9-cis Epoxycarotenoid Dioxygenase Research Articles

Dual PGPR-AMF Inoculation Offsets Salinity Stress Impact on the Fodder Halophyte Sulla carnosa by Concomitantly Modulating Plant ABA Content and Leaf Antioxidant Response

AbstractSalt-tolerant microbes are known to mitigate various biotic and abiotic stresses in plants. However, the intimate mechanisms involved, as well as their effects on the production of signaling molecules associated with the host plant–microbe interaction remain largely unknown. The present work aimed to investigate the role and potential uses of arbuscular mycorrhizal fungi (AMF) Rhizophagus intraradices and/or halotolerant plant growth-promoting rhizobacteria (PGPR) Bacillus subtilis in improving plant growth, functional biochemical synthesis and signaling of endogenous abscisic acid during plant response to short- and long-term salt stress in the forage halophyte Sulla carnosa. Plant growth attributes and biochemical traits were determined at 2 different time intervals (45 and 60 d after transplanting time) when salinity was raised from 100 to 200 mM NaCl. S. carnosa showed significant reduction in dry biomass in response to NaCl stress at the second harvest (200 mM NaCl); however inoculating plants with B. subtilis alone or associated with R. intraradices offset salt impact. Leaf electrolyte leakage was significantly increased by salinity but was significantly reduced following dual microbial inoculation. The applied bacterial inoculants also mitigated oxidative stress as reflected by the higher activities of catalase (APX) and superoxide dismutase (SOD) antioxidant enzymes and reduced H2O2 level. Inoculation with B. subtilis and R. intraradices upregulated 9-cisepoxycarotenoid dioxygenase 1 (NCED1) and SOD genes expression in S. carnosa plants upon salinity treatment. Furthermore, dual AMF-PGPR -inoculated plants accumulated significantly higher levels of abscisic acid (ABA) in both leaves and roots than non-inoculated and single inoculated plants under salinity stress at both harvest times, thereby accounting for their higher salt tolerance of salt-challenged S. carnosa plants. As a whole, the use of halophytic plants associated with beneficial soil microorganisms could improve the effectiveness of biological methods for saline soil rehabilitation. At the mechanistic level, ABA might represent a key player in the attenuation of salt impact in inoculated plants.

Expression profile of the NCED/CCD genes in chickpea and lentil during abiotic stress reveals a positive correlation with increased plant tolerance

Carotenoid cleavage dioxygenase (CCD) gene family is organized in two subfamilies: (i) 9-cis epoxycarotenoid dioxygenase (NCED) genes and (ii) CCD genes. NCED genes are essential for catalyzing the first step of the abscisic-acid (ABA) biosynthesis, while CCD genes produce precursors of the strigolactones hormone. The functional characterization of these gene subfamilies has not been yet performed in chickpea and lentil. Herein, were identified and systematically characterized two NCED and five CCD genes in the chickpea and two NCED and six CCD genes in lentil. After in silico sequence analysis and phylogeny, the expression profile of the NCED/CCD genes was determined by meta-analysis and real-time PCR in plants under different stress conditions. Sequence data revealed that NCED/CCD genes are highly conserved between chickpea and lentil. This conservation was observed both at gene and protein sequence levels and phylogenetic relationships. Analysis of the promoter sequences revealed that all NCED/CCD genes have a considerable number of cis-regulatory elements responsive to biotic and abiotic stress. Protein sequence analysis evidenced that NCED/CCD genes share several conserved motifs and that they have a highly interconnected interaction network. Furthermore, the three-dimensional structure of these proteins was determined and indicated that some proteins have structures with considerable similarity. The meta-analysis revealed that NCED/CCD genes are dynamically modulated in different organs and under different stress conditions, but they have a positive correlation with plant tolerance. In accordance, real-time PCR data showed that both NCED and CCD genes are differentially modulated in plants under drought stress. In particular, CaNCED2, CaCCD5, LcNCED2, LcCCD1, and LcCCD2 genes have a positive correlation with improved plant tolerance to drought stress. Therefore, this study presented a detailed characterization of the chickpea and lentil NCED/CCD genes and provided new insights to improve abiotic stress tolerance in these two important crops.

