Overexpression of a <Emphasis Type="Italic">Miscanthus sacchariflorus</Emphasis> yellow stripe-like transporter <Emphasis Type="Italic">MsYSL1</Emphasis> enhances resistance of <Emphasis Type="Italic">Arabidopsis</Emphasis> to cadmium by mediating metal ion reallocation

The yellow stripe-like (YSL) family of transporters mediates the uptake, translocation, and distribution of various mineral elements in vivo by transferring metal ions chelated with phytosiderophore or nicotianamine (NA). However, little is known about the roles of the YSL genes against cadmium in planta. In this study, we first cloned and characterized a vital member of the YSL gene family, MsYSL1, from the bioenergy plant Miscanthus sacchariflorus. MsYSL1 localized in the plasma membrane and was widely expressed throughout the whole seedling with the highest expression level in the stem. In addition, its expression in the root was stimulated by excess manganese (Mn), cadmium (Cd), and lead, and a shortage of iron (Fe), zinc (Zn), and copper. Functional complementation in yeast indicated that MsYSL1 showed transport activity for Fe(II)–NA and Zn–NA, but not for Cd–NA. Although they exhibited no significant differences versus the wild type under normal cultivation conditions, MsYSL1-overexpressing Arabidopsis lines displayed a higher resistance to Cd accompanied by longer root lengths, lower Cd, Zn, and Mn levels in roots, and higher Cd, Fe, and Mn translocation ratios under Cd stress. Moreover, genes related to NA synthesis, metal translocation, long-distance transport, and Cd exclusion were highly induced in transgenic lines under Cd stress. Thus, MsYSL1 may be an essential transporter for diverse metal–NAs to participate in the Cd detoxification by mediating the reallocation of other metal ions.     

Variation in cadmium accumulation and translocation among peanut cultivars as affected by iron deficiency

Purpose

The current study aimed to test the hypothesis that the variations in shoot Cd accumulation among peanut cultivars was ascribed to the difference in capacity of competition with Fe transport, xylem loading and transpiration.

Methods

A hydroponics experiment was conducted to determine the plant biomass, gas exchange, and Cd accumulation in Fe-sufficient or -deficient plants of 12 peanut cultivars, at low Cd level (0.2 μM CdCl₂).

Results

Peanut varied among cultivars in morpho-physiological response to Cd stress as well as Cd accumulation, translocation and distribution. Qishan 208 and Xvhua 13 showed a higher capacity for accumulating Cd in their shoots. Fe deficiency increased the concentration and amount of Cd in plant organs, but decreased TF _{root to shoot} and TF _{root to stem}, while TF _{stem to leaf} remained unaffected. Fe deficiency-induced increase rates of Cd concentration and total Cd amount in roots and leaves were negatively correlated with the values in Fe-sufficient plants. Transpiration rate was positively correlated with leaf Cd concentration, TF _{root to shoot}, TF _{root to stem} and TF _{stem to leaf}.

Conclusions

The difference in shoot Cd concentration among peanut cultivars was mainly ascribed to the difference in Fe transport system, xylem loading capacity and transpiration.     

Mutation of mpk6 enhances cadmium tolerance in Arabidopsis plants by alleviating oxidative stress

Background and aims

Cadmium (Cd) could activate activity of mitogen-activated protein kinase MPK6 in plants. In this study, we investigated the role of MPK6 in mediating Cd toxicity in plants.

Methods

The wild type Arabidopsis plants (WT) and the mpk6-2 mutants were subjected either 0 (Control) or 10 μM Cd treatment. Kinase activity of MPK6, nitric oxide (NO) level, Cd concentration, and oxidative stress were measured.

Results

In WT plants, Cd exposure rapidly stimulated kinase activity of MPK6. However, upon Cd exposure, mpk6-2 showed better growth than the WT. Although Cd-induced production of NO in roots was greater in WT than in mpk6-2, there was no difference in Cd concentration between the two plants. Nevertheless, the Cd-induced hydroperoxide burst, lipid peroxidation and loss of membrane integrity, were all more severe in the WT than in mpk6-2. Foliar applications of antioxidant ascorbic acid, vigorously improved the growth of both the WT and mpk6-2 under Cd exposure. Thereby the growth difference between these two plants was minimized.

Conclusions

Mutation of mpk6 enhances Cd tolerance in plants by alleviating oxidative stress, but did not affect cadmium accumulation in plants.     

Predictability of the Zn and Cd phytoextraction efficiency of a Salix smithiana clone by DGT and conventional bioavailability assays

Aims

Phytomanagement of metal-polluted soils requires information on plant responses to metal availability in soil, but the predictability of metal accumulation in plant shoots and/or roots may be limited by metal toxicity and inherent shortfalls of the bioavailability assays.

Methods

We measured the uptake of Cd and Zn in a Salix smithiana clone grown in a pot experiment on soils with different characteristics and metal availabilities, determined by conventional soil single extractions (0.05 M Na₂-EDTA and 1 M NH₄NO₃), soil solution obtained by centrifugation, and diffusive gradients in thin films (DGT). The Cd and Zn phytoavailability after a 2-year phytoextraction by willow was assessed by metal accumulation in the straw of the following barley culture.

