Indole-3-acetic Acid Metabolism Research Articles

Analytical methods for stable isotope labeling to elucidate rapid auxin kinetics in Arabidopsis thaliana.

The phytohormone auxin plays a critical role in plant growth and development. Despite significant progress in elucidating metabolic pathways of the primary bioactive auxin, indole-3-acetic acid (IAA), over the past few decades, key components such as intermediates and enzymes have not been fully characterized, and the dynamic regulation of IAA metabolism in response to environmental signals has not been completely revealed. In this study, we established a protocol employing a highly sensitive liquid chromatography-mass spectrometry (LC-MS) instrumentation and a rapid stable isotope labeling approach. We treated Arabidopsis seedlings with two stable isotope labeled precursors ([13C6]anthranilate and [13C8, 15N1]indole) and monitored the label incorporation into proposed indolic compounds involved in IAA biosynthetic pathways. This Stable Isotope Labeled Kinetics (SILK) method allowed us to trace the turnover rates of IAA pathway precursors and product concurrently with a time scale of seconds to minutes. By measuring the entire pathways over time and using different isotopic tracer techniques, we demonstrated that these methods offer more detailed information about this complex interacting network of IAA biosynthesis, and should prove to be useful for studying auxin metabolic network in vivo in a variety of plant tissues and under different environmental conditions.

Exploring the Relationship between Gut Microbiome Composition and Blood Indole-3-acetic Acid in Hemodialysis Patients.

Indole-3-acetic acid (IAA), a protein-bound uremic toxin resulting from gut microbiota-driven tryptophan metabolism, increases in hemodialysis (HD) patients. IAA may induce endothelial dysfunction, inflammation, and oxidative stress, elevating cardiovascular and cognitive risk in HD patients. However, research on the microbiome-IAA association is limited. This study aimed to explore the gut microbiome's relationship with plasma IAA levels in 72 chronic HD patients aged over 18 (August 2016-January 2017). IAA levels were measured using tandem mass spectrometry, and gut microbiome analysis utilized 16s rRNA next-generation sequencing. Linear discriminative analysis effect size and random forest analysis distinguished microbial species linked to IAA levels. Patients with higher IAA levels had reduced microbial diversity. Six microbial species significantly associated with IAA levels were identified; Bacteroides clarus, Bacteroides coprocola, Bacteroides massiliensi, and Alisteps shahii were enriched in low-IAA individuals, while Bacteroides thetaiotaomicron and Fusobacterium varium were enriched in high-IAA individuals. This study sheds light on specific gut microbiota species influencing IAA levels, enhancing our understanding of the intricate interactions between the gut microbiota and IAA metabolism.

NnWOX1-1, NnWOX4-3, and NnWOX5-1 of lotus (Nelumbo nucifera Gaertn)promote root formation and enhance stress tolerance in transgenic Arabidopsis thaliana

BackgroundAdventitious roots (ARs) represent an important organ system for water and nutrient uptake in lotus plants because of degeneration of the principal root. The WUSCHEL-related homeobox (WOX) gene regulates plant development and growth by affecting the expression of several other genes. In this study, three WOX genes, NnWOX1-1, NnWOX4-3, and NnWOX5-1, were isolated and their functions were assessed in Arabidopsis plants.ResultsThe full lengths of NnWOX1-1, NnWOX4-3, and NnWOX5-1 were 1038, 645, and 558 bp, encoding 362, 214, and 185 amino acid residues, respectively. Phylogenetic analysis classified NnWOX1-1 and NnWOX4-3 encoding proteins into one group, and NnWOX5-1 and MnWOX5 encoding proteins exhibited strong genetic relationships. The three genes were induced by sucrose and indoleacetic acid (IAA) and exhibited organ-specific expression characteristics. In addition to improving root growth and salt tolerance, NnWOX1-1 and NnWOX4-3 promoted stem development in transgenic Arabidopsis plants. A total of 751, 594, and 541 genes, including 19, 19, and 13 respective genes related to ethylene and IAA metabolism and responses, were enhanced in NnWOX1-1, NnWOX4-3, and NnWOX5-1 transgenic plants, respectively. Further analysis showed that ethylene production rates in transgenic plants increased, whereas IAA, peroxidase, and lignin content did not significantly change. Exogenous application of ethephon on lotus seedlings promoted AR formation and dramatically increased the fresh and dry weights of the plants.ConclusionsNnWOX1-1, NnWOX4-3, and NnWOX5-1 influence root formation, stem development, and stress adaptation in transgenic Arabidopsis plants by affecting the transcription of multiple genes. Among these, changes in gene expression involving ethylene metabolism and responses likely critically affect the development of Arabidopsis plants. In addition, ethylene may represent an important factor affecting AR formation in lotus seedlings.

