (2) resting cells induced by cold and dark acclimation and resting cells, where (3) phosphorus or (4) nitrogen deficiency were used in addition to cold and dark acclimation. Tolerance was evaluated by measurement of basal cell fluorescence of chlorophyll and determination of physiological cell status using a multiparameter fluorescent staining. Four strains out of 12 were able to tolerate freezing in at least some of the treatments. The minority of cells appeared to be active immediately after thawing process, while most cells were inactive, injured or dead. Overall, the results showed a high sensitivity of vegetative and resting cells to freezing stress among strains originating from polar areas. However, the importance of resting cells for survival was emphasized by a slight but statistically significant increase of freezing tolerance of nutrient-depleted cells. Low numbers of surviving cells in our experimental setup could indicate their importance for the overwintering of diatom populations in harsh polar conditions. [ABSTRACT FROM AUTHOR]

Freezing stress affects the geographic distribution, growth, and development of potato, resulting in yield loss. Solanum malmeanum, a diploid wild species with strong freezing tolerance, was fused with the freezing sensitive dihaploid S. tuberosum by somatic hybridization. In our study, 980 calli were obtained, and 248 differentiated shoots were obtained from the calli. Parental-specific SSR markers were used to analyse the chromosome composition of the 80 randomly selected regenerated plants, obtaining 51 somatic hybrids. Among them, 44 somatic hybrids were tested with ploidy analysis in the years 2016 and 2020. During subculture, the genomic ploidy levels changed due to the composition of the unstable chromosome in 56.82% of the somatic hybrids. The somatic hybrids showed better freezing tolerance than the cultivated parent. Then, freezing-tolerant somatic hybrids were selected to backcross with cultivars, and we obtained valuable breeding resources with enhanced freezing tolerance and tuberization capacity similar to that of cultivars. The correlation analysis showed that freezing tolerance has no relation with tuberization capacity, which indicates that they are controlled by independent genetic loci. Key message: Freezing tolerance was transferred to cultivated potato from S. malmeanum by protoplast fusion for the first time, and valuable resources for freezing tolerance breeding were obtained. [ABSTRACT FROM AUTHOR]

Plant defense and adaptation to adverse environmental conditions rely on gene expression control, such as mRNA transcription, processing, stability, and translation. Sudden temperature changes are common in the era of global warming; thus, understanding plant acclimation responses at the molecular level becomes imperative. mRNA translation initiation regulation has a pivotal role in achieving the synthesis of the appropriate battery of proteins needed to cope with temperature stress. In this study, we analyzed the role of translation initiation factors belonging to the eIF4E family in Arabidopsis acclimation to cold temperatures and freezing tolerance. Using knockout (KO) and overexpressing mutants of AteIF4E1 or AteIF(iso)4E , we found that AteIF4E1 but not AteIF(iso)4E overexpressing lines displayed enhanced tolerance to freezing without previous acclimation at 4°C. However, KO mutant lines, eif(iso)4e-1 and eif4e1-KO , were more sensitive to the stress. Cold acclimation in wild-type plants was accompanied by increased levels of eIF4E1 and eIF(iso)4E transcript levels, polysomes (P) enrichment, and shifts of these factors from translationally non-active to active fractions. Transcripts, previously found as candidates for eIF(iso)4E or eIF4E1 selective translation, changed their distribution in both P and total RNA in the presence of cold. Some of these transcripts changed their polysomal distribution in the mutant and one eIF4E1 overexpressing line. According to this, we propose a role of eIF4E1 and eIF(iso)4E in cold acclimation and freezing tolerance by regulating the expression of stress-related genes. [ABSTRACT FROM AUTHOR]

Background: Winter freezing temperature impacts alfalfa (Medicago sativa L.) persistence and seasonal yield and can lead to the death of the plant. Understanding the genetic mechanisms of alfalfa freezing tolerance (FT) using high-throughput phenotyping and genotyping is crucial to select suitable germplasm and develop winter-hardy cultivars. Several clones of an alfalfa F1 mapping population (3010 x CW 1010) were tested for FT using a cold chamber. The population was genotyped with SNP markers identified using genotyping-by-sequencing (GBS) and the quantitative trait loci (QTL) associated with FT were mapped on the parent-specific linkage maps. The ultimate goal is to develop non-dormant and winter-hardy alfalfa cultivars that can produce extended growth in the areas where winters are often mild. Results: Alfalfa FT screening method optimized in this experiment comprises three major steps: clone preparation, acclimation, and freezing test. Twenty clones of each genotype were tested, where 10 samples were treated with freezing temperature, and 10 were used as controls. A moderate positive correlation (r ~ 0.36, P < 0.01) was observed between indoor FT and field-based winter hardiness (WH), suggesting that the indoor FT test is a useful indirect selection method for winter hardiness of alfalfa germplasm. We detected a total of 20 QTL associated with four traits; nine for visual rating-based FT, five for percentage survival (PS), four for treated to control regrowth ratio (RR), and two for treated to control biomass ratio (BR). Some QTL positions overlapped with WH QTL reported previously, suggesting a genetic relationship between FT and WH. Some favorable QTL from the winter-hardy parent (3010) were from the potential genic region for a cold tolerance gene CBF. The BLAST alignment of a CBF sequence of M. truncatula, a close relative of alfalfa, against the alfalfa reference showed that the gene's ortholog resides around 75 Mb on chromosome 6. Conclusions: The indoor freezing tolerance selection method reported is useful for alfalfa breeders to accelerate breeding cycles through indirect selection. The QTL and associated markers add to the genomic resources for the research community and can be used in marker-assisted selection (MAS) for alfalfa cold tolerance improvement. [ABSTRACT FROM AUTHOR]

Plant defense and adaptation to adverse environmental conditions rely on gene expression control, such as mRNA transcription, processing, stability, and translation. Sudden temperature changes are common in the era of global warming; thus, understanding plant acclimation responses at the molecular level becomes imperative. mRNA translation initiation regulation has a pivotal role in achieving the synthesis of the appropriate battery of proteins needed to cope with temperature stress. In this study, we analyzed the role of translation initiation factors belonging to the eIF4E family in Arabidopsis acclimation to cold temperatures and freezing tolerance. Using knockout (KO) and overexpressing mutants of AteIF4E1 or AteIF(iso)4E, we found that AteIF4E1 but not AteIF(iso)4E overexpressing lines displayed enhanced tolerance to freezing without previous acclimation at 4°C. However, KO mutant lines, eif(iso)4e-1 and eif4e1-KO, were more sensitive to the stress. Cold acclimation in wild-type plants was accompanied by increased levels of eIF4E1 and eIF(iso)4E transcript levels, polysomes (P) enrichment, and shifts of these factors from translationally non-active to active fractions. Transcripts, previously found as candidates for eIF(iso)4E or eIF4E1 selective translation, changed their distribution in both P and total RNA in the presence of cold. Some of these transcripts changed their polysomal distribution in the mutant and one eIF4E1 overexpressing line. According to this, we propose a role of eIF4E1 and eIF(iso)4E in cold acclimation and freezing tolerance by regulating the expression of stress-related genes.

