Search

Highlight Rice only harbors one functional gene for DMA synthesis, and the knock-down of OsDMAS1 significantly up-regulates the genes involved in Fe uptake and homeostasis., Rice (Oryza sativa) secretes 2′-deoxymugineic acid (DMA) to acquire insoluble iron (Fe) from the rhizosphere. In rice, DMA is synthesized by DMA synthase 1 (OsDMAS1), a member of the aldo-keto reductase super family. We screened OsDMAS1 paralogs for DMA synthesis. None of these paralogs displayed in vitro DMA synthesis activity, suggesting that rice only harbors one functional DMAS. We further characterized OsDMAS1 mutant plants. We failed to screen homozygous knock-out plants (dmas-1), so we characterized DMAS knock-down plants (dmas-kd1 and dmas-kd2). Under Fe-deficient conditions, dmas-kd1 plants were more chlorotic compared to the wild-type (WT) plants, and the expression of OsNAS3, OsYSL2, OsIRT1, and OsIRO2 was significantly up-regulated in the dmas-kd1 mutant, indicating that metal homeostasis was significantly disturbed. The secretion of DMA in dmas-kd1 was not significantly reduced. The dmas-kd1 plants accumulated less Fe in their roots compared to WT plants when grown with 10 μM FeSO4. The dmas-kd1 plants accumulated more Zn in their roots compared to WT plants under Fe-deficient, Fe-EDTA, and FeSO4 conditions. In both dehusked rice seeds (brown rice) and polished rice, no differences were observed for Fe, Cu, or Mn accumulation, whereas dmas-kd1 seeds significantly accumulated more Zn in brown rice. Our data suggests that rice only harbors one functional gene for DMA synthesis. In addition, the knock-down of OsDMAS1 significantly up-regulates the genes involved in Fe uptake and homeostasis.

Plant mechanisms responding to iron (Fe) deficiency have been widely described; it is well known that Strategy II plants, as durum wheat, cope with this stress by increasing both the synthesis and secretion of phytosiderophores (PS). The important contribution of the sulfate assimilatory pathway has been also demonstrated to improve Fe use efficiency in several grasses, such as maize, barley and wheat, most likely because PS are produced from nicotianamine, whose precursor is methionine. Here, the physiological response of durum wheat ( T. durum L.) plants - in terms of plant ionome, PS release, thiols content and S pathway-related enzymes – was investigated by gradually decreasing Fe availability that allowed the identification of three specific limit Fe concentrations: 75 μM, 25 μM and 0 μM Fe, i.e . the complete Fe deprivation. At each limit, plants begin to induce different and specific adaptive responses to improve Fe acquisition or to reduce the damage resulting from limited Fe availability. The identification of the Fe availability level below which durum wheat plants start an expensive metabolic reorganization of S and several other elements, could be of benefit not only for an effective cultivation of the crop but also for the grain quality.

Main conclusion The ratio of nicotianamine to deoxymugenic acid controls tissue-specific metal homeostasis in rice and regulates metal delivery to the endosperm. Abstract The metal-chelating phytosiderophores nicotianamine (NA) and 2′deoxymugenic acid (DMA) are significant factors for the control of metal homeostasis in graminaceous plants. These compounds are thought to influence metal homeostasis, but their individual roles and the effect of altering the NA:DMA ratio are unknown. We purposely generated rice lines with high and low NA:DMA ratios (HND and LND lines, respectively). The HND lines accumulated more iron (Fe), zinc (Zn), manganese (Mn) and copper (Cu) in the endosperm through the mobilization of Fe, Zn and Mn from the seed husk to the endosperm. In contrast, Fe, Zn and Mn were mobilized to the husk in the LND lines, whereas Cu accumulated in the endosperm. Different groups of metals are, therefore, taken up, transported and sequestered in vegetative tissues in the HND and LND lines to achieve this metal distribution pattern in the seeds. We found that different sets of endogenous metal homeostasis genes were modulated in the HND and LND lines to achieve differences in metal homeostasis. Our findings demonstrate that the NA:DMA ratio is a key factor regulating metal homeostasis in graminaceous plants. These findings can help formulate refined strategies to improve nutrient composition and nutrient use efficiency in crop plants., We acknowledge support from the European Research Council IDEAS Advanced Grant Program (BIOFORCE) to P.C., Generalitat de Catalunya Grant 2017 SGR 828 to ABBU (BIO2014-54426-P) and the Spanish Ministry of Economy and Competitivity (MINECO; projects AGL2016-75226-R, co-financed with FEDER) and the Aragón Government (Group A09_17R) to J.A. R.B was supported by a PhD fellowship from the University of Lleida, Spain.