Effects of Zinc Oxide Particles with Different Sizes on Root Development in Oryza sativa

Given the consistent release of zinc oxide (ZnO) nanoparticles into the environment, it is urgent to study their impact on plants in depth. In this study, grains of rice were treated with two different concentrations of ZnO nanoparticles (NP-ZnO, 10 and 100 mg/L), and their bulk counterpart (B-ZnO) were used to evaluate whether ZnO action could depend on particle size. To test this hypothesis, root growth and development assessment, oxidative stress parameters, indole-3-acetic acid (IAA) content and molecules/enzymes involved in IAA metabolism were analyzed. In situ localization of Zn in control and treated roots was also performed. Though Zn was visible inside root cells only following nanoparticle treatment, both materials (NP-ZnO and B-ZnO) were able to affect seedling growth and root morphology, with alteration in the concentration/pattern of localization of oxidative stress markers and with a different action depending on particle size. In addition, only ZnO supplied as bulk material induced a significant increase in both IAA concentration and lateral root density, supporting our hypothesis that bulk particles might enhance lateral root development through the rise of IAA concentration. Apparently, IAA concentration was influenced more by the activity of the catabolic peroxidases than by the protective action of phenols.

The Nutrient Zinc in Soil and Plant: A Review

Zinc is an element found in low amounts in the soil, which may limit the development of agricultural crops. In addition, the absorption of this nutrient by plants is directly influenced by several factors such as: high pH, ​​amount of organic matter, high humidity associated with low temperatures and soil microbiota. Zinc is important for enzymatic activity, being a constituent part of the enzymes alcohol dehydrogenase, carbonic anhydrase, superoxide dismutase enzyme and polymer RNA, in addition to participating in the synthesis of precursor tryptophan in the metabolism of indoleacetic acid, which is a plant hormone directly related to the development of plants. In plants, the symptoms of zinc deficiency appear with the shortening of the internodes, reduction of the leaf area, reduction of its size, appearance of rosettes, chlorosis and necrosis. Thus, the objective of this review is to present the main aspects of the zinc nutrient in the soil and the functions performed in plants.

Key factors for differential drought tolerance in two contrasting wild materials of Artemisia wellbyi identified using comparative transcriptomics

BackgroundDrought is a significant condition that restricts vegetation growth on the Tibetan Plateau. Artemisia wellbyi is a unique semi-shrub-like herb in the family Compositae, which distributed in northern and northwest of Tibetan Plateau. It is a dominant species in the community that can well adapt to virous environment stress, such as drought and low temperature. Therefore, A. wellbyi. has a potential ecological value for soil and water conservation of drought areas. Understanding the molecular mechanisms of A. wellbyi. that defense drought stress can acquire the key genes for drought resistance breeding of A. wellbyi. and provide a theoretical basis for vegetation restoration of desertification area. However, they remain unclear. Thus, our study compared the transcriptomic characteristics of drought-tolerant “11” and drought-sensitive “6” material of A. wellbyi under drought stress.ResultsA total of 4875 upregulated and 4381 downregulated differentially expressed genes (DEGs) were induced by drought in the tolerant material; however, only 1931 upregulated and 4174 downregulated DEGs were induced by drought in the sensitive material. The photosynthesis and transcriptional regulation differed significantly with respect to the DEGs number and expression level. We found that CDPKs (calmodulin-like domain protein kinases), SOS3 (salt overly sensitive3), MAPKs (mitogen-activated protein kinase cascades), RLKs (receptor like kinase), and LRR-RLKs (repeat leucine-rich receptor kinase) were firstly involved in response to drought stress in drought tolerant A. wellbyi. Positive regulation of genes associated with the metabolism of ABA (abscisic acid), ET (ethylene), and IAA (indole acetic acid) could play a crucial role in the interaction with other transcriptional regulatory factors, such as MYBs (v-myb avian myeloblastosis viral oncogene homolog), AP2/EREBPs (APETALA2/ethylene-responsive element binding protein family), WRKYs, and bHLHs (basic helix-loop-helix family members) and receptor kinases, and regulate downstream genes for defense against drought stress. In addition, HSP70 (heat shock protein70) and MYB73 were considered as the hub genes because of their strong association with other DEGs.ConclusionsPositive transcriptional regulation and negative regulation of photosynthesis could be associated with better growth performance under drought stress in the drought-tolerant material. In addition, the degradation of sucrose and starch in the tolerant A. wellbyi to alleviate osmotic stress and balance excess ROS. These results highlight the candidate genes that are involved in enhancing the performance of drought-tolerant A. wellbyi and provide a theoretical basis for improving the performance of drought-resistant A. wellbyi.

