Immunocytochemical localization of indole-3-acetic acid in primary roots of Zea mays

Colloidal gold-labelled antibody was used to localize indole-3-acetic acid in caps of primary roots of Z. mays cv. Kys. Gold particles accumulated on the nucleus, vacuoles, mitochondria, and some dictyosomes and dictyosome-derived vesicles. This is the first localization of indole-3-acetic acid in dictyosomes and dictyosome-derived vesicles and indicates that dictyosomes and vesicles constitute a pathway for indole-3-acetic acid movement in and secretion from root cap cells. Our findings provide cytochemical evidence to support the hypothesis that indole-3-acetic acid plays an important role in root gravitropism.     

Regulation of enzymic oxidation of indole-3-acetic acid by phenols: Structure-activity relationships

Mono- and diphenols were tested for their effects on the decarboxylation of [1-¹⁴C]IAA catalysed by purified horseradish peroxidase (EC 1.11.1.7) in the presence or absence of 2,4-dichlorophenol (DCP). The number of hydroxyl groups and their position relative to each other and the nature and position of other substituents on the aromatic ring were found to affect the activity. Although the effects were complex, the following generalizations may be made. (1) Monophenols produce activation when no other cofactor is present. p-Substituted monophenols are more active than o- or m-compounds. In the presence of DCP, the activity varies from slight activation to strong inhibition. (2) m-Diphenols also produce activation in the absence of other cofactors while o- and p-diphenols, with the exception of 3,4-dihydroxyacetophenone and 3,4-dihydroxypropiophenone, produce strong inhibition in the presence or absence of DCP. The o-diphenolsare degraded in the IAA-oxidizing enzyme system and thus produce only a temporary inhibition. (3) m-Diphenols and 3,4-dihydroxyacetophenone produce a sustained inhibition in the presence of DCP. (4) Substitution at position 2 significantly alters the activity of m-diphenols. (5) O-Methylation alters the activity of most o-diphenols. In the absence of DCP, o-methoxyphenols and certain other phenols such as 3,4-dihydroxyacetophenone and 2,6-dihydroxyacetophenone either promote or inhibit IAA oxidation depending on concentration.     

New phenolic inhibitors of the peroxidse-catalysed oxidation of indole-3-acetic acid

Previously we reported two metabolites of the insecticide carbofuran as persistent inhibitors of the peroxidase-catalysed oxidtion ofindole-3-acetic acid. In searching for more active inhibitors of this type, we have found that 5-hydroxy-2,2-dimethylchromene (β-tubanol), 2′,6′-dihydroxycetophenone oxime, 5-hydroxy-2,2-dimethylchroman, 2′,6′-dihydroxyacetophenone and 2,6-dihydroxybenzoic acid methyl ester were more active than the carbofuran metabolite 7-hydroxy-2,2-dimethyl-3-oxo-2,3-dihydrobenzofuran. Resorcinol, 5-hydroxy-2,2-dimethylchroman-4-one, 3-hydroxy-5-methoxy-2,2-dimethylchroman-4-one and 5-hydroxy-2-methylchrom-4-one were also inhibitory but with less activity. The new inhibitors differed from the well-known phenolic inhibitors such as caffeic acid in inhibition kinetics as demonstrated by the rate of disappearance of indole-3-acetic acid, the rate of formation of the oxidation products, and the transient spectral change in the enzyme.     

Effects of indole-3-acetic acid on arsenic uptake and antioxidative enzymes in Pteris cretica var. nervosa and Pteris ensiformis

A hydroponic experiment was conducted to investigate the effects of indole-3-acetic acid (IAA) on arsenic (As) uptake and antioxidative enzymes in fronds of Pteris cretica var. nervosa (As hyperaccumulator) and Pteris ensiformis (non-hyperaccumulator). Plants were exposed to 2 mg L^?1 As(III), As(V) or dimethylarsinic acid (DMA) and IAA concentrations for 14 d. The biomass and total As in the plants significantly increased at 30 mg L^?1 IAA. Superoxide dismutase (SOD) activities significantly increased with IAA addition. Catalase (CAT) activities showed a significant increase in P. ensiformis exposed to three As species at 30 or 50 mg L^?1 IAA but varied in P. cretica var. nervosa. Peroxidase (POD) activities were unchanged in P. ensiformis except for a significant decrease at 50 mg L^?1 IAA under As(III) treatment. However, a significant increase was observed in P. cretica var. nervosa at 10 mg L^?1 IAA under As(III) or DMA treatment and at 50 mg L^?1 IAA under As(V) treatment. Under DMA stress, malondialdehyde contents in fronds of P. cretica var. nervosa showed a significant decrease at 10 mg L^?1 IAA but remained unchanged in P. ensiformis. Therefore, IAA enhanced As uptake and frond POD activity in P. cretica var. nervosa under As stress.     