Genome-wide identification of the NCED gene family and functional characterization of PavNCED5 related to bud dormancy in sweet cherry

Abscisic acid (ABA) is involved in responding to various stresses and regulating the dormancy of flower buds in fruit trees. The cleavage of carotenoids is catalyzed by 9-cis epoxy carotenoid dioxygenase (NCED), and is a key step in ABA biosynthesis. Although NCED genes have been identified in a variety of plants, the characteristics of these genes in sweet cherry (Prunus avium L.) trees have been seldom reported. In the present study, 10 PavNCED genes were identified in the sweet cherry genome and distributed on three chromosomes. Cis-element analysis of PavNCEDs indicated that they might be related to hormone regulation, stress responses, and plant development. Through co-expression network analysis, the transcription factors were identified, such as ERF, bHLH and TCP, which might be involved in the regulation of PavNCEDs. Expression pattern analysis of the PavNCED family genes showed that PavNCED1–5 genes were highly expressed during the flower bud dormancy. Importantly, the expression of PavNCED5 was decreased with the decline of ABA content in the sweet cherry. Additionally, the overexpression of PavNCED5 in Arabidopsis showed dormancy deficiency of seeds. This implies that PavNCED5 may play an important role during the dormancy stage of sweet cherry. In general, we characterized the NCED family in sweet cherry and revealed the essential role of PavNCED5 in flower bud dormancy. These findings open up future research directions and provide valuable information for the further functional study of PavNCEDs in sweet cherry.

Molecular and Enzymatic Characterization of 9-Cis-epoxycarotenoid Dioxygenases from Mulberry.

Abscisic acid (ABA) is involved in many physiological regulatory processes in plants, such as leaf shedding, stomatal closure, inhibition of cell elongation, as well as responses to multi-abiotic stress, and 9-cis epoxy carotenoid dioxygenase (NCED) is related to the indirect synthesis of ABA. However, NCED genes involved in multi-abiotic stress and ABA synthesis pathway in mulberry (Morus alba L.) are still unknown. Here, two NCED genes cloned from mulberry (MaNCED) and their function were preliminarily identified. Interestingly, MaNCED2 responded strongly to drought stress while MaNCED1 responded strongly to pathogen stress. Then, two MaNCED proteins were successfully obtained by prokaryotic expression, and the degradation products of MaNCED1 and MaNCED2 were analyzed using UPLC-MS. The results show that recombinant MaNCED1 and MaNCED2 both cleave 9-cis-violaxanthin to form C15 xanthoxin,involved in the formation of the precursor of ABA.

DcCCD4 catalyzes the degradation of α-carotene and β-carotene to affect carotenoid accumulation and taproot color in carrot.

Carotenoids are important natural pigments that give bright colors to plants. The difference in the accumulation of carotenoids is one of the key factors in the formation of various colors in carrot taproots. Carotenoid cleavage dioxygenases (CCDs), including CCD and 9-cis epoxycarotenoid dioxygenase, are the main enzymes involved in the cleavage of carotenoids in plants. Seven CCD genes have been annotated from the carrot genome. In this study, through expression analysis, we found that the expression level of DcCCD4 was significantly higher in the taproot of white carrot (low carotenoid content) than orange carrot (high carotenoid content). The overexpression of DcCCD4 in orange carrots caused the taproot color to be pale yellow, and the contents of α- and β-carotene decreased sharply. Mutant carrot with loss of DcCCD4 function exhibited yellow color (the taproot of the control carrot was white). The accumulation of β-carotene was also detected in taproot. Functional analysis of the DcCCD4 enzyme in vitro showed that it was able to cleave α- and β-carotene at the 9, 10 (9', 10') double bonds. In addition, the number of colored chromoplasts in the taproot cells of transgenic carrots overexpressing DcCCD4 was significantly reduced compared with that in normal orange carrots. Results showed that DcCCD4 affects the accumulation of carotenoids through cleavage of α- and β-carotene in carrot taproot.

The abscisic acid receptor OsPYL6 confers drought tolerance to indica rice through dehydration avoidance and tolerance mechanisms.

