Root exudation and rhizosphere acidification by two lines of Medicago ciliaris in response to lime-induced iron deficiency

The main objective of this work was to compare the tolerance to lime-induced Fe deficiency of two lines of Medicago ciliaris (TN11.11 and TN8.7). We studied the effects of Fe deficiency on: (1) root biomass and rhizosphere acidification, (2) accumulation in the roots and the exudation into the rhizosphere of organic compounds (citric acid, malic acid and phenols), (3) changes under Fe deficient conditions in the activities of two enzymes, the first related to organic acid metabolism (malate dehydrogenase: MDH) and the other to proton extrusion (H⁺-ATPase activity). After a pre-treatment of one week, plants were transferred into hydroponic culture under three treatments: +Fe, ?Fe and +Fe +lime. Iron deficiency led to 40% increase in root biomass in TN11.11 line in the presence of lime. Both the omission of Fe and the addition of lime to the nutrient solution increased the H⁺-ATPase activity more in TN11.11 than in TN8.7. In addition, Fe deficiency increased accumulation of organic acids as well as phenols in roots, and stimulated the MDH activity more in TN11.11 than in TN8.7 (+75% and +41% in TN11.11 and TN8.7, respectively). Iron deficiency also increased the amounts of citrate, malate and phenols in root exudates. Our data allowed us to note that the TN11.11 line is more effective in overcoming Fe deficiency than TN8.7.     

Responses of two lines of <Emphasis Type="Italic">Medicago ciliaris</Emphasis> to Fe deficiency under saline conditions

The aim of this research was to study the responses of two lines of Medicago ciliaris: TN11.11 and TN8.7 to iron deficiency under saline conditions. However; the paper showed also the results of a preliminary study which report the contrastive responses of the two lines to salinity. We found that plant growth and chlorophyll content of TN11.11 line were more affected by salinity than TN8.7. The severity of symptoms was linked to the sodium accumulation in shoots as well as a limitation of potassium uptake. Our data allowed us to note that TN8.7 line is less sensitive and can better cope with the salinity. Concerning the effect of salinity on iron deficiency responses, we noted that root PM H⁺-ATPase and FCR activities were reduced when iron deficiency was associated with salinity. This probably explained the decrease of Fe uptake. On the contrary, PEPC activity was not affected.     

Metabolic responses in iron deficient tomato plants

The effects of Fe deficiency on different metabolic processes were characterized in roots, xylem sap and leaves of tomato. The total organic acid pool increased significantly with Fe deficiency in xylem sap and leaves of tomato plants, whereas it did not change in roots. However, the composition of the pool changed with Fe deficiency, with major increases in citrate concentrations in roots (20-fold), leaves (2-fold) and xylem sap (17-fold). The activity of phosphoenolpyruvate carboxylase, an enzyme leading to anaplerotic C fixation, increased 10-fold in root tip extracts with Fe deficiency, whereas no change was observed in leaf extracts. The activities of the organic acid synthesis-related enzymes malate dehydrogenase, citrate synthase, isocitrate dehydrogenase, fumarase and aconitase, as well as those of the enzymes lactate dehydrogenase and pyruvate carboxylase, increased with Fe deficiency in root extracts, whereas only citrate synthase increased significantly with Fe deficiency in leaf extracts. These results suggest that the enhanced C fixation capacity in Fe-deficient tomato roots may result in producing citrate that could be used for Fe xylem transport. Total pyridine nucleotide pools did not change significantly with Fe deficiency in roots or leaves, although NAD(P)H/NAD(P) ratios were lower in Fe-deficient roots than in controls. Rates of O(2) consumption were similar in Fe-deficient and Fe-sufficient roots, but the capacity of the alternative oxidase pathway was decreased by Fe deficiency. Also, increases in Fe reductase activity with Fe deficiency were only 2-fold higher when measured in tomato root tips. These values are significantly lower than those found in other plant species, where Fe deficiency leads to larger increases in organic acid synthesis-related enzyme activities and flavin accumulation. These data support the hypothesis that the extent of activation of different metabolic pathways, including carbon fixation via PEPC, organic acid synthesis-related enzymes and oxygen consumption is different among species, and this could modulate the different levels of efficiency in Strategy I plants.     

Physiological and biochemical responses of the iron chlorosis tolerant grapevine rootstock 140 Ruggeri to iron deficiency and bicarbonate

Background and aims

Iron (Fe) deficiency chlorosis associated with high levels of soil bicarbonate is one of the main nutritional disorders observed in sensitive grapevine genotypes. The aim of the experiment was to assess both the independent and combined effects of Fe and bicarbonate nutrition in grapevine.

