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BACKGROUND: Calcium-binding proteins that contain EF-hand motifs have been reported to play important roles in transduction of signals associated with biotic and abiotic stresses. To functionally characterize genes of EF-hand family in response to abiotic stress, an MtCaMP1 gene belonging to EF-hand family from legume model plant Medicago truncatula was isolated and its function in response to drought and salt stress was investigated by expressing MtCaMP1 in Arabidopsis. METHODOLOGY/PRINCIPAL FINDINGS: Transgenic Arabidopsis seedlings expressing MtCaMP1 exhibited higher survival rate than wild-type seedlings under drought and salt stress, suggesting that expression of MtCaMP1 confers tolerance of Arabidopsis to drought and salt stress. The transgenic plants accumulated greater amounts of Pro due to up-regulation of P5CS1 and down-regulation of ProDH than wild-type plants under drought stress. There was a less accumulation of Na(+) in the transgenic plants than in WT plants due to reduced up-regulation of AtHKT1 and enhanced regulation of AtNHX1 in the transgenic plants compared to WT plants under salt stress. There was a reduced accumulation of H2O2 and malondialdehyde in the transgenic plants than in WT plants under both drought and salt stress. CONCLUSIONS/SIGNIFICANCE: The expression of MtCaMP1 in Arabidopsis enhanced tolerance of the transgenic plants to drought and salt stress by effective osmo-regulation due to greater accumulation of Pro and by minimizing toxic Na(+) accumulation, respectively. The enhanced accumulation of Pro and reduced accumulation of Na(+) under drought and salt stress would protect plants from water default and Na(+) toxicity, and alleviate the associated oxidative stress. These findings demonstrate that MtCaMP1 encodes a stress-responsive EF-hand protein that plays a regulatory role in response of plants to drought and salt stress.

Background: Long non-coding RNAs (lncRNAs) have been shown to play crucially regulatory roles in diverse biological processes involving complex mechanisms. However, information regarding the number, sequences, characteristics and potential functions of lncRNAs in plants is so far overly limited. Results: Using high-throughput sequencing and bioinformatics analysis, we identified a total of 23,324 putative lncRNAs from control, osmotic stress- and salt stress-treated leaf and root samples of Medicago truncatula, a model legume species. Out of these lncRNAs, 7,863 and 5,561 lncRNAs were identified from osmotic stress-treated leaf and root samples, respectively. While, 7,361 and 7,874 lncRNAs were identified from salt stress-treated leaf and root samples, respectively. To reveal their potential functions, we analyzed Gene Ontology (GO) terms of genes that overlap with or are neighbors of the stress-responsive lncRNAs. Enrichments in GO terms in biological processes such as signal transduction, energy synthesis, molecule metabolism, detoxification, transcription and translation were found. Conclusions: LncRNAs are likely involved in regulating plant's responses and adaptation to osmotic and salt stresses in complex regulatory networks with protein-coding genes. These findings are of importance for our understanding of the potential roles of lncRNAs in responses of plants in general and M. truncatula in particular to abiotic stresses. [ABSTRACT FROM AUTHOR]

Background Resequencing can be used to identify genome variations underpinning many morphological and physiological phenotypes. Legume model plant Medicago truncatula ecotypes Jemalong A17 (J. A17) and R108 differ in their responses to mineral toxicity of aluminum and sodium, and mineral deficiency of iron in growth medium. The difference may result from their genome variations, but no experimental evidence supports this hypothesis. Results A total of 12,750 structure variations, 135,045 short insertions/deletions and 764,154 single nucleotide polymorphisms were identified by resequencing the genome of R108. The suppressed expression of MtAACT that encodes a putative aluminum-induced citrate efflux transporter by deletion of partial sequence of the second intron may account for the less aluminum-induced citrate exudation and greater accumulation of aluminum in roots of R108 than in roots of J. A17, thus rendering R108 more sensitive to aluminum toxicity. The higher expression-level of MtZpt2-1 encoding a TFIIIA-related transcription factor in J. A17 than R108 under conditions of salt stress can be explained by the greater number of stressresponsive elements in its promoter sequence, thus conferring J. A17 more tolerant to salt stress than R108 plants by activating the expression of downstream stress-responsive genes. YSLs (Yellow Stripe-Likes) are involved in long-distance transport of iron in plants. We found that an YSL gene was deleted in the genome of R108 plants, thus rendering R108 less tolerance to iron deficiency than J. A17 plants. Conclusions The deletion or change in several genes may account for the different responses of M. truncatula ecotypes J. A17 and R108 to mineral toxicity of aluminum and sodium as well as iron deficiency. Uncovering genome variations by resequencing is an effective method to identify different traits between species/ecotypes that are genetically related. These findings demonstrate that analyses of genome variations by resequencing can shed important light on differences in responses of M. truncatula ecotypes to abiotic stress in general and mineral stress in particular. [ABSTRACT FROM AUTHOR]

