Freezing Tolerance Research Articles

Chromosome-level genome provides new insight into the overwintering process of Korla pear (Pyrus sinkiangensis Yu)

Korla pear has a unique taste and aroma and is a breeding parent of numerous pear varieties. It is susceptible to Valsa mali var. pyri, which invades bark wounded by freezing injury. Its genetic relationships have not been fully defined and could offer insight into the mechanism for freezing tolerance and disease resistance. We generated a high-quality, chromosome-level genome assembly for Korla pear via the Illumina and PacBio circular consensus sequencing (CCS) platforms and high-throughput chromosome conformation capture (Hi-C). The Korla pear genome is ~ 496.63 Mb, and 99.18% of it is assembled to 17 chromosomes. Collinearity and phylogenetic analyses indicated that Korla might be derived from Pyrus pyrifolia and that it diverged ~ 3.9-4.6 Mya. During domestication, seven late embryogenesis abundant (LEA), two dehydrin (DHN), and 54 disease resistance genes were lost from Korla pear compared with P. betulifolia. Moreover, 21 LEA and 31 disease resistance genes were common to the Korla pear and P. betulifolia genomes but were upregulated under overwintering only in P. betulifolia because key cis elements were missing in Korla pear. Gene deletion and downregulation during domestication reduced freezing tolerance and disease resistance in Korla pear. These results could facilitate the breeding of novel pear varieties with high biotic and abiotic stress resistance.

Assembly and analysis of sequence from a spring and winter type Camelina sativa by whole genome PacBio HiFi technologies

Camelina sativa is an emerging oilseed crop with potential for multi-cropping with more traditional cash crops. Until very recently, the only reference genome for camelina was produced using short read technologies and was considerably fragmented. To facilitate the mapping of traits associated with vernalization requirement and freezing tolerance in an F7 recombinant inbred line (RIL) population developed by crossing a spring- and winter-biotype of camelina, we performed long read sequencing using PacBio HiFi technology. Here we report on the genome assembly and gene annotation obtained from the long read sequencing of the two parental lines. Both assemblies formed 20 chromosomal units with a genome size of 667,897,934 and 714,129,675 bases for C046 and Joelle, respectively. Assessment of completeness by BUSCO analysis indicated the C046 and Joelle genomes were both 99.5 complete with 98.1 % of the conserved genes being duplicated. Skim-sequencing of the F6:7 RILs resulted in 823,142 markers that mapped to variants identified by comparisons of the parental genome sequences. These sequences provide a valuable resource for breeders seeking to improve the food and industrial attributes of camelina.

CaMYB80 enhances the cold tolerance of pepper by directly targeting CaPOA1.

Cold temperatures negatively impact crop yield and quality, posing significant limitations to the advancement of the vegetable industry. MYB transcription factors are pivotal in enhancing plant resilience against various abiotic stresses, including low-temperature stress. Pepper (Capsicum annuum L.) is a nutrient-rich vegetable crop sensitive to low temperatures. This study aimed to determine the function of CaMYB80 in the cold stress response of pepper through virus-induced silencing. The study also conducted heterologous expression of CaMYB80 in Arabidopsis and tomato plants. The results showed that CaMYB80 could respond to low-temperature stress in pepper. CaMYB80 was localized in the nucleus and cytoplasm and exhibited transcriptional activation ability. Moreover, CaMYB80 silencing decreased cold tolerance in pepper, while its heterologous overexpression increased cold tolerance in Arabidopsis and tomato. Further analysis showed that CaMYB80 interacted with CaPOA1 (peroxidase N1-like). Similarly, the expression of CaPOA1 also responded to low-temperature stress. Overexpression of CaPOA1 enhanced freezing tolerance in Arabidopsis, while its silencing reduced cold stress tolerance in pepper. Furthermore, overexpression of CaMYB80 in Arabidopsis and tomato could increase the activity of peroxidases and the expression levels of genes in the ICE-CBF-COR (inducer of CBF expression, C-repeat binding factor, cold-responsive) regulatory network. In conclusion, our research results indicate that CaMYB80 enhances pepper cold tolerance by interacting with CaPOA1 to increase peroxidase activity and influence the expression of ICE-CBF-COR related genes.

