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Sample records for arabidopsis freezing tolerance

  1. Natural Genetic Variation of Freezing Tolerance in Arabidopsis[W][OA

    PubMed Central

    Hannah, Matthew A.; Wiese, Dana; Freund, Susanne; Fiehn, Oliver; Heyer, Arnd G.; Hincha, Dirk K.

    2006-01-01

    Low temperature is a primary determinant of plant growth and survival. Using accessions of Arabidopsis (Arabidopsis thaliana) originating from Scandinavia to the Cape Verde Islands, we show that freezing tolerance of natural accessions correlates with habitat winter temperatures, identifying low temperature as an important selective pressure for Arabidopsis. Combined metabolite and transcript profiling show that during cold exposure, global changes of transcripts, but not of metabolites, correlate with the ability of Arabidopsis to cold acclimate. There are, however, metabolites and transcripts, including several transcription factors, that correlate with freezing tolerance, indicating regulatory pathways that may be of primary importance for this trait. These data identify that enhanced freezing tolerance is associated with the down-regulation of photosynthesis and hormonal responses and the induction of flavonoid metabolism, provide evidence for naturally increased nonacclimated freezing tolerance due to the constitutive activation of the C-repeat binding factors pathway, and identify candidate transcriptional regulators that correlate with freezing tolerance. PMID:16844837

  2. Comparison of freezing tolerance, compatible solutes and polyamines in geographically diverse collections of Thellungiella sp. and Arabidopsis thaliana accessions

    PubMed Central

    2012-01-01

    Background Thellungiella has been proposed as an extremophile alternative to Arabidopsis to investigate environmental stress tolerance. However, Arabidopsis accessions show large natural variation in their freezing tolerance and here the tolerance ranges of collections of accessions in the two species were compared. Results Leaf freezing tolerance of 16 Thellungiella accessions was assessed with an electrolyte leakage assay before and after 14 days of cold acclimation at 4°C. Soluble sugars (glucose, fructose, sucrose, raffinose) and free polyamines (putrescine, spermidine, spermine) were quantified by HPLC, proline photometrically. The ranges in nonacclimated freezing tolerance completely overlapped between Arabidopsis and Thellungiella. After cold acclimation, some Thellungiella accessions were more freezing tolerant than any Arabidopsis accessions. Acclimated freezing tolerance was correlated with sucrose levels in both species, but raffinose accumulation was lower in Thellungiella and only correlated with freezing tolerance in Arabidopsis. The reverse was true for leaf proline contents. Polyamine levels were generally similar between the species. Only spermine content was higher in nonacclimated Thellungiella plants, but decreased during acclimation and was negatively correlated with freezing tolerance. Conclusion Thellungiella is not an extremophile with regard to freezing tolerance, but some accessions significantly expand the range present in Arabidopsis. The metabolite data indicate different metabolic adaptation strategies between the species. PMID:22863402

  3. Transcriptional and metabolomic analysis of Ascophyllum nodosum mediated freezing tolerance in Arabidopsis thaliana

    PubMed Central

    2012-01-01

    Background We have previously shown that lipophilic components (LPC) of the brown seaweed Ascophyllum nodosum (ANE) improved freezing tolerance in Arabidopsis thaliana. However, the mechanism(s) of this induced freezing stress tolerance is largely unknown. Here, we investigated LPC induced changes in the transcriptome and metabolome of A. thaliana undergoing freezing stress. Results Gene expression studies revealed that the accumulation of proline was mediated by an increase in the expression of the proline synthesis genes P5CS1 and P5CS2 and a marginal reduction in the expression of the proline dehydrogenase (ProDH) gene. Moreover, LPC application significantly increased the concentration of total soluble sugars in the cytosol in response to freezing stress. Arabidopsis sfr4 mutant plants, defective in the accumulation of free sugars, treated with LPC, exhibited freezing sensitivity similar to that of untreated controls. The 1H NMR metabolite profile of LPC-treated Arabidopsis plants exposed to freezing stress revealed a spectrum dominated by chemical shifts (δ) representing soluble sugars, sugar alcohols, organic acids and lipophilic components like fatty acids, as compared to control plants. Additionally, 2D NMR spectra suggested an increase in the degree of unsaturation of fatty acids in LPC treated plants under freezing stress. These results were supported by global transcriptome analysis. Transcriptome analysis revealed that LPC treatment altered the expression of 1113 genes (5%) in comparison with untreated plants. A total of 463 genes (2%) were up regulated while 650 genes (3%) were down regulated. Conclusion Taken together, the results of the experiments presented in this paper provide evidence to support LPC mediated freezing tolerance enhancement through a combination of the priming of plants for the increased accumulation of osmoprotectants and alteration of cellular fatty acid composition. PMID:23171218

  4. Flavonoids are determinants of freezing tolerance and cold acclimation in Arabidopsis thaliana

    PubMed Central

    Schulz, Elisa; Tohge, Takayuki; Zuther, Ellen; Fernie, Alisdair R.; Hincha, Dirk K.

    2016-01-01

    In plants from temperate climates such as Arabidopsis thaliana low, non-freezing temperatures lead to increased freezing tolerance in a process termed cold acclimation. This process is accompanied by massive changes in gene expression and in the content of primary metabolites and lipids. In addition, most flavonols and anthocyanins accumulate upon cold exposure, along with most transcripts encoding transcription factors and enzymes of the flavonoid biosynthetic pathway. However, no evidence for a functional role of flavonoids in plant freezing tolerance has been shown. Here, we present a comprehensive analysis using qRT-PCR for transcript, LC-MS for flavonoid and GC-MS for primary metabolite measurements, and an electrolyte leakage assay to determine freezing tolerance of 20 mutant lines in two Arabidopsis accessions that are affected in different steps of the flavonoid biosynthetic pathway. This analysis provides evidence for a functional role of flavonoids in plant cold acclimation. The accumulation of flavonoids in the activation tagging mutant line pap1-D improved, while reduced flavonoid content in different knock-out mutants impaired leaf freezing tolerance. Analysis of the different knock-out mutants suggests redundancy of flavonoid structures, as the lack of flavonols or anthocyanins could be compensated by other compound classes. PMID:27658445

  5. Acquisition of freezing tolerance in Arabidopsis and two contrasting ecotypes of the extremophile Eutrema salsugineum (Thellungiella salsuginea).

    PubMed

    Khanal, Nityananda; Moffatt, Barbara A; Gray, Gordon R

    2015-05-15

    Eutrema salsugineum (Thellungiella salsuginea) is an extremophile, a close relative of Arabidopsis, but possessing much higher constitutive levels of tolerance to abiotic stress. This study aimed to characterize the freezing tolerance of Arabidopsis (Columbia ecotype) and two ecotypes of Eutrema (Yukon and Shandong) isolated from contrasting geographical locations. Under our growth conditions, maximal freezing tolerance was observed after two- and three-weeks of cold acclimation for Arabidopsis and Eutrema, respectively. The ecotypes of Eutrema and Arabidopsis do not differ in their constitutive level of freezing tolerance or short-term cold acclimation capacity. However Eutrema remarkably outperforms Arabidopsis in long-term acclimation capacity suggesting a wider phenotypic plasticity for the trait of freezing tolerance. The combination of drought treatment and one-week of cold acclimation was more effective than long-term cold acclimation in achieving maximum levels of freezing tolerance in Eutrema, but not Arabidopsis. Furthermore, it was demonstrated growth conditions, particularly irradiance, are determinates of the level of freezing tolerance attained during cold acclimation suggesting a role for photosynthetic processes in adaptive stress responses. Copyright © 2015 Elsevier GmbH. All rights reserved.

  6. Heterosis in the freezing tolerance, and sugar and flavonoid contents of crosses between Arabidopsis thaliana accessions of widely varying freezing tolerance

    PubMed Central

    KORN, MARINA; PETEREK, SILKE; MOCK, HANS-PETER; HEYER, ARND G; HINCHA, DIRK K

    2008-01-01

    Heterosis is defined as the increased vigour of hybrids in comparison to their parents. We investigated 24 F1 hybrid lines of Arabidopsis thaliana generated by reciprocally crossing either C24 or Col with six other parental accessions (Can, Co, Cvi, Ler, Rsch, Te) that differ widely in their freezing tolerance. The crosses differed in the degree of heterosis for freezing tolerance, both in the non-acclimated state and after a 14 d cold acclimation period. Crosses with C24 showed more heterosis than crosses with Col, and heterosis was stronger in acclimated than in non-acclimated plants. Leaf content of soluble sugars and proline showed more deviation from mid-parent values in crosses involving C24 than in those involving Col, and deviations were larger in acclimated than in non-acclimated plants. There were significant correlations between the content of different sugars and leaf freezing tolerance, as well as between heterosis effects in freezing tolerance and sugar content. Flavonoid content and composition varied between accessions, and between non-acclimated and acclimated plants. In the crosses, large deviations from the mid-parent values in the contents of different flavonols occurred, and there were strikingly strong correlations between both flavonol content and freezing tolerance, and between heterosis effects in freezing tolerance and flavonol content. PMID:18284584

  7. ICE1: a regulator of cold-induced transcriptome and freezing tolerance in Arabidopsis

    PubMed Central

    Chinnusamy, Viswanathan; Ohta, Masaru; Kanrar, Siddhartha; Lee, Byeong-ha; Hong, Xuhui; Agarwal, Manu; Zhu, Jian-Kang

    2003-01-01

    Cold temperatures trigger the expression of the CBF family of transcription factors, which in turn activate many downstream genes that confer chilling and freezing tolerance to plants. We report here the identification of ICE1 (inducer of CBF expression 1), an upstream transcription factor that regulates the transcription of CBF genes in the cold. An Arabidopsis ice1 mutant was isolated in a screen for mutations that impair cold-induced transcription of a CBF3 promoter-luciferase reporter gene. The ice1 mutation blocks the expression of CBF3 and decreases the expression of many genes downstream of CBFs, which leads to a significant reduction in plant chilling and freezing tolerance. ICE1 encodes a MYC-like bHLH transcriptional activator. ICE1 binds specifically to the MYC recognition sequences in the CBF3 promoter. ICE1 is expressed constitutively, and its overexpression in wild-type plants enhances the expression of the CBF regulon in the cold and improves freezing tolerance of the transgenic plants. PMID:12672693

  8. Cold shock domain protein 3 regulates freezing tolerance in Arabidopsis thaliana.

    PubMed

    Kim, Myung-Hee; Sasaki, Kentaro; Imai, Ryozo

    2009-08-28

    In response to cold, Escherichia coli produces cold shock proteins (CSPs) that have essential roles in cold adaptation as RNA chaperones. Here, we demonstrate that Arabidopsis cold shock domain protein 3 (AtCSP3), which shares a cold shock domain with bacterial CSPs, is involved in the acquisition of freezing tolerance in plants. AtCSP3 complemented a cold-sensitive phenotype of the E. coli CSP quadruple mutant and displayed nucleic acid duplex melting activity, suggesting that AtCSP3 also functions as an RNA chaperone. Promoter-GUS transgenic plants revealed tissue-specific expression of AtCSP3 in shoot and root apical regions. When exposed to low temperature, GUS activity was extensively induced in a broader region of the roots. In transgenic plants expressing an AtCSP3-GFP fusion, GFP signals were detected in both the nucleus and cytoplasm. An AtCSP3 knock-out mutant (atcsp3-2) was sensitive to freezing compared with wild-type plants under non-acclimated and cold-acclimated conditions, whereas expression of C-repeat-binding factors and their downstream genes during cold acclimation was not altered in the atcsp3-2 mutant. Overexpression of AtCSP3 in transgenic plants conferred enhanced freezing tolerance over wild-type plants. Together, the data demonstrated an essential role of RNA chaperones for cold adaptation in higher plants.

  9. Ice-binding proteins confer freezing tolerance in transgenic Arabidopsis thaliana.

    PubMed

    Bredow, Melissa; Vanderbeld, Barbara; Walker, Virginia K

    2017-01-01

    Lolium perenne is a freeze-tolerant perennial ryegrass capable of withstanding temperatures below -13 °C. Ice-binding proteins (IBPs) presumably help prevent damage associated with freezing by restricting the growth of ice crystals in the apoplast. We have investigated the expression, localization and in planta freezing protection capabilities of two L. perenne IBP isoforms, LpIRI2 and LpIRI3, as well as a processed IBP (LpAFP). One of these isoforms, LpIRI2, lacks a conventional signal peptide and was assumed to be a pseudogene. Nevertheless, both LpIRI2 and LpIRI3 transcripts were up-regulated following cold acclimation. LpIRI2 also demonstrated ice-binding activity when produced recombinantly in Escherichia coli. Both the LpIRI3 and LpIRI2 isoforms appeared to accumulate in the apoplast of transgenic Arabidopsis thaliana plants. In contrast, the fully processed isoform, LpAFP, remained intracellular. Transgenic plants expressing either LpIRI2 or LpIRI3 showed reduced ion leakage (12%-39%) after low-temperature treatments, and significantly improved freezing survival, while transgenic LpAFP-expressing lines did not confer substantial subzero protection. Freeze protection was further enhanced by with the introduction of more than one IBP isoform; ion leakage was reduced 26%-35% and 10% of plants survived temperatures as low as -8 °C. Our results demonstrate that apoplastic expression of multiple L. perenne IBP isoforms shows promise for providing protection to crops susceptible to freeze-induced damage. © 2016 The Authors. Plant Biotechnology Journal published by Society for Experimental Biology and The Association of Applied Biologists and John Wiley & Sons Ltd.

  10. Disruption of the Arabidopsis Defense Regulator Genes SAG101, EDS1, and PAD4 Confers Enhanced Freezing Tolerance

    PubMed Central

    Chen, Qin-Fang; Xu, Le; Tan, Wei-Juan; Chen, Liang; Qi, Hua; Xie, Li-Juan; Chen, Mo-Xian; Liu, Bin-Yi; Yu, Lu-Jun; Yao, Nan; Zhang, Jian-Hua; Shu, Wensheng; Xiao, Shi

    2017-01-01

    In Arabidopsis, three lipase-like regulators, SAG101, EDS1, and PAD4, act downstream of resistance protein-associated defense signaling. Although the roles of SAG101, EDS1, and PAD4 in biotic stress have been extensively studied, little is known about their functions in plant responses to abiotic stresses. Here, we show that SAG101, EDS1, and PAD4 are involved in the regulation of freezing tolerance in Arabidopsis. With or without cold acclimation, the sag101, eds1, and pad4 single mutants, as well as their double mutants exhibited similarly enhanced tolerance to freezing temperatures. Upon cold exposure, the sag101, eds1, and pad4 mutants showed increased transcript levels of C-REPEAT/DRE BINDING FACTORs and their regulons, compared with wild type. Moreover, freezing-induced cell death and accumulation of hydrogen peroxide were ameliorated in sag101, eds1, and pad4 mutants. The sag101, eds1, and pad4 mutants had much lower salicylic acid (SA) and diacylglycerol (DAG) contents than wild type and exogenous application of SA and DAG compromised the freezing tolerance of the mutants. Furthermore, SA suppressed the cold-induced expression of DGATs and DGKs in wild-type leaves. These findings indicate that SAG101, EDS1, and PAD4 are involved in freezing response in Arabidopsis, at least in part, by modulating the homeostasis of SA and DAG. PMID:26149542

  11. Disruption of the Arabidopsis Defense Regulator Genes SAG101, EDS1, and PAD4 Confers Enhanced Freezing Tolerance.

    PubMed

    Chen, Qin-Fang; Xu, Le; Tan, Wei-Juan; Chen, Liang; Qi, Hua; Xie, Li-Juan; Chen, Mo-Xian; Liu, Bin-Yi; Yu, Lu-Jun; Yao, Nan; Zhang, Jian-Hua; Shu, Wensheng; Xiao, Shi

    2015-10-05

    In Arabidopsis, three lipase-like regulators, SAG101, EDS1, and PAD4, act downstream of resistance protein-associated defense signaling. Although the roles of SAG101, EDS1, and PAD4 in biotic stress have been extensively studied, little is known about their functions in plant responses to abiotic stresses. Here, we show that SAG101, EDS1, and PAD4 are involved in the regulation of freezing tolerance in Arabidopsis. With or without cold acclimation, the sag101, eds1, and pad4 single mutants, as well as their double mutants, exhibited similarly enhanced tolerance to freezing temperatures. Upon cold exposure, the sag101, eds1, and pad4 mutants showed increased transcript levels of C-REPEAT/DRE BINDING FACTORs and their regulons compared with the wild type. Moreover, freezing-induced cell death and accumulation of hydrogen peroxide were ameliorated in sag101, eds1, and pad4 mutants. The sag101, eds1, and pad4 mutants had much lower salicylic acid (SA) and diacylglycerol (DAG) contents than the wild type, and exogenous application of SA and DAG compromised the freezing tolerance of the mutants. Furthermore, SA suppressed the cold-induced expression of DGATs and DGKs in the wild-type leaves. These findings indicate that SAG101, EDS1, and PAD4 are involved in the freezing response in Arabidopsis, at least in part, by modulating the homeostasis of SA and DAG. Copyright © 2015 The Author. Published by Elsevier Inc. All rights reserved.

  12. Identification of ICE2, a gene involved in cold acclimation which determines freezing tolerance in Arabidopsis thaliana.

    PubMed

    Fursova, Oksana V; Pogorelko, Gennady V; Tarasov, Valentin A

    2009-01-15

    Several transcription factors are presently known to regulate the response to cold stress. Here we describe a new positive regulator, ICE2, which is a transcription factor of the bHLH family that participates in the response to deep freezing through the cold acclimation-dependent pathway in Arabidopsis thaliana plants. An overexpression of ICE2 (as we named the At1g12860 locus) in transgenic Arabidopsis plants results in increased tolerance to deep freezing stress after cold acclimation. The seeds of transgenic lines that overexpressed ICE2 were characterized by decreased levels of carbohydrate and increased levels of lipids. The analysis of expression of CBF1 gene (also known as DREB1B), which have been shown to be required for the complete development of cold acclimation response in Arabidopsis indicated a difference between expression of the CBF1 gene in transgenic plants and the wild-type control plants, Col-0. These results suggested that the CBF1 transcription factor, known as one of the regulators of the cold stress response, has a dominant role in providing freezing tolerance in transgenic plants characterized by overexpression of ICE2.

  13. PpCBF3 from Cold-Tolerant Kentucky Bluegrass Involved in Freezing Tolerance Associated with Up-Regulation of Cold-Related Genes in Transgenic Arabidopsis thaliana

    PubMed Central

    Chen, Yu; Xu, Bin; Yang, Zhimin; Huang, Bingru

    2015-01-01

    Dehydration-Responsive Element Binding proteins (DREB)/C-repeat (CRT) Binding Factors (CBF) have been identified as transcriptional activators during plant responses to cold stress. The objective of this study was to determine the physiological roles of a CBF gene isolated from a cold-tolerant perennial grass species, Kentucky bluegrass (Poa pratensis L.), which designated as PpCBF3, in regulating plant tolerance to freezing stress. Transient transformation of Arabidopsis thaliana mesophyll protoplast with PpCBF3-eGFP fused protein showed that PpCBF3 was localized to the nucleus. RT-PCR analysis showed that PpCBF3 was specifically induced by cold stress (4°C) but not by drought stress [induced by 20% polyethylene glycol 6000 solution (PEG-6000)] or salt stress (150 mM NaCl). Transgenic Arabidopsis overexpressing PpCBF3 showed significant improvement in freezing (-20°C) tolerance demonstrated by a lower percentage of chlorotic leaves, lower cellular electrolyte leakage (EL) and H2O2 and O2.- content, and higher chlorophyll content and photochemical efficiency compared to the wild type. Relative mRNA expression level analysis by qRT-PCR indicated that the improved freezing tolerance of transgenic Arabidopsis plants overexpressing PpCBF3 was conferred by sustained activation of downstream cold responsive (COR) genes. Other interesting phenotypic changes in the PpCBF3-transgenic Arabidopsis plants included late flowering and slow growth or ‘dwarfism’, both of which are desirable phenotypic traits for perennial turfgrasses. Therefore, PpCBF3 has potential to be used in genetic engineering for improvement of turfgrass freezing tolerance and other desirable traits. PMID:26177510

  14. Jasmonate Regulates the INDUCER OF CBF EXPRESSION–C-REPEAT BINDING FACTOR/DRE BINDING FACTOR1 Cascade and Freezing Tolerance in Arabidopsis[W

    PubMed Central

    Hu, Yanru; Jiang, Liqun; Wang, Fang; Yu, Diqiu

    2013-01-01

    The INDUCER OF CBF EXPRESSION (ICE)–C-REPEAT BINDING FACTOR/DRE BINDING FACTOR1 (CBF/DREB1) transcriptional pathway plays a critical role in modulating cold stress responses in Arabidopsis thaliana. Dissecting crucial upstream regulatory signals or components of the ICE-CBF/DREB1 cascade will enhance our understanding of plant cold-tolerance mechanisms. Here, we show that jasmonate positively regulates plant responses to freezing stress in Arabidopsis. Exogenous application of jasmonate significantly enhanced plant freezing tolerance with or without cold acclimation. By contrast, blocking endogenous jasmonate biosynthesis and signaling rendered plants hypersensitive to freezing stress. Consistent with the positive role of jasmonate in freezing stress, production of endogenous jasmonate was triggered by cold treatment. In addition, cold induction of genes acting in the CBF/DREB1 signaling pathway was upregulated by jasmonate. Further investigation revealed that several JASMONATE ZIM-DOMAIN (JAZ) proteins, the repressors of jasmonate signaling, physically interact with ICE1 and ICE2 transcription factors. JAZ1 and JAZ4 repress the transcriptional function of ICE1, thereby attenuating the expression of its regulon. Consistent with this, overexpression of JAZ1 or JAZ4 represses freezing stress responses of Arabidopsis. Taken together, our study provides evidence that jasmonate functions as a critical upstream signal of the ICE-CBF/DREB1 pathway to positively regulate Arabidopsis freezing tolerance. PMID:23933884

  15. Overexpression of Arabidopsis NADPH-dependent thioredoxin reductase C (AtNTRC) confers freezing and cold shock tolerance to plants

    SciTech Connect

    Moon, Jeong Chan; Lee, Sangmin; Shin, Su Young; Chae, Ho Byoung; Jung, Young Jun; Jung, Hyun Suk; Lee, Kyun Oh; Lee, Jung Ro; Lee, Sang Yeol

    2015-08-07

    Overexpression of AtNTRC (AtNTRC{sup OE}) in Arabidopsis thaliana led to a freezing and cold stress tolerance, whereas a knockout mutant (atntrc) showed a stress-sensitive phenotype. Biochemical analyses showed that the recombinant AtNTRC proteins exhibited a cryoprotective activity for malate dehydrogenase and lactic dehydrogenase. Furthermore, conclusive evidence of its interaction with nucleic acids in vitro is provided here on the basis of gel shift and electron microscopy analysis. Recombinant AtNTRC efficiently protected RNA and DNA from RNase A and metal catalyzed oxidation damage, respectively. The C-terminal thioredoxin domain is required for the nucleic acid–protein complex formation. From these results, it can be hypothesized that AtNTRC, which is known to be an electron donor of peroxiredoxin, contributes the stability of macromolecules under cold stress. - Highlights: • AtNTRC has a cryoprotective activity in vitro. • Overexpression of AtNTRC increases tolerance to freezing and cold shock stresses. • Thioredoxin domain of AtNTRC protects nucleic acids in vitro. • AtNTRC inhibits protein aggregation under freezing stress in vitro.

  16. A single-repeat R3-MYB transcription factor MYBC1 negatively regulates freezing tolerance in Arabidopsis

    SciTech Connect

    Zhai, Hong; Bai, Xi; Zhu, Yanming; Li, Yong; Cai, Hua; Ji, Wei; Ji, Zuojun; Liu, Xiaofei; Liu, Xin; Li, Jing

    2010-04-16

    We had previously identified the MYBC1 gene, which encodes a single-repeat R3-MYB protein, as a putative osmotic responding gene; however, no R3-MYB transcription factor has been reported to regulate osmotic stress tolerance. Thus, we sought to elucidate the function of MYBC1 in response to osmotic stresses. Real-time RT-PCR analysis indicated that MYBC1 expression responded to cold, dehydration, salinity and exogenous ABA at the transcript level. mybc1 mutants exhibited an increased tolerance to freezing stress, whereas 35S::MYBC1 transgenic plants exhibited decreased cold tolerance. Transcript levels of some cold-responsive genes, including CBF/DREB genes, KIN1, ADC1, ADC2 and ZAT12, though, were not altered in the mybc1 mutants or the 35S::MYBC1 transgenic plants in response to cold stress, as compared to the wild type. Microarray analysis results that are publically available were investigated and found transcript level of MYBC1 was not altered by overexpression of CBF1, CBF2, and CBF3, suggesting that MYBC1 is not down regulated by these CBF family members. Together, these results suggested that MYBC1is capable of negatively regulating the freezing tolerance of Arabidopsis in the CBF-independent pathway. In transgenic Arabidopsis carrying an MYBC1 promoter driven {beta}-glucuronidase (GUS) construct, GUS activity was observed in all tissues and was relatively stronger in the vascular tissues. Fused MYBC1 and GFP protein revealed that MYBC1 was localized exclusively in the nuclear compartment.

  17. Ethylene Signaling Negatively Regulates Freezing Tolerance by Repressing Expression of CBF and Type-A ARR Genes in Arabidopsis[W][OA

    PubMed Central

    Shi, Yiting; Tian, Shouwei; Hou, Lingyan; Huang, Xiaozhen; Zhang, Xiaoyan; Guo, Hongwei; Yang, Shuhua

    2012-01-01

    The phytohormone ethylene regulates multiple aspects of plant growth and development and responses to environmental stress. However, the exact role of ethylene in freezing stress remains unclear. Here, we report that ethylene negatively regulates plant responses to freezing stress in Arabidopsis thaliana. Freezing tolerance was decreased in ethylene overproducer1 and by the application of the ethylene precursor 1-aminocyclopropane-1-carboxylic acid but increased by the addition of the ethylene biosynthesis inhibitor aminoethoxyvinyl glycine or the perception antagonist Ag+. Furthermore, ethylene-insensitive mutants, including etr1-1, ein4-1, ein2-5, ein3-1, and ein3 eil1, displayed enhanced freezing tolerance. By contrast, the constitutive ethylene response mutant ctr1-1 and EIN3-overexpressing plants exhibited reduced freezing tolerance. Genetic and biochemical analyses revealed that EIN3 negatively regulates the expression of CBFs and type-A Arabidopsis response regulator5 (ARR5), ARR7, and ARR15 by binding to specific elements in their promoters. Overexpression of these ARR genes enhanced the freezing tolerance of plants. Thus, our study demonstrates that ethylene negatively regulates cold signaling at least partially through the direct transcriptional control of cold-regulated CBFs and type-A ARR genes by EIN3. Our study also provides evidence that type-A ARRs function as key nodes to integrate ethylene and cytokinin signaling in regulation of plant responses to environmental stress. PMID:22706288

  18. Putrescine accumulation in Arabidopsis thaliana transgenic lines enhances tolerance to dehydration and freezing stress.

    PubMed

    Alet, Analía I; Sanchez, Diego H; Cuevas, Juan C; Del Valle, Secundino; Altabella, Teresa; Tiburcio, Antonio F; Marco, Francisco; Ferrando, Alejandro; Espasandín, Fabiana D; González, María E; Ruiz, Oscar A; Carrasco, Pedro

    2011-02-01

    Polyamines have been globally associated to plant responses to abiotic stress. Particularly, putrescine has been related to a better response to cold and dehydration stresses. It is known that this polyamine is involved in cold tolerance, since Arabidopsis thaliana plants mutated in the key enzyme responsible for putrescine synthesis (arginine decarboxilase, ADC; EC 4.1.1.19) are more sensitive than the wild type to this stress. Although it is speculated that the over-expression of ADC genes may confer tolerance, this is hampered by pleiotropic effects arising from the constitutive expression of enzymes from the polyamine metabolism. Here, we present our work using A. thaliana transgenic plants harboring the ADC gene from oat under the control of a stress-inducible promoter (pRD29A) instead of a constitutive promoter. The transgenic lines presented in this work were more resistant to both cold and dehydration stresses, associated with a concomitant increment in endogenous putrescine levels under stress. Furthermore, the increment in putrescine upon cold treatment correlated with the induction of known stress-responsive genes, and suggested that putrescine may be directly or indirectly involved in ABA metabolism and gene expression.

  19. Putrescine accumulation in Arabidopsis thaliana transgenic lines enhances tolerance to dehydration and freezing stress

    PubMed Central

    Alet, Analía I; Sanchez, Diego H; Cuevas, Juan C; del Valle, Secundino; Altabella, Teresa; Tiburcio, Antonio F; Marco, Francisco; Ferrando, Alejandro; Espasandín, Fabiana D; González, María E; Carrasco, Pedro

    2011-01-01

    Polyamines have been globally associated to plant responses to abiotic stress. Particularly, putrescine has been related to a better response to cold and dehydration stresses. It is known that this polyamine is involved in cold tolerance, since Arabidopsis thaliana plants mutated in the key enzyme responsible for putrescine synthesis (arginine decarboxilase, ADC; EC 4.1.1.19) are more sensitive than the wild type to this stress. Although it is speculated that the overexpression of ADC genes may confer tolerance, this is hampered by pleiotropic effects arising from the constitutive expression of enzymes from the polyamine metabolism. Here, we present our work using A. thaliana transgenic plants harboring the ADC gene from oat under the control of a stress-inducible promoter (pRD29A) instead of a constitutive promoter. The transgenic lines presented in this work were more resistant to both cold and dehydration stresses, associated with a concomitant increment in endogenous putrescine levels under stress. Furthermore, the increment in putrescine upon cold treatment correlates with the induction of known stress-responsive genes, and suggests that putrescine may be directly or indirectly involved in ABA metabolism and gene expression. PMID:21330789

  20. A C-repeat binding factor transcriptional activator (CBF/DREB1) from European bilberry (Vaccinium myrtillus) induces freezing tolerance when expressed in Arabidopsis thaliana.

    PubMed

    Oakenfull, Rachael J; Baxter, Robert; Knight, Marc R

    2013-01-01

    Freezing stress affects all plants from temperate zones to the poles. Global climate change means such freezing events are becoming less predictable. This in turn reduces the ability of plants to predict the approaching low temperatures and cold acclimate. This has consequences for crop yields and distribution of wild plant species. C-repeat binding factors (CBFs) are transcription factors previously shown to play a vital role in the acclimation process of Arabidopsis thaliana, controlling the expression of hundreds of genes whose products are necessary for freezing tolerance. Work in other plant species cements CBFs as key determinants in the trait of freezing tolerance in higher plants. To test the function of CBFs from highly freezing tolerant plants species we cloned and sequenced CBF transcription factors from three Vaccinium species (Vaccinium myrtillus, Vaccinium uliginosum and Vaccinium vitis-idaea) which we collected in the Arctic. We tested the activity of CBF transcription factors from the three Vaccinium species by producing transgenic Arabidopsis lines overexpressing them. Only the Vaccinium myrtillus CBF was able to substantially activate COR (CBF-target) gene expression in the absence of cold. Correspondingly, only the lines expressing the Vaccinium myrtillus CBF were constitutively freezing tolerant. The basis for the differences in potency of the three Vaccinium CBFs was tested by observing cellular localisation and protein levels. All three CBFs were correctly targeted to the nucleus, but Vaccinium uliginosum CBF appeared to be relatively unstable. The reasons for lack of potency for Vaccinium vitis-idaea CBF were not due to stability or targeting, and we speculate that this was due to altered transcription factor function.

  1. The Arabidopsis RCC1 Family Protein TCF1 Regulates Freezing Tolerance and Cold Acclimation through Modulating Lignin Biosynthesis

    PubMed Central

    Jenkins, Gareth I.; Wang, Shuangfeng; Shang, Zhonglin; Shi, Yiting; Yang, Shuhua; Li, Xia

    2015-01-01

    Abstract Cell water permeability and cell wall properties are critical to survival of plant cells during freezing, however the underlying molecular mechanisms remain elusive. Here, we report that a specifically cold-induced nuclear protein, Tolerant to Chilling and Freezing 1 (TCF1), interacts with histones H3 and H4 and associates with chromatin containing a target gene, BLUE-COPPER-BINDING PROTEIN (BCB), encoding a glycosylphosphatidylinositol-anchored protein that regulates lignin biosynthesis. Loss of TCF1 function leads to reduced BCB transcription through affecting H3K4me2 and H3K27me3 levels within the BCB gene, resulting in reduced lignin content and enhanced freezing tolerance. Furthermore, plants with knocked-down BCB expression (amiRNA-BCB) under cold acclimation had reduced lignin accumulation and increased freezing tolerance. The pal1pal2 double mutant (lignin content reduced by 30% compared with WT) also showed the freezing tolerant phenotype, and TCF1 and BCB act upstream of PALs to regulate lignin content. In addition, TCF1 acts independently of the CBF (C-repeat binding factor) pathway. Our findings delineate a novel molecular pathway linking the TCF1-mediated cold-specific transcriptional program to lignin biosynthesis, thus achieving cell wall remodeling with increased freezing tolerance. PMID:26393916

  2. Responses of antioxidant enzymes to cold and high light are not correlated to freezing tolerance in natural accessions of Arabidopsis thaliana.

    PubMed

    Distelbarth, H; Nägele, T; Heyer, A G

    2013-11-01

    Low temperatures and high light cause imbalances in primary and secondary reactions of photosynthesis, and thus can result in oxidative stress. Plants employ a range of low-molecular weight antioxidants and antioxidant enzymes to prevent oxidative damage, and antioxidant defence is considered an important component of stress tolerance. To figure out whether oxidative stress and antioxidant defence are key factors defining the different cold acclimation capacities of natural accessions of the model plant Arabidopsis thaliana, we investigated hydrogen peroxide (H2 O2 ) production, antioxidant enzyme activity and lipid peroxidation during a time course of cold treatment and exposure to high light in four differentially cold-tolerant natural accessions of Arabidopsis (C24, Nd, Rsch, Te) that span the European distribution range of the species. All accessions except Rsch (from Russia) had elevated H2 O2 in the cold, indicating that production of reactive oxygen species is part of the cold response in Arabidopsis. Glutathione reductase activity increased in all but Rsch, while ascorbate peroxidase and superoxide dismutase were unchanged and catalase decreased in all but Rsch. Under high light, the Scandinavian accession Te had elevated levels of H2 O2 . Te appeared most sensitive to oxidative stress, having higher malondialdehyde (MDA) levels in the cold and under high light, while only high light caused elevated MDA in the other accessions. Although the most freezing-tolerant, Te had the highest sensitivity to oxidative stress. No correlation was found between freezing tolerance and activity of antioxidant enzymes in the four accessions investigated, arguing against a key role for antioxidant defence in the differential cold acclimation capacities of Arabidopsis accessions. © 2013 German Botanical Society and The Royal Botanical Society of the Netherlands.

  3. The GI-CDF module of Arabidopsis affects freezing tolerance and growth as well as flowering.

    PubMed

    Fornara, Fabio; de Montaigu, Amaury; Sánchez-Villarreal, Alfredo; Takahashi, Yasuyuki; Ver Loren van Themaat, Emiel; Huettel, Bruno; Davis, Seth J; Coupland, George

    2015-03-01

    Plants monitor and integrate temperature, photoperiod and light quality signals to respond to continuous changes in their environment. The GIGANTEA (GI) protein is central in diverse signaling pathways, including photoperiodic, sugar and light signaling pathways, stress responses and circadian clock regulation. Previously, GI was shown to activate expression of the key floral regulators CONSTANS (CO) and FLOWERING LOCUS T (FT) by facilitating degradation of a family of CYCLING DOF FACTOR (CDF) transcriptional repressors. However, whether CDFs are implicated in other processes affected by GI remains unclear. We investigated the contribution of the GI-CDF module to traits that depend on GI. Transcriptome profiling indicated that mutations in GI and the CDF genes have antagonistic effects on expression of a wider set of genes than CO and FT, whilst other genes are regulated by GI independently of the CDFs. Detailed expression studies followed by phenotypic assays showed that the CDFs function downstream of GI, influencing responses to freezing temperatures and growth, but are not necessary for proper clock function. Thus GI-mediated regulation of CDFs contributes to several processes in addition to flowering, but is not implicated in all of the traits influenced by GI. © 2015 The Authors The Plant Journal published by Society for Experimental Biology and John Wiley & Sons Ltd.

  4. Comparison of plasma membrane proteomic changes of Arabidopsis suspension-cultured cells (T87 Line) after cold and ABA treatment in association with freezing tolerance development.

    PubMed

    Li, Bin; Takahashi, Daisuke; Kawamura, Yukio; Uemura, Matsuo

    2012-03-01

    The plasma membrane (PM) is the primary site of freezing injury in plants. To determine global changes in PM protein profiles in association with freezing tolerance development, proteome analysis of the purified PM of Arabidopsis suspension-cultured cells (T87 line) was conducted with label-free protein quantification technology. Freezing tolerance of Arabidopsis cells at the lag growth phase (8 d old) increased after cold acclimation (CA) or ABA treatment. Proteome analysis assigned 658 proteins in the PM in total, of which 45.3% (298 proteins) were predicted to have transmembrane domains. They were classified into several functional categories, with the primary categories being proteins in transporters, signal transduction, protein destination and storage, and cell structure. After CA, 271 proteins increased and 111 proteins decreased. ABA treatment resulted in 185 increased and 56 decreased proteins. Of these, 139 increased and 49 decreased proteins were identified in common after both CA and ABA treatment. In addition, there were proteins specifically expressed in cold- (132 increased and 62 decreased) or ABA- (46 increased and 7 decreased) treated cells. Collectively, our results clearly show that (i) responses of the PM proteome to CA and ABA treatment overlap substantially but, at the same time, some proteins exhibited different response patterns in each treatment; and (ii) the majority of ABA-responsive proteins are CA-responsive proteins but not vice versa, suggesting complex interactions of CA and ABA signaling pathways in the PM proteome responses.

  5. The unified ICE-CBF pathway provides a transcriptional feedback control of freezing tolerance during cold acclimation in Arabidopsis.

    PubMed

    Kim, Ye Seul; Lee, Minyoung; Lee, Jae-Hyung; Lee, Hyo-Jun; Park, Chung-Mo

    2015-09-01

    During cold acclimation, C-repeat binding factors (CBFs) activate downstream targets, such as cold-regulated genes, leading to the acquisition of freezing tolerance in plants. Inducer of CBF expression 1 (ICE1) plays a key role by activating CBF3 expression in shaping the cold-induced transcriptome. While the ICE1-CBF3 regulon constitutes a major cold acclimation pathway, gene regulatory networks governing the CBF signaling are poorly understood. Here, we demonstrated that ICE1 and its paralog ICE2 induce CBF1, CBF2, and CBF3 by binding to the gene promoters. ICE2, like ICE1, was ubiquitinated by the high expression of osmotically responsive gene 1 (HOS1) E3 ubiquitin ligase. Whereas ICE2-defective ice2-2 mutant did not exhibit any discernible freezing-sensitive phenotypes, ice1-2 ice2-2/+ plant, which is defective in ICE1 and has a heterozygotic ice2 mutation, exhibited significantly reduced freezing tolerance. Accordingly, all three CBF genes were markedly down-regulated in the ice1-2 ice2-2/+ plant, indicating that ICE1 and ICE2 are functionally redundant with different implementations in inducing CBF genes. Together with the negative regulation of CBF3 by CBF2, we propose that the unified ICE-CBF pathway provides a transcriptional feedback of freezing tolerance to sustain plant development and survival during cold acclimation.

  6. A comparison of the low temperature transcriptomes and CBF regulons of three plant species that differ in freezing tolerance: Solanum commersonii, Solanum tuberosum, and Arabidopsis thaliana

    PubMed Central

    Pino, María-Teresa; Jeknić, Zoran; Zou, Cheng; Shiu, Shin-Han; Chen, Tony H. H.; Thomashow, Michael F.

    2011-01-01

    Solanum commersonii and Solanum tuberosum are closely related plant species that differ in their abilities to cold acclimate; whereas S. commersonii increases in freezing tolerance in response to low temperature, S. tuberosum does not. In Arabidopsis thaliana, cold-regulated genes have been shown to contribute to freezing tolerance, including those that comprise the CBF regulon, genes that are controlled by the CBF transcription factors. The low temperature transcriptomes and CBF regulons of S. commersonii and S. tuberosum were therefore compared to determine whether there might be differences that contribute to their differences in ability to cold acclimate. The results indicated that both plants alter gene expression in response to low temperature to similar degrees with similar kinetics and that both plants have CBF regulons composed of hundreds of genes. However, there were considerable differences in the sets of genes that comprised the low temperature transcriptomes and CBF regulons of the two species. Thus differences in cold regulatory programmes may contribute to the differences in freezing tolerance of these two species. However, 53 groups of putative orthologous genes that are cold-regulated in S. commersonii, S. tuberosum, and A. thaliana were identified. Given that the evolutionary distance between the two Solanum species and A. thaliana is 112–156 million years, it seems likely that these conserved cold-regulated genes—many of which encode transcription factors and proteins of unknown function—have fundamental roles in plant growth and development at low temperature. PMID:21511909

  7. A comparison of the low temperature transcriptomes and CBF regulons of three plant species that differ in freezing tolerance: Solanum commersonii, Solanum tuberosum, and Arabidopsis thaliana.

    PubMed

    Carvallo, Marcela A; Pino, María-Teresa; Jeknic, Zoran; Zou, Cheng; Doherty, Colleen J; Shiu, Shin-Han; Chen, Tony H H; Thomashow, Michael F

    2011-07-01

    Solanum commersonii and Solanum tuberosum are closely related plant species that differ in their abilities to cold acclimate; whereas S. commersonii increases in freezing tolerance in response to low temperature, S. tuberosum does not. In Arabidopsis thaliana, cold-regulated genes have been shown to contribute to freezing tolerance, including those that comprise the CBF regulon, genes that are controlled by the CBF transcription factors. The low temperature transcriptomes and CBF regulons of S. commersonii and S. tuberosum were therefore compared to determine whether there might be differences that contribute to their differences in ability to cold acclimate. The results indicated that both plants alter gene expression in response to low temperature to similar degrees with similar kinetics and that both plants have CBF regulons composed of hundreds of genes. However, there were considerable differences in the sets of genes that comprised the low temperature transcriptomes and CBF regulons of the two species. Thus differences in cold regulatory programmes may contribute to the differences in freezing tolerance of these two species. However, 53 groups of putative orthologous genes that are cold-regulated in S. commersonii, S. tuberosum, and A. thaliana were identified. Given that the evolutionary distance between the two Solanum species and A. thaliana is 112-156 million years, it seems likely that these conserved cold-regulated genes-many of which encode transcription factors and proteins of unknown function-have fundamental roles in plant growth and development at low temperature.

  8. Putrescine Is Involved in Arabidopsis Freezing Tolerance and Cold Acclimation by Regulating Abscisic Acid Levels in Response to Low Temperature1

    PubMed Central

    Cuevas, Juan C.; López-Cobollo, Rosa; Alcázar, Rubén; Zarza, Xavier; Koncz, Csaba; Altabella, Teresa; Salinas, Julio; Tiburcio, Antonio F.; Ferrando, Alejandro

    2008-01-01

    The levels of endogenous polyamines have been shown to increase in plant cells challenged with low temperature; however, the functions of polyamines in the regulation of cold stress responses are unknown. Here, we show that the accumulation of putrescine under cold stress is essential for proper cold acclimation and survival at freezing temperatures because Arabidopsis (Arabidopsis thaliana) mutants defective in putrescine biosynthesis (adc1, adc2) display reduced freezing tolerance compared to wild-type plants. Genes ADC1 and ADC2 show different transcriptional profiles upon cold treatment; however, they show similar and redundant contributions to cold responses in terms of putrescine accumulation kinetics and freezing sensitivity. Our data also demonstrate that detrimental consequences of putrescine depletion during cold stress are due, at least in part, to alterations in the levels of abscisic acid (ABA). Reduced expression of NCED3, a key gene involved in ABA biosynthesis, and down-regulation of ABA-regulated genes are detected in both adc1 and adc2 mutant plants under cold stress. Complementation analysis of adc mutants with ABA and reciprocal complementation tests of the aba2-3 mutant with putrescine support the conclusion that putrescine controls the levels of ABA in response to low temperature by modulating ABA biosynthesis and gene expression. PMID:18701673

  9. Ectopic Overexpression of SsCBF1, a CRT/DRE-Binding Factor from the Nightshade Plant Solanum lycopersicoides, Confers Freezing and Salt Tolerance in Transgenic Arabidopsis

    PubMed Central

    Zhang, Lili; Li, Zhenjun; Li, Jingfu; Wang, Aoxue

    2013-01-01

    The C-repeat (CRT)/dehydration-responsive element (DRE) binding factor (CBF/DREB1) transcription factors play a key role in cold response. However, the detailed roles of many plant CBFs are far from fully understood. A CBF gene (SsCBF1) was isolated from the cold-hardy plant Solanum lycopersicoides. A subcellular localization study using GFP fusion protein indicated that SsCBF1 is localized in the nucleus. We delimited the SsCBF1 transcriptional activation domain to the C-terminal segment comprising amino acid residues 193–228 (SsCBF1193–228). The expression of SsCBF1 could be dramatically induced by cold, drought and high salinity. Transactivation assays in tobacco leaves revealed that SsCBF1 could specifically bind to the CRT cis-elements in vivo to activate the expression of downstream reporter genes. The ectopic overexpression of SsCBF1 conferred increased freezing and high-salinity tolerance and late flowering phenotype to transgenic Arabidopsis. RNA-sequencing data exhibited that a set of cold and salt stress responsive genes were up-regulated in transgenic Arabidopsis. Our results suggest that SsCBF1 behaves as a typical CBF to contribute to plant freezing tolerance. Increased resistance to high-salinity and late flowering phenotype derived from SsCBF1 OE lines lend more credence to the hypothesis that plant CBFs participate in diverse physiological and biochemical processes related to adverse conditions. PMID:23755095

  10. Cold tolerance in Arabidopsis kamchatica.

    PubMed

    Armstrong, Jessica J; Takebayashi, Naoki; Sformo, Todd; Wolf, Diana E

    2015-03-01

    • Cold tolerance is a critically important factor determining how plants will be influenced by climate change, including changes in snowcover and extreme weather events. Although a great deal is known about cold tolerance in Arabidopsis thaliana, it is not highly cold tolerant. This study examined cold tolerance and its genetic diversity in an herbaceous subarctic relative, Arabidopsis kamchatica, which generally occurs in much colder climates.• Thermal analysis and electrolyte leakage were used to estimate supercooling points and lethal temperatures (LT50) in cold-acclimated and nonacclimated families from three populations of A. kamchatica.• Arabidopsis kamchatica was highly cold tolerant, with a mean LT50 of -10.8°C when actively growing, and -21.8°C when cold acclimated. It also was able to supercool to very low temperatures. Surprisingly, actively growing plants supercooled more than acclimated plants (-14.7 vs. -12.7°C). There was significant genetic variation for cold tolerance both within and among populations. However, both cold tolerance and genetic diversity were highest in the midlatitude population rather than in the far north, indicating that adaptations to climate change are most likely to arise in the center of the species range rather than at the edges.• Arabidopsis kamchatica is highly cold tolerant throughout its range. It is far more freeze tolerant than A. thaliana, and supercooled to lower temperatures, suggesting that A. kamchatica provides a valuable complement to A. thaliana for cold tolerance research. © 2015 Botanical Society of America, Inc.

  11. Phosphorylation of the transcriptional repressor MYB15 by mitogen-activated protein kinase 6 is required for freezing tolerance in Arabidopsis

    PubMed Central

    Kim, Sun Ho; Kim, Ho Soo; Bahk, Sunghwa; An, Jonguk; Yoo, Yeji; Kim, Jae-Yean

    2017-01-01

    Abstract The expression of CBF (C-repeat-binding factor) genes is required for freezing tolerance in Arabidopsis thaliana. CBFs are positively regulated by INDUCER OF CBF EXPRESSION1 (ICE1) and negatively regulated by MYB15. These transcription factors directly interact with specific elements in the CBF promoters. Mitogen-activated protein kinase (MAPK/MPK) cascades function upstream to regulate CBFs. However, the mechanism by which MPKs control CBF expression during cold stress signaling remains unknown. This study showed that the activity of MYB15, a transcriptional repressor of cold signaling, is regulated by MPK6-mediated phosphorylation. MYB15 specifically interacts with MPK6, and MPK6 phosphorylates MYB15 on Ser168. MPK6-induced phosphorylation reduced the affinity of MYB15 binding to the CBF3 promoter and mutation of its phosphorylation site (MYB15S168A) enhanced the transcriptional repression of CBF3 by MYB15. Furthermore, transgenic plants overexpressing MYB15S168A showed significantly reduced CBF transcript levels in response to cold stress, compared with plants overexpressing MYB15. The MYB15S168A-overexpressing plants were also more sensitive to freezing than MYB15-overexpressing plants. These results suggest that MPK6-mediated regulation of MYB15 plays an important role in cold stress signaling in Arabidopsis. PMID:28510716

  12. A novel Zea mays ssp. mexicana L. MYC-type ICE-like transcription factor gene ZmmICE1, enhances freezing tolerance in transgenic Arabidopsis thaliana.

    PubMed

    Lu, Xiang; Yang, Lei; Yu, Mengyuan; Lai, Jianbin; Wang, Chao; McNeil, David; Zhou, Meixue; Yang, Chengwei

    2017-04-01

    The annual Zea mays ssp. mexicana L., a member of the teosinte group, is a close wild relative of maize and thus can be effectively used in maize improvement. In this study, an ICE-like gene, ZmmICE1, was isolated from a cDNA library of RNA-Seq from cold-treated seedling tissues of Zea mays ssp. mexicana L. The deduced protein of ZmmICE1 contains a highly conserved basic helix-loop-helix (bHLH) domain and C-terminal region of ICE-like proteins. The ZmmICE1 protein localizes to the nucleus and shows sumoylation when expressed in an Escherichia coli reconstitution system. In addition, yeast one hybrid assays indicated that ZmmICE1 has transactivation activities. Moreover, ectopic expression of ZmmICE1 in the Arabidopsis ice1-2 mutant increased freezing tolerance. The ZmmICE1 overexpressed plants showed lower electrolyte leakage (EL), reduced contents of malondialdehyde (MDA). The expression of downstream cold related genes of Arabidopsis C-repeat-binding factors (AtCBF1, AtCBF2 and AtCBF3), cold-responsive genes (AtCOR15A and AtCOR47), kinesin-1 member gene (AtKIN1) and responsive to desiccation gene (AtRD29A) was significantly induced when compared with wild type under low temperature treatment. Taken together, these results indicated that ZmmICE1 is the homolog of Arabidopsis inducer of CBF expression genes (AtICE1/2) and plays an important role in the regulation of freezing stress response. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

  13. ACYL-LIPID DESATURASE2 Is Required for Chilling and Freezing Tolerance in Arabidopsis[C][W

    PubMed Central

    Chen, Mingjie; Thelen, Jay J.

    2013-01-01

    Fatty acid desaturation of membrane lipids is a strategy for plants to survive chilling or freezing temperature. To further characterize enzymes involved in this stress response pathway, ACYL-LIPID DESATURASE2 (ADS2; Enzyme Commission 1.14.99) was studied using genetic, cell, and biochemical approaches. ads2 mutant plants appear similar to the wild type under standard growth conditions but display a dwarf and sterile phenotype when grown at 6°C and also show increased sensitivity to freezing temperature. Fatty acid composition analysis demonstrated that ads2 mutant plants at 6°C have reduced levels of 16:1, 16:2, 16:3, and 18:3 and higher levels of 16:0 and 18:0 fatty acids compared with the wild type. Lipid profiling revealed that 34C species of phosphatidylglycerol (PG) and monogalactosyl diacylglycerol (MGDG) content in ads2 mutants were lower and phosphatidic acid, phosphatidylinositol, phosphatidylethanolamine, phosphatidylcholine, lyso-phosphatidylcholine, and phosphatidylserine were higher than the wild type. Subcellular localization of C- and N-terminal enhanced fluorescence fusion proteins indicated that ADS2 localized primarily to the endoplasmic reticulum, although signal was also confirmed in Golgi and plastids. A double mutation with a putative plastid ADS3 paralog exacerbates the growth defects of ads2 mutant plants under low temperature. These observations suggest that ADS2 encodes a 16:0 desaturase of MGDG and PG. We hypothesize that a low temperature–induced shift from the plastid to endoplasmic reticulum pathway for membrane lipid biosynthesis is required for the cold stress response in Arabidopsis thaliana, and ADS2 is essential to adjust the acyl composition of organelle membrane lipid composition in response to cold stress. PMID:23585650

  14. The Arabidopsis 14-3-3 Protein RARE COLD INDUCIBLE 1A Links Low-Temperature Response and Ethylene Biosynthesis to Regulate Freezing Tolerance and Cold Acclimation[C][W

    PubMed Central

    Catalá, Rafael; López-Cobollo, Rosa; Mar Castellano, M.; Angosto, Trinidad; Alonso, José M.; Ecker, Joseph R.; Salinas, Julio

    2014-01-01

    In plants, the expression of 14-3-3 genes reacts to various adverse environmental conditions, including cold, high salt, and drought. Although these results suggest that 14-3-3 proteins have the potential to regulate plant responses to abiotic stresses, their role in such responses remains poorly understood. Previously, we showed that the RARE COLD INDUCIBLE 1A (RCI1A) gene encodes the 14-3-3 psi isoform. Here, we present genetic and molecular evidence implicating RCI1A in the response to low temperature. Our results demonstrate that RCI1A functions as a negative regulator of constitutive freezing tolerance and cold acclimation in Arabidopsis thaliana by controlling cold-induced gene expression. Interestingly, this control is partially performed through an ethylene (ET)-dependent pathway involving physical interaction with different ACC SYNTHASE (ACS) isoforms and a decreased ACS stability. We show that, consequently, RCI1A restrains ET biosynthesis, contributing to establish adequate levels of this hormone in Arabidopsis under both standard and low-temperature conditions. We further show that these levels are required to promote proper cold-induced gene expression and freezing tolerance before and after cold acclimation. All these data indicate that RCI1A connects the low-temperature response with ET biosynthesis to modulate constitutive freezing tolerance and cold acclimation in Arabidopsis. PMID:25122152

  15. Freeze tolerance and avoidance in plants

    USDA-ARS?s Scientific Manuscript database

    Cold acclimation is a multigenic, quantitative trait that involves biochemical and structural changes that effect the physiology of a plant. Mechanisms associated with freeze tolerance or freeze avoidance develop and are lost on an annual cycle. When conducting studies to characterize and determin...

  16. Ice recrystallization inhibition proteins of perennial ryegrass enhance freezing tolerance.

    PubMed

    Zhang, Chunzhen; Fei, Shui-zhang; Arora, Rajeev; Hannapel, David J

    2010-06-01

    Ice recrystallization inhibition (IRI) proteins are thought to play an important role in conferring freezing tolerance in plants. Two genes encoding IRI proteins, LpIRI-a and LpIRI-b, were isolated from a relatively cold-tolerant perennial ryegrass cv. Caddyshack. Amino acid alignments among the IRI proteins revealed the presence of conserved repetitive IRI-domain motifs (NxVxxG/NxVxG) in both proteins. Quantitative reverse transcriptase PCR (qRT-PCR) analysis indicated that LpIRI-a was up-regulated approximately 40-fold while LpIRI-b was up-regulated sevenfold after just 1 h of cold acclimation, and by 7 days of cold acclimation the transcripts had increased 8,000-fold for LpIRI-a and 1,000-fold for LpIRI-b. Overexpression of either LpIRI-a or LpIRI-b gene in Arabidopsis increased survival rates of the seedlings following a freezing test under both cold-acclimated and nonacclimated conditions. For example, without cold acclimation a -4 degrees C treatment reduced the wild type's survival rate to an average of 73%, but resulted in survival rates of 85-100% for four transgenic lines. With cold acclimation, a -12 degrees C treatment reduced the wild type's survival rate to an average of 38.7%, while it resulted in a survival rate of 51-78.5% for transgenic lines. After cold acclimation, transgenic Arabidopsis plants overexpressing either LpIRI-a or LpIRI-b gene exhibited a consistent reduction in freezing-induced ion leakage at -8, -9, and -10 degrees C. Furthermore, the induced expression of the LpIRI-a and LpIRI-b proteins in transgenic E. coli enhanced the freezing tolerance in host cells. Our results suggest that IRI proteins play an important role in freezing tolerance in plants.

  17. Physiological responses of freeze-tolerant and -intolerant frogs: clues to evolution of anuran freeze tolerance.

    PubMed

    Costanzo, J P; Lee, R E; Lortz, P H

    1993-10-01

    Freeze tolerance in the wood frog, Rana sylvatica, is promoted by multiple, integrated physiological responses to ice forming within body tissues. By analyzing the freezing responses of the sympatric, but freeze intolerant, leopard frog (R. pipiens), we sought clues to the evolution of anuran freeze tolerance. Physiological responses critical to R. sylvatica's freeze tolerance, such as the synthesis and distribution of the cryoprotectant glucose, protective dehydration of organs, and deferred cardiac failure, were present, but comparatively less pronounced, in R. pipiens. Both species were innately tolerant of hyperglycemia. Glucose supplements did not enhance the freezing viability of R. pipiens, although in vitro tests of cryoprotectant efficacy revealed that glucose and glycerol provided comparable protection to erythrocytes of both species. We conclude that the evolution of freeze tolerance in R. sylvatica is not only promoted by its desiccation tolerance and the fortuitous biophysical consequences of freezing (e.g., exothermic induction of cardioacceleration and moderation of cooling rate) but also involves a progressive enhancement of fundamental physiological stress responses.

  18. Over-expression of JcDREB, a putative AP2/EREBP domain-containing transcription factor gene in woody biodiesel plant Jatropha curcas, enhances salt and freezing tolerance in transgenic Arabidopsis thaliana.

    PubMed

    Tang, Mingjuan; Liu, Xiaofei; Deng, Huaping; Shen, Shihua

    2011-12-01

    Jatropha curcas L. is an all-purpose biodiesel plant and is widely distributed in tropical and subtropical climates. It can grow well on poor quality soil which is not qualified for crop cultivation. This is very important for relieving land, food and energy crises. However, tropical and subtropical distribution limits the production of J. curcas seed. So it is valuable to know the molecular mechanism of J. curcas response to adverse abiotic environmental factors, especially freezing stress, in order to change the plant's characteristics. Until now there are just a few reports about J. curcas molecular biology. In this paper, we cloned and characterized a DNA binding protein from this plant, designated as JcDREB. Sequence analysis and yeast one-hybrid assays show that JcDREB can effectively function as a transcription factor of DREB protein family belonging to A-6 subgroup member. Expression patterns of JcDREB showed that it was induced by cold, salt and drought stresses, not by ABA. Over-expression of JcDREB in transgenic Arabidopsis exhibited enhanced salt and freezing stresses. Understanding the molecular mechanisms of J. curcas responses to environmental stresses, for example, high salinity, drought and low temperature, is crucial for improving their stress tolerance and productivity. This work provides more information about A-6 subgroup members of DREB subfamily.

  19. Molecular Physiology of Freeze Tolerance in Vertebrates.

    PubMed

    Storey, Kenneth B; Storey, Janet M

    2017-04-01

    Freeze tolerance is an amazing winter survival strategy used by various amphibians and reptiles living in seasonally cold environments. These animals may spend weeks or months with up to ∼65% of their total body water frozen as extracellular ice and no physiological vital signs, and yet after thawing they return to normal life within a few hours. Two main principles of animal freeze tolerance have received much attention: the production of high concentrations of organic osmolytes (glucose, glycerol, urea among amphibians) that protect the intracellular environment, and the control of ice within the body (the first putative ice-binding protein in a frog was recently identified), but many other strategies of biochemical adaptation also contribute to freezing survival. Discussed herein are recent advances in our understanding of amphibian and reptile freeze tolerance with a focus on cell preservation strategies (chaperones, antioxidants, damage defense mechanisms), membrane transporters for water and cryoprotectants, energy metabolism, gene/protein adaptations, and the regulatory control of freeze-responsive hypometabolism at multiple levels (epigenetic regulation of DNA, microRNA action, cell signaling and transcription factor regulation, cell cycle control, and anti-apoptosis). All are providing a much more complete picture of life in the frozen state.

  20. Potential role of salicylic acid in modulating diacylglycerol homeostasis in response to freezing temperatures in Arabidopsis.

    PubMed

    Tan, Wei-Juan; Xiao, Shi; Chen, Qin-Fang

    2015-01-01

    In our recent article in Molecular Plant, we reported that 3 lipase-like defense regulators SENESCENCE-ASSOCIATED GENE101 (SAG101), ENHANCED DISEASE SUSCEPTIBILITY1 (EDS1) and PHYTOALEXIN DEFICIENT4 (PAD4) are involved in the regulation of freezing tolerance in Arabidopsis. The transcripts of SAG101, EDS1 and PAD4 were inducible by cold stress and their knockout or knockdown mutants exhibited enhanced chilling and freezing tolerance in comparison to the wild type. The freezing tolerance phenotype showed in the sag101, eds1 and pad4 mutants was correlated with the transcriptional upregulation of C-REPEAT/DRE BINDING FACTORs (CBFs) and their regulons as well as increased levels of proline. Upon cold exposure, the sag101, eds1 and pad4 mutants showed ameliorated cell death and accumulation of hydrogen peroxide, which were highly induced by freezing stress in the wild-type leaves. Moreover, the contents of salicylic acid (SA) and diacylglycerol (DAG) were significantly decreased in the sag101, eds1 and pad4 mutants compared to the wild type. Taken together, our results suggest that the SAG101, EDS1 and PAD4 are negative regulators in the freezing response and function, at least in part, by modulating the homeostasis of SA and DAG in Arabidopsis.

  1. QTL mapping of freezing tolerance: links to fitness and adaptive trade-offs.

    PubMed

    Oakley, Christopher G; Ågren, Jon; Atchison, Rachel A; Schemske, Douglas W

    2014-09-01

    Local adaptation, defined as higher fitness of local vs. nonlocal genotypes, is commonly identified in reciprocal transplant experiments. Reciprocally adapted populations display fitness trade-offs across environments, but little is known about the traits and genes underlying fitness trade-offs in reciprocally adapted populations. We investigated the genetic basis and adaptive significance of freezing tolerance using locally adapted populations of Arabidopsis thaliana from Italy and Sweden. Previous reciprocal transplant studies of these populations indicated that subfreezing temperature is a major selective agent in Sweden. We used quantitative trait locus (QTL) mapping to identify the contribution of freezing tolerance to previously demonstrated local adaptation and genetic trade-offs. First, we compared the genomic locations of freezing tolerance QTL to those for previously published QTL for survival in Sweden, and overall fitness in the field. Then, we estimated the contributions to survival and fitness across both field sites of genotypes at locally adaptive freezing tolerance QTL. In growth chamber studies, we found seven QTL for freezing tolerance, and the Swedish genotype increased freezing tolerance for five of these QTL. Three of these colocalized with locally adaptive survival QTL in Sweden and with trade-off QTL for overall fitness. Two freezing tolerance QTL contribute to genetic trade-offs across environments for both survival and overall fitness. A major regulator of freezing tolerance, CBF2, is implicated as a candidate gene for one of the trade-off freezing tolerance QTL. Our study provides some of the first evidence of a trait and gene that mediate a fitness trade-off in nature. © 2014 John Wiley & Sons Ltd.

  2. Freeze tolerance in an arctic Alaska stonefly.

    PubMed

    Walters, Kent R; Sformo, Todd; Barnes, Brian M; Duman, John G

    2009-01-01

    Most aquatic insects do not survive subzero temperatures and, for those that do, the physiology has not been well characterized. Nemoura arctica is a species of stonefly widely distributed throughout arctic and subarctic Alaska. We collected nymphs from the headwaters of the Chandalar River, where we recorded streambed temperatures as low as -12.7 degrees C in midwinter. When in contact with ice, autumn-collected N. arctica cool to -1.5+/-0.4 degrees C before freezing, but individuals survived temperatures as low as -15 degrees C, making this the first described species of freeze-tolerant stonefly. N. arctica clearly survive freezing in nature, as winter-collected nymphs encased in ice demonstrated high survivorship when thawed. In the laboratory, 87% of N. arctica nymphs frozen to -15 degrees C for 2.5 weeks survived and, within one month of thawing, 95% of the last-instar nymphs emerged. N. arctica produce both glycerol and ice-binding factors (e.g. antifreeze protein) in response to low temperature. Hemolymph glycerol concentrations increased from 3 mmol l(-1) to 930+/-114 mmol l(-1) when temperatures were decreased from 4 degrees C to -8 degrees C, and N. arctica continued to produce glycerol even while frozen. Although the hemolymph of individual cold-acclimated nymphs occasionally exhibited more than a degree of thermal hysteresis, typically the hemolymph exhibited only hexagonal crystal growth, indicating a low concentration of ice-binding factor. Hemolymph of nymphs acclimated to subzero temperatures had recrystallization inhibition. These results demonstrate that, in the face of freezing conditions, N. arctica exhibit overwintering adaptations similar to those of terrestrial insects.

  3. Freeze tolerance of soil chytrids from temperate climates in Australia.

    PubMed

    Gleason, Frank H; Letcher, Peter M; McGee, Peter A

    2008-08-01

    Very little is known about the capacity of soil chytrids to withstand freezing in the field. Tolerance to freezing was tested in 21 chytrids isolated from cropping and undisturbed soils in temperate Australia. Samples of thalli grown on peptone-yeast-glucose (PYG) agar were incubated for seven days at -15 degrees C. Recovery of growth after thawing and transferring to fresh medium at 20 degrees C indicated survival. All isolates in the Blastocladiales and Spizellomycetales survived freezing in all tests. All isolates in the Chytridiales also survived freezing in some tests. None of the isolates in the Rhizophydiales survived freezing in any of the tests. However, some isolates in the Rhizophydiales recovered growth after freezing if they were grown on PYG agar supplemented with either 1% sodium chloride or 1% glycerol prior to freezing. After freezing, the morphology of the thalli of all isolates was observed under LM. In those isolates that recovered growth after transfer to fresh media, mature zoosporangia were observed in the monocentric isolates and resistant sporangia or resting spores in the polycentric isolates. Encysted zoospores in some monocentric isolates also survived freezing. In some of the experiments the freezing and thawing process caused visible structural damage to the thalli. The production of zoospores after freezing and thawing was also used as an indicator of freeze tolerance. The chytrids in this study responded differently to freezing. These data add significantly to our limited knowledge of freeze tolerance in chytrids but leave many questions unanswered.

  4. Glycerolipidome responses to freezing- and chilling-induced injuries: examples in Arabidopsis and rice.

    PubMed

    Zheng, Guowei; Li, Lixia; Li, Weiqi

    2016-03-22

    Glycerolipids are the principal constituent of cellular membranes; remodelling of glycerolipids plays important roles in temperature adaptation in plants. Temperate plants can endure freezing stress, but even chilling at above-zero temperatures can induce death in tropical species. However, little is known about the differences in glycerolipid response to low temperatures between chilling-sensitive and freezing-tolerant plants. Using ESI-MS/MS-based lipidomic analysis, we compared the glycerolipidome of chilling (4 and 10 °C)-treated rice with that of freezing (-6 and -12 °C)-treated Arabidopsis, both immediately after these low-temperature treatments and after a subsequent recovery culture period. Arabidopsis is a 16:3 plant that harbours both eukaryotic and prokaryotic-type lipid synthesis pathways, while rice is an 18:3 plant that harbours only the eukaryotic lipid synthesis pathway. Arabidopsis contains higher levels of galactolipids than rice and has a higher double bond index (DBI). Arabidopsis contains lower levels of high melting point phosphatidylglycerol (PG) molecules and has a lower average acyl chain length (ACL). Marked phospholipid degradation occurred during the recovery culture period of non-lethal chilling treated rice, but did not occur in non-lethal freezing treated Arabidopsis. Glycerolipids with larger head groups were synthesized more in Arabidopsis than in rice at sub-lethal low-temperatures. Levels of phosphatidic acid (PA) and phosphatidylinositol (PI) rose in both plants after low-temperature treatment. The DBI and ACL of total lipids did not change during low-temperature treatment. A higher DBI and a lower ACL could make the membranes of Arabidopsis more fluid at low temperatures. The ability to synthesize glycerolipids containing a larger head group may correlate with low-temperature tolerance. The low-temperature-induced increase of PA may play a dual role in plant responses to low temperatures: as a lipid signal that initiates

  5. Trehalose levels and survival ratio of freeze-tolerant versus freeze-sensitive yeasts.

    PubMed Central

    Hino, A; Mihara, K; Nakashima, K; Takano, H

    1990-01-01

    Five freeze-tolerant yeast strains suitable for frozen dough were compared with ordinary commercial bakers' yeast. Kluyveromyces thermotolerans FRI 501 cells showed high survival ability after freezing when their resting cells were fermented for 0 to 180 min in modified liquid medium, and they grew to log and stationary phases. Among the freeze-tolerant strains of Saccharomyces cerevisiae, FRI 413 and FRI 869 showed higher surviving and trehalose-accumulating abilities than other S. cerevisiae strains, but were affected by a prolonged prefermentation period and by growth phases. The freeze tolerance of the yeasts was, to some extent, associated with the basal amount of intracellular trehalose after rapid degradation at the onset of the prefermentation period. In the freeze-sensitive yeasts, the degree of hydrolysis of trehalose may thus be affected by the kind of saccharide, unlike in freeze-tolerant yeasts. PMID:2339891

  6. Trehalose levels and survival ratio of freeze-tolerant versus freeze-sensitive yeasts.

    PubMed

    Hino, A; Mihara, K; Nakashima, K; Takano, H

    1990-05-01

    Five freeze-tolerant yeast strains suitable for frozen dough were compared with ordinary commercial bakers' yeast. Kluyveromyces thermotolerans FRI 501 cells showed high survival ability after freezing when their resting cells were fermented for 0 to 180 min in modified liquid medium, and they grew to log and stationary phases. Among the freeze-tolerant strains of Saccharomyces cerevisiae, FRI 413 and FRI 869 showed higher surviving and trehalose-accumulating abilities than other S. cerevisiae strains, but were affected by a prolonged prefermentation period and by growth phases. The freeze tolerance of the yeasts was, to some extent, associated with the basal amount of intracellular trehalose after rapid degradation at the onset of the prefermentation period. In the freeze-sensitive yeasts, the degree of hydrolysis of trehalose may thus be affected by the kind of saccharide, unlike in freeze-tolerant yeasts.

  7. Freezing tolerance in plants requires lipid remodeling at the outer chloroplast membrane.

    PubMed

    Moellering, Eric R; Muthan, Bagyalakshmi; Benning, Christoph

    2010-10-08

    Plants show complex adaptations to freezing that prevent cell damage caused by cellular dehydration. Lipid remodeling of cell membranes during dehydration is one critical mechanism countering loss of membrane integrity and cell death. SENSITIVE TO FREEZING 2 (SFR2), a gene essential for freezing tolerance in Arabidopsis, encodes a galactolipid remodeling enzyme of the outer chloroplast envelope membrane. SFR2 processively transfers galactosyl residues from the abundant monogalactolipid to different galactolipid acceptors, forming oligogalactolipids and diacylglycerol, which is further converted to triacylglycerol. The combined activity of SFR2 and triacylglycerol-biosynthetic enzymes leads to the removal of monogalactolipids from the envelope membrane, changing the ratio of bilayer- to non-bilayer-forming membrane lipids. This SFR2-based mechanism compensates for changes in organelle volume and stabilizes membranes during freezing.

  8. Induction of Freeze-sensitive Mutants from a Freeze-tolerant Yeast, Torulaspora delbrueckii.

    PubMed

    Murakami, Y; Hahn, Y S; Yokoigawa, K; Endo, K; Kawai, H

    1994-01-01

    Freeze-sensitive strains of yeast were induced from a freeze-tolerant yeast Torulaspora delbrueckii by incubation with ethyl-methane sulfonate as a mutagen. A maximum ratio of mutation was attained by the incubation at 30°C for 75min. One-hundred and fifty strains of freeze-sensitive yeast were selected by plating-culture for the first screening. The freeze-tolerance ratio of each strain was examined based on the fermentative activity before and after freezing in liquid medium and dough. Strain 60B3 showed the highest freeze-sensitivity in a pre-fermented frozen dough (pre-fermented at 30°C for 2h, and frozen at -20°C for 7 days) among eight strains finally selected.

  9. Avoidance and tolerance of freezing in ectothermic vertebrates.

    PubMed

    Costanzo, Jon P; Lee, Richard E

    2013-06-01

    Ectothermic vertebrates have colonized regions that are seasonally or perpetually cold, and some species, particularly terrestrial hibernators, must cope with temperatures that fall substantially below 0°C. Survival of such excursions depends on either freeze avoidance through supercooling or freeze tolerance. Supercooling, a metastable state in which body fluids remain liquid below the equilibrium freezing/melting point, is promoted by physiological responses that protect against chilling injury and by anatomical and behavioral traits that limit risk of inoculative freezing by environmental ice and ice-nucleating agents. Freeze tolerance evolved from responses to fundamental stresses to permit survival of the freezing of a substantial amount of body water under thermal and temporal conditions of ecological relevance. Survival of freezing is promoted by a complex suite of molecular, biochemical and physiological responses that limit cell death from excessive shrinkage, damage to macromolecules and membranes, metabolic perturbation and oxidative stress. Although freeze avoidance and freeze tolerance generally are mutually exclusive strategies, a few species can switch between them, the mode used in a particular instance of chilling depending on prevailing physiological and environmental conditions.

  10. Aquaporin Expression Correlates with Freeze Tolerance in Baker's Yeast, and Overexpression Improves Freeze Tolerance in Industrial Strains

    PubMed Central

    Tanghe, An; Van Dijck, Patrick; Dumortier, Françoise; Teunissen, Aloys; Hohmann, Stefan; Thevelein, Johan M.

    2002-01-01

    Little information is available about the precise mechanisms and determinants of freeze resistance in baker's yeast, Saccharomyces cerevisiae. Genomewide gene expression analysis and Northern analysis of different freeze-resistant and freeze-sensitive strains have now revealed a correlation between freeze resistance and the aquaporin genes AQY1 and AQY2. Deletion of these genes in a laboratory strain rendered yeast cells more sensitive to freezing, while overexpression of the respective genes, as well as heterologous expression of the human aquaporin gene hAQP1, improved freeze tolerance. These findings support a role for plasma membrane water transport activity in determination of freeze tolerance in yeast. This appears to be the first clear physiological function identified for microbial aquaporins. We suggest that a rapid, osmotically driven efflux of water during the freezing process reduces intracellular ice crystal formation and resulting cell damage. Aquaporin overexpression also improved maintenance of the viability of industrial yeast strains, both in cell suspensions and in small doughs stored frozen or submitted to freeze-thaw cycles. Furthermore, an aquaporin overexpression transformant could be selected based on its improved freeze-thaw resistance without the need for a selectable marker gene. Since aquaporin overexpression does not seem to affect the growth and fermentation characteristics of yeast, these results open new perspectives for the successful development of freeze-resistant baker's yeast strains for use in frozen dough applications. PMID:12450819

  11. Aquaporin expression correlates with freeze tolerance in baker's yeast, and overexpression improves freeze tolerance in industrial strains.

    PubMed

    Tanghe, An; Van Dijck, Patrick; Dumortier, Françoise; Teunissen, Aloys; Hohmann, Stefan; Thevelein, Johan M

    2002-12-01

    Little information is available about the precise mechanisms and determinants of freeze resistance in baker's yeast, Saccharomyces cerevisiae. Genomewide gene expression analysis and Northern analysis of different freeze-resistant and freeze-sensitive strains have now revealed a correlation between freeze resistance and the aquaporin genes AQY1 and AQY2. Deletion of these genes in a laboratory strain rendered yeast cells more sensitive to freezing, while overexpression of the respective genes, as well as heterologous expression of the human aquaporin gene hAQP1, improved freeze tolerance. These findings support a role for plasma membrane water transport activity in determination of freeze tolerance in yeast. This appears to be the first clear physiological function identified for microbial aquaporins. We suggest that a rapid, osmotically driven efflux of water during the freezing process reduces intracellular ice crystal formation and resulting cell damage. Aquaporin overexpression also improved maintenance of the viability of industrial yeast strains, both in cell suspensions and in small doughs stored frozen or submitted to freeze-thaw cycles. Furthermore, an aquaporin overexpression transformant could be selected based on its improved freeze-thaw resistance without the need for a selectable marker gene. Since aquaporin overexpression does not seem to affect the growth and fermentation characteristics of yeast, these results open new perspectives for the successful development of freeze-resistant baker's yeast strains for use in frozen dough applications.

  12. Savanna Tree Seedlings are Physiologically Tolerant to Nighttime Freeze Events

    PubMed Central

    O’Keefe, Kimberly; Nippert, Jesse B.; Swemmer, Anthony M.

    2016-01-01

    Freeze events can be important disturbances in savanna ecosystems, yet the interactive effect of freezing with other environmental drivers on plant functioning is unknown. Here, we investigated physiological responses of South African tree seedlings to interactions of water availability and freezing temperatures. We grew widely distributed South African tree species (Colophospermum mopane, Combretum apiculatum, Acacia nigrescens, and Cassia abbreviata) under well-watered and water-limited conditions and exposed individuals to nighttime freeze events. Of the four species studied here, C. mopane was the most tolerant of lower water availability. However, all species were similarly tolerant to nighttime freezing and recovered within one week following the last freezing event. We also show that water limitation somewhat increased freezing tolerance in one of the species (C. mopane). Therefore, water limitation, but not freezing temperatures, may restrict the distribution of these species, although the interactions of these stressors may have species-specific impacts on plant physiology. Ultimately, we show that unique physiologies can exist among dominant species within communities and that combined stresses may play a currently unidentified role in driving the function of certain species within southern Africa. PMID:26870065

  13. Physiological responses to freezing in hatchlings of freeze-tolerant and -intolerant turtles.

    PubMed

    Costanzo, Jon P; Baker, Patrick J; Lee, Richard E

    2006-09-01

    Freeze tolerance is a complex cold-hardiness adaptation that has independently evolved in a diverse group of organisms, including several ectothermic vertebrates. Because little is known about the mechanistic basis for freeze tolerance in reptiles, we compared responses to experimental freezing in winter-acclimatized hatchlings representing nine taxa of temperate North American turtles, including ones that tolerated freezing and others that did not. Viability rates of hatchlings frozen to -3 degrees C for 72 h ranged from 0 to 100%. Tolerance to freezing was poor in Sternotherus odoratus, Graptemys geographica and Trachemys scripta, intermediate in Chelydra serpentina, and high in Emydoidea blandingii, Chrysemys picta bellii, C. p. marginata, Malaclemys terrapin, and Terrapene ornata, and generally reflected the winter thermal ecology of each taxon. Plasma activity of lactate dehydrogenase (LDH), a novel in vivo index of freeze/thaw damage, corroborated viability assessments and demonstrated that cryoinjury occurred even in surviving turtles. Irrespective of taxon, cryoinjury tended to be higher in smaller individuals and in those having relatively low water contents; however, bases for these associations were not apparent. Screening for certain organic osmolytes that might promote freezing survival by colligatively reducing ice content and limiting cell dehydration showed that the plasma of unfrozen (control) turtles contained small quantities of glucose (1.3-5.8 mmol l(-1)) and lactate (0.6-3.2 mmol l(-1)) and modest amounts of urea (range of mean values for all taxa 8.2-52.3 mmol l(-1)). Frozen/thawed turtles of all taxa accumulated modest amounts of glucose and lactate that jointly raised the plasma solute concentration by 30-100 mmol l(-1). We conclude that organic osmolytes accumulated both before and during freezing may promote survival in species that have evolved a tolerance to freezing, but are not necessarily accumulated for that purpose.

