Root uptake of cationic amino acids by Arabidopsis depends on functional expression of amino acid permease 5

* 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.     

Arabidopsis LHT1 is a high-affinity transporter for cellular amino acid uptake in both root epidermis and leaf mesophyll

Amino acid transport in plants is mediated by at least two large families of plasma membrane transporters. Arabidopsis thaliana, a nonmycorrhizal species, is able to grow on media containing amino acids as the sole nitrogen source. Arabidopsis amino acid permease (AAP) subfamily genes are preferentially expressed in the vascular tissue, suggesting roles in long-distance transport between organs. We show that the broad-specificity, high-affinity amino acid transporter LYSINE HISTIDINE TRANSPORTER1 (LHT1), an AAP homolog, is expressed in both the rhizodermis and mesophyll of Arabidopsis. Seedlings deficient in LHT1 cannot use Glu or Asp as sole nitrogen sources because of the severe inhibition of amino acid uptake from the medium, and uptake of amino acids into mesophyll protoplasts is inhibited. Interestingly, lht1 mutants, which show growth defects on fertilized soil, can be rescued when LHT1 is reexpressed in green tissue. These findings are consistent with two major LHT1 functions: uptake in roots and supply of leaf mesophyll with xylem-derived amino acids. The capacity for amino acid uptake, and thus nitrogen use efficiency under limited inorganic N supply, is increased severalfold by LHT1 overexpression. These results suggest that LHT1 overexpression may improve the N efficiency of plant growth under limiting nitrogen, and the mutant analyses may enhance our understanding of N cycling in plants.     

Transporters in Arabidopsis roots mediating uptake of amino acids at naturally occurring concentrations

Recent studies of Arabidopsis have identified several transporters as being important for amino acid uptake. We used Arabidopsis plants with altered expression of lysine histidine transporter 1 (LHT1), amino acid permease 1 (AAP1) and amino acid permease 5 (AAP5) with the aim of disentangling the roles of each transporter in the uptake of different amino acids at naturally occurring concentrations (2-50 μM). LHT1 mutants displayed reduced uptake rates of L-Gln, L-Ala, L-Glu and L-Asp but not of L-Arg or L-Lys, while AAP5 mutants were affected in the uptake of L-Arg and L-Lys only. Double mutants (lht1aap5) exhibited reduced uptake of all tested amino acids. In the concentration range tested, AAP1 mutants did not display altered uptake rates for any of the studied amino acids. Expression analysis of amino acid transporter genes with important root functions revealed no major differences in the individual mutants other than for genes targeted for mutation. We conclude that LHT1 and AAP5, but not AAP1, are crucial for amino acid uptake at concentrations typically found in soils. LHT1 and AAP5 displayed complementary affinity spectra, and no redundancy with respect to gene expression was found between the two transporters, suggesting these two transporters have separate roles in amino acid uptake.     

Capacities and constraints of amino acid utilization in Arabidopsis

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.     

Amino acid transporter mutants of Arabidopsis provides evidence that a non‐mycorrhizal plant acquires organic nitrogen from agricultural soil

Although organic nitrogen (N) compounds are ubiquitous in soil solutions, their potential role in plant N nutrition has been questioned. We performed a range of experiments on Arabidopsis thaliana genetically modified to enhance or reduce root uptake of amino acids. Plants lacking expression of the Lysine Histidine Transporter 1 (LHT1) displayed significantly lower contents of ¹³C and ¹⁵N label and of U‐¹³C₅,¹⁵N₂ L‐glutamine, as determined by liquid chromatography–mass spectrometry when growing in pots and supplied with dually labelled L‐glutamine compared to wild type plants and LHT1‐overexpressing plants. Slopes of regressions between accumulation of ¹³C‐labelled carbon and ¹⁵N‐labelled N were higher for LHT1‐overexpressing plants than wild type plants, while plants lacking expression of LHT1 did not display a significant regression between the two isotopes. Uptake of labelled organic N from soil tallied with that of labelled ammonium for wild type plants and LHT1‐overexpressing plants but was significantly lower for plants lacking expression of LHT1. When grown on agricultural soil plants lacking expression of LHT1 had the lowest, and plants overexpressing LHT1 the highest C/N ratios and natural δ¹⁵N abundance suggesting their dependence on different N pools. Our data show that LHT1 expression is crucial for plant uptake of organic N from soil.     