Strong Tough Ionic Organohydrogels with Negative‐Thermopower via the Synergy of Coordination Interaction and Hofmeister Effect

AbstractIonic thermoelectric (i‐TE) gels have attracted widespread attention in flexible electronics due to their flexibility, leakage‐free, low toxicity, and converting low‐grade waste heat into electricity. For practical application, it is strongly desirable to fabricate negative‐thermopower (n‐type) i‐TE gels with excellent mechanical performances, which is still a big challenge. Herein, a multi‐hierarchical network structure is proposed to simultaneously improve the TE and mechanical performance. Such structure including hierarchical hydrogen bonds, coordination bonds, and dense polymer chains is realized via the synergy of the coordination effect of polymer to cations and the Hofmeister effect of anions to polymer. As a result, the optimized gel exhibits not only negative thermopower up to −3.69 mV K−1 with a conductivity of 0.15 S m−1 at room temperature, but also outstanding tensile strength (>6.7 MPa), elongation at break (>1100%), and toughness (>43 MJ m−3), which is the toughest n‐type i‐TE gel reported to date. In addition, these n‐type i‐TE gels present good freeze tolerance (−58.53 °C) and dry resistance. Furthermore, the application potentials of the i‐TE device are proven in converting human thermal power into electricity. These findings provide a new way to obtain high‐performance n‐type i‐TE materials by synergistic anion and cation action on polymers.

Floral freezing tolerance is tied to flowering time in North American woody plant species.

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. 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. 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.

Waterlogging‐Induced Changes in ABA, Carbohydrates and CBF6 Modify Freezing Tolerance in Prehardened Festuca pratensis

ABSTRACTChanges in precipitation and snow melt during warmer winters can increase low‐temperature waterlogging. Such conditions may bring about different effects when compared with a single stress trigger, such as low‐temperature or water excess. The effects of waterlogging are clearly related to water temperature, and the consequences of water excess might be less severe, as more oxygen is dissolved in colder water. The effect of waterlogging during cold acclimation (CA) is poorly understood; most experiments concerning water excess are performed at relatively high‐temperatures. In this study, we examined the effect of 3 weeks of waterlogging (approx. 2 cm above the soil level) on CA in Festuca pratensis Huds. (Fp), a cool‐season grass. Measurements were taken before CA (after prehardening, before flooding) and after 3 weeks of CA in waterlogged (treated) and non‐waterlogged (control) plants. The work included: (i) freezing tolerance test (regrowth after freezing), (ii) analysis of abscisic acid (ABA) content in the leaf, (iii) leaf stomatal conductance, (iv) leaf water content, (v) carbohydrates analysis, including fructans, and (vi) transcript levels of selected genes involved in freezing tolerance, ABA signalling and fructan biosynthesis. The aim of the study was to test a hypothesis that low‐temperature waterlogging in Fp enhances freezing tolerance (plant regrowth after freezing) related to increased ABA accumulation, increased C‐repeat‐binding transcription factor expression and/or increased carbohydrate accumulation, including fructans. Two out of four genotypes exhibited enhanced regrowth following freezing due to waterlogging relative to control. Principal component analysis (PCA) revealed a positive correlation between ABA levels and freezing tolerance in both treatments, with a more pronounced effect observed in the waterlogged plants. However, the phytohormone played different roles in these two treatments. In the context of low‐temperature waterlogging, ABA may be involved in the dehydration tolerance response in genotypes suffering from physiological drought, as well as the induction of C‐repeat‐binding transcription factors (CBFs) and sucrose, which may improve freezing tolerance. The increased fructan amount and polymerisation degree due to waterlogging may provide a carbohydrate sink to maintain a high photosynthetic efficiency, but are not directly responsible for freezing tolerance changes. The study indicates that tolerance mechanisms of Fp exposed to low‐temperature waterlogging involve maintaining a high photosynthetic rate, as well as oxidative and dehydration stress tolerance.