Abscisic acid (ABA) is a key regulator of plant development and stress tolerance. Here we report functional validation of the ABA receptor OsPYL6 by constitutive and stress-inducible overexpression and RNAi silencing, in an indica rice cultivar 'Pusa Sugandh 2'. Overexpression of OsPYL6 conferred ABA hypersensitivity during germination and promoted total root length. Overexpression and RNAi silencing of OsPYL6 resulted in enhanced accumulation of ABA in seedlings under non-stress conditions, at least, in part through up-regulation of different 9-cis epoxycarotenoid dioxygenase (NCED )genes. This suggests that PYL6 expression is crucial for ABA homeostasis. Analysis of drought tolerance of OsPYL6 transgenic and wild type plants showed that OsPYL6 overexpression enhanced the expression of stress-responsive genes and dehydration tolerance. Transgenic rice plants overexpressing OsPYL6 with AtRD29A (Arabidopsis thaliana Responsive to Dehydration 29A) promoter also exhibited about 25% less whole plant transpiration, compared with wild type plants under drought, confirming its role in activation of dehydration avoidance mechanisms. However, overexpression of PYL6 reduced grain yield under non-stress conditions due to reduction in height, biomass, panicle branching and spikelet fertility. RNAi silencing of OsPYL6 also reduced grain yield under drought. These results showed that rice OsPYL6 is a key regulator of plant development and drought tolerance, and fine-tuning of its expression is critical for improving yield and stress tolerance.

Transcriptional Analysis of Carotenoids Accumulation and Metabolism in a Pink-Fleshed Lemon Mutant.

Pink lemon is a spontaneous bud mutation of lemon (Citrus limon, L. Burm. f) characterized by the production of pink-fleshed fruits due to an unusual accumulation of lycopene. To elucidate the genetic determinism of the altered pigmentation, comparative carotenoid profiling and transcriptional analysis of both the genes involved in carotenoid precursors and metabolism, and the proteins related to carotenoid-sequestering structures were performed in pink-fleshed lemon and its wild-type. The carotenoid profile of pink lemon pulp is characterized by an increased accumulation of linear carotenoids, such as lycopene, phytoene and phytofluene, from the early stages of development, reaching their maximum in mature green fruits. The distinctive phenotype of pink lemon is associated with an up-regulation and down-regulation of the genes upstream and downstream the lycopene cyclase, respectively. In particular, 9-cis epoxycarotenoid dioxygenase genes were overexpressed in pink lemon compared with the wild-type, suggesting an altered regulation of abscisic acid biosynthesis. Similarly, during early development of the fruits, genes of the carotenoid-associated proteins heat shock protein 21, fibrillin 1 and 2 and orange gene were overexpressed in the pulp of the pink-fleshed lemon compared to the wild-type, indicating its increased capacity for sequestration, stabilization or accumulation of carotenes. Altogether, the results highlighted significant differences at the transcriptomic level between the pink-fleshed lemon and its wild-type, in terms of carotenoid metabolism and the capacity of stabilization in storage structures between the two accessions. Such changes may be either responsible for the altered carotenoid accumulation or in contrast, a metabolic consequence.

Induced Tolerance to Salinity Stress by Halotolerant Bacteria Bacillus aryabhattai H19-1 and B. mesonae H20-5 in Tomato Plants.

Salinity is one of the major abiotic stresses that cause reduction of plant growth and crop productivity. It has been reported that plant growth-promoting bacteria (PGPB) could confer abiotic stress tolerance to plants. In a previous study, we screened bacterial strains capable of enhancing plant health under abiotic stresses and identified these strains based on 16s rRNA sequencing analysis. In this study, we investigated the effects of two selected strains, Bacillus aryabhattai H19-1 and B. mesonae H20-5, on responses of tomato plants against salinity stress. As a result, they alleviated decrease in plant growth and chlorophyll content; only strain H19-1 increased carotenoid content compared to that in untreated plants under salinity stress. Strains H19-1 and H20-5 significantly decreased electrolyte leakage, whereas they increased Ca2+ content compared to that in the untreated control. Our results also indicated that H20-5-treated plants accumulated significantly higher levels of proline, abscisic acid (ABA), and antioxidant enzyme activities compared to untreated and H19-1-treated plants during salinity stress. Moreover, strain H20-5 upregulated 9-cisepoxycarotenoid dioxygenase 1 (NCED1) and abscisic acid-response element-binding proteins 1 (AREB1) genes, otherwise strain H19-1 downregulated AREB1 in tomato plants after the salinity challenge. These findings demonstrated that strains H19-1 and H20-5 induced ABA-independent and -dependent salinity tolerance, respectively, in tomato plants, therefore these strains can be used as effective bio-fertilizers for sustainable agriculture.

