Precise Regulation of the TAA1/TAR-YUCCA Auxin Biosynthesis Pathway in Plants.

The indole-3-pyruvic acid (IPA) pathway is the main auxin biosynthesis pathway in the plant kingdom. Local control of auxin biosynthesis through this pathway regulates plant growth and development and the responses to biotic and abiotic stresses. During the past decades, genetic, physiological, biochemical, and molecular studies have greatly advanced our understanding of tryptophan-dependent auxin biosynthesis. The IPA pathway includes two steps: Trp is converted to IPA by TRYPTOPHAN AMINOTRANSFERASE OF ARABIDOPSIS/TRYPTOPHAN AMINOTRANSFERASE RELATED PROTEINs (TAA1/TARs), and then IPA is converted to IAA by the flavin monooxygenases (YUCCAs). The IPA pathway is regulated at multiple levels, including transcriptional and post-transcriptional regulation, protein modification, and feedback regulation, resulting in changes in gene transcription, enzyme activity and protein localization. Ongoing research indicates that tissue-specific DNA methylation and miRNA-directed regulation of transcription factors may also play key roles in the precise regulation of IPA-dependent auxin biosynthesis in plants. This review will mainly summarize the regulatory mechanisms of the IPA pathway and address the many unresolved questions regarding this auxin biosynthesis pathway in plants.

Rice EIL1 interacts with OsIAAs to regulate auxin biosynthesis mediated by the tryptophan aminotransferase MHZ10/OsTAR2 during root ethylene responses.

Ethylene plays essential roles in adaptive growth of rice (Oryza sativa). Understanding of the crosstalk between ethylene and auxin (Aux) is limited in rice. Here, from an analysis of the root-specific ethylene-insensitive rice mutant mao hu zi 10 (mhz10), we identified the tryptophan aminotransferase (TAR) MHZ10/OsTAR2, which catalyzes the key step in indole-3-pyruvic acid-dependent Aux biosynthesis. Genetically, OsTAR2 acts downstream of ethylene signaling in root ethylene responses. ETHYLENE INSENSITIVE3 like1 (OsEIL1) directly activated OsTAR2 expression. Surprisingly, ethylene induction of OsTAR2 expression still required the Aux pathway. We also show that Os indole-3-acetic acid (IAA)1/9 and OsIAA21/31 physically interact with OsEIL1 and show promotive and repressive effects on OsEIL1-activated OsTAR2 promoter activity, respectively. These effects likely depend on their EAR motif-mediated histone acetylation/deacetylation modification. The special promoting activity of OsIAA1/9 on OsEIL1 may require both the EAR motifs and the flanking sequences for recruitment of histone acetyltransferase. The repressors OsIAA21/31 exhibit earlier degradation upon ethylene treatment than the activators OsIAA1/9 in a TIR1/AFB-dependent manner, allowing OsEIL1 activation by activators OsIAA1/9 for OsTAR2 expression and signal amplification. This study reveals a positive feedback regulation of ethylene signaling by Aux biosynthesis and highlights the crosstalk between ethylene and Aux pathways at a previously underappreciated level for root growth regulation in rice.

The strigolactone receptor SlDWARF14 plays a role in photosynthetic pigment accumulation and photosynthesis in tomato.

KEY MESSAGE: Tomato DWARF14 regulates the development of roots, shoot branches and leaves, and also plays a role in photosynthetic pigment accumulation and photosynthetic capacity. Strigolactones (SLs) are a novel class of plant hormones. DWARF14 (D14) is the only SL receptor identified to date, but it is not functionally analyzed in tomato (Solanum lycopersicum). In the present study, we identified the potential SL receptor in tomato by bioinformatic analysis, which was designated as SlD14. SlD14 was expressed in roots, stems, flowers and developing fruits, with the highest expression level in leaves. sld14 mutant plants produced by the CRISPR/Cas9 system displayed reduced plant height and root biomass, increased shoot branching and altered leaf shape comparing with WT plants. The cytokinin biosynthetic gene ISOPENTENYLTRANSFERASE 3 (SlIPT3), auxin biosynthetic genes FLOOZY (SlFZY) and TRYPTOPHAN AMINOTRANSFERASE RELATED 1 (SlTAR1) and several auxin transport genes SlPINs, which are involved in branch formation, showed higher expression levels in the sld14 plant stem. In addition, sld14 plants exhibited light-green leaves, reduced chlorophyll and carotenoid contents, abnormal chloroplast structure and reduced photosynthetic capacity. Transcriptomic analysis showed that the transcript levels of six chlorophyll biosynthetic genes, three carotenoid biosynthetic genes and numerous chlorophyll a/b-binding protein genes were decreased in sld14 plants. These results suggest that tomato SL receptor gene SlD14 not only regulates the development of roots, shoot branches and leaves, but also plays a role in regulating photosynthetic pigment accumulation and photosynthetic capacity.

