Synthesis In Roots Research Articles

Overexpression of phosphatidylserine synthase IbPSS1 enhances salt tolerance by stimulating ethylene signaling-dependent lignin synthesis in sweetpotato roots

Phosphatidylserine (PS) is an important lipid signaling required for plant growth regulation and salt stress adaptation. However, how PS positively regulate plant salt tolerance is still largely unknown. In this study, IbPSS1-overexpressed sweetpotato plants that exhibited overproduction of PS was employed to explore the mechanisms underlying the PS stimulation of plant salt tolerance. The results revealed that the IbPSS1-overexpressed sweetpotato accumulated less Na+ in the stem and leaf tissues compared with the wild type plants. Proteomic profile of roots showed that lignin synthesis-related proteins over-accumulated in IbPSS1-overexpressed sweetpotato. Correspondingly, the lignin content was enhanced but the influx of Na + into the stele was significantly blocked in IbPSS1-overexpressed sweetpotato. The results further revealed that ethylene synthesis and signaling related genes were upregulated in IbPSS1-overexpressed sweetpotato. Ethylene imaging experiment revealed the enhancement of ethylene mainly localized in the root stele. Inhibition of ethylene synthesis completely reversed the PS-overproduction induced lignin synthesis and Na+ influx pattern in stele tissues. Taken together, our findings demonstrate a mechanism by which PS regulates ethylene signaling and lignin synthesis in the root stele, thus helping sweetpotato plants to block the loading of Na+ into the xylem and to minimize the accumulation of Na+ in the shoots.

Hydrolysis of riboflavins in root exudates under iron deficiency and alkaline stress

Riboflavins are secreted under iron deficiency as a part of the iron acquisition Strategy I, mainly when the external pH is acidic. In plants growing under Fe-deficiency and alkaline conditions, riboflavins have been reported to accumulate inside the roots, with very low or negligible secretion. However, the fact that riboflavins may undergo hydrolysis under alkaline conditions has been so far disregarded. In this paper, we report the presence of riboflavin derivatives and products of their alkaline hydrolysis (lumichrome, lumiflavin and carboxymethylflavin) in nutrient solutions of Cucumis sativus plants grown under different iron regimes (soluble Fe-EDDHA in the nutrient solution, total absence of iron in the nutrient solution, or two different doses of FeSO4 supplied as a foliar spray), either cultivated in slightly acidic (pH 6) or alkaline (pH 8.8, 10 mM bicarbonate) nutrient solutions. The results show that root synthesis and exudation of riboflavins is controlled by shoot iron status, and that exuded riboflavins undergo hydrolysis, especially at alkaline pH, with lumichrome being the main product of hydrolysis.

Phosphate starvation regulates cellulose synthesis to modify root growth.

In the model plant Arabidopsis (Arabidopsis thaliana), the absence of the essential macro-nutrient phosphate reduces primary root growth through decreased cell division and elongation, requiring alterations to the polysaccharide-rich cell wall surrounding the cells. Despite its importance, the regulation of cell wall synthesis in response to low phosphate levels is not well understood. In this study, we show that plants increase cellulose synthesis in roots under limiting phosphate conditions, which leads to changes in the thickness and structure of the cell wall. These changes contribute to the reduced growth of primary roots in low-phosphate conditions. Furthermore, we found that the cellulose synthase complex (CSC) activity at the plasma membrane increases during phosphate deficiency. Moreover, we show that this increase in the activity of the CSC is likely due to alterations in the phosphorylation status of cellulose synthases in low-phosphate conditions. Specifically, phosphorylation of CELLULOSE SYNTHASE 1 (CESA1) at the S688 site decreases in low-phosphate conditions. Phosphomimic versions of CESA1 with an S688E mutation showed significantly reduced cellulose induction and primary root length changes in low-phosphate conditions. Protein structure modeling suggests that the phosphorylation status of S688 in CESA1 could play a role in stabilizing and activating the CSC. This mechanistic understanding of root growth regulation under limiting phosphate conditions provides potential strategies for changing root responses to soil phosphate content.