Methods

Plants of the Fe chlorosis tolerant 140 Ruggeri rootstock were grown with and without Fe(III)-EDTA and bicarbonate in the nutrient solution. SPAD index, plant growth, root enzyme (PEPC, MDH, CS, NADP⁺ ?IDH) activities, kinetic properties of root PEPC, organic acid concentrations in roots and xylem sap and xylem sap pH were determined. A factorial statistical design with two factors (Fe and BIC) and two levels of each factor was adopted: +Fe and ?Fe, and +BIC and ?BIC.

Results

This rootstock strongly reacted to Fe deficiency by activating several response mechanisms at different physiological levels. The presence of bicarbonate in the nutrient solution changed the activity of PEPC and TCA related enzymes (CS, NADP⁺-IDH) and the accumulation/translocation of organic acids in roots of Fe-deprived plants. Moreover, this genotype increased root biomass and root malic acid concentration in response to high bicarbonate levels in the substrate. Bicarbonate also enhanced leaf chlorophyll content.

Conclusions

Along with a clear independent effect on Fe nutrition, our data support a modulating role of bicarbonate on Fe deficiency response mechanisms at root level.     

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.     

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.     

Induction of the Root Cell Plasma Membrane Ferric Reductase (An Exclusive Role for Fe and Cu)

Induction of ferric reductase activity in dicots and nongrass monocots is a well-recognized response to Fe deficiency. Recent evidence has shown that Cu deficiency also induces plasma membrane Fe reduction. In this study we investigated whether other nutrient deficiencies could also induce ferric reductase activity in roots of pea (Pisum sativum L. cv Sparkle) seedlings. Of the nutrient deficiencies tested (K, Mg, Ca, Mn, Zn, Fe, and Cu), only Cu and Fe deficiencies elicited a response. Cu deficiency induced an activity intermediate between Fe-deficient and control plant activities. To ascertain whether the same reductase is induced by Fe and Cu deficiency, concentration- and pH-dependent kinetics of root ferric reduction were compared in plants grown under control, -Fe, and -Cu conditions. Additionally, rhizosphere acidification, another process induced by Fe deficiency, was quantified in pea seedlings grown under the three regimes. Control, Fe-deficient, and Cu-deficient plants exhibited no major differences in pH optima or Km for the kinetics of ferric reduction. However, the Vmax for ferric reduction was dramatically influenced by plant nutrient status, increasing 16- to 38-fold under Fe deficiency and 1.5- to 4-fold under Cu deficiency, compared with that of control plants. These results are consistent with a model in which varying amounts of the same enzyme are deployed on the plasma membrane in response to plant Fe or Cu status. Rhizosphere acidification rates in the Cu-deficient plants were similarly intermediate between those of the control and Fe-deficient plants. These results suggest that Cu deficiency induces the same responses induced by Fe deficiency in peas.     

The effect of sodium bicarbonate on plant performance and iron acquisition system of FA-5 (Forner-Alcaide 5) citrus seedlings

This work studies the effect of bicarbonate on plant performance and the iron acquisition system of Forner-Alcaide 5 (FA-5) seedlings, a citrus genotype known for its tolerance to calcareous soils. Plants were irrigated for 6 weeks with or without 10 mM NaHCO₃. Treatment significantly decreased shoot growth, photosynthetic levels and iron concentration in shoots and roots. o,o-⁵⁷FeEDDHA experiments indicated that ⁵⁷Fe uptake by roots was inhibited in treated plants. Moreover, those seedlings accumulated more ⁵⁷Fe in roots, and enhanced mRNA accumulation of ferric reductase genes FRO1 and FRO2 and FC-R activity in roots. H⁺-ATPase activity and HA1 gene expression were also increased, while HA2 was not affected. In addition, expression of the iron transporter gene IRT1 was increased, while IRT2 was not significantly affected. Finally, according to PEPC enzymatic activity, PEPC1 gene expression was higher in treated roots. In conclusion, it appears that bicarbonate prevents medium acidification by roots, thus reducing Fe²⁺ uptake. Accordingly, Fe deficiency enhanced the expression of some genes related with the Fe acquisition system (IRT1, FRO1, FRO2, HA1 and PEPC1) and the activity of the corresponding enzymes, which appear to constitute an adaptive mechanism of FA-5 in these soils.     

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.     

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.     

Leaf responses to iron nutrition and low cadmium in peanut: anatomical properties in relation to gas exchange

Aims

This study evaluated how iron nutrition affect leaf anatomical and photosynthetic responses to low cadmium and its accumulation in peanut plants.