Results

The phytoextraction efficiency was largest on a moderately polluted acid soil. Biomass and shoot Zn concentrations of S. smithiana were better predicted by DGT-measured Zn concentrations in soil solution (C _DGT) than by Zn concentrations in the soil solution and extractable soil fractions. The weaker correlation for Cd in shoots may be related to relative Cd enrichment in the plant tissues. The metal accumulation in barley straw was unaffected or increased after a 2-year phytoextraction.

Conclusions

The shoot Zn and Cd removal of the tested Salix clone can be predicted by C _DGT concentrations and is highest on either calcareous or moderately polluted acid soils. Single extraction with NH₄NO₃ and the C _DGT value of Cd were not able to predict shoot Cd removal on the tested soils. Only shoot removal of Zn was predicted fairly well by the C _DGT value.     

Metallothioneins BcMT1 and BcMT2 from Brassica campestris enhance tolerance to cadmium and copper and decrease production of reactive oxygen species in Arabidopsis thaliana

Aims

Metallothioneins are cysteine-rich, metal-binding proteins, but their exact functions are not fully understood. In this study, we isolated two metallothionein genes, BcMT1 and BcMT2 from Brassica campestris to increase our understanding of metal tolerance mechanisms in Brassica plants.

Methods

Semi-quantitative RT-PCR was used to analyze expression of the two BcMTs genes. BcMT1 and BcMT2 were ectopically expressed in Arabidopsis thaliana. Quantitative real-time RT-PCR and GUS-staining method were used to select transgenic Arabidopsis plants. Cd and Cu concentrations were analyzed by flame atomic absorption spectrometry. Histochemical detection of H₂O₂ and O₂ ^?? were conducted by 3,3-diaminobenzidine and nitroblue tetrazoliu-staining methods.

Results

BcMT1 is expressed predominantly in roots, whereas BcMT2 is expressed mainly in leaves of B. campestris. Expression of BcMT1 was induced by both Cd and Cu, but expression of BcMT2 was enhanced only by Cd. Ectopic expression of BcMT1 and BcMT2 in Arabidopsis thaliana enhanced the tolerance to Cd and Cu and increased the Cu concentration in the shoots of the transgenic plants. Transgenic Arabidopsis accumulated less reactive oxygen species (ROS) than wild-type plants.

Conclusions

BcMT1 and BcMT2 increased Cd and Cu tolerance in transgenic Arabidopsis, and decreased production of Cd- and Cu-induced ROS, thereby protecting plants from oxidative damage.     

Physiological responses and antioxidant enzyme changes in <Emphasis Type="Italic">Sulla coronaria</Emphasis> inoculated by cadmium resistant bacteria

Plant growth promoting bacteria (PGPB) may help to reduce the toxicity of heavy metals on plants growing in polluted soils. In this work, Sulla coronaria inoculated with four Cd resistant bacteria (two Pseudomonas spp. and two Rhizobium sullae) were cultivated in hydroponic conditions treated by Cd; long time treatment 50 µM CdCl₂ for 30 days and short time treatment; 100 µM CdCl₂ for 7 days. Results showed that inoculation with Cd resistant PGPB enhanced plant biomass, thus shoot and root dry weights of control plants were enhanced by 148 and 35% respectively after 7 days. Co-inoculation of plants treated with 50 and 100 µM Cd increased plant biomasses as compared to Cd-treated and uninoculated plants. Cadmium treatment induced lipid peroxidation in plant tissues measured through MDA content in short 7 days 100 µM treatment. Antioxidant enzyme studies showed that inoculation of control plants enhanced APX, SOD and CAT activities after 30 days in shoots and SOD, APX, SOD, GPOX in roots. Application of 50 µM CdCl₂ stimulated all enzymes in shoots and decreased SOD and CAT activities in roots. Moreover, 100 µM of CdCl₂ increased SOD, APX, CAT and GPOX activities in shoots and increased significantly CAT activity in roots. Metal accumulation depended on Cd concentration, plant organ and time of treatment. Furthermore, the inoculation enhanced Cd uptake in roots by 20% in all treatments. The cultivation of this symbiosis in Cd contaminated soil or in heavy metal hydroponically treated medium, showed that inoculation improved plant biomass and increased Cd uptake especially in roots. Therefore, the present study established that co-inoculation of S. coronaria by a specific consortium of heavy metal resistant PGPB formed a symbiotic system useful for soil phytostabilization.     

