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SummaryThe major constraint to plant growth in acid soils is the presence of toxic aluminum (Al) cations, which inhibit root elongation. The enhanced Al tolerance exhibited by some cultivars of wheat is associated with the Al-dependent ef¯ux of malate from root apices. Malate forms a stable complex with Al that is harmless to plants and, therefore, this ef¯ux of malate forms the basis of a hypothesis to explain Al tolerance in wheat. Here, we report on the cloning of a wheat gene, ALMT1 (aluminum-activated malate transporter), that co-segregates with Al tolerance in F 2 and F 3 populations derived from crosses between near-isogenic wheat lines that differ in Al tolerance. The ALMT1 gene encodes a membrane protein, which is constitutively expressed in the root apices of the Al-tolerant line at greater levels than in the near-isogenic but Alsensitive line. Heterologous expression of ALMT1 in Xenopus oocytes, rice and cultured tobacco cells conferred an Al-activated malate ef¯ux. Additionally, ALMT1 increased the tolerance of tobacco cells to Al treatment. These ®ndings demonstrate that ALMT1 encodes an Al-activated malate transporter that is capable of conferring Al tolerance to plant cells.

Aluminum (Al) is the most abundant metal in the earths crust, comprising about 7% of its mass. Since many plant species are sensitive to micromolar concentrations of Al, the potential for soils to be A1 toxic is considerable. Fortunately, most of the A1 is bound by ligands or occurs in other nonphytotoxic forms such as aluminosilicates and precipitates. However, solubilization of this A1 is enhanced by low pH and A1 toxicity is a major factor limiting plant production on acid soils. Soil acidification can develop naturally when basic cations are leached from soils, but it can be accelerated by some farming practices and by acid rain (Kennedy, 1986). Strategies to maintain production on these soils include the application of lime to raise the soil pH and the use of plants that are tolerant of acid soils. Although A1 toxicity has been identified as a problem of acid soils for over 70 years, our knowledge about the primary sites of toxicity and the chain of events that finally affects plant growth remains largely speculative. In this paper we review recent progress that has been made in our understanding of A1 toxicity and the mechanisms of A1 tolerance in plants. ALUMINUM TOXlClTYThe most easily recognized symptom of A1 toxicity is the inhibition of root growth, and this has become a widely accepted measure of A1 stress in plants. In simple nutrient solutions micromolar concentrations of A1 can begin to inhibit root growth within 60 min. However, the inhibition of growth per se offers little information about the causes of stress that will either precede or coincide with changes in growth. To understand the mechanisms of A1 toxicity, it is essential to identify the primary sites involved, both anatomical and metabolic, being mindful that A1 could have diverse effects and act differently in different species. Severa1 reviews on AI toxicity are available (see Haug, 1984;Taylor, 1988; Rengel, 1992a); here we limit our discussion to the sites of A1 toxicity in higher plants and to the possible role of Ca in the primary mechanism of A1 toxicity.The Phytotoxic Form of AI Part of the difficulty of studying Al-related processes in plants can be attributed to the complex chemistry of A1 * Corresponding author; e-mail manny@pican.pi.csiro.au; fax 61-6-246-5000. 315 (Martin, 1988;Kinraide, 1991). A1 hydrolyzes in solution such that the trivalent A1 species, A13+, dominates in acid conditions (pH < 5), whereas the A1(OH)2+ and Al(OH)2+ species form as the pH increases. At near-neutra1 pH the solid phase Al(OH),, or gibbsite, occurs, whereas Al(OH),-, or aluminate, dominates in alkaline conditions. Many of these monomeric A1 cations bind to various organic and inorganic ligands such as POb-, S02-, F-, organic acids, proteins, and lipids. Equilibrium constants are available for many of these reactions and these can be used to predict the relative concentrations of the monomeric A1 species and other A1 compounds in solution. A very toxic polynuclear A1 species, Al,,, can also form when A1 solutions are partially neutralized with...

We investigated the role of organic acids i n conferring AI tolerance in near-isogenic wheat (Trificum aestivum 1.) lines differing i n AI tolerance at the AI tolerance locus (Altl). Addition of AI to nutrient solutions stimulated excretion of malic and succinic acids from roots of wheat seedlings, and AI-tolerant genotypes excreted 5-to 10-fold more malic acid than AI-sensitive genotypes. Malic acid excretion was detectable after 15 min of exposure to 200 p~ AI, and the amount excreted increased linearly over 24 h. The amount of malic acid excreted was dependent on the externa1 AI concentration, and excretion was stimulated by as little as 10 p~ AI. Malic acid added to nutrient solutions was able to protect Alsensitive seedlings from normally phytotoxic AI concentrations. Root apices (terminal 3-5 mm of root) were the primary source of the malic acid excreted. Root apices of AI-tolerant and AI-sensitive seedlings contained similar amounts of malic acid before and after a 2-h exposure to 200 p~ AI. During this treatment, AI-tolerant seedlings excreted about four times the total amount of malic acid initially present within root apices, indicating that continua1 synthesis of malic acid was occurring. Malic acid excretion was specifically stimulated by AI, and neither La, Fe, nor the absence of Pi was able to elicit this response. There was a consistent correlation of AI tolerance with high rates of malic acid excretion stimulated by AI in a population of seedlings segregating for AI tolerance. These data are consistent with the hypothesis that the Altl locus in wheat encodes an AI tolerance mechanism based on AI-stimulated excretion of malic acid.

