Identification of mutants in phosphorus metabolism

Phosphorus availability is often limiting for plant growth. However, little is known of the pathways and mechanisms that regulate phosphorus (P) uptake and distribution in plants. We have developed a screen based on the induction of secreted root acid phosphatase activity by low‐P stress to identify mutants of Arabidopsis thaliana with defects in P metabolism. Acid phosphatase activity was detected visually in the roots of A. thaliana seedlings grown in vitro on low‐P medium, using the chromogenic substrate, 5‐bromo‐4‐chloro‐3‐indolyl‐phosphate (BCIP). In low‐P stress conditions the roots of wild‐type plants stained blue, as the induced root acid phosphatase cleaved BCIP to release the coloured product. Potential mutants were identified as having white, or pale blue, roots under these conditions. Out of approximately 79 000 T‐DNA mutagenised seedlings screened, two mutants with reduced acid phosphatase staining were further characterised. Both exhibited reduced growth and differences in their P contents when compared to wild‐type A. thaliana. The mutant with the most severe phenotype, pho3, accumulated high levels of anthocyanins and starch in a distinctive visual pattern within the leaves. The phenotypes of these mutants are distinct from two previously identified phosphorus mutants (phol and pho2) and from an acid phosphatase deficient mutant (pupl) of A. thaliana. This suggested that the screening method was robust and might lead to the identification of further mutants with the potential for increasing our understanding of P nutrition.     

The role of cell wall components from groundnut roots in solubilizing sparingly soluble phosphorus in low fertility soils

Groundnuts have a superior ability to take up P from soils with low P fertility compared to sorghum and soybean. Previous experiments showed that this ability was neither attributable to better root development nor to root exudates capable of solubilizing Fe- and Al-bound P, the sparingly soluble P forms in soils. Direct "contact reactions" between cell wall components from these 3 plant species (groundnut, soybean and sorghum) and P-fixing Fe and Al minerals were examined. Cell wall preparations from groundnut roots showed a superior P solubilizing ability than those of soybean and sorghum. Cell wall activity of groundnut roots may thus at least partly explain the superior growth of this crop under P-deficient conditions. To characterize the active site responsible for P solubilization, effects of pH, heat, addition of cations, and digestion with enzymes (pectinase and cellulase) or HCl on P solubilization were investigated. Conclusion are 1) Solubilizing ability is not related to root CEC because soybean with higher root CEC showed an inferior solubilizing ability compared to groundnut. 2) The reaction site of cell-walls of groundnut roots is stable against heating and digestion with cellulase and pectinase. 3) Solubilizing ability was severely reduced by digestion with HCl. 4) Pre-treating cell walls with either Al3+, Fe3+, or Ga3+ decreased solubilizing ability but cations with lower valency such as Na+, K+, Ca2+ or Mg2+ had no effect. Soaking roots of groundnuts grown in solution culture in 0.5 M NaOH for 30 seconds prior to cell wall preparation led to a 30% reduction in solubilization of P from FePO4 without permanently damaging plants. This suggests that 5) the active component of the cell walls was located on the root epidermal cell surfaces. Based on these results a phosphorus solubilizing mechanism is proposed.     

