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In situ characterization of mercury-resistant growth-promoting fluorescent pseudomonads.

Pseudomonas fluorescens strains PRS9 and GRS1 (wild type) were made mercury resistant PRS9Hg(r) (147 microM HgCl2) and GRS1Hg(r) (55 microM HgCl2), respectively, in King's medium by enrichment selection and their in situ root colonization studies were carried out. Mercury resistant mutant of PRS9 was stable and resulted in significant increase in root and shoot fresh weight (P < 0.05). Both the mutants are positive for indoleacetic acid (IAA), 'P' solubilization and siderophore production. PRS9, potent 'P' solubilizer, exhibited higher 'P' solubilization as compared to GRS1. After 2 weeks of inoculation, the population level of wild type PRS9 and its mercury resistant mutants has increased (50 fold). Mercury resistance has no adverse effect on the growth promoting properties of mutants besides being comparable in its morphological and physiological properties with their wild type counterpart. Furthermore, mercury resistant character facilitates rhizospheric competition and thus helpful for establishment of growth promoting strains where metal ions are either limiting and/or present at toxic level.

Biomass↗

INDOLE COMPOUNDS: ISOLATION FROM PINEAL TISSUE.

Five indole compounds have been isolated from bovine pineal tissue and characterized as 5-methoxytryptophol, N-acetyl-5-methoxytryptamine (melatonin), 5-hydroxytryptophol, 5-methoxyindole-3-acetic acid, and 5-hydroxyindole-3-acetic acid. Pineal hydroxyindole-O-methyltransferase, with S-adenosylmethionine, converts 5-hydroxytryptophol to 5-methoxytryptophol.

Animals↗

OsAGAP, an ARF-GAP from rice, regulates root development mediated by auxin in Arabidopsis.

Arf (ADP-ribosylation factor) proteins, which mediate vesicular transport, have little or no intrinsic GTPase activity. They rely on the action of GTPase-activating proteins (GAPs) and guanine nucleotide exchange factors (GEFs) for their function. In the present study the OsAGAP gene in rice, which encoded a protein with predicted structure similar to ArfGAP, was identified. The purified OsAGAP-GST fusion protein was able to stimulate the GTPase activity of rice Arf. Furthermore, OsAGAP can rescue the defect of vesicular transport in the yeast gcs1 delta glo3 delta double-mutant cells. Transgenic Arabidopsis with OsAGAP constitutively expression showed reduced apical dominance, shorter primary roots, increasing number of longer adventitious roots. Many of the phenotypes can be phenocopied by treatment of exogenous indoleacetic acid level (IAA) in wild-type plants. Determination of whole-plant IAA level showed that there is a sharp increase of free IAA in OsAGAP transgenic Arabidopsis seedlings. In addition, removal of the 4-day-old shoot apex could inhibit the adventitious root formation in the transgenic seedlings. These results suggest OsAGAP, an ARF-GAP of rice, maybe involved in the mediation of plant root development by regulating auxin level.

ADP-Ribosylation Factors↗

CNS tryptamine metabolism in hepatic coma.

Lumbar CSF indoleacetic acid (IAA) was higher in patients with cirrhosis of the liver than in controls. It was also higher in CSF of patients in coma than in those with hepatic cirrhosis but not in coma. There was a strong correlation (r = 0.89, p less than 0.01) between the grade of hepatic coma and CSF IAA. These data indicate that there is an association between elevated CNS tryptamine metabolism and hepatic coma. How far changes in the metabolism of tryptamine and other trace amines are relevant to the induction of hepatic coma or are simply a reflection of advanced liver dysfunction is unclear.

Adult↗

Some tryptophan pathways in the phytopathogen Xanthomonas oryzae pv. oryzae.

