Roser Tolrà

Zinc hyperaccumulation in Thlaspi caerulescens. II. Influence on organic acids

Abstract The influence of different zinc (Zn) concentrations (1.5 to 1500 μM) on organic acid levels in roots and shoots of the Zn‐hyperaccumulator plant Thlaspi caerulescens was investigated. In shoots, malate was the most abundant organic acid (164 to 248 μmol/g f.w.), followed by citrate, succinate, and oxalate. A significant correlation between soluble Zn and both malate and oxalate was observed in shoots, but not in roots. In shoots, a significant correlation between inorganic cation equivalents and organic acid anion equivalents was found. These observations and the finding, that organic acid concentrations were high even under suboptimal Zn supply (1.5 μM) suggest that in T. caerulescens the high organic acid concentration in shoots is a constitutive property. The variation of the organic acid concentrations seem to be a consequence of the cation‐anion balance rather than a specific Zn tolerance mechanism. The constitutively high organic acid concentration may be responsible for the high Zn and iro...

Zinc hyperaccumulation in Thlaspi caerulescens. I. Influence on growth and mineral nutrition

Abstract Thlaspi caerulescens, a metallophyte that is able to accumulate up to 4% zinc (Zn) in leaf dry matter, has attracted much attention for its possible use in phytoremediation of metal contaminated soils. In the present study, the influence of Zn supply on mineral nutrition in T. caerulescens was investigated, in order to establish the extent to which growth stimulation by high Zn supply is related to changes in the levels of other essential nutrients. The plants were exposed to nutrient solutions containing 1.5, 100, 500, 750, 1000, or 1500 μM Zn. Zinc supply significantly influenced root and shoot concentrations of essential nutrients, but excepting Zn, the concentrations stayed within the range considered adequate for optimum growth in Brassicaceae crops. Best performance was achieved with the supply of 500 μM Zn. Growth stimulation by this treatment was accompanied by increased translocation of iron (Fe) from root to shoot and a significant correlation between shoot dry weight and Fe concentrati...

Determination of glucosinolates in rapeseed and Thlaspi caerulescens plants by liquid chromatography-atmospheric pressure chemical ionization mass spectrometry.

Liquid chromatography-atmospheric pressure chemical ionization mass spectrometry was used to identify glucosinolates in plant extracts. Optimization of the analytical conditions and the determination of the method detection limit was performed using commercial 2-propenylglucosinolate (sinigrin). Optimal values for the following parameters were determined: nebulization pressure, gas temperature, flux of drying gas, capillar voltage, corona current and fragmentor conditions. The method detection limit for sinigrin was 2.85 ng. For validation of the method the glucosinolates in reference material (rapeseed) from the Community Bureau of Reference Materials (BCR) were analyzed. The method was applied for the determination of glucosinolates in Thlaspi caerulescens plants.

Constitutive and aluminium-induced patterns of phenolic compounds in two maize varieties differing in aluminium tolerance.

Aluminium tolerance in maize is mainly due to more efficient Al exclusion. Nonetheless, even in tolerant varieties Al can gain access into the cells. Detoxification by binding to strong organic ligands should therefore play a role also in plants with high Al exclusion capacity. To test this hypothesis in this study the concentrations of soluble, free and bound, phenolics were analyzed in roots of two maize varieties differing in Al tolerance. Exposure for 24 h to 50 microM Al in nutrient solution strongly inhibited root elongation in the sensitive variety 16 x 36, but not in the Al-tolerant variety Cateto. Cateto accumulated about half the concentration of Al in roots than 16 x 36 (analysis performed after root desorption with citrate). Roots of Al-tolerant Cateto contained higher concentrations of caffeic acid, catechol and catechin than roots of the sensitive variety. Exposure to Al induced the accumulation of taxifolin in roots of both varieties. However, Al-tolerant Cateto accumulated about twice the concentration than Al-sensitive 16 x 36 of this pentahydroxyfavonol. The molar ratio for phenolics with catecholate groups to Al was about unity in roots of Cateto, while in those of 16 x 36 the ratio was ten times lower. Both the fact that these phenolics are strong ligands for Al and their high antioxidant and antiradical activity suggest that these compounds may provide protection against the Al fraction that is able to surpass the exclusion mechanisms operating in the tolerant maize variety.