The amino acid permease AAP8 is important for early seed development in <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis>

The development of seeds depends on the import of carbohydrates and amino acids supplied by the maternal tissue via the phloem. Several amino acid transporters have been reported to be expressed during seed and silique development in Arabidopsis thaliana (L.) Heynh. Here we show that mutants lacking the high affinity amino acid permease 8 (At1g10010) display a severe seed phenotype. The overall number of seeds and the number of normally developed seed is reduced by ∼50% in siliques of the Ataap8 T-DNA insertion mutant. This result could be reproduced in plants where expression of AtAAP8 is targeted with an RNAi approach. The seed phenotype is correlated with a specifically altered amino acid composition of young siliques. Aspartic acid and glutamic acid are significantly reduced in young siliques of the mutants. In correlation with the fact that AAP8 is a high affinity transporter for acidic amino acids, translocation of ¹⁴C-labelled aspartate fed via the root system to seeds of the mutants is reduced. AAP8 plays a crucial role for the uptake of amino acids into the endosperm and supplying the developing embryo with amino acids during early embryogenesis.     

AAP1 regulates import of amino acids into developing Arabidopsis embryos

The embryo of Arabidopsis seeds is symplasmically isolated from the surrounding seed coat and endosperm, and uptake of nutrients from the seed apoplast is required for embryo growth and storage reserve accumulation. With the aim of understanding the importance of nitrogen (N) uptake into developing embryos, we analysed two mutants of AAP1 (At1g58360), an amino acid transporter that was localized to Arabidopsis embryos. In mature and desiccated aap1 seeds the total N and carbon content was reduced while the total free amino acid levels were strongly increased. Separately analysed embryos and seed coats/endosperm of mature seeds showed that the elevated amounts in amino acids were caused by an accumulation in the seed coat/endosperm, demonstrating that a decrease in uptake of amino acids by the aap1 embryo affects the N pool in the seed coat/endosperm. Also, the number of protein bodies was increased in the aap1 endosperm, suggesting that the accumulation of free amino acids triggered protein synthesis. Analysis of seed storage compounds revealed that the total fatty acid content was unchanged in aap1 seeds, but storage protein levels were decreased. Expression analysis of genes of seed N transport, metabolism and storage was in agreement with the biochemical data. In addition, seed weight, as well as total silique and seed number, was reduced in the mutants. Together, these results demonstrate that seed protein synthesis and seed weight is dependent on N availability and that AAP1-mediated uptake of amino acids by the embryo is important for storage protein synthesis and seed yield.     

AAP1 transports uncharged amino acids into roots of Arabidopsis

Amino acids are available to plants in some soils in significant amounts, and plants frequently make use of these nitrogen sources. The goal of this study was to identify transporters involved in the uptake of amino acids into root cells. Based on the fact that high concentrations of amino acids inhibit plant growth, we hypothesized that mutants tolerating toxic levels of amino acids might be deficient in the uptake of amino acids from the environment. To test this hypothesis, we employed a forward genetic screen for Arabidopsis thaliana mutants tolerating toxic concentrations of amino acids in the media. We identified an Arabidopsis mutant that is deficient in the amino acid permease 1 (AAP1, At1g58360) and resistant to 10 mm phenylalanine and a range of other amino acids. The transporter was localized to the plasma membrane of root epidermal cells, root hairs, and throughout the root tip of Arabidopsis. Feeding experiments with [(14)C]-labeled neutral, acidic and basic amino acids showed significantly reduced uptake of amino acids in the mutant, underscoring that increased tolerance of aap1 to high levels of amino acids is coupled with reduced uptake by the root. The growth and uptake studies identified glutamate, histidine and neutral amino acids, including phenylalanine, as physiological substrates for AAP1, whereas aspartate, lysine and arginine are not. We also demonstrate that AAP1 imports amino acids into root cells when these are supplied at ecologically relevant concentrations. Together, our data indicate an important role of AAP1 for efficient use of nitrogen sources present in the rhizosphere.     