Induction of maize acid phosphatase activities under phosphorus starvation

Large variation in phosphorus-(P) acquisition efficiency exists among maize inbred and hybrid genotypes. Acid phosphatases are a type of enzyme that affects P acquisition and P-use efficiency in plants. The objectives of this research were (1) to characterize acid phosphatase activity in maize grown hydroponically under P starvation, and (2) to determine if there is differential induction of acid phosphatases in two maize genotypes previously characterized as P efficient (Mo17) and P inefficient (B73). B73 and Mo17 seedlings were grown hydroponically and both intracellular and secreted acid phosphatase activities were characterized. Fresh seedling weight of both genotypes decreased under P starvation, but percent fresh weight allocated to roots increased 14 days after P starvation in B73. Soluble protein concentration in shoots and roots was affected little, but secreted protein decreased by 40 and 20% in B73 and Mo17 seedlings grown without P for 14 days. Intracellular and secreted acid phosphate activity increased substantially in leaves and roots in B73 and Mo17 in response to P starvation. Secreted APase activity per unit protein increased 310 and 300% in B73 and Mo17, respectively, 7 days after P withdrawal. One of the minor isozymes identified on non-denaturing PAGE, was increased specifically in response to P starvation in both maize genotypes. The patterns and levels of change in APase activities in B73 and Mo17 were not sufficiently different to account for the diverse growth response of these genotypes in low-P conditions. The results suggest that APases may not be a major mechanism for scavenging or acquiring P and changes in APases may reflect a state of P stress in both varieties. Other factors such as root architecture, secretion of low-molecular weight carboxylates and microbial interactions might explain the difference between these two genotypes.     

Root interactions and tomato growth in tomato/potato onion companion-cropping system under different phosphorus levels

When two plants interact, changes in plant growth are usually related to variations in root distribution and phosphorus (P) levels. However, root distributions and root tendencies are difficult to study because root systems grow beneath the soil surface. In this study, a transparent root box was used to observe interactions between root systems in situ, and the relation between tomato growth and root proliferation at different depths and distance from the rows at no P added and 120?mg kg^-1 P added levels were also tested. We found that tomato shoot and total biomass increased and roots grew deeper when companion cropped with potato onion under both P levels. Moreover, tomato roots tended to grow away from the potato onion roots. Our results suggest that a deeper and more evasive root distribution may be related to the increased plant growth of tomato when companion cropped with potato onion.     

不同基因型苦荞幼苗对低磷胁迫的响应

采用沙培法,以4个不同耐低磷苦荞(Fagopyrum tataricum(L.) Gaertn)品种为材料,设正常磷处理(P1,2 mmol/L对照)、低磷胁迫(P2,1 mmol/L)和极低磷胁迫(P3,0.2 mmol/L) 3个处理,研究低磷胁迫对苦荞苗期农艺性状、生理生化指标以及植株磷利用的影响。结果显示:(1)低磷胁迫下,苦荞苗期株高、茎粗、叶面积、地上部干重、根系干重、根系平均直径、根系表面积、根系体积等指标均有所下降;主根伸长、根冠比有所升高,但不同品种的升降幅度有所不同。(2)低磷胁迫使苦荞叶绿素含量、可溶性蛋白含量和根系活力均有所下降,根系的SOD活性、POD活性、酸性磷酸酶活性、可溶性糖含量、游离脯氨酸含量显著增加,且表现为耐低磷苦荞品种的增幅大于不耐低磷苦荞。(3)低磷胁迫使苦荞植株全磷含量和单株磷积累量下降,却使磷利用效率升高。研究结果表明耐低磷品种通过主根伸长下扎以及分泌较多的酸性磷酸酶,合理吸收与利用土壤磷素,通过保持叶片较高的叶绿素含量维持较强的光合能力,通过保持较高的抗氧化酶活性降低膜脂过氧化伤害,最大程度的适应低磷环境。     

Functional significance of dauciform roots: exudation of carboxylates and acid phosphatase under phosphorus deficiency in Caustis blakei (Cyperaceae)

Caustis blakei produces an intriguing morphological adaptation by inducing dauciform roots in response to phosphorus (P) deficiency. We tested the hypothesis that these hairy, swollen lateral roots play a similar role to cluster roots in the exudation of organic chelators and ectoenzymes known to aid the chemical mobilization of sparingly available soil nutrients, such as P. Dauciform-root development and exudate composition (carboxylates and acid phosphatase activity) were analysed in C. blakei plants grown in nutrient solution under P-starved conditions. The distribution of dauciform roots in the field was determined in relation to soil profile depth and matrix. The percentage of dauciform roots of the entire root mass was greatest at the lowest P concentration ([P]) in solution, and was suppressed with increasing solution [P], while in the field dauciform roots were predominantely located in the upper soil horizons, and decreased with increasing soil depth. Citrate was the major carboxylate released in an exudative burst from mature dauciform roots, which also produced elevated levels of acid phosphatase activity. Malonate was the dominant internal carboxylate present, with the highest concentration in young dauciform roots. The high concentration of carboxylates and phosphatases released from dauciform roots, combined with their prolific distribution in the organic surface layer of nutrient-impoverished soils, provides an ecophysiological advantage for enhancing nutrient acquisition.     

