Plants have evolved a plethora of responses to cope with phosphate (Pi) deficiency, including the transcriptional activation of a large set of genes. Among Pi-responsive genes, the expression of the Arabidopsis phospholipase DZ2 (PLDZ2) is activated to participate in the degradation of phospholipids in roots in order to release Pi to support other cellular activities. A deletion analysis was performed to identify the regions determining the strength, tissue-specific expression, and Pi responsiveness of this regulatory region. This study also reports the identification and characterization of a transcriptional enhancer element that is present in the PLDZ2 promoter and able to confer Pi responsiveness to a minimal, inactive 35S promoter. This enhancer also shares the cytokinin and sucrose responsive properties observed for the intact PLDZ2 promoter. The EZ2 element contains two P1BS motifs, each of which is the DNA binding site of transcription factor PHR1. Mutation analysis showed that the P1BS motifs present in EZ2 are necessary but not sufficient for the enhancer function, revealing the importance of adjacent sequences. The structural organization of EZ2 is conserved in the orthologous genes of at least eight families of rosids, suggesting that architectural features such as the distance between the two P1BS motifs are also important for the regulatory properties of this enhancer element.
enhancer element; gene regulation; phosphate deprivation; phospholipase; root
Although the mechanisms of nodule N2 fixation in legumes are now well documented, some uncertainty remains on the metabolic consequences of water deficit. In most cases, little consideration is given to other organs and, therefore, the coordinated changes in metabolism in leaves, roots, and nodules are not well known. Here, the effect of water restriction on exclusively N2-fixing alfalfa (Medicago sativa L.) plants was investigated, and proteomic, metabolomic, and physiological analyses were carried out. It is shown that the inhibition of nitrogenase activity caused by water restriction was accompanied by concerted alterations in metabolic pathways in nodules, leaves, and roots. The data suggest that nodule metabolism and metabolic exchange between plant organs nearly reached homeostasis in asparagine synthesis and partitioning, as well as the N demand from leaves. Typically, there was (i) a stimulation of the anaplerotic pathway to sustain the provision of C skeletons for amino acid (e.g. glutamate and proline) synthesis; (ii) re-allocation of glycolytic products to alanine and serine/glycine; and (iii) subtle changes in redox metabolites suggesting the implication of a slight oxidative stress. Furthermore, water restriction caused little change in both photosynthetic efficiency and respiratory cost of N2 fixation by nodules. In other words, the results suggest that under water stress, nodule metabolism follows a compromise between physiological imperatives (N demand, oxidative stress) and the lower input to sustain catabolism.
Alfalfa; C/N; drought; metabolomic; nodule; proteomic
Glutathione is a tripeptide involved in various aspects of plant metabolism. This study investigated the effects of the reduced form of glutathione (GSH) applied to specific organs (source leaves, sink leaves, and roots) on cadmium (Cd) distribution and behaviour in the roots of oilseed rape plants (Brassica napus) cultured hydroponically. The translocation ratio of Cd from roots to shoots was significantly lower in plants that had root treatment of GSH than in control plants. GSH applied to roots reduced the Cd concentration in the symplast sap of root cells and inhibited root-to-shoot Cd translocation via xylem vessels significantly. GSH applied to roots also activated Cd efflux from root cells to the hydroponic solution. Inhibition of root-to-shoot translocation of Cd was visualized, and the activation of Cd efflux from root cells was also shown by using a positron-emitting tracer imaging system (PETIS). This study investigated a similar inhibitory effect on root-to-shoot translocation of Cd by the oxidized form of glutathione, GSSG. Inhibition of Cd accumulation by GSH was abolished by a low-temperature treatment. Root cells of plants exposed to GSH in the root zone had less Cd available for xylem loading by actively excluding Cd from the roots. Consequently, root-to-shoot translocation of Cd was suppressed and Cd accumulation in the shoot decreased.
Cadmium (Cd); glutathione; oilseed rape (Brassica napus); oxidized form of glutathione (GSSG); positron-emitting tracer imaging system (PETIS); reduced form of glutathione; transport; xylem.