Regulation of root development in nitrogen-susceptible and nitrogen-tolerant sweet potato cultivars under different nitrogen and soil moisture conditions

BackgroundDue to unreasonable nitrogen (N) application and water supply, sweet potato vines tend to grow excessively. Early development of storage roots is conducive to inhibiting vine overgrowth. Hence, we investigated how N and soil moisture affect early root growth and development.ResultsA pot experiment was conducted using the sweet potato cultivars Jishu26 (J26, N-susceptible) and Xushu32 (X32, N-tolerant). Two N application rates of 50 (N1) and 150 mg kg− 1 (N2) and two water regimes, drought stress (DS) (W1) and normal moisture (W2), were applied to each cultivar. For J26, the lowest expansion root weight was observed in the N2W2 treatment, while for X32, the N1W2 and N2W2 treatments resulted in higher root weights compared to other treatments. The interaction between N rates and water regimes significantly affected root surface area and volume in J26. Root cross-sections revealed that N2W2 increased the percentage of root area covered by xylem vessels and decreased the amount of secondary xylem vessels (SXV) in J26. However, in X32, it increased the number of SXV. A high N rate reduced the 13 C distribution ratio in J26 expansion roots, but had no significant effect on X32. In J26, N2W2 inhibited starch synthesis in roots by downregulating the expression of AGPa, AGPb, GBSS I, and SBE I.ConclusionThe observed effects were more pronounced in J26. For X32, relatively high N and moisture levels did not significantly impact storage root development. Therefore, special attention should be paid to N supply and soil moisture for N-susceptible cultivars during the early growth stage.

Maize OPR2 and LOX10 Mediate Defense against Fall Armyworm and Western Corn Rootworm by Tissue-Specific Regulation of Jasmonic Acid and Ketol Metabolism.

Foliage-feeding fall armyworm (FAW; Spodoptera frugiperda) and root-feeding western corn rootworm (WCR; Diabrotica virgifera virgifera) are maize (Zea mays L.) pests that cause significant yield losses. Jasmonic acid (JA) plays a pivotal defense role against insects. 12-oxo-phytodienoic acid (12-OPDA) is converted into JA by peroxisome-localized OPDA reductases (OPR). However, little is known about the physiological functions of cytoplasmic OPRs. Here, we show that disruption of ZmOPR2 reduced wound-induced JA production and defense against FAW while accumulating more JA catabolites. Overexpression of ZmOPR2 in Arabidopsis enhanced JA production and defense against beet armyworm (BAW; Spodoptera exigua). In addition, lox10opr2 double mutants were more susceptible than either single mutant, suggesting that ZmOPR2 and ZmLOX10 uniquely and additively contributed to defense. In contrast to the defensive roles of ZmOPR2 and ZmLOX10 in leaves, single mutants did not display any alteration in root herbivory defense against WCR. Feeding on lox10opr2 double mutants resulted in increased WCR mortality associated with greater herbivory-induced production of insecticidal death acids and ketols. Thus, ZmOPR2 and ZmLOX10 cooperatively inhibit the synthesis of these metabolites during herbivory by WCR. We conclude that ZmOPR2 and ZmLOX10 regulate JA-mediated resistance in leaves against FAW while suppressing insecticidal oxylipin synthesis in roots during WCR infestation.

CRISPR/Cas9 disruption of MYB134 and MYB115 in transgenic poplar leads to differential reduction of proanthocyanidin synthesis in roots and leaves.

Proanthocyanidins (PAs) are common specialized metabolites and particularly abundant in trees and woody plants. In poplar (Populus spp.), PA biosynthesis is stress-induced and regulated by two previously studied transcription factors MYB115 and MYB134. To determine the relative contribution of these regulators to PA biosynthesis, we created single and double knockout mutants for both genes in transgenic poplars using CRISPR/Cas9. Knocking out either MYB134 or MYB115 showed reduced PA accumulation and down-regulated flavonoid genes in leaves, but MYB134 disruption had the greatest impact and reduced PAs to 30% of controls. In roots, by contrast, only the MYB134/MYB115 double knockouts showed a significant change in PA concentration. The loss of PAs paralleled the lower expression of PA biosynthesis genes, and concentrations of flavan-3-ol PA precursors catechin and epicatechin. Interestingly, salicinoids were also affected in double knock-outs, with distinct patterns in roots and shoots. We conclude that the regulatory pathways for PA biosynthesis differs in poplar leaves and roots. The residual PA content in the double-KO plants indicates that other transcription factors must also be involved in control of the PA pathway.