Phenotypic plasticity is often considered important for invasive plant success, yet relatively few studies have assessed plasticity in both native and invasive populations of the same species. We examined the plastic response to temperature for Bromus tectorum populations collected from similar shrub-steppe environments in the Republics of Armenia and Georgia, where it is native, and along an invasive front in California and Nevada. Plants were grown in growth chambers in either 'warm' (30/20 °C, day/night) or 'cold' (10/5 °C) conditions. Invasive populations exhibited greater adaptive plasticity than natives for freezing tolerance (as measured by chlorophyll a fluorescence), such that invasive populations grown in the cold treatment exhibited the highest tolerance. Invasive populations also exhibited more rapid seedling emergence in response to warm temperatures compared to native populations. The climatic conditions of population source locations were related to emergence timing for invasive populations and to freezing tolerance across all populations combined. Plasticity in growth-related traits such as biomass, allocation, leaf length, and photosynthesis did not differ between native and invasive populations. Rather, some growth-related traits were very plastic across all populations, which may help to dampen differences in biomass in contrasting environments. Thus, invasive populations were found to be particularly plastic for some important traits such as seedling emergence and freezing tolerance, but plasticity at the species level may also be an important factor in the invasive ability of B. tectorum. © 2014 Springer International Publishing Switzerland.

Resurrection plants are able to deal with complete dehydration of their leaves and then recover normal metabolic activity after rehydration. Only a few resurrection species are exposed to freezing temperatures in their natural environments, making them interesting models to study the key metabolic adjustments of freezing tolerances. Here, we investigate the effect of cold and freezing temperatures on physiological and biochemical changes in the leaves of Haberlea rhodopensis under natural and controlled environmental conditions. Our data shows that leaf water content affects its thermodynamical properties during vitrification under low temperatures. The changes in membrane lipid composition, accumulation of sugars, and synthesis of stress-induced proteins were significantly activated during the adaptation of H. rhodopensis to both cold and freezing temperatures. In particular, the freezing tolerance of H. rhodopensis relies on a sucrose/hexoses ratio in favor of hexoses during cold acclimation, while there is a shift in favor of sucrose upon exposure to freezing temperatures, especially evident when leaf desiccation is relevant. This pattern was paralleled by an elevated ratio of unsaturated/saturated fatty acids and significant quantitative and compositional changes in stress-induced proteins, namely dehydrins and early light-induced proteins (ELIPs). Taken together, our data indicate that common responses of H. rhodopensis plants to low temperature and desiccation involve the accumulation of sugars and upregulation of dehydrins/ELIP protein expression. Further studies on the molecular mechanisms underlying freezing tolerance (genes and genetic regulatory mechanisms) may help breeders to improve the resistance of crop plants.

Cold-acclimated and non-acclimated contrasting Camelina (Camelina sativa L.) biotypes were investigated for changes in stress-associated biomarkers, including antioxidant enzyme activity, lipid peroxidation, protein, and proline content. In addition, a well-known freezing tolerance pathway participant known as C-repeat/DRE-binding factors (CBFs), an inducer of CBF expression (ICE1), and a cold-regulated (COR6.6) genes of the ICE-CBF-COR pathway were studied at the transcriptional level on the doubled-haploid (DH) lines. Freezing stress had significant effects on all studied parameters. The cold-acclimated DH34 (a freezing-tolerant line) showed an overall better performance under freezing stress than non-acclimated plants. The non-cold-acclimated DH08 (a frost-sensitive line) showed the highest electrolyte leakage after freezing stress. The highest activity of antioxidant enzymes (glutathione peroxidase, superoxide dismutase, and catalase) was also detected in non-acclimated plants, whereas the cold-acclimated plants showed lower enzyme activities upon stress treatment. Cold acclimation had a significantly positive effect on the total protein and proline content of stressed plants. The qRT-PCR analysis revealed significant differences in the expression and cold-inducibility of CsCBF1-3, CsICE1, and CsCOR6.6 genes among the samples of different treatments. The highest expression of all CBF genes was recorded in the non-acclimated frost-tolerant biotype after freezing stress. Interestingly a significantly higher expression of COR6.6 was detected in cold-acclimated samples of both frost-sensitive and -tolerant biotypes after freezing stress. The presented results provide more insights into freezing tolerance mechanisms in the Camelina plant from both a biochemical point of view and the expression of the associated genes.

Low temperature during cold acclimation (CA) leads to the accumulation of detrimental reactive oxygen species (ROS) in plant tissues, which are scavenged by antioxidants, such as ascorbate and glutathione. However, there is a lack of studies examining the dynamics of antioxidants throughout CA, deacclimation (DEA), and reacclimation (REA) in winter wheat. Six winter wheat genotypes were selected to assess the effect of CA, DEA, and REA on the concentrations of ascorbate and glutathione in leaf and crown tissues under two CA temperature treatments. Higher levels of total, reduced, and oxidised ascorbate were determined in leaves, whereas crowns accumulated higher concentrations of nicotinamide adenine dinucleotide (NAD+) after CA, DEA, and REA. Constant low temperature (CLT) during CA led to higher contents of ascorbate and glutathione in both tissues at all stages of acclimation, in comparison with prolonged higher low temperature (PHLT). The concentrations of antioxidants increased after CA, tended to decrease after DEA, and returned to CA levels after REA. Significant positive correlations between freezing tolerance (FT) and antioxidants were only determined under the CA at CLT treatment, thus, affirming the negative effect of PHLT during CA on the FT of winter wheat.

