Molecular cloning of the gene for indolepyruvate decarboxylase from Enterobacter cloacae

Summary Although indole-3-acetic acid (IAA) is a well-known plant hormone, the main IAA biosynthetic pathway from l-tryptophan (Trp) via indole-3-pyruvic acid (IPyA) has yet to be elucidated. Previous studies have suggested that IAA is produced by Enterobacter cloacae isolated from the rhizosphere of cucumbers and its biosynthetic pathway may possibly be the same as that in plants. To elucidate this pathway, the IAA biosynthetic gene was isolated from a genomic library of E. cloacae by assaying for the ability to convert Trp to IAA. DNA sequence analysis showed that this gene codes for only one enzyme and its predicted protein sequence has extensive homology with pyruvate decarboxylase in yeast and Zymomonas mobilis. Cell-free extracts prepared from Escherichia coli harboring this gene could convert IPyA to indole-3-acetaldehyde (IAAld). These results clearly show that this pathway is mediated only by indolepyruvate decarboxylase, which catalyzes the conversion of IPyA to IAAld.     

Biosynthesis of indole-3-acetic acid in tomato shoots: Measurement,mass-spectral identification and incorporation of −2H from −2H2O into indole-3-acetic acid,d- and l-tryptophan,indole-3-pyruvate and tryptamine

Indole-3-acetic acid (IAA) and its putative precursors, l- and d-tryptophan, indole-3-pyruvate, and tryptamine were isolated from tomato (Lycopersicon esculentum (L.) Mill.) shoots, identified by mass spectrometry, and measured using capillary gas chromatography with an electron capture detector and radioactive internal standards. Average amounts present were 7.9ng · (g FW)^–-1 IAA, 5.7ng · (g FW)^–-1 indole-3-pyruvate, 132 ng · (g FW)^–-1 tryptamine, 103 ng · (g FW)^–-1 d-tryptophan, and 2250 ng · (g FW)^–-1 l-tryptophan. Indole-3-acetaldoxime was not found; detection limits were less than 1ng · (g FW)^–-1. When tomato shoots were incubated for 6, 10 and 21 h in 30% ^–2H₂O, up to four positions in IAA, l- and d-tryptophan, tryptamine and indole-3-pyruvate became labelled with ^–2H. Compounds became labelled rapidly with 10% of IAA molecules containing ^–2H after 6 h. The percentage of labelled molecules of IAA and l-tryptophan increased up to 10 h but then decreased again, correlating with an increase in the total shoot tryptophan and presumably a result of protein hydrolysis in the excised, slowly senescing tissue. The amount of ^–2H in d-tryptophan also showed an increase followed by a decrease, but the proportion of labelled molecules was much less than in l-tryptophan and IAA. Tryptamine became labelled initially at a similar rate to IAA but continued to accumulate ^–2H up to 21 h. We conclude that tryptamine is synthesized from a different pool of tryptophan from that used in IAA synthesis, and is not a major endogenous precursor of IAA in tomato shoots. Indole-3-pyruvate was the most heavily labelled compound after 6 and 10 h incubation (21-h data not available). Furthermore, the proportion of ^–2H-labelled indole-3-pyruvate molecules was quantitatively consistent with the amount of label in IAA. On the other hand, a quantitative comparison of the IAA turnover rate and the rate of ^–2H incorporation into both l- and d-tryptophan indicates that IAA is not made from the total shoot pool of either l- or d-tryptophan. Instead IAA appears to be synthesized from a restricted pool which is turning over rapidly and which has access to both newly synthesized tryptophan and that from protein hydrolysis.Abbreviations GC-ecd gas chromatography with electroncapture detector - GC-MS combined gas chromatography-mass spectrometry - HPLC high-performance liquid chromatography - IAA indole-3-acetic acid - IAOX indole-3-acetaldoxime - IPyA indole-3-pyruvate - PFB pentafluorobenzyl - R_T retention time - TNH₂ tryptamine - Trp tryptophan - SIM selected ion monitoring We wish to thank Ms. Sue Alford for running the mass spectra and Dr Harry Young for advice with the mass spectrometry. The work was supported by grants from the University of Auckland Research Committee and the C. Alma Baker Trust fund. The mass spectrometer was supported jointly by the University Grants Commitee (NZ) and the DSIR Division of Horticulture and Processing.     

