Real-time [11C]methionine translocation in barley in relation to mugineic acid phytosiderophore biosynthesis

[11C]Methionine was supplied through barley roots and the 11C signal was followed for 90 min using a real-time imaging system (PETIS), with subsequent development of autoradiographic images of the whole plant. In all cases, [11C]methionine was first translocated to the 'discrimination center', the basal part of the shoot, and this part was most strongly labeled. Methionine absorbed by the roots of the plants was subsequently translocated to other parts of the plant. In Fe-deficient barley plants, a drastic reduction in [11C]methionine translocation from the roots to the shoot was observed, while a greater amount of 11C was found in the leaves of Fe-sufficient or methionine-pretreated Fe-deficient plants. Treatment of Fe-deficient plants with aminooxyacetic acid, an inhibitor of nicotianamine aminotransferase, increased the translocation of [11C]methionine to the shoot. The retention of exogenously supplied [11C]methionine in the roots of Fe-deficient barley indicates that the methionine is used in the biosynthesis of mugineic acid phytosiderophores in barley roots. This and the absence of methionine movement from shoots to the roots suggest that the mugineic acid precursor methionine originates in the roots of plants.     

52Fe Translocation in Barley as Monitored by a Positron-Emitting Tracer Imaging System (PETIS): Evidence for the Direct Translocation of Fe from Roots to Young Leaves via Phloem

The real-time translocation of iron (Fe) in barley (Hordeumvulgare L. cv. Ehimehadaka no. 1) was visualized using the positron-emittingtracer ⁵²Fe and a positron-emitting tracer imaging system (PETIS).PETIS allowed us to monitor Fe translocation in barley non-destructivelyunder various conditions. In all cases, ⁵²Fe first accumulatedat the basal part of the shoot, suggesting that this regionmay play an important role in Fe distribution in graminaceousplants. Fe-deficient barley showed greater translocation of⁵²Fe from roots to shoots than did Fe-sufficient barley, demonstratingthat Fe deficiency causes enhanced ⁵²Fe uptake and translocationto shoots. In the dark, translocation of ⁵²Fe to the youngestleaf was equivalent to or higher than that under the light condition,while the translocation of ⁵²Fe to the older leaves was decreased,in both Fe-deficient and Fe-sufficient barley. This suggeststhe possibility that the mechanism and/or pathway of Fe translocationto the youngest leaf may be different from that to the olderleaves. When phloem transport in the leaf was blocked by steamtreatment, ⁵²Fe translocation from the roots to older leaveswas not affected, while ⁵²Fe translocation to the youngest leafwas reduced, indicating that Fe is translocated to the youngestleaf via phloem in addition to xylem. We propose a novel modelin which root-absorbed Fe is translocated from the basal partof the shoots and/or roots to the youngest leaf via phloem ingraminaceous plants.     

Iron-Stress Induced Redox Activity in Tomato (Lycopersicum esculentum Mill.) Is Localized on the Plasma Membrane

Tomato plants (Lycopersicum esculentum Mill.) were grown for 21-days in a complete hydroponic nutrient solution including Fe³⁺-ethylenediamine-di(o-hydroxyphenylacetate) and subsequently switched to nutrient solution withholding Fe for 8 days to induce Fe stress. The roots of Fe-stressed plants reduced chelated Fe at rates sevenfold higher than roots of plants grown under Fe-sufficient conditions. The response in intact Fe-deficient roots was localized to root hairs, which developed on secondary roots during the period of Fe stress. Plasma membranes (PM) isolated by aqueous two-phase partitioning from tomato roots grown under Fe stress exhibited a 94% increase in rates of NADH-dependent Fe³⁺-citrate reduction compared to PM isolated from roots of Fe-sufficient plants. Optimal detection of the reductase activity required the presence of detergent indicating structural latency. In contrast, NADPH-dependent Fe³⁺-citrate reduction was not significantly different in root PM isolated from Fe-deficient versus Fe-sufficient plants and proceeded at substantially lower rates than NADH-dependent reduction. Mg²⁺-ATPase activity was increased 22% in PM from roots of Fe-deficient plants compared to PM isolated from roots of Fe-sufficient plants. The results localized the increase in Fe reductase activity in roots grown under Fe stress to the PM.     