  14. Living in the cold: freeze-induced gene responses in freeze-tolerant vertebrates.

    PubMed

    Storey, K B

    1999-01-01

    1. Winter survival for numerous cold-blooded animals includes freeze tolerance: the ability to endure the conversion of as much as 65% of total body water into extracellular ice. Selected molecular adaptations underlying freeze tolerance (e.g. cryoprotectants, ice nucleating proteins) have been widely studied, but the full range of metabolic adjustments needed for freeze endurance remains unknown. 2. Recent studies using gene screening techniques are providing a different approach to the search for biochemical responses that support freezing survival by identifying genes and proteins that are up-regulated by freezing or thawing in freeze-tolerant amphibians and reptiles. 3. Screening of a cDNA library from wood frog liver revealed the freeze-induced up-regulation of genes coding for the alpha- and gamma-subunits of fibrinogen (a plasma clotting protein), the mitochondrial ADP/ATP translocase and a novel 10 kDa protein containing a nuclear exporting sequence. 4. Northern blotting revealed that these genes were differentially responsive to two of the component stresses of freezing (dehydration and anoxia), indicating that different genes are induced by signals radiating either from cell volume change or oxygen deprivation during freezing. 5. Freeze up-regulation of fibrinogen synthesis in liver and other organs appears to be a damage repair response that anticipates a need for enhanced plasma clotting capacity to deal with ice crystal damage to capillary beds. 6. Up-regulation of ADP/ATP translocase in frog liver is linked with ischaemia resistance and studies with freeze-tolerant turtles have shown that other genes encoding proteins involved in mitochondrial energetics (NADH-ubiquinone oxido-reductase subunit 5, cytochrome C oxidase subunit 1) are also up-regulated by both anoxia and freezing exposures. 7. These studies are making major advances in our understanding of freeze tolerance as a natural phenomenon and also highlight new key areas that can be targeted by

  15. Freezing tolerance of winter wheat as influenced by extended growth at low temperature and exposure to freeze-thaw cycles

    USDA-ARS?s Scientific Manuscript database

    As the seasons progress, autumn-planted winter wheat plants (Triticum aestivum L.) first gain, then progressively lose freezing tolerance. Exposing the plants to freeze-thaw cycles of -3/3°C results in increased ability to tolerate subsequent freezing to potentially damaging temperatures. This stu...

  16. Burkholderia phytofirmans PsJN reduces impact of freezing temperatures on photosynthesis in Arabidopsis thaliana

    PubMed Central

    Su, Fan; Jacquard, Cédric; Villaume, Sandra; Michel, Jean; Rabenoelina, Fanja; Clément, Christophe; Barka, Essaid A.; Dhondt-Cordelier, Sandrine; Vaillant-Gaveau, Nathalie

    2015-01-01

    Several plant growth-promoting rhizobacteria (PGPR) are known to improve plant tolerance to multiple stresses, including low temperatures. However, mechanisms underlying this protection are still poorly understood. The aim of this study was to evaluate the role of the endophytic PGPR, Burkholderia phytofirmans strain PsJN (Bp PsJN), on Arabidopsis thaliana cold tolerance using photosynthesis parameters as physiological markers. Under standard conditions, our results indicated that Bp PsJN inoculation led to growth promotion of Arabidopsis plants without significant modification on photosynthesis parameters and chloroplast organization. However, bacterial colonization induced a cell wall strengthening in the mesophyll. Impact of inoculation modes (either on seeds or by soil irrigation) and their effects overnight at 0, -1, or -3°C, were investigated by following photosystem II (PSII) activity and gas exchanges. Following low temperatures stress, a decrease of photosynthesis parameters was observed. In addition, during three consecutive nights or days at -1°C, PSII activity was monitored. Pigment contents, RuBisCO protein abundance, expression of several genes including RbcS, RbcL, CBF1, CBF2, CBF3, ICE1, COR15a, and COR78 were evaluated at the end of exposure. To assess the impact of the bacteria on cell ultrastructure under low temperatures, microscopic observations were achieved. Results indicated that freezing treatment induced significant changes in PSII activity as early as the first cold day, whereas the same impact on PSII activity was observed only during the third cold night. The significant effects conferred by PsJN were differential accumulation of pigments, and reduced expression of RbcL and COR78. Microscopical observations showed an alteration/disorganization in A. thaliana leaf mesophyll cells independently of the freezing treatments. The presence of bacteria during the three successive nights or days did not significantly improved A. thaliana

  17. Cryoprotectant Production in Freeze-Tolerant Wood Frogs Is Augmented by Multiple Freeze-Thaw Cycles.

    PubMed

    Larson, Don J; Barnes, Brian M

    2016-01-01

    Ice nucleation across the skin of wood frogs (Lithobates sylvaticus) rapidly induces endogenous production of glucose, a cryoprotectant necessary for freeze tolerance. In laboratory studies of freeze tolerance, wood frogs are cooled slowly, often at -0.05°C h(-1), to facilitate high cryoprotectant production and survival. Under natural conditions in Alaska, however, wood frogs accumulate maximal tissue glucose concentrations while cooling at much faster rates, -0.35° to -1.6°C h(-1), and in addition undergo multiple successive freeze-thaw cycles before remaining frozen for the winter. We examined whether simulating these ecologically relevant cooling rates and repeated freeze-thaw events in captive wood frogs results in the high glucose concentrations found in naturally frozen wood frogs. We found that over successive freezing and thawing events, glucose concentrations increased stepwise in all measured tissues. Short thawing periods did not result in a statistically significant decline of glucose concentrations. Wood frogs that experienced three freeze-thaw events had fresh weight glucose concentrations that approached values found in tissues of wood frogs frozen in natural conditions. Laboratory wood frogs survive frozen for 2 mo, while wood frogs frozen under natural conditions survive frozen for up to 7 mo at temperatures below -18°C. We hypothesize that repeated freeze-thaw cycles with rapid cooling and warming rates allow for greater survival in Alaskan wood frogs through enhanced cryoprotectant production.

  18. Freezing Tolerance of Citrus, Spinach, and Petunia Leaf Tissue 1

    PubMed Central

    Yelenosky, George; Guy, Charles L.

    1989-01-01

    Seasonal variations in freezing tolerance, water content, water and osmotic potential, and levels of soluble sugars of leaves of field-grown Valencia orange (Citrus sinensis) trees were studied to determine the ability of citrus trees to cold acclimate under natural conditions. Controlled environmental studies of young potted citrus trees, spinach (Spinacia pleracea), and petunia (Petunia hybrids) were carried out to study the water relations during cold acclimation under less variable conditions. During the coolest weeks of the winter, leaf water content and osmotic potential of field-grown trees decreased about 20 to 25%, while soluble sugars increased by 100%. At the same time, freezing tolerance increased from lethal temperature for 50% (LT50) of −2.8 to −3.8°C. In contrast, citrus leaves cold acclimated at a constant 10°C in growth chambers were freezing tolerant to about −6°C. The calculated freezing induced cellular dehydration at the LT50 remained relatively constant for field-grown leaves throughout the year, but increased for leaves of plants cold acclimated at 10°C in a controlled environment. Spinach leaves cold acclimated at 5°C tolerated increased cellular dehydration compared to nonacclimated leaves. Cold acclimated petunia leaves increased in freezing tolerance by decreasing osmotic potential, but had no capacity to change cellular dehydration sensitivity. The result suggest that two cold acclimation mechanisms are involved in both citrus and spinach leaves and only one in petunia leaves. The common mechanism in all three species tested was a minor increase in tolerance (about −1°C) resulting from low temperature induced osmotic adjustment, and the second in citrus and spinach was a noncolligative mechanism that increased the cellular resistance to freeze hydration. PMID:16666563

  19. Overexpression of VOZ2 confers biotic stress tolerance but decreases abiotic stress resistance in Arabidopsis

    PubMed Central

    Nakai, Yusuke; Fujiwara, Sumire; Kubo, Yasuyuki; Sato, Masa H.

    2013-01-01

    VOZ (vascular plant one zinc-finger protein) is a plant specific one-zinc finger type transcriptional activator, which is highly conserved through land plant evolution. We have previously shown that loss-of-function mutations in VOZ1 and VOZ2 showed increased cold and drought stress tolerances whereas decreased biotic stress resistance in Arabidopsis. Here, we demonstrate that transgenic plants overexpressing VOZ2 impairs freezing and drought stress tolerances but increases resistance to a fungal pathogen, Colletoricum higginsianum. Consistent with changes in the tolerance to biotic and abiotic stresses, the expression of marker genes for these stresses is significantly altered compared with those of the wild-type plant. These results indicate that a overexpression of VOZ2 confers biotic stress tolerance but impairs abiotic stress tolerances in Arabidopsis. PMID:23299334

  20. Overexpression of VOZ2 confers biotic stress tolerance but decreases abiotic stress resistance in Arabidopsis.

    PubMed

    Nakai, Yusuke; Fujiwara, Sumire; Kubo, Yasuyuki; Sato, Masa H

    2013-03-01

    VOZ (vascular plant one zinc-finger protein) is a plant specific one-zinc finger type transcriptional activator, which is highly conserved through land plant evolution. We have previously shown that loss-of-function mutations in VOZ1 and VOZ2 showed increased cold and drought stress tolerances whereas decreased biotic stress resistance in Arabidopsis. Here, we demonstrate that transgenic plants overexpressing VOZ2 impairs freezing and drought stress tolerances but increases resistance to a fungal pathogen, Colletoricum higginsianum. Consistent with changes in the tolerance to biotic and abiotic stresses, the expression of marker genes for these stresses is significantly altered compared with those of the wild-type plant. These results indicate that a overexpression of VOZ2 confers biotic stress tolerance but impairs abiotic stress tolerances in Arabidopsis.

  1. Aquaporin-Mediated Improvement of Freeze Tolerance of Saccharomyces cerevisiae Is Restricted to Rapid Freezing Conditions

    PubMed Central

    Tanghe, An; Van Dijck, Patrick; Colavizza, Didier; Thevelein, Johan M.

    2004-01-01

    Previous observations that aquaporin overexpression increases the freeze tolerance of baker's yeast (Saccharomyces cerevisiae) without negatively affecting the growth or fermentation characteristics held promise for the development of commercial baker's yeast strains used in frozen dough applications. In this study we found that overexpression of the aquaporin-encoding genes AQY1-1 and AQY2-1 improves the freeze tolerance of industrial strain AT25, but only in small doughs under laboratory conditions and not in large doughs under industrial conditions. We found that the difference in the freezing rate is apparently responsible for the difference in the results. We tested six different cooling rates and found that at high cooling rates aquaporin overexpression significantly improved the survival of yeast cells, while at low cooling rates there was no significant effect. Differences in the cultivation conditions and in the thawing rate did not influence the freeze tolerance under the conditions tested. Survival after freezing is determined mainly by two factors, cellular dehydration and intracellular ice crystal formation, which depend in an inverse manner on the cooling velocity. In accordance with this so-called two-factor hypothesis of freezing injury, we suggest that water permeability is limiting, and therefore that aquaporin function is advantageous, only under rapid freezing conditions. If this hypothesis is correct, then aquaporin overexpression is not expected to affect the leavening capacity of yeast cells in large, industrial frozen doughs, which do not freeze rapidly. Our results imply that aquaporin-overexpressing strains have less potential for use in frozen doughs than originally thought. PMID:15184134

  2. Aquaporin-mediated improvement of freeze tolerance of Saccharomyces cerevisiae is restricted to rapid freezing conditions.

    PubMed

    Tanghe, An; Van Dijck, Patrick; Colavizza, Didier; Thevelein, Johan M

    2004-06-01

    Previous observations that aquaporin overexpression increases the freeze tolerance of baker's yeast (Saccharomyces cerevisiae) without negatively affecting the growth or fermentation characteristics held promise for the development of commercial baker's yeast strains used in frozen dough applications. In this study we found that overexpression of the aquaporin-encoding genes AQY1-1 and AQY2-1 improves the freeze tolerance of industrial strain AT25, but only in small doughs under laboratory conditions and not in large doughs under industrial conditions. We found that the difference in the freezing rate is apparently responsible for the difference in the results. We tested six different cooling rates and found that at high cooling rates aquaporin overexpression significantly improved the survival of yeast cells, while at low cooling rates there was no significant effect. Differences in the cultivation conditions and in the thawing rate did not influence the freeze tolerance under the conditions tested. Survival after freezing is determined mainly by two factors, cellular dehydration and intracellular ice crystal formation, which depend in an inverse manner on the cooling velocity. In accordance with this so-called two-factor hypothesis of freezing injury, we suggest that water permeability is limiting, and therefore that aquaporin function is advantageous, only under rapid freezing conditions. If this hypothesis is correct, then aquaporin overexpression is not expected to affect the leavening capacity of yeast cells in large, industrial frozen doughs, which do not freeze rapidly. Our results imply that aquaporin-overexpressing strains have less potential for use in frozen doughs than originally thought.

  3. Hibernation physiology, freezing adaptation and extreme freeze tolerance in a northern population of the wood frog.

    PubMed

    Costanzo, Jon P; do Amaral, M Clara F; Rosendale, Andrew J; Lee, Richard E

    2013-09-15

    We investigated hibernation physiology and freeze tolerance in a population of the wood frog, Rana sylvatica, indigenous to Interior Alaska, USA, near the northernmost limit of the species' range. Winter acclimatization responses included a 233% increase in the hepatic glycogen depot that was subsidized by fat body and skeletal muscle catabolism, and a rise in plasma osmolality that reflected accrual of urea (to 106±10 μmol ml(-1)) and an unidentified solute (to ~73 μmol ml(-1)). In contrast, frogs from a cool-temperate population (southern Ohio, USA) amassed much less glycogen, had a lower uremia (28±5 μmol ml(-1)) and apparently lacked the unidentified solute. Alaskan frogs survived freezing at temperatures as low as -16°C, some 10-13°C below those tolerated by southern conspecifics, and endured a 2-month bout of freezing at -4°C. The profound freeze tolerance is presumably due to their high levels of organic osmolytes and bound water, which limits ice formation. Adaptive responses to freezing (-2.5°C for 48 h) and subsequent thawing (4°C) included synthesis of the cryoprotectants urea and glucose, and dehydration of certain tissues. Alaskan frogs differed from Ohioan frogs in retaining a substantial reserve capacity for glucose synthesis, accumulating high levels of cryoprotectants in brain tissue, and remaining hyperglycemic long after thawing. The northern phenotype also incurred less stress during freezing/thawing, as indicated by limited cryohemolysis and lactate accumulation. Post-glacial colonization of high latitudes by R. sylvatica required a substantial increase in freeze tolerance that was at least partly achieved by enhancing their cryoprotectant system.

  4. Freeze Tolerant Radiator for an Advanced EMU

    NASA Technical Reports Server (NTRS)

    Copeland, Robert J.; Elliott, Jeannine; Weislogel, Mark

    2004-01-01

    During an Extravehicular Activity (EVA), the astronaut s metabolic heat and the heat produced by the Portable Life Support Unit (PLSS) must be rejected. This heat load is currently rejected by a sublimator, which vents up to eight pounds of water each EVA. However, for advanced space missions of the future, water venting to space needs to be minimized because resupply impacts from earth will be prohibitive. If this heat load could be radiated to space from the PLSS, which has enough surface area to radiate most of the heat, the amount of water now vented could be greatly reduced. Unfortunately, a radiator rejects heat at a relatively constant rate, but the astronauts generate a variable heat load depending on how hard they are working. Without a way to vary the heat removal rate, the astronaut would experience cold discomfort or even frostbite. A proven method allowing a radiator to be turned-down is to sequentially allow tubes that carry the heat transfer fluid to the radiator to freeze. A drawback of current freezable radiators using this method is that they are far to heavy for use on a PLSS, because they use heavy construction to prevent the tubes from bursting as they freeze and thaw. This creates the need for a large radiator to reject most of the heat but with a lightweight tube that doesn t burst as it freezes and thaws. The new freezable radiator for the Extravehicular Mobility Unit (EMU) has features to accommodate the expansion of the radiator fluid when it freezes, and still have the high tube to fin conductance needed to minimize the number and weight of the tubes. Radiator fluid candidates are water and a propylene glycol-water mixture. This design maintains all materials within their elastic limits so that large volume changes can be achieved without breaking the tube. This concept couples this elastic expansion with an extremely lightweight, extremely high conductivity carbon fiber fin that can carry the heat needed to thaw a frozen tube. By using

  5. Freeze Tolerant Radiator for an Advanced EMU

    NASA Technical Reports Server (NTRS)

    Copeland, Robert J.; Elliott, Jeannine; Weislogel, Mark

    2004-01-01

    During an Extravehicular Activity (EVA), the astronaut s metabolic heat and the heat produced by the Portable Life Support Unit (PLSS) must be rejected. This heat load is currently rejected by a sublimator, which vents up to eight pounds of water each EVA. However, for advanced space missions of the future, water venting to space needs to be minimized because resupply impacts from earth will be prohibitive. If this heat load could be radiated to space from the PLSS, which has enough surface area to radiate most of the heat, the amount of water now vented could be greatly reduced. Unfortunately, a radiator rejects heat at a relatively constant rate, but the astronauts generate a variable heat load depending on how hard they are working. Without a way to vary the heat removal rate, the astronaut would experience cold discomfort or even frostbite. A proven method allowing a radiator to be turned-down is to sequentially allow tubes that carry the heat transfer fluid to the radiator to freeze. A drawback of current freezable radiators using this method is that they are far to heavy for use on a PLSS, because they use heavy construction to prevent the tubes from bursting as they freeze and thaw. This creates the need for a large radiator to reject most of the heat but with a lightweight tube that doesn t burst as it freezes and thaws. The new freezable radiator for the Extravehicular Mobility Unit (EMU) has features to accommodate the expansion of the radiator fluid when it freezes, and still have the high tube to fin conductance needed to minimize the number and weight of the tubes. Radiator fluid candidates are water and a propylene glycol-water mixture. This design maintains all materials within their elastic limits so that large volume changes can be achieved without breaking the tube. This concept couples this elastic expansion with an extremely lightweight, extremely high conductivity carbon fiber fin that can carry the heat needed to thaw a frozen tube. By using

  6. Overexpression of a proton-coupled vacuolar glucose exporter impairs freezing tolerance and seed germination.

    PubMed

    Klemens, Patrick A W; Patzke, Kathrin; Trentmann, Oliver; Poschet, Gernot; Büttner, Michael; Schulz, Alexander; Marten, Irene; Hedrich, Rainer; Neuhaus, H Ekkehard

    2014-04-01

    Arabidopsis vacuoles harbor, besides sugar transporter of the TMT-type, an early response to dehydration like 6 (ERDL6) protein involved in glucose export into the cytosol. However, the mode of transport of ERDL6 and the plant's feedback to overexpression of its activity on essential properties such as, for example, seed germination or freezing tolerance, remain unexplored. Using patch-clamp studies on vacuoles expressing AtERDL6 we demonstrated directly that this carrier operates as a proton-driven glucose exporter. Overexpression of BvIMP, the closest sugar beet (Beta vulgaris) homolog to AtERDL6, in Arabidopsis leads surprisingly to impaired seed germination under both conditions, sugar application and low environmental temperatures, but not under standard conditions. Upon cold treatment, BvIMP overexpressor plants accumulated lower quantities of monosaccharides than the wild-type, a response in line with the reduced frost tolerance of the transgenic Arabidopsis plants, and the fact that cold temperatures inhibits BvIMP transcription in sugar beet leaves. With these findings we show that the tight control of vacuolar sugar import and export is a key requisite for cold tolerance and seed germination of plants.

  7. Cytokinin response factor 4 (CRF4) is induced by cold and involved in freezing tolerance.

    PubMed

    Zwack, Paul J; Compton, Margaret A; Adams, Cami I; Rashotte, Aaron M

    2016-03-01

    Cytokinin response factor 4 (CRF4) shows a short-term induction by cold (4 °C) that appears to play a role in non-acclimated freezing tolerance as seen in mutant and overexpression lines. Responses to abiotic stresses, such as cold stress, are critical to plant growth and optimal production. Examination of Arabidopsis cytokinin response factors (CRFs) showed transcriptional induction after exposure to cold (4 °C). In particular, CRF4 was strongly induced in both root and shoot tissues. As CRF4 is one of several CRFs not transcriptionally regulated by cytokinin, we further investigated its response to cold. Peak CRF4 induction occurred 6 h post cold exposure, after which expression was maintained at moderately elevated levels during extended cold and subsequent treatment recovery. Examination of CRF4 mutant and overexpression lines under standard (non-cold) conditions revealed little difference from WT. One exception was a small, but significant increase in primary root growth of overexpression plants (CRF4OX). Under cold conditions, the only phenotype observed was a reduction in the rate of germination of CRF4OX seeds. The pattern of CRF4 expression along with the lack of strong phenotype at 4 °C led us to hypothesize that cold induction of CRF4 could play a role in short-term cold acclimation leading to increased freeze tolerance. Examination of CRF4OX and crf4 plants exposed to freezing temperatures revealed mutants lacking expression of CRF4 were more sensitive to freezing, while CRF4OXs with increased levels CRF4 levels were more tolerant. Altered transcript expression of CBF and COR15a cold signaling pathway genes in crf4 mutant and overexpression lines suggest that CRF4 may be potentially connected to this pathway. Overall this indicates that CRF4 plays an important role in both cold response and freezing stress.

  8. Regulation of SMAD transcription factors during freezing in the freeze tolerant wood frog, Rana sylvatica.

    PubMed

    Aguilar, Oscar A; Hadj-Moussa, Hanane; Storey, Kenneth B

    2016-11-01

    The wood frog, Rana sylvatica, survives sub-zero winter temperatures by undergoing full body freezing for weeks at a time, during which it displays no measurable brain activity, no breathing, and a flat-lined heart. Freezing is a hypometabolic state characterized by a global suppression of gene expression that is elicited in part by transcription factors that coordinate the activation of vital pro-survival pathways. Smad transcription factors respond to TGF-β signalling and are involved in numerous cellular functions from development to stress. Given the identity of genes they regulate, we hypothesized that they may be involved in coordinating gene expression during freezing. Protein expression of Smad1/2/3/4/5 in response to freezing was examined in 24h frozen and 8h thawed wood frog tissues using western immunoblotting, with the determination of subcellular localization in muscle and liver tissues. Transcript levels of smad2, smad4 and downstream genes (serpine1, myostatin, and tsc22d3) were measured by RT-PCR. Tissue-specific responses were observed during freezing where brain, heart, and liver had elevated levels of pSmad3, and skeletal muscle and kidneys had increased levels of pSmad1/5 and pSmad2 during freeze/thaw cycle, while protein and transcript levels remained constant. There were increases in nuclear levels of pSmad2 in muscle and pSmad3 in liver. Transcript levels of serpine1 were induced in heart, muscle, and liver, myostatin in muscle, and tsc22d3 in heart, and liver during freezing. These results suggest a novel freeze-responsive activation of Smad proteins that may play an important role in coordinating pro-survival gene networks necessary for freeze tolerance.

  9. Engineering carpel-specific cold stress tolerance: a case study in Arabidopsis.

    PubMed

    Artlip, Timothy S; Wisniewski, Michael E; Takatsuji, Hiroshi; Bassett, Carole L

    2016-08-01

    Climate change predictions forecast an increase in early spring frosts that could result in severe damage to perennial crops. For example, the Easter freeze of April 2007 left several states in the United States reporting a complete loss of that year's peach crop. The most susceptible organ to early frost damage in fruit trees is the carpel, particularly during bloom opening. In this study, we explored the use of a carpel-specific promoter (ZPT2-10) from petunia (Petunia hybrida var. Mitchell) to drive expression of the peach dehydrin PpDhn1. In peach, this gene is exceptionally responsive to low temperature but has not been observed to be expressed in carpels. This study examined carpel-specific properties of a petunia promoter driving the expression of the GUS gene (uidA) in transgenic Arabidopsis flowers and developed a carpel-specific ion leakage test to assess freezing tolerance. A homozygous Arabidopsis line (line 1-20) carrying the petunia ZPT2-10 promoter::PpDhn1 construct was obtained and freezing tolerance in the transgenic line was compared with an untransformed control. Overexpression of PpDhn1 in line 1-20 provided as much as a 1.9°C increase in carpel freezing tolerance as measured by electrolyte leakage. © 2016 Scandinavian Plant Physiology Society.

  10. Brassinosteroids participate in the control of basal and acquired freezing tolerance of plants.

    PubMed

    Eremina, Marina; Unterholzner, Simon J; Rathnayake, Ajith I; Castellanos, Marcos; Khan, Mamoona; Kugler, Karl G; May, Sean T; Mayer, Klaus F X; Rozhon, Wilfried; Poppenberger, Brigitte

    2016-10-04

    Brassinosteroids (BRs) are growth-promoting plant hormones that play a role in abiotic stress responses, but molecular modes that enable this activity remain largely unknown. Here we show that BRs participate in the regulation of freezing tolerance. BR signaling-defective mutants of Arabidopsis thaliana were hypersensitive to freezing before and after cold acclimation. The constitutive activation of BR signaling, in contrast, enhanced freezing resistance. Evidence is provided that the BR-controlled basic helix-loop-helix transcription factor CESTA (CES) can contribute to the constitutive expression of the C-REPEAT/DEHYDRATION-RESPONSIVE ELEMENT BINDING FACTOR (CBF) transcriptional regulators that control cold responsive (COR) gene expression. In addition, CBF-independent classes of BR-regulated COR genes are identified that are regulated in a BR- and CES-dependent manner during cold acclimation. A model is presented in which BRs govern different cold-responsive transcriptional cascades through the posttranslational modification of CES and redundantly acting factors. This contributes to the basal resistance against freezing stress, but also to the further improvement of this resistance through cold acclimation.

  11. Brassinosteroids participate in the control of basal and acquired freezing tolerance of plants

    PubMed Central

    Eremina, Marina; Unterholzner, Simon J.; Rathnayake, Ajith I.; Castellanos, Marcos; Khan, Mamoona; Kugler, Karl G.; May, Sean T.; Mayer, Klaus F. X.; Rozhon, Wilfried; Poppenberger, Brigitte

    2016-01-01

    Brassinosteroids (BRs) are growth-promoting plant hormones that play a role in abiotic stress responses, but molecular modes that enable this activity remain largely unknown. Here we show that BRs participate in the regulation of freezing tolerance. BR signaling-defective mutants of Arabidopsis thaliana were hypersensitive to freezing before and after cold acclimation. The constitutive activation of BR signaling, in contrast, enhanced freezing resistance. Evidence is provided that the BR-controlled basic helix–loop–helix transcription factor CESTA (CES) can contribute to the constitutive expression of the C-REPEAT/DEHYDRATION-RESPONSIVE ELEMENT BINDING FACTOR (CBF) transcriptional regulators that control cold responsive (COR) gene expression. In addition, CBF-independent classes of BR-regulated COR genes are identified that are regulated in a BR- and CES-dependent manner during cold acclimation. A model is presented in which BRs govern different cold-responsive transcriptional cascades through the posttranslational modification of CES and redundantly acting factors. This contributes to the basal resistance against freezing stress, but also to the further improvement of this resistance through cold acclimation. PMID:27655893

  12. Air-Cooled Stack Freeze Tolerance Freeze Failure Modes and Freeze Tolerance Strategies for GenDriveTM Material Handling Application Systems and Stacks Final Scientific Report

    SciTech Connect

    Hancock, David, W.

    2012-02-14

    Air-cooled stack technology offers the potential for a simpler system architecture (versus liquid-cooled) for applications below 4 kilowatts. The combined cooling and cathode air allows for a reduction in part count and hence a lower cost solution. However, efficient heat rejection challenges escalate as power and ambient temperature increase. For applications in ambient temperatures below freezing, the air-cooled approach has additional challenges associated with not overcooling the fuel cell stack. The focus of this project was freeze tolerance while maintaining all other stack and system requirements. Through this project, Plug Power advanced the state of the art in technology for air-cooled PEM fuel cell stacks and related GenDrive material handling application fuel cell systems. This was accomplished through a collaborative work plan to improve freeze tolerance and mitigate freeze-thaw effect failure modes within innovative material handling equipment fuel cell systems designed for use in freezer forklift applications. Freeze tolerance remains an area where additional research and understanding can help fuel cells to become commercially viable. This project evaluated both stack level and system level solutions to improve fuel cell stack freeze tolerance. At this time, the most cost effective solutions are at the system level. The freeze mitigation strategies developed over the course of this project could be used to drive fuel cell commercialization. The fuel cell system studied in this project was Plug Power's commercially available GenDrive platform providing battery replacement for equipment in the material handling industry. The fuel cell stacks were Ballard's commercially available FCvelocity 9SSL (9SSL) liquid-cooled PEM fuel cell stack and FCvelocity 1020ACS (Mk1020) air-cooled PEM fuel cell stack.

  13. Breeding of Freeze-tolerant Yeast and the Mechanisms of Stress-tolerance

    NASA Astrophysics Data System (ADS)

    Hino, Akihiro

    Frozen dough method have been adopted in the baking industry to reduce labor and to produce fresh breads in stores. New freeze-tolerant yeasts for frozen dough preparations were isolated from banana peel and identified. To obtain strains that have fermentative ability even after several months of frozen storage in fermented dough, we attempted to breed new freeze-tolerantstrain. The hybrid between S.cerevisiae, which is a isolated freeze-tolerant strain, and a strain isolated from bakers' yeast with sexual conjugation gave a good quality bread made from frozen dough method. Freeze-tolerant strains showed higher surviving and trehalose accumulating abilities than freeze-sensitive strains. The freeze tolerance of the yeasts was associated with the basal amount of intracellular trehalose after rapid degradation at the onset of the prefermentation period. The complicated metabolic pathway and the regulation system of trehalose in yeast cells are introduced. The trehalose synthesis may act as a metabolic buffer system which contribute to maintain the intracellular inorganic phosphate and as a feedback regulation system in the glycolysis. However, it is not known enough how the trehalose protects yeast cells from stress.

  14. Low Temperature Induces the Accumulation of Alcohol Dehydrogenase mRNA in Arabidopsis thaliana, a Chilling-Tolerant Plant.

    PubMed Central

    Jarillo, J. A.; Leyva, A.; Salinas, J.; Martinez-Zapater, J. M.

    1993-01-01

    mRNA encoding alcohol dehydrogenase (ADH) increases in etiolated seedlings and leaves of Arabidopsis thaliana (L.) Heynh. upon exposure to low temperature. The analysis of this response after water stress and abscisic acid (ABA) treatments in Arabidopsis wild type and ABA-deficient and -insensitive mutants indicates that cold accumulation of ADH mRNA could be induced by both anaerobic metabolism and increase of ABA concentration resulting from low temperature exposure. By using one Arabidopsis ADH null mutant, we show that ADH activity is not required for successful development of freezing tolerance in this species. PMID:12231733

  15. Overexpression of the Calcineurin Target CRZ1 Provides Freeze Tolerance and Enhances the Fermentative Capacity of Baker's Yeast▿

    PubMed Central

    Panadero, Joaquín; Hernández-López, Maria José; Prieto, José Antonio; Randez-Gil, Francisca

    2007-01-01

    Recent years have shown a huge growth in the market of industrial baker's yeasts (Saccharomyces cerevisiae), with the need for strains affording better performance in prefrozen dough. Evidence suggests that during the freezing process, cells can suffer biochemical damage caused by osmotic stress. Nevertheless, the involvement of ion-responsive transcriptional factors and pathways in conferring freeze resistance has not yet been examined. Here, we have investigated the role of the salt-responsive calcineurin-Crz1p pathway in mediating tolerance to freezing by industrial baker's yeast. Overexpression of CRZ1 in the industrial HS13 strain increased both salt and freeze tolerance and improved the leavening ability of baker's yeast in high-sugar dough. Moreover, engineered cells were able to produce more gas during fermentation of prefrozen dough than the parental strain. Similar effects were observed for overexpression of TdCRZ1, the homologue to CRZ1 in Torulaspora delbrueckii, suggesting that expression of calcineurin-Crz1p target genes can alleviate the harmful effects of ionic stress during freezing. However, overexpression of STZ and FTZ, two unrelated Arabidopsis thaliana genes encoding Cys2/His2-type zinc finger proteins, also conferred freeze resistance in yeast. Furthermore, experiments with Δcnb1 and Δcrz1 mutants failed to show a freeze-sensitive phenotype, even in cells pretreated with NaCl. Overall, our results demonstrate that overexpression of CRZ1 has the potential to be a useful tool for increasing freeze tolerance and fermentative capacity in industrial strains. However, these effects do not appear to be mediated through activation of known salt-responding pathways. PMID:17557846

  16. Overexpression of the calcineurin target CRZ1 provides freeze tolerance and enhances the fermentative capacity of baker's yeast.

    PubMed

    Panadero, Joaquín; Hernández-López, Maria José; Prieto, José Antonio; Randez-Gil, Francisca

    2007-08-01

    Recent years have shown a huge growth in the market of industrial baker's yeasts (Saccharomyces cerevisiae), with the need for strains affording better performance in prefrozen dough. Evidence suggests that during the freezing process, cells can suffer biochemical damage caused by osmotic stress. Nevertheless, the involvement of ion-responsive transcriptional factors and pathways in conferring freeze resistance has not yet been examined. Here, we have investigated the role of the salt-responsive calcineurin-Crz1p pathway in mediating tolerance to freezing by industrial baker's yeast. Overexpression of CRZ1 in the industrial HS13 strain increased both salt and freeze tolerance and improved the leavening ability of baker's yeast in high-sugar dough. Moreover, engineered cells were able to produce more gas during fermentation of prefrozen dough than the parental strain. Similar effects were observed for overexpression of TdCRZ1, the homologue to CRZ1 in Torulaspora delbrueckii, suggesting that expression of calcineurin-Crz1p target genes can alleviate the harmful effects of ionic stress during freezing. However, overexpression of STZ and FTZ, two unrelated Arabidopsis thaliana genes encoding Cys(2)/His(2)-type zinc finger proteins, also conferred freeze resistance in yeast. Furthermore, experiments with Deltacnb1 and Deltacrz1 mutants failed to show a freeze-sensitive phenotype, even in cells pretreated with NaCl. Overall, our results demonstrate that overexpression of CRZ1 has the potential to be a useful tool for increasing freeze tolerance and fermentative capacity in industrial strains. However, these effects do not appear to be mediated through activation of known salt-responding pathways.

  17. Allometry of cooling, supercooling, and freezing in the freeze-tolerant turtle Chrysemys picta.

    PubMed

    Claussen, D L; Zani, P A

    1991-09-01

    Although several vertebrates are freeze tolerant, little is known of the relationship between body size and the kinetics of cooling and freezing. We compared these responses for six hatchling and eight adult Chrysemys picta from an Ohio population. All turtles initially recovered from freezing, and all adults, but only two hatchlings (which experienced ice contents of approximately 35%), exhibited long-term survival. Rapid thawing may have compromised hatchling survival. Turtle water content was inversely related to body mass, but we found no significant correlation between the extent of supercooling and body size. Prefreezing and postfreezing cooling rates scaled with body mass to the -0.55 and -0.40 power, respectively, but the latter rate was more than two orders of magnitude slower. Theoretical (assuming 20% bound water) and calorimetric estimates of body ice agreed reasonably well. Ice contents were both body mass and time dependent. The absolute rate of ice formation scaled with body mass to the 0.4 power. Body size strongly influences the freezing response of ectotherms and deserves more attention.

  18. Use of a stress inducible promoter to drive ectopic AtCBF expression improves potato freezing tolerance while minimizing negative effects on tuber yield.

    PubMed

    Pino, María-Teresa; Skinner, Jeffrey S; Park, Eung-Jun; Jeknić, Zoran; Hayes, Patrick M; Thomashow, Michael F; Chen, Tony H H

    2007-09-01

    Solanum tuberosum is a frost-sensitive species incapable of cold acclimation. A brief exposure to frost can significantly reduce its yields, while hard frosts can completely destroy entire crops. Thus, gains in freezing tolerance of even a few degrees would be of considerable benefit relative to frost damage. The S. tuberosum cv. Umatilla was transformed with three Arabidopsis CBF genes (AtCBF1-3) driven by either a constitutive CaMV35S or a stress-inducible Arabidopsis rd29A promoter. AtCBF1 and AtCBF3 over-expression via the 35S promoter increased freezing tolerance about 2 degrees C, whereas AtCBF2 over-expression failed to increase freezing tolerance. Transgenic plants of AtCBF1 and AtCBF3 driven by the rd29A promoter reached the same level of freezing tolerance as the 35S versions within a few hours of exposure to low but non-freezing temperatures. Constitutive expression of AtCBF genes was associated with negative phenotypes, including smaller leaves, stunted plants, delayed flowering, and reduction or lack of tuber production. While imparting the same degree of freezing tolerance, control of AtCBF expression via the stress-inducible promoter ameliorated these negative phenotypic effects and restored tuber production to levels similar to wild-type plants. These results suggest that use of a stress-inducible promoter to direct CBF transgene expression can yield significant gains in freezing tolerance without negatively impacting agronomically important traits in potato.

  19. The Siberian timberman Acanthocinus aedilis: a freeze-tolerant beetle with low supercooling points.

    PubMed

    Kristiansen, E; Li, N G; Averensky, A I; Laugsand, A E; Zachariassen, K E

    2009-07-01

    Larvae of the Siberian timberman beetle Acanthocinus aedilis display a number of unique features, which may have important implications for the field of cold hardiness in general. Their supercooling points are scattered over a wide temperature range, and some individuals have supercooling points in the low range of other longhorn beetles. However, they differ from other longhorn beetles in being tolerant to freezing, and in the frozen state they tolerate cooling to below -37 degrees C. In this respect they also differ from the European timberman beetles, which have moderate supercooling capacity and die if they freeze. The combination of freezing tolerance and low supercooling points is unusual and shows that freezing at a high subzero temperature is not an absolute requirement for freezing tolerance. Like other longhorn beetles, but in contrast to other freeze-tolerant insects, the larvae of the Siberian timberman have a low cuticular water permeability and can thus stay supercooled for long periods without a great water loss. This suggests that a major function of the extracellular ice nucleators of some freeze-tolerant insects may be to prevent intolerable water loss in insects with high cuticular water permeability, rather than to create a protective extracellular freezing as has generally been assumed. The freezing tolerance of the Siberian timberman larvae is likely to be an adaptation to the extreme winter cold of Siberia.

  20. Life in a frozen state: adaptive strategies for natural freeze tolerance in amphibians and reptiles.

    PubMed

    Storey, K B

    1990-03-01

    Winter survival for various species of amphibians and reptiles that hibernate on land depends on freeze tolerance, the ability to survive for long periods of time with up to 65% of total body water as extracellular ice. Freeze tolerance has been described for four species of frogs, one salamander, and hatchlings of the painted turtle. A very limited tolerance also occurs in garter snakes. Studies of freeze tolerance in vertebrates have primarily focused on the wood frog Rana sylvatica and have assessed the regulation of cryoprotectant synthesis, cryoprotectant action in freezing preservation of isolated cells and tissues, metabolism and energetics under the ischemic conditions imposed by freezing, and the role of ice-nucleating agents in blood. The adaptations that preserve life at subzero temperatures for these animals illustrate the principles of vertebrate organ cryopreservation and may have important applications in the development of technology for the freezing preservation of transplantable human organs.

  1. A whole-plant screening test to identify genotypes with superior freezing tolerance.

    PubMed

    Bertrand, Annick; Castonguay, Yves; Bourassa, Josée

    2014-01-01

    Freezing tolerance is a determinant factor of persistence of perennials grown in northern climate. Selection for winterhardiness in field nurseries is difficult because of the unpredictability of occurrence of test winters allowing the identification of hardy genotypes. Here we describe a whole-plant assay entirely performed indoor in growth chambers and walk-in freezers to identify genotypes with superior tolerance to freezing within populations of open pollinated species. Three successive freezing stresses are applied to progressively eliminate 90 % of the population and to retain only the 10 % best performing genotypes. This approach can be used to generate recurrently selected populations more tolerant to freezing in different species.

  2. Freezing and Desiccation Tolerance in Entomopathogenic Nematodes: Diversity and Correlation of Traits

    USDA-ARS?s Scientific Manuscript database

    The ability of entomopathogenic nematodes to tolerate environmental stress such as desiccating or freezing conditions, can contribute significantly to biocontrol efficacy. Our objective was to compare inter and intraspecific variation in freeze and desiccation tolerance among a broad array of entom...

  3. Variation in Arabidopsis flooding responses identifies numerous putative "tolerance genes".

    PubMed

    Vashisht, Divya; van Veen, Hans; Akman, Melis; Sasidharan, Rashmi

    2016-11-01

    Plant survival in flooded environments requires a combinatory response to multiple stress conditions such as limited light availability, reduced gas exchange and nutrient uptake. The ability to fine-tune the molecular response at the transcriptional and/or post-transcriptional level that can eventually lead to metabolic and anatomical adjustments are the underlying requirements to confer tolerance. Previously, we compared the transcriptomic adjustment of submergence tolerant, intolerant accessions and identified a core conserved and genotype-specific response to flooding stress, identifying numerous 'putative' tolerance genes. Here, we performed genome wide association analyses on 81 natural Arabidopsis accessions that identified 30 additional SNP markers associated with flooding tolerance. We argue that, given the many genes associated with flooding tolerance in Arabidopsis, improving resistance to submergence requires numerous genetic changes.

  4. Responses to freezing exposure of hatchling turtles Trachemys scripta elegans: factors influencing the development of freeze tolerance by reptiles.

    PubMed

    Churchill, T A; Storey, K B

    1992-06-01

    Hatchling red-eared turtles Trachemys (= Pseudemys) scripta elegans (Wied) from a Louisiana population display a significant ability to withstand the freezing of extracellular body fluids. All animals survived at least 2 h of freezing at -2.5 or -4 degrees C. At -2.5 degrees C, survival declined to 50% after 6 h of freezing and no animals recovered after 24 h or longer, when mean ice content reached 54.7 +/- 1.4% of total body water. At -4 degrees C, all turtles recovered from 4 h of freezing exposure with a mean ice content of 49.6 +/- 2.4%, but survival dropped sharply thereafter with no animals recovering after 8 h, when ice content had reached 64.5 +/- 0.7%. Survival times were substantially shorter and percentage ice values greater than comparable values for hatchling painted turtles (Chrysemys picta (Schneider)) from northern populations subjected to identical freezing exposures. The ability to synthesize cryoprotectants in response to freezing was poorly developed in T. s. elegans; maximal accumulation of glucose was only 3.2 mumol g-1 wet mass in liver. Lactate content increased two- to threefold in oxygen-sensitive organs (heart and brain) during freezing, but levels of lactate and other putative cryoprotectants were unchanged in other organs. Total free amino acid content rose significantly in liver, muscle and blood during freezing; increased taurine concentration was primarily responsible for the changes in liver and blood. The capacity for freezing survival by T. s. elegans hatchlings from southern populations would be of limited use for hibernation in a cold climate, but the metabolic responses to freezing displayed by these animals might be enhanced by northern populations to increase their freeze tolerance.

  5. Chlorophyll fluorescence emission as a reporter on cold tolerance in Arabidopsis thaliana accessions

    PubMed Central

    Mishra, Anamika; Höermiller, Imke I; Heyer, Arnd G; Nedbal, Ladislav

    2011-01-01

    Non-invasive, high-throughput screening methods are valuable tools in breeding for abiotic stress tolerance in plants. Optical signals such as chlorophyll fluorescence emission can be instrumental in developing new screening techniques. In order to examine the potential of chlorophyll fluorescence to reveal plant tolerance to low temperatures, we used a collection of nine Arabidopsis thaliana accessions and compared their fluorescence features with cold tolerance quantified by the well established electrolyte leakage method on detached leaves. We found that, during progressive cooling, the minimal chlorophyll fluorescence emission rose strongly and that this rise was highly dependent on the cold tolerance of the accessions. Maximum quantum yield of PSII photochemistry and steady state fluorescence normalized to minimal fluorescence were also highly correlated to the cold tolerance measured by the electrolyte leakage method. In order to further increase the capacity of the fluorescence detection to reveal the low temperature tolerance, we applied combinatorial imaging that employs plant classification based on multiple fluorescence features. We found that this method, by including the resolving power of several fluorescence features, can be well employed to detect cold tolerance already at mild sub-zero temperatures. Therefore, there is no need to freeze the screened plants to the largely damaging temperatures of around −15°C. This, together with the method's easy applicability, represents a major advantage of the fluorescence technique over the conventional electrolyte leakage method. PMID:21427532

  6. High-pressure freezing and freeze substitution of Arabidopsis for electron microscopy.

    PubMed

    Austin, Jotham R

    2014-01-01

    The objectives of electron microscopy ultrastructural studies are to examine cellular architecture and relate the cell's structural machinery to dynamic functional roles. This aspiration is difficult to achieve if specimens have not been adequately preserved in a "living state"; hence specimen preparation is of the utmost importance for the success of any electron micrographic study. High-pressure freezing (HPF)/freeze substitution (FS) has long been recognized as the primer technique for the preservation of ultrastructure in biological samples. In most cases a basic HPF/freeze substitution protocol is sufficient to obtain superior ultrastructural preservation and structural contrast, which allows one to use more advanced microscopy techniques such as 3D electron tomography. However, for plant tissues, which have a thick cell wall, large water-filled vacuoles, and air spaces (all of which are detrimental to cryopreservation), these basic HPF/FS protocols often yield undesirable results. In particular, ice crystal artifacts and the staining of membrane systems are often poorly or negatively stained, which make 3D segmentation of a tomogram difficult. To overcome these problems, various aspects of the HPF/FS protocol can be altered, including the cryo-filler(s) used, freeze substitution cocktail, and the resin infiltration process. This chapter will describe these modifications for the preparation of plant tissues for routine electron microscopic studies, immunocytochemistry, and 3D tomographic electron imaging.

  7. Over-expression of ThpI from Choristoneura fumiferana enhances tolerance to cold in Arabidopsis.

    PubMed

    Zhu, Bo; Xiong, Ai-Sheng; Peng, Ri-He; Xu, Jing; Jin, Xiao-Fen; Meng, Xiu-Rong; Yao, Quan-Hong

    2010-02-01

    Thermal hysteresis proteins (Thps) known as antifreeze proteins for their antifreeze activity, depress the freezing point of water below the melting point in many polar marine fishes, terrestrial arthropods and plants. For the purpose of breeding cold-resistant plants, we designed to introduce the Thp gene into the plants. The physiological and biochemical effect of high-lever expression of the modified Choristoneura fumiferana Thp (ThpI) in Arabidopsis thaliana plants was analyzed. Under low temperature stress, the ThpI transgenic plants exhibited stronger growth than wild-type plants. The elevated cold tolerance of the ThpI over-expressing plants was confirmed by the changes of electrolyte leakage activity, malonyldialdehyde and proline contents. These results preliminarily showed that the Thp possibly be used to enhance the low temperature-tolerant ability of plants.

  8. Improvement of tolerance to freeze-thaw stress of baker's yeast by cultivation with soy peptides.

    PubMed

    Izawa, Shingo; Ikeda, Kayo; Takahashi, Nobuyuki; Inoue, Yoshiharu

    2007-06-01

    The tolerance to freeze-thaw stress of yeast cells is critical for frozen-dough technology in the baking industry. In this study, we examined the effects of soy peptides on the freeze-thaw stress tolerance of yeast cells. We found that the cells cultured with soy peptides acquired improved tolerance to freeze-thaw stress and retained high leavening ability in dough after frozen storage for 7 days. The final quality of bread regarding its volume and texture was also improved by using yeast cells cultured with soy peptides. These findings promote the utilization of soy peptides as ingredients of culture media to improve the quality of baker's yeast.

  9. Anhydrobiosis and Freezing-Tolerance: Adaptations That Facilitate the Establishment of Panagrolaimus Nematodes in Polar Habitats

    PubMed Central

    McGill, Lorraine M.; Shannon, Adam J.; Pisani, Davide; Félix, Marie-Anne; Ramløv, Hans; Dix, Ilona; Wharton, David A.; Burnell, Ann M.

    2015-01-01

    Anhydrobiotic animals can survive the loss of both free and bound water from their cells. While in this state they are also resistant to freezing. This physiology adapts anhydrobiotes to harsh environments and it aids their dispersal. Panagrolaimus davidi, a bacterial feeding anhydrobiotic nematode isolated from Ross Island Antarctica, can survive intracellular ice formation when fully hydrated. A capacity to survive freezing while fully hydrated has also been observed in some other Antarctic nematodes. We experimentally determined the anhydrobiotic and freezing-tolerance phenotypes of 24 Panagrolaimus strains from tropical, temperate, continental and polar habitats and we analysed their phylogenetic relationships. We found that several other Panagrolaimus isolates can also survive freezing when fully hydrated and that tissue extracts from these freezing-tolerant nematodes can inhibit the growth of ice crystals. We show that P. davidi belongs to a clade of anhydrobiotic and freezing-tolerant panagrolaimids containing strains from temperate and continental regions and that P. superbus, an early colonizer at Surtsey island, Iceland after its volcanic formation, is closely related to a species from Pennsylvania, USA. Ancestral state reconstructions show that anhydrobiosis evolved deep in the phylogeny of Panagrolaimus. The early-diverging Panagrolaimus lineages are strongly anhydrobiotic but weakly freezing-tolerant, suggesting that freezing tolerance is most likely a derived trait. The common ancestors of the davidi and the superbus clades were anhydrobiotic and also possessed robust freezing tolerance, along with a capacity to inhibit the growth and recrystallization of ice crystals. Unlike other endemic Antarctic nematodes, the life history traits of P. davidi do not show evidence of an evolved response to polar conditions. Thus we suggest that the colonization of Antarctica by P. davidi and of Surtsey by P. superbus may be examples of recent “ecological fitting

  10. Anhydrobiosis and freezing-tolerance: adaptations that facilitate the establishment of Panagrolaimus nematodes in polar habitats.

    PubMed

    McGill, Lorraine M; Shannon, Adam J; Pisani, Davide; Félix, Marie-Anne; Ramløv, Hans; Dix, Ilona; Wharton, David A; Burnell, Ann M

    2015-01-01

    Anhydrobiotic animals can survive the loss of both free and bound water from their cells. While in this state they are also resistant to freezing. This physiology adapts anhydrobiotes to harsh environments and it aids their dispersal. Panagrolaimus davidi, a bacterial feeding anhydrobiotic nematode isolated from Ross Island Antarctica, can survive intracellular ice formation when fully hydrated. A capacity to survive freezing while fully hydrated has also been observed in some other Antarctic nematodes. We experimentally determined the anhydrobiotic and freezing-tolerance phenotypes of 24 Panagrolaimus strains from tropical, temperate, continental and polar habitats and we analysed their phylogenetic relationships. We found that several other Panagrolaimus isolates can also survive freezing when fully hydrated and that tissue extracts from these freezing-tolerant nematodes can inhibit the growth of ice crystals. We show that P. davidi belongs to a clade of anhydrobiotic and freezing-tolerant panagrolaimids containing strains from temperate and continental regions and that P. superbus, an early colonizer at Surtsey island, Iceland after its volcanic formation, is closely related to a species from Pennsylvania, USA. Ancestral state reconstructions show that anhydrobiosis evolved deep in the phylogeny of Panagrolaimus. The early-diverging Panagrolaimus lineages are strongly anhydrobiotic but weakly freezing-tolerant, suggesting that freezing tolerance is most likely a derived trait. The common ancestors of the davidi and the superbus clades were anhydrobiotic and also possessed robust freezing tolerance, along with a capacity to inhibit the growth and recrystallization of ice crystals. Unlike other endemic Antarctic nematodes, the life history traits of P. davidi do not show evidence of an evolved response to polar conditions. Thus we suggest that the colonization of Antarctica by P. davidi and of Surtsey by P. superbus may be examples of recent "ecological fitting

  11. Importance of freeze-thaw events in low temperature ecotoxicology of cold tolerant enchytraeids.

    PubMed

    Silva, Ana L Patrício; Enggrob, Kirsten; Slotsbo, Stine; Amorim, Mónica J B; Holmstrup, Martin

    2014-08-19

    Due to global warming it is predicted that freeze-thaw cycles will increase in Arctic and cold temperate regions. The effects of this variation becomes of particular ecological importance to freeze-tolerant species when it is combined with chemical pollutants. We compared the effect of control temperature (2 °C), daily freeze-thaw cycles (2 to -4 °C) and constant freezing (-2 °C) temperatures on the cold-tolerance of oligochaete worms (Enchytraeus albidus) and tested how survival was influenced by pre-exposure to 4-nonylphenol (4-NP), a common nonionic detergent found in sewage sludge amended soils. Results showed that combined effect of 4-NP and daily freeze-thaw cycles can cause higher mortality to worms as compared with sustained freezing or control temperature. Exposure to 4-NP caused a substantial depletion of glycogen reserves which is catabolized during freezing to produce cryoprotective concentrations of free glucose. Further, exposure to freeze-thaw cycles resulted in higher concentrations of 4-NP in worm tissues as compared to constant freezing or control temperature (2 °C). Thus, worms exposed to combined effect of freeze-thaw cycles and 4-NP suffer higher consequences, with the toxic effect of the chemical potentiating the deleterious effects of freezing and thawing.

  12. Overexpression of SOD2 increases salt tolerance of Arabidopsis.

    PubMed

    Gao, Xiuhua; Ren, Zhonghai; Zhao, Yanxiu; Zhang, Hui

    2003-12-01

    The yeast (Schizosaccharomyces pombe) SOD2 (Sodium2) gene was introduced into Arabidopsis under the control of the cauliflower mosaic virus 35S promoter. Transformants were selected for their ability to grow on medium containing kanamycin. Southern- and northern-blot analyses confirmed that SOD2 was transferred into the Arabidopsis genome. There were no obvious morphological or developmental differences between the transgenic and wild-type (wt) plants. Several transgenic homozygous lines and wt plants (control) were evaluated for salt tolerance and gene expression. Overexpression of SOD2 in Arabidopsis improved seed germination and seedling salt tolerance. Analysis of Na+ and K+ contents of the symplast and apoplast in the parenchyma cells of the root cortex and mesophyll cells in the spongy tissue of the leaf showed that transgenic lines accumulated less Na+ and more K+ in the symplast than the wt plants did. The photosynthetic rate and the fresh weight of the transgenic lines were distinctly higher than that of wt plants after NaCl treatment. Results from different tests indicated that the expression of the SOD2 gene promoted a higher level of salt tolerance in vivo in transgenic Arabidopsis plants.

  13. Using Arabidopsis to explore zinc tolerance and hyperaccumulation.

    PubMed

    Roosens, Nancy H C J; Willems, Glenda; Saumitou-Laprade, Pierre

    2008-05-01

    Identifying the particular gene or genes underlying a specific adaptation is a major challenge in modern biology. Currently, the study of naturally occurring variation in Arabidopsis thaliana provides a bridge between functional genetics and evolutionary analyses. Nevertheless, the use of A. thaliana to study adaptation is limited to those traits that have undergone selection. Therefore, to understand fully the genetics of adaptation, the vast arsenal of genetic resources developed in A. thaliana must be extended to other species that display traits absent in this model species. Here, we discuss how A. thaliana resources can significantly enhance the study of heavy-metal tolerance and hyperaccumulation in the wild species Arabidopsis halleri.

  14. Arginine and proline applied as food additives stimulate high freeze tolerance in larvae of Drosophila melanogaster.

    PubMed

    Koštál, Vladimír; Korbelová, Jaroslava; Poupardin, Rodolphe; Moos, Martin; Šimek, Petr

    2016-08-01

    The fruit fly Drosophila melanogaster is an insect of tropical origin. Its larval stage is evolutionarily adapted for rapid growth and development under warm conditions and shows high sensitivity to cold. In this study, we further developed an optimal acclimation and freezing protocol that significantly improves larval freeze tolerance (an ability to survive at -5°C when most of the freezable fraction of water is converted to ice). Using the optimal protocol, freeze survival to adult stage increased from 0.7% to 12.6% in the larvae fed standard diet (agar, sugar, yeast, cornmeal). Next, we fed the larvae diets augmented with 31 different amino compounds, administered in different concentrations, and observed their effects on larval metabolomic composition, viability, rate of development and freeze tolerance. While some diet additives were toxic, others showed positive effects on freeze tolerance. Statistical correlation revealed tight association between high freeze tolerance and high levels of amino compounds involved in arginine and proline metabolism. Proline- and arginine-augmented diets showed the highest potential, improving freeze survival to 42.1% and 50.6%, respectively. Two plausible mechanisms by which high concentrations of proline and arginine might stimulate high freeze tolerance are discussed: (i) proline, probably in combination with trehalose, could reduce partial unfolding of proteins and prevent membrane fusions in the larvae exposed to thermal stress (prior to freezing) or during freeze dehydration; (ii) both arginine and proline are exceptional among amino compounds in their ability to form supramolecular aggregates which probably bind partially unfolded proteins and inhibit their aggregation under increasing freeze dehydration. © 2016. Published by The Company of Biologists Ltd.

  15. Projecting potential adoption of genetically engineered freeze-tolerant Eucalyptus in the United States

    Treesearch

    David N. Wear; Ernest Dixon IV; Robert C. Abt; Navinder Singh

    2015-01-01

    Development of commercial Eucalyptus plantations has been limited in the United States because of the species’ sensitivity to freezing temperatures. Recently developed genetically engineered clones of a Eucalyptus hybrid, which confer freeze tolerance, could expand the range of commercial plantations. This study explores how...

  16. Population genetics of freeze tolerance among natural populations of Populus balsamifera across the growing season.

    PubMed

    Menon, Mitra; Barnes, William J; Olson, Matthew S

    2015-08-01

    Protection against freeze damage during the growing season influences the northern range limits of plants. Freeze tolerance and freeze avoidance are the two major freeze resistance strategies. Winter survival strategies have been extensively studied in perennials, but few have addressed them and their genetic basis during the growing season. We examined intraspecific phenotypic variation in freeze resistance of Populus balsamifera across latitude and the growing season. To investigate the molecular basis of this variation, we surveyed nucleotide diversity and examined patterns of gene expression in the poplar C-repeat binding factor (CBF) gene family. Foliar freeze tolerance exhibited latitudinal and seasonal variation indicative of natural genotypic variation. CBF6 showed signatures of recent selective sweep. Of the 46 SNPs surveyed across the six CBF homologs, only CBF2_619 exhibited latitudinal differences consistent with increased freeze tolerance in the north. All six CBF genes were cold inducible, but showed varying patterns of expression across the growing season. Some Poplar CBF homologs exhibited patterns consistent with historical selection and clinal variation in freeze tolerance documented here. However, the CBF genes accounted for only a small amount of the variation, indicating that other genes in this and other molecular pathways likely play significant roles in nature.

  17. Mapping salinity tolerance during Arabidopsis thaliana germination and seedling growth.

    PubMed

    DeRose-Wilson, Leah; Gaut, Brandon S

    2011-01-01

    To characterize and dissect genetic variation for salinity tolerance, we assessed variation in salinity tolerance during germination and seedling growth for a worldwide sample of Arabidopsis thaliana accessions. By combining QTL mapping, association mapping and expression data, we identified genomic regions involved in salinity response. Among the worldwide sample, we found germination ability within a moderately saline environment (150 mM NaCl) varied considerable, from >90% among the most tolerant lines to complete inability to germinate among the most susceptible. Our results also demonstrated wide variation in salinity tolerance within A. thaliana RIL populations and identified multiple genomic regions that contribute to this variation. These regions contain known candidate genes, but at least four of the regions contain loci not yet associated with salinity tolerance response phenotypes. Our observations suggest A. thaliana natural variation may be an underutilized resource for investigating salinity stress response.

  18. A proteome analysis of freezing tolerance in red clover (Trifolium pratense L.).

    PubMed

    Bertrand, Annick; Bipfubusa, Marie; Castonguay, Yves; Rocher, Solen; Szopinska-Morawska, Aleksandra; Papadopoulos, Yousef; Renaut, Jenny

    2016-03-10

    Improvement of freezing tolerance of red clover (Trifolium pratense L.) would increase its persistence under cold climate. In this study, we assessed the freezing tolerance and compared the proteome composition of non-acclimated and cold-acclimated plants of two initial cultivars of red clover: Endure (E-TF0) and Christie (C-TF0) and of populations issued from these cultivars after three (TF3) and four (TF4) cycles of phenotypic recurrent selection for superior freezing tolerance. Through this approach, we wanted to identify proteins that are associated with the improvement of freezing tolerance in red clover. Freezing tolerance expressed as the lethal temperature for 50 % of the plants (LT50) increased markedly from approximately -2 to -16 °C following cold acclimation. Recurrent selection allowed a significant 2 to 3 °C increase of the LT50 after four cycles of recurrent selection. Two-dimensional difference gel electrophoresis (2D-DIGE) was used to study variations in protein abundance. Principal component analysis based on 2D-DIGE revealed that the largest variability in the protein data set was attributable to the cold acclimation treatment and that the two genetic backgrounds had differential protein composition in the acclimated state only. Vegetative storage proteins (VSP), which are essential nitrogen reserves for plant regrowth, and dehydrins were among the most striking changes in proteome composition of cold acclimated crowns of red clovers. A subset of proteins varied in abundance in response to selection including a dehydrin that increased in abundance in TF3 and TF4 populations as compared to TF0 in the Endure background. Recurrent selection performed indoor is an effective approach to improve the freezing tolerance of red clover. Significant improvement of freezing tolerance by recurrent selection was associated with differential accumulation of a small number of cold-regulated proteins that may play an important role in the determination of

  19. DEAR1, a transcriptional repressor of DREB protein that mediates plant defense and freezing stress responses in Arabidopsis.

    PubMed

    Tsutsui, Tomokazu; Kato, Wataru; Asada, Yutaka; Sako, Kaori; Sato, Takeo; Sonoda, Yutaka; Kidokoro, Satoshi; Yamaguchi-Shinozaki, Kazuko; Tamaoki, Masanori; Arakawa, Keita; Ichikawa, Takanari; Nakazawa, Miki; Seki, Motoaki; Shinozaki, Kazuo; Matsui, Minami; Ikeda, Akira; Yamaguchi, Junji

    2009-11-01

    Plants have evolved intricate mechanisms to respond and adapt to a wide variety of biotic and abiotic stresses in their environment. The Arabidopsis DEAR1 (DREB and EAR motif protein 1; At3g50260) gene encodes a protein containing significant homology to the DREB1/CBF (dehydration-responsive element binding protein 1/C-repeat binding factor) domain and the EAR (ethylene response factor-associated amphiphilic repression) motif. We show here that DEAR1 mRNA accumulates in response to both pathogen infection and cold treatment. Transgenic Arabidopsis overexpressing DEAR1 (DEAR1ox) showed a dwarf phenotype and lesion-like cell death, together with constitutive expression of PR genes and accumulation of salicylic acid. DEAR1ox also showed more limited P. syringae pathogen growth compared to wild-type, consistent with an activated defense phenotype. In addition, transient expression experiments revealed that the DEAR1 protein represses DRE/CRT (dehydration-responsive element/C-repeat)-dependent transcription, which is regulated by low temperature. Furthermore, the induction of DREB1/CBF family genes by cold treatment was suppressed in DEAR1ox, leading to a reduction in freezing tolerance. These results suggest that DEAR1 has an upstream regulatory role in mediating crosstalk between signaling pathways for biotic and abiotic stress responses.

  20. Hepatic changes in the freeze-tolerant turtle Chrysemys picta marginata in response to freezing and thawing.

    PubMed

    Hemmings, S J; Storey, K B

    2000-09-01

    Select hepatic changes in the freeze-tolerant hatchling turtle, Chrysemys picta marginata, were studied in response to freezing at -2.5 degrees C and thawing. Upon freezing, a small, selective increase in the liver weight with no increase in body weight was seen suggestive of an hepatic capacitance response. In all turtles studies, lobular differences in the hepatic content of glycogen were evident: the smaller lobe contained twice as much glycogen as the larger lobe. The response to freezing and thawing was comparable. Total hepatic glycogen levels of turtles were reduced approximately 60 per cent from control levels in the frozen state and recovered to >80 per cent of control levels in the thawed state. Compared to the control state, turtle blood glucose levels were: unchanged after 12 h in the cool state; reduced 28 per cent after 24 h and increased two-fold after 48 h in the frozen state; and increased 4.5-fold in the thawed state. Thus, changes in hepatic glycogen metabolism occur without large changes in blood glucose levels. In turtle liver plasma membranes, the hepatic alpha(1)-adrenergic receptor was barely detectable and did not change. The beta(2)-adrenergic receptor was expressed at high levels and, compared to control levels, was: unchanged after 12 h in the cool state; reduced 20 per cent after 24 h and 40 per cent after 48 h in the frozen state. On thawing, this receptor was 50 per cent of control levels. While catecholamines working through the beta(2)-adrenergic receptor may effect early hepatic glycogen breakdown in response to freezing, other factors must be involved to complete the process. The plasma membrane-bound enzyme gamma-glutamyltranspeptidase displayed a different pattern of changes indicative of selective modulation: it was increased 2.7-fold over control levels in the cool state; unchanged in the frozen state; and increased 1.8-fold in the thawed state. The activity of the kidney enzyme was decreased in the cool state and slightly

  1. Overexpression of a Panax ginseng tonoplast aquaporin alters salt tolerance, drought tolerance and cold acclimation ability in transgenic Arabidopsis plants.

    PubMed

    Peng, Yanhui; Lin, Wuling; Cai, Weiming; Arora, Rajeev

    2007-08-01

    Water movement across cellular membranes is regulated largely by a family of water channel proteins called aquaporins (AQPs). Since several abiotic stresses such as, drought, salinity and freezing, manifest themselves via altering water status of plant cells and are linked by the fact that they all result in cellular dehydration, we overexpressed an AQP (tonoplast intrinsic protein) from Panax ginseng, PgTIP1, in transgenic Arabidopsis thaliana plants to test its role in plant's response to drought, salinity and cold acclimation (induced freezing tolerance). Under favorable conditions, PgTIP1 overexpression significantly increased plant growth as determined by the biomass production, and leaf and root morphology. PgTIP1 overexpression had beneficial effect on salt-stress tolerance as indicated by superior growth status and seed germination of transgenic plants under salt stress; shoots of salt-stressed transgenic plants also accumulated greater amounts of Na(+) compared to wild-type plants. Whereas PgTIP1 overexpression diminished the water-deficit tolerance of plants grown in shallow (10 cm deep) pots, the transgenic plants were significantly more tolerant to water stress when grown in 45 cm deep pots. The rationale for this contrasting response, apparently, comes from the differences in the root morphology and leaf water channel activity (speed of dehydration/rehydration) between the transgenic and wild-type plants. Plants overexpressed with PgTIP1 exhibited lower (relative to wild-type control) cold acclimation ability; however, this response was independent of cold-regulated gene expression. Our results demonstrate a significant function of PgTIP1 in growth and development of plant cells, and suggest that the water movement across tonoplast (via AQP) represents a rate-limiting factor for plant vigor under favorable growth conditions and also significantly affect responses of plant to drought, salt and cold stresses.

  2. The Effect of Abscisic Acid on the Freezing Tolerance of Callus Cultures of Lotus corniculatus L.

    PubMed

    Keith, C N; McKersie, B D

    1986-03-01

    The effects of growth temperature (2 degrees C and 24 degrees C), abscisic acid (ABA) concentration, duration of exposure to ABA, and light were assessed for their ability to induce acclimation to freezing temperatures in callus cultures of Lotus corniculatus L. cv Leo, a perennial forage legume. The maximal expression of freezing tolerance was achieved on B(5) media containing 10(-5) molar ABA, at 24 degrees C for 7 or 14 days. Under these culture conditions, the freezing tolerance of the callus approximated that observed in field grown plants. In contrast, low temperatures (2 degrees C) induced only a limited degree of freezing tolerance in these cultures. Viability was assessed by tetrazolium reduction and by regrowth of the callus. The two assays often differed in their estimates of absolute freezing tolerance. Regression analysis of the temperature profile suggested that there may be two or more distinct populations of cells differing in freezing tolerance, which may have contributed to the variability between viability assays.

  3. Genomic Regions Associated with Tolerance to Freezing Stress and Snow Mold in Winter Wheat.

    PubMed

    Kruse, Erika B; Carle, Scott W; Wen, Nuan; Skinner, Daniel Z; Murray, Timothy D; Garland-Campbell, Kimberly A; Carter, Arron H

    2017-03-10

    Plants grown through the winter are subject to selective pressures that vary with each year's unique conditions, necessitating tolerance of numerous abiotic and biotic stress factors. The objective of this study was to identify molecular markers in winter wheat (Triticum aestivum L.) associated with tolerance of two of these stresses, freezing temperatures and snow mold-a fungal disease complex active under snow cover. A population of 155 F2:5 recombinant inbred lines from a cross between soft white wheat cultivars "Finch" and "Eltan" was evaluated for snow mold tolerance in the field, and for freezing tolerance under controlled conditions. A total of 663 molecular markers was used to construct a genetic linkage map and identify marker-trait associations. One quantitative trait locus (QTL) associated with both freezing and snow mold tolerance was identified on chromosome 5A. A second, distinct, QTL associated with freezing tolerance also was found on 5A, and a third on 4B. A second QTL associated with snow mold tolerance was identified on chromosome 6B. The QTL on 5A associated with both traits was closely linked with the Fr-A2 (Frost-Resistance A2) locus; its significant association with both traits may have resulted from pleiotropic effects, or from greater low temperature tolerance enabling the plants to better defend against snow mold pathogens. The QTL on 4B associated with freezing tolerance, and the QTL on 6B associated with snow mold tolerance have not been reported previously, and may be useful in the identification of sources of tolerance for these traits. Copyright © 2017 Kruse et al.

  4. Genomic Regions Associated with Tolerance to Freezing Stress and Snow Mold in Winter Wheat

    PubMed Central

    Kruse, Erika B.; Carle, Scott W.; Wen, Nuan; Skinner, Daniel Z.; Murray, Timothy D.; Garland-Campbell, Kimberly A.; Carter, Arron H.

    2017-01-01

    Plants grown through the winter are subject to selective pressures that vary with each year’s unique conditions, necessitating tolerance of numerous abiotic and biotic stress factors. The objective of this study was to identify molecular markers in winter wheat (Triticum aestivum L.) associated with tolerance of two of these stresses, freezing temperatures and snow mold—a fungal disease complex active under snow cover. A population of 155 F2:5 recombinant inbred lines from a cross between soft white wheat cultivars “Finch” and “Eltan” was evaluated for snow mold tolerance in the field, and for freezing tolerance under controlled conditions. A total of 663 molecular markers was used to construct a genetic linkage map and identify marker-trait associations. One quantitative trait locus (QTL) associated with both freezing and snow mold tolerance was identified on chromosome 5A. A second, distinct, QTL associated with freezing tolerance also was found on 5A, and a third on 4B. A second QTL associated with snow mold tolerance was identified on chromosome 6B. The QTL on 5A associated with both traits was closely linked with the Fr-A2 (Frost-Resistance A2) locus; its significant association with both traits may have resulted from pleiotropic effects, or from greater low temperature tolerance enabling the plants to better defend against snow mold pathogens. The QTL on 4B associated with freezing tolerance, and the QTL on 6B associated with snow mold tolerance have not been reported previously, and may be useful in the identification of sources of tolerance for these traits. PMID:28143950

  5. Population Structure, Genetic Variation, and Linkage Disequilibrium in Perennial Ryegrass Populations Divergently Selected for Freezing Tolerance

    PubMed Central

    Kovi, Mallikarjuna Rao; Fjellheim, Siri; Sandve, Simen R.; Larsen, Arild; Rudi, Heidi; Asp, Torben; Kent, Matthew Peter; Rognli, Odd Arne

    2015-01-01

    Low temperature is one of the abiotic stresses seriously affecting the growth of perennial ryegrass (Lolium perenne L.), and freezing tolerance is a complex trait of major agronomical importance in northern and central Europe. Understanding the genetic control of freezing tolerance would aid in the development of cultivars of perennial ryegrass with improved adaptation to frost. The plant material investigated in this study was an experimental synthetic population derived from pair-crosses among five European perennial ryegrass genotypes, representing adaptations to a range of climatic conditions across Europe. A total number of 80 individuals (24 of High frost [HF]; 29 of Low frost [LF], and 27 of Unselected [US]) from the second generation of the two divergently selected populations and an unselected (US) control population were genotyped using 278 genome-wide SNPs derived from perennial ryegrass transcriptome sequences. Our studies investigated the genetic diversity among the three experimental populations by analysis of molecular variance and population structure, and determined that the HF and LF populations are very divergent after selection for freezing tolerance, whereas the HF and US populations are more similar. Linkage disequilibrium (LD) decay varied across the seven chromosomes and the conspicuous pattern of LD between the HF and LF population confirmed their divergence in freezing tolerance. Furthermore, two Fst outlier methods; finite island model (fdist) by LOSITAN and hierarchical structure model using ARLEQUIN, both detected six loci under directional selection. These outlier loci are most probably linked to genes involved in freezing tolerance, cold adaptation, and abiotic stress. These six candidate loci under directional selection for freezing tolerance might be potential marker resources for breeding perennial ryegrass cultivars with improved freezing tolerance. PMID:26617611

  6. Freezing and desiccation tolerance in entomopathogenic nematodes: diversity and correlation of traits.

    PubMed

    Shapiro-Ilan, David I; Brown, Ian; Lewis, Edwin E

    2014-03-01

    The ability of entomopathogenic nematodes to tolerate environmental stress such as desiccating or freezing conditions, can contribute significantly to biocontrol efficacy. Thus, in selecting which nematode to use in a particular biocontrol program, it is important to be able to predict which strain or species to use in target areas where environmental stress is expected. Our objectives were to (i) compare inter- and intraspecific variation in freeze and desiccation tolerance among a broad array of entomopathogenic nematodes, and (ii) determine if freeze and desiccation tolerance are correlated. In laboratory studies we compared nematodes at two levels of relative humidity (RH) (97% and 85%) and exposure periods (24 and 48 h), and nematodes were exposed to freezing temperatures (-2°C) for 6 or 24 h. To assess interspecific variation, we compared ten species including seven that are of current or recent commercial interest: Heterorhabditis bacteriophora (VS), H. floridensis, H. georgiana, (Kesha), H. indica (HOM1), H. megidis (UK211), Steinernema carpocapsae (All), S. feltiae (SN), S. glaseri (VS), S. rarum (17C&E), and S. riobrave (355). To assess intraspecific variation we compared five strains of H. bacteriophora (Baine, Fl1-1, Hb, Oswego, and VS) and four strains of S. carpocapsae (All, Cxrd, DD136, and Sal), and S. riobrave (355, 38b, 7-12, and TP). S. carpocapsae exhibited the highest level of desiccation tolerance among species followed by S. feltiae and S. rarum; the heterorhabditid species exhibited the least desiccation tolerance and S. riobrave and S. glaseri were intermediate. No intraspecific variation was observed in desiccation tolerance; S. carpocapsae strains showed higher tolerance than all H. bacteriophora or S. riobrave strains yet there was no difference detected within species. In interspecies comparisons, poor freeze tolerance was observed in H. indica, and S. glaseri, S. rarum, and S. riobrave whereas H. georgiana and S. feltiae exhibited the

  7. Freezing and Desiccation Tolerance in Entomopathogenic Nematodes: Diversity and Correlation of Traits

    PubMed Central

    Shapiro-Ilan, David I.; Brown, Ian; Lewis, Edwin E.

    2014-01-01

    The ability of entomopathogenic nematodes to tolerate environmental stress such as desiccating or freezing conditions, can contribute significantly to biocontrol efficacy. Thus, in selecting which nematode to use in a particular biocontrol program, it is important to be able to predict which strain or species to use in target areas where environmental stress is expected. Our objectives were to (i) compare inter- and intraspecific variation in freeze and desiccation tolerance among a broad array of entomopathogenic nematodes, and (ii) determine if freeze and desiccation tolerance are correlated. In laboratory studies we compared nematodes at two levels of relative humidity (RH) (97% and 85%) and exposure periods (24 and 48 h), and nematodes were exposed to freezing temperatures (-2°C) for 6 or 24 h. To assess interspecific variation, we compared ten species including seven that are of current or recent commercial interest: Heterorhabditis bacteriophora (VS), H. floridensis, H. georgiana, (Kesha), H. indica (HOM1), H. megidis (UK211), Steinernema carpocapsae (All), S. feltiae (SN), S. glaseri (VS), S. rarum (17C&E), and S. riobrave (355). To assess intraspecific variation we compared five strains of H. bacteriophora (Baine, Fl1-1, Hb, Oswego, and VS) and four strains of S. carpocapsae (All, Cxrd, DD136, and Sal), and S. riobrave (355, 38b, 7-12, and TP). S. carpocapsae exhibited the highest level of desiccation tolerance among species followed by S. feltiae and S. rarum; the heterorhabditid species exhibited the least desiccation tolerance and S. riobrave and S. glaseri were intermediate. No intraspecific variation was observed in desiccation tolerance; S. carpocapsae strains showed higher tolerance than all H. bacteriophora or S. riobrave strains yet there was no difference detected within species. In interspecies comparisons, poor freeze tolerance was observed in H. indica, and S. glaseri, S. rarum, and S. riobrave whereas H. georgiana and S. feltiae exhibited the

  8. Higher daytime leaf temperatures contribute to lower freeze tolerance under elevated CO2.

    PubMed

    Loveys, Beth R; Egerton, John J G; Ball, Marilyn C

    2006-06-01

    Elevated atmospheric CO2 adversely affects freezing tolerance in many evergreens, but the underlying mechanism(s) have been elusive. We compared effects of elevated CO2 with those of daytime warming on acclimation of snow gum (Eucalyptus pauciflora) to freezing temperatures under field conditions. Reduction in stomatal conductance g(c) under elevated CO2 was shown to cause leaf temperature to increase by up to 3 degrees C. In this study, this increase in leaf temperature was simulated under ambient CO2 conditions by using a free air temperature increase (FATI) system to warm snow gum leaves during daytime, thereby increasing the diurnal range in temperature without affecting temperature minima. Acclimation to freezing temperatures was assessed using measures of electrolyte leakage and photosynthetic efficiency of leaf discs exposed to different nadir temperatures. Here, we show that both elevated CO2 and daytime warming delayed acclimation to freezing temperatures for 2-3 weeks after which time freeze tolerance of the treated plants in both the FATI and open top chamber (OTC) experiments did not differ from control plants. Our results support the hypothesis that delayed development of freezing tolerance under elevated CO2 is because of higher daytime leaf temperatures under elevated CO2. Thus, potential gains in productivity in response to increasing atmospheric CO2 and prolonging the growing season may be reduced by an increase in freezing stress in frost-prone area.

  9. Lipid composition of commercial bakers' yeasts having different freeze-tolerance in frozen dough.

    PubMed

    Murakami, Y; Yokoigawa, K; Kawai, F; Kawai, H

    1996-11-01

    The lipid composition of some commercial bakers' yeasts having different freeze-sensitivity in frozen dough was investigated to clarify the correlation between their lipid composition and freeze-tolerance. The total lipid content including neutral lipid, free fatty acid, sterol, and phospholipid ranged between 23.0 to 32.2 mg/100 mg protein of the yeasts tested. Phosphatidylcholine, phosphatidylethanolamine, phosphatidylinositol, and phosphatidylserine were the main phospholipids found in all yeast strains, but no distinct difference in these components between freeze-tolerant and freeze-sensitive strains was observed. Palmitoleic (C16:1), oleic (C18:1), palmitic (16:0), and stearic (C18:0) acids were the major fatty acids present in total lipid and phospholipid, and unsaturation indices of fatty acid in these lipid components were almost equal by the strains. The molar ratios of sterol to phospholipid of freeze-sensitive strains were higher than those of freeze-tolerant strains. The difference in the sterol-phospholipid ratio that influences the fluidity of plasma membranes in yeast cells was supposed to reflect the difference in freeze-sensitivity of bakers' yeast.