A T-DNA insertion knockout of the bifunctional lysine-ketoglutarate reductase/saccharopine dehydrogenase gene elevates lysine levels in Arabidopsis seeds

Plants possess both anabolic and catabolic pathways for the essential amino acid lysine (Lys). However, although the biosynthetic pathway was clearly shown to regulate Lys accumulation in plants, the functional significance of Lys catabolism has not been experimentally elucidated. To address this issue, we have isolated an Arabidopsis knockout mutant with a T-DNA inserted into exon 13 of the gene encoding Lys ketoglutarate reductase/saccharopine dehydrogenase. This bifunctional enzyme controls the first two steps of Lys catabolism. The phenotype of the LKR/SDH knockout was indistinguishable from wild-type plants under normal growth conditions, suggesting that Lys catabolism is not an essential pathway under standard growth conditions. However, mature seeds of the knockout mutant over-accumulated Lys compared with wild-type plants. This report provides the first direct evidence for the functional significance of Lys catabolism in regulating Lys accumulation in seeds. Such a knockout mutant may also provide new perspectives to improve the level of the essential amino acid Lys in plant seeds.     

Role of a single aquaporin isoform in root water uptake

Aquaporins are ubiquitous channel proteins that facilitate the transport of water across cell membranes. Aquaporins show a typically high isoform multiplicity in plants, with 35 homologs in Arabidopsis. The integrated function of plant aquaporins and the function of each individual isoform remain poorly understood. Matrix-assisted laser desorption/ionization time-of-flight analyses suggested that Plasma Membrane Intrinsic Protein2;2 (PIP2;2) is one of the abundantly expressed aquaporin isoforms in Arabidopsis root plasma membranes. Two independent Arabidopsis knockout mutants of PIP2;2 were isolated using a PCR-based strategy from a library of plant lines mutagenized by the insertion of Agrobacterium tumefaciens T-DNA. Expression in transgenic Arabidopsis of a PIP2;2 promoter-beta-glucuronidase gene fusion indicated that PIP2;2 is expressed predominantly in roots, with a strong expression in the cortex, endodermis, and stele. The hydraulic conductivity of root cortex cells, as measured with a cell pressure probe, was reduced by 25 to 30% in the two allelic PIP2;2 mutants compared with the wild type. In addition, free exudation measurements revealed a 14% decrease, with respect to wild-type values, in the osmotic hydraulic conductivity of roots excised from the two PIP2;2 mutants. Together, our data provide evidence for the contribution of a single aquaporin gene to root water uptake and identify PIP2;2 as an aquaporin specialized in osmotic fluid transport. PIP2;2 has a close homolog, PIP2;3, showing 96.8% amino acid identity. The phenotype of PIP2;2 mutants demonstrates that, despite their high homology and isoform multiplicity, plant aquaporins have evolved with nonredundant functions.     

ATMYB123和ATKOR1基因参与拟南芥根发育的调控

从拟南芥T-DNA插入突变体库中筛选到2个根发育相关基因ATMYB123和ATKOR1表达缺失的突变体atmyb123和atkor1,通过杂交构建这两个基因表达缺失的双突变体atmyb123/atkor1,以明确这两个基因在根发育中的作用。结果显示:(1)ATMYB123表达缺失突变体atmyb123植株地上部分发育减缓,种皮颜色变黄,而ATKOR1表达缺失突变体atkor1植株在这两方面与其野生型没有明显差异;两基因缺失均显著影响了拟南芥根的发育,根生长受到了严重抑制。(2)双突变体atmyb123/atkor1在植株形态和种皮颜色方面表现出单突变体AT-MYB123的特点,而其根长却介于两单突变体的中间。(3)进一步研究发现,培养基pH改变、NaCl处理、外源GA施用均没有改变突变体根生长趋势,显示这3种因素与两基因缺失突变引起的根发育抑制无关。研究表明,AT-MYB123和ATKOR1基因参与拟南芥根的发育调控,转录因子ATMYB123可能作为主调控因子参与ATKOR1对拟南芥根发育的调控。     