Aluminium-induced changes in root epidermal cell patterning, a distinctive feature of hyperresistance to Al in Brachiaria decumbens

Brachiaria, a genus of forage grasses of African origin, is gaining considerable importance because of both its nutritional value and its high stress resistance. An extraordinary resistance to Al toxicity has been reported in B. decumbens. The mechanisms of this hyperresistance are still unknown. This study explores the localization of Al in two contrasting Brachiaria species, the hyperresistant B. decumbens and the less resistant B. brizantha. Scanning Electron Microscope/Energy Dispersive Spectrometry, confocal fluorescence microscopy and optical microscopy of lumogallion or morin-stained roots was performed. Species differences in Al resistance were evident at 32 μM Al(3+) activity in low ionic strength full nutrient solution containing Si. Roots of B. decumbens accumulated less Al than those of B. brizantha. Moreover, location and Al form seemed different. In B. decumbens Al accumulation was localized in hot spots of high Al concentrations. These sites with high Al accumulation mainly correspond to root hairs. B. brizantha exhibited a more even distribution of Al in cell walls of the root tip. Analysis of soluble phenolic substances in roots revealed species differences in response to Al. An Al-induced increase of chlorogenic acid concentrations was found in B. decumbens but not in B. brizantha. Taken together the results suggest a possible role for chlorogenic acid as a primer for changes in root epidermal cell patterning that may contribute to the Al hyperresistance in B. decumbens.

Amelioration of iron toxicity: A mechanism for aluminum-induced growth stimulation in tea plants

Tea plants (Camellia sinensis) are well adapted to acid soils with high Al availability. These plants not only accumulate high leaf Al concentrations, but also respond to Al with growth stimulation. Decreased oxidative stress has been associated with this effect. Why tea plants not exposed to Al suffer from oxidative stress has not been clarified. In this study, hydroponically grown tea plants treated with 0 to 300 μM Al were analyzed for growth, Al and Fe accumulation, and Al distribution by means of morin and hematoxylin staining. Roots of control plants stained black with hematoxylin. This indicates the formation of a Fe-hematoxylin complex. Young leaves of controls accumulated more than 1000 mg Fe kg(-1) dry weight. This concentration is above the Fe-toxicity threshold in most species. Supply of Al stimulated growth and reduced Fe uptake and transport. These results indicate that Al-induced growth stimulation might be due to alleviation of a latent Fe toxicity occurring in tea plants without Al supply.

Glucosinolate Profiles Change During the Life Cycle and Mycorrhizal Colonization in a Cd/Zn Hyperaccumulator Thlaspi praecox (Brassicaceae)

Thlaspi praecox Wulfen (Brassicaceae) is a perennial Cd/Zn hyperaccumulating plant species that forms functional arbuscular mycorrhizal (AM) symbiosis. Glucosinolates (GS) were studied in different organs of field-collected T. praecox at differing plant developmental stages. Additionally, AM colonization was recorded. Total GS concentrations and profiles of nine individual GS varied during the plant life cycle. Novel individual GS that were related to specific developmental phases, mainly to flowering and seed production, were identified. The highest total GS and sinalbin concentrations in rosette leaves were found in the vegetative phase, possibly contributing to protection of young, palatable leaves. The lowest were found in roots during the flowering and the seeding phases. Increased total GS concentrations in roots and enhanced aliphatic GS, especially glucobrassicanapin, in the senescence phase may protect roots from herbivory during winter and early spring. The presence of glucotropaeolin and the absence of glucobrassicanapin in the flowering phase coincided with peak AM colonization. This is the first report on GS profiles in an AM and metal-hyperaccumulating plant.