Enhancement of ferric-mugineic acid uptake by iron deficient barley roots in the presence of excess free mugineic acid in the medium

To investigate the mechanism of mugineic acid-Fe^III uptake by barley roots, plasma membrane fractions were isolated from Fe-deficient barley roots using an aqueous two-phase partition method. Utilizing the plasma membrane vesicles, we developed an assay system for studying mugineic acid-⁵⁵Fe^III binding to the plasma membrane. However, no efficient active transport of mugineic acid-⁵⁵Fe^III into the plasma membrane vesicle was detected, because of large amount of non-specific adsorption of ⁵⁵Fe^III onto the vesicle. And the adsorption could be decreased by adding excess amount of free mugineic acid to the assay system. From the results it is speculated that an excess of free mugineic acids is necessary in the medium for effective uptake of mugineic acid-Fe^III by Fe-deficient barley roots. Support for this speculation came from a multi-compartment transport box experiment with excised roots of Fe-deficient barley.Abbreviations CCCP carbonylcyanide-m-chlorophenylhydrazone - MA mugineic acid     

Interrelationships between trans-Plasma Membrane Electron/Proton Transfer Stoichiometry, Organic Acid Metabolism, and Nitrate Reduction in Dwarf Bean (Phaseolus vulgaris)

Iron deficiency in dwarf bean (Phaseolus vulgaris L.) induces an increased activity of a system in the rhizodermal cells, which reduces extracellular ferric salts, and an active proton efflux from the roots, which is coupled to accumulation of citrate and malate in the roots and subsequent export of these compounds in the xylem. During reduction of extracellular ferricyanide by Fe-deficient plants, the stoichiometry of electron transport to proton efflux is 2e⁻/1H⁺, and citrate and malate levels in the roots are strongly decreased. Reduction of ferricyanide by Fe-sufficient plants has no influence on root and shoot levels of citrate and malate, but in such plants the process is characterized by a e⁻/H⁺ efflux stoichiometry close to unity. Apparently, organic acid metabolism and transport are closely associated with the e⁻/H⁺ efflux ratio. To assess the significance of organic acid metabolism as one of the direct intracellular components of the induced unbalanced e⁻/H⁺ efflux by roots, we studied NO₃⁻ reduction in shoots and roots of Fe-deficient and Fe-sufficient plants. Nitrate reductase activity in the roots was positively correlated with the level of citrate and malate, whereas the enzyme activity in the leaves responded positively to the import of these organic acid anions.     

Characterization of mugineic-acid-Fe transporter in Fe-deficient barley roots using the multi-compartment transport box method

Summary We have investigated the mugineicacid-Fe transport activity of Fe-deficient barley roots, using the multi-compartment transport box system. The roots maintained Fe transport activity for 20 h after excision. The following results were obtained. (1) In Fe-deficient roots, mugineic acid addition enhanced the transport of Fe by 32.2 times over that of the control (with FeC1₃ addition). (2) The mugineic-acid-⁵⁵Fe transport activity of Fe-deficient roots was 18.4-fold higher than that of the Fe-sufficient roots. (3) The mugineic-acid-⁵⁵Fe transport activity was decreased (7.13% based on the control) by treatment with 5 M carbonylcyanidem-chlorophenyl hydrazone (CCCP). Pretreatment with 0.1 mM dicyclohexyl carbodiimide (DCCD) lowered the transport activity (10.7% based on the control) and 1 mMN-ethylmaleimide (NEM) pretreatment reduced the transport activity to a value equivalent to 2.41% of that in the control. It is concluded that mugineicacid-Fe transporter is induced in its activity and/or amount by Fe-deficiency treatment and has an SH residue at its active site, and that the transporter needs the proton motive force produced by ATPase. We detected three polypeptides (14, 28 and 40 kDa) in the root plasma membrane that were induced under Fe-deficiency treatment.Abbreviations p-APMSF (p-amidinophenyl)methanesulfonyl fluoride hydrochloride - CCCP carbonylcyanide m-chlorophenylhydrazone - DCCD dicyclohexylcabodiimide - DMSO dimethyl sulfoxide - MA mugineic acid - NEM N-ethylmaleimide     

Formate Dehydrogenase, an Enzyme of Anaerobic Metabolism, Is Induced by Iron Deficiency in Barley Roots

