Ureide Transport Research Articles

The selenium-promoted daidzein production contributes to its induced nodulation in soybean plants

Selenium (Se) is an essential micronutrient to humans. Se biofortification in crops provides an effective means to enhance dietary Se intake. However, the effects of Se treatment on plant growth and development remain to be fully studied. In this study, we applied Se to soybean plants grown hydroponically and in soil. We observed that selenate treatment increased nodule number and fresh weight. Furthermore, Se supplementation increased isoflavone daidzein accumulation in roots and nodules. An exogenous supply of daidzein but not kaempferol was found to increase nodule number and fresh weight significantly, suggesting a role of daidzein in the Se-induced nodulation in soybean plants. In addition, the total sugar levels in nodules, roots, and leaves were significantly enhanced following selenate treatment. Subsequently, the N-compounds were greatly increased after Se treatment in all tissues examined. Transcriptome analysis revealed that Se treatment altered various metabolic and cellular processes to affect nodulation and nodule development. Se treatment upregulated GmNIN2b, an important nodule organogenesis gene, as well as genes associated with sugar metabolism and ureide transport. Our study provides insights underlying the Se-induced nodulation in soybean plants and further documents that Se biofortification in soybean can simultaneously improve not only the essential nutrient content but also nodulation for biological nitrogen fixation, a critically important agronomic trait in soybean plants.

Multi-scale analysis provides insights into the roles of ureide permeases in wheat nitrogen use efficiency.

The ureides allantoin and allantoate serve as nitrogen (N) transport compounds in plants, and more recently, allantoin has been shown to play a role in signaling. In planta, tissue ureide levels are controlled by the activity of enzymes of the purine degradation pathway and by ureide transporters called ureide permeases (UPS). Little is known about the physiological function of UPS proteins in crop plants, and especially in monocotyledon species. Here, we identified 13 TaUPS genes in the wheat (Triticum aestivum L.) genome. Phylogenetic and genome location analyses revealed a close relationship of wheat UPSs to orthologues in other grasses and a division into TaUPS1, TaUPS2.1, and TaUPS2.2 groups, each consisting of three homeologs, with a total of four tandem duplications. Expression, localization, and biochemical analyses resolved spatio-temporal expression patterns of TaUPS genes, transporter localization at the plasma membrane, and a role for TaUPS2.1 proteins in cellular import of ureides and phloem and seed loading. In addition, positive correlations between TaUPS1 and TaUPS2.1 transcripts and ureide levels were found. Together the data support that TaUPSs function in regulating ureide pools at source and sink, along with source-to-sink transport. Moreover, comparative studies between wheat cultivars grown at low and high N strengthened a role for TaUPS1 and TaUPS2.1 transporters in efficient N use and in controlling primary metabolism. Co-expression, protein-protein interaction, and haplotype analyses further support TaUPS involvement in N partitioning, N use efficiency, and domestication. Overall, this work provides a new understanding on UPS transporters in grasses as well as insights for breeding resilient wheat varieties with improved N use efficiency.

Altering ureide transport in nodulated soybean results in whole-plant adjustments of metabolism, assimilate partitioning, and sink strength

Legumes develop a symbiotic relationship with bacteria that are housed in root nodules and fix atmospheric di-nitrogen (N2) to ammonia. In soybean (Glycine max (L.) Merr.) nodules, the final products of nitrogen (N) fixation are amino acids, and the ureides allantoin and allantoic acid that also serve as the major long-distance N transport forms. Recently, we have shown that increased expression of UPS1 (ureide permease 1) in soybean nodules results in enhanced ureide export from nodules with positive effects on N fixation and seed yield. Here, we demonstrate that changes in the ureide transport processes trigger alterations in allantoin and allantoic acid pools and partitioning throughout the transgenic plants. They further result in adjustments in amino acid availability in, and translocation to, root and shoot sinks. In addition, leaf carbon (C) capture, assimilation and allocation to sinks are improved, accommodating the increased nodule function, and root and shoot growth. Overall, we demonstrate that enhanced ureide partitioning in nodulated soybean leads to a complex rebalancing of N and C acquisition, metabolism, and transport processes with positive consequences for above- and below-ground vegetative biomass, and whole-plant N and C gains.