Iron is an essential metal for all living organisms that is absorbed in the intestinal cells as a heme-chelated or free form. It is unclear how important plant-derived chelators, such as nicotianamine (NA), an organic small molecule that is ubiquitous in crops, vegetables, and various other foods, contribute to iron bioavailability in mammals. We performed electrophysiological assays with Xenopus laevis oocytes and radioactive tracer experiments with Caco-2 cells. The findings revealed that the proton-coupled amino acid transporter SLC36A1 (PAT1) transports iron in the form of NA-Fe (II) complex in vitro. Decreased expression of hPAT1 by RNA interference in Caco-2 cells reduced the uptake of NA-59Fe (II) complex. The uptake of inorganic 59Fe (II) was relatively unaffected. These results imply that PAT1 transports iron as a NA-Fe (II) complex. The rate of 59Fe absorption in the spleen, liver, and kidney was higher when mice were orally administered NA-59Fe (II) compared with free 59Fe (II). The profile of site-specific PAT1 expression in the mouse intestine coincided with those of NA and iron contents, which were the highest in the proximal jejunum. Orally administered NA-59Fe (II) complex in mice was detected in the proximal jejunum by thin layer chromatography. In contrast, much less 59Fe (or NA) was detected in the duodenum, where the divalent metal transporter SLC11A2 (DMT1) absorbs free Fe (II). The collective results revealed the role of PAT1 in NA-Fe (II) absorption in the intestine and potential implication of NA in iron uptake in mammals.

In recent years it has been established a significant relationship between sulfur (S) and iron (Fe) nutrition. In particular, it has been demonstrated that S deprivation can hinder Fe acquisition in barley, maize and wheat. This can be explained by assuming that, in order to cope with low Fe availability in the soil, grasses have evolved a phytosiderophores (PS)-based Fe chelation system (Strategy II) and PS are synthesized from methionine, through a nicotianamine intermediate. On the other hand, it has been demonstrated that wheat plants exhibit a higher Fe accumulation when supplied with excess S concentration, this effect being especially beneficial under severely limited Fe supply. The goal of this study was to explore whether the higher ability to acquire Fe, induced by a higher S supply, might be a general response of graminaceous species. The response of durum wheat ( Triticum durum L.), barley ( Hordeum vulgare L.), and maize ( Zea mays L.) to the excess of S availability (2.4 mM vs 1.2 mM which is considered as optimal) was studied as a function of Fe availability (limited and sufficient, 20 and 80 μM, respectively). At the end of the experimental period, which lasted 11 days, growth parameters (shoot and root fresh weight and chlorophyll content), total S and Fe concentrations, and PS release rate were compared among the three species. Furthermore, we evaluated plant sulfate uptake capability, by analysing the expression of genes coding for high affinity sulfate transporter ( TdST1.1, HvST1.1 and ZmST1.1 ) in roots of each graminaceous plant. Our preliminary findings are largely consistent with the apparent divergence among the three species. In particular, an excess S supply may result in the improvement of Fe use efficiency in durum wheat plants, but not in both barley and maize. The use of higher S supply seems to be a promising approach, at least for wheat plants, which can both reduce agricultural demand for Fe fertilizers and improve the Fe use efficiency of plants.