Methods

Seedlings were treated with Cd (0 and 0.2 μM CdCl₂) and Fe (0, 10, 25, 50 or 100 μM EDTA-Na₂Fe) in hydroponic culture.

Results

Cadmium accumulation is highest in Fe-deficient plants, and dramatically decreased with increasing Fe supply. The biomass, gas exchange, and reflectance indices were highest at 25 μM Fe²⁺ treatments, indicating the concentration is favorable for the growth of peanut plants. Both Fe deficiency and Cd exposure impair photosynthesis and reduce reflectance indices. However, they show different effects on leaf anatomical traits. Fe deficiency induces more and smaller stomata in the leaf surface, but does not affect the inner structure. Low Cd results in a thicker lamina with smaller stomata, thicker palisade and spongy tissues, and lower palisade to spongy thickness ratio. The stomatal length and length/width ratio in the upper epidermis, spongy tissue thickness, and palisade to spongy thickness ratio were closely correlated with net photosynthetic rate, stomatal conductance, and transpiration rate.

Conclusions

Cd accumulation rather than Fe deficiency alters leaf anatomy that may increase water use efficiency but inhibit photosynthesis.     

Inhibition of iron deficiency stress response in cucumber by rare earth elements.

We investigated the influence of the trivalent scandium (Sc), chromium (Cr), gallium (Ga), yttrium (Y) and lanthanum (La) on both the function and activity of ferric chelate reductase (FCR) in cucumber (Cucumis sativus L.) roots. Cucumber seedlings were grown for 1week in a nutrient solution without Fe or in some experiments with 10microM FeEDTA. Intact root systems were assayed for FCR activity in a medium at pH 5.0 containing 100microM FeEDTA with the ferrous chelating agent Ferrozine. Addition of 100microM concentrations of the EDTA complexes of Sc, Cr, Ga, Y and La did not inhibit FCR in Fe-deficient roots. When Fe-deficient roots were grown with 10microM LaCl(3), ScCl(3), or YCl(3) for 3days, FCR activity decreased to 23%, 15% and 1%, respectively, of the activity of Fe-deficient plants grown without trivalent metal addition. Additionally, these trivalent metals suppressed proton secretion. Growth of Fe-deficient plants with 80microM Ga(2)(SO(4))(3) decreased FCR activity to 35% of the control activity while 80microM CrEDTA did not affect FCR activity. With the addition of either FeEDTA or YCl(3), FCR activity decreased to less than 5% of the activity of the Fe-deficient control roots in 3days. Addition of FeEDTA, but not Y, resulted in recovery from Fe deficiency as indicated by increasing chlorophyll content of leaves.     

Water-extractable humic substances enhance iron deficiency responses by Fe-deficient cucumber plants

The ability of Fe-deficient cucumber plants to use iron complexed to a water-extractable humic substances fraction (WEHS), was investigated. Seven-day-old Fe-deficient plants were transferred to a nutrient solution supplemented daily for 5 days with 0.2 μM Fe as Fe-WEHS (5 μg org. C mL^-1), Fe-EDTA, Fe-citrate or FeCl₃. These treatments all allowed re-greening of the leaf tissue, and partial recovery of dry matter accumulation, chlorophyll and iron contents. However, the recovery was faster in plants supplied with Fe-WEHS and was already evident 48 h after Fe supply. The addition of 0.2 μM Fe to the nutrient solution caused also a partial recovery of the dry matter and iron accumulation in roots of Fe-deficient cucumber plants, particularly in those supplied with Fe-WEHS. The addition of WEHS alone (5 μg org. C mL^-1, 0.04 μM Fe) to the nutrient solution slightly but significantly increased iron and chlorophyll contents in leaves of Fe-deficient plants; in these plants, dry matter accumulation in leaves and roots was comparable or even higher than that measured in plants treated with Fe-citrate or FeCl₃. After addition of the different iron sources for 5 days to Fe-deficient roots, morphological modifications (proliferation of lateral roots, increase in the diameter of the sub-apical zones and amplified root-hair formation) and physiological responses (enhanced Fe(III)-chelate reductase and acidification of the nutrient solution) induced by Fe deficiency, were still evident, particularly in plants treated with the humic molecules. The presence of WEHS caused also a further acidification of the nutrient medium by Fe-deficient plants. The Fe-WEHS complex (1 μM Fe) could be reduced by intact cucumber roots, at rates of reduction higher than those measured for Fe-EDTA at equimolar iron concentration. Plasma membrane vesicles, purified by two-phase partition from root microsomes of Fe-deficient plants, were also able to reduce Fe-WEHS. Results show that Fe-deficient cucumber plants can use iron complexed to water soluble humic substances, at least in part via reduction of complexed Fe(III) by the plasma membrane Fe(III)-chelate reductase of root cells. In addition, the stimulating effect of humic substances on H⁺ release might be of relevance for the overall response of the plants to iron shortage. This revised version was published online in June 2006 with corrections to the Cover Date.     