Indole-Acetic Acid Promotes Ammonia Removal Through Heterotrophic Nitrification, Aerobic Denitrification With Mixed Enterobacter sp. Z1 and Klebsiella sp. Z2.

Mixed Enterobacter sp. Z1 and Klebsiella sp. Z2 displayed an outstanding ammonia removal capacity than using a single strain. Metabolomics, proteomics, and RNA interference analysis demonstrated that the HNAD process was closely related to indole-acetic acid (IAA). Under the cocultured conditions, the excess IAA produced by Z2 could be absorbed by Z1 to compensate for the deficiency of IAA in the cells. IAA directly induced the expression of denitrifying enzymes and further activated the IAA metabolism level, thus greatly improving the nitrogen removal ability of Z1. In turn, nitrate and nitrite induced the expression of key enzymes in the IAA pathways. Moreover, Z1 and Z2 enhanced two IAA metabolic pathways in the process of mixed removal process. The activated hydrolysis-redox pathway in Z1 reduced the oxidative stress level, and the activated decarboxylation pathway in Z2 promoted intracellular energy metabolism, which indirectly promoted the process of HNAD in the system.

Proline-rich protein MdPRP6 alters low nitrogen stress tolerance by regulating lateral root formation and anthocyanin accumulation in transgenic apple (Malus domestica)

Nitrogen (N) is the main nutrient element limiting plant growth and crop yield. Plant proline-rich proteins (PRPs) are a group of structural proteins that are rich in proline and hydroxyproline . Previously, we identified 9 PRPs from the apple genome and found that they were induced by different stress treatments. In this study, we showed that the expression of MdPRP6 was inhibited by low N stress. Transgenic apple plants with decreased MdPRP6 expression were generated by RNA interference (Ri) to explore the biological role of MdPRP6 under low N stress. Ri transgenic apple plants grew better than wild-type (WT) plants under low N stress, as indicated by their well-developed roots; greater height, stem diameter, number of leaves, and biomass; and higher photosynthetic capacity and N content. N and starch metabolism were more active in Ri plants than in WT plants. Ri plants accumulated greater amounts of starch and soluble sugar under low N stress, which was consistent with the higher expression levels of related genes in Ri plants relative to WT plants. The accumulation of anthocyanins was higher in Ri plants than in WT plants under low N stress, which helped plants resist the effects of low N stress. The indole acetic acid (IAA) content was higher in the roots of Ri plants, which strengthened the root system. Overall, our data suggested that MdPRP6 negatively regulates low N stress tolerance by modulating anthocyanin accumulation and IAA metabolism in the roots of apple plants. • RNAi of MdPRP6 resulted in more LR in transgenic apples under LN stress. • RNAi of MdPRP6 conferred more accumulation of anthocyanins in transgenic apples under LN stress. • The tolerance of LN stress conferred by MdPRP6 was related to the soluble sugars content.

Inactivation of the entire Arabidopsis group II GH3s confers tolerance to salinity and water deficit.