Background The release of phytosiderephores (PS) to the rhizosphere is the main root response to iron (Fe) deficiency in graminaceous plants. We have investigated the role of the Fe status in the shoot as well as of the signaling pathways controlled by three relevant phytoregulators – indolacetic acid (IAA), ethylene and nitric oxide (NO) – in the regulation of this root response in Fe-starved wheat plants. To this end, the PS accumulation in the nutrient solution and the root expression of the genes encoding the nicotianamine aminotransferase (TaNAAT) and ferritin (TaFER) have been evaluated in plants subjected to different treatments. Results The application of Fe to leaves of Fe-deficient plants prevented the increase in both PS root release and TaNAAT gene expression thus showing the relevant role of the shoot to root communication in the regulation of PS root release and some steps of PS biosynthesis. Experiments with specific hormone inhibitors showed that while ethylene and NO did not positively regulate Fe-deficiency induced PS root release, auxin plays an essential role in the regulation of this process. Moreover, the application of IAA to Fe-sufficient plants promoted both PS root release and TaNAAT gene expression thus indicating that auxin might be involved in the shoot to root signaling network regulating Fe-deficiency root responses in wheat. Conclusions These results therefore indicate that PS root release in Fe-deficient wheat plants is directly modulated by the shoot Fe status through signaling pathways involving, among other possible effectors, auxin.

The micronutrient iron (Fe) is essential for photosynthesis, respiration, and many other processes, but it is only sparingly soluble in aqueous solution, making adequate acquisition by plants a serious challenge. Fe is a limiting factor for plant growth on approximately 30% of the world’s arable lands. Moreover, Fe deficiency in humans is a global health issue, affecting 1.62 billion people, or about 25% of the world’s population. It is imperative that we gain a better understanding of the mechanisms that plants use to regulate iron homeostasis, since these will be important targets for future biofortification and crop improvement strategies. Grasses and non-grasses have evolved independent mechanisms for primary iron uptake from the soil. The grasses, which include most of the world’s staple grains, have evolved a distinct ‘chelation’ mechanism to acquire iron from the soil. Strong iron chelators called phytosiderophores (PSs) are synthesized by grasses and secreted into the rhizosphere where they bind and solubilize Fe(III). The Fe(III)-PS complex is then taken up into root cells via transporters specific for the Fe(III)-PS complex. In this study, 31 novel, uncharacterized striped maize mutants available through the Maize Genetics Cooperation Stock Center (MGCSC) were analyzed to determine whether their mutant phenotypes are caused by decreased iron. Many of these proved to be either pale yellow or white striped mutants. Complementation tests were performed by crossing the MGCSC mutants to ys1 and ys3 reference mutants. This allowed assignment of 10 ys1 alleles and 4 ys3 alleles among the novel mutants. In addition, four ys∗ mutant lines were identified that are not allelic to either ys1 or ys3. Three of these were characterized as being non-allelic to each other and as having low iron in leaves. These represent new genes involved in iron acquisition by maize, and future cloning of these genes may reveal novel aspects of the grass iron acquisition mechanism.

Many chemicals, including herbicides, are routinely applied to crops for weed management and food production improvement. However, the intensive use of herbicides could lead eventually to a great environmental threat due to their persistence and accumulation in the ecosystems and contamination of soils. Furthermore, the possible effect of these chemicals on nutrient uptake and assimilation in crops has only recently been discussed. The present study aimed at understanding the effect of the herbicide terbuthylazine (TBA), a herbicide commonly applied to control weeds in leguminous species and triazine tolerant crops, on the capability of maize plants to cope with iron (Fe) shortage. The application of 2 and 5 mg L−1 TBA caused a significant reduction of root Fe concentration. This reduction might be attributed to a decreased release of phytosiderophores, which in turn could be ascribed to a reduced sulfur assimilation. Results provide evidence that TBA impairs Fe uptake and accumulation in non-target plants most likely interacting with sulfur-assimilating enzymes [ATP sulfurylase and O-acetylserine(thiol)lyase].