NAD(H)-dependent glutamate dehydrogenase is essential for the survival of Arabidopsis thaliana during dark-induced carbon starvation

Interconversion between glutamate and 2-oxoglutarate, which can be catalysed by glutamate dehydrogenase (GDH), is a key reaction in plant carbon (C) and nitrogen (N) metabolism. However, the physiological role of plant GDH has been a controversial issue for several decades. To elucidate the function of GDH, the expression of GDH in various tissues of Arabidopsis thaliana was studied. Results suggested that the expression of two Arabidopsis GDH genes was differently regulated depending on the organ/tissue types and cellular C availability. Moreover, Arabidopsis mutants defective in GDH genes were identified and characterized. The two isolated mutants, gdh1-2 and gdh2-1, were crossed to make a double knockout mutant, gdh1-2/gdh2-1, which contained negligible levels of NAD(H)-dependent GDH activity. Phenotypic analysis on these mutants revealed an increased susceptibility of gdh1-2/gdh2-1 plants to C-deficient conditions. This conditional phenotype of the double knockout mutant supports the catabolic role of GDH and its role in fuelling the TCA cycle during C starvation. The reduced rate of glutamate catabolism in the gdh2-1 and gdh1-2/gdh2-1 plants was also evident by the growth retardation of these mutants when glutamate was supplied as the alternative N source. Furthermore, amino acid profiles during prolonged dark conditions were significantly different between WT and the gdh mutant plants. For instance, glutamate levels increased in WT plants but decreased in gdh1-2/gdh2-1 plants, and aberrant accumulation of several amino acids was detected in the gdh1-2/gdh2-1 plants. These results suggest that GDH plays a central role in amino acid breakdown under C-deficient conditions.     

Arabidopsis stromal 70-kD heat shock proteins are essential for plant development and important for thermotolerance of germinating seeds

The 70-kD heat shock proteins (Hsp70s) have been shown to be important for protein folding, protein translocation, and stress responses in almost all organisms and in almost all subcellular compartments. However, the function of plastid stromal Hsp70s in higher plants is still uncertain. Genomic surveys have revealed that there are two putative stromal Hsp70s in Arabidopsis thaliana, denoted cpHsc70-1 (At4g24280) and cpHsc70-2 (At5g49910). In this study, we show that cpHsc70-1 and cpHsc70-2 could indeed be imported into the chloroplast stroma. Their corresponding T-DNA insertion knockout mutants were isolated and designated as Deltacphsc70-1 and Deltacphsc70-2. No visible phenotype was observed in the Deltacphsc70-2 mutant under normal growth conditions. In contrast, Deltacphsc70-1 mutant plants exhibited variegated cotyledons, malformed leaves, growth retardation, and impaired root growth, even though the protein level of cpHsc70-2 was up-regulated in the Deltacphsc70-1 mutant. After heat shock treatment of germinating seeds, root growth from Deltacphsc70-1 seeds was further impaired, indicating that cpHsc70-1 is important for thermotolerance of germinating seeds. No Deltacphsc70-1 Deltacphsc70-2 double mutant could be obtained, suggesting that the Deltacphsc70 double knockout was lethal. Genotype analyses of F(1) seedlings from various crosses indicated that double-knockout mutation was lethal to the female gametes and reduced the transmission efficiency of the male gametes. These results indicate that cpHsc70s are essential for plant development and the two cpHsc70s most likely have redundant but also distinct functions.     