A double-edged sword: Phosphorylation of Ca2+ channel CNGC20 fine-tunes plant freezing tolerance.

A double-edged sword: Phosphorylation of Ca2+ channel CNGC20 fine-tunes plant freezing tolerance.

Small Stomates and Xylem Vessels Associated With Freeze Tolerance in Winter Barley

ABSTRACTFreeze tolerance is a complex agronomic trait that is difficult to evaluate in the field because of year‐to‐year variation in weather. Discovering plant characteristics closely related to freeze tolerance would enable more effective selection for this important trait. To explore possible physiological mechanisms and search for useful characteristics related to freeze tolerance in winter barley, we conducted field and growth chamber experiments with seven freeze‐tolerant and seven freeze‐susceptible genotypes that exhibited contrasting winter survival in preliminary field screenings. In a 2‐year field experiment, malondialdehyde, proline and water‐soluble carbohydrate concentrations were measured during cold acclimation and deacclimation to investigate differences in osmoregulation and membrane stability between freeze‐tolerant and freeze‐sensitive genotypes. All parameters varied by sampling year, and significant differences between freeze tolerance groups were found mainly during cold deacclimation in the spring. In growth chamber experiments, the size of xylem vessels and stomates was measured with and without cold acclimation. Freeze‐tolerant genotypes had smaller xylem vessels and stomates than freeze‐sensitive genotypes with and without cold acclimation, and small stomatal length was associated with a small xylem vessel area. Thus, it may be possible to improve freeze tolerance in winter barley by selecting smaller xylem and stomate cells. This study also validated germplasms of winter barley with differential freeze tolerance for future projects on breeding for improving winter hardiness and on plant physiology and genetics in response to freezing stress.

Physiological, transcriptomic and metabolomic insights of three extremophyte woody species living in the multi-stress environment of the Atacama Desert.

In contrast to Neltuma species, S. tamarugo exhibited higher stress tolerance, maintaining photosynthetic performance through enhanced gene expression and metabolites. Differentially accumulated metabolites include chlorophyll and carotenoids and accumulation of non-nitrogen osmoprotectants. Plant species have developed different adaptive strategies to live under extreme environmental conditions. Hypothetically, extremophyte species present a unique configuration of physiological functions that prioritize stress-tolerance mechanisms while carefully managing resource allocation for photosynthesis. This could be particularly challenging under a multi-stress environment, where the synthesis of multiple and sequential molecular mechanisms is induced. We explored this hypothesis in three phylogenetically related woody species co-occurring in the Atacama Desert, Strombocarpa tamarugo, Neltuma alba, and Neltuma chilensis, by analyzing their leaf dehydration and freezing tolerance and by characterizing their photosynthetic performance under natural growth conditions. Besides, the transcriptomic profiling, biochemical analyses of leaf pigments, and metabolite analysis by untargeted metabolomics were conducted to study gene expression and metabolomic landscape within this challenging multi-stress environment. S. tamarugo showed a higher photosynthetic capacity and leaf stress tolerance than the other species. In this species, a multifactorial response was observed, which involves high photochemical activity associated with a higher content of chlorophylls and β-carotene. The oxidative damage of the photosynthetic apparatus is probably attenuated by the synthesis of complex antioxidant molecules in the three species, but S. tamarugo showed the highest antioxidant capacity. Comparative transcriptomic and metabolomic analyses among the species showed the differential expression of genes involved in the biosynthetic pathways of key stress-related metabolites. Moreover, the synthesis of non-nitrogen osmoprotectant molecules, such as ciceritol and mannitol in S. tamarugo, would allow the nitrogen allocation to support its high photosynthetic capacity without compromising leaf dehydration tolerance and freezing stress avoidance.

