Effect of phosphorus and zinc nutrition on soybean seed phytic Acid and zinc

The relationships between nutrient P and Zn levels and the phytic acid, P, and Zn concentrations in soybean (Glycine max L. Merr. cv `Williams 79') seed were studied. Phytic acid increased linearly from 4.2 to 19.2 milligrams per gram as nutrient P treatment was varied from 2.0 to 50 milligrams per liter and Zn was held constant at 0.05 milligrams per liter. Leaf P concentration during seed development was found to be closely related to the concentrations of seed P and phytic acid. Leaf and seed Zn concentrations both responded positively to increasing nutrient Zn treatment. The effects of P treatment on plant and seed P and phytic acid were largely independent of the effects of Zn treatment on leaf and seed Zn. Phytic acid to Zn molar ratios ranging from 3.6 to 33.8 were observed.

The effects of nutrient P treatments on the concentrations of phytic acid, seed P, and leaf P were also studied in the P-sensitive (gene np) cultivars `Harosoy' and `Clark' and their respective P-tolerant (gene Np) near-isogenic lines L66-704 and L63-1677. In general, the positive relationships observed among nutrient P, leaf P, seed P, and phytic acid concentrations were similar to those observed in the studies with Williams 79. When fertilized with low or moderate nutrient P (2.5 and 25.0 milligrams P per liter, respectively) no significant differences in any parameter were observed between Harosoy or Clark and their respective P-tolerant isolines. When fertilized with high nutrient P (100 milligrams P per liter), Harosoy seed had a significantly higher concentration of phytic acid (30 milligrams per gram) than did seed of its P-tolerant near-isogenic line L66-704 (24.2 milligrams per gram phytic acid), whereas no significant difference was observed between Clark and its P-tolerant near-isogenic line L63-1677 (22.8 and 21.6 milligrams per gram, respectively). Variation in the phytic acid concentrations in the mature seed of the cultivars and isolines more closely paralleled leaf P concentrations observed during seed development (49 days after flowering), than those observed at the onset of seed development (14 days after flowering). Electrophoresis and ion-exchange chromatography revealed that partially phosphorylated intermediates do not appear when phytic acid accumulation is greatly reduced by limiting the nutrient P or when accumulation is greatly accelerated by excess P.

     

Relationship of endogenous abscisic Acid to sucrose level and seed growth rate of soybeans

It has been proposed that abscisic acid (ABA) may stimulate sucrose transport into filling seeds of legumes, potentially regulating seed growth rate. The objective of this study was to determine whether the rate of dry matter accumulation in seeds of soybeans (Glycine max L.) is correlated with the endogenous levels of ABA and sucrose in those sinks. The levels of ABA and sucrose in seed tissues were compared in nine diverse Plant Introduction lines having seed growth rates ranging from 2.5 to 10.0 milligrams dry weight per seed per day. At 14 days after anthesis (DAA), seeds of all genotypes contained less than 2 micrograms of ABA per gram fresh weight. Levels of ABA increased rapidly, however, reaching maxima at 20 to 30 DAA, depending upon tissue type and genotype. ABA accumulated first in seed coats and then in embryos, and ABA maxima were higher in seed coats (8 to 20 micrograms per gram fresh weight) than in embryos (4 to 9 micrograms per gram fresh weight. From 30 to 50 DAA, ABA levels in both tissues decreased to less than 2 micrograms per gram fresh weight. Levels of sucrose were also low early in development, less than 10 milligrams per gram fresh weight at 14 DAA. However, by 30 DAA, sucrose levels in seed coats had increased to 20 milligrams per gram fresh weight and remained fairly constant for the remainder of the filling period. In contrast, sucrose accumulated in embryos throughout the filling period, reaching levels greater than 40 milligrams per gram fresh weight by 50 DAA. Correlation analyses indicated that the level of ABA in seed coats and embryos was not directly correlated to the level of sucrose measured in those tissues or to the rate of seed dry matter accumulation during the linear filling period. Rather, the ubiquitous pattern of ABA accumulation early in development appeared to coincide with water uptake and the rapid expansion of cotyledons occurring at that time. Whole tissue sucrose levels in embryos and seed coats, as well as sucrose levels in the embryo apoplast, were generally not correlated with the rate of dry matter accumulation. Thus, it appears that, in this set of diverse soybean genotypes, seed growth rate was not limited by endogenous concentrations of ABA or sucrose in reproductive tissues.     