The fruit of melting-flesh peach (Prunus persica L. Batsch) cultivars produce high levels of ethylene caused by high expression of PpACS1 (an isogene of 1-aminocyclopropane-1-carboxylic acid synthase), resulting in rapid fruit softening at the late-ripening stage. In contrast, the fruit of stony hard peach cultivars do not soften and produce little ethylene due to low expression of PpACS1. To elucidate the mechanism for suppressing PpACS1 expression in stony hard peaches, a microarray analysis was performed. Several genes that displayed similar expression patterns as PpACS1 were identified and shown to be indole-3-acetic acid (IAA)-inducible genes (Aux/IAA, SAUR). That is, expression of IAA-inducible genes increased at the late-ripening stage in melting flesh peaches; however, these transcripts were low in mature fruit of stony hard peaches. The IAA concentration increased suddenly just before harvest time in melting flesh peaches exactly coinciding with system 2 ethylene production. In contrast, the IAA concentration did not increase in stony hard peaches. Application of 1-naphthalene acetic acid, a synthetic auxin, to stony hard peaches induced a high level of PpACS1 expression, a large amount of ethylene production and softening. Application of an anti-auxin, α-(phenylethyl-2-one)-IAA, to melting flesh peaches reduced levels of PpACS1 expression and ethylene production. These observations indicate that suppression of PpACS1 expression at the late-ripening stage of stony hard peach may result from a low level of IAA and that a high concentration of IAA is required to generate a large amount of system 2 ethylene in peaches.
ACC synthase; auxin; ethylene; softening; stony hard peach.
Protein tyrosine nitration is a post-translational modification mediated by reactive nitrogen species (RNS) that is associated with nitro-oxidative damage. No information about this process is available in relation to higher plants during development and senescence. Using pea plants at different developmental stages (ranging from 8 to 71 days), tyrosine nitration in the main organs (roots, stems, leaves, flowers, and fruits) was analysed using immunological and proteomic approaches. In the roots of 71-day-old senescent plants, nitroproteome analysis enabled the identification a total of 16 nitrotyrosine-immunopositive proteins. Among the proteins identified, NADP-isocitrate dehydrogenase (ICDH), an enzyme involved in the carbon and nitrogen metabolism, redox regulation, and responses to oxidative stress, was selected to evaluate the effect of nitration. NADP-ICDH activity fell by 75% during senescence. Analysis showed that peroxynitrite inhibits recombinant cytosolic NADP-ICDH activity through a process of nitration. Of the 12 tyrosines present in this enzyme, mass spectrometric analysis of nitrated recombinant cytosolic NADP-ICDH enabled this study to identify the Tyr392 as exclusively nitrated by peroxynitrite. The data as a whole reveal that protein tyrosine nitration is a nitric oxide-derived PTM prevalent throughout root development and intensifies during senescence.
Nitration; nitric oxide; nitrotyrosine; peroxynitrite; plant development; reactive nitrogen species; senescence
The coordination of plant cell division and expansion controls plant morphogenesis, development, and growth. Cyclin-dependent kinases (CDKs) are not only key regulators of cell division but also play an important role in cell differentiation. In plants, CDK activity is modulated by the binding of INHIBITOR OF CDK/KIP-RELATED PROTEIN (ICK/KRP). Previously, ICK2/KRP2 has been shown to mediate auxin responses in lateral root initiation. Here are analysed the roles of all ICK/KRP genes in root growth. Analysis of ick/krp null-mutants revealed that only ick3/krp5 was affected in primary root growth. ICK3/KRP5 is strongly expressed in the root apical meristem (RAM), with lower expression in the expansion zone. ick3/krp5 roots grow more slowly than wildtype controls, and this results not from reduction of division in the proliferative region of the RAM but rather reduced expansion as cells exit the meristem. This leads to shorter final cell lengths in different tissues of the ick3/krp5 mutant root, particularly the epidermal non-hair cells, and this reduction in cell size correlates with reduced endoreduplication. Loss of ICK3/KRP5 also leads to delayed germination and in the mature embryo ICK3/KRP5 is specifically expressed in the transition zone between root and hypocotyl. Cells in the transition zone were smaller in the ick3/krp5 mutant, despite the absence of endoreduplication in the embryo suggesting a direct effect of ICK3/KRP5 on cell growth. It is concluded that ICK3/KRP5 is a positive regulator of both cell growth and endoreduplication.