The role of rice phenolics efflux transporter in solubilizing apoplasmic iron

Iron (Fe) is an essential micronutrient for plants whose deficiency presents a major worldwide agricultural problem. Moreover, Fe is not easily available in neutral to alkaline soils, rendering plants deficient in Fe despite its abundance. Plants secrete phenolics, such as protocatechuic acid (PCA) and caffeic acid (CA), to take up and utilize apoplasmic precipitated Fe, but despite the rapid progress in understanding cellular and subcellular Fe transport, the molecular mechanisms of phenolics synthesis and secretion are not clear. Recently, we isolated and characterized a phenolics efflux transporter in rice by characterizing a mutant in which the amount of PCA and CA in the xylem sap was dramatically reduced, which we hence named phenolics efflux zero 1 (pez1). PEZ1 is a plasma membrane protein that transports PCA when expressed in Xenopus laevis oocytes, and characterization of PEZ1 knockdown and overexpressing plants revealed that it plays an essential role in solubilizing precipitated apoplasmic Fe. The identification of PEZ1 will increase our understanding of apoplasmic Fe solubilization as well as promote research on phenolics efflux mechanisms in different organisms.Key words: iron, Oryza sativa, phenolics transport, protocatechuic acid, xylem sapAlthough mineral soils contain over 6% iron (Fe),¹ it predominantly exists as Fe(III) chelates, and plants ultimately cannot absorb Fe under various physiological conditions such as high soil pH in alkaline soils.² Thus, plants growing in high-pH soils are not very efficient in developing and stabilizing chlorophyll, resulting in the yellowing of leaves, poor growth and reduced yield. Plants, however, have developed sophisticated mechanisms to take up the small amount of soluble Fe. Non-graminaceous plants release protons, secrete phenolics, reduce Fe(III), and finally, take up Fe²⁺.^3–5 Once Fe is solubilized, Fe(III) is reduced to Fe²⁺ by a membrane-bound Fe(III) reductase oxidase.⁶ Then Fe²⁺ is transported into the root by an iron-regulated transporter (IRT1). In contrast, graminaceous plants rely on an Fe(III) chelation system through the secretion of mugineic acid (MA) family phytosiderophores.^3,7,8 The MAs are secreted to the rhizosphere through TOM1 ⁹ and then they chelate Fe(III); the resulting Fe-MA complex is transported by the Yellow Stripe family transporters (OsYSL15 in the case of rice¹⁰). Rice plants also have the ability to take up Fe²⁺ through the OsIRT1 transporter.¹¹In plants, Fe uptake from the apoplasm is well documented at the molecular level, with the exception of phenolics synthesis and efflux. Phenolics, such as protocatechuic acid (PCA), are reported to chelate Fe(III) solubilization and reduce it to Fe²⁺ in vitro.¹² Moreover, removing the secreted phenolics in hydroponic culture solution triggers Fe deficiency responses in roots by inhibiting the solubilization and utilization of apoplasmic Fe.¹³ In this manner, phenolics play a major role in Fe solubilization, besides which PCA and other phenolics play a diverse role in biological systems, such as acting as antioxidants and free radical scavengers, and in nitric oxide synthase.^14–17 Phenolics are also converted to lignin and suberin through the action of peroxidases.² The activity of peroxidases, as well as the formation of lignin, decreases under Fe deficient conditions.^2,18 As suberin plays an important role in controlling the movement of solutes,¹⁹ the role of phenolics in controlling water and mineral transport cannot be overlooked. Thus, understanding the molecular mechanism of phenolics efflux transport is crucial for developing strategies to mitigate widespread Fe deficiency.PEZ1 was isolated in an effort to characterize T-DNA mutants for genes regulated by cadmium (Cd). PEZ1 belongs to the multidrug and toxic compound extrusion transporter family whose members transport small organic compounds.²⁰ The substrates of PEZ1 were identified by analyzing liquid chromatography/mass spectrometry data profiles of the xylem sap of pez1-1 and pez1-2 mutants. The data indicated that PEZ1 transports PCA and caffeic acid (CA). Furthermore, PEZ1 transported radiolabeled PCA when expressed in Xenopus laevis oocytes. PEZ1 localizes to the plasma membrane in rice root cells, as well as in rice root hairs and onion epidermal cells. The pez1-2 mutant accumulated more Fe in the roots, but not in the leaves, compared to wild-type (WT) plants; the differences were greater in the presence of Cd, while no difference was observed in the accumulation of other metals. No significant difference was observed in zinc, manganese (Mn), and copper concentration between WT and pez1-2, in both the roots and shoots, with or without Cd. Fe concentration in the xylem sap was lower than in the WT, while no difference was observed for xylem Cd and Mn. Significant differences in the localization of insoluble Fe were observed when leaf samples were stained with Perl''s solution to examine the localization of Fe. These results suggested a clear role of PEZ1 in solubilizing precipitated apoplasmic Fe.²¹Secretion of excess PCA strongly solubilizes Fe precipitated in the stele, leading to symptoms of Fe excess. The analysis of PEZ1 overexpression lines confirmed this hypothesis. PEZ1 overexpression lines accumulated higher amounts of Fe in roots and leaves owing to the high solubilization of precipitated apoplasmic stele Fe, and as a result, the growth of these lines was severely restricted. In contrast, PEZ1 overexpression lines grew better than the WT in calcareous soil, showing that in these lines, PCA-solubilized Fe is available under Fe-limiting conditions.The expression of PEZ1 is regulated by Cd, and both of the PEZ1 knockdown mutants accumulated higher Cd amounts in leaves and seeds when grown in soil, without compromising morphological or physiological characteristics, like the SPAD value, leaf dry weight, yield, and the concentration of other metals in seeds. Why pez1 accumulates Cd is not clear. PCA has a lower affinity for Cd compared to glutathione, and PEZ1 does not transport Cd.²¹ Cd is partly transported through the Fe uptake system in plants.^22–26 Thus, in pez1, Cd accumulation seems to be triggered by the upregulation of OsIRT1. OsIRT1 localization in the phloem, its substrate specificity, and increased expression in pez1 mutants suggests that Fe and Cd uptake and translocation in pez1 mutants could be enhanced through OsIRT1,¹¹ and that an increased Cd accumulation in pez1 mutants may be due to the increase in OsIRT activity in a decreased Fe environment in which Cd will have reduced competition. PEZ1 localizes to the stele in root cells. The localization of different genes involved in Fe transport is summarized in Figure 1.Open in a separate windowFigure 1Tissue-specific expression of Fe homeostasis-related genes in rice root.In short, phenolics secretion affects Fe acquisition in rice. Reduced secretion of PCA in the pez1-2 mutant impairs the solubilization of precipitated apoplasmic Fe in the stele, and thus, the low availability of Fe leads to the induction of OsIRT1. As PEZ1 and OsIRT1 co-localize in the stele, the PCA secretion may complement Fe²⁺ uptake by OsIRT1 and seems to be an integral part of the Fe²⁺ uptake system in rice (Fig. 2). In contrast, the increase in phenolics secretion in PEZ1-overexpressing plants increases the solubilization of apoplasmic Fe, and plants showed an increased tolerance to Fe deficiency in alkaline soils. The identification of PEZ1 is an important step that helps in better understanding the solubilization of apoplasmic Fe and will generate research on phenolics efflux mechanisms in other plants.Open in a separate windowFigure 2Model of Fe and Cd uptake mechanisms in rice xylem. P.M., plasma membrane.     