Role of ureides in source-to-sink transport of photoassimilates in non-fixing soybean.

Nitrogen (N)-fixing soybean plants use the ureides allantoin and allantoic acid as major long-distance transport forms of N, but in non-fixing, non-nodulated plants amino acids mainly serve in source-to-sink N allocation. However, some ureides are still synthesized in roots of non-fixing soybean, and our study addresses the role of ureide transport processes in those plants. In previous work, legume ureide permeases (UPSs) were identified that are involved in cellular import of allantoin and allantoic acid. Here, UPS1 from common bean was expressed in the soybean phloem, which resulted in enhanced source-to-sink transport of ureides in the transgenic plants. This was accompanied by increased ureide synthesis and elevated allantoin and allantoic acid root-to-sink transport. Interestingly, amino acid assimilation, xylem transport, and phloem partitioning to sinks were also strongly up-regulated. In addition, photosynthesis and sucrose phloem transport were improved in the transgenic plants. These combined changes in source physiology and assimilate partitioning resulted in increased vegetative growth and improved seed numbers. Overall, the results support that ureide transport processes in non-fixing plants affect source N and carbon acquisition and assimilation as well as source-to-sink translocation of N and carbon assimilates with consequences for plant growth and seed development.

Disruption of ureide degradation affects plant growth and development during and after transition from vegetative to reproductive stages

BackgroundThe ureides allantoin and allantoate are major metabolic intermediates of purine catabolism with high nitrogen-to-carbon ratios. Ureides play a key role in nitrogen utilization in ureide-type legumes, but their effects on growth and development in non-legume plants are poorly understood. Here, we examined the effects of knocking out genes encoding ureide-degrading enzymes, allantoinase (ALN) and allantoate amidohydrolase (AAH), on the vegetative-to-reproductive transition and subsequent growth of Arabidopsis plants.ResultsThe ureide-degradation mutants (aln and aah) showed symptoms similar to those of nitrogen deficiency: early flowering, reduced size at maturity, and decreased fertility. Consistent with these phenotypes, carbon-to-nitrogen ratios and nitrogen-use efficiencies were significantly decreased in ureide-degradation mutants; however, adding nitrogen to irrigation water did not alleviate the reduced growth of these mutants. In addition to nitrogen status, levels of indole-3-acetic acid and gibberellin in five-week-old plants were also affected by the aln mutations. To test the possibility that ureides are remobilized from source to sink organs, we measured ureide levels in various organs. In wild-type plants, allantoate accumulated predominantly in inflorescence stems and siliques; this accumulation was augmented by disruption of its catabolism. Mutants lacking ureide transporters, ureide permeases 1 and 2 (UPS1 and UPS2), exhibited phenotypes similar to those of the ureide-degradation mutants, but had decreased allantoate levels in the reproductive organs. Transcript analysis in wild-type plants suggested that genes involved in allantoate synthesis and ureide transport were coordinately upregulated in senescing leaves.ConclusionsThis study demonstrates that ureide degradation plays an important role in supporting healthy growth and development in non-legume Arabidopsis during and after transition from vegetative to reproductive stages.

Transporters involved in source to sink partitioning of amino acids and ureides: opportunities for crop improvement

In most plant species, amino acids are the predominant chemical forms in which nitrogen is transported. However, in nodulated tropical or subtropical legumes, ureides are the main nitrogen transport compounds. This review describes the partitioning of amino acids and ureides within the plant, and follows their movement from the location of synthesis (source) to the sites of usage (sink). Xylem and phloem connect source and sink organs and serve as routes for long-distance transport of the organic nitrogen. Loading and unloading of these transport pathways might require movement of amino acids and ureides across cell membranes, a task that is mediated by membrane proteins (i.e. transporters) functioning as export or import systems. The current knowledge on amino acid and ureide transporters involved in long-distance transport of nitrogen is provided and their importance for source and sink physiology discussed. The review concludes by exploring possibilities for genetic manipulation of organic nitrogen transporter activities to confer increases in crop productivity.

