Tradeoffs between metal hyperaccumulation ... - Wiley Online Library

Plant Pathology (2013) 62 (Suppl. 1), 63–71

Doi: 10.1111/ppa.12171

Trade-offs between metal hyperaccumulation and induced disease resistance in metal hyperaccumulator plants H. N. Fonesa and G. M. Prestonb* a

Biosciences, University of Exeter, Stocker Road, Exeter, EX4 4QD; and bDepartment of Plant Sciences, University of Oxford, South Parks Road, Oxford, OX1 3RB, UK

Metal hyperaccumulation is an unusual trait involving the uptake and storage of high concentrations of metals in the aerial tissues of plants. A number of hypotheses have been proposed to explain the evolution of the metal hyperaccumulation trait, of which the hypothesis that accumulated metal provides a defence against herbivores or pathogens has received most attention and support. Metal hyperaccumulation requires a range of physiological adaptations that enable plants to take up, transport, sequester and tolerate high concentrations of metal. Such adaptations may confer a fitness cost, and it has been suggested that metal hyperaccumulator plants may compensate for this cost by reducing investment in other traits such as induced disease resistance. This is supported by recent work that shows that metal hyperaccumulators such as Noccaea spp. show reduced or altered production of key components of induced disease resistance. However, an alternative explanation exists, which is that the physiological adaptations involved in metal hyperaccumulation require alterations to, or compromise, functional aspects of plant defence mechanisms. Here, the evidence for trade-offs between metal hyperaccumulation and disease resistance mechanisms is reviewed, and the nature of the physiological adaptations involved in metal hyperaccumulation and their potential to impact other forms of plant defence is discussed. It is suggested that defensive trade-offs may have been key to the evolution of the metal hyperaccumulation trait, resulting in increased dependence upon the protection conferred by metals. Keywords: metal hyperaccumulator, mineral nutrition, nickel, Noccaea caerulescens, reactive oxygen species, zinc

*E-mail: [email protected]

evolved on multiple occasions in different plant lineages, including 45 angiosperm families and the Pteridophyta (Salt & Kr€ amer, 2000; Ma et al., 2001). It therefore provides a useful framework in which to explore questions about plant evolution, plant population genetics and the trade-offs that occur during plant evolution. Recent research has greatly advanced the understanding of the molecular mechanisms that underpin the metal hyperaccumulation trait (Verbruggen et al., 2009; Freeman et al., 2010; Kr€ amer, 2010). However, many questions about the selective processes and the sequence of molecular events that occurred during the evolution of this trait remain unanswered. A number of hypotheses have been proposed to explain the evolution of the metal hyperaccumulation trait (Boyd & Martens, 1992; Boyd et al., 1994; Poschenrieder et al., 2006), of which the theory that accumulated metal provides a defence against herbivores or pathogens has received most attention and support (Boyd & Martens, 1992; Boyd, 2007, 2012; Vesk & Reichman, 2009). This hypothesis, sometimes known as the ‘elemental defence hypothesis’ is supported by the authors’ own work, which has shown that the metal accumulator Noccaea caerulescens displays increased resistance to pathogens such as the bacterial pathogen Pseudomonas syringae pv. maculicola when grown on increasing levels of zinc, nickel or cadmium. In plants grown on moderately high metal concentrations, the concentration of metal in the apoplastic compart-

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Introduction Metal hyperaccumulation is an unusual plant trait in which plants accumulate exceptionally high concentrations of metal in their foliar tissues (Pollard, 2000). For example, a metal hyperaccumulator plant may accumulate in excess of 10 000 lg g 1 (1%) zinc, while for other plants 30 lg g 1 may be sufficient and 300 lg g 1 toxic (Brooks et al., 1977; Assuncß~ao et al., 2003). Plants that exhibit this trait are typically found growing in naturally metal-rich or polluted soil, and may show reduced competitive fitness in normal or metal-deficient soil (Dechamps et al., 2007, 2008; Maestri et al., 2010). There are several reasons why these plants are of interest to researchers. First, they can be studied to understand the mechanisms used by plants to take up, store and tolerate metals, and thereby provide useful insight into potential strategies for biofortification of crops (Cakmak, 2008; White & Broadley, 2009; Zhao et al., 2012). Secondly, they can have practical applications in both phytoremediation of metal-polluted soils and phytoextraction of valuable metals from the environment (Chaney et al., 1998; Zhao & McGrath, 2009; Terry & Ba~ nuelos, 2010). Thirdly, metal hyperaccumulation is an intrinsically interesting trait, which appears to have