The HvNramp5 Transporter Mediates Uptake of Cadmium and Manganese,But Not Iron

The Natural Resistance Associated Macrophage Protein (Nramp) represents a transporter family for metal ions in all organisms. Here, we functionally characterized a member of Nramp family in barley (Hordeum vulgare), HvNramp5. This member showed different expression patterns, transport substrate specificity, and cellular localization from its close homolog in rice (Oryza sativa), OsNramp5, although HvNramp5 was also localized to the plasma membrane. HvNramp5 was mainly expressed in the roots and its expression was not affected by Cd and deficiency of Zn, Cu, and Mn, but slightly up-regulated by Fe deficiency. Spatial expression analysis showed that the expression of HvNramp5 was higher in the root tips than that in the basal root regions. Furthermore, analysis with laser microdissection revealed higher expression of HvNramp5 in the outer root cell layers. HvNramp5 showed transport activity for both Mn²⁺ and Cd²⁺, but not for Fe²⁺ when expressed in yeast. Immunostaining with a HvNramp5 antibody showed that this protein was localized in the root epidermal cells without polarity. Knockdown of HvNramp5 in barley resulted in a significant reduction in the seedling growth at low Mn supply, but this reduction was rescued at high Mn supply. The concentration of Mn and Cd, but not other metals including Cu, Zn, and Fe, was decreased in both the roots and shoots of knockdown lines compared with the wild-type barley. These results indicate that HvNramp5 is a transporter required for uptake of Mn and Cd, but not for Fe, and that barley has a distinct uptake system from rice.Transport of mineral elements from soil to different organs and tissues of plants requires different types of transporters (Hall and Williams, 2003; Nevo and Nelson, 2006; Yokosho et al., 2009; Olsen and Palmgren, 2014; Sasaki et al., 2016), which include the zinc-regulated transporters, iron-regulated transporter-like protein family; the natural resistance-associated macrophage protein (Nramp) family of transporters; the multidrug and toxic compound extrusion protein transporters; the heavy metal ATPase transporters; the oligopeptide transporters family; the ATP-binding cassette family of transporters; and the cation-diffusion facilitator family of transporters. Among them, Nramp represents a transporter family for metal ion in all organisms including bacteria, animals, and plants (Curie et al., 2000; Nevo and Nelson, 2006). Some members of this family in plants have been functionally characterized, especially in model plants such as Arabidopsis (Arabidopsis thaliana) and rice (Oryza sativa). In Arabidopsis, there are six members of Nramp transporter proteins. AtNramp1 is localized to the plasma membrane of root cells and functions as a high-affinity transporter for Mn uptake (Cailliatte et al., 2010). Both AtNramp3 and AtNramp4 are localized to the tonoplast and play redundant roles in Fe, exporting from the vacuole during seed germination and in Mn homeostasis at the adult stage (Thomine et al., 2000; Lanquar et al., 2005, 2010). AtNramp6 is targeted to a vesicular-shaped endomembrane compartment and is implicated in the distribution or availability of Cd within cells (Cailliatte et al., 2009). However, the function of AtNramp2 and AtNramp5 has not been characterized.On the other hand, there are seven members of Nramp transporter family in the rice genome, of which four have been functionally characterized. They all are localized to the plasma membrane but show different roles. OsNramp1 shows transport activity for Fe and Cd in yeast and is proposed to be involved in Cd accumulation (Takahashi et al., 2011). OsNramp3 is localized at the vascular tissues of nodes and plays an important role in distribution of Mn, but not Fe and Cd (Yamaji et al., 2013). On the other hand, OsNrat1 (OsNramp4) transports trivalent Al ion (Xia et al., 2010) and is required for high Al tolerance in rice roots. Finally, OsNramp5 functions as a major transporter responsible for root Mn and Cd uptake (Ishimaru et al., 2012; Ishikawa