Potassium deficiency causes more nitrate nitrogen to be stored in leaves for low-K sensitive sweet potato genotypes.

In order to explore the effect of potassium (K) deficiency on nitrogen (N) metabolism in sweet potato (Ipomoea batatas L.), a hydroponic experiment was conducted with two genotypes (Xushu 32, low-K-tolerant; Ningzishu 1, low-K-sensitive) under two K treatments (-K, <0.03 mM of K+; +K, 5 mM of K+) in the greenhouse of Jiangsu Normal University. The results showed that K deficiency decreased root, stem, and leaf biomass by 13%-58% and reduced whole plant biomass by 24%-35%. Compared to +K, the amount of K and K accumulation in sweet potato leaves and roots was significantly decreased by increasing root K+ efflux in K-deficiency-treated plants. In addition, leaf K, N, ammonium nitrogen (NH4 +-N), or nitrate nitrogen (NO3 --N) in leaves and roots significantly reduced under K deficiency, and leaf K content had a significant quadratic relationship with soluble protein, NO3 --N, or NH4 +-N in leaves and roots. Under K deficiency, higher glutamate synthase (GOGAT) activity did not increase amino acid synthesis in roots; however, the range of variation in leaves was larger than that in roots with increased amino acid in roots, indicating that the transformation of amino acids into proteins in roots and the amino acid export from roots to leaves were not inhibited. K deficiency decreased the activity of nitrate reductase (NR) and nitrite reductase (NiR), even if the transcription level of NR and NiR increased, decreased, or remained unchanged. The NO3 -/NH4 + ratio in leaves and roots under K deficiency decreased, except in Ningzishu 1 leaves. These results indicated that for Ningzishu 1, more NO3 - was stored under K deficiency in leaves, and the NR and NiR determined the response to K deficiency in leaves. Therefore, the resistance of NR and NiR activities to K deficiency may be a dominant factor that ameliorates the growth between Xushu 32 and Ningzishu 1 with different low-K sensitivities.

Tomato growth promotion by the fungal endophytes Serendipita indica and Serendipita herbamans is associated with sucrose de-novo synthesis in roots and differential local and systemic effects on carbohydrate metabolisms and gene expression

Plant growth-promoting and stress resilience-inducing root endophytic fungi represent an additional carbohydrate sink. This study aims to test if such root endophytes affect the sugar metabolism of the host plant to divert the flow of resources for their purposes. Fresh and dry weights of roots and shoots of tomato (Solanum lycopersicum) colonised by the closely related Serendipita indica and Serendipita herbamans were recorded. Plant carbohydrate metabolism was analysed by measuring sugar levels, by determining activity signatures of key enzymes of carbohydrate metabolism, and by quantifying mRNA levels of genes involved in sugar transport and turnover. During the interaction with the tomato plants, both fungi promoted root growth and shifted shoot biomass from stem to leaf tissues, resulting in increased leaf size. A common effect induced by both fungi was the inhibition of phosphofructokinase (PFK) in roots and leaves. This glycolytic-pacing enzyme shows how the glycolysis rate is reduced in plants and, eventually, how sugars are allocated to different tissues. Sucrose phosphate synthase (SPS) activity was strongly induced in colonised roots. This was accompanied by increased SPS-A1 gene expression in S. herbamans-colonised roots and by increased sucrose amounts in roots colonised by S. indica. Other enzyme activities were barely affected by S. indica, but mainly induced in leaves of S. herbamans-colonised plants and decreased in roots. This study suggests that two closely related root endophytic fungi differentially influence plant carbohydrate metabolism locally and systemically, but both induce a similar increase in plant biomass. Notably, both fungal endophytes induce an increase in SPS activity and, in the case of S. indica, sucrose resynthesis in roots. In leaves of S. indica-colonised plants, SWEET11b expression was enhanced, thus we assume that excess sucrose was exported by this transporter to the roots. ‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬

Cyanogenesis in cassava and its molecular manipulation for crop improvement.