To identify the proteins induced by Fe deficiency, we have compared the proteins of Fe-sufficient and Fe-deficient barley (Hordeum vulgare L.) roots by two-dimensional polyacrylamide gel electrophoresis. Peptide sequence analysis of induced proteins revealed that formate dehydrogenase (FDH), adenine phosphoribosyltransferase, and the Ids3 gene product (for Fe deficiency-specific) increased in Fe-deficient roots. FDH enzyme activity was detected in Fe-deficient roots but not in Fe-sufficient roots. A cDNA encoding FDH (Fdh) was cloned and sequenced. Fdh expression was induced by Fe deficiency. Fdh was also expressed under anaerobic stress and its expression was more rapid than that induced by Fe deficiency. Thus, the expression of Fdh observed in Fe-deficient barley roots appeared to be a secondary effect caused by oxygen deficiency in Fe-deficient plants.     

Cloning two genes for nicotianamine aminotransferase, a critical enzyme in iron acquisition (Strategy II) in graminaceous plants

Nicotianamine aminotransferase (NAAT), the key enzyme involved in the biosynthesis of mugineic acid family phytosiderophores (MAs), catalyzes the amino transfer of nicotianamine (NA). MAs are found only in graminaceous plants, although NA has been detected in every plant so far investigated. Therefore, this amino transfer reaction is the first step in the unique biosynthesis of MAs that has evolved in graminaceous plants. NAAT activity is dramatically induced by Fe deficiency and suppressed by Fe resupply. Based on the protein sequence of NAAT purified from Fe-deficient barley (Hordeum vulgare) roots, two distinct cDNA clones encoding NAAT, naat-A and naat-B, were identified. Their deduced amino acid sequences were homologous to several aminotransferases, and shared consensus sequences for the pyridoxal phosphate-binding site lysine residue and its surrounding residues. The expression of both naat-A and naat-B is increased in Fe-deficient barley roots, while naat-B has a low level of constitutive expression in Fe-sufficient barley roots. No detectable mRNA from either naat-A or naat-B was present in the leaves of either Fe-deficient or Fe-sufficient barley. One genomic clone with a tandem array of naat-B and naat-A in this order was identified. naat-B and naat-A each have six introns at the same locations. The isolation of NAAT genes will pave the way to understanding the mechanism of the response to Fe in graminaceous plants, and may lead to the development of cultivars tolerant to Fe deficiency that can grow in calcareous soils.     

Use of positron-emitting tracer imaging system for measuring the effect of salinity on temporal and spatial distribution of C tracer and coupling between source and sink organs

Salinity stress affects photosynthate partitioning between sources and sinks of plants, but how it affects these systems is less well understood. Because sources and sinks are closely tied, any adverse effect under suboptimal conditions on one of these is often misinterpreted for an effect on the other. Carbon partitioning is indispensable for stress resistance and good plant growth. In the present study, carbon partitioning in tomato plants (Lycopersicon esculentum L. cv. Momotarou) in a saline (NaCl) environment was studied by feeding radioactive ¹¹C and stable ¹³C isotopes. Pulse-chases were conducted to measure the spatial and temporal distribution of ¹³C. ¹³C was measured by a standard conventional technique, but ¹¹C distribution was monitored using a positron-emitting tracer imaging system (PETIS). Salt stress resulted in reduced carbon translocation toward roots. The majority of the photosynthate accumulated in the leaf. We also observed that the reduction in translocation of carbon occurred well before the salt stress symptoms of reduced photosynthesis and reduced plant growth in salt-exposed plants. The effect on sink activity was also shown by a decrease in stem diameter. In addition, PETIS analysis of ¹¹C translocation indicated that carbon translocation to roots was inhibited under salt conditions without a direct effect on leaf Na accumulation or osmotic stress. These results suggest that NaCl has direct effects on plants, inhibiting carbon partitioning within a few hours of salt exposure without inhibition of source activity.     