Medicago falcata is a legume species that exhibits great capacity of tolerance to abiotic stresses. To elucidate the mechanism underlying tolerance of M. falcata to freezing, we compared the characteristics of M. falcata in response to cold acclimation and freezing with those of the legume model plant Medicago truncatula. M. falcata seedlings were more tolerant to freezing than M. truncatula, as evidenced by a lower value of EL (temperature at which 50% electrolyte leakage after freezing) and greater survival rate for M. falcata than M. truncatula. Cold acclimation led to greater reduction in EL for M. falcata than M. truncatula. Sucrose was the most abundant sugar in both M. falcta and M. truncatula, and a greater accumulation of sucrose and Pro in M. falcata than in M. truncatula during cold acclimation was observed. Cold acclimation induced small amounts of raffinose and stachyose in M. falcata, but not in M. truncatula. The activities of sucrose phosphate synthase and sucrose synthase were greater in M. falcata than in M. truncatula. In contrast, the activity of acid invertase was higher in M. truncatula than in M. falcata. There was an increase in transcript of CRT binding factor ( CBF) upon exposure to low temperature in the two species. The low temperature-induced increase in transcript of CBF2 was much higher in M. truncatula than in M. falcata, while transcript of CBF3 in M. falcata was greater than that in M. truncatula. There were sustained increases in transcripts of cold acclimation specific ( CAS), a downstream target of CBF, during cold acclimation and the increases were greater in M. falcata than in M. truncatula. These results demonstrate that accumulation of greater amounts of soluble sugars coupled with higher CBF3 and CAS transcript levels in M. falcata may play a role in conferring greater tolerance of M. falcata to freezing than that of M. truncatula. [ABSTRACT FROM AUTHOR]

Nitric oxide (NO) is an important signaling molecule involved in many physiological processes in plants. We evaluated the role of NO in cold acclimation and freezing tolerance using Arabidopsis (Arabidopsis thaliana) wild type and mutants nia1nia2 (for nitrate reductase [NR]-defective double mutant) and Atnoa1/rif1 (for nitric oxide associatedl/resistant to inhibition by fosinidomycin1) that exhibit defects in NR and reduced NO production, respectively. Cold acclimation induced an increase in endogenous NO production in wild-type and Atnoa1/rif1 leaves, while endogenous NO level in nia1nia2 leaves was lower than in wild-type ones and was little changed during cold acclimation. Cold acclimation stimulated NR activity and induced up-regulation of NIA1 gene expression. In contrast, cold acclimation reduced the quantity of NOA1/RIF1 protein and inhibited NO synthase (NOS) activity. These results indicate that up-regulation of NR-dependent NO synthesis underpins cold acclimation-induced NO production. Seedlings of nia1nia2 were less tolerant to freezing than wild-type plants. Pharmacological studies using NR inhibitor, NO scavenger, and NO donor showed that NR-dependent NO level was positively correlated with freezing tolerance. Furthermore, cold acclimation up- and down-regulated expression of P5CS1 and ProDH genes, respectively, resulting in enhanced accumulation of proline (Pro) in wild-type plants. The stimulation of Pro accumulation by cold acclimation was reduced by NR inhibitor and NO scavenger, while Pro accumulation by cold acclimation was not affected by the NOS inhibitor. In contrast to wild-type plants, cold acclimation up-regulated ProDH gene expression in nia1nia2 plants, leading to less accumulation in nia1nia2 plants than in wild-type plants. These findings demonstrate that NR-dependent NO production plays an important role in cold acclimation-induced increase in freezing tolerance by modulating Pro accumulation in Arabidopsis. [ABSTRACT FROM AUTHOR]

Nitric oxide (NO) is an important signaling molecule involved in many physiological processes in plants. We evaluated the role of NO in cold acclimation and freezing tolerance using Arabidopsis (Arabidopsis thaliana) wild type and mutants nia1nia2 (for nitrate reductase [NR]-defective double mutant) and Atnoa1/rif1 (for nitric oxide associatedl/resistant to inhibition by fosinidomycin1) that exhibit defects in NR and reduced NO production, respectively. Cold acclimation induced an increase in endogenous NO production in wild-type and Atnoa1/rif1 leaves, while endogenous NO level in nia1nia2 leaves was lower than in wild-type ones and was little changed during cold acclimation. Cold acclimation stimulated NR activity and induced up-regulation of NIA1 gene expression. In contrast, cold acclimation reduced the quantity of NOA1/RIF1 protein and inhibited NO synthase (NOS) activity. These results indicate that up-regulation of NR-dependent NO synthesis underpins cold acclimation-induced NO production. Seedlings of nia1nia2 were less tolerant to freezing than wild-type plants. Pharmacological studies using NR inhibitor, NO scavenger, and NO donor showed that NR-dependent NO level was positively correlated with freezing tolerance. Furthermore, cold acclimation up- and down-regulated expression of P5CS1 and ProDH genes, respectively, resulting in enhanced accumulation of proline (Pro) in wild-type plants. The stimulation of Pro accumulation by cold acclimation was reduced by NR inhibitor and NO scavenger, while Pro accumulation by cold acclimation was not affected by the NOS inhibitor. In contrast to wild-type plants, cold acclimation up-regulated ProDH gene expression in nia1nia2 plants, leading to less accumulation in nia1nia2 plants than in wild-type plants. These findings demonstrate that NR-dependent NO production plays an important role in cold acclimation-induced increase in freezing tolerance by modulating Pro accumulation in Arabidopsis. [ABSTRACT FROM AUTHOR]