Xanthomonas oryzae pv. oryzae, the causal organism of bacterial blight of rice which produces leaf blight as well as kresek (wilt) symptoms in plants were tested for indole, auxin production in culture supplemented with L-tryptophan. On the basis of indoleacetic acid (IAA) production the isolates were grouped into IAA-positive and IAA-negative. Out of 17 isolates, 11 were IAA-positive while 6 were IAA-negative. The isolates metabolized tryptophan through two different routes and the isolates vary in the pathway of tryptophan utilization. The IAA-positive isolates converted tryptophan to IAA as the end product, whereas the IAA-negative isolates formed anthranilate as an intermediate metabolite and finally produced pyrocatechol via the kynurenine pathway. Quantification of tryptophan metabolism revealed that the maximum production of IAA and pyrocatechol in culture occurred during 2-d incubation at 30 +/- 2 degrees C.

Catechols↗

Physical organization of phytobeneficial genes nifH and ipdC in the plant growth-promoting rhizobacterium Azospirillum lipoferum 4VI.

The physical organization of phytobeneficial genes was investigated in the plant growth-promoting rhizobacterium Azospirillum lipoferum 4VI by hybridization screening of a bacterial artificial chromosome (BAC) library. Pulsed-field gel electrophoresis gave an estimated 5.7-Mb genome size for strain 4VI and a coverage level of 9 for the BAC library. The phytobeneficial genes nifH (associative nitrogen fixation) and ipdC (synthesis of the phytohormone indoleacetic acid) are chromosomal, but no BAC clone containing both genes was found, pointing to the absence of any genetic island containing nifH and ipdC. A 11.8-kb fragment containing nifH was analyzed. Neighboring genes implicated in nitrogen fixation (nifH, draT, draG) or not (arsC, yafJ and acpD) were organized as in A. brasilense. In contrast, the region located downstream of acpD contained four housekeeping genes (i.e. genes encoding DapF-, MiaB- and FtsY-like proteins, as well as gene amn) and differed totally from the one found in A. brasilense.

Azospirillum lipoferum↗

Studies on the biochemical aspects of the 'disulfiram-like' reaction induced by oral hypoglycemics.

In vitro experiments, using rat liver homogenates, were designed to examine certain of the proposed enzymatic mechanisms for the interaction of oral hypoglycemic drugs with monoamine and ethanol metabolism. The oxidative degradation of tryptamine was studied by measuring indoleacetic acid (IAA) production and conclusions were drawn with regard to the activity of monoamine oxidase, aldehyde dehydrogenase and ethanol dehydrogenase. Acetohexamide, hydroxyhexamide, tolazamide, tolbutamide and chlorpropamide failed to reveal any specific inhibition of the three enzymes. Ethanol (0.2% w/v) and disulfiram decreased IAA formation, as did a lack of available aldehyde dehydrogenase and NAD, but these reductions were not enhanced by the hypoglycemic agents. The results suggest that the 'disulfiram-like' reaction which occurs in certain patients imbibing ethanol while receiving oral hypoglycemic drugs, depends upon some factor(s) other than, or additional to, a specific interference with monoamine and/or ethanol metabolism.

Administration, Oral↗

Partial purification of an enzyme hydrolyzing indole-3-acetamide from rice cells.

The activity of indole-3-acetamide (IAM) hydrolase from rice cells was enriched ca. 628-fold by gel filtration and anion exchange column chromatography. The molecular masses of the IAM hydrolase estimated by gel filtration and sodium dodecyl sulfate polyacrylamide gel electrophoresis were approximately 50.5 kD and 50.0 kD, respectively. The enzyme exhibited maximum activity at pH 6.0-6.5. The enzyme was stable against heat treatments between 4 and 50 degrees C and works optimally at 52 degrees C. The activity remained constant at 4 degrees C for at least 143 days. The purified enzyme fraction hydrolyzed indoleacetic acid ethyl ester (Et-IAA) in addition to IAM and its homologue, 1-naphthalene-acetamide, but not indole-3-acetonitrile. Km values of the enzyme were 0.96 mM and 0.55 mM for IAM and Et-IAA, respectively. Although the molecular mass of the enzyme was very similar to that of IAM hydrolase of Agrobacterium tumefaciens involved in tumor formation, the biochemical properties of the enzyme including its high Km value were considerably different from those of the A. tumefaciens enzyme. Based on these enzyme properties, we will discuss whether the amidohydrolase is involved in auxin biosynthesis in rice cells.