Local auxin biosynthesis acts downstream of brassinosteroids to trigger root foraging for nitrogen.

Lateral roots (LRs) dominate the overall root surface of adult plants and are crucial for soil exploration and nutrient acquisition. When grown under mild nitrogen (N) deficiency, flowering plants develop longer LRs to enhance nutrient acquisition. This response is partly mediated by brassinosteroids (BR) and yet unknown mechanisms. Here, we show that local auxin biosynthesis modulates LR elongation while allelic coding variants of YUCCA8 determine the extent of elongation under N deficiency. By up-regulating the expression of YUCCA8/3/5/7 and of Tryptophan Aminotransferase of Arabidopsis 1 (TAA1) under mild N deficiency auxin accumulation increases in LR tips. We further demonstrate that N-dependent auxin biosynthesis in LRs acts epistatic to and downstream of a canonical BR signaling cascade. The uncovered BR-auxin hormonal module and its allelic variants emphasize the importance of fine-tuning hormonal crosstalk to boost adaptive root responses to N availability and offer a path to improve soil exploration by expanded root systems in plants.

Auxin biosynthesis in the phytopathogenic fungus Leptosphaeria maculans is associated with enhanced transcription of indole-3-pyruvate decarboxylase LmIPDC2 and tryptophan aminotransferase LmTAM1.

Auxins are hormones that regulate growth and development in plants. Besides plants, various microorganisms also produce auxins. Here we investigate whether and how the phytopathogenic fungus Leptosphaeria maculans biosynthesizes auxins. We characterized the auxin profile of in vitro grown L. maculans. The culture was further supplied with the auxin biosynthetic-precursors tryptophan and tryptamine and gene expression and phytohormone content was analyzed. L. maculans in vitro produced IAA (indole-3-acetic acid) as the predominant auxin metabolite. IAA production could be further stimulated by supplying precursors. Expression of indole-3-pyruvate decarboxylase LmIPDC2, tryptophan aminotransferase LmTAM1 and nitrilase LmNIT1 genes was mainly upregulated after adding tryptophan and correlated with IAA production, suggesting that these genes are the key components of auxin biosynthesis in L. maculans. Tryptamine acted as a potent inducer of IAA production, though a pathway independent of LmIPDC2/LmTAM1 may be involved. Despite L. maculans being a rich source of bioactive IAA, the auxin metabolic profile of host plant Brassica napus was not altered upon infection. Exogenous IAA inhibited the growth of L. maculans in vitro when supplied in high concentration. Altogether, we showed that L. maculans is capable of IAA production and we have identified biosynthetic genes that were responsive to tryptophan treatment.

Genome-Wide Identification and Analysis on YUCCA Gene Family in Isatis indigotica Fort. and IiYUCCA6-1 Functional Exploration.

Auxin is one of the most critical hormones in plants. YUCCA (Tryptophan aminotransferase of Arabidopsis (TAA)/YUCCA) enzymes catalyze the key rate-limiting step of the tryptophan-dependent auxin biosynthesis pathway, from IPA (Indole-3-pyruvateacid) to IAA (Indole-3-acetic acid). Here, 13 YUCCA family genes were identified from Isatis indigotica, which were divided into four categories, distributing randomly on chromosomes (2n = 14). The typical and conservative motifs, including the flavin adenine dinucleotide (FAD)-binding motif and flavin-containing monooxygenases (FMO)-identifying sequence, existed in the gene structures. IiYUCCA genes were expressed differently in different organs (roots, stems, leaves, buds, flowers, and siliques) and developmental periods (7, 21, 60, and 150 days after germination). Taking IiYUCCA6-1 as an example, the YUCCA genes functions were discussed. The results showed that IiYUCCA6-1 was sensitive to PEG (polyethylene glycol), cold, wounding, and NaCl treatments. The over-expressed tobacco plants exhibited high auxin performances, and some early auxin response genes (NbIAA8, NbIAA16, NbGH3.1, and NbGH3.6) were upregulated with increased IAA content. In the dark, the contents of total chlorophyll and hydrogen peroxide in the transgenic lines were significantly lower than in the control group, with NbSAG12 downregulated and some delayed leaf senescence characteristics, which delayed the senescence process to a certain extent. The findings provide comprehensive insight into the phylogenetic relationships, chromosomal distributions, and expression patterns and functions of the YUCCA gene family in I. indigotica.