Indolic constituents and indole-3-acetic acid biosynthesis in the wild-type and a tryptophan auxotroph mutant of Arabidopsis thaliana

 The tryptophan auxotroph mutant trp3-1 of Arabidopsis thaliana (L.) Heynh., despite having reduced levels of l-tryptophan, accumulates the tryptophan-derived glucosinolate, glucobrassicin and, thus, does not appear to be tryptophan-limited. However, due to the block in tryptophan synthase, the mutant hyperaccumulates the precursor indole-3-glycerophosphate (up to 10 mg per g FW). Instability of indole-3-glycerophosphate leads to release of indole-3-acetic acid (IAA) from this metabolite during standard workup of samples for determination of conjugated IAA. The apparent increase in “conjugated IAA” in trp3-1 mutant plants can be traced back entirely to indole-3-glycerophosphate degradation. Thus, the levels of neither free IAA nor conjugated IAA increase detectably in the trp3-1 mutant compared to wild-type plants. Precursor-feeding experiments to shoots of sterile-grown wild-type plants using [²H]₅-l-tryptophan have shown incorporation of label from this precursor into indole-3-acetonitrile and indole-3-acetic acid with very little isotope dilution. It is concluded that Arabidopsis thaliana shoots synthesize IAA from l-tryptophan and that the non-tryptophan pathway is probably an artifact. Received: 1 March 2000 / Accepted: 10 April 2000     

Genes encoding nitrilase-like proteins from tobacco.

Nitrilase (nitrile aminohydrolase, EC 3.5.5.1) catalyzes the hydrolysis of indole-3-acetonitrile (IAN) to indole-3-acetic acid (IAA). Arabidopsis thaliana genome has four nitrilase genes (NIT1, NIT2, NIT3 and NIT4). Three (NIT1, NIT2 and NIT3) of the four genes have high similarity. We have cloned two NIT4 homologs (TNIT4A and TNIT4B) from tobacco (Nicotiana tabacum). Genomic Southern hybridization, among other experiments, strongly suggests that tobacco has NIT4 homologs but not NIT1 to NIT3 homologs. Introduction of Arabidopsis NIT2 into tobacco conferred IAN-mediated growth inhibition, probably due to hydrolysis of IAN to IAA, while ectopic expression of TNIT4A had little effect on the sensitivity of transgenic plants to IAN. Nitrilase activity of TNIT4 proteins is discussed.     

Fungal auxin antagonist hypaphorine competitively inhibits indole-3-acetic acid-dependent superoxide generation by horseradish peroxidase.

Plant peroxidases (EC 1.11.1.7) including horseradish peroxidase (HRP-C), but not the nonplant peroxidases, are known to be highly specific indole-3-acetic acid (IAA) oxygenases which oxidize IAA in the absence of H2O2, and superoxide anion radicals (O2*-) are produced as by-products. Hypaphorine, a putative auxin antagonist isolated from ectomycorrhizal fungi, inhibited the IAA-dependent generation of O2*- by HRP-C, which occurs in the absence of H2O2. Hypaphorine has no effect on the nonspecific heme-catalyzed O2*- generation induced by high concentration of ethanol. It is probable that the inhibitory effect of hypaphorine on O2*- generation is highly specific to the IAA-dependent reaction. The mode of inhibition of the IAA-dependent O2*--generating reaction by hypaphorine was analyzed with a double-reciprocal plot and determined to be competitive inhibition, indicating that hypaphorine competes with IAA by binding to the putative IAA binding site on HRP-C. This implies the importance of structural similarity between hypaphorine and IAA. This work presented the first evidence for antagonism between IAA and a structurally related fungal alkaloid on binding to a purified protein which shares some structural similarity with auxin-binding proteins.     

Current aspects of auxin biosynthesis in plants

Auxin is an important plant hormone essential for many aspects of plant growth and development. Indole-3-acetic acid (IAA) is the most studied auxin in plants, and its biosynthesis pathway has been investigated for over 70 years. Although the complete picture of auxin biosynthesis remains to be elucidated, remarkable progress has been made recently in understanding the mechanism of IAA biosynthesis. Genetic and biochemical studies demonstrate that IAA is mainly synthesized from l-tryptophan (Trp) via indole-3-pyruvate by two-step reactions in Arabidopsis. While IAA is also produced from Trp via indole-3-acetaldoxime in Arabidopsis, this pathway likely plays an auxiliary role in plants of the family Brassicaceae. Recent studies suggest that the Trp-independent pathway is not a major route for IAA biosynthesis, but they reveal an important role for a cytosolic indole synthase in this pathway. In this review, I summarize current views and future prospects of IAA biosynthesis research in plants.     

