Some aspects of the control of root growth and georeaction: the involvement of indoleacetic Acid and abscisic Acid

Apical segments of roots of Zea mays L. cv. Orla and cv. Anjou show a strong georeaction during 7 hours geostimulation. This is abolished by detipping the segments and restored by replacing the tips upon the apical cut surfaces. After exodiffusion of endogenous indoleacetic acid (IAA) the retipped segments showed a significantly lower geocurvature. Application of low concentrations of IAA to the basal cut surface of root segments from which endogenous IAA had not been allowed to exodiffuse increased the geocurvature of retipped Orla segments but decreased geocurvature of Anjou segments. At appropriate concentration basally applied IAA restored the georeaction capacity of root segments from which the endogenous auxin had exodiffused. The implications of the interaction between exogenous and endogenous IAA in the control of root georeaction are discussed with special reference to the normal role of endogenous IAA in the regulation of root georeaction and the variation in endogenous IAA content of roots of different cultivars of maize. The probability that the normal control of root growth and georeaction involves concomitant actions in the elongation zone of IAA moving preferentially in the acropetal direction and basipetally transported growth inhibitors (such as abscisic acid) produced in the cap cells is stressed.     

Effect of applied and endogenous indol-3-yl-acetic acid on maize root growth

The level of endogenous Indol-3-yl-acetic acid (IAA) measured by gas chromatography-mass spectrometry in the elongating zone of intact primary roots of Zea mays showed a good linear correlation with the growth rate of these roots. When they were treated with IAA, their relative elongation decreased; this indicates a supraoptimal content of endogenous IAA. However, the growth of some of the relatively rapidly extending roots was enhanced by such treatment. Interactions between endogenous and applied IAA in the control of root growth are discussed.Abbreviations GC-MS gas chromatography-mass spectrometry - IAA Indol-3-yl-acetic acid     

An alternative explanation for plant growth promotion by bacteria of the genus Azospirillum

Experiments were performed to identify the substances that are excreted by the soil bacterium Azospirillum brasilense Sp7 and that were reported to stimulate the formation of lateral roots and of root hairs of grasses. Azospirillum forms indole-3 acetic acid (IAA) but only in the late stationary growth phase or when tryptophan is present in the medium, but not in continuous cultures or in the logarithmic growth phase of batch cultures. Formation of IAA by Azospirillum requires aerobic conditions. Nitrite can replace IAA in several phytohormone assay, and is even more active than IAA in a test with wheat root segments in which the increase of wet weight is determined. Higher amounts of nitrite are necessary for activity in other classical auxin assays. Nitrite shows 40–60% of the activity of IAA in the straight-growth test of Avena coleoptiles and in the formation of C₂H₄ by pea epicotyl segments. Like IAA, nitrite is inactive in promoting C₂H₄ formation by ripe apple tissues. Since nitrite alone can hardly exert phytohormonal effects, it is postulated that nitrite reacts with a substance in the cells and that a product formed by this reaction functions as auxin. Such a substance could be ascorbate. Exogenously added ascorbate enhances the rate of nitrite-dependent C₂H₄ formation by pea epicotyl sections and the nitrite-dependent increase in the wet weight of wheat root segments. Nitrite is formed by nitrate respiration of Azospirillum. The findings that nitrite can have phytohormonal effects offers an alternative explanation of the promotion of the growth of roots and the enhancement of mineral uptake of grasses by Azospirillum. Indole-acetic acid completely and nitrite partly substitute for an inoculation with Azospirillum in an assay where the increase of the dry weight of intact wheat roots is determined after an incubation for 10 d. Nitrite and IAA are, therefore, possibly the only factors causing an enhancement of the growth of roots of grasses.Abbreviations HPLC high-performance liquid chromatography - IAA indole-3-acetic acid     

Stimulation of indoleacetic acid production in a <Emphasis Type="Italic">Rhizobium</Emphasis> isolate of <Emphasis Type="Italic">Vigna mungo</Emphasis> by root nodule phenolic acids