Ice in the intertidal: patterns and processes of freeze tolerance in intertidal invertebrates.

Many intertidal invertebrates are freeze tolerant, meaning that they can survive ice formation within their body cavity. Freeze tolerance is a fascinating trait, and understanding its mechanisms is important for predicting the survival of intertidal animals during extreme cold weather events. In this Review, we bring together current research on the ecology, biochemistry and physiology of this group of freeze-tolerant organisms. We first introduce the ecology of the intertidal zone, then highlight the strong geographic and taxonomic biases within the current body of literature on this topic. Next, we detail current knowledge on the mechanisms of freeze tolerance used by intertidal invertebrates. Although the mechanisms of freeze tolerance in terrestrial arthropods have been well-explored, marine invertebrate freeze tolerance is less well understood and does not appear to work similarly because of the osmotic differences that come with living in seawater. Freeze tolerance mechanisms thought to be utilized by intertidal invertebrates include: (1) low molecular weight cryoprotectants, such as compatible osmolytes and anaerobic by-products; (2) high molecular weight cryoprotectants, such as ice-binding proteins; as well as (3) other molecular mechanisms involving heat shock proteins and aquaporins. Lastly, we describe untested hypotheses, methods and approaches that researchers can use to fill current knowledge gaps. Understanding the mechanisms and consequences of freeze tolerance in the intertidal zone has many important ecological implications, but also provides an opportunity to broaden our understanding of the mechanisms of freeze tolerance more generally.

High interspecific variability in ice nucleation activity suggests pollen ice nucleators are incidental

Abstract. Ice-nucleating macromolecules (INMs) produced by plant pollen can nucleate ice at warm temperatures and may play an important role in weather- and climate-relevant cloud glaciation. INMs have also proved useful for mammalian cell and tissue model cryopreservation. The high ice nucleation (IN) activity of some INMs indicates an underlying biological function, either freezing tolerance or bioprecipitation-mediated dispersal. Here, using the largest study of pollen ice nucleation to date, we show that phylogenetic proximity, spermatophyte subdivision, primary growth biome, pollination season, primary pollination method, desiccation tolerance and native growth elevation do not account for the IN activity of INMs released from different plant species' pollen. The results suggest that these macromolecules are produced by plants for a purpose unrelated to ice nucleation and have an incidental ability to nucleate ice. This ability may have been adapted by some species for specific biological purposes, producing exceptional ice nucleators. Pollen INMs may be more active, widespread in nature, and diverse than previously thought.

Reversible Histone Modifications Contribute to the Frozen and Thawed Recovery States of Wood Frog Brains.

Epigenetic regulation, notably histone post-translational modification (PTM), has emerged as a major transcriptional control of gene expression during cellular stress adaptation. In the present study, we use an acid extraction method to isolate total histone protein and investigate dynamic changes in 23 well-characterized histone methylations/acetylations in the brains of wood frogs subject to 24-h freezing and subsequent 8-h thawed recovery conditions. Our results identify four histone PTMs (H2BK5ac, H3K14ac, H3K4me3, H3K9me2) and three histone proteins (H1.0, H2B, H4) that were significantly (p < 0.05) responsive to freeze-thaw in freeze-tolerant R. sylvatica brains. Two other permissive modifications (H3R8me2a, H3K9ac) also trended downwards following freezing stress. Together, these data are strongly supportive of the proposed global transcriptional states of hypometabolic freeze tolerance and rebounded thawed recovery. Our findings shed light on the intricate interplay between epigenetic regulation, gene transcription and energy metabolism in wood frogs' adaptive response to freezing stress.