et al., 2012; Sasaki et al., 2012). However, the function of OsNramp2, OsNramp6, and OsNramp7 is unknown.In addition to Nramp members characterized in rice and Arabidopsis, some members in other plant species have also been characterized. For example, a soybean (Glycine max) Nramp transporter, GmDMT1 is implicated in the ferrous iron transport (Kaiser et al., 2003). Nramp1 isolated from Noccaea caerulescens, a Zn/Cd hyperaccumulator, is involved in the influx of Cd across the endodermal plasma membrane and plays a key role in Cd flux into the stele and root-to-shoot Cd transport (Milner et al., 2014). In Malus baccata, Nramp1 is capable of mediating the distribution of ions as well as transport of Fe, Mn, and Cd (Xiao et al., 2008). Besides, Nramp1 and Nramp3 in tomato (Solanum lycopersicum) have also been suggested to be involved in Mn transport (Bereczky et al., 2003). When NcNramp3 and NcNramp4 from Noccaea caerulescens were expressed in yeast, NcNramp3 transported Fe, Mn, and Cd, while NcNramp4 also transported Zn in addition to Fe, Mn, and Cd (Oomen et al., 2009). However, Nramp4 isolated from Thlaspi japonicum, a Ni hyperaccumulator, showed transport activity for Ni but not for Zn, Cd, or Mn in yeast (Mizuno et al., 2005). These findings indicate that Nramp members have a diverse role in metal transport in plants.Barley (Hordeum vulgare) is the fourth most important cereal crop in the world; however, less progress has been made in understating of molecular mechanisms on mineral element transport in barley due to its large genome size. For example, no Nramp members in barley have been functionally characterized so far. In this study, we first isolated barley Nramp member, HvNramp5, which is a close homolog of rice OsNramp5. Detailed functional analysis revealed that HvNramp5 is involved in the uptake of both Mn and Cd, but not of Fe in barley roots. Furthermore, we found that different from OsNramp5, HvNramp5 showed a distinct pattern in the gene expression, cellular localization, and transport substrate.     

Arabidopsis desaturase 2 gene is involved in the regulation of cadmium and lead resistance

Aims

It was shown previously that Arabidopsis (Arabidopsis thaliana) desaturase 2 (ADS2) cDNA was isolated and it was shown that the expression of ADS2 was organ-dependent and up-regulated by low temperature. However, little is known about the role of ADS2 gene in heavy metal resistance in plants. In this study, we showed that ADS2 gene is involved in the regulation of cadmium (Cd) and lead (Pb) resistance.

Methods

For heavy metal resistance tests, seeds were germinated and grown on 1/2 MS media supplemented with the indicated concentrations of metal ions. To quantify root length, plants were grown vertically in plates. For heavy metal treatments, two-week old wild-type seedlings grown on MS media were treated with cadmium (Cd) or lead (Pb) for 24 h, and then sampled for metal content measurement and qPCR analysis.

Results

ADS2 was strongly repressed by Cd(II), and ads2-1 mutant plants showed increased Cd(II) resistance. A lower Cd content was detected in ads2-1 plants than in wild-type plants subjected to Cd(II) treatment, which was associated with activation in expression of AtPDR8 gene, a pump excluding Cd(II) and/or Cd(II)-containing toxic compounds from the cytoplasm, suggesting that ADS2-mediated Cd(II) resistance is AtPDR8 dependent. We also found that ads2-1 plants showed increased Pb(II) sensitivity, and ADS2 was strongly repressed by hydrogen peroxide (H₂O₂) but not by Pb(II). The ads2-1 mutant showed increased sensitivity to oxidative stresses mediated by H₂O₂ and paraquat, and higher levels of H₂O₂ accumulation were observed in leaves of ads2-1 plants than those of wild-type plants when subjected to Pb(II) and H₂O₂, indicating that ADS2 mediates Pb(II) resistance indirectly by impaired ROS scavenging.

Conclusions

ADS2 gene mediates Cd(II) and Pb(II) resistance, at least in part, through two distinct mechanisms, an AtPDR8-dependent mechanism and a ROS detoxification system-mediated mechanism, respectively.     