Identification and localisation of the rice nicotianamine aminotransferase gene OsNAAT1 expression suggests the site of phytosiderophore synthesis in rice

Rice plants (Oryza sativa L.) take up iron using iron-chelating compounds known as mugineic acid family phytosiderophores (MAs). In the biosynthetic pathway of MAs, nicotianamine aminotransferase (NAAT) catalyses the key step from nicotianamine to the 3′′-keto form. In the present study, we identified six rice NAAT genes (OsNAAT1–6) by screening a cDNA library made from Fe-deficient rice roots and by searching databases. Among the NAAT homologues, OsNAAT1 belongs to a subgroup containing barley functional NAAT (HvNAAT-A and HvNAAT-B) as well as a maize homologue cloned by cDNA library screening (ZmNAAT1). Northern blot and RT-PCR analysis showed that OsNAAT1, but not OsNAAT2–6, was strongly up-regulated by Fe deficiency, both in roots and shoots. The OsNAAT1 protein had NAAT enzyme activity in vitro, confirming that the OsNAAT1 gene encodes functional NAAT. Promoter–GUS analysis revealed that OsNAAT1 was expressed in companion and pericycle cells adjacent to the protoxylem of Fe-sufficient roots. In addition, expression was induced in all cells of Fe-deficient roots, with particularly strong GUS activity evident in the companion and pericycle cells. OsNAAT1 expression was also observed in the companion cells of Fe-sufficient shoots, and was clearly induced in all the cells of Fe-deficient leaves. These expression patterns highly resemble those of OsNAS1, OsNAS2 and OsDMAS1, the genes responsible for MAs biosynthesis for Fe acquisition. These findings strongly suggest that rice synthesises MAs in whole Fe-deficient roots to acquire Fe from the rhizosphere, and also in phloem cells to maintain metal homeostasis facilitated by MAs-mediated long-distance transport.     

Translocation of nitrogen in a vegetative wheat plant (Triticum aestivum)

The translocation of nitrogen was studied in vegetative wheat plants ( Triticum aestivum L. cv. SUN 9E) grown with a limited supply of nitrogen. The concentration of nitrogen in xylem sap exuding from the excised roots was the same as the nitrogen concentration in the transpiration stream. Translocation of nitrogen to the shoot was, therefore, calculated as the product of the transpiration rate and the concentration of nitrogen in xylem exudates. On the 22nd day from sowing more nitrogen was translo-cated to the shoot than it incorporated, and 56% of the nitrogen translocated to the shoot was retranslocated to the roots. The nitrogen retranslocated to the roots was more than adequate to supply the requirements of the roots for growth, and the balance of the retranslocated nitrogen was reloaded into the xylem stream. Expressed as a proportion of the total increment of nitrogen in the plant on day 22, between 79 and 100% of the nitrogen absorbed by the plant was "cycled' in the plant (root → shoot → root → shoot). It is suggested that the size of this mobile reserve of nitrogen may vary depending on the growth requirement of the plant, its nitrogen-uptake capacity and the contribution of nitrogen from mobilisation of leaf protein during senescence.     

Tillers Do Not Influence Phytotoxicity of Imazamethabenz in Wild Oat (Avena fatua)

The response of wild oat to imazamethabenz varies with the growth stage, but the role of tillers in this regard is unclear. Removal of tillers at the three-leaf stage before spraying with imazamethabenz did not significantly affect the total shoot fresh weight measured 3 weeks later. The leaf area and dry weight of intact plants at the three-leaf stage were 17–21% greater than for plants with coleoptilar and first leaf main shoot tillers (T0 and T1) removed. The greater leaf area may have increased herbicide interception per plant. Similar fresh weight reductions in main shoot, total tillers, and total shoots were found whether imazamethabenz was applied to the plant at the two-leaf without tillers or the three-leaf with two tillers stage. Imazamethabenz applied only to the main shoot reduced total shoot dry weight more than an equivalent amount of imazamethabenz applied only to tiller T1 or applied over the whole shoot. Imazamethabenz had the least inhibitory effect on whole plant growth when applied only to T1. When ¹⁴C-herbicide was applied to the first main shoot leaf of plants at the three-leaf stage with two tillers, the ¹⁴C translocated 38% to roots, 33% to the main shoot, and nearly 30% to all tillers. When ¹⁴C-herbicide was applied to the first leaf of T1 then the ¹⁴C translocated 50% to T1, 25% to the main shoot, 20% to roots, and 5% to all other tillers. The translocation pattern and fresh weight values suggested that the presence of early tillers during herbicide application neither increased nor decreased imazamethabenz efficacy in wild oat. Received June 4, 1997; accepted June 5, 1997     