Proteomic analysis of Rana sylvatica reveals differentially expressed proteins in liver in response to anoxia, dehydration or freezing stress

Ectothermic animals that live in seasonally cold regions must adapt to seasonal variation and specific environmental conditions. During the winter, some amphibians hibernate on land and encounter limited environmental water, deficient oxygen, and extremely low temperatures that can cause the whole body freezing. These stresses trigger physiological and biochemical adaptations in amphibians that allow them to survive. Rana sylvatica, commonly known as the wood frog, shows excellent freeze tolerance. They can slow their metabolic activity to a near halt and endure freezing of 65–70% of their total body water as extracellular ice during hibernation, returning to normal when the temperatures rise again. To investigate the molecular adaptations of freeze-tolerant wood frogs, a comprehensive proteomic analysis was performed on frog liver tissue after anoxia, dehydration, or freezing exposures using a label-free LC–MS/MS proteomic approach. Quantitative proteomic analysis revealed that 87, 118, and 86 proteins were significantly upregulated in dehydrated, anoxic, and frozen groups, suggesting potential protective functions. The presence of three upregulated enzymes, glutathione S-transferase (GST), aldolase (ALDOA), and sorbitol dehydrogenase (SORD), was also validated. For all enzymes, the specific enzymatic activity was significantly higher in the livers of frozen and anoxic groups than in the controls. This study reveals that GST, ALDOA, and SORD might participate in the freeze tolerance mechanism by contributing to regulating cellular detoxification and energy metabolism.

ATBS1-INTERACTING FACTOR 2 Positively Regulates Freezing Tolerance via INDUCER OF CBF EXPRESSION 1/C-REPEAT BINDING FACTOR-Induced Cold Acclimation Pathway.

The INDUCER OF CBF EXPRESSION 1/C-REPEAT BINDING FACTOR (ICE1/CBF) pathway plays a crucial role in plant responses to cold stress, impacting growth and development. Here, we demonstrated that ATBS1-INTERACTING FACTOR 2 (AIF2), a non-DNA-binding basic helix-loop-helix transcription factor, positively regulates freezing tolerance through the ICE1/CBF-induced cold tolerance pathway in Arabidopsis. Cold stress transcriptionally upregulated AIF2 expression and induced AIF2 phosphorylation, thereby stabilizing the AIF2 protein during early stages of cold acclimation. The AIF2 loss-of-function mutant, aif2-1, exhibited heightened sensitivity to freezing before and after cold acclimation. In contrast, ectopic expression of AIF2, but not the C-terminal-deleted AIF2 variant, restored freezing tolerance. AIF2 enhanced ICE1 stability during cold acclimation and promoted the transcriptional expression of CBFs and downstream cold-responsive genes, ultimately enhancing plant tolerance to freezing stress. MITOGEN-ACTIVATED PROTEIN KINASES 3 and 6 (MPK3/6), known negative regulators of freezing tolerance, interacted with and phosphorylated AIF2, subjecting it to protein degradation. Furthermore, transient co-expression of MPK3/6 with AIF2 and ICE1 downregulated AIF2/ICE1-induced transactivation of CBF2 expression. AIF2 interacted preferentially with BRASSINOSTEROID-INSENSITIVE 2 (BIN2) and MPK3/6 during the early and later stages of cold acclimation, respectively, thereby differentially regulating AIF2 activity in a cold acclimation time-dependent manner. Moreover, AIF2 acted additively in a gain-of-function mutant of BRASSINAZOLE-RESISTANT 1 (BZR1; bzr1-1D) and a triple knockout mutant of BIN2 and its homologs (bin2bil1bil2) to induce CBFs-mediated freezing tolerance. This suggests that cold-induced AIF2 coordinates freezing tolerance along with BZR1 and BIN2, key positive and negative components, respectively, of brassinosteroid signaling pathways.

HOS15-mediated turnover of PRR7 enhances freezing tolerance.