Mechanistic studies of perfluorooctane sulfonate, perfluorooctanoic acid uptake by maize (Zea mays L. cv. TY2)

Aims

There is a need to predict trace metal concentration in plant organs at given development stages. The aim of this work was to describe the Cd hyperaccumulation kinetics in the different plant organs, throughout the complete cultivation cycle, independently of a possible soil effect.

Methods

Plants of Noccaea caerulescens were exposed in aeroponics to three constantly low Cd concentrations and harvested at 6 to 11 dates, until siliquae formation.

Results

Dry matter allocation between roots and shoots was constant over time and exposure concentrations, as well as Cd allocation. However 86 % of the Cd taken up was allocated to the shoots. Senescent rosette leaves showed similar Cd concentrations to the living ones, suggesting no redistribution from old to young organs. The Cd root influx was proportional to the exposure concentration and constant over time, indicating that plant development had no effect on this. The bio-concentration factor (BCF), i.e. [Cd]_plant/[Cd²⁺]_solution for the whole plant, roots or shoots was independent of the exposure concentration and of the plant stage.

Conclusions

Cadmium uptake in a given plant part could therefore be predicted at any plant stage by multiplying the plant part dry matter by the corresponding BCF and the Cd²⁺ concentration in the exposure solution.     

Overexpression of barley nicotianamine synthase 1 confers tolerance in the sweet potato to iron deficiency in calcareous soil

Background and aims

Iron (Fe) is an essential micronutrient for all higher organisms. Fe is sparingly available in calcareous soils and Fe deficiency is a major agricultural problem worldwide. Nicotianamine (NA) is a metal chelator involved in metal translocation in plants. Sweet potato is an attractive crop that can grow in poor soil and thus is useful for planting in uncultivated soil. In addition, the sweet potato has recently been suggested as a source of bioethanol. Our aim is to increase NA concentration in sweet potato to ameliorate Fe deficiency.

Method

Sweet potato plants expressing the barley NA synthase 1 (HvNAS1) gene under the control of CaMV 35S promoter were produced by Agrobacterium-mediated transformation.

Results

The transgenic sweet potato exhibited tolerance to low Fe availability when grown in calcareous soil. The level of tolerance to low Fe availability was positively correlated with the HvNAS1 expression level. The NA concentration of the transgenic sweet potato leaves was up to 7.9-fold greater than that of the non-transgenic (NT) plant leaves. Furthermore, the Fe and zinc concentrations were 3- and 2.9-fold greater, respectively, in transgenic sweet potato than in NT plant leaves.

Conclusions

Our results suggest that increasing the NA concentration of sweet potato by overexpression of HvNAS1 could significantly improve agricultural productivity and energy source.

     

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.     

The crucial role of roots in increased cadmium-tolerance and Cd-accumulation in the pea mutant <Emphasis Type="Italic">SGECd</Emphasis><Superscript><Emphasis Type="Italic">t</Emphasis></Superscript>

Elucidation of mechanisms underlying plant tolerance to cadmium, a widespread toxic soil pollutant, and accumulation of Cd in plants are urgent tasks. For this purposes, the pea (Pisum sativum L.) mutant SGECd^t (obtained by treatment of the laboratory pea line SGE with ethylmethane sulfonate) was reciprocally grafted with the parental line SGE, and four scion/rootstock combinations were obtained: SGE/SGE, SGECd^t/SGECd^t, SGE/SGECd^t, and SGECd^t/SGE. They were grown in hydroponics in the presence of 1 μM CdCl₂ for 30 d. The SGE and SGECd^t scions on the SGECd^t rootstock had a higher root and shoot biomass and an elevated root and shoot Cd content compared with the grafts having SGE rootstock. Only the grafts with the SGE rootstock showed chlorosis and roots demonstrating symptoms of Cd toxicity. The content of nutrient elements in roots (Fe, K, Mg, Mn, Na, P, and Zn) was higher in the grafts having the SGECd^t rootstock, and three elements, namely Ca, Fe, and Mn, were efficiently transported by the SGECd^t root to the shoot of these grafts. The content of other measured elements (K, Mg, Na, P, and Zn) was similar in the root and shoot in all the grafts. Then, the non-grafted plants were grown in the presence of Cd and subjected to deficit or excess concentrations of Ca, Fe, or Mn. Exclusion of these elements from the nutrient solution retained or increased differences between SGE and SGECd^t in growth response to Cd toxicity, whereas excess of Ca, Fe, or Mn decreased or eliminated such differences. The obtained results assign a principal role of roots to realizing the increased Cd-tolerance and Cdaccumulation in the SGECd^t mutant. Efficient translocation of Ca, Fe, and Mn from roots to shoots appeared to counteract Cd toxicity, although Cd was actively taken up by roots and accumulated in shoots.     