Improvement in drought tolerance of lemon balm, Melissa officinalis L. under the pre-treatment of LED lighting

Stress priming (pre-exposure of plants to various types of moderate stresses) could affect plant responses to subsequent severe stresses. Drought stress is one of the major threats to plants which reduces the global agricultural productions. Here we demonstrated that light emitting diodes (LEDs)-driven tolerant to drought stress in lemon balm plantlets was highly correlated with priming with these lighting sources. Plantlets of the two genotypes of M. officinalis L. were first grown in 4 incubators with different LED lamps, including white LEDs (380–760 nm), blue LEDs (460 nm), red LEDs (650 nm) and red + blue LEDs (70%:30%), in a greenhouse for 4 weeks. The potted plants were then subjected to drought stress. Under drought stress, LED-primed plants maintained significantly higher fresh and dry weight, relative water content (RWC), concentration of soluble sugars, antioxidant activity and higher content of proline, H2O2, abscisic acid (ABA) and rosmarinic acid than non-primed plants. The results of Real-Time RT-PCR confirmed that LED pretreatment up-regulated the expression levels of respiratory burst oxidase homologues (RBOHs) or NADPH oxidase, 9-cis epoxy carotenoid dioxygenase (NCED), and rosmarinic acid synthase (RAS), while down-regulated that of ABA 8′-hydroxylase (ABA8Ox). These findings suggest, for the first time, that pre-treatment of plants with red + blue LEDs could improve their growth and quality under drought stress.

Multi-omics Analysis Reveals Sequential Roles for ABA during Seed Maturation.

Abscisic acid (ABA) is an important hormone for seed development and germination whose physiological action is modulated by its endogenous levels. Cleavage of carotenoid precursors by 9-cis epoxycarotenoid dioxygenase (NCED) and inactivation of ABA by ABA 8'-hydroxylase (CYP707A) are key regulatory metabolic steps. In Arabidopsis (Arabidopsis thaliana), both enzymes are encoded by multigene families, having distinctive expression patterns. To evaluate the genome-wide impact of ABA deficiency in developing seeds at the maturation stage when dormancy is induced, we used a nced2569 quadruple mutant in which ABA deficiency is mostly restricted to seeds, thus limiting the impact of maternal defects on seed physiology. ABA content was very low in nced2569 seeds, similar to the severe mutant aba2; unexpectedly, ABA Glc ester was detected in aba2 seeds, suggesting the existence of an alternative metabolic route. Hormone content in nced2569 seeds compared with nced259 and wild type strongly suggested that specific expression of NCED6 in the endosperm is mainly responsible for ABA production. In accordance, transcriptome analyses revealed broad similarities in gene expression between nced2569 and either wild-type or nced259 developing seeds. Gene ontology enrichments revealed a large spectrum of ABA activation targets involved in reserve storage and desiccation tolerance, and repression of photosynthesis and cell cycle. Proteome and metabolome profiles in dry nced2569 seeds, compared with wild-type and cyp707a1a2 seeds, also highlighted an inhibitory role of ABA on remobilization of reserves, reactive oxygen species production, and protein oxidation. Down-regulation of these oxidative processes by ABA may have an essential role in dormancy control.

The roles of cell wall invertase inhibitor in regulating chilling tolerance in tomato

BackgroundHexoses are important metabolic signals that respond to abiotic and biotic stresses. Cold stress adversely affects plant growth and development, limiting productivity. The mechanism by which sugars regulate plant cold tolerance remains elusive.ResultsWe examined the function of INVINH1, a cell wall invertase inhibitor, in tomato chilling tolerance. Cold stress suppressed the transcription of INVINH1 and increased that of cell wall invertase genes, Lin6 and Lin8 in tomato seedlings. Silencing INVINH1 expression in tomato increased cell wall invertase activity and enhanced chilling tolerance. Conversely, transgenic tomatoes over-expressing INVINH1 showed reduced cell wall invertase activity and were more sensitive to cold stress. Chilling stress increased glucose and fructose levels, and the hexoses content increased or decreased by silencing or overexpression INVINH1. Glucose applied in vitro masked the differences in chilling tolerance of tomato caused by the different expressions of INVINH1. The repression of INVINH1 or glucose applied in vitro regulated the expression of C-repeat binding factors (CBFs) genes. Transcript levels of NCED1, which encodes 9-cisepoxycarotenoid dioxygenase (NCED), a key enzyme in the biosynthesis of abscisic acid, were suppressed by INVINH1 after exposure to chilling stress. Meanwhile, application of ABA protected plant from chilling damage caused by the different expression of INVINH1.ConclusionsIn tomato, INVINH1 plays an important role in chilling tolerance by adjusting the content of glucose and expression of CBFs.