Soybean ureide transporters play a critical role in nodule development, function and nitrogen export

Legumes can access atmospheric nitrogen through a symbiotic relationship with nitrogen-fixing bacteroids that reside in root nodules. In soybean, the products of fixation are the ureides allantoin and allantoic acid, which are also the dominant long-distance transport forms of nitrogen from nodules to the shoot. Movement of nitrogen assimilates out of the nodules occurs via the nodule vasculature; however, the molecular mechanisms for ureide export and the importance of nitrogen transport processes for nodule physiology have not been resolved. Here, we demonstrate the function of two soybean proteins - GmUPS1-1 (XP_003516366) and GmUPS1-2 (XP_003518768) - in allantoin and allantoic acid transport out of the nodule. Localization studies revealed the presence of both transporters in the plasma membrane, and expression in nodule cortex cells and vascular endodermis. Functional analysis in soybean showed that repression of GmUPS1-1 and GmUPS1-2 in nodules leads to an accumulation of ureides and decreased nitrogen partitioning to roots and shoot. It was further demonstrated that nodule development, nitrogen fixation and nodule metabolism were negatively affected in RNAi UPS1 plants. Together, we conclude that export of ureides from nodules is mediated by UPS1 proteins, and that activity of the transporters is not only essential for shoot nitrogen supply but also for nodule development and function.

Amino acid and ureide transport in the xylem of symbiotic soybean plants during short-term flooding of the root system in the presence of different sources of nitrogen

The transport of organic N compounds to the shoot in the xylem sap of nodulated soybean plants was investigated in an attempt to better understand the changes in N metabolism under root hypoxia (first 5 days of flooding), with different sources of N in the medium. NO3- is beneficial for tolerance of plants to waterlogging, whereas other N sources such as NH4+ and NH4NO3, are not. Nevertheless, in the presence of NH4+ high levels of amino acids were transported in the xylem, consistent with its assimilation. Some increase in the transport of amino acids was also seen with NO3- nutrition during waterlogging, but not with N-free medium. Ureide transport in the xylem was severely reduced during waterlogging, consistent with impaired N2 fixation under these conditions. The relative proportions of some amino acids in the xylem showed dramatic changes during treatment. Alanine increased tremendously under root hypoxia, especially with NH4+ as N source, where it reached near 70 % of the total amino acids present. Aspartic acid, on the other hand, dropped to very low levels and was inversely related to alanine levels, consistent with this amino acid being the immediate source of N for alanine synthesis. Glutamine levels also fell to a larger or lesser extent, depending on the N source present. The changes in asparagine, one of the prominent amino acids of the xylem sap, were most outstanding in the treatment with NO3-, where they showed a large increase, characteristic of plants switching from dependence on N2 fixation to NO3- assimilation. The data indicate that the lesser effectiveness of NH4+ during waterlogging, in contrast to NO3-, involves restricted amino acids metabolism, and may result from energy metabolism being directed towards NH4+ detoxification.

Heterogeneity of sucrose synthase genes in bean (Phaseolus vulgaris L.): evidence for a nodule-enhanced sucrose synthase gene.