Safeners are agrochemicals used in agriculture to protect crops from herbicide injuries. They act by stimulating herbicide metabolism. As graminaceous plants, to cope with iron (Fe) deficiency, activate sulfur (S) metabolism and release huge amounts of Fe-chelating compounds, or phytosiderophores (PSs), we investigated, in barley plants (Hordeum vulgare, L.) grown in Fe deficiency, the effects of three safeners on two enzymes of S assimilation, cysteine (Cys) and glutathione (GSH), and PS release. Finally, we monitored the root Fe content in plants treated with the most effective safener.Generally, all the safeners activated S metabolism and increased Cys and GSH contents. In addition, the safened plants excreted higher levels of PSs. Given that mefenpyr-diethyl (Mef) was the most effective in causing these effects, we assessed the Fe concentration in Mef-treated barley and found higher Fe levels than those in untreated plants.The three safeners, in different ways but specifically, activated S reductive metabolism and regulated Cys and GSH contents, PS release rate and Fe content (Mef-treated barley). The results of this research provide new indications of the biochemical and physiological mechanisms involved in the safening action. © 2016 Society of Chemical Industry.

Growth and development of plants mainly depend on the availability of water and nutrients in their habitat. Increasing food production to face the rise of global population will require to exploit uncultivated or abandoned arable land, some of which with unfavorable soil mineral composition or availability, and to reduce the use of fertilizers for more sustainable practices. Therefore, understanding the mechanisms underlying nutrients acquisition by plants and improving plant nutrient use efficiency (NUE) is of social, economic, agricultural, and ecological importance. The availability of nutrients in different soils and how it influences the physiology of the plants has been widely described at the level of single nutrient. However, this picture is most likely confounded by nutrient interactions. In nature it is unlikely that plants are exposed to a single nutrient deficiency. Multiple nutrient deficiency situations are more likely to occur. For example, agricultural soils are becoming depleted for important elements, such as sulfur (S), while iron (Fe), although being highly abundant in the soil, is poorly available for uptake due to its insolubility in the soil. Therefore, interactions among the various mineral elements need to be investigated urgently in order to understand how the different sensing and signaling pathways activated in response to changes in availability of one element are coordinately integrated with those of other elements. The main problem in studying nutrient interactions is that, in some cases, stress responses are antagonistic, while in other ones are either additive or synergistic, thus combined nutrient deficiencies very often result as a new condition, that requires different responses from those observed when plant response to deficiency of a single nutrient is described. The aim of this thesis was to provide a comprehensive understanding of the molecular, physiological, and biochemical interactions underlying S and Fe nutrition and how these two essential nutrients are associated within the plant and one may impact the uptake, transport and storage of another. In particular, the present thesis deal with three key questions for this critical research area: 1) New functions for S in improving plant efficiency to uptake, transport or accumulate Fe; 2) New mechanisms underlying Fe homeostasis and how these are impacted by S; 3) Crop management strategies that minimize Fe deficiency without additional input of Fe fertilizers. Since monocots and dicots have, for the most part, distinct mechanisms for regulating Fe acquisition, both Strategy I and Strategy II plants (tomato and durum wheat, respectively) were considered. As a result of the different approaches used in the research, several novel acquisitions have been achieved from the analysis of S/Fe interaction. As expected, it has to be assumed a complex regulatory system, wherein S and Fe starvation induce partially overlapping and partially distinct response mechanisms, which coordinate plant response to either or to joint starvations. A high degree of common and even synergistic response patterns as well as nutrient specific responses were identified by the metabolomic approach in tomato. Furthermore, it appears that tomato plants exposed to Fe deficiency are able to change S metabolic balance mimicking S starvation responses through an increased sulfate uptake and root-to-shoot translocation to meet the increased demand for ethylene and nicotianamine, allowing them to cope with this stress. On the other hand, the finding that in wheat plants changes in S accumulation were closely related to plant capability to release phytosiderophores and correspondingly to accumulate Fe could be significant to overcome a key challenge in plant Fe nutrition: the optimization of Fe acquisition and allocation based on a sustainable new approach with low cost and decreased requirements for Fe fertilizers. Besides being a promising approach, whereby targeted application of one nutrient (S) could be used to overcome the deficiency of another (Fe), it could be also a promising tool to increase the micronutrient content (Fe) in food crops. This could lead to a decrease, in the long term, of chemical inputs, getting healthier food and respecting the surrounding environment of the cultivated crops, as well as the health of the agricultural workers. Furthermore, the identification of the Fe availability level, below which durum wheat plants start an expensive metabolic reorganization of S and several other elements, could be particularly important not exclusively for the cost of the metabolic reorganization underlying the adaptive responses to the Fe shortage but also for the grain quality that this process may