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H. N. Fones & G. M. Preston

ment of plants, in which this bacterium multiplies, is in excess of the concentrations required to inhibit bacterial growth in vitro. Additionally, bacterial mutants with reduced metal tolerance displayed a reduced ability to infect these plants (Fones et al., 2010). It is therefore plausible that these plants have evolved to exploit the high concentrations of metal present in some soils as an environmentally available microbicide or pesticide. If metal hyperaccumulation has evolved as, or is currently, a defensive trait, it is logical to ask whether the evolution of this trait has had an impact on other defence mechanisms. In a recent publication (Fones et al., 2013), it was reported that N. caerulescens lacks the ability to produce several ‘typical’ induced defences in response to P. syringae, including the oxidative burst, callose deposition and PR gene expression. Metal hyperaccumulation requires a range of physiological adaptations that enable plants to take up, transport, sequester and tolerate high concentrations of metal (Lasat et al., 1996, 2000; Salt & Kr€amer, 2000; Assuncß~ ao et al., 2001; Dr€ager et al., 2004; Ueno et al., 2011). Such adaptations may confer a substantial fitness cost, and it has been suggested that metal hyperaccumulator plants may compensate for this cost by reducing investment in other costly traits, which could include induced disease resistance (Maestri et al., 2010; Fones et al., 2013). Alternatively, it has been suggested that the physiological adaptations involved in metal hyperaccumulation necessitate alterations to, or lead to compromise of, functional aspects of plant defence mechanisms (Freeman et al., 2005; Fones et al., 2013; Llugany et al., 2013). Here, the evidence for trade-offs between metal hyperaccumulation and disease resistance mechanisms is reviewed, and the nature of the physiological adaptations involved in metal hyperaccumulation and their potential to impact on other forms of plant defence is discussed.

Trade-offs Evidence of costliness Metal hyperaccumulation is a complex trait, requiring many adaptations to the biochemistry of the plant. The a priori supposition that such adaptation must confer some degree of expense in terms of resource allocation has received support from a number of different empirical findings. Even before the term ‘metal hyperaccumulation’ was coined (Jaffre et al., 1976), metal tolerance, an important facet of the hyperaccumulation trait, was shown to become negatively selected as soil metal concentrations fell at the periphery of former mine sites (McNeilly, 1968). On non-metalliferous soils, metal hyperaccumulators have been found to be at a competitive disadvantage to non-accumulating plants (K€ upper et al., 1999). In reciprocal transplant experiments, ecotypes of the hyperaccumulator N. caerulescens, native to metalliferous soils, showed increased mortality and a reduction in seed production on non-metalliferous soils, due perhaps to higher metal requirements or higher costs

associated with adaptation to the metalliferous environment, and thus reduced competitiveness. However, it should be noted that metalliferous plants may also lack adaptations to the more pest- and pathogen-rich, nonmetalliferous environments (Dechamps et al., 2007). In present-day populations of metal hyperaccumulating species, edaphic endemism is frequently observed (K€ upper et al., 1999), indicating a lack of fitness to grow on ‘normal’ soils in competition with non-accumulator plants. One plausible reason for this disadvantage is that the highly efficient metal transport and sequestration mechanisms of metal hyperaccumulators lead to a greater requirement for the hyperaccumulated metal or other essential metals (Kr€ amer et al., 1996; Shen et al., 1997; K€ upper et al., 2001). The physiological and mechanistic adaptations of hyperaccumulators are discussed in more detail next, with an exploration of the ways in which each might lead to trade-offs in defence (Fig. 1).

