Arsenic (As) exposure from consumption of rice can be substantial, particularly for the population on a subsistence rice diet in South Asia. Paddy rice has a much enhanced As accumulation compared with other cereal crops, and practical measures are urgently needed to decrease As transfer from soil to grain. We investigated the dynamics of As speciation in the soil solution under both flooded and aerobic conditions and compared As accumulation in rice shoot and grain in a greenhouse experiment. Flooding of soil led to a rapid mobilization of As, mainly as arsenite, in the soil solution. Arsenic concentrations in the soil solution were 7-16 and 4-13 times higher under the flooded than under the aerobic conditions in the control without As addition and in the +As treatments (10 mg As kg -1 as arsenite or arsenate), respectively. Arsenate was the main As species in the aerobic soil. Arsenic accumulation in rice shoots and grain was markedly increased under flooded conditions; grain As concentrations were 10-15-fold higher in flooded than in aerobically grown rice. With increasing total As concentrations in grain, the proportion of inorganic As decreased, while that of dimethylarsinic acid (DMA) increased. The concentration of inorganic As was 2.6-2.9 fold higher in the grain from the flooded treatment than in that from the aerobic treatment. The results demonstrate that a greatly increased bioavailability of As under the flooded conditions is the main reason for an enhanced As accumulation by flooded rice, and growing rice aerobically can dramatically decrease the As transfer from soil to grain.

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Nickel uptake and cellular compartmentation were investigated in three Ni hyperaccumulators: Alyssum bertolonii (Desv), Alyssum lesbiacum (Candargy) and Thlaspi goesingense (Hálácsy). The three species showed similar hyperaccumulation of Ni, but T. goesingense was less tolerant to Ni than the two Alyssum species. An addition of 500 mg Ni kg(-1) to a nutrient-rich growth medium significantly increased shoot biomass of all three species, suggesting that the Ni hyperaccumulators have a higher requirement for Ni than normal plants. Energy-dispersive X-ray microanalysis (EDXA) was performed on frozen-hydrated tissues of leaves (all species) and stems (Alyssum only). In all species analysed, Ni was distributed preferentially in the epidermal cells, most likely in the vacuoles, of the leaves and stems. In stems, there was a second peak of Ni in the boundary cells between the cortical parenchyma and the vascular cylinder. The non-glandular trichomes on the leaf surfaces of the two Alyssum species were highly enriched with Ca, but contained little Ni except in the base. In the leaves of T. goesingense, the large elongated epidermal cells contained more Ni than the cells of the stomatal complexes. The role of cellular compartmentation in Ni hyperaccumulation is discussed.

Rice represents a major route of As exposure in populations that depend on a rice diet. Practical measures are needed to mitigate the problem of excessive As accumulation in paddy rice. Two potential mitigation methods, management of the water regime and Si fertilization, were investigated under greenhouse conditions. Growing rice aerobically during the entire rice growth duration resulted in the leastAs accumulation. Maintaining aerobic conditions during either vegetative or reproductive stage of rice growth also decreased As accumulation in rice straw and grain significantly compared with rice grown under flooded conditions. The effect of water management regimes was consistent with the observed effect of flooding-induced arsenite mobilization in the soil solution. Aerobic treatments increased the percentage of inorganic As in grain, but the concentrations of inorganic As remained lower than in the flooded rice. Silicon fertilization decreased the total As concentration in straw and grain by 78 and 16%, respectively, even though Si addition increased As concentration in the soil solution. Silicon also significantly influenced As speciation in rice grain and husk by enhancing methylation. Silicon decreased the inorganic As concentration in grain by 59% while increasing the concentration of dimethylarsinic acid (DMA) by 33%. There were also significant differences between two rice genotypes in grain As speciation. This study demonstrated that water management Si fertilization, and selection of rice cultivars are effective measures that can be used to reduce As accumulation in rice.