Carbon mineralization in soil of roots from twenty crop species,as affected by their chemical composition and botanical family

Background and aims

Plant growth-promoting rhizobacteria (PGPR) have been widely studied for agricultural applications. One aim of this study was to isolate cadmium (Cd)-tolerant bacteria from nodules of Glycine max (L.) Merr. grown in heavy metal-contaminated soil in southwest of China. The plant growth-promoting (PGP) traits and the effects of the isolate on plant growth and Cd uptake by legume and non-legume plants in Cd-polluted soil were investigated.

Methods

Cd-tolerant bacteria were isolated by selective media. The isolates were identified by 16S rRNA gene and phylogenetic analysis. The PGR traits of the isolates were evaluated in vitro. Cd in soil and plant samples was determined by ICP-MS.

Results

One of the most Cd-tolerant bacteria simultaneously exhibited several PGP traits. Inoculation with the PGPR strain had positive impacts on contents of photosynthesis pigments and mineral nutrients (Fe or Mg) in plant leaves. The shoot dry weights of Lolium multiflorum Lam. increased significantly compared to uninoculated control. Furthermore, inoculation with the PGPR strain increased the Cd concentrations in root of L. multiflorum Lam. and extractable Cd concentrations in the rhizosphere, while the Cd concentrations in root and shoot of G. max (L.) Merr. significantly decreased.

Conclusions

This study indicates that inoculation with Cd-tolerant PGPR can alleviate Cd toxicity to the plants, increase Cd accumulation in L. multiflorum Lam. by enhancing Cd availability in soils and plant biomass, but decrease Cd accumulation in G. max (L.) Merr. by increasing Fe availability, thus highlighting new insight into the exploration of PGPR on Cd-contaminated soil.     

Integration of metal chemical forms and subcellular partitioning to understand metal toxicity in two lettuce (Lactuca sativa L.) cultivars

Aims

Metal chemical forms and subcellular partitioning model (SPM) in organisms can provide valuable insights into metal toxicity.

Methods

Two cultivars of lettuce (Lactuca sativa L.) were grown in Cd and Cu contaminated soils and chemical forms and subcellular distribution of Cd and Cu within the lettuce shoots were determined.

Results

Examination of the inhibition of superoxide dismutase (SOD) and catalase (CAT) activities, as well as the production of H₂O₂ showed that Lactuca sativa L. var. longifolia is more sensitive to metal-stress than is Lactuca sativa L. var. crispa. In L. crispa, the majority of accumulated Cd was in the pectate- and protein-integrated forms (53.7–62.9 %), while in L. longifolia, a higher proportion of the Cd was in the water soluble forms (33.0–39.2 %) and in the organelles fraction – these forms being potentially associated with toxicity. The chemically-based chemical form approach agreed closely with independent biologically-based SPM, as demonstrated by their significant linear relationships.

Conclusions

This study provides a first step towards the integration of chemical form approach and SPM into a common mechanistic framework, which is important for predicting the likelihood of toxic effects of metals in the environment of interest.     

Impact of active transport and transpiration on nickel and cadmium accumulation in the leaves of the Ni-hyperaccumulator Leptoplax emarginata: a biophysical approach

Aims

The primary aim of this study was to investigate the impact of active nickel and cadmium transport, transpiration and shoot biomass production on Ni and Cd accumulation in the leaves of the Ni-hyperaccumulator Leptoplax emarginata. A secondary objective was to observe the effects of various concentrations of nickel and cadmium in solutions on the plant growth and ecophysiological characteristics of these plants. Finally, the study sought to identify possible nickel and cadmium concentration gradients in solution as a function of the root distance.

Methods

The Intact Plant Transpiration Stream Concentration Factor (TSCF=xylem/solution solute concentration ratio) was determined for both Ni and Cd and for the selected intact transpiring Ni-hyperaccumulator Leptoplax emarginata, cultivated on two contrasting fertilized and Ni-Cd-contaminated sandy porous media (rhizotrons with central root compartments, linked to Mariotte tubes operated at ?1?kPa). IPTSCF_Ni and IPTSCF_Cd were calculated as the ratios between the hyperaccumulator plant’s nickel or cadmium mass in the leaves and the nickel or cadmium concentration in solution by the volume of water transpired during the period of culture. Plant growth characteristics and gas exchanges were also recorded.

Results

IPTSCF values were much greater than 1 (IPTSCF_Ni?=?5.2?±?0.9 and IPTSCF_Cd?=?4.4?±?0.6) whatever the amount of available Ni and Cd. This characterized a predominantly active plant metal uptake. Moreover, biological regulation was reported: plant growth and transpiration were significantly lower for hyperaccumulator plants cultivated in sand which was rich in available Ni and Cd, than for hyperaccumulator plants cultivated in topsoil, poor in available Ni and Cd. In the soil rhizosphere, capillary flow was related to transpiration and a depletion pattern was developed for Ni and sometimes for Cd.

Conclusions

Overall, the Intact Plant Transpiration Stream Concentration Factor appeared to be a relevant metal bioconcentration factor taking into account the predominant type of metal transport from roots to leaves, plant growth and transpiration coupling and metal availability. IPTSCF_Ni and IPTSCF_Cd values were much greater than 1 and similar whatever the amount of available Ni and Cd. This characterized a predominantly active plant combining Ni and Cd uptake and biological regulations dependent of the Ni and Cd concentrations in solution.     