Summary Indole‐3‐acetic acid (IAA) controls a plethora of developmental processes. Thus, regulation of its concentration is of great relevance for plant performance. Cellular IAA concentration depends on its transport, biosynthesis and the various pathways for IAA inactivation, including oxidation and conjugation.Group II members of the GRETCHEN HAGEN 3 (GH3) gene family code for acyl acid amido synthetases catalysing the conjugation of IAA to amino acids. However, the high degree of functional redundancy among them has hampered thorough analysis of their roles in plant development.In this work, we generated an Arabidopsis gh3.1,2,3,4,5,6,9,17 (gh3oct) mutant to knock out the group II GH3 pathway. The gh3oct plants had an elaborated root architecture, showed an increased tolerance to different osmotic stresses, including an IAA‐dependent tolerance to salinity, and were more tolerant to water deficit. Indole‐3‐acetic acid metabolite quantification in gh3oct plants suggested the existence of additional GH3‐like enzymes in IAA metabolism. Moreover, our data suggested that 2‐oxindole‐3‐acetic acid production depends, at least in part, on the GH3 pathway. Targeted stress‐hormone analysis further suggested involvement of abscisic acid in the differential response to salinity of gh3oct plants.Taken together, our data provide new insights into the roles of group II GH3s in IAA metabolism and hormone‐regulated plant development.

Uniconazole Augments Abscisic Acid in Promoting Somatic Embryogenesis in Cotton (Gossypium hirsutum L.).

During somatic embryogenesis (SE), somatic cells initiate embryogenic development under appropriate conditions. Uniconazole, a plant growth regulator, was found to inhibit the proliferation of callus but promoted the conversion of callus into an embryogenic callus (EC) in cotton. The supplementation of uniconazole in the culture medium significantly suppressed the endogenous auxin [indole acetic acid (IAA)] level in callus tissues in both the callus initiation and proliferation stage but enhanced the abscisic acid (ABA) level only in the callus proliferation stage. Exogenous ABA and uniconazole showed cooperative effects on promoting the differentiation rate of callus into EC. These findings were verified by RNA-seq analysis, which elucidated that the genes involved in the IAA biosynthesis, metabolism, and signaling, and ABA metabolism pathways were regulated by uniconazole during the callus development and SE. Overall, the results suggest that uniconazole could modulate callus proliferation and callus differentiation rate by regulating the endogenous levels of IAA and ABA.

Molecular mechanisms and hormonal regulation underpinning morphological dormancy: a case study using Apium graveolens (Apiaceae).

Underdeveloped (small) embryos embedded in abundant endosperm tissue, and thus having morphological dormancy (MD) or morphophysiological dormancy (MPD), are considered to be the ancestral state in seed dormancy evolution. This trait is retained in the Apiaceae family, which provides excellent model systems for investigating the underpinning mechanisms. We investigated Apium graveolens (celery) MD by combined innovative imaging and embryo growth assays with the quantification of hormone metabolism, as well as the analysis of hormone and cell-wall related gene expression. The integrated experimental results demonstrated that embryo growth occurred inside imbibed celery fruits in association with endosperm degradation, and that a critical embryo size was required for radicle emergence. The regulation of these processes depends on gene expression leading to gibberellin and indole-3-acetic acid (IAA) production by the embryo and on crosstalk between the fruit compartments. ABA degradation associated with distinct spatiotemporal patterns in ABA sensitivity control embryo growth, endosperm breakdown and radicle emergence. This complex interaction between gibberellins, IAA and ABA metabolism, and changes in the tissue-specific sensitivities to these hormones is distinct from non-MD seeds. We conclude that the embryo growth to reach the critical size and the associated endosperm breakdown inside MD fruits constitute a unique germination programme.

Aldoximes are precursors of auxins in Arabidopsis and maize.

Two natural auxins, phenylacetic acid (PAA) and indole-3-acetic acid (IAA), play crucial roles in plant growth and development. One route of IAA biosynthesis uses the glucosinolate intermediate indole-3-acetaldoxime (IAOx) as a precursor, which is thought to occur only in glucosinolate-producing plants in Brassicales. A recent study showed that overproducing phenylacetaldoxime (PAOx) in Arabidopsis increases PAA production. However, it remains unknown whether this increased PAA resulted from hydrolysis of PAOx-derived benzyl glucosinolate or, like IAOx-derived IAA, is directly converted from PAOx. If glucosinolate hydrolysis is not required, aldoxime-derived auxin biosynthesis may occur beyond Brassicales. To better understand aldoxime-derived auxin biosynthesis, we conducted an isotope-labelled aldoxime feeding assay using an Arabidopsis glucosinolate-deficient mutant sur1 and maize, and transcriptomics analysis. Our study demonstrated that the conversion of PAOx to PAA does not require glucosinolates in Arabidopsis. Furthermore, maize produces PAA and IAA from PAOx and IAOx, respectively, indicating that aldoxime-derived auxin biosynthesis also occurs in maize. Considering that aldoxime production occurs widely in the plant kingdom, aldoxime-derived auxin biosynthesis is likely to be more widespread than originally believed. A genome-wide transcriptomics study using PAOx-overproduction plants identified complex metabolic networks among IAA, PAA, phenylpropanoid and tryptophan metabolism.