Tillering and small grain 1 dominates the tryptophan aminotransferase family required for local auxin biosynthesis in rice.

Auxin is a crucial phytohormone, controlling multiple aspects of plant growth and responses to the changing environment. However, the role of local auxin biosynthesis in specific developmental programs remains unknown in crops. This study characterized the rice tillering and small grain 1 (tsg1) mutant, which has more tillers but a smaller panicle and grain size resulting from a reduction in endogenous auxin. TSG1 encodes a tryptophan aminotransferase that is allelic to the FISH BONE (FIB) gene. The tsg1 mutant showed hypersensitivity to indole-3-acetic acid and the competitive inhibitor of aminotransferase, L-kynurenine. TSG1 knockout resulted in an increased tiller number but reduction in grain number and size, and decrease in height. Meanwhile, deletion of the TSG1 homologs OsTAR1, OsTARL1, and OsTARL2 caused no obvious changes, although the phenotype of the TSG1/OsTAR1 double mutant was intensified and infertile, suggesting gene redundancy in the rice tryptophan aminotransferase family. Interestingly, TSG1 and OsTAR1, but not OsTARL1 and OsTARL2, displayed marked aminotransferase activity. Meanwhile, subcellular localization was identified as the endoplasmic reticulum, while phylogenetic analysis revealed functional divergence of TSG1 and OsTAR1 from OsTARL1 and OsTARL2. These findings suggest that TSG1 dominates the tryptophan aminotransferase family, playing a prominent role in local auxin biosynthesis in rice.

GmGRP-like gene confers Al tolerance in Arabidopsis.

Aluminium (Al) toxicity restrains water and nutrient uptake and is toxic to plant roots, ultimately inhibiting crop production. Here, we isolated and characterized a soybean glycine-rich protein-like gene (GmGRPL) that is mainly expressed in the root and that is regulated by Al treatment. Overexpression of GmGRPL can alleviate Al-induced root growth inhibition in Arabidopsis. The levels of IAA and ethylene in GmGRPL-overexpressing hairy roots were lower than those in control and RNA interference-exposed GmGRPL hairy roots with or without Al stress, which were mainly regulated by TAA1 and ACO, respectively. In transgenic soybean hairy roots, the MDA, H2O2 and O2-·content in GmGRPL-overexpressing hairy roots were less than that in control and RNA interference-exposed GmGRPL hairy roots under Al stress. In addition, IAA and ACC can enhance the expression level of the GmGRPL promoter with or without Al stress. These results indicated that GmGRPL can alleviate Al-induced root growth inhibition by regulating the level of IAA and ethylene and improving antioxidant activity.

Bioinformatics Analysis of Phylogeny and Transcription of TAA/YUC Auxin Biosynthetic Genes.

Auxin is a main plant growth hormone crucial in a multitude of developmental processes in plants. Auxin biosynthesis via the tryptophan aminotransferase of arabidopsis (TAA)/YUCCA (YUC) route involving tryptophan aminotransferases and YUC flavin-dependent monooxygenases that produce the auxin indole-3-acetic acid (IAA) from tryptophan is currently the most researched auxin biosynthetic pathway. Previous data showed that, in maize and arabidopsis, TAA/YUC-dependent auxin biosynthesis can be detected in endoplasmic reticulum (ER) microsomal fractions, and a subset of auxin biosynthetic proteins are localized to the ER, mainly due to transmembrane domains (TMD). The phylogeny presented here for TAA/TAR (tryptophan aminotransferase related) and YUC proteins analyses phylogenetic groups as well as transmembrane domains for ER-membrane localisation. In addition, RNAseq datasets are analysed for transcript abundance of YUC and TAA/TAR proteins in Arabidopsis thaliana. We show that ER membrane localisation for TAA/YUC proteins involved in auxin biosynthesis is already present early on in the evolution of mosses and club mosses. ER membrane anchored YUC proteins can mainly be found in roots, while cytosolic proteins are more abundant in the shoot. The distribution between the different phylogenetic classes in root and shoot may well originate from gene duplications, and the phylogenetic groups detected also overlap with the biological function.