Arabidopsis; CDK; cell cycle; cell division; cell expansion; development; root growth
Symbiotic nitrogen fixation (SNF) involves global changes in gene expression and metabolite accumulation in both rhizobia and the host plant. In order to study the metabolic changes mediated by leaf–root interaction, photosynthesis was limited in leaves by exposure of plants to darkness, and subsequently gene expression was profiled by real-time reverse transcription–PCR (RT–PCR) and metabolite levels by gas chromatography–mass spectrometry in the nodules of the model legume Lotus japonicus. Photosynthetic carbon deficiency caused by prolonged darkness affected many metabolic processes in L. japonicus nodules. Most of the metabolic genes analysed were down-regulated during the extended dark period. In addition to that, the levels of most metabolites decreased or remained unaltered, although accumulation of amino acids was observed. Reduced glycolysis and carbon fixation resulted in lower organic acid levels, especially of malate, the primary source of carbon for bacteroid metabolism and SNF. The high amino acid concentrations together with a reduction in total protein concentration indicate possible protein degradation in nodules under these conditions. Interestingly, comparisons between amino acid and protein content in various organs indicated systemic changes in response to prolonged darkness between nodulated and non-nodulated plants, rendering the nodule a source organ for both C and N under these conditions.
Carbon starvation; Lotus japonicus; metabolomic analysis; nodule; symbiosis; transcript profile.
In several dicotyledonous species, NAC transcription factors act as master switches capable of turning on programmes of secondary cell-wall synthesis and cell death. This work used an oestradiol-inducible system to overexpress the NAC transcription factor BdSWN5 in the monocot model Brachypodium distachyon. This resulted in ectopic secondary cell-wall formation in both roots and shoots. Some of the genes upregulated in the process were a secondary cell-wall cellulose synthase (BdCESA4), a xylem-specific protease (BdXCP1) and an orthologue of AtMYB46 (BdMYB1). While activation of BdMYB1 may not be direct, this study showed that BdSWN5 is capable of transactivating the BdXCP1 promoter through two conserved binding sites. In the course of Brachypodium development, the BdXCP1 promoter was observed to be active in all types of differentiating tracheary elements. Together, these results suggest that Brachypodium SWNs can act as switches that turn on secondary cell-wall synthesis and programmed cell death.
Brachypodium distachyon; cell death; monocot; NAC transcription factor; secondary wall; xylem
The adaptations of root morphology, physiology, and biochemistry to phosphorus supply have been characterized intensively. However, characterizing these adaptations at molecular level is largely neglected under field conditions. Here, two consecutive field experiments were carried out to investigate the agronomic traits and root traits of wheat (Triticum aestivum L.) at six P-fertilizer rates. Root samples were collected at flowering to investigate root dry weight, root length density, arbusular-mycorrhizal colonization rate, acid phosphatase activity in rhizosphere soil, and expression levels of genes encoding phosphate transporter, phosphatase, ribonucleases, and expansin. These root traits exhibited inducible, inhibitory, or combined responses to P deficiency, and the change point for responses to P supply was at or near the optimal P supply for maximum grain yield. This research improves the understanding of mechanisms of plant adaptation to soil P in intensive agriculture and provides useful information for optimizing P management based on the interactions between soil P dynamics and root processes.
Agronomic trait; phosphatase; phosphate-starvation response; phosphate transporter; phosphorus fertilizer; root morphology; Triticum aestivum L.
Arginine is an important medium for the transport and storage of nitrogen, and arginase (also known as arginine amidohydrolase, ARGAH) is responsible for catalyse of arginine into ornithine and urea in plants. In this study, the impact of AtARGAHs on abiotic stress response was investigated by manipulating AtARGAHs expression. In the knockout mutants of AtARGAHs, enhanced tolerances were observed to multiple abiotic stresses including water deficit, salt, and freezing stresses, while AtARGAH1- and AtARGAH2-overexpressing lines exhibited reduced abiotic stress tolerances compared to the wild type. Consistently, the enhanced tolerances were confirmed by the changes of physiological parameters including electrolyte leakage, water loss rate, stomatal aperture, and survival rate. Interestingly, the direct downstream products of arginine catabolism including polyamines and nitric oxide (NO) concentrations significantly increased in the AtARGAHs-knockout lines, but decreased in overexpressing lines under control conditions. Additionally, the AtARGAHs-overexpressing and -knockout lines displayed significantly reduced relative arginine (% of total free amino acids) relative to the wild type. Similarly, reactive oxygen species accumulation was remarkably regulated by AtARGAHs under abiotic stress conditions, as shown from hydrogen peroxide (H2O2), superoxide radical () concentrations, and antioxidant enzyme activities. Taken together, this is the first report, as far as is known, to provide evidence that AtARGAHs negatively regulate many abiotic stress tolerances, at least partially, attribute to their roles in modulating arginine metabolism and reactive oxygen species accumulation. Biotechnological strategy based on manipulation of AtARGAHs expression will be valuable for future crop breeding.