Cold stress poses a serious treat to cultivated kiwifruit since this plant generally has a weak ability to tolerate freezing tolerance temperatures. Surprisingly, however, the underlying mechanism of kiwifruit’s freezing tolerance remains largely unexplored and unknown, especially regarding the key pathways involved in conferring this key tolerance trait. Here, we studied the metabolome and transcriptome profiles of the freezing-tolerant genotype KL (Actinidia arguta) and freezing-sensitive genotype RB (A. arguta), to identify the main pathways and important metabolites related to their freezing tolerance. A total of 565 metabolites were detected by a wide-targeting metabolomics method. Under (−25°C) cold stress, KEGG (Kyoto Encyclopedia of Genes and Genomes) pathway annotations showed that the flavonoid metabolic pathways were specifically upregulated in KL, which increased its ability to scavenge for reactive oxygen species (ROS). The transcriptome changes identified in KL were accompanied by the specific upregulation of a codeinone reductase gene, a chalcone isomerase gene, and an anthocyanin 5-aromatic acyltransferase gene. Nucleotides metabolism and phenolic acids metabolism pathways were specifically upregulated in RB, which indicated that RB had a higher energy metabolism and weaker dormancy ability. Since the LPCs (LysoPC), LPEs (LysoPE) and free fatty acids were accumulated simultaneously in both genotypes, these could serve as biomarkers of cold-induced frost damages. These key metabolism components evidently participated in the regulation of freezing tolerance of both kiwifruit genotypes. In conclusion, the results of this study demonstrated the inherent differences in the composition and activity of metabolites between KL and RB under cold stress conditions.

During autumn, evergreen conifers utilize the decrease in daylength and temperature as environmental signals to trigger cold acclimation, a process that involves the downregulation of photosynthesis, upregulation of photoprotection, and development of cold hardiness. Global warming will delay the occurrence of autumn low temperatures while daylength remains unaffected. The impact of autumn warming on cold acclimation and the length of the carbon uptake period of species with ranges that encompass diverse climates, such as Douglas-fir (Pseudotsuga menziesii), remains unclear. Our study investigated intraspecific variation in the effects of autumn warming on photosynthetic activity, photosynthetic pigments, and freezing tolerance in two interior (var. glauca) and two coastal (var. menziesii) Douglas-fir provenances. Following growth under simulated summer conditions with long days (16 h photoperiod) and summer temperatures (22/13°C day/night), Douglas-fir seedlings were acclimated to simulated autumn conditions with short days (8 h photoperiod) and either low temperatures (cool autumn, CA; 4/−4°C day/night) or elevated temperatures (warm autumn, WA; 19/11°C day/night). Exposure to low temperatures in the CA treatment induced the downregulation of photosynthetic carbon assimilation and photosystem II efficiency, increased the size and de-epoxidation of the xanthophyll cycle pigment pool, and caused the development of sustained nonphotochemical quenching (NPQ). Seedlings in the WA treatment exhibited no downregulation of photosynthesis, no change in xanthophyll cycle pigment de-epoxidation, and no development of sustained NPQ. Albeit these changes, freezing tolerance was not impaired under WA conditions compared with CA conditions. Interior Douglas-fir seedlings developed greater freezing tolerance than coastal seedlings. Our findings suggest that autumn warming, i.e., short photoperiod alone, does not induce the downregulation of photosynthesis in Douglas-fir. Although autumn warming delays the downregulation of photosynthesis, the prolonged period of photosynthetic activity does not bear a trade-off of impaired freezing tolerance.

Background: Kiwifruit (Actinidia Lindl.) is considered an important fruit species worldwide. Due to its temperate origin, this species is highly vulnerable to freezing injury while under low-temperature stress. To obtain further knowledge of the mechanism underlying freezing tolerance, we carried out a hybrid transcriptome analysis of two A. arguta (Actinidi arguta) genotypes, KL and RB, whose freezing tolerance is high and low, respectively. Both genotypes were subjected to − 25 °C for 0 h, 1 h, and 4 h. Results: SMRT (single-molecule real-time) RNA-seq data were assembled using the de novo method, producing 24,306 unigenes with an N50 value of 1834 bp. Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis of DEGs showed that they were involved in the 'starch and sucrose metabolism', the 'mitogen-activated protein kinase (MAPK) signaling pathway', the 'phosphatidylinositol signaling system', the 'inositol phosphate metabolism', and the 'plant hormone signal transduction'. In particular, for 'starch and sucrose metabolism', we identified 3 key genes involved in cellulose degradation, trehalose synthesis, and starch degradation processes. Moreover, the activities of beta-GC (beta-glucosidase), TPS (trehalose-6-phosphate synthase), and BAM (beta-amylase), encoded by the abovementioned 3 key genes, were enhanced by cold stress. Three transcription factors (TFs) belonging to the AP2/ERF, bHLH (basic helix-loop-helix), and MYB families were involved in the low-temperature response. Furthermore, weighted gene coexpression network analysis (WGCNA) indicated that beta-GC, TPS5, and BAM3.1 were the key genes involved in the cold response and were highly coexpressed together with the CBF3, MYC2, and MYB44 genes. Conclusions: Cold stress led various changes in kiwifruit, the 'phosphatidylinositol signaling system', 'inositol phosphate metabolism', 'MAPK signaling pathway', 'plant hormone signal transduction', and 'starch and sucrose metabolism' processes were significantly affected by low temperature. Moreover, starch and sucrose metabolism may be the key pathway for tolerant kiwifruit to resist low temperature damages. These results increase our understanding of the complex mechanisms involved in the freezing tolerance of kiwifruit under cold stress and reveal a series of candidate genes for use in breeding new cultivars with enhanced freezing tolerance. [ABSTRACT FROM AUTHOR]

