Miroslav Nikolic

Uptake of 59Fe from soluble 59Fe-humate complexes by cucumber and barley plants

The capability of cucumber (Cucumis sativus L., cv. Serpente cinese), a Strategy I plant and barley (Hordeum vulgaris L., cv. Europa), a Strategy II plant to use Fe complexed by a water-soluble humic fraction (WEHS) extracted from a peat, was studied. Uptake of 59Fe from 59Fe-WEHS by cucumber plants was higher at pH 6.0 than at pH 7.5. Roots of intact cucumber plants were able to reduce the FeIII-WEHS complex either at pH 6.0 or 7.5, rates being higher in the assay medium buffered at pH 6.0. After supply of 59Fe-WEHS, a large pool of root extraplasmatic 59Fe was formed, which could be used to a large extent by Fe-deficient plants, particularly under acidic conditions. Uptake of 59Fe from 59Fe-WEHS by Fe-sufficient and Fe-deficient barley plants was examined during periods of high (morning) and low (evening) PS release. Uptake paralleled the diurnal rhythm of PS release. Furthermore, 59Fe uptake was strongly enhanced by addition of PS to the uptake solution in both Fe-sufficient and Fe-deficient plants. High amount of root extraplasmatic 59Fe was formed upon supply of Fe-WEHS, particularly in the evening experiment. Fe-deficient barley plants were able to utilize Fe from the root extraplasmatic pool, conceivably as a result of high rates of PS release. The results of the present work together with previous observations indicate that cucumber plants (Strategy I) utilize Fe complexed to WEHS, presumably via reduction of FeIII-WEHS by the plasma membrane-bound reductase, while barley plants (Strategy II) use an indirect mechanism involving ligand exchange between WEHS and PS.

Mechanism of Fe uptake by the leaf symplast : Is Fe inactivation in leaf a cause of Fe deficiency chlorosis?

The mechanism of iron (Fe) uptake from the leaf apoplast into leaf mesophyll cells was studied to evaluate the putative Fe inactivation as a possible cause of Fe deficiency chlorosis. For this purpose, sunflower (Helianthus annuus L.) and faba bean plants (Vicia faba L.) were precultured with varied Fe and bicarbonate (HCO3-) supply in nutrient solution. After 2–3 weeks preculture, FeIII reduction and 59Fe uptake by leaf discs were measured in solutions with Fe supplied as citrate or synthetic chelates in darkness. The data clearly indicate that FeIII reduction is a prerequisite for Fe uptake into leaf cells and that the Fe nutritional status of plants does not affect either process. In addition, varied supply of Fe and HCO3- to the root medium during preculture had no effect on pH of the xylem sap and leaf apoplastic fluid. A varied pH of the incubation solution had no significant effect on FeIII reduction and Fe uptake by leaf discs in the physiologically relevant pH range of 5.0–6.0 as measured in the apoplastic leaf fluid. It is concluded that Fe inactivation in the leaf apoplast is not a primary cause of Fe deficiency chlorosis induced by bicarbonate.

Silicon alleviates iron deficiency in cucumber by promoting mobilization of iron in the root apoplast

· Root responses to lack of iron (Fe) have mainly been studied in nutrient solution experiments devoid of silicon (Si). Here we investigated how Si ameliorates Fe deficiency in cucumber (Cucumis sativus) with focus on the storage and utilization of Fe in the root apoplast. · A combined approach was performed including analyses of apoplastic Fe, reduction-based Fe acquisition and Fe-mobilizing compounds in roots along with the expression of related genes. · Si-treated plants accumulated higher concentrations of root apoplastic Fe, which rapidly decreased when Fe was withheld from the nutrient solution. Under Fe-deficient conditions, Si also increased the accumulation of Fe-mobilizing compounds in roots. Si supply stimulated root activity of Fe acquisition at the early stage of Fe deficiency stress through regulation of gene expression levels of proteins involved in Fe acquisition. However, when the period of Fe deprivation was extended, these reactions further decreased as a consequence of Si-induced enhancement of the Fe status of the plants. · This work provides new evidence for the beneficial role of Si in plant nutrition and clearly indicates that Si-mediated alleviation of Fe deficiency includes an increase of the apoplastic Fe pool in roots and an enhancement of Fe acquisition.

