Arsenic‐eaters: by accident or by design (2008)

In late June 2008 over 70 people gathered at the Douglas Hotel in Aberdeen, Scotland to discuss the metabolism and chemical speciation of arsenic (As). For the participants originating closer to the equator, the surprising midnight blue skies were certainly a treat, particularly when leaving the pub around midnight after some ‘off-site’ scientific discussion. But putting aside the pleasures of Aberdeen in early summer, what is so interesting about arsenic that would persuade over 70 people to spend 2 days discussing it? Well there are many interesting aspects of arsenic biogeochemistry, some better known than others. For example, numerous reports exist in the literature of arsenic being consumed for medical reasons, the most extreme of which are the ‘arsenic-eaters’ from the mid-1800s who consumed arsenic compounds to improve their general well-being (for a review see Przygoda et al., 2001). However, there is no doubt that arsenic is best known for its toxicity, which was recognized even by the ancient Greeks. Deliberate arsenic poisoning is now generally a thing of the past. However, as a result of natural and anthropogenic processes arsenic still threatens human societies through its entrainment in the food chain. It was to discuss this issue that participants attended the meeting. Topics ranged across multiple plant species and scales, from the molecular to the field, and covered a diverse set of processes, from arsenic soil chemistry, plant biological and analytical chemistry, to arsenic contamination of food and the practical approaches that could be taken to reduce such contamination. It was also clear from the meeting that research into the biogeochemistry of arsenic is perhaps one of a handful of research areas that is naturally moving towards a model in which researchers utilize numerous experimental systems, from traditional genetic models such as Arabidopsis thaliana, and ‘natural’ populations such as the grass Holcus lanatus and the fern Pteris vittata, to crops such as rice (Oryza sativa), for both research and final deployment of new traits such as reduced grain arsenic accumulation. Using these diverse approaches rapid progress is being made in this field, and highlights of this work presented at the meeting are summarized below. ‘Not only is the amount of arsenic in the grain crucial to its toxicity, but also the chemical form of the arsenic in the grain plays a critical role.’ It is now well established that plants take up arsenate (As(V)) as a phosphate analog and rapidly reduce it to arsenite (As(III)) in roots. In most plants the As(III) produced by this reductive process is chelated by phytochelatins (PCs) and the majority is stored in the root vacuoles. Also it has recently been demonstrated that As(III) can be effluxed out of the roots in hydroponic systems (Xu et al., 2007). Reduction, PC chelation and storage also occur on uptake of methylarsenate. However, dimethylarsenate appears not to be reduced or complexed, and this may explain its preferential translocation from roots to shoots (Jörg Feldmann, University of Aberdeen, UK). Interestingly, in arsenic-hyperaccumulating plants such as the fern P. vittata, after reduction As(III) is preferentially translocated from the roots to the shoots as the As(III) oxyanion arsenite in which form it is stored in the shoot. Plant enzymes capable of reducing arsenate to arsenite have been cloned and enzymatically characterized from A. thaliana, H. lanatus, P. vittata and rice (Bleeker et al., 2006; Ellis et al., 2006; Duan et al., 2007). However, the function of these enzymes in vivo remains to be fully understood. In A. thaliana knockdown and loss-of-function mutants are both hypersensitive to As(V) (Bleeker et al., 2006; Dhankher et al., 2006) as would be expected if arsenate reduction is necessary to resistance. Furthermore, A. thaliana lines ectopically overexpressing the gene that encodes arsenate reductase are more resistant to arsenate. Interestingly, Henk Schat (Vrije Universiteit Amsterdam, the Netherlands) reported at the meeting that these arsenic phenotypes are dependent on the phosphorus (P) status of the plants, with high P reversing these phenotypes. This gave rise to the suggestion that under low P, where loss-of-function of arsenate reductase causes arsenate hypersensitivity, it is As(V) that is the toxic species. However, under elevated P nutrition, where loss-of-function of arsenate reductase causes enhanced As(V) resistance, As(III) is the likely toxic species. Unpublished data on arsenic speciation in both yeast and A. thaliana lacking arsenate reductase show that the majority of arsenic is still reduced to As(III) in these organisms, suggesting that there may be several ways to reduce As(V) to As(III) in vivo. These issues await further clarification. In aerobic environments arsenic is thought to be taken up by a phosphate transporter(s). However, in anaerobic environments, such as during rice cultivation, arsenite is the main species present in the soil and therefore taken up by plants. However, the transport proteins for both arsenite uptake from the soil and transport from roots to shoots were unknown. As reported at the meeting (Fangjie Zhao, Rothamsted Research, UK), and recently published (Ma et al., 2008), this mystery is now solved, at least for rice. Ma and co-authors have elegantly established that arsenite uptake into roots and transport into the xylem occur via the previously characterized silicon transporters OsLsi1 and OsLsi2, which are, respectively, an aquaglyceroporin (a member of the subfamily of nodulin26-like intrinsic proteins (NIPs)) and a silicon/arsenite efflux carrier. Mutation of OsLsi1 causes a significant decrease in arsenite uptake in rice. Furthermore, mutation of the silicon efflux transporter OsLsi2 reduces arsenite transport to the xylem and also accumulation of arsenic in the shoot and grain. Arsenite transport in rice therefore appears to share the same transport pathway as silicon, which explains why rice, a silicon accumulator, also accumulates elevated arsenic. Data reported at the meeting from several different research groups (Thomas Jahn (University of Copenhagen, Denmark), Toru Fujiwara (University of Tokyo, Japan) and Frans Maathuis (University of York, UK)) now also establish that members of the NIP subfamily can act as bidirectional arsenite transporters in other species, including A. thaliana and Lotus japonicus. Specifically, loss-of-function mutants nip1;1 and nip7;1 in A. thaliana both show increased resistance to arsenite, and NIP1;1 directly transports arsenite in oocytes, suggesting