  10. Glucose concentration regulates freeze tolerance in the wood frog Rana sylvatica.

    PubMed

    Costanzo, J P; Lee, R E; Lortz, P H

    1993-08-01

    In spring, the lowest temperature during freezing that can be survived by wood frogs (Rana sylvatica) from southern Ohio is approximately -3 degrees C. We investigated whether the thermal limit of freeze tolerance in these frogs is regulated by tissue levels of glucose, a putative cryoprotectant that is distributed to tissues during freezing. Frogs receiving exogenous glucose injections prior to freezing showed dose-dependent increases in glucose within the heart, liver, skeletal muscle and blood. Tissue glucose concentrations were further elevated during freezing by the production of endogenous glucose. Most glucose-loaded frogs survived freezing to -5 degrees C, whereas all control (saline-injected) frogs succumbed. Further, we investigated some mechanisms by which glucose might function as a cryoprotectant in R. sylvatica. Organ dehydration, a normal, beneficial response that reduces freezing injury to tissues, occurred independently of tissue glucose concentrations. However, elevated glucose levels reduced both body ice content and in vivo erythrocyte injury. These results not only provided conclusive evidence for glucose's cryoprotective role in R. sylvatica, but also revealed that tissue glucose level is a critical determinant of freeze tolerance capacity in this species.

  11. Improving the freeze tolerance of bakers' yeast by loading with trehalose.

    PubMed

    Hirasawa, R; Yokoigawa, K; Isobe, Y; Kawai, H

    2001-03-01

    We examined the freeze tolerance of bakers' yeast loaded with exogenous trehalose. Freeze-tolerant and freeze-sensitive compressed bakers' yeast samples were soaked at several temperatures in 0.5 M and 1 M trehalose and analyzed. The intracellular trehalose contents in both types of bakers' yeast increased with increasing soaking period. The initial trehalose-accumulation rate increased with increasing exogenous trehalose concentration and soaking temperature. The maximum trehalose content was almost identical (200-250 mg/g of dry cells) irrespective of the soaking temperature and the type of bakers' yeast, but depended on the exogenous trehalose concentration. The leavening ability of both types of bakers' yeast loaded with trehalose was almost identical to that of the respective original cells, irrespective of the soaking conditions. The freeze-tolerant ratio (FTR) of both types of bakers' yeast increased with increasing intracellular trehalose content. However, FTR decreased during over-soaking after the maximum amount of trehalose had accumulated. FTR of the freeze-sensitive bakers' yeast was more efficiently improved than that of the freeze-tolerant type.

  12. Freezing tolerance and water relations of Opuntia fragilis from Canada and the United States

    SciTech Connect

    Loik, M.E.; Nobel, P.S. )

    1993-09-01

    To investigate the influence of winter climate on freezing tolerance at the population level, minimum January air temperatures in the field and cold acclimation determined in the laboratory were compared for Opuntia fragilis. Populations occurred at 20 locations as far north as 56[degrees]46' N latitude and at elevations up to 3029 m in Canada and the United States, most of which experience extreme freezing temperatures each winter. Low-temperature responses and water relations of stems were examined in the laboratory at day/night air temperatures of 25[degrees]/15[degrees]C and 14 d after the plants were shifted to a 5[degrees]/[minus]5[degrees]C temperature cycle. Cold acclimation averaged 17[degrees]C and freezing tolerance averaged [minus]29[degrees]C for the 20 populations following a shift to low day/night air temperatures, indicating that O. fragilis has the greatest cold acclimation ability and the greatest freezing tolerance reported for any cactus. Moreover, freezing tolerance and cold acclimation were both positively correlated (r[sup 2] [congruent] 0.7) with the minimum temperatures at the 20 locations. Plants lost water during low-temperature acclimation, leading to 30% decreases in cladode and chlorenchyma thickness; the decrease in water content was greater for the five warmest populations than for the five coldest ones. Over the same period, the average osmotic pressure of the chlorenchyma increased from 1.42 to 1.64 MPa, and the relative water content (RWC) decreased from 0.58 to 0.49, but the average osmotic pressure of saturated chlorenchyma was unchanged, indicating no net change in solute content during acclimation. Although the role of water relations in freezing tolerance is unclear, the substantial freezing tolerance and cold acclimation ability of O. fragilis leads to its distribution into regions of Canada and the United States that experience minimum temperatures below [minus]40[degrees]C during the winter. 47 refs., 3 figs., 5 tabs.

  13. The relationship of freeze tolerance with intracellular compounds in baker's yeasts.

    PubMed

    Shi, Xiaojian; Miao, Yelian; Chen, Jie Yu; Chen, Jun; Li, Wenli; He, Xun; Wang, Jining

    2014-03-01

    Freeze-tolerant baker's yeasts are required for the processing of frozen doughs. The present study was carried out to investigate the cell survival rate after frozen storage and the change of fermentability in dough due to frozen storage, and to discuss quantitatively the relationship of freeze tolerance with intracellular trehalose, amino acids, and glycerol, using six types of baker's yeasts as the test materials. The experimental results showed that the fermentability of yeast cells in frozen dough was strongly correlated with the cell survival rate. The baker's yeast with a higher level of cell survival rate had a larger increase in the total intracellular compound content after frozen storage, and the cell survival rate increased linearly with increasing total intracellular compound content in frozen yeast cells. Trehalose was a primary compound affecting freeze tolerance, followed by glutamic acid, arginine, proline, asparagic acid, and glycerol. The basic information provided by the present study is useful for exploring the freeze-tolerance mechanisms of baker's yeast cells, breeding better freeze-tolerant baker's yeast strains, and developing more effective cryoprotectants.

  14. Habitat-Associated Life History and Stress-Tolerance Variation in Arabidopsis arenosa1[OPEN

    PubMed Central

    Baduel, Pierre; Arnold, Brian; Weisman, Cara M.; Hunter, Ben

    2016-01-01

    Weediness in ephemeral plants is commonly characterized by rapid cycling, prolific all-in flowering, and loss of perenniality. Many species made transitions to weediness of this sort, which can be advantageous in high-disturbance or human-associated habitats. The molecular basis of this shift, however, remains mostly mysterious. Here, we use transcriptome sequencing, genome resequencing scans for selection, and stress tolerance assays to study a weedy population of the otherwise nonweedy Arabidopsis arenosa, an obligately outbreeding relative of Arabidopsis thaliana. Although weedy A. arenosa is widespread, a single genetic lineage colonized railways throughout central and northern Europe. We show that railway plants, in contrast to plants from sheltered outcrops in hill/mountain regions, are rapid cycling, have lost the vernalization requirement, show prolific flowering, and do not return to vegetative growth. Comparing transcriptomes of railway and mountain plants across time courses with and without vernalization, we found that railway plants have sharply abrogated vernalization responsiveness and high constitutive expression of heat- and cold-responsive genes. Railway plants also have strong constitutive heat shock and freezing tolerance compared with mountain plants, where tolerance must be induced. We found 20 genes with good evidence of selection in the railway population. One of these, LATE ELONGATED HYPOCOTYL, is known in A. thaliana to regulate many stress-response genes that we found to be differentially regulated among the distinct habitats. Our data suggest that, beyond life history regulation, other traits like basal stress tolerance also are associated with the evolution of weediness in A. arenosa. PMID:26941193

  15. Identification of Quantitative Trait Loci and a candidate locus for freezing tolerance in controlled and outdoor environments in the overwintering crucifer Boechera stricta

    PubMed Central

    Heo, Jae-Yun; Feng, Dongsheng; Niu, Xiaomu; Mitchell-Olds, Thomas; van Tienderen, Peter H.; Tomes, Dwight; Schranz, M. Eric

    2015-01-01

    Development of chilling and freezing tolerance is complex and can be affected by photoperiod, temperature and photosynthetic performance; however, there has been limited research on the interaction of these three factors. We evaluated 108 recombinant inbred lines of Boechera stricta, derived from a cross between lines originating from Idaho and Colorado, under controlled Long-Day (LD), Short-Day (SD) and in an Outdoor Environment (OE). We measured maximum quantum yield of photosystem II, lethal temperature for 50% survival and electrolyte leakage of leaves. Our results revealed significant variation for chilling and freezing tolerance and photosynthetic performance in different environments. Using both single and multi-trait analyses, three main-effect Quantitative Trait Loci (QTL) were identified. QTL on LG3 were SD-specific, whereas QTL on LG4 were found under both LD and SD. Under all conditions, QTL on LG7 were identified, but were particularly predictive for the Outdoor Experiment. The co-localization of photosynthetic performance and freezing tolerance effects supports these traits being co-regulated. Finally, the major QTL on LG7 is syntenic to the Arabidopsis CBF locus, known regulators of chilling and freezing responses in A. thaliana and other species. PMID:24811132

  16. Chloroplast RNA-Binding Protein RBD1 Promotes Chilling Tolerance through 23S rRNA Processing in Arabidopsis

    PubMed Central

    Yang, Leiyun; Yang, Fen; Wang, Yi; Zhu, Jian-Kang; Hua, Jian

    2016-01-01

    Plants have varying abilities to tolerate chilling (low but not freezing temperatures), and it is largely unknown how plants such as Arabidopsis thaliana achieve chilling tolerance. Here, we describe a genome-wide screen for genes important for chilling tolerance by their putative knockout mutants in Arabidopsis thaliana. Out of 11,000 T-DNA insertion mutant lines representing half of the genome, 54 lines associated with disruption of 49 genes had a drastic chilling sensitive phenotype. Sixteen of these genes encode proteins with chloroplast localization, suggesting a critical role of chloroplast function in chilling tolerance. Study of one of these proteins RBD1 with an RNA binding domain further reveals the importance of chloroplast translation in chilling tolerance. RBD1 is expressed in the green tissues and is localized in the chloroplast nucleoid. It binds directly to 23S rRNA and the binding is stronger under chilling than at normal growth temperatures. The rbd1 mutants are defective in generating mature 23S rRNAs and deficient in chloroplast protein synthesis especially under chilling conditions. Together, our study identifies RBD1 as a regulator of 23S rRNA processing and reveals the importance of chloroplast function especially protein translation in chilling tolerance. PMID:27138552

  17. Hepatocyte responses to in vitro freezing and β-adrenergic stimulation: Insights into the extreme freeze tolerance of subarctic Rana sylvatica.

    PubMed

    do Amaral, M Clara F; Lee, Richard E; Costanzo, Jon P

    2015-02-01

    The wood frog, Rana sylvatica LeConte 1825, is a freeze-tolerant amphibian widely distributed in North America. Subarctic populations of this species can survive experimental freezing to temperatures below -16 °C, whereas temperate populations tolerate freezing only at temperatures above -6 °C. We investigated whether hepatocytes isolated from frogs indigenous to Interior Alaska (subarctic) or southern Ohio (temperate) had distinct characteristics that could contribute to this variation in freeze tolerance capacity. Following in vitro freezing, cell damage, as assessed from lactate dehydrogenase leakage, was similar between samples from Alaskan and Ohioan frogs. Preincubation of cells in media containing glucose or urea, the two primary cryoprotectants used by R. sylvatica, markedly reduced freezing damage to hepatocytes; however, results suggested that cells of the northern phenotype were comparatively more amenable to cryoprotection by urea. Stimulation of isolated hepatocytes with β-adrenergic agonists, which simulates the freezing-induced cryoprotectant mobilization response, gave rates of glucose production from endogenous glycogen reserves that were similar between the populations. Our findings suggest that extreme freeze tolerance in subarctic R. sylvatica does not require an enhanced ability of the liver to resist freezing stress or rapidly mobilize cryoprotectant. © 2015 Wiley Periodicals, Inc.

  18. GBF3 transcription factor imparts drought tolerance in Arabidopsis thaliana.

    PubMed

    Ramegowda, Venkategowda; Gill, Upinder Singh; Sivalingam, Palaiyur Nanjappan; Gupta, Aarti; Gupta, Chirag; Govind, Geetha; Nataraja, Karaba N; Pereira, Andy; Udayakumar, Makarla; Mysore, Kirankumar S; Senthil-Kumar, Muthappa

    2017-08-22

    Drought transcriptome analysis of finger millet (Eleusine coracana) by cDNA subtraction identified drought responsive genes that have a potential role in drought tolerance. Through virus-induced gene silencing (VIGS) in a related crop species, maize (Zea mays), several genes, including a G-BOX BINDING FACTOR 3 (GBF3) were identified as candidate drought stress response genes and the role of GBF3 in drought tolerance was studied in Arabidopsis thaliana. Overexpression of both EcGBF3 and AtGBF3 in A. thaliana resulted in improved tolerance to osmotic stress, salinity and drought stress in addition to conferring insensitivity to ABA. Conversely, loss of function of this gene increased the sensitivity of A. thaliana plants to drought stress. EcGBF3 transgenic A. thaliana results also suggest that drought tolerance of sensitive plants can be improved by transferring genes from far related crops like finger millet. Our results demonstrate the role of GBF3 in imparting drought tolerance in A. thaliana and indicate the conserved role of this gene in drought and other abiotic stress tolerance in several plant species.

  19. The interaction between freezing tolerance and phenology in temperate deciduous trees

    PubMed Central

    Vitasse, Yann; Lenz, Armando; Körner, Christian

    2014-01-01

    Temperate climates are defined by distinct temperature seasonality with large and often unpredictable weather during any of the four seasons. To thrive in such climates, trees have to withstand a cold winter and the stochastic occurrence of freeze events during any time of the year. The physiological mechanisms trees adopt to escape, avoid, and tolerate freezing temperatures include a cold acclimation in autumn, a dormancy period during winter (leafless in deciduous trees), and the maintenance of a certain freezing tolerance during dehardening in early spring. The change from one phase to the next is mediated by complex interactions between temperature and photoperiod. This review aims at providing an overview of the interplay between phenology of leaves and species-specific freezing resistance. First, we address the long-term evolutionary responses that enabled temperate trees to tolerate certain low temperature extremes. We provide evidence that short term acclimation of freezing resistance plays a crucial role both in dormant and active buds, including re-acclimation to cold conditions following warm spells. This ability declines to almost zero during leaf emergence. Second, we show that the risk that native temperate trees encounter freeze injuries is low and is confined to spring and underline that this risk might be altered by climate warming depending on species-specific phenological responses to environmental cues. PMID:25346748

  20. Cross-tolerance between osmotic and freeze-thaw stress in microbial assemblages from temperate lakes.

    PubMed

    Wilson, Sandra L; Frazer, Corey; Cumming, Brian F; Nuin, Paulo A S; Walker, Virginia K

    2012-11-01

    Osmotic stress can accompany increases in solute concentrations because of freezing or high-salt environments. Consequently, microorganisms from environments with a high-osmotic potential may exhibit cross-tolerance to freeze stress. To test this hypothesis, enrichments derived from the sediment and water of temperate lakes with a range of salt concentrations were subjected to multiple freeze-thaw cycles. Surviving isolates were identified and metagenomes were sampled prior to and following selection. Enrichments from alkali lakes were typically the most freeze-thaw resistant with only 100-fold losses in cell viability, and those from freshwater lakes were most susceptible, with cell numbers reduced at least 100,000-fold. Metagenomic analysis suggested that selection reduced assemblage diversity more in freshwater samples than in those from saline lakes. Survivors included known psychro-, halo- and alkali-tolerant bacteria. Characterization of freeze-thaw-resistant isolates from brine and alkali lakes showed that few isolates had ice-associating activities such as antifreeze or ice nucleation properties. However, all brine- and alkali-derived isolates had high intracellular levels of osmolytes and/or appeared more likely to form biofilms. Conversely, these phenotypes were infrequent amongst the freshwater-derived isolates. These observations are consistent with microbial cross-tolerance between osmotic and freeze-thaw stresses. © 2012 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.

  1. Adaptive and freeze-tolerant heteronetwork organohydrogels with enhanced mechanical stability over a wide temperature range

    NASA Astrophysics Data System (ADS)

    Gao, Hainan; Zhao, Ziguang; Cai, Yudong; Zhou, Jiajia; Hua, Wenda; Chen, Lie; Wang, Li; Zhang, Jianqi; Han, Dong; Liu, Mingjie; Jiang, Lei

    2017-06-01

    Many biological organisms with exceptional freezing tolerance can resist the damages to cells from extra-/intracellular ice crystals and thus maintain their mechanical stability at subzero temperatures. Inspired by the freezing tolerance mechanisms found in nature, here we report a strategy of combining hydrophilic/oleophilic heteronetworks to produce self-adaptive, freeze-tolerant and mechanically stable organohydrogels. The organohydrogels can simultaneously use water and oil as a dispersion medium, and quickly switch between hydrogel- and organogel-like behaviours in response to the nature of the surrounding phase. Accordingly, their surfaces display unusual adaptive dual superlyophobic in oil/water system (that is, they are superhydrophobic under oil and superoleophobic under water). Moreover, the organogel component can inhibit the ice crystallization of the hydrogel component, thus enhancing the mechanical stability of organohydrogel over a wide temperature range (-78 to 80 °C). The organohydrogels may have promising applications in complex and harsh environments.

  2. Abscisic acid induced freezing tolerance in chilling-sensitive suspension cultures and seedlings of rice

    PubMed Central

    2013-01-01

    Background The role of abscisic acid (ABA) as a possible activator of cold acclimation process was postulated since endogenous levels of ABA increase temporarily or constitutively during cold-hardening. Exogenous application of ABA has been known to induce freezing tolerance at ambient temperatures in in vitro systems derived from cold hardy plants. Yet, some cell cultures acquired much greater freezing tolerance by ABA than by cold whilst maintaining active growth. This raises questions about the relationships among ABA, cold acclimation and growth cessation. To address this question, we attempted to 1) determine whether exogenous ABA can confer freezing tolerance in chilling-sensitive rice suspension cells and seedlings, which obviously lack the mechanisms to acquire freezing tolerance in response to cold; 2) characterize this phenomenon by optimizing the conditions and compare with the case of cold hardy bromegrass cells. Results Non-embryogenic suspension cells of rice suffered serious chilling injury when exposed to 4°C. When incubated with ABA at the optimal conditions (0.5-1 g cell inoculum, 75 μM ABA, 25-30°C, 7–10 days), they survived slow freezing (2°C/h) to −9.0 ~ −9.3°C (LT50: 50% killing temperature) while control cells were mostly injured at −3°C (LT50: -0.5 ~ −1.5°C). Ice-inoculation of the cell suspension at −3°C and survival determination by regrowth confirmed that ABA-treated rice cells survived extracellular freezing at −9°C. ABA-induced freezing tolerance did not require any exposure to cold and was best achieved at 25-30°C where the rice cells maintained high growth even in the presence of ABA. ABA treatment also increased tolerance to heat (43°C) as determined by regrowth. ABA-treated cells tended to have more augmented cytoplasm and/or reduced vacuole sizes compared to control cultures with a concomitant increase in osmolarity and a decrease in water content. ABA-treated (2–7 days) in vitro grown

  3. The intrinsically disordered protein LEA7 from Arabidopsis thaliana protects the isolated enzyme lactate dehydrogenase and enzymes in a soluble leaf proteome during freezing and drying.

    PubMed

    Popova, Antoaneta V; Rausch, Saskia; Hundertmark, Michaela; Gibon, Yves; Hincha, Dirk K

    2015-10-01

    The accumulation of Late Embryogenesis Abundant (LEA) proteins in plants is associated with tolerance against stresses such as freezing and desiccation. Two main functions have been attributed to LEA proteins: membrane stabilization and enzyme protection. We have hypothesized previously that LEA7 from Arabidopsis thaliana may stabilize membranes because it interacts with liposomes in the dry state. Here we show that LEA7, contrary to this expectation, did not stabilize liposomes during drying and rehydration. Instead, it partially preserved the activity of the enzyme lactate dehydrogenase (LDH) during drying and freezing. Fourier-transform infrared (FTIR) spectroscopy showed no evidence of aggregation of LDH in the dry or rehydrated state under conditions that lead to complete loss of activity. To approximate the complex influence of intracellular conditions on the protective effects of a LEA protein in a convenient in-vitro assay, we measured the activity of two Arabidopsis enzymes (glucose-6-P dehydrogenase and ADP-glucose pyrophosphorylase) in total soluble leaf protein extract (Arabidopsis soluble proteome, ASP) after drying and rehydration or freezing and thawing. LEA7 partially preserved the activity of both enzymes under these conditions, suggesting its role as an enzyme protectant in vivo. Further FTIR analyses indicated the partial reversibility of protein aggregation in the dry ASP during rehydration. Similarly, aggregation in the dry ASP was strongly reduced by LEA7. In addition, mixtures of LEA7 with sucrose or verbascose reduced aggregation more than the single additives, presumably through the effects of the protein on the H-bonding network of the sugar glasses.

  4. Low Temperature-Induced Decrease in trans-Δ3-Hexadecenoic Acid Content Is Correlated with Freezing Tolerance in Cereals 1

    PubMed Central

    Huner, Norman P. A.; Williams, John P.; Maissan, Ellen E.; Myscich, Elizabeth G.; Krol, Marianna; Laroche, Andre; Singh, Jasbir

    1989-01-01

    The effect of growth at 5°C on the trans-Δ3-hexadecenoic acid content of phosphatidyl(d)glycerol was examined in a total of eight cultivars of rye (Secale cereale L.) and what (Triticum aestivum L.) of varying freezing tolerance. In these monocots, low temperature growth caused decreases in the trans-Δ3-hexadecenoic acid content of between 0 and 74% with concomitant increases in the palmitic acid content of phosphatidyl(d)glycerol. These trends were observed for whole leaf extracts as well as isolated thylakoids. The low growth temperature-induced decrease in the trans-Δ3-hexadecenoic acid content was shown to be a linear function (r2 = 0.954) of freezing tolerance in these cultivars. Of the six cold tolerant dicotyledonous species examined, only Brassica and Arabidopsis thaliana L. cv Columbia exhibited a 42% and 65% decrease, respectively, in trans-Δ3-hexadecenoic acid content. Thus, the relationship between the change in trans-Δ3-hexadecenoic acid content of phosphatidyl(d)glycerol and freezing tolerance cannot be considered a general one for all cold tolerant plant species. However, species which exhibited a low growth temperature-induced decrease in trans-Δ3-hexadecenoic acid also exhibited a concomitant shift in the in vitro organization of the light harvesting complex II from a predominantly oligomeric form to the monomeric form. We conclude that the proposed role of phosphatidyl(d)glycerol in modulating the organization of light harvesting complex II as a function of growth temperature manifests itself to varying degrees in different plant species. A possible physiological role for this phenomenon with respect to low temperature acclimation and freezing tolerance in cereals is discussed. PMID:16666505

  5. Cryoprotectants and Extreme Freeze Tolerance in a Subarctic Population of the Wood Frog

    PubMed Central

    Costanzo, Jon P.; Reynolds, Alice M.; do Amaral, M. Clara F.; Rosendale, Andrew J.; Lee, Richard E.

    2015-01-01

    Wood frogs (Rana sylvatica) exhibit marked geographic variation in freeze tolerance, with subarctic populations tolerating experimental freezing to temperatures at least 10-13 degrees Celsius below the lethal limits for conspecifics from more temperate locales. We determined how seasonal responses enhance the cryoprotectant system in these northern frogs, and also investigated their physiological responses to somatic freezing at extreme temperatures. Alaskan frogs collected in late summer had plasma urea levels near 10 μmol ml-1, but this level rose during preparation for winter to 85.5 ± 2.9 μmol ml-1 (mean ± SEM) in frogs that remained fully hydrated, and to 186.9 ± 12.4 μmol ml-1 in frogs held under a restricted moisture regime. An osmolality gap indicated that the plasma of winter-conditioned frogs contained an as yet unidentified osmolyte(s) that contributed about 75 mOsmol kg-1 to total osmotic pressure. Experimental freezing to –8°C, either directly or following three cycles of freezing/thawing between –4 and 0°C, or –16°C increased the liver’s synthesis of glucose and, to a lesser extent, urea. Concomitantly, organs shed up to one-half (skeletal muscle) or two-thirds (liver) of their water, with cryoprotectant in the remaining fluid reaching concentrations as high as 0.2 and 2.1 M, respectively. Freeze/thaw cycling, which was readily survived by winter-conditioned frogs, greatly increased hepatic glycogenolysis and delivery of glucose (but not urea) to skeletal muscle. We conclude that cryoprotectant accrual in anticipation of and in response to freezing have been greatly enhanced and contribute to extreme freeze tolerance in northern R. sylvatica. PMID:25688861

  6. Cryoprotectants and extreme freeze tolerance in a subarctic population of the wood frog.

    PubMed

    Costanzo, Jon P; Reynolds, Alice M; do Amaral, M Clara F; Rosendale, Andrew J; Lee, Richard E

    2015-01-01

    Wood frogs (Rana sylvatica) exhibit marked geographic variation in freeze tolerance, with subarctic populations tolerating experimental freezing to temperatures at least 10-13 degrees Celsius below the lethal limits for conspecifics from more temperate locales. We determined how seasonal responses enhance the cryoprotectant system in these northern frogs, and also investigated their physiological responses to somatic freezing at extreme temperatures. Alaskan frogs collected in late summer had plasma urea levels near 10 μmol ml-1, but this level rose during preparation for winter to 85.5 ± 2.9 μmol ml-1 (mean ± SEM) in frogs that remained fully hydrated, and to 186.9 ± 12.4 μmol ml-1 in frogs held under a restricted moisture regime. An osmolality gap indicated that the plasma of winter-conditioned frogs contained an as yet unidentified osmolyte(s) that contributed about 75 mOsmol kg-1 to total osmotic pressure. Experimental freezing to -8°C, either directly or following three cycles of freezing/thawing between -4 and 0°C, or -16°C increased the liver's synthesis of glucose and, to a lesser extent, urea. Concomitantly, organs shed up to one-half (skeletal muscle) or two-thirds (liver) of their water, with cryoprotectant in the remaining fluid reaching concentrations as high as 0.2 and 2.1 M, respectively. Freeze/thaw cycling, which was readily survived by winter-conditioned frogs, greatly increased hepatic glycogenolysis and delivery of glucose (but not urea) to skeletal muscle. We conclude that cryoprotectant accrual in anticipation of and in response to freezing have been greatly enhanced and contribute to extreme freeze tolerance in northern R. sylvatica.

  7. Deciphering the molecular bases for drought tolerance in Arabidopsis autotetraploids.

    PubMed

    del Pozo, Juan C; Ramirez-Parra, Elena

    2014-12-01

    Whole genome duplication (autopolyploidy) is common in many plant species and often leads to better adaptation to adverse environmental conditions. However, little is known about the physiological and molecular mechanisms underlying these adaptations. Drought is one of the major environmental conditions limiting plant growth and development. Here, we report that, in Arabidopsis thaliana, tetraploidy promotes alterations in cell proliferation and organ size in a tissue-dependent manner. Furthermore, it potentiates plant tolerance to salt and drought stresses and decreases transpiration rate, likely through controlling stomata density and closure, abscisic acid (ABA) signalling and reactive oxygen species (ROS) homeostasis. Our transcriptomic analyses revealed that tetraploidy mainly regulates the expression of genes involved in redox homeostasis and ABA and stress response. Taken together, our data have shed light on the molecular basis associated with stress tolerance in autopolyploid plants.

  8. Overexpression of Muscadinia rotundifolia CBF2 gene enhances biotic and abiotic stress tolerance in Arabidopsis.

    PubMed

    Wu, Jiao; Folta, Kevin M; Xie, Yifan; Jiang, Wenming; Lu, Jiang; Zhang, Yali

    2017-01-01

    C-repeat-binding factor dehydration-responsive element-binding factor 1C (CBF2/DREB1C) gene encodes a small family of transcriptional activator that has been described as playing an important role in freezing tolerance and cold acclimation of plants. We here report that CBF2 gene also plays an important role in the early response to the pathogen infection of grapevine downy mildew disease. The expression level of CBF2 increased dramatically and reached a peak at 7 h after infection in immune grapevine Muscadinia rotundifolia 'Noble', which was much faster than moderate resistant Vitis amurensis 'PI1288' and susceptible Vitis vinifera 'Cabernet Sauvignon'. Muscadinia rotundifolia MrCBF2 exhibited amino acid domains characteristic of Vitis CBF2 proteins with unique features including rich serine repeats and slight differences in NLS, DSAWRL, and AP2 domains. The MrCBF2 gene was introduced to Arabidopsis 'COL0' which are susceptible to downy mildew pathogen. The transgenic lines showed an increased resistance to downy mildew disease and more accumulation of SA as well as higher expression of pathogenesis-related (PR) genes (AtPR1, AtPR4, and AtPR5) as a consequence of MrCBF2 overexpression. Besides, constitutive expression of MrCBF2 enhanced phytohormone abscisic acid (ABA)-independent drought tolerance of transgenic plants. Freezing tolerance of transgenic lines was also enhanced accompanied with an increase in the expression of the cold-regulated genes AtCOR, AtCOR15A, AtKIN1, AtRD29A, and AtSuSy. In addition, the development of MrCBF2-overexpressing plants was seen to be altered and resulted in growth retardation, dwarfism, late flowering, and prone rosette leaves, which may be because of an increase in the gene expression of partial DELLA proteins and DDF1.

  9. Re-Evaluation of Reportedly Metal Tolerant Arabidopsis thaliana Accessions

    PubMed Central

    Silva-Guzman, Macarena; Addo-Quaye, Charles; Dilkes, Brian P.

    2016-01-01

    Santa Clara, Limeport, and Berkeley are Arabidopsis thaliana accessions previously identified as diversely metal resistant. Yet these same accessions were determined to be genetically indistinguishable from the metal sensitive Col-0. We robustly tested tolerance for Zn, Ni and Cu, and genetic relatedness by growing these accessions under a range of Ni, Zn and Cu concentrations for three durations in multiple replicates. Neither metal resistance nor variance in growth were detected between them and Col-0. We re-sequenced the genomes of these accessions and all stocks available for each accession. In all cases they were nearly indistinguishable from the standard laboratory accession Col-0. As Santa Clara was allegedly collected from the Jasper Ridge serpentine outcrop in California, USA we investigated the possibility of extant A. thaliana populations adapted to serpentine soils. Botanically vouchered Arabidopsis accessions in the Jepson database were overlaid with soil maps of California. This provided no evidence of A. thaliana collections from serpentine sites in California. Thus, our work demonstrates that the Santa Clara, Berkeley and Limeport accessions are not metal tolerant, not genetically distinct from Col-0, and that there are no known serpentine adapted populations or accessions of A. thaliana. PMID:27467746

  10. The Vitis vinifera C-repeat binding protein 4 (VvCBF4) transcriptional factor enhances freezing tolerance in wine grape

    PubMed Central

    Tillett, Richard L.; Wheatley, Matthew D.; Tattersall, Elizabeth A.R.; Schlauch, Karen A.; Cramer, Grant R.; Cushman, John C.

    2014-01-01

    Summary Chilling and freezing can reduce significantly vine survival and fruit set in Vitis vinifera wine grape. To overcome such production losses, a recently identified grapevine C-repeat binding factor (CBF) gene, VvCBF4, was overexpressed in grape vine cv. “Freedom” and found to improve freezing survival and reduced freezing-induced electrolyte leakage by up to 2°C in non-cold-acclimated vines. In addition, overexpression of this transgene caused a reduced growth phenotype similar to that observed for CBF overexpression in Arabidopsis and other species. Both freezing tolerance and reduced growth phenotypes were manifested in a transgene dose-dependent manner. To understand the mechanistic basis of VvCBF4 transgene action, one transgenic line (9–12) was genotyped using microarray-based mRNA expression profiling. Forty-seven and 12 genes were identified in unstressed transgenic shoots with either a greater than 1.5-fold increase or decrease in mRNA abundance, respectively. Comparison of mRNA changes with characterized CBF regulons in woody and herbaceous species revealed partial overlaps suggesting that CBF-mediated cold acclimation responses are widely conserved. Putative VvCBF4-regulon targets included genes with functions in cell wall structure, lipid metabolism, epicuticular wax formation, and stress-responses suggesting that the observed cold tolerance and dwarf phenotypes are the result of a complex network of diverse functional determinants. PMID:21914113

  11. The Vitis vinifera C-repeat binding protein 4 (VvCBF4) transcriptional factor enhances freezing tolerance in wine grape.

    PubMed

    Tillett, Richard L; Wheatley, Matthew D; Tattersall, Elizabeth A R; Schlauch, Karen A; Cramer, Grant R; Cushman, John C

    2012-01-01

    Chilling and freezing can reduce significantly vine survival and fruit set in Vitis vinifera wine grape. To overcome such production losses, a recently identified grapevine C-repeat binding factor (CBF) gene, VvCBF4, was overexpressed in grape vine cv. 'Freedom' and found to improve freezing survival and reduced freezing-induced electrolyte leakage by up to 2 °C in non-cold-acclimated vines. In addition, overexpression of this transgene caused a reduced growth phenotype similar to that observed for CBF overexpression in Arabidopsis and other species. Both freezing tolerance and reduced growth phenotypes were manifested in a transgene dose-dependent manner. To understand the mechanistic basis of VvCBF4 transgene action, one transgenic line (9-12) was genotyped using microarray-based mRNA expression profiling. Forty-seven and 12 genes were identified in unstressed transgenic shoots with either a >1.5-fold increase or decrease in mRNA abundance, respectively. Comparison of mRNA changes with characterized CBF regulons in woody and herbaceous species revealed partial overlaps, suggesting that CBF-mediated cold acclimation responses are widely conserved. Putative VvCBF4-regulon targets included genes with functions in cell wall structure, lipid metabolism, epicuticular wax formation and stress-responses suggesting that the observed cold tolerance and dwarf phenotypes are the result of a complex network of diverse functional determinants.

  12. Determining factors for cryoprotectant accumulation in the freeze-tolerant earthworm, Dendrobaena octaedra.

    PubMed

    Overgaard, Johannes; Slotsbo, Stine; Holmstrup, Martin; Bayley, Mark

    2007-10-01

    The freeze-tolerant earthworm Dendrobaena octaedra is found in most of the European forest and tundra, Siberia, North America and Greenland where it over-winters in the top soil and encounters winter frost. In response to freezing this earthworm rapidly synthesises glucose which acts as a cryoprotectant. Frost tolerance varies extensively between geographical populations, and of the populations studied so far, the Finnish worms are most and the Danish worms least frost tolerant. Little is known about the determining factors for glucose synthesis and this study therefore investigated possible roles of acclimation and the cues for synthesis of glucose, in Finnish and Danish worms. The Finnish population had significantly larger glycogen reserves than the Danish during acclimation and in all worms, glucose synthesis was the result of an almost stoichemical reduction in glycogen stores. Maximum glucose levels were reached after the onset of freezing and were significantly higher in Finnish worms where the sugar accounted for as much as 5% of the fresh weight. On average, both the total glycogen phosphorylase activity and the active enzyme pool increased during acclimation in the Finnish but not the Danish populations. However, the increase in this enzyme was only significant during the freezing process. In this study, we show contrary to previous theory that glucose synthesis is initiated before the onset of freezing and that in this species, cryoprotectant synthesis is sensitive to very small temperature changes below 0 degrees C without the presence of ice.

  13. Disruption of the CAR1 gene encoding arginase enhances freeze tolerance of the commercial baker's yeast Saccharomyces cerevisiae.

    PubMed

    Shima, Jun; Sakata-Tsuda, Yuko; Suzuki, Yasuo; Nakajima, Ryouichi; Watanabe, Hajime; Kawamoto, Shinichi; Takano, Hiroyuki

    2003-01-01

    The effect of intracellular charged amino acids on freeze tolerance in dough was determined by constructing homozygous diploid arginase-deficient mutants of commercial baker's yeast. An arginase mutant accumulated higher levels of arginine and/or glutamate and showed increased leavening ability during the frozen-dough baking process, suggesting that disruption of the CAR1 gene enhances freeze tolerance.

  14. Freezing survival and cryoprotective dehydration as cold tolerance mechanisms in the Antarctic nematode Panagrolaimus davidi.

    PubMed

    Wharton, David A; Goodall, Gordon; Marshall, Craig J

    2003-01-01

    The relative importance of freezing tolerance and cryoprotective dehydration in the Antarctic nematode Panagrolaimus davidi has been investigated. If nucleation of the medium is initiated at a high subzero temperature (-1 degree C), the nematodes do not freeze but dehydrate. This effect occurs in deionised water, indicating that the loss of water is driven by the difference in vapour pressure of ice and supercooled water at the same temperature. If the nematodes are held above their nucleation temperature for a sufficient time, or are cooled slowly, enough water is lost to prevent freezing (cryoprotective dehydration). However, if the medium is nucleated at lower temperatures or if the sample is cooled at a faster cooling rate, the nematodes freeze and can survive intracellular ice formation. P. davidi thus has a variety of mechanisms that ensure its survival in its harsh terrestrial Antarctic habitat.

  15. The PSE1 gene modulates lead tolerance in Arabidopsis

    PubMed Central

    Fan, Tingting; Yang, Libo; Wu, Xi; Ni, Jiaojiao; Jiang, Haikun; Zhang, Qi’an; Fang, Ling; Sheng, Yibao; Ren, Yongbing; Cao, Shuqing

    2016-01-01

    Lead (Pb) is a dangerous heavy metal contaminant with high toxicity to plants. However, the regulatory mechanism of plant Pb tolerance is poorly understood. Here, we showed that the PSE1 gene confers Pb tolerance in Arabidopsis. A novel Pb-sensitive mutant pse1-1 (Pb-sensitive1) was isolated by screening T-DNA insertion mutants. PSE1 encodes an unknown protein with an NC domain and was localized in the cytoplasm. PSE1 was induced by Pb stress, and the pse1-1 loss-of-function mutant showed enhanced Pb sensitivity; overexpression of PSE1 resulted in increased Pb tolerance. PSE1-overexpressing plants showed increased Pb accumulation, which was accompanied by the activation of phytochelatin (PC) synthesis and related gene expression. In contrast, the pse1-1 mutant showed reduced Pb accumulation, which was associated with decreased PC synthesis and related gene expression. In addition, the expression of PDR12 was also increased in PSE1-overexpressing plants subjected to Pb stress. Our results suggest that PSE1 regulates Pb tolerance mainly through glutathione-dependent PC synthesis by activating the expression of the genes involved in PC synthesis and at least partially through activating the expression of the ABC transporter PDR12/ABCG40. PMID:27335453

  16. SENSITIVE TO FREEZING2 Aides in Resilience to Salt and Drought in Freezing-Sensitive Tomato.

    PubMed

    Wang, Kun; Hersh, Hope Lynn; Benning, Christoph

    2016-11-01

    SENSITIVE TO FREEZING2 (SFR2) is crucial for protecting chloroplast membranes following freezing in Arabidopsis (Arabidopsis thaliana). It has been shown that SFR2 homologs are present in all land plants, including freezing-sensitive species, raising the question of SFR2 function beyond freezing tolerance. Similar to freezing, salt and drought can cause dehydration. Thus, it is hypothesized that in freezing-sensitive plants SFR2 may play roles in their resilience to salt or drought. To test this hypothesis, SlSFR2 RNAi lines were generated in the cold/freezing-sensitive species tomato (Solanum lycopersicum [M82 cv]). Hypersensitivity to salt and drought of SlSFR2-RNAi lines was observed. Higher tolerance of wild-type tomatoes was correlated with the production of trigalactosyldiacylglycerol, a product of SFR2 activity. Tomato SFR2 in vitro activity is Mg(2+)-dependent and its optimal pH is 7.5, similar to that of Arabidopsis SFR2, but the specific activity of tomato SFR2 in vitro is almost double that of Arabidopsis SFR2. When salt and drought stress were applied to Arabidopsis, no conditions could be identified at which SFR2 was induced prior to irreversibly impacting plant growth, suggesting that SFR2 protects Arabidopsis primarily against freezing. Discovery of tomato SFR2 function in drought and salt resilience provides further insights into general membrane lipid remodeling-based stress tolerance mechanisms and together with protection against freezing in freezing-resistant plants such as Arabidopsis, it adds lipid remodeling as a possible target for the engineering of abiotic stress-resilient crops. © 2016 American Society of Plant Biologists. All Rights Reserved.

  17. Supercooling Point Plasticity During Cold Storage in the Freeze-tolerant Sugarbeet Root Maggot Tetanops myopaeformis

    USDA-ARS?s Scientific Manuscript database

    The sugarbeet root maggot, Tetanops myopaeformis (Röder), overwinters as a freeze-tolerant 3rd instar larva. While most larvae are thought to overwinter for only one year, some may exhibit prolonged diapause in the field. In the laboratory, they can live for over five years using a combination of ...

  18. Freeze tolerance of perennial ryegrass and implications for future species distribution.

    USDA-ARS?s Scientific Manuscript database

    Winter hardiness is one of the factors limiting the use of the palatable and productive cool-season forage grass Lolium perenne L, perennial ryegrass, in the northeastern United States. We performed a screening study to compare freeze tolerance, one component of winter hardiness, among thirteen comm...

  19. Sucrose incubation increases freezing tolerance of asparagus (Asparagus officinalis L.) embryogenic cell suspensions.

    PubMed

    Jitsuyama, Y; Suzuki, T; Harada, T; Fujikawa, S

    2002-01-01

    The freezing tolerance of asparagus (Asparagus officinalis L.) embryogenic cells, as determined by electrolyte leakage, was increased by the incubation of samples in medium containing 0.8 M sucrose. To elucidate the mechanism involved, we investigated the changes in soluble carbohydrates, cell ultrastructure and proteins accompanying the increase in freezing tolerance following incubation in sugar-rich medium. During sugar incubation, the intracellular sucrose content increased from 67 mol g-1FW to 429 mol g-1FW; it was also metabolized into fructose and glucose, as determined by high-performance liquid chromatography. Microscopy revealed that sugar incubation induced plasmolysis of embryogenic cells and drastic changes in cell ultrastructure with the appearance of rough endoplasmic reticulum (rER). Furthermore, immunoblotting analysis with anti-dehydrin antiserum revealed that a dehydrin-like protein appeared only when maximal freezing tolerance was induced by sugar incubation. These results suggest that freezing tolerance of asparagus embryogenic cells is increased by a complex mechanism involving notably changes in cell ultrastructure and accumulation of certain sugars and proteins during sugar incubation.

  20. Physiological and molecular characterization of lucerne (Medicago sativa L.) germplasm with improved seedling freezing tolerance

    USDA-ARS?s Scientific Manuscript database

    We conducted greenhouse experiments to compare 14 alfalfa germplasms for their survival following freezing. Among these germplasms are collections adapted to the Grand River National Grasslands in South Dakota. Our hypothesis was that these collections developed tolerance to survive the frigid gro...

  1. MicroRNA regulation in heart and skeletal muscle over the freeze-thaw cycle in the freeze tolerant wood frog.

    PubMed

    Bansal, Saumya; Luu, Bryan E; Storey, Kenneth B

    2016-02-01

    The North American wood frog, Rana sylvatica, is one of just a few anuran species that tolerates whole body freezing during the winter and has been intensely studied to identify the biochemical adaptations that support freeze tolerance. Among these adaptations is the altered expression of many genes, making freeze-responsive changes to gene regulatory mechanisms a topic of interest. The present study focuses on the potential involvement of microRNAs as one such regulatory mechanism and aims to better understand freeze/thaw stress-induced microRNA responses in the freeze-tolerant wood frog. Using quantitative PCR, relative levels of 53 microRNAs were measured in heart and skeletal muscle of control, 24 h frozen, and 8 h thawed frogs. MicroRNAs showed tissue specific expression patterns: 21 microRNAs decreased in the heart during thawing, whereas 16 microRNAs increased during freezing stress in skeletal muscle. These findings suggest that select genes may be activated and suppressed in heart and skeletal muscle, respectively, in response to freezing. Bioinformatics analysis using the DIANA miRPath program (v.2.0) predicted that the differentially expressed microRNAs may collectively regulate tissue-specific cellular pathways to promote survival of wood frogs undergoing freezing and thawing.

  2. Comparative analysis of the cold acclimation and freezing tolerance capacities of seven diploid Brachypodium distachyon accessions

    PubMed Central

    Colton-Gagnon, Katia; Ali-Benali, Mohamed Ali; Mayer, Boris F.; Dionne, Rachel; Bertrand, Annick; Do Carmo, Sonia; Charron, Jean-Benoit

    2014-01-01

    Background and Aims Cold is a major constraint for cereal cultivation under temperate climates. Winter-hardy plants interpret seasonal changes and can acquire the ability to resist sub-zero temperatures. This cold acclimation process is associated with physiological, biochemical and molecular alterations in cereals. Brachypodium distachyon is considered a powerful model system to study the response of temperate cereals to adverse environmental conditions. To date, little is known about the cold acclimation and freezing tolerance capacities of Brachypodium. The main objective of this study was to evaluate the cold hardiness of seven diploid Brachypodium accessions. Methods An integrated approach, involving monitoring of phenological indicators along with expression profiling of the major vernalization regulator VRN1 orthologue, was followed. In parallel, soluble sugars and proline contents were determined along with expression profiles of two COR genes in plants exposed to low temperatures. Finally, whole-plant freezing tests were performed to evaluate the freezing tolerance capacity of Brachypodium. Key Results Cold treatment accelerated the transition from the vegetative to the reproductive phase in all diploid Brachypodium accessions tested. In addition, low temperature exposure triggered the gradual accumulation of BradiVRN1 transcripts in all accessions tested. These accessions exhibited a clear cold acclimation response by progressively accumulating proline, sugars and COR gene transcripts. However, whole-plant freezing tests revealed that these seven diploid accessions only have a limited capacity to develop freezing tolerance when compared with winter varieties of temperate cereals such as wheat and barley. Furthermore, little difference in terms of survival was observed among the accessions tested despite their previous classification as either spring or winter genotypes. Conclusions This study is the first to characterize the freezing tolerance capacities

  3. Comparative analysis of the cold acclimation and freezing tolerance capacities of seven diploid Brachypodium distachyon accessions.

    PubMed

    Colton-Gagnon, Katia; Ali-Benali, Mohamed Ali; Mayer, Boris F; Dionne, Rachel; Bertrand, Annick; Do Carmo, Sonia; Charron, Jean-Benoit

    2014-03-01

    Cold is a major constraint for cereal cultivation under temperate climates. Winter-hardy plants interpret seasonal changes and can acquire the ability to resist sub-zero temperatures. This cold acclimation process is associated with physiological, biochemical and molecular alterations in cereals. Brachypodium distachyon is considered a powerful model system to study the response of temperate cereals to adverse environmental conditions. To date, little is known about the cold acclimation and freezing tolerance capacities of Brachypodium. The main objective of this study was to evaluate the cold hardiness of seven diploid Brachypodium accessions. An integrated approach, involving monitoring of phenological indicators along with expression profiling of the major vernalization regulator VRN1 orthologue, was followed. In parallel, soluble sugars and proline contents were determined along with expression profiles of two COR genes in plants exposed to low temperatures. Finally, whole-plant freezing tests were performed to evaluate the freezing tolerance capacity of Brachypodium. Cold treatment accelerated the transition from the vegetative to the reproductive phase in all diploid Brachypodium accessions tested. In addition, low temperature exposure triggered the gradual accumulation of BradiVRN1 transcripts in all accessions tested. These accessions exhibited a clear cold acclimation response by progressively accumulating proline, sugars and COR gene transcripts. However, whole-plant freezing tests revealed that these seven diploid accessions only have a limited capacity to develop freezing tolerance when compared with winter varieties of temperate cereals such as wheat and barley. Furthermore, little difference in terms of survival was observed among the accessions tested despite their previous classification as either spring or winter genotypes. This study is the first to characterize the freezing tolerance capacities of B. distachyon and provides strong evidence that

  4. Relationship between freezing tolerance and shoot water relations of western red cedar.

    PubMed

    Grossnickle, S C

    1992-10-01

    Freezing tolerance and shoot water relations parameters of western red cedar (Thuja plicata Donn) seedlings were measured every 2 weeks from October 1989 to April 1990. Freezing tolerance, measured by freeze-induced electrolyte leakage, showed seasonal shifts in the temperature causing 50% foliage electrolyte leakage (LT(50)). The LT(50) value was -4 degrees C in October, it decreased to -20 degrees C in February and then increased to -6 degrees C in April. The foliage index of injury at -10 degrees C (II(-10)) also showed seasonal shifts from a high of 98% in October to a low of 18% in February followed by an increase to 82% in April. Osmotic potentials at saturation (Psi(s(sat))) and turgor loss point (Psi(s(tlp))) were, respectively, -1.07 and -1.26 MPa in October, -1.57 and -2.43 MPa in January, and -1.04 and -1.86 MPa in April. Dry weight fraction (DWF) increased and symplastic volume at full turgor (V(o)) decreased during the fall-winter acclimation phase, whereas DWF decreased and V(o) increased during the late winter-spring deacclimation phase. Relationships between seasonal patterns of freezing tolerance and shoot water relations parameters showed that LT(50) and II(-10) decreased linearly as Psi(s(tlp)) and V(o) decreased and DWF increased. There was no discernible difference in the relationship during fall acclimation or spring deacclimation. The freezing dehydration index at -10 degrees C (FDI(-10)) declined from 0.69 in November to 0.41 in February and increased to 0.56 in April. The value of II(-10) decreased linearly as FDI(-10) decreased, although a measurement made on actively growing spring foliage did not fit this relationship. The results indicate that seasonal changes in freezing tolerance of western red cedar are partially due to changes in tissue water content, symplastic volume, passive osmotic adjustment and FDI(-10).

  5. Simple improvement in freeze-tolerance of bakers' yeast with poly-gamma-glutamate.

    PubMed

    Yokoigawa, Kumio; Sato, Machiko; Soda, Kenji

    2006-09-01

    We examined the effect of poly-gamma-glutamate (PGA) on the freeze-tolerance of four types of commercial bakers' yeast (freeze-tolerant, osmotic-tolerant, low-temperature-sensitive, and ordinary bakers' yeasts). The survival ratio of ordinary bakers' yeast cells frozen at -30 degrees C for 3 d in a medium (0.5% yeast extract, 0.5% peptone, and 2% glucose: YPD medium) was improved by adding more than 1% PGA to the medium; the survival ratio increased from about 10% to more than 70%. All PGA preparations, which differed in average molecular mass (50, 2,000, 4,000, 6,000, 8,000, and 10,000 kDa), showed a similar cryoprotecive effect on the cells. Similar results were also obtained with other types of bakers' yeast, sake yeast and beer yeast. When the four types of bakers' yeast cell were frozen at -30 degrees C for 3 d in dough supplemented with more than 1% PGA, the cells (after freezing and thawing) showed higher leavening ability than those frozen in dough without PGA, irrespective of the molecular mass of PGA. Thus, PGA appears to protect bakers' yeast from lethal freeze injury, leading to a high leavening ability after freezing and thawing. PGA did not decrease the original leavening ability of the bakers' yeast, and was not decomposed by the yeast cells. PGA suppressed the decrease in leavening ability during a prolonged fermentation time, probably because PGA adsorbed inhibitory metabolites accumulated in the dough. PGA could prove useful for improving the freeze-tolerance of bakers' yeast by its addition to dough.

  6. Down-regulating alpha-galactosidase enhances freezing tolerance in transgenic petunia.

    PubMed

    Pennycooke, Joyce C; Jones, Michelle L; Stushnoff, Cecil

    2003-10-01

    Alpha-galactosidase (alpha-Gal; EC 3.2.1.22) is involved in many aspects of plant metabolism, including hydrolysis of the alpha-1,6 linkage of raffinose oligosaccharides during deacclimation. To examine the relationship between endogenous sugars and freezing stress, the expression of alpha-Gal was modified in transgenic petunia (Petunia x hybrida cv Mitchell). The tomato (Lycopersicon esculentum) Lea-Gal gene under the control of the Figwort Mosaic Virus promoter was introduced into petunia in the sense and antisense orientations using Agrobacterium tumefaciens-mediated transformation. RNA gel blots confirmed that alpha-Gal transcripts were reduced in antisense lines compared with wild type, whereas sense plants had increased accumulation of alpha-Gal mRNAs. alpha-Gal activity followed a similar trend, with reduced activity in antisense lines and increased activity in all sense lines evaluated. Raffinose content of nonacclimated antisense plants increased 12- to 22-fold compared with wild type, and 22- to 53-fold after cold acclimation. Based upon electrolyte leakage tests, freezing tolerance of the antisense lines increased from -4 degrees C for cold-acclimated wild-type plants to -8 degrees C for the most tolerant antisense line. Down-regulating alpha-Gal in petunia results in an increase in freezing tolerance at the whole-plant level in nonacclimated and cold-acclimated plants, whereas overexpression of the alpha-Gal gene caused a decrease in endogenous raffinose and impaired freezing tolerance. These results suggest that engineering raffinose metabolism by transformation with alpha-Gal provides an additional method for improving the freezing tolerance of plants.

  7. Down-Regulating α-Galactosidase Enhances Freezing Tolerance in Transgenic Petunia1

    PubMed Central

    Pennycooke, Joyce C.; Jones, Michelle L.; Stushnoff, Cecil

    2003-01-01

    α-Galactosidase (α-Gal; EC 3.2.1.22) is involved in many aspects of plant metabolism, including hydrolysis of the α-1,6 linkage of raffinose oligosaccharides during deacclimation. To examine the relationship between endogenous sugars and freezing stress, the expression of α-Gal was modified in transgenic petunia (Petunia × hybrida cv Mitchell). The tomato (Lycopersicon esculentum) Lea-Gal gene under the control of the Figwort Mosaic Virus promoter was introduced into petunia in the sense and antisense orientations using Agrobacterium tumefaciens-mediated transformation. RNA gel blots confirmed that α-Gal transcripts were reduced in antisense lines compared with wild type, whereas sense plants had increased accumulation of α-Gal mRNAs. α-Gal activity followed a similar trend, with reduced activity in antisense lines and increased activity in all sense lines evaluated. Raffinose content of nonacclimated antisense plants increased 12- to 22-fold compared with wild type, and 22- to 53-fold after cold acclimation. Based upon electrolyte leakage tests, freezing tolerance of the antisense lines increased from –4°C for cold-acclimated wild-type plants to –8°C for the most tolerant antisense line. Down-regulating α-Gal in petunia results in an increase in freezing tolerance at the whole-plant level in nonacclimated and cold-acclimated plants, whereas overexpression of the α-Gal gene caused a decrease in endogenous raffinose and impaired freezing tolerance. These results suggest that engineering raffinose metabolism by transformation with α-Gal provides an additional method for improving the freezing tolerance of plants. PMID:14500789

  8. Seasonal variation in freeze tolerance and ice content of the tree frog Hyla versicolor.

    PubMed

    Layne, J R; Lee, R E

    1989-02-01

    Freeze tolerance and ice content of Hyla versicolor showed pronounced variation between summer (June) and winter (December). Summer frogs survived freezing at -3 degrees C for up to 9 hr and ice accumulation up to 50% of their total body water. A time course of ice formation indicated that an equilibrium level was reached in approximately 15 hr. Thus, the lethal ice content was less than the equilibrium ice content for these conditions (63.1%). A second group was induced to enter an overwintering condition by holding them through the summer and then subjecting them to a progressive reduction in temperature and photoperiod for 2 months. These frogs survived freezing for 48 hr at -3 degrees C. Their equilibrium ice content at this temperature was significantly lower (52.5%) than comparably treated summer animals. In the winter acclimatized group, frozen frogs had substantially higher blood glucose levels than unfrozen frogs (22.7 mumol/ml vs. 1.33 mumol/ml), but glycerol levels were not elevated after freezing. Freezing frogs conditioned for overwintering at -7 degrees C resulted in a higher equilibrium ice content (62.6%), but none survived. It is evident that in preparation for overwintering, frogs reduce the amount of ice formed at a given subzero temperature, but there is little indication of a substantial change in the total amount of ice tolerated.

  9. Farinose flavonoids are associated with high freezing tolerance in fairy primrose (Primula malacoides) plants.

    PubMed

    Isshiki, Ryutaro; Galis, Ivan; Tanakamaru, Shigemi

    2014-02-01

    The deposition of surface (farinose) flavonoids on aerial parts of some Primula species is a well-documented but poorly understood phenomenon. Here, we show that flavonoid deposition on the leaves and winter buds may contribute strongly to preventing freezing damage in these plants. The ice nucleation temperature of fairy primrose (Primula malacoides) leaves covered with natural flavone was approximately 6 °C lower compared to those that had their flavone artificially removed. Additionally, farinose flavonoids on the leaves reduced subsequent electrolyte leakage (EL) from the cells exposed to freezing temperatures. Interestingly, exogenous application of flavone at 4 mg/g fresh weight to P. malacoides leaves, which had the original flavone mechanically removed, restored freezing tolerance, and diminished EL from the cells to pretreatment values. Our results suggest that farinose flavonoids may function as mediators of freezing tolerance in P. malacoides, and exogenous application of flavone could be used to reduce freezing damage during sudden but predictable frost events in other plant species. © 2013 Institute of Botany, Chinese Academy of Sciences.

  10. Freezing tolerance of the European water frogs: the good, the bad, and the ugly.

    PubMed

    Voituron, Yann; Joly, Pierre; Eugène, Michel; Barré, Hervé

    2005-06-01

    Survival and some physiological responses to freezing were investigated in three European water frogs (Rana lessonae, Rana ridibunda, and their hybridogen Rana esculenta). The three species exhibited different survival times during freezing (from 10 h for R. lessonae to 20 h for R. ridibunda). The time courses of percent water frozen were similar; however, because of the huge differences in body mass among species (from 10 g for Rana lessonae to nearly 100 g for Rana ridibunda), the ice mass accumulation rate varied markedly (from 0.75 +/- 0.12 to 1.43 +/- 0.11 g ice/h, respectively) and was lowest in the terrestrial hibernator Rana lessonae. The hybrid Rana esculenta exhibited an intermediate response between the two parental species; furthermore, within-species correlation existed between body mass and ice mass accumulation rates, suggesting the occurrence of subpopulations in this species (0.84 +/- 0.08 g ice/h for small R. esculenta and 1.78 +/- 0.09 g ice/h for large ones). Biochemical analyses showed accumulation of blood glucose and lactate, liver glucose (originating from glycogen), and liver alanine in Rana lessonae and Rana esculenta but not in Rana ridibunda in response to freezing. The variation of freeze tolerance between these three closely related species could bring understanding to the physiological processes involved in the evolution of freeze tolerance in vertebrates.

  11. Transformation of beta-lycopene cyclase genes from Salicornia europaea and Arabidopsis conferred salt tolerance in Arabidopsis and tobacco.

    PubMed

    Chen, Xianyang; Han, Heping; Jiang, Ping; Nie, Lingling; Bao, Hexigeduleng; Fan, Pengxiang; Lv, Sulian; Feng, Juanjuan; Li, Yinxin

    2011-05-01

    Inhibition of lycopene cyclization decreased the salt tolerance of the euhalophyte Salicornia europaea L. We isolated a β-lycopene cyclase gene SeLCY from S. europaea and transformed it into Arabidopsis with stable expression. Transgenic Arabidopsis on post-germination exhibited enhanced tolerance to oxidative and salt stress. After 8 and 21 d recovery from 200 mM NaCl treatment, transgenic lines had a higher survival ratio than wild-type (WT) plants. Three-week-old transgenic plants treated with 200 mM NaCl showed better growth than the WT with higher photosystem activity and less H(2)O(2) accumulation. Determination of endogenous pigments of Arabidopsis treated with 200 mM NaCl for 0, 2 or 4 d demonstrated that the transgenic plants retained higher contents of carotenoids than the WT. Furthermore, to compare the difference between SeLCY and AtLCY from Arabidopsis, we used viral vector mediating ectopic expression of SeLCY and AtLCY in Nicotiana benthamiana. Although LCY genes transformation increased the salt tolerance in tobacco, there is no significant difference between SeLCY- and AtLCY-transformed plants. These findings indicate that SeLCY transgenic Arabidopsis improved salt tolerance by increasing synthesis of carotenoids, which impairs reactive oxygen species and protects the photosynthesis system under salt stress, and as a single gene, SeLCY functionally showed no advantage for salt tolerance improvement compared with AtLCY.

  12. Freeze tolerance and cryoprotectant mobilization in the gray treefrog (Hyla versicolor).

    PubMed

    Layne, J R

    1999-02-15

    Interstudy differences have been reported in the cryobiology of Hyla versicolor, especially for southern vs. northern populations, although laboratory conditioning likely was a contributing factor. This study measured freeze tolerance and cryoprotectant levels in H. versicolor from a southern population (Illinois). Frogs fully tolerated freezing at -1.5 and -3.5 degrees C for 24-48 hr but their survival rate declined at -5.5 degrees C (< 50%). Calorimetry revealed that 46% of the body water froze after 24 hr at -1.5 degrees C. Levels of plasma glucose and glycerol were substantially elevated, 14x and 5x respectively, in recently thawed frogs vs. unfrozen frogs. Plasma osmolality correspondingly rose from 242 to 304 mOsmol/L. Three unfrozen frogs had levels of plasma glycerol ranging between 17.1-36.8 mmol/L, suggesting an anticipatory response to freezing, but another three unfrozen frogs had a glycerol level of 1.1 mmol/L. A direct relationship existed between glycerol content and plasma osmolality and an inverse relationship was observed between plasma osmolality and ice content. Glycerol clearly was a major component of cryo-protectant production in these frogs, which was likely essential to their freeze tolerance. The cryobiology of Illinois frogs was only marginally less developed than seen in northern populations of this species.

  13. Wood frog adaptations to overwintering in Alaska: new limits to freezing tolerance.

    PubMed

    Larson, Don J; Middle, Luke; Vu, Henry; Zhang, Wenhui; Serianni, Anthony S; Duman, John; Barnes, Brian M

    2014-06-15

    We investigated the ecological physiology and behavior of free-living wood frogs [Lithobates (Rana) sylvaticus] overwintering in Interior Alaska by tracking animals into natural hibernacula, recording microclimate, and determining frog survival in spring. We measured cryoprotectant (glucose) concentrations and identified the presence of antifreeze glycolipids in tissues from subsamples of naturally freezing frogs. We also recorded the behavior of wood frogs preparing to freeze in artificial hibernacula, and tissue glucose concentrations in captive wood frogs frozen in the laboratory to -2.5°C. Wood frogs in natural hibernacula remained frozen for 193 ± 11 consecutive days and experienced average (October-May) temperatures of -6.3°C and average minimum temperatures of -14.6 ± 2.8°C (range -8.9 to -18.1°C) with 100% survival (N=18). Mean glucose concentrations were 13-fold higher in muscle, 10-fold higher in heart and 3.3-fold higher in liver in naturally freezing compared with laboratory frozen frogs. Antifreeze glycolipid was present in extracts from muscle and internal organs, but not skin, of frozen frogs. Wood frogs in Interior Alaska survive freezing to extreme limits and durations compared with those described in animals collected in southern Canada or the Midwestern United States. We hypothesize that this enhancement of freeze tolerance in Alaskan wood frogs is due to higher cryoprotectant levels that are produced by repeated freezing and thawing cycles experienced under natural conditions during early autumn.

  14. Skin ice nucleators and glycerol in the freezing-tolerant frog Litoria ewingii.

    PubMed

    Rexer-Huber, Kalinka M J; Bishop, Phillip J; Wharton, David A

    2011-08-01

    The brown tree frog (Litoria ewingii) is the only known Southern Hemisphere vertebrate that can survive full-body freezing. Freezing challenges living organisms in many ways, with ice formation in the body producing a suite of physical and metabolic stresses which can damage cells and tissues. The present study looked at two mechanisms that address some of these stresses: cryoprotectants and ice nucleating agents (INAs). Skin secretions from L. ewingii were sampled along with microhabitat substrate and tested for the presence of INAs, which help control ice formation in the body. L. ewingii plasma was tested for seasonal and freezing-induced changes in both glucose and glycerol, which may have a cryoprotective role in freezing-tolerant frogs. Glycerol levels increased on freezing and decreased on thawing, while glucose levels did not change on freezing but increased on thawing. This suggests that glycerol may be acting as a cryoprotectant, although levels are low compared to other frogs. A clear seasonal change was seen in INA activity, with greater activity in winter than in summer. While potent INAs came from the microhabitat substrate, this work has shown for the first time that skin secretions also contain active INAs.

  15. Threshold temperatures mediate the impact of reduced snow cover on overwintering freeze-tolerant caterpillars.

    PubMed

    Marshall, Katie E; Sinclair, Brent J

    2012-01-01

    Decreases in snow cover due to climate change could alter the energetics and physiology of ectothermic animals that overwinter beneath snow, yet how snow cover interacts with physiological thresholds is unknown. We applied numerical simulation of overwintering metabolic rates coupled with field validation to determine the importance of snow cover and freezing to the overwintering lipid consumption of the freeze-tolerant Arctiid caterpillar Pyrrharctia isabella. Caterpillars that overwintered above the snow experienced mean temperatures 1.3°C lower than those below snow and consumed 18.36 mg less lipid of a total 68.97-mg reserve. Simulations showed that linear temperature effects on metabolic rate accounted for only 30% of the difference in lipid consumption. When metabolic suppression by freezing was included, 93% of the difference between animals that overwintered above and below snow was explained. Our results were robust to differences in temperature sensitivity of metabolic rate, changes in freezing point, and the magnitude of metabolic suppression by freezing. The majority of the energy savings was caused by the non-continuous reduction in metabolic rate due to freezing, the first example of the importance of temperature thresholds in the lipid use of overwintering insects.

  16. Threshold temperatures mediate the impact of reduced snow cover on overwintering freeze-tolerant caterpillars

    NASA Astrophysics Data System (ADS)

    Marshall, Katie E.; Sinclair, Brent J.

    2012-01-01

    Decreases in snow cover due to climate change could alter the energetics and physiology of ectothermic animals that overwinter beneath snow, yet how snow cover interacts with physiological thresholds is unknown. We applied numerical simulation of overwintering metabolic rates coupled with field validation to determine the importance of snow cover and freezing to the overwintering lipid consumption of the freeze-tolerant Arctiid caterpillar Pyrrharctia isabella. Caterpillars that overwintered above the snow experienced mean temperatures 1.3°C lower than those below snow and consumed 18.36 mg less lipid of a total 68.97-mg reserve. Simulations showed that linear temperature effects on metabolic rate accounted for only 30% of the difference in lipid consumption. When metabolic suppression by freezing was included, 93% of the difference between animals that overwintered above and below snow was explained. Our results were robust to differences in temperature sensitivity of metabolic rate, changes in freezing point, and the magnitude of metabolic suppression by freezing. The majority of the energy savings was caused by the non-continuous reduction in metabolic rate due to freezing, the first example of the importance of temperature thresholds in the lipid use of overwintering insects.

  17. Transcriptome Sequencing Identified Genes and Gene Ontologies Associated with Early Freezing Tolerance in Maize

    PubMed Central

    Li, Zhao; Hu, Guanghui; Liu, Xiangfeng; Zhou, Yao; Li, Yu; Zhang, Xu; Yuan, Xiaohui; Zhang, Qian; Yang, Deguang; Wang, Tianyu; Zhang, Zhiwu

    2016-01-01

    Originating in a tropical climate, maize has faced great challenges as cultivation has expanded to the majority of the world's temperate zones. In these zones, frost and cold temperatures are major factors that prevent maize from reaching its full yield potential. Among 30 elite maize inbred lines adapted to northern China, we identified two lines of extreme, but opposite, freezing tolerance levels—highly tolerant and highly sensitive. During the seedling stage of these two lines, we used RNA-seq to measure changes in maize whole genome transcriptome before and after freezing treatment. In total, 19,794 genes were expressed, of which 4550 exhibited differential expression due to either treatment (before or after freezing) or line type (tolerant or sensitive). Of the 4550 differently expressed genes, 948 exhibited differential expression due to treatment within line or lines under freezing condition. Analysis of gene ontology found that these 948 genes were significantly enriched for binding functions (DNA binding, ATP binding, and metal ion binding), protein kinase activity, and peptidase activity. Based on their enrichment, literature support, and significant levels of differential expression, 30 of these 948 genes were selected for quantitative real-time PCR (qRT-PCR) validation. The validation confirmed our RNA-Seq-based findings, with squared correlation coefficients of 80% and 50% in the tolerance and sensitive lines, respectively. This study provided valuable resources for further studies to enhance understanding of the molecular mechanisms underlying maize early freezing response and enable targeted breeding strategies for developing varieties with superior frost resistance to achieve yield potential. PMID:27774095

  18. Freeze-thaw tolerance and clues to the winter survival of a soil community.

    PubMed

    Walker, Virginia K; Palmer, Gerald R; Voordouw, Gerrit

    2006-03-01

    Although efforts have been made to sample microorganisms from polar regions and to investigate a few of the properties that facilitate survival at freezing or subzero temperatures, soil communities that overwinter in areas exposed to alternate freezing and thawing caused by Foehn or Chinook winds have been largely overlooked. We designed and constructed a cryocycler to automatically subject soil cultures to alternating freeze-thaw cycles. After 48 freeze-thaw cycles, control Escherichia coli and Pseudomonas chlororaphis isolates were no longer viable. Mixed cultures derived from soil samples collected from a Chinook zone showed that the population complexity and viability were reduced after 48 cycles. However, when bacteria that were still viable after the freeze-thaw treatments were used to obtain selected cultures, these cultures proved to be >1,000-fold more freeze-thaw tolerant than the original consortium. Single-colony isolates obtained from survivors after an additional 48 freeze-thaw cycles were putatively identified by 16S RNA gene fragment sequencing. Five different genera were recognized, and one of the cultures, Chryseobacterium sp. strain C14, inhibited ice recrystallization, a property characteristic of antifreeze proteins that prevents the growth of large, potentially damaging ice crystals at temperatures close to the melting temperature. This strain was also notable since cell-free medium derived from cultures of it appeared to enhance the multiple freeze-thaw survival of another isolate, Enterococcus sp. strain C8. The results of this study and the development of a cryocycler should allow further investigations into the biochemical and soil community adaptations to the rigors of a Chinook environment.

  19. Freeze-Thaw Tolerance and Clues to the Winter Survival of a Soil Community

    PubMed Central

    Walker, Virginia K.; Palmer, Gerald R.; Voordouw, Gerrit

    2006-01-01

    Although efforts have been made to sample microorganisms from polar regions and to investigate a few of the properties that facilitate survival at freezing or subzero temperatures, soil communities that overwinter in areas exposed to alternate freezing and thawing caused by Foehn or Chinook winds have been largely overlooked. We designed and constructed a cryocycler to automatically subject soil cultures to alternating freeze-thaw cycles. After 48 freeze-thaw cycles, control Escherichia coli and Pseudomonas chlororaphis isolates were no longer viable. Mixed cultures derived from soil samples collected from a Chinook zone showed that the population complexity and viability were reduced after 48 cycles. However, when bacteria that were still viable after the freeze-thaw treatments were used to obtain selected cultures, these cultures proved to be >1,000-fold more freeze-thaw tolerant than the original consortium. Single-colony isolates obtained from survivors after an additional 48 freeze-thaw cycles were putatively identified by 16S RNA gene fragment sequencing. Five different genera were recognized, and one of the cultures, Chryseobacterium sp. strain C14, inhibited ice recrystallization, a property characteristic of antifreeze proteins that prevents the growth of large, potentially damaging ice crystals at temperatures close to the melting temperature. This strain was also notable since cell-free medium derived from cultures of it appeared to enhance the multiple freeze-thaw survival of another isolate, Enterococcus sp. strain C8. The results of this study and the development of a cryocycler should allow further investigations into the biochemical and soil community adaptations to the rigors of a Chinook environment. PMID:16517623

  20. The Influence of Light Quality, Circadian Rhythm, and Photoperiod on the CBF-Mediated Freezing Tolerance

    PubMed Central

    Maibam, Punyakishore; Nawkar, Ganesh M.; Park, Joung Hun; Sahi, Vaidurya Pratap; Lee, Sang Yeol; Kang, Chang Ho

    2013-01-01

    Low temperature adversely affects crop yields by restraining plant growth and productivity. Most temperate plants have the potential to increase their freezing tolerance upon exposure to low but nonfreezing temperatures, a process known as cold acclimation. Various physiological, molecular, and metabolic changes occur during cold acclimation, which suggests that the plant cold stress response is a complex, vital phenomenon that involves more than one pathway. The C-Repeat Binding Factor (CBF) pathway is the most important and well-studied cold regulatory pathway that imparts freezing tolerance to plants. The regulation of freezing tolerance involves the action of phytochromes, which play an important role in light-mediated signalling to activate cold-induced gene expression through the CBF pathway. Under normal temperature conditions, CBF expression is regulated by the circadian clock through the action of a central oscillator and also day length (photoperiod). The phytochrome and phytochrome interacting factor are involved in the repression of the CBF expression under long day (LD) conditions. Apart from the CBF regulon, a novel pathway involving the Z-box element also mediates the cold acclimation response in a light-dependent manner. This review provides insights into the progress of cold acclimation in relation to light quality, circadian regulation, and photoperiodic regulation and also explains the underlying molecular mechanisms of cold acclimation for introducing the engineering of economically important, cold-tolerant plants. PMID:23722661

  1. Urea is not a universal cryoprotectant among hibernating anurans: evidence from the freeze-tolerant boreal chorus frog (Pseudacris maculata).

    PubMed

    Higgins, Steven A; Swanson, David L

    2013-02-01

    Freeze-tolerant organisms accumulate a diversity of low molecular weight compounds to combat negative effects of ice formation. Previous studies of anuran freeze tolerance have implicated urea as a cryoprotectant in the wood frog (Lithobates sylvatica). However, a cryoprotective role for urea has been identified only for wood frogs, though urea accumulation is an evolutionarily conserved mechanism for coping with osmotic stress in amphibians. To identify whether multiple solutes are involved in freezing tolerance in the boreal chorus frog (Pseudacris maculata), we examined seasonal and freezing-induced variation in several potential cryoprotectants. We further tested for a cryoprotective role for urea by comparing survival and recovery from freezing in control and urea-loaded chorus frogs. Tissue levels of glucose, urea, and glycerol did not vary significantly among seasons for heart, liver, or leg muscle. Furthermore, no changes in urea or glycerol levels were detected with exposure to freezing temperatures in these tissues. Urea-loading increased tissue urea concentrations, but failed to enhance freezing survival or facilitate recovery from freezing in chorus frogs in this study, suggesting little role for urea as a natural cryoprotectant in this species. These data suggest that urea may not universally serve as a primary cryoprotectant among freeze-tolerant, terrestrially hibernating anurans.

  2. Osmoregulation and salinity tolerance in the Antarctic midge, Belgica antarctica: seawater exposure confers enhanced tolerance to freezing and dehydration.

    PubMed

    Elnitsky, Michael A; Benoit, Joshua B; Lopez-Martinez, Giancarlo; Denlinger, David L; Lee, Richard E

    2009-09-01

    Summer storms along the Antarctic Peninsula can cause microhabitats of the terrestrial midge Belgica antarctica to become periodically inundated with seawater from tidal spray. As microhabitats dry, larvae may be exposed to increasing concentrations of seawater. Alternatively, as a result of melting snow or following rain, larvae may be immersed in freshwater for extended periods. The present study assessed the tolerance and physiological response of B. antarctica larvae to salinity exposure, and examined the effect of seawater acclimation on their subsequent tolerance of freezing, dehydration and heat shock. Midge larvae tolerated extended exposure to hyperosmotic seawater; nearly 50% of larvae survived a 10-day exposure to 1000 mOsm kg(-1) seawater and approximately 25% of larvae survived 6 days in 2000 mOsm kg(-1) seawater. Exposure to seawater drastically reduced larval body water content and increased hemolymph osmolality. By contrast, immersion in freshwater did not affect water content or hemolymph osmolality. Hyperosmotic seawater exposure, and the accompanying osmotic dehydration, resulted in a significant correlation between the rate of oxygen consumption and larval water content and induced the de novo synthesis and accumulation of several organic osmolytes. A 3-day exposure of larvae to hyperosmotic seawater increased freezing tolerance relative to freshwater-acclimated larvae. Even after rehydration, the freezing survival of larvae acclimated to seawater was greater than freshwater-acclimated larvae. Additionally, seawater exposure increased the subsequent tolerance of larvae to dehydration. Our results further illustrate the similarities between these related, yet distinct, forms of osmotic stress and add to the suite of physiological responses used by larvae to enhance survival in the harsh and unpredictable Antarctic environment.

  3. 6-Phosphofructo-2-kinase and control of cryoprotectant synthesis in freeze tolerant frogs.

    PubMed

    Vazquez Illanes, M D; Storey, K B

    1993-08-20

    A critical part of natural freeze tolerance is the production of low molecular weight cryoprotectants; in freeze tolerant frogs this involves a freezing-induced activation of liver glycogenolysis that leads to the accumulation of glucose as the cryoprotectant, in amounts up to 300 mM, in all organs. The present study shows that the synthesis and maintenance of high organ glucose pools is facilitated by changes in the levels of fructose-2,6-bisphosphate (F2,6P2) and an inhibition of liver 6-phosphofructo-2-kinase (PFK-2) activity that blocks the catabolism of glucose by glycolysis. Freezing exposure (24 h at -2.5 degrees C) resulted in a sharp drop in F2,6P2 levels in four organs, to 23-75% of control values, but F2,6P2 rebounded when frogs were thawed. Freezing also stimulated changes in the properties of liver PFK-2 including a decrease in maximal velocity, a basic shift in pH optimum, a 10-fold increase in Km for fructose-6-phosphate, and increased I50 values for enzyme inhibitors. I50 values for glycerol-3-phosphate and phosphoenolpyruvate were 60- and 2.4-fold higher, respectively, for liver PFK-2 from frozen frogs compared with controls. Changes in liver PFK-2 properties are consistent with a freezing-induced phosphorylation of the enzyme to produce a less active enzyme form, resulting in reduced organ F2,6P2 levels and a decrease in 6-phosphofructo-1-kinase activity.