Amidohydrolases↗

Effect of dark pretreatment on the kinetics of response of barley pulvini to gravistimulation and hormones.

Starch in pulvinus amyloplasts of barley (Hordeum vulgare cv Larker) disappears when 45-day-old, light-grown plants are given 5 days of continuous darkness. The effect of this loss on the pulvinus graviresponse was evaluated by following changes in the kinetics of response during the 5-day dark period. Over 5 days of dark pretreatment, the lag to initial graviresponse and the subsequent half-time to maximum steady state bending rate increased significantly while the maximum bending rate did not change. The change in response to applied indoleacetic acid (100 micromolar) plus gibberellic acid (10 micromolar) without gravistimulation, under identical dark pretreatments, was used as a model system for the response component of gravitropism. Dark pretreatment did not change the lag to initial response following hormone application to vertical pulvini, but both the maximum bending rate and the half-time to the maximum rate were significantly reduced. Also, after dark pretreatment, significant bending responses following hormone application were observed in vertical segments with or without added sucrose, while gravistimulation produced a response only if segments were given sucrose. These results indicate that starch-filled amyloplasts are required for the graviresponse of barley pulvini and suggest that they function in the stimulus perception and signal transduction components of gravitropism.

Darkness↗

Establishment of a highly efficient transformation system for pepper (Capsicum annuum L.).

Application of modern genetic manipulation has been limited in pepper ( Capsicum annuum L.) due to the lack of an efficient transformation system. Following the development of an efficient protocol for in vitro regeneration of pepper cotyledons, we investigated the key factors affecting transformation and established a highly efficient genetic transformation system using the pepper cotyledon as starting material. In this system, cotyledon explants are preconditioned for 2 days on kanamycin (km)-free DM1 medium [Murashige and Skoog (MS) salts/Gamborg B5 vitamins basal medium supplemented with 20 g/l sucrose, 5,000 mg/l DJ nutrients and a hormone combination of 1.0 mg/l indoleacetic acid (IAA) and 5.0 mg/l 6-benzyladenine (BA) solidified with 0.7% agar, pH 5.8], followed by co-cultivation with Agrobacterium tumefaciens on DM1 for 2 days and delay selection on DM1 with 500 mg/l carbenicillin (carb) for 2 days. The explants are then placed on DM1 containing 10 mg/l AgNO(3), 50 mg/l km-sulfate and 500 mg/l carb. After 4-5 weeks, the explants with buds are transferred to EM1 medium (MS salts/Gamborg B5 vitamins basal medium supplemented with 20 g/l sucrose, 5,000 mg/l DJ nutrients, 10 mg/l AgNO(3) and a hormone combination of 1.0 mg/l IAA, 3.0 mg/l BA and 2.0 mg/l gibberellic acid, solidified with 0.7% agar, pH 5.8) with 50 mg/l kanamycin and 500 mg/l carbenicillin for the elongation of buds. After 3-6 weeks, 1- to 2-cm-long elongated shoots are excised and planted on RM1 medium (MS basal medium supplemented with a hormone combination of 0.2 mg/l NAA and 0.1 mg/l IAA, solidified with 0.8% agar, pH 5.8) with 25 mg/l km and 200 mg/l carb for rooting. We tested four genotypes of pepper, and all presented a high differentiation efficiency (81.3% on average), elongation rate (61.5%) and rooting efficiency (89.5%). Polymerase chain reaction analysis results showed that 40.8% of the regenerated plantlets were transgenic plants.

Adenine↗

[Excretion of 5-hydroxyindole-3-acetic and 5-methoxyindole-3-acetic acids in cancer patients].