In Antarctica, multiple stresses (low temperatures, drought and excessive irradiance) hamper photosynthesis even in summer. We hypothesize that controlled inactivation of PSII reaction centres, a mechanism widely studied by pioneer work of Fred Chow and co-workers, may effectively guarantee functional photosynthesis under these conditions. Thus, we analysed the energy partitioning through photosystems in response to temperature in 15 bryophyte species presenting different worldwide distributions but all growing in Livingston Island, under controlled and field conditions. We additionally tested their tolerance to desiccation and freezing and compared those with their capability for sexual reproduction in Antarctica (as a proxy to overall fitness). Under field conditions, when irradiance rules air temperature by the warming of shoots (up to 20 °C under sunny days), a predominance of sustained photoinhibition beyond dynamic heat dissipation was observed at low temperatures. Antarctic endemic and polar species showed the largest increases of photoinhibition at low temperatures. On the contrary, the variation of thermal dissipation with temperature was not linked to species distribution. Instead, maximum non-photochemical quenching at 20 °C was related (strongly and positively) with desiccation tolerance, which also correlated with fertility in Antarctica, but not with freezing tolerance. Although all the analysed species tolerated − 20 °C when dry, the tolerance to freezing in hydrated state ranged from the exceptional ability of Schistidium rivulare (that survived for 14 months at − 80 °C) to the susceptibility of Bryum pseudotriquetrum (that died after 1 day at − 20 °C unless being desiccated before freezing). [ABSTRACT FROM AUTHOR]

Exposure to low, non-freezing temperatures develops freezing tolerance in many plant species. Such process is called cold acclimation. Molecular changes undergone during cold acclimation are orchestrated by signalling networks including MAP kinases. Structure and function of chloroplasts are affected by low temperatures. The aim of this work was to study how the MAP kinases MPK3 and MPK6 are involved in the chloroplast performance upon a long period of cold acclimation. We used Arabidopsis thaliana wild type and mpk3 and mpk6 mutants. Adult plants were acclimated during 7 days at 4 °C and then measurements of PSII performance and chloroplast ultrastructure were carried out. Only the mpk6 acclimated plants showed a high freezing sensitivity. No differences in the PSII function were observed in the plants from the three genotypes exposed to non-acclimated or acclimated conditions. The acclimation of wild-type plants produced severe alterations in the ultrastructure of chloroplast and thylakoids, which was more accentuated in the mpk plants. However, only the mpk6 mutant was unable to internalize the damaged chloroplasts into the vacuole. These results indicate that cold acclimation induces alterations in the chloroplast architecture leading to preserve an optimal performance of PSII. MPK3 and MPK6 are necessary to regulate these morphological changes, but besides, MPK6 is needed to the vacuolization of the damaged chloroplasts, suggesting a role in the chloroplast recycling during cold acclimation. The latter could be quite relevant, since it could explain why this mutant is the only one showing an extremely low freezing tolerance. [ABSTRACT FROM AUTHOR]

Recently, we reported that tuber-derived Solanum tuberosum plants have significantly higher freezing tolerance than tissue-culture-derived plants after cold acclimation. This difference in freezing tolerance were thought to be induced by the presence or absence of the nutrient supply from seed tubers. In this study, in order to check its generality in potato plants, we investigated and compared freezing tolerance of tissue-culture-derived plants and tuber-derived plants of two freezing-tolerant interspecific hybrid potato lines, CB05 04.25 and CB05 24.14. In both hybrid lines, soluble protein contents in above-ground tissues were higher in tuber-derived plants than in tissueculture-derived plants as observed in S. tuberosum. However, levels of freezing tolerance of the two hybrid lines were similar between tissue-culture-derived plants and tuber-derived plants both before and after cold acclimation. The reason why the effects of source of plants on freezing tolerance were different between S. tuberosum and hybrid lines are still unclear. The difference in the response of cells to the freezing stress between freezing-sensitive S. tuberosum and tolerant hybrid lines might cause the difference in the effects of source of plants on freezing tolerance. [ABSTRACT FROM AUTHOR]

This study was conducted to investigate the changes in the levels of soluble carbohydrate, antioxidant capacity, total phenolic acid and fatty acid composition of olive during cold acclimation and deacclimation related with freezing tolerance (FT). Three olive cultivars, Amphisis, Gorgan and Manzanilla were selected and leaf samples were prepared from fieldgrown trees from autumn to summer at six times (November, December, December, February, April and July). The FT was measured by exposure to artificial freezing (0ºC to -25ºC). Result showed that there was a significant difference between the olive cultivars in terms of FT at each stage of sampling. The FT of cultivars increased during the cold acclimation stage and declined in deacclimation stage. Soluble carbohydrates, antioxidant capacity, total phenolic content, leaf dry matter, especially from the beginning of cold acclimation until full acclimation, in cold-tolerant -Amphisis- cultivar was more than the Gorgan and Manzanilla cultivars. The highest ratio of unsaturated to saturated fatty acids in both January and July was observed in Amphisis cultivar. Therefore, compounds such as soluble carbohydrates, total phenolic content, and fatty acids can be used as biochemical indicator of FT in screening of olive cultivars. [ABSTRACT FROM AUTHOR]

Red clover (Trifolium pratense L.) is the most important forage legume in the Nordic region, but its utilization is limited by poor persistency. The improvement of cultivated red clover can potentially take advantage of the numerous wild populations and landraces conserved in gene banks; however, there is often limited information available on the phenotypic and genetic characteristics of this material. We characterized 48 populations conserved at NordGen for a number of traits and compared them to commercial cultivars. The material was evaluated in field trials at four locations over two years and in an experiment under controlled conditions. Considerable variation was identified, with stem length, growth type and flowering date having the highest broad sense heritabilities. Traits related to plant size were strongly associated with late flowering and upright growth and differed between landraces/cultivars on the one hand and wild populations on the other. There was a large genotype by environment interaction on winter survival, which only partially correlated with freezing tolerance under controlled conditions. A majority of gene bank accessions exceeded the commercial cultivars in winter survival and freezing tolerance and can therefore be a genetic resource for future improvement of these traits. The phenotypic variation among accessions was associated with two main axes of climatic variation at the collection site. Petiole length of young plants under controlled conditions as well as plant size in the field increased with increasing summer temperature and decreasing summer precipitation, while number of leaves and an apparent vernalization requirement, recorded under controlled conditions, increased with decreasing annual and winter temperature. We discuss the implications these results have for collection, conservation and utilization of red clover genetic resources in the Nordic region. [ABSTRACT FROM AUTHOR]