  4. Novel NAC Transcription Factor TaNAC67 Confers Enhanced Multi-Abiotic Stress Tolerances in Arabidopsis

    PubMed Central

    Mao, Xinguo; Chen, Shuangshuang; Li, Ang; Zhai, Chaochao; Jing, Ruilian

    2014-01-01

    Abiotic stresses are major environmental factors that affect agricultural productivity worldwide. NAC transcription factors play pivotal roles in abiotic stress signaling in plants. As a staple crop, wheat production is severely constrained by abiotic stresses whereas only a few NAC transcription factors have been characterized functionally. To promote the application of NAC genes in wheat improvement by biotechnology, a novel NAC gene designated TaNAC67 was characterized in common wheat. To determine its role, transgenic Arabidopsis overexpressing TaNAC67-GFP controlled by the CaMV-35S promoter was generated and subjected to various abiotic stresses for morphological and physiological assays. Gene expression showed that TaNAC67 was involved in response to drought, salt, cold and ABA treatments. Localization assays revealed that TaNAC67 localized in the nucleus. Morphological analysis indicated the transgenics had enhanced tolerances to drought, salt and freezing stresses, simultaneously supported by enhanced expression of multiple abiotic stress responsive genes and improved physiological traits, including strengthened cell membrane stability, retention of higher chlorophyll contents and Na+ efflux rates, improved photosynthetic potential, and enhanced water retention capability. Overexpression of TaNAC67 resulted in pronounced enhanced tolerances to drought, salt and freezing stresses, therefore it has potential for utilization in transgenic breeding to improve abiotic stress tolerance in crops. PMID:24427285

  5. High tolerance to repeated cycles of freezing and thawing in different Trichinella nativa isolates.

    PubMed

    Davidson, Rebecca K; Handeland, Kjell; Kapel, Christian M O

    2008-10-01

    Trichinella nativa is the most frequent Trichinella species in arctic wildlife and also the predominating species seen in Norwegian fauna. The adaptation of T. nativa to a cold climate is reflected by the well-documented freeze tolerance of its muscle larvae. The ability of the larvae to survive repeated freezing and thawing events has not however been elucidated and was investigated in the present study, using an Alaskan isolate and two isolates from coastal and inland Norway, respectively. Each T. nativa isolate was inoculated in foxes (Vulpes vulpes) and the muscle tissue obtained after 20 weeks was minced and divided into freezer bags. In the initial part of the study, the bags were exposed to either continuous freezing (-5 degrees C) for up to 7 weeks or freezing (-5 degrees C) for up to 7 weeks with seven overnight (+21 degrees C) thawing events. Once a week a bag was removed from each group, the meat was digested and muscle larvae isolated. In vitro assessment of larval viability was carried out based on larval motility (active vs non-active) and morphology, coiled (alive) or C-shaped (dead). Larval infectivity was subsequently bioassayed in mice, administering 500 larvae per mouse. The mice were euthanised 4 weeks post inoculation, the muscle digested and larvae per gram (lpg) and reproductive capacity index (RCI) were calculated. During the second part of the study, some of the minced fox muscle, exposed to the initial freeze protocol, was stored for a further 23 weeks at -18 degrees C prior to in vitro and in vivo assessment of larval viability and infectivity. The study demonstrated that Trichinella isolates originating from carnivores from higher northern latitudes expressed highest tolerance to freezing and that temperature fluctuations around freezing point, for up to 7 weeks, had little effect on larval infectivity. A negative effect of the initial repeated freeze-thaw events could be demonstrated once the larvae were exposed to longer periods of

  6. Freeze-tolerance of Trichinella muscle larvae in experimentally infected wild boars.

    PubMed

    Lacour, Sandrine A; Heckmann, Aurélie; Macé, Pauline; Grasset-Chevillot, Aurélie; Zanella, Gina; Vallée, Isabelle; Kapel, Christian M O; Boireau, Pascal

    2013-05-20

    Freeze-tolerance of encapsulated Trichinella muscle larvae (ML) is mainly determined by Trichinella species, but is also influenced by host species, the age of the infection and the storage time and temperature of the infected meat. Moreover, the freeze-tolerance of the encapsulated species appears to be correlated to the development of thick capsule walls which increases with age. An extended infection period and the muscle composition in some hosts (e.g. herbivores) may provide freeze-avoiding matrices due to high carbohydrate contents. The present experiment compares freeze-tolerance of Trichinella spiralis and Trichinella britovi ML in wild boar meat 24 weeks post inoculation (wpi). Three groups of four wild boars were infected with 200, 2000 or 20,000 ML of T. britovi (ISS 1575), respectively. Additionally, three wild boars were inoculated with 20,000 ML of T. spiralis (ISS 004) and two animals served as negative controls. All wild boars were sacrificed 24 wpi. Muscle samples of 70 g were stored at -21°C for 19, 30 and 56 h, and for 1-8 weeks. Larvae were recovered by artificial digestion. Their mobilities were recorded using Saisam(®) image analysis software and their infectivities were evaluated using mouse bioassays. Samples frozen for 19, 30 and 56 h allowed recovery of mobile ML, but samples frozen for 1 or 2 weeks did not. Correspondingly, only T. spiralis and T. britovi larvae isolated from wild boar meat frozen for 19, 30 and 56 h established in mice. This study showed that freezing at -21°C for 1 week inactivated T. spiralis and T. britovi ML encapsulated in wild boar meat for 24 weeks.

  7. Proteome dynamics of cold-acclimating Rhododendron species contrasting in their freezing tolerance and thermonasty behavior

    PubMed Central

    Arora, Rajeev; Rowland, Lisa J.

    2017-01-01

    To gain a better understanding of cold acclimation in rhododendron and in woody perennials in general, we used the 2D-DIGE technique to analyze the rhododendron proteome during the seasonal development of freezing tolerance. We selected two species varying in their cold acclimation ability as well as their thermonasty response (folding of leaves in response to low temperature). Proteins were extracted from leaves of non-acclimated (NA) and cold acclimated (CA) plants of the hardier thermonastic species, R. catawbiense (Cata.), and from leaves of cold acclimated plants of the less hardy, non-thermonastic R. ponticum (Pont.). All three protein samples (Cata.NA, Cata.CA, and Pont.CA) were labeled with different CyDyes and separated together on a single gel. Triplicate gels were run and protein profiles were compared resulting in the identification of 72 protein spots that consistently had different abundances in at least one pair-wise comparison. From the 72 differential spots, we chose 56 spots to excise and characterize further by mass spectrometry (MS). Changes in the proteome associated with the seasonal development of cold acclimation were identified from the Cata.CA—Cata.NA comparisons. Differentially abundant proteins associated with the acquisition of superior freezing tolerance and with the thermonastic response were identified from the Cata.CA—Pont.CA comparisons. Our results indicate that cold acclimation in rhododendron involves increases in abundance of several proteins related to stress (freezing/desiccation tolerance), energy and carbohydrate metabolism, regulation/signaling, secondary metabolism (possibly involving cell wall remodeling), and permeability of the cell membrane. Cold acclimation also involves decreases in abundance of several proteins involved in photosynthesis. Differences in freezing tolerance between genotypes can probably be attributed to observed differences in levels of proteins involved in these functions. Also differences in

  8. Proteome dynamics of cold-acclimating Rhododendron species contrasting in their freezing tolerance and thermonasty behavior.

    PubMed

    Die, Jose V; Arora, Rajeev; Rowland, Lisa J

    2017-01-01

    To gain a better understanding of cold acclimation in rhododendron and in woody perennials in general, we used the 2D-DIGE technique to analyze the rhododendron proteome during the seasonal development of freezing tolerance. We selected two species varying in their cold acclimation ability as well as their thermonasty response (folding of leaves in response to low temperature). Proteins were extracted from leaves of non-acclimated (NA) and cold acclimated (CA) plants of the hardier thermonastic species, R. catawbiense (Cata.), and from leaves of cold acclimated plants of the less hardy, non-thermonastic R. ponticum (Pont.). All three protein samples (Cata.NA, Cata.CA, and Pont.CA) were labeled with different CyDyes and separated together on a single gel. Triplicate gels were run and protein profiles were compared resulting in the identification of 72 protein spots that consistently had different abundances in at least one pair-wise comparison. From the 72 differential spots, we chose 56 spots to excise and characterize further by mass spectrometry (MS). Changes in the proteome associated with the seasonal development of cold acclimation were identified from the Cata.CA-Cata.NA comparisons. Differentially abundant proteins associated with the acquisition of superior freezing tolerance and with the thermonastic response were identified from the Cata.CA-Pont.CA comparisons. Our results indicate that cold acclimation in rhododendron involves increases in abundance of several proteins related to stress (freezing/desiccation tolerance), energy and carbohydrate metabolism, regulation/signaling, secondary metabolism (possibly involving cell wall remodeling), and permeability of the cell membrane. Cold acclimation also involves decreases in abundance of several proteins involved in photosynthesis. Differences in freezing tolerance between genotypes can probably be attributed to observed differences in levels of proteins involved in these functions. Also differences in

  9. Small heat-shock protein Hsp12 contributes to yeast tolerance to freezing stress.

    PubMed

    Pacheco, A; Pereira, C; Almeida, M J; Sousa, M J

    2009-06-01

    The HSP12 gene encodes one of the two major small heat-shock proteins of Saccharomyces cerevisiae and is induced under different conditions, such as low and high temperatures, osmotic or oxidative stress and high sugar or ethanol concentrations. However, few studies could demonstrate any correlation between HSP12 deletion or overexpression and a phenotype of sensitivity/resistance, making it difficult to attribute a role for Hsp12p under several of these stress conditions. We investigated the possible role of Hsp12p in yeast freezing tolerance. Contrary to what would be expected, the hsp12 null mutant when subjected to prolonged storage at -20 degrees C showed an increased resistance to freezing when compared with the isogenic wild-type strain. Because the mutant strain displayed a higher intracellular trehalose concentration than the wild-type, which could mask the effect of manipulating HSP12, we overexpressed the HSP12 gene in a trehalose-6-phosphate synthase (TPS1) null mutant. The tps1Delta strain overexpressing HSP12 showed an increase in resistance to freezing storage, indicating that Hsp12p plays a role in freezing tolerance in a way that seems to be interchangeable with trehalose. In addition, we show that overexpression of HSP12 in this tps1Delta strain also increased resistance to heat shock and that absence of HSP12 compromises the ability of yeast cells to accumulate high levels of trehalose in response to a mild heat stress.

  10. Strategy of freeze tolerance in adult of sunn pest bug (Eurygaster integriceps Put.).

    PubMed

    Baghdadi, Ahmed

    2007-01-01

    To investigate cold hardiness of Eurygaster integriceps Put, super cooling points of whole bodies during December to March 2006-2007 were measured on samples collected from altitude of Ghara-aghaj varamin-Iran. In addition, the lower lethal temperature (LLT) was determined for adult insect. The super cooling points were observed about -5 in cold months. Since the minimum temperature under natural condition is reached to-7 degreesC or lower in January and LLT99 (-21.65 degrees C), --11 degreesC lower than their minimum SCP (-10.5 degrees C), therefore I concluded that adult insects are freeze tolerant. Furthermore, super cooling points of different sex, weight and attitude were not significantly different. Freeze tolerance in these insects may be a strategy to provide protection from long-term exposures to ice crystals in microhabitat in alititude.

  11. Freezing tolerance revisited-effects of variable temperatures on gene regulation in temperate grasses and legumes.

    PubMed

    Kovi, Mallikarjuna Rao; Ergon, Åshild; Rognli, Odd Arne

    2016-10-01

    Climate change creates new patterns of seasonal climate variation with higher temperatures, longer growth seasons and more variable winter climates. This is challenging the winter survival of perennial herbaceous plants. In this review, we focus on the effects of variable temperatures during autumn/winter/spring, and its interactions with light, on the development and maintenance of freezing tolerance. Cold temperatures induce changes at several organizational levels in the plant (cold acclimation), leading to the development of freezing tolerance, which can be reduced/lost during warm spells (deacclimation) in winters, and attained again during cold spells (reacclimation). We summarize how temperature interacts with components of the light regime (photoperiod, PSII excitation pressure, irradiance, and light quality) in determining changes in the transcriptome, proteome and metabolome.

  12. Drought increases freezing tolerance of both leaves and xylem of Larrea tridentata.

    PubMed

    Medeiros, Juliana S; Pockman, William T

    2011-01-01

    Drought and freezing are both known to limit desert plant distributions, but the interaction of these stressors is poorly understood. Drought may increase freezing tolerance in leaves while decreasing it in the xylem, potentially creating a mismatch between water supply and demand. To test this hypothesis, we subjected Larrea tridentata juveniles grown in a greenhouse under well-watered or drought conditions to minimum temperatures ranging from -8 to -24 °C. We measured survival, leaf retention, gas exchange, cell death, freezing point depression and leaf-specific xylem hydraulic conductance (k₁). Drought-exposed plants exhibited smaller decreases in gas exchange after exposure to -8 °C compared to well-watered plants. Drought also conferred a significant positive effect on leaf, xylem and whole-plant function following exposure to -15 °C; drought-exposed plants exhibited less cell death, greater leaf retention, higher k₁ and higher rates of gas exchange than well-watered plants. Both drought-exposed and well-watered plants experienced 100% mortality following exposure to -24 °C. By documenting the combined effects of drought and freezing stress, our data provide insight into the mechanisms determining plant survival and performance following freezing and the potential for shifts in L. tridentata abundance and range in the face of changing temperature and precipitation regimes. © 2010 Blackwell Publishing Ltd.

  13. Association between the presence of protein bands in ram seminal plasma and sperm tolerance to freezing.

    PubMed

    Goularte, K L; Gastal, G D A; Schiavon, R S; Gonçalves, A O; Schneider, J R; Corcini, C D; Lucia, T

    2014-05-01

    This study evaluated associations between the presence of protein bands in ram seminal plasma and the quality of sperm frozen with distinct extenders. Ejaculates were frozen in a Tris-egg yolk based extender, including either 5% glycerol or 100mM trehalose. Seminal plasma samples were submitted to unidimensional electrophoresis. Pre-freezing and post-thawing sperm quality was similar between extenders (P>0.05). A total of 26 bands were identified in ram seminal plasma. Pre-freezing sperm motility was increased when the 15, 19 and 80kDa bands were present in seminal plasma (P<0.05). The presence of an 11kDa band in seminal plasma was associated with reduced pre-freezing membrane integrity (P<0.05). After thawing, both sperm motility and membrane integrity were reduced when a 24kDa band was present in seminal plasma (P<0.05). Post-thawing acrosome integrity was greater in the presence of a 31kDa band in seminal plasma (P<0.05). Regardless of the cryoprotectant included in the freezing extender, these six bands may be potential markers for ram sperm tolerance to freezing.

  14. Adaptive evolution of baker's yeast in a dough‐like environment enhances freeze and salinity tolerance

    PubMed Central

    Aguilera, Jaime; Andreu, Pasqual; Randez‐Gil, Francisca; Prieto, Jose Antonio

    2010-01-01

    Summary We used adaptive evolution to improve freeze tolerance of industrial baker's yeast. Our hypothesis was that adaptation to low temperature is accompanied by enhanced resistance of yeast to freezing. Based on this hypothesis, yeast was propagated in a flour‐free liquid dough model system, which contained sorbitol and NaCl, by successive batch refreshments maintained constantly at 12°C over at least 200 generations. Relative to the parental population, the maximal growth rate (µmax) under the restrictive conditions, increased gradually over the time course of the experiment. This increase was accompanied by enhanced freeze tolerance. However, these changes were not the consequence of genetic adaptation to low temperature, a fact that was confirmed by prolonged selection of yeast cells in YPD at 12°C. Instead, the experimental populations showed a progressive increase in NaCl tolerance. This phenotype was likely achieved at the expense of others traits, since evolved cells showed a ploidy reduction, a defect in the glucose derepression mechanism and a loss in their ability to utilize gluconeogenic carbon sources. We discuss the genetic flexibility of S. cerevisiae in terms of adaptation to the multiple constraints of the experimental design applied to drive adaptive evolution and the technologically advantageous phenotype of the evolved population. PMID:21255321

  15. Adaptive evolution of baker's yeast in a dough-like environment enhances freeze and salinity tolerance.

    PubMed

    Aguilera, Jaime; Andreu, Pasqual; Randez-Gil, Francisca; Prieto, Jose Antonio

    2010-03-01

    We used adaptive evolution to improve freeze tolerance of industrial baker's yeast. Our hypothesis was that adaptation to low temperature is accompanied by enhanced resistance of yeast to freezing. Based on this hypothesis, yeast was propagated in a flour-free liquid dough model system, which contained sorbitol and NaCl, by successive batch refreshments maintained constantly at 12°C over at least 200 generations. Relative to the parental population, the maximal growth rate (µ(max)) under the restrictive conditions, increased gradually over the time course of the experiment. This increase was accompanied by enhanced freeze tolerance. However, these changes were not the consequence of genetic adaptation to low temperature, a fact that was confirmed by prolonged selection of yeast cells in YPD at 12°C. Instead, the experimental populations showed a progressive increase in NaCl tolerance. This phenotype was likely achieved at the expense of others traits, since evolved cells showed a ploidy reduction, a defect in the glucose derepression mechanism and a loss in their ability to utilize gluconeogenic carbon sources. We discuss the genetic flexibility of S. cerevisiae in terms of adaptation to the multiple constraints of the experimental design applied to drive adaptive evolution and the technologically advantageous phenotype of the evolved population.

  16. Influence of simulated snow cover on the cold tolerance and freezing injury of yellow-cedar seedlings

    Treesearch

    Paul G. Schaberg; Paul E. Hennon; David V. D' amore; Gary J.  Hawley

    2008-01-01

    It has been hypothesized that yellow-cedar [Chamaecyparis nootkatensis (D. Don) Spach] decline may result from root freezing injury following climate change-induced reductions in protective snow cover. To test this hypothesis, we measured the freezing tolerance and injury expression of yellow-cedar seedlings in three treatments that differed in the...

  17. The Alfin-like homeodomain finger protein AL5 suppresses multiple negative factors to confer abiotic stress tolerance in Arabidopsis.

    PubMed

    Wei, Wei; Zhang, Yu-Qin; Tao, Jian-Jun; Chen, Hao-Wei; Li, Qing-Tian; Zhang, Wan-Ke; Ma, Biao; Lin, Qing; Zhang, Jin-Song; Chen, Shou-Yi

    2015-03-01

    Plant homeodomain (PHD) finger proteins affect processes of growth and development by changing transcription and reading epigenetic histone modifications, but their functions in abiotic stress responses remain largely unclear. Here we characterized seven Arabidopsis thaliana Alfin1-like PHD finger proteins (ALs) in terms of the responses to abiotic stresses. ALs localized to the nucleus and repressed transcription. Except AL6, all the ALs bound to G-rich elements. Mutations of the amino acids at positions 34 and 35 in AL6 caused loss of ability to bind to G-rich elements. Expression of the AL genes responded differentially to osmotic stress, salt, cold and abscisic acid treatments. AL5-over-expressing plants showed higher tolerance to salt, drought and freezing stress than Col-0. Consistently, al5 mutants showed reduced stress tolerance. We used ChIP-Seq assays to identify eight direct targets of AL5, and found that AL5 binds to the promoter regions of these genes. Knockout mutants of five of these target genes exhibited varying tolerances to stresses. These results indicate that AL5 inhibits multiple signaling pathways to confer stress tolerance. Our study sheds light on mechanisms of AL5-mediated signaling in abiotic stress responses, and provides tools for improvement of stress tolerance in crop plants.

  18. Ice recrystallization inhibition proteins (IRIPs) and freeze tolerance in the cryophilic Antarctic hair grass Deschampsia antarctica E. Desv.

    PubMed

    John, Ulrik P; Polotnianka, Renatam M; Sivakumaran, Kailayapillai A; Chew, Orinda; Mackin, Leanne; Kuiper, Micheal J; Talbot, Jonathan P; Nugent, Gregory D; Mautord, Julie; Schrauf, Gustavo E; Spangenberg, German C

    2009-04-01

    Antarctic hair grass (Deschampsia antarctica E. Desv.), the only grass indigenous to Antarctica, has well-developed freezing tolerance, strongly induced by cold acclimation. Here, we show that in response to low temperatures, D. antarctica expresses potent recrystallization inhibition (RI) activity that, inhibits the growth of small ice crystals into potentially damaging large ones, is proteinaceous and localized to the apoplasm. A gene family from D. antarctica encoding putative homologs of an ice recrystallization inhibition protein (IRIP) has been isolated and characterized. IRIPs are apoplastically targeted proteins with two potential ice-binding motifs: 1-9 leucine-rich repeats (LRRs) and c. 16 'IRIP' repeats. IRIP genes appear to be confined to the grass subfamily Pooideae and their products, exhibit sequence similarity to phytosulphokine receptors and are predicted to adopt conformations with two ice-binding surfaces. D. antarctica IRIP (DaIRIP) transcript levels are greatly enhanced in leaf tissue following cold acclimation. Transgenic Arabidopsis thaliana expressing a DaIRIP has novel RI activity, and purified DaIRIP, when added back to extracts of leaves from non-acclimated D. antarctica, can reconstitute the activity found in acclimated plants. We propose that IRIP-mediated RI activity may contribute to the cryotolerance of D. antarctica, and thus to its unique ability to have colonized Antarctica.

  19. SENSITIVE TO FREEZING2 Aids in Resilience to Salt and Drought in Freezing-Sensitive Tomato1[OPEN

    PubMed Central

    Hersh, Hope Lynn

    2016-01-01

    SENSITIVE TO FREEZING2 (SFR2) is crucial for protecting chloroplast membranes following freezing in Arabidopsis (Arabidopsis thaliana). It has been shown that SFR2 homologs are present in all land plants, including freezing-sensitive species, raising the question of SFR2 function beyond freezing tolerance. Similar to freezing, salt and drought can cause dehydration. Thus, it is hypothesized that in freezing-sensitive plants SFR2 may play roles in their resilience to salt or drought. To test this hypothesis, SlSFR2 RNAi lines were generated in the cold/freezing-sensitive species tomato (Solanum lycopersicum [M82 cv]). Hypersensitivity to salt and drought of SlSFR2-RNAi lines was observed. Higher tolerance of wild-type tomatoes was correlated with the production of trigalactosyldiacylglycerol, a product of SFR2 activity. Tomato SFR2 in vitro activity is Mg2+-dependent and its optimal pH is 7.5, similar to that of Arabidopsis SFR2, but the specific activity of tomato SFR2 in vitro is almost double that of Arabidopsis SFR2. When salt and drought stress were applied to Arabidopsis, no conditions could be identified at which SFR2 was induced prior to irreversibly impacting plant growth, suggesting that SFR2 protects Arabidopsis primarily against freezing. Discovery of tomato SFR2 function in drought and salt resilience provides further insights into general membrane lipid remodeling-based stress tolerance mechanisms and together with protection against freezing in freezing-resistant plants such as Arabidopsis, it adds lipid remodeling as a possible target for the engineering of abiotic stress-resilient crops. PMID:27600812

  20. Exogenous Abscisic Acid Mimics Cold Acclimation for Cacti Differing in Freezing Tolerance.

    PubMed

    Loik, M. E.; Nobel, P. S.

    1993-11-01

    The responses to low temperature were determined for two species of cacti sensitive to freezing, Ferocactus viridescens and Opuntia ficus-indica, and a cold hardy species, Opuntia fragilis. Fourteen days after shifting the plants from day/night air temperatures of 30/20[deg]C to 10/0[deg]C, the chlorenchyma water content decreased only for O. fragilis. This temperature shift caused the freezing tolerance (measured by vital stain uptake) of chlorenchyma cells to be enhanced only by about 2.0[deg]C for F. viridescens and O. ficus-indica but by 14.6[deg]C for O. fragilis. Also, maintenance of high water content by injection of water into plants at 10/0[deg]C reversed the acclimation. The endogenous abscisic acid (ABA) concentration was below 0.4 pmol g-1 fresh weight at 30/20[deg]C, but after 14 d at 10/0[deg]C it increased to 84 pmol g-1 fresh weight for O. ficus-indica and to 49 pmol g-1 fresh weight for O. fragilis. Four days after plants were sprayed with 7.5 x 10-5 M ABA at 30/20[deg]C, freezing tolerance was enhanced by 0.5[deg]C for F. viridescens, 4.1[deg]C for O. ficus-indica, and 23.4[deg]C for O. fragilis. Moreover, the time course for the change in freezing tolerance over 14 d was similar for plants shifted to low temperatures as for plants treated with exogenous ABA at moderate temperatures. Decreases in plant water content and increases in ABA concentration may be important for low-temperature acclimation by cacti, especially O. fragilis, which is widely distributed in Canada and the United States.

  1. Exogenous Abscisic Acid Mimics Cold Acclimation for Cacti Differing in Freezing Tolerance.

    PubMed Central

    Loik, M. E.; Nobel, P. S.

    1993-01-01

    The responses to low temperature were determined for two species of cacti sensitive to freezing, Ferocactus viridescens and Opuntia ficus-indica, and a cold hardy species, Opuntia fragilis. Fourteen days after shifting the plants from day/night air temperatures of 30/20[deg]C to 10/0[deg]C, the chlorenchyma water content decreased only for O. fragilis. This temperature shift caused the freezing tolerance (measured by vital stain uptake) of chlorenchyma cells to be enhanced only by about 2.0[deg]C for F. viridescens and O. ficus-indica but by 14.6[deg]C for O. fragilis. Also, maintenance of high water content by injection of water into plants at 10/0[deg]C reversed the acclimation. The endogenous abscisic acid (ABA) concentration was below 0.4 pmol g-1 fresh weight at 30/20[deg]C, but after 14 d at 10/0[deg]C it increased to 84 pmol g-1 fresh weight for O. ficus-indica and to 49 pmol g-1 fresh weight for O. fragilis. Four days after plants were sprayed with 7.5 x 10-5 M ABA at 30/20[deg]C, freezing tolerance was enhanced by 0.5[deg]C for F. viridescens, 4.1[deg]C for O. ficus-indica, and 23.4[deg]C for O. fragilis. Moreover, the time course for the change in freezing tolerance over 14 d was similar for plants shifted to low temperatures as for plants treated with exogenous ABA at moderate temperatures. Decreases in plant water content and increases in ABA concentration may be important for low-temperature acclimation by cacti, especially O. fragilis, which is widely distributed in Canada and the United States. PMID:12231985

  2. Crucial Role of Extracellular Polysaccharides in Desiccation and Freezing Tolerance in the Terrestrial Cyanobacterium Nostoc commune

    PubMed Central

    Tamaru, Yoshiyuki; Takani, Yayoi; Yoshida, Takayuki; Sakamoto, Toshio

    2005-01-01

    The cyanobacterium Nostoc commune is adapted to the terrestrial environment and has a cosmopolitan distribution. In this study, the role of extracellular polysaccharides (EPS) in the desiccation tolerance of photosynthesis in N. commune was examined. Although photosynthetic O2 evolution was not detected in desiccated colonies, the ability of the cells to evolve O2 rapidly recovered after rehydration. The air-dried colonies contained approximately 10% (wt/wt) water, and field-isolated, natural colonies with EPS were highly water absorbent and were rapidly hydrated by atmospheric moisture. The cells embedded in EPS in Nostoc colonies were highly desiccation tolerant, and O2 evolution was not damaged by air drying. Although N. commune was determined to be a mesophilic cyanobacterium, the cells with EPS were heat tolerant in a desiccated state. EPS could be removed from cells by homogenizing colonies with a blender and filtering with coarse filter paper. This treatment to remove EPS did not damage Nostoc cells or their ability to evolve O2, but O2 evolution was significantly damaged by desiccation treatment of the EPS-depleted cells. Similar to the EPS-depleted cells, the laboratory culture strain KU002 had only small amount of EPS and was highly sensitive to desiccation. In the EPS-depleted cells, O2 evolution was also sensitive to freeze-thaw treatment. These results strongly suggest that EPS of N. commune is crucial for the stress tolerance of photosynthesis during desiccation and during freezing and thawing. PMID:16269775

  3. Crucial role of extracellular polysaccharides in desiccation and freezing tolerance in the terrestrial cyanobacterium Nostoc commune.

    PubMed

    Tamaru, Yoshiyuki; Takani, Yayoi; Yoshida, Takayuki; Sakamoto, Toshio

    2005-11-01

    The cyanobacterium Nostoc commune is adapted to the terrestrial environment and has a cosmopolitan distribution. In this study, the role of extracellular polysaccharides (EPS) in the desiccation tolerance of photosynthesis in N. commune was examined. Although photosynthetic O2 evolution was not detected in desiccated colonies, the ability of the cells to evolve O2 rapidly recovered after rehydration. The air-dried colonies contained approximately 10% (wt/wt) water, and field-isolated, natural colonies with EPS were highly water absorbent and were rapidly hydrated by atmospheric moisture. The cells embedded in EPS in Nostoc colonies were highly desiccation tolerant, and O2 evolution was not damaged by air drying. Although N. commune was determined to be a mesophilic cyanobacterium, the cells with EPS were heat tolerant in a desiccated state. EPS could be removed from cells by homogenizing colonies with a blender and filtering with coarse filter paper. This treatment to remove EPS did not damage Nostoc cells or their ability to evolve O2, but O2 evolution was significantly damaged by desiccation treatment of the EPS-depleted cells. Similar to the EPS-depleted cells, the laboratory culture strain KU002 had only small amount of EPS and was highly sensitive to desiccation. In the EPS-depleted cells, O2 evolution was also sensitive to freeze-thaw treatment. These results strongly suggest that EPS of N. commune is crucial for the stress tolerance of photosynthesis during desiccation and during freezing and thawing.

  4. Leavening ability and freeze tolerance of yeasts isolated from traditional corn and rye bread doughs.

    PubMed

    Almeida, M J; Pais, C

    1996-12-01

    Strains of Saccharomyces cerevisiae and Torulaspora delbrueckii isolated from traditional bread doughs displayed dough-raising capacities similar to the ones found in baker's yeasts. During storage of frozen doughs, strains of T. delbrueckii (IGC 5321, IGC 5323, and IGC 4478) presented approximately the same leavening ability for 30 days. Cell viability was not significantly affected by freezing, but when the dough was submitted to a bulk fermentation before being stored at -20 degrees C, there was a decrease in the survival ratio which depended on the yeast strain. Furthermore, the leavening ability after 4 days of storage decreased as the prefermentation period of the dough before freezing increased, except for strains IGC 5321 and IGC 5323. These two strains retained their fermentative activity after 15 days of storage and 2.5 h of prefermentation, despite showing a reduction of viable cells under the same conditions. The intracellular trehalose content was higher than 20% (wt/wt) in four of the yeasts tested: the two commercial strains of baker's yeast (S. cerevisiae IGC 5325 and IGC 5326) and the two mentioned strains of T. delbrueckii (IGC 5321 and IGC 5323). However, the strains of S. cerevisiae were clearly more susceptible to freezing damages, indicating that other factors may contribute to the freeze tolerance of these yeasts.

  5. Leavening ability and freeze tolerance of yeasts isolated from traditional corn and rye bread doughs.

    PubMed Central

    Almeida, M J; Pais, C

    1996-01-01

    Strains of Saccharomyces cerevisiae and Torulaspora delbrueckii isolated from traditional bread doughs displayed dough-raising capacities similar to the ones found in baker's yeasts. During storage of frozen doughs, strains of T. delbrueckii (IGC 5321, IGC 5323, and IGC 4478) presented approximately the same leavening ability for 30 days. Cell viability was not significantly affected by freezing, but when the dough was submitted to a bulk fermentation before being stored at -20 degrees C, there was a decrease in the survival ratio which depended on the yeast strain. Furthermore, the leavening ability after 4 days of storage decreased as the prefermentation period of the dough before freezing increased, except for strains IGC 5321 and IGC 5323. These two strains retained their fermentative activity after 15 days of storage and 2.5 h of prefermentation, despite showing a reduction of viable cells under the same conditions. The intracellular trehalose content was higher than 20% (wt/wt) in four of the yeasts tested: the two commercial strains of baker's yeast (S. cerevisiae IGC 5325 and IGC 5326) and the two mentioned strains of T. delbrueckii (IGC 5321 and IGC 5323). However, the strains of S. cerevisiae were clearly more susceptible to freezing damages, indicating that other factors may contribute to the freeze tolerance of these yeasts. PMID:8953712

  6. Anthocyanin biosynthesis for cold and freezing stress tolerance and desirable color in Brassica rapa.

    PubMed

    Ahmed, Nasar Uddin; Park, Jong-In; Jung, Hee-Jeong; Hur, Yoonkang; Nou, Ill-Sup

    2015-07-01

    Flavonoids are divided into several structural classes, including anthocyanins, which provide flower and leaf colors and other derivatives that play diverse roles in plant development and interactions with the environment. This study characterized four anthocyanidin synthase (ANS) genes of Brassica rapa, a structural gene of the anthocyanin biosynthetic pathway, and investigated their association with pigment formation, cold and freezing tolerance in B. rapa. Sequences of these genes were analyzed and compared with similar gene sequences from other species, and a high degree of homology with their respective functions was found. Organ-specific expression analysis revealed that these genes were only expressed in the colored portion of leaves of different lines of B. rapa. Conversely, B. rapa anthocyanidin synthase (BrANS) genes also showed responses to cold and freezing stress treatment in B. rapa. BrANSs were also shown to be regulated by two transcription factors, BrMYB2-2 and BrTT8, contrasting with anthocyanin accumulation and cold stress. Thus, the above results suggest the association of these genes with anthocyanin biosynthesis and cold and freezing stress tolerance and might be useful resources for development of cold-resistant Brassica crops with desirable colors as well.

  7. A Multi-Environment Thermal Control System With Freeze-Tolerant Radiator

    NASA Technical Reports Server (NTRS)

    Chen, Weibo; Fogg, David; Mancini, Nick; Steele, John; Quinn, Gregory; Bue, Grant; Littibridge, Sean

    2013-01-01

    Future space exploration missions require advanced thermal control systems (TCS) to dissipate heat from spacecraft, rovers, or habitats operating in environments that can vary from extremely hot to extremely cold. A lightweight, reliable TCS is being developed to effectively control cabin and equipment temperatures under widely varying heat loads and ambient temperatures. The system uses freeze-tolerant radiators, which eliminate the need for a secondary circulation loop or heat pipe systems. Each radiator has a self-regulating variable thermal conductance to its ambient environment. The TCS uses a nontoxic, water-based working fluid that is compatible with existing lightweight aluminum heat exchangers. The TCS is lightweight, compact, and requires very little pumping power. The critical characteristics of the core enabling technologies were demonstrated. Functional testing with condenser tubes demonstrated the key operating characteristics required for a reliable, freeze-tolerant TCS, namely (1) self-regulating thermal conductance with short transient responses to varying thermal loads, (2) repeatable performance through freeze-thaw cycles, and (3) fast start-up from a fully frozen state. Preliminary coolant tests demonstrated that the corrosion inhibitor in the water-based coolant can reduce the corrosion rate on aluminum by an order of magnitude. Performance comparison with state-of-the-art designs shows significant mass and power saving benefits of this technology.

  8. Genomic regions associated with freezing tolerance and snow mold tolerance in winter wheat

    USDA-ARS?s Scientific Manuscript database

    Crops grown through the winter are subject to selective pressures that vary with each year’s unique conditions, necessitating tolerance of numerous stress factors. The objective of this study was to identify molecular markers in winter wheat (Triticum aestivum L. em Thell) associated with tolerance...

  9. Cell cycle regulation in the freeze tolerant wood frog, Rana sylvatica.

    PubMed

    Zhang, Jing; Storey, Kenneth B

    2012-05-01

    The wood frog (Rana sylvatica) is one of only a few vertebrate species that can survive extensive freezing of its body fluids during the winter. The mechanisms of natural freeze tolerance include metabolic rate depression to conserve energy and the implementation of cryoprotective strategies, especially the synthesis of huge amounts of glucose as a cryoprotectant. Liver is the main source of glucose production/export (and other cryoprotective actions) and plays a central role in freezing survival of the whole animal. Freezing is a multi-component stress that includes anoxia/ischemia due to the cessation of blood flow and dehydration of cells caused by ice accumulation in extracellular spaces. To help endure these stresses, cells need to suppress and reprioritize ATP-expensive cell functions. One of these is cell growth and proliferation, and we hypothesized that cell cycle arrest would be key to freezing survival. The present study examines the responses by key cell cycle components to freezing, anoxia and dehydration stresses in wood frog liver. Immunoblotting was used to investigate protein expression of Cdc 2, Cdks (2, 4, 6), and cyclins (A, B1, D1, E) as well as the phosphorylation states of Cdks (Thr14/Tyr15), the phosphatases Cdc25a (Ser76) and Cdc25c (Ser216) and the CIP/KIP Cdk inhibitors p21 (Thr145) and p27 (Thr187). Responses to 24 h freezing, 24 h anoxia and 40% dehydration as well as recovery from these stresses were analyzed. The results showed very similar responses by cell cycle components to anoxia or dehydration and were consistent with cell cycle suppression under stress and reversal during recovery. Freezing showed elements of cell cycle suppression, including reduced protein levels of Cdks and cyclins A and B1, but also showed unique responses by cyclin D1, Cdc25 phosphatases and p21/p27. These may be linked with alternative actions by these proteins that contribute to cryoprotection; e.g., an alternative action of cyclin D1 as a transcription

  10. Lipid transfer protein 3 as a target of MYB96 mediates freezing and drought stress in Arabidopsis

    PubMed Central

    Yang, Shuhua

    2013-01-01

    Several lipid-transfer proteins were reported to modulate the plant response to biotic stress; however, whether lipid-transfer proteins are also involved in abiotic stress remains unknown. This study characterized the function of a lipid-transfer protein, LTP3, during freezing and drought stress. LTP3 was expressed ubiquitously and the LTP3 protein was localized to the cytoplasm. A biochemical study showed that LTP3 was able to bind to lipids. Overexpression of LTP3 resulted in constitutively enhanced freezing tolerance without affecting the expression of CBFs and their target COR genes. Further analyses showed that LTP3 was positively regulated by MYB96 via the direct binding to the LTP3 promoter; consistently, transgenic plants overexpressing MYB96 exhibited enhanced freezing tolerance. This study also found that the loss-of-function mutant ltp3 was sensitive to drought stress, whereas overexpressing plants were drought tolerant, phenotypes reminiscent of myb96 mutant plants and MYB96-overexpressing plants. Taken together, these results demonstrate that LTP3 acts as a target of MYB96 to be involved in plant tolerance to freezing and drought stress. PMID:23404903

  11. Ice-Active Substances from the Infective Juveniles of the Freeze Tolerant Entomopathogenic Nematode, Steinernema feltiae.

    PubMed

    Ali, Farman; Wharton, David A

    2016-01-01

    Steinernema feltiae is a moderately freezing tolerant nematode, that can withstand intracellular ice formation. We investigated recrystallization inhibition, thermal hysteresis and ice nucleation activities in the infective juveniles of S. feltiae. Both the splat cooling assay and optical recrystallometry indicate the presence of ice active substances that inhibit recrystallization in the nematode extract. The substance is relatively heat stable and largely retains the recrystallization inhibition activity after heating. No thermal hysteresis activity was detected but the extract had a typical hexagonal crystal shape when grown from a single seed crystal and weak ice nucleation activity. An ice active substance is present in a low concentration, which may be involved in the freezing survival of this species by inhibiting ice recrystallization.

  12. Ice-Active Substances from the Infective Juveniles of the Freeze Tolerant Entomopathogenic Nematode, Steinernema feltiae

    PubMed Central

    Ali, Farman; Wharton, David A.

    2016-01-01

    Steinernema feltiae is a moderately freezing tolerant nematode, that can withstand intracellular ice formation. We investigated recrystallization inhibition, thermal hysteresis and ice nucleation activities in the infective juveniles of S. feltiae. Both the splat cooling assay and optical recrystallometry indicate the presence of ice active substances that inhibit recrystallization in the nematode extract. The substance is relatively heat stable and largely retains the recrystallization inhibition activity after heating. No thermal hysteresis activity was detected but the extract had a typical hexagonal crystal shape when grown from a single seed crystal and weak ice nucleation activity. An ice active substance is present in a low concentration, which may be involved in the freezing survival of this species by inhibiting ice recrystallization. PMID:27227961

  13. Reversion of nitrate tolerance in rat aorta rings by freeze-dried red wine.

    PubMed

    Fusi, Fabio; Sgaragli, Giampietro

    2015-04-01

    Chronically administered organic nitrates induce nitrate tolerance and endothelial dysfunction, which limit their therapeutic use. eNOS uncoupling, ROS over-production, aldehyde dehydrogenase-2 as well as superoxide dismutase (SOD) oxidative inhibition, and cGMP desensitization are thought to play an important role. Natural polyphenols are effective antioxidants, which might counteract the mechanisms leading to nitrate tolerance. The aim of this work was to verify whether freeze-dried (dealcoholized) red wine (FDRW) was able to revert glyceryl trinitrate (GTN) tolerance and endothelial dysfunction induced in rat aorta rings with either GTN or diethyldithiocarbamate (DETCA), an irreversible inhibitor of Cu/Zn SOD. GTN induced a concentration-dependent relaxation of rings pre-contracted with phenylephrine. GTN spasmolysis was significantly reduced in rings pre-incubated with either GTN or DETCA. FDRW, at 2.8 µg of gallic acid equivalents (GAE)/mL concentration, was able to revert partially, though significantly, GTN-induced tolerance but not tolerance and endothelial dysfunction induced by DETCA. This work provides the first evidence in vitro that red wine components, at concentrations comparable to those achieved in human blood after moderate consumption of red wine, revert tolerance to nitrates with a mechanism possibly mediated by SOD. Copyright © 2015 John Wiley & Sons, Ltd.

  14. Superoxide dismutase enhances tolerance of freezing stress in transgenic alfalfa (Medicago sativa L.).

    PubMed Central

    McKersie, B D; Chen, Y; de Beus, M; Bowley, S R; Bowler, C; Inzé, D; D'Halluin, K; Botterman, J

    1993-01-01

    Activated oxygen or oxygen free radicals have been implicated in a number of physiological disorders in plants including freezing injury. Superoxide dismutase (SOD) catalyzes the dismutation of superoxide into O2 and H2O2 and thereby reduces the titer of activated oxygen molecules in the cell. To further examine the relationship between oxidative and freezing stresses, the expression of SOD was modified in transgenic alfalfa (Medicago sativa L.). The Mn-SOD cDNA from Nicotiana plumbaginifolia under the control of the cauliflower mosaic virus 35S promoter was introduced into alfalfa using Agrobacterium tumefaciens-mediated transformation. Two plasmid vectors, pMitSOD and pChlSOD, contained a chimeric Mn-SOD construct with a transit peptide for targeting to the mitochondria or one for targeting to the chloroplast, respectively. The putatively transgenic plants were selected for resistance to kanamycin and screened for neomycin phosphotransferase activity and the presence of an additional Mn-SOD isozyme. Detailed analysis of a set of four selected transformants indicated that some had enhanced SOD activity, increased tolerance to the diphenyl ether herbicide, acifluorfen, and increased regrowth after freezing stress. The F1 progeny of one line, RA3-ChlSOD-30, were analyzed by SOD isozyme activity, by polymerase chain reaction for the Mn-SOD gene, and by polymerase chain reaction for the neo gene. RA3-ChlSOD-30 had three sites of insertion of pChlSOD, but only one gave a functional Mn-SOD isozyme; the other two were apparently partial insertions. The progeny with a functional Mn-SOD transgene had more rapid regrowth following freezing stress than those progeny lacking the functional Mn-SOD transgene, suggesting that Mn-SOD serves a protective role by minimizing oxygen free radical production after freezing stress. PMID:8290627

  15. Metabolic mechanisms for anoxia tolerance and freezing survival in the intertidal gastropod, Littorina littorea.

    PubMed

    Storey, Kenneth B; Lant, Benjamin; Anozie, Obiajulu O; Storey, Janet M

    2013-08-01

    The gastropod mollusk, Littorina littorea L., is a common inhabitant of the intertidal zone along rocky coastlines of the north Atlantic. This species has well-developed anoxia tolerance and freeze tolerance and is extensively used as a model for exploring the biochemical adaptations that support these tolerances as well as for toxicological studies aimed at identifying effective biomarkers of aquatic pollution. This article highlights our current understanding of the molecular mechanisms involved in anaerobiosis and freezing survival of periwinkles, particularly with respect to anoxia-induced metabolic rate depression. Analysis of foot muscle and hepatopancreas metabolism includes anoxia-responsive changes in enzyme regulation, signal transduction, gene expression, post-transcriptional regulation of mRNA, control of translation, and cytoprotective strategies including chaperones and antioxidant defenses. New studies describe the regulation of glucose-6-phosphate dehydrogenase by reversible protein phosphorylation, the role of microRNAs in suppressing mRNA translation in the hypometabolic state, modulation of glutathione S-transferase isozyme patterns, and the regulation of the unfolded protein response. Copyright © 2013 Elsevier Inc. All rights reserved.

  16. The Arabidopsis-related halophyte Thellungiella halophila: boron tolerance via boron complexation with metabolites?

    PubMed

    Lamdan, Netta Li; Attia, Ziv; Moran, Nava; Moshelion, Menachem

    2012-04-01

    Tolerance to boron (B) is still not completely understood. We tested here the hypothesis that Thellungiella halophila, an Arabidopsis thaliana-related 'extremophile' plant, with abundance of B in its natural environment, is tolerant to B, and examined the potential mechanisms of this tolerance. With 1-10 mm B applied ([B](ext)) to Thellungiella and Arabidopsis grown in hydroponics, the steady-state accumulated B concentration ([B](int)) in the root was below [B](ext), and was similar in both, suggesting both extrude B actively. Whether grown in soil or hydroponically, the shoot [B](int) was higher in Arabidopsis than in Thellungiella, suggesting more effective net B exclusion by Thellungiella root. Arabidopsis exhibited toxicity symptoms including reduced shoot fresh weight (FW), but Thellungiella was not affected, even at similar levels of shoot-accumulated [B](int) (about 10 to 40 mm B in 'shoot water'), suggesting additional B tolerance mechanism in Thellungiella shoot. At [B](ext) = 5 mm, the summed shoot concentration of the potentially B-binding polyhydroxyl metabolites (malic acid, fructose, glucose, sucrose and citric acid) in Arabidopsis was below [B](int) , but in Thellungiella it was over twofold higher than [B](int) , and therefore likely to allow appreciable 1:2 boron-metabolite complexation in the shoot. This, we suggest, is an important component of Thellungiella B tolerance mechanism.

  17. Overexpression of the Vacuolar Sugar Carrier AtSWEET16 Modifies Germination, Growth, and Stress Tolerance in Arabidopsis1[W

    PubMed Central

    Klemens, Patrick A.W.; Patzke, Kathrin; Deitmer, Joachim; Spinner, Lara; Le Hir, Rozenn; Bellini, Catherine; Bedu, Magali; Chardon, Fabien; Krapp, Anne; Neuhaus, H. Ekkehard

    2013-01-01

    Here, we report that SUGARS WILL EVENTUALLY BE EXPORTED TRANSPORTER (SWEET16) from Arabidopsis (Arabidopsis thaliana) is a vacuole-located carrier, transporting glucose (Glc), fructose (Fru), and sucrose (Suc) after heterologous expression in Xenopus laevis oocytes. The SWEET16 gene, similar to the homologs gene SWEET17, is mainly expressed in vascular parenchyma cells. Application of Glc, Fru, or Suc, as well as cold, osmotic stress, or low nitrogen, provoke the down-regulation of SWEET16 messenger RNA accumulation. SWEET16 overexpressors (35SPro:SWEET16) showed a number of peculiarities related to differences in sugar accumulation, such as less Glc, Fru, and Suc at the end of the night. Under cold stress, 35SPro:SWEET16 plants are unable to accumulate Fru, while under nitrogen starvation, both Glc and Fru, but not Suc, were less abundant. These changes of individual sugars indicate that the consequences of an increased SWEET16 activity are dependent upon the type of external stimulus. Remarkably, 35SPro:SWEET16 lines showed improved germination and increased freezing tolerance. The latter observation, in combination with the modified sugar levels, points to a superior function of Glc and Suc for frost tolerance. 35SPro:SWEET16 plants exhibited increased growth efficiency when cultivated on soil and showed improved nitrogen use efficiency when nitrate was sufficiently available, while under conditions of limiting nitrogen, wild-type biomasses were higher than those of 35SPro:SWEET16 plants. Our results identify SWEET16 as a vacuolar sugar facilitator, demonstrate the substantial impact of SWEET16 overexpression on various critical plant traits, and imply that SWEET16 activity must be tightly regulated to allow optimal Arabidopsis development under nonfavorable conditions. PMID:24028846

  18. Overexpression of the vacuolar sugar carrier AtSWEET16 modifies germination, growth, and stress tolerance in Arabidopsis.

    PubMed

    Klemens, Patrick A W; Patzke, Kathrin; Deitmer, Joachim; Spinner, Lara; Le Hir, Rozenn; Bellini, Catherine; Bedu, Magali; Chardon, Fabien; Krapp, Anne; Neuhaus, H Ekkehard

    2013-11-01

    Here, we report that SUGARS WILL EVENTUALLY BE EXPORTED TRANSPORTER (SWEET16) from Arabidopsis (Arabidopsis thaliana) is a vacuole-located carrier, transporting glucose (Glc), fructose (Fru), and sucrose (Suc) after heterologous expression in Xenopus laevis oocytes. The SWEET16 gene, similar to the homologs gene SWEET17, is mainly expressed in vascular parenchyma cells. Application of Glc, Fru, or Suc, as well as cold, osmotic stress, or low nitrogen, provoke the down-regulation of SWEET16 messenger RNA accumulation. SWEET16 overexpressors (35SPro:SWEET16) showed a number of peculiarities related to differences in sugar accumulation, such as less Glc, Fru, and Suc at the end of the night. Under cold stress, 35SPro:SWEET16 plants are unable to accumulate Fru, while under nitrogen starvation, both Glc and Fru, but not Suc, were less abundant. These changes of individual sugars indicate that the consequences of an increased SWEET16 activity are dependent upon the type of external stimulus. Remarkably, 35SPro:SWEET16 lines showed improved germination and increased freezing tolerance. The latter observation, in combination with the modified sugar levels, points to a superior function of Glc and Suc for frost tolerance. 35SPro:SWEET16 plants exhibited increased growth efficiency when cultivated on soil and showed improved nitrogen use efficiency when nitrate was sufficiently available, while under conditions of limiting nitrogen, wild-type biomasses were higher than those of 35SPro:SWEET16 plants. Our results identify SWEET16 as a vacuolar sugar facilitator, demonstrate the substantial impact of SWEET16 overexpression on various critical plant traits, and imply that SWEET16 activity must be tightly regulated to allow optimal Arabidopsis development under nonfavorable conditions.

  19. Annual variation in glycerol mobilization and effect of freeze rigor on post-thaw locomotion in the freeze-tolerant frog Hyla versicolor.

    PubMed

    Layne, Jack R; Stapleton, Michael G

    2009-02-01

    This study documents post-thaw recovery of jump distance and cryoprotectant mobilization in the freeze-tolerant frog Hyla versicolor over two successive years. Cold acclimated frogs had plasma glycerol levels near 1.0 mM in 2004 but it was nearly 70x higher during 2005. Freezing of frogs induced nearly identical levels of plasma glycerol (ca. 177 mM) during 2004 and 2005. Plasma glucose was only mobilized upon somatic freezing, with averages ranging between 21 and 36 mM. Control jump distance showed no difference between the two years of the study. The post-thaw jump response was identical during the first 2 years despite large differences in glycerol mobilization between these 2 years. Recovery proceeded much faster in 2005 when frogs mobilized glycerol prior to freeze exposure. Frogs were more impaired in their locomotion performance during the initial stages of recovery period when they were frozen at a lower temperature (-3 vs. -1.5 degrees C) but they eventually recovered. Moderate lengthening of the freeze duration (3 vs. 7 days) with the 2004 collection group did not affect recovery of jump distance when frogs were frozen at -1.5 degrees C. Hence, postfreeze impairment of locomotion is dependent of the intensity of the freeze temperature but it is a reversible process that is mitigated when glycerol is more freely distributed to body tissues.

  20. Identification of two hydrophilins that contribute to the desiccation and freezing tolerance of yeast (Saccharomyces cerevisiae) cells.

    PubMed

    Dang, Nghiem X; Hincha, Dirk K

    2011-06-01

    Hydrophilins are a group of proteins that are present in all organisms and that have been defined as being highly hydrophilic and rich in glycine. They are assumed to play important roles in cellular dehydration tolerance. There are 12 genes in the yeast Saccharomyces cerevisiae that encode hydrophilins and most of these genes are stress responsive. However, the functional role of yeast hydrophilins, especially in desiccation and freezing tolerance, is largely unknown. Here, we selected six candidate hydrophilins for further analysis. All six proteins were predicted to be intrinsically disordered, i.e. to have no stable structure in solution. The contribution of these proteins to the desiccation and freezing tolerance of yeast was investigated in the respective knock-out strains. Only the disruption of the genes YJL144W and YMR175W (SIP18) resulted in significantly reduced desiccation tolerance, while none of the strains was affected in its freezing tolerance under our experimental conditions. Complementation experiments showed that yeast cells overexpressing these two genes were both more desiccation and freezing tolerant, confirming the role of these two hydrophilins in yeast dehydration stress tolerance.

  1. Accumulation of lactate by frozen painted turtles (Chrysemys picta) and its relationship to freeze tolerance.

    PubMed

    Packard, Mary J; Packard, Gary C

    2004-01-01

    Hatchling painted turtles (Chrysemys picta) survived freezing at -2 degrees C for 4 d, few recovered from freezing lasting 6 d, and none survived being frozen for 8 d. Whole-body glucose and lactate were low in animals that had not been subjected to cold and ice but increased precipitously in animals that were frozen for 2 d. Both metabolites continued to increase, but at a somewhat lower rate, in animals frozen for 4, 6, or 8 d. The increase in whole-body lactate reflects a reliance by frozen hatchlings on anaerobiosis, whereas the increase in glucose presumably results from mobilization of glycogen reserves to support anaerobic metabolism. Mortality of frozen hatchlings is correlated with the increase in whole-body lactate. Factors that may contribute to the observed correlation include a compromised capacity for individual organs to cope with the lactic acidosis that accompanies anaerobic metabolism and organ-specific depletion of energy reserves. Individual organs must rely on buffering and glucose reserves available in situ because blood of frozen hatchlings does not circulate. Thus, buffer from the shell cannot be transported to other organs, lactate cannot be sequestered in the shell, and glucose mobilized from liver glycogen is not available to supplement glucose reserves of other tissues. This integrated suite of physiological disruptions may limit tolerance of freezing to conditions with little or no ecological relevance.

  2. Aluminum-activated citrate and malate transporters from the MATE and ALMT families function independently to confer Arabidopsis aluminum tolerance

    USDA-ARS?s Scientific Manuscript database

    Aluminum (Al) activated root malate and citrate exudation play an important role in Al tolerance in many plant species. AtALMT1, an Al-activated malate transporter, is a major contributor to Arabidopsis Al tolerance. Here, we demonstrate that a second, unrelated gene, AtMATE, encodes an Arabidopsi...

  3. Fluidization of membrane lipids enhances the tolerance of Saccharomyces cerevisiae to freezing and salt stress.

    PubMed

    Rodríguez-Vargas, Sonia; Sánchez-García, Alicia; Martínez-Rivas, Jose Manuel; Prieto, Jose Antonio; Randez-Gil, Francisca

    2007-01-01

    Unsaturated fatty acids play an essential role in the biophysical characteristics of cell membranes and determine the proper function of membrane-attached proteins. Thus, the ability of cells to alter the degree of unsaturation in their membranes is an important factor in cellular acclimatization to environmental conditions. Many eukaryotic organisms can synthesize dienoic fatty acids, but Saccharomyces cerevisiae can introduce only a single double bond at the Delta(9) position. We expressed two sunflower (Helianthus annuus) oleate Delta(12) desaturases encoded by FAD2-1 and FAD2-3 in yeast cells of the wild-type W303-1A strain (trp1) and analyzed their effects on growth and stress tolerance. Production of the heterologous desaturases increased the content of dienoic fatty acids, especially 18:2Delta(9,12), the unsaturation index, and the fluidity of the yeast membrane. The total fatty acid content remained constant, and the level of monounsaturated fatty acids decreased. Growth at 15 degrees C was reduced in the FAD2 strains, probably due to tryptophan auxotrophy, since the trp1 (TRP1) transformants that produced the sunflower desaturases grew as well as the control strain did. Our results suggest that changes in the fluidity of the lipid bilayer affect tryptophan uptake and/or the correct targeting of tryptophan transporters. The expression of the sunflower desaturases, in either Trp(+) or Trp(-) strains, increased NaCl tolerance. Production of dienoic fatty acids increased the tolerance to freezing of wild-type cells preincubated at 30 degrees C or 15 degrees C. Thus, membrane fluidity is an essential determinant of stress resistance in S. cerevisiae, and engineering of membrane lipids has the potential to be a useful tool of increasing the tolerance to freezing in industrial strains.

  4. Fluidization of Membrane Lipids Enhances the Tolerance of Saccharomyces cerevisiae to Freezing and Salt Stress▿

    PubMed Central

    Rodríguez-Vargas, Sonia; Sánchez-García, Alicia; Martínez-Rivas, Jose Manuel; Prieto, Jose Antonio; Randez-Gil, Francisca

    2007-01-01

    Unsaturated fatty acids play an essential role in the biophysical characteristics of cell membranes and determine the proper function of membrane-attached proteins. Thus, the ability of cells to alter the degree of unsaturation in their membranes is an important factor in cellular acclimatization to environmental conditions. Many eukaryotic organisms can synthesize dienoic fatty acids, but Saccharomyces cerevisiae can introduce only a single double bond at the Δ9 position. We expressed two sunflower (Helianthus annuus) oleate Δ12 desaturases encoded by FAD2-1 and FAD2-3 in yeast cells of the wild-type W303-1A strain (trp1) and analyzed their effects on growth and stress tolerance. Production of the heterologous desaturases increased the content of dienoic fatty acids, especially 18:2Δ9,12, the unsaturation index, and the fluidity of the yeast membrane. The total fatty acid content remained constant, and the level of monounsaturated fatty acids decreased. Growth at 15°C was reduced in the FAD2 strains, probably due to tryptophan auxotrophy, since the trp1 (TRP1) transformants that produced the sunflower desaturases grew as well as the control strain did. Our results suggest that changes in the fluidity of the lipid bilayer affect tryptophan uptake and/or the correct targeting of tryptophan transporters. The expression of the sunflower desaturases, in either Trp+ or Trp− strains, increased NaCl tolerance. Production of dienoic fatty acids increased the tolerance to freezing of wild-type cells preincubated at 30°C or 15°C. Thus, membrane fluidity is an essential determinant of stress resistance in S. cerevisiae, and engineering of membrane lipids has the potential to be a useful tool of increasing the tolerance to freezing in industrial strains. PMID:17071783

  5. Water Relations and Low-Temperature Acclimation for Cactus Species Varying in Freezing Tolerance.

    PubMed

    Goldstein, G.; Nobel, P. S.

    1994-02-01

    Opuntia ficus-indica and Opuntia streptacantha are widely cultivated cacti that can tolerate temperatures no lower than -10[deg]C, whereas Opuntia humifusa, which is native to southern Canada and the eastern United States, can tolerate -24[deg]C. As day/night air temperatures were decreased from 30/20 to 10/0[deg]C, the osmotic pressure increased 0.10 MPa for O. ficus-indica and O. streptacantha but 0.38 MPa for O. humifusa. The increases in osmotic pressures were due mostly to the synthesis of fructose, glucose, and sucrose. In addition, O. humifusa produced a substantial amount of mannitol during exposure to low temperatures. Substantial accumulation of sugars and mannitol in cells of O. humifusa may help prevent intracellular freeze dehydration and ice formation as well as provide noncolligative protection to its membranes. Mucilage was slightly higher in all three species at the lower temperatures. Extracellular nucleation of ice occurred closer to the equilibrium freezing temperature for plants at 10/0[deg]C compared with 30/20[deg]C, which could make the cellular dehydration more gradual and, thus, less damaging. Results from nuclear magnetic resonance indicated a restricted mobility of intracellular water at the lower temperatures, especially for O. humifusa, which is consistent with its lower water content and higher levels of low molecular weight solutes.

  6. Water Relations and Low-Temperature Acclimation for Cactus Species Varying in Freezing Tolerance.

    PubMed Central

    Goldstein, G.; Nobel, P. S.

    1994-01-01

    Opuntia ficus-indica and Opuntia streptacantha are widely cultivated cacti that can tolerate temperatures no lower than -10[deg]C, whereas Opuntia humifusa, which is native to southern Canada and the eastern United States, can tolerate -24[deg]C. As day/night air temperatures were decreased from 30/20 to 10/0[deg]C, the osmotic pressure increased 0.10 MPa for O. ficus-indica and O. streptacantha but 0.38 MPa for O. humifusa. The increases in osmotic pressures were due mostly to the synthesis of fructose, glucose, and sucrose. In addition, O. humifusa produced a substantial amount of mannitol during exposure to low temperatures. Substantial accumulation of sugars and mannitol in cells of O. humifusa may help prevent intracellular freeze dehydration and ice formation as well as provide noncolligative protection to its membranes. Mucilage was slightly higher in all three species at the lower temperatures. Extracellular nucleation of ice occurred closer to the equilibrium freezing temperature for plants at 10/0[deg]C compared with 30/20[deg]C, which could make the cellular dehydration more gradual and, thus, less damaging. Results from nuclear magnetic resonance indicated a restricted mobility of intracellular water at the lower temperatures, especially for O. humifusa, which is consistent with its lower water content and higher levels of low molecular weight solutes. PMID:12232118

  7. Freezing tolerance of sea urchin embryonic cells: Differentiation commitment and cytoskeletal disturbances in culture.

    PubMed

    Odintsova, Nelly A; Ageenko, Natalya V; Kipryushina, Yulia O; Maiorova, Mariia A; Boroda, Andrey V

    2015-08-01

    This study focuses on the freezing tolerance of sea urchin embryonic cells. To significantly reduce the loss of physiological activity of these cells that occurs after cryopreservation and to study the effects of ultra-low temperatures on sea urchin embryonic cells, we tested the ability of the cells to differentiate into spiculogenic or pigment directions in culture, including an evaluation of the expression of some genes involved in pigment differentiation. A morphological analysis of cytoskeletal disturbances after freezing in a combination of penetrating (dimethyl sulfoxide and ethylene glycol) and non-penetrating (trehalose and polyvinylpyrrolidone) cryoprotectants revealed that the distribution pattern of filamentous actin and tubulin was similar to that in the control cultures. In contrast, very rare spreading cells and a small number of cells with filamentous actin and tubulin were detected after freezing in the presence of only non-penetrating cryoprotectants. The largest number of pigment cells was found in cultures frozen with trehalose or trehalose and dimethyl sulfoxide. The ability to induce the spicule formation was lost in the cells frozen only with non-penetrating cryoprotectants, while it was maximal in cultures frozen in a cryoprotective mixture containing both non-penetrating and penetrating cryoprotectants (particularly, when ethylene glycol was present). Using different markers for cell state assessment, an effective cryopreservation protocol for sea urchin cells was developed: three-step freezing with a low cooling rate (1-2°C/min) and a combination of non-penetrating and penetrating cryoprotectants made it possible to obtain a high level of cell viability (up to 65-80%). Copyright © 2015 Elsevier Inc. All rights reserved.

  8. Transgenic expression of TaMYB2A confers enhanced tolerance to multiple abiotic stresses in Arabidopsis.

    PubMed

    Mao, Xinguo; Jia, Dongsheng; Li, Ang; Zhang, Hongying; Tian, Shanjun; Zhang, Xiaoke; Jia, Jizeng; Jing, Ruilian

    2011-09-01

    Osmotic stresses such as drought, salinity, and cold are major environmental factors that limit agricultural productivity. Transcription factors play essential roles in abiotic stress signaling in plants. Three TaMYB2 members were identified and designated TaMYB2A, TaMYB2B, and TaMYB2D based on their genomic origins. The cis-regulatory elements in the promoter regions were compared, and their diverse expression patterns under different abiotic stress conditions were identified. TaMYB2A was further characterized because of its earlier response to stresses. Subcellular localization revealed that TaMYB2A localized in the nucleus. To examine the role of TaMYB2A under various environmental stresses, transgenic Arabidopsis plants carrying TaMYB2A controlled by the CaMV 35S promoter were generated and subjected to severe abiotic stress. TaMYB2A transgenics had enhanced tolerance to drought, salt, and freezing stresses, which were confirmed by the enhanced expressions of abiotic stress-responsive genes and several physiological indices, including decreased rate of water loss, enhanced cell membrane stability, improved photosynthetic potential, and reduced osmotic potential. TaMYB2A is a multifunctional regulatory factor. Its overexpression confers enhanced tolerance to multiple abiotic stresses while having no obvious negative effects on phenotype under well-watered and stressed conditions; thus, TaMYB2A has the potential for utilization in transgenic breeding to improve abiotic stress tolerances in crops.

  9. Enzymatic Regulation of Glycogenolysis in a Subarctic Population of the Wood Frog: Implications for Extreme Freeze Tolerance

    PubMed Central

    do Amaral, M. Clara F.; Lee, Richard E.; Costanzo, Jon P.

    2013-01-01

    The wood frog, Rana sylvatica, from Interior Alaska survives freezing at –16°C, a temperature 10–13°C below that tolerated by its southern conspecifics. We investigated the hepatic freezing response in this northern phenotype to determine if its profound freeze tolerance is associated with an enhanced glucosic cryoprotectant system. Alaskan frogs had a larger liver glycogen reserve that was mobilized faster during early freezing as compared to conspecifics from a cool-temperate region (southern Ohio, USA). In Alaskan frogs the rapid glucose production in the first hours of freezing was associated with a 7-fold increase in glycogen phosphorylase activity above unfrozen frog levels, and the activity of this enzyme was higher than that of frozen Ohioan frogs. Freezing of Ohioan frogs induced a more modest (4-fold) increase in glycogen phosphorylase activity above unfrozen frog values. Relative to the Ohioan frogs, Alaskan frogs maintained a higher total protein kinase A activity throughout an experimental freezing/thawing time course, and this may have potentiated glycogenolysis during early freezing. We found populational variation in the activity and protein level of protein kinase A which suggested that the Alaskan population had a more efficient form of this enzyme. Alaskan frogs modulated their glycogenolytic response by decreasing the activity of glycogen phosphorylase after cryoprotectant mobilization was well under way, thereby conserving their hepatic glycogen reserve. Ohioan frogs, however, sustained high glycogen phosphorylase activity until early thawing and consumed nearly all their liver glycogen. These unique hepatic responses of Alaskan R. sylvatica likely contribute to this phenotype’s exceptional freeze tolerance, which is necessary for their survival in a subarctic climate. PMID:24236105

  10. Latitudinal variation of freeze tolerance in intertidal marine snails of the genus Melampus (Gastropoda: Ellobiidae).

    PubMed

    Dennis, A B; Loomis, S H; Hellberg, M E

    2014-01-01

    Abstract Low temperatures limit the poleward distribution of many species such that the expansion of geographic range can only be accomplished via evolutionary innovation. We have tested for physiological differences among closely related species to determine whether their poleward latitudinal ranges are limited by tolerance to cold. We measured lower temperature tolerance (LT50) among a group of intertidal pulmonate snails from six congeneric species and nine locales. Differences in tolerance are placed in the context of a molecular phylogeny based on one mitochondrial (cytochrome oxidase subunit I) and two nuclear (histone 3 and a mitochondrial phosphate carrier protein) markers. Temperate species from two separate lineages had significantly lower measures of LT50 than related tropical species. Range differences within the temperate zone, however, were not explained by LT50. These results show that multiple adaptations to cold and freezing may have enabled range expansions out of the tropics in Melampus. However, northern range limits within temperate species are not governed by cold tolerance alone.

  11. Improving freeze-tolerance of baker's yeast through seamless gene deletion of NTH1 and PUT1.

    PubMed

    Dong, Jian; Chen, Didi; Wang, Guanglu; Zhang, Cuiying; Du, Liping; Liu, Shanshan; Zhao, Yu; Xiao, Dongguang

    2016-06-01

    Baker's yeast strains with freeze-tolerance are highly desirable to maintain high leavening ability after freezing. Enhanced intracellular concentration of trehalose and proline in yeast is linked with freeze-tolerance. In this study, we constructed baker's yeast with enhanced freeze-tolerance by simultaneous deletion of the neutral trehalase-encoded gene NTH1 and the proline oxidase-encoded gene PUT1. We first used the two-step integration-based seamless gene deletion method to separately delete NTH1 and PUT1 in haploid yeast. Subsequently, through two rounds of hybridization and sporulation-based allelic exchange and colony PCR-mediated tetrad analysis, we obtained strains with restored URA3 and deletion of NTH1 and/or PUT1. The resulting strain showed higher cell survival and dough-leavening ability after freezing compared to the wild-type strain due to enhanced accumulation of trehalose and/or proline. Moreover, mutant with simultaneous deletion of NTH1 and PUT1 exhibits the highest relative dough-leavening ability after freezing compared to mutants with single-gene deletion perhaps due to elevated levels of both trehalose and proline. These results verified that it is applicable to construct frozen dough baker's yeast using the method proposed in this paper.

  12. Freezing tolerance and avoidance in high tropical Andean plants: Is it equally represented in species with different plant height?

    PubMed

    Squeo, F A; Rada, F; Azocar, A; Goldstein, G

    1991-05-01

    Freezing tolerance and avoidance were studied in several different sized species of the tropical high Andes (4200 m) to determine whether there was a relationship between plant height and cold resistance mechanisms. Freezing injury and supercooling capacity were determined in ground level plants (i.e. cushions, small rosettes and a perennial herb), intermediate height plants (shrubs and perennial herbs) and arborescent forms (i.e. giant rosettes and small trees). All ground-level plants showed tolerance as the main mechanism of resistance to cold temperatures. Arborescent forms showed avoidance mechanisms mainly through supercooling, while intermediate plants exhibited both. Insulation mechanisms to avoid low temperatures were present in the two extreme life-forms. We suggest that a combination of freezing tolerance and avoidance by insulation is least expensive and is a more secure mechanism for high tropical mountain plants than supercooling alone.

  13. Aquaporins play a role in desiccation and freeze tolerance in larvae of the goldenrod gall fly, Eurosta solidaginis.

    PubMed

    Philip, Benjamin N; Yi, Shu-Xia; Elnitsky, Michael A; Lee, Richard E

    2008-04-01

    Survival of freezing not only requires organisms to tolerate ice formation within their body, but also depends on the rapid redistribution of water and cryoprotective compounds between intra- and extracellular compartments. Aquaporins are transmembrane proteins that serve as the major pathway through which water and small uncharged solutes (e.g. glycerol) enter and leave the cell. Consequently, we examined freeze-tolerant larvae of the goldenrod gall fly, Eurosta solidaginis, to determine whether aquaporins are present and if their presence promotes freeze tolerance of specific tissues. Immunoblotting with mammalian anti-AQP2, -AQP3 and -AQP4 revealed corresponding aquaporin homologues in E. solidaginis, whose patterns of expression varied depending on acclimation temperature and desiccation treatment. To examine the role of aquaporins in freeze tolerance, we froze fat body, midgut and salivary gland tissues in the presence and absence of mercuric chloride, an aquaporin inhibitor. Survival of fat body and midgut cells was significantly reduced when mercuric chloride was present. In contrast, survival of the salivary gland did not decrease when it was frozen with mercuric chloride. Overall, this study supports our hypothesis that naturally occurring aquaporins in E. solidaginis are regulated during desiccation and promote cell survival during freezing.

  14. Overexpression of a bacterial mercury transporter MerT in Arabidopsis enhances mercury tolerance.

    PubMed

    Xu, Sheng; Sun, Bin; Wang, Rong; He, Jia; Xia, Bing; Xue, Yong; Wang, Ren

    2017-08-19

    The phytoremediation by using of green plants in the removal of environmental pollutant is an environment friendly, green technology that is cost effective and energetically inexpensive. By using Agrobacterium-mediated gene transfer, we generated transgenic Arabidopsis plants ectopically expressing mercuric transport protein gene (merT) from Pseudomonas alcaligenes. Compared with wild-type (WT) plants, overexpressing PamerT in Arabidopsis enhanced the tolerance to HgCl2. Further results showed that the enhanced total activities or corresponding transcripts of antioxidant enzymes, including superoxide dismutase (SOD), catalase (CAT) and guaiacol peroxidase (POD) were observed in transgenic Arabidopsis under HgCl2 stress. These results were confirmed by the alleviation of oxidative damage, as indicated by the decrease of thiobarbituric acid reactive substances (TBARS) contents and reactive oxygen species (ROS) accumulation. In addition, localization analysis of PaMerT in Arabidopsis protoplast showed that it is likely to be associated with vacuole. In all, PamerT increased mercury (Hg) tolerance in transgenic Arabidopsis, and decreased production of Hg-induced ROS, thereby protecting plants from oxidative damage. The present study has provided further evidence that bacterial MerT plays an important role in the plant tolerance to HgCl2 and in reducing the production of ROS induced by HgCl2. Copyright © 2017 Elsevier Inc. All rights reserved.

  15. Comparison of the ultrastructure of conventionally fixed and high pressure frozen/freeze substituted root tips of Nicotiana and Arabidopsis

    NASA Technical Reports Server (NTRS)

    Kiss, J. Z.; Giddings, T. H. Jr; Staehelin, L. A.; Sack, F. D.

    1990-01-01

    To circumvent the limitations of chemical fixation (CF) and to gain more reliable structural information about higher plant tissues, we have cryofixed root tips of Nicotiana and Arabidopsis by high pressure freezing (HPF). Whereas other freezing techniques preserve tissue to a relatively shallow depth, HPF in conjunction with freeze substitution (FS) resulted in excellent preservation of entire root tips. Compared to CF, in tissue prepared by HPF/FS: (1) the plasmalemma and all internal membranes were much smoother and often coated on the cytoplasmic side by a thin layer of stained material, (2) the plasmalemma was appressed to the cell wall, (3) organelle profiles were rounder, (4) the cytoplasmic, mitochondrial, and amyloplast matrices were denser, (5) vacuoles contained electron dense material, (6) microtubules appeared to be more numerous and straighter, with crossbridges observed between them, (7) cisternae of endoplasmic reticulum (ER) were wider and filled with material, (8) Golgi intercisternal elements were more clearly resolved and were observed between both Golgi vesicles and cisternae, and (9) larger vesicles were associated with Golgi stacks. This study demonstrates that HPF/FS can be used to successfully preserve the ultrastructure of relatively large plant tissues without the use of intracellular cryoprotectants.

  16. Comparison of the ultrastructure of conventionally fixed and high pressure frozen/freeze substituted root tips of Nicotiana and Arabidopsis

    NASA Technical Reports Server (NTRS)

    Kiss, J. Z.; Giddings, T. H. Jr; Staehelin, L. A.; Sack, F. D.

    1990-01-01

    To circumvent the limitations of chemical fixation (CF) and to gain more reliable structural information about higher plant tissues, we have cryofixed root tips of Nicotiana and Arabidopsis by high pressure freezing (HPF). Whereas other freezing techniques preserve tissue to a relatively shallow depth, HPF in conjunction with freeze substitution (FS) resulted in excellent preservation of entire root tips. Compared to CF, in tissue prepared by HPF/FS: (1) the plasmalemma and all internal membranes were much smoother and often coated on the cytoplasmic side by a thin layer of stained material, (2) the plasmalemma was appressed to the cell wall, (3) organelle profiles were rounder, (4) the cytoplasmic, mitochondrial, and amyloplast matrices were denser, (5) vacuoles contained electron dense material, (6) microtubules appeared to be more numerous and straighter, with crossbridges observed between them, (7) cisternae of endoplasmic reticulum (ER) were wider and filled with material, (8) Golgi intercisternal elements were more clearly resolved and were observed between both Golgi vesicles and cisternae, and (9) larger vesicles were associated with Golgi stacks. This study demonstrates that HPF/FS can be used to successfully preserve the ultrastructure of relatively large plant tissues without the use of intracellular cryoprotectants.

  17. Glycogen synthase kinase-3: cryoprotection and glycogen metabolism in the freeze-tolerant wood frog.

    PubMed

    Dieni, Christopher A; Bouffard, Melanie C; Storey, Kenneth B

    2012-02-01

    The terrestrial anuran Rana sylvatica tolerates extended periods of whole-body freezing during the winter. Freezing survival is facilitated by extensive glycogen hydrolysis and distribution of high concentrations of the cryoprotectant glucose into blood and all tissues. As glycogenesis is both an energy-expensive process and counter-productive to maintaining sustained high cryoprotectant levels, we proposed that glycogen synthase kinase-3 (GSK-3) would be activated when wood frogs froze and would phosphorylate its downstream substrates to inactivate glycogen synthesis. Western blot analysis determined that the amount of phosphorylated (inactive) GSK-3 decreased in all five tissues tested in 24 h frozen frogs compared with unfrozen controls. Total GSK-3 protein levels did not change, with the exception of heart GSK-3, indicating that post-translational modification was the primary regulatory mechanism for this kinase. Kinetic properties of skeletal muscle GSK-3 from control and frozen frogs displayed differential responses to a temperature change (22 versus 4°C) and high glucose. For example, when assayed at 4°C, the K(m) for the GSK-3 substrate peptide was ∼44% lower for frozen frogs than the corresponding value in control frogs, indicating greater GSK-3 affinity for its substrates in the frozen state. This indicates that at temperatures similar to the environment encountered by frogs, GSK-3 in frozen frogs will phosphorylate its downstream targets more readily than in unfrozen controls. GSK-3 from skeletal muscle of control frogs was also allosterically regulated. AMP and phosphoenolpyruvate activated GSK-3 whereas inhibitors included glucose, glucose 6-phosphate, pyruvate, ATP, glutamate, glutamine, glycerol, NH(4)Cl, NaCl and KCl. The combination of phosphorylation and allosteric control argues for a regulatory role of GSK-3 in inactivating glycogenesis to preserve high glucose cryoprotectant levels throughout each freezing bout.

  18. Novel control of lactate dehydrogenase from the freeze tolerant wood frog: role of posttranslational modifications.

    PubMed

    Abboud, Jean; Storey, Kenneth B

    2013-01-01

    Lactate dehydrogenase (LDH), the terminal enzyme of anaerobic glycolysis, plays a crucial role both in sustaining glycolytic ATP production under oxygen-limiting conditions and in facilitating the catabolism of accumulated lactate when stress conditions are relieved. In this study, the effects on LDH of in vivo freezing and dehydration stresses (both of which impose hypoxia/anoxia stress on tissues) were examined in skeletal muscle of the freeze-tolerant wood frog, Rana sylvatica. LDH from muscle of control, frozen and dehydrated wood frogs was purified to homogeneity in a two-step process. The kinetic properties and stability of purified LDH were analyzed, revealing no significant differences in V max, K m and I 50 values between control and frozen LDH. However, control and dehydrated LDH differed significantly in K m values for pyruvate, lactate, and NAD, I 50 urea, and in temperature, glucose, and urea effects on these parameters. The possibility that posttranslational modification of LDH was responsible for the stable differences in enzyme behavior between control and dehydrated states was assessed using ProQ diamond staining to detect phosphorylation and immunoblotting to detect acetylation, methylation, ubiquitination, SUMOylation and nitrosylation of the enzyme. LDH from muscle of dehydrated wood frogs showed significantly lower levels of acetylation, providing one of the first demonstrations of a potential role for protein acetylation in the stress-responsive control of a metabolic enzyme.

  19. Novel control of lactate dehydrogenase from the freeze tolerant wood frog: role of posttranslational modifications

    PubMed Central

    Abboud, Jean

    2013-01-01

    Lactate dehydrogenase (LDH), the terminal enzyme of anaerobic glycolysis, plays a crucial role both in sustaining glycolytic ATP production under oxygen-limiting conditions and in facilitating the catabolism of accumulated lactate when stress conditions are relieved. In this study, the effects on LDH of in vivo freezing and dehydration stresses (both of which impose hypoxia/anoxia stress on tissues) were examined in skeletal muscle of the freeze-tolerant wood frog, Rana sylvatica. LDH from muscle of control, frozen and dehydrated wood frogs was purified to homogeneity in a two-step process. The kinetic properties and stability of purified LDH were analyzed, revealing no significant differences in Vmax, Km and I50 values between control and frozen LDH. However, control and dehydrated LDH differed significantly in Km values for pyruvate, lactate, and NAD, I50 urea, and in temperature, glucose, and urea effects on these parameters. The possibility that posttranslational modification of LDH was responsible for the stable differences in enzyme behavior between control and dehydrated states was assessed using ProQ diamond staining to detect phosphorylation and immunoblotting to detect acetylation, methylation, ubiquitination, SUMOylation and nitrosylation of the enzyme. LDH from muscle of dehydrated wood frogs showed significantly lower levels of acetylation, providing one of the first demonstrations of a potential role for protein acetylation in the stress-responsive control of a metabolic enzyme. PMID:23638346

  20. Physiological Mechanisms Only Tell Half Story: Multiple Biological Processes are involved in Regulating Freezing Tolerance of Imbibed Lactuca sativa Seeds.

    PubMed

    Jaganathan, Ganesh K; Han, Yingying; Li, Weijie; Song, Danping; Song, Xiaoyan; Shen, Mengqi; Zhou, Qiang; Zhang, Chenxue; Liu, Baolin

    2017-03-13

    The physiological mechanisms by which imbibed seeds survive freezing temperatures in their natural environment have been categorized as freezing avoidance by supercooling and freezing tolerance by extracellular freeze-desiccation, but the biochemical and molecular mechanisms conferring seed freezing tolerance is unexplored. In this study, using imbibed Lactuca sativa seeds we show that fast cooled seeds (60 °C h(-1)) suffered significantly higher membrane damage at temperature between -20 °C and -10 °C than slow cooled (3 °Ch(-1)) seeds (P < 0.05), presumably explaining viability loss during fast cooling when temperature approaches -20 °C. Total soluble sugars increase in low temperature environment, but did not differ significantly between two cooling rates (P > 0.05). However, both SOD activity and accumulation of free proline were induced significantly after slow cooling to -20 °C compared with fast cooling. RNA-seq demonstrated that multiple pathways were differentially regulated between slow and fast cooling. Real-time verification of some differentially expressed genes (DEGs) revealed that fast cooling caused mRNA level changes of plant hormone and ubiquitionation pathways at higher sub-zero temperature, whilst slow cooling caused mRNA level change of those pathways at lower sub-zero ttemperatures. Thus, we conclude that imbibed seed tolerate low temperature not only by physiological mechanisms but also by biochemical and molecular changes.