Sucrose synthase (SS), the key sucrose hydrolytic enzyme (EC 2.4.1.13), plays an important role in N(2)-fixing nodule metabolism. It has also been proposed that N(2) fixation in soybean nodules could be mediated by the potential to metabolize sucrose. The isolation and characterization of a nodule-enhanced SS full-length cDNA clone from the bean Phaseolus vulgaris is reported here. Southern blot analysis indicated that there are at least two SS genes in beans. Using a 3' specific probe from this SS cDNA clone, it was possible to identify a nodule-enhanced SS gene (PvSSn), which is expressed almost exclusively in nodules. A second gene (PvSS), which is expressed in all tissues tested, was detected using a coding region probe. Nodule-enhanced PvSSn transcript levels, but not the enzyme activity or protein amount, is reduced during nodule development. These data indicated that this reduction could be due to a limitation on the carbon availability in the nodule. PvSSn expression is reduced in the asparagine-treated nodules. By contrast, PvSSn transcript levels in nodules increased in the presence of glutamine, allantoin and allopurinol. This result suggests a relationship between ureide transport and SS regulation and could help in understanding why the ureide transport mechanism is activated during nitrogen fixation in bean.

Rhizobium etli mutant modulates carbon and nitrogen metabolism in Phaseolus vulgaris nodules.

The aim of this study was to evaluate the biochemical events in root nodules which lead to increased yield when bean is inoculated with a Rhizobium etli mutant (CFN037) having increased respiratory capacity. CFN037-inoculated plants had 22% more nitrogen (N) than did wild-type (CE3)-inoculated plants. Root nodule enzymes involved in nodule carbon and nitrogen assimilation as well as in ureides and amides synthesis were assessed in plants inoculated with CFN037 and the CE3. Our results show that the xylem ureides content was lower while that of amino acids was higher in CFN037- compared with CE3-inoculated plants. Supporting these results, enzymes involved in ureide synthesis were reduced while activity of aspartate aminotransferase, glutamate synthase, sucrose synthase, and glucose-6-P dehydrogenase were increased in CFN037-induced nodules. Glutamate synthase and phosphoenolpyruvate carboxylase transcripts were detected early in the development of nodules induced by CFN037 compared with CE3. However, plants inoculated with strain CE3-vhb, which express the Vitreoscilla sp. hemoglobin and also displays increased respiratory capacity, did not have altered ureide transport in N2-fixing plants. The data suggest that inoculation with special selected mutant strains of R. etli can modulate nodule N assimilation and N transport compounds.

Nitrogen Assimilation in Nitrogen‐Fixing Soybean Plants during Phosphorus Deficiency

This study evaluated the hypothesis that P deficiency alters N assimilation in N2‐fixing soybean plants (Glycine max L. Merr.) through effects on nodule carbohydrate status. Transport of N constituents in xylem sap, concentrations and total quantities of N fractions in plant organs, and nonstructural carbohydrate status of nodules were compared in P‐sufficient (1.0 mM P) and P‐deficient (0.05 mM P) greenhouse‐grown soybean plants dependent solely on symbiotic N2 fixation. Increased ureide concentration in nodules and decreased ureide concentration in the xylem sap indicated that P deficiency restricted ureide transport from nodules to the xylem. Decreased ureide and total N concentration and flux in xylem sap, and decreased concentrations of ureide and soluble and insoluble reduced N in leaves and stems indicated that P deficiency decreased N delivery to the shoot. Phosphorus deficiency decreased the sap exudation rate per unit root mass by 75%. Thus, decreased exudation rate, as well as decreased constituent concentrations, contributed to decreased flux of N compounds in the xylem. The decreased exudation rate may have resulted from decreased hydraulic conductivity of the root system and/or decreased ion transport into the xylem. Phosphorus deficiency caused decreased hexose and sucrose concentrations and increased starch concentration in nodules, indicating impaired carbohydrate utilization. This impaired carbohydrate utilization and the resulting decrease in energy status may have contributed to the decreased movement of ureide, and other N constituents, from host cells into the xylem of nodules.