its direct involvement in arsenite uptake by roots. Furthermore, NIP5;1 and NIP6;1 were also shown to increase arsenite sensitivity when heterologously expressed in yeast, although the in vivo function of these proteins is not yet clear. Interestingly, NIPs do not appear to be the only means by which plants can transport arsenite. As reported at the meeting (David E. Salt, Purdue University, IN, USA) it would appear that the arsenic-hyperaccumulating fern P. vittata transports arsenite via a protein in the bile/arsenite/riboflavin (BART) superfamily, with close homology to the yeast arsenite effluxer arsenic compounds resistance 3 (ACR3). When heterologously expressed in yeast the P. vittata PvACR3 gene fully complements the yeast Δacr3 mutation, and knockdown of PvACR3 by RNAi in P. vittata causes severe arsenite hypersensitivity. Interestingly, orthologs of PvACR3 are absent from all sequenced angiosperm genomes, suggesting that this gene has been lost during the evolution of angiosperms. A major route of arsenic poisoning in humans is through the consumption of arsenic-tainted food. There is substantial variation in the natural concentrations of arsenic in soils and groundwater around the world. However, arsenic has also been introduced into the environment by a number of human activities. These include industrial and mining activities, as one might expect, along with other less obvious activities such as the agricultural use of arsenic-based pesticides or the sinking of tube wells in countries such as Bangladesh, which tap into arsenic-contaminated groundwater which is used to irrigate crops such as rice. Andy Meharg (University of Aberdeen, UK) and co-workers demonstrated that rice was far more adept than other cereal crops at taking up arsenic from the soil. This is probably attributable in part to the anaerobic conditions of the soil in the flooded paddy fields where rice is grown, which promote the reduction of arsenate to arsenite, allowing its efficient uptake via aquaglyceroporin/silicon transporters. Arsenic has been shown to accumulate to elevated concentrations in grain (0.3–0.5 µg g−1) from various regions around the world, including Bangladesh (from irrigation water) and the southern central USA (potentially from arsenic-containing pesticides) (Williams et al., 2006, 2007). Meharg indicated that, on the basis of the US Environmental Protection Agencies 2002 recommendations for arsenic toxicity (developed from epidemiological studies performed in Taiwan), people subsisting primarily on rice with these elevated concentrations of arsenic are at a higher risk of cancer. Not only is the amount of arsenic in the grain crucial to its toxicity, but also the chemical form of the arsenic in the grain plays a critical role. Grain with higher concentrations of inorganic arsenic (arsenate and arsenite) is more toxic than grain with a higher proportion of organic arsenic (dimethyl arsinic acid). A major question discussed at the meeting is how to decrease the concentration of arsenic in the rice grain, and specifically how to decrease the concentrations of inorganic arsenic without altering grain yield. These issues were addressed by Adam Price (University of Aberdeen, UK) when he described efforts he is involved in to identify rice varieties with reduced arsenic accumulation in the grain. These experiments included using local rice varieties from various countries, recombinant inbred mapping populations, and an association mapping population grown on arsenic-contaminated fields in a number of countries in Asia. It is hoped that such low-grain-arsenic varieties can be used in breeding programs to introduce low-grain-arsenic traits into elite production varieties. There is also potential for the identification of genes involved in regulating grain arsenic loading which will gives us a greater understanding of both mechanisms of and natural variation in plant–arsenic interactions. The idea of using plants for the restoration of trace element-contaminated environments can be traced back at least 35 yr to the pioneering work of Tony Bradshaw and co-workers who used metal-tolerant grass races for the restoration of mine tailings. The recent discovery of the arsenic-hyperaccumulating fern P. vittata by Ma et al. (2001) has opened the door to the possibility of using this fern to remediate, or phytoextract, arsenic from polluted soils. Such phytoextraction strategies have already been extensively tested for both cadmium and nickel using various Thlaspi and Alyssum species, and work has also been done on developing plants that directly volatilize Hg° as a mercury remediation strategy. Interestingly, the idea of developing plants engineered for the phytovolatilization of trimethyarsine gas as an arsenic remediation strategy was also discussed at the meeting by Barry Rosen (Wayne State University, MI, USA). He also reported the identification of an arsenite methyltransferase (ArsM) from the hot spring acidophilic alga Cyanidoschyzon merolae capable of methylating arsenite to trimethyarsine, bringing this potential approach one step closer to application. As was the case with the phytovolatilization of mercury, the toxicology of such an approach for arsenic needs to be carefully evaluated before it can be deployed. It was clear from this meeting that integration of arsenic research groups focused on basic abiotic/biotic mechanisms, through environmental and food chain arsenic accumulation to restoration technologies, is allowing rapid progress to be made, with translation of basic scientific understanding into practical applications for limiting arsenic uptake into the food chain. The broad significance and general public interest of this work, and the optimum size of the research community, will also facilitate this process. Development of a community cyberinfrastructure was also discussed. This would further enhance community interactions through sharing of data sets, analysis tools, predictive models, educational material and collaborative tools. On a final note, we would also like to take this opportunity to acknowledge the significant contributions Prof. Mark Macnair has made over the last 30 yr in the field of the ecological genetics of arsenic, copper and zinc tolerance and hyperaccumulation in plants. Mark will be retiring from the University of Exeter this year to play croquet, among other things, and the talk he gave at this meeting on ‘The ecological genetics of arsenic tolerance in Holcus lanatus’ was his last as a University Professor. We wish Mark all the best and expect that his postdoc Prof. Andy Meharg (Meeting co-organizer) will continue the good work.