OPT3 Is a Component of the Iron-Signaling Network between Leaves and Roots and Misregulation of OPT3 Leads to an Over-Accumulation of Cadmium in Seeds

Plants and seeds are the main dietary sources of zinc, iron, manganese, and copper, but are also the main entry point for toxic elements such as cadmium into the food chain. We report here that an Arabidopsis oligopeptide transporter mutant, opt3-2, over-accumulates cadmium （Cd） in seeds and roots but, unexpectedly, under-accumulates Cd in leaves. The cadmium distribution in opt3-2 differs from iron, zinc, and manganese, suggesting a metal-specific mechanism for metal partitioning within the plant. The opt3-2 mutant constitutively up-regulates the Fe/Zn/Cd transporter IRT1 and FRO2 in roots, indicative of an iron-deficiency response. No genetic mutants that impair the shoot-to-root signaling of iron status in leaves have been identified. Interestingly, shoot-specific expression of OPT3 rescues the Cd sensitivity and complements the aberrant expression of IRT1 in opt3-2 roots, suggesting that OPT3 is required to relay the iron status from leaves to roots. OPT3 expression was found in the vasculature with preferential expression in the phloem at the plasma membrane. Using radioisotope experiments, we found that mobilization of Fe from leaves is severely affected in opt3-2, suggesting that Fe mobilization out of leaves is required for proper trace-metal homeostasis. When expressed in yeast, OPT3 does not localize to the plasma membrane, precluding the identification of the OPT3 substrate. Our in planta results show that OPT3 is important for leaf phloem-loading of iron and plays a key role regulating Fe, Zn, and Cd distribution within the plant. Furthermore, ferric chelate reductase activity analyses provide evidence that iron is not the sole signal transferred from leaves to roots in leaf iron status signaling.     

Possible roles of phytochelatins and glutathione metabolism in cadmium tolerance in chickpea roots

The possible roles of phytochelatin (PC) and glutathione (GSH) in the heavy metal detoxification in plants were examined using two varieties (CSG-8962 and C-235) of chickpea (Cicer arietinum L.). The seedlings were grown for 5 days and the roots were treated with 0–20 μM CdSO₄ for 3 days. The CSG-8962 seedlings exhibited more Cd-tolerant characteristics than did the C-235, where the roots, rather than shoots, suffered from more toxic effects by Cd. Both the seedlings synthesized the large amounts of PCs and homo-phytochelatins (hPCs) in roots, but only a little in shoots in response to Cd. The Cd treatments also caused a marked increase in the levels of GSH and cysteine in both the root and shoot tissues, suggesting that Cd may activate the GSH biosynthesis and, hence, enhance PC synthesis in the plants. Such a Cd-sensitive PC synthesis in chickpea plants does not explain the difference in Cd sensitivity in the varieties, but can be used as a biochemical indicator for Cd contamination in various environments. In the chickpea plants, possible PC-dependent and independent mechanisms for Cd tolerance are discussed. Electronic Publication     

Upland rice and lowland rice exhibited different PIP expression under water deficit and ABA treatment

Aquaporins play a significant role in plant water relations. To further understand the aquaporin function in plants under water stress, the expression of a subgroup of aquaporins, plasma membrane intrinsic proteins （PIPs）, was studied at both the protein and mRNA level in upland rice （Oryza sativa L. cv. Zhonghan 3） and lowland rice （Oryza sativa L. cv. Xiushui 63） when they were water stressed by treatment with 20% polyethylene glycol （PEG）. Plants responded differently to 20% PEG treatment. Leaf water content of upland rice leaves was reduced rapidly. PIP protein level increased markedly in roots of both types, but only in leaves of upland rice after 10 h of PEG treatment. At the mRNA level, OsPIP1,2, OsPIP1,3, OsPIP2;1 and OsPIP2;5 in roots as well as OsPIP1,2 and OsPIP1;3 in leaves were significantly up-regulated in upland rice, whereas the corresponding genes remained unchanged or down-regulated in lowland rice. Meanwhile, we observed a significant increase in the endogenous abscisic acid （ABA） level in upland rice but not in lowland rice under water deficit. Treatment with 60 μM ABA enhanced the expression of OsPIP1;2, OsPIP2;5 and OsPIP2;6 in roots and OsPIP1;2, OsPIP2;4 and OsPIP2;6 in leaves of upland rice. The responsiveness of PIP genes to water stress and ABA were different, implying that the regulation of PIP genes involves both ABA-dependent and ABA-independent signaling oathways during water deficit.     