The bHLH transcription factor SPATULA enables cytokinin signaling, and both activate auxin biosynthesis and transport genes at the medial domain of the gynoecium.

Fruits and seeds are the major food source on earth. Both derive from the gynoecium and, therefore, it is crucial to understand the mechanisms that guide the development of this organ of angiosperm species. In Arabidopsis, the gynoecium is composed of two congenitally fused carpels, where two domains: medial and lateral, can be distinguished. The medial domain includes the carpel margin meristem (CMM) that is key for the production of the internal tissues involved in fertilization, such as septum, ovules, and transmitting tract. Interestingly, the medial domain shows a high cytokinin signaling output, in contrast to the lateral domain, where it is hardly detected. While it is known that cytokinin provides meristematic properties, understanding on the mechanisms that underlie the cytokinin signaling pattern in the young gynoecium is lacking. Moreover, in other tissues, the cytokinin pathway is often connected to the auxin pathway, but we also lack knowledge about these connections in the young gynoecium. Our results reveal that cytokinin signaling, that can provide meristematic properties required for CMM activity and growth, is enabled by the transcription factor SPATULA (SPT) in the medial domain. Meanwhile, cytokinin signaling is confined to the medial domain by the cytokinin response repressor ARABIDOPSIS HISTIDINE PHOSPHOTRANSFERASE 6 (AHP6), and perhaps by ARR16 (a type-A ARR) as well, both present in the lateral domains (presumptive valves) of the developing gynoecia. Moreover, SPT and cytokinin, probably together, promote the expression of the auxin biosynthetic gene TRYPTOPHAN AMINOTRANSFERASE OF ARABIDOPSIS 1 (TAA1) and the gene encoding the auxin efflux transporter PIN-FORMED 3 (PIN3), likely creating auxin drainage important for gynoecium growth. This study provides novel insights in the spatiotemporal determination of the cytokinin signaling pattern and its connection to the auxin pathway in the young gynoecium.

Biochemical and Chemical Biology Study of Rice OsTAR1 Revealed that Tryptophan Aminotransferase is Involved in Auxin Biosynthesis: Identification of a Potent OsTAR1 Inhibitor, Pyruvamine2031.

IAA, a major form of auxin, is biosynthesized from l-tryptophan via the indole-3-pyruvic acid (IPyA) pathway in Arabidopsis. Tryptophan aminotransferases (TAA1/TARs) catalyze the first step from l-tryptophan to IPyA. In rice, the importance of TAA/TARs or YUC homologs in auxin biosynthesis has been suggested, but the enzymatic activities and involvement of the intermediate IPyA in auxin biosynthesis remain elusive. In this study, we obtained biochemical evidence that the rice tryptophan aminotransferase OsTAR1 converts l-tryptophan to IPyA, and has a Km of 82.02 µM and a Vmax of 10.92 µM min-1 m-1, comparable with those in Arabidopsis. Next, we screened for an effective inhibitor of OsTAR1 from our previously reported inhibitor library for TAA1/TARs, designated pyruvamine (PVM). Differing from previous observations in Arabidopsis, hydroxy-type PVMs, e.g. PVM2031 (previous name KOK2031), had stronger inhibitory effects in rice than the methoxy-type. PVM2031 inhibited recombinant OsTAR1 in vitro. The Ki of PVM2031 was 276 nM. PVM2031 treatment of rice seedlings resulted in morphological changes in vivo, such as reduced lateral root density. Exogenous IAA rescued this growth inhibition, suggesting that the inhibitory effect is auxin specific. Furthermore, rice roots showed reduced IAA levels concomitant with reduced levels of IPyA in the presence of the inhibitors, suggesting that the IPyA pathway is an auxin biosynthesis pathway in rice. Since PVM2031 showed stronger inhibitory effects on rice auxin biosynthesis than known tryptophan aminotransferase inhibitors, we propose that the hydroxy-type PVM2031 is an effective tool for biochemical analysis of the function of auxin biosynthesis in rice roots.

Metabolic engineering of indole pyruvic acid biosynthesis in Escherichia coli with tdiD.