Effect of iron plaque outside roots on nutrient uptake by rice (Oryza sativa L.): Phosphorus uptake

Under anaerobic conditions, ferric hydroxide deposits on the surface of rice roots have been shown to affect the uptake of some nutrients. In the present experiment, different amount of this iron plaque were induced on the roots of rice (Oryza sativa L. cv. TZ88-145) by supplying different Fe(OH)₃ concentrations in nutrient solutions, and the effect of the iron plaque on phosphorus uptake was investigated. Results showed that 1) iron plaque adsorbed phosphorus from the growth medium, and that the amount of phosphorus adsorbed by the plaque was correlated with the amount of plaque; 2) the phosphorus concentration in the shoot increased by up to 72% after 72 h at concentration of Fe(OH)₃ in the nutrient solution from 0 to 30 mg Fe/L, corresponding with amounts of iron plaque from 0.2 to 24.5 mg g^-1 (root d. wt); 3) the phosphorus concentration in the shoots of rice with iron plaque was higher than that without iron plaque though the concentration in the shoot decreased when Fe(OH)₃ was added at 50 mg Fe/L producing 28.3 mg g^-1 (root d. wt) of plaque; and 4) the phosphorus concentrations in Fe-deficient and Fe-sufficient rice plants with iron plaque were the same, although phytosiderophores were released from the Fe-deficient roots. The phytosiderophores evidently did not mobilise phosphorus adsorbed on plaque. The results suggest that iron plaque on rice plant roots might be considered a phosphorus reservoir. This revised version was published online in June 2006 with corrections to the Cover Date.     

The absorption, translocation and distribution of the herbicide glyphosate in maize expressing the CP-4 transgene

Maize (Zea mays L. var. Bonnie) transformed with a gene encoding a 5-enolpyruvylshikimate 3-phosphate synthase with altered sensitivity showed over 100-fold greater resistance to the herbicide glyphosate (N-[phosphonomethyl]glycine) in comparison with its non-transformed progenitor (parental control) at the third-leaf stage. Studies with [¹⁴C]-glyphosate at a dosage lethal to the parental control, but sublethal to the transgenic, revealed that a maximum of 45-65% of the applied dose was absorbed, with greater absorption occurring in transgenic plants. Translocation of glyphosate was closely related to its absorption (r value 0.956) with approximately 15% more of the applied dose being mobilized in transgenic plants than the parental controls. Analysis of electronic autoradiograms along the treated leaf lamina found discrete internal regions of glyphosate accumulation closely associated with the site of application. These regions contained lower amounts of glyphosate present in the treated leaf lamina was almost completely translocated in transgenic plants, while in the parental controls more remained and the leaf became necrotic. In both types of maize there was a small accumulation of herbicide in the tip region of the leaf which was not mobilized. Younger shoot tissues and roots were major sinks for translocated glyphosate accumulating approximately 25-40% of the applied dose depending upon treatment. In the parental control, equal amounts of glyphosate were found distributed between young shoot tissues and roots; while in transgenic plants, the young shoot tissue accumulated around three times more glyphosate than the roots. In both plant types, glyphosate was localized in the meristems and young, actively growing leaves. Specific glyphosate activity (the amount of glyphosate per unit dry weight of tissue) in the major sinks of the transgenic declined towards the end of the treatment period but remained relatively constant in the parental control. In conclusion, enhancing glyphosate resistance by genetic transformation influenced the absorption, translocation and distribution of this herbicide in whole plants.Keywords: Zea mays, glyphosate (N-[phosphonomethyl]-glycine), transgenic, absorption, translocation, source-sink.      

Effect of primary leaves on 59Fe uptake by roots and 59Fe distribution in the shoot of iron sufficient and iron deficient bean (Phaseolus vulgaris L.) plants