Auxin Metabolism in Plants.

The major natural auxin in plants, indole-3-acetic acid (IAA), orchestrates a plethora of developmental responses that largely depend on the formation of auxin concentration gradients within plant tissues. Together with inter- and intracellular transport, IAA metabolism-which comprises biosynthesis, conjugation, and degradation-modulates auxin gradients and is therefore critical for plant growth. It is now very well established that IAA is mainly produced from Trp and that the IPyA pathway is a major and universally conserved biosynthetic route in plants, while other redundant pathways operate in parallel. Recent findings have shown that metabolic inactivation of IAA is also redundantly performed by oxidation and conjugation processes. An exquisite spatiotemporal expression of the genes for auxin synthesis and inactivation have been shown to drive several plant developmental processes. Moreover, a group of transcription factors and epigenetic regulators controlling the expression of auxin metabolic genes have been identified in past years, which are illuminating the road to understanding the molecular mechanisms behind the coordinated responses of local auxin metabolism to specific cues. Besides transcriptional regulation, subcellular compartmentalization of the IAA metabolism and posttranslational modifications of the metabolic enzymes are emerging as important contributors to IAA homeostasis. In this review, we summarize the current knowledge on (1) the pathways for IAA biosynthesis and inactivation in plants, (2) the influence of spatiotemporally regulated IAA metabolism on auxin-mediated responses, and (3) the regulatory mechanisms that modulate IAA levels in response to external and internal cues during plant development.

UDP-glucosyltransferase UGT84B1 regulates the levels of indole-3-acetic acid and phenylacetic acid in Arabidopsis

Auxin is a key plant growth regulator for diverse developmental processes in plants. Indole-3-acetic acid (IAA) is a primary plant auxin that regulates the formation of various organs. Plants also produce phenylacetic acid (PAA), another natural auxin, which occurs more abundantly than IAA in various plant species. Although it has been demonstrated that the two auxins have distinct transport characteristics, the metabolic pathways and physiological roles of PAA in plants remain unsolved. In this study, we investigated the role of Arabidopsis UDP-glucosyltransferase UGT84B1 in IAA and PAA metabolism. We demonstrated that UGT84B1, which converts IAA to IAA-glucoside (IAA-Glc), can also catalyze the conversion of PAA to PAA-glucoside (PAA-Glc), with a higher catalytic activity in vitro. Furthermore, we showed a significant increase in both the IAA and PAA levels in the ugt84b1 null mutants. However, no obvious developmental phenotypes were observed in the ugt84b1 mutants under laboratory growth conditions. Moreover, the overexpression of UGT84B1 resulted in auxin-deficient root phenotypes and changes in the IAA and PAA levels. Our results indicate that UGT84B1 plays an important role in IAA and PAA homeostasis in Arabidopsis.

Role of Arabidopsis INDOLE-3-ACETIC ACID CARBOXYL METHYLTRANSFERASE 1 in auxin metabolism

The phytohormone auxin regulates a wide range of developmental processes in plants. Indole-3-acetic acid (IAA) is the main auxin that moves in a polar manner and forms concentration gradients, whereas phenylacetic acid (PAA), another natural auxin, does not exhibit polar movement. Although these auxins occur widely in plants, the differences between IAA and PAA metabolism remain largely unknown. In this study, we investigated the role of Arabidopsis IAA CARBOXYL METHYLTRANSFERASE 1 (IAMT1) in IAA and PAA metabolism. IAMT1 proteins expressed in Escherichia coli could convert both IAA and PAA to their respective methyl esters. Overexpression of IAMT1 caused severe auxin-deficient phenotypes and reduced the levels of IAA, but not PAA, in the root tips of Arabidopsis, suggesting that IAMT1 exclusively metabolizes IAA in vivo. We generated iamt1 null mutants via CRISPR/Cas9-mediated genome editing and found that the single knockout mutants had normal auxin levels and did not exhibit visibly altered phenotypes. These results suggest that other proteins, namely the IAMT1 homologs in the SABATH family of carboxyl methyltransferases, may also regulate IAA levels in Arabidopsis.