Cold stress poses a serious treat to cultivated kiwifruit since this plant generally has a weak ability to tolerate freezing tolerance temperatures. Surprisingly, however, the underlying mechanism of kiwifruit's freezing tolerance remains largely unexplored and unknown, especially regarding the key pathways involved in conferring this key tolerance trait. Here, we studied the metabolome and transcriptome profiles of the freezing-tolerant genotype KL (Actinidia arguta) and freezing-sensitive genotype RB (A. arguta), to identify the main pathways and important metabolites related to their freezing tolerance. A total of 565 metabolites were detected by a wide-targeting metabolomics method. Under (−25°C) cold stress, KEGG (Kyoto Encyclopedia of Genes and Genomes) pathway annotations showed that the flavonoid metabolic pathways were specifically upregulated in KL, which increased its ability to scavenge for reactive oxygen species (ROS). The transcriptome changes identified in KL were accompanied by the specific upregulation of a codeinone reductase gene, a chalcone isomerase gene, and an anthocyanin 5-aromatic acyltransferase gene. Nucleotides metabolism and phenolic acids metabolism pathways were specifically upregulated in RB, which indicated that RB had a higher energy metabolism and weaker dormancy ability. Since the LPCs (LysoPC), LPEs (LysoPE) and free fatty acids were accumulated simultaneously in both genotypes, these could serve as biomarkers of cold-induced frost damages. These key metabolism components evidently participated in the regulation of freezing tolerance of both kiwifruit genotypes. In conclusion, the results of this study demonstrated the inherent differences in the composition and activity of metabolites between KL and RB under cold stress conditions. [ABSTRACT FROM AUTHOR]

Freezing temperature during overwintering often kills plants; plants have thus, developed a defense mechanism called 'cold acclimation', in which a number of genes are involved in increasing cell protection and gene expression. Mitogen-activated protein kinase (MAPK) controls proteins' activities by phosphorylation and is involved in numerous metabolic pathways. In this study, we identified the protein interaction between TaMAPK3 and the proteins in the cold response pathway, ICE41, ICE87, and CBFIVd-D9. The subcellular localization and bimolecular fluorescence complement (BiFC) assays revealed that these proteins interact in the nucleus or in the plasma membrane. Furthermore, MAPK3-mediated phosphorylation of ICE41, ICE87, and CBFIVd-D9 was verified using an in vitro phosphorylation assay. TaMAPK3 -overexpressing transgenic Brachypodium showed a lower survival rate upon freezing stress and lower proline content during cold acclimation, compared to wild-type plants. Furthermore, cold response gene expression analysis revealed that the expression of these genes was suppressed in the transgenic lines under cold treatment. It was further elucidated that MAPK3 mediates the degradation of ICE and CBF proteins, which implies the negative impact of MAPK3 on the freezing tolerance of plants. This study will help to elucidate the molecular mechanisms of cold tolerance and the activity of MAPK3 in wheat. [ABSTRACT FROM AUTHOR]

Elevation gradients provide powerful study systems for examining the influence of environmental filters in shaping species assemblages. High-mountain habitats host specific high-elevation assemblages, often comprising specialist species adapted to endure pronounced abiotic stress, while such harsh conditions prevent lowland species from colonizing or establishing. While thermal tolerance may drive the altitudinal segregation of ectotherms, its role in structuring aquatic insect communities remains poorly explored. This study investigates the role of thermal physiology in shaping the current distribution of high-mountain diving beetles from the Sierra Nevada Iberian mountain range and closely related lowland species. Cold tolerance of five species from each altitudinal zone was measured estimating the supercooling point (SCP), lower lethal temperature (LLT) and tolerance to ice enclosure, while heat tolerance was assessed from the heat coma temperature (HCT). Alpine species exhibited wider fundamental thermal niches than lowland species, likely associated with the broader range of climatic conditions in high-mountain areas. Cold tolerance did not seem to prevent lowland species from colonizing higher elevations, as most studied species were moderately freeze-tolerant. Therefore, fundamental thermal niches seem not to fully explain species segregation along elevation gradients, suggesting that other thermal tolerance traits, environmental factors, and biotic interactions may also play important roles. • Elevation gradients shape species assemblages. • Alpine lakes host specialist species. • This study investigates thermal physiology in diving beetles. • Alpine diving beetles exhibit wider thermal niches than lowland species. • Cold tolerance did not fully explain species distribution. [ABSTRACT FROM AUTHOR]

As the direct recipients of plant abscisic acid (ABA), pyrabactin resistance/pyrabactin resistance-like/regulatory component of ABA receptor proteins (PYR/PYL/RCAR; hereinafter called PYLs) play pivotal roles in plant coercive responses. However, the PYL genes in Acer palmatum have yet to be investigated. In this study, using a genome search method, we identified 14 A. palmatum PYL genes (ApPYLs) and clustered them into three clades based on the phylogenetic, gene structure, and conserved motif analyses. The ApPYLs were dispersed on eight chromosomes of A. palmatum ; cis -acting element analysis indicated that the ApPYLs were involved in biological processes, including resistance, hormone regulation, and growth. Gene expression profiling of cold-treated A. palmatum plants revealed that ApPYL1 and ApPYL2 may be related to cold stress response. ApPYL1 and ApPYL2 interacted directly with ApMYB44 , and the ApPYL1 , ApPYL2 , and ApMYB44 genes were found to enhance freezing tolerance in Arabidopsis. These findings illustrate that ApPYL1 and ApPYL2 respond to the cold signalling pathway by interacting with ApMYB44 and provide a reference for improving cold resistance of plants. • Fourteen PYL genes in three subgroups were identified in the A. palmatum genome. • ApPYL1 and ApPYL2 were highly expressed after exposure to cold stress. • ApPYL1 and ApPYL2 overexpression improved frost resistance in A. thaliana. • ApPYL1/ApPYL2–ApMYB44 complex may be involved in cold signalling response. [ABSTRACT FROM AUTHOR]