Secreted acid phosphatase is expressed in cluster roots of lupin in response to phosphorus deficiency

The roots of white lupin (Lupinus albus L. cv. Kievskij mutant) secrete acid phosphatase, S-APase, when they grow under conditions of low available phosphorus (P). S-APases hydrolyze organic phosphate compounds in the rhizosphere and supply inorganic phosphate to the plants. Low phosphorus availability also induces vigorous growth of cluster roots. In this study, the function of cluster roots was investigated with reference to S-APase secretion. White lupins were grown in hydroponic culture in a greenhouse under P-deficient and P-sufficient conditions. S-APase in the excised roots after treatment was detected by staining with 4-methylumbelliferone phosphate (MUP). Gene expression of S-APase in cluster and normal roots was also investigated. Activity was greatest in the roots of plants grown under conditions of P -deficiency, particularly in cluster roots. S-APase gene expression was induced by a decrease in internal P concentrations, and was especially high in cluster roots formed under conditions of P -deficiency. It was suggested that decrease of internal P concentration stimulated both of the S-APase expression and cluster root formation.     

甘蔗幼苗对难溶性磷的吸收及其根系对低磷胁迫的响应

为明确甘蔗适应低磷胁迫的生理生化机制,挖掘甘蔗对磷素的利用潜力,揭示甘蔗对低磷胁迫适应的可能机制,该研究以ROC22和ROC10两个甘蔗品种为材料,采用水培和土培的方法研究了甘蔗幼苗对难溶性磷的吸收及其在低磷胁迫下根构型和根系的生理反应。结果表明:(1)培养在以难溶性磷(Ca-P和Al-P)为磷源的培养液中的甘蔗的叶片数、地上部干重、生物量较缺磷(-P)处理显著增加,与对照(+P)的相当,甘蔗总磷积累量也显著提高,达到对照(+P)处理磷积累量的30%～77%。(2)在低磷条件下,甘蔗幼苗的根系有向土壤深层分布的趋势,根的总体积增大、最长根长变长、浅根系分布增多。(3)甘蔗幼苗在低磷环境下,根际环境明显酸化,且根系分泌物能溶解难溶性的铝磷,植株体内酸性磷酸酶的活性也明显增强。以上表明甘蔗幼苗有较强的吸收利用难溶性磷的能力,而低磷条件下根系数量增加、主根的向地性、浅根系分布增多、根际酸化以及植株体内酸性磷酸酶活性的增强可能是甘蔗幼苗适应缺磷环境的重要机制。     

缺磷条件下的小麦根系酸性磷酸酶活性研究

1　引　　言植物根可向根际分泌许多有机化合物 ,其中有许多物质都能促进植物对矿质养分的吸收 .作为必需大量营养元素的Ｐ ,在土壤中以无机磷酸盐阴离子的形式被吸收 ,而有机磷酸酯必须被水解成无机Ｐ后才能进入植物根 ,在这一过程中有一非常重要的步骤 ,就是由微生物、菌根外真菌和植物根分泌酸性磷酸酶 .土壤中的有机Ｐ一般占全Ｐ的 30 %～ 5 0 % ,有的可达95 % .因此 ,如何发挥植物自身利用土壤有机Ｐ的潜力已成为目前植物营养学研究的热点之一 .Ｇｏｌｄｓｔｅｉｎ等[3 ] 研究Ｐ胁迫条件下悬浮培养细胞时发现 ,抑制植物生长和诱导酸性…     