The influence of endogenous root nodules phenolic acids on indoleacetic acid (IAA) production by its symbiont (Rhizobium) was examined. The root nodules contain higher amount of IAA and phenolic acids than non-nodulated roots. Presence of IAA metabolizing enzymes, IAA oxidase, peroxidase, and polyphenol oxidase indicate the metabolism of IAA in the nodules and roots. Three most abundant endogenous root nodule phenolic acids (protocatechuic acid, 4-hydroxybenzaldehyde and p-coumaric acid) have been identified and their effects on IAA production by the symbiont have been studied in l-tryptophan supplemented yeast extract basal medium. Protocatechuic acid (1.5 μg ml⁻¹) showed maximum stimulation (2.15-fold over control) of IAA production in rhizobial culture. These results indicate that the phenolic acids present in the nodule might serve as a stimulator for IAA production by the symbiont (Rhizobium). Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users. An erratum to this article can be found at     

Effects of chlorogenic and caffeic acids on IAA oxidase preparations from sweet potato roots

IAA oxidase preparations from sweet potato (Ipomoea batatas) roots oxidised IAA in the absence of added phenolics. Activity was optimal around pH 6·8 and a minor pH optimum occurred around pH 4·3. Both chlorogenic and caffeic acids inhibited IAA oxidase activity at high concentrations (0·6–5·7 nmol/ml) but stimulated enzyme activity at low concentrations (0·10-0·55 nmol/ml); these effects were dependent on IAA and enzyme concentration and on pH. The activities of both substances are compared with those of other phenolics known to stimulate and inhibit plant IAA oxidases.     

Water stress and indol-3yl-acetic acid content of maize roots

Water-stress conditions were applied to the apical 12 mm of intact or excised roots ofZea mays L. (cv. LG 11) using mannitol solutions (0 to 0.66 M) and changes in weight, water content, growth and IAA level of these roots were investigated. With increasing stress a decrease in growth, correlated with an increased IAA level, was observed. The largest increase in IAA (about 2.7-fold) was found in the apical 5 mm of the root and was obtained under a stress corresponding to an osmotic potential of −1.39 MPa in the solution. This stress led to an isotonic state in the cells after 1 h. When the duration of water stress (−1.09 MPa) was increased to 2 or 3 h, no further increase in the IAA content was observed in the root segments. This indicated that there was no correlation between a hypothetical passive penetration of mannitol in the cells and IAA content. Indol-3yl-acetic acid rose to the same level in excised as in intact roots. In both cases, IAA accumulation was apparently independent of the hydrolysis of the conjugated form. The caryopsis and shoot seem not to be necessary to induce the increase of the IAA level in the roots during water stress (−1.09 MPa). Therefore, there seems to be a high rate of IAA biosynthesis in excised maize roots under water-stress conditions. Exodiffusion of IAA was observed during an immersion in either buffer or stress (−1.09 MPa) solution. In both cases, this IAA efflux into the medium represented about 50% of the endogenous level. Considering the present results, IAA appears to play an important part in the regulation of maize root metabolism and growth under water deficiency.     

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     

Synergistic effect of auxins and polyamines in hairy roots of <Emphasis Type="Italic">Cichorium intybus</Emphasis> L. during growth,coumarin production and morphogenesis