Pentavalent methylated arsenic (As) species such as monomethylarsonic acid [MMA(V)] and dimethylarsinic acid [DMA(V)] are used as herbicides or pesticides, and can also be synthesized by soil microorganisms or algae through As methylation. The mechanism of MMA(V) and DMA(V) uptake remains unknown. Recent studies have shown that arsenite is taken up by rice (Oryza sativa) roots through two silicon transporters, Lsi1 (the aquaporin NIP2;1) and Lsi2 (an efflux carrier). Here we investigated whether these two transporters also mediate the uptake of MMA(V) and DMA(V). MMA(V) was partly reduced to trivalent MMA(III) in rice roots, but only MMA(V) was translocated to shoots. DMA(V) was stable in plants. The rice lsi1 mutant lost about 80% and 50% of the uptake capacity for MMA(V) and DMA(V), respectively, compared with the wild-type rice, whereas Lsi2 mutation had little effect. The short-term uptake kinetics of MMA(V) can be described by a MichaelisMenten plus linear model, with the wild type having 3.5-fold higher V max than the lsi1 mutant. The uptake kinetics of DMA(V) were linear with the slope being 2.8-fold higher in the wild type than the lsi1 mutant. Heterologous expression of Lsi1 in Xenopus laevis oocytes significantly increased the uptake of MMA(V) but not DMA(V), possibly because of a very limited uptake of the latter. Uptake of MMA(V) and DMA(V) by wild-type rice was increased as the pH of the medium decreased, consistent with an increasing proportion of the undissociated species. The results demonstrate that Lsi1 mediates the uptake of undissociated methylated As in rice roots.

Growth and zinc uptake of the hyperaccumulator species Thiaspi caerulescens ,1. & C. Presl and the non-hyperaccumulator species Thiaspi ochroleucum Boiss. & Heldr. were compared in solution culture experiments. T. caerulesecns was able to tolerate 500 mmol m"' (32-5 g m"') Zn in solution without growth reduction, and up to 1000 mmol m""^ (65 g m~"^) Zn without showing visible toxic symptoms but with a 25% decrease in dry matter (DM) yield. Up to 28 g kg"' of Zn in shoot DM was obtained in healthy plants of T. caerulescens. In contrast, 7'. ochroleucum suffered severe phytotoxicity at 500 mmol m~^ Zn. Marked differences were shown in Zn uptake, distribution and re-distribution between the two species. T. caerulescens had much higher concentrations of Zn in the shoots, whereas T. ochroleucum accumulated higher concentrations of Zn in the roots. When an external supply of 500 mmol m"^ Zn was withheld, 89% of the Zn accumulated previously in the roots of T. caerulescens was transported to tbe shoots over a 33 d period, whereas in 7". ochroleucum only 32% was transported. T. caerulescens was shown to have a greater internal requirement for Zn than other plants. Increasing the supply of Zn from 1 to 10 mmol m~' gave a 19% increase in the total DM of this species. Even the shoots from the 1 mmol m'^ Zn treatment which showed Zn deficiency contained 10 times greater Zn concentrations than the widely reported critical value for Zn deficiency to occur in many other plant species. The results obtained suggest that strongly expressed constitutive sequestration mechanisms exist in the hyperaccumulator 7'. caeruleseens, wbicb detoxify the large amount of Zn present in shoot tissues and decrea.se its physiological availability in tbe cytosol. Both T. caerulescens and T. ochroleiietim had constitutively high concentrations of malate in shoots, which were little affected by different Zn treatments. Although malate may play a role in Zn chelation because of tbe high concentrations present, it cannot explain the species specificity of Zn tolerance and hyperaccumulation.

Nodulin-26-like intrinsic proteins (NIPs) of the aquaporin family are involved in the transport of diverse solutes, but the mechanisms controlling the selectivity of transport substrates are poorly understood. The purpose of this study was to investigate how the aromatic/arginine (ar/R) selectivity filter influences the substrate selectivity of two NIP aquaporins; the silicic acid (Si) transporter OsLsi1 (OsNIP2;1) from rice and the boric acid (B) transporter AtNIP5;1 from Arabidopsis; both proteins are also permeable to arsenite. Native and site-directed mutagenized variants of the two genes were expressed in Xenopus oocytes and the transport activities for Si, B, arsenite, and water were assayed. Substitution of the amino acid at the ar/R second helix (H2) position of OsLsi1 did not affect the transport activities for Si, B, and arsenite, but that at the H5 position resulted in a total loss of Si and B transport activities and a partial loss of arsenite transport activity. Conversely, changes of the AtNIP5;1 ar/R selectivity filter and the NPA motifs to the OsLsi1 type did not result in a gain of Si transport activity. B transport activity was partially lost in the H5 mutant but unaffected in the H2 mutant of AtNIP5;1. In contrast, both the single and double mutations at the H2 and/or H5 positions of AtNIP5;1 did not affect arsenite transport activity. The results reveal that the residue at the H5 position of the ar/R filter of both OsLsi1 and AtNIP5;1 plays a key role in the permeability to Si and B, but there is a relatively low selectivity for arsenite.