Subzero temperatures are often lethal to plants. Many temperate herbaceous plants have a cold acclimation mechanism that allows them to sense a drop in temperature and prepare for freezing stress through accumulation of soluble sugars and cryoprotective proteins. As ice formation primarily occurs in the apoplast (the cell wall space), cell wall functional properties are important for plant freezing tolerance. Although previous studies have shown that the amounts of constituent sugars of the cell wall, in particular those of pectic polysaccharides, are altered by cold acclimation, the significance of this change during cold acclimation has not been clarified. We found that β-1,4-galactan, which forms neutral side chains of the acidic pectic rhamnogalacturonan-I, accumulates in the cell walls of Arabidopsis and various freezing-tolerant vegetables during cold acclimation. The gals1 gals2 gals3 triple mutant, which has reduced β-1,4-galactan in the cell wall, exhibited impaired freezing tolerance compared with wild-type Arabidopsis during initial stages of cold acclimation. Expression of genes involved in the galactan biosynthesis pathway, such as galactan synthases and UDP-glucose 4-epimerases , was induced during cold acclimation in Arabidopsis, explaining the galactan accumulation. Cold acclimation resulted in a decrease in extensibility and an increase in rigidity of the cell wall in the wild type, whereas these changes were not observed in the gals1 gals2 gals3 triple mutant. These results indicate that the accumulation of pectic β-1,4-galactan contributes to acquired freezing tolerance by cold acclimation, likely via changes in cell wall mechanical properties. [Display omitted] • Cold acclimation leads to an increase in pectic galactan within plant cell walls • Reduced galactan levels result in a decrease in plant freezing tolerance • Galactan plays a role in altering wall mechanical properties during cold acclimation β-1,4-galactan constitutes a side chain of pectin in plant cell walls. Takahashi et al. show an accumulation of pectic galactan with cold acclimation treatment in plants. This increase in pectic galactan may be involved in the enhancement of freezing tolerance and changes in cell wall mechanical properties during cold acclimation. [ABSTRACT FROM AUTHOR]

This study explores and compares the limits for photosynthesis in subzero temperatures of six Antarctic lichens: Sphaerophorus globosus, Caloplaca regalis , Umbilicaria antarctica , Pseudephebe minuscula , Parmelia saxatilis and Lecania brialmontii combining linear cooling and chlorophyll fluorescence methods. The results revealed triphasic S-curves in the temperature response of the maximum quantum yield (F V /F M) and effective quantum yield of photosystem II (Φ PSII) for all species. All investigated species showed a high level of cryoresistance with critical temperatures (T c) below −20 °C. However, record low T c temperatures have been discovered for L. brialmotii (−54 °C for F V /F M and −40 °C for Φ PSII) and C. regalis (−52 °C for F V /F M and −38 °C for Φ PSII). Additionally, the yield differentials (F V /F M − Φ PSII) in functions of temperature revealed one or two peaks, with the larger one occurring for temperatures below −20 °C for the above-mentioned species. Finally, Kautsky kinetics were measured and compared at different temperatures (20 °C, 10 °C, 0 °C and −10 °C and then −10 °C after 1 h of incubation). This research serves as a foundation for further developing investigations into the biophysical mechanisms by which photosynthesis is carried out at subzero temperatures. [Display omitted] • Antarctic lichens are cryo-resistant, their critical (cooling) temperature was found below -20°C. • Among 6 tested lichens, Lecania brialmontii showed highest frost-resistance. • Critical (cooling) temperature is lower for maximum quantum yield (F V /F M) than effective quantum yield of PSII (Φ PSII). • Differential between normalized F V /F M and Φ PSII is suggested as a new parameter for cold-resistance studies. [ABSTRACT FROM AUTHOR]

In order to predict future range shifts for invasive species it is important to explore their ability to acclimate to the new environment and understand physiological and reproductive constraints controlling their distribution. My dissertation studied mechanisms by which temperature may affect the distribution of two aggressive plant invaders in North America, Bromus tectorum and Bromus rubens. I first evaluated winter freezing tolerance of Bromus species and demonstrated that the mechanism explaining their distinct northern range limits is different acquisition time of freezing tolerance. While B. rubens has a slower rate of freezing acclimation that leads to intolerance of sudden, late-autumn drops in temperature below -12°C, B. tectorum rapidly hardens and so is not impacted by the sudden onset of severe late-autumn cold. In addition, the analysis of male reproductive development and seed production showed that neither species produces seed at or above 36°C, due to complete pollen sterility, which might trigger climate-mediated range contractions at B. tectorum and B. rubens southern margins. Finally, a detailed gas-exchange analysis combined with biochemical modelling demonstrated that both species acclimate to a broad range of temperatures and photosynthetic response to temperature does not explain their current range separation.

Tardigrades are small micrometazoans able to resist several environmental stresses in any stage of their life cycle. An integrated analysis of tardigrade specimens collected in Tsukuba (Japan) revealed a peculiar morphology and a new sensory field in the cloaca. Molecular taxonomy and phylogenetic analysis on different genes (COI, ITS2, 18S and 28S) confirmed that this population is a new species, Macrobiotus kyoukenus sp. nov., belonging to the widespread Macrobiotus hufelandi group. The stress resistance capabilities of M. kyoukenus sp. nov. have been tested by submitting animals to extreme desiccation, rapid freezing, and high levels of ultraviolet radiations (UVB and UVC). Animals were able to survive desiccation (survivorship 95.71 ± 7.07%) and freezing up to −80 °C (82.33 ± 17.11%). Both hydrated and desiccated animals showed a high tolerance to increasing UV radiations: hydrated animals survived to doses up to 152.22 kJ m−2 (UVB) and up to 15.00 kJ m−2 (UVC), while desiccated specimens persisted to radiations up to 165.12 kJ m−2 (UVB) and up to 35.00 kJ m−2 (UVC). Present data contribute to the discovery of a larger tardigrade biodiversity in Japan, and the tolerance capabilities of M. kyoukenus sp. nov. show that it could become a new emerging model for stress resistance studies.