affect. In fact, it is well known that S is relevant for bread and pasta-making qualities through its effect on grain protein composition. In conclusion, to extend these concepts, we need to develop a better understanding of how Fe/S interactions take place at a mechanistic level and, for that, a multi-stage comprehensive approach is necessary. La crescita e lo sviluppo delle piante dipende principalmente dalla disponibilità di acqua e nutrienti presenti nell’ambiente. La necessità di incrementare la produzione agricola per far fronte alla crescita della popolazione mondiale richiederà di sfruttare come aree agricole anche i suoli non coltivati e/o abbandonati, sebbene caratterizzati da condizioni sfavorevoli per la crescita delle piante (scarsa disponibilità di nutrienti) e di ridurre l’impiego di fertilizzanti chimici, perseguendo pratiche agricole più sostenibili. Migliorare la conoscenza dei meccanismi di acquisizione dei nutrienti per aumentare l’efficienza d’uso dei nutrienti (NUE) delle piante, è quindi di fondamentale importanza sociale, economica, agricola ed ecologica. Le risposte fisiologiche delle piante alle fluttuazioni della disponibilità dei nutrienti nel suolo è stata ampiamente studiata e discussa a livello della singola carenza nutrizionale. Tuttavia, tali studi non rispecchiano la situazione reale di campo in quanto non tengono conto delle interazioni che si verificano tra i nutrienti a livello rizosferico. Negli ambienti naturali, raramente le piante sono esposte ad una sola carenza nutrizionale, mentre è più frequente il verificarsi di carenze nutrizionali multiple. Recentemente è stato osservato un graduale impoverimento dei suoli agricoli di un importante nutriente, lo zolfo (S), mentre il ferro (Fe), sebbene sia molto abbondante nel suolo, a causa della scarsa solubilità non risulta facilmente disponibile per le piante. Lo studio delle interazioni tra i vari nutrienti risulta quindi di primaria importanza per comprendere come i meccanismi attivati in risposta alla carenza di un elemento si coordinano con quelli di altri elementi. Il problema principale, in questo tipo di studio, deriva dal fatto che le carenze multiple in alcuni casi generano delle risposte antagonistiche mentre, in altri, additive o sinergiche, per cui molto spesso determinano la comparsa di una situazione nuova che richiede l’attivazione di una risposta completamente diversa da quella osservata per la singola carenza nutrizionale. Lo scopo di questa tesi è stato di analizzare l’interazione tra lo S e il Fe a livello molecolare, fisiologico e biochimico al fine di comprendere come i meccanismi di assorbimento, trasporto e stoccaggio dell’uno siano coordinati con quelli dell’altro. In particolare, in questa tesi sono state affrontate tre questioni fondamentali: 1) il meccanismo dell’omeostasi del Fe e come questo sia influenzato dallo S; 2) il ruolo dello S nel migliorare l’efficienza di assorbimento, trasporto e accumulo di Fe nella pianta; 3) ipotesi di strategie di gestione colturale finalizzate a ridurre il problema della Fe carenza limitando l’uso di fertilizzanti ferrici. Dato che i meccanismi per l’acquisizione del Fe sono diversi nelle dicotiledoni e nelle graminacee (Strategia I e II, rispettivamente), lo studio è stato condotto su entrambi i tipi di piante, prendendo come piante modello pomodoro e frumento duro, rispettivamente. I risultati ottenuti hanno mostrato l’esistenza di un complesso sistema di regolazione, in cui la carenza combinata di S e di Fe induce nelle piante meccanismi di risposta, in parte sovrapposti ed in parte distinti. Attraverso l’analisi metabolica in piante di pomodoro sono state evidenziate risposte comuni e anche sinergiche indotte dalla carenza combinata dei due elementi, ma anche risposte specifiche del singolo nutriente. Inoltre, il pomodoro in condizioni di Fe carenza sembra in grado di indurre l’assorbimento e l’assimilazione dello S simulando quanto avviene in risposta alla S carenza, al fine di soddisfare la maggiore richiesta di etilene e di nicotianamina, necessaria per affrontare lo stress. D’altra parte, l’osservazione in frumento che la maggiore capacità della pianta di accumulare S è strettamente correlata ad un maggiore rilascio di fitosiderofori e quindi ad un maggiore accumulo di Fe a livello fogliare, potrebbe essere importante per affrontare una delle sfide principali della nutrizione ferrica delle piante: l’ottimizzazione dell’acquisizione e dell’allocazione del Fe tramite un approccio innovativo e sostenibile, che richieda bassi costi ed un minore uso di fertilizzanti contenenti Fe. Questo, oltre ad essere un promettente approccio, in cui la somministrazione di un nutriente (S) potrebbe permettere di superare la carenza di un altro nutriente (Fe), potrebbe anche rappresentare una strategia promettente per incrementare la quantità del micronutriente (Fe) nelle parti edibili delle colture agrarie. Tutto ciò potrebbe determinare una riduzione, a lungo termine, dell’impiego di prodotti chimici, la produzione di alimenti più sani, il rispetto dell’ambiente e la salute degli agricoltori. Inoltre, l’identificazione del livello soglia di disponibilità di Fe, sotto il quale le piante di frumento duro cominciano a riorganizzare il metabolismo dello S e di altri elementi con elevato dispendio energetico, potrebbe rappresentare un vantaggio economico non solo dal punto di vista della quantità, ma anche della qualità della granella dalla coltivazione di questa pianta. Infatti, è noto che lo S riveste un ruolo importante non solo nell’aumentare l’efficienza di acquisizione del Fe nelle graminacee, ma anche nel determinare la qualità delle farine utilizzate nei processi di panificazione e pastificazione. Tuttavia, molto lavoro è ancora necessario per una comprensione dettagliata della relazione tra S e Fe, soprattutto a livello meccanicistico e, per questo, è sicuramente utile seguire un approccio multidisciplinare. Dottorato di ricerca in Scienze e tecnologie per la gestione forestale e ambientale