Molecular characterization of the citrate transporter gene TaMATE1 and expression analysis of upstream genes involved in organic acid transport under Al stress in bread wheat (Triticum aestivum)

In bread wheat, besides malate, the importance of citrate efflux for Al tolerance has also been reported. For better understanding the Al tolerance mechanism in bread wheat, here, we performed both a molecular characterization of the citrate transporter gene TaMATE1 and an investigation on the upstream variations in citrate and malate transporter genes. TaMATE1 belong to multidrug transporter protein family, which are located on the long arm of homoeologous group 4 chromosomes (TaMATE1-4A, TaMATE1-4B TaMATE1-4D). TaMATE1 homoeologues transcript expression study exhibited the preponderance of homoeologue TaMATE1-4B followed by TaMATE1-4D whereas homoeologue TaMATE1-4A seemed to be silenced. TaMATE1, particularly homoeologue TaMATE1-4B and TaALMT1 transcripts were much more expressed in the root apices than in shoots of Al tolerant genotype Barbela 7/72/92 under both control and Al stress conditions. In addition, in both tissues of Barbela 7/72/92, higher basal levels of these gene transcripts were observed than in Anahuac (Al sensitive). Noticeably, the presence of a transposon in the upstream of TaMATE1-4B in Barbela 7/72/92 seems to be responsible for its higher transcript expression where it may confer citrate efflux. Thus, promoter variations (transposon in TaMATE1-4B upstream and type VI promoter in TaALMT1) associated with higher basal transcript expression of TaMATE1-4B and TaALMT1 clearly show how different mechanisms for Al tolerance operate simultaneously in a single genotype. In conclusion, our results demonstrate that Barbela 7/72/92 has favorable alleles for these organic acids transporter genes which could be utilized through genomic assisted selection to develop improved cultivars for acidic soils.

A proteomic approach to the mechanisms underlying activation of aluminium resistance in roots of Urochloa decumbens

The mechanisms of extreme Al-resistance in Urochloa decumbens are not established. Full resistance expression requires a lag time of 72-96h and is preceded by a sensitive phase (24-48h) with Al-induced root growth inhibition. The aim here was to identify key processes of the activation phase of Al-resistance analysing both root exudates and comparative root proteome. Samples were taken after 0, 24 and 96h exposure to 0 or 200μM Al. Al-induced stimulation of citrate and oxalate efflux was limited to the sensitive phase. Only 11 proteins revealed Al-induced abundance differences; six were identified. After 24h, phenylalanine ammonium lyase (PAL), methionine synthase (MS), and deoxymugineic acid synthase (DMAS) decreased, while acid phosphatase (APase) abundance increased. Coincident with growth recovering, PAL and MS, but not DMAS, returned to initial levels. After 96h, γ‑carbonic anhydrase (γ‑CA) and adenylate kinase (AK) along with two unidentified proteins were more abundant. In conclusion, few protein changes characterize the initial response to Al in signalgrass. During the alarm phase, changes are related to P-mobilization, downregulation of Fe-acquisition, reduction of phenolic biosynthesis, and small stimulation of organic acid exudation. After recovering (resistant phase), biosynthesis of phenolics and methionine, but not Fe-mobilization are re-established. Full expression of Al-resistance is characterized by enhanced γ‑CA mediating mitochondrial complex I assembly and increased AK abundance indicating higher root respiration and better provision of ADP and Mg2+ to ATP synthase, respectively. The unidentified proteins and the specific role of γ‑CA in Al resistance of U. decumbens will centre future research.

Mechanisms of Hyper-resistance and Hyper-tolerance to Aluminum in Plants

As a widespread, permanent stress factor in acid soils, Aluminum toxicity has driven the evolution of different mechanisms that allow plants to colonize these adverse environments. Even more, Al-induced stimulation of growth has frequently been observed in highly adapted plants. Plant strategies for handling excess Al span from highly efficient exclusion (hyper-resistance) to the tolerance of extremely high Al accumulation within leaf tissues (hyper-tolerance). This chapter, after considering potential mechanisms for Al-induced growth stimulation, gives an overview of the current knowledge on Al hyper-resistance and Al hyper-tolerance mechanisms in plants with special focus on both the highly efficient excluder species of the genus Urochloa (former Brachiaria) and the most studied Al accumulators, tea and buckwheat.