Abiotic stress; arginase; arginine metabolism; nitric oxide; polyamines; reactive oxygen species.
Reactive oxygen species (ROS) production is a common denominator in a variety of biotic and abiotic stresses, including salinity. In recent years, haem oxygenase (HO; EC 18.104.22.168) has been described as an important component of the antioxidant defence system in both mammalian and plant systems. Moreover, a recent report on Arabidopsis demonstrated that HO overexpression resulted in an enhanced salinity tolerance in this species. However, physiological mechanisms and downstream targets responsible for the observed salinity tolerance in these HO mutants remain elusive. To address this gap, ion transport characteristics (K+ and H+ fluxes and membrane potentials) and gene expression profiles in the roots of Arabidopsis thaliana HO-overexpressing (35S:HY1-1/2/3/4) and loss-of-function (hy-100, ho2, ho3, and ho4) mutants were compared during salinity stress. Upon acute salt stress, HO-overexpressing mutants retained more K+ (less efflux), and exhibited better membrane potential regulation (maintained more negative potential) and higher H+ efflux activity in root epidermis, compared with loss-of-function mutants. Pharmacological experiments suggested that high activity of the plasma membrane H+-ATPase in HO overexpressor mutants provided the proton-motive force required for membrane potential maintenance and, hence, better K+ retention. The gene expression analysis after 12h and 24h of salt stress revealed high expression levels of H+-ATPases (AHA1/2/3) and Na+/H+ antiporter [salt overly sensitive1 (SOS1)] transcripts in the plasma membrane of HO overexpressors. It is concluded that HO modifies salinity tolerance in Arabidopsis by controlling K+ retention via regulation of the plasma membrane H+-ATPase and by altering SOS1 transcript levels in roots.
Gene expression; H+-ATPase; haem oxygenase; ion fluxes; membrane potential; potassium; sodium.
Maximum and minimum stomatal conductance, as well as stomatal size and rate of response, are known to vary widely across plant species, but the functional relationship between these static and dynamic stomatal properties is unknown. The objective of this study was to test three hypotheses: (i) operating stomatal conductance under standard conditions (g
op) correlates with minimum stomatal conductance prior to morning light [g
min(dawn)]; (ii) stomatal size (S) is negatively correlated with g
op and the maximum rate of stomatal opening in response to light, (dg/dt)max; and (iii) g
op correlates negatively with instantaneous water-use efficiency (WUE) despite positive correlations with maximum rate of carboxylation (Vc
max) and light-saturated rate of electron transport (J
max). Using five closely related species of the genus Banksia, the above variables were measured, and it was found that all three hypotheses were supported by the results. Overall, this indicates that leaves built for higher rates of gas exchange have smaller stomata and faster dynamic characteristics. With the aid of a stomatal control model, it is demonstrated that higher g
op can potentially expose plants to larger tissue water potential gradients, and that faster stomatal response times can help offset this risk.
Maximum stomatal conductance; night-time conductance; stomatal control; stomatal size; transpiration; water-use efficiency.
Upward leaf movement, called hyponastic growth, is employed by plants to cope with adverse environmental conditions. Ethylene is a key regulator of this process and, in Arabidopsis thaliana, hyponasty is induced by this phytohormone via promotion of epidermal cell expansion in a proximal zone of the abaxial side of the petiole. ROTUNDIFOLIA3/CYP90C1 encodes an enzyme which was shown to catalyse C-23 hydroxylation of several brassinosteroids (BRs) – phytohormones involved in, for example, organ growth, cell expansion, cell division, and responses to abiotic and biotic stresses. This study tested the interaction between ethylene and BRs in regulating hyponastic growth. A mutant isolated in a forward genetic screen, with reduced hyponastic response to ethylene treatment, was allelic to rot3. The cause of the reduced hyponastic growth in this mutant was examined by studying ethylene–BR interaction during local cell expansion, pharmacological inhibition of BR synthesis and ethylene effects on transcription of BR-related genes. This work demonstrates that rot3 mutants are impaired in local cell expansion driving hyponasty. Moreover, the inhibition of BR biosynthesis reduces ethylene-induced hyponastic growth and ethylene increases sensitivity to BR in promoting cell elongation in Arabidopsis hypocotyls. Together, the results show that ROT3 modulates ethylene-induced petiole movement and that this function is likely BR related.