Genome-based identification and analysis of the genes involved in auxin biosynthesis and signal transduction during tea plant leaf development

Auxin plays crucial roles in the process of plant growth and development. Among all the organs of the tea plant, the leaves are the main edible part for humans. The appearance and endoplasm of tea leaves directly affects taste, quality, and economic value. The metabolism of Indole-3-acetic acid (IAA) in tea plants changes the growth and development of leaves in certain aspects. However, the molecular mechanism of IAA metabolism during the growth and development in tea leaves remains unclear. To explore the molecular mechanism of IAA metabolism in tea leaves, the samples were collected from the three different development stages of tea leaves. Cell morphology and IAA contents were also investigated during tea leaves development. A total of 16 genes involved in IAA biosynthesis and signal transduction pathways were identified based on the genome sequence of Camellia sinensis. CsTSA, CsIAA27, CsTIR1 and CsCH3-6 expression patterns exhibited positive correlation with IAA concentration in tea leaves. In contrast, CsYUC1, CsCYP83B1 and CsLAX1 transcriptions were negatively correlated with IAA contents during tea leaves ripening. The interaction of IAA metabolism related proteins was also discussed. The results indicated that the growth and development of tea leaves could be regulated by IAA metabolism in the form of related-genes differentially expression and protein-protein interaction. This study provided a further glimpse to IAA metabolic pathways in leaf during tea plant growth and development.

Computationally Guided Discovery and Experimental Validation of Indole-3-acetic Acid Synthesis Pathways.

Elucidating the interaction networks associated with secondary metabolite production in microorganisms is an ongoing challenge made all the more daunting by the rate at which DNA sequencing technology reveals new genes and potential pathways. Developing the culturing methods, expression conditions, and genetic systems needed for validating pathways in newly discovered microorganisms is often not possible. Therefore, new tools and techniques are needed for defining complex metabolic pathways. Here, we describe an in vitro computationally assisted pathway description approach that employs bioinformatic searches of genome databases, protein structural modeling, and protein-ligand-docking simulations to predict the gene products most likely to be involved in a particular secondary metabolite production pathway. This information is then used to direct in vitro reconstructions of the pathway and subsequent confirmation of pathway activity using crude enzyme preparations. As a test system, we elucidated the pathway for biosynthesis of indole-3-acetic acid (IAA) in the plant-associated microbe Pantoea sp. YR343. This organism is capable of metabolizing tryptophan into the plant phytohormone IAA. BLAST analyses identified a likely three-step pathway involving an amino transferase, an indole pyruvate decarboxylase, and a dehydrogenase. However, multiple candidate enzymes were identified at each step, resulting in a large number of potential pathway reconstructions (32 different enzyme combinations). Our approach shows the effectiveness of crude extracts to rapidly elucidate enzymes leading to functional pathways. Results are compared to affinity purified enzymes for select combinations and found to yield similar relative activities. Further, in vitro testing of the pathway reconstructions revealed the "underground" nature of IAA metabolism in Pantoea sp. YR343 and the various mechanisms used to produce IAA. Importantly, our experiments illustrate the scalable integration of computational tools and cell-free enzymatic reactions to identify and validate metabolic pathways in a broadly applicable manner.

The Effects of DNA Methylation Inhibition on Flower Development in the Dioecious Plant Salix Viminalis

DNA methylation, an important epigenetic modification, regulates the expression of genes and is therefore involved in the transitions between floral developmental stages in flowering plants. To explore whether DNA methylation plays different roles in the floral development of individual male and female dioecious plants, we injected 5-azacytidine (5-azaC), a DNA methylation inhibitor, into the trunks of female and male basket willow (Salix viminalis L.) trees before flower bud initiation. As expected, 5-azaC decreased the level of DNA methylation in the leaves of both male and female trees during floral development; however, it increased DNA methylation in the leaves of male trees at the flower transition stage. Furthermore, 5-azaC increased the number, length and diameter of flower buds in the female trees but decreased these parameters in the male trees. The 5-azaC treatment also decreased the contents of soluble sugars, starch and reducing sugars in the leaves of the female plants, while increasing them in the male plants at the flower transition stage; however, this situation was largely reversed at the flower development stage. In addition, 5-azaC treatment decreased the contents of auxin indoleacetic acid (IAA) in both male and female trees at the flower transition stage. These results indicate that hypomethylation in leaves at the flower transition stage promotes the initiation of flowering and subsequent floral growth in Salix viminalis, suggesting that DNA methylation plays a similar role in vegetative–reproductive transition and early floral development. Furthermore, methylation changes during the vegetative–reproductive transition and floral development were closely associated with the biosynthesis, metabolism and transportation of carbohydrates and IAA. These results provide insight into the epigenetic regulation of carbohydrate accumulation.