Silicon ameliorates manganese toxicity in cucumber by decreasing hydroxyl radical accumulation in the leaf apoplast

This work was focused on the role of silicon (Si) in amelioration of manganese (Mn) toxicity caused by elevated production of hydroxyl radicals (·OH) in the leaf apoplast of cucumber (Cucumis sativus L.). The plants were grown in nutrient solutions with adequate (0.5 μM) or excessive (100 μM) Mn concentrations with or without Si being supplied. The symptoms of Mn toxicity were absent in the leaves of Si-treated plants subjected to excess Mn, although the leaf Mn concentration remained extremely high. The apoplastic concentration of free Mn(2+) and H(2)O(2) of high Mn-treated plants was significantly decreased by Si treatment. Si supply suppressed the Mn-induced increased abundance of peroxidase (POD) isoforms in the leaf apoplastic fluid, and led to a rapid suppression of guaiacol-POD activity under excess Mn. The spin-trapping reagent 5-(diethoxyphosphoryl)-5-methyl-1-pyrroline-N-oxide was used to detect ·OH by electron paramagnetic resonance spectroscopy. Although supplying Si markedly decreased the accumulation of ·OH in the leaf apoplast with excess Mn, adding monosilicic acid to the Mn(2+)/H(2)O(2) reaction mixture did not directly affect the Fenton reaction in vitro. The results indicate that Si contributes indirectly to a decrease in ·OH in the leaf apoplast by decreasing the free apoplastic Mn(2+), thus regulating the Fenton reaction. A direct inhibitory effect of Si on guaiacol-POD activity (demonstrated in vitro) may also contribute to decreasing the POD-mediated generation of ·OH.

Germanium-68 as an Adequate Tracer for Silicon Transport in Plants. Characterization of Silicon Uptake in Different Crop Species

A basic problem in silicon (Si) uptake studies in biology is the lack of an appropriate radioactive isotope. Radioactive germanium-68 (68Ge) has been used previously as a Si tracer in biological materials, but its suitability for the study of Si transport in higher plants is still untested. In this study, we investigated 68Ge-traced Si uptake by four crop species differing widely in uptake capacity for Si, including rice (Oryza sativa), barley (Hordeum vulgare), cucumber (Cucumis sativus), and tomato (Lycopersicon esculentum). Maintenance of a 68Ge:Si molar ratio that was similar in the plant tissues of all four plant species to that supplied in the nutrient solution over a wide range of Si concentrations demonstrated the absence of discrimination between 68Ge and Si. Further, using the 68Ge tracer, a typical Michaelis-Menten uptake kinetics for Si was found in rice, barley, and cucumber. Compared to rice, the relative proportion of root-to-shoot translocated Si was lower in barley and cucumber and especially in tomato (only 30%). Uptake and translocation of Si in rice, barley, and cucumber (Si accumulators) were strongly inhibited by 2,4-dinitrophenol and HgCl2, but in tomato, as a Si-excluding species, both inhibitors produced the opposite effect. In conclusion, our results suggest the use of the 68Ge tracer method as an appropriate choice for future studies of Si transport in plants. Our findings also indicate that the restriction of Si from symplast to apoplast in the cortex of Si excluders is a metabolically active process.

Uptake and transport of foliar applied zinc (65Zn) in bread and durum wheat cultivars differing in zinc efficiency

Using two bread wheat (Triticum aestivum) and two durum wheat (Triticum durum) cultivars differing in zinc (Zn) efficiency, uptake and translocation of foliar-applied 65Zn were studied to characterize the role of Zn nutritional status of plants on the extent of phloem mobility of Zn and to determine the relationship between phloem mobility of Zn and Zn efficiency of the used wheat cultivars. Irrespective of leaf age and Zn nutritional status of plants, all cultivars showed similar Zn uptake rates with application of 65ZnSO4 to leaf strips in a short-term experiment. Also with supply of 65ZnSO4 by immersing the tip (3 cm) of the oldest leaf of intact plants, no differences in Zn uptake were observed among and within both wheat species. Further, Zn nutritional status did not affect total uptake of foliar applied Zn. However, Zn-deficient plants translocated more 65Zn from the treated leaf to the roots and remainder parts of shoots. In Zn-deficient plants about 40% of the total absorbed 65Zn was translocated from the treated leaf to the roots and remainder parts of shoots within 8 days while in Zn-sufficient plants the proportion of the translocated 65Zn of the total absorbed 65Zn was about 25%. Although differences in Zn efficiency existed between the cultivars did not affect the translocation and distribution of 65Zn between roots and shoots. Bread wheats compared to durum wheats, tended to accumulate more 65Zn in shoots and less 65Zn in roots, particularly under Zn-deficient conditions. The results indicate that differences in expression of Zn efficiency between and within durum and bread wheats are not related to translocation or distribution of foliar-applied 65Zn within plants. Differential compartementation of Zn at the cellular levels is discussed as a possible factor determining genotypic variation in Zn efficiency within wheat.

Effect of bicarbonate and Fe supply on Fe nutrition of grapevine.