Quantitative Determination of Indole-3-Acetaldehyde. II.

The method by Larsen and Klungsöyr (1964) for the quantitative determination of indole-3-acetaldehyde (IAAld) was modified for the purpose of eliminating the need for filtration after oxidation of the IAAld to indole-3-acetic acid (IAA). The essentials of the modified method are as follows: Samples of IAAld or IAA containing 0.015 to 0.15 μmol (ca. 2.5 to 25 μg) dissolved in peroxide-free ether are evaporated to dryness and redissolved in 1.5 ml 0.02 M Ag2SO4. The oxidation is carried out in dim light by adding 0.5 ml 0.12 N NaOH. After 1.5 min, 2 ml of a modified Salkowski reagent are added. The optical density at 525 nm is read on a spectrophotometer after 75 min. The modified Salkowski reagent consists of 100 ml 0.05 M Fe2(SO4)3 in 1.5 N H2SO4; 240 ml H2O; and 160 ml cone. H2SO4 (sp. gr. 1.84). O.D. readings are identical for equal samples of IAAld and IAA (the latter used as a standard) up to 0.08 μmol (O.D. = 0.32). Larger quantities of IAAld may be determined when using pure IAAld as a standard, but at 0.20 μmol the O.D. for IAAld is lower than for IAA (0.69 as against 0.72). Indole-3-acetonitrile, tryptophol, indole-3-carboxylic acid, and indole-3-aldehyde all give O.D. values lower than 0.1 when tested at 0.20 μmol under the same conditions as described for IAAld and IAA.     

Identification of an aldehyde oxidase involved in indole-3-acetic acid synthesis in Bombyx mori silk gland

Auxin is thought to be an important factor in the induction of galls by galling insects. We have previously shown that both galling and nongalling insects synthesize indole-3-acetic acid (IAA) from tryptophan (Trp) via two intermediates, indole-3-acetaldoxime (IAOx) and indole-3-acetaldehyde (IAAld). In this study, we isolated an enzyme that catalyzes the last step “IAAld → IAA” from a silk-gland extract of Bombyx mori. The enzyme, designated “BmIAO1”, contains two 2Fe–2S iron–sulfur-cluster-binding domains, an FAD-binding domain, and a molybdopterin-binding domain, which are conserved in aldehyde oxidases. BmIAO1 causes the nonenzymatic conversion of Trp to IAAld and the enzymatic conversion of IAOx to IAA, suggesting that BmIAO1 alone is responsible for IAA production in B. mori. However, a detailed comparison of pure BmIAO1 and the crude silk-gland extract suggested the presence of other enzymes involved in IAA production from Trp.

Abbreviations: BA: benzoic acid; CE: collision energy; CXP: collision cell exit potential; DP: declustering potential; IAA: indole-3-acetic acid; IBI1: IAA biosynthetic inhibitor-1; IAAld: indole-3-acetaldehyde; ICA: indole-3-carboxylic acid; IAOx: indole-3-acetaldoxime; IEtOH: indole-3-ethanol; LC–MS/MS: liquid chromatography–tandem mass spectrometry; Trp: tryptophan     

Effect of Lanthanum on Rooting of In Vitro Regenerated Shoots of Saussurea involucrata Kar. et Kir

In present study, the effect of lanthanum (La) on the rooting of regenerated shoots of Saussurea involucrata Kar. et Kir was analyzed. Rooting occurred from regenerated shoots inoculated on a medium supplemented with La, the plant rooting hormone indole-3-acetic acid (IAA), or both La and IAA together. The highest rooting efficiency (96%), root number/shoot (8.5), and root length (63 mm) were recorded in shoots cultured on medium containing 2.5 μM IAA combined with 100 μM La(3+). In order to elucidate the mechanism of rooting enhancement by La, we examined dynamic changes in antioxidant enzyme activities in plant tissue over time in culture. We found that the activities of peroxidase (POX) and superoxide dismutase (SOD) were significantly higher in plant tissue cultured in IAA plus La than in La or IAA alone. At the same time, the highest H(2)O(2) content was detected in plant tissue in the presence of 2.5 μM IAA plus 100 μM La(3+). In light of these data and previous results, we speculate that La enhanced IAA-induced rooting by acting as a mild abiotic stress to stimulate POX and SOD activities in plant cells. Then, IAA reacted with oxygen and POX to form the ternary complex enzyme-IAA-O(2) that dissociated into IAA radicals and O(2)(-). Subsequently, IAA-induced O(2)(-) readily converted to hydroxyl radical (HO·) via SOD-catalyzed dismutation. Finally, cell wall loosening and cell elongation occurred as a consequence of HO-dependent scission of wall components, leading to root growth. The treatment of IAA combined with La resulted in the highest plantlet survival (80%) compared to single treatments with IAA or La alone. These data suggest that rare earth elements enhance root morphogenesis and the growth of S. involucrata.     