Mechanisms: transport and sequestration One of the most widely recognized features of metal hyperaccumulating and metal tolerant organisms is increased activity of metal transporters, which act to extrude toxic metal ions from the cytoplasm and to load metals into the xylem and vacuole for transport and storage respectively. For example, N. caerulescens shows increased expression of zinc transporters, P-type ATPases and cation diffusion facilitator (CDF) proteins (Lasat et al., 2000; Assuncß~ ao et al., 2001; Hammond et al., 2006). Indeed, repeated duplication of the gene encoding the P-type ATPase, HMA4, which is responsible for xylem loading of zinc and cadmium, has been proposed as a key event in the evolution of metal hyperaccumulation in both Arabidopsis halleri (Hanikenne et al., 2008) Lochlainn et al., 2011). In the and N. caerulescens (O zinc hyperaccumulator, Noccaea (formerly Thlaspi) goesingense, the tonoplast transporter MTP1 has been demonstrated to be of importance in loading zinc into the vacuole, playing a role both in sequestering the metal to increase tolerance (Dr€ ager et al., 2004) and, importantly, in inducing a systemic zinc-deficiency response which both drives further metal accumulation and may also explain the elevated metal requirements observed in metal hyperaccumulators (Gustin et al., 2009). A similar phenomenon has been reported for the nickel hyperaccumulators of the genus Alyssum, in which heightened production of histidine allows for both increased tolerance and increased root-to-shoot transport of nickel (Kr€ amer et al., 1996). Transport into the vacuole, as well as complexation of metals with amino acids (Callahan et al., 2007; Maestri et al., 2010), organic acids (Boominathan & Doran, 2003; Lu et al., 2013) or specialized molecules such as metallothioneins (Maestri et al., 2010), are also important adaptations for metal hyperaccumulation and hypertolerance (Kr€ amer et al., 1997; K€ upper et al., 1999, 2000, 2004). Recent studies of yeast and Arabidopsis clearly illustrate how altered metal transport and sequestration can Plant Pathology (2013) 62 (Suppl. 1), 63–71

Metal hyperaccumulation and disease resistance

Capacity to hyperaccumulate metal allows deterrence even of metal tolerant pathogens

Investment in costly inducible defences no longer required Accumulated metals deter or are toxic to pests and pathogens

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Costs Quenching of ROS signals required for expression of inducible defences? Reliance upon metals for defence? Reduced competitiveness on non-metalliferous soils

Increased ROS production requiring increased antioxidant production Increased tolerance to metals; colonization of metalliferous soils possible

Benefits

Increased expression of metal uptake pumps and translocation into vacuole: increased basal metal requirements

Figure 1 Potential costs and benefits associated with the development of the metal hyperaccumulation phenotype. The ability to tolerate high soil metal concentrations opens a novel ecological niche, but metal accumulation requires expenditure on uptake, transport and sequestration, as well as increasing basal metal requirements, so that plants adapted to metalliferous sites are often endemic to them. Set against the costs of metal accumulation may be the benefit accrued as a result of reducing the cost of inducible defences against pathogens and herbivores. However, it is also possible that the loss of inducible defences follows from conflict between the physiological adaptations needed for hyperaccumulation and defence signals. An increasing reliance on metal-based defences may mean that increasing concentrations of metal must be accumulated in order to overcome increased metal tolerance in co-evolved pathogens and herbivores.

positively or negatively impact fitness. For example, cadmium tolerant Saccharomyces cerevisiae strains isolated from natural environments were found to show increased expression of the metal efflux pump gene PCA1. However, strains with high PCA1 expression were found to be at a selective disadvantage in low cadmium conditions compared to strains with low PCA1 activity (Chang & Leu, 2011). The authors speculated that over-expression of PCA1 in low cadmium conditions could result in depletion of other essential metals, thereby compromising growth. Similar processes could help to account for the reduced fitness shown by some metal hyperaccumulating plants when grown in soils with low concentrations of the accumulated metal. High levels of metal uptake or sequestration may also reduce the availability of other metals. For example, Nishida and collaborators demonstrated that excess Ni can up-regulate expression of the Fe transporter AtIRT1 in Arabidopsis, increasing uptake of both Ni and Fe, but causing changes in gene expression that are consistent with Ni-induced Fe deficiency (Nishida et al., 2011, 2012). Similarly, Arabidopsis plants engineered to have elevated levels of the metal-binding ligand nicotianamine have been demonstrated to show increased Ni tolerance, but also increased sensitivity to Fe deficiency (Pianelli et al., 2005; Cassin et al., 2009). In addition to these effects upon metal homeostasis, there is good reason to believe that depletion or sequestration of essential metals could specifically impact on plant disease resistance mechanisms. For example, zinc deficiency is one of the most widespread plant nutrient deficiencies worldwide, and has been shown to increase susceptibility to a range of pathogens (Dordas, 2009). Zinc deficiency has been reported to cause an increase in Plant Pathology (2013) 62 (Suppl. 1), 63–71