Aluminum (Al) tolerance in Arabidopsis is a genetically complex trait, yet it is mediated by a single physiological mechanism based on Al-activated root malate efflux. We investigated a possible molecular determinant for Al tolerance involving a homolog of the wheat Al-activated malate transporter, ALMT1. This gene, named AtALMT1 (At1g08430), was the best candidate from the 14-member AtALMT family to be involved with Al tolerance based on expression patterns and genomic location. Physiological analysis of a transferred DNA knockout mutant for AtALMT1 as well as electrophysiological examination of the protein expressed in Xenopus oocytes showed that AtALMT1 is critical for Arabidopsis Al tolerance and encodes the Al-activated root malate efflux transporter associated with tolerance. However, gene expression and sequence analysis of AtALMT1 alleles from tolerant Columbia (Col), sensitive Landsberg erecta (Ler), and other ecotypes that varied in Al tolerance suggested that variation observed at AtALMT1 is not correlated with the differences observed in Al tolerance among these ecotypes. Genetic complementation experiments indicated that the Ler allele of AtALMT1 is equally effective as the Col allele in conferring Al tolerance and Al-activated malate release. Finally, fine-scale mapping of a quantitative trait locus (QTL) for Al tolerance on chromosome 1 indicated that AtALMT1 is located proximal to this QTL. These results indicate that AtALMT1 is an essential factor for Al tolerance in Arabidopsis but does not represent the major Al tolerance QTL also found on chromosome 1. abiotic stress ͉ electrophysiology ͉ genetics ͉ organic acid exudation

Background Agricultural production is often limited by low phosphorus (P) availability. In developing countries, which have limited access to P fertiliser, there is a need to develop plants that are more efficient at low soil P. In fertilised and intensive systems, P-efficient plants are required to minimise inefficient use of P-inputs and to reduce potential for loss of P to the environment. Scope Three strategies by which plants and microorganisms may improve P-use efficiency are outlined: (i) Root-foraging strategies that improve P acquisition by lowering the critical P requirement of plant growth and allowing agriculture to operate at lower levels of soil P; (ii) P-mining strategies to enhance the desorption, solubilisation or mineralisation of P from sparingly-available sources in soil using root exudates (organic anions, phosphatases), and (iii) improving internal P-utilisation efficiency through the use of plants that yield more per unit of P uptake.Conclusions We critically review evidence that more P-efficient plants can be developed by modifying root growth and architecture, through manipulation of root exudates or by managing plant-microbial associations such as arbuscular mycorrhizal fungi and microbial inoculants. Opportunities to develop P-efficient plants through breeding or genetic modification are described and issues that may limit success including potential trade-offs and trait interactions are discussed. Whilst demonstrable progress has been made by selecting plants for root morphological traits, the potential for manipulating root physiological traits or selecting plants for low internal P concentration has yet to be realised.

Aluminium (A1) stimulates the efflux of malate from the apices of wheat (Triticum aestivum L.) roots (Delhaize et al. 1993, Plant Physiol. 103, 695-702). The response was five to tenfold higher in Al-tolerant seedlings than Al-sensitive seedlings and the capacity for Al-stimulated malate efflux was found to co-segregate with A1 tolerance in a pair of near-isogenic wheat lines differing in Al-tolerance at a single dominant locus. We have investigated this response further using excised root apices. Half-maximal efflux of malate from apices of A1-tolerant seedlings was measured with 30 gM A1 in 0.2raM CaC12, pH4.2, while saturating rates of 2.0 nmol. apex -~. h 1 occurred with concentrations above 100 gM A1. The stimulation of malate efflux by A1 was accompanied by an increase in K + efflux which appeared to account for electroneutrality. The greater stimulation of malate efflux from Al-tolerant apices compared to A1-sensitive apices could not be explained by differences in the activities of phosphoenolpyruvate carboxylase or NAD-malate dehydrogenase. Several other polyvalent cations, including gallium, indium and the tridecamer Al13, failed to elicit malate efflux. Aluminium-stimulated efflux of malate was correlated with the measured concentration of total monomeric AI present, and with the predicted concentrations of A13 + and A1OH 2 + ions in the solution. Several antagonists of anion channels inhibited Al-stimulated efflux of malate with the following order of effectiveness: niflumic acid ~NPPB > IAA-94 ~ A-9-C>ethacrynic acid. Lanthanum, chlorate, perchlorate, zinc and ~-cyano-4-hydroxycinnamic acid inhibited malate release by less than 30% at 100 gM while 4,4'-diisothiocyanatostilbene-2,2'-disulphonate (DIDS) had no Abbreviations: A-9-C = anthracene-9-carboxylic acid; DIDS = 4,4'-diisothiocyanatostilbene-2,2'-disulphonate; ET3, ET8, ES3, and ES8 = Al-tolerant and Al-sensitive lines of wheat; IAA-94 = [6, 7-effect. These results suggest that the AI 3+ cation stimulates malate efflux via anion channels in apical cells of Al-tolerant wheat roots.

The non-protein amino acid, gamma-aminobutyric acid (GABA) rapidly accumulates in plant tissues in response to biotic and abiotic stress, and regulates plant growth. Until now it was not known whether GABA exerts its effects in plants through the regulation of carbon metabolism or via an unidentified signalling pathway. Here, we demonstrate that anion flux through plant aluminium-activated malate transporter (ALMT) proteins is activated by anions and negatively regulated by GABA. Site-directed mutagenesis of selected amino acids within ALMT proteins abolishes GABA efficacy but does not alter other transport properties. GABA modulation of ALMT activity results in altered root growth and altered root tolerance to alkaline pH, acid pH and aluminium ions. We propose that GABA exerts its multiple physiological effects in plants via ALMT, including the regulation of pollen tube and root growth, and that GABA can finally be considered a legitimate signalling molecule in both the plant and animal kingdoms.