Occurrence and formation of indole-3-acetamide in Arabidopsis thaliana

An HPLC/GC-MS/MS technique (high-pressure liquid chromatography in combination with gas chromatography-tandem mass spectrometry) has been worked out to analyze indole-3-acetamide (IAM) with very high sensitivity, using isotopically labelled IAM as an internal standard. Using this technique, the occurrence of IAM in sterile-grown Arabidopsis thaliana (L.) Heynh. was demonstrated unequivocally. In comparison, plants grown under non-sterile conditions in soil in a greenhouse showed approximately 50% higher average levels of IAM, but the differences were not statistically significant. Thus, microbial contributions to the IAM extracted from the tissue are likely to be minor. Levels of IAM in sterile-grown seedlings were highest in imbibed seeds and then sharply declined during the first 24 h of germination and further during early seedling development to remain below 20-30 pmol g(-1) fresh weight throughout the rosette stage. The decline in indole-3-aetic acid (IAA) levels during germination was paralleled by a similar decline in IAM levels. Recombinant nitrilase isoforms 1, 2 and 3, known to synthesize IAA from indole-3-acetonitrile, were shown to produce significant amounts of IAM in vitro as a second end product of the reaction besides IAA. NIT2 was earlier shown to be highly expressed in developing and in mature A. thaliana embryos, and NIT3 is the dominantly active gene in the hypocotyl and the cotyledons of young, germinating seedlings. Collectively, these data suggest that the elevated levels of IAM in seeds and germinating seedlings result from nitrilase action on indole-3-acetonitrile, a metabolite produced in the plants presumably from glucobrassicin turnover.     

Indole-3-glycerol phosphate, a branchpoint of indole-3-acetic acid biosynthesis from the tryptophan biosynthetic pathway in Arabidopsis thaliana

The phytohormone indole-3-acetic acid (IAA) plays a vital role in plant growth and development as a regulator of numerous biological processes. Its biosynthetic pathways have been studied for decades. Recent genetic and in vitro labeling evidence indicates that IAA in Arabidopsis thaliana and other plants is primarily synthesized from a precursor that is an intermediate in the tryptophan (Trp) biosynthetic pathway. To determine which intermediate(s) acts as the possible branchpoint for the Trp-independent IAA biosynthesis in plants, we took an in vivo approach by generating antisense indole-3-glycerol phosphate synthase (IGS) RNA transgenic plants and using available Arabidopsis Trp biosynthetic pathway mutants trp2-1 and trp3-1. Antisense transgenic plants display some auxin deficient-like phenotypes including small rosettes and reduced fertility. Protein gel blot analysis indicated that IGS expression was greatly reduced in the antisense lines. Quantitative analyses of IAA and Trp content in antisense IGS transgenic plants and Trp biosynthetic mutants revealed striking differences. Compared with wild-type plants, the Trp content in all the transgenic and mutant plants decreased significantly. However, total IAA levels were significantly decreased in antisense IGS transgenic plants, but remarkably increased in trp3-1 and trp2-1 plants. These results suggest that indole-3-glycerol phosphate (IGP) in the Arabidopsis Trp biosynthetic pathway serves as a branchpoint compound in the Trp-independent IAA de novo biosynthetic pathway.     