Physiological and structural traits of the congeneric species Styrax ferrugineus and S. pohlii occurring in contrasting environments

Abscisic acid (ABA) attenuates stomatal conductance (gs) and transpiration (E) in response to drought. Broadly known as ‘Brazilian savanna’, the Cerrado sensu lato is a mosaic of landscapes, including savanna-type vegetation, such as the cerrado sensu stricto (s. str.), and riparian forests occurring along rivers. These habitats share floristic elements, and congeneric species are adapted to grow in these drought- and flood-prone environments. We assessed leaf concentration of ABA and its metabolites, as well as gs, E and leaf water potential (Ψw) in shrubs of Styrax ferrugineus in a cerrado s. str. (savanna species) and in trees of S. pohlii in a riparian forest (forest species), in both the wet and dry seasons. We also quantified 9-cis epoxycarotenoid dioxygenase (NCED) gene expression in leaves, and wood anatomical features in stems. We expected that the forest species would not increase their leaf ABA content during the dry season because of the moist soil from the riparian forest. Regardless of the environment, both species expressed NCED, synthesized ABA, and exhibited reductions in both Ψw and gs in the dry season, confirming these as universal responses to the dry season. The savanna species had more xylem vessels per mm2 and these vessels were narrower than those observed in stems of the forest species, resulting in a low vulnerability to cavitation (lvul) estimated for both species. Our results demonstrate that, despite inhabiting contrasting habitats, these species display similar ecophysiological responses to the dry season and their wood anatomical features may explain their geographical distribution.

From pond slime to rain forest: the evolution of ABA signalling and the acquisition of dehydration tolerance.

Keywords: abscisic acid (ABA); comparative genomics; dehydration tolerance; land plant evolution; Physcomitrella patens ; zeaxanthin epoxidase

Light and abscisic acid independently regulated FaMYB10 in Fragaria × ananassa fruit.

Light and ABA independently regulated anthocyanin biosynthesis via activation of FaMYB10 expression. FaMYB10 accelerated anthocyanin synthesis of pelargonidin 3-glucoside and cyanidin 3-glucoside during strawberry fruit ripening. Light is an integral factor in fruit ripening. Ripening in non-climacteric fruit is also effected by the plant hormone abscisic acid (ABA). However, how light and/or ABA regulate fruit ripening processes, such as strawberry color development remains elusive. Results of the present study showed light and ABA regulated strawberry fruit coloration via activation of FaMYB10 expression, an R2R3 MYB transcription factor. Light exposure increased FaMYB10 transcript levels, flavonoid pathway genes, and anthocyanin content. Exogenous ABA promoted FaMYB10 expression, and anthocyanin content, accompanied by increased ABA-responsive transcript levels and flavonoid pathway genes. ABA biosynthesis inhibitor treatment, and RNAi-mediated down-regulation of the ABA biosynthetic gene (9-cis epoxycarotenoid dioxygenase: FaNCED1), and ABA receptor (magnesium chelatase H subunit: FaCHLH/ABAR) showed inverse ABA effects. Furthermore, additive effects were observed in anthocyanin accumulation under combined light and ABA, indicating independent light and ABA signaling pathways. FaMYB10 down-regulation by Agrobacterium-mediated RNA interference (RNAi) in strawberry fruits showed decreased pelargonidin 3-glucoside and cyanidin 3-glucoside levels, accompanied by consistent flavonoid pathway gene expression levels. FaMYB10 over-expression showed opposite FaMYB10 RNAi phenotypes, particularly cyanidin 3-glucoside synthesis by FaMYB10, which was correlated with FaF3'H transcript levels. These data provided evidence that light and ABA promoted FaMYB10 expression, resulting in anthocyanin accumulation via acceleration of flavonoid pathway gene expression. Finally, our results suggested FaMYB10 serves a role as a signal transduction mediator from light and ABA perception to anthocyanin synthesis in strawberry fruit.

