Plant nitrogen (N) uptake is a key process in the global N cycle and is usually considered a “bottleneck” for biomass production in land ecosystems. Earlier, mineral N was considered the only form available to plants. Recent studies have questioned this dogma and shown that plants may access organic N sources such as amino acids. The actual mechanism enabling plants to access amino acid N is still unknown. However, a recent study suggested the Lysine Histidine Transporter 1 (LHT1) to be involved in root amino acid uptake. In this study, we isolated mutants defective in root amino acid uptake by screening Arabidopsis (Arabidopsis thaliana) seeds from ethyl methanesulfonate-treated plants and seeds from amino acid transporter T-DNA knockout mutants for resistance against the toxic d-enantiomer of alanine (Ala). Both ethyl methanesulfonate and T-DNA knockout plants identified as d-Ala resistant were found to be mutated in the LHT1 gene. LHT1 mutants displayed impaired capacity for uptake of a range of amino acids from solutions, displayed impaired growth when N was supplied in organic forms, and acquired substantially lower amounts of amino acids than wild-type plants from solid growth media. LHT1 mutants grown on mineral N did not display a phenotype until at the stage of flowering, when premature senescence of old leaf pairs occurred, suggesting that LHT1 may fulfill an important function at this developmental stage. Based on the broad and unbiased screening of mutants resistant to d-Ala, we suggest that LHT1 is an important mediator of root uptake of amino acids. This provides a molecular background for plant acquisition of organic N from the soil.

Various amino acids, including both L- and D-enantiomers, may be present in soils, and recent studies have indicated that plants may access such nitrogen (N) forms. Here, the capacity of Arabidopsis to utilize different L- and D-amino acids is investigated and the constraints on this process are explored. Mutants defective in the lysine histidine transporter 1 (LHT1) and transgenic plants overexpressing LHT1 as well as plants expressing D-amino acid-metabolizing enzymes, were used in studies of uptake and growth on various N forms. Arabidopsis absorbed all tested N-forms, but D-enantiomers at lower rates than L-forms. Several L- but no D-forms were effective as N sources. Plants deficient in LHT1 displayed strong growth reductions and plants overexpressing LHT1 showed strong growth enhancement when N was supplied as amino acids, in particular when these were supplied at low concentrations. Several D- amino acids inhibited growth of wild-type plants, while transgenic Arabidopsis-expressing genes encoding D-amino acid-metabolizing enzymes could efficiently utilize such compounds for growth. These results suggest that several amino acids, and in particular L-Gln and L-Asn, promote growth of Arabidopsis, and increased expression of specific amino acid transporters enhances growth on amino acids. The efficiency by which transgenic plants exploit D-amino acids illustrates how plants can be engineered to utilize specific N sources otherwise inaccessible to them.

Summary• Specific transporters mediate uptake of amino acids by plant roots. Earlier studies have indicated that the lysine histidine transporter 1 and amino acid permease 1 participate in this process, but although plant roots have been shown to absorb cationic amino acids with high affinity, neither of these transporters seems to mediate transport of L-arginine (L-Arg) or L-lysine (L-Lys).• Here, a collection of T-DNA knockout mutants were screened for alterations in Arabidopsis root uptake rates of L-Arg and it was found that only the AAP5 mutant displayed clear phenotypic divergence on high concentrations of L-Arg. A second screen using low concentrations of 15 N-labelled L-Arg in the growth media also identified AAP5 as being involved in L-Arg acquisition.• Momentaneous root uptake of basic amino acids was strongly affected in AAP5 mutant lines, but their uptake of other types of amino acids was only marginally affected. Comparisons of the root uptake characteristics of AAP5 and LHT1 mutants corroborated the hypothesis that the two transporters have distinct affinity spectra in planta.• Root uptake of all tested amino acids, except L-aspartic acid (L-Asp), was significantly affected in double AAP5*LHT1 mutants, suggesting that these two transporters account for a major proportion of roots' uptake of amino acids at low concentrations.Key words: amino acid, amino acid permease 5, double mutant, lysine histidine transporter 1, nitrogen uptake.New Phytologist (2008) 180: 620-630