Double agent indole-3-acetic acid: mechanistic analysis of indole-3-acetaldehyde dehydrogenase AldA that synthesizes IAA,an auxin that aids bacterial virulence

The large diversity of organisms inhabiting various environmental niches on our planet are engaged in a lively exchange of biomolecules, including nutrients, hormones, and vitamins. In a quest to survive, organisms that we define as pathogens employ innovative methods to extract valuable resources from their host leading to an infection. One such instance is where plant-associated bacterial pathogens synthesize and deploy hormones or their molecular mimics to manipulate the physiology of the host plant. This commentary describes one such specific example—the mechanism of the enzyme AldA, an aldehyde dehydrogenase (ALDH) from the bacterial plant pathogen Pseudomonas syringae which produces the plant auxin hormone indole-3-acetic acid (IAA) by oxidizing the substrate indole-3-acetaldehyde (IAAld) using the cofactor nicotinamide adenine dinucleotide (NAD⁺) (Bioscience Reports (2020) 40(12), https://doi.org/10.1042/BSR20202959). Using mutagenesis, enzyme kinetics, and structural analysis, Zhang et al. established that the progress of the reaction hinges on the formation of two distinct conformations of NAD(H) during the reaction course. Additionally, a key mutation in the AldA active site ‘aromatic box’ changes the enzyme’s preference for an aromatic substrate to an aliphatic one. Our commentary concludes that such molecular level investigations help to establish the nature of the dynamics of NAD(H) in ALDH-catalyzed reactions, and further show that the key active site residues control substrate specificity. We also contemplate that insights from the present study can be used to engineer novel ALDH enzymes for environmental, health, and industrial applications.     

IAA-induced adventitious root formation in greenwood cuttings of Populus tremula and formation of 2-indolone-3-acetylaspartic acid, a new metabolite of exogeneously applied indole-3-acetic acid

When indole-3-acetic acid (IAA) is applied through the basal cut surface of greenwood cuttings from Populus tremula L. with the aim to induce adventitious roots, it is observed that a positive correlation between the number of new roots and the duration of the application exists only for the first 5 to 6 hours. This is most likely due to the induction, during this time, of a metabolic system that transforms IAA to compounds unable to provoke new roots. The most important of these compounds was identified as 2-indolone-3-acetylaspartic acid (OxlAasp). The metabolic pathway from IAA to OxIAasp via indole-3-acetylaspartic acid was demonstrated by thin layer chromatography.     

打顶后施用生长素(IAA)和钾肥对烤烟碳氮代谢的影响

研究了不打顶(T1)、打顶(T2)、打顶 追施K_2SO_4(T_3)、打顶 涂抹生长素1次(T4)、打顶 追施K2SO4 涂抹生长素1次(T5)、打顶 追施K2SO4 涂抹生长素2次(T6)等6种调控措施对烤烟碳氮代谢的影响。结果表明:由打顶当天到打顶后30d,烤烟淀粉酶活性总体上表现为下降后又略有上升的趋势,转化酶(INV)活性则逐渐降低;2种酶活性均以T6最高,T1最低,且差异达到极显著水平;除T1外,各处理在打顶后的淀粉含量逐渐升高,总糖含量则呈现上升后又逐步下降的变化,至打顶后30d,淀粉和总糖含量均以T6最高,T1最低;随生育时期延长,硝酸还原酶(NR)活性和蛋白质含量均表现为逐渐下降的趋势。打顶后30d,T6的NR活性和蛋白质含量均为最高;在打顶后30d,以T5的NR/INV比值最大。打顶当天在顶端涂抹生长素,同时追施K2SO4肥,可促进烤烟生长和碳氮代谢。     

Endogenous levels of abscisic acid and indole-3-acetic acid during in vitro rooting of Wild Cherry explants produced by micropropagation

Levels of endogenous ABA and IAA were quantified during the first week of in vitro rooting of Wild Cherry (Prunus avium L.) using IBA in the culture medium. Hormones were measured in the apical, median and basal parts of the explants using an avidin-biotin based enzyme linked immunosorbent assay (ELISA), after a purification of the methanolic extracts by high-performance liquid chromatography (HPLC).Root primordia started to differentiate from day 5 at the basal part of the explants. ABA and IAA showed considerable changes and high levels were detected during the first week of culture. ABA levels increased transiently mainly in the apical part during root formation. Exogenous IBA was possibly transformed into IAA mainly in the basal part of the explants.     