Abstract Grapevine grafts (Vitis vinifera L. cv. Riesling, on rootstocks of Vitis sp. L. cv. 5BB) were grown hydroponically in complete nutrient solution (control), Fe‐free nutrient solution and complete nutrient solution with added 10 mM HCO3 −. The concentration of total chlorophyll was significantly reduced in Fe deficient plants, particularly in HCO3 − supplied nutrient solution. The concentration of extracellular (extraplasmatic) root Fe decreased in the case of ‐Fe treatment and increased in that of HCO3 − treatment, while the concentration of symplastic Fe decreased in both treatments. However, the concentration of total root Fe decreased in roots of plants grown in ‐Fe solution and increased in those grown in HCO3 − supplied solution. Fe‐deficient plants showed lowering of total Fe and “active Fe” (extractable in 1 M HCl or o‐phenanthroline) concentration in leaves. These results indicate that bicarbonate in the nutrient solution may be a major inducing factor of Fe‐deficiency chlorosis in grapevine presumably due to inhibited Fe acquisition by roots, but do not confirm Fe inactivation in leaves.

Nitrate does not result in iron inactivation in the apoplast of sunflower leaves.

It has been hypothesized that nitrate (NO3–) nutrition might induce iron (Fe) deficiency chlorosis by inactivation of Fe in the leaf apoplast (H.U. Kosegarten, B. Hoffmann, K. Mengel [1999] Plant Physiol 121: 1069–1079). To test this hypothesis, sunflower (Helianthus annuus L. cv Farnkasol) plants were grown in nutrient solutions supplied with various nitrogen (N) forms (NO3–, NH4+ and NH4NO3), with or without pH control by using pH buffers [2-(N-morpholino)ethanesulfonic acid or 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid]. It was shown that high pH in the nutrient solution restricted uptake and shoot translocation of Fe independently of N form and, therefore, induced Fe deficiency chlorosis at low Fe supply [1 μm ferric ethylenediaminedi(O-hydroxyphenylacetic acid)]. Root NO3– supply (up to 40 mm) did not affect the relative distribution of Fe between leaf apoplast and symplast at constant low external pH of the root medium. Although perfusion of high pH-buffered solution (7.0) into the leaf apoplast restricted 59Fe uptake rate as compared with low apoplastic solution pH (5.0 and 6.0, respectively), loading of NO3– (6 mm) showed no effect on 59Fe uptake by the symplast of leaf cells. However, high light intensity strongly increased 59Fe uptake, independently of apoplastic pH or of the presence of NO3– in the apoplastic solution. Finally, there are no indications in the present study that NO3– supply to roots results in the postulated inactivation of Fe in the leaf apoplast. It is concluded that NO3– nutrition results in Fe deficiency chlorosis exclusively by inhibited Fe acquisition by roots due to high pH at the root surface.

Silicon in Agriculture

Although silicon (Si) is not yet listed among the essential elements for the growth of higher plants, it has been well documented to play an important role in providing benefi cial effects on growth and yield, especially in plants under stressful environments. From a practical perspective, the use of slag-based silicate fertilizers in agriculture can be dated back to the Middle Ages in Europe. Over the last decade, the discovery of specifi c Si transporters in rice roots has allowed great progress in the understanding of Si uptake by plants at the molecular level. In the same manner, important advancements have been made in dissecting the molecular mechanisms by which Si enhances plant resistance to fungal and bacterial diseases and insect pest damage. In contrast, more efforts are needed to explain at the molecular level the numerous reports showing Si benefi ts against abiotic stresses. In this chapter, a brief review is presented focusing on the most important historical points and general introduction of worldwide Si research.

Contrasting effect of silicon on iron, zinc and manganese status and accumulation of metal-mobilizing compounds in micronutrient-deficient cucumber.

Although the beneficial role of silicon (Si) in alleviation of abiotic stress is well established, little is known of the relevance of Si nutrition under microelement deficiency. The aim of our work was to investigate the physiological role of Si in relation to micronutrient (Fe, Zn and Mn) deficiencies in cucumber (Cucumis sativus L.). Cucumber (cv. Semkross) plants were grown hydroponically in a complete nutrient solution (control) and in nutrient solutions free from Fe, Zn or Mn, with or without Si supply. Plant tissue concentrations of microelements, organic acids and phenolics were measured. Si supply effectively mitigated the symptoms of Fe deficiency, but only in part, the symptoms of Zn- or Mn deficiency. Leaf Fe concentration significantly increased in plants deprived of Fe but treated with Si, whereas the concentrations of other microelements were not affected by Si supply. The effects of Si supply in increasing accumulation of both organic acids and phenolic compounds in cucumber tissues were exclusively related to Fe nutrition. Enhancement of Fe distribution towards apical shoot parts, along with the tissue accumulation of Fe-mobilizing compounds such as citrate (in leaves and roots) or cathechin (in roots) appears to be the major alleviating effect of Si. Si nutrition, however, was without effect on the mobility and tissue distribution of either Zn or Mn.