Daily urinary excretion of 5-hydroxy-3-indoleacetic acid (5-HIAA) and 5-methoxyindole-3-acetic acid (5-MIAA) was studied by chromatography-mass-spectrometry in patients with cancer and healthy subjects. A considerable increase in urinary 5-MIAA-excretion was shown in patients with cancer of the stomach, rectum and lung. In some cases, a correlation was found between high urine levels of carcinogenic 5-MIAA and similarly high levels of 5-HIAA excretion with urine.

Humans↗

Flavonoids induced in cells undergoing nodule organogenesis in white clover are regulators of auxin breakdown by peroxidase.

It was tested whether flavonoids that specifically accumulate in cells undergoing early nodule organogenesis could affect auxin turnover by a peroxidase to explain local changes in auxin distribution that occur during nodule formation in white clover (Trifolium repens cv. Haifa). A fluorometric assay was developed to determine the kinetics of indoleacetic acid (IAA) breakdown rates by a horseradish peroxidase in vitro. Three flavonoid compounds, which had previously been localized and identified, were purified from root tissue and their tissue concentrations estimated. A derivative of 7,4'-dihydroxyflavone (DHF), as well as free DHF, strongly inhibited auxin breakdown by peroxidase at concentrations estimated in the root tissue. Formononetin, an isoflavonoid accumulating in nodule primordia, accelerated auxin breakdown by peroxidase at concentrations estimated to be present in the roots. These results suggest that local changes in flavonoid accumulation could regulate local auxin levels during nodule organogenesis. The results are consistent with previous observations on the localization of auxin during nodule organogenesis. A model for the interaction of flavonoids with peroxidases is proposed to explain changes auxin during nodule development. A similar mechanism could be involved in lateral root and root gall formation.

Flavonoids↗

Radioautographic study of cell wall deposition in growing plant cells.

Segments cut from growing oat coleoptiles and pea stems were fed glucose-(3)H in presence and absence of the growth hormone indoleacetic acid (IAA). By means of electron microscope radioautography it was demonstrated that new cell wall material is deposited both at the wall surface (apposition) and within the preexisting wall structure (internally). Quantitative profiles for the distribution of incorporation with position through the thickness of the wall were obtained for the thick outer wall of epidermal cells. With both oat coleoptile and pea stem epidermal outer walls, it was found that a larger proportion of the newly synthesized wall material appeared to become incorporated within the wall in the presence of IAA. Extraction experiments on coleoptile tissue showed that activity that had been incorporated into the cell wall interior represented noncellulosic constituents, mainly hemicelluloses, whereas cellulose was deposited largely or entirely by apposition. It seems possible that internal incorporation of hemicelluloses plays a role in the cell wall expansion process that is involved in cell growth.

Autoradiography↗

Isolation and characterization of soybean-associated bacteria and their potential for plant growth promotion.

Endophytic and epiphytic bacteria were isolated from two soybean cultivars (Foscarin and Cristalina). Significant differences were observed in bacterial population densities in relation to season of isolation, soybean growth phase and the tissues from which the isolates were obtained. The isolates were identified by partial 16S rDNA sequence analysis, with most of the isolates belonging to the Pseudomonaceae, Burkholderiacea and Enterobacteriaceae groups. The potential of the isolates for plant growth promotion was evaluated by screening for indoleacetic acid (IAA) production and mineral phosphate solubilization; 34% of endophytic bacteria produced IAA and 49% were able to solubilize mineral phosphate whereas only 21% of epiphytic bacteria produced IAA although 52% were able to solubilize mineral phosphate. A high frequency of IAA producing isolates occurred in the early ripening Foscarin cultivar whereas a high percentage of phosphate solubilizing isolates were obtained from plants in the initial development stage (V6). We also found that 60% of endophytic and 69% of epiphytic isolates that produced IAA and solubilized mineral phosphate were also able to fix nitrogen in vitro. The soybean-associated bacteria showing characteristics related to plant growth promotion were identified as belonging to the genera Pseudomonas, Ralstonia, Enterobacter, Pantoea and Acinetobacter.

Bacteria↗

Auxin and light control of adventitious rooting in Arabidopsis require ARGONAUTE1.