Changes in ureide synthesis, transport and assimilation following ammonium nitrate fertilization of nodulated Soybeans

Abstract The effect of ammonium nitrate fertilization on ureide synthesis, xylem transport and assimilation was examined in four week old nodulated soybean plants. In nodules the activity of 5‐phosphoribosylpyrophosphate amidotransferase, a key enzyme of ureide biosynthesis, declined 75%, while enzymes of purine oxidation (xanthine dehydrogenase and uricase) showed no response to nitrogen treatment. Xylem sap concentrations of allantoin and allantoate, as compared to untreated controls, were reduced by about 85% and 65%, respectively. Despite the substantial decline in ureide synthesis and transport, allantoate concentration in leaves of ammonium nitrate treated plants increased by about three‐fold. No ureide accumulation was observed in leaf tissue following a suppression of ureide formation in nodules by allopurinol, an inhibitor of xanthine dehydrogenase. In the pathway of ureide assimilation in leaves, application of inorganic nitrogen had no effect on activity of allantoinase, but caused a 50% drop i...

Relative efficiency, ureide transport and harvest index in soybeans inoculated with isogenic HUP mutants of Bradyrhizobium japonicum

Differences between isogenic uptake hydrogenase (HUP) mutants of Bradyrhizobium japonicum in terms of nodule efficiency, N2 fixation and N incorporation into various plant parts were studied in a monoxenic greenhouse experiment in order to confirm previous results with soybeans and beans inoculated with various HUP+ and HUP− strains. The HUP+ revertant PJ17-1 of a HUP− mutant (PJ17) of strain USDA DES 122 showed a completely restored relative efficiency (100% versus 78±2% for the HUP− mutant), higher nodule efficiency (N2 fixed per g nodules), higher ureide-N transport rates, higher N contents in pods and higher N harvest indices. All these observations confirm previous experiments with HUP+ and HUP− strains.

Growth ofPhaseolus vulgarison Various Nitrogen Sources: The Importance of Urease

Hine, J. C. and Sprent, J. I. 1988. Growth of Phaseolus vulgaris on various nitrogen sources: the importance of urease.—J. exp. Bot. 39: 1505 -1512. Rhizobium-moculated plants of Phaseolus vulgaris L. were growo with different N-sources (nitrate, ammonium, urea) and different concentrations of urea. The distribution of growth between plant parts varied with N-sources. Nitrate and ammonium were more inhibitory to nodulation than urea, which at 4 0 mol m-3 N had no effect. Urease activity varied in amount and location over a range of urea concentrations. At higher concentrations, more urea was transported to and increased urease activity was found in the shoot. Lower levels of activity in plants reliant on N2-fixation were consistent with a ureide-degradation pathway not involving urea. Moderate doses of urea could be assimilated concomitantly with N2-fixation. At higher levels of applied urea, nodulation and ureide transport to the shoots were reduced, although increased growth could not be maintained at concentrations of applied urea greater than 60 mol m-3 urea N.

Effect of Rhizobium japonicum Mutants with Enhanced N2 Fixation Activity on N Transport and Photosynthesis of Soybeans during Vegetative Growth1

The effects of two mutant strains of Rhizobium japonicum with enhanced capacities N2 fixation, on ureide transport, photosynthesis,root respiration, and growth in soybeans [Glycine max (L.) Merr. ‘Wells’] were examined during vegetative growth. Three‐week‐old plants inoculated with the mutants were larger and had higher rates of acetylene reduction, photosynthesis, xylary ureide transport, leaf allantoinase activity and higher concentrations of ureides in shoot tissues than did plants inoculated with the wild‐type strain. The relative composition of xylem sap N was unaffected by Rhizobium strain. Ureides comprised over 90% of the sap N in all treatments. After 5 weeks growth, however, there were no statistically significant differences among treatments in rates of the physiological processes and growth parameters examined. Cross contamination of nodules containing the mutant strains with the wild‐type strain was not responsible for the apparent loss of the initial advantages to the host plant (in terms of N2 fixation and growth) brought about by the use of the mutant strains. The use of modifications in Rhizobium genetics for improving the dependency of the host plant on N2 fixation for its N requirements is briefly discussed.