Roles of the reactive oxygen species-generating peroxidase reactions in plant defense and growth induction

Extracellularly secreted plant peroxidases (POXs) are considered to catalyze the generation of reactive oxygen species (ROS) coupled to oxidation of plant hormone indole-3-acetic acid (IAA) and defense-related compounds salicylic acid (SA), aromatic monoamines (AMAs) and chitooligosaccharides (COSs). This review article consists of two parts, which describe H(2)O(2)-dependent and H(2)O(2)-independent mechanisms for ROS generation, respectively. Recent studies have shown that plant POXs oxidize SA, AMAs and COSs in the presence of H(2)O(2) via a conventional POX cycle, yielding the corresponding radical species, such as SA free radicals. These radical species may react with oxygen, and superoxide (O(2)(.-)) is produced. Through the series of reactions 2 moles of O(2)(.-) can be formed from 1 moles of H(2)O(2), thus leading to oxidative burst. It has been revealed that the ROS induced by SA, AMAs and COSs triggers the increase in cytosolic Ca(2+) concentration. Actually POXs transduce the extracellular signals into the redox signals that eventually stimulate the intracellular Ca(2+) signaling required for induction of defense responses. On the other hand, IAA can react with oxygen and plant POXs in the absence of H(2)O(2), by forming the ternary complex enzyme-IAA-O(2), which readily dissociates into enzyme, IAA radicals and O(2)(.-). This article covers the recent reports showing that extracellularly produced hydroxy radicals derived from O(2)(.-) mediate the IAA-induced cell elongation. Here a novel model for IAA signaling pathway mediated by extracellular ROS produced by cell-wall POXs is proposed. In addition, possible controls of the IAA-POX reactions by a fungal alkaloid are discussed.     

Hydrogen peroxide is a mediator of indole-3-acetic acid/horseradish peroxidase-induced apoptosis

Recently, we reported that a combination of indole-3-acetic acid (IAA) and horseradish peroxidase (HRP) induces apoptosis in G361 human melanoma cells. However, the apoptotic mechanism involved has been poorly studied. It is known that when IAA is oxidized by HRP, free radicals are produced, and since oxidative stress can induce apoptosis, we investigated whether reactive oxygen species (ROS) are involved in IAA/HRP-induced apoptosis. Our results show that IAA/HRP-induced free radical production is inhibited by catalase, but not by superoxide dismutase or sodium formate. Furthermore, catalase was found to prevent IAA/HRP-induced apoptotic cell death, indicating that IAA/HRP-produced hydrogen peroxide (H2O2) may be involved in the apoptotic process. Moreover, the antiapoptotic effect of catalase is potentiated by NADPH, which is known to protect catalase. On further investigating the IAA/HRP-mediated apoptotic pathway, we found that the IAA/HRP reaction leads to caspase-3 activation and poly(ADP-ribose) polymerase (PARP) cleavage, which was also blocked by catalase. Additionally, we found that IAA/HRP produces H2O2 and induces peroxiredoxin (Prx) sulfonylation. Consequently, our results suggest that H2O2 plays a major role in IAA/HRP-induced apoptosis.     

Identical amino acid substitutions in the repression domain of auxin/indole-3-acetic acid proteins have contrasting effects on auxin signaling

Auxin/indole-3-acetic acid (Aux/IAA) proteins function as repressors of auxin response gene expression when auxin concentrations in a cell are low. At elevated auxin concentrations, these repressors are destroyed via the ubiquitin-proteasome pathway, resulting in derepression/activation of auxin response genes. Most Aux/IAA repressors contain four conserved domains, with one of these being an active, portable repression domain (domain I) and a second being an auxin-dependent instability domain (domain II). Here, we have analyzed the effects of amino acid substitutions in the repression domain of selected Aux/IAA proteins. We show that stabilized versions of Aux/IAA proteins with amino acid substitutions in domain I display contrasting phenotypes when expressed in transformed Arabidopsis (Arabidopsis thaliana) plants. An alanine-for-leucine substitution in the LxLxL (where L is leucine and x is another amino acid) repression domain of IAA3, IAA6, or IAA19 confers enhanced auxin response gene expression and "high-auxin" phenotypes when expressed from the 35S or IAA19 promoter (as tested with IAA19) in transformed Arabidopsis plants. In marked contrast, a single alanine-for-leucine substitution in domain I of IAA12 or IAA17 confers repression of auxin response genes and "low-auxin" phenotypes. These results point to intrinsic differences in the repression domain(s) of IAA proteins and suggest that some IAA proteins have stronger or more complex repression domains than others.     

Biosynthesis of indole-3-acetic acid in Azospirillum brasilense. Insights from quantum chemistry.