Adventitious rooting is a quantitative genetic trait regulated by both environmental and endogenous factors. To better understand the physiological and molecular basis of adventitious rooting, we took advantage of two classes of Arabidopsis thaliana mutants altered in adventitious root formation: the superroot mutants, which spontaneously make adventitious roots, and the argonaute1 (ago1) mutants, which unlike superroot are barely able to form adventitious roots. The defect in adventitious rooting observed in ago1 correlated with light hypersensitivity and the deregulation of auxin homeostasis specifically in the apical part of the seedlings. In particular, a clear reduction in endogenous levels of free indoleacetic acid (IAA) and IAA conjugates was shown. This was correlated with a downregulation of the expression of several auxin-inducible GH3 genes in the hypocotyl of the ago1-3 mutant. We also found that the Auxin Response Factor17 (ARF17) gene, a potential repressor of auxin-inducible genes, was overexpressed in ago1-3 hypocotyls. The characterization of an ARF17-overexpressing line showed that it produced fewer adventitious roots than the wild type and retained a lower expression of GH3 genes. Thus, we suggest that ARF17 negatively regulates adventitious root formation in ago1 mutants by repressing GH3 genes and therefore perturbing auxin homeostasis in a light-dependent manner. These results suggest that ARF17 could be a major regulator of adventitious rooting in Arabidopsis.

Arabidopsis↗

[Growth-regulating activity of N-benzyl- and O-benzyl-containing compounds belonging to a new group of synthetic analogues of natural auxins].

We studied the effect of benzylamine, benzyl alcohol, and their derivatives (constituting a new group of synthetic analogues of natural auxins) on rooting of leaf and stem cuttings, rhizogenesis and growth of barley plantlets and tomato seedlings, and tomato plant productivity. These compounds promoted rooting of leaf and stem bean cuttings, increased rhizogenic activity, and stimulated the development of root systems in barley and tomato seeds. The activity of the compounds studied was similar to that of standard substances (3-indoleacetic acid potassium salt and 2-naphthylacetic acid). The benzyl group attached to the oxygen or nitrogen atom was shown to be the smallest molecular structure which provided auxin activity of the compounds. Derivatives of benzyl alcohol containing the quaternary ammonium fragment possessed auxin and anti-gibberellin (retardant) properties. They were selected by chemical synthesis of low-molecular-weight bioregulators with desired properties (a combination of chemical fragments with complementary physiological activity in the molecule). Auxin and anti-gibberellin (retardant) activities produced a synergistic effect. Germination of seeds treated with these compounds was accompanied by a more significant increase in the weight and length of roots (compared to standard auxins). The rate of seedling establishment reached 100%. The development of fruits and accumulation of reserve nutrient substances were synchronized and accelerated after spraying vegetating plants with solutions of studied compounds. The synergistic effect underlay a significant increase in the amount and quality of the crop (e.g., tomatoes).

Benzene Derivatives↗

Effect of ethanol on the oxidative metabolism of tryptamine by rat liver homogenate.

1 The effect of a wide range of ethanol concentrations (v/v) on indoleacetic acid (IAA) formation from the oxidative deamination of tryptamine was studied in vitro, in rat whole liver homogenate.2 IAA production was inhibited progressively by ethanol in concentrations between 0.01% to 0.2%, but the inhibition declined when the ethanol concentration was increased further to 6%.3 Ethanol-induced inhibition of IAA formation was only partially reversed by excess aldehyde dehydrogenase, whereas reductions in IAA formation were completely prevented by pyrazole or ethanol (6% and 10%) itself.4 Excess nicotinamide adenine dinucleotide failed to alter the inhibitory effect of ethanol and no evidence was obtained for inhibition of monoamine oxidase by ethanol or its metabolite, acetaldehyde.5 We conclude that ethanol indirectly inhibits IAA production as a result of oxidation of ethanol by alcohol dehydrogenase, during which the oxidative metabolism of tryptamine is shifted towards the reductive pathway, thus favouring the formation of tryptophol in place of IAA.

Acetaldehyde↗