Arabidopsis PSEUDO RESPONSE REGULATOR7 (PRR7) is a core component of the circadian oscillator which also plays a crucial role in freezing tolerance. PRR7 undergoes proteasome-dependent degradation to discretely phase maximal expression in early evening. While its transcriptional repressive activity on downstream genes is integral to cold regulation, the mechanism of the conditional regulation of the PRR7 protein activity is unknown. We used double mutant analysis, protein interaction and ubiquitylation assays to establish that the ubiquitin ligase adaptor, HIGH EXPRESSION OF OSMOTICALLY RESPONSIVE GENE 15 (HOS15), controls the protein accumulation pattern of PRR7 through direct protein-protein interactions. Freezing tolerance and electrolyte leakage assays show that PRR7 enhances cold temperature sensitivity, supported by ChIP-qPCR at C-REPEAT BINDING FACTOR (CBF) and COLD REGULATED 15A (COR15A) promoters where PRR7 levels were higher in hos15 mutants. We establish that HOS15 mediates PRR7 protein turnover through enhanced ubiquitylation at low temperature in the dark. Under the same conditions, increased PRR7 association with the promoter regions of CBFs and COR15A in hos15 correlates with decreased CBF1 and COR15A transcription and enhanced freezing sensitivity. We propose a novel mechanism whereby HOS15-mediated regulation of PRR7 provides an intersection between the circadian system and other cold acclimation pathways leading to freezing tolerance through upregulation of CBF1 and COR15A.

Historical and contemporary climate jointly determine angiosperm plant diversity patterns across east Eurasia

Mechanisms underlying large‐scale spatial patterns of species richness are one of the central issues in ecology. Although contemporary climate, evolutionary history, and historical climate change have been proposed as drivers of species richness patterns, variation in the relative importance of different factors remains a major challenge. Here, using newly compiled distribution data with a spatial resolution of 100 × 100 km for 43 023 angiosperms plant species in east Eurasia, we mapped species richness patterns for plants with different growth forms (i.e. woody versus herbaceous) and range sizes (i.e. wide‐ranged versus narrow‐ranged species), and compared the relative importance of the four hypotheses in explaining these patterns, i.e. freezing tolerance hypothesis, historical climate change hypothesis, Janzen's hypothesis (predicting that climate seasonality and topography determine species richness patterns), and diversification rate hypothesis. We found that species richness of all angiosperm plants presented a clear latitudinal gradient and was highest in southwestern China and Central Asian mountains. Notably, species richness patterns and their dominant drivers differed between species groups. Historical climate change was the dominant driver for richness patterns of all and herbaceous species. The freezing tolerance hypothesis dominated the drivers for all woody species, while Janzen's hypothesis dominated narrow‐ranged woody and herbaceous species. Our study suggests that different hypotheses contribute to large‐scale plant richness patterns via their effects on different plant groups. Our results did not support the diversification rate hypothesis, but demonstrated the high importance of historical climate change to plant diversity in east Eurasia, providing new perspectives on the mechanisms of plant diversity patterns in this continent.

Autophagy is suppressed by low temperatures and is dispensable for cold acclimation in Arabidopsis.

Plants have evolved various mechanisms to adapt to the ever-changing external environment. Autophagy is one such mechanism and has been suggested to play a key role in responding to and adapting to abiotic stresses in plants. However, the role of autophagy in adaptation to cold and freezing stresses remains to be characterized in detail. Here, we investigated the role of autophagy in the low-temperature response of Arabidopsis using atg mutants. Both the atg5-1 and atg10-1 mutants exhibited normal freezing tolerance, regardless of cold acclimation. A comparison of fresh weights indicated that the difference in growth between the wild-type and atg plants under cold conditions was rather small compared with that under normal conditions. Analysis of COLD-REGULATED gene expression showed no significant differences between the atg mutants and wild type. Treatment with 3-methyladenine, an inhibitor of autophagy, did not impair the induction of COR15Apro::LUC expression upon exposure to low temperature. Evaluation of autophagic activity using transgenic plants expressing RBCS-mRFP demonstrated that autophagy was rarely induced by cold exposure, even in the dark. Taken together, these data suggest that autophagy is suppressed by low temperatures and is dispensable for cold acclimation and freezing tolerance in Arabidopsis.