Revealing the roles of y4wF and tidC genes in Rhizobium tropici CIAT 899: biosynthesis of indolic compounds and impact on symbiotic properties.

Rhizobium tropici CIAT 899 is a strain known by its ability to nodulate a broad range of legume species, to synthesize a variety of Nod factors, its tolerance of abiotic stresses, and its high capacity to fix atmospheric N2, especially in symbiosis with common bean (Phaseolus vulgaris L.). Genes putatively related to the synthesis of indole acetic acid (IAA) have been found in the symbiotic plasmid of CIAT 899, in the vicinity of the regulatory nodulation gene nodD5, and, in this study, we obtained mutants for two of these genes, y4wF and tidC (R. tropiciindole-3-pyruvic acid decarboxylase), and investigated their expression in the absence and presence of tryptophan (TRP) and apigenin (API). In general, mutations of both genes increased exopolysaccharide (EPS) synthesis and did not affect swimming or surface motility; mutations also delayed nodule formation, but increased competitiveness. We found that the indole-3-acetamide (IAM) pathway was active in CIAT 899 and not affected by the mutations, and-noteworthy-that API was required to activate the tryptamine (TAM) and the indol-3-pyruvic acid (IPyA) pathways in all strains, particularly in the mutants. High up-regulation of y4wF and tidC genes was observed in both the wild-type and the mutant strains in the presence of API. The results obtained revealed an intriguing relationship between IAA metabolism and nod-gene-inducing activity in R. tropici CIAT 899. We discuss the IAA pathways, and, based on our results, we attribute functions to the y4wF and tidC genes of R. tropici.

New insights into indole-3-acetic acid metabolism in Azospirillum brasilense.

The aim of this research was to analyse the global indole-3-acetic acid (IAA) metabolism in three commercially used strains of Azospirillum brasilense. Azospirillum brasilense Sp245, Az39 and Cd, containing a plasmid with the ipdC-gusA fusion (pFAJ64), were cultured in minimal medium MMAB with or without 10mgl-1 of l-trp till exponential or stationary growth phase. The cultures were then split into 10ml tubes and individually treated with 10mgml-1 IAA, IBA or NAA (auxin catabolism and homeostasis); IAPhe, IALeu, IAA-ala, IAA-glucose (IAA conjugate hydrolysis); or l-lys, l-leu, l-ileu, l-phe, l-ala, l-val, l-arg, l-glu, l-his, l-met, l-asp, l-cys, l-ser, l-pro, l-thr and l-trp (regulation of IAA biosynthesis and IAA conjugation). Bacterial growth, IAA production and ipdC expression were evaluated. None of the A. brasilense strains were able to hydrolyse IAA conjugates, catabolize auxins, or conjugate IAA with amino acids or glucose. l-amino acids l-met, l-val, l-cys and l-ser inhibited bacterial growth and decreased IAA biosynthesis. The expression of ipdC and IAA biosynthesis but not bacterial growth was affected by l-leu, l-phe, l-ala, l-ile, l-pro. l-arg, l-glu, l-his, l-lys, l-asp and l-thr did not affect any of the measured parameters. In this paper, we confirmed that A. brasilense produces IAA only in presence of l-trp is not able to degrade auxins, conjugate IAA with sugars and/or l-amino acids, or hydrolyse such conjugates to release free IAA. Finally, we found that bacterial growth and/or IAA biosynthesis were inhibited by the presence of several l-amino acids probably by diversion of the cellular metabolism. We propose a renewed model to explain IAA metabolism in A. brasilense, one of the most studied phytostimulatory bacteria.