Arabidopsis; brassinosteroids; cell expansion; ethylene; hyponasty; ROTUNDIFOLIA3
Polyploidy is very common within angiosperms, and several studies are in progress to ascertain the effects of early polyploidization at the molecular, physiological, and phenotypic level. Extensive studies are available only in synthetic allopolyploids. By contrast, less is known about the consequences of autopolyploidization. The current study aimed to assess the occurrence and extent of genetic, epigenetic, and anatomical changes occurring after oryzaline-induced polyploidization of Solanum commersonii Dunal and Solanum bulbocastanum Dunal, two diploid (2n=2×=24) potato species widely used in breeding programmes. Microsatellite analysis showed no polymorphisms between synthetic tetraploids and diploid progenitors. By contrast, analysis of DNA methylation levels indicated that subtle alterations at CG and CHG sites were present in tetraploids of both species. However, no change occurred concurrently in all tetraploids analysed with respect to their diploid parent, revealing a stochastic trend in the changes observed. The morpho-anatomical consequences of polyploidization were studied in leaf main veins and stomata. With only a few exceptions, analyses showed no clear superiority of tetraploids in terms of leaf thickness and area, vessel number, lumen size and vessel wall thickness, stomata pore length and width, guard cell width, and stomatal density compared with their diploid progenitors. These results are consistent with the hypothesis that there are no traits systematically associated with autopolyploidy.
chromosome doubling; methylation; microsatellite; stomata; vessels; potato.
Three phytohormone molecules – ethylene (ET), jasmonic acid (JA) and salicylic acid (SA) – play key roles in mediating disease response to necrotrophic fungal pathogens. This study investigated the roles of the ET, JA, and SA pathways as well as their crosstalk during the interaction between tomato (Solanum lycopersicum) plants and a necrotrophic fungal pathogen Alternaria alternata f. sp. lycopersici (AAL). Both the ET and JASMONIC ACID INSENSITIVE1 (JAI1) receptor-dependent JA signalling pathways are necessary for susceptibility, while SA response promotes resistance to AAL infection. In addition, the role of JA in susceptibility to AAL is partly dependent on ET biosynthesis and perception, while the SA pathway enhances resistance to AAL and antagonizes the ET response. Based on these results, it is proposed that ET, JA, and SA each on their own can influence the susceptibility of tomato to AAL. Furthermore, the functions of JA and SA in susceptibility to the pathogen are correlated with the enhanced or decreased action of ET, respectively. This study has revealed the functional relationship among the three key hormone pathways in tomato defence against AAL.
AAL; ethylene; hormonal interactions; jasmonic acid; salicylic acid; tomato
Seed germination and flowering initiation are both transitions responding to similar seasonal cues. This study shows that ABSCISIC ACID-INSENSITIVE MUTANT 5 (ABI5), a bZIP transcription factor, which plays an important role in the abscisic acid (ABA)-arrested seed germination, is robustly associated with the floral transition in Arabidopsis. Under long-day conditions, overexpression of ABI5 could delay floral transition through upregulating FLOWERING LOCUS C (FLC) expression. In contrast, ectopically overexpressing FLC in an abi5 mutant reversed the earlier flowering phenotype. Further analysis indicated that transactivation of FLC could be promoted by ABI5 and/or other abscisic acid-responsive element (ABRE)-binding factors (ABFs). The expression of FLC that was promoted by ABI5 and/or other ABFs could be blocked in a triple SNF1-related protein kinase (SnRK) mutant, snrk2.2/2.3/2.6, despite the presence of ABA. In sharp contrast, when SnRK2.6 was coexpressed, the reduction of transactivity of FLC was reverted in mesophyll protoplasts of snrk2.2/2.3/2.6. Additional results from analysing transgenic plants carrying mutations of phosphoamino acids (ABI5
S42AS145AT201A), which are conserved in ABI5, suggested that SnRK2-mediated ABI5 and/or ABF phosphorylation may be crucial for promoting FLC expression. The transgenic plants ABI5
S42AS145AT201A were insensitive to ABA in seed germination, in addition to having an earlier flowering phenotype. Direct binding of ABI5 to the ABRE/G-box promoter elements existing in FLC was demonstrated by chromatin immunoprecipitation. Mutations at the ABRE/G-box regions in FLC promoter sequences abolished the ABI5-promoted transactivation of FLC. In summary, these results may decipher the inhibitory effect of ABA on floral transition in Arabidopsis.
ABA; ABFs; ABI5; chromatin immunoprecipitation; FLC; flowering time; SnRK2s.