Matricaria chamomilla is not a hyperaccumulator, but tolerant to cadmium stress

The influence of low (3 μM) and high (60 and 120 μM) cadmium (Cd) concentrations were studied on selected aspects of metabolism in 4-week-old chamomile (Matricaria chamomilla L.) plants. After 10 days’ exposure, dry mass accumulation and nitrogen content were not significantly altered under any of the levels of Cd. However, there was a significant decline in chlorophyll and water content in the leaves. Among coumarin-related compounds, herniarin was not affected by Cd, while its precursors (Z)- and (E)-2-β-d-glucopyranosyloxy-4-methoxycinnamic acids (GMCAs) increased significantly at all the levels of Cd tested. Cd did not have any effect on umbelliferone, a stress metabolite of chamomile. Lipid peroxidation was also not affected by even 120 μM Cd. Cd accumulation was approximately seven- (60 μM Cd treatment) to eleven- (120 μM Cd treatment) fold higher in the roots than that in the leaves. At high concentrations, it stimulated potassium leakage from the roots, while at the lowest concentration it could stimulate potassium uptake. The results supported the hypothesis that metabolism was altered only slightly under high Cd stress, indicating that chamomile is tolerant to this metal. Preferential Cd accumulation in the roots indicated that chamomile could not be classified as a hyperaccumulator and, therefore, it is unsuitable for phytoremediation.     

OsHT, a Rice Gene Encoding for a Plasma-Membrane Localized Histidine Transporter

Using a degenerative probe designed according to the most conservative region of a known Lys- and His-specific amino acid transporter (LHT 1) from Arabidopsis, we isolated a full-length cDNA named OsHT (histidine transporter of Oryza sativa L.) by screening the rice cDNA library. The cDNA is 1.3 kb in length and the open reading frame encodes for a 441 amino acid protein with a calculated molecular mass of 49 kDa. Multiple sequence alignments showed that OsHT shares a high degree of sequence conservation at the deduced amino acid level with the Arabidopsis LHT1 and six putative lysine and histidine transporters. Computational analysis indicated that OsHT is an integral membrane protein with 11 putative transmembrane helices. This was confirmed by the transient expression assay because the OsHT-GFP fusion protein was, indeed, localized mainly in the plasma membrane of onion epidermal cells. Functional complementation experiments demonstrated that OsHT was able to work as a histidine transporter in Saccharomyces cerevisiae, suggesting that OsHT is a gene that encodes for a histidine transporter from rice.This is the first time that an LHT-type amino acid transporter gene has been cloned from higher plants other than Arabidopsis.     

Upland rice and lowland rice exhibited different P/P expression under water deficit and ABA treatment

Aquaporins play a significant role in plant water relations.To further understand the aquaporin function in plants underwater stress,the expression of a subgroup of aquaporins,plasma membrane intrinsic proteins(PIPs),was studied at boththe protein and mRNA level in upland rice(Oryza sativa L.cv.Zhonghan 3)and lowland rice(Oryza sativa L.cv.Xiushui63)when they were water stressed by treatment with 20% polyethylene glycol(PEG).Plants responded differently to20% PEG treatment.Leaf water content of upland rice leaves was reduced rapidly.PIP protein level increased markedlyin roots of both types,but only in leaves of upland rice after 10h of PEG treatment.At the mRNA level,OsPIP1;2,Os-PIP1;3,OsPIP2;1 and OsPIP2;5 in roots as well as OsPIP1;2 and OsPIP1;3 in leaves were significantly up-regulatedin upland rice,whereas the corresponding genes remained unchanged or down-regulated in lowland rice.Meanwhile,weobserved a significant increase in the endogenous abscisic acid(ABA)level in upland rice but not in lowland rice underwater deficit.Treatment with 60μM ABA enhanced the expression of OsPIP1;2,OsPIP2;5 and OsPIP2;6 in roots andOsPIP1;2,OsPIP2;4 and OsPIP2;6 in leaves of upland rice.The responsiveness of PIP genes to water stress and ABAwere different,implying that the regulation of PIP genes involves both ABA-dependent and ABA-independent signalingpathways during water deficit.     

Iron deprivation results in a rapid but not sustained increase of the expression of genes involved in iron metabolism and sulfate uptake in tomato （Solanum lycopersicum L.） seedlings

Characterization of the relationship between sulfur and iron in both Strategy I and Strategy II plants, has proven that low sulfur availability often limits plant capability to cope with iron shortage. Here it was investigated whether the adaptation to iron deficiency in tomato （Solanum lycopersicum L.） plants was associated with an increased root sulfate uptake and translocation capacity, and modified dynamics of total sulfur and thiols accumulation between roots and shoots. Most of the tomato sulfate transporter genes belonging to Groups 1, 2, and 4 were significantly upregulated in iron-deficient roots, as it commonly occurs under S-deficient conditions. The upregulation of the two high affinity sulfate transporter genes, SlST1.1 and SlST1.2, by iron deprivation clearly suggests an increased root capability to take up sulfate. Furthermore, the upregulation of the two low affinity sulfate transporter genes SlST2.1 and SlST4.1 in iron-deficient roots, accompanied by a substantial accumulation of total sulfur and thiols in shoots of＆amp;nbsp;iron-starved plants, likely supports an increased root-to-shoot translocation of sulfate. Results suggest that tomato plants exposed to iron-deficiency are able to change sulfur metabolic balance mimicking sulfur starvation responses to meet the increased demand for methionine and its derivatives, al owing them to cope with this stress.     