A study has been made on the effect of primary leaves on iron (Fe) distribution in the shoot. Bean (Phaseolus vulgaris L.) seedlings were precultured in nutrient solution with 8×10^-5 M FeEDTA for 4 days, and then grown further with either 8×10^-5 M FeEDTA (+Fe) or without Fe supply (-Fe) for another 5 days. Thereafter, both +Fe and -Fe plants were treated in three different ways: undisturbed; one primary leaf removed; or one primary leaf shaded, starting two hours before supply ⁵⁹FeEDTA to the roots. The +Fe plants were supplied with 8×10^-5 M ⁵⁹FeEDTA, and the -Fe plants with only 1×10^-6 M ⁵⁹FeEDTA. After 1 to 8 hour uptake periods, plants were harvested and ⁵⁹Fe in different organs was determined. Removal or shading of one primary leaf did not affect ⁵⁹Fe uptake by roots and ⁵⁹Fe translocation to the shoot in +Fe plants. In the -Fe plants, however, removal of one primary leaf decreased ⁵⁹Fe uptake by roots, whereas shading of one primary leaf had no effect on ⁵⁹Fe uptake but slightly enhanced ⁵⁹Fe translocation from roots to the shoot. The quantity of ⁵⁹Fe in primary leaves was positively correlated with quantity of ⁵⁹Fe in the stem in the -Fepplants, but not in the +Fe plants. In both, the +Fe and -Fe plants, the quantity of ⁵⁹Fe in the shoot apex was positively correlated with ⁵⁹Fe in primary leaves. The results suggest that irrespective of the Fe nutritional status of plants, the source of Fe for the shoot apex is Fe retranslocated from primary leaves.     

Increases in phosphoenolpyruvate carboxylase activity in iron-deficient sugar beet roots: Analysis of spatial localization and post-translational modification

Root tips of Fe-deficient and Fe-sufficient sugar beet plants grown in hydroponics have been used to study the changes in the amount and activity of the cytosolic enzyme phosphoenolpyruvate carboxylase (PEPC, EC 4.1.1.31). Phosphoenolpyruvate carboxylase activity in extracts of the yellow Fe-deficient root tips was, at pH 7.3, 30-fold higher (when expressed on a FW basis) and 7.1-fold higher (when expressed on a protein basis) than that found in the extracts of Fe-sufficient root tips. The amount of phosphoenolpyruvate carboxylase protein determined by immuno-blotting was, on a protein basis, 35-fold larger in the yellow zone of Fe-deficient root tips than in the Fe-sufficient root tips. The inhibition of the phosphoenolpyruvate carboxylase activity by 500 m malate was 41 and 58% in the extracts Fe-deficient and Fe-sufficient roots. The possibility that post-translational regulation of phosphoenolpyruvate carboxylase may occur mediated through phosphorylation, was studied by immunological detection of phosphoserine residues in root tip extracts.     

An Exogenous Source of Nitric Oxide Modulates Iron Nutritional Status in Peanut Seedlings (<Emphasis Type="Italic">Arachis hypogaea</Emphasis> L.)

Iron (Fe) deficiency chlorosis is a common and severe nutritional deficiency in plants, and nitric oxide (NO) is an important signaling molecule in regulating Fe homeostasis in plants. We studied the effect of sodium nitroprusside (SNP, an NO donor) on Fe uptake, translocation, storage, and activation in a greenhouse. The concentrations of active Fe, total Fe, and the ratio of active Fe to total Fe, the activities of key enzymes, and chlorophyll concentration were determined, and resistance to oxidative stress and mineral element distribution in peanut plants grown in Fe sufficiency and Fe deficiency (an absence of Fe and low level of Fe concentration) conditions were also investigated. The results showed that NO significantly increased the concentration of active Fe and the ratio of active Fe to total Fe in Fe-deficient plants, and increased active Fe concentration in leaves and stems of Fe-sufficient plants. NO application also increased Fe translocation from roots to the shoots and the accumulation of Fe in cell organelles and the soluble fraction in leaves, especially in the low-level Fe concentration condition, thus increased available Fe and chlorophyll concentration in leaves of Fe-deficient plants. The activities of key enzymes were regulated by NO, which effectively mitigated oxidative damages by enhancing the activities of antioxidant enzymes (SOD, POD, CAT), increasing H⁺-ATPase and Ca²⁺-ATPase activities to balance the ion (Fe, Ca, Mg and Zn) uptake and distribution in Fe-deficient plants. However, NO application had no obvious effect on these variables in Fe-sufficient plants. These results indicated that NO application can improve Fe uptake, translocation, and activation of related enzymes in Fe-deficient plants, thus mitigating the adverse effect of Fe deficiency.     