AtPTR1 and AtPTR5 transport dipeptides in planta

Transporters for di- and tripeptides belong to the large and poorly characterized PTR/NRT1 (peptide transporter/nitrate transporter 1) family. A new member of this gene family, AtPTR5, was isolated from Arabidopsis (Arabidopsis thaliana). Expression of AtPTR5 was analyzed and compared with tissue specificity of the closely related AtPTR1 to discern their roles in planta. Both transporters facilitate transport of dipeptides with high affinity and are localized at the plasma membrane. Mutants, double mutants, and overexpressing lines were exposed to several dipeptides, including toxic peptides, to analyze how the modified transporter expression affects pollen germination, growth of pollen tubes, root, and shoot. Analysis of atptr5 mutants and AtPTR5-overexpressing lines showed that AtPTR5 facilitates peptide transport into germinating pollen and possibly into maturating pollen, ovules, and seeds. In contrast, AtPTR1 plays a role in uptake of peptides by roots indicated by reduced nitrogen (N) levels and reduced growth of atptr1 mutants on medium with dipeptides as the sole N source. Furthermore, overexpression of AtPTR5 resulted in enhanced shoot growth and increased N content. The function in peptide uptake was further confirmed with toxic peptides, which inhibited growth. The results show that closely related members of the PTR/NRT1 family have different functions in planta. This study also provides evidence that the use of organic N is not restricted to amino acids, but that dipeptides should be considered as a N source and transport form in plants.     

Uptake of intact amino acids by plants depends on soil amino acid concentrations

Studies in different ecosystems have shown that plants take up intact amino acids directly but little is known about the influence of free amino acid concentrations in the soil on this process. We investigated the effect of three different soil amino acid N concentrations (0.025, 0.13 and 2.5 μg N g^?1 soil) on direct uptake of four dual labelled (¹⁵N, ¹³C) amino acids (glycine, tyrosine, lysine, valine) in a greenhouse experiment using Anthoxantum odoratum as a model plant.Our results revealed that 8–45% of applied ¹⁵N was incorporated into plant root and shoot tissue 48 h after labelling. Additional ¹³C enrichment showed that 2–70% of this incorporated ¹⁵N was taken up as intact amino acid. Total ¹⁵N uptake and ¹⁵N uptake as intact amino acids were significantly affected by soil amino acid N concentrations and significantly differed between the four amino acids tested.We found a positive effect of soil amino acid concentrations on uptake of mineralized ¹⁵N relative to amino acid concentrations for all amino acids which was presumably due to higher diffusion rates of mineralized tracer to the root surface. However, intact amino acid uptake relative to amino acid concentrations as well as the proportion of total ¹⁵N taken up directly decreased with increasing soil amino acid N concentrations for all amino acids, irrespective of their microbial degradability. This effect is most likely controlled by the mineral N concentration in soil and perhaps in plants which inhibits direct amino acids uptake.Overall, we conclude that plant internal regulation of amino acid uptake controlled by mineral N is the main mechanism determining direct uptake of amino acids and thus a lower contribution of intact amino acid uptake to the plants N nutrition has to be expected for higher amino acid concentrations accompanied by mineralization in soil.     

Protein geranylgeranyltransferase I is involved in specific aspects of abscisic acid and auxin signaling in Arabidopsis

Arabidopsis (Arabidopsis thaliana) mutants lacking a functional ERA1 gene, which encodes the beta-subunit of protein farnesyltransferase (PFT), exhibit pleiotropic effects that establish roles for protein prenylation in abscisic acid (ABA) signaling and meristem development. Here, we report the effects of T-DNA insertion mutations in the Arabidopsis GGB gene, which encodes the beta-subunit of protein geranylgeranyltransferase type I (PGGT I). Stomatal apertures of ggb plants were smaller than those of wild-type plants at all concentrations of ABA tested, suggesting that PGGT I negatively regulates ABA signaling in guard cells. However, germination of ggb seeds in response to ABA was similar to the wild type. Lateral root formation in response to exogenous auxin was increased in ggb seedlings compared to the wild type, but no change in auxin inhibition of primary root growth was observed, suggesting that PGGT I is specifically involved in negative regulation of auxin-induced lateral root initiation. Unlike era1 mutants, ggb mutants exhibited no obvious developmental phenotypes. However, era1 ggb double mutants exhibited more severe developmental phenotypes than era1 mutants and were indistinguishable from plp mutants lacking the shared alpha-subunit of PFT and PGGT I. Furthermore, overexpression of GGB in transgenic era1 plants partially suppressed the era1 phenotype, suggesting that the relatively weak phenotype of era1 plants is due to partial redundancy between PFT and PGGT I. These results are discussed in the context of Arabidopsis proteins that are putative substrates of PGGT I.     