Quantum chemical methods AM1 and PM3 and chromatographic methods were used to qualitatively characterize pathways of bacterial production of indole-3-acetic acid (IAA). The standard free energy changes (delta G(o)'sum) for the synthesis of tryptophan (Trp) from chorismic acid via anthranilic acid and indole were calculated, as were those for several possible pathways for the synthesis of IAA from Trp, namely via indole-3-acetamide (IAM), indole-3-pyruvic acid (IPyA), and indole-3-acetonitrile (IAN). The delta G(o)'sum for Trp synthesis from chorismic acid was -402 (-434) kJ.mol-1 (values in parentheses were calculated by PM3). The delta G(o)'sum for IAA synthesis from Trp were -565 (-548) kJ.mol-1 for the IAN pathway, -481 (-506) kJ.mol-1 for the IAM pathway, and -289 (-306) kJ.mol-1 for the IPyA pathway. By HPLC analysis, the possibility was assessed that indole, anthranilic acid, and Trp might be utilized as precursors for IAA synthesis by Azospirillum brasilense strain Sp 245. The results indicate that there is a high motive force for Trp synthesis from chorismic acid and for IAA synthesis from Trp, and make it unlikely that anthranilic acid and indole act as the precursors to IAA in a Trp-independent pathway.     

Isolation and characterization of mutants of Arabidopsis thaliana with increased resistance to growth inhibition by indoleacetic acid-amino acid conjugates.

Two mutants of Arabidopsis thaliana that are resistant to growth inhibition by indole-3-acetic acid (IAA)-phenylalanine have been isolated. Both mutants were 2- to 3-fold more resistant than wild type to inhibition by IAA-phenylalanine, IAA-alanine, and IAA-glycine in root growth assays. The mutant icr1 (but not icr2) also shows some resistance to IAA-aspartate. Studies using 3H-labeled IAA-phenylalanine showed that the uptake of conjugate from the medium by icr1 was the same as wild type and was reduced by about 25% in icr2. No differences in hydrolysis of the exogenous conjugate were detected between the mutants and their wild-type parents. There was no significant metabolism of the IAA released from the [3H]IAA-phenylalanine, whereas exogenous [3H]IAA was rapidly metabolized to two unidentified products considerably more polar than IAA. Analysis of a cross between icr1 and icr2 indicated that these mutations were at distinct loci and that their effects were additive, and preliminary mapping data indicated that icr1 and icr2 were located at the top and bottom of chromosome V, respectively.     

Biosynthesis and metabolism of indole-3-ethanol and indole-3-acetic acid by Pinus sylvestris L. needles

Combined gas chromatography-mass spectrometry has been used to identify indole-3-ethanol (IEt) in a purified extract from needles of Pinus sylvestris L. Quantitative estimates obtained by high-performance liquid chromatography with fluorescence detection, corrected for samples losses occurring during purification, indicate that Pinus needles contain 46±4 ng g^-1 IEt. This compares with 24.5±6.5 ng g^-1 indole-3-acetic acid (IAA) and 2.3±0.4 ng g^-1 indole-3-carboxylic acid (ICA) (Sandberg et al. 1984, Phytochemistry, 23, 99–102). Metabolism studies with needles incubated in a culture medium in darkness revealed that both [3-¹⁴C]-tryptophan and [2-¹⁴C]tryptamine mine are converted to [¹⁴C]IEt. It was also shown that [3-¹⁴C]IEt acted as a precursor of [¹⁴C]IAA. The observed metabolism appears to be enzymic in nature. The [2-¹⁴C]IAA was not catabolised to [¹⁴C]ICA in detectable quantities implying that, at best, only a minor portion of the endogenous ICA pool in the Pinus needles originates from IAA.Abbreviations DEAE diethylaminoethyl - GC-MS gas chromatography-mass spectrometry - HPLC high-performance liquid chromatography - IAA indole-3-acetic acid - ICA indole-3-carboxylic acid - IEt indole-3-ethanol - PVP polyvinylpyrrolidone     

Deuterium-labelled indole-3-acetic acid neo-synthesis in plantlets and excised roots of maize

Synthesis of indole-3-acetic acid (IAA), using stable-isotope incorporation, was investigated in Zea mays L. Incorporation of ²H from ²H₂O into IAA molecules was shown to occur in intact plantlets and excised primary roots cultured in vitro. This demonstrates the de-novo formation of IAA, a process which is quantitatively well defined and is initiated early in germination.Abbreviations IAA indole-3-acetic acid