Validation of reference genes for accurate normalization of gene expression for real time-quantitative PCR in strawberry fruits using different cultivars and osmotic stresses

The increasing demand of strawberry (Fragaria×ananassa Duch) fruits is associated mainly with their sensorial characteristics and the content of antioxidant compounds. Nevertheless, the strawberry production has been hampered due to its sensitivity to abiotic stresses. Therefore, to understand the molecular mechanisms highlighting stress response is of great importance to enable genetic engineering approaches aiming to improve strawberry tolerance. However, the study of expression of genes in strawberry requires the use of suitable reference genes. In the present study, seven traditional and novel candidate reference genes were evaluated for transcript normalization in fruits of ten strawberry cultivars and two abiotic stresses, using RefFinder, which integrates the four major currently available software programs: geNorm, NormFinder, BestKeeper and the comparative delta-Ct method. The results indicate that the expression stability is dependent on the experimental conditions. The candidate reference gene DBP (DNA binding protein) was considered the most suitable to normalize expression data in samples of strawberry cultivars and under drought stress condition, and the candidate reference gene HISTH4 (histone H4) was the most stable under osmotic stresses and salt stress. The traditional genes GAPDH (glyceraldehyde-3-phosphate dehydrogenase) and 18S (18S ribosomal RNA) were considered the most unstable genes in all conditions. The expression of phenylalanine ammonia lyase (PAL) and 9-cis epoxycarotenoid dioxygenase (NCED1) genes were used to further confirm the validated candidate reference genes, showing that the use of an inappropriate reference gene may induce erroneous results. This study is the first survey on the stability of reference genes in strawberry cultivars and osmotic stresses and provides guidelines to obtain more accurate RT-qPCR results for future breeding efforts.

Molecular characterization of carotenoid biosynthetic genes and carotenoid accumulation in Lycium chinense.

Lycium chinense is a shrub that has health benefits and is used as a source of medicines in Asia. In this study, a full-length cDNA clone encoding β-ring carotene hydroxylase (LcCHXB) and partial-length cDNA clones encoding phytoene synthase (LcPSY), phytoene desaturase (LcPDS), ξ-carotene desaturase (LcZDS), lycopene β-cyclase (LcLCYB), lycopene ε-cyclase (LcLCYE), ε-ring carotene hydroxylase (LcCHXE), zeaxanthin epoxidase (LcZEP), carotenoid cleavage dioxygenase (LcCCD1), and 9-cis epoxycarotenoid dioxygenase (LcNCED) were identified in L. chinense. The transcripts were constitutively expressed at high levels in leaves, flowers and red fruits, where the carotenoids are mostly distributed. In contrast, most of the carotenoid biosynthetic genes were weakly expressed in the roots and stems, which contained only small amounts of carotenoids. The level of LcLCYE transcripts was very high in leaves and correlated with the abundance of lutein in this plant tissue. During maturation, the levels of lutein and zeaxanthin in L. chinense fruits dramatically increased, concomitant with a rise in the level of β-cryptoxanthin. LcPSY, LcPDS, LcZDS, LcLCYB, and LcCHXE were highly expressed in red fruits, leading to their substantially higher total carotenoid content compared to that in green fruits. Total carotenoid content was high in both the leaves and red fruits of L. chinense. Our findings on the biosynthesis of carotenoids in L. chinense provide insights into the molecular mechanisms involved in carotenoid biosynthesis and may facilitate the optimization of carotenoid production in L. chinense.