While cassava is one of the most important staple crops worldwide, it has received the least investment per capita consumption of any of the major global crops. This is in part due to cassava being a crop of subsistence farmers that is grown in countries with limited resources for crop improvement. While its starchy roots are rich in calories, they are poor in protein and other essential nutrients. In addition, they contain potentially toxic levels of cyanogenic glycosides which must be reduced to safe levels before consumption. Furthermore, cyanogens compromise the shelf life of harvested roots due to cyanide-induced inhibition of mitochondrial respiration, and associated production of reactive oxygen species that accelerate root deterioration. Over the past two decades, the genetic, biochemical, and developmental factors that control cyanogen synthesis, transport, storage, and turnover have largely been elucidated. It is now apparent that cyanogens contribute substantially to whole-plant nitrogen metabolism and protein synthesis in roots. The essential role of cyanogens in root nitrogen metabolism, however, has confounded efforts to create acyanogenic varieties. This review proposes alternative molecular approaches that integrate accelerated cyanogen turnover with nitrogen reassimilation into root protein that may offer a solution to creating a safer, more nutritious cassava crop.

Seed Yield and Nitrogen Efficiency in Oilseed Rape After Ammonium Nitrate or Urea Fertilization.

In agricultural plant production, nitrate, ammonium, and urea are the major fertilized nitrogen forms, which differ in root uptake and downstream signaling processes in plants. Nitrate is known to stimulate cytokinin synthesis in roots, while for urea no hormonal effect has been described yet. Elevated cytokinin levels can delay plant senescence favoring prolonged nitrogen uptake. As the cultivation of winter oilseed rape provokes high nitrogen-balance surpluses, we tested the hypotheses whether nitrogen use efficiency increases under ammonium nitrate- relative to urea-based nutrition and whether this is subject to genotypic variation. In a 2-year field study, 15 oilseed rape lines were fertilized either with ammonium nitrate or with urease inhibitor-stabilized urea and analyzed for seed yield and nitrogen-related yield parameters. Despite a significant environmental impact on the performance of the individual lines, which did not allow revealing consistent impact of the genotype, ammonium nitrate-based nutrition tended to increase seed yield in average over all lines. To resolve whether the fertilizer N forms act on grain yield via phytohormones, we collected xylem exudates at three developmental stages and determined the translocation rates of cytokinins and N forms. Relative to urea, ammonium nitrate-based nutrition enhanced the translocation of nitrate or total nitrogen together with cytokinins, whereas in the urea treatment translocation rates were lower as long as urea remained stable in the soil solution. At later developmental stages, i.e., when urea became hydrolyzed, nitrogen and cytokinin translocation increased. In consequence, urea tended to increase nitrogen partitioning in the shoot toward generative organs. However, differences in overall nitrogen accumulation in shoots were not present at the end of the vegetation period, and neither nitrogen uptake nor utilization efficiency was consistently different between the two applied nitrogen forms.

Cocultivating rye with berseem clover affects benzoxazinoid production and expression of related genes

AbstractBenzoxazinoids (BXs) are secondary metabolites synthesized mainly by gramineous plants, including rye (Secale cereale L.), that play an important role in stress resistance and allelopathy. In the present work, the influence of cocultivation of rye with berseem clover (Trifolium alexandrinum L.) on the expression of BX synthesis‐regulating genes (ScBx1–ScBx5, ScIgl, and ScGT) and the content of six BXs (HBOA, DIBOA, GDIBOA, DIMBOA, GDIMBOA, and MBOA) in roots and the aerial portion of three rye inbred lines (ILs) (L318, D33, and D39) was investigated. Cocultivation of rye with berseem clover influenced its gene expression levels and BX contents. The response was strongly affected by rye genotype, plant part, time point, gene, and metabolite. The most frequently observed changes of gene expression concerned IL D33, aerial plant parts, the second time point (4 wk after germination), and ScBx3. For BX synthesis, the most frequently observed changes for IL D33 were in roots, the third time point (6 wk after germination), GDIBOA, and DIBOA. In 18.3% of cases, gene expression was correlated with BX synthesis. The coregulation of gene expression and BX synthesis in roots and aerial parts of rye plants affected by clover was observed relatively rarely. Cocultivation of rye with clover led, after 6 wk, to BX increases in roots of all tested ILs. Despite the lack of clearly universal response, the cocultivation of rye with clover may strengthen rye defense capability, at least against soil pathogens.