production of reactive oxygen species (ROS) and result in reduced detoxification of superoxide and hydrogen peroxide (Cakmak, 2000). Studies have also reported that low amounts of zinc in plant cells may enhance production of oxygen radicals during photosynthetic electron transport and induce membrane-bound NADPH oxidase activity (Cakmak, 2000), all of which could affect the activity and evolution of ROS-dependent signalling in plant immune responses, as discussed further below. Similarly, manganese deficiency has been reported to affect chloroplast functions and the activity of manganese superoxide dismutase (SOD; Yu & Rengel, 1999; Allen et al., 2007; Cao et al., 2011; Yruela, 2013). Nickel deficiency can impair the activity of the urea-hydrolysing enzyme urease, resulting in the accumulation of toxic levels of urea and oxalic and lactic acids (Bai et al., 2006; Seregin & Kozhevnikova, 2006) and impairing ammonium assimilation via glutamine synthetase (GS; Witte, 2011). GS is known to play an important role in plant metabolism during plant–pathogen interactions (Rico et al., 2011; Seifi et al., 2013), and is directly targeted by pathogens such as P. syringae pv. tabaci, which produces the GS-inhibiting toxin tabtoxin (Turner, 1981).

Mechanisms: tolerance – antioxidants Accumulated metals have the potential to become toxic when present in high concentrations through a variety of mechanisms that include causing or inducing oxidative stress and damage, interacting with thiol groups in proteins, or competing with other metals for binding sites in proteins (Fones & Preston, 2012). Free metal concentrations are known to be subject to exquisitely fine homeo-

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static control, with cytosolic zinc concentrations, for instance, measured as femtomolar in Escherichia coli (Outten & O’Halloran, 2001) and nanomolar in mammalian cells (Vinkenborg et al., 2009). Despite this, it has been demonstrated that superoxide production increases with zinc treatment in the hyperaccumulator N. caerulescens under hydroponic growth conditions, although without any symptoms of stress in these plants (Fones et al., 2013). It could be hypothesized that this ‘extra’ ROS is produced during the process of transport or sequestration of the metal. Supporting the concept that metal hyperaccumulation requires plants to tolerate increased ROS, genes encoding the key antioxidant enzymes catalase and peroxidase, as well as proteins involved in glutathione metabolism, have been found to be over-represented in hyperaccumulator transcriptomes when compared to the transcriptomes of congeneric non-accumulators (Becher et al., 2004; Chiang et al., 2006; Hammond et al., 2006) and this is supported by higher activities of antioxidant enzymes including catalase, ascorbate peroxidase and SOD (Srivastava et al., 2005; Hammond et al., 2006; Sharma & Dietz, 2009), and higher glutathione concentrations (Boominathan & Doran, 2003; Freeman et al., 2004; Sun et al., 2007; Jin et al., 2008; Tian et al., 2011). In addition, studies have shown hyperaccumulators to produce higher concentrations of organic and amino acids, such as nicotianamine and histidine, which may provide protection against metal-induced stress (Ingle et al., 2005; W ojcik et al., 2006; Sun et al., 2007). Taken together, these results provide a compelling picture of biochemical adaptation to control and contain enhanced production of ROS in metal hyperaccumulators.

Cross-talk and conflict Heightened ROS production, along with efficient mechanisms to protect against this, could have far-reaching effects for metal hyperaccumulators. Plants respond to a wide variety of stresses, including biotic stresses such as pathogen or herbivore attack and abiotic stresses such as cold, drought, salinity or excess light, via a tightly regulated signalling network in which ROS are central. Along with other shared signalling components such as ion fluxes, MAPK (mitogen-activated protein kinase; Romeis et al., 1999; Kovtun et al., 2000; Ludwig et al., 2005) and CDPK (calcium-dependent protein kinase; Romeis et al., 2001; Ludwig et al., 2004; Harper & Harmon, 2005) cascades, elevated ROS form a node in plant signalling networks where multiple signalling pathways can potentially interact (Narusaka et al., 2004; Fujita et al., 2006; Chmielowska et al., 2010; Choudhury et al., 2013; Fig. 2). ROS signalling has been shown to be important in the response of plants to abiotic stresses (Miller et al., 2008; Suzuki et al., 2012) as well as in the plant response to pathogens (Wojtaszek, 1997; Alvarez et al., 1998; Navarro et al., 2004), being an early step in pathways such as salicylic acid signalling of defence gene (PR gene) induction and the hypersensitive response (HR; Fobert & Despres, 2005).