Effect of 3,4-dihydroxyacetophenone and some related phenols on the peroxidase-catalysed oxidation of indole-3-acetic acid

3,4-Dihydroxyacetophenone, 3,4-dihydroxypropiophenone and acetovanillone differed in activity in the enzymic oxidative reaction of indole-3-acetic acid     

Circadian changes in endogenous concentrations of indole-3-acetic acid,melatonin, serotonin,abscisic acid and jasmonic acid in Characeae (Chara australis Brown)

Giant-celled Characeae (Chara australis Brown), grown for 4 months on 12/12 hr day/night cycle and summer/autumn temperatures, exhibited distinct concentration maxima in auxin (indole-3-acetic acid; IAA), melatonin and serotonin about 4 hr after subjective daybreak. These concentration peaks persisted after 3 day pretreatment in continuous darkness: confirming a circadian rhythm, rather than a response to “light on.” The plants pretreated for 3 d in continuous light exhibited several large IAA concentration maxima throughout the 24 hr. The melatonin and serotonin concentrations decreased and were less synchronized with IAA. Chara plants grown on 9/15 hr day/night cycle for 4 months and winter/spring temperatures contained much smaller concentrations of IAA, melatonin and serotonin. The IAA concentration maxima were observed in subjective dark phase. Serotonin concentration peaks were weakly correlated with those of IAA. Melatonin concentration was low and mostly independent of circadian cycle. The “dark” IAA concentration peaks persisted in plants treated for 3 d in the dark. The plants pretreated for 3 d in the light again developed more IAA concentration peaks. In this case the concentration maxima in melatonin and serotonin became more synchronous with those in IAA. The abscisic acid (ABA) and jasmonic acid (JA) concentrations were also measured in plants on winter regime. The ABA concentration did not exhibit circadian pattern, while JA concentration peaks were out of phase with those of IAA. The data are discussed in terms of crosstalk between metabolic pathways.     

The relative importance of tryptophan-dependent and tryptophan-independent biosynthesis of indole-3-acetic acid in tobacco during vegetative growth

A quantitative study of indole-3-acetic acid (IAA) turnover, and the contribution of tryptophan-dependent and tryptophan-independent IAA-biosynthesis pathways, was carried out using protoplast preparations and shoot apices obtained from wild-type and transgenic, IAA-overproducing tobacco (Nicotiana tabacum L.) plants, during a phase of growth when the level of endogenous IAA was stable. Based on the rate of disappearance of [¹³C₆]IAA, the half-life of the IAA pool was calculated to be 1.1 h in wild-type protoplasts and 0.8 h in protoplasts from the IAA-overproducing line, corresponding to metabolic rates of 59 and 160 pg IAA (μg Chl)⁻¹ h⁻¹, respectively. The rate of conversion of tryptophan to IAA was 15 pg IAA (μg Chl)⁻¹ h⁻¹ in wild-type protoplasts and 101 pg IAA (μg Chl)⁻¹ h⁻¹ in protoplasts from IAA-overproducing plants. In both instances, IAA was metabolised more rapidly than it was synthesised from tryptophan. As the endogenous IAA pools were in a steady state, these findings indicate that IAA biosynthesis via the tryptophan-independent pathway was 44 pg IAA (μg Chl)⁻¹ h⁻¹ and 59 pg IAA (μg Chl)⁻¹ h⁻¹, respectively, in the wild-type and transformed protoplast preparations. In a parallel study with apical shoot tissue, the presumed site of IAA biosynthesis, the rate of tryptophan-dependent IAA biosynthesis exceeded the rate of metabolism of [¹³C₆]IAA despite the steady state of the endogenous IAA pool. The most likely explanation for this anomaly is that, unlike the protoplast system, injection of substrates into the apical tissues did not result in uniform distribution of label, and that at least some of the [²H₅]tryptophan was metabolised in compartments not normally active in IAA biosynthesis. This demonstrates the importance of using experimental systems where labelling of the precursor pool can be strictly controlled. Received: 18 January 2000 / Accepted 24 February 2000     

热胁迫对不同发育阶段大豆生长素和脱落酸含量的影响(英文)