Profiling microRNAs and their targets in an important fleshy fruit: Tomato (Solanum lycopersicum)

Tomato (Solanum lycopersicum) is an important and the most useful plant based diet. It is widely used for its antioxidant property. Presently, only two digits, tomato microRNAs (miRNAs) are reported in miRBase: a miRNA database. This study is aimed to profile and characterize more miRNAs and their targets in tomato. A comprehensive comparative genomic approach is applied and a total of 109 new miRNAs belonging to 106 families are identified and characterized from the tomato expressed sequence tags (ESTs). All these potential miRNAs are profiled for the first time in tomato. The profiled miRNAs are also observed with stable stem-loop structures (Precursor-miRNAs), whose length ranges from 45to 329 nucleotides (nt) with an average of 125 nt. The mature miRNAs are found in the stem of pre-miRNAs and their length ranges from 19 to 24 nt with an average of 21 nt. Furthermore, twelve miRNAs are randomly selected and experimentally validated through RT-PCR. A total of 406 putative targets are also predicted for the newly 109 tomato miRNAs. These targets are involved in structural protein, metabolism, transcription factor, growth & development, stress related, signaling pathways, storage proteins and other vital processes. Some important proteins like; 9-cisepoxycarotenoid dioxygenase (NCED), transcription factor MYB, ATP-binding cassette transporters, terpen synthase, 14-3-3 and TIR-NBS proteins are also predicted as putative targets for tomato miRNAs. These findings improve a baseline data of miRNAs and their targets in tomato. This baseline data can be utilized to fine tune this important fleshy fruit for nutritional & antioxidant properties and also under biotic & abiotic stresses.

Tissue specific distribution of ABA from Arachis hyopgaea L. contribute to expression of Arachis hypogaea 9-cis epoxycarotenoid dioxygenase 1 (Ahnced1) during development

Tissue specific distribution of ABA from Arachis hyopgaea L. contribute to expression of Arachis hypogaea 9-cis epoxycarotenoid dioxygenase 1 (Ahnced1) during development

Comparative Study of the Tissue-Specific Distribution of ABA from Arachis Hypogaea L. and Expression of the 9-CIS Epoxycarotenoid Dioxygenase 1 (AhNCED1) During Plant Development

ABSTRACTThe oxidative cleavage of cis-epoxycarotenoids catalyzed by 9-cis-epoxycarotenoid dioxygenase (NCED) is considered to be the rate-limiting step in abscisic acid (ABA) biosynthesis. In this study the tissue-specific distribution of ABA in peanut plants during development, and the relationship and influence between tissue-specific distribution of ABA and tissue-specific expression of AhNCED1 were studied by immunoenzyme localization. At the four-leaf stage, ABA and AhNCED1 immunohistochemical specific staining was detected in the root vascular parenchyma. During the blooming stage, there was no ABA and AhNCED1 immunohistochemical specific staining in any of the analyzed organs (leaf, stem, including root). However, ABA immunohistochemical specific staining was detected in the leaf vascular parenchyma during the fruiting stage. The organ-specific pattern of AhNCED1 mRNA and protein expression under water stress was consistent with that of ABA accumulation. The results suggest that ABA biosynthesis in peanut plants occurs predominantly in the vascular parenchyma cells. In peanut plants, initial ABA biosynthesis occurs in different organs during the different developmental stages: the root is the initial organ of ABA biosynthesis at the four-leaf stage, whereas leaves play this role at the fruiting stages.

The plant cuticle is required for osmotic stress regulation of abscisic acid biosynthesis and osmotic stress tolerance in Arabidopsis.

Osmotic stress activates the biosynthesis of abscisic acid (ABA). One major step in ABA biosynthesis is the carotenoid cleavage catalyzed by a 9-cis epoxycarotenoid dioxygenase (NCED). To understand the mechanism for osmotic stress activation of ABA biosynthesis, we screened for Arabidopsis thaliana mutants that failed to induce the NCED3 gene expression in response to osmotic stress treatments. The ced1 (for 9-cis epoxycarotenoid dioxygenase defective 1) mutant isolated in this study showed markedly reduced expression of NCED3 in response to osmotic stress (polyethylene glycol) treatments compared with the wild type. Other ABA biosynthesis genes are also greatly reduced in ced1 under osmotic stress. ced1 mutant plants are very sensitive to even mild osmotic stress. Map-based cloning revealed unexpectedly that CED1 encodes a putative α/β hydrolase domain-containing protein and is allelic to the BODYGUARD gene that was recently shown to be essential for cuticle biogenesis. Further studies discovered that other cutin biosynthesis mutants are also impaired in osmotic stress induction of ABA biosynthesis genes and are sensitive to osmotic stress. Our work demonstrates that the cuticle functions not merely as a physical barrier to minimize water loss but also mediates osmotic stress signaling and tolerance by regulating ABA biosynthesis and signaling.