Insect egg deposition activates plant defence, but very little is known about signalling events that control this response. In Arabidopsis thaliana, oviposition by Pieris brassicae triggers salicylic acid (SA) accumulation and induces the expression of defence genes. This is similar to the recognition of pathogen-associated molecular patterns (PAMPs), which are involved in PAMP-triggered immunity (PTI). Here, the involvement of known signalling components of PTI in response to oviposition was studied. Treatment with P. brassicae egg extract caused a rapid induction of early PAMP-responsive genes. In addition, expression of the defence gene PR-1 required EDS1, SID2, and, partially, NPR1, thus implicating the SA pathway downstream of egg recognition. PR-1 expression was triggered by a non-polar fraction of egg extract and by an oxidative burst modulated through the antagonistic action of EDS1 and NUDT7, but which did not depend on the NADPH oxidases RBOHD and RBOHF. Searching for receptors of egg-derived elicitors, a receptor-like kinase mutant, lecRK-I.8, was identified which shows a much reduced induction of PR-1 in response to egg extract treatment. These results demonstrate the importance of the SA pathway in response to egg-derived elicitor(s) and unravel intriguing similarities between the detection of insect eggs and PTI in Arabidopsis.
Arabidopsis thaliana; oviposition; PAMP-triggered immunity; Pieris brassicae; PR-1 expression; SA pathway.
Herbivory initiates a shift in plant metabolism from growth to defence that may reduce fitness in the absence of further herbivory. However, the defence-induced changes in carbon assimilation that precede this reallocation in resources remain largely undetermined. This study characterized the response of photosynthesis to herbivore induction of jasmonic acid (JA)-related defences in Nicotiana attenuata to increase understanding of these mechanisms. It was hypothesized that JA-induced defences would immediately reduce the component processes of photosynthesis upon attack and was predicted that wild-type plants would suffer greater reductions in photosynthesis than plants lacking JA-induced defences. Gas exchange, chlorophyll fluorescence, and thermal spatial patterns were measured together with the production of defence-related metabolites after attack and through recovery. Herbivore damage immediately reduced electron transport and gas exchange in wild-type plants, and gas exchange remained suppressed for several days after attack. The sustained reductions in gas exchange occurred concurrently with increased defence metabolites in wild-type plants, whereas plants lacking JA-induced defences suffered minimal suppression in photosynthesis and no increase in defence metabolite production. This suppression in photosynthesis occurred only after sustained defence signalling and defence chemical mobilization, whereas a short bout of feeding damage only transiently altered components of photosynthesis. It was identified that lipoxygenase signalling interacted with photosynthetic electron transport and that the resulting JA-related metabolites reduced photosynthesis. These data represent a metabolic cost to mounting a chemical defence against herbivory and link defence-signalling networks to the differential effects of herbivory on photosynthesis in remaining leaf tissues in a time-dependent manner.
chlorophyll fluorescence; defence; lipoxygenase; nicotine; plant–insect interaction
Tomato is a model and economically important crop plant with little information available about gene expression in roots. Currently, there have only been a few studies that examine hormonal responses in tomato roots and none at a genome-wide level. This study examined the transcriptome atlas of tomato root regions (root tip, lateral roots, and whole roots) and the transcriptional regulation of each root region in response to the plant hormones cytokinin and auxin using Illumina RNA sequencing. More than 165 million 1×54 base pair reads were mapped onto the Solanum lycopersicum reference genome and differential expression patterns in each root region in response to each hormone were assessed. Many novel cytokinin- and auxin-induced and -repressed genes were identified as significantly differentially expressed and the expression levels of several were confirmed by qPCR. A number of these regulated genes represent tomato orthologues of cytokinin- or auxin-regulated genes identified in other species, including CKXs, type-A RRs, Aux/IAAs, and ARFs. Additionally, the data confirm some of the hormone regulation studies for recently examined genes in tomato such as SlIAAs and SlGH3s. Moreover, genes expressed abundantly in each root region were identified which provide a spatial distribution of many classes of genes, including plant defence, secondary metabolite production, and general metabolism across the root. Overall this study presents the first global expression patterns of hormone-regulated transcripts in tomato roots, which will be functionally relevant for future studies directed towards tomato root growth and development.
Auxin; cytokinin; lateral root; RNA sequencing; root; root tip; tomato.