Single-nucleotide polymorphisms and copy number variations drive adaptive evolution to freezing stress in a subtropical evergreen broad-leaved tree: Hexaploid wild Camellia oleifera

Subtropical evergreen broad-leaved trees are usually vulnerable to freezing stress, while hexaploid wild Camellia oleifera shows strong freezing tolerance. As a valuable genetic resource of woody oil crop C. oleifera, wild C. oleifera can serve as a case for studying the molecular bases of adaptive evolution to freezing stress. Here, 47 wild C. oleifera from 11 natural distribution sites in China and 4 relative species of C. oleifera were selected for genome sequencing. “Min Temperature of Coldest Month” (BIO6) had the highest comprehensive contribution to wild C. oleifera distribution. The population genetic structure of wild C. oleifera could be divided into two groups: in cold winter (BIO6 ≤ 0 °C) and warm winter (BIO6 > 0 °C) areas. Wild C. oleifera in cold winter areas might have experienced stronger selection pressures and population bottlenecks with lower Ne than those in warm winter areas. 155 single-nucleotide polymorphisms (SNPs) were significantly correlated with the key bioclimatic variables (106 SNPs significantly correlated with BIO6). Twenty key SNPs and 15 key copy number variation regions (CNVRs) were found with genotype differentiation > 50% between the two groups of wild C. oleifera. Key SNPs in cis-regulatory elements might affect the expression of key genes associated with freezing tolerance, and they were also found within a CNVR suggesting interactions between them. Some key CNVRs in the exon regions were closely related to the differentially expressed genes under freezing stress. The findings suggest that rich SNPs and CNVRs in polyploid trees may contribute to the adaptive evolution to freezing stress.

Effects of osmoprotective compounds on freezing tolerance in chestnut (Castanea sativa) leaves

Effects of osmoprotective compounds on freezing tolerance in chestnut (<i>Castanea sativa</i>) leaves

Evolution of PIN gene family between monocotyledons and dicotyledons and VvPIN1 negatively regulates freezing tolerance in transgenic Arabidopsis.

The PIN-FORMED (PIN) proteins mediate the auxin flow throughout the plant and have been identified in many species. However, evolution differences in the PIN gene families have not been systematically analyzed, and their functions under abiotic stresses in grape are largely unexplored. In this study, 373 PIN genes were identified from 25 species and divided into 3 subgroups. Physicochemical properties analysis indicated that most of the PIN proteins were unstable alkaline hydrophobic proteins in nature. The synteny analysis showed that the PINs contained strong gene duplication. Motif composition revealed that PIN gene sequence differences between monocotyledons and dicotyledons were due to evolutionary-induced base loss, and the loss was more common in dicotyledonous. Meanwhile, the codon usage bias showed that the PINs showed stronger codon preference in monocotyledons, monocotyledons biased towards C3s and G3s, and dicotyledons biased towards A3s and T3s. In addition, the VvPIN1 can interact with VvCSN5. Significantly, under freezing treatment, the ion leakage, , H2O2, and malondialdehyde (MDA) were obviously increased, while the proline (Pro) content, peroxidase (POD) activity, and glutathione (GSH) content were decreased in VvPIN1-overexpressing Arabidopsis compared to the wild type (WT). And quantitative real-time PCR (qRT-PCR) showed that AtICE1, AtICE2, AtCBF1, AtCBF2, and AtCBF3 were down-regulated in overexpression lines. These results demonstrated that VvPIN1 negatively regulated the freezing tolerance in transgenic Arabidopsis. Collectively, this study provides a novel insight into the evolution and a basis for further studies on the biological functions of PIN genes in monocotyledons and dicotyledons.