BACKGROUND: Indole pyruvic acid (IPA) is a versatile platform intermediate and building block for a number of high-value products in the pharmaceutical and food industries. It also has a wide range of applications, such as drugs for the nervous system, cosmetics, and luminophores. Chemical synthesis of IPA is a complicated and costly process. Moreover, through the biosynthesis route employing L-amino acid oxidase, the byproduct hydrogen peroxide leads the degradation of IPA. TdiD, identified as a specific tryptophan aminotransferase, could be an alternative solution for efficient IPA biosynthesis. RESULTS: Escherichia coli strain W3110, which demonstrates basic production when supplied with tryptophan, was engineered for IPA biosynthesis. Several strategies were implemented to improve IPA production. First, through incorporating the codon-optimized tdiD into W3110, IPA levels increased from 41.54 ± 1.26 to 52.54 ± 2.08 mg/L. Second, after verifying the benefit of an increased phenylpyruvate pool, a YL03 strain was constructed based on a previously reported mutant strain of W3110 with a plasmid carrying aroF fbr and pheA fbr to further improve IPA production. The recombinant YL03 strain accumulated IPA at 158.85 ± 5.36 mg/L, which was 3.82-fold higher than that of the wild-type W3110 strain. Third, optimization of tdiD co expression was carried out by replacing the Trc promoter with a series of constitutively active promoters along with increasing the plasmid copy numbers. The highest IPA production was observed in YL08, which achieved 236.42 ± 17.66 mg/L and represented a greater than 5-fold increase as compared to W3110. Finally, the effects of deletion and overexpression of tnaA on IPA biosynthesis were evaluated. The removal of tnaA led to slightly reduced IPA levels, whereas the overexpression of tnaA resulted in a considerable decline in production. CONCLUSIONS: This study illustrates the feasibility of IPA biosynthesis in E. coli through tdiD. An efficient IPA producing strain, YL08, was developed, which provides a new possibility for biosynthesis of IPA. Although the final production was limited, this study demonstrates a convenient method of IPA synthesis.

Forward genetic screen for auxin-deficient mutants by cytokinin.

Identification of mutants with impairments in auxin biosynthesis and dynamics by forward genetic screening is hindered by the complexity, redundancy and necessity of the pathways involved. Furthermore, although a few auxin-deficient mutants have been recently identified by screening for altered responses to shade, ethylene, N-1-naphthylphthalamic acid (NPA) or cytokinin (CK), there is still a lack of robust markers for systematically isolating such mutants. We hypothesized that a potentially suitable phenotypic marker is root curling induced by CK, as observed in the auxin biosynthesis mutant CK-induced root curling 1 / tryptophan aminotransferase of Arabidopsis 1 (ckrc1/taa1). Phenotypic observations, genetic analyses and biochemical complementation tests of Arabidopsis seedlings displaying the trait in large-scale genetic screens showed that it can facilitate isolation of mutants with perturbations in auxin biosynthesis, transport and signaling. However, unlike transport/signaling mutants, the curled (or wavy) root phenotypes of auxin-deficient mutants were significantly induced by CKs and could be rescued by exogenous auxins. Mutants allelic to several known auxin biosynthesis mutants were re-isolated, but several new classes of auxin-deficient mutants were also isolated. The findings show that CK-induced root curling provides an effective marker for discovering genes involved in auxin biosynthesis or homeostasis.

Auxin Import and Local Auxin Biosynthesis Are Required for Mitotic Divisions, Cell Expansion and Cell Specification during Female Gametophyte Development in Arabidopsis thaliana.

The female gametophyte of flowering plants, called the embryo sac, develops from a haploid cell named the functional megaspore, which is specified after meiosis by the diploid sporophyte. In Arabidopsis, the functional megaspore undergoes three syncitial mitotic divisions followed by cellularization to form seven cells of four cell types including two female gametes. The plant hormone auxin is important for sporophytic developmental processes, and auxin levels are known to be regulated by biosynthesis and transport. Here, we investigated the role of auxin biosynthetic genes and auxin influx carriers in embryo sac development. We find that genes from the YUCCA/TAA pathway (YUC1, YUC2, YUC8, TAA1, TAR2) are expressed asymmetrically in the developing ovule and embryo sac from the two-nuclear syncitial stage until cellularization. Mutants for YUC1 and YUC2 exhibited defects in cell specification, whereas mutations in YUC8, as well as mutations in TAA1 and TAR2, caused defects in nuclear proliferation, vacuole formation and anisotropic growth of the embryo sac. Additionally, expression of the auxin influx carriers AUX1 and LAX1 were observed at the micropylar pole of the embryo sac and in the adjacent cells of the ovule, and the aux1 lax1 lax2 triple mutant shows multiple gametophyte defects. These results indicate that both localized auxin biosynthesis and auxin import, are required for mitotic divisions, cell expansion and patterning during embryo sac development.