Changes in carbohydrate metabolism during grape berry development play a central role in shaping the final composition of the fruit. The present work aimed to identify metabolic switches during grape development and to provide insights into the timing of developmental regulation of carbohydrate metabolism. Metabolites from central carbon metabolism were measured using high-pressure anion-exchange chromatography coupled to tandem mass spectrometry and enzymatic assays during the development of grape berries from either field-grown vines or fruiting cuttings grown in the greenhouse. Principal component analysis readily discriminated the various stages of berry development, with similar trajectories for field-grown and greenhouse samples. This showed that each stage of fruit development had a characteristic metabolic profile and provided compelling evidence that the fruit-bearing cuttings are a useful model system to investigate regulation of central carbon metabolism in grape berry. The metabolites measured showed tight coordination within their respective pathways, clustering into sugars and sugar-phosphate metabolism, glycolysis, and the tricarboxylic acid cycle. In addition, there was a pronounced shift in metabolism around veraison, characterized by rapidly increasing sugar levels and decreasing organic acids. In contrast, glycolytic intermediates and sugar phosphates declined before veraison but remained fairly stable post-veraison. In summary, these detailed and comprehensive metabolite analyses revealed the timing of important switches in primary carbohydrate metabolism, which could be related to transcriptional and developmental changes within the berry to achieve an integrated understanding of grape berry development. The results are discussed in a meta-analysis comparing metabolic changes in climacteric versus non-climacteric fleshy fruits.
berry development; central carbon metabolism; fruit quality; grape ripening; metabolic coordination; metabolite profiling; Vitis vinifera.
In a non-model staple crop like wheat (Triticum aestivumI L.), functional validation of potential drought stress responsive genes identified in Arabidopsis could provide gene targets for breeding. Virus-induced gene silencing (VIGS) of genes of interest can overcome the inherent problems of polyploidy and limited transformation potential that hamper functional validation studies in wheat. In this study, three potential candidate genes shown to be involved in abiotic stress response pathways in Arabidopsis thaliana were selected for VIGS experiments in wheat. These include Era1 (enhanced response to abscisic acid), Cyp707a (ABA 8’-hydroxylase), and Sal1 (inositol polyphosphate 1-phosphatase). Gene homologues for these three genes were identified in wheat and cloned in the viral vector barley stripe mosaic virus (BSMV) in the antisense direction, followed by rub inoculation of BSMV viral RNA transcripts onto wheat plants. Quantitative real-time PCR showed that VIGS-treated wheat plants had significant reductions in target gene transcripts. When VIGS-treated plants generated for Era1 and Sal1 were subjected to limiting water conditions, they showed increased relative water content, improved water use efficiency, reduced gas exchange, and better vigour compared to water-stressed control plants inoculated with RNA from the empty viral vector (BSMV0). In comparison, the Cyp707a-silenced plants showed no improvement over BSMV0-inoculated plants under limited water condition. These results indicate that Era1 and Sal1 play important roles in conferring drought tolerance in wheat. Other traits affected by Era1 silencing were also studied. Delayed seed germination in Era1-silenced plants suggests this gene may be a useful target for developing resistance to pre-harvest sprouting.
Arabidopsis; drought; Era1; Sal1; virus-induced gene silencing; wheat.
Recent in vitro, in vivo, and theoretical experiments strongly suggest that sugar-(like) molecules counteract oxidative stress by acting as genuine reactive oxygen species (ROS) scavengers. A concept was proposed to include the vacuole as a part of the cellular antioxidant network. According to this view, sugars and sugar-like vacuolar compounds work in concert with vacuolar phenolic compounds and the ‘classic’ cytosolic antioxidant mechanisms. Among the biologically relevant ROS (H2O2, O2·–, and ·OH), hydroxyl radicals are the most reactive and dangerous species since there are no enzymatic systems known to neutralize them in any living beings. Therefore, it is important to study in more detail the radical reactions between ·OH and different biomolecules, including sugars. Here, Fenton reactions were used to compare the ·OH-scavenging capacities of a range of natural vacuolar compounds to establish relationships between antioxidant capacity and chemical structure and to unravel the mechanisms of ·OH–carbohydrate reactions. The in vitro work on the ·OH-scavenging capacity of sugars and phenolic compounds revealed a correlation between structure and ·OH-scavenging capacity. The number and position of the C=C type of linkages in phenolic compounds greatly influence antioxidant properties. Importantly, the splitting of disaccharides and oligosaccharides emerged as a predominant outcome of the ·OH–carbohydrate interaction. Moreover, non-enzymatic synthesis of new fructan oligosaccharides was found starting from 1-kestotriose. Based on these and previous findings, a working model is proposed describing the putative radical reactions involving fructans and secondary metabolites at the inner side of the tonoplast and in the vacuolar lumen.