The APETALA2 (AP2) and ethylene-responsive element-binding factor (ERF) gene family is one of the largest plant-specific transcription factor gene families, which plays a critical role in plant development and evolution, as well as response to various stresses. The TARGET OF EAT3 (TOE3) gene is derived from Halostachys caspica and belongs to the AP2 subfamily with two AP2 DNA-binding domains. Currently, AP2 family mainly plays crucial roles in plant growth and evolution, yet there are few reports about the role of AP2 in abiotic stress tolerance. Here, we report HcTOE3, a new cold-regulated transcription factor gene, which has an important contribution to freezing tolerance. The main results showed that the expression of HcTOE3 in the H. caspica assimilating branches was strongly induced by different abiotic stresses, including high salinity, drought, and extreme temperature (heat, chilling, and freezing), as well as abscisic acid and methyl viologen treatments. Overexpressing HcTOE3 gene (OE) induced transgenic Arabidopsis plant tolerance to freezing stress. Under freezing treatment, the OE lines showed lower content of malondialdehyde and electrolyte leakage and less accumulation of reactive oxygen species compared with the wild type. However, the survival rates, antioxidant enzyme activities, and contents of osmotic adjustment substance proline were enhanced in transgenic plants. Additionally, the OE lines increased freezing tolerance by up-regulating the transcription level of cold responsive genes (CBF1, CBF2, COR15, COR47, KIN1, and RD29A) and abscisic acid signal transduction pathway genes (ABI1, ABI2, ABI5, and RAB18). Our results suggested that HcTOE3 positively regulated freezing stress and has a great potential as a candidate gene to improve plant freezing tolerance.

Winter rapeseed is susceptible to low temperature during winter in Northwest China, which could lead to a severe reduction of crop production. The freezing temperature could stress the whole plant, especially the leaf, and ultimately harm the survival rate of winter rapeseed. However, the molecular mechanism underlying freezing tolerance is still unclear in winter rapeseed. In this study, a comprehensive investigation of winter rapeseed freezing tolerance was conducted at the levels of transcript, protein, and physiology and biochemistry, using a pair of freezing-sensitive and freezing-resistant cultivars NQF24 and 17NTS57. There were 4,319 unique differentially expressed genes (DEGs) and 137 unique differentially abundant proteins (DAPs) between two cultivars identified in leaf under freezing stress. Function enrichment analysis showed that most of the enriched DEGs and DAPs were involved in plant hormone signal transduction, alpha-linolenic/linoleic acid metabolism, peroxisome, glutathione metabolism, fatty acid degradation, and secondary metabolite biosynthesis pathways. Based on our findings, it was speculated that freezing tolerance formation is caused by increased signal transduction, enhanced biosynthesis of protein, secondary metabolites, and plant hormones, elevated energy supply, greater reactive oxygen species scavenging, and lower lipid peroxidation as well as stronger cell stability in leaf under freezing stress. These results provide a comprehensive profile of leaf response under freezing stress, which have potential to be used as selection indicators of breeding programs to improve freezing tolerance in rapeseed.

Background: A variety of basic and applied research programs in plant biology require the accurate and reliable determination of plant tissue cold hardiness. Over the past 50 years, the electrolyte leakage method has emerged as a popular and practical method for quantifying the amount of damage inflicted on plant tissue by exposure to freezing temperatures. Numerous approaches for carrying out this method and analyzing the resultant data have emerged. These include multiple systems for standardizing and modeling raw electrolyte leakage data and multiple protocols for boiling or autoclaving samples in order to maximize leakage as a positive control. We compare four different routines for standardization of leakage data and assess a novel control method—immersion in liquid nitrogen in lieu of traditional autoclaving—and apply them to woody twigs collected from 12 maple (Acer) species in early spring. We compare leakage data from these samples using each of four previously published forms of data analysis and autoclaving vs. liquid nitrogen controls and validate each of these approaches against visual estimates of freezing damage and differential thermal analysis. Results: Through presentation of our own data and re-analysis of previously published findings, we show that standardization of raw data against estimates of both minimum and maximum attainable freezing damage allows for reliable estimation of cold hardiness at the species level and across studies in diverse systems. Furthermore, use of our novel liquid nitrogen control produces data commensurate across studies and enhances the consistency and realism of the electrolyte leakage method, especially for very cold hardy samples. Conclusion: Future leakage studies that relativize data against minimum and maximum leakage and that employ our updated liquid nitrogen control will contribute generalizable, repeatable, and realistic data to the existing body of cold hardiness research in woody plants. Data from studies conducted using a liquid nitrogen (and not an autoclaving) control can still be compared to previously published data, especially when raw data are standardized using the best-performing approach among those we assessed. Electrolyte leakage of woody twigs emerges as a useful technique for quickly assessing the probability of tissue death in response to freezing in dormant plants. Differential thermal analysis may provide different and complementary information on cold hardiness. [ABSTRACT FROM AUTHOR]

The APETALA2 (AP2) and ethylene-responsive element-binding factor (ERF) gene family is one of the largest plant-specific transcription factor gene families, which plays a critical role in plant development and evolution, as well as response to various stresses. The TARGET OF EAT3 (TOE3) gene is derived from Halostachys caspica and belongs to the AP2 subfamily with two AP2 DNA-binding domains. Currently, AP2 family mainly plays crucial roles in plant growth and evolution, yet there are few reports about the role of AP2 in abiotic stress tolerance. Here, we report HcTOE3 , a new cold-regulated transcription factor gene, which has an important contribution to freezing tolerance. The main results showed that the expression of HcTOE3 in the H. caspica assimilating branches was strongly induced by different abiotic stresses, including high salinity, drought, and extreme temperature (heat, chilling, and freezing), as well as abscisic acid and methyl viologen treatments. Overexpressing HcTOE3 gene (OE) induced transgenic Arabidopsis plant tolerance to freezing stress. Under freezing treatment, the OE lines showed lower content of malondialdehyde and electrolyte leakage and less accumulation of reactive oxygen species compared with the wild type. However, the survival rates, antioxidant enzyme activities, and contents of osmotic adjustment substance proline were enhanced in transgenic plants. Additionally, the OE lines increased freezing tolerance by up-regulating the transcription level of cold responsive genes (CBF1 , CBF2 , COR15 , COR47 , KIN1 , and RD29A) and abscisic acid signal transduction pathway genes (ABI1 , ABI2 , ABI5 , and RAB18). Our results suggested that HcTOE3 positively regulated freezing stress and has a great potential as a candidate gene to improve plant freezing tolerance. [ABSTRACT FROM AUTHOR]