• Here, the effects of the ethylene-releasing compound, ethephon, and the ethylene precursor, 1-aminocyclopropane-1-carboxylic acid (ACC), on ionic currents across plasma membranes and on the cytosolic Ca2+ activity ([Ca2+]c) of tobacco ( Nicotiana tabacum) suspension cells were characterized using a patch-clamp technique and confocal laser scanning microscopy. • Exposure of tobacco protoplasts to ethephon and ACC led to activation of a plasma membrane cation channel that was permeable to Ba2+, Mg2+ and Ca2+, and inhibited by La3+, Gd3+ and Al3+. • The ethephon- and ACC-induced Ca2+-permeable channel was abolished by the antagonist of ethylene perception (1-metycyclopropene) and by the inhibitor of ACC synthase (aminovinylglycin), indicating that activation of the Ca2+-permeable channels results from ethylene. Ethephon elicited an increase in the [Ca2+]c of tobacco suspension cells, as visualized by the Ca2+-sensitive probe Fluo-3 and confocal microscopy. The ethephon-induced elevation of [Ca2+]c was markedly inhibited by Gd3+ and BAPTA, suggesting that an influx of Ca2+ underlies the elevation of [Ca2+]c. • These results indicate that an elevation of [Ca2+]c, resulting from activation of the plasma membrane Ca2+-permeable channels by ethylene, is an essential component in ethylene signaling in plants. [ABSTRACT FROM AUTHOR]

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 (Atnoal) plants with an impaired in vivo NO synthase (NOS) activity and a reduced endogenous NO level was investigated. Atnoal 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 Atnoal 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 Atnoal 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 Atnoal plants with NO donor sodium nitroprusside attenuated the NaCI-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. [ABSTRACT FROM AUTHOR]

• Aluminum (Al) is toxic to plants when solubilized into Al3+ in acidic soils, and becomes a major factor limiting plant growth. However, the primary cause for Al toxicity remains unknown. • Nitric oxide (NO) is an important signaling molecule modulating numerous physiological processes in plants. Here, we investigated the role of NO in Al toxicity to Hibiscus moscheutos. • Exposure of H. moscheutos to Al3+ led to a rapid inhibition of root elongation, and the inhibitory effect was alleviated by NO donor sodium nitroprusside (SNP). NO scavenger and inhibitors of NO synthase (NOS) and nitrate reductase had a similar inhibitory effect on root elongation. The inhibition of root elongation by these treatments was ameliorated by SNP. • Aluminum inhibited activity of NOS and reduced endogenous NO concentrations. The alleviation of inhibition of root elongation induced by Al, NO scavenger and NOS inhibitor was correlated with endogenous NO concentrations in root apical cells, suggesting that reduction of endogenous NO concentrations resulting from inhibition of NOS activity could underpin Al-induced arrest of root elongation in H. moscheutos. [ABSTRACT FROM AUTHOR]

Inhibition of root elongation by toxic aluminum (Al3+) occurs rapidly and is one of the most distinct and earliest symptoms of Al toxicity. To elucidate mechanism underlying Al3+-induced inhibition of root elongation, we investigated the involvement of ethylene in Al3+-induced inhibition of root elongation using the legume model plants Lotus japonicus and Medicago truncatula. Root elongation of L. japonicus and M. truncatula was rapidly inhibited by exposure to AlCl3. A similar rapid inhibition of root elongation by the ethylene-releasing substance, ethephon, and the ethylene precursor, 1-aminocyclopropane-1-carboxylic acid (ACC), was also observed. The Al3+-induced inhibition of root elongation was substantially ameliorated in the presence of antagonists of ethylene biosynthesis [Co2+ and aminoethoxyvinylglycine (AVG)]. Al3+ increased the activity of ACC oxidase (ACO), and induced a rapid evolution of ethylene from root apices and expression of genes of ACC synthase (ACS) and ACO. These findings suggest that induction of ethylene evolution resulting from up-regulation of ACS and ACO plays a critical role in Al3+-induced inhibition of root elongation. [ABSTRACT FROM AUTHOR]