Origin and seed phenotype of maize low phytic acid 1-1 and low phytic acid 2-1

Phytic acid (myo-inositol-1, 2, 3, 4, 5, 6-hexakisphosphate or Ins P(6)) typically represents approximately 75% to 80% of maize (Zea mays) seed total P. Here we describe the origin, inheritance, and seed phenotype of two non-lethal maize low phytic acid mutants, lpa1-1 and lpa2-1. The loci map to two sites on chromosome 1S. Seed phytic acid P is reduced in these mutants by 50% to 66% but seed total P is unaltered. The decrease in phytic acid P in mature lpa1-1 seeds is accompanied by a corresponding increase in inorganic phosphate (P(i)). In mature lpa2-1 seed it is accompanied by increases in P(i) and at least three other myo-inositol (Ins) phosphates (and/or their respective enantiomers): D-Ins(1,2,4,5,6) P(5); D-Ins (1,4,5,6) P(4); and D-Ins(1,2,6) P(3). In both cases the sum of seed P(i) and Ins phosphates (including phytic acid) is constant and similar to that observed in normal seeds. In both mutants P chemistry appears to be perturbed throughout seed development. Homozygosity for either mutant results in a seed dry weight loss, ranging from 4% to 23%. These results indicate that phytic acid metabolism during seed development is not solely responsible for P homeostasis and indicate that the phytic acid concentration typical of a normal maize seed is not essential to seed function.     

Embryo-specific silencing of a transporter reduces phytic acid content of maize and soybean seeds

Phytic acid in cereal grains and oilseeds is poorly digested by monogastric animals and negatively affects animal nutrition and the environment. However, breeding programs involving mutants with less phytic acid and more inorganic phosphate (P(i)) have been frustrated by undesirable agronomic characteristics associated with the phytic acid-reducing mutations. We show that maize lpa1 mutants are defective in a multidrug resistance-associated protein (MRP) ATP-binding cassette (ABC) transporter that is expressed most highly in embryos, but also in immature endosperm, germinating seed and vegetative tissues. Silencing expression of this transporter in an embryo-specific manner produced low-phytic-acid, high-Pi transgenic maize seeds that germinate normally and do not show any significant reduction in seed dry weight. This dominant transgenic approach obviates the need for incorporating recessive lpa1 mutations to create maize hybrids with reduced phytic acid. Suppressing the homologous soybean MRP gene also generated low-phytic-acid seed, suggesting that the strategy might be feasible for many crops.     

A novel phytase with sequence similarity to purple acid phosphatases is expressed in cotyledons of germinating soybean seedlings

Phytic acid (myo-inositol hexakisphosphate) is the major storage form of phosphorus in plant seeds. During germination, stored reserves are used as a source of nutrients by the plant seedling. Phytic acid is degraded by the activity of phytases to yield inositol and free phosphate. Due to the lack of phytases in the non-ruminant digestive tract, monogastric animals cannot utilize dietary phytic acid and it is excreted into manure. High phytic acid content in manure results in elevated phosphorus levels in soil and water and accompanying environmental concerns. The use of phytases to degrade seed phytic acid has potential for reducing the negative environmental impact of livestock production. A phytase was purified to electrophoretic homogeneity from cotyledons of germinated soybeans (Glycine max L. Merr.). Peptide sequence data generated from the purified enzyme facilitated the cloning of the phytase sequence (GmPhy) employing a polymerase chain reaction strategy. The introduction of GmPhy into soybean tissue culture resulted in increased phytase activity in transformed cells, which confirmed the identity of the phytase gene. It is surprising that the soybean phytase was unrelated to previously characterized microbial or maize (Zea mays) phytases, which were classified as histidine acid phosphatases. The soybean phytase sequence exhibited a high degree of similarity to purple acid phosphatases, a class of metallophosphoesterases.     

Response of sugar interconversion, starch and protein accumulation to supplied metabolites during pod filling in chickpea

Detached chickpea inflorescences bearing pods at 20 days after flowering (DAF) were cultured for 5 days in complete liquid medium supplemented separately with asparate, myo-inositol, alpha-ketoglutarate and phytic acid. Effect of these metabolites on sugar interconvestion and starch and protein accumulation in developing pods was studied. Substituting asparate (62.5 mM) for glutamine in culture medium decreased relative proportion of sucrose in all pod tissues but increased the level of sugars, starch and protein in pod wall and cotyledons. In cotyledons, whereas myo-inositol (75 mM) reduced the accumulation of starch without affecting protein level, alpha-ketoglutarate (44 mM) increased both starch and protein accumulation. Both myo-inositol and alpha-ketoglutarate increased relative proportion of sucrose in cotyledons. Phytic acid (1 mM) decreased in cotyledons 14C incorporation from glucose into EtOH extract (principally constituted by sugars), amino acids and proteins but increased the same into starch. In cotyledons, phytic acid also increased 14C incorporation from glutamate into amino acids but this increase was negatively correlated with protein synthesis. Phytic acid decreased the relative distribution of 14C from glucose and glutamate into sucrose from pod wall but enhanced the same into EtOH extract from embryo. Based on the results, it is suggested that mode of metabolic response to exogenously supplied metabolites widely differs in pod tissues of chickpea.     