Iron (Fe) is essential for rice growth and humans consuming as their staple food but is often deficient because of insoluble Fe(III) in soil for rice growth and limited assimilation for human bodies, while cadmium (Cd) is non-essential and toxic for rice growth and humans if accumulating at high levels. Over-accumulated Cd can cause damage to human bodies. Selecting and breeding Fe-rich but Cd-free rice cultivars are ambitious, challenging and meaningful tasks for researchers. Although evidences show that the mechanisms of Fe/Cd uptake and accumulation in rice are common to some extent as a result of similar entry routes within rice, an increasing number of researchers have discovered distinct mechanisms between Fe/Cd uptake and accumulation in rice. This comprehensive review systematically elaborates and compares cellular mechanisms of Fe/Cd uptake and accumulation in rice, respectively. Mechanisms for maintaining Fe homeostasis and Cd detoxicification are also elucidated. Then, effects of different fertilizer management on Fe/Cd accumulation in rice are discussed. Finally, this review enumerates various approaches for reducing grain Cd accumulation and enhancing Fe content in rice. In summary, understanding of discrepant cellular mechanisms of Fe/Cd accumulation in rice provides guidance for cultivating Fe-fortified rice and has paved the way to develop rice that are tolerant to Cd stress, aiming at breeding Fe-rich but Cd-free rice.