Fructan; phenols; radicals; ROS; stress; sugars; vacuole.
Jasmonates and phytoprostanes are oxylipins that regulate stress responses and diverse physiological and developmental processes. 12-Oxo-phytodienoic acid (OPDA) and phytoprostanes are structurally related electrophilic cyclopentenones, which activate similar gene expression profiles that are for the most part different from the action of the cyclopentanone jasmonic acid (JA) and its biologically active amino acid conjugates. Whereas JA–isoleucine signals through binding to COI1, the bZIP transcription factors TGA2, TGA5, and TGA6 are involved in regulation of gene expression in response to phytoprostanes. Here root growth inhibition and target gene expression were compared after treatment with JA, OPDA, or phytoprostanes in mutants of the COI1/MYC2 pathway and in different TGA factor mutants. Inhibition of root growth by phytoprostanes was dependent on COI1 but independent of jasmonate biosynthesis. In contrast, phytoprostane-responsive gene expression was strongly dependent on TGA2, TGA5, and TGA6, but not dependent on COI1, MYC2, TGA1, and TGA4. Different mutant and overexpressing lines were used to determine individual contributions of TGA factors to cyclopentenone-responsive gene expression. Whereas OPDA-induced expression of the cytochrome P450 gene CYP81D11 was primarily regulated by TGA2 and TGA5, the glutathione S-transferase gene GST25 and the OPDA reductase gene OPR1 were regulated by TGA5 and TGA6, but less so by TGA2. These results support the model that phytoprostanes and OPDA regulate differently (i) growth responses, which are COI1 dependent but jasmonate independent; and (ii) lipid stress responses, which are strongly dependent on TGA2, TGA5, and TGA6. Identification of molecular components in cyclopentenone signalling provides an insight into novel oxylipin signal transduction pathways.
Arabidopsis thaliana; biotic and abiotic stress; class II TGA factors; detoxification; lipid signaling; reactive electrophile oxylipins.
Production of maternal haploids via a male inducer can greatly accelerate maize breeding and is an interesting biological phenomenon in double fertilization. However, the mechanism behind haploid induction remains elusive. Segregation distortion, which is increasingly recognized as a potentially powerful evolutionary force, has recently been observed during maternal haploid induction in maize. The results present here showed that both male gametophytic and zygotic selection contributed to severe segregation distortion of a locus, named segregation distortion 1 (sed1), during maternal haploid induction in maize. Interestingly, analysis of reciprocal crosses showed that sed1 is expressed in the male gametophyte. A novel mapping strategy based on segregation distortion has been used to fine-map this locus. Strong selection for the presence of the sed1 haplotype from inducers in kernels with haploid formation and defects could be detected in the segregating population. Dual-pollination experiments showed that viable pollen grains from inducers had poor pollen competitive ability against pollen from normal genotypes. Although defective kernels and haploids have different phenotypes, they are most probably caused by the sed1 locus, and possible mechanisms for production of maternal haploids and the associated segregation distortion are discussed. This research also provides new insights into the process of double fertilization.
Defective kernel; haploid; maize; male gametophyte; pollen competitive ability; segregation distortion.
Autophagy is one of the main mechanisms of degradation and remobilization of macromolecules, and it appears to play an important role in petal senescence. However, little is known about the regulatory mechanisms of autophagy in petal senescence. Autophagic processes were observed by electron microscopy and monodansylcadaverine staining of senescing petals of petunia (Petunia hybrida); autophagy-related gene 8 (ATG8) homologues were isolated from petunia and the regulation of expression was analysed. Nutrient remobilization was also examined during pollination-induced petal senescence. Active autophagic processes were observed in the mesophyll cells of senescing petunia petals. Pollination induced the expression of PhATG8 homologues and was accompanied by an increase in ethylene production. Ethylene inhibitor treatment in pollinated flowers delayed the induction of PhATG8 homologues, and ethylene treatment rapidly upregulated PhATG8 homologues in petunia petals. Dry weight and nitrogen content were decreased in the petals and increased in the ovaries after pollination in detached flowers. These results indicated that pollination induces autophagy and that ethylene is a key regulator of autophagy in petal senescence of petunia. The data also demonstrated the translocation of nutrients from the petals to the ovaries during pollination-induced petal senescence.
autophagy; ethylene; flower; nutrient remobilization; petunia; pollination; senescence.