Reduced phytic Acid content does not have an adverse effect on germination of soybean seeds

Altering the level of phytic acid phosphorus by nutritional means had no effect on the ability of soybean (Glycine max L. [Merr.], cv `Williams 79') seeds to germinate under laboratory or greenhouse conditions. Dry matter moved out of the cotyledons at similar rates whether the germinating seeds initially contained low (0.19), medium (0.59), or high (1.00 milligram per seed) phytic acid phosphorus. Growth of roots and shoots from 3 to 9 days after planting was similar for seeds containing low and medium levels of phytic acid phosphorus. The medium level of phytic acid resembles that found in field-grown seed, so it is clear that soybean seeds normally contain a phosphorus reserve far above that needed for germination and early seedling growth.     

Approaches and challenges to engineering seed phytate and total phosphorus

About 75% of seed total phosphorus (P) is found in a single compound, phytic acid (myo-inositol-1,2,3,4,5,6-hexakisphosphate or InsP₆). Phytic acid is not efficiently utilized by monogastric animals (poultry, swine, fish), which creates phosphorus management and environmental impact problems in animal production. Phytic acid also functions as an antinutrient when consumed in human and animal diets. These problems can be addressed via feed or food supplementation with P and other minerals or phytase, or more efficiently and sustainably at their source by crop breeding or bioengineering of low-phytic acid/high-available P crops. However, since phytic acid and its synthetic pathways are central to a number of metabolic, developmental and signaling pathways important to plant function and productivity, low-phytate can translate into low-yield or stress susceptibility. The biological functions of phytic acid and identification of genetic resources and strategies useful in engineering high-yielding, stress-tolerant low-phytate germplasm will be reviewed here. One promising approach that can avoid undesirable outcomes due to impacts on phytic acid metabolism is to engineer “high-phytase” seeds. In contrast to the issue of seed phytic acid, there has been relatively little interest in seed total P as a trait of agricultural importance. However, seed total P is very important to the long-term goal of sustainable and environmentally friendly agricultural production. Certain low-phytate genotypes are also “low-total P”, which might represent the ideal seed P trait for nearly all end-uses, including uses in ruminant and non-ruminant feeds and in biofuels production. Future research directions will include screening for additional genetic resources such as seed total P mutants.     

Seed abortion in Pongamia pinnata (Fabaceae)

In Pongamia pinnata only one of the two ovules develops into a seed in most of the pods. Since pollen was not found to be limiting and reduced fertilization could not completely explain the observed frequency of seed abortion, it implied an effect of postfertilization factors. Aqueous extracts of developing seeds and maternal tissue (placenta) did not influence abortion in vitro, suggesting that abortion may not be mediated by a chemical. Experimental uptake of ¹⁴C sucrose in vitro indicated that both the stigmatic and the peduncular seed have similar inherent capacities of drawing resources, but the peduncular seed is deprived of resources in the presence of the stigmatic seed. This deprivation of the peduncular seed could be offset by supplying an excess of hormones leading to the subsequent formation of two seeds in a pod. The prevalence of single-seeded pods in P. pinnata seems therefore to be a result of competition between the two seeds for maternal resources. The evolutionary significance of single-seeded pods in P. pinnata is discussed with respect to possible dispersal advantage enjoyed by such pods.     

The maize low-phytic acid 3 encodes a myo-inositol kinase that plays a role in phytic acid biosynthesis in developing seeds

Phytic acid, myo-inositol-1,2,3,4,5,6-hexakisphosphate or Ins P6, is the most abundant myo-inositol phosphate in plant cells, but its biosynthesis is poorly understood. Also uncertain is the role of myo-inositol as a precursor of phytic acid biosynthesis. We identified a low-phytic acid mutant, lpa3, in maize. The Mu-insertion mutant has a phenotype of reduced phytic acid, increased myo-inositol and lacks significant amounts of myo-inositol phosphate intermediates in seeds. The gene responsible for the mutation encodes a myo-inositol kinase (MIK). Maize MIK protein contains conserved amino acid residues found in pfkB carbohydrate kinases. The maize lpa3 gene is expressed in developing embryos, where phytic acid is actively synthesized and accumulates to a large amount. Characterization of the lpa3 mutant provides direct evidence for the role of myo-inositol and MIK in phytic acid biosynthesis in developing seeds. Recombinant maize MIK phosphorylates myo-inositol to produce multiple myo-inositol monophosphates, Ins1/3P, Ins4/6P and possibly Ins5P. The characteristics of the lpa3 mutant and MIK suggest that MIK is not a salvage enzyme for myo-inositol recycling and that there are multiple phosphorylation routes to phytic acid in developing seeds. Analysis of the lpa2/lpa3 double mutant implies interactions between the phosphorylation routes.     