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.     

Formation of Root Epidermal Transfer Cells in Plantago

The root ultrastructure and transmembrane electron transport activities of two Plantago species have been examined with respect to alterations in response to Fe deficiency, exogenously supplied auxin, and the presence of chromium in the external medium. Both species showed increased ferric reductase activity upon Fe starvation, but they differed in the maximum rates. The addition of chromium to the nutrient solution led to a further enhancement in Fe-ethylenediaminetetraacetate reduction by Fe-deficient plants. In roots of Plantago lanceolata, the enhanced redox activity is associated with the formation of transfer cells in the epidermis. Similar characteristics of rhizodermal cells were observed in Fe-sufficient roots 3 d after application of the auxin analog 2,4-dichlorophenoxy-acetic acid. No structural adaptations occurred in roots of Plantago maritima. A quantitative estimation of the frequencies of transfer cells in root segments of Fe-deficient plants that differ in reduction activity revealed no correlation between the two phenomena. It is concluded that the area of plasmalemma infoldings is not specialized for the enhanced reduction of extracytoplasmatic Fe in response to Fe deficiency. The role of transfer cells in the adaptation to suboptimal Fe availability and the mechanisms triggering their formation are discussed.     

Carboxylate metabolism changes induced by Fe deficiency in barley, a Strategy II plant species

The effects of iron (Fe) deficiency on carboxylate metabolism were investigated in barley (Hordeum vulgare L.) using two cultivars, Steptoe and Morex, which differ in their Fe efficiency response. In both cultivars, root extracts of plants grown in Fe-deficient conditions showed higher activities of enzymes related to organic acid metabolism, including citrate synthase, malate dehydrogenase and phosphoenolpyruvate carboxylase, compared to activities measured in root extracts of Fe-sufficient plants. Accordingly, the concentration of total carboxylates was higher in Fe-deficient roots of both cultivars, with citrate concentration showing the greatest increase. In xylem sap, the concentration of total carboxylates was also higher with Fe deficiency in both cultivars, with citrate and malate being the major organic acids. Leaf extracts of Fe-deficient plants also showed increases in citric acid concentration and in the activities of glucose-6-phosphate dehydrogenase and fumarase activities, and decreases in aconitase activity. Our results indicate that changes in root carboxylate metabolism previously reported in Strategy I species also occur in barley, a Strategy II plant species, supporting the existence of anaplerotic carbon fixation via increases in the root activities of these enzymes, with citrate playing a major role. However, these changes occur less intensively than in Strategy I plants. Activities of the anaerobic metabolism enzymes pyruvate decarboxylase and lactate dehydrogenase did not change in barley roots with Fe deficiency, in contrast to what occurs in Strategy I plants, suggesting that these changes may be Strategy I-specific. No significant differences were observed in overall carboxylate metabolism between cultivars, for plants challenged with high or low Fe treatments, suggesting that carboxylate metabolism changes are not behind the Fe-efficiency differences between these cultivars. Citrate synthase was the only measured enzyme with constitutively higher activity in Steptoe relative to Morex leaf extracts.     

Characterization of cadmium concentration and translocation among ramie cultivars as affected by zinc and iron deficiency

Zn and Fe are essential nutritional elements in plants and play important roles in various physiological processes of plants. Zn and Fe are chemically similar to cadmium (Cd); therefore, Zn and Fe may mediate Cd-induced physiological or metabolic changes in plants. In order to evaluate the interaction between Cd, Zn and Fe, we conducted a hydroponics experiment to determine the plant biomass, photosynthetic characteristics, and Cd accumulation of ten ramie cultivars under Zn/Fe-sufficient or Zn/Fe-deficient conditions in the presence of 32 µM CdCl₂. Ramie varied among cultivars in morpho-physiological response to Cd stress as well as Cd accumulation, translocation and distribution. Zn and Fe deficiency increased the concentration and amount of Cd in plant organs, but decreased TF_{stem to leaf} and TF_{root to stem}. Cultivars with more Cd in roots and shoots showed smaller increase in Cd accumulation under Zn and Fe-deficiency stress. Xiangzhu 7 and Duobeiti 1 showed a higher capacity of Cd accumulation in their shoots. Zn and Fe deficiency decreased Pn, but increased Ci, Gs, and E in most cultivars. The difference in Cd translocation among ramie cultivars was mainly ascribed to the difference in plant transpiration.     