Вплив системи догляду за кроною на декоративність і довговічність рослин сортів Syringa vulgaris L.

Досліджено вплив способів обрізування крони рослин Syringa vulgaris L. на їхню декоративність і довговічність. Встановлено, що досягти найкращого результату можна тільки за регулярного догляду за кроною. Спонукає до цього, щонайперше, дихотомічне (несправжнє) гілкування, за яким кожна однорічна гілка продукує навесні два пагони з верхівкової пари бруньок та ще кілька – з бруньок, що розташовані нижче по осі цієї гілки. Отже, щороку загальна їхня кількість щонайменше потроюється. Коренева система не зможе за цей час збільшити в таких обсягах свою масу і фізіологічну активність, що корелятивно спричиняє значне зменшення розмірів пагонів і суцвіть. Якщо догляд був нерегулярний або відсутній, виникає потреба інтенсивного прорідження або кардинального омолодження крони. Доведено, що цими заходами можна поліпшити декоративність рослин, але на довговічність вони вплинуть негативно. Зумовлює це дві причини. Перша – запобігання ран внаслідок видалення гілок на кільце. Встановлено, що навіть 1,5-2,0-сантиметрові зрізи на стовбурі чи скелетних гілках ніколи не загояться, а за 10-15 років їхня деревина струхліє, утворивши порожнину. Деревина великих ран руйнується швидше, що призводить до виникнення дупел та повільного відмирання всієї рослини. Другою причиною вкорочення довговічності є раптово порушений функціональний корелятивний коренево-листковий зв'язок, що є наслідком різкого зменшення в омолодженій рослині надземної частини. Спочатку це призведе до сповільнення фотосинтезу (крона майже відсутня), а потім, корелятивно, й до затухання кореневого синтезу: через мізерне надходження асимілятів до коренів. Унаслідок цього загальна маса фізіологічно активної частини кореневої системи теж значно зменшиться, що негативно вплине на довговічність рослини загалом. Декоративність кущів через це з року в рік погіршується. Також встановлено, що до обрізування крони змушує й загущена посадка бузків. Для відновлення її декоративності спочатку варто видалити кволі рослини, а в решти – знизити крону приблизно на третину аби наблизити її до кореневої системи, що забезпечить рослинам фізичну витривалість та дещо внормує в них функціональний коренево-листковий зв'язок. Здійснити цей захід треба якомога раніше в часовому вимірі, бо, окрім згаданого, рослини почнуть страждати й від хвороб. Потужні гілки потрібно вкорочувати, а не вирізати на кільце, аби не нанести стовбурам і гілкам першого порядку великих ран. За такого підходу до обрізування крони бузки будуть декоративними й здебільшого випадків – довговічними.

Effects of UV-A radiation on organ-specific accumulation and gene expression of isoflavones and flavonols in soybean sprout

Organ-specific flavonoid destination in soybean sprouts following UV irradiation is still unclear although the metabolic pathway of flavonoid synthesis and UV responded flavonoid accumulation have been well investigated. We report the identification of organ-specific localization and specific gene expression of isoflavones and kaempferol glycosides in the soybean sprouts responded to UV-A irradiation. UV-A irradiation stimulated only root isoflavones, especially increase of genistein types. The daidzein types predominated in non-UV-A treated roots. Kaempferol glycosides were not increased in roots by UV-A, but distinctly increased in aerial organs, especially in the cotyledons. These results demonstrate that UV-A upregulates the naringenin pathway synthesizing genistin and kaempferol rather than the liquiritigenin pathway synthesizing daidzin and glycitin. High GmUGT9 and other gene expression related to isoflavone synthesis in roots clearly demonstrate the UV-A-induced isoflavone accumulation. Aerial organ specific increase of GmF3H, GmFLS1, and GmDFR1 expression by UV-A distinctly demonstrates the flavonol increase in aerial organs.