  1. Physiological Mechanisms Only Tell Half Story: Multiple Biological Processes are involved in Regulating Freezing Tolerance of Imbibed Lactuca sativa Seeds

    PubMed Central

    Jaganathan, Ganesh K.; Han, Yingying; Li, Weijie; Song, Danping; Song, Xiaoyan; Shen, Mengqi; Zhou, Qiang; Zhang, Chenxue; Liu, Baolin

    2017-01-01

    The physiological mechanisms by which imbibed seeds survive freezing temperatures in their natural environment have been categorized as freezing avoidance by supercooling and freezing tolerance by extracellular freeze-desiccation, but the biochemical and molecular mechanisms conferring seed freezing tolerance is unexplored. In this study, using imbibed Lactuca sativa seeds we show that fast cooled seeds (60 °C h−1) suffered significantly higher membrane damage at temperature between −20 °C and −10 °C than slow cooled (3 °Ch−1) seeds (P < 0.05), presumably explaining viability loss during fast cooling when temperature approaches −20 °C. Total soluble sugars increase in low temperature environment, but did not differ significantly between two cooling rates (P > 0.05). However, both SOD activity and accumulation of free proline were induced significantly after slow cooling to −20 °C compared with fast cooling. RNA-seq demonstrated that multiple pathways were differentially regulated between slow and fast cooling. Real-time verification of some differentially expressed genes (DEGs) revealed that fast cooling caused mRNA level changes of plant hormone and ubiquitionation pathways at higher sub-zero temperature, whilst slow cooling caused mRNA level change of those pathways at lower sub-zero ttemperatures. Thus, we conclude that imbibed seed tolerate low temperature not only by physiological mechanisms but also by biochemical and molecular changes. PMID:28287125

  2. A novel wheat bZIP transcription factor, TabZIP60, confers multiple abiotic stress tolerances in transgenic Arabidopsis.

    PubMed

    Zhang, Lina; Zhang, Lichao; Xia, Chuan; Zhao, Guangyao; Liu, Ji; Jia, Jizeng; Kong, Xiuying

    2015-04-01

    The basic region/leucine zipper (bZIP) transcription factors (TFs) play vital roles in the response to abiotic stress. However, little is known about the function of bZIP genes in wheat abiotic stress. In this study, we report the isolation and functional characterization of the TabZIP60 gene. Three homologous genome sequences of TabZIP60 were isolated from hexaploid wheat and mapped to the wheat homoeologous group 6. A subcellular localization analysis indicated that TabZIP60 is a nuclear-localized protein that activates transcription. Furthermore, TabZIP60 gene transcripts were strongly induced by polyethylene glycol, salt, cold and exogenous abscisic acid (ABA) treatments. Further analysis showed that the overexpression of TabZIP60 in Arabidopsis resulted in significantly improved tolerances to drought, salt, freezing stresses and increased plant sensitivity to ABA in seedling growth. Meanwhile, the TabZIP60 was capable of binding ABA-responsive cis-elements that are present in promoters of many known ABA-responsive genes. A subsequent analysis showed that the overexpression of TabZIP60 led to enhanced expression levels of some stress-responsive genes and changes in several physiological parameters. Taken together, these results suggest that TabZIP60 enhances multiple abiotic stresses through the ABA signaling pathway and that modifications of its expression may improve multiple stress tolerances in crop plants.

  3. Effect of exogenous extracellular polysaccharides on the desiccation and freezing tolerance of rock-inhabiting phototrophic microorganisms.

    PubMed

    Knowles, Emily J; Castenholz, Richard W

    2008-11-01

    Two major stresses that threaten rock-inhabiting microbial communities are desiccation and freezing; both result in a loss of liquid water in the cells. The mechanisms necessary to tolerate these extremes may be similar, but are not well understood. In both cases extracellular polysaccharides (EPS) seem to play an important role. This study examines whether the EPS released by a rock-inhabiting phototroph can have a protective effect on other members of similar and neighboring microbial communities. This interaction was modeled by adding EPS isolated from the cryptoendolithic cyanobacterium Nostoc sp. to cells of the cryptoendolithic green alga Chlorella sp. and to cells of the epilithic cyanobacterium Chroococcidiopsis sp. The cells were then subjected to desiccation and freezing and the survival rates were determined by vital staining, using membrane integrity as a measure of viability. The results clearly demonstrate the importance of exogenous EPS in the desiccation tolerance of both species, while mixed results were found for the freezing trials.

  4. Potential implications for expansion of freeze-tolerant eucalyptus plantations on water resources in the southern United States

    Treesearch

    James M. Vose; Chelcy F. Miniat; Ge Sun; Peter V. Caldwell

    2014-01-01

    The potential expansion of freeze-tolerant (FT) Eucalyptus plantations in the United States has raised concerns about the implications for water resources. Modeling was used to examine the potential effects of expanding the distribution of FT Eucalyptus plantations in US Department of Agriculture Plant Hardiness Zones 8b and...

  5. Genome Wide Association Mapping for the Tolerance to the Polyamine Oxidase Inhibitor Guazatine in Arabidopsis thaliana

    PubMed Central

    Atanasov, Kostadin E.; Barboza-Barquero, Luis; Tiburcio, Antonio F.; Alcázar, Rubén

    2016-01-01

    Guazatine is a potent inhibitor of polyamine oxidase (PAO) activity. In agriculture, guazatine is used as non-systemic contact fungicide efficient in the protection of cereals and citrus fruits against disease. The composition of guazatine is complex, mainly constituted by a mixture of synthetic guanidated polyamines (polyaminoguanidines). Here, we have studied the effects from exposure to guazatine in the weed Arabidopsis thaliana. We report that micromolar concentrations of guazatine are sufficient to inhibit growth of Arabidopsis seedlings and induce chlorosis, whereas germination is barely affected. We observed the occurrence of quantitative variation in the response to guazatine between 107 randomly chosen Arabidopsis accessions. This enabled us to undertake genome-wide association (GWA) mapping that identified a locus on chromosome one associated with guazatine tolerance. CHLOROPHYLLASE 1 (CLH1) within this locus was studied as candidate gene, together with its paralog (CLH2). The analysis of independent clh1-2, clh1-3, clh2-3, clh2-2, and double clh1-2 clh2-3 mutant alleles indicated that CLH1 and/or CLH2 loss-of-function or expression down-regulation promote guazatine tolerance in Arabidopsis. We report a natural mechanism by which Arabidopsis populations can overcome toxicity by the fungicide guazatine. PMID:27092150

  6. Genome Wide Association Mapping for the Tolerance to the Polyamine Oxidase Inhibitor Guazatine in Arabidopsis thaliana.

    PubMed

    Atanasov, Kostadin E; Barboza-Barquero, Luis; Tiburcio, Antonio F; Alcázar, Rubén

    2016-01-01

    Guazatine is a potent inhibitor of polyamine oxidase (PAO) activity. In agriculture, guazatine is used as non-systemic contact fungicide efficient in the protection of cereals and citrus fruits against disease. The composition of guazatine is complex, mainly constituted by a mixture of synthetic guanidated polyamines (polyaminoguanidines). Here, we have studied the effects from exposure to guazatine in the weed Arabidopsis thaliana. We report that micromolar concentrations of guazatine are sufficient to inhibit growth of Arabidopsis seedlings and induce chlorosis, whereas germination is barely affected. We observed the occurrence of quantitative variation in the response to guazatine between 107 randomly chosen Arabidopsis accessions. This enabled us to undertake genome-wide association (GWA) mapping that identified a locus on chromosome one associated with guazatine tolerance. CHLOROPHYLLASE 1 (CLH1) within this locus was studied as candidate gene, together with its paralog (CLH2). The analysis of independent clh1-2, clh1-3, clh2-3, clh2-2, and double clh1-2 clh2-3 mutant alleles indicated that CLH1 and/or CLH2 loss-of-function or expression down-regulation promote guazatine tolerance in Arabidopsis. We report a natural mechanism by which Arabidopsis populations can overcome toxicity by the fungicide guazatine.

  7. Freeze tolerance, supercooling points and ice formation: comparative studies on the subzero temperature survival of limno-terrestrial tardigrades.

    PubMed

    Hengherr, S; Worland, M R; Reuner, A; Brümmer, F; Schill, R O

    2009-03-01

    Many limno-terrestrial tardigrades live in unstable habitats where they experience extreme environmental conditions such as drought, heat and subzero temperatures. Although their stress tolerance is often related only to the anhydrobiotic state, tardigrades can also be exposed to great daily temperature fluctuations without dehydration. Survival of subzero temperatures in an active state requires either the ability to tolerate the freezing of body water or mechanisms to decrease the freezing point. Considering freeze tolerance in tardigrades as a general feature, we studied the survival rate of nine tardigrade species originating from polar, temperate and tropical regions by cooling them at rates of 9, 7, 5, 3 and 1 degrees C h(-1) down to -30 degrees C then returning them to room temperature at 10 degrees C h(-1). The resulting moderate survival after fast and slow cooling rates and low survival after intermediate cooling rates may indicate the influence of a physical effect during fast cooling and the possibility that they are able to synthesize cryoprotectants during slow cooling. Differential scanning calorimetry of starved, fed and cold acclimatized individuals showed no intraspecific significant differences in supercooling points and ice formation. Although this might suggest that metabolic and biochemical preparation are non-essential prior to subzero temperature exposure, the increased survival rate with slower cooling rates gives evidence that tardigrades still use some kind of mechanism to protect their cellular structure from freezing injury without influencing the freezing temperature. These results expand our current understanding of freeze tolerance in tardigrades and will lead to a better understanding of their ability to survive subzero temperature conditions.

  8. A comparative study of salt tolerance parameters in 11 wild relatives of Arabidopsis thaliana

    PubMed Central

    Orsini, Francesco; D'Urzo, Matilde Paino; Inan, Gunsu; Serra, Sara; Oh, Dong-Ha; Mickelbart, Michael V.; Consiglio, Federica; Jeong, Jae Cheol; Yun, Dae-Jin; Bohnert, Hans J.; Bressan, Ray A.; Maggio, Albino

    2010-01-01

    Salinity is an abiotic stress that limits both yield and the expansion of agricultural crops to new areas. In the last 20 years our basic understanding of the mechanisms underlying plant tolerance and adaptation to saline environments has greatly improved owing to active development of advanced tools in molecular, genomics, and bioinformatics analyses. However, the full potential of investigative power has not been fully exploited, because the use of halophytes as model systems in plant salt tolerance research is largely neglected. The recent introduction of halophytic Arabidopsis-Relative Model Species (ARMS) has begun to compare and relate several unique genetic resources to the well-developed Arabidopsis model. In a search for candidates to begin to understand, through genetic analyses, the biological bases of salt tolerance, 11 wild relatives of Arabidopsis thaliana were compared: Barbarea verna, Capsella bursa-pastoris, Hirschfeldia incana, Lepidium densiflorum, Malcolmia triloba, Lepidium virginicum, Descurainia pinnata, Sisymbrium officinale, Thellungiella parvula, Thellungiella salsuginea (previously T. halophila), and Thlaspi arvense. Among these species, highly salt-tolerant (L. densiflorum and L. virginicum) and moderately salt-tolerant (M. triloba and H. incana) species were identified. Only T. parvula revealed a true halophytic habitus, comparable to the better studied Thellungiella salsuginea. Major differences in growth, water transport properties, and ion accumulation are observed and discussed to describe the distinctive traits and physiological responses that can now be studied genetically in salt stress research. PMID:20595237

  9. ABA inducible rice protein phosphatase 2C confers ABA insensitivity and abiotic stress tolerance in Arabidopsis.

    PubMed

    Singh, Amarjeet; Jha, Saroj K; Bagri, Jayram; Pandey, Girdhar K

    2015-01-01

    Arabidopsis PP2C belonging to group A have been extensively worked out and known to negatively regulate ABA signaling. However, rice (Oryza sativa) orthologs of Arabidopsis group A PP2C are scarcely characterized functionally. We have identified a group A PP2C from rice (OsPP108), which is highly inducible under ABA, salt and drought stresses and localized predominantly in the nucleus. Genetic analysis revealed that Arabidopsis plants overexpressing OsPP108 are highly insensitive to ABA and tolerant to high salt and mannitol stresses during seed germination, root growth and overall seedling growth. At adult stage, OsPP108 overexpression leads to high tolerance to salt, mannitol and drought stresses with far better physiological parameters such as water loss, fresh weight, chlorophyll content and photosynthetic potential (Fv/Fm) in transgenic Arabidopsis plants. Expression profile of various stress marker genes in OsPP108 overexpressing plants revealed interplay of ABA dependent and independent pathway for abiotic stress tolerance. Overall, this study has identified a potential rice group A PP2C, which regulates ABA signaling negatively and abiotic stress signaling positively. Transgenic rice plants overexpressing this gene might provide an answer to the problem of low crop yield and productivity during adverse environmental conditions.

  10. ABA Inducible Rice Protein Phosphatase 2C Confers ABA Insensitivity and Abiotic Stress Tolerance in Arabidopsis

    PubMed Central

    Singh, Amarjeet; Jha, Saroj K.; Bagri, Jayram; Pandey, Girdhar K.

    2015-01-01

    Arabidopsis PP2C belonging to group A have been extensively worked out and known to negatively regulate ABA signaling. However, rice (Oryza sativa) orthologs of Arabidopsis group A PP2C are scarcely characterized functionally. We have identified a group A PP2C from rice (OsPP108), which is highly inducible under ABA, salt and drought stresses and localized predominantly in the nucleus. Genetic analysis revealed that Arabidopsis plants overexpressing OsPP108 are highly insensitive to ABA and tolerant to high salt and mannitol stresses during seed germination, root growth and overall seedling growth. At adult stage, OsPP108 overexpression leads to high tolerance to salt, mannitol and drought stresses with far better physiological parameters such as water loss, fresh weight, chlorophyll content and photosynthetic potential (Fv/Fm) in transgenic Arabidopsis plants. Expression profile of various stress marker genes in OsPP108 overexpressing plants revealed interplay of ABA dependent and independent pathway for abiotic stress tolerance. Overall, this study has identified a potential rice group A PP2C, which regulates ABA signaling negatively and abiotic stress signaling positively. Transgenic rice plants overexpressing this gene might provide an answer to the problem of low crop yield and productivity during adverse environmental conditions. PMID:25886365

  11. Polyploids exhibit higher potassium uptake and salinity tolerance in Arabidopsis.

    PubMed

    Chao, Dai-Yin; Dilkes, Brian; Luo, Hongbing; Douglas, Alex; Yakubova, Elena; Lahner, Brett; Salt, David E

    2013-08-09

    Genome duplication (or polyploidization) has occurred throughout plant evolutionary history and is thought to have driven the adaptive radiation of plants. We found that the cytotype of the root, and not the genotype, determined the majority of heritable natural variation in leaf potassium (K) concentration in Arabidopsis thaliana. Autopolyploidy also provided resistance to salinity and may represent an adaptive outcome of the enhanced K accumulation of plants with higher ploidy.

  12. Electrophoretic analysis of liver glycogen phosphorylase activation in the freeze-tolerant wood frog.

    PubMed

    Crerar, M M; David, E S; Storey, K B

    1988-08-19

    As an adaptation for overwinter survival, the wood frog, Rana sylvatica is able to tolerate the freezing of extracellular body fluids. Tolerance is made possible by the production of very high amounts of glucose in liver which is then sent to other organs where it acts as a cryoprotectant. Cryoprotectant synthesis is under the control of glycogen phosphorylase which in turn is activated in response to ice formation. To determine the mechanism of phosphorylase activation, a quantitative analysis of phosphorylase protein concentration and enzymatic activity in liver was carried out following separation of the phosphorylated a and nonphosphorylated b forms of the enzyme on native polyacrylamide gels. The results suggest that in gels, the b form is completely inactive, even in the presence of AMP and sodium sulfate, whereas the a form is active and stimulated 3-fold by these substances. Further, phosphorylase activation appears to arise solely from conversion of the b to a form of the enzyme without an increase in phosphorylase concentration or activation of a second isozyme. The quantitative analysis presented here should prove generally useful as a simple and rapid method for examining the physiological and genetic regulation of phosphorylase in animal cells.

  13. MAL62 overexpression and NTH1 deletion enhance the freezing tolerance and fermentation capacity of the baker's yeast in lean dough.

    PubMed

    Sun, Xi; Zhang, Cui-Ying; Wu, Ming-Yue; Fan, Zhi-Hua; Liu, Shan-Na; Zhu, Wen-Bi; Xiao, Dong-Guang

    2016-04-04

    Trehalose is related to several types of stress responses, especially freezing response in baker's yeast (Saccharomyces cerevisiae). It is desirable to manipulate trehalose-related genes to create yeast strains that better tolerate freezing-thaw stress with improved fermentation capacity, which are in high demand in the baking industry. The strain overexpressing MAL62 gene showed increased trehalose content and cell viability after prefermention-freezing and long-term frozen. Deletion of NTH1 in combination of MAL62 overexpression further strengthens freezing tolerance and improves the leavening ability after freezing-thaw stress. The mutants of the industrial baker's yeast with enhanced freezing tolerance and leavening ability in lean dough were developed by genetic engineering. These strains had excellent potential industrial applications.

  14. Nitric Oxide and Hydrogen Peroxide Mediate Wounding-Induced Freezing Tolerance through Modifications in Photosystem and Antioxidant System in Wheat.

    PubMed

    Si, Tong; Wang, Xiao; Wu, Lin; Zhao, Chunzhao; Zhang, Lini; Huang, Mei; Cai, Jian; Zhou, Qin; Dai, Tingbo; Zhu, Jian-Kang; Jiang, Dong

    2017-01-01

    Mechanical wounding is a common stress caused by herbivores or manual and natural manipulations, whereas its roles in acclimation response to a wide spectrum of abiotic stresses remain unclear. The present work showed that local mechanical wounding enhanced freezing tolerance in untreated systemic leaves of wheat plants (Triticum aestivum L.), and meanwhile the signal molecules hydrogen peroxide (H2O2) and nitric oxide (NO) were accumulated systemically. Pharmacological study showed that wounding-induced NO synthesis was substantially arrested by pretreatment with scavengers of reactive oxygen species and an inhibitor of NADPH oxidase (respiratory burst oxidase homolog, RBOH). On the contrary, wounding-induced H2O2 accumulation was not sensitive to NO synthetic inhibitors or scavenger, indicating that H2O2 acts upstream of NO in wounding signal transduction pathways. Cytochemical and vascular tissues localizations approved that RBOH-dependent H2O2 acts as long-distance signal in wounding response. Transcriptome analysis revealed that 279 genes were up-regulated in plants treated with wounding and freezing, but not in plants treated with freezing alone. Importantly, freezing- and wounding-induced genes were significantly enriched in the categories of "photosynthesis" and "signaling." These results strongly supported that primary mechanical wounding can induce freezing tolerance in wheat through the systemic accumulation of NO and H2O2, and further modifications in photosystem and antioxidant system.

  15. Cadmium tolerance and phytochelatin content of Arabidopsis seedlings over-expressing the phytochelatin synthase gene AtPCS1

    PubMed Central

    Brunetti, Patrizia; Zanella, Letizia; Proia, Alessandra; De Paolis, Angelo; Falasca, Giuseppina; Altamura, Maria Maddalena; Sanità di Toppi, Luigi; Costantino, Paolo; Cardarelli, Maura

    2011-01-01

    Previous studies demonstrated that expression of the Arabidopsis phytochelatin (PC) biosynthetic gene AtPCS1 in Nicotiana tabacum plants increases the Cd tolerance in the presence of exogenous glutathione (GSH). In this paper, the Cd tolerance of Arabidopsis plants over-expressing AtPCS1 (AtPCSox lines) has been analysed and the differences between Arabidopsis and tobacco are shown. Based on the analysis of seedling fresh weight, primary root length, and alterations in root anatomy, evidence is provided that, at relatively low Cd concentrations, the Cd tolerance of AtPCSox lines is lower than the wild type, while AtPCS1 over-expressing tobacco is more tolerant to Cd than the wild type. At higher Cd concentrations, Arabidopsis AtPCSox seedlings are more tolerant to Cd than the wild type, while tobacco AtPCS1 seedlings are as sensitive as the wild type. Exogenous GSH, in contrast to what was observed in tobacco, did not increase the Cd tolerance of AtPCSox lines. The PC content in wild-type Arabidopsis at low Cd concentrations is more than three times higher than in tobacco and substantial differences were also found in the PC chain lengths. These data indicate that the differences in Cd tolerance and in its dependence on exogenous GSH between Arabidopsis and tobacco are due to species-specific differences in the endogenous content of PCs and GSH and may be in the relative abundance of PCs of different length. PMID:21841172

  16. Cadmium tolerance and phytochelatin content of Arabidopsis seedlings over-expressing the phytochelatin synthase gene AtPCS1.

    PubMed

    Brunetti, Patrizia; Zanella, Letizia; Proia, Alessandra; De Paolis, Angelo; Falasca, Giuseppina; Altamura, Maria Maddalena; Sanità di Toppi, Luigi; Costantino, Paolo; Cardarelli, Maura

    2011-11-01

    Previous studies demonstrated that expression of the Arabidopsis phytochelatin (PC) biosynthetic gene AtPCS1 in Nicotiana tabacum plants increases the Cd tolerance in the presence of exogenous glutathione (GSH). In this paper, the Cd tolerance of Arabidopsis plants over-expressing AtPCS1 (AtPCSox lines) has been analysed and the differences between Arabidopsis and tobacco are shown. Based on the analysis of seedling fresh weight, primary root length, and alterations in root anatomy, evidence is provided that, at relatively low Cd concentrations, the Cd tolerance of AtPCSox lines is lower than the wild type, while AtPCS1 over-expressing tobacco is more tolerant to Cd than the wild type. At higher Cd concentrations, Arabidopsis AtPCSox seedlings are more tolerant to Cd than the wild type, while tobacco AtPCS1 seedlings are as sensitive as the wild type. Exogenous GSH, in contrast to what was observed in tobacco, did not increase the Cd tolerance of AtPCSox lines. The PC content in wild-type Arabidopsis at low Cd concentrations is more than three times higher than in tobacco and substantial differences were also found in the PC chain lengths. These data indicate that the differences in Cd tolerance and in its dependence on exogenous GSH between Arabidopsis and tobacco are due to species-specific differences in the endogenous content of PCs and GSH and may be in the relative abundance of PCs of different length.

  17. Ectopic expression of Arabidopsis glutaredoxin gene AtGRXS17 in tomato (Solanum lycopersicum) confers tolerance to chilling stress

    USDA-ARS?s Scientific Manuscript database

    The monothiol glutaredoxin AtGRXS17 from "Arabidopsis" confers thermotolerance in yeast, "Arabidopsis", and tomato plants. Here, we report that AtGRXS17 also enhances tolerance to chilling stress in tomato and is associated with elevation of antioxidant enzyme activities, which are known to be invol...

  18. Dual roles of glucose in the freeze-tolerant earthworm Dendrobaena octaedra: cryoprotection and fuel for metabolism.

    PubMed

    Calderon, Sofia; Holmstrup, Martin; Westh, Peter; Overgaard, Johannes

    2009-03-01

    Ectothermic animals inhabiting the subarctic and temperate regions have evolved strategies to deal with periods of continuous frost during winter. The earthworm Dendrobaena octaedra is freeze tolerant and accumulates large concentrations of glucose upon freezing. The present study investigates the roles of glucose accumulation for long-term freeze tolerance in worms kept frozen at -2 degrees C for 47 days. During this period, worms were sampled periodically for determination of survival and for measurements of glucose, glycogen, lactate, alanine and succinate. In addition we performed calorimetric measurements to assess metabolic rate of frozen and unfrozen worms. Long-term freezing was associated with a gradual depletion of glucose and worms that succumbed during this period were always characterised by low glucose and glycogen levels. The anaerobic waste products lactate and alanine increased slightly whereas succinate levels remained constant. However, it is argued that other waste products (particularly propionate) could be the primary end product of a continued anaerobic metabolism. Calorimetric measures of the metabolic rate of frozen worms were in accord with values calculated from the reduction in glucose assuming that most ( approximately 90%) glucose was metabolised anaerobically. Both estimates of metabolic rate demonstrated a 10-fold metabolic depression associated with freezing. Thus, in addition to the suspected role of glucose as cryoprotectant, the present study demonstrates that glucose accumulation is vital to ensure substrate for long-term anaerobic metabolism in frozen worms. On the basis of the estimated metabolite levels, we calculate that the combined effect of metabolic depression and large glucose stores enables a projected 3 months survival of freezing at -2 degrees C of the ;average' D. octaedra. Such conditions are very likely to occur in the northern distribution ranges of this stress-tolerant earthworm.

  19. A nonprotein thermal hysteresis-producing xylomannan antifreeze in the freeze-tolerant Alaskan beetle Upis ceramboides

    PubMed Central

    Walters, Kent R.; Serianni, Anthony S.; Sformo, Todd; Barnes, Brian M.; Duman, John G.

    2009-01-01

    Thermal hysteresis (TH), a difference between the melting and freezing points of a solution that is indicative of the presence of large-molecular-mass antifreezes (e.g., antifreeze proteins), has been described in animals, plants, bacteria, and fungi. Although all previously described TH-producing biomolecules are proteins, most thermal hysteresis factors (THFs) have not yet been structurally characterized, and none have been characterized from a freeze-tolerant animal. We isolated a highly active THF from the freeze-tolerant beetle, Upis ceramboides, by means of ice affinity. Amino acid chromatographic analysis, polyacrylamide gel electrophoresis, UV-Vis spectrophotometry, and NMR spectroscopy indicated that the THF contained little or no protein, yet it produced 3.7 ± 0.3 °C of TH at 5 mg/ml, comparable to that of the most active insect antifreeze proteins. Compositional and structural analyses indicated that this antifreeze contains a β-mannopyranosyl-(1→4) β-xylopyranose backbone and a fatty acid component, although the lipid may not be covalently linked to the saccharide. Consistent with the proposed structure, treatment with endo-β-(1→4)xylanase ablated TH activity. This xylomannan is the first TH-producing antifreeze isolated from a freeze-tolerant animal and the first in a new class of highly active THFs that contain little or no protein. PMID:19934038

  20. Stability evaluation of freeze-dried Lactobacillus paracasei subsp. tolerance and Lactobacillus delbrueckii subsp. bulgaricus in oral capsules.

    PubMed

    Jalali, M; Abedi, D; Varshosaz, J; Najjarzadeh, M; Mirlohi, M; Tavakoli, N

    2012-01-01

    Freeze-drying is a common preservation technology in the pharmaceutical industry. Various studies have investigated the effect of different cryoprotectants on probiotics during freeze-drying. However, information on the effect of cryoprotectants on the stability of some Lactobacillus strains during freeze-drying seems scarce. Therefore, the aim of the present study was to establish production methods for preparation of oral capsule probiotics containing Lactobacillus paracasei subsp. tolerance and Lactobacillus delbrueckii subsp. Bulgaricus. It was also of interest to examine the effect of various formulations of cryoprotectant media containing skim milk, trehalose and sodium ascorbate on the survival rate of probiotic bacteria during freeze-drying at various storage temperatures. Without any cryoprotectant, few numbers of microorganisms survived. However, microorganisms tested maintained higher viability after freeze-drying in media containing at least one of the cryoprotectants. Use of skim milk in water resulted in an increased viability after lyophilization. Media with a combination of trehalose and skim milk maintained a higher percentage of live microorganisms, up to 82%. In general, bacteria retained a higher number of viable cells in capsules containing freeze-dried bacteria with sodium ascorbate after three months of storage. After this period, a marked decline was observed in all samples stored at 23°C compared to those stored at 4°C. The maximum survival rate (about 72-76%) was observed with media containing 6% skim milk, 8% trehalose and 4% sodium ascorbate.

  1. Stability evaluation of freeze-dried Lactobacillus paracasei subsp. tolerance and Lactobacillus delbrueckii subsp. bulgaricus in oral capsules

    PubMed Central

    Jalali, M.; Abedi, D.; Varshosaz, J.; Najjarzadeh, M.; Mirlohi, M.; Tavakoli, N.

    2012-01-01

    Freeze-drying is a common preservation technology in the pharmaceutical industry. Various studies have investigated the effect of different cryoprotectants on probiotics during freeze-drying. However, information on the effect of cryoprotectants on the stability of some Lactobacillus strains during freeze-drying seems scarce. Therefore, the aim of the present study was to establish production methods for preparation of oral capsule probiotics containing Lactobacillus paracasei subsp. tolerance and Lactobacillus delbrueckii subsp. Bulgaricus. It was also of interest to examine the effect of various formulations of cryoprotectant media containing skim milk, trehalose and sodium ascorbate on the survival rate of probiotic bacteria during freeze-drying at various storage temperatures. Without any cryoprotectant, few numbers of microorganisms survived. However, microorganisms tested maintained higher viability after freeze-drying in media containing at least one of the cryoprotectants. Use of skim milk in water resulted in an increased viability after lyophilization. Media with a combination of trehalose and skim milk maintained a higher percentage of live microorganisms, up to 82%. In general, bacteria retained a higher number of viable cells in capsules containing freeze-dried bacteria with sodium ascorbate after three months of storage. After this period, a marked decline was observed in all samples stored at 23°C compared to those stored at 4°C. The maximum survival rate (about 72-76%) was observed with media containing 6% skim milk, 8% trehalose and 4% sodium ascorbate. PMID:23181077

  2. Enzymatic regulation of seasonal glycogen cycling in the freeze-tolerant wood frog, Rana sylvatica.

    PubMed

    do Amaral, M Clara F; Lee, Richard E; Costanzo, Jon P

    2016-12-01

    Liver glycogen is an important energy store in vertebrates, and in the freeze-tolerant wood frog, Rana sylvatica, this carbohydrate also serves as a major source of the cryoprotectant glucose. We investigated how variation in the levels of the catalytic subunit of protein kinase A (PKAc), glycogen phosphorylase (GP), and glycogen synthase (GS) relates to seasonal glycogen cycling in a temperate (Ohioan) and subarctic (Alaskan) populations of this species. In spring, Ohioan frogs had reduced potential for glycogen synthesis, as evidenced by low GS activity and high PKAc protein levels. In addition, glycogen levels in spring were the lowest of four seasonal samples, as energy input was likely directed towards metabolism and somatic growth during this period. Near-maximal glycogen levels were reached by mid-summer, and remained unchanged in fall and winter, suggesting that glycogenesis was curtailed during this period. Ohioan frogs had a high potential for glycogenolysis and glycogenesis in winter, as evidenced by large glycogen reserves, high levels of GP and GS proteins, and high GS activity, which likely allows for rapid mobilization of cryoprotectant during freezing and replenishing of glycogen reserves during thawing. Alaskan frogs also achieved a near-maximal liver glycogen concentration by summer and displayed high glycogenic and glycogenolytic potential in winter, but, unlike Ohioan frogs, started replenishing their energy reserves early in spring. We conclude that variation in levels of both glycogenolytic and glycogenic enzymes likely happens in response to seasonal changes in energetic strategies and demands, with winter survival being a key component to understanding the regulation of glycogen cycling in this species.

  3. Regulation of 5'-adenosine monophosphate deaminase in the freeze tolerant wood frog, Rana sylvatica

    PubMed Central

    Dieni, Christopher A; Storey, Kenneth B

    2008-01-01

    Background The wood frog, Rana sylvatica, is one of a few vertebrate species that have developed natural freeze tolerance, surviving days or weeks with 65–70% of its total body water frozen in extracellular ice masses. Frozen frogs exhibit no vital signs and their organs must endure multiple stresses, particularly long term anoxia and ischemia. Maintenance of cellular energy supply is critical to viability in the frozen state and in skeletal muscle, AMP deaminase (AMPD) plays a key role in stabilizing cellular energetics. The present study investigated AMPD control in wood frog muscle. Results Wood frog AMPD was subject to multiple regulatory controls: binding to subcellular structures, protein phosphorylation, and effects of allosteric effectors, cryoprotectants and temperature. The percentage of bound AMPD activity increased from 20 to 35% with the transition to the frozen state. Bound AMPD showed altered kinetic parameters compared with the free enzyme (S0.5 AMP was reduced, Hill coefficient fell to ~1.0) and the transition to the frozen state led to a 3-fold increase in S0.5 AMP of the bound enzyme. AMPD was a target of protein phosphorylation. Bound AMPD from control frogs proved to be a low phosphate form with a low S0.5 AMP and was phosphorylated in incubations that stimulated PKA, PKC, CaMK, or AMPK. Bound AMPD from frozen frogs was a high phosphate form with a high S0.5 AMP that was reduced under incubation conditions that stimulated protein phosphatases. Frog muscle AMPD was activated by Mg·ATP and Mg·ADP and inhibited by Mg·GTP, KCl, NaCl and NH4Cl. The enzyme product, IMP, uniquely inhibited only the bound (phosphorylated) enzyme from muscle of frozen frogs. Activators and inhibitors differentially affected the free versus bound enzyme. S0.5 AMP of bound AMPD was also differentially affected by high versus low assay temperature (25 vs 5°C) and by the presence/absence of the natural cryoprotectant (250 mM glucose) that accumulates during freezing

  4. Natural variation among Arabidopsis accessions reveals malic acid as a key mediator of Nickel (Ni) tolerance.

    PubMed

    Agrawal, Bhavana; Lakshmanan, Venkatachalam; Kaushik, Shail; Bais, Harsh P

    2012-08-01

    Plants have evolved various mechanisms for detoxification that are specific to the plant species as well as the metal ion chemical properties. Malic acid, which is commonly found in plants, participates in a number of physiological processes including metal chelation. Using natural variation among Arabidopsis accessions, we investigated the function of malic acid in Nickel (Ni) tolerance and detoxification. The Ni-induced production of reactive oxygen species was found to be modulated by intracellular malic acid, indicating its crucial role in Ni detoxification. Ni tolerance in Arabidopsis may actively involve malic acid and/or complexes of Ni and malic acid. Investigation of malic acid content in roots among tolerant ecotypes suggested that a complex of Ni and malic acid may be involved in translocation of Ni from roots to leaves. The exudation of malic acid from roots in response to Ni treatment in either susceptible or tolerant plant species was found to be partially dependent on AtALMT1 expression. A lower concentration of Ni (10 µM) treatment induced AtALMT1 expression in the Ni-tolerant Arabidopsis ecotypes. We found that the ecotype Santa Clara (S.C.) not only tolerated Ni but also accumulated more Ni in leaves compared to other ecotypes. Thus, the ecotype S.C. can be used as a model system to delineate the biochemical and genetic basis of Ni tolerance, accumulation, and detoxification in plants. The evolution of Ni hyperaccumulators, which are found in serpentine soils, is an interesting corollary to the fact that S.C. is also native to serpentine soils.

  5. Transcript Differences Associated With Non-Acclimated Freezing Tolerance in Two Barley (Hordeum Vulgare L.) Cultivars

    USDA-ARS?s Scientific Manuscript database

    Barley periodically suffers from late spring freezes in area throughout the world, with significant losses to yield. To better understand the response of barley to spring freezes, we examined the response of Dicktoo and Keunal barley varieties in their jointing stage to non-acclimated freezing (NAF...

  6. Overexpression of Late Embryogenesis Abundant 14 enhances Arabidopsis salt stress tolerance

    SciTech Connect

    Jia, Fengjuan Qi, Shengdong Li, Hui Liu, Pu Li, Pengcheng Wu, Changai Zheng, Chengchao Huang, Jinguang

    2014-11-28

    Highlights: • It is the first time to investigate the biological function of AtLEA14 in salt stress response. • AtLEA14 enhances the salt stress tolerance both in Arabidopsis and yeast. • AtLEA14 responses to salt stress by stabilizing AtPP2-B11, an E3 ligase, under normal or salt stress conditions. - Abstract: Late embryogenesis abundant (LEA) proteins are implicated in various abiotic stresses in higher plants. In this study, we identified a LEA protein from Arabidopsis thaliana, AtLEA14, which was ubiquitously expressed in different tissues and remarkably induced with increased duration of salt treatment. Subcellular distribution analysis demonstrated that AtLEA14 was mainly localized in the cytoplasm. Transgenic Arabidopsis and yeast overexpressing AtLEA14 all exhibited enhanced tolerance to high salinity. The transcripts of salt stress-responsive marker genes (COR15a, KIN1, RD29B and ERD10) were overactivated in AtLEA14 overexpressing lines compared with those in wild type plants under normal or salt stress conditions. In vivo and in vitro analysis showed that AtLEA14 could effectively stabilize AtPP2-B11, an important E3 ligase. These results suggested that AtLEA14 had important protective functions under salt stress conditions in Arabidopsis.

  7. Wheat TaSP gene improves salt tolerance in transgenic Arabidopsis thaliana.

    PubMed

    Ma, Xiaoli; Cui, Weina; Liang, Wenji; Huang, Zhanjing

    2015-12-01

    A novel salt-induced gene with unknown functions was cloned through analysis of gene expression profile of a salt-tolerant wheat mutant RH8706-49 under salt stress. The gene was named Triticum aestivum salt-related protein (TaSP) and deposited in GenBank (Accession No. KF307326). Quantitative polymerase chain reaction (qPCR) results showed that TaSP expression was induced under salt, abscisic acid (ABA), and polyethylene glycol (PEG) stresses. Subcellular localization revealed that TaSP was mainly localized in cell membrane. Overexpression of TaSP in Arabidopsis could improve salt tolerance of 35S::TaSP transgenic Arabidopsis. 35S::TaSP transgenic Arabidopsis lines after salt stress presented better physiological indexes than the control group. In the non-invasive micro-test (NMT), an evident Na(+) excretion was observed at the root tip of salt-stressed 35S::TaSP transgenic Arabidopsis. TaSP promoter was cloned, and its beta-glucuronidase (GUS) activities before and after ABA, salt, cold, heat, and salicylic acid (SA) stresses were determined. Full-length TaSP promoter contained ABA and salt response elements.

  8. Chinese Wild-Growing Vitis amurensis ICE1 and ICE2 Encode MYC-Type bHLH Transcription Activators that Regulate Cold Tolerance in Arabidopsis

    PubMed Central

    Xu, Weirong; Jiao, Yuntong; Li, Ruimin; Zhang, Ningbo; Xiao, Dongming; Ding, Xiaoling; Wang, Zhenping

    2014-01-01

    Winter hardiness is an important trait for grapevine breeders and producers, so identification of the regulatory mechanisms involved in cold acclimation is of great potential value. The work presented here involves the identification of two grapevine ICE gene homologs, VaICE1 and VaICE2, from an extremely cold-tolerant accession of Chinese wild-growing Vitis amurnensis, which are phylogenetically related to other plant ICE1 genes. These two structurally different ICE proteins contain previously reported ICE-specific amino acid motifs, the bHLH-ZIP domain and the S-rich motif. Expression analysis revealed that VaICE1 is constitutively expressed but affected by cold stress, unlike VaICE2 that shows not such changed expression as a consequence of cold treatment. Both genes serve as transcription factors, potentiating the transactivation activities in yeasts and the corresponding proteins localized to the nucleus following transient expression in onion epidermal cells. Overexpression of either VaICE1 or VaICE2 in Arabidopsis increase freezing tolerance in nonacclimated plants. Moreover, we show that they result in multiple biochemical changes that were associated with cold acclimation: VaICE1/2-overexpressing plants had evaluated levels of proline, reduced contents of malondialdehyde (MDA) and decreased levels of electrolyte leakage. The expression of downstream cold responsive genes of CBF1, COR15A, and COR47 were significantly induced in Arabidopsis transgenically overexpressing VaICE1 or VaICE2 upon cold stress. VaICE2, but not VaICE1 overexpression induced KIN1 expression under cold-acclimation conditions. Our results suggest that VaICE1 and VaICE2 act as key regulators at an early step in the transcriptional cascade controlling freezing tolerance, and modulate the expression levels of various low-temperature associated genes involved in the C-repeat binding factor (CBF) pathway. PMID:25019620

  9. Heterologous expression of mitochondria-targeted microbial nitrilase enzymes increases cyanide tolerance in Arabidopsis.

    PubMed

    Molojwane, E; Adams, N; Sweetlove, L J; Ingle, R A

    2015-07-01

    Anthropogenic activities have resulted in cyanide (CN) contamination of both soil and water in many areas of the globe. While plants possess a detoxification pathway that serves to degrade endogenously generated CN, this system is readily overwhelmed, limiting the use of plants in bioremediation. Genetic engineering of additional CN degradation pathways in plants is one potential strategy to increase their tolerance to CN. Here we show that heterologous expression of microbial nitrilase enzymes targeted to the mitochondria increases CN tolerance in Arabidopsis. Root length in seedlings expressing either a CN dihydratase from Bacillus pumilis or a CN hydratase from Neurospora crassa was increased by 45% relative in wild-type plants in the presence of 50 μm KCN. We also demonstrate that in contrast to its strong inhibitory effects on seedling establishment, seed germination of the Col-0 ecotype of Arabidopsis is unaffected by CN. © 2015 German Botanical Society and The Royal Botanical Society of the Netherlands.

  10. Functional characterization of aroA from Rhizobium leguminosarum with significant glyphosate tolerance in transgenic Arabidopsis.

    PubMed

    Han, Jing; Tian, Yong-Sheng; Xu, Jing; Wang, Li-Juan; Wang, Bo; Peng, Ri-He; Yao, Quan-Hong

    2014-09-01

    Glyphosate is the active component of the top-selling herbicide, the phytotoxicity of which is due to its inhibition of the shikimic acid pathway. 5-Enolpyruvylshikimate-3-phosphate synthase (EPSPS) is a key enzyme in the shikimic acid pathway. Glyphosate tolerance in plants can be achieved by the expression of a glyphosate-insensitive aroA gene (EPSPS). In this study, we used a PCR-based two-step DNA synthesis method to synthesize a new aroA gene (aroAR. leguminosarum) from Rhizobium leguminosarum. In vitro glyphosate sensitivity assays showed that aroAR. leguminosarum is glyphosate tolerant. The new gene was then expressed in E. coli and key kinetic values of the purified enzyme were determined. Furthermore, we transformed the aroA gene into Arabidopsis thaliana by the floral dip method. Transgenic Arabidopsis with the aroAR. leguminosarum gene was obtained to prove its potential use in developing glyphosate-resistant crops.

  11. Freeze-tolerant condenser for a closed-loop heat-transfer system

    NASA Technical Reports Server (NTRS)

    Crowley, Christopher J. (Inventor); Elkouh, Nabil A. (Inventor)

    2002-01-01

    A freeze tolerant condenser (106) for a two-phase heat transfer system is disclosed. The condenser includes an enclosure (110) and a porous artery (112) located within and extending along the length of the enclosure. A vapor space (116) is defined between the enclosure and the artery, and a liquid space (114) is defined by a central passageway within the artery. The artery includes a plurality of laser-micromachined capillaries (130) extending from the outer surface of the artery to its inner surface such that the vapor space is in fluid communication with the liquid space. In one embodiment of the invention, the capillaries (130) are cylindrical holes having a diameter of no greater than 50 microns. In another embodiment, the capillaries (130') are slots having widths of no greater than 50 microns. A method of making an artery in accordance with the present invention is also disclosed. The method includes providing a solid-walled tube and laser-micromachining a plurality of capillaries into the tube along a longitudinal axis, wherein each capillary has at least one cross-sectional dimension transverse to the longitudinal axis of less than 50 microns.

  12. Metabolomic analysis of extreme freezing tolerance in Siberian spruce (Picea obovata).

    PubMed

    Angelcheva, Liudmila; Mishra, Yogesh; Antti, Henrik; Kjellsen, Trygve D; Funk, Christiane; Strimbeck, Richard G; Schröder, Wolfgang P

    2014-11-01

    Siberian spruce (Picea obovata) is one of several boreal conifer species that can survive at extremely low temperatures (ELTs). When fully acclimated, its tissues can survive immersion in liquid nitrogen. Relatively little is known about the biochemical and biophysical strategies of ELT survival. We profiled needle metabolites using gas chromatography coupled with mass spectrometry (GC-MS) to explore the metabolic changes that occur during cold acclimation caused by natural temperature fluctuations. In total, 223 metabolites accumulated and 52 were depleted in fully acclimated needles compared with pre-acclimation needles. The metabolite profiles were found to develop in four distinct phases, which are referred to as pre-acclimation, early acclimation, late acclimation and fully acclimated. Metabolite changes associated with carbohydrate and lipid metabolism were observed, including changes associated with increased raffinose family oligosaccharide synthesis and accumulation, accumulation of sugar acids and sugar alcohols, desaturation of fatty acids, and accumulation of digalactosylglycerol. We also observed the accumulation of protein and nonprotein amino acids and polyamines that may act as compatible solutes or cryoprotectants. These results provide new insight into the mechanisms of freezing tolerance development at the metabolite level and highlight their importance in rapid acclimation to ELT in P. obovata.

  13. Brassica oleracea MATE encodes a citrate transporter and enhances aluminum tolerance in Arabidopsis thaliana.

    PubMed

    Wu, Xinxin; Li, Ren; Shi, Jin; Wang, Jinfang; Sun, Qianqian; Zhang, Haijun; Xing, Yanxia; Qi, Yan; Zhang, Na; Guo, Yang-Dong

    2014-08-01

    The secretion of organic acid anions from roots is an important mechanism for plant aluminum (Al) tolerance. Here we report cloning and characterizing BoMATE (KF031944), a multidrug and toxic compound extrusion (MATE) family gene from cabbage (Brassica oleracea). The expression of BoMATE was more abundant in roots than in shoots, and it was highly induced by Al treatment. The (14)C-citrate efflux experiments in oocytes demonstrated that BoMATE is a citrate transporter. Electrophysiological analysis and SIET analysis of Xenopus oocytes expressing BoMATE indicated BoMATE is activated by Al. Transient expression of BoMATE in onion epidermal cells demonstrated that it localized to the plasma membrane. Compared with the wild-type Arabidopsis, the transgenic lines constitutively overexpressing BoMATE enhanced Al tolerance and increased citrate secretion. In addition, Arabidopsis transgenic lines had a lower K(+) efflux and higher H(+) efflux, in the presence of Al, than control wild type in the distal elongation zone (DEZ). This is the first direct evidence that MATE protein is involved in the K(+) and H(+) flux in response to Al treatment. Taken together, our results show that BoMATE is an Al-induced citrate transporter and enhances aluminum tolerance in Arabidopsis thaliana.

  14. Potassium Retention under Salt Stress Is Associated with Natural Variation in Salinity Tolerance among Arabidopsis Accessions

    PubMed Central

    Sun, Yanling; Kong, Xiangpei; Li, Cuiling; Liu, Yongxiu; Ding, Zhaojun

    2015-01-01

    Plants are exposed to various environmental stresses during their life cycle such as salt, drought and cold. Natural variation mediated plant growth adaptation has been employed as an effective approach in response to the diverse environmental cues such as salt stress. However, the molecular mechanism underlying this process is not well understood. In the present study, a collection of 82 Arabidopsis thaliana accessions (ecotypes) was screened with a view to identify variation for salinity tolerance. Seven accessions showed a higher level of tolerance than Col-0. The young seedlings of the tolerant accessions demonstrated a higher K+ content and a lower Na+/K+ ratio when exposed to salinity stress, but its Na+ content was the same as that of Col-0. The K+ transporter genes AtHAK5, AtCHX17 and AtKUP1 were up-regulated significantly in almost all the tolerant accessions, even in the absence of salinity stress. There was little genetic variation or positive transcriptional variation between the selections and Col-0 with respect to Na+-related transporter genes, as AtSOS genes, AtNHX1 and AtHKT1;1. In addition, under salinity stress, these selections accumulated higher compatible solutes and lower reactive oxygen species than did Col-0. Taken together, our results showed that natural variation in salinity tolerance of Arabidopsis seems to have been achieved by the strong capacity of K+ retention. PMID:25993093

  15. Azetidine-2-carboxylic acid resistant mutants of Arabidopsis thaliana with increased salt tolerance

    SciTech Connect

    Lehle, F.R.; Murphy, M.A.; Khan, R.A. )

    1989-04-01

    Nineteen mutant Arabidopsis families resistant to the proline analog azetidine-2-carboxylic acid (ACA) were characterized in terms of NaCl tolerance and proline content. Mutants were selected from about 64,000 progeny of about 16,000 self-pollinated Columbia parents which had been mutated with ethyl methane sulfonate during seed imbibition. Selections were performed during seed germination on aseptic agar medium containing 0.2 to 0.25 mM ACA. Nineteen mutant families, 12 clearly independent, retained resistance to ACA in the M{sub 4} generation. Based on germination on 150 mM NaCl, 13 of the mutant families were more tolerant than the wild type. Two mutants of intermediate resistance to ACA were markedly more salt tolerant than the others. Four mutant families appeared to overproduce proline. Of these, only 3 showed slight increases in salt tolerance.

  16. Group 1 LEA proteins contribute to the desiccation and freeze tolerance of Artemia franciscana embryos during diapause.

    PubMed

    Toxopeus, Jantina; Warner, Alden H; MacRae, Thomas H

    2014-11-01

    Water loss either by desiccation or freezing causes multiple forms of cellular damage. The encysted embryos (cysts) of the crustacean Artemia franciscana have several molecular mechanisms to enable anhydrobiosis-life without water-during diapause. To better understand how cysts survive reduced hydration, group 1 late embryogenesis abundant (LEA) proteins, hydrophilic unstructured proteins that accumulate in the stress-tolerant cysts of A. franciscana, were knocked down using RNA interference (RNAi). Embryos lacking group 1 LEA proteins showed significantly lower survival than control embryos after desiccation and freezing, or freezing alone, demonstrating a role for group 1 LEA proteins in A. franciscana tolerance of low water conditions. In contrast, regardless of group 1 LEA protein presence, cysts responded similarly to hydrogen peroxide (H2O2) exposure, indicating little to no function for these proteins in diapause termination. This is the first in vivo study of group 1 LEA proteins in an animal and it contributes to the fundamental understanding of these proteins. Knowing how LEA proteins protect A. franciscana cysts from desiccation and freezing may have applied significance in aquaculture, where Artemia is an important feed source, and in the cryopreservation of cells for therapeutic applications.

  17. Improvement of the multiple-stress tolerance of an ethanologenic Saccharomyces cerevisiae strain by freeze-thaw treatment.

    PubMed

    Wei, Pingying; Li, Zilong; Lin, Yuping; He, Peng; Jiang, Ning

    2007-10-01

    An effective, simple, and convenient method to improve yeast's multiple-stress tolerance, and ethanol production was developed. After an ethanologenic Saccharomyces cerevisiae strain SC521 was treated by nine cycles of freeze-thaw, a mutant FT9-11 strain with higher multiple-stress tolerance was isolated, whose viabilities under acetic acid, ethanol, freeze-thaw, H(2)O(2), and heat-shock stresses were, respectively, 23-, 26-, 10- and 7-fold more than the parent strain at an initial value 2 x 10(7) c.f.u. per ml. Ethanol production of FT9-11 was similar (91.5 g ethanol l(-1)) to SC521 at 30 degrees C with 200 g glucose l(-1), and was better than the parent strain at 37 degrees C (72.5 g ethanol l(-1)), with 300 (111 g ethanol l(-1)) or with 400 (85 g ethanol l(-1)) g glucose l(-1).

  18. Zinc finger protein STOP1 is critical for proton tolerance in Arabidopsis and coregulates a key gene in aluminum tolerance.

    PubMed

    Iuchi, Satoshi; Koyama, Hiroyuki; Iuchi, Atsuko; Kobayashi, Yasufumi; Kitabayashi, Sadako; Kobayashi, Yuriko; Ikka, Takashi; Hirayama, Takashi; Shinozaki, Kazuo; Kobayashi, Masatomo

    2007-06-05

    Acid soil syndrome causes severe yield losses in various crop plants because of the rhizotoxicities of ions, such as aluminum (Al(3+)). Although protons (H(+)) could be also major rhizotoxicants in some soil types, molecular mechanisms of their tolerance have not been identified yet. One mutant that was hypersensitive to H(+) rhizotoxicity was isolated from ethyl methanesulfonate mutagenized seeds, and a single recessive mutation was found on chromosome 1. Positional cloning followed by genomic sequence analysis revealed that a missense mutation in the zinc finger domain in a predicted Cys(2)His(2)-type zinc finger protein, namely sensitive to proton rhizotoxicity (STOP)1, is the cause of hypersensitivity to H(+) rhizotoxicity. The STOP1 protein belongs to a functionally unidentified subfamily of zinc finger proteins, which consists of two members in Arabidopsis based on a Blast search. The stop1 mutation resulted in no effects on cadmium, copper, lanthanum, manganese and sodium chloride sensitivitities, whereas it caused hypersensitivity to Al(3+) rhizotoxicity. This stop1 mutant lacked the induction of the AtALMT1 gene encoding a malate transporter, which is concomitant with Al-induced malate exudation. There was no induction of AtALMT1 by Al(3+) treatment in the stop1 mutant. These results indicate that STOP1 plays a critical role in Arabidopsis tolerance to major stress factors in acid soils.

  19. Irradiation with low-dose gamma ray enhances tolerance to heat stress in Arabidopsis seedlings.

    PubMed

    Zhang, Liang; Zheng, Fengxia; Qi, Wencai; Wang, Tianqi; Ma, Lingyu; Qiu, Zongbo; Li, Jingyuan

    2016-06-01

    Gamma irradiation at low doses can stimulate the tolerance to environmental stress in plants. However, the knowledge regarding the mechanisms underlying the enhanced tolerance induced by low-dose gamma irradiation is far from fully understood. In this study, to investigate the physiological and molecular mechanisms of heat stress alleviated by low-dose gamma irradiation, the Arabidopsis seeds were exposed to a range of doses before subjected to heat treatment. Our results showed that 50-Gy gamma irradiation maximally promoted seedling growth in response to heat stress. The production rate of superoxide radical and contents of hydrogen peroxide and malondialdehyde in the seedlings irradiated with 50-Gy dose under heat stress were significantly lower than those of controls. The activities of antioxidant enzymes, glutathione (GSH) content and proline level in the gamma-irradiated seedlings were significantly increased compared with the controls. Furthermore, transcriptional expression analysis of selected genes revealed that some components related to heat tolerance were stimulated by low-dose gamma irradiation under heat shock. Our results suggest that low-dose gamma irradiation can modulate the physiological responses as well as gene expression related to heat tolerance, thus alleviating the stress damage in Arabidopsis seedlings. Copyright © 2016 Elsevier Inc. All rights reserved.

  20. A dehydrin gene isolated from feral olive enhances drought tolerance in Arabidopsis transgenic plants

    PubMed Central

    Chiappetta, Adriana; Muto, Antonella; Bruno, Leonardo; Woloszynska, Magdalena; Lijsebettens, Mieke Van; Bitonti, Maria B.

    2015-01-01

    Dehydrins belong to a protein family whose expression may be induced or enhanced by developmental process and environmental stresses that lead to cell dehydration. A dehydrin gene named OesDHN was isolated and characterized from oleaster (Olea europaea L. subsp. europaea, var. sylvestris), the wild form of olive. To elucidate the contribution of OesDHN in the development of drought tolerance, its expression levels were investigated in oleaster plants during development and under drought stress condition. The involvement of OesDHN in plant stress response was also evaluated in Arabidopsis transgenic lines, engineered to overexpress this gene, and exposed to a controlled mild osmotic stress. OesDHN expression was found to be modulated during development and induced under mild drought stress in oleaster plants. In addition, the Arabidopsis transgenic plants showed a better tolerance to osmotic stress than wild-type plants. The results demonstrated that OesDHN expression is induced by drought stress and is able to confer osmotic stress tolerance. We suggest a role for OesDHN, as a putative functional marker of plant stress tolerance. PMID:26175736

  1. Arabidopsis AINTEGUMENTA mediates salt tolerance by trans-repressing SCABP8.

    PubMed

    Meng, Lai-Sheng; Wang, Yi-Bo; Yao, Shun-Qiao; Liu, Aizhong

    2015-08-01

    The Arabidopsis AINTEGUMENTA (ANT) gene, which encodes an APETALA2 (AP2)-like transcription factor, controls plant organ cell number and organ size throughout shoot development. ANT is thus a key factor in the development of plant shoots. Here, we have found that ANT plays an essential role in conferring salt tolerance in Arabidopsis. ant-knockout mutants presented a salt-tolerant phenotype, whereas transgenic plants expressing ANT under the 35S promoter (35S:ANT) exhibited more sensitive phenotypes under high salt stress. Further analysis indicated that ANT functions mainly in the shoot response to salt toxicity. Target gene analysis revealed that ANT bound to the promoter of SOS3-LIKE CALCIUM BINDING PROTEIN 8 (SCABP8), which encodes a putative Ca(2+) sensor, thereby inhibiting expression of SCABP8 (also known as CBL10). It has been reported that the salt sensitivity of scabp8 is more prominent in shoot tissues. Genetic experiments indicated that the mutation of SCABP8 suppresses the ant-knockout salt-tolerant phenotype, implying that ANT functions as a negative transcriptional regulator of SCABP8 upon salt stress. Taken together, the above results reveal that ANT is a novel regulator of salt stress and that ANT binds to the SCABP8 promoter, mediating salt tolerance.

  2. Transcriptome analyses give insights into selenium-stress responses and selenium tolerance mechanisms in Arabidopsis.

    PubMed

    Van Hoewyk, Doug; Takahashi, Hideki; Inoue, Eri; Hess, Ann; Tamaoki, Masanori; Pilon-Smits, Elizabeth A H

    2008-02-01

    Selenate is chemically similar to sulfate and can be taken up and assimilated by plants via the same transporters and enzymes. In contrast to many other organisms, selenium (Se) has not been shown to be essential for higher plants. In excess, Se is toxic and restricts development. Both Se deficiency and toxicity pose problems worldwide. To obtain better insights into the effects of Se on plant metabolism and into plant mechanisms involved in Se tolerance, the transcriptome of Arabidopsis plants grown with or without selenate was studied and Se-responsive genes identified. Roots and shoots exhibited different Se-related changes in gene regulation and metabolism. Many genes involved in sulfur (S) uptake and assimilation were upregulated. Accordingly, Se treatment enhanced sulfate levels in plants, but the quantity of organic S metabolites decreased. Transcripts regulating the synthesis and signaling of ethylene and jasmonic acid were also upregulated by Se. Arabidopsis mutants defective in ethylene or jasmonate response pathways exhibited reduced tolerance to Se, suggesting an important role for these two stress hormones in Se tolerance. Selenate upregulated a variety of transcripts that were also reportedly induced by salt and osmotic stress. Selenate appeared to repress plant development, as suggested by the downregulation of genes involved in cell wall synthesis and auxin-regulated proteins. The Se-responsive genes discovered in this study may help create plants that can better tolerate and accumulate Se, which may enhance the effectiveness of Se phytoremediation or serve as Se-fortified food.

  3. An autophosphorylation site of the protein kinase SOS2 is important for salt tolerance in Arabidopsis.

    PubMed

    Fujii, Hiroaki; Zhu, Jian-Kang

    2009-01-01

    The protein kinase SOS2 (Salt Overly Sensitive 2) is essential for salt-stress signaling and tolerance in Arabidopsis. SOS2 is known to be activated by calcium-SOS3 and by phosphorylation at its activation loop. SOS2 is autophosphorylated in vitro, but the autophosphorylation site and its role in salt tolerance are not known. In this study, we identified an autophosphorylation site in SOS2 and analyzed its role in the responses of Arabidopsis to salt stress. Mass spectrometry analysis showed that Ser 228 of SOS2 is autophosphorylated. When this site was mutated to Ala, the autophosphorylation rate of SOS2 decreased. The substrate phosphorylation by the mutated SOS2 was also less than that by the wild-type SOS2. In contrast, changing Ser228 to Asp to mimic the autophosphorylation enhanced substrate phosphorylation by SOS2. Complementation tests in a sos2 mutant showed that the S228A but not the S228D mutation partially disrupted the function of SOS2 in salt tolerance. We also show that activation loop phosphorylation at Thr168 and autophosphorylation at Ser228 cannot substitute for each other, suggesting that both are required for salt tolerance. Our results indicate that Ser 228 of SOS2 is autophosphorylated and that this autophosphorylation is important for SOS2 function under salt stress.

  4. An Autophosphorylation Site of the Protein Kinase SOS2 Is Important for Salt Tolerance in Arabidopsis

    PubMed Central

    Fujii, Hiroaki; Zhu, Jian-Kang

    2009-01-01

    The protein kinase SOS2 (Salt Overly Sensitive 2) is essential for salt-stress signaling and tolerance in Arabidopsis. SOS2 is known to be activated by calcium-SOS3 and by phosphorylation at its activation loop. SOS2 is autophosphorylated in vitro, but the autophosphorylation site and its role in salt tolerance are not known. In this study, we identified an autophosphorylation site in SOS2 and analyzed its role in the responses of Arabidopsis to salt stress. Mass spectrometry analysis showed that Ser 228 of SOS2 is autophosphorylated. When this site was mutated to Ala, the autophosphorylation rate of SOS2 decreased. The substrate phosphorylation by the mutated SOS2 was also less than that by the wild-type SOS2. In contrast, changing Ser228 to Asp to mimic the autophosphorylation enhanced substrate phosphorylation by SOS2. Complementation tests in a sos2 mutant showed that the S228A but not the S228D mutation partially disrupted the function of SOS2 in salt tolerance. We also show that activation loop phosphorylation at Thr168 and autophosphorylation at Ser228 cannot substitute for each other, suggesting that both are required for salt tolerance. Our results indicate that Ser 228 of SOS2 is autophosphorylated and that this autophosphorylation is important for SOS2 function under salt stress. PMID:19529820

  5. Sm-Like Protein-Mediated RNA Metabolism Is Required for Heat Stress Tolerance in Arabidopsis

    PubMed Central

    Okamoto, Masanori; Matsui, Akihiro; Tanaka, Maho; Morosawa, Taeko; Ishida, Junko; Iida, Kei; Mochizuki, Yoshiki; Toyoda, Tetsuro; Seki, Motoaki

    2016-01-01

    Sm-like proteins play multiple functions in RNA metabolism, which is essential for biological processes such as stress responses in eukaryotes. The Arabidopsis thaliana sad1 mutant has a mutation of sm-like protein 5 (LSM5) and shows impaired drought and salt stress tolerances. The lsm5/sad1 mutant also showed hypersensitivity to heat stress. GFP-fused LSM5/SAD1 was localized in the nucleus under optimal growth conditions. After heat stress treatment, GFP-fused LSM5/SAD1 fluorescence was also observed as small cytoplasmic dots, in addition to nuclear localization. Whole genome transcriptome analysis revealed that many genes in Arabidopsis were drastically changed in response to heat stress. More heat-responsive genes were highly expressed in lsm5/sad1 mutant at both 2 and 6 h after heat stress treatment. Additionally, intron-retained and capped transcripts accumulated in the lsm5/sad1 mutant after heat stress treatment. In this study, we also identified non-Arabidopsis Genome Initiative transcripts that were expressed from unannotated regions. Most of these transcripts were antisense transcripts, and many capped non-AGI transcripts accumulated in the lsm5/sad1 mutant during heat stress treatment. These results indicated that LSM5/SAD1 functions to degrade aberrant transcripts through appropriate mRNA splicing and decapping, and precise RNA metabolic machinery is required for heat stress tolerance. PMID:27493656

  6. The Arabidopsis PLAT domain protein1 promotes abiotic stress tolerance and growth in tobacco.

    PubMed

    Hyun, Tae Kyung; Albacete, Alfonso; van der Graaff, Eric; Eom, Seung Hee; Großkinsky, Dominik K; Böhm, Hannah; Janschek, Ursula; Rim, Yeonggil; Ali, Walid Wahid; Kim, Soo Young; Roitsch, Thomas

    2015-08-01

    Plant growth and consequently crop yield can be severely compromised by abiotic and biotic stress conditions. Transgenic approaches that resulted in increased tolerance against abiotic stresses often were typically accompanied by adverse effects on plant growth and fitness under optimal growing conditions. Proteins that belong to the PLAT-plant-stress protein family harbour a single PLAT (Polycystin, Lipoxygenase, Alpha-toxin and Triacylglycerol lipase) domain and are ubiquitously present in monocot and dicot plant species. Until now, only limited data is available for PLAT-plant-stress family members, which suggested that these proteins in general could promote tolerance towards stress responses. We studied the function of the Arabidopsis PLAT-plant-stress protein AtPLAT1 employing heterologous gain-of-function analysis in tobacco. AtPLAT1 conferred increased abiotic stress tolerance in tobacco, evident by improved tolerance towards cold, drought and salt stresses, and promoted growth, reflected by a faster development under non-stressed conditions. However, the overexpression of AtPLAT1 in tobacco reduced the tolerance towards biotic stress conditions and, therefore, could be involved in regulating the crosstalk between abiotic and biotic stress responses. Thus, we showed that heterologously expressed AtPLAT1 functions as positive regulator of abiotic stress tolerance and plant growth, which could be an important new asset for strategies to develop plants with improved abiotic stress tolerance, without growth and subsequent yield penalties under optimal growth conditions.

  7. Growth characteristics of freeze-tolerant baker's yeast Saccharomyces cerevisiae AFY in aerobic batch culture.

    PubMed

    Ji, Meng; Miao, Yelian; Chen, Jie Yu; You, Yebing; Liu, Feilong; Xu, Lin

    2016-01-01

    Saccharomyces cerevisiae AFY is a novel baker's yeast strain with strong freeze-tolerance, and can be used for frozen-dough processing. The present study armed to clarify the growth characteristics of the yeast AFY. Aerobic batch culture experiments of yeast AFY were carried out using media with various initial glucose concentrations, and the culture process was analyzed kinetically. The growth of the yeast AFY exhibited a diauxic pattern with the first growth stage consuming glucose and the second growth stage consuming ethanol. The cell yield decreased with increasing initial glucose concentration in the first growth stage, and also decreased with increasing initial ethanol concentration in the second growth stage. In the initial glucose concentration range of 5.0-40.0 g/L, the simultaneous equations of Monod equation, Luedeking-Piret equation and pseudo-Luedeking-Piret equation could be used to describe the concentrations of cell, ethanol and glucose in either of the two exponential growth phases. At the initial glucose concentrations of 5.0, 10.0 and 40.0 g/L, the first exponential growth phase had a maximal specific cell growth rate of 0.52, 0.98 and 0.99 h(-1), while the second exponential growth phase had a maximal specific cell growth rate of 0.11, 0.06 and 0.07 h(-1), respectively. It was indicated that the efficiency of the yeast production could be improved by reducing the ethanol production in the first growth stage.

  8. [Freezing tolerance of winter wheat plants depends on adaptation of photosynthesis and respiration in different time intervals].

    PubMed

    Klimov, S V

    2009-01-01

    This study is devoted to CO2 gas exchange (true photosynthesis at light saturation (P), dark respiration (R), and P/R ratio) in vegetating and cold-hardened winter wheat (Triticum aestivum L.) plants (cultivar Mironovskaya 808) in relation to their freezing tolerance. Under natural cultivation conditions, freezing tolerance of plants depended on adaptive changes in the shape of P and R curves in the temperature range from 20 to -2 degrees C. These changes, induced by cold hardening and treatment of plants with the photosynthesis inhibitor diuron, were observed within month and week ranges. Under laboratory conditions, the P/R ratio in vegetating plants increased three times within an hour range as the temperature decreased from 22 to 0 degree C. The P/R ratio also decreased within a minute range as a result of partial inhibition of photosynthesis with diuron and immediately decreased when CO2 concentration in the air was reduced from 419 to 0 microl/l. The P/R ratio decreased primarily at the expense of a decrease in P. The decrease in P/R was more pronounced at low temperatures, indicating variability of low-temperature tolerance of photosynthesis within a minute range. The possibility of plant adaptation to nonsimultaneous temperature changes under natural conditions via adaptive changes in temperature tolerance of the photosynthetic apparatus is discussed.

  9. Increased salt and drought tolerance by D-ononitol production in transgenic Arabidopsis thaliana.

    PubMed

    Ahn, Chulhyun; Park, Uhnmee; Park, Phun Bum

    2011-12-02

    The methylation of myo-inositol forms O-methyl inositol (D-ononitol) when plants are under abiotic stress in a reaction catalyzed by myo-inositol methyltransferase (IMT). D-Ononitol can serve as an osmoprotectant that prevents water loss in plants. We isolated the IMT cDNA from Glycine max and found by RT-PCR analysis that GmIMT transcripts are induced by drought and salinity stress treatments in the leaves of soybean seedlings. We confirmed the protein product of GmIMT and its substrate using a recombinant system in E. coli. Transgenic Arabidopsis plants over-expressing GmIMT displayed improved tolerance to dehydration stress treatment and to a lesser extent high salinity stress treatment. These results indicate that GmIMT is functional in heterologous Arabidopsis plants.

  10. Nitric oxide synthase-dependent nitric oxide production is associated with salt tolerance in Arabidopsis.

    PubMed

    Zhao, Min-Gui; Tian, Qiu-Ying; Zhang, Wen-Hao

    2007-05-01

    Nitric oxide (NO) has emerged as a key molecule involved in many physiological processes in plants. To characterize roles of NO in tolerance of Arabidopsis (Arabidopsis thaliana) to salt stress, effect of NaCl on Arabidopsis wild-type and mutant (Atnoa1) plants with an impaired in vivo NO synthase (NOS) activity and a reduced endogenous NO level was investigated. Atnoa1 mutant plants displayed a greater Na+ to K+ ratio in shoots than wild-type plants due to enhanced accumulation of Na+ and reduced accumulation of K+ when exposed to NaCl. Germination of Atnoa1 seeds was more sensitive to NaCl than that of wild-type seeds, and wild-type plants exhibited higher survival rates than Atnoa1 plants when grown under salt stress. Atnoa1 plants had higher levels of hydrogen peroxide than wild-type plants under both control and salt stress, suggesting that Atnoa1 is more vulnerable to salt and oxidative stress than wild-type plants. Treatments of wild-type plants with NOS inhibitor and NO scavenger reduced endogenous NO levels and enhanced NaCl-induced increase in Na+ to K+ ratio. Exposure of wild-type plants to NaCl inhibited NOS activity and reduced quantity of NOA1 protein, leading to a decrease in endogenous NO levels measured by NO-specific fluorescent probe. Treatment of Atnoa1 plants with NO donor sodium nitroprusside attenuated the NaCl-induced increase in Na+ to K+ ratio. Therefore, these findings provide direct evidence to support that disruption of NOS-dependent NO production is associated with salt tolerance in Arabidopsis.

  11. Freezing tolerance and low molecular weight cryoprotectants in an invasive parasitic fly, the deer ked (Lipoptena cervi).

    PubMed

    Nieminen, Petteri; Paakkonen, Tommi; Eerilä, Harri; Puukka, Katri; Riikonen, Joakim; Lehto, Vesa-Pekka; Mustonen, Anne-Mari

    2012-01-01

    Insect cold hardiness is often mediated by low molecular weight cryoprotectants, such as sugars, polyols, and amino acids (AA). While many free-living northern insects must cope with extended periods of freezing ambient temperatures (Ta), the ectoparasitic deer ked Lipoptena cervi imago can encounter subfreezing Ta only during a short autumnal period between hatching and host location. Subsequently, it benefits from the body temperature of the cervid host for survival in winter. This study investigated the cold tolerance of the species by determining its lower lethal temperature (100% mortality, LLT100) during faster and slower cold acclimation, by determining the supercooling point (SCP) and by measuring the concentrations of potential low molecular weight cryoprotectants. The LLT100 of the deer ked was approximately -16 ° C, which would enable it to survive freezing nighttime Ta not only in its current area of distribution but also further north. The SCP was -7.8 ° C, clearly higher than the LLT100 , indicating that the deer ked displays freezing tolerance. The concentrations of free AA, especially nonessential AA, were higher in the cold-acclimated deer keds similar to several other insects. The concentrations of proline increased together with γ-aminobutyrate, arginine, asparagine, cystine, glutamate, glutamine, hydroxylysine, sarcosine, serine, and taurine. AA could be hypothesized to act as cryoprotectants by, e.g., protecting enzymes and lipid membranes from damage caused by cold. Copyright © 2011 Wiley Periodicals, Inc., A Wiley Company.

  12. Expression of OsCAS (Calcium-Sensing Receptor) in an Arabidopsis Mutant Increases Drought Tolerance.

    PubMed

    Zhao, Xin; Xu, Mengmeng; Wei, Rongrong; Liu, Yang

    2015-01-01

    that helps to compensate for the absence of CaS in Arabidopsis and increases the drought stress tolerance of transgenic plants.

  13. Suppression of Arabidopsis RING E3 ubiquitin ligase AtATL78 increases tolerance to cold stress and decreases tolerance to drought stress.

    PubMed

    Kim, Soo Jin; Kim, Woo Taek

    2013-08-19

    AtATL78 is an Arabidopsis RING E3 ubiquitin ligase. RT-PCR and promoter-GUS assays revealed that AtATL78 was up-regulated by cold stress and down-regulated by drought. AtATL78 was localized at the plasma-membrane. Suppression of AtATL78 increased tolerance to cold stress but decreased tolerance to drought. Our data suggests that AtATL78 is a negative regulator of cold stress response and a positive regulator of drought stress response in Arabidopsis. These results further suggest that AtATL78 plays opposing roles in cold and drought stress responses.

  14. Quantifying the dynamics of light tolerance in Arabidopsis plants during ontogenesis.

    PubMed

    Carvalho, Fabricio E L; Ware, Maxwell A; Ruban, Alexander V

    2015-12-01

    The amount of light plants can tolerate during different phases of ontogenesis remains largely unknown. This was addressed here employing a novel methodology that uses the coefficient of photochemical quenching (qP) to assess the intactness of photosystem II reaction centres. Fluorescence quenching coefficients, total chlorophyll content and concentration of anthocyanins were determined weekly during the juvenile, adult, reproductive and senescent phases of plant ontogenesis. This enabled quantification of the protective effectiveness of non-photochemical fluorescence quenching (NPQ) and determination of light tolerance. The light intensity that caused photoinhibition in 50% of leaf population increased from ∼70 μmol m(-2)  s(-1) , for 1-week-old seedlings, to a maximum of 1385 μmol m(-2)  s(-1) for 8-week-old plants. After 8 weeks, the tolerated light intensity started to gradually decline, becoming only 332 μmol m(-2)  s(-1) for 13-week-old plants. The dependency of light tolerance on plant age was well-related to the amplitude of protective NPQ (pNPQ) and the electron transport rates (ETRs). Light tolerance did not, however, show a similar trend to chlorophyll a/b ratios and content of anthocyanins. Our data suggest that pNPQ is crucial in defining the capability of high light tolerance by Arabidopsis plants during ontogenesis. © 2015 John Wiley & Sons Ltd.

  15. Identification of the Submergence Tolerance QTL Come Quick Drowning1 (CQD1) in Arabidopsis thaliana.

    PubMed

    Akman, Melis; Kleine, Rogier; van Tienderen, Peter H; Schranz, M Eric

    2017-02-16

    Global climate change is predicted to increase water precipitation fluctuations and lead to localized prolonged floods in agricultural fields and natural plant communities. Thus, understanding the genetic basis of submergence tolerance is crucial in order to improve plant survival under these conditions. In this study, we performed a quantitative trait locus (QTL) analysis in Arabidopsis to identify novel candidate genes for increased submergence tolerance by using Kas-1 and Col (gl1) parental accessions and their derived recombinant inbred lines (RILs). We measured survival after submergence in dark for a 13-day period and used median lethal time, LT50 values for the QTL analysis. A major QTL, the Come Quick, Drowning (CQD1) locus, was detected in 2 independent experiments on the lower arm of chromosome 5 involved in higher submergence tolerance in the parental accession Kas-1. For fine-mapping, we then constructed near isogenic lines (NILs) by backcrossing the CQD1 QTL region. We also analyzed QTL regions related to size, leaf number, flowering, or survival in darkness and none of the QTL related to these traits overlapped with CQD1. The submergence tolerance QTL, CQD1, region detected in this study includes genes that have potential to be novel candidates effecting submergence tolerance such as trehalose-6-phosphate phosphatase and respiratory burst oxidase protein D. Gene expression and functional analysis for these genes under submergence would reveal the significance of these novel candidates and provide new perspectives for understanding genetic basis of submergence tolerance.

  16. Overexpression of quinone reductase from Salix matsudana Koidz enhances salt tolerance in transgenic Arabidopsis thaliana.

    PubMed

    Song, Xixi; Fang, Jie; Han, Xiaojiao; He, Xuelian; Liu, Mingying; Hu, Jianjun; Zhuo, Renying

    2016-01-15

    Quinone reductase (QR) is an oxidative-related gene and few studies have focused on its roles concerning salt stress tolerance in plants. In this study, we cloned and analyzed the QR gene from Salix matsudana, a willow with tolerance of moderate salinity. The 612-bp cDNA corresponding to SmQR encodes 203 amino acids. Expression of SmQR in Escherichia coli cells enhanced their tolerance under salt stress. In addition, transgenic Arabidopsis thaliana lines overexpressing SmQR exhibited higher salt tolerance as compared with WT, with higher QR activity and antioxidant enzyme activity as well as higher chlorophyll content, lower methane dicarboxylic aldehyde (MDA) content and electric conductivity under salt stress. Nitro blue tetrazolium (NBT) and 3,3'-diaminobenzidine (DAB) staining also indicated that the transgenic plants accumulated less reactive oxygen species compared to WT when exposed to salt stress. Overall, our results suggested that SmQR plays a significant role in salt tolerance and that this gene may be useful for biotechnological development of plants with improved tolerance of salinity.

  17. Assessing Tolerance to Heavy-Metal Stress in Arabidopsis thaliana Seedlings.

    PubMed

    Remy, Estelle; Duque, Paula

    2016-01-01

    Heavy-metal soil contamination is one of the major abiotic stress factors that, by negatively affecting plant growth and development, severely limit agricultural productivity worldwide. Plants have evolved various tolerance and detoxification strategies in order to cope with heavy-metal toxicity while ensuring adequate supply of essential micronutrients at the whole-plant as well as cellular levels. Genetic studies in the model plant Arabidopsis thaliana have been instrumental in elucidating such mechanisms. The root assay constitutes a very powerful and simple method to assess heavy-metal stress tolerance in Arabidopsis seedlings. It allows the simultaneous determination of all the standard growth parameters affected by heavy-metal stress (primary root elongation, lateral root development, shoot biomass, and chlorophyll content) in a single experiment. Additionally, this protocol emphasizes the tips and tricks that become particularly useful when quantifying subtle alterations in tolerance to a given heavy-metal stress, when simultaneously pursuing a large number of plant lines, or when testing sensitivity to a wide range of heavy metals for a single line.

  18. Overexpression of AtCpNifS enhances selenium tolerance and accumulation in Arabidopsis.

    PubMed

    Van Hoewyk, Douglas; Garifullina, Gulnara F; Ackley, Ashley R; Abdel-Ghany, Salah E; Marcus, Matthew A; Fakra, Sirine; Ishiyama, Keiki; Inoue, Eri; Pilon, Marinus; Takahashi, Hideki; Pilon-Smits, Elizabeth A H

    2005-11-01

    Selenium (Se) is an essential element for many organisms but is toxic at higher levels. CpNifS is a chloroplastic NifS-like protein in Arabidopsis (Arabidopsis thaliana) that can catalyze the conversion of cysteine into alanine and elemental sulfur (S0) and of selenocysteine into alanine and elemental Se (Se0). We overexpressed CpNifS to investigate the effects on Se metabolism in plants. CpNifS overexpression significantly enhanced selenate tolerance (1.9-fold) and Se accumulation (2.2-fold). CpNifS overexpressors showed significantly reduced Se incorporation into protein, which may explain their higher Se tolerance. Also, sulfur accumulation was enhanced by approximately 30% in CpNifS overexpressors, both on media with and without selenate. Root transcriptome changes in response to selenate mimicked the effects observed under sulfur starvation. There were only a few transcriptome differences between CpNifS-overexpressing plants and wild type, besides the 25- to 40-fold increase in CpNifS levels. Judged from x-ray analysis of near edge spectrum, both CpNifS overexpressors and wild type accumulated mostly selenate (Se(VI)). In conclusion, overexpression of this plant NifS-like protein had a pronounced effect on plant Se metabolism. The observed enhanced Se accumulation and tolerance of CpNifS overexpressors show promise for use in phytoremediation.