Responses of Arabidopsis thaliana to bicarbonate-induced iron deficiency

Morpho-physiological and biochemical responses of Arabidopsis thaliana (accession N1438) to bicarbonate-induced iron deficiency were investigated. Plants were grown in cabinet under controlled conditions, in a nutrient solution containing 5 μM Fe, added or not with 10 mM NaHCO₃. After 30 days, bicarbonate-treated plants displayed significantly lower biomass, leaf number and leaf surface area as compared to control plants, and slight yellowing of their younger leaves was observed. Potassium (K⁺) content was not modified by bicarbonate treatment in roots, whereas it was significantly diminished in shoots. Their content in ferrous iron (Fe²⁺) and in leaf total chlorophylls was noticeably lower than in control plants. Root Fe(III)-chelate reductase and phosphoenolpyruvate carboxylase (PEPC) activities were significantly enhanced, but leaf ribulose 1.5-bisphosphate carboxylase (Rubisco) activity was decreased.     

Effects of short term iron citrate treatments at different pH values on roots of iron-deficient cucumber: A Mössbauer analysis

Alkaline pH values and bicarbonate greatly reduce the mobility and uptake of Fe, causing Fe deficiency chlorosis. In the present work, the effects of pH and bicarbonate on the uptake and accumulation of Fe in the roots of cucumber were studied by Mössbauer spectroscopy combined with physiological tests and diaminobenzidine enhanced Perls staining. Mössbauer spectra of Fe-deficient cucumber roots supplied with 500 μM ⁵⁷Fe(III)-citrate at different pH values showed the presence of an Fe(II) and an Fe(III) component. As the pH was increased from 4.5 to 7.5, the root ferric chelate reductase (FCR) activity decreased significantly and a structural change in the Fe(III) component was observed. While at pH 4.5 the radial intrusion of Fe reached the endodermis, at pH 7.5, Fe was found only in the outer cortical cell layers. The Mössbauer spectra of Fe-deficient plants supplied with Fe(III)-citrate in the presence of bicarbonate (pH 7.0 and 7.5) showed similar Fe components, but the relative Fe(II) concentration compared to that measured at pH values 6.5 and 7.5 was greater. The Mössbauer parameters calculated for the Fe(II) component in the presence of bicarbonate were slightly different from those of Fe(II) alone at pH 6.5–7.5, whereas the FCR activity was similarly low. Fe incorporation into the root apoplast involved only the outer cortical cell layers, as in the roots treated at pH 7.5. In Fe-sufficient plants grown with Fe(III)-citrate and 1 mM bicarbonate, Fe precipitated as granules and was in diffusely scattered grains on the root surface. The “bicarbonate effect” may involve a pH component, decreasing both the FCR activity and the acidification of the apoplast and a mineralization effect leading to the slow accumulation of extraplasmatic Fe particles, forming an Fe plaque and trapping Fe and other minerals in biologically unavailable forms.     

Interspecies compatibility of NAS1 gene promoters.

Nicotianamine and nicotianamine synthase (NAS) play key roles in iron nutrition in all higher plants. However, the mechanism underlying the regulation of NAS expression differs among plant species. Sequences homologous to iron deficiency-responsive elements (IDEs), i.e., cis-acting elements, are found on the promoters of these genes. We aimed to verify the interspecies compatibility of the Fe-deficiency response of NAS1 genes and understand the universal mechanisms that regulate their expression patterns in higher plants. Therefore, we introduced the graminaceous (Hordeum vulgare L. and Oryza sativa L.) NAS1 promoter::GUS into dicots (Nicotiana tabacum L. and Arabidopsis thaliana L.). Fe deficiency induced HvNAS1 expression in the shoots and roots when introduced into rice. HvNAS1 promoter::GUS and OsNAS1 promoter::GUS induced strong expression of GUS under Fe-deficient conditions in transformed tobacco. In contrast, these promoters only definitely functioned in Arabidopsis transformants. These results suggest that some Fe nutrition-related trans-factors are not compatible between graminaceous plants and Arabidopsis. HvNAS1 promoter::GUS induced GUS activity only in the roots of transformed tobacco under Fe-deficient conditions. On the other hand, OsNAS1 promoter::GUS induced GUS activity in both the roots and shoots of transformed tobacco under conditions of Fe deficiency. In tobacco transformants, the induction of GUS activity was induced earlier in the shoots than roots. These results suggest that the HvNAS1 and OsNAS1 promoters are compatible with Fe-acquisition-related trans-factors in the roots of tobacco and that the OsNAS1 promoter is also compatible with some shoot-specific Fe deficiency-related trans-factors in tobacco.