Abiotic stress

Biotic stress

ROS

Altered gene expression

n p k Nutrient stress

Kinase cascades

Plant hormones (SA, JA, GA, ABA)

Ni

Zn

Metal stress

Figure 2 Shared signalling pathways provide capacity for cross-talk. Many stresses, including biotic and abiotic stress, activate components of a shared signalling network: reactive oxygen species (ROS) production and changes in cellular redox states; ion fluxes across membranes and kinase cascades; and a network of plant hormones – salicylic (SA), jasmonic (JA), gibberellic (GA) and abscisic (ABA) acid, which is itself multiply interconnected. The fine-balancing and integration of this signalling network means that cross-talk is not only possible, but is often an integral feature of signalling events.

The central role of ROS in plant stress tolerance raises two distinct possibilities for an effect of elevated ROS production on signalling in metal hyperaccumulating plants. On the one hand, it is possible that metal-induced ROS might stimulate signalling in other stress pathways, thus ‘priming’ the plant to withstand those stresses. Examples of such ‘cross-protection’ between abiotic and biotic stresses have been documented, including, for example, an increase in disease resistance in pepper plants exposed to cold stress (Mengiste et al., 2003; Fujita et al., 2006; Chmielowska et al., 2010). There is also evidence that metals can directly protect hyperaccumulator plants from pathogen attack (Boyd et al., 1994; Ghaderian et al., 2000; Hanson et al., 2004; Fones et al., 2010). Under these circumstances, plants may have been subject to selection for defensive signalling to become ‘uncoupled’ from elevated ROS in order to limit constitutive activation of defence pathways, which is known to carry a fitness cost (Heidel et al., 2004; van Hulten et al., 2006), and thus have become increasingly reliant on metal-based defences (Fones et al., 2013). It is also possible that the requirement for enhanced ROS protection systems in metal hyperaccumulating plants leads to the damping of ROS signals generated in response to stress such as pathogens. It is even possible to postulate a more direct effect, as zinc has been shown to inhibit NADPH oxidase, slowing the production of superoxide in response to salt stress in tobacco cell culture (Kawano et al., 2002). It is therefore possible that a trade-off between metal-based defences and ROS-signalled Plant Pathology (2013) 62 (Suppl. 1), 63–71


defences occurs as a result of physiological incompatibility between metal hyperaccumulation and defence (Fones et al., 2013).

Context When considering the evolution of metal hyperaccumulation and possible trade-offs between defence mechanisms in these plants, it is necessary to consider the environments in which they are found and the often unique challenges these provide. Metal hyperaccumulators are generally found on metal-contaminated soils, although in certain species, including N. caerulescens, the trait is facultative and populations do occur on non-metalliferous soils (Escarre et al., 2000). Metal-contaminated soils arise in two ways: natural and anthropogenic (Broadley et al., 2007; Kumar & Maiti, 2013). Natural contamination occurs via the weathering of metals from the underlying rocks, and is particularly notable in the case of serpentine (ultramafic) soils, which are high in metals including nickel and chromium (Gambi, 1992; Jaffre et al., 2013; Kumar & Maiti, 2013), and of calamine soils, which are high in zinc and lead (Brooks, 1987; Pawlowska et al., 1996; Broadley et al., 2007). Anthropogenic contamination is the result of activities such as mining and smelting (Broadley et al., 2007), the application of sewage sludge to soils as fertilizer (Broadley et al., 2007), or the use of certain pesticides (Russell, 2005; Fones & Preston, 2012). A common feature of both naturally metalliferous sites, and the older of the anthropogenic metalliferous soils – those that have been contaminated by mining (e.g. Southwood & Bevins, 1995) – is that they are often poor in other nutrients (Pawlowska et al., 1996; Robinson et al., 1996). It has been proposed that this nutrient poverty is important in shaping both the evolution of the plants native to these soils, and the evolution of defensive traits in those plants (Noret et al., 2007), and some evidence has been provided in favour of the idea that metalliferous sites, as environments rather hostile to the fostering of life, present the plants that can survive them with a reduced herbivory pressure (Noret et al., 2007; Dechamps et al., 2008). If true, this effect of metalliferous soil on herbivore populations will inevitably impact on the evolution of antiherbivory defences in plants native to such soils, including metal hyperaccumulators. The question of metal hyperaccumulation as a defence against herbivores has been fairly thoroughly explored, and recent reviews indicate that the balance of evidence is in favour of a defensive effect of hyperaccumulated metals (Boyd, 2007, 2012; Vesk & Reichman, 2009), even in plants in which metal concentrations are below the limits used to define hyperaccumulation (Cheruiyot et al., 2013). Understandably, metal hyperaccumulation appears to be most effective as a defence against those herbivores exposed to the total metal content of the leaves (Jhee et al., 2005; Boyd, 2007). Whether this provides evidence for defence against herbivory as an explanation for the evolution of the trait is a more Plant Pathology (2013) 62 (Suppl. 1), 63–71