Increased gibberellin contents contribute to accelerated growth and development of transgenic tobacco overexpressing a wheat ubiquitin gene

Key message

Overexpressing TaUb2 promoted stem growth and resulted in early flowering in transgenic tobacco plants. Ubiquitin are involved in the production, metabolism and proper function of gibberellin.

Abstract

The ubiquitin–26S proteasome system (UPS), in which ubiquitin (Ub) functions as a marker, is a post-translational regulatory system that plays a prominent role in various biological processes. To investigate the impact of different Ub levels on plant growth and development, transgenic tobacco (Nicotiana tabacum L.) plants were engineered to express an Ub gene (TaUb2) from wheat (Triticum aestivum L.) under the control of cauliflower mosaic virus 35S promoter. Transgenic tobacco plants overexpressing TaUb2 demonstrated an accelerated growth rate at early stage and an early flowering phenotype in development. The preceding expression of MADS-box genes also corresponded to the accelerated developmental phenotypes of the transgenic tobacco plants compared to that of wild-type (WT). Total gibberellin (GA) and active GA contents in transgenic tobacco plants were higher than those in WT at the corresponding developmental stages, and some GA metabolism genes were upregulated. Treatment with GA₃ conferred a similarly accelerated grown rate in WT plants to that of transgenic tobacco plants, while growth was inhibited when transgenic tobacco plants were treated with a GA biosynthesis inhibitor. Thus, the results suggest that Ub are involved in the production, metabolism and proper function of GA, which is important in the regulation of plant growth and development.     

Endogenous jasmonic and salicylic acids levels in the Cd-hyperaccumulator Noccaea (Thlaspi) praecox exposed to fungal infection and/or mechanical stress

Key message

Sensitivity to Erysiphe in Noccaea praecox with low metal supply is related to the failure in enhancing SA. Cadmium protects against fungal-infection by direct toxicity and/or enhanced fungal-induced JA signaling.

Abstract

Metal-based defense against biotic stress is an attractive hypothesis on evolutionary advantages of plant metal hyperaccumulation. Metals may compensate for a defect in biotic stress signaling in hyperaccumulators (metal-therapy) by either or both direct toxicity to pathogens and by metal-induced alternative signaling pathways. Jasmonic acid (JA) and salicylic acid (SA) are well-established components of stress signaling pathways. However, few studies evaluate the influence of metals on endogenous concentrations of these defense-related hormones. Even less data are available for metal hyperaccumulators. To further test the metal-therapy hypothesis we analyzed endogenous SA and JA concentrations in Noccaea praecox, a cadmium (Cd) hyperaccumulator. Plants treated or not with Cd, were exposed to mechanical wounding, expected to enhance JA signaling, and/or to infection by biotrophic fungus Erysiphe cruciferarum for triggering SA. JA and SA were analyzed in leaf extracts using LC–ESI(?)–MS/MS. Plants without Cd were more susceptible to fungal attack than plants receiving Cd. Cadmium alone tended to increase leaf SA but not JA. Either or both fungal attack and mechanical wounding decreased SA levels and enhanced JA in the Cd-rich leaves of plants exposed to Cd. High leaf Cd in N. praecox seems to hamper biotic-stress-induced SA, while triggering JA signaling in response to fungal attack and wounding. To the best of our knowledge, this is the first report on the endogenous JA and SA levels in a Cd-hyperaccumulator exposed to different biotic and abiotic stresses. Our results support the view of a defect in SA stress signaling in Cd hyperaccumulating N. praecox.     

Silencing of omega-5 gliadins in transgenic wheat eliminates a major source of environmental variability and improves dough mixing properties of flour

Background

Silicon (Si) application has been known to enhance the tolerance of plants against abiotic stresses. However, the protective mechanism of Si under heavy metals contamination is poorly understood. The aim of this study was to assess the role of Si in counteracting toxicity due to cadmium (Cd) and copper (Cu) in rice plants (Oryza sativa).

Results

Si significantly improved the growth and biomass of rice plants and reduced the toxic effects of Cd/Cu after different stress periods. Si treatment ameliorated root function and structure compared with non-treated rice plants, which suffered severe root damage. In the presence of Si, the Cd/Cu concentration was significantly lower in rice plants, and there was also a reduction in lipid peroxidation and fatty acid desaturation in plant tissues. The reduced uptake of metals in the roots modulated the signaling of phytohormones involved in responses to stress and host defense, such as abscisic acid, jasmonic acid, and salicylic acid. Furthermore, the low concentration of metals significantly down regulated the mRNA expression of enzymes encoding heavy metal transporters (OsHMA2 and OsHMA3) in Si-metal-treated rice plants. Genes responsible for Si transport (OsLSi1 and OsLSi2), showed a significant up-regulation of mRNA expression with Si treatment in rice plants.

Conclusion

The present study supports the active role of Si in the regulation of stresses from heavy metal exposure through changes in root morphology.