  19. Abscisic acid (ABA) sensitivity regulates desiccation tolerance in germinated Arabidopsis seeds.

    PubMed

    Maia, Julio; Dekkers, Bas J W; Dolle, Miranda J; Ligterink, Wilco; Hilhorst, Henk W M

    2014-07-01

    During germination, orthodox seeds lose their desiccation tolerance (DT) and become sensitive to extreme drying. Yet, DT can be rescued, in a well-defined developmental window, by the application of a mild osmotic stress before dehydration. A role for abscisic acid (ABA) has been implicated in this stress response and in DT re-establishment. However, the path from the sensing of an osmotic cue and its signaling to DT re-establishment is still largely unknown. Analyses of DT, ABA sensitivity, ABA content and gene expression were performed in desiccation-sensitive (DS) and desiccation-tolerant Arabidopsis thaliana seeds. Furthermore, loss and re-establishment of DT in germinated Arabidopsis seeds was studied in ABA-deficient and ABA-insensitive mutants. We demonstrate that the developmental window in which DT can be re-established correlates strongly with the window in which ABA sensitivity is still present. Using ABA biosynthesis and signaling mutants, we show that this hormone plays a key role in DT re-establishment. Surprisingly, re-establishment of DT depends on the modulation of ABA sensitivity rather than enhanced ABA content. In addition, the evaluation of several ABA-insensitive mutants, which can still produce normal desiccation-tolerant seeds, but are impaired in the re-establishment of DT, shows that the acquisition of DT during seed development is genetically different from its re-establishment during germination. © 2014 The Authors. New Phytologist © 2014 New Phytologist Trust.

  20. Temperature-stress resistance and tolerance along a latitudinal cline in North American Arabidopsis lyrata.

    PubMed

    Wos, Guillaume; Willi, Yvonne

    2015-01-01

    The study of latitudinal gradients can yield important insights into adaptation to temperature stress. Two strategies are available: resistance by limiting damage, or tolerance by reducing the fitness consequences of damage. Here we studied latitudinal variation in resistance and tolerance to frost and heat and tested the prediction of a trade-off between the two strategies and their costliness. We raised plants of replicate maternal seed families from eight populations of North American Arabidopsis lyrata collected along a latitudinal gradient in climate chambers and exposed them repeatedly to either frost or heat stress, while a set of control plants grew under standard conditions. When control plants reached maximum rosette size, leaf samples were exposed to frost and heat stress, and electrolyte leakage (PEL) was measured and treated as an estimate of resistance. Difference in maximum rosette size between stressed and control plants was used as an estimate of tolerance. Northern populations were more frost resistant, and less heat resistant and less heat tolerant, but-unexpectedly-they were also less frost tolerant. Negative genetic correlations between resistance and tolerance to the same and different thermal stress were generally not significant, indicating only weak trade-offs. However, tolerance to frost was consistently accompanied by small size under control conditions, which may explain the non-adaptive latitudinal pattern for frost tolerance. Our results suggest that adaptation to frost and heat is not constrained by trade-offs between them. But the cost of frost tolerance in terms of plant size reduction may be important for the limits of species distributions and climate niches.

  1. Temperature-Stress Resistance and Tolerance along a Latitudinal Cline in North American Arabidopsis lyrata

    PubMed Central

    Wos, Guillaume; Willi, Yvonne

    2015-01-01

    The study of latitudinal gradients can yield important insights into adaptation to temperature stress. Two strategies are available: resistance by limiting damage, or tolerance by reducing the fitness consequences of damage. Here we studied latitudinal variation in resistance and tolerance to frost and heat and tested the prediction of a trade-off between the two strategies and their costliness. We raised plants of replicate maternal seed families from eight populations of North American Arabidopsis lyrata collected along a latitudinal gradient in climate chambers and exposed them repeatedly to either frost or heat stress, while a set of control plants grew under standard conditions. When control plants reached maximum rosette size, leaf samples were exposed to frost and heat stress, and electrolyte leakage (PEL) was measured and treated as an estimate of resistance. Difference in maximum rosette size between stressed and control plants was used as an estimate of tolerance. Northern populations were more frost resistant, and less heat resistant and less heat tolerant, but—unexpectedly—they were also less frost tolerant. Negative genetic correlations between resistance and tolerance to the same and different thermal stress were generally not significant, indicating only weak trade-offs. However, tolerance to frost was consistently accompanied by small size under control conditions, which may explain the non-adaptive latitudinal pattern for frost tolerance. Our results suggest that adaptation to frost and heat is not constrained by trade-offs between them. But the cost of frost tolerance in terms of plant size reduction may be important for the limits of species distributions and climate niches. PMID:26110428

  2. MAN3 gene regulates cadmium tolerance through the glutathione-dependent pathway in Arabidopsis thaliana.

    PubMed

    Chen, Jian; Yang, Libo; Gu, Ju; Bai, Xiaoya; Ren, Yongbin; Fan, Tingting; Han, Yi; Jiang, Li; Xiao, Fangming; Liu, Yongsheng; Cao, Shuqing

    2015-01-01

    Pollution of soil by the heavy metal cadmium (Cd) is a global environmental problem. The glutathione (GSH)-dependent phytochelatin (PC) synthesis pathway is one of the most important mechanisms contributing to Cd accumulation and tolerance. However, the regulation of this pathway is poorly understood. Here, we identified an Arabidopsis thaliana cadmium-tolerant dominant mutant xcd1-D (XVE system-induced cadmium-tolerance 1) and cloned XCD1 gene (previously called MAN3), which encodes an endo-β-mannanase. Overexpression of MAN3 led to enhanced Cd accumulation and tolerance, whereas loss-of-function of MAN3 resulted in decreased Cd accumulation and tolerance. In the presence of estradiol, enhanced Cd accumulation and tolerance in xcd1-D was associated with GSH-dependent, Cd-activated synthesis of PCs, which was correlated with coordinated activation of gene expression. Cd stress-induced expression of MAN3 and the consequently increased mannanase activity, led to increased mannose content in cell walls. Moreover, mannose treatment not only rescued the Cd-sensitive phenotype of the xcd1-2 mutant, but also improved the Cd tolerance of wild-type plants. Significantly, this mannose-mediated Cd accumulation and tolerance is dependent on GSH-dependent PC concentrations via coordinated control of expression of genes involved in PC synthesis. Our results suggest that MAN3 regulates the GSH-dependent PC synthesis pathway that contributes to Cd accumulation and tolerance in A. thaliana by coordinated control of gene expression. © 2014 The Authors. New Phytologist © 2014 New Phytologist Trust.

  3. Ectopic expression of Arabidopsis RCI2A gene contributes to cold tolerance in tomato.

    PubMed

    Sivankalyani, Velu; Geetha, Mahalingam; Subramanyam, Kondeti; Girija, Shanmugam

    2015-04-01

    Cold is a major stress that limits the quality and productivity of economically important crops such as tomato (Solanum lycopersicum L.). Generating a cold-stress-tolerant tomato by expressing cold-inducible genes would increase agricultural strategies. Rare cold-inducible 2a (RCI2A) is expressed in Arabidopsis, but its molecular function during cold stress is not fully understood. Here we ectopically expressed Arabidopsis RCI2A in transgenic tomato to evaluate tolerance to cold stress without altering agronomic traits. Biochemical and physiological study demonstrated that expression of RCI2A in transgenic tomato enhanced the activity of peroxidase and ascorbate peroxidase (APX) and reduced the accumulation of H2O2, alleviated lipid peroxidation, increased the accumulation of chlorophyll, reduced chilling-induced membrane damage, retained relative water content and enhanced cold tolerance. A motif search revealed that the motifs of photosystem II (PSII) phosphoproteins PsbJ and PsbH and reaction-center proteins PsbL and PsbK were common to cold-inducible RCI2A and peroxidase proteins RCI3A, tomato peroxidase (TPX1), TPX2, tomato ascorbate peroxidase (APX1), and horseradish peroxidase (HRP-c). In addition to membrane protection, RCI2A may cross talk with PSII-associated proteins or peroxidase family enzymes in response to cold stress. Our findings may strengthen the understanding of the molecular function of RCI2A in cold-stress tolerance. RCI2A could be used to improve abiotic stress tolerance in agronomic crops.

  4. Arabidopsis sos1 mutant in a salt-tolerant accession revealed an importance of salt acclimation ability in plant salt tolerance.

    PubMed

    Ariga, Hirotaka; Katori, Taku; Yoshihara, Ryouhei; Hase, Yoshihiro; Nozawa, Shigeki; Narumi, Issay; Iuchi, Satoshi; Kobayashi, Masatomo; Tezuka, Kenji; Sakata, Yoichi; Hayashi, Takahisa; Taji, Teruaki

    2013-07-01

    An analysis of the salinity tolerance of 354 Arabidopsis thaliana accessions showed that some accessions were more tolerant to salt shock than the reference accession, Col-0, when transferred from 0 to 225 mM NaCl. In addition, several accessions, including Zu-0, showed marked acquired salt tolerance after exposure to moderate salt stress. It is likely therefore that Arabidopsis plants have at least two types of tolerance, salt shock tolerance and acquired salt tolerance. To evaluate a role of well-known salt shock tolerant gene SOS1 in acquired salt tolerance, we isolated a sos1 mutant from ion-beam-mutagenized Zu-0 seedlings. The mutant showed severe growth inhibition under salt shock stress owing to a single base deletion in the SOS1 gene and was even more salt sensitive than Col-0. Nevertheless, it was able to survive after acclimation on 100 mM NaCl for 7 d followed by 750 mM sorbitol for 20 d, whereas Col-0 became chlorotic under the same conditions. We propose that genes for salt acclimation ability are different from genes for salt shock tolerance and play an important role in the acquisition of salt or osmotic tolerance.

  5. Arabidopsis sos1 mutant in a salt-tolerant accession revealed an importance of salt acclimation ability in plant salt tolerance

    PubMed Central

    Ariga, Hirotaka; Katori, Taku; Yoshihara, Ryouhei; Hase, Yoshihiro; Nozawa, Shigeki; Narumi, Issay; Iuchi, Satoshi; Kobayashi, Masatomo; Tezuka, Kenji; Sakata, Yoichi; Hayashi, Takahisa; Taji, Teruaki

    2013-01-01

    An analysis of the salinity tolerance of 354 Arabidopsis thaliana accessions showed that some accessions were more tolerant to salt shock than the reference accession, Col-0, when transferred from 0 to 225 mM NaCl. In addition, several accessions, including Zu-0, showed marked acquired salt tolerance after exposure to moderate salt stress. It is likely therefore that Arabidopsis plants have at least two types of tolerance, salt shock tolerance and acquired salt tolerance. To evaluate a role of well-known salt shock tolerant gene SOS1 in acquired salt tolerance, we isolated a sos1 mutant from ion-beam-mutagenized Zu-0 seedlings. The mutant showed severe growth inhibition under salt shock stress owing to a single base deletion in the SOS1 gene and was even more salt sensitive than Col-0. Nevertheless, it was able to survive after acclimation on 100 mM NaCl for 7 d followed by 750 mM sorbitol for 20 d, whereas Col-0 became chlorotic under the same conditions. We propose that genes for salt acclimation ability are different from genes for salt shock tolerance and play an important role in the acquisition of salt or osmotic tolerance. PMID:23656872

  6. Expression of Arabidopsis Bax Inhibitor-1 in transgenic sugarcane confers drought tolerance.

    PubMed

    Ramiro, Daniel Alves; Melotto-Passarin, Danila Montewka; Barbosa, Mariana de Almeida; Santos, Flavio Dos; Gomez, Sergio Gregorio Perez; Massola Júnior, Nelson Sidnei; Lam, Eric; Carrer, Helaine

    2016-09-01

    The sustainability of global crop production is critically dependent on improving tolerance of crop plants to various types of environmental stress. Thus, identification of genes that confer stress tolerance in crops has become a top priority especially in view of expected changes in global climatic patterns. Drought stress is one of the abiotic stresses that can result in dramatic loss of crop productivity. In this work, we show that transgenic expression of a highly conserved cell death suppressor, Bax Inhibitor-1 from Arabidopsis thaliana (AtBI-1), can confer increased tolerance of sugarcane plants to long-term (>20 days) water stress conditions. This robust trait is correlated with an increased tolerance of the transgenic sugarcane plants, especially in the roots, to induction of endoplasmic reticulum (ER) stress by the protein glycosylation inhibitor tunicamycin. Our findings suggest that suppression of ER stress in C4 grasses, which include important crops such as sorghum and maize, can be an effective means of conferring improved tolerance to long-term water deficit. This result could potentially lead to improved resilience and yield of major crops in the world.

  7. Allantoin Increases Cadmium Tolerance in Arabidopsis via Activation of Antioxidant Mechanisms.

    PubMed

    Nourimand, Maryam; Todd, Christopher D

    2016-12-01

    Plants apply various molecular, physiological and morphological strategies in response to undesirable environmental conditions. One of the possible responses which may contribute to surviving stressful conditions is the accumulation of ureides. Ureides are recognized as important nitrogen-rich compounds involved in recycling nitrogen in plants to support growth and reproduction. Amongst them, allantoin not only serves as a transportable nitrogen-rich compound, but has also been suggested to protect plants from abiotic stresses via minimizing oxidative damage. This work focuses on the effect of cadmium (Cd) on ureide metabolism in Arabidopsis, in order to clarify the potential role of allantoin in plant tolerance to heavy metals. In response to Cd treatment, allantoin levels increase in Arabidopsis thaliana, ecotype Col-0, due to reduced allantoinase (ALN) gene expression and enzyme activity. This coincides with increases in uricase (UO) transcripts. UO and ALN encode the enzymes for the production and degradation of allantoin, respectively. ALN-negative aln-3 Arabidopsis mutants with elevated allantoin levels demonstrate resistance to soil-applied CdCl2, up to 1,500 μM. Although aln-3 mutants take up and store more Cd within their leaf tissue, they contain less damaging superoxide radicals. The protective mechanism of aln-3 mutants appears to involve enhancing the activity of antioxidant enzymes such as superoxide dismutase and ascorbate peroxidase. © The Author 2016. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oup.com.

  8. The re-establishment of desiccation tolerance in germinated Arabidopsis thaliana seeds and its associated transcriptome.

    PubMed

    Maia, Julio; Dekkers, Bas J W; Provart, Nicholas J; Ligterink, Wilco; Hilhorst, Henk W M

    2011-01-01

    The combination of robust physiological models with "omics" studies holds promise for the discovery of genes and pathways linked to how organisms deal with drying. Here we used a transcriptomics approach in combination with an in vivo physiological model of re-establishment of desiccation tolerance (DT) in Arabidopsis thaliana seeds. We show that the incubation of desiccation sensitive (DS) germinated Arabidopsis seeds in a polyethylene glycol (PEG) solution re-induces the mechanisms necessary for expression of DT. Based on a SNP-tile array gene expression profile, our data indicates that the re-establishment of DT, in this system, is related to a programmed reversion from a metabolic active to a quiescent state similar to prior to germination. Our findings show that transcripts of germinated seeds after the PEG-treatment are dominated by those encoding LEA, seed storage and dormancy related proteins. On the other hand, a massive repression of genes belonging to many other classes such as photosynthesis, cell wall modification and energy metabolism occurs in parallel. Furthermore, comparison with a similar system for Medicago truncatula reveals a significant overlap between the two transcriptomes. Such overlap may highlight core mechanisms and key regulators of the trait DT. Taking into account the availability of the many genetic and molecular resources for Arabidopsis, the described system may prove useful for unraveling DT in higher plants.

  9. The Re-Establishment of Desiccation Tolerance in Germinated Arabidopsis thaliana Seeds and Its Associated Transcriptome

    PubMed Central

    Maia, Julio; Dekkers, Bas J. W.; Provart, Nicholas J.; Ligterink, Wilco; Hilhorst, Henk W. M.

    2011-01-01

    The combination of robust physiological models with “omics” studies holds promise for the discovery of genes and pathways linked to how organisms deal with drying. Here we used a transcriptomics approach in combination with an in vivo physiological model of re-establishment of desiccation tolerance (DT) in Arabidopsis thaliana seeds. We show that the incubation of desiccation sensitive (DS) germinated Arabidopsis seeds in a polyethylene glycol (PEG) solution re-induces the mechanisms necessary for expression of DT. Based on a SNP-tile array gene expression profile, our data indicates that the re-establishment of DT, in this system, is related to a programmed reversion from a metabolic active to a quiescent state similar to prior to germination. Our findings show that transcripts of germinated seeds after the PEG-treatment are dominated by those encoding LEA, seed storage and dormancy related proteins. On the other hand, a massive repression of genes belonging to many other classes such as photosynthesis, cell wall modification and energy metabolism occurs in parallel. Furthermore, comparison with a similar system for Medicago truncatula reveals a significant overlap between the two transcriptomes. Such overlap may highlight core mechanisms and key regulators of the trait DT. Taking into account the availability of the many genetic and molecular resources for Arabidopsis, the described system may prove useful for unraveling DT in higher plants. PMID:22195004

  10. Identification and classification of genes required for tolerance to freeze-thaw stress revealed by genome-wide screening of Saccharomyces cerevisiae deletion strains.

    PubMed

    Ando, Akira; Nakamura, Toshihide; Murata, Yoshinori; Takagi, Hiroshi; Shima, Jun

    2007-03-01

    Yeasts used in bread making are exposed to freeze-thaw stress during frozen-dough baking. To clarify the genes required for freeze-thaw tolerance, genome-wide screening was performed using the complete deletion strain collection of diploid Saccharomyces cerevisiae. The screening identified 58 gene deletions that conferred freeze-thaw sensitivity. These genes were then classified based on their cellular function and on the localization of their products. The results showed that the genes required for freeze-thaw tolerance were frequently involved in vacuole functions and cell wall biogenesis. The highest numbers of gene products were components of vacuolar H(+)-ATPase. Next, the cross-sensitivity of the freeze-thaw-sensitive mutants to oxidative stress and to cell wall stress was studied; both of these are environmental stresses closely related to freeze-thaw stress. The results showed that defects in the functions of vacuolar H(+)-ATPase conferred sensitivity to oxidative stress and to cell wall stress. In contrast, defects in gene products involved in cell wall assembly conferred sensitivity to cell wall stress but not to oxidative stress. Our results suggest the presence of at least two different mechanisms of freeze-thaw injury: oxidative stress generated during the freeze-thaw process, and defects in cell wall assembly.

  11. Vernalization Requirement and the Chromosomal VRN1-Region can Affect Freezing Tolerance and Expression of Cold-Regulated Genes in Festuca pratensis

    PubMed Central

    Ergon, Åshild; Melby, Tone I.; Höglind, Mats; Rognli, Odd A.

    2016-01-01

    Plants adapted to cold winters go through annual cycles of gain followed by loss of freezing tolerance (cold acclimation and deacclimation). Warm spells during winter and early spring can cause deacclimation, and if temperatures drop, freezing damage may occur. Many plants are vernalized during winter, a process making them competent to flower in the following summer. In winter cereals, a coincidence in the timing of vernalization saturation, deacclimation, downregulation of cold-induced genes, and reduced ability to reacclimate, occurs under long photoperiods and is under control of the main regulator of vernalization requirement in cereals, VRN1, and/or closely linked gene(s). Thus, the probability of freezing damage after a warm spell may depend on both vernalization saturation and photoperiod. We investigated the role of vernalization and the VRN1-region on freezing tolerance of meadow fescue (Festuca pratensis Huds.), a perennial grass species. Two F2 populations, divergently selected for high and low vernalization requirement, were studied. Each genotype was characterized for the copy number of one of the four parental haplotypes of the VRN1-region. Clonal plants were cold acclimated for 2 weeks or vernalized/cold acclimated for a total of 9 weeks, after which the F2 populations reached different levels of vernalization saturation. Vernalized and cold acclimated plants were deacclimated for 1 week and then reacclimated for 2 weeks. All treatments were given at 8 h photoperiod. Flowering response, freezing tolerance and expression of the cold-induced genes VRN1, MADS3, CBF6, COR14B, CR7 (BLT14), LOS2, and IRI1 was measured. We found that some genotypes can lose some freezing tolerance after vernalization and a deacclimation–reacclimation cycle. The relationship between vernalization and freezing tolerance was complex. We found effects of the VRN1-region on freezing tolerance in plants cold acclimated for 2 weeks, timing of heading after 9 weeks of

  12. The Miscanthus NAC transcription factor MlNAC9 enhances abiotic stress tolerance in transgenic Arabidopsis.

    PubMed

    Zhao, Xun; Yang, Xuanwen; Pei, Shengqiang; He, Guo; Wang, Xiaoyu; Tang, Qi; Jia, Chunlin; Lu, Ying; Hu, Ruibo; Zhou, Gongke

    2016-07-15

    NAC (NAM, ATAF1/2, and CUC2) transcription factors are known to play important roles in responses to abiotic stresses in plants. Currently, little information regarding the functional roles of NAC genes in stress tolerance is available in Miscanthus lutarioriparius, a promising bioenergy plant for cellulosic ethanol production. In this study, we carried out the functional characterization of MlNAC9 in abiotic stresses. MlNAC9 was shown to act as a nuclear localized transcription activator with the activation domain in its C-terminus. The overexpression of MlNAC9 in Arabidopsis conferred hypersensitivity to abscisic acid (ABA) at seed germination and root elongation stages. In addition, the overexpression of MlNAC9 led to increased seed germination rate and root growth under salt (NaCl) treatment. Meanwhile, the transgenic Arabidopsis overexpressing MlNAC9 showed enhanced tolerance to drought and cold stresses. The expression of stress-responsive marker genes was significantly increased in MlNAC9 overexpression lines compared to that of WT under ABA, drought, salt, and cold stresses. Correspondingly, the activities of antioxidant enzymes superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT) were significantly increased and the malondialdehyde (MDA) content was lower accumulated in MlNAC9 overexpression lines under drought and salt treatments. These results indicated that the overexpression of MlNAC9 improved the tolerance to abiotic stresses via an ABA-dependent pathway, and the enhanced tolerance of transgenic plants was mainly attributed to the increased expression of stress-responsive genes and the enhanced scavenging capability of reactive oxygen species (ROS).

  13. The Arabidopsis PLAT domain protein1 is critically involved in abiotic stress tolerance.

    PubMed

    Hyun, Tae Kyung; van der Graaff, Eric; Albacete, Alfonso; Eom, Seung Hee; Großkinsky, Dominik K; Böhm, Hannah; Janschek, Ursula; Rim, Yeonggil; Ali, Walid Wahid; Kim, Soo Young; Roitsch, Thomas

    2014-01-01

    Despite the completion of the Arabidopsis genome sequence, for only a relatively low percentage of the encoded proteins experimental evidence concerning their function is available. Plant proteins that harbour a single PLAT (Polycystin, Lipoxygenase, Alpha-toxin and Triacylglycerol lipase) domain and belong to the PLAT-plant-stress protein family are ubiquitously present in monocot and dicots. However, the function of PLAT-plant-stress proteins is still poorly understood. Therefore, we have assessed the function of the uncharacterised Arabidopsis PLAT-plant-stress family members through a combination of functional genetic and physiological approaches. PLAT1 overexpression conferred increased abiotic stress tolerance, including cold, drought and salt stress, while loss-of-function resulted in opposite effects on abiotic stress tolerance. Strikingly, PLAT1 promoted growth under non-stressed conditions. Abiotic stress treatments induced PLAT1 expression and caused expansion of its expression domain. The ABF/ABRE transcription factors, which are positive mediators of abscisic acid signalling, activate PLAT1 promoter activity in transactivation assays and directly bind to the ABRE elements located in this promoter in electrophoretic mobility shift assays. This suggests that PLAT1 represents a novel downstream target of the abscisic acid signalling pathway. Thus, we showed that PLAT1 critically functions as positive regulator of abiotic stress tolerance, but also is involved in regulating plant growth, and thereby assigned a function to this previously uncharacterised PLAT domain protein. The functional data obtained for PLAT1 support that PLAT-plant-stress proteins in general could be promising targets for improving abiotic stress tolerance without yield penalty.

  14. Reduced tolerance to abiotic stress in transgenic Arabidopsis overexpressing a Capsicum annuum multiprotein bridging factor 1

    PubMed Central

    2014-01-01

    Background The pepper fruit is the second most consumed vegetable worldwide. However, low temperature affects the vegetative development and reproduction of the pepper, resulting in economic losses. To identify cold-related genes regulated by abscisic acid (ABA) in pepper seedlings, cDNA representational difference analysis was previously performed using a suppression subtractive hybridization method. One of the genes cloned from the subtraction was homologous to Solanum tuberosum MBF1 (StMBF1) encoding the coactivator multiprotein bridging factor 1. Here, we have characterized this StMBF1 homolog (named CaMBF1) from Capsicum annuum and investigated its role in abiotic stress tolerance. Results Tissue expression profile analysis using quantitative RT-PCR showed that CaMBF1 was expressed in all tested tissues, and high-level expression was detected in the flowers and seeds. The expression of CaMBF1 in pepper seedlings was dramatically suppressed by exogenously supplied salicylic acid, high salt, osmotic and heavy metal stresses. Constitutive overexpression of CaMBF1 in Arabidopsis aggravated the visible symptoms of leaf damage and the electrolyte leakage of cell damage caused by cold stress in seedlings. Furthermore, the expression of RD29A, ERD15, KIN1, and RD22 in the transgenic plants was lower than that in the wild-type plants. On the other hand, seed germination, cotyledon greening and lateral root formation were more severely influenced by salt stress in transgenic lines compared with wild-type plants, indicating that CaMBF1-overexpressing Arabidopsis plants were hypersensitive to salt stress. Conclusions Overexpression of CaMBF1 in Arabidopsis displayed reduced tolerance to cold and high salt stress during seed germination and post-germination stages. CaMBF1 transgenic Arabidopsis may reduce stress tolerance by downregulating stress-responsive genes to aggravate the leaf damage caused by cold stress. CaMBF1 may be useful for genetic engineering of novel

  15. Constitutive overexpression of the calcium sensor CBL5 confers osmotic or drought stress tolerance in Arabidopsis.

    PubMed

    Cheong, Yong Hwa; Sung, Sun Jin; Kim, Beom-Gi; Pandey, Girdhar K; Cho, Ju-Sik; Kim, Kyung-Nam; Luan, Sheng

    2010-02-28

    Calcium serves as a critical messenger in many adaptation and developmental processes. Cellular calcium signals are detected and transmitted by sensor molecules such as calcium-binding proteins. In plants, the calcineurin B-like protein (CBL) family represents a unique group of calcium sensors and plays a key role in decoding calcium transients by specifically interacting with and regulating a family of CBL-interacting protein kinases (CIPKs). In this study, we report the role of Arabidopsis CBL5 gene in high salt or drought tolerance. CBL5 gene is expressed significantly in green tissues, but not in roots. CBL5 was not induced by abiotic stress conditions such as high salt, drought or low temperature. To determine whether the CBL5 gene plays a role in stress response pathways, we ectopically expressed the CBL5 protein in transgenic Arabidopsis plants (35S-CBL5) and examined plant responses to abiotic stresses. CBL5-overexpressing plants displayed enhanced tolerance to high salt or drought stress. CBL5 overexpression also rendered plants more resistant to high salt or hyperosmotic stress during early development (i.e., seed germination) but did not alter their response to abiscisic acid (ABA). Furthermore, overexpression of CBL5 alters the gene expression of stress gene markers, such as RD29A, RD29B and Kin1 etc. These results suggest that CBL5 may function as a positive regulator of salt or drought responses in plants.

  16. Expression of Vitis amurensis NAC26 in Arabidopsis enhances drought tolerance by modulating jasmonic acid synthesis

    PubMed Central

    Fang, Linchuan; Su, Lingye; Sun, Xiaoming; Li, Xinbo; Sun, Mengxiang; Karungo, Sospeter Karanja; Fang, Shuang; Chu, Jinfang; Li, Shaohua; Xin, Haiping

    2016-01-01

    The growth and fruit quality of grapevines are widely affected by abnormal climatic conditions such as water deficits, but many of the precise mechanisms by which grapevines respond to drought stress are still largely unknown. Here, we report that VaNAC26, a member of the NAC transcription factor family, was upregulated dramatically during cold, drought and salinity treatments in Vitis amurensis, a cold and drought-hardy wild Vitis species. Heterologous overexpression of VaNAC26 enhanced drought and salt tolerance in transgenic Arabidopsis. Higher activities of antioxidant enzymes and lower concentrations of H2O2 and O2 − were found in VaNAC26-OE lines than in wild type plants under drought stress. These results indicated that scavenging by reactive oxygen species (ROS) was enhanced by VaNAC26 in transgenic lines. Microarray-based transcriptome analysis revealed that genes related to jasmonic acid (JA) synthesis and signaling were upregulated in VaNAC26-OE lines under both normal and drought conditions. VaNAC26 showed a specific binding ability on the NAC recognition sequence (NACRS) motif, which broadly exists in the promoter regions of upregulated genes in transgenic lines. Endogenous JA content significantly increased in the VaNAC26-OE lines 2 and 3. Our data suggest that VaNAC26 responds to abiotic stresses and may enhance drought tolerance by transcriptional regulation of JA synthesis in Arabidopsis. PMID:27162276

  17. Musa paradisica RCI complements AtRCI and confers Na+ tolerance and K+ sensitivity in Arabidopsis.

    PubMed

    Liu, Bing; Feng, Dongru; Zhang, Bipei; Mu, Peiqiang; Zhang, Yang; He, Yanming; Qi, Kangbiao; Wang, Jinfa; Wang, Hongbin

    2012-03-01

    The mechanisms involved in Na⁺/K⁺ uptake and extrusion are important in plant salt tolerance. In this study, we investigated the physiological role of a plasma membrane (PM)-localized protein, MpRCI, from plantain in transgenic Arabidopsis under NaCl and KCl stress and determined its effect on PM fluidity and H⁺-ATPase activity. The MpRCI gene exhibited high homology to the AtRCI2 gene family in Arabidopsis and was therefore able to complement for loss of the yeast AtRCI2-related PMP3 gene. Results of phenotypic espial and atomic emission spectrophotometer (AES) assays indicated that MpRCI overexpression in the AtRCI2A knockout mutant with reduced shoot Na⁺ and increased K⁺ exhibited increased Na⁺-tolerance and K⁺-sensitivity under NaCl or KCl treatments, respectively. Furthermore, comparisons of PM fluidity and H⁺-ATPase activity in shoots, with expression or absence of MpRCI/AtRCI2A expression under NaCl or KCl stress, showed MpRCI maintained PM fluidity and H⁺-ATPase activity under stress conditions. Results suggest that MpRCI plays an essential role in Na⁺/K⁺ flux in plant cells.

  18. Ectopic expression of a tobacco vacuolar invertase inhibitor in guard cells confers drought tolerance in Arabidopsis.

    PubMed

    Chen, Su-Fen; Liang, Ke; Yin, Dong-Mei; Ni, Di-An; Zhang, Zhi-Guo; Ruan, Yong-Ling

    2016-12-01

    There are several hypotheses that explain stomatal behavior. These include the concept of osmoregulation mediated by potassium and its counterions malate and chlorine and the more recent starch-sugar hypothesis. We have previously reported that the activity of the sucrose cleavage enzyme, vacuolar invertase (VIN), is significantly higher in guard cells than in other leaf epidermal cells and its activity is correlated with stomatal aperture. Here, we examined whether VIN indeed controls stomatal movement under normal and drought conditions by transforming Arabidopsis with a tobacco vacuolar invertase inhibitor homolog (Nt-inhh) under the control of an abscisic acid-sensitive and guard cell-specific promoter (AtRab18). The data obtained showed that guard cells of transgenic Arabidopsis plants had lower VIN activity, stomatal aperture and conductance than that of wild-type plants. Moreover, the transgenic plants also displayed higher drought tolerance than wild-type plants. The data indicate that VIN is a promising target for manipulating stomatal function to increase drought tolerance.

  19. Drought tolerance in Arabidopsis is controlled by the OCP3 disease resistance regulator.

    PubMed

    Ramírez, Vicente; Coego, Alberto; López, Ana; Agorio, Astrid; Flors, Víctor; Vera, Pablo

    2009-05-01

    Water scarcity and corresponding abiotic drought stress is one of the most important factors limiting plant performance and yield. In addition, plant productivity is severely compromised worldwide by infection with microbial pathogens. Two of the most prominent pathways responsible for drought tolerance and disease resistance to fungal pathogens in Arabidopsis are those controlled by the phytohormones abscisic acid (ABA) and the oxylipin methyl jasmonate (MeJA), respectively. Here, we report on the functional characterization of OCP3, a transcriptional regulator from the homeodomain (HD) family. The Arabidopsis loss-of-function ocp3 mutant exhibits both drought resistance and enhanced disease resistance to necrotrophic fungal pathogens. Double-mutant analysis revealed that these two resistance phenotypes have different genetic requirements. Whereas drought tolerance in ocp3 is ABA-dependent but MeJA-independent, the opposite holds true for the enhanced disease resistance characteristics. These observations lead us to propose a regulatory role of OCP3 in the adaptive responses to these two stresses, functioning as a modulator of independent and specific aspects of the ABA- and MeJA-mediated signal transduction pathways.

  20. Heterologous Overexpression of Poplar SnRK2 Genes Enhanced Salt Stress Tolerance in Arabidopsis thaliana

    PubMed Central

    Song, Xueqing; Yu, Xiang; Hori, Chiaki; Demura, Taku; Ohtani, Misato; Zhuge, Qiang

    2016-01-01

    Subfamily 2 of SNF1-related protein kinase (SnRK2) plays important roles in plant abiotic stress responses as a global positive regulator of abscisic acid signaling. In the genome of the model tree Populus trichocarpa, 12 SnRK2 genes have been identified, and some are upregulated by abiotic stresses. In this study, we heterologously overexpressed the PtSnRK2 genes in Arabidopsis thaliana and found that overexpression of PtSnRK2.5 and PtSnRK2.7 genes enhanced stress tolerance. In the PtSnRK2.5 and PtSnRK2.7 overexpressors, chlorophyll content, and root elongation were maintained under salt stress conditions, leading to higher survival rates under salt stress compared with those in the wild type. Transcriptomic analysis revealed that PtSnRK2.7 overexpression affected stress-related metabolic genes, including lipid metabolism and flavonoid metabolism, even under normal growth conditions. However, the stress response genes reported to be upregulated in Arabidopsis SRK2C/SnRK2.6 and wheat SnRK2.8 overexpressors were not changed by PtSnRK2.7 overexpression. Furthermore, PtSnRK2.7 overexpression widely and largely influenced the transcriptome in response to salt stress; genes related to transport activity, including anion transport-related genes, were characteristically upregulated, and a variety of metabolic genes were specifically downregulated. We also found that the salt stress response genes were greatly upregulated in the PtSnRK2.7 overexpressor. Taken together, poplar subclass 2 PtSnRK2 genes can modulate salt stress tolerance in Arabidopsis, through the activation of cellular signaling pathways in a different manner from that by herbal subclass 2 SnRK2 genes. PMID:27242819

  1. Lipid profiles of detergent resistant fractions of the plasma membrane in oat and rye in association with cold acclimation and freezing tolerance

    PubMed Central

    Takahashi, Daisuke; Imai, Hiroyuki; Kawamura, Yukio; Uemura, Matsuo

    2016-01-01

    Cold acclimation (CA) results in alteration of the plasma membrane (PM) lipid composition in plants, which plays a crucial role in the acquisition of freezing tolerance via membrane stabilization. Recent studies have indicated that PM structure is consistent with the fluid mosaic model but is laterally non-homogenous and contains microdomains enriched in sterols, sphingolipids and specific proteins. In plant cells, the function of these microdomains in relation to CA and freezing tolerance is not yet fully understood. The present study aimed to investigate the lipid compositions of detergent resistant fractions of the PM (DRM) which are considered to represent microdomains. They were prepared from leaves of low-freezing tolerant oat and high-freezing tolerant rye. The DRMs contained higher proportions of sterols, sphingolipids and saturated phospholipids than the PM. In particular, one of the sterol lipid classes, acylated sterylglycoside, was the predominant sterol in oat DRM while rye DRM contained free sterol as the major sterol. Oat and rye showed different patterns (or changes) of sterols and 2-hydroxy fatty acids of sphingolipids of DRM lipids during CA. Taken together, these results suggest that CA-induced changes of lipid classes and molecular species in DRMs are associated with changes in the thermodynamic properties and physiological functions of microdomains during CA and hence, influence plant freezing tolerance. PMID:26904981

  2. Lipid profiles of detergent resistant fractions of the plasma membrane in oat and rye in association with cold acclimation and freezing tolerance.

    PubMed

    Takahashi, Daisuke; Imai, Hiroyuki; Kawamura, Yukio; Uemura, Matsuo

    2016-04-01

    Cold acclimation (CA) results in alteration of the plasma membrane (PM) lipid composition in plants, which plays a crucial role in the acquisition of freezing tolerance via membrane stabilization. Recent studies have indicated that PM structure is consistent with the fluid mosaic model but is laterally non-homogenous and contains microdomains enriched in sterols, sphingolipids and specific proteins. In plant cells, the function of these microdomains in relation to CA and freezing tolerance is not yet fully understood. The present study aimed to investigate the lipid compositions of detergent resistant fractions of the PM (DRM) which are considered to represent microdomains. They were prepared from leaves of low-freezing tolerant oat and high-freezing tolerant rye. The DRMs contained higher proportions of sterols, sphingolipids and saturated phospholipids than the PM. In particular, one of the sterol lipid classes, acylated sterylglycoside, was the predominant sterol in oat DRM while rye DRM contained free sterol as the major sterol. Oat and rye showed different patterns (or changes) of sterols and 2-hydroxy fatty acids of sphingolipids of DRM lipids during CA. Taken together, these results suggest that CA-induced changes of lipid classes and molecular species in DRMs are associated with changes in the thermodynamic properties and physiological functions of microdomains during CA and hence, influence plant freezing tolerance. Copyright © 2016 Elsevier Inc. All rights reserved.

  3. Soybean Salt Tolerance 1 (GmST1) Reduces ROS Production, Enhances ABA Sensitivity, and Abiotic Stress Tolerance in Arabidopsis thaliana

    PubMed Central

    Ren, Shuxin; Lyle, Chimera; Jiang, Guo-liang; Penumala, Abhishek

    2016-01-01

    Abiotic stresses, including high soil salinity, significantly reduce crop production worldwide. Salt tolerance in plants is a complex trait and is regulated by multiple mechanisms. Understanding the mechanisms and dissecting the components on their regulatory pathways will provide new insights, leading to novel strategies for the improvement of salt tolerance in agricultural and economic crops of importance. Here we report that soybean salt tolerance 1, named GmST1, exhibited strong tolerance to salt stress in the Arabidopsis transgenic lines. The GmST1-overexpressed Arabidopsis also increased sensitivity to ABA and decreased production of reactive oxygen species under salt stress. In addition, GmST1 significantly improved drought tolerance in Arabidopsis transgenic lines. GmST1 belongs to a 3-prime part of Glyma.03g171600 gene in the current version of soybean genome sequence annotation. However, comparative reverse transcription-polymerase chain reaction analysis around Glyma.03g171600 genomic region confirmed that GmST1 might serve as an intact gene in soybean leaf tissues. Unlike Glyma.03g171600 which was not expressed in leaves, GmST1 was strongly induced by salt treatment in the leaf tissues. By promoter analysis, a TATA box was detected to be positioned close to GmST1 start codon and a putative ABRE and a DRE cis-acting elements were identified at about 1 kb upstream of GmST1 gene. The data also indicated that GmST1-transgenic lines survived under drought stress and showed a significantly lower water loss than non-transgenic lines. In summary, our results suggest that overexpression of GmST1 significantly improves Arabidopsis tolerance to both salt and drought stresses and the gene may be a potential candidate for genetic engineering of salt- and drought-tolerant crops. PMID:27148284

  4. Interact to Survive: Phyllobacterium brassicacearum Improves Arabidopsis Tolerance to Severe Water Deficit and Growth Recovery

    PubMed Central

    Bresson, Justine; Vasseur, François; Dauzat, Myriam; Labadie, Marc; Varoquaux, Fabrice; Touraine, Bruno; Vile, Denis

    2014-01-01

    Mutualistic bacteria can alter plant phenotypes and confer new abilities to plants. Some plant growth-promoting rhizobacteria (PGPR) are known to improve both plant growth and tolerance to multiple stresses, including drought, but reports on their effects on plant survival under severe water deficits are scarce. We investigated the effect of Phyllobacterium brassicacearum STM196 strain, a PGPR isolated from the rhizosphere of oilseed rape, on survival, growth and physiological responses of Arabidopsis thaliana to severe water deficits combining destructive and non-destructive high-throughput phenotyping. Soil inoculation with STM196 greatly increased the survival rate of A. thaliana under several scenarios of severe water deficit. Photosystem II efficiency, assessed at the whole-plant level by high-throughput fluorescence imaging (Fv/Fm), was related to the probability of survival and revealed that STM196 delayed plant mortality. Inoculated surviving plants tolerated more damages to the photosynthetic tissues through a delayed dehydration and a better tolerance to low water status. Importantly, STM196 allowed a better recovery of plant growth after rewatering and stressed plants reached a similar biomass at flowering than non-stressed plants. Our results highlight the importance of plant-bacteria interactions in plant responses to severe drought and provide a new avenue of investigations to improve drought tolerance in agriculture. PMID:25226036

  5. Improved growth and stress tolerance in the Arabidopsis oxt1 mutant triggered by altered adenine metabolism.

    PubMed

    Sukrong, Suchada; Yun, Kil-Young; Stadler, Patrizia; Kumar, Charan; Facciuolo, Tony; Moffatt, Barbara A; Falcone, Deane L

    2012-11-01

    Plants perceive and respond to environmental stresses with complex mechanisms that are often associated with the activation of antioxidant defenses. A genetic screen aimed at isolating oxidative stress-tolerant lines of Arabidopsis thaliana has identified oxt1, a line that exhibits improved tolerance to oxidative stress and elevated temperature but displays no apparent deleterious growth effects under non-stress conditions. Oxt1 harbors a mutation that arises from the altered expression of a gene encoding adenine phosphoribosyltransferase (APT1), an enzyme that converts adenine to adenosine monophosphate (AMP), indicating a link between purine metabolism, whole-plant growth responses, and stress acclimation. The oxt1 mutation results in decreased APT1 expression that leads to reduced enzymatic activity. Correspondingly, oxt1 plants possess elevated levels of adenine. Decreased APT enzyme activity directly correlates with stress resistance in transgenic lines that ectopically express APT1. The metabolic alteration in oxt1 plants also alters the expression of several antioxidant defense genes and the response of these genes to oxidative challenge. Finally, it is shown that manipulation of adenine levels can induce stress tolerance to wild-type plants. Collectively, these results show that alterations in cellular adenine levels can trigger stress tolerance and improve growth, leading to increases in plant biomass. The results also suggest that adenine might play a part in the signals that modulate responses to abiotic stress and plant growth.

  6. Ky-2, a Histone Deacetylase Inhibitor, Enhances High-Salinity Stress Tolerance in Arabidopsis thaliana.

    PubMed

    Sako, Kaori; Kim, Jong-Myong; Matsui, Akihiro; Nakamura, Kotaro; Tanaka, Maho; Kobayashi, Makoto; Saito, Kazuki; Nishino, Norikazu; Kusano, Miyako; Taji, Teruaki; Yoshida, Minoru; Seki, Motoaki

    2016-04-01

    Adaptation to environmental stress requires genome-wide changes in gene expression. Histone modifications are involved in gene regulation, but the role of histone modifications under environmental stress is not well understood. To reveal the relationship between histone modification and environmental stress, we assessed the effects of inhibitors of histone modification enzymes during salinity stress. Treatment with Ky-2, a histone deacetylase inhibitor, enhanced high-salinity stress tolerance in Arabidopsis. We confirmed that Ky-2 possessed inhibition activity towards histone deacetylases by immunoblot analysis. To investigate how Ky-2 improved salt stress tolerance, we performed transcriptome and metabolome analysis. These data showed that the expression of salt-responsive genes and salt stress-related metabolites were increased by Ky-2 treatment under salinity stress. A mutant deficient in AtSOS1(Arabidopis thaliana SALT OVERLY SENSITIVE 1), which encodes an Na(+)/H(+)antiporter and was among the up-regulated genes, lost the salinity stress tolerance conferred by Ky-2. We confirmed that acetylation of histone H4 at AtSOS1 was increased by Ky-2 treatment. Moreover, Ky-2 treatment decreased the intracellular Na(+)accumulation under salinity stress, suggesting that enhancement of SOS1-dependent Na(+)efflux contributes to increased high-salinity stress tolerance caused by Ky-2 treatment.

  7. Transgenic poplar expressing Arabidopsis NDPK2 enhances growth as well as oxidative stress tolerance.

    PubMed

    Kim, Yun-Hee; Kim, Myoung Duck; Choi, Young Im; Park, Sung-Chul; Yun, Dae-Jin; Noh, Eun Woon; Lee, Haeng-Soon; Kwak, Sang-Soo

    2011-04-01

    Nucleoside diphosphate kinase 2 (NDPK2) is known to regulate the expression of antioxidant genes in plants. Previously, we reported that overexpression of Arabidopsis NDPK2 (AtNDPK2) under the control of an oxidative stress-inducible SWPA2 promoter in transgenic potato and sweetpotato plants enhanced tolerance to various abiotic stresses. In this study, transgenic poplar (Populus alba × Poplus glandulosa) expressing the AtNDPK2 gene under the control of a SWPA2 promoter (referred to as SN) was generated to develop plants with enhanced tolerance to oxidative stress. The level of AtNDPK2 expression and NDPK activity in SN plants following methyl viologen (MV) treatment was positively correlated with the plant's tolerance to MV-mediated oxidative stress. We also observed that antioxidant enzyme activities such as ascorbate peroxidase, catalase and peroxidase were increased in MV-treated leaf discs of SN plants. The growth of SN plants was substantially increased under field conditions including increased branch number and stem diameter. SN plants exhibited higher transcript levels of the auxin-response genes IAA2 and IAA5. These results suggest that enhanced AtNDPK2 expression affects oxidative stress tolerance leading to improved plant growth in transgenic poplar. © 2010 The Authors. Plant Biotechnology Journal © 2010 Society for Experimental Biology, Association of Applied Biologists and Blackwell Publishing Ltd.

  8. Elevated atmospheric CO2 and strain of rhizobium alter freezing tolerance and cold-induced molecular changes in alfalfa (Medicago sativa).

    PubMed

    Bertrand, Annick; Prévost, Danielle; Bigras, Francine J; Castonguay, Yves

    2007-02-01

    The objective of the study was to assess the impact of elevated CO2 in interaction with rhizobial strains on freezing tolerance and cold-induced molecular changes in alfalfa. Alfalfa inoculated with two different strains of rhizobium (A2 and NRG34) was grown and cold acclimated (2 weeks at 2 degrees C) under either 400 (ambient) or 800 micromol mol(-1) (elevated) CO2. Plants acclimated under 400 micromol mol(-1) CO2 were more freezing tolerant than those maintained under 800 micromol mol(-1). Cryoprotective sugars typically linked with the acquisition of freezing tolerance such as sucrose, stachyose and raffinose increased in roots in response to low temperature but did not differ between CO2 treatments. Similarly high CO2 did not alter the expression of many cold-regulated (COR) genes although it significantly increased the level of transcripts encoding a COR gene homologous to glyceraldehyde-3-phosphate-dehydrogenase (GAPDH). A significant effect of rhizobial strain was observed on both freezing tolerance and gene expression. Plants of alfalfa inoculated with strain A2 were more freezing tolerant than those inoculated with strain NRG34. Transcripts of COR genes homologous to a pathogenesis-related protein (PR-10) and to a nuclear-targeted protein were markedly enhanced in roots of alfalfa inoculated with strain A2 as compared with strain NRG34. Transcripts encoding the vegetative storage proteins (VSPs) beta-amylase and chitinase were more abundant in roots of non-acclimated plants inoculated with strain NRG34 than with strain A2. Taken together, the results suggest that elevated CO2 stimulates plant growth and reduces freezing tolerance. The acquisition of cold tolerance is also influenced by the rhizobial strain, as indicated by lower levels of expression of COR genes and sustained accumulation of VSP-encoding transcripts in alfalfa inoculated with strain NRG34 as compared with strain A2.

  9. Abscisic acid-induced rearrangement of intracellular structures associated with freezing and desiccation stress tolerance in the liverwort Marchantia polymorpha.

    PubMed

    Akter, Khaleda; Kato, Masahiro; Sato, Yuki; Kaneko, Yasuko; Takezawa, Daisuke

    2014-09-15

    The plant growth regulator abscisic acid (ABA) is known to be involved in triggering responses to various environmental stresses such as freezing and desiccation in angiosperms, but little is known about its role in basal land plants, especially in liverworts, representing the earliest land plant lineage. We show here that survival rate after freezing and desiccation of Marchantia polymorpha gemmalings was increased by pretreatment with ABA in the presence of increasing concentrations of sucrose. ABA treatment increased accumulation of soluble sugars in gemmalings, and sugar accumulation was further increased by addition of sucrose to the culture medium. ABA treatment of gemmalings also induced accumulation of transcripts for proteins with similarity to late embryogenesis abundant (LEA) proteins, which accumulate in association with acquisition of desiccation tolerance in maturing seeds. Observation by light and electron microscopy indicated that the ABA treatment caused fragmentation of vacuoles with increased cytosolic volume, which was more prominent in the presence of a high concentration of external sucrose. ABA treatment also increased the density of chloroplast distribution and remarkably enlarged their volume. These results demonstrate that ABA induces drastic physiological changes in liverwort cells for stress tolerance, accompanied by accumulation of protectants against dehydration and rearrangement and morphological alterations of cellular organelles. Copyright © 2014 Elsevier GmbH. All rights reserved.

  10. Anti-apoptotic response during anoxia and recovery in a freeze-tolerant wood frog (Rana sylvatica)

    PubMed Central

    Gerber, Victoria E.M.; Wijenayake, Sanoji

    2016-01-01

    The common wood frog, Rana sylvatica, utilizes freeze tolerance as a means of winter survival. Concealed beneath a layer of leaf litter and blanketed by snow, these frogs withstand subzero temperatures by allowing approximately 65–70% of total body water to freeze. Freezing is generally considered to be an ischemic event in which the blood oxygen supply is impeded and may lead to low levels of ATP production and exposure to oxidative stress. Therefore, it is as important to selectively upregulate cytoprotective mechanisms such as the heat shock protein (HSP) response and expression of antioxidants as it is to shut down majority of ATP consuming processes in the cell. The objective of this study was to investigate another probable cytoprotective mechanism, anti-apoptosis during oxygen deprivation and recovery in the anoxia tolerant wood frog. In particular, relative protein expression levels of two important apoptotic regulator proteins, Bax and p-p53 (S46), and five anti-apoptotic/pro-survival proteins, Bcl-2, p-Bcl-2 (S70), Bcl-xL, x-IAP, and c-IAP in response to normoxic, 24 Hr anoxic exposure, and 4 Hr recovery stages were assessed in the liver and skeletal muscle using western immunoblotting. The results suggest a tissue-specific regulation of the anti-apoptotic pathway in the wood frog, where both liver and skeletal muscle shows an overall decrease in apoptosis and an increase in cell survival. This type of cytoprotective mechanism could be aimed at preserving the existing cellular components during long-term anoxia and oxygen recovery phases in the wood frog. PMID:27042393

  11. Isolation and characterization of low-sulphur-tolerant mutants of Arabidopsis

    PubMed Central

    Wu, Yu; Zhao, Qing; Gao, Lei; Yu, Xiao-Min; Fang, Ping; Oliver, David J.; Xiang, Cheng-Bin

    2010-01-01

    Sulphur is an essential element for plant growth and development as well as for defence against biotic and abiotic stresses. Increasing sulphate utilization efficiency (SUE) is an important issue for crop improvement. Little is known about the genetic determinants of sulphate utilization efficiency. No gain-of-function mutants with improved SUE have been reported to date. Here the isolation and characterization of two low-sulphur-tolerant mutants, sue3 and sue4 are reported using a high-throughput genetic screen where a ‘sulphur-free’ solid medium was devised to give the selection pressure necessary to suppress the growth of the wild-type seedlings. Both mutants showed improved tolerance to low sulphur conditions and well-developed root systems. The mutant phenotype of both sue3 and sue4 was specific to sulphate deficiency and the mutants displayed enhanced tolerance to heavy metal and oxidative stress. Genetic analysis revealed that sue3 was caused by a single recessive nuclear mutation while sue4 was caused by a single dominant nuclear mutation. The recessive locus in sue3 is the previously identified VirE2-interacting Protein 1. The dominant locus in sue4 is a function-unknown locus activated by the four enhancers on the T-DNA. The function of SUE3 and SUE4 in low sulphur tolerance was confirmed either by multiple mutant alleles or by recapitulation analysis. Taken together, our results demonstrate that this genetic screen is a reasonable approach to isolate Arabidopsis mutants with improved low sulphur tolerance and potentially with enhanced sulphate utilization efficiency. The two loci identified in sue3 and sue4 should assist in understanding the molecular mechanisms of low sulphur tolerance. PMID:20547563

  12. Transcriptome profiling of genes and pathways associated with arsenic toxicity and tolerance in Arabidopsis

    PubMed Central

    2014-01-01

    Background Arsenic (As) is a toxic metalloid found ubiquitously in the environment and widely considered an acute poison and carcinogen. However, the molecular mechanisms of the plant response to As and ensuing tolerance have not been extensively characterized. Here, we report on transcriptional changes with As treatment in two Arabidopsis accessions, Col-0 and Ws-2. Results The root elongation rate was greater for Col-0 than Ws-2 with As exposure. Accumulation of As was lower in the more tolerant accession Col-0 than in Ws-2. We compared the effect of As exposure on genome-wide gene expression in the two accessions by comparative microarray assay. The genes related to heat response and oxidative stresses were common to both accessions, which indicates conserved As stress-associated responses for the two accessions. Most of the specific response genes encoded heat shock proteins, heat shock factors, ubiquitin and aquaporin transporters. Genes coding for ethylene-signalling components were enriched in As-tolerant Col-0 with As exposure. A tolerance-associated gene candidate encoding Leucine-Rich Repeat receptor-like kinase VIII (LRR-RLK VIII) was selected for functional characterization. Genetic loss-of-function analysis of the LRR-RLK VIII gene revealed altered As sensitivity and the metal accumulation in roots. Conclusions Thus, ethylene-related pathways, maintenance of protein structure and LRR-RLK VIII-mediated signalling may be important mechanisms for toxicity and tolerance to As in the species. Here, we provide a comprehensive survey of global transcriptional regulation for As and identify stress- and tolerance-associated genes responding to As. PMID:24734953

  13. Isolation and characterization of low-sulphur-tolerant mutants of Arabidopsis.

    PubMed

    Wu, Yu; Zhao, Qing; Gao, Lei; Yu, Xiao-Min; Fang, Ping; Oliver, David J; Xiang, Cheng-Bin

    2010-07-01

    Sulphur is an essential element for plant growth and development as well as for defence against biotic and abiotic stresses. Increasing sulphate utilization efficiency (SUE) is an important issue for crop improvement. Little is known about the genetic determinants of sulphate utilization efficiency. No gain-of-function mutants with improved SUE have been reported to date. Here the isolation and characterization of two low-sulphur-tolerant mutants, sue3 and sue4 are reported using a high-throughput genetic screen where a 'sulphur-free' solid medium was devised to give the selection pressure necessary to suppress the growth of the wild-type seedlings. Both mutants showed improved tolerance to low sulphur conditions and well-developed root systems. The mutant phenotype of both sue3 and sue4 was specific to sulphate deficiency and the mutants displayed enhanced tolerance to heavy metal and oxidative stress. Genetic analysis revealed that sue3 was caused by a single recessive nuclear mutation while sue4 was caused by a single dominant nuclear mutation. The recessive locus in sue3 is the previously identified VirE2-interacting Protein 1. The dominant locus in sue4 is a function-unknown locus activated by the four enhancers on the T-DNA. The function of SUE3 and SUE4 in low sulphur tolerance was confirmed either by multiple mutant alleles or by recapitulation analysis. Taken together, our results demonstrate that this genetic screen is a reasonable approach to isolate Arabidopsis mutants with improved low sulphur tolerance and potentially with enhanced sulphate utilization efficiency. The two loci identified in sue3 and sue4 should assist in understanding the molecular mechanisms of low sulphur tolerance.

  14. Exploring ammonium tolerance in a large panel of Arabidopsis thaliana natural accessions

    PubMed Central

    Sarasketa, Asier; González-Moro, María Begoña; González-Murua, Carmen; Marino, Daniel

    2014-01-01

    Plants are dependent on exogenous nitrogen (N) supply. Ammonium (NH4 +), together with nitrate (NO3 –), is one of the main nitrogenous compounds available in the soil. Paradoxically, although NH4 + assimilation requires less energy than that of NO3 –, many plants display toxicity symptoms when grown with NH4 + as the sole N source. However, in addition to species-specific ammonium toxicity, intraspecific variability has also been shown. Thus, the aim of this work was to study the intraspecific ammonium tolerance in a large panel of Arabidopsis thaliana natural accessions. Plants were grown with either 1mM NO3 – or NH4 + as the N source, and several parameters related to ammonium tolerance and assimilation were determined. Overall, high variability was observed in A. thaliana shoot growth under both forms of N nutrition. From the parameters determined, tissue ammonium content was the one with the highest impact on shoot biomass, and interestingly this was also the case when N was supplied as NO3 –. Enzymes of nitrogen assimilation did not have an impact on A. thaliana biomass variation, but the N source affected their activity. Glutamate dehydrogenase (GDH) aminating activity was, in general, higher in NH4 +-fed plants. In contrast, GDH deaminating activity was higher in NO3 –-fed plants, suggesting a differential role for this enzyme as a function of the N form supplied. Overall, NH4 + accumulation seems to be an important player in Arabidopsis natural variability in ammonium tolerance rather than the cell NH4 + assimilation capacity. PMID:25205573

  15. Expression of an Arabidopsis sodium/proton antiporter gene (AtNHX1)in peanut to improve salt tolerance

    USDA-ARS?s Scientific Manuscript database

    Salinity is a major environmental stress that affects agricultural productivity worldwide. One approach to improving salt tolerance in crops is through high expression of the Arabidopsis gene AtNHX1, which encodes a vacuolar sodium/proton antiporter that sequesters excess sodium ion into the large i...

  16. Construction from a single parent of baker's yeast strains with high freeze tolerance and fermentative activity in both lean and sweet doughs.

    PubMed

    Nakagawa, S; Ouchi, K

    1994-10-01

    From a freeze-tolerant baker's yeast (Saccharomyces cerevisiae), 2,333 spore clones were obtained. To improve the leavening ability in lean dough of the parent strain, we selected 555 of the high-maltose-fermentative spore clones by using a method in which a soft agar solution containing maltose and bromocresol purple was overlaid on yeast colonies. By measuring the gassing power in the dough, we selected 66 spore clones with a good leavening ability in lean dough and a total of 694 hybrids were constructed by crossing them. Among these hybrids, we obtained 50 novel freeze-tolerant strains with good leavening ability in all lean, regular, and sweet doughs comparable to that of commercial baker's yeast. Hybrids with improved leavening ability or freeze tolerance compared with the parent yeast and commercial baker's yeasts were also obtained. These results suggest that hybridization between spore clones derived from a single parent strain is effective for improving the properties of baker's yeasts.

  17. Deficiency in the glycerol channel Fps1p confers increased freeze tolerance to yeast cells: application of the fps1delta mutant to frozen dough technology.

    PubMed

    Izawa, Shingo; Ikeda, Kayo; Maeta, Kazuhiro; Inoue, Yoshiharu

    2004-12-01

    Intracellular glycerol content affects the freeze-thaw stress tolerance of Saccharomyces cerevisiae. We have recently reported that intracellular-glycerol-enriched cells cultured in glycerol medium acquire tolerance to freeze stress and retain high leavening ability even in dough after frozen storage [Izawa et al. (2004) Appl Microbiol Biotechnol http://dx.doi.org/10.1007/s00253-004-1624-4]. A deletion mutant of the FPS1 gene, which encodes a glycerol channel, accumulates glycerol inside the cell without an exogenous supply of glycerol into the medium. We found that the fps1delta cells acquired tolerance to freeze stress and retained high leavening ability in dough after frozen storage for 7 days. These results suggest that the fps1delta mutant is a useful strain for developing better frozen-dough with a commercial advantage.

  18. Tolerance of brown bear spermatozoa to conditions of pre-freezing cooling rate and equilibration time.

    PubMed

    López-Urueña, E; Alvarez, M; Gomes-Alves, S; Martínez-Rodríguez, C; Borragan, S; Anel-López, L; de Paz, P; Anel, L

    2014-06-01

    Specific protocols for the cryopreservation of endangered Cantabrian brown bear spermatozoa are critical to create a genetic resource bank. The aim of this study was to assess the effect of cooling rates and equilibration time before freezing on post-thawed brown bear spermatozoa quality. Electroejaculates from 11 mature bears were extended to 100 × 10(6) spermatozoa/mL in a TES-Tris-Fructose-based extender, cryopreserved following performance of the respective cooling/equilibration protocol each sample was assigned to, and stored at -196 °C for further assessment. Before freezing, after thawing, and after 1 hour's incubation post-thawing at 37 °C (thermal stress test), the quality of the samples was assessed for motility by computer-assisted semen analysis, and for viability (SYBR-14/propidium iodide), acrosomal status (peanut agglutinin-fluorescein isothiocyanate /propidium iodide), and sperm chromatin stability (SCSA) by flow cytometry. In experiment 1, three cooling rates (0.25 °C/min, 1 °C/min, and 4 °C/min) to 5 °C were assessed. After thawing, total motility (%TM) was higher and percentage of damaged acrosomes (%dACR) was lower (P < 0.05) for 0.25 °C/min than for 4 °C/min. The thermal stress test data indicated equally poor quality (P < 0.05) for the 4 °C/min cooled samples in viability (%VIAB), %dACR, %TM, and progressive motility (%PM). In experiment 2, the effect of a pre-freezing equilibration period at 5 °C for 1 hour (cooling at 0.25 °C/min) was evaluated. Samples kept at 5 °C for 1 hour showed higher (P < 0.05) values than the nonequilibrated ones for both thawing (%dACR) and thermal stress test (%VIAB, %TM, and %PM). In experiment 3, samples stored without cooling and equilibration (direct freezing) were compared with the samples cooled at 0.25 °C/min and equilibrated for 1 hour (control freezing). Using thermal stress test, we observed that direct freezing causes damage in viability, acrosomal status, and motility of spermatozoa

  19. Is the OJIP Test a Reliable Indicator of Winter Hardiness and Freezing Tolerance of Common Wheat and Triticale under Variable Winter Environments?

    PubMed Central

    Rapacz, Marcin; Sasal, Monika; Kalaji, Hazem M.; Kościelniak, Janusz

    2015-01-01

    OJIP analysis, which explores changes in photosystem II (PSII) photochemical performance, has been used as a measure of plant susceptibility to stress. However, in the case of freezing tolerance and winter hardiness, which are highly environmentally variable, the use of this method can give ambiguous results depending on the species as well as the sampling year and time. To clarify this issue, we performed chlorophyll fluorescence measurements over three subsequent winters (2010/11, 2011/12 and 2012/13) on 220 accessions of common winter wheat and 139 accessions of winter triticale. After freezing, leaves were collected from cold-acclimated plants in the laboratory and field-grown plants. Observations of field survival in seven locations across Poland and measurements of freezing tolerance of the studied plants were also recorded. Our results confirm that the OJIP test is a reliable indicator of winter hardiness and freezing tolerance of common wheat and triticale under unstable winter environments. Regardless of species, the testing conditions giving the most reliable results were identical, and the reliability of the test could be easily checked by analysis of some relationships between OJIP-test parameters. We also found that triticale is more winter hardy and freezing tolerant than wheat. In addition, the two species were characterized by different patterns of photosynthetic apparatus acclimation to cold. PMID:26230839

  20. Co-Expression of Monodehydroascorbate Reductase and Dehydroascorbate Reductase from Brassica rapa Effectively Confers Tolerance to Freezing-Induced Oxidative Stress

    PubMed Central

    Shin, Sun-Young; Kim, Myung-Hee; Kim, Yul-Ho; Park, Hyang-Mi; Yoon, Ho-Sung

    2013-01-01

    Plants are exposed to various environmental stresses and have therefore developed antioxidant enzymes and molecules to protect their cellular components against toxicity derived from reactive oxygen species (ROS). Ascorbate is a very important antioxidant molecule in plants, and monodehydroascorbate reductase (MDHAR; EC 1.6.5.4) and dehydroascorbate reductase (DHAR; EC 1.8.5.1) are essential to regeneration of ascorbate for maintenance of ROS scavenging ability. The MDHAR and DHAR genes from Brassica rapa were cloned, transgenic plants overexpressing either BrMDHAR and BrDHAR were established, and then, each transgenic plant was hybridized to examine the effects of co-expression of both genes conferring tolerance to freezing. Transgenic plants co-overexpressing BrMDHAR and BrDHAR showed activated expression of relative antioxidant enzymes, and enhanced levels of glutathione and phenolics under freezing condition. Then, these alteration caused by co-expression led to alleviated redox status and lipid peroxidation and consequently conferred improved tolerance against severe freezing stress compared to transgenic plants overexpressing single gene. The results of this study suggested that although each expression of BrMDHAR or BrDHAR was available to according tolerance to freezing, the simultaneous expression of two genes generated synergistic effects conferring improved tolerance more effectively even severe freezing. PMID:24170089

  1. Line differences in Cor/Lea and fructan biosynthesis-related gene transcript accumulation are related to distinct freezing tolerance levels in synthetic wheat hexaploids.

    PubMed

    Yokota, Hirokazu; Iehisa, Julio C M; Shimosaka, Etsuo; Takumi, Shigeo

    2015-03-15

    In common wheat, cultivar differences in freezing tolerance are considered to be mainly due to allelic differences at two major loci controlling freezing tolerance. One of the two loci, Fr-2, is coincident with a cluster of genes encoding C-repeat binding factors (CBFs), which induce downstream Cor/Lea genes during cold acclimation. Here, we conducted microarray analysis to study comprehensive changes in gene expression profile under long-term low-temperature (LT) treatment and to identify other LT-responsive genes related to cold acclimation in leaves of seedlings and crown tissues of a synthetic hexaploid wheat line. The microarray analysis revealed marked up-regulation of a number of Cor/Lea genes and fructan biosynthesis-related genes under the long-term LT treatment. For validation of the microarray data, we selected four synthetic wheat lines that contain the A and B genomes from the tetraploid wheat cultivar Langdon and the diverse D genomes originating from different Aegilops tauschii accessions with distinct levels of freezing tolerance after cold acclimation. Quantitative RT-PCR showed increased transcript levels of the Cor/Lea, CBF, and fructan biosynthesis-related genes in more freezing-tolerant lines than in sensitive lines. After a 14-day LT treatment, a significant difference in fructan accumulation was observed among the four lines. Therefore, the fructan biosynthetic pathway is associated with cold acclimation in development of wheat freezing tolerance and is another pathway related to diversity in freezing tolerance, in addition to the CBF-mediated Cor/Lea expression pathway. Copyright © 2014 Elsevier GmbH. All rights reserved.

  2. GaMYB85, an R2R3 MYB gene, in transgenic Arabidopsis plays an important role in drought tolerance.

    PubMed

    Butt, Hamama Islam; Yang, Zhaoen; Gong, Qian; Chen, Eryong; Wang, Xioaqian; Zhao, Ge; Ge, Xiaoyang; Zhang, Xueyan; Li, Fuguang

    2017-08-22

    MYB transcription factors (TFs) are one of the largest families of TFs in higher plants and are involved in diverse biological, functional, and structural processes. Previously, very few functional validation studies on R2R3 MYB have been conducted in cotton in response to abiotic stresses. In the current study, GaMYB85, a cotton R2R3 MYB TF, was ectopically expressed in Arabidopsis thaliana (Col-0) and was functionally characterized by overexpression in transgenic plants. The in-silico analysis of GaMYB85 shows the presence of a SANT domain with a conserved R2R3 MYB imperfect repeat. The GaMYB85 protein has a 257-amino acid sequence, a molecular weight of 24.91 kD, and an isoelectric point of 5.58. Arabidopsis plants overexpressing GaMYB85 exhibited a higher seed germination rate in response to mannitol and salt stress, and higher drought avoidance efficiency than wild-type plants upon water deprivation. These plants had notably higher levels of free proline and chlorophyll with subsequent lower water loss rates and higher relative water content. Germination of GaMYB85 transgenics was more sensitive to abscisic acid (ABA) and extremely liable to ABA-induced inhibition of primary root elongation. Moreover, when subjected to treatment with different concentrations of ABA, transgenic plants with ectopically expressed GaMYB85 showed reduced stomatal density, with greater stomatal size and lower stomatal opening rates than those in wild-type plants. Ectopic expression of GaMYB85 led to enhanced transcript levels of stress-related marker genes such as RD22, ADH1, RD29A, P5CS, and ABI5. Our results indicate previously unknown roles of GaMYB85, showing that it confers good drought, salt, and freezing tolerance, most probably via an ABA-induced pathway. These findings can potentially be exploited to develop improved abiotic stress tolerance in cotton plants.

  3. Freezing tolerance and the histology of recovering nodes in St. Augustinegrass

    USDA-ARS?s Scientific Manuscript database

    St. Augustinegrass [Stenataphrum secundatum (Walt.) Kuntze] is a coarse-textured turfgrass commonly utilized for its excellent shade tolerance. However, inferior cold tolerance in comparison to other warm-season grasses limits its range primarily to the southeastern U. S., The objectives of this stu...

  4. Cadmium uptake, translocation, and tolerance in AHA1OX Arabidopsis thaliana.

    PubMed

    Hou, Lingyan; Shi, Weiming; Wei, Wenhui; Shen, Hong

    2011-02-01

    Information on cadmium (Cd) uptake and transport is essential to understand better the physiology of Cd tolerance in plants. In this study, Cd uptake, translocation, and tolerance were investigated in AHA1 (Arabidopsis plasma membrane H(+)-ATPase gene) overexpressed plants. Exposed to 10 µM CdCl(2), AHA1OX showed a higher root elongation, accumulated more Cd, and maintained better integrity of nucleus membrane of root tips in comparison to the control plant (WT), suggesting that AHA1OX was more Cd tolerant than WT. To investigate Cd tolerance mechanism of AHA1OX plants, we measured the activity of plasma membrane H(+)-ATPase and the secretion of citrate. Results indicated that treatment with 10 µM of Cd stimulated the activity of plasma membrane H(+)-ATPase and the secretion of citrate, while 30 µM of Cd inhibited them. AHA1OX had higher activity of H(+)-ATPase and secretion of citrate than WT. Addition of citrate enhanced root-to-shoot translocation of Cd significantly. A higher root-to-shoot Cd translocation was observed in AHA1OX than in WT plants. Treatment with low temperature or metabolic inhibitor (carbonyl cyanide m-chlorophenylhydrazone) inhibited Cd uptake and translocation. The study of Cd forms using sequential extraction indicated that Cd was mainly present as a protein-bound form, and AHA1OX had more water-soluble Cd than WT. Taken together, our results suggested that the Cd tolerance of AHA1OX was associated with its root-to-shoot Cd translocation and secretion of citrate, which converts Cd(2+) into less toxic and more easily transportable forms in plant cells.

  5. Aluminium-induced ion transport in Arabidopsis: the relationship between Al tolerance and root ion flux.

    PubMed

    Bose, Jayakumar; Babourina, Olga; Shabala, Sergey; Rengel, Zed

    2010-06-01

    Aluminium (Al) rhizotoxicity coincides with low pH; however, it is unclear whether plant tolerance to these two factors is controlled by the same mechanism. To address this question, the Al-resistant alr104 mutant, two Al-sensitive mutants (als3 and als5), and wild-type Arabidopsis thaliana were compared in long-term exposure (solution culture) and in short-term exposure experiments (H(+) and K(+) fluxes, rhizosphere pH, and plasma membrane potential, E(m)). Based on biomass accumulation, als5 and alr104 showed tolerance to low pH, whereas alr104 was tolerant to the combined low-pH/Al treatment. The sensitivity of the als5 and als3 mutants to the Al stress was similar. The Al-induced decrease in H(+) influx at the distal elongation zone (DEZ) and Al-induced H(+) efflux at the mature zone (MZ) were higher in the Al-sensitive mutants (als3 and als5) than in the wild type and the alr104 mutant. Under combined low-pH/Al treatment, alr104 and the wild type had depolarized plasma membranes for the entire 30 min measurement period, whereas in the Al-sensitive mutants (als3 and als5), initial depolarization to around -60 mV became hyperpolarization at -110 mV after 20 min. At the DEZ, the E(m) changes corresponded to the changes in K(+) flux: K(+) efflux was higher in alr104 and the wild type than in the als3 and als5 mutants. In conclusion, Al tolerance in the alr104 mutant correlated with E(m) depolarization, higher K(+) efflux, and higher H(+) influx, which led to a more alkaline rhizosphere under the combined low-pH/Al stress. Low-pH tolerance (als5) was linked to higher H(+) uptake under low-pH stress, which was abolished by Al exposure.

  6. Increased β-cyanoalanine nitrilase activity improves cyanide tolerance and assimilation in Arabidopsis.

    PubMed

    O'Leary, Brendan; Preston, Gail M; Sweetlove, Lee J

    2014-01-01

    Plants naturally produce cyanide (CN) which is maintained at low levels in their cells by a process of rapid assimilation. However, high concentrations of environmental CN associated with activities such as industrial pollution are toxic to plants. There is thus an interest in increasing the CN detoxification capacity of plants as a potential route to phytoremediation. Here, Arabidopsis seedlings overexpressing the Pseudomonas fluorescens β-cyanoalanine nitrilase pinA were compared with wild-type and a β-cyanoalanine nitrilase knockout line (ΔAtnit4) for growth in the presence of exogenous CN. After incubation with CN, +PfpinA seedlings had increased root length, increased fresh weight, and decreased leaf bleaching compared with wild-type, indicating increased CN tolerance. The increased tolerance was achieved without an increase in β-cyanoalanine synthase activity, the other enzyme in the cyanide assimilation pathway, suggesting that nitrilase activity is the limiting factor for cyanide detoxification. Labeling experiments with [¹³C]KCN demonstrated that the altered CN tolerance could be explained by differences in flux from CN to Asn caused by altered β-cyanoalanine nitrilase activity. Metabolite profiling after CN treatment provided new insight into downstream metabolism, revealing onward metabolism of Asn by the photorespiratory nitrogen cycle and accumulation of aromatic amino acids.

  7. Ectopic expression of Arabidopsis glycosyltransferase UGT85A5 enhances salt stress tolerance in tobacco.

    PubMed

    Sun, Yan-Guo; Wang, Bo; Jin, Shang-Hui; Qu, Xiao-Xia; Li, Yan-Jie; Hou, Bing-Kai

    2013-01-01

    Abiotic stresses greatly influence plant growth and productivity. While glycosyltransferases are widely distributed in plant kingdom, their biological roles in response to abiotic stresses are largely unknown. In this study, a novel Arabidopsis glycosyltransferase gene UGT85A5 was identified as significantly induced by salt stress. Ectopic expression of UGT85A5 in tobacco enhanced the salt stress tolerance in the transgenic plants. There were higher seed germination rates, better plant growth and less chlorophyll loss in transgenic lines compared to wild type plants under salt stress. This enhanced tolerance of salt stress was correlated with increased accumulations of proline and soluble sugars, but with decreases in malondialdehyde accumulation and Na(+)/K(+) ratio in UGT85A5-expressing tobacco. Furthermore, during salt stress, expression of several carbohydrate metabolism-related genes including those for sucrose synthase, sucrose-phosphate synthase, hexose transporter and a group2 LEA protein were obviously upregulated in UGT85A5-expressing transgenic plants compared with wild type controls. Thus, these findings suggest a specific protective role of this glycosyltransferase against salt stress and provide a genetic engineering strategy to improve salt tolerance of crops.

  8. Release of SOS2 kinase from sequestration with GIGANTEA determines salt tolerance in Arabidopsis.

    PubMed

    Kim, Woe-Yeon; Ali, Zahir; Park, Hee Jin; Park, Su Jung; Cha, Joon-Yung; Perez-Hormaeche, Javier; Quintero, Francisco Javier; Shin, Gilok; Kim, Mi Ri; Qiang, Zhang; Ning, Li; Park, Hyeong Cheol; Lee, Sang Yeol; Bressan, Ray A; Pardo, Jose M; Bohnert, Hans J; Yun, Dae-Jin

    2013-01-01

    Environmental challenges to plants typically entail retardation of vegetative growth and delay or cessation of flowering. Here we report a link between the flowering time regulator, GIGANTEA (GI), and adaptation to salt stress that is mechanistically based on GI degradation under saline conditions, thus retarding flowering. GI, a switch in photoperiodicity and circadian clock control, and the SNF1-related protein kinase SOS2 functionally interact. In the absence of stress, the GI:SOS2 complex prevents SOS2-based activation of SOS1, the major plant Na(+)/H(+)-antiporter mediating adaptation to salinity. GI overexpressing, rapidly flowering, plants show enhanced salt sensitivity, whereas gi mutants exhibit enhanced salt tolerance and delayed flowering. Salt-induced degradation of GI confers salt tolerance by the release of the SOS2 kinase. The GI-SOS2 interaction introduces a higher order regulatory circuit that can explain in molecular terms, the long observed connection between floral transition and adaptive environmental stress tolerance in Arabidopsis.

  9. Enhancement of oxidative and drought tolerance in Arabidopsis by overaccumulation of antioxidant flavonoids

    PubMed Central

    Nakabayashi, Ryo; Yonekura-Sakakibara, Keiko; Urano, Kaoru; Suzuki, Makoto; Yamada, Yutaka; Nishizawa, Tomoko; Matsuda, Fumio; Kojima, Mikiko; Sakakibara, Hitoshi; Shinozaki, Kazuo; Michael, Anthony J; Tohge, Takayuki; Yamazaki, Mami; Saito, Kazuki

    2014-01-01

    The notion that plants use specialized metabolism to protect against environmental stresses needs to be experimentally proven by addressing the question of whether stress tolerance by specialized metabolism is directly due to metabolites such as flavonoids. We report that flavonoids with radical scavenging activity mitigate against oxidative and drought stress in Arabidopsis thaliana. Metabolome and transcriptome profiling and experiments with oxidative and drought stress in wild-type, single overexpressors of MYB12/PFG1 (PRODUCTION OF FLAVONOL GLYCOSIDES1) or MYB75/PAP1 (PRODUCTION OF ANTHOCYANIN PIGMENT1), double overexpressors of MYB12 and PAP1, transparent testa4 (tt4) as a flavonoid-deficient mutant, and flavonoid-deficient MYB12 or PAP1 overexpressing lines (obtained by crossing tt4 and the individual MYB overexpressor) demonstrated that flavonoid overaccumulation was key to enhanced tolerance to such stresses. Antioxidative activity assays using 2,2-diphenyl-1-picrylhydrazyl, methyl viologen, and 3,3′-diaminobenzidine clearly showed that anthocyanin overaccumulation with strong in vitro antioxidative activity mitigated the accumulation of reactive oxygen species in vivo under oxidative and drought stress. These data confirm the usefulness of flavonoids for enhancing both biotic and abiotic stress tolerance in crops. PMID:24274116

  10. The rice Mybleu transcription factor increases tolerance to oxygen deprivation in Arabidopsis plants.

    PubMed

    Mattana, Monica; Vannini, Candida; Espen, Luca; Bracale, Marcella; Genga, Annamaria; Marsoni, Milena; Iriti, Marcello; Bonazza, Veronica; Romagnoli, Francesco; Baldoni, Elena; Coraggio, Immacolata; Locatelli, Franca

    2007-09-01

    Mybleu is a natural incomplete transcription factor of rice (Oryza sativa), consisting of a partial Myb repeat followed by a short leucine zipper. We previously showed its localization to the apical region of rice roots and coleoptiles. Specifically, in coleoptiles, Mybleu is expressed under both aerobic and anaerobic conditions, whereas in roots, it is expressed only under aerobic conditions. Mybleu is able to dimerize with canonical leucine zippers and to activate transcription selectively. To investigate Mybleu function in vivo, we transformed Arabidopsis thaliana and evaluated several morphological, physiological and biochemical parameters. In agreement with a hypothesized role of Mybleu in cell elongation in the differentiation zone, we found that the constitutive expression of this transcription factor in Arabidopsis induced elongation in the primary roots and in the internodal region of the floral stem; we also observed a modification of the root apex morphology in transformed lines. Based on the high expression of Mybleu in anaerobic rice coleoptiles, we studied the role of this transcription factor in transgenic plants grown under low-oxygen conditions. We found that overexpression of this transcription factor increased tolerance to oxygen deficit. In transgenic plants, this effect may depend both on the maintenance of a higher metabolism during stress and on the higher expression levels of certain genes involved in the anaerobic response.