拟南芥胼胝质合酶基因GSL8参与细胞壁形成和根端静止中心建立与维持

用T-DNA插入和RNA干扰技术敲除拟南芥(Arabidopsis thaliana)胼胝质合酶基因GSL8, 在光学和透射电子显微镜下观察野生型和突变体种子的细胞壁和胚根结构, 比较未敲除和敲除该基因幼苗细胞壁及根端分生组织结构。结果表明, 敲除该基因可导致细胞壁发育不良, 壁上出现大小不等的缺口, 缺口处没有质膜将相邻细胞分隔。用T-DNA插入法抑制该基因表达, 发现在种子发育阶段胚根不能形成正常的静止中心。用小RNA干扰技术抑制该基因表达, 结果显示根端失去静止中心。综合以上结果, 认为该基因不仅参与细胞壁发育, 也参与根端静止中心的建立与维持。     

The chloroplastic 2-oxoglutarate/malate transporter has dual function as the malate valve and in carbon/nitrogen metabolism

Transport of dicarboxylates across the chloroplast envelope plays an important role in transferring carbon skeletons to the nitrogen assimilation pathway and exporting reducing equivalent to the cytosol to prevent photo-inhibition (the malate valve). It was previously shown that the Arabidopsis plastidic 2-oxoglutarate/malate transporter (AtpOMT1) and the general dicarboxylate transporter (AtpDCT1) play crucial roles at the interface between carbon and nitrogen metabolism. However, based on the in vitro transport properties of the recombinant transporters, it was hypothesized that AtpOMT1 might play a dual role, also functioning as an oxaloacetate/malate transporter, which is a crucial but currently unidentified component of the chloroplast malate valve. Here, we test this hypothesis using Arabidopsis T-DNA insertional mutants of AtpOMT1. Transport studies revealed a dramatically reduced rate of oxaloacetate uptake into chloroplasts isolated from the knockout plant. CO(2) -dependent O(2) evolution assays showed that cytosolic oxaloacetate is efficiently transported into chloroplasts mainly by AtpOMT1, and supported the absence of additional oxaloacetate transporters. These findings strongly indicate that the high-affinity oxaloacetate transporter in Arabidopsis chloroplasts is AtpOMT1. Further, the knockout plants showed enhanced photo-inhibition under high light due to greater accumulation of reducing equivalents in the stroma, indicating malfunction of the malate valve in the knockout plants. The knockout mutant showed a phenotype consistent with reductions in 2-oxoglutarate transport, glutamine synthetase/glutamate synthase activity, subsequent amino acid biosynthesis and photorespiration. Our results demonstrate that AtpOMT1 acts bi-functionally as an oxaloacetate/malate transporter in the malate valve and as a 2-oxoglutarate/malate transporter mediating carbon/nitrogen metabolism.     

Altered xylem-phloem transfer of amino acids affects metabolism and leads to increased seed yield and oil content in Arabidopsis

Seed development and nitrogen (N) storage depend on delivery of amino acids to seed sinks. For efficient translocation to seeds, amino acids are loaded into the phloem in source leaves and along the long distance transport pathway through xylem-phloem transfer. We demonstrate that Arabidopsis thaliana AMINO ACID PERMEASE2 (AAP2) localizes to the phloem throughout the plant. AAP2 T-DNA insertion lines showed changes in source-sink translocation of amino acids and a decrease in the amount of seed total N and storage proteins, supporting AAP2 function in phloem loading and amino acid distribution to the embryo. Interestingly, in aap2 seeds, total carbon (C) levels were unchanged, while fatty acid levels were elevated. Moreover, branch and silique numbers per plant and seed yield were strongly increased. This suggests changes in N and C delivery to sinks and subsequent modulations of sink development and seed metabolism. This is supported by tracer experiments, expression studies of genes of N/C transport and metabolism in source and sink, and by phenotypic and metabolite analyses of aap2 plants. Thus, AAP2 is key for xylem to phloem transfer and sink N and C supply; moreover, modifications of N allocation can positively affect C assimilation and source-sink transport and benefit sink development and oil yield.