Winter rapeseed is susceptible to low temperature during winter in Northwest China, which could lead to a severe reduction of crop production. The freezing temperature could stress the whole plant, especially the leaf, and ultimately harm the survival rate of winter rapeseed. However, the molecular mechanism underlying freezing tolerance is still unclear in winter rapeseed. In this study, a comprehensive investigation of winter rapeseed freezing tolerance was conducted at the levels of transcript, protein, and physiology and biochemistry, using a pair of freezing-sensitive and freezing-resistant cultivars NQF24 and 17NTS57. There were 4,319 unique differentially expressed genes (DEGs) and 137 unique differentially abundant proteins (DAPs) between two cultivars identified in leaf under freezing stress. Function enrichment analysis showed that most of the enriched DEGs and DAPs were involved in plant hormone signal transduction, alpha-linolenic/linoleic acid metabolism, peroxisome, glutathione metabolism, fatty acid degradation, and secondary metabolite biosynthesis pathways. Based on our findings, it was speculated that freezing tolerance formation is caused by increased signal transduction, enhanced biosynthesis of protein, secondary metabolites, and plant hormones, elevated energy supply, greater reactive oxygen species scavenging, and lower lipid peroxidation as well as stronger cell stability in leaf under freezing stress. These results provide a comprehensive profile of leaf response under freezing stress, which have potential to be used as selection indicators of breeding programs to improve freezing tolerance in rapeseed. [ABSTRACT FROM AUTHOR]

Freezing tolerance in an organism that rarely experiences freezing could provide support for the hypothesis that the DT trait inherently confers some degree of freezing tolerance. During freezing, plant cells are dehydrated, but it remains unclear whether freezing DT plant cells typically (i) suffer partial dehydration or (ii) transition into the desiccated, inactive state (but this could be species dependent, see Fernández-Marín et al., 2018; Georgieva et al., 2020). However, if freezing tolerance of DT plants is merely a mechanistic side effect of desiccation tolerance, then freezing tolerance in DT plants may be de-coupled from the minimum temperatures of their habitats. [Extracted from the article]

An altitudinal gradient of leaf water repellency is often observed between and within species. In a previous study of Arabidopsis halleri, cauline leaves (stem leaves that wrap flowering buds) showed higher water repellency in exposed semi-alpine plants than in understory low-elevation plants. Here, we examined altitudinal variations in the cuticular wax content of the leaf surface and experimentally evaluated the role of high water repellency of cauline leaves. Leaf cuticular wax was analysed using comprehensive two-dimensional gas chromatography (GC)-mass spectrometry and a GC-flame ionisation detector. Young flowering buds wrapped by cauline leaves were exposed to freezing temperatures with or without water, and frost damage to the flowering buds was compared between plants from semi-alpine and low-elevation habitats. Higher amounts of C29, C31, and C33 alkanes were observed in the cauline leaves of semi-alpine plants than in those of low-elevation plants. In the freezing experiment, water application increased damage to the flowering buds of low-elevation plants, and the extent of damage to the flowering buds was lower in semi-alpine plants than in low-elevation plants when water was applied to the plant surface. Genetic variations in the amounts of alkanes on the leaf surface depending on the altitude occurred specifically in cauline leaves. Our results indicate that the water repellency of cauline leaves presumably minimises frost damage to flowering buds at high altitudes. [ABSTRACT FROM AUTHOR]

Theoretical and experimental studies have demonstrated that temperature is an important environmental factor that affects the regional distribution of plants. However, how to modify the distribution pattern of plants in different regions is a focus of current research. Obtain the information of cold tolerance genes from cold tolerance species, cloning genes with real cold tolerance effects is one of the most important ways to find the genes related to cold tolerance. In this study, we investigated whether transferring the VHA-c gene from Antarctic notothenioid fishes into Arabidopsis enhances freezing tolerance of Arabidopsis. The physiological response and molecular changes of VHA-c overexpressing pedigree and wildtype Arabidopsis were studied at −20 °C. The results showed that the malondialdehyde (MDA) and membrane leakage rates of WT plants were significantly higher than those of VHA-c8 and VHA-c11 plants, but the soluble sugar, soluble protein, proline and ATP contents of WT plants were significantly lower than those of VHA-c8 and VHA-c11 plants under −20 °C freezing treatment. The survival rate, VHA-c gene expression level and VHA-c protein contents of WT plants were significantly lower than those of VHA-c8 and VHA-c11 plants under −20 °C freezing treatment. Correlation analysis showed that ATP content was significantly negatively correlated with MDA and membrane leakage rate, and positively correlated with soluble sugar, soluble protein and proline content under −20 °C freezing treatment. These results demonstrated that overexpression of the VHA-c gene provided strong freezing tolerance to Arabidopsis by increasing the synthesis of ATP and improved the adaptability of plants in low temperature environment. • Transgenic Arabidopsis showed stronger resistance to freezing than WT. • Transgenic Arabidopsis can increase the content of ATP. • ATP content was negatively correlated with MDA and membrane leakage rate. • ATP content was positively correlated with osmoregulation substances. • VHA-c gene has potential application prospect in genetic improvement of cold tolerance. [ABSTRACT FROM AUTHOR]

The moor frog Rana arvalis is one of a few amphibians that can tolerate freezing to low temperatures, up to −16 °C. In this study, we performed metabolomic analysis of the liver and hindlimb muscles of frozen and control R. arvalis. We found that the moor frog synthesizes glucose and glycerol in similar concentrations as low molecular weight cryoprotectants. This is the first such case reported for the genus Rana, which was believed to use glucose only. We found that freezing upregulates glycolysis, with the accumulation of several end products: lactate, alanine, ethanol, and, possibly, 2,3-butanediol. To our knowledge, this is also the first report of ethanol as an end product of glycolysis in terrestrial vertebrates. We observed highly increased concentrations of nucleotide degradation products, implying high level of stress. The Krebs cycle arrest resulted in high concentrations of succinate, which is common for animals. However, we found almost no signs of adaptations to reoxygenation stress, with overall low levels of antioxidants. We also performed metabolomics analysis of subcutaneous ice that was found to contain glucose, glycerol, and several other substances.