Isolation and characterisation of an lpa (low phytic acid) mutant in common bean (Phaseolus vulgaris L.)

Phytic acid is considered as one of the major antinutritional compounds in cereal and legume seeds. The development of lpa (low phytic acid) grains, resulting in increased mineral cation availability, is considered a major goal in the improvement of the nutritional quality of seed crops, especially those largely consumed in developing countries. From a mutagenised population of common bean we isolated a homozygous lpa mutant line (lpa-280-10) showing, compared to wild type, a 90% reduction of phytic acid, a 25% reduction of raffinosaccharides and a much higher amount of free or weakly bound iron cations in the seed. Genetic analysis showed that the lpa character is due to a recessive mutation that segregates in a monogenic, Mendelian fashion. Germination tests performed using varying ageing or stress conditions, clearly showed that the bean line lpa-280-10 has a better germination response than the wild type. These data, together with those obtained from 2 years of agronomic trials showing that the mutant seed yield is close to that of its parents and other evidence, indicate that the new lpa-280-10 mutation might be the first devoid of visible macroscopic negative effects in plants, pods and seeds. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Phytic acid and raffinose series oligosaccharides metabolism in developing chickpea seeds

Phytic acid and raffinose series oligosaccharides (RFOs) have anti-nutritional properties where phytic acid chelates minerals and reduces their bioavailability to humans and other animals, and RFOs cause flatulence. Both phytic acid and RFOs cannot be digested by monogastric animals and are released as pollutant-wastes. Efforts are being made to reduce the contents of these factors without affecting the viability of seeds. This will require a thorough understanding of their metabolism in different crops. Biosynthetic pathways of both metabolites though are interlinked but not well described. This study was made on metabolism of these two contents in developing chickpea (Cicer arietinum L cv GL 769) seeds. In this study, deposition of RFOs was found to occur before deposition of phytic acid. A decline in inorganic phosphorus and increase in phospholipid phosphorus and phytic acid was observed in seeds during development. Acid phosphatase was the major phosphatase in seed as well as podwall and its activity was highest at early stage of development, thereafter it decreased. Partitioning of ¹⁴ C label from ¹⁴ C-glucose and ¹⁴ C-sucrose into RFOs and phytic acid was studied in seeds in presence of inositol, galactose and iositol and galactose, which favored the view that galactinol synthase is not the key enzyme in RFOs synthesis.     

Quantitative trait loci associated with soybean seed weight and composition under different phosphorus levels

Seed size and composition are important traits in food crops and can be affected by nutrient availability in the soil. Phosphorus (P) is a non‐renewable, essential macronutrient, and P deficiency limits soybean (Glycine max) yield and quality. To investigate the associations of seed traits in low‐ and high‐P environments, soybean recombinant inbred lines (RILs) from a cross of cultivars Fiskeby III and Mandarin (Ottawa) were grown under contrasting P availability environments. Traits including individual seed weight, seed number, and intact mature pod weight were significantly affected by soil P levels and showed transgressive segregation among the RILs. Surprisingly, P treatments did not affect seed composition or weight, suggesting that soybean maintains sufficient P in seeds even in low‐P soil. Quantitative trait loci (QTLs) were detected for seed weight, intact pods, seed volume, and seed protein, with five significant QTLs identified in low‐P environments and one significant QTL found in the optimal‐P environment. Broad‐sense heritability estimates were 0.78 (individual seed weight), 0.90 (seed protein), 0.34 (seed oil), and 0.98 (seed number). The QTLs identified under low P point to genetic regions that may be useful to improve soybean performance under limiting P conditions.     