Morphogenetic Aspects of a Leafless Mutant in Tomato: III. GROWTH AND PEROXIDASE ACTIVITY OF ROOTS

Using morphological and biochemical criteria, comparisons weremade between intact and excised roots of normal tomato (Lycopersiconesculentum Mill.) and the reduced form of the homozygous lanceolatemutant. Intact normal roots showed greater growth as reflectedin length of the main root axis, number of lateral roots, andprotodermal cell size. Excised normal roots grew more rapidlythan those of the mutant only during the first 24-h intervalof the first week in culture. Intact mutant roots revealed agreater activity of peroxidase, but excised mutant roots showedno increase in enzyme activity. It is concluded that the primarysite of action of the mutant allele is the shoot system, andin particular the leaf marginal meristem. The effects of thelanceolate gene on the root system in tomato are interpretedas being of secondary importance with regard to gene action.     

供磷水平及方式对杉木幼苗根系生长和磷利用效率的影响

为研究杉木幼苗根系生长、形态学指标及养分利用效率对土壤磷素异质分布的响应规律,选择杉木种子园单株采种培育的半同胞家系实生幼苗为研究对象,采用室内沙培控磷盆栽试验,设计低浓度供磷(8 mg/kg KH_2PO_4)、正常供磷(16 mg/kg KH_2PO_4)和高浓度供磷(32 mg/kg KH_2PO_4)3个供磷水平,每个供磷水平分别采用2种供磷方式(局部供磷和均匀供磷)进行根部施磷。结果表明:(1)从供磷水平来看,低浓度供磷下的杉木根长、根系生物量、根冠比、根系及全株的磷素利用效率均显著大于正常供磷和高浓度供磷,而根平均直径相反;随着供磷水平的提高,杉木苗高和地上部生物量无显著差异,而比根长表现出逐渐降低的趋势。(2)从供磷方式来看,局部供磷处理的杉木苗高、根长、根系表面积、比根长、地上部生物量、根系及全株的磷素利用效率均显著大于均匀供磷处理,而根平均直径和根冠比则相反。总体上,低浓度局部供磷处理下杉木可明显增强其根系的形态可塑性,从而优化根系在养分异质土壤里的空间分布,并通过提高根系磷素利用效率以维持地上部的正常生长。     

Root cell-wall properties are proposed to contribute to phosphorus (P) mobilization by groundnut and pigeonpea

Groundnuts showed a superior ability to take up phosphorus (P) from two soils of extremely low fertility, where sorghum and soybean died of P deficiency. This ability could not be attributed to differences in root development, to P uptake parameters such as C_min, or to the excretion of root exudates capable of solubilizing iron- (Fe-P) and aluminum-bound P (Al-P), the sparingly soluble P forms in soils. A new P solubilizing mechanism (called `contact reaction') which occurs at the interface between root surface and soil particles, is therefore proposed. Isolated cell walls from groundnut roots solubilized more P from P-fixing minerals than those from sorghum and soybean roots. The P-solubilizing activity of groundnut root cell-walls might therefore be related to the superior growth of this crop under P-deficient conditions. The P-solubilizing active sites in groundnut root cell walls were located at the root surface and could act as chelating agent with Fe(III). This P-solubilizing active component in the cell walls could be extracted by NaOH, but not by HCl, and was identified as a small molecule through column chromatography with Sephadex LH-20. The P-solubilizing ability of pigeonpea root cell-walls was examined and found to be as high as that of groundnut. As pigeonpea plants excrete significant amount of root exudates with Fe-P solubilizing ability only after they flower, the P-solubilizing ability of root cell-walls may partially explain the high P efficiency of this species before it flowers.     