TAA1-regulated local auxin biosynthesis in the root-apex transition zone mediates the aluminum-induced inhibition of root growth in Arabidopsis.

The transition zone (TZ) of the root apex is the perception site of Al toxicity. Here, we show that exposure of Arabidopsis thaliana roots to Al induces a localized enhancement of auxin signaling in the root-apex TZ that is dependent on TAA1, which encodes a Trp aminotransferase and regulates auxin biosynthesis. TAA1 is specifically upregulated in the root-apex TZ in response to Al treatment, thus mediating local auxin biosynthesis and inhibition of root growth. The TAA1-regulated local auxin biosynthesis in the root-apex TZ in response to Al stress is dependent on ethylene, as revealed by manipulating ethylene homeostasis via the precursor of ethylene biosynthesis 1-aminocyclopropane-1-carboxylic acid, the inhibitor of ethylene biosynthesis aminoethoxyvinylglycine, or mutant analysis. In response to Al stress, ethylene signaling locally upregulates TAA1 expression and thus auxin responses in the TZ and results in auxin-regulated root growth inhibition through a number of auxin response factors (ARFs). In particular, ARF10 and ARF16 are important in the regulation of cell wall modification-related genes. Our study suggests a mechanism underlying how environmental cues affect root growth plasticity through influencing local auxin biosynthesis and signaling.

Plant roots use a patterning mechanism to position lateral root branches toward available water.

The architecture of the branched root system of plants is a major determinant of vigor. Water availability is known to impact root physiology and growth; however, the spatial scale at which this stimulus influences root architecture is poorly understood. Here we reveal that differences in the availability of water across the circumferential axis of the root create spatial cues that determine the position of lateral root branches. We show that roots of several plant species can distinguish between a wet surface and air environments and that this also impacts the patterning of root hairs, anthocyanins, and aerenchyma in a phenomenon we describe as hydropatterning. This environmental response is distinct from a touch response and requires available water to induce lateral roots along a contacted surface. X-ray microscale computed tomography and 3D reconstruction of soil-grown root systems demonstrate that such responses also occur under physiologically relevant conditions. Using early-stage lateral root markers, we show that hydropatterning acts before the initiation stage and likely determines the circumferential position at which lateral root founder cells are specified. Hydropatterning is independent of endogenous abscisic acid signaling, distinguishing it from a classic water-stress response. Higher water availability induces the biosynthesis and transport of the lateral root-inductive signal auxin through local regulation of tryptophan aminotransferase of Arabidopsis 1 and PIN-formed 3, both of which are necessary for normal hydropatterning. Our work suggests that water availability is sensed and interpreted at the suborgan level and locally patterns a wide variety of developmental processes in the root.

The rice FISH BONE gene encodes a tryptophan aminotransferase, which affects pleiotropic auxin-related processes.

Auxin is a fundamental plant hormone and its localization within organs plays pivotal roles in plant growth and development. Analysis of many Arabidopsis mutants that were defective in auxin biosynthesis revealed that the indole-3-pyruvic acid (IPA) pathway, catalyzed by the TRYPTOPHAN AMINOTRANSFERASE OF ARABIDOPSIS (TAA) and YUCCA (YUC) families, is the major biosynthetic pathway of indole-3-acetic acid (IAA). In contrast, little information is known about the molecular mechanisms of auxin biosynthesis in rice. In this study, we identified a auxin-related rice mutant, fish bone (fib). FIB encodes an orthologue of TAA genes and loss of FIB function resulted in pleiotropic abnormal phenotypes, such as small leaves with large lamina joint angles, abnormal vascular development, small panicles, abnormal organ identity and defects in root development, together with a reduction in internal IAA levels. Moreover, we found that auxin sensitivity and polar transport activity were altered in the fib mutant. From these results, we suggest that FIB plays a pivotal role in IAA biosynthesis in rice and that auxin biosynthesis, transport and sensitivity are closely interrelated.