Isoflavonoid biosynthesis and accumulation in developing soybean seeds

Isoflavonoids are biologically active natural products that accumulate in soybean seeds during development. The amount of isoflavonoids present in soybean seed is variable, depending on genetic and environmental factors that are not fully understood. Experiments were conducted to determine whether isoflavonoids are synthesized within seed tissues during development, or made in other plant organs and transported to the seeds where they accumulate. An analysis of isoflavonoids by HPLC detected the compounds in all organs of soybean plant, but the amount of isoflavonoids present varied depending on the tissue and developmental stage. The greatest concentrations were found in mature seeds and leaves. The 2-hydroxyisoflavanone synthase genes IFS1 and IFS2 were studied to determine their pattern of expression in different tissues and developmental stages. The highest level of expression of IFS1 was observed in the root and seed coat, while IFS2 was most highly expressed in embryos and pods, and in elicitor-treated or pathogen-challenged tissues. Incorporation of radiolabel into isoflavonoids was observed when developing embryos and other plant organs were fed with [(14)C]phenylalanine. Embryos excised from developing soybean seeds also accumulated isoflavonoids from a synthetic medium. A maternal effect on seed isoflavonoid content was noted in reciprocal crosses between soybean cultivars that differ in seed isoflavonoids. From these results, we propose that developing soybean embryos have an ability to synthesize isoflavonoids de novo, but that transport from maternal tissues may in part contribute to the accumulation of these natural products in the seed.     

Synthesis and accumulation of nitrite in soybean nodules supplied with nitrate

Nodulated soybean plants (Glycine max [L.] Merr) were grown in sand culture without combined N or with a continuous supply of nitrate in nutrient solution. Moderate nitrate concentration (30 milligrams N per liter) had little effect on nodule weight/plant while high nitrate concentration (100 milligrams N per liter) depressed nodule weight/plant by 70 to 80% with harvests 30 to 60 days after planting and initiation of nitrate treatments.     

Phytic acid in green leaves

Phytic acid or phytate, the free‐acid form of myo‐inositolhexakiphosphate, is abundant in many seeds and fruits, where it represents the major storage form of phosphorus. Although also known from other plant tissues, available reports on the occurrence of phytic acid, e.g. in leaves, have never been compiled, nor have they been critically reviewed. We found 45 published studies with information on phytic acid content in leaves. Phytic acid was almost always detected when studies specifically tried to detect it, and accounted for up to 98% of total P. However, we argue that such extreme values, which rival findings from storage organs, are dubious and probably result from measurement errors. Excluding these high values from further quantitative analysis, foliar phytic acid‐P averaged 2.3 mg·g^?1, and represented, on average, 7.6% of total P. Remarkably, the ratio of phytic acid‐P to total P did not increase with total P, we even detected a negative correlation of the two variables within one species, Manihot esculenta. This enigmatic finding warrants further attention.     

玉米中植酸代谢研究的策略

作物尤其是玉米的种子中积累了丰富的植酸。早先的研究侧重于降低种子中植酸的含量,但是随着人们对植酸认识的深入,发现植酸对于动、植物而言具有不可替代的生物功能。对于人和动物而言,植酸有抗营养作用,但也是重要的健康因子;对于植物而言,植酸及其代谢中间体的生物学功能却缺乏明确的研究。若要明确把握植酸的育种方向,就必须对植酸在植物中的合成过程有明确的认识。但自植酸被发现至今,人们对于其在高等植物中的合成过程仍然知之甚少,对其生物学功能更是缺乏全面的了解。本文综述了植酸代谢研究的现状,分析并总结了植酸的代谢通路,指出了植酸代谢研究的突破点,结合植酸代谢的研究特点和进展,比较了基因同源克隆、关联分析等4种最具潜力的研究策略。     

Phytic acid mobilization is an early response to chilling of the embryonic axes from dormant oilseed of hazel (Corylus avellana)

Dormancy of hazel (Corylus avellana L.) seeds is alleviated by a chilling treatment during which cytological, hormonal, and biochemical changes occur. Phytic acid and phosphate mobilization have been examined during this treatment. Phytic acid accounted for 0.7% and up to 3.2% of dry weight in axiferous and cotyledonary tissue, respectively. Phytic acid levels in embryonic axes were reduced by 60% within the first 3 weeks of chilling, with little subsequent change, in contrast to warm-imbibed tissue where levels did not change significantly. In cotyledons, phytic acid was mobilized to a lesser extent. Phosphate levels expressed on a fresh weight basis remained almost unaltered suggesting either the operation of a homeostatic mechanism for intracellular concentration or rapid utilization due to active metabolism. Phytase activity increased during stratification in both axiferous and cotyledonary tissue. The initial rise observed was associated with dormancy alleviation, since it occurred before the realization of full germination potential by the seeds and not in warm-imbibed tissue. Protein bodies were isolated from hazel seeds by non-aqueous density gradients. Phytase activity was closely associated with the purified organelles, where phytic acid was located by light microscopy. Overall, these findings suggest that phytic acid mobilization by phytase and previously described processes associated with protein bodies, such as considerable proteolysis, are early participants in the plethora of events leading to seed dormancy relief and germination in hazel.