Root surface phosphatase activity in ecotypes of Aegilops peregrina

The relationships between root surface phosphatase activity and the edaphic factors of their native habitats were investigated in four ecotypes of Aegilops peregrina (Hack.) Maire et Weil. In one set of experiments plants were grown in phosphate-deficient nutrient solution cultures (5 μ M ) with three pH values: 5.5, 6.5 and 7.5. In a second series, plants were grown in both P-poor and P-rich soils.
Results showed an optimal activity of the commonly-described root surface acid phosphatase of pH 4.5–5.0 in the ecotypes Meron (a P-poor montmorillonitic, typical mediterranean Terra-Rossa soil) and Har-Hurshan (a P-rich calcareous soil). However, in the ecotypes Malkiya (a P-rich kaolinitic Terra-Rossa) and Bet-Guvrin (a P-rich calcareous soil) the optimal activity of the phosphatase occurred at pH 6.0. The pH level of the growth solution had no effect on the pH of optimal activity of the phosphatase in the ecotypes Malkiya and Bet-Guvrin, but it somewhat affected their level of activity.
Phosphatase activity was stimulated when plant roots were grown in a P-poor soil, as compared to the activity of those which were grown in a P-rich soil. Plants of the Malkiya ecotype exhibited the strongest activation of phosphatase as compared to the other three ecotypes. It seems that ecotypes which have evolved in P-rich soils may regulate their root surface phosphatase activity better than those which have evolved in P-poor soils.     

Long‐term nitrogen loading alleviates phosphorus limitation in terrestrial ecosystems

Increased human‐derived nitrogen (N) deposition to terrestrial ecosystems has resulted in widespread phosphorus (P) limitation of net primary productivity. However, it remains unclear if and how N‐induced P limitation varies over time. Soil extracellular phosphatases catalyze the hydrolysis of P from soil organic matter, an important adaptive mechanism for ecosystems to cope with N‐induced P limitation. Here we show, using a meta‐analysis of 140 studies and 668 observations worldwide, that N stimulation of soil phosphatase activity diminishes over time. Whereas short‐term N loading (≤5 years) significantly increased soil phosphatase activity by 28%, long‐term N loading had no significant effect. Nitrogen loading did not affect soil available P and total P content in either short‐ or long‐term studies. Together, these results suggest that N‐induced P limitation in ecosystems is alleviated in the long‐term through the initial stimulation of soil phosphatase activity, thereby securing P supply to support plant growth. Our results suggest that increases in terrestrial carbon uptake due to ongoing anthropogenic N loading may be greater than previously thought.     

A root hairless barley mutant for elucidating genetic of root hairs and phosphorus uptake

This paper reports a new barley mutant missing root hairs. The mutant was spontaneously discovered among the population of wild type (Pallas, a spring barley cultivar), producing normal, 0.8 mm long root hairs. We have called the mutant bald root barley (brb). Root anatomical studies confirmed the lack of root hairs on mutant roots. Amplified Fragment Length Polymorphism (AFLP) analyses of the genomes of the mutant and Pallas supported that the brb mutant has its genetic background in Pallas. The segregation ratio of selfed F₂ plants, resulting from mutant and Pallas outcross, was 1:3 (–root hairs:+root hairs), suggesting a monogenic recessive mode of inheritance.In rhizosphere studies, Pallas absorbed nearly two times more phosphorus (P) than the mutant. Most of available inorganic P in the root hair zone (0.8 mm) of Pallas was depleted, as indicated by the uniform P depletion profile near its roots. The acid phosphatase (Apase) activity near the roots of Pallas was higher and Pallas mobilised more organic P in the rhizosphere than the mutant. The higher Apase activity near Pallas roots also suggests a link between root hair formation and rhizosphere Apase activity. Hence, root hairs are important for increasing plant P uptake of inorganic as well as mobilisation of organic P in soils.Laboratory, pot and field studies showed that barley cultivars with longer root hairs (1.10 mm), extracted more P from rhizosphere soil, absorbed more P in low-P field (Olsen P=14 mg P kg^–1 soil), and produced more shoot biomass than shorter root hair cultivars (0.63 mm). Especially in low-P soil, the differences in root hair length and P uptake among the cultivars were significantly larger. Based on the results, the perspectives of genetic analysis of root hairs and their importance in P uptake and field performance of cereals are discussed.     