Decreased expression of fructosyltransferase genes in asparagus roots may contribute to efficient fructan degradation during asparagus spear harvesting

Asparagus (Asparagus officinalis L.) accumulates inulin and inulin neoseries-type fructans in root, which are synthesized by three fructosyltransferases—sucrose:sucrose 1-fructosyltransferase (1-SST, EC 2.4.1.99), fructan:fructan 1-fructosyltransferase (1-FFT, EC 2.4.1.100), and fructan:fructan 6G-fructosyltransferase (6G-FFT, EC 2.4.1.243). Fructans in roots are considered as energy sources for emerging of spears, and it has been demonstrated that a gradual decrease in root fructan content occurs during the spear harvesting season (budding and shooting up period). However, the roles of certain three fructosyltransferases during the harvest season have not yet been elucidated. Here, we investigated the variation in enzymatic activities and gene expression levels of three fructosyltransferases and examined sugar contents in roots before and during the spear harvest period. Two cDNAs, aoft2 and aoft3, were isolated from the cDNA library of roots. The respective recombinant proteins (rAoFT2 and rAoFT3), produced by Pichia pastoris, were characterized: rAoFT2 showed 1-FFT activity (producing nystose from 1-kestose), whereas rAoFT3 showed 1-SST activity (producing 1-kestose from sucrose). These reaction profiles of recombinant proteins were similar to those of native enzymes purified previously. These results indicate that aoft2 and aoft3 encoding 1-FFT and 1-SST are involved in fructan synthesis in roots. A gradual downregulation of fructosyltransferase genes and activity of respective enzymes was observed in roots during the harvest period, which also coincided with the decrease in fructooligosaccharides and increase in fructose due to fructan exohydrolase activity. These findings suggest that downregulation of fructosyltransferases genes during harvest time may contribute to efficient degradation of fructan required for the emergence of spears.

Effects of root abscisic acid on Na+ transport and photosystem 2 in Helianthus tuberosus under salt stress

The effects of root abscisic acid (ABA) signal on Na+ transport and photosystem 2 (PS2) in Jerusalem artichoke (Helianthus tuberosus) under salt stress (150 mmol·L-1 NaCl) were examined by applying ABA synthesis inhibitor sodium tungstate to roots. Sodium tungstate inhibited ABA synthesis in roots, reduced root Na+ efflux, and increased the efficiency of Na+ transport from roots to leaves under salt stress. Salt stress increased leaf Na+ content and did not affect leaf membrane lipid peroxidation, PS2 reaction center protein and PS2 maximum photochemical efficiency (Fv/Fm ). The inhibition on root ABA synthesis significantly increased leaf Na+ accumulation, aggravated leaf membrane lipid peroxidation, impaired PS2 reaction center protein, decreased Fv/Fm, and induced PS2 photoinhibition. In conclusion, root ABA signal was beneficial to reducing leaf Na+ accumulation and preventing PS2 oxidative damage by inducing root Na+ efflux and inhibiting Na+ transport to the aerial part in H. tuberosus under salt stress.

Providing carbon skeletons to sustain amide synthesis in roots underlines the suitability of Brachypodium distachyon for the study of ammonium stress in cereals.