Functional characterization of a putative nitrate transporter gene promoter from rice

Drought is one of the most significant abiotic stresses that influence plant growth anddevelopment.Expression analysis revealed that OsNRT1.3,a putative nitrate transporter gene in rice,wasinduced by drought.To confirm if the OsNRT1.3 promoter can respond to drought stress,a 2019 bpupstream sequence of OsNRT1.3 was cloned.Three OsNRT1.3 promoter fragments were generated by5′-deletion,and fused to the β-glucuronidase (GUS) gene.The chimeric genes were introduced into riceplants.NRT2019::GUS,NRT1196::GUS and NRT719::GUS showed similar expression patterns in seeds,roots,leaves and flowers in all transgenic rice,and GUS activity conferred by different OsNRT1.3 promoterfragments was significantly upregulated by drought stress,indicating that OsNRT1.3 promoter responds todrought stress and the 719 bp upstream sequence of OsNRT1.3 contains the drought response elements.     

Cloning,expression and function of phosphate transporter encoded gene in Oryza sativa L.

OsPT6:1,a phosphate transporter encoding gene from the leaf samples of Oryza sativa, was identified through PCR with specifically designed primers.The phylogenetic analysis and the conserved amino acid residue site detection suggested OsPT6:1 a possible high-affinity phosphate transporter encoding gene.In situ hybridization and RT-PCR demonstrated the expression of OsPT6:1 in both roots and leaves.The peak expression signal was observed in mesophyll cells under low phosphorus(P)induction.A homologous recombination study indicated that OsPT6:1 can enhance the Pi uptake efficiency of Pichia pastoris.At the meantime,the introduction of OsPT6:1 was able to complement the Pi uptake function of yeast cells with high-affinity phosphate transporters de- ficient.Those results substantiated our contention that OsPT6:1 encoded a high-affinity phosphate transporter of Oryza sativa.     

Effects of Interaction Between Cadmium and Plumbum on Phytochelatins and Glutathione Production in Wheat （Triticum aestivum L.）

Phytochelatins (PCs) may function as a potential biomarker for metal toxicity. However, less attention has been paid to the effects of metal interactions on the production of PCs and glutathione (GSH), the most prominent cellular thiol. In the present study, the effects of interactions between cadmium (Cd) and plumbum (Pb) on the production of PCs and GSH were monitored over a period of 14 d in wheat (Triticum aestivum L.) tissues. The results showed that combination of Cd and Pb led to synergistic growth inhibition in wheat. Exposure to Cd or Pb increased levels of PCs in a concentration-, tissue-, and time-dependent manner. Cadmium was more effective that Pb in increasing PCs production. Compared with the effects of Cd or Pb alone on the production of PCs, the combination of Cd and Pb acted synergistically, resulting in an enhanced production of PCs. Cadmium also stimulated GSH production in a concentration-, tissue-, and time-dependent manner. However, Pb had no obvious effects on GSH levels. The combination of Pb and Cd antagonized GSH production over the course of the growth period. The results of the present study suggest that metal interactions should be considered in the application of PCs and GSH as potential biomarkers for the evaluation of metal toxicity.     

Effects of Enhanced UV-B Radiation on the Activity and Expression of Alternative Oxidase in Red Kidney Bean Leaves

An increase in ultraviolet （UV） B radiation on the earth＇s surface is a feature of current global climate changes. It has been reported that alternative oxidase （AOX） may have a protective role against oxidative stress induced by environmental stresses, such as UV-B. To better understand the characteristic tolerance of plants to UV-B radiation, the effects of enhanced UV-B radiation on the activity and expression of AOX in red kidney bean （Phaseolus vulgaris） leaves were investigated in the present study. The results show that the total respiration rate and AOX activity in red kidney bean leaves increased significantly during treatment with enhanced UV-B. However, cytochrome oxidase （COX） activity did not change significantly. The H2O2 content was also markedly increased and reached a maximum of 4.45 mmol·L^-1·g^-1 DW （dry weight） at 24 h of UV-B treatment, before dropping rapidly. Both alternative pathway content and alternative pathway activity were increased in the presence of exogenous H2O2. Immunoblotting analysis with anti-AOX monoclonal antibody revealed that expression of the AOX protein increased in red kidney bean leaves under enhanced UV-B radiation, reaching a peak at 72 h. In addition, AOX expression in red kidney bean leaves was induced by exogenous H2O2. These data indicate that the increase in AOX activity in red kidney bean leaves under enhanced UV-B radiation was mainly due to H2O2-induced AOX expression.