大豆等植物体内细胞受热或受其它理化因素（如：重金属离子、乙醇、氨基酸类似物）、以及缺氧、DNA损伤、病毒感染等病理因素刺激后,促发应激反应,启动某些基因表达,能产生各种生理活性物质以及各种酶类,共同调控代谢过程和某些激素的活动,如：吲哚乙酸（IAA）、脱落酸（ABA）等。这些内源IAA和ABA共同作用,调节着大豆的抗逆性,从而影响着大豆的农艺性状。本试验对华北生态型的六个大豆栽培种,进行热激处理;取其第三片展开叶,测其内源IAA和ABA含量。这些品种分别是：早熟17,诱处4号,诱变31,耐阴黑豆、科丰6号和科丰34（Tan．1）。初花期,第一天热激（43～45℃,4h）后,它们的IAA和ABA水平均显著高于对照（30～33℃）（Fig．1）。然而,在连续一天热激后（43～45℃,4h／d）,大多数品种的IAA和ABA比第一天减少（Fig．2）。盛花期连续热激处理二天（43～45℃,4h／d）,IAA水平一般低于对照（3～33℃）,半数品种ABA水平也低于对照（Fig．3）。结荚期连续两天热激后（45℃,4h／d）,IAA和ABA含量均显著高于对照（30～33℃）（Fig．4）。     

Effects of water stress on cellular ultrastructure and on concentrations of endogenous abscisic acid and indole-3-acetic acid in Fatsia japonica leaves

Ultrastructural alterations in mesophyll cells as well as variations in bulk leaf endogenous ABA and IAA concentrations were studied in water-stressed field-grown plants of Fatsia japonica. Under water deficit cellular membranes were modified and an increase in vesicles was observed. The main damage to the chloroplasts included thylakoid swelling and disruption of the chloroplast envelope. Concomitant variations in abscisic acid and indole-3-acetic acid were observed. Despite the expected increased in endogenous ABA concentration in relation to water stress, after the highest concentration of ABA, observed at predawn in severely stressed plants (29-1), there was a sharp decline from 2768 pmol g fw^–1 to 145 pmol g fw^–1; thus in severely stressed plants ABA levels were not related to changes in bulk leaf ABA contents. Water stress did not influence the concentrations of indole-3-acetic acid, although the increase in the endogenous abscisic acid concentration could be related with the ultrastructural changes.Abbreviations ABA abscisic acid - IAA indole-3-acetic acid - leaf water potential     

The vacuolar localization of basic isoperoxidases in grapevine suspension cell cultures and its significance in indole-3-acetic acid catabolism

The study of the subcellular localization of the basic isoperoxidases in grapevines was carried out by using cells cultured in suspension as a model system. Results from subcellular fractionation, isoenzyme analysis, enzyme binding and cytochemical probes suggest that basic isoperoxidases are localized mainly in the vacuolar sap of the suspension cultured cells, probably in equilibrium with a pool of the same basic isoperoxidases bound to the internal face of tonoplast membranes through a Ca²⁺-saline bridge. This vacuolar location of basic isoperoxidases raised the question of their function, since indole-3-acetic acid (IAA) oxidase activity of these isoperoxidases is almost totally inhibited by vacuolar anthocyanins in the in vivo concentration range of these compounds. Thus, a central role is proposed for these isoenzymes in the H₂O₂-dependent oxidative phenol metabolism which occurs in grapevines, discarding therefore a possible role of these isoperoxidases in the control of IAA levels during the later stages of development of anthocyanin-rich grapes.     

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.     

Endogenous indoles and the biosynthesis and metabolism of indole-3-acetic acid in cultures of Rhizobium phaseoli

Gas chromatography-mass spectrometric analyses of purified extracts from cultures of Rhizobium phaseoli wild-type strain 8002, grown in a non-tryptophan-supplemented liquid medium, demonstrated the presence of indole-3-acetic acid (IAA), indole-3-ethanol (IEt), indole-3-aldehyde and indole-3-methanol (IM). In metabolism studies with ³H-, ¹⁴C- and ²H-labelled substrates the bacterium was shown to convert tryptophan to IEt, IAA and IM; IEt to IAA and IM; and IAA to IM. Indole-3-acetamide (IAAm) could not be detected as either an endogenous constituent or a metabolite of [³H]tryptophan nor did cultures convert [¹⁴C]IAAm to IAA. Biosynthesis of IAA in R. phaseoli, thus, involves a different pathway from that operating in Pseudomonas savastanio and Agrobacterium tumefaciens-induced crown-gall tumours.Abbreviations IAA indole-3-acetic acid - IAld indole-3-aldehyde - IAAm indole-3-acetamide - IEt indole-3-ethanol - IM indole-3-methanol - HPLC-RC high-performance liquid chromatography-radio counting - GC-MS gas chromatography-mass spectrometry     

Effect of triacontanol on the lipid composition of cotton (Gossypium hirsutum L.) leaves and its interaction with indole-3-acetic acid and benzyladenine