Deoxymugineic acid increases Zn translocation in Zn-deficient rice plants

Deoxymugineic acid (DMA) is a member of the mugineic acid family phytosiderophores (MAs), which are natural metal chelators produced by graminaceous plants. Rice secretes DMA in response to Fe deficiency to take up Fe in the form of Fe(III)–MAs complex. In contrast with barley, the roots of which secrete MAs in response to Zn deficiency, the amount of DMA secreted by rice roots was slightly decreased under conditions of low Zn supply. There was a concomitant increase in endogenous DMA in rice shoots, suggesting that DMA plays a role in the translocation of Zn within Zn-deficient rice plants. The expression of OsNAS1 and OsNAS2 was not increased in Zn-deficient roots but that of OsNAS3 was increased in Zn-deficient roots and shoots. The expression of OsNAAT1 was also increased in Zn-deficient roots and dramatically increased in shoots; correspondingly, HPLC analysis was unable to detect nicotianamine in Zn-deficient shoots. The expression of OsDMAS1 was increased in Zn-deficient shoots. Analyses using the positron-emitting tracer imaging system (PETIS) showed that Zn-deficient rice roots absorbed less ⁶²Zn-DMA than ⁶²Zn²⁺. Importantly, supply of ⁶²Zn-DMA rather than ⁶²Zn²⁺ increased the translocation of ⁶²Zn into the leaves of Zn-deficient plants. This was especially evident in the discrimination center (DC). These results suggest that DMA in Zn-deficient rice plants has an important role in the distribution of Zn within the plant rather than in the absorption of Zn from the soil. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users. Motofumi Suzuki and Takashi Tsukamoto equally contributed to this work.     

Distribution of assimilate during the flush cycle of growth in Theobroma cacao L.

The growth of the shoot and roots of seedling plants of cocoa (Theobroma cacao L.) under constant glasshouse conditions showed a rhythmic cycle, with the maximum growth stages of each alternating in a regular sequence. When the growth cycle of the shoot was upset by removing all new leaves immediately after unfolding, the roots showed a high constant growth rate during this period, suggesting that normally the rapidly expanding leaves exert an inhibitory influence on the roots. Conversely removal of portions of the root delayed the production of new leaves in the shoot. The level of soluble and starch carbohydrate in the mature leaves, roots and stem declined during the period of expansion of the flush leaves, but accumulated again at the end of the leaf expansion stage. It is likely that this reserve carbohydrate was remobilised and translocated to the flush leaves during their period of expansion. A large proportion of newly formed photoassimilate, as shown by the distribution of ¹⁴C radioactivity from different source leaves, was also translocated to the young leaves during expansion. The large sink created by these leaves may cause photoassimilate and reserve carbohydrate to be diverted from the roots, thereby inhibiting root growth during the stage of leaf expansion. It is suggested that the rhythmic leaf production at the apex may control the growth cycle of the roots.     

Induced activity of adenine phosphoribosyltransferase (APRT) in iron-deficiency barley roots: a possible role for phytosiderophore production

To isolate the genes involved in the response of graminaceous plants to Fe-deficient stress, a protein induced by Fe-deficiency treatment was isolated from barley (Hordeum vulgare L.) roots. Based on the partial amino acid sequence of this protein, a cDNA (HvAPT1) encoding adenine phosphoribosyltransferase (APRT: EC 2.4.2.7) was cloned from a cDNA library prepared from Fe-deficient barley roots. Southern analysis suggested that there were at least two genes encoding APRT in barley. Fe deficiency increased HvAPT1 expression in barley roots and resupplying Fe to the Fe-deficient plants rapidly negated the increase in HvAPT1 mRNA. Analysis of localization of HvAPT1-sGFP fusion proteins in tobacco BY-2 cells indicated that the protein from HvAPT1 was localized in the cytoplasm of cells. Consistent with the results of Northern analysis, the enzymatic activity of APRT in barley roots was remarkably increased by Fe deficiency. This induction of APRT activity by Fe deficiency was also observed in roots of other graminaceous plants such as rye, maize, and rice. In contrast, the induction was not observed to occur in the roots of a non-graminaceous plant, tobacco. Graminaceous plants generally synthesize the mugineic acid family phytosiderophores (MAs) in roots under Fe-deficient conditions. In this paper, a possible role of HvAPT1 in the biosynthesis of MAs related to adenine salvage in the methionine cycle is discussed.