One of the main purposes of modern agriculture is to raise crop production by means of a proper application of herbicides. Nonetheless, the use of herbicides is causing some concerns associated to their persistence and accumulation in the environment. Notwithstanding the relevance of these aspects, little or no attention has been paid to the interferences that these chemicals and their residues can exert on iron (Fe) mineral nutrition of plants. To this purpose, we studied the effect of terbuthylazine (TBA) on the Fe-acquisition processes of Fe-deficient barley plants, by investigating some aspects related to phytosiderophores (PS) exudation and sulfur (S) metabolism. Results showed that plant growth, chlorophyll content expressed as SPAD index, PS release and the expression of genes involved in their secretion, uptake and biosynthesis were negatively affected. This response was associated with reduced cysteine concentrations, and it suggests that the TBA interferences on Fe-acquisition in barley are the consequence of the induced changes in S metabolism.

Bor ve demir yüksek bitkilerin büyümesi ve gelişimi için elzem besin maddeleridir. Bu maddelerin eksikliği ve aşırı fazlalığı bitkilerin fizyolojik, hücresel ve moleküler yapılarında kayda değer değişikliklere yol açar. Arpada demir alımı başlıca fitosideroforların üretimi ve çevreye salınımı ile olur. Diğer yandan fitosiderofor üretimi bor ve demir konsantrasyonlarından da etkilenir. Bitkiler farklı demir ve bor konsantrasyonlarında büyütüldüklerinde, fitosiderofor hidroksimugineik asit (HMA) üretiminden sorumlu dört enzim (NAS, NAAT, IDS2 ve IDS3) farklı ekspresyon profilleri göstermektedir. Bu çalışmada arpada demir, bor ve fitosiderofor üretimi arasındaki olası bağlantıyı araştırılmıştır. Bu amaçla, arpa fideleri bor ve demir eksikliği, fazlalığı ve bu durumların kombinasyonlarıyla muamele edilmiş ardından daha önce adı geçen genlerin ekspresyon seviyeleri incelenmiştir. Bunun yanısıra, bitkilerin fizyolojik parametreleri, toplam bor, demir ve çinko içerikleri ve fotosentetik aktiviteleri göz önünde bulundurulmuştur. Boron and iron are essential nutrients for growth and development of higher plants. Deficiency and toxicity of them cause significant changes in physiological, cellular and molecular make-up of plants. Iron uptake in barley is mainly achieved by production and release of phytosiderophores to the environment. On the other hand, production of phytosiderophores are affected by concentrations of boron and iron. Four enzymes which function in phytosiderophore hydroxymugineic acid (HMA) production namely NAS, NAAT, IDS2, and IDS3 showed differential expression profiles when plants were treated with different concentrations of iron and boron. In this study, we investigated the potential link between iron, boron, and phytosiderophore production in barley. For this purpose the barley seedlings were treated with deficiency or toxicity of boron and iron as well as combinations of these and expression levels of aforementioned genes were examined.In addition to gene expression analysis, physiological parameters, total boron, iron, and zinc content and photosynthetic performance of seedlings were considered. 120

Chlorosis-susceptible fruit trees growing on calcareous soils have been observed to recover in the presence of grass cover species. However, the physiological mechanisms behind this phenomenon are only scarcely understood. An investigation was carried out to verify whether citrus plants can use 59Fe solubilized from a sparingly soluble source by the phytosiderophores (PS) released from graminaceous species. Experiments were performed in hydroponics, using two citrus rootstocks differing in their sensitivity to Fe-deficiency in the field (Poncirus trifoliata · Citrus paradisi, citrumelo ‘‘Swingle’’, highly susceptible, and Citrus aurantium L., moderately tolerant). Barley (Hordeum vulgare L., cv Europa) was used as a model species for PS-releasing graminaceous plants. Fe-deficient citrus plants increased 59Fe uptake from 59Fe-hydroxide supplied inside a dialysis tube, when Fe-deficient barley plants or PS-containing barley root exudates were present in the uptake solution, this effect being particularly evident for the susceptible rootstock. 59Fe-uptake from 59Fe-hydroxide was also enhanced in Fe-deficient citrumelo ‘‘Swingle’’ in the presence of Fe-deficient Poa pratensis L. and Festuca rubra L., two perennial grasses normally grown in association with fruit trees; no effect was found when Fe-sufficient grasses were employed. The uptake of 59Fe by the susceptible citrus rootstock increased in proportion to the amount of 2¢-deoxymugineic acid (DMA), the major PS released by Fe-deficient F. rubra, present in the uptake solution. The beneficial effect of F. rubra or P. pratensis was evident from the leaf re-greening observed when Fe-deficient citrumelo ‘‘Swingle’’ plants were grown in association with the grasses in pots filled with a calcareous soil. Leaf regreening was not observed when citrumelo ‘‘Swingle’’ plants and yellow stripe 3 (ys3) maize (Zea mays L.) mutant plants, unable to release PS, were co-cultivated in pots filled with calcareous soil, unless exogenous PS were added to the soil. Results indicate that graminaceous cover species can improve the Fe-nutrition of fruit trees grown on calcareous soils by enhancing Fe-availability.