  11. Ascorbate peroxidase from Jatropha curcas enhances salt tolerance in transgenic Arabidopsis.

    PubMed

    Chen, Y; Cai, J; Yang, F X; Zhou, B; Zhou, L R

    2015-05-11

    Ascorbate peroxidase (APX) plays a central role in the ascorbate-glutathione cycle and is a key enzyme in cellular H2O2 me-tabolism. It includes a family of isoenzymes with different character-istics, which are identified in many higher plants. In the present study, we isolated the APX gene from Jatropha curcas L, which is similar with other previously characterized APXs as revealed by alignment and phylogenetic analysis of its deduced amino acid sequence. Real-time qPCR analysis showed that the expression level of JcAPX transcript significantly increased under NaCl stress. Subsequently, to elucidate the contribution of JcAPX to the protection against salt-induced oxi-dative stress, the expression construct p35S: JcAPX was created and transformed into Arabidopsis and transcribed. Under 150-mM NaCl stress, compared with wild type (WT), the overexpression of JcAPX in Arabidopsis increased the germination rate, the number of leaves, and the rosette area. In addition, the transgenic plants had longer roots, higher total chlorophyll content, higher total APX activity, and lower H2O2 content than the WT under NaCl stress conditions. These results suggested that higher APX activity in transgenic lines increases the salt tolerance by enhancing scavenging capacity for reactive oxygen spe-cies under NaCl stress conditions.

  12. Knock-out of Arabidopsis AtNHX4 gene enhances tolerance to salt stress

    SciTech Connect

    Li, Hong-Tao; Liu, Hua; Gao, Xiao-Shu; Zhang, Hongxia

    2009-05-08

    AtNHX4 belongs to the monovalent cation:proton antiporter-1 (CPA1) family in Arabidopsis. Several members of this family have been shown to be critical for plant responses to abiotic stress, but little is known on the biological functions of AtNHX4. Here, we provide the evidence that AtNHX4 plays important roles in Arabidopsis responses to salt stress. Expression of AtNHX4 was responsive to salt stress and abscisic acid. Experiments with CFP-AtNHX4 fusion protein indicated that AtNHX4 is vacuolar localized. The nhx4 mutant showed enhanced tolerance to salt stress, and lower Na{sup +} content under high NaCl stress compared with wild-type plants. Furthermore, heterologous expression of AtNHX4 in Escherichia coli BL21 rendered the transformants hypersensitive to NaCl. Deletion of the hydrophilic C-terminus of AtNHX4 dramatically increased the hypersensitivity of transformants, indicating that AtNHX4 may function in Na{sup +} homeostasis in plant cell, and its C-terminus plays a role in regulating the AtNHX4 activity.

  13. Photosynthetic capacity of Arabidopsis plants at the reproductive stage tolerates γ irradiation.

    PubMed

    Kim, Jin-Hong; Moon, Yu Ran; Lee, Min Hee; Kim, Ji Hong; Wi, Seung Gon; Park, Bong-Ju; Kim, Cha Soon; Chung, Byung Yeoup

    2011-01-01

    The developmental stage has an influence on the overall responses of plants under biotic or abiotic stress conditions. However, there is a lack of data about the effects of ionizing radiation in plants at different developmental stages. We examined radiation sensitivity of Arabidopsis plants in terms of photosynthetic ability and oxidative stress resistance at two distinct vegetative and reproductive stages, which correspond to 23 and 43 d after seeding (DAS), respectively. When plants were exposed to γ rays at a dose rate 50 Gy h(-1) for 4 h, they were characterized as various common or differential cellular responses depending on the developmental stage. Radial expansion of leaves, inhibition of non-photochemical quenching, and production of •O(2)(-) and H(2)O(2) under methyl viologen-induced photooxidative stress were commonly more conspicuous in the irradiated leaves of both plants than in the respective control. In contrast, the 23 and 43-DAS plants were explicitly discriminated in growth, chloroplast number & ultrastructure, photosynthetic pigment content & activity, and protein damage after γ irradiation. Natural leaf senescence was thereby enhanced in the irradiated leaves of the 23-DAS plants, while it was reversely alleviated in those of the 43-DAS ones. These results suggest that photosynthetic machineries of Arabidopsis plants at the reproductive stage can be relatively tolerant to γ rays of 200 Gy.

  14. The RdDM pathway is required for basal heat tolerance in Arabidopsis.

    PubMed

    Popova, Olga V; Dinh, Huy Q; Aufsatz, Werner; Jonak, Claudia

    2013-03-01

    Heat stress affects epigenetic gene silencing in Arabidopsis. To test for a mechanistic involvement of epigenetic regulation in heat-stress responses, we analyzed the heat tolerance of mutants defective in DNA methylation, histone modifications, chromatin-remodeling, or siRNA-based silencing pathways. Plants deficient in NRPD2, the common second-largest subunit of RNA polymerases IV and V, and in the Rpd3-type histone deacetylase HDA6 were hypersensitive to heat exposure. Microarray analysis demonstrated that NRPD2 and HDA6 have independent roles in transcriptional reprogramming in response to temperature stress. The misexpression of protein-coding genes in nrpd2 mutants recovering from heat correlated with defective epigenetic regulation of adjacent transposon remnants which involved the loss of control of heat-stress-induced read-through transcription. We provide evidence that the transcriptional response to temperature stress, at least partially, relies on the integrity of the RNA-dependent DNA methylation pathway.

  15. Overexpression of Selenocysteine Methyltransferase in Arabidopsis and Indian Mustard Increases Selenium Tolerance and Accumulation1

    PubMed Central

    LeDuc, Danika L.; Tarun, Alice S.; Montes-Bayon, Maria; Meija, Juris; Malit, Michele F.; Wu, Carol P.; AbdelSamie, Manal; Chiang, Chih-Yuan; Tagmount, Abderrhamane; deSouza, Mark; Neuhierl, Bernhard; Böck, August; Caruso, Joseph; Terry, Norman

    2004-01-01

    A major goal of phytoremediation is to transform fast-growing plants with genes from plant species that hyperaccumulate toxic trace elements. We overexpressed the gene encoding selenocysteine methyltransferase (SMT) from the selenium (Se) hyperaccumulator Astragalus bisulcatus in Arabidopsis and Indian mustard (Brassica juncea). SMT detoxifies selenocysteine by methylating it to methylselenocysteine, a nonprotein amino acid, thereby diminishing the toxic misincorporation of Se into protein. Our Indian mustard transgenic plants accumulated more Se in the form of methylselenocysteine than the wild type. SMT transgenic seedlings tolerated Se, particularly selenite, significantly better than the wild type, producing 3- to 7-fold greater biomass and 3-fold longer root lengths. Moreover, SMT plants had significantly increased Se accumulation and volatilization. This is the first study, to our knowledge, in which a fast-growing plant was genetically engineered to overexpress a gene from a hyperaccumulator in order to increase phytoremediation potential. PMID:14671009

  16. The Vacuolar Manganese Transporter MTP8 Determines Tolerance to Iron Deficiency-Induced Chlorosis in Arabidopsis.

    PubMed

    Eroglu, Seckin; Meier, Bastian; von Wirén, Nicolaus; Peiter, Edgar

    2016-02-01

    Iron (Fe) deficiency is a widespread nutritional disorder on calcareous soils. To identify genes involved in the Fe deficiency response, Arabidopsis (Arabidopsis thaliana) transfer DNA insertion lines were screened on a high-pH medium with low Fe availability. This approach identified METAL TOLERANCE PROTEIN8 (MTP8), a member of the Cation Diffusion Facilitator family, as a critical determinant for the tolerance to Fe deficiency-induced chlorosis, also on soil substrate. Subcellular localization to the tonoplast, complementation of a manganese (Mn)-sensitive Saccharomyces cerevisiae yeast strain, and Mn sensitivity of mtp8 knockout mutants characterized the protein as a vacuolar Mn transporter suitable to prevent plant cells from Mn toxicity. MTP8 expression was strongly induced on low-Fe as well as high-Mn medium, which were both strictly dependent on the transcription factor FIT, indicating that high-Mn stress induces Fe deficiency. mtp8 mutants were only hypersensitive to Fe deficiency when Mn was present in the medium, which further suggested an Mn-specific role of MTP8 during Fe limitation. Under those conditions, mtp8 mutants not only translocated more Mn to the shoot than did wild-type plants but suffered in particular from critically low Fe concentrations and, hence, Fe chlorosis, although the transcriptional Fe deficiency response was up-regulated more strongly in mtp8. The diminished uptake of Fe from Mn-containing low-Fe medium by mtp8 mutants was caused by an impaired ability to boost the ferric chelate reductase activity, which is an essential process in Fe acquisition. These findings provide a mechanistic explanation for the long-known interference of Mn in Fe nutrition and define the molecular processes by which plants alleviate this antagonism. © 2016 American Society of Plant Biologists. All Rights Reserved.

  17. Expression of Vitis amurensis NAC26 in Arabidopsis enhances drought tolerance by modulating jasmonic acid synthesis.

    PubMed

    Fang, Linchuan; Su, Lingye; Sun, Xiaoming; Li, Xinbo; Sun, Mengxiang; Karungo, Sospeter Karanja; Fang, Shuang; Chu, Jinfang; Li, Shaohua; Xin, Haiping

    2016-04-01

    The growth and fruit quality of grapevines are widely affected by abnormal climatic conditions such as water deficits, but many of the precise mechanisms by which grapevines respond to drought stress are still largely unknown. Here, we report that VaNAC26, a member of the NAC transcription factor family, was upregulated dramatically during cold, drought and salinity treatments in Vitis amurensis, a cold and drought-hardy wild Vitis species. Heterologous overexpression of VaNAC26 enhanced drought and salt tolerance in transgenic Arabidopsis. Higher activities of antioxidant enzymes and lower concentrations of H2O2 and O2 (-) were found in VaNAC26-OE lines than in wild type plants under drought stress. These results indicated that scavenging by reactive oxygen species (ROS) was enhanced by VaNAC26 in transgenic lines. Microarray-based transcriptome analysis revealed that genes related to jasmonic acid (JA) synthesis and signaling were upregulated in VaNAC26-OE lines under both normal and drought conditions. VaNAC26 showed a specific binding ability on the NAC recognition sequence (NACRS) motif, which broadly exists in the promoter regions of upregulated genes in transgenic lines. Endogenous JA content significantly increased in the VaNAC26-OE lines 2 and 3. Our data suggest that VaNAC26 responds to abiotic stresses and may enhance drought tolerance by transcriptional regulation of JA synthesis in Arabidopsis. © The Author 2016. Published by Oxford University Press on behalf of the Society for Experimental Biology.

  18. Impairment of Sulfite Reductase Decreases Oxidative Stress Tolerance in Arabidopsis thaliana

    PubMed Central

    Wang, Meiping; Jia, Yunli; Xu, Ziwei; Xia, Zongliang

    2016-01-01

    As an essential enzyme in the sulfate assimilation reductive pathway, sulfite reductase (SiR) plays important roles in diverse metabolic processes such as sulfur homeostasis and cysteine metabolism. However, whether plant SiR is involved in oxidative stress response is largely unknown. Here, we show that SiR functions in methyl viologen (MV)-induced oxidative stress in Arabidopsis. The transcript levels of SiR were higher in leaves, immature siliques, and roots and were markedly and rapidly up-regulated by MV exposure. The SiR knock-down transgenic lines had about 60% residual transcripts and were more susceptible than wild-type when exposed to oxidative stress. The severe damage phenotypes of the SiR-impaired lines were accompanied by increases of hydrogen peroxide (H2O2), malondialdehyde (MDA), and sulfite accumulations, but less amounts of glutathione (GSH). Interestingly, application of exogenous GSH effectively rescued corresponding MV hypersensitivity in SiR-impaired plants. qRT-PCR analysis revealed that there was significantly increased expression of several sulfite metabolism-related genes in SiR-impaired lines. Noticeably, enhanced transcripts of the three APR genes were quite evident in SiR-impaired plants; suggesting that the increased sulfite in the SiR-impaired plants could be a result of the reduced SiR coupled to enhanced APR expression during oxidative stress. Together, our results indicate that SiR is involved in oxidative stress tolerance possibly by maintaining sulfite homeostasis, regulating GSH levels, and modulating sulfite metabolism-related gene expression in Arabidopsis. SiR could be exploited for engineering environmental stress-tolerant plants in molecular breeding of crops. PMID:27994615

  19. CATION EXCHANGER1 Cosegregates with Cadmium Tolerance in the Metal Hyperaccumulator Arabidopsis halleri and Plays a Role in Limiting Oxidative Stress in Arabidopsis Spp.

    PubMed

    Baliardini, Cecilia; Meyer, Claire-Lise; Salis, Pietrino; Saumitou-Laprade, Pierre; Verbruggen, Nathalie

    2015-09-01

    Arabidopsis halleri is a model species for the study of plant adaptation to extreme metallic conditions. In this species, cadmium (Cd) tolerance seems to be constitutive, and the mechanisms underlying the trait are still poorly understood. A previous quantitative trait loci (QTL) analysis performed on A. halleri × Arabidopsis lyrata backcross population1 identified the metal-pump gene Heavy Metal ATPase4 as the major genetic determinant for Cd tolerance. However, although necessary, Heavy Metal ATPase4 alone is not sufficient for determining this trait. After fine mapping, a gene encoding a calcium(2+)/hydrogen(+) antiporter, cation/hydrogen(+) exchanger1 (CAX1), was identified as a candidate gene for the second QTL of Cd tolerance in A. halleri. Backcross population1 individuals displaying the A. halleri allele for the CAX1 locus exhibited significantly higher CAX1 expression levels compared with the ones with the A. lyrata allele, and a positive correlation between CAX1 expression and Cd tolerance was observed. Here, we show that this QTL is conditional and that it is only detectable at low external Ca concentration. CAX1 expression in both roots and shoots was higher in A. halleri than in the close Cd-sensitive relative species A. lyrata and Arabidopsis thaliana. Moreover, CAX1 loss of function in A. thaliana led to higher Cd sensitivity at low concentration of Ca, higher sensitivity to methylviologen, and stronger accumulation of reactive oxygen species after Cd treatment. Overall, this study identifies a unique genetic determinant of Cd tolerance in the metal hyperaccumulator A. halleri and offers a new twist for the function of CAX1 in plants. © 2015 American Society of Plant Biologists. All Rights Reserved.

  20. Arabidopsis thaliana NIP7;1 is involved in tissue arsenic distribution and tolerance in response to arsenate.

    PubMed

    Lindsay, Emma R; Maathuis, Frans J M

    2016-03-01

    The Arabidopsis aquaglyceroporin NIP7;1 is involved in uptake and tolerance to the trivalent arsenic species arsenite. Here, we show that NIP7;1 is also involved in the response to pentavalent arsenate. Loss of function of NIP7;1 improved tolerance to arsenate and reduced arsenic levels in both the phloem and xylem, resulting in altered arsenic distribution between tissues. There was no clear correlation between growth and shoot arsenic concentration. This is the first report detailing the involvement of a NIP transporter in response to arsenate. The data suggest that these proteins are relevant targets for breeding and engineering arsenic tolerance in crops.

  1. Enhanced freeze tolerance of baker's yeast by overexpressed trehalose-6-phosphate synthase gene (TPS1) and deleted trehalase genes in frozen dough.

    PubMed

    Tan, Haigang; Dong, Jian; Wang, Guanglu; Xu, Haiyan; Zhang, Cuiying; Xiao, Dongguang

    2014-08-01

    Several recombinant strains with overexpressed trehalose-6-phosphate synthase gene (TPS1) and/or deleted trehalase genes were obtained to elucidate the relationships between TPS1, trehalase genes, content of intracellular trehalose and freeze tolerance of baker's yeast, as well as improve the fermentation properties of lean dough after freezing. In this study, strain TL301(TPS1) overexpressing TPS1 showed 62.92 % higher trehalose-6-phosphate synthase (Tps1) activity and enhanced the content of intracellular trehalose than the parental strain. Deleting ATH1 exerted a significant effect on trehalase activities and the degradation amount of intracellular trehalose during the first 30 min of prefermentation. This finding indicates that acid trehalase (Ath1) plays a role in intracellular trehalose degradation. NTH2 encodes a functional neutral trehalase (Nth2) that was significantly involved in intracellular trehalose degradation in the absence of the NTH1 and/or ATH1 gene. The survival ratio, freeze-tolerance ratio and relative fermentation ability of strain TL301(TPS1) were approximately twice as high as those of the parental strain (BY6-9α). The increase in freeze tolerance of strain TL301(TPS1) was accompanied by relatively low trehalase activity, high Tps1 activity and high residual content of intracellular trehalose. Our results suggest that overexpressing TPS1 and deleting trehalase genes are sufficient to improve the freeze tolerance of baker's yeast in frozen dough. The present study provides guidance for the commercial baking industry as well as the research on the intracellular trehalose mobilization and freeze tolerance of baker's yeast.

  2. Isolation and characterization of a freeze-tolerant diploid derivative of an industrial baker's yeast strain and its use in frozen doughs.

    PubMed

    Teunissen, Aloys; Dumortier, Françoise; Gorwa, Marie-Françoise; Bauer, Jürgen; Tanghe, An; Loïez, Annie; Smet, Peter; Van Dijck, Patrick; Thevelein, Johan M

    2002-10-01

    The routine production and storage of frozen doughs are still problematic. Although commercial baker's yeast is highly resistant to environmental stress conditions, it rapidly loses stress resistance during dough preparation due to the initiation of fermentation. As a result, the yeast loses gassing power significantly during storage of frozen doughs. We obtained freeze-tolerant mutants of polyploid industrial strains following screening for survival in doughs prepared with UV-mutagenized yeast and subjected to 200 freeze-thaw cycles. Two strains in the S47 background with a normal growth rate and the best freeze tolerance under laboratory conditions were selected for production in a 20-liter pilot fermentor. Before frozen storage, the AT25 mutant produced on the 20-liter pilot scale had a 10% higher gassing power capacity than the S47 strain, while the opposite was observed for cells produced under laboratory conditions. AT25 also retained more freeze tolerance during the initiation of fermentation in liquid cultures and more gassing power during storage of frozen doughs. Other industrially important properties (yield, growth rate, nitrogen assimilation, and phosphorus content) were very similar. AT25 had only half of the DNA content of S47, and its cell size was much smaller. Several diploid segregants of S47 had freeze tolerances similar to that of AT25 but inferior performance for other properties, while an AT25-derived tetraploid, TAT25, showed only slightly improved freeze tolerance compared to S47. When AT25 was cultured in a 20,000-liter fermentor under industrial conditions, it retained its superior performance and thus appears to be promising for use in frozen dough production. Our results also show that a diploid strain can perform at least as well as a tetraploid strain for commercial baker's yeast production and usage.

  3. Isolation and Characterization of a Freeze-Tolerant Diploid Derivative of an Industrial Baker's Yeast Strain and Its Use in Frozen Doughs

    PubMed Central

    Teunissen, Aloys; Dumortier, Françoise; Gorwa, Marie-Françoise; Bauer, Jürgen; Tanghe, An; Loïez, Annie; Smet, Peter; Van Dijck, Patrick; Thevelein, Johan M.

    2002-01-01

    The routine production and storage of frozen doughs are still problematic. Although commercial baker's yeast is highly resistant to environmental stress conditions, it rapidly loses stress resistance during dough preparation due to the initiation of fermentation. As a result, the yeast loses gassing power significantly during storage of frozen doughs. We obtained freeze-tolerant mutants of polyploid industrial strains following screening for survival in doughs prepared with UV-mutagenized yeast and subjected to 200 freeze-thaw cycles. Two strains in the S47 background with a normal growth rate and the best freeze tolerance under laboratory conditions were selected for production in a 20-liter pilot fermentor. Before frozen storage, the AT25 mutant produced on the 20-liter pilot scale had a 10% higher gassing power capacity than the S47 strain, while the opposite was observed for cells produced under laboratory conditions. AT25 also retained more freeze tolerance during the initiation of fermentation in liquid cultures and more gassing power during storage of frozen doughs. Other industrially important properties (yield, growth rate, nitrogen assimilation, and phosphorus content) were very similar. AT25 had only half of the DNA content of S47, and its cell size was much smaller. Several diploid segregants of S47 had freeze tolerances similar to that of AT25 but inferior performance for other properties, while an AT25-derived tetraploid, TAT25, showed only slightly improved freeze tolerance compared to S47. When AT25 was cultured in a 20,000-liter fermentor under industrial conditions, it retained its superior performance and thus appears to be promising for use in frozen dough production. Our results also show that a diploid strain can perform at least as well as a tetraploid strain for commercial baker's yeast production and usage. PMID:12324320

  4. Overexpression of ARGOS Genes Modifies Plant Sensitivity to Ethylene, Leading to Improved Drought Tolerance in Both Arabidopsis and Maize.

    PubMed

    Shi, Jinrui; Habben, Jeffrey E; Archibald, Rayeann L; Drummond, Bruce J; Chamberlin, Mark A; Williams, Robert W; Lafitte, H Renee; Weers, Ben P

    2015-09-01

    Lack of sufficient water is a major limiting factor to crop production worldwide, and the development of drought-tolerant germplasm is needed to improve crop productivity. The phytohormone ethylene modulates plant growth and development as well as plant response to abiotic stress. Recent research has shown that modifying ethylene biosynthesis and signaling can enhance plant drought tolerance. Here, we report novel negative regulators of ethylene signal transduction in Arabidopsis (Arabidopsis thaliana) and maize (Zea mays). These regulators are encoded by the ARGOS gene family. In Arabidopsis, overexpression of maize ARGOS1 (ZmARGOS1), ZmARGOS8, Arabidopsis ARGOS homolog ORGAN SIZE RELATED1 (AtOSR1), and AtOSR2 reduced plant sensitivity to ethylene, leading to enhanced drought tolerance. RNA profiling and genetic analysis suggested that the ZmARGOS1 transgene acts between an ethylene receptor and CONSTITUTIVE TRIPLE RESPONSE1 in the ethylene signaling pathway, affecting ethylene perception or the early stages of ethylene signaling. Overexpressed ZmARGOS1 is localized to the endoplasmic reticulum and Golgi membrane, where the ethylene receptors and the ethylene signaling protein ETHYLENE-INSENSITIVE2 and REVERSION-TO-ETHYLENE SENSITIVITY1 reside. In transgenic maize plants, overexpression of ARGOS genes also reduces ethylene sensitivity. Moreover, field testing showed that UBIQUITIN1:ZmARGOS8 maize events had a greater grain yield than nontransgenic controls under both drought stress and well-watered conditions.

  5. Membrane adaptation in phospholipids and cholesterol in the widely distributed, freeze-tolerant wood frog, Rana sylvatica.

    PubMed

    Reynolds, Alice M; Lee, Richard E; Costanzo, Jon P

    2014-04-01

    Maintaining proper membrane phase and fluidity is important for preserving membrane structure and function, and by altering membrane lipid composition many organisms can adapt to changing environmental conditions. We compared the phospholipid and cholesterol composition of liver and brain plasma membranes in the freeze-tolerant wood frog, Rana sylvatica, from southern Ohio and Interior Alaska during summer, fall, and winter. We also compared membranes from winter-acclimatized frogs from Ohio that were either acclimated to 0, 4, or 10 °C, or frozen to -2.5 °C and sampled before or after thawing. Lipids were extracted from isolated membranes, separated by one-dimensional thin-layer chromatography, and analyzed via densitometry. Liver membranes underwent seasonal changes in phospholipid composition and lipid ratios, including a winter increase in phosphatidylethanolamine, which serves to increase fluidity. However, whereas Ohioan frogs decreased phosphatidylcholine and increased sphingomyelin, Alaskan frogs only decreased phosphatidylserine, indicating that these phenotypes use different adaptive strategies to meet the functional needs of their membranes. Liver membranes showed no seasonal variation in cholesterol abundance, though membranes from Alaskan frogs contained relatively less cholesterol, consistent with the need for greater fluidity in a colder environment. No lipid changed seasonally in brain membranes in either population. In the thermal acclimation experiment, cold exposure induced an increase in phosphatidylethanolamine in liver membranes and a decrease in cholesterol in brain membranes. No changes occurred during freezing and thawing in membranes from either organ. Wood frogs use tissue-specific membrane adaptation of phospholipids and cholesterol to respond to changing environmental factors, particularly temperature, though not with freezing.

  6. Does acute lead (Pb) contamination influence membrane fatty acid composition and freeze tolerance in intertidal blue mussels in arctic Greenland?

    PubMed

    Thyrring, Jakob; Juhl, Bodil Klein; Holmstrup, Martin; Blicher, Martin E; Sejr, Mikael K

    2015-11-01

    In their natural habitats, organisms are exposed to multiple stressors. Heavy metal contamination stresses the cell membrane due to increased peroxidation of lipids. Likewise, sub-zero air temperatures potentially reduce membrane functionality in ectothermal animals. We tested if acute lead (Pb) exposure for 7 days would influence survival in intertidal blue mussels (Mytilus edulis) after exposure to realistic sub-zero air temperatures. A full factorial experiment with five tissue Pb concentrations between 0 and 3500 μg Pb/g and six sub-zero temperatures from 0 to -17 °C were used to test the hypothesis that sub-lethal effects of Pb may increase the lethality caused by freezing in blue mussels exposed to temperatures simulating Greenland winter conditions. We found a significant effect of temperature on mortality. However, the short-term exposure to Pb did not result in any effects of Pb, nor did we find interactions between Pb and temperature. We analysed the relative abundance of major phospholipid fatty acids (PLFAs) in the gill tissue, but we found no significant effect of Pb tissue concentration on PLFA composition. Results suggest that Pb accumulation has limited effects on freeze tolerance and does not induce membrane damage in terms of persistent lipid peroxidation.

  7. Identification of a Retroelement from the Resurrection Plant Boea hygrometrica That Confers Osmotic and Alkaline Tolerance in Arabidopsis thaliana

    PubMed Central

    Shen, Chun-Ying; Xu, Guang-Hui; Chen, Shi-Xuan; Song, Li-Zhen; Li, Mei-Jing; Wang, Li-Li; Zhu, Yan; Lv, Wei-Tao; Gong, Zhi-Zhong; Liu, Chun-Ming; Deng, Xin

    2014-01-01

    Functional genomic elements, including transposable elements, small RNAs and non-coding RNAs, are involved in regulation of gene expression in response to plant stress. To identify genomic elements that regulate dehydration and alkaline tolerance in Boea hygrometrica, a resurrection plant that inhabits drought and alkaline Karst areas, a genomic DNA library from B. hygrometrica was constructed and subsequently transformed into Arabidopsis using binary bacterial artificial chromosome (BIBAC) vectors. Transgenic lines were screened under osmotic and alkaline conditions, leading to the identification of Clone L1-4 that conferred osmotic and alkaline tolerance. Sequence analyses revealed that L1-4 contained a 49-kb retroelement fragment from B. hygrometrica, of which only a truncated sequence was present in L1-4 transgenic Arabidopsis plants. Additional subcloning revealed that activity resided in a 2-kb sequence, designated Osmotic and Alkaline Resistance 1 (OAR1). In addition, transgenic Arabidopsis lines carrying an OAR1-homologue also showed similar stress tolerance phenotypes. Physiological and molecular analyses demonstrated that OAR1-transgenic plants exhibited improved photochemical efficiency and membrane integrity and biomarker gene expression under both osmotic and alkaline stresses. Short transcripts that originated from OAR1 were increased under stress conditions in both B. hygrometrica and Arabidopsis carrying OAR1. The relative copy number of OAR1 was stable in transgenic Arabidopsis under stress but increased in B. hygrometrica. Taken together, our results indicated a potential role of OAR1 element in plant tolerance to osmotic and alkaline stresses, and verified the feasibility of the BIBAC transformation technique to identify functional genomic elements from physiological model species. PMID:24851859

  8. Identification of a retroelement from the resurrection plant Boea hygrometrica that confers osmotic and alkaline tolerance in Arabidopsis thaliana.

    PubMed

    Zhao, Yan; Xu, Tao; Shen, Chun-Ying; Xu, Guang-Hui; Chen, Shi-Xuan; Song, Li-Zhen; Li, Mei-Jing; Wang, Li-Li; Zhu, Yan; Lv, Wei-Tao; Gong, Zhi-Zhong; Liu, Chun-Ming; Deng, Xin

    2014-01-01

    Functional genomic elements, including transposable elements, small RNAs and non-coding RNAs, are involved in regulation of gene expression in response to plant stress. To identify genomic elements that regulate dehydration and alkaline tolerance in Boea hygrometrica, a resurrection plant that inhabits drought and alkaline Karst areas, a genomic DNA library from B. hygrometrica was constructed and subsequently transformed into Arabidopsis using binary bacterial artificial chromosome (BIBAC) vectors. Transgenic lines were screened under osmotic and alkaline conditions, leading to the identification of Clone L1-4 that conferred osmotic and alkaline tolerance. Sequence analyses revealed that L1-4 contained a 49-kb retroelement fragment from B. hygrometrica, of which only a truncated sequence was present in L1-4 transgenic Arabidopsis plants. Additional subcloning revealed that activity resided in a 2-kb sequence, designated Osmotic and Alkaline Resistance 1 (OAR1). In addition, transgenic Arabidopsis lines carrying an OAR1-homologue also showed similar stress tolerance phenotypes. Physiological and molecular analyses demonstrated that OAR1-transgenic plants exhibited improved photochemical efficiency and membrane integrity and biomarker gene expression under both osmotic and alkaline stresses. Short transcripts that originated from OAR1 were increased under stress conditions in both B. hygrometrica and Arabidopsis carrying OAR1. The relative copy number of OAR1 was stable in transgenic Arabidopsis under stress but increased in B. hygrometrica. Taken together, our results indicated a potential role of OAR1 element in plant tolerance to osmotic and alkaline stresses, and verified the feasibility of the BIBAC transformation technique to identify functional genomic elements from physiological model species.

  9. The sunflower transcription factor HaWRKY76 confers drought and flood tolerance to Arabidopsis thaliana plants without yield penalty.

    PubMed

    Raineri, Jesica; Ribichich, Karina F; Chan, Raquel L

    2015-12-01

    Arabidopsis transgenic plants expressing the sunflower transcription factor HaWRKY76 exhibit increased yield and tolerance to drought and flood stresses. The genetic construct containing HaWRKY76 is proposed as a potential biotechnological tool to improve crops. Water deficit and water excess are abiotic stress factors that seriously affect crops worldwide. To increase the tolerance to such stresses without causing yield penalty constitutes a major goal for biotechnologists. In this survey, we report that HaWRKY76, a divergent sunflower WRKY transcription factor, is able to confer both dehydration and submergence tolerance to Arabidopsis transgenic plants without yield penalty. The expression pattern of HaWRKY76 was analyzed in plants grown in standard conditions and under different watering regimes indicating a regulation by water availability. The corresponding cDNA was isolated and cloned under the control of a constitutive promoter and Arabidopsis plants were transformed with this construct. These transgenic plants presented higher biomass, seed production and sucrose content than controls in standard growth conditions. Moreover, they exhibited tolerance to mild drought or flood (complete submergence/waterlogging) stresses as well as the same or increased yield, depending on the stress severity and plant developmental stage, compared with controls. Drought tolerance occurred via an ABA-independent mechanism and induction of stomatal closure. Submergence tolerance can be explained by the carbohydrate (sucrose and starch) preservation achieved through the repression of fermentation pathways. Higher cell membrane stability and chlorenchyma maintenance could be the nexus between tolerance responses in front of both stresses. Altogether, the obtained results indicated that HaWRKY76 can be a potential biotechnological tool to improve crops yield as well as drought and flood tolerances.

  10. Differential remodeling of the lipidome during cold acclimation in natural accessions of Arabidopsis thaliana.

    PubMed

    Degenkolbe, Thomas; Giavalisco, Patrick; Zuther, Ellen; Seiwert, Bettina; Hincha, Dirk K; Willmitzer, Lothar

    2012-12-01

    Freezing injury is a major factor limiting the geographical distribution of plant species and the growth and yield of crop plants. Plants from temperate climates are able to increase their freezing tolerance during exposure to low but non-freezing temperatures in a process termed cold acclimation. Damage to cellular membranes is the major cause of freezing injury in plants, and membrane lipid composition is strongly modified during cold acclimation. Forward and reverse genetic approaches have been used to probe the role of specific lipid-modifying enzymes in the freezing tolerance of plants. In the present paper we describe an alternative ecological genomics approach that relies on the natural genetic variation within a species. Arabidopsis thaliana has a wide geographical range throughout the Northern Hemisphere with significant natural variation in freezing tolerance that was used for a comparative analysis of the lipidomes of 15 Arabidopsis accessions using ultra-performance liquid chromatography coupled to Fourier-transform mass spectrometry, allowing the detection of 180 lipid species. After 14 days of cold acclimation at 4°C the plants from most accessions had accumulated massive amounts of storage lipids, with most of the changes in long-chain unsaturated triacylglycerides, while the total amount of membrane lipids was only slightly changed. Nevertheless, major changes in the relative amounts of different membrane lipids were also evident. The relative abundance of several lipid species was highly correlated with the freezing tolerance of the accessions, allowing the identification of possible marker lipids for plant freezing tolerance.

  11. Comparison of Dehydrin Gene Expression and Freezing Tolerance in Bromus inermis and Secale cereale Grown in Controlled Environments, Hydroponics, and the Field.

    PubMed Central

    Robertson, A. J.; Weninger, A.; Wilen, R. W.; Fu, P.; Gusta, L. V.

    1994-01-01

    There have been very few reports on the expression of stress-responsive genes in field-grown material. A barley dehydrin cDNA was used to investigate the expression of dehydrin-like transcripts after low-temperature and abscisic acid-induced acclimation of bromegrass (Bromus inermis Leyss) suspension cells and of bromegrass and rye (Secale cereale) plants grown in the field and under controlled environmental conditions. Field-acclimated plants accumulated high levels of dehydrin transcripts and were very freezing tolerant. Plants grown in pots and hydroponics under controlled environments also accumulated dehydrin transcripts and showed increased freezing tolerance. Simulation of a combined drought and freezing stress in pots resulted in expression of dehydrin-like transcripts comparable to those observed in field-acclimated material. PMID:12232403

  12. Differential expression and regulation of iron-regulated metal transporters in Arabidopsis halleri and Arabidopsis thaliana--the role in zinc tolerance.

    PubMed

    Shanmugam, Varanavasiappan; Lo, Jing-Chi; Wu, Chia-Lin; Wang, Shan-Li; Lai, Chong-Cheong; Connolly, Erin L; Huang, Jing-Ling; Yeh, Kuo-Chen

    2011-04-01

    To avoid zinc (Zn) toxicity, plants have developed a Zn homeostasis mechanism to cope with Zn excess in the surrounding soil. In this report, we uncovered the difference of a cross-homeostasis system between iron (Fe) and Zn in dealing with Zn excess in the Zn hyperaccumulator Arabidopsis halleri ssp. gemmifera and nonhyperaccumulator Arabidopsis thaliana. Arabidopsis halleri shows low expression of the Fe acquisition and deficiency response-related genes IRT1 and IRT2 compared with A. thaliana. In A. thaliana, lowering the expression of IRT1 and IRT2 through the addition of excess Fe to the medium increases Zn tolerance. Excess Zn induces significant Fe deficiency in A. thaliana and reduces Fe accumulation in shoots. By contrast, the accumulation of Fe in shoots of A. halleri was stable under various Zn treatments. Root ferric chelate reductase (FRO) activity and expression of FIT are low in A. halleri compared with A. thaliana. Overexpressing a ZIP family member IRT3 in irt1-1, rescues the Fe-deficient phenotype. A fine-tuned Fe homeostasis mechanism in A. halleri maintains optimum Fe level by Zn-regulated ZIP transporters and prevents high Zn uptake through Fe-regulated metal transporters, and in part be responsible for Zn tolerance.

  13. Seasonal differences in freezing tolerance of yellow-cedar and western hemlock trees at a site affected by yellow-cedar decline

    Treesearch

    Paul G. Schaberg; Paul E. Hennon; Amore, David V. D; Gary J. Hawley; Catherine H. Borer; Catherine H. Borer

    2005-01-01

    To assess whether inadequate cold hardiness could be a contributor to yellow-cedar (Chamaecyparis nootkatensis (D. Don) Spach) decline, we measured the freezing tolerance of foliage from yellow-cedar trees in closed-canopy (nondeclining) and open-canopy (declining at elevations below 130 m) stands at three sites along an elevational gradient in the heart of the decline...

  14. Hydrogen sulfide regulates abiotic stress tolerance and biotic stress resistance in Arabidopsis.

    PubMed

    Shi, Haitao; Ye, Tiantian; Han, Ning; Bian, Hongwu; Liu, Xiaodong; Chan, Zhulong

    2015-07-01

    Hydrogen sulfide (H2S) is an important gaseous molecule in various plant developmental processes and plant stress responses. In this study, the transgenic Arabidopsis thaliana plants with modulated expressions of two cysteine desulfhydrases, and exogenous H2S donor (sodium hydrosulfide, NaHS) and H2S scavenger (hypotaurine, HT) pre-treated plants were used to dissect the involvement of H2S in plant stress responses. The cysteine desulfhydrases overexpressing plants and NaHS pre-treated plants exhibited higher endogenous H2S level and improved abiotic stress tolerance and biotic stress resistance, while cysteine desulfhydrases knockdown plants and HT pre-treated plants displayed lower endogenous H2S level and decreased stress resistance. Moreover, H2S upregulated the transcripts of multiple abiotic and biotic stress-related genes, and inhibited reactive oxygen species (ROS) accumulation. Interestingly, MIR393-mediated auxin signaling including MIR393a/b and their target genes (TIR1, AFB1, AFB2, and AFB3) was transcriptionally regulated by H2S, and was related with H2S-induced antibacterial resistance. Moreover, H2S regulated 50 carbon metabolites including amino acids, organic acids, sugars, sugar alcohols, and aromatic amines. Taken together, these results indicated that cysteine desulfhydrase and H2S conferred abiotic stress tolerance and biotic stress resistance, via affecting the stress-related gene expressions, ROS metabolism, metabolic homeostasis, and MIR393-targeted auxin receptors. © 2014 Institute of Botany, Chinese Academy of Sciences.

  15. Cyclic nucleotide gated channel 10 negatively regulates salt tolerance by mediating Na+ transport in Arabidopsis.

    PubMed

    Jin, Yakang; Jing, Wen; Zhang, Qun; Zhang, Wenhua

    2015-01-01

    A number of cyclic nucleotide gated channel (CNGC) genes have been identified in plant genomes, but their functions are mainly undefined. In this study, we identified the role of CNGC10 in the response of Arabidopsis thaliana to salt stress. The cngc10 T-DNA insertion mutant showed greater tolerance to salt than wild-type A. thaliana during seed germination and seedling growth. The cngc10 mutant accumulated less Na(+) and K(+), but not less Ca(2+), in shoots in response to salt stress. By contrast, overexpression of CNGC10 resulted in greater sensitivity to salt stress, and complementation of this gene recovered salt sensitivity. In response to salt stress, heterologous expression of CNGC10 in the Na(+) sensitive yeast mutant strain B31 inhibited growth due to accumulation of Na(+) at a rate greater than that of yeast transformed with an empty vector. Quantitative RT-PCR analysis demonstrated that CNGC10 was expressed mainly in roots and flowers. GUS analysis of a root cross section indicated that CNGC10 was expressed mainly in the endodermis and epidermis. Furthermore, the expression of CNGC10 in roots was dramatically inhibited by exposure to 200 mM NaCl for 6 h. These data suggest that CNGC10 negatively regulates salt tolerance in A. thaliana and may be involved in mediating Na(+) transport.

  16. Overexpression of Arabidopsis molybdenum cofactor sulfurase gene confers drought tolerance in maize (Zea mays L.).

    PubMed

    Lu, Yao; Li, Yajun; Zhang, Jiachang; Xiao, Yitao; Yue, Yuesen; Duan, Liusheng; Zhang, Mingcai; Li, Zhaohu

    2013-01-01

    Abscisic acid (ABA) is a key component of the signaling system that integrates plant adaptive responses to abiotic stress. Overexpression of Arabidopsis molybdenum cofactor sulfurase gene (LOS5) in maize markedly enhanced the expression of ZmAO and aldehyde oxidase (AO) activity, leading to ABA accumulation and increased drought tolerance. Transgenic maize (Zea mays L.) exhibited the expected reductions in stomatal aperture, which led to decreased water loss and maintenance of higher relative water content (RWC) and leaf water potential. Also, transgenic maize subjected to drought treatment exhibited lower leaf wilting, electrolyte leakage, malondialdehyde (MDA) and H(2)O(2) content, and higher activities of antioxidative enzymes and proline content compared to wild-type (WT) maize. Moreover, overexpression of LOS5 enhanced the expression of stress-regulated genes such as Rad 17, NCED1, CAT1, and ZmP5CS1 under drought stress conditions, and increased root system development and biomass yield after re-watering. The increased drought tolerance in transgenic plants was associated with ABA accumulation via activated AO and expression of stress-related gene via ABA induction, which sequentially induced a set of favorable stress-related physiological and biochemical responses.

  17. Overexpression of Arabidopsis Molybdenum Cofactor Sulfurase Gene Confers Drought Tolerance in Maize (Zea mays L.)

    PubMed Central

    Zhang, Jiachang; Xiao, Yitao; Yue, Yuesen; Duan, Liusheng; Zhang, Mingcai; Li, Zhaohu

    2013-01-01

    Abscisic acid (ABA) is a key component of the signaling system that integrates plant adaptive responses to abiotic stress. Overexpression of Arabidopsis molybdenum cofactor sulfurase gene (LOS5) in maize markedly enhanced the expression of ZmAO and aldehyde oxidase (AO) activity, leading to ABA accumulation and increased drought tolerance. Transgenic maize (Zea mays L.) exhibited the expected reductions in stomatal aperture, which led to decreased water loss and maintenance of higher relative water content (RWC) and leaf water potential. Also, transgenic maize subjected to drought treatment exhibited lower leaf wilting, electrolyte leakage, malondialdehyde (MDA) and H2O2 content, and higher activities of antioxidative enzymes and proline content compared to wild-type (WT) maize. Moreover, overexpression of LOS5 enhanced the expression of stress-regulated genes such as Rad 17, NCED1, CAT1, and ZmP5CS1 under drought stress conditions, and increased root system development and biomass yield after re-watering. The increased drought tolerance in transgenic plants was associated with ABA accumulation via activated AO and expression of stress-related gene via ABA induction, which sequentially induced a set of favorable stress-related physiological and biochemical responses. PMID:23326325

  18. Regulatory network analysis reveals novel regulators of seed desiccation tolerance in Arabidopsis thaliana

    PubMed Central

    González-Morales, Sandra Isabel; Chávez-Montes, Ricardo A.; Hayano-Kanashiro, Corina; Alejo-Jacuinde, Gerardo; Rico-Cambron, Thelma Y.; de Folter, Stefan; Herrera-Estrella, Luis

    2016-01-01

    Desiccation tolerance (DT) is a remarkable process that allows seeds in the dry state to remain viable for long periods of time that in some instances exceed 1,000 y. It has been postulated that seed DT evolved by rewiring the regulatory and signaling networks that controlled vegetative DT, which itself emerged as a crucial adaptive trait of early land plants. Understanding the networks that regulate seed desiccation tolerance in model plant systems would provide the tools to understand an evolutionary process that played a crucial role in the diversification of flowering plants. In this work, we used an integrated approach that included genomics, bioinformatics, metabolomics, and molecular genetics to identify and validate molecular networks that control the acquisition of DT in Arabidopsis seeds. Two DT-specific transcriptional subnetworks were identified related to storage of reserve compounds and cellular protection mechanisms that act downstream of the embryo development master regulators LEAFY COTYLEDON 1 and 2, FUSCA 3, and ABSCICIC ACID INSENSITIVE 3. Among the transcription factors identified as major nodes in the DT regulatory subnetworks, PLATZ1, PLATZ2, and AGL67 were confirmed by knockout mutants and overexpression in a desiccation-intolerant mutant background to play an important role in seed DT. Additionally, we found that constitutive expression of PLATZ1 in WT plants confers partial DT in vegetative tissues. PMID:27551092

  19. Prefoldins 3 and 5 play an essential role in Arabidopsis tolerance to salt stress.

    PubMed

    Rodríguez-Milla, Miguel A; Salinas, Julio

    2009-05-01

    During the last years, our understanding of the mechanisms that control plant response to salt stress has been steadily progressing. Pharmacological studies have allowed the suggestion that the cytoskeleton may be involved in regulating such a response. Nevertheless, genetic evidence establishing that the cytoskeleton has a role in plant tolerance to salt stress has not been reported yet. Here, we have characterized Arabidopsis T-DNA mutants for genes encoding proteins orthologous to prefoldin (PFD) subunits 3 and 5 from yeast and mammals. In these organisms, PFD subunits, also known as Genes Involved in Microtubule biogenesis (GIM), form a heterohexameric PFD complex implicated in tubulin and actin folding. We show that, indeed, PFD3 and PFD5 can substitute for the loss of their yeast orthologs, as they are able to complement yeast gim2Delta and gim5Delta mutants, respectively. Our results indicate that pfd3 and pfd5 mutants have reduced levels of alpha- and beta-tubulin compared to the wild-type plants when growing under both control and salt-stress conditions. In addition, pfd3 and pfd5 mutants display alterations in their developmental patterns and microtubule organization, and, more importantly, are hypersensitive to high concentrations of NaCl but not of LiCl or mannitol. These results demonstrate that the cytoskeleton plays an essential role in plant tolerance to salt stress.

  20. GmGBP1, a homolog of human ski interacting protein in soybean, regulates flowering and stress tolerance in Arabidopsis

    PubMed Central

    2013-01-01

    Background SKIP is a transcription cofactor in many eukaryotes. It can regulate plant stress tolerance in rice and Arabidopsis. But the homolog of SKIP protein in soybean has been not reported up to now. Results In this study, the expression patterns of soybean GAMYB binding protein gene (GmGBP1) encoding a homolog of SKIP protein were analyzed in soybean under abiotic stresses and different day lengths. The expression of GmGBP1 was induced by polyethyleneglycol 6000, NaCl, gibberellin, abscisic acid and heat stress. GmGBP1 had transcriptional activity in C-terminal. GmGBP1 could interact with R2R3 domain of GmGAMYB1 in SKIP domain to take part in gibberellin flowering pathway. In long-day (16 h-light) condition, transgenic Arabidopsis with the ectopic overexpression of GmGBP1 exhibited earlier flowering and less number of rosette leaves; Suppression of AtSKIP in Arabidopsis resulted in growth arrest, flowering delay and down-regulation of many flowering-related genes (CONSTANS, FLOWERING LOCUS T, LEAFY); Arabidopsis myb33 mutant plants with ectopic overexpression of GmGBP1 showed the same flowering phenotype with wild type. In short-day (8 h-light) condition, transgenic Arabidopsis plants with GmGBP1 flowered later and showed a higher level of FLOWERING LOCUS C compared with wild type. When treated with abiotic stresses, transgenic Arabidopsis with the ectopic overexpression of GmGBP1 enhanced the tolerances to heat and drought stresses but reduced the tolerance to high salinity, and affected the expressions of several stress-related genes. Conclusions In Arabidopsis, GmGBP1 might positively regulate the flowering time by affecting CONSTANS, FLOWERING LOCUS T, LEAFY and GAMYB directly or indirectly in photoperiodic and gibberellin pathways in LDs, but GmGBP1 might represse flowering by affecting FLOWERING LOCUS C and SHORT VEGETATIVE PHASE in autonomous pathway in SDs. GmGBP1 might regulate the activity of ROS-eliminating to improve the resistance to heat and

  1. Hv-CBF2A overexpression in barley accelerates COR gene transcript accumulation and acquisition of freezing tolerance during cold acclimation.

    PubMed

    Jeknić, Zoran; Pillman, Katherine A; Dhillon, Taniya; Skinner, Jeffrey S; Veisz, Ottó; Cuesta-Marcos, Alfonso; Hayes, Patrick M; Jacobs, Andrew K; Chen, Tony H H; Stockinger, Eric J

    2014-01-01

    C-Repeat Binding Factors (CBFs) are DNA-binding transcriptional activators of gene pathways imparting freezing tolerance. Poaceae contain three CBF subfamilies, two of which, HvCBF3/CBFIII and HvCBF4/CBFIV, are unique to this taxon. To gain mechanistic insight into HvCBF4/CBFIV CBFs we overexpressed Hv-CBF2A in spring barley (Hordeum vulgare) cultivar 'Golden Promise'. The Hv-CBF2A overexpressing lines exhibited stunted growth, poor yield, and greater freezing tolerance compared to non-transformed 'Golden Promise'. Differences in freezing tolerance were apparent only upon cold acclimation. During cold acclimation freezing tolerance of the Hv-CBF2A overexpressing lines increased more rapidly than that of 'Golden Promise' and paralleled the freezing tolerance of the winter hardy barley 'Dicktoo'. Transcript levels of candidate CBF target genes, COR14B and DHN5 were increased in the overexpressor lines at warm temperatures, and at cold temperatures they accumulated to much higher levels in the Hv-CBF2A overexpressors than in 'Golden Promise'. Hv-CBF2A overexpression also increased transcript levels of other CBF genes at FROST RESISTANCE-H2-H2 (FR-H2) possessing CRT/DRE sites in their upstream regions, the most notable of which was CBF12. CBF12 transcript levels exhibited a relatively constant incremental increase above levels in 'Golden Promise' both at warm and cold. These data indicate that Hv-CBF2A activates target genes at warm temperatures and that transcript accumulation for some of these targets is greatly enhanced by cold temperatures.

  2. Seed tolerance to deterioration in arabidopsis is affected by virus infection.

    PubMed

    Bueso, Eduardo; Serrano, Ramón; Pallás, Vicente; Sánchez-Navarro, Jesús A

    2017-07-01

    Seed longevity is the period during which the plant seed is able to germinate. This property is strongly influenced by environment conditions experienced by seeds during their formation and storage. In the present study we have analyzed how the biotic stress derived from the infection of Cauliflower mosaic virus (CaMV), Turnip mosaic virus (TuMV), Cucumber mosaic virus (CMV) and Alfalfa mosaic virus (AMV) affects seed tolerance to deterioration measuring germination rates after an accelerated aging treatment. Arabidopsis wild type plants infected with AMV and CMV rendered seeds with improved tolerance to deterioration when compared to the non-inoculated plants. On the other hand, CaMV infection generated seeds more sensitive to deterioration. No seeds were obtained from TuMV infected plants. Similar pattern of viral effects was observed in the double mutant athb22 athb25, which is more sensitive to accelerated seed aging than wild type. However, we observed a significant reduction of the seed germination for CMV (65% vs 55%) and healthy (50% vs 30%) plants in these mutants. The seed quality differences were overcomed using the A. thaliana athb25-1D dominant mutant, which over accumulated gibberellic acid (GA), except for TuMV which generated some siliques with low seed tolerance to deterioration. For AMV and TuMV (in athb25-1D), the seed quality correlated with the accumulation of the messengers of the gibberellin 3-oxidase family, the mucilage of the seed and the GA1. For CMV and CaMV it was not a good correlation suggesting that other factors are affecting seed viability. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

  3. Expression of wild soybean WRKY20 in Arabidopsis enhances drought tolerance and regulates ABA signalling.

    PubMed

    Luo, Xiao; Bai, Xi; Sun, Xiaoli; Zhu, Dan; Liu, Baohui; Ji, Wei; Cai, Hua; Cao, Lei; Wu, Jing; Hu, Mengran; Liu, Xin; Tang, Lili; Zhu, Yanming

    2013-05-01

    The WRKY-type transcription factors are involved in plant development and stress responses, but how the regulation of stress tolerance is related to plant development is largely unknown. GsWRKY20 was initially identified as a stress response gene using large-scale Glycine soja microarrays. Quantitative reverse transcription-PCR (qRT-PCR) showed that the expression of this gene was induced by abscisic acid (ABA), salt, cold, and drought. Overexpression of GsWRKY20 in Arabidopsis resulted in a decreased sensitivity to ABA during seed germination and early seedling growth. However, compared with the wild type, GsWRKY20 overexpression lines were more sensitive to ABA in stomatal closure, and exhibited a greater tolerance to drought stress, a decreased water loss rate, and a decreased stomatal density. Moreover, microarray and qRT-PCR assays showed that GsWRKY20 mediated ABA signalling by promoting the expression of negative regulators of ABA signalling, such as AtWRKY40, ABI1, and ABI2, while repressing the expression of the positive regulators of ABA, for example ABI5, ABI4, and ABF4. Interestingly, GsWRKY20 also positively regulates the expression of a group of wax biosynthetic genes. Further, evidence is provided to support that GsWRKY20 overexpression lines have more epicuticular wax crystals and a much thicker cuticle, which contribute to less chlorophyll leaching compared with the wild type. Taken together, the findings reveal an important role for GsWRKY20 in enhancing drought tolerance and regulating ABA signalling.

  4. Constitutive S-adenosylmethionine decarboxylase gene expression increases drought tolerance through inhibition of reactive oxygen species accumulation in Arabidopsis.

    PubMed

    Wi, Soo Jin; Kim, Soo Jin; Kim, Woo Taek; Park, Ky Young

    2014-05-01

    Using subtractive hybridization analysis, the S-adenosylmethionine decarboxylase (SAMDC) gene from Capsicum annuum was isolated and renamed CaSAMDC. We generated independent transgenic Arabidopsis (Arabidopsis thaliana) lines constitutively expressing a 35S::CaSAMDC construct. Drought tolerance was significantly enhanced in Arabidopsis T4 transgenic homozygous lines as compared to wild-type (WT) plants. The levels of main polyamines (PAs) were more significantly increased in CaSAMDC-overexpressing transgenic plants after 6 h of drought stress as compared to stressed WT plants. Basal transcription of polyamine oxidase (PAO) showed at a much higher level in unstressed-transgenic plants as compared to unstressed WT plants. However, the difference in PAO transcription level between WT and transgenic plants was reduced after drought stress. Cellular accumulation of reactive oxygen species (ROS) was significantly reduced following drought stress in transgenic Arabidopsis plants as compared to WT plants. These results were in agreement with additional observations that stress-induced ROS generation, as determined by qRT-PCR analysis of NADPH oxidase (RbohD and RbohF), was significantly suppressed while transcription of ROS-detoxifying enzymes was notably elevated in transgenic lines in response to drought stress. Further, ROS-induced transcription of the metacaspase II gene was remarkably inhibited in transgenic plants. Collectively, these results suggest that drought stress tolerance due to reduction of ROS production and enhancement of ROS detoxification can be attributed to elevation of PAs.

  5. Improvement of water use efficiency in rice by expression of HARDY, an Arabidopsis drought and salt tolerance gene

    PubMed Central

    Karaba, Aarati; Dixit, Shital; Greco, Raffaella; Aharoni, Asaph; Trijatmiko, Kurniawan R.; Marsch-Martinez, Nayelli; Krishnan, Arjun; Nataraja, Karaba N.; Udayakumar, Makarla; Pereira, Andy

    2007-01-01

    Freshwater is a limited and dwindling global resource; therefore, efficient water use is required for food crops that have high water demands, such as rice, or for the production of sustainable energy biomass. We show here that expression of the Arabidopsis HARDY (HRD) gene in rice improves water use efficiency, the ratio of biomass produced to the water used, by enhancing photosynthetic assimilation and reducing transpiration. These drought-tolerant, low-water-consuming rice plants exhibit increased shoot biomass under well irrigated conditions and an adaptive increase in root biomass under drought stress. The HRD gene, an AP2/ERF-like transcription factor, identified by a gain-of-function Arabidopsis mutant hrd-D having roots with enhanced strength, branching, and cortical cells, exhibits drought resistance and salt tolerance, accompanied by an enhancement in the expression of abiotic stress associated genes. HRD overexpression in Arabidopsis produces thicker leaves with more chloroplast-bearing mesophyll cells, and in rice, there is an increase in leaf biomass and bundle sheath cells that probably contributes to the enhanced photosynthesis assimilation and efficiency. The results exemplify application of a gene identified from the model plant Arabidopsis for the improvement of water use efficiency coincident with drought resistance in the crop plant rice. PMID:17881564

  6. A DEAD Box RNA Helicase Is Critical for Pre-mRNA Splicing, Cold-Responsive Gene Regulation, and Cold Tolerance in Arabidopsis[C][W

    PubMed Central

    Guan, Qingmei; Wu, Jianmin; Zhang, Yanyan; Jiang, Changhua; Liu, Renyi; Chai, Chenglin; Zhu, Jianhua

    2013-01-01

    Cold stress resulting from chilling and freezing temperatures substantially reduces crop production worldwide. To identify genes critical for cold tolerance in plants, we screened Arabidopsis thaliana mutants for deregulated expression of a firefly luciferase reporter gene under the control of the C-REPEAT BINDING FACTOR2 (CBF2) promoter (CBF2:LUC). A regulator of CBF gene expression1 (rcf1-1) mutant that is hypersensitive to cold stress was chosen for in-depth characterization. RCF1 encodes a cold-inducible DEAD (Asp-Glu-Ala-Asp) box RNA helicase. Unlike a previously reported DEAD box RNA helicase (LOW EXPRESSION OF OSMOTICALLY RESPONSIVE GENES4 [LOS4]) that regulates mRNA export, RCF1 does not play a role in mRNA export. Instead, RCF1 functions to maintain proper splicing of pre-mRNAs; many cold-responsive genes are mis-spliced in rcf1-1 mutant plants under cold stress. Functional characterization of four genes (PSEUDO-RESPONSE REGULATOR5 [PRR5], SHAGGY-LIKE SERINE/THREONINE KINASE12 [SK12], MYB FAMILY TRANSCRIPTION FACTOR CIRCADIAN1 [CIR1], and SPFH/PHB DOMAIN-CONTAINING MEMBRANE-ASSOCIATED PROTEIN [SPFH]) that are mis-spliced in rcf1-1 revealed that these genes are cold-inducible positive (CIR1 and SPFH) and negative (PRR5 and SK12) regulators of cold-responsive genes and cold tolerance. Together, our results suggest that the cold-inducible RNA helicase RCF1 is essential for pre-mRNA splicing and is important for cold-responsive gene regulation and cold tolerance in plants. PMID:23371945

  7. Suppressed expression of the apoplastic ascorbate oxidase gene increases salt tolerance in tobacco and Arabidopsis plants.

    PubMed

    Yamamoto, Atsuko; Bhuiyan, Md Nazmul H; Waditee, Rungaroon; Tanaka, Yoshito; Esaka, Muneharu; Oba, Kazuko; Jagendorf, André T; Takabe, Teruhiro

    2005-07-01

    Transgenic tobacco plants expressing the ascorbate oxidase (AAO) gene in sense and antisense orientations, and an Arabidopsis mutant in which the T-DNA was inserted into a putative AAO gene, were used to examine the potential roles of AAO for salt-stress tolerance in plants. AAO activities in the transgenic tobacco plants expressing the gene in sense and antisense orientations were, respectively, about 16-fold and 0.2-fold of those in the wild type. Under normal growth conditions, no significant differences in phenotypes were observed, except for a delay in flowering time in the antisense plants. However, at high salinity, the percentage germination, photosynthetic activity, and seed yields were higher in antisense plants, with progressively lower levels in the wild type and the sense plants. The redox state of apoplastic ascorbate in sense plants was very low even under normal growth conditions. Upon salt stress, the redox state of symplastic and apoplastic ascorbate decreased among the three types of plants, but was lowest in the sense plants. The hydrogen peroxide contents in the symplastic and apoplastic spaces were higher in sense plants, progressively lower in the wild type, followed by the antisense plants. The Arabidopsis T-DNA inserted mutant exhibited very low ascorbate oxidase activity, and its phenotype was similar to that of antisense tobacco plants. These results suggest that the suppressed expression of apoplastic AAO under salt-stress conditions leads to a relatively low level of hydrogen peroxide accumulation and a high redox state of symplastic and apoplastic ascorbate which, in turn, permits a higher seed yield.

  8. Splicing factor SR34b mutation reduces cadmium tolerance in Arabidopsis by regulating iron-regulated transporter 1 gene

    SciTech Connect

    Zhang, Wentao; Du, Bojing; Liu, Di; Qi, Xiaoting

    2014-12-12

    Highlights: • Arabidopsis splicing factor SR34b gene is cadmium-inducible. • SR34b T-DNA insertion mutant is sensitive to cadmium due to high cadmium uptake. • SR34b is a regulator of cadmium transporter IRT1 at the posttranscription level. • These results highlight the roles of splicing factors in cadmium tolerance of plant. - Abstract: Serine/arginine-rich (SR) proteins are important splicing factors. However, the biological functions of plant SR proteins remain unclear especially in abiotic stresses. Cadmium (Cd) is a non-essential element that negatively affects plant growth and development. In this study, we provided clear evidence for SR gene involved in Cd tolerance in planta. Systemic expression analysis of 17 Arabidopsis SR genes revealed that SR34b is the only SR gene upregulated by Cd, suggesting its potential roles in Arabidopsis Cd tolerance. Consistent with this, a SR34b T-DNA insertion mutant (sr34b) was moderately sensitive to Cd, which had higher Cd{sup 2+} uptake rate and accumulated Cd in greater amounts than wild-type. This was due to the altered expression of iron-regulated transporter 1 (IRT1) gene in sr34b mutant. Under normal growth conditions, IRT1 mRNAs highly accumulated in sr34b mutant, which was a result of increased stability of IRT1 mRNA. Under Cd stress, however, sr34b mutant plants had a splicing defect in IRT1 gene, thus reducing the IRT1 mRNA accumulation. Despite of this, sr34b mutant plants still constitutively expressed IRT1 proteins under Cd stress, thereby resulting in Cd stress-sensitive phenotype. We therefore propose the essential roles of SR34b in posttranscriptional regulation of IRT1 expression and identify it as a regulator of Arabidopsis Cd tolerance.

  9. The effect of cold acclimation on the water relations and freezing tolerance of Hordeum vulgare L.

    PubMed

    Burchett, S; Niven, S; Fuller, M P

    2006-01-01

    During a 5 degree C and a 5/-1 degree C cold acclimation (CA) regime there was a significant decline in the water potential of winter barley, and a concurrent decline in tissue water content of the 5/-1 degree C CA plants. Results of carbohydrate analysis illustrated a significant (P < 0.001) accumulation of sucrose, fructose and glucose in the 5/-1 degree C CA plants, which was inversely correlated to water potential. Using an infrared imaging radiometer during a convection frost test the water release time (WRT) of 5/-1 degree C CA was demonstrated to be significantly (P < 0.001) longer than that observed in non-cold acclimated plants. This observation is consistent with visual analysis of exotherm curves where the rate of cellular water release to extracellular ice is reduced in the 5/-1 degree C CA plants, compared to the non-cold acclimated plants. These biochemical and physiological changes were correlated to increased plant health following a non-lethal freezing test to -5 degree C, where non-cold acclimated plants produced 2.3 +/- 0.3 tillers and 5 degree C and 5/-1 degree C CA plants produced 2.4 +/- 0.3 and 4.7 +/- 0.7 tillers, respectively. Results from this study imply that cold acclimation leads to changes in the physical state of water that result in a less osmotically responsive cellular environment and subsequently significantly less damage to meristematic tissue.

  10. Arabidopsis fatty acid desaturase FAD2 is required for salt tolerance during seed germination and early seedling growth.

    PubMed

    Zhang, Jiantao; Liu, Hua; Sun, Jian; Li, Bei; Zhu, Qiang; Chen, Shaoliang; Zhang, Hongxia

    2012-01-01

    Fatty acid desaturases play important role in plant responses to abiotic stresses. However, their exact function in plant resistance to salt stress is unknown. In this work, we provide the evidence that FAD2, an endoplasmic reticulum localized ω-6 desaturase, is required for salt tolerance in Arabidopsis. Using vacuolar and plasma membrane vesicles prepared from the leaves of wild-type (Col-0) and the loss-of-function Arabidopsis mutant, fad2, which lacks the functional FAD2, we examined the fatty acid composition and Na+-dependent H+ movements of the isolated vesicles. We observed that, when compared to Col-0, the level of vacuolar and plasma membrane polyunsaturation was lower, and the Na+/H+ exchange activity was reduced in vacuolar and plasma membrane vesicles isolated from fad2 mutant. Consistent with the reduced Na+/H+ exchange activity, fad2 accumulated more Na+ in the cytoplasm of root cells, and was more sensitive to salt stress during seed germination and early seedling growth, as indicated by CoroNa-Green staining, net Na+ efflux and salt tolerance analyses. Our results suggest that FAD2 mediated high-level vacuolar and plasma membrane fatty acid desaturation is essential for the proper function of membrane attached Na+/H+ exchangers, and thereby to maintain a low cytosolic Na+ concentration for salt tolerance during seed germination and early seedling growth in Arabidopsis.

  11. Overproduction of the membrane-bound receptor-like protein kinase 1, RPK1, enhances abiotic stress tolerance in Arabidopsis.

    PubMed

    Osakabe, Yuriko; Mizuno, Shinji; Tanaka, Hidenori; Maruyama, Kyonoshin; Osakabe, Keishi; Todaka, Daisuke; Fujita, Yasunari; Kobayashi, Masatomo; Shinozaki, Kazuo; Yamaguchi-Shinozaki, Kazuko

    2010-03-19

    RPK1 (receptor-like protein kinase 1) localizes to the plasma membrane and functions as a regulator of abscisic acid (ABA) signaling in Arabidopsis. In our current study, we investigated the effect of RPK1 disruption and overproduction upon plant responses to drought stress. Transgenic Arabidopsis overexpressing the RPK1 protein showed increased ABA sensitivity in their root growth and stomatal closure and also displayed less transpirational water loss. In contrast, a mutant lacking RPK1 function, rpk1-1, was found to be resistant to ABA during these processes and showed increased water loss. RPK1 overproduction in these transgenic plants thus increased their tolerance to drought stress. We performed microarray analysis of RPK1 transgenic plants and observed enhanced expression of several stress-responsive genes, such as Cor15a, Cor15b, and rd29A, in addition to H(2)O(2)-responsive genes. Consistently, the expression levels of ABA/stress-responsive genes in rpk1-1 had decreased compared with wild type. The results suggest that the overproduction of RPK1 enhances both the ABA and drought stress signaling pathways. Furthermore, the leaves of the rpk1-1 plants exhibit higher sensitivity to oxidative stress upon ABA-pretreatment, whereas transgenic plants overproducing RPK1 manifest increased tolerance to this stress. Our current data suggest therefore that RPK1 overproduction controls reactive oxygen species homeostasis and enhances both water and oxidative stress tolerance in Arabidopsis.

  12. Physiological and molecular characterization of the enhanced salt tolerance induced by low-dose gamma irradiation in Arabidopsis seedlings

    SciTech Connect

    Qi, Wencai; Zhang, Liang; Xu, Hangbo; Wang, Lin; Jiao, Zhen

    2014-07-25

    Highlights: • 50-Gy gamma irradiation markedly promotes the seedling growth under salt stress in Arabidopsis. • The contents of H{sub 2}O{sub 2} and MDA are obviously reduced by low-dose gamma irradiation under salt stress. • Low-dose gamma irradiation stimulates the activities of antioxidant enzymes under salt stress. • Proline accumulation is required for the low-gamma-ray-induced salt tolerance. • Low gamma rays differentially regulate the expression of genes related to salt stress. - Abstract: It has been established that gamma rays at low doses stimulate the tolerance to salt stress in plants. However, our knowledge regarding the molecular mechanism underlying the enhanced salt tolerance remains limited. In this study, we found that 50-Gy gamma irradiation presented maximal beneficial effects on germination index and root length in response to salt stress in Arabidopsis seedlings. The contents of H{sub 2}O{sub 2} and MDA in irradiated seedlings under salt stress were significantly lower than those of controls. The activities of antioxidant enzymes and proline levels in the irradiated seedlings were markedly increased compared with the controls. Furthermore, transcriptional expression analysis of selected genes revealed that some components of salt stress signaling pathways were stimulated by low-dose gamma irradiation under salt stress. Our results suggest that gamma irradiation at low doses alleviates the salt stress probably by modulating the physiological responses as well as stimulating the stress signal transduction in Arabidopsis seedlings.

  13. The Opuntia streptacantha OpsHSP18 gene confers salt and osmotic stress tolerance in Arabidopsis thaliana.

    PubMed

    Salas-Muñoz, Silvia; Gómez-Anduro, Gracia; Delgado-Sánchez, Pablo; Rodríguez-Kessler, Margarita; Jiménez-Bremont, Juan Francisco

    2012-01-01

    Abiotic stress limits seed germination, plant growth, flowering and fruit quality, causing economic decrease. Small Heat Shock Proteins (sHSPs) are chaperons with roles in stress tolerance. Herein, we report the functional characterization of a cytosolic class CI sHSP (OpsHSP18) from Opuntia streptacantha during seed germination in Arabidopsis thaliana transgenic lines subjected to different stress and hormone treatments. The over-expression of the OpsHSP18 gene in A. thaliana increased the seed germination rate under salt (NaCl) and osmotic (glucose and mannitol) stress, and in ABA treatments, compared with WT. On the other hand, the over-expression of the OpsHSP18 gene enhanced tolerance to salt (150 mM NaCl) and osmotic (274 mM mannitol) stress in Arabidopsis seedlings treated during 14 and 21 days, respectively. These plants showed increased survival rates (52.00 and 73.33%, respectively) with respect to the WT (18.75 and 53.75%, respectively). Thus, our results show that OpsHSP18 gene might have an important role in abiotic stress tolerance, in particular in seed germination and survival rate of Arabidopsis plants under unfavorable conditions.

  14. Splicing factor SR34b mutation reduces cadmium tolerance in Arabidopsis by regulating iron-regulated transporter 1 gene.

    PubMed

    Zhang, Wentao; Du, Bojing; Liu, Di; Qi, Xiaoting

    2014-12-12

    Serine/arginine-rich (SR) proteins are important splicing factors. However, the biological functions of plant SR proteins remain unclear especially in abiotic stresses. Cadmium (Cd) is a non-essential element that negatively affects plant growth and development. In this study, we provided clear evidence for SR gene involved in Cd tolerance in planta. Systemic expression analysis of 17 Arabidopsis SR genes revealed that SR34b is the only SR gene upregulated by Cd, suggesting its potential roles in Arabidopsis Cd tolerance. Consistent with this, a SR34b T-DNA insertion mutant (sr34b) was moderately sensitive to Cd, which had higher Cd(2+) uptake rate and accumulated Cd in greater amounts than wild-type. This was due to the altered expression of iron-regulated transporter 1 (IRT1) gene in sr34b mutant. Under normal growth conditions, IRT1 mRNAs highly accumulated in sr34b mutant, which was a result of increased stability of IRT1 mRNA. Under Cd stress, however, sr34b mutant plants had a splicing defect in IRT1 gene, thus reducing the IRT1 mRNA accumulation. Despite of this, sr34b mutant plants still constitutively expressed IRT1 proteins under Cd stress, thereby resulting in Cd stress-sensitive phenotype. We therefore propose the essential roles of SR34b in posttranscriptional regulation of IRT1 expression and identify it as a regulator of Arabidopsis Cd tolerance.

  15. The Opuntia streptacantha OpsHSP18 Gene Confers Salt and Osmotic Stress Tolerance in Arabidopsis thaliana

    PubMed Central

    Salas-Muñoz, Silvia; Gómez-Anduro, Gracia; Delgado-Sánchez, Pablo; Rodríguez-Kessler, Margarita; Jiménez-Bremont, Juan Francisco

    2012-01-01

    Abiotic stress limits seed germination, plant growth, flowering and fruit quality, causing economic decrease. Small Heat Shock Proteins (sHSPs) are chaperons with roles in stress tolerance. Herein, we report the functional characterization of a cytosolic class CI sHSP (OpsHSP18) from Opuntia streptacantha during seed germination in Arabidopsis thaliana transgenic lines subjected to different stress and hormone treatments. The over-expression of the OpsHSP18 gene in A. thaliana increased the seed germination rate under salt (NaCl) and osmotic (glucose and mannitol) stress, and in ABA treatments, compared with WT. On the other hand, the over-expression of the OpsHSP18 gene enhanced tolerance to salt (150 mM NaCl) and osmotic (274 mM mannitol) stress in Arabidopsis seedlings treated during 14 and 21 days, respectively. These plants showed increased survival rates (52.00 and 73.33%, respectively) with respect to the WT (18.75 and 53.75%, respectively). Thus, our results show that OpsHSP18 gene might have an important role in abiotic stress tolerance, in particular in seed germination and survival rate of Arabidopsis plants under unfavorable conditions. PMID:22949853

  16. Characterization of Arabidopsis thaliana cDNAs that render yeasts tolerant toward the thiol-oxidizing drug diamide.

    PubMed Central

    Kushnir, S; Babiychuk, E; Kampfenkel, K; Belles-Boix, E; Van Montagu, M; Inzé, D

    1995-01-01

    Diamide oxidizes cellular thiols and induces oxidative stress. To isolate plant genes which may, when overexpressed, increase tolerance of plants toward oxidative damage, an in vivo diamide tolerance screening in yeasts was used. An Arabidopsis cDNA library in a yeast expression vector was used to transform a yeast strain with intact antioxidant defense. Cells from approximately 10(5) primary transformants were selected for resistance to diamide. Three Arabidopsis cDNAs which confer diamide tolerance were isolated. This drug tolerance was specific and no cross tolerance toward hydroperoxides was found. One cDNA (D3) encodes a polypeptide which has an amino-terminal J domain characteristic of a divergent family of DnaJ chaperones. Another (D18) encodes a putative dTDP-D-glucose 4,6-dehydratase. Surprisingly, the third cDNA (D22) encodes a plant homolog of gamma-glutamyltransferases. It would have been difficult to predict that the expression of those genes would lead to an improved survival under conditions of depletion of cellular thiols. Hence, we suggest that this cloning approach may be a useful contribution to the isolation of plant genes that can help to cope with oxidative stress. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 Fig. 5 Fig. 6 PMID:7479844

  17. Development, molecular composition and freeze tolerance of bovine embryos cultured in TCM-199 supplemented with hyaluronan.

    PubMed

    Palasz, A T; Breña, P Beltrán; Martinez, Marcelo F; Perez-Garnelo, S S; Ramirez, M A; Gutiérrez-Adán, A; De la Fuente, J

    2008-02-01

    Hyaluronan (HA) is glycosaminoglycan that is present from the start of embryonic development and its role and concentration increases with embryo development. The objective of this study was to evaluate if the presence of HA in TCM-199 culture medium had an effect on the development and quality of bovine embryos. There was no effect of HA on the total number of zygotes developing to blastocysts on day 7, however more expanded and hatched blastocyst stages were observed on days 8 and 9 in the group supplemented with HA (p<0.05). Following freeze/thawing, significantly more (p<0.05) embryos cultured in medium supplemented with HA hatched than those cultured in TCM-199 alone or those with BSA. Medium supplemented with HA and BSA significantly increased the level of expression of glucose metabolism Glut-1 gene and embryo compaction Cx43 gene (p<0.05), and had no effect on Glut-5 and IGF-II expression. In addition, HA presence in culture decreased the level of expression of apoptosis Bax and oxidative stress SOX genes (p<0.05). There was significant difference in total number of nuclei between TCM-199 medium only and the remaining media containing BSA or HA plus BSA, between which there was no difference. In summary, our results indicate that the addition of high molecular weight HA to TCM-199 medium that contains BSA on day 4 of culture improved embryo development to hatching and hatched blastocysts and the quality of produced embryos, which were superior to embryos cultured without HA addition.

  18. Arsenic tolerance in Arabidopsis is mediated by two ABCC-type phytochelatin transporters

    PubMed Central

    Song, Won-Yong; Park, Jiyoung; Mendoza-Cózatl, David G.; Suter-Grotemeyer, Marianne; Shim, Donghwan; Hörtensteiner, Stefan; Geisler, Markus; Weder, Barbara; Rea, Philip A.; Rentsch, Doris; Schroeder, Julian I.; Lee, Youngsook; Martinoia, Enrico

    2010-01-01

    Arsenic is an extremely toxic metalloid causing serious health problems. In Southeast Asia, aquifers providing drinking and agricultural water for tens of millions of people are contaminated with arsenic. To reduce nutritional arsenic intake through the consumption of contaminated plants, identification of the mechanisms for arsenic accumulation and detoxification in plants is a prerequisite. Phytochelatins (PCs) are glutathione-derived peptides that chelate heavy metals and metalloids such as arsenic, thereby functioning as the first step in their detoxification. Plant vacuoles act as final detoxification stores for heavy metals and arsenic. The essential PC–metal(loid) transporters that sequester toxic metal(loid)s in plant vacuoles have long been sought but remain unidentified in plants. Here we show that in the absence of two ABCC-type transporters, AtABCC1 and AtABCC2, Arabidopsis thaliana is extremely sensitive to arsenic and arsenic-based herbicides. Heterologous expression of these ABCC transporters in phytochelatin-producing Saccharomyces cerevisiae enhanced arsenic tolerance and accumulation. Furthermore, membrane vesicles isolated from these yeasts exhibited a pronounced arsenite [As(III)]–PC2 transport activity. Vacuoles isolated from atabcc1 atabcc2 double knockout plants exhibited a very low residual As(III)–PC2 transport activity, and interestingly, less PC was produced in mutant plants when exposed to arsenic. Overexpression of AtPCS1 and AtABCC1 resulted in plants exhibiting increased arsenic tolerance. Our findings demonstrate that AtABCC1 and AtABCC2 are the long-sought and major vacuolar PC transporters. Modulation of vacuolar PC transporters in other plants may allow engineering of plants suited either for phytoremediation or reduced accumulation of arsenic in edible organs. PMID:21078981

  19. Role of Abscisic Acid in the Induction of Desiccation Tolerance in Developing Seeds of Arabidopsis thaliana

    PubMed Central

    Meurs, Cor; Basra, Amarjit S.; Karssen, Cees M.; van Loon, Leendert C.