Plants mainly acquire N from the soil in the form of nitrate (NO3−) or ammonium (NH4+). Ammonium-based nutrition is gaining interest because it helps to avoid the environmental pollution associated with nitrate fertilization. However, in general, plants prefer NO3− and indeed, when growing only with NH4+ they can encounter so-called ammonium stress. Since Brachypodium distachyon is a useful model species for the study of monocot physiology and genetics, we chose it to characterize performance under ammonium nutrition. Brachypodium distachyon Bd21 plants were grown hydroponically in 1 or 2.5 mM NO3− or NH4+. Nitrogen and carbon metabolism associated with NH4+ assimilation was evaluated in terms of tissue contents of NO3−, NH4+, K, Mg, Ca, amino acids and organic acids together with tricarboxylic acid (TCA) cycle and NH4+-assimilating enzyme activities and RNA transcript levels. The roots behaved as a physiological barrier preventing NH4+ translocation to aerial parts, as indicated by a sizeable accumulation of NH4+, Asn and Gln in the roots. A continuing high NH4+ assimilation rate was made possible by a tuning of the TCA cycle and its associated anaplerotic pathways to match 2-oxoglutarate and oxaloacetate demand for Gln and Asn synthesis. These results show B. distachyon to be a highly suitable tool for the study of the physiological, molecular and genetic basis of ammonium nutrition in cereals.

Development of an efficient root transgenic system for pigeon pea and its application to other important economically plants.

SummaryFor non‐model plants, functional characterization of genes is still hampered by lack of efficient stable transformation procedures. Here, we report a simple, fast and efficient transformation technique with Agrobacterium rhizogenes for generating stable transgenic roots in living plants to facilitate functional studies in vivo. We showed that injection of A. rhizogenes into stems of various plant species lead to stable transgenic root generation, which can sustain plant growth after the original, non‐transgenic roots were cut off. A transformation system was established for pigeon pea, a major woody food crop, after optimizing the selection of A. rhizogenes strains, bacterium concentration, injection position and seedling age. RT‐PCR and fluorescence observation indicated a transgenic root induction efficiency of about 39% in pigeon pea. Furthermore, induction of hairy roots was achieved in nine out of twelve tested economically important plants at an efficiency of 15–39%. As proof of concept, bimolecular fluorescence complementation (BiFC) assay was applied to test the interaction between CcCIPK14 and CcCBL1/2 in pigeon pea. Additionally, ectopic expression of the bZIP transcription factor MdHY5 from apple confirmed the utility of the transformation technique for engineering anthocyanin synthesis in roots. Taken together, we show that this method allows fast in vivo studies of gene function in a wide range of plant species.

Nitric oxide synthase-mediated early nitric oxide burst alleviates water stress-induced oxidative damage in ammonium-supplied rice roots

BackgroundNutrition with ammonium (NH4+) can enhance the drought tolerance of rice seedlings in comparison to nutrition with nitrate (NO3−). However, there are still no detailed studies investigating the response of nitric oxide (NO) to the different nitrogen nutrition and water regimes. To study the intrinsic mechanism underpinning this relationship, the time-dependent production of NO and its protective role in the antioxidant defense system of NH4+- or NO3−-supplied rice seedlings were studied under water stress.ResultsAn early NO burst was induced by 3 h of water stress in the roots of seedlings subjected to NH4+ treatment, but this phenomenon was not observed under NO3− treatment. Root oxidative damage induced by water stress was significantly higher for treatment with NO3− than with NH4+ due to reactive oxygen species (ROS) accumulation in the former. Inducing NO production by applying the NO donor 3 h after NO3− treatment alleviated the oxidative damage, while inhibiting the early NO burst by applying the NO scavenger 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (c-PTIO) increased root oxidative damage in NH4+ treatment. Application of the nitric oxide synthase (NOS) inhibitor N(G)-nitro-L-arginine methyl ester(L-NAME) completely suppressed NO synthesis in roots 3 h after NH4+ treatment and aggravated water stress-induced oxidative damage. Therefore, the aggravation of oxidative damage by L-NAME might have resulted from changes in the NOS-mediated early NO burst. Water stress also increased the activity of root antioxidant enzymes (catalase, superoxide dismutase, and ascorbate peroxidase). These were further induced by the NO donor but repressed by the NO scavenger and NOS inhibitor in NH4+-treated roots.ConclusionThese findings demonstrate that the NOS-mediated early NO burst plays an important role in alleviating oxidative damage induced by water stress by enhancing the antioxidant defenses in roots supplemented with NH4+.