不同浓度和形态磷处理下内生真菌感染对高羊茅的影响

以感染内生真菌(endophyte-infected,EI)和不感染内生真菌(endophyte-free,EF)的高羊茅(Festuca arundinacea Schreb.)为材料,在温室沙培条件下研究内生真菌对高羊茅适应缺磷及利用不同形态磷肥的影响。结果表明,1)缺磷条件下,高羊茅EI和EF植株生长差异不显著;正常供磷条件下,高羊茅EI植株拥有更多分蘖数和绿叶数。说明正常供磷条件下内生真菌改善了宿主高羊茅的生长。2)与水溶性磷相比,高羊茅根有机酸和酸性磷酸酶(acid phosphatase,APase)活性在难溶性磷条件下显著增加,而根总酚含量无显著变化。在水溶性磷条件下,高羊茅EI植株根总酚含量显著高于EF植株,此时EI植株比EF植株拥有更多分蘖数和绿叶数,说明在水溶性磷条件下内生真菌对宿主地上部生长具有一定贡献。在难溶性磷条件下,虽然高羊茅EI植株根总酚含量仍然高于EF植株,但同时EI植株根有机酸含量显著低于EF植株,因此内生真菌感染只是增大了宿主植物的根冠比,而对分蘖数和绿叶数等无显著影响,说明内生真菌对宿主利用难溶性磷贡献不大。可见,内生真菌对宿主植物的生长在水溶性磷条件下更有利。     

Genotypic variability in phosphorus solubilizing activity of root exudates by pigeonpea grown in low-nutrient environments

A pot experiment confirmed that pigeonpea could efficiently utilize various sources of phosphorus (P) (aluminium phosphate, iron phosphate and apatite), irrespective of genotype. A qualitative assay method for iron (Fe)-P solubilizing activity showed that root exudates collected from P-deficient pigeonpea contained Fe-P solubilizing substances and that they were released mainly from root tips. Citric, malic, malonic, succinic and piscidic acids were identified in root exudates. Citric and piscidic acids release from roots was increased by low-P treatment in all the genotypes tested. The release rates of citric and piscidic acids were affected by the P concentration of shoots rather than that of roots. The pigeonpea roots released approximately 5–100 times more piscidic acid than citric acid depending on P stress status, plant age and genotype. When organic acids were added to Alfisols, citric acid was most capable of mobilizing P from the soil, followed by piscidic acid and malic acid. No correlation was found between genotypic variability in the release rates of citric and piscidic acids from the roots under low-P treatment at hydroponic culture and in the growth and P uptake of plants on Alfisols. Although citric and piscidic acids released from pigeonpea roots may play a partial role in solubilizing unavailable insoluble P in soils, the releases were thought to be an unsatisfactory strategy for explaining genotypic variation in low P availability of pigeonpea.     

Distribution in excised Lycopersicum esculentum roots of the principal enzymes involved in sucrose metabolism

Sucrose synthetase and sucrose phosphate synthetase could not be detected in 7-day-old excised tomato roots grown in sucrose. These roots, however, possessed a highly active acid invertase and a neutral invertase of low activity. The distribution of the cell wall-located acid invertase along the root axis appeared to be related to growth. This was not the case for the soluble enzyme. The possible functions of these two enzymes are discussed.     

Availability of organic and inorganic forms of phosphorus to lupins (Lupinus spp.)

Inositol phosphate is at least equal to KH₂PO₄ as a source of P for the growth of lupins in sand but a much poorer source in soil. RNA and glycerophosphate were excellent sources of P for lupin growth in a P-fixing soil. Soil and root phosphatase activity were not altered by amendment of soils with either inorganic- or organic-P. The difference in availability of differing P-sources is related to their solubility in soils rather than susceptibility to phosphatases.