    1992-01-01

    In contrast to wild-type seeds of Arabidopsis thaliana and to seeds deficient in (aba) or insensitive to (abi3) abscisic acid (ABA), maturing seeds of recombinant (aba,abi3) plants fail to desiccate, remain green, and lose viability upon drying. These double-mutant seeds acquire only low levels of the major storage proteins and are deficient in several low mol wt polypeptides, both soluble and bound, and some of which are heat stable. A major heat-stable glycoprotein of more than 100 kilodaltons behaves similarly; during seed development, it shows a decrease in size associated with the abi3 mutation. In seeds of the double mutant from 14 to 20 days after pollination, the low amounts of various maturation-specific proteins disappear and many higher mol wt proteins similar to those occurring during germination are induced, but no visible germination is apparent. It appears that in the aba,abi3 double mutant seed development is not completed and the program for seed germination is initiated prematurely in the absence of substances protective against dehydration. Seeds may be made desiccation tolerant by watering the plants with the ABA analog LAB 173711 or by imbibition of isolated immature seeds, 11 to 15 days after pollination, with ABA and sucrose. Whereas sucrose stimulates germination and may protect dehydration-sensitive structures from desiccation damage, ABA inhibits precocious germination and is required to complete the program for seed maturation and the associated development of desiccation tolerance. ImagesFigure 1Figure 2Figure 4Figure 5Figure 6Figure 8 PMID:16668818

  20. Arsenic tolerance in Arabidopsis is mediated by two ABCC-type phytochelatin transporters.

    PubMed

    Song, Won-Yong; Park, Jiyoung; Mendoza-Cózatl, David G; Suter-Grotemeyer, Marianne; Shim, Donghwan; Hörtensteiner, Stefan; Geisler, Markus; Weder, Barbara; Rea, Philip A; Rentsch, Doris; Schroeder, Julian I; Lee, Youngsook; Martinoia, Enrico

    2010-12-07

    Arsenic is an extremely toxic metalloid causing serious health problems. In Southeast Asia, aquifers providing drinking and agricultural water for tens of millions of people are contaminated with arsenic. To reduce nutritional arsenic intake through the consumption of contaminated plants, identification of the mechanisms for arsenic accumulation and detoxification in plants is a prerequisite. Phytochelatins (PCs) are glutathione-derived peptides that chelate heavy metals and metalloids such as arsenic, thereby functioning as the first step in their detoxification. Plant vacuoles act as final detoxification stores for heavy metals and arsenic. The essential PC-metal(loid) transporters that sequester toxic metal(loid)s in plant vacuoles have long been sought but remain unidentified in plants. Here we show that in the absence of two ABCC-type transporters, AtABCC1 and AtABCC2, Arabidopsis thaliana is extremely sensitive to arsenic and arsenic-based herbicides. Heterologous expression of these ABCC transporters in phytochelatin-producing Saccharomyces cerevisiae enhanced arsenic tolerance and accumulation. Furthermore, membrane vesicles isolated from these yeasts exhibited a pronounced arsenite [As(III)]-PC(2) transport activity. Vacuoles isolated from atabcc1 atabcc2 double knockout plants exhibited a very low residual As(III)-PC(2) transport activity, and interestingly, less PC was produced in mutant plants when exposed to arsenic. Overexpression of AtPCS1 and AtABCC1 resulted in plants exhibiting increased arsenic tolerance. Our findings demonstrate that AtABCC1 and AtABCC2 are the long-sought and major vacuolar PC transporters. Modulation of vacuolar PC transporters in other plants may allow engineering of plants suited either for phytoremediation or reduced accumulation of arsenic in edible organs.

  1. The role of glutathione in mercury tolerance resembles its function under cadmium stress in Arabidopsis.

    PubMed

    Sobrino-Plata, Juan; Carrasco-Gil, Sandra; Abadía, Javier; Escobar, Carolina; Álvarez-Fernández, Ana; Hernández, Luis E

    2014-02-01

    Recent research efforts have highlighted the importance of glutathione (GSH) as a key antioxidant metabolite for metal tolerance in plants. Little is known about the mechanisms involved in stress due to mercury (Hg), one of the most hazardous metals to the environment and human health. To understand the implication of GSH metabolism for Hg tolerance, we used two γ-glutamylcysteine synthetase (γECS) Arabidopsis thaliana allele mutants (rax1-1 and cad2-1) and a phytochelatin synthase (PCS) mutant (cad1-3). The leaves of these mutants and of wild type (Col-0) were infiltrated with a solution containing Cd or Hg (0, 3 and 30 μM) and incubated for 24 and 48 h. The formation of phytochelatins (PCs) in the leaf extracts was followed by two different HPLC-based methods and occurred in Col-0, cad2-1 and rax1-1 plants exposed to Cd, whereas in the Hg treatments, PCs accumulated mainly in Col-0 and rax1-1, where Hg-PC complexes were also detected. ASA and GSH/GSSG levels increased under moderate metal stress conditions, accompanied by increased GSH reductase (GR) activity and expression. However, higher metal doses led to a decrease in the analysed parameters, and stronger toxic effects appeared with 30 μM Hg. The GSH concentration was significantly higher in rax1-1 (70% of Col-0) than in cad2-1 (40% of Col-0). The leaves of rax1-1 were less sensitive than cad2-1, in accordance with the greater expression of γECS in rax1-1. Our results underline the existence of a minimal GSH concentration threshold needed to minimise the toxic effects exerted by Hg.

  2. Low-temperature-induced transcription factors in grapevine enhance cold tolerance in transgenic Arabidopsis plants.

    PubMed

    Takuhara, Yuki; Kobayashi, Masayuki; Suzuki, Shunji

    2011-06-15

    We report the characterization of low-temperature-induced transcription factors in grapevine (Vitis vinifera). Four transcription factors were identified in low-temperature-treated grapevine. The expression of V. vinifera C-repeat-binding factors, VvCBF2, VvCBF4, and VvCBFL, and V. vinifera B-box-type zinc finger protein, VvZFPL, was immediately induced and upregulated in leaves by the low-temperature treatment. Similar induction of the gene expression was observed in low-temperature-treated stems and flowers, although VvZFPL was constitutively expressed in flowers. Tendrils expressed all the four genes constitutively. In berry skin, VvCBF2 and VvCBFL were induced by the low-temperature treatment before the onset of véraison, while only VvCBF2 was induced under the low-temperature condition after the onset of véraison. The overexpression of VvCBF2 and VvZFPL in Arabidopsis plants led to longer hypocotyls than the control plants. The rosette leaves of these plants were smaller and had lower chlorophyll contents than those of the control plants, resulting in a pale green color. Finally, the VvCBF2- and VvZFPL-overexpressing plants revealed growth retardation. These results suggest that VvCBF2 and VvZFPL may affect photomorphogenesis and growth in grapevine. Meanwhile, no morphological changes were detected in the VvCBF4- and VvCBFL-overexpressing plants. The cold tolerance test demonstrated that all of the overexpressing plants remained viable and noticeably healthy compared with the control plants even after exposure to severe cold treatment, suggesting that VvCBF2, VvCBF4, VvCBFL, or VvZFPL may enhance cold tolerance in grapevine.

  3. A moso bamboo WRKY gene PeWRKY83 confers salinity tolerance in transgenic Arabidopsis plants.

    PubMed

    Wu, Min; Liu, Huanlong; Han, Guomin; Cai, Ronghao; Pan, Feng; Xiang, Yan

    2017-09-15

    The WRKY family are transcription factors, involved in plant development, and response to biotic and abiotic stresses. Moso bamboo is an important bamboo that has high ecological, economic and cultural value and is widely distributed in the south of China. In this study, we performed a genome-wide identification of WRKY members in moso bamboo and identified 89 members. By comparative analysis in six grass genomes, we found the WRKY gene family may have experienced or be experiencing purifying selection. Based on relative expression levels among WRKY IIc members under three abiotic stresses, PeWRKY83 functioned as a transcription factor and was selected for detailed analysis. The transgenic Arabidopsis of PeWRKY83 showed superior physiological properties compared with the WT under salt stress. Overexpression plants were less sensitive to ABA at both germination and postgermination stages and accumulated more endogenous ABA under salt stress conditions. Further studies demonstrated that overexpression of PeWRKY83 could regulate the expression of some ABA biosynthesis genes (AtAAO3, AtNCED2, AtNCED3), signaling genes (AtABI1, AtPP2CA) and responsive genes (AtRD29A, AtRD29B, AtABF1) under salt stress. Together, these results suggested that PeWRKY83 functions as a novel WRKY-related TF which plays a positive role in salt tolerance by regulating stress-induced ABA synthesis.

  4. Regulation of flooding tolerance of SAG12:ipt Arabidopsis plants by cytokinin.

    PubMed

    Huynh, Le Nguyen; Vantoai, Tara; Streeter, John; Banowetz, Gary

    2005-05-01

    A SAG12:ipt gene construct, which increases cytokinin biosynthesis in response to senescence, was introduced into Arabidopsis plants to delay senescence induced by flooding stress. Two forms of flooding stress, including total submergence and root waterlogging, were applied to SAG12:ipt (IPT) and wild-type (WT) plants for 1, 3, and 5 d. A separate experiment compared the recovery of WT and IPT plants subjected to flooding stress. Biomass accumulation, carbohydrate and chlorophyll contents, and cytokinin and abscisic acid were quantified to compare genotypic responses to flooding stress and post-flooding recovery. Real-time RT-PCR studies were performed to quantify ipt and SAG12 gene expression. IPT plants exposed to waterlogging accumulated greater quantities of cytokinins more rapidly than WT plants or those exposed to total submergence. Cytokinin accumulation was accompanied by phenotypic adaptations, including chlorophyll retention and increased biomass and carbohydrate content relative to WT plants. Abscisic acid accumulated rapidly in WT and IPT plants under waterlogging stress but remained low in all genotypes exposed to total submergence. IPT plants showed improved recovery after waterlogging stress was removed. Expression of ipt in submerged plants did not result in cytokinin accumulation until submergence stress was removed. At that point, IPT plants accumulated greater quantities of cytokinin and recovered to a greater extent than WT plants. This study established the relationship between flooding tolerance and cytokinin accumulation in IPT plants and suggested that translation of ipt transcripts and subsequent cytokinin accumulation were delayed under submergence stress.

  5. Protein S-ACYL Transferase10 is critical for development and salt tolerance in Arabidopsis.

    PubMed

    Zhou, Liang-Zi; Li, Sha; Feng, Qiang-Nan; Zhang, Yu-Ling; Zhao, Xinying; Zeng, Yong-lun; Wang, Hao; Jiang, Liwen; Zhang, Yan

    2013-03-01

    Protein S-acylation, commonly known as palmitoylation, is a reversible posttranslational modification that catalyzes the addition of a saturated lipid group, often palmitate, to the sulfhydryl group of a Cys. Palmitoylation regulates enzyme activity, protein stability, subcellular localization, and intracellular sorting. Many plant proteins are palmitoylated. However, little is known about protein S-acyl transferases (PATs), which catalyze palmitoylation. Here, we report that the tonoplast-localized PAT10 is critical for development and salt tolerance in Arabidopsis thaliana. PAT10 loss of function resulted in pleiotropic growth defects, including smaller leaves, dwarfism, and sterility. In addition, pat10 mutants are hypersensitive to salt stresses. We further show that PAT10 regulates the tonoplast localization of several calcineurin B-like proteins (CBLs), including CBL2, CBL3, and CBL6, whose membrane association also depends on palmitoylation. Introducing a C192S mutation within the highly conserved catalytic motif of PAT10 failed to complement pat10 mutants, indicating that PAT10 functions through protein palmitoylation. We propose that PAT10-mediated palmitoylation is critical for vacuolar function by regulating membrane association or the activities of tonoplast proteins.

  6. Overexpression of cotton PYL genes in Arabidopsis enhances the transgenic plant tolerance to drought stress.

    PubMed

    Chen, Yun; Feng, Li; Wei, Ning; Liu, Zhi-Hao; Hu, Shan; Li, Xue-Bao

    2017-03-30

    PYR/PYL/RCAR proteins are putative abscisic acid (ABA) receptors that play important roles in plant responses to biotic and abiotic stresses. In this study, 27 predicted PYL proteins were identified in cotton (Gossypium hirsutum). Sequence analysis showed they are conserved in structures. Phylogenetic analysis showed that cotton PYL family could be categorized into three groups. Yeast two-hybrid assay revealed that the GhPYL proteins selectively interacted with some GhPP2C proteins. Quantitative RT-PCR analysis indicated that the most of nine GhPYL genes were down-regulated, while the other three were up-regulated in cotton under drought stress. Overexpression of GhPYL10/12/26 in Arabidopsis conferred the transgenic plants increased ABA sensitivity during seed germination and early seedling growth. On the contrary, the transgenic seedlings displayed better growth status and longer primary roots under normal conditions and mannitol stress, compared with wild type. Furthermore, the transgenic plants showed the enhanced drought tolerance, relative to wild type, when they were suffered from drought stress. Expression of some stress-related genes in transgenic plants was significant higher than that in wild type under osmotic stress. Thus, our data suggested that these cotton PYL genes may be involved in plant response and defense to drought/osmotic stress.

  7. RhEXPA4, a rose expansin gene, modulates leaf growth and confers drought and salt tolerance to Arabidopsis.

    PubMed

    Lü, Peitao; Kang, Mei; Jiang, Xinqiang; Dai, Fanwei; Gao, Junping; Zhang, Changqing

    2013-06-01

    Drought and high salinity are major environmental conditions limiting plant growth and development. Expansin is a cell-wall-loosening protein known to disrupt hydrogen bonds between xyloglucan and cellulose microfibrils. The expression of expansin increases in plants under various abiotic stresses, and plays an important role in adaptation to these stresses. We aimed to investigate the role of the RhEXPA4, a rose expansin gene, in response to abiotic stresses through its overexpression analysis in Arabidopsis. In transgenic Arabidopsis harboring the Pro RhEXPA4 ::GUS construct, RhEXPA4 promoter activity was induced by abscisic acid (ABA), drought and salt, particularly in zones of active growth. Transgenic lines with higher RhEXPA4 level developed compact phenotypes with shorter stems, curly leaves and compact inflorescences, while the lines with relatively lower RhEXPA4 expression showed normal phenotypes, similar to the wild type (WT). The germination percentage of transgenic Arabidopsis seeds was higher than that of WT seeds under salt stress and ABA treatments. Transgenic plants showed enhanced tolerance to drought and salt stresses: they displayed higher survival rates after drought, and exhibited more lateral roots and higher content of leaf chlorophyll a under salt stress. Moreover, high-level RhEXPA4 overexpressors have multiple modifications in leaf blade epidermal structure, such as smaller, compact cells, fewer stomata and midvein vascular patterning in leaves, which provides them with more tolerance to abiotic stresses compared to mild overexpressors and the WT. Collectively, our results suggest that RhEXPA4, a cell-wall-loosening protein, confers tolerance to abiotic stresses through modifying cell expansion and plant development in Arabidopsis.

  8. Volatile-Mediated Effects Predominate in Paraburkholderia phytofirmans Growth Promotion and Salt Stress Tolerance of Arabidopsis thaliana

    PubMed Central

    Ledger, Thomas; Rojas, Sandy; Timmermann, Tania; Pinedo, Ignacio; Poupin, María J.; Garrido, Tatiana; Richter, Pablo; Tamayo, Javier

    2016-01-01

    Abiotic stress has a growing impact on plant growth and agricultural activity worldwide. Specific plant growth promoting rhizobacteria have been reported to stimulate growth and tolerance to abiotic stress in plants, and molecular mechanisms like phytohormone synthesis and 1-aminocyclopropane-1-carboxylate deamination are usual candidates proposed to mediate these bacterial effects. Paraburkholderia phytofirmans PsJN is able to promote growth of several plant hosts, and improve their tolerance to chilling, drought and salinity. This work investigated bacterial determinants involved in PsJN stimulation of growth and salinity tolerance in Arabidopsis thaliana, showing bacteria enable plants to survive long-term salinity treatment, accumulating less sodium within leaf tissues relative to non-inoculated controls. Inactivation of specific bacterial genes encoding ACC deaminase, auxin catabolism, N-acyl-homoserine-lactone production, and flagellin synthesis showed these functions have little influence on bacterial induction of salinity tolerance. Volatile organic compound emission from strain PsJN was shown to reproduce the effects of direct bacterial inoculation of roots, increasing plant growth rate and tolerance to salinity evaluated both in vitro and in soil. Furthermore, early exposure to VOCs from P. phytofirmans was sufficient to stimulate long-term effects observed in Arabidopsis growth in the presence and absence of salinity. Organic compounds were analyzed in the headspace of PsJN cultures, showing production of 2-undecanone, 7-hexanol, 3-methylbutanol and dimethyl disulfide. Exposure of A. thaliana to different quantities of these molecules showed that they are able to influence growth in a wide range of added amounts. Exposure to a blend of the first three compounds was found to mimic the effects of PsJN on both general growth promotion and salinity tolerance. To our knowledge, this is the first report on volatile compound-mediated induction of plant abiotic

  9. Volatile-Mediated Effects Predominate in Paraburkholderia phytofirmans Growth Promotion and Salt Stress Tolerance of Arabidopsis thaliana.

    PubMed

    Ledger, Thomas; Rojas, Sandy; Timmermann, Tania; Pinedo, Ignacio; Poupin, María J; Garrido, Tatiana; Richter, Pablo; Tamayo, Javier; Donoso, Raúl

    2016-01-01

    Abiotic stress has a growing impact on plant growth and agricultural activity worldwide. Specific plant growth promoting rhizobacteria have been reported to stimulate growth and tolerance to abiotic stress in plants, and molecular mechanisms like phytohormone synthesis and 1-aminocyclopropane-1-carboxylate deamination are usual candidates proposed to mediate these bacterial effects. Paraburkholderia phytofirmans PsJN is able to promote growth of several plant hosts, and improve their tolerance to chilling, drought and salinity. This work investigated bacterial determinants involved in PsJN stimulation of growth and salinity tolerance in Arabidopsis thaliana, showing bacteria enable plants to survive long-term salinity treatment, accumulating less sodium within leaf tissues relative to non-inoculated controls. Inactivation of specific bacterial genes encoding ACC deaminase, auxin catabolism, N-acyl-homoserine-lactone production, and flagellin synthesis showed these functions have little influence on bacterial induction of salinity tolerance. Volatile organic compound emission from strain PsJN was shown to reproduce the effects of direct bacterial inoculation of roots, increasing plant growth rate and tolerance to salinity evaluated both in vitro and in soil. Furthermore, early exposure to VOCs from P. phytofirmans was sufficient to stimulate long-term effects observed in Arabidopsis growth in the presence and absence of salinity. Organic compounds were analyzed in the headspace of PsJN cultures, showing production of 2-undecanone, 7-hexanol, 3-methylbutanol and dimethyl disulfide. Exposure of A. thaliana to different quantities of these molecules showed that they are able to influence growth in a wide range of added amounts. Exposure to a blend of the first three compounds was found to mimic the effects of PsJN on both general growth promotion and salinity tolerance. To our knowledge, this is the first report on volatile compound-mediated induction of plant abiotic

  10. Overexpression of MuHSP70 gene from Macrotyloma uniflorum confers multiple abiotic stress tolerance in transgenic Arabidopsis thaliana.

    PubMed

    Masand, Shikha; Yadav, Sudesh Kumar

    2016-02-01

    A 70-KD heat shock protein (HSP70) is one of the most conserved chaperones. It is involved in de novo protein folding and prevents the aggregation of unfolded proteins under lethal environmental factors. The purpose of this study is to characterise a MuHSP70 from horsegram (Macrotyloma uniflorum) and elucidating its role in stress tolerance of plants. A MuHSP70 was cloned and characterised from a natural drought stress tolerant HPK4 variety of horsegram (M. uniflorum). For functional characterization, MuHSP70 was overexpressed in transgenic Arabidopsis. Overexpression of MuHSP70 was found to provide tolerance to the transgenic Arabidopsis against various stresses such as heat, cold, drought, salinity and oxidative stress. MuHSP70 transgenics were observed to maintain the shoot biomass, root length, relative water content, and chlorophyll content during exposure to multi-stresses relative to non-transgenic control. Transgenic lines have further shown the reduced levels of MDA, H2O2, and proteolytic activity. Together, these findings suggest that overexpression of MuHSP70 plays an important role in improving abiotic stress tolerance and could be a crucial candidate gene for exploration in crop improvement program.

  11. A maize stress-responsive NAC transcription factor, ZmSNAC1, confers enhanced tolerance to dehydration in transgenic Arabidopsis.

    PubMed

    Lu, Min; Ying, Sheng; Zhang, Deng-Feng; Shi, Yun-Su; Song, Yan-Chun; Wang, Tian-Yu; Li, Yu

    2012-09-01

    NAC proteins are plant-specific transcription factors that play essential roles in stress responses. However, only little information regarding stress-related NAC genes is available in maize. In this study, a maize NAC gene, ZmSNAC1, was cloned and functionally characterized. Expression analysis revealed that ZmSNAC1 was strongly induced by low temperature, high-salinity, drought stress, and abscisic acid (ABA) treatment, but downregulated by salicylic acid treatment. Subcellular localization experiments in Arabidopsis protoplast cells indicated that ZmSNAC1 was localized in the nucleus. Transactivation assays demonstrated that ZmSNAC1 functioned as a transcriptional activator. Overexpression of ZmSNAC1 in Arabidopsis led to hypersensitivity to ABA and osmotic stress at the germination stage, but enhanced tolerance to dehydration compared to wild-type seedlings. These results suggest that ZmSNAC1 functions as a stress-responsive transcription factor in positive modulation of abiotic stress tolerance, and may have applications in the engineering of drought-tolerant crops. ZmSNAC1 functioned as a stress-responsive transcription factor in response to abiotic stresses, and might be useful for crop tolerance improvement.

  12. ZAT11, a zinc finger transcription factor, is a negative regulator of nickel ion tolerance in Arabidopsis.

    PubMed

    Liu, Xiao-Min; An, Jonguk; Han, Hay Ju; Kim, Sun Ho; Lim, Chae Oh; Yun, Dae-Jin; Chung, Woo Sik

    2014-12-01

    ZAT11, a Zinc Finger of Arabidopsis Thaliana 11, is a dual-function transcriptional regulator that positively regulates primary root growth but negatively regulates Ni (2+) tolerance. Zinc Finger of Arabidopsis Thaliana 11 (ZAT11) is a C2H2-type zinc finger protein that has been reported to function as an active transcriptional repressor. However, the biological function of ZAT11 remains unknown. Here we show that GFP-tagged ZAT11 is targeted to the nucleus. Analysis of plants expressing ZAT11 promoter-GUS showed that ZAT11 is highly expressed in roots and particularly in root tips. To identify the biological function of ZAT11, we constructed three independent lines of ZAT11 overexpressing transgenic plant (ZAT11 OE). ZAT11 OE enhanced the elongation of primary root but reduced the metal tolerance against nickel ion (Ni(2+)). The reduced Ni(2+) tolerance of ZAT11 OE was correlated with decreased accumulation of Ni(2+) in plants. The decreased accumulation of Ni(2+) in ZAT11 OE was caused by the reduced transcription of a vacuolar Ni(2+) transporter gene. Taken together, our results suggest that ZAT11 is a dual function transcriptional regulator that positively regulates primary root growth but negatively regulates Ni(2+) tolerance.

  13. Overexpression of the ascorbate peroxidase gene from eggplant and sponge gourd enhances flood tolerance in transgenic Arabidopsis.

    PubMed

    Chiang, Chih-Ming; Chen, Chiu-Chen; Chen, Shi-Peng; Lin, Kuan-Hung; Chen, Li-Ru; Su, Yu-Huei; Yen, His-Cheng

    2017-03-01

    Previously, we found that the flood resistance of eggplant (Solanum melongena) and sponge gourd (Luffa cylindrica) enhanced ascorbate peroxidase (APX) activity under flooding, and consequently, both the SmAPX and LcAPX genes were cloned. In this study, the SmAPX and LcAPX genes were transferred under a ubiquitin promoter to Arabidopsis (At) via Agrobacterium tumefaciens. The expression and amount of APX and APX activities of the SmAPX and LcAPX transgenic lines were significantly higher than those of non-transgenic (NT) plants under a waterlogged condition. Furthermore, the SmAPX, LcAPX, At-sucrose synthases (SUS)-1, phosphoenolpyruvate carboxylase (PEPC), and lactate dehydrogenase (LDH) genes were overexpressed in all transgenic Arabidopsis lines after flooding treatment. Compared to NT plants, the malondialdehyde (MDA) contents and H2O2 accumulation were significantly lower, but germination rates were significantly higher in all transgenic lines with higher APX activity, indicating that the overexpression of SmAPX and LcAPX in Arabidopsis could enhance flood tolerance by eliminating H2O2. Moreover, Arabidopsis seedlings overexpressing SmAPX and LcAPX also displayed greater resistance to flooding and less oxidative injury than NT plants subjected to flooding condition.

  14. Verticillium infection triggers VASCULAR-RELATED NAC DOMAIN7-dependent de novo xylem formation and enhances drought tolerance in Arabidopsis.

    PubMed

    Reusche, Michael; Thole, Karin; Janz, Dennis; Truskina, Jekaterina; Rindfleisch, Sören; Drübert, Christine; Polle, Andrea; Lipka, Volker; Teichmann, Thomas

    2012-09-01

    The soilborne fungal plant pathogen Verticillium longisporum invades the roots of its Brassicaceae hosts and proliferates in the plant vascular system. Typical aboveground symptoms of Verticillium infection on Brassica napus and Arabidopsis thaliana are stunted growth, vein clearing, and leaf chloroses. Here, we provide evidence that vein clearing is caused by pathogen-induced transdifferentiation of chloroplast-containing bundle sheath cells to functional xylem elements. In addition, our findings suggest that reinitiation of cambial activity and transdifferentiation of xylem parenchyma cells results in xylem hyperplasia within the vasculature of Arabidopsis leaves, hypocotyls, and roots. The observed de novo xylem formation correlates with Verticillium-induced expression of the VASCULAR-RELATED NAC DOMAIN (VND) transcription factor gene VND7. Transgenic Arabidopsis plants expressing the chimeric repressor VND7-SRDX under control of a Verticillium infection-responsive promoter exhibit reduced de novo xylem formation. Interestingly, infected Arabidopsis wild-type plants show higher drought stress tolerance compared with noninfected plants, whereas this effect is attenuated by suppression of VND7 activity. Together, our results suggest that V. longisporum triggers a tissue-specific developmental plant program that compensates for compromised water transport and enhances the water storage capacity of infected Brassicaceae host plants. In conclusion, we provide evidence that this natural plant-fungus pathosystem has conditionally mutualistic features.

  15. Soybean GmMYB76, GmMYB92, and GmMYB177 genes confer stress tolerance in transgenic Arabidopsis plants.

    PubMed

    Liao, Yong; Zou, Hong-Feng; Wang, Hui-Wen; Zhang, Wan-Ke; Ma, Biao; Zhang, Jin-Song; Chen, Shou-Yi

    2008-10-01

    MYB-type transcription factors contain the conserved MYB DNA-binding domain of approximately 50 amino acids and are involved in the regulation of many aspects of plant growth, development, metabolism and stress responses. From soybean plants, we identified 156 GmMYB genes using our previously obtained 206 MYB unigenes, and 48 were found to have full-length open-reading frames. Expressions of all these identified genes were examined, and we found that expressions of 43 genes were changed upon treatment with ABA, salt, drought and/or cold stress. Three GmMYB genes, GmMYB76, GmMYB92 and GmMYB177, were chosen for further analysis. Using the yeast assay system, GmMYB76 and GmMYB92 were found to have transactivation activity and can form homodimers. GmMYB177 did not appear to have transactivation activity but can form heterodimers with GmMYB76. Yeast one-hybrid assay revealed that all the three GmMYBs could bind to cis-elements TAT AAC GGT TTT TT and CCG GAA AAA AGG AT, but with different affinity, and GmMYB92 could also bind to TCT CAC CTA CC. The transgenic Arabidopsis plants overexpressing GmMYB76 or GmMYB177 showed better performance than the GmMYB92-transgenic plants in salt and freezing tolerance. However, these transgenic plants exhibited reduced sensitivity to ABA treatment at germination stage in comparison with the wild-type plants. The three GmMYB genes differentially affected a subset of stress-responsive genes in addition to their regulation of a common subset of stress-responsive genes. These results indicate that the three GmMYB genes may play differential roles in stress tolerance, possibly through regulation of stress-responsive genes.

  16. Overexpression of a Soybean Ariadne-Like Ubiquitin Ligase Gene GmARI1 Enhances Aluminum Tolerance in Arabidopsis

    PubMed Central

    Zhang, Xiaolian; Wang, Ning; Chen, Pei; Gao, Mengmeng; Liu, Juge; Wang, Yufeng; Zhao, Tuanjie; Li, Yan; Gai, Junyi

    2014-01-01

    Ariadne (ARI) subfamily of RBR (Ring Between Ring fingers) proteins have been found as a group of putative E3 ubiquitin ligases containing RING (Really Interesting New Gene) finger domains in fruitfly, mouse, human and Arabidopsis. Recent studies showed several RING-type E3 ubiquitin ligases play important roles in plant response to abiotic stresses, but the function of ARI in plants is largely unknown. In this study, an ariadne-like E3 ubiquitin ligase gene was isolated from soybean, Glycine max (L.) Merr., and designated as GmARI1. It encodes a predicted protein of 586 amino acids with a RBR supra-domain. Subcellular localization studies using Arabidopsis protoplast cells indicated GmARI protein was located in nucleus. The expression of GmARI1 in soybean roots was induced as early as 2–4 h after simulated stress treatments such as aluminum, which coincided with the fact of aluminum toxicity firstly and mainly acting on plant roots. In vitro ubiquitination assay showed GmARI1 protein has E3 ligase activity. Overexpression of GmARI1 significantly enhanced the aluminum tolerance of transgenic Arabidopsis. These findings suggest that GmARI1 encodes a RBR type E3 ligase, which may play important roles in plant tolerance to aluminum stress. PMID:25364908

  17. Enhancement of naphthalene tolerance in transgenic Arabidopsis plants overexpressing the ferredoxin-like protein (ADI1) from rice.

    PubMed

    Fu, Xiao-Yan; Zhu, Bo; Han, Hong-Juan; Zhao, Wei; Tian, Yong-Sheng; Peng, Ri-He; Yao, Quan-Hong

    2016-01-01

    The ADI1 Arabidopsis plants enhanced tolerance and degradation efficiency to naphthalene and had great potential for phytoremediation of naphthalene in the plant material before composting or harvesting and removal. Naphthalene is a global environmental concern, because this substance is assumed to contribute considerably to human cancer risk. Cleaning up naphthalene contamination in the environment is crucial. Phytoremediation is an efficient technology to clean up contaminants. However, no gene that can efficiently degrade exogenous recalcitrant naphthalene in plants has yet been discovered. Ferredoxin (Fd) is a key player of biological electron transfer reaction in the PAH degradation process. The biochemical pathway for bacterial degradation of naphthalene has been well investigated. In this study, a rice gene, ADI1, which codes for a putative photosynthetic-type Fd, has been transformed into Arabidopsis thaliana. The transgenic Arabidopsis plants enhanced tolerance and degradation efficiency of naphthalene. Compared with wild-type plants, transgenic plants assimilated naphthalene from the culture media faster and removed more of this substance. When taken together, our findings suggest that breeding plants with overexpressed ADI1 gene is an effective strategy to degrade naphthalene in the environment.

  18. S-methylmethionine is involved in the salinity tolerance of Arabidopsis thaliana plants at germination and early growth stages.

    PubMed

    Ogawa, Saori; Mitsuya, Shiro

    2012-01-01

    Methionine (Met) is biosynthesized by the activated methyl cycle and S-methylmethionine (SMM) cycle in one-carbon (C1) metabolism in plants. It is converted to S-adenosylmethionine (SAM) which serves as a precursor for many metabolites including glycinebetaine, methylated polyols, polyamines and ethylene which accumulate in plants in response to salinity. We have investigated how the Met biosynthetic pathway is regulated under saline conditions at the transcriptional level in Arabidopsis thaliana plants. Within Met biosynthesis-related genes, the expression of homocysteine methyltransferase (HMT) and methionine methyltransferase (MMT) genes in SMM cycle had altered toward increasing Met production by the presence of NaCl. We have determined the salinity tolerance of an Arabidopsis mmt mutant with an insertional mutation in the single copy of the AtMMT gene. Although the mmt mutant showed comparable germination and shoot growth with wild type under normal conditions, NaCl treatment caused severe repression of germination rate and shoot growth in the mmt mutant compared with in the wild type. These results indicate that the utilization of SMM is important for the salinity tolerance of Arabidopsis plants at the germination and early growth stages.

  19. Overexpression of a soybean ariadne-like ubiquitin ligase gene GmARI1 enhances aluminum tolerance in Arabidopsis.

    PubMed

    Zhang, Xiaolian; Wang, Ning; Chen, Pei; Gao, Mengmeng; Liu, Juge; Wang, Yufeng; Zhao, Tuanjie; Li, Yan; Gai, Junyi

    2014-01-01

    Ariadne (ARI) subfamily of RBR (Ring Between Ring fingers) proteins have been found as a group of putative E3 ubiquitin ligases containing RING (Really Interesting New Gene) finger domains in fruitfly, mouse, human and Arabidopsis. Recent studies showed several RING-type E3 ubiquitin ligases play important roles in plant response to abiotic stresses, but the function of ARI in plants is largely unknown. In this study, an ariadne-like E3 ubiquitin ligase gene was isolated from soybean, Glycine max (L.) Merr., and designated as GmARI1. It encodes a predicted protein of 586 amino acids with a RBR supra-domain. Subcellular localization studies using Arabidopsis protoplast cells indicated GmARI protein was located in nucleus. The expression of GmARI1 in soybean roots was induced as early as 2-4 h after simulated stress treatments such as aluminum, which coincided with the fact of aluminum toxicity firstly and mainly acting on plant roots. In vitro ubiquitination assay showed GmARI1 protein has E3 ligase activity. Overexpression of GmARI1 significantly enhanced the aluminum tolerance of transgenic Arabidopsis. These findings suggest that GmARI1 encodes a RBR type E3 ligase, which may play important roles in plant tolerance to aluminum stress.

  20. The sunflower transcription factor HaHB11 improves yield, biomass and tolerance to flooding in transgenic Arabidopsis plants.

    PubMed

    Cabello, Julieta V; Giacomelli, Jorge I; Piattoni, Claudia V; Iglesias, Alberto A; Chan, Raquel L

    2016-03-20

    HaHB11 is a member of the sunflower homeodomain-leucine zipper I subfamily of transcription factors. The analysis of a sunflower microarray hybridized with RNA from HaHB11-transformed leaf-disks indicated the regulation of many genes encoding enzymes from glycolisis and fermentative pathways. A 1300bp promoter sequence, fused to the GUS reporter gene, was used to transform Arabidopsis plants showing an induction of expression after flooding treatments, concurrently with HaHB11 regulation by submergence in sunflower. Arabidopsis transgenic plants expressing HaHB11 under the control of the CaMV 35S promoter and its own promoter were obtained and these plants exhibited significant increases in rosette and stem biomass. All the lines produced more seeds than controls and particularly, those of high expression level doubled seeds yield. Transgenic plants also showed tolerance to flooding stress, both to submergence and waterlogging. Carbohydrates contents were higher in the transgenics compared to wild type and decreased less after submergence treatments. Finally, transcript levels of selected genes involved in glycolisis and fermentative pathways as well as the corresponding enzymatic activities were assessed both, in sunflower and transgenic Arabidopsis plants, before and after submergence. Altogether, the present work leads us to propose HaHB11 as a biotechnological tool to improve crops yield, biomass and flooding tolerance. Copyright © 2016 Elsevier B.V. All rights reserved.

  1. Genetic variation in photosynthetic performance and tolerance to osmotic stress (desiccation, freezing, hyposalinity) in the rocky littoral foundation species Fucus vesiculosus (Fucales, Phaeophyceae).

    PubMed

    Rothäusler, Eva; Sjöroos, Joakim; Heye, Katharina; Jormalainen, Veijo

    2016-10-01

    Genetic diversity may play an analogous role to species diversity, as it can contribute to ecosystem function and stability, and provision of ecosystem services. In the Baltic Sea, perennial algal beds are often comprised of only Fucus vesiculosus and the amount of genetic variation in fitness-related traits (i.e., the ability of the alga to photosynthesize or withstand stress) will thus determine the alga's local persistence in a changing environment. To study genetic variation in the crucial traits behind persistence we grew replicate vegetative branches that came from the same genotype in common gardens. We quantified osmotic stress tolerance and recovery responses by exposing branches to desiccation, freezing, and hyposalinity regimens. Our results show that genetic variation among genotypes was apparent for some photosynthetic parameters (maximal electron transport rate, saturation irradiance for electron transport, nonphotochemical quenching) and growth. Algae tolerated freezing (1,440 min at -2.5°C) and hyposalinity (1,560 min at 2.5) well, but did not recover from desiccation (70 min at 12°C, causing ~94% water loss). Furthermore, we found very little if any evidence on genetic variation in tolerance to these stressors. Our results suggest that low salinity and cold winters in the northern marginal populations selected for hyposalinity and freezing tolerant genotypes, possibly eroding genetic variation in tolerance, but that tolerance to harsh desiccation has been lost, likely due to relaxed selection. The overall availability of genetic variation in fitness related traits might be supportive for F. vesiculosus during adaptation to gradual changes of its environment. © 2016 Phycological Society of America.

  2. Nitric Oxide Synthase-Dependent Nitric Oxide Production Is Associated with Salt Tolerance in Arabidopsis1[OA

    PubMed Central

    Zhao, Min-Gui; Tian, Qiu-Ying; Zhang, Wen-Hao

    2007-01-01

    Nitric oxide (NO) has emerged as a key molecule involved in many physiological processes in plants. To characterize roles of NO in tolerance of Arabidopsis (Arabidopsis thaliana) to salt stress, effect of NaCl on Arabidopsis wild-type and mutant (Atnoa1) plants with an impaired in vivo NO synthase (NOS) activity and a reduced endogenous NO level was investigated. Atnoa1 mutant plants displayed a greater Na+ to K+ ratio in shoots than wild-type plants due to enhanced accumulation of Na+ and reduced accumulation of K+ when exposed to NaCl. Germination of Atnoa1 seeds was more sensitive to NaCl than that of wild-type seeds, and wild-type plants exhibited higher survival rates than Atnoa1 plants when grown under salt stress. Atnoa1 plants had higher levels of hydrogen peroxide than wild-type plants under both control and salt stress, suggesting that Atnoa1 is more vulnerable to salt and oxidative stress than wild-type plants. Treatments of wild-type plants with NOS inhibitor and NO scavenger reduced endogenous NO levels and enhanced NaCl-induced increase in Na+ to K+ ratio. Exposure of wild-type plants to NaCl inhibited NOS activity and reduced quantity of NOA1 protein, leading to a decrease in endogenous NO levels measured by NO-specific fluorescent probe. Treatment of Atnoa1 plants with NO donor sodium nitroprusside attenuated the NaCl-induced increase in Na+ to K+ ratio. Therefore, these findings provide direct evidence to support that disruption of NOS-dependent NO production is associated with salt tolerance in Arabidopsis. PMID:17351048

  3. 5-Formyltetrahydrofolate is an inhibitory but well tolerated metabolite in Arabidopsis leaves.

    PubMed

    Goyer, Aymeric; Collakova, Eva; Díaz de la Garza, Rocío; Quinlivan, Eoin P; Williamson, Jerry; Gregory, Jesse F; Shachar-Hill, Yair; Hanson, Andrew D

    2005-07-15

    5-Formyltetrahydrofolate (5-CHO-THF) is formed via a second catalytic activity of serine hydroxymethyltransferase (SHMT) and strongly inhibits SHMT and other folate-dependent enzymes in vitro. The only enzyme known to metabolize 5-CHO-THF is 5-CHO-THF cycloligase (5-FCL), which catalyzes its conversion to 5,10-methenyltetrahydrofolate. Because 5-FCL is mitochondrial in plants and mitochondrial SHMT is central to photorespiration, we examined the impact of an insertional mutation in the Arabidopsis 5-FCL gene (At5g13050) under photorespiratory (30 and 370 micromol of CO2 mol(-1)) and non-photorespiratory (3200 micromol of CO2 mol(-1)) conditions. The mutation had only mild visible effects at 370 micromol of CO2 mol(-1), reducing growth rate by approximately 20% and delaying flowering by 1 week. However, the mutation doubled leaf 5-CHO-THF level under all conditions and, under photorespiratory conditions, quadrupled the pool of 10-formyl-/5,10-methenyltetrahydrofolates (which could not be distinguished analytically). At 370 micromol of CO2 mol(-1), the mitochondrial 5-CHO-THF pool was 8-fold larger in the mutant and contained most of the 5-CHO-THF in the leaf. In contrast, the buildup of 10-formyl-/5,10-methenyltetrahydrofolates was extramitochondrial. In photorespiratory conditions, leaf glycine levels were up to 46-fold higher in the mutant than in the wild type. Furthermore, when leaves were supplied with 5-CHO-THF, glycine accumulated in both wild type and mutant. These data establish that 5-CHO-THF can inhibit SHMT in vivo and thereby influence glycine pool size. However, the near-normal growth of the mutant shows that even exceptionally high 5-CHO-THF levels do not much affect fluxes through SHMT or any other folate-dependent reaction, i.e. that 5-CHO-THF is well tolerated in plants.

  4. The cyclic nucleotide-gated channel, AtCNGC10, influences salt tolerance in Arabidopsis.

    PubMed

    Guo, Kun-Mei; Babourina, Olga; Christopher, David A; Borsics, Tamas; Rengel, Zed

    2008-11-01

    Cyclic nucleotide-gated channels (CNGCs) in the plasma membrane transport K+ and other cations; however, their roles in the response and adaptation of plants to environmental salinity are unclear. Growth, cation contents, salt tolerance and K+ fluxes were assessed in wild-type and two AtCNGC10 antisense lines (A2 and A3) of Arabidopsis thaliana (L.) Heynh. Compared with the wild-type, mature plants of both antisense lines had altered K+ and Na+ concentrations in shoots and were more sensitive to salt stress, as assessed by biomass and Chl fluorescence. The shoots of A2 and A3 plants contained higher Na+ concentrations and significantly higher Na+/K+ ratios compared with wild-type, whereas roots contained higher K+ concentrations and lower Na+/K+ ratios. Four-day-old seedlings of both antisense lines exposed to salt stress had smaller Na+/K+ ratios and longer roots than the wild-type. Under sudden salt treatment, the Na+ efflux was higher and the K+ efflux was smaller in the antisense lines, indicating that AtCNGC10 might function as a channel providing Na+ influx and K+ efflux at the root/soil interface. We conclude that the AtCNGC10 channel is involved in Na+ and K+ transport during cation uptake in roots and in long-distance transport, such as phloem loading and/or xylem retrieval. Mature A2 and A3 plants became more salt sensitive than wild-type plants because of impaired photosynthesis induced by a higher Na+ concentration in the leaves.

  5. Increased Drought Tolerance through the Suppression of ESKMO1 Gene and Overexpression of CBF-Related Genes in Arabidopsis

    PubMed Central

    Xu, Fuhui; Liu, Zhixue; Xie, Hongyan; Zhu, Jian; Zhang, Juren; Kraus, Josef; Blaschnig, Tasja; Nehls, Reinhard; Wang, Hong

    2014-01-01

    Improved drought tolerance is always a highly desired trait for agricultural plants. Significantly increased drought tolerance in Arabidopsis thaliana (Columbia-0) has been achieved in our work through the suppression of ESKMO1 (ESK1) gene expression with small-interfering RNA (siRNA) and overexpression of CBF genes with constitutive gene expression. ESK1 has been identified as a gene linked to normal development of the plant vascular system, which is assumed directly related to plant drought response. By using siRNA that specifically targets ESK1, the gene expression has been reduced and drought tolerance of the plant has been enhanced dramatically in the work. However, the plant response to external abscisic acid application has not been changed. ICE1, CBF1, and CBF3 are genes involved in a well-characterized plant stress response pathway, overexpression of them in the plant has demonstrated capable to increase drought tolerance. By overexpression of these genes combining together with suppression of ESK1 gene, the significant increase of plant drought tolerance has been achieved in comparison to single gene manipulation, although the effect is not in an additive way. Accompanying the increase of drought tolerance via suppression of ESK1 gene expression, the negative effect has been observed in seeds yield of transgenic plants in normal watering conditions comparing with wide type plant. PMID:25184213

  6. A wheat lipid transfer protein (TdLTP4) promotes tolerance to abiotic and biotic stress in Arabidopsis thaliana.

    PubMed

    Safi, Hela; Saibi, Walid; Alaoui, Meryem Mrani; Hmyene, Abdelaziz; Masmoudi, Khaled; Hanin, Moez; Brini, Faïçal

    2015-04-01

    Lipid transfer proteins (LTPs) are members of the family of pathogenesis-related proteins (PR-14) that are believed to be involved in plant defense responses. In this study, we report the isolation and characterization of a novel gene TdLTP4 encoding an LTP protein from durum wheat [Triticum turgidum L. subsp. Durum Desf.]. Molecular Phylogeny analyses of wheat TdLTP4 gene showed a high identity to other plant LTPs. Predicted three-dimensional structural model revealed the presence of six helices and nine loop turns. Expression analysis in two local durum wheat varieties with marked differences in salt and drought tolerance, revealed a higher transcript accumulation of TdLTP4 under different stress conditions in the tolerant variety, compared to the sensitive one. The overexpression of TdLTP4 in Arabidopsis resulted in a promoted plant growth under various stress conditions including NaCl, ABA, JA and H2O2 treatments. Moreover, the LTP-overexpressing lines exhibit less sensitivity to jasmonate than wild-type plants. Furthermore, detached leaves from transgenic Arabidopsis expressing TdLTP4 gene showed enhanced fungal resistance against Alternaria solani and Botrytis cinerea. Together, these data provide the evidence for the involvement of TdLTP4 gene in the tolerance to both abiotic and biotic stresses in crop plants.

  7. Heterologous expression of the yeast arsenite efflux system ACR3 improves Arabidopsis thaliana tolerance to arsenic stress.

    PubMed

    Ali, Waqar; Isner, Jean-Charles; Isayenkov, Stanislav V; Liu, Wenju; Zhao, Fang-Jie; Maathuis, Frans J M

    2012-05-01

    • Arsenic contamination has a negative impact on crop cultivation and on human health. As yet, no proteins have been identified in plants that mediate the extrusion of arsenic. Here, we heterologously expressed the yeast (Saccharomyces cerevisiae) arsenite efflux transporter ACR3 into Arabidopsis to evaluate how this affects plant tolerance and tissue arsenic contents. • ACR3 was cloned from yeast and transformed into wild-type and nip7;1 Arabidopsis. Arsenic tolerance was determined at the cellular level using vitality stains in protoplasts, in intact seedlings grown on agar plates and in mature plants grown hydroponically. Arsenic efflux was measured from protoplasts and from intact plants, and arsenic levels were measured in roots and shoots of plants exposed to arsenate. • At the cellular level, all transgenic lines showed increased tolerance to arsenite and arsenate and a greater capacity for arsenate efflux. With intact plants, three of four stably transformed lines showed improved growth, whereas only transgenic lines in the wild-type background showed increased efflux of arsenite into the external medium. The presence of ACR3 hardly affected tissue arsenic levels, but increased arsenic translocation to the shoot. • Heterologous expression of yeast ACR3 endows plants with greater arsenic resistance, but does not lower significantly arsenic tissue levels. © 2012 The Authors. New Phytologist © 2012 New Phytologist Trust.

  8. Expression of the ggpPS gene for glucosylglycerol biosynthesis from Azotobacter vinelandii improves the salt tolerance of Arabidopsis thaliana.

    PubMed

    Klähn, Stephan; Marquardt, Daniel M; Rollwitz, Inga; Hagemann, Martin

    2009-01-01

    Many organisms accumulate compatible solutes in response to salt or desiccation stress. Moderate halotolerant cyanobacteria and some heterotrophic bacteria synthesize the compatible solute glucosylglycerol (GG) as their main protective compound. In order to analyse the potential of GG to improve salt tolerance of higher plants, the model plant Arabidopsis thaliana was transformed with the ggpPS gene from the gamma-proteobacterium Azotobacter vinelandii coding for a combined GG-phosphate synthase/phosphatase. The heterologous expression of the ggpPS gene led to the accumulation of high amounts of GG. Three independent Arabidopsis lines showing different GG contents were characterized in growth experiments. Plants containing a low (1-2 micromol g(-1) FM) GG content in leaves showed no altered growth performance under control conditions but an increased salt tolerance, whereas plants accumulating a moderate (2-8 micromol g(-1) FM) or a high GG content (around 17 micromol g(-1) FM) showed growth retardation and no improvement of salt resistance. These results indicate that the synthesis of the compatible solute GG has a beneficial effect on plant stress tolerance as long as it is accumulated to an extent that does not negatively interfere with plant metabolism.

  9. Populus euphratica APYRASE2 Enhances Cold Tolerance by Modulating Vesicular Trafficking and Extracellular ATP in Arabidopsis Plants1[OPEN

    PubMed Central

    Deng, Shurong; Sun, Jian; Zhao, Rui; Ding, Mingquan; Zhang, Yinan; Sun, Yuanling; Wang, Wei; Tan, Yeqing; Liu, Dandan; Ma, Xujun; Hou, Peichen; Wang, Meijuan; Lu, Cunfu; Shen, Xin; Chen, Shaoliang

    2015-01-01

    Apyrase and extracellular ATP play crucial roles in mediating plant growth and defense responses. In the cold-tolerant poplar, Populus euphratica, low temperatures up-regulate APYRASE2 (PeAPY2) expression in callus cells. We investigated the biochemical characteristics of PeAPY2 and its role in cold tolerance. We found that PeAPY2 predominantly localized to the plasma membrane, but punctate signals also appeared in the endoplasmic reticulum and Golgi apparatus. PeAPY2 exhibited broad substrate specificity, but it most efficiently hydrolyzed purine nucleotides, particularly ATP. PeAPY2 preferred Mg2+ as a cofactor, and it was insensitive to various, specific ATPase inhibitors. When PeAPY2 was ectopically expressed in Arabidopsis (Arabidopsis thaliana), cold tolerance was enhanced, based on root growth measurements and survival rates. Moreover, under cold stress, PeAPY2-transgenic plants maintained plasma membrane integrity and showed reduced cold-elicited electrolyte leakage compared with wild-type plants. These responses probably resulted from efficient plasma membrane repair via vesicular trafficking. Indeed, transgenic plants showed accelerated endocytosis and exocytosis during cold stress and recovery. We found that low doses of extracellular ATP accelerated vesicular trafficking, but high extracellular ATP inhibited trafficking and reduced cell viability. Cold stress caused significant increases in root medium extracellular ATP. However, under these conditions, PeAPY2-transgenic lines showed greater control of extracellular ATP levels than wild-type plants. We conclude that Arabidopsis plants that overexpressed PeAPY2 could increase membrane repair by accelerating vesicular trafficking and hydrolyzing extracellular ATP to avoid excessive, cold-elicited ATP accumulation in the root medium and, thus, reduced ATP-induced inhibition of vesicular trafficking. PMID:26224801

  10. Populus euphratica APYRASE2 Enhances Cold Tolerance by Modulating Vesicular Trafficking and Extracellular ATP in Arabidopsis Plants.

    PubMed

    Deng, Shurong; Sun, Jian; Zhao, Rui; Ding, Mingquan; Zhang, Yinan; Sun, Yuanling; Wang, Wei; Tan, Yeqing; Liu, Dandan; Ma, Xujun; Hou, Peichen; Wang, Meijuan; Lu, Cunfu; Shen, Xin; Chen, Shaoliang

    2015-09-01

    Apyrase and extracellular ATP play crucial roles in mediating plant growth and defense responses. In the cold-tolerant poplar, Populus euphratica, low temperatures up-regulate APYRASE2 (PeAPY2) expression in callus cells. We investigated the biochemical characteristics of PeAPY2 and its role in cold tolerance. We found that PeAPY2 predominantly localized to the plasma membrane, but punctate signals also appeared in the endoplasmic reticulum and Golgi apparatus. PeAPY2 exhibited broad substrate specificity, but it most efficiently hydrolyzed purine nucleotides, particularly ATP. PeAPY2 preferred Mg(2+) as a cofactor, and it was insensitive to various, specific ATPase inhibitors. When PeAPY2 was ectopically expressed in Arabidopsis (Arabidopsis thaliana), cold tolerance was enhanced, based on root growth measurements and survival rates. Moreover, under cold stress, PeAPY2-transgenic plants maintained plasma membrane integrity and showed reduced cold-elicited electrolyte leakage compared with wild-type plants. These responses probably resulted from efficient plasma membrane repair via vesicular trafficking. Indeed, transgenic plants showed accelerated endocytosis and exocytosis during cold stress and recovery. We found that low doses of extracellular ATP accelerated vesicular trafficking, but high extracellular ATP inhibited trafficking and reduced cell viability. Cold stress caused significant increases in root medium extracellular ATP. However, under these conditions, PeAPY2-transgenic lines showed greater control of extracellular ATP levels than wild-type plants. We conclude that Arabidopsis plants that overexpressed PeAPY2 could increase membrane repair by accelerating vesicular trafficking and hydrolyzing extracellular ATP to avoid excessive, cold-elicited ATP accumulation in the root medium and, thus, reduced ATP-induced inhibition of vesicular trafficking. © 2015 American Society of Plant Biologists. All Rights Reserved.

  11. A salt-regulated peptide derived from the CAP superfamily protein negatively regulates salt-stress tolerance in Arabidopsis.

    PubMed

    Chien, Pei-Shan; Nam, Hong Gil; Chen, Yet-Ran

    2015-08-01

    High salinity has negative impacts on plant growth through altered water uptake and ion-specific toxicities. Plants have therefore evolved an intricate regulatory network in which plant hormones play significant roles in modulating physiological responses to salinity. However, current understanding of the plant peptides involved in this regulatory network remains limited. Here, we identified a salt-regulated peptide in Arabidopsis. The peptide was 11 aa and was derived from the C terminus of a cysteine-rich secretory proteins, antigen 5, and pathogenesis-related 1 proteins (CAP) superfamily. This peptide was found by searching homologues in Arabidopsis using the precursor of a tomato CAP-derived peptide (CAPE) that was initially identified as an immune signal. In searching for a CAPE involved in salt responses, we screened CAPE precursor genes that showed salt-responsive expression and found that the PROAtCAPE1 (AT4G33730) gene was regulated by salinity. We confirmed the endogenous Arabidopsis CAP-derived peptide 1 (AtCAPE1) by mass spectrometry and found that a key amino acid residue in PROAtCAPE1 is critical for AtCAPE1 production. Moreover, although PROAtCAPE1 was expressed mainly in the roots, AtCAPE1 was discovered to be upregulated systemically upon salt treatment. The salt-induced AtCAPE1 negatively regulated salt tolerance by suppressing several salt-tolerance genes functioning in the production of osmolytes, detoxification, stomatal closure control, and cell membrane protection. This discovery demonstrates that AtCAPE1, a homologue of tomato immune regulator CAPE1, plays an important role in the regulation of salt stress responses. Our discovery thus suggests that the peptide may function in a trade-off between pathogen defence and salt tolerance.

  12. Overexpression of MzASMT improves melatonin production and enhances drought tolerance in transgenic Arabidopsis thaliana plants.

    PubMed

    Zuo, Bixiao; Zheng, Xiaodong; He, Pingli; Wang, Lin; Lei, Qiong; Feng, Chao; Zhou, Jingzhe; Li, Qingtian; Han, Zhenhai; Kong, Jin

    2014-11-01

    Melatonin is a potent naturally occurring reactive oxygen species (ROS) and reactive nitrogen species (RNS) scavenger in plants. Melatonin protects plants from oxidative stress and, therefore, it improves their tolerance against a variety of environmental abiotic stressors. N-acetylserotonin-O-methyltransferase (ASMT) is a specific enzyme required for melatonin synthesis. In this report, an ASMT gene was cloned from apple rootstock (Malus zumi Mats) and designated as MzASMT1 (KJ123721). The MzASMT1 expression was induced by drought stress in apple leaves. The upregulation of MzASMT1 in the apple leaf positively relates to melatonin production over a 24-hr dark/light cycle. Purified MzASMT1 protein expressed in E. coli converted its substrates to melatonin with an activity of approximately 5.5 pmol/min/mg protein. The transient transformation in tobacco identified that MzASMT1 is located in cytoplasm of the cell. When MzASMT1 gene driven by 35S promoter was transferred to Arabidopsis, melatonin levels in transgenic Arabidopsis plants were 2-4 times higher than those in the wild type. The transgenic Arabidopsis plants had significantly lower intrinsic ROS than the wild type and therefore these plants exhibited greater tolerance to drought stress than that of wild type. This is, at least partially, attributed to the elevated melatonin levels resulting from the overexpression of MzASMT1. The results elucidated the important role that membrane-located melatonin synthase plays in drought tolerance. These findings have significant implications in agriculture.

  13. A salt-regulated peptide derived from the CAP superfamily protein negatively regulates salt-stress tolerance in Arabidopsis

    PubMed Central

    Chien, Pei-Shan; Nam, Hong Gil; Chen, Yet-Ran

    2015-01-01

    High salinity has negative impacts on plant growth through altered water uptake and ion-specific toxicities. Plants have therefore evolved an intricate regulatory network in which plant hormones play significant roles in modulating physiological responses to salinity. However, current understanding of the plant peptides involved in this regulatory network remains limited. Here, we identified a salt-regulated peptide in Arabidopsis. The peptide was 11 aa and was derived from the C terminus of a cysteine-rich secretory proteins, antigen 5, and pathogenesis-related 1 proteins (CAP) superfamily. This peptide was found by searching homologues in Arabidopsis using the precursor of a tomato CAP-derived peptide (CAPE) that was initially identified as an immune signal. In searching for a CAPE involved in salt responses, we screened CAPE precursor genes that showed salt-responsive expression and found that the PROAtCAPE1 (AT4G33730) gene was regulated by salinity. We confirmed the endogenous Arabidopsis CAP-derived peptide 1 (AtCAPE1) by mass spectrometry and found that a key amino acid residue in PROAtCAPE1 is critical for AtCAPE1 production. Moreover, although PROAtCAPE1 was expressed mainly in the roots, AtCAPE1 was discovered to be upregulated systemically upon salt treatment. The salt-induced AtCAPE1 negatively regulated salt tolerance by suppressing several salt-tolerance genes functioning in the production of osmolytes, detoxification, stomatal closure control, and cell membrane protection. This discovery demonstrates that AtCAPE1, a homologue of tomato immune regulator CAPE1, plays an important role in the regulation of salt stress responses. Our discovery thus suggests that the peptide may function in a trade-off between pathogen defence and salt tolerance. PMID:26093145

  14. Cold acclimation-induced up-regulation of the ribosomal protein L7 gene in the freeze tolerant wood frog, Rana sylvatica.

    PubMed

    Wu, Shaobo; De Croos, J N Amritha; Storey, Kenneth B

    2008-11-15

    Natural freezing survival by the wood frog, Rana sylvatica, involves multiple organ-specific changes in gene expression. The present study used differential display PCR to find cold-responsive genes in wood frog skin. A cDNA was retrieved from skin that was in higher amounts in cold- versus warm-acclimated frogs. The cDNA was used to probe a wood frog liver cDNA library and retrieve a long sequence that, after the further application of 5'RACE, was shown to encode the full sequence of the ribosomal large subunit protein 7 (RPL7) (GenBank accession number AF175983). Wood frog RPL7 contained 246 amino acids and shared 90% identity with Xenopus laevis RPL7, 82-83% with chicken and zebrafish homologues, and 79% with mammalian RPL7. Multiple binding domains found in human RPL7 showed differing degrees of conservation in the frog protein. Transcript levels of rpl7 were elevated up to 4-fold in skin of cold-acclimated frogs as compared with warm-acclimated animals. Organ-specific responses by rpl7 transcripts also occurred when frogs were given survivable freezing exposures. Transcripts rose by 1.8-3.3 fold in brain and skeletal muscle during freezing but were unaffected in central organs such as liver and heart. Up-regulation of rpl7 also occurred in brain of anoxia-exposed frogs and RPL7 protein levels increased strongly in heart under both freezing and dehydration stresses. Cold- and freezing-responsive up-regulation of the rpl7 gene and RPL7 protein in selected organs suggests that targeted changes in selected ribosomal proteins may be an integral part of natural freeze tolerance.

  15. Co-overexpressing a Plasma Membrane and a Vacuolar Membrane Sodium/Proton Antiporter Significantly Improves Salt Tolerance in Transgenic Arabidopsis Plants.

    PubMed

    Pehlivan, Necla; Sun, Li; Jarrett, Philip; Yang, Xiaojie; Mishra, Neelam; Chen, Lin; Kadioglu, Asim; Shen, Guoxin; Zhang, Hong

    2016-05-01

    The Arabidopsis gene AtNHX1 encodes a vacuolar membrane-bound sodium/proton (Na(+)/H(+)) antiporter that transports Na(+) into the vacuole and exports H(+) into the cytoplasm. The Arabidopsis gene SOS1 encodes a plasma membrane-bound Na(+)/H(+) antiporter that exports Na(+) to the extracellular space and imports H(+) into the plant cell. Plants rely on these enzymes either to keep Na(+) out of the cell or to sequester Na(+) into vacuoles to avoid the toxic level of Na(+) in the cytoplasm. Overexpression of AtNHX1 or SOS1 could improve salt tolerance in transgenic plants, but the improved salt tolerance is limited. NaCl at concentration >200 mM would kill AtNHX1-overexpressing or SOS1-overexpressing plants. Here it is shown that co-overexpressing AtNHX1 and SOS1 could further improve salt tolerance in transgenic Arabidopsis plants, making transgenic Arabidopsis able to tolerate up to 250 mM NaCl treatment. Furthermore, co-overexpression of AtNHX1 and SOS1 could significantly reduce yield loss caused by the combined stresses of heat and salt, confirming the hypothesis that stacked overexpression of two genes could substantially improve tolerance against multiple stresses. This research serves as a proof of concept for improving salt tolerance in other plants including crops. © The Author 2016. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists.

  16. Co-overexpressing a Plasma Membrane and a Vacuolar Membrane Sodium/Proton Antiporter Significantly Improves Salt Tolerance in Transgenic Arabidopsis Plants

    PubMed Central

    Pehlivan, Necla; Sun, Li; Jarrett, Philip; Yang, Xiaojie; Mishra, Neelam; Chen, Lin; Kadioglu, Asim; Shen, Guoxin; Zhang, Hong

    2016-01-01

    The Arabidopsis gene AtNHX1 encodes a vacuolar membrane-bound sodium/proton (Na+/H+) antiporter that transports Na+ into the vacuole and exports H+ into the cytoplasm. The Arabidopsis gene SOS1 encodes a plasma membrane-bound Na+/H+ antiporter that exports Na+ to the extracellular space and imports H+ into the plant cell. Plants rely on these enzymes either to keep Na+ out of the cell or to sequester Na+ into vacuoles to avoid the toxic level of Na+ in the cytoplasm. Overexpression of AtNHX1 or SOS1 could improve salt tolerance in transgenic plants, but the improved salt tolerance is limited. NaCl at concentration >200 mM would kill AtNHX1-overexpressing or SOS1-overexpressing plants. Here it is shown that co-overexpressing AtNHX1 and SOS1 could further improve salt tolerance in transgenic Arabidopsis plants, making transgenic Arabidopsis able to tolerate up to 250 mM NaCl treatment. Furthermore, co-overexpression of AtNHX1 and SOS1 could significantly reduce yield loss caused by the combined stresses of heat and salt, confirming the hypothesis that stacked overexpression of two genes could substantially improve tolerance against multiple stresses. This research serves as a proof of concept for improving salt tolerance in other plants including crops. PMID:26985021

  17. Macromolecular differentiation of Golgi stacks in root tips of Arabidopsis and Nicotiana seedlings as visualized in high pressure frozen and freeze-substituted samples

    NASA Technical Reports Server (NTRS)

    Staehelin, L. A.; Giddings, T. H. Jr; Kiss, J. Z.; Sack, F. D.

    1990-01-01

    The plant root tip represents a fascinating model system for studying changes in Golgi stack architecture associated with the developmental progression of meristematic cells to gravity sensing columella cells, and finally to "young" and "old", polysaccharide-slime secreting peripheral cells. To this end we have used high pressure freezing in conjunction with freeze-substitution techniques to follow developmental changes in the macromolecular organization of Golgi stacks in root tips of Arabidopsis and Nicotiana. Due to the much improved structural preservation of all cells under investigation, our electron micrographs reveal both several novel structural features common to all Golgi stacks, as well as characteristic differences in morphology between Golgi stacks of different cell types. Common to all Golgi stacks are clear and discrete differences in staining patterns and width of cis, medial and trans cisternae. Cis cisternae have the widest lumina (approximately 30 nm) and are the least stained. Medial cisternae are narrower (approximately 20 nm) and filled with more darkly staining products. Most trans cisternae possess a completely collapsed lumen in their central domain, giving rise to a 4-6 nm wide dark line in cross-sectional views. Numerous vesicles associated with the cisternal margins carry a non-clathrin type of coat. A trans Golgi network with clathrin coated vesicles is associated with all Golgi stacks except those of old peripheral cells. It is easily distinguished from trans cisternae by its blebbing morphology and staining pattern. The zone of ribosome exclusion includes both the Golgi stack and the trans Golgi network. Intercisternal elements are located exclusively between trans cisternae of columella and peripheral cells, but not meristematic cells. In older peripheral cells only trans cisternae exhibit slime-related staining. Golgi stacks possessing intercisternal elements also contain parallel rows of freeze-fracture particles in their trans

  18. Macromolecular differentiation of Golgi stacks in root tips of Arabidopsis and Nicotiana seedlings as visualized in high pressure frozen and freeze-substituted samples

    NASA Technical Reports Server (NTRS)

    Staehelin, L. A.; Giddings, T. H. Jr; Kiss, J. Z.; Sack, F. D.

    1990-01-01

    The plant root tip represents a fascinating model system for studying changes in Golgi stack architecture associated with the developmental progression of meristematic cells to gravity sensing columella cells, and finally to "young" and "old", polysaccharide-slime secreting peripheral cells. To this end we have used high pressure freezing in conjunction with freeze-substitution techniques to follow developmental changes in the macromolecular organization of Golgi stacks in root tips of Arabidopsis and Nicotiana. Due to the much improved structural preservation of all cells under investigation, our electron micrographs reveal both several novel structural features common to all Golgi stacks, as well as characteristic differences in morphology between Golgi stacks of different cell types. Common to all Golgi stacks are clear and discrete differences in staining patterns and width of cis, medial and trans cisternae. Cis cisternae have the widest lumina (approximately 30 nm) and are the least stained. Medial cisternae are narrower (approximately 20 nm) and filled with more darkly staining products. Most trans cisternae possess a completely collapsed lumen in their central domain, giving rise to a 4-6 nm wide dark line in cross-sectional views. Numerous vesicles associated with the cisternal margins carry a non-clathrin type of coat. A trans Golgi network with clathrin coated vesicles is associated with all Golgi stacks except those of old peripheral cells. It is easily distinguished from trans cisternae by its blebbing morphology and staining pattern. The zone of ribosome exclusion includes both the Golgi stack and the trans Golgi network. Intercisternal elements are located exclusively between trans cisternae of columella and peripheral cells, but not meristematic cells. In older peripheral cells only trans cisternae exhibit slime-related staining. Golgi stacks possessing intercisternal elements also contain parallel rows of freeze-fracture particles in their trans

  19. Metal transport protein 8 in Camellia sinensis confers superior manganese tolerance when expressed in yeast and Arabidopsis thaliana

    PubMed Central

    Li, Qinghui; Li, Yue; Wu, Xiayuan; Zhou, Lin; Zhu, Xujun; Fang, Wanping

    2017-01-01

    Manganese (Mn) is an important micronutrient element required for plant growth and development, playing catalytic roles in enzymes, membranes and DNA replication. The tea plant (Camellia sinensis) is able to accumulate high concentration of Mn without showing signs of toxicity, but the molecular mechanisms underlying this remain largely unknown. In this study, the C. sinensis cultivar ‘LJCY’ had higher Mn tolerance than cultivar ‘YS’, because chlorophyll content reduction was lower under the high Mn treatment. Proteomic analysis of the leaves revealed that C. sinensis Metal Tolerance Protein 8 (CsMTP8) accumulated in response to Mn toxicity in cultivar ‘LJCY’. The gene encoding CsMTP8, designated as CsMTP8 was also isolated, and its expression enhanced Mn tolerance in Saccharomyces cerevisiae. Similarly, the overexpression of CsMTP8 in Arabidopsis thaliana increased plant tolerance and reduced Mn accumulation in plant tissues under excess Mn conditions. Subcellular localization analysis of green florescence fused protein indicated that CsMTP8 was localized to the plasma membranes. Taken together, the results suggest that CsMTP8 is a Mn-specific transporter, which is localized in the plasma membrane, and transports excess Mn out of plant cells. The results also suggest that it is needed for Mn tolerance in shoots. PMID:28051151

  20. The limits of drought-induced rapid cold-hardening: extremely brief, mild desiccation triggers enhanced freeze-tolerance in Eurosta solidaginis larvae.

    PubMed

    Gantz, J D; Lee, Richard E

    2015-02-01

    Rapid cold-hardening (RCH) is a highly conserved response in insects that induces physiological changes within minutes to hours of exposure to low temperature and provides protection from chilling injury. Recently, a similar response, termed drought-induced RCH, was described following as little as 6h of desiccation, producing a loss of less than 10% of fresh mass. In this study, we investigated the limits and mechanisms of this response in larvae of the goldenrod gall fly Eurosta solidaginis (Diptera, Tephritidae). The cold-hardiness of larvae increased markedly after as few as 2h of desiccation and a loss of less than 1% fresh mass, as organismal survival increased from 8% to 41% following exposure to -18 °C. Tissue-level effects of desiccation were observed within 1h, as 87% of midgut cells from desiccated larvae remained viable following freezing compared to 57% of controls. We also demonstrated that drought-induced RCH occurs independently of neuroendocrine input, as midgut tissue desiccated ex vivo displayed improved freeze-tolerance relative to control tissue (78-11% survival, respectively). Finally, though there was an increase in hemolymph osmolality beyond the expected effects of the osmo-concentration of solutes during dehydration, we determined that this increase was not due to the synthesis of glycerol, glucose, sorbitol, or trehalose. Our results indicate that E. solidaginis larvae are extremely sensitive to desiccation, which is a triggering mechanism for one or more physiological pathways that confer enhanced freeze-tolerance.

  1. Central role of the flowering repressor ZCCT2 in the redox control of freezing tolerance and the initial development of flower primordia in wheat

    PubMed Central

    2014-01-01

    Background As both abiotic stress response and development are under redox control, it was hypothesised that the pharmacological modification of the redox environment would affect the initial development of flower primordia and freezing tolerance in wheat (Triticum aestivum L.). Results Pharmacologically induced redox changes were monitored in winter (T. ae. ssp. aestivum cv. Cheyenne, Ch) and spring (T. ae. ssp. spelta; Tsp) wheat genotypes grown after germination at 20/17°C for 9 d (chemical treatment: last 3 d), then at 5°C for 21 d (chemical treatment: first 4 d) and subsequently at 20/17°C for 21 d (recovery period). Thiols and their disulphide forms were measured and based on these data reduction potentials were calculated. In the freezing-tolerant Ch the chemical treatments generally increased both the amount of thiol disulphides and the reduction potential after 3 days at 20/17°C. In the freezing-sensitive Tsp a similar effect of the chemicals on these parameters was only observed after the continuation of the treatments for 4 days at 5°C. The applied chemicals slightly decreased root fresh weight and increased freezing tolerance in Ch, whereas they increased shoot fresh weight in Tsp after 4 days at 5°C. As shown after the 3-week recovery at 20/17°C, the initial development of flower primordia was accelerated in Tsp, whereas it was not affected by the treatments in Ch. The chemicals differently affected the expression of ZCCT2 and that of several other genes related to freezing tolerance and initial development of flower primordia in Ch and Tsp after 4 d at 5°C. Conclusions Various redox-altering compounds and osmotica had differential effects on glutathione disulphide content and reduction potential, and consequently on the expression of the flowering repressor ZCCT2 in the winter wheat Ch and the spring wheat Tsp. We propose that the higher expression of ZCCT2 in Ch may be associated with activation of genes of cold acclimation and its lower

  2. Overexpression of CaDSR6 increases tolerance to drought and salt stresses in transgenic Arabidopsis plants.

    PubMed

    Kim, Eun Yu; Seo, Young Sam; Park, Ki Youl; Kim, Soo Jin; Kim, Woo Taek

    2014-11-15

    The partial CaDSR6 (Capsicum annuum Drought Stress Responsive 6) cDNA was previously identified as a drought-induced gene in hot pepper root tissues. However, the cellular role of CaDSR6 with regard to drought stress tolerance was unknown. In this report, full-length CaDSR6 cDNA was isolated. The deduced CaDSR6 protein was composed of 234 amino acids and contained an approximately 30 amino acid-long Asp-rich domain in its central region. This Asp-rich domain was highly conserved in all plant DSR6 homologs identified and shared a sequence identity with the N-terminal regions of yeast p23(fyp) and human hTCTP, which contain Rab protein binding sites. Transgenic Arabidopsis plants overexpressing CaDSR6 (35S:CaDSR6-sGFP) were tolerant to high salinity, as identified by more vigorous root growth and higher levels of total chlorophyll than wild type plants. CaDSR6-overexpressors were also more tolerant to drought stress compared to wild type plants. The 35S:CaDSR6-sGFP leaves retained their water content and chlorophyll more efficiently than wild type leaves in response to dehydration stress. The expression of drought-induced marker genes, such as RD20, RD22, RD26, RD29A, RD29B, RAB18, KIN2, ABF3, and ABI5, was markedly increased in CaDSR6-overexpressing plants relative to wild type plants under both normal and drought conditions. These results suggest that overexpression of CaDSR6 is associated with increased levels of stress-induced genes, which, in turn, conferred a drought tolerant phenotype in transgenic Arabidopsis plants. Overall, our data suggest that CaDSR6 plays a positive role in the response to drought and salt stresses.

  3. Overexpression of PSK1, a SKP1-like gene homologue, from Paeonia suffruticosa, confers salinity tolerance in Arabidopsis.

    PubMed

    Hao, Qing; Ren, Hongxu; Zhu, Jin; Wang, Liangsheng; Huang, Shouchen; Liu, Zheng'an; Gao, Zhimin; Shu, Qingyan

    2017-01-01

    Our study is the first to demonstrate that PSK1 , a SKP1 -like gene homologue, is involved in salinity tolerance. Our functional characterization of PSK1 provides new insights into tree peony development. A homologous gene of S-phase kinase-associated protein1 (SKP1) was cloned from tree peony (Paeonia suffruticosa) and denoted as PSK1. The 462-bp open reading frame of PSK1 was predicted to encode a protein comprising 153 amino acids, with a molecular mass of 17 kDa. The full-length gene was 1,634 bp long and included a large 904-bp intron. PSK1 transcription was detected in all tissues, with the highest level observed in sepals, followed by leaves. Under salinity stress, overexpression of PSK1 in Arabidopsis resulted in increased germination percentages, cotyledon greening, and fresh weights relative to wild-type plants. Furthermore, transgenic Arabidopsis lines containing 35S::PSK1 displayed increased expression of genes that would be essential for reproduction and growth under salinity stress: ASK1, LEAFY, FT, and CO involved in flower development and flowering time as well as P5CS, RAB18, DREB, and SOD1-3 contributing to salinity tolerance. Our functional characterization of PSK1 adds to global knowledge of the multiple functions of previously explored SKP1-like genes in plants and sheds light on the molecular mechanism underlying its role in salinity tolerance. Our findings also provide information on the function and molecular mechanism of PSK1 in tree peony flower development, thereby revealing a theoretical basis for regulation of flowering and conferral of salinity tolerance in tree peony.

  4. Overexpression of a peroxidase gene (AtPrx64) of Arabidopsis thaliana in tobacco improves plant's tolerance to aluminum stress.

    PubMed

    Wu, Yuanshuang; Yang, Zhili; How, Jingyi; Xu, Huini; Chen, Limei; Li, Kunzhi

    2017-08-16

    AtPrx64 is one of the peroxidases gene up-regulated in Al stress and has some functions in the formation of plant second cell wall. Its overexpression may improve plant tolerance to Al by some ways. Studies on its function under Al stress may help us to understand the mechanism of plant tolerance to Al stress. In Arabidopsis thaliana, the expressions of some genes (AtPrxs) encoding class III plant peroxidases have been found to be either up-regulated or down-regulated under aluminum (Al) stress. Among 73 genes that encode AtPrxs in Arabidopsis, AtPrx64 is always up-regulated by Al stress, suggesting this gene plays protective roles in response to such stress. In this study, transgenic tobacco plants were generated to examine the effects of overexpressing of AtPrx64 gene on the tolerance to Al stress. The results showed that overexpression of AtPrx64 gene increased the root growth and reduced the accumulation of Al and ROS in the roots. Compared with wild type controls, transgenic tobaccos had much less soluble proteins and malondialdehyde in roots and much more root citrate exudation. The activity of plasma membrane (PM) H(+)-ATPase, the phosphorylation of PM H(+)-ATPase and its interaction with 14-3-3 proteins increased in transgenic tobaccos; moreover, the content of lignin in root tips also increased. Taken together, these results showed that overexpression of AtPrx64 gene might enhance the tolerance of tobacco to Al stress.

  5. Involvement of elevated proline accumulation in enhanced osmotic stress tolerance in Arabidopsis conferred by chimeric repressor gene silencing technology.

    PubMed

    Kazama, Daisuke; Kurusu, Takamitsu; Mitsuda, Nobutaka; Ohme-Takagi, Masaru; Tada, Yuichi

    2014-01-01

    Arabidopsis plants transformed with a chimeric repressor for 6 transcription factors (TFs), including ADA2b, Msantd, DDF1, DREB26, AtGeBP, and ATHB23, that were converted by Chimeric REpressor gene Silencing Technology (CRES-T), show elevated salt and osmotic stress tolerance compared with wild type (WT) plants. However, the roles of TFs in salt and osmotic signaling remain largely unknown. Their hyper-osmotic stress tolerance was evaluated using 3 criteria: germination rate, root length, and rate of seedlings with visible cotyledons at the germination stage. All CRES-T lines tested exhibited better performance than WT, at least for one criterion under stress conditions. Under 600 mM mannitol stress, 3-week-old CRES-T lines accumulated proline, which is a major compatible solute involved in osmoregulation, at higher levels than WT. Expression levels of the delta 1-pyrroline-5-carboxylate synthase gene in CRES-T lines were similar to or lower than those in WT. In contrast, expression of the proline dehydrogenase (PHD) gene in DREB26-SRDX was significantly downregulated and that in ADA2b-SRDX and AtGeBP-SRDX was also rather downregulated compared with that in WT. Although plants at different stages were used for stress tolerance test and proline measurement in this study, we previously reported that 4 out of the 6 CRES-T lines showed better growth than WT after 4 weeks of incubation under 400 mM mannitol. These results suggest that proline accumulation caused by PHD gene suppression may be involved in enhanced osmotic stress tolerance in the CRES-T lines, and that these TFs may be involved in regulating proline metabolism in Arabidopsis.

  6. The Arabidopsis transcriptional regulator DPB3-1 enhances heat stress tolerance without growth retardation in rice.

    PubMed

    Sato, Hikaru; Todaka, Daisuke; Kudo, Madoka; Mizoi, Junya; Kidokoro, Satoshi; Zhao, Yu; Shinozaki, Kazuo; Yamaguchi-Shinozaki, Kazuko

    2016-08-01

    The enhancement of heat stress tolerance in crops is an important challenge for food security to facilitate adaptation to global warming. In Arabidopsis thaliana, the transcriptional regulator DNA polymerase II subunit B3-1 (DPB3-1)/nuclear factor Y subunit C10 (NF-YC10) has been reported as a positive regulator of Dehydration-responsive element binding protein 2A (DREB2A), and the overexpression of DPB3-1 enhances heat stress tolerance without growth retardation. Here, we show that DPB3-1 interacts with DREB2A homologues