Application of triacontanol (TRIA), a long chain aliphatic alcohol (C-30), to cotton (Gossypium hirsutum L.) leaves resulted in an increase in dry weight and an alteration in lipid composition. A significant increase in monogalactosyldiacylglycerol (MGDG) and digalactosyldiacylglycerol (DGDG) was attained 24 h after TRIA treatment. However, no significant change in any of the individual phospholipids was observed. Benzyladenine (BA) treatment increased only phosphatidylcholine (PC) levels without having any effect on either glycolipids or other phospholipids. Indole-3-acetic acid (IAA) initiated no significant change in the lipid composition. Combined treatment with TRIA and BA resulted in an increase of MGDG, DGDG and PC, indicating that the individual effects of these two growth regulators were not altered.The combined treatment of IAA and TRIA did not bring about any change in the levels of MGDG and DGDG indicating that the effect of TRIA was nullified by IAA. MDGD is known to be involved in the packaging of photosystem I proteins. Whether TRIA-induced increase in dry weight which is due to the enhanced photosynthetic rate, is related to increased MGDG levels is not yet discernible.Abbreviations BA benzyladenine - DGDG digalactosyldiacylglycerol - IAA indole-3-acetic acid - MGDG monogalactosyldiacylglycerol - PC phosphatidylcholine - PE phosphatidylethanolamine - PG phosphatidylglycerol - PI phosphatidylinositol - PS phosphatidylserine - SQDG sulfoquinovosyldiacylglycerol - TRIA triacontanol     

Endogeneous levels of indole-3-acetic acid and abscisic acid during the rooting of Cotinus coggygria cuttings taken at different times of the year

Cuttings of Cotinus coggyria cv Royal Purple rooted well in the spring but not at all later in the season. Levels of free and conjugated IAA and ABA were measured in cuttings taken at different times of the year. Hormones were measured in the leaf, the upper stem and the lower stem (rooting zone). In cuttings taken in early June the level of IAA was much higher than that of conjugated IAA. In late July the opposite was found. No significant differences in ABA levels were found although the ABA/IAA ratio changed dramatically.     

Temperature effects on endogenous indole-3-acetic acid levels in leaves and stamens of the normal and male sterile 'stamenless-2' mutant of tomato (Lycopersicon esculentum Mill.)

The indole-3-acetic acid (IAA) concentration in leaves and stamens of the normal and a temperature-sensitive male sterile ‘stamenless-2′ (sl-2/sl-2) mutant of tomato (Lycopersicon esculentum Mill.), grown under three temperature conditions, was measured by gas chromatography — mass spectrometry — selected ion monitoring (GC-MS-SIM) and by enzyme-linked immunosorbant assay (ELISA). At low (LTR, 18°C day/15°C night), intermediate (ITR, 23°C day/18°C night), and high temperatures (HTR, 28°C day/23°C night), the mutant leaves had approximately 10 to 20 times higher IAA concentrations, respectively, than the normal leaves under these temperature regimes. Similarly, the stamens of mutant flowers had approximately five and eight times higher IAA concentration at ITR and HTR, respectively, than the normal flowers. In the low temperature reverted mutant stamens, however, the level of IAA was similar to that in normal stamens. Also, with an increase in temperature, there was an increase in the level of IAA in the leaves and stamens of mutant plants. However, different temperatures had no appreciable effect on the IAA content of leaves and stamens of normal plants. It is suggested that the high IAA content in leaves and stamens of the stamenless-2 mutant is one of the factors associated with male sterility and carpellization of stamens in this mutant.     

Metabolism of [14C]indole-3-acetic acid by the cortical and stelar tissues of Zea mays L. roots

Reverse-phase high-performance liquid chromatography was used to analyse ¹⁴C-labelled metabolites of indole-3-acetic acid (IAA) formed in the cortical and stelar tissues of Zea mays roots. After a 2-h incubation in [¹⁴C]IAA, stelar segments had metabolised between 1–6% of the methanol-extractable radioactivity compared with 91–92% by the cortical segments. The pattern of metabolites produced by cortical segments was similar to that produced by intact segments bathed in aqueous solutions of [¹⁴C]IAA. In contrast, when IAA was supplied in agar blocks to stelar tissue protruding from the basal ends of segments, negligible metabolism was evident. On the basis of its retention characteristics both before and after methylation, the major metabolite of [¹⁴C]IAA in Zea mays root segments was tentatively identified by high-performance liquid chromatography as oxindole-3-acetic acid.Abbreviations HPLC High-performance liquid chromatography - IAA Indole-3-acetic acid