This study was implemented to determine the appropriate methods to be used in evaluation of iron (Fe) chlorosis in peach trees. For this purpose, three peach orchards having trees various visual chlorosis ratings were selected, and the leaf samples were collected during a 2-year period. The leaf samples were collected from two different positions in the annual shoots: 1) 5(th) 6(th), and 7(th) leaves in midpoint position of annual shoots and 2) extremity leaves of annual shoots. The leaf samples were only taken from the trees at 3 stages in the first year of the experiment to determine most convenient date of sampling. The experiment had 3 chlorosis categories (green, slightly chlorotic, and severely chlorotic) in the first year and 5 chlorosis categories in the second year (green, slightly chlorotic, slightly green, chlorotic, and severely chlorotic) with 3 replicates in each of the orchards. One M HCl, 1.5% o-phenanthroline, 0.5 M oxalic acid, and 0.1 M HCl methods in the fresh leaf samples, 1 M HCl, 0.5 M oxalic acid, 0.1 M HCl, and total Fe methods in oven-dried leaves were used for Fe analysis. The data were statistically analyzed by analysis of variance (ANOVA) and the least significant difference (LSD) values were calculated at the 5% level. According to the results obtained from the experiment, 1 M HCl and 1.5% o-phenanthroline in the fresh leaves and 1 M HCl in oven-dried leaves were well related to visual chlorosis ratings and chlorophyll content of the leaves as compared with other methods. The 1 M HCl method seemed to be the method of choice for producing a suitable index of Fe status of plants due to lower cost of analysis and ease in handling dry samples. Concentration of Fe determined by this method from green leaves at the midpoint position was generally greater than or equal to 30 mg kg(-1), which may be accepted as the critical index value for Fe in peach trees. Active Fe in the midpoint leaves was markedly and significantly higher than extremity leaves. To sample midpoint leaves may be recommended in determination of active Fe. There was no evidence to sample leaves on a certain date. This might have resulted from the wide range of sampling interval.

Grasses are more efficient than dicots in acquiring Fe from calcareous soils. We studied whether intercropping with grasses alleviates Fe chlorosis in olive and whether the effect persists in succeeding dicot crops. Three different pot experiments were conducted. In the first, olive plants were intercropped with 6 different grass species (purple false brome, annual ryegrass, compact brome, goatgrass, barley and red fescue); in the second, the two species best performing in the previous experiment were studied in various calcareous soils and; in the third, chickpea and peanut were grown in pots previously used to cultivate the two grasses. Intercropping with purple false brome and barley increased leaf chlorophyll concentrations and/or boosted growth of olive trees on three different calcareous soils. Olive growth was adversely affected by intercropping in one soil as a result of competition for water. Intercropping increased Fe, Mn, Cu and Zn leaf contents in olive. Also, grass cropping generally raised available levels of soil Fe, Mn, Cu and Zn; this effect, however, resulted in no substantial alleviation of Fe chlorosis in succeeding chickpea or peanut crops. Intercropping with purple false brome and barley appears to be a promising remedy for Fe chlorosis in olive orchards affected by Fe chlorosis.