Species-Wide Variation in Shoot Nitrate Concentration, and Genetic Loci Controlling Nitrate, Phosphorus and Potassium Accumulation in Brassica napus L.

Large nitrogen, phosphorus and potassium fertilizer inputs are used in many crop systems. Identifying genetic loci controlling nutrient accumulation may be useful in crop breeding strategies to increase fertilizer use efficiency and reduce financial and environmental costs. Here, variation in leaf nitrate concentration across a diversity population of 383 genotypes of Brassica napus was characterized. Genetic loci controlling variation in leaf nitrate, phosphorus and potassium concentration were then identified through Associative Transcriptomics using single nucleotide polymorphism (SNP) markers and gene expression markers (GEMs). Leaf nitrate concentration varied over 8-fold across the diversity population. A total of 455 SNP markers were associated with leaf nitrate concentration after false-discovery-rate (FDR) correction. In linkage disequilibrium of highly associated markers are a number of known nitrate transporters and sensors, including a gene thought to mediate expression of the major nitrate transporter NRT1.1. Several genes influencing root and root-hair development co-localize with chromosomal regions associated with leaf P concentration. Orthologs of three ABC-transporters involved in suberin synthesis in roots also co-localize with association peaks for both leaf nitrate and phosphorus. Allelic variation at nearby, highly associated SNPs confers large variation in leaf nitrate and phosphorus concentration. A total of five GEMs associated with leaf K concentration after FDR correction including a GEM that corresponds to an auxin-response family protein. Candidate loci, genes and favorable alleles identified here may prove useful in marker-assisted selection strategies to improve fertilizer use efficiency in B. napus.

Functional characterization of soybean strigolactone biosynthesis and signaling genes in Arabidopsis MAX mutants and GmMAX3 in soybean nodulation

BackgroundStrigolactones (SLs) play important roles in controlling root growth, shoot branching, and plant-symbionts interaction. Despite the importance, the components of SL biosynthesis and signaling have not been unequivocally explored in soybean.ResultsHere we identified the putative components of SL synthetic enzymes and signaling proteins in soybean genome. Soybean genome contains conserved MORE AXILLARY BRANCHING (MAX) orthologs, GmMAX1s, GmMAX2s, GmMAX3s, and GmMAX4s. The tissue expression patterns are coincident with SL synthesis in roots and signaling in other tissues under normal conditions. GmMAX1a, GmMAX2a, GmMAX3b, and GmMAX4a expression in their Arabidopsis orthologs’ mutants not only restored most characteristic phenotypes, such as shoot branching and shoot height, leaf shape, primary root length, and root hair growth, but also restored the significantly changed hormone contents, such as reduced JA and ABA contents in all mutant leaves, but increased auxin levels in atmax1, atmax3 and atmax4 mutants. Overexpression of these GmMAXs also altered the hormone contents in wild-type Arabidopsis. GmMAX3b was further characterized in soybean nodulation with overexpression and knockdown transgenic hairy roots. GmMAX3b overexpression (GmMAX3b-OE) lines exhibited increased nodule number while GmMAX3b knockdown (GmMAX3b-KD) decreased the nodule number in transgenic hairy roots. The expression levels of several key nodulation genes were also altered in GmMAX3b transgenic hairy roots. GmMAX3b overexpression hairy roots had reduced ABA, but increased JA levels, with no significantly changed auxin content, while the contrast changes were observed in GmMAX3b-KD lines. Global gene expression in GmMAX3b-OE or GmMAX3b-KD hairy roots also revealed that altered expression of GmMAX3b in soybean hairy roots changed several subsets of genes involved in hormone biosynthesis and signaling and transcriptional regulation of nodulation processes.ConclusionsThis study not only revealed the conservation of SL biosynthesis and signaling in soybean, but also showed possible interactions between SL and other hormone synthesis and signaling during controlling plant development and soybean nodulation. GmMAX3b-mediated SL biosynthesis and signaling may be involved in soybean nodulation by affecting both root hair formation and its interaction with rhizobia.