Auxin biosynthetic gene TAR2 is involved in low nitrogen-mediated reprogramming of root architecture in Arabidopsis.

In plants, the plasticity of root architecture in response to nitrogen availability largely determines nitrogen acquisition efficiency. One poorly understood root growth response to low nitrogen availability is an observed increase in the number and length of lateral roots (LRs). Here, we show that low nitrogen-induced Arabidopsis LR growth depends on the function of the auxin biosynthesis gene TAR2 (tryptophan aminotransferase related 2). TAR2 was expressed in the pericycle and the vasculature of the mature root zone near the root tip, and was induced under low nitrogen conditions. In wild type plants, low nitrogen stimulated auxin accumulation in the non-emerged LR primordia with more than three cell layers and LR emergence. Conversely, these low nitrogen-mediated auxin accumulation and root growth responses were impaired in the tar2-c null mutant. Overexpression of TAR2 increased LR numbers under both high and low nitrogen conditions. Our results suggested that TAR2 is required for reprogramming root architecture in response to low nitrogen conditions. This finding suggests a new strategy for improving nitrogen use efficiency through the engineering of TAR2 expression in roots.

Genetic aspects of auxin biosynthesis and its regulation.

Auxin is an essential plant hormone that controls nearly every aspect of a plant's life, from embryo development to organ senescence. In the last decade the key genes involved in auxin transport, perception, signaling and response have been identified and characterized, but the elucidation of auxin biosynthesis has proven to be especially challenging. In plants, a significant amount of indole-3-acetic acid (IAA), the predominant biologically active form of auxin, is synthesized via a simple two-step route where indole-3-pyruvic acid (IPyA) produced from l-tryptophan by tryptophan aminotransferases (TAA1/TAR) is converted to IAA by the YUC family of flavin monooxygenases. The TAA1/TAR and YUC gene families constitute the first complete auxin biosynthetic pathway described in plants. Detailed characterization of these genes' expression patterns suggested a key role of local auxin biosynthesis in plant development. This has prompted an active search for the molecular mechanisms that regulate the spatiotemporal activity of the IPyA route. In addition to the TAA1/TAR and YUC-mediated auxin biosynthesis, several alternative routes of IAA production have been postulated to function in plants, but their biological significance is yet to be demonstrated. Herein, we take a genetic perspective to describe the current view of auxin biosynthesis and its regulation in plants, focusing primarily on Arabidopsis.

Yucasin is a potent inhibitor of YUCCA, a key enzyme in auxin biosynthesis.

Indole-3-acetic acid (IAA), an auxin plant hormone, is biosynthesized from tryptophan. The indole-3-pyruvic acid (IPyA) pathway, involving the tryptophan aminotransferase TAA1 and YUCCA (YUC) enzymes, was recently found to be a major IAA biosynthetic pathway in Arabidopsis. TAA1 catalyzes the conversion of tryptophan to IPyA, and YUC produces IAA from IPyA. Using a chemical biology approach with maize coleoptiles, we identified 5-(4-chlorophenyl)-4H-1,2,4-triazole-3-thiol (yucasin) as a potent inhibitor of IAA biosynthesis in YUC-expressing coleoptile tips. Enzymatic analysis of recombinant AtYUC1-His suggested that yucasin strongly inhibited YUC1-His activity against the substrate IPyA in a competitive manner. Phenotypic analysis of Arabidopsis YUC1 over-expression lines (35S::YUC1) demonstrated that yucasin acts in IAA biosynthesis catalyzed by YUC. In addition, 35S::YUC1 seedlings showed resistance to yucasin in terms of root growth. A loss-of-function mutant of TAA1, sav3-2, was hypersensitive to yucasin in terms of root growth and hypocotyl elongation of etiolated seedlings. Yucasin combined with the TAA1 inhibitor l-kynurenine acted additively in Arabidopsis seedlings, producing a phenotype similar to yucasin-treated sav3-2 seedlings, indicating the importance of IAA biosynthesis via the IPyA pathway in root growth and leaf vascular development. The present study showed that yucasin is a potent inhibitor of YUC enzymes that offers an effective tool for analyzing the contribution of IAA biosynthesis via the IPyA pathway to plant development and physiological processes.