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controversial question. To answer this, it will be necessary, among other requirements, to reach an understanding of the population structure of both metal hyperaccumulating plants and the herbivores that attack them in their native environments. The same questions must, of course, be faced for the case of the influence of metal hyperaccumulation upon the population biology of plants and their pathogens. The fact that many pathogens are capable of rapid evolution, with short generation times and the capacity to exchange genetic material by horizontal transfer (Ochman et al., 2000), makes them attractive candidates to take part in an arms race, driving the transition from metal accumulation to metal hyperaccumulation. However, the question remains as to whether microorganisms can evolve metal tolerance at such an increased rate relative to plants as to render metal hyperaccumulation incapable of providing durable resistance to locally adapted pathogens. Indeed, it has been demonstrated that plant pathogenic bacteria isolated from the leaves of metal hyperaccumulating plants, as well as bacteria adapted to the rhizosphere of metal hyperaccumulating plants, or simply to the high metal environment of a metalliferous soil, are highly metal tolerant (Idris et al., 2004; Barzanti et al., 2007; Fones et al., 2010) and that bacteria can develop metal tolerance with relative ease (Mergeay et al., 2003; Fones et al., 2010). Of course, the existence of metal-tolerant pathogens, like the existence of metaltolerant herbivores (e.g. Freeman et al., 2006) does not necessarily represent the tolerance levels of organisms associated with the ancestors of metal hyperaccumulating plants. Under the model proposed, both high metal concentrations at metalliferous sites and metal accumulation by plants could drive pathogens to acquire increased metal tolerance. Meanwhile, however, even modest levels of metal accumulation could protect plants from opportunistic pathogens not local to metalliferous sites, while locally adapted, metal-tolerant pathogens might provide a selection pressure for further accumulation. However, in a model of hyperaccumulator evolution in which the advantage of defence against pathogens drives the development of the metal hyperaccumulation phenotype, the concept of a trade-off between metalbased and other defences is key (Fones et al., 2013). All plants are exposed to pathogens and a variety of defensive signalling pathways and responses have been characterized in model species. Importantly, as discussed above, metal stress signals and responses could interact with these defensive pathways, either reinforcing their effects or leading them to be uncoupled to avoid spurious defence signals and their associated costs. If ‘ordinary’ defence signals are down-regulated, uncoupled, or quenched, the plant may become increasingly reliant on metals for defence. It is this reliance that could provide both the selection pressure for more extensive metal accumulation on the part of the plant, and for metal tolerance in the pathogen. In this scenario, the evolutionary plasticity of the pathogen becomes a relevant factor, and pathogens become players in a co-evolutionary arms race

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driving the emergence of metal hyperaccumulating plants.

Summary In this article, the physiologically complex trait of metal hyperaccumulation has been discussed with a view to exploring the potential for trade-offs between hyperaccumulated metal as an antimicrobial defence, and the retention and use of other forms of disease resistance. There is evidence that both metal hyperaccumulation and pathogen-induced defences incur costs to the plant, setting the scene for potential trade-offs in defensive resource allocation. Moreover, there are a number of reasons for hypothesizing that metal hyperaccumulation may be physiologically incompatible with some of the signal transduction mechanisms involved in plant defences, making the trade-off not a beneficial saving in resources but an integral feature of the hyperaccumulation lifestyle. A thorough mechanistic and physiological understanding of such tradeoffs will be valuable in the further development of theories of the evolution of metal hyperaccumulation. Any trade-off between defences that leaves the plants to any extent reliant upon metals for defence opens the door to an evolutionary arms race in which the plant becomes dependent on everheightening metal concentrations, to counter ever more metal-tolerant enemies. In this scenario, the ubiquity and evolutionary plasticity of pathogens renders them strong candidates to fill the role of ‘metal-tolerant enemy’.

Conflicts of interest The authors have no conflicts of interests to declare.

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