Hairy roots of Cichorium intybus obtained by infecting with different Agrobacterium rhizogenes strains (LMG-150 and A20/83) were studied for total endogenous indole-3-acetic acid (IAA) levels and indole-3-acetic acid oxidase (IAAO) activity. The roots initiated by LMG- 150 showed higher endogenous IAA levels as well as IAAO activity as compared to the roots from A20/83. Coumarin production in roots obtained by both of these strains strictly correlated with growth, with higher content in the roots obtained by LMG- 150. Moreover roots from LMG-150 showed increased growth index, length of primary roots and number of secondary and tertiary roots. The roots derived from LMG-150 were studied for total endogenous IAA and IAAO activity under the exogenous administration of polyamines and fungal elicitors. The treatment with putrescine (Put) at 1.5 mM level showed maximum endogenous IAA levels and IAAO activity as compared to the control and other polyamine administration, it also supported faster growth in terms of biomass accumulation, and total coumarin production. Of the various treatments, mycelial extract (ME) and culture media filtrate (CMF) of Pythium aphanidermatum and Phytopthora parasitica var. nicotiana, the treatment with 1 % CMF of P. parasitica var. nicotiana, resulted in maximum IAA levels and IAAO activity, which was supported by maximum biomass, coumarin production as compared to the control and other elicitor treatments. Two different regenerants of chicory obtained through A. rhizogenes LMG-150 designated as T-I and T-II, were studied for total endogenous IAA levels and IAAO activity. T-II showed higher titers of IAA with higher activity of IAAO as compared to T-I. Endogenous titer of IAA and IAAO activity was found to be maximum in transformed roots as compared to T-I, T-II, normal roots and normal plants. Our work showed a variation in endogenous auxin levels in these transformed plants. There exists a synergistic effect of endogenous IAA titers and polyamines in regulating root morphogenesis. Fungal elicitors influenced growth and coumarin production and an elicitor preparation of 1 % CMF of P. parasitica var. nicotiana gave spontaneous regeneration of shoots. The implications of these results are discussed.     

Endogenous and exogenous auxin in the control of root growth

The endogenous indol-3yl-acetic acid (IAA) of detipped apical segments from roots of maize (cv ORLA) was greatly reduced by an exodiffusion technique which depended upon the preferential acropetal transport of the phytohormone into buffered agar. When IAA was applied to the basal cut ends of freshly prepared root segments only growth inhibitions were demonstrable but after the endogenous auxin concentration had been reduced by the exodiffusion technique it became possible to stimulate growth by IAA application. The implications of the interaction between exogenous and endogenous IAA in the control of root segment growth are discussed with special reference to the role of endogenous IAA in the regulation of root growth and geotropism.Abbreviations IAA indol-3yl-acetic acid - GC-MS gas chromatography-mass spectrometry     

Comparative investigations of IAA metabolism in suspension cultures and plant organs fromBeta vulgaris (sugar beet) andChenopodium album L. (common lamb’s quarters)

Exogenoualy applicated indol-3-ylaeetic acid (IAA) is metabolized mainly to IAA aspartate in intact plants and plant segments and to IAA glucose in suspension cultures fromBeta vulgaris andChenopodium album. Main metabolic product of D-tryptophan is N-malonyltryptophan in both suspension cultures and hypocotyl segments of both species. The turnover rate of L-tryptophan to IAA is comparatively low (0.1 %); inBeta the turnover rate is higher than inChenopodium. In sugar beets phenmedipham leads to a decrease in the IAA. biosynthesis rate in suspension cultures of both plant species. There is, however, an increase in the IAA content in all intact plants. The metabolic activity is substantially higher in suspension cultures than in intact plants and plant segments.     

Comparison of some kinetic parameters of peroxidase and IAA oxidase in the course of growth and differentiation of plant cells

An attempt is made to characterize the functional activity of the protein moleculo possessing both peroxidase and IAA oxidase activity by comparing the kinetic parameters for the two types of enzyme activity with regard to the following substrates: H₂O₂, benzidine, guaiacol and IAA. The curves expressing the dependence of the enzyme reaction velocity on the concentration of the enzyme or the substrate are different depending on the enzyme extract origin and the type of the substrate. It is established that the K_m of peroxidase for IAA decreases while its K_m for H₂O₂ increases during cell development. Both types of enzyme activity show similar pH and temperature dependence. The presented data show that IAA oxidase activity of the peroxidase develops as extension and differentiation of the root cells proceed. This is one of the possible mechanisms through which peroxidase may participate in the regulation of growth and differentiation of the primary root cells of maize (Zea mays L.)     

The Effect of Pseudomonas putida Colonization on Root Surface Peroxidase

Increased activities of peroxidase and indole 3-acetic acid (IAA) oxidase were detected on root surfaces of bean (Phaseolus vulgaris) seedlings colonized with a soil saprophytic bacterium, Pseudomonas putida. IAA oxidase activity increased over 250-fold and peroxidase 8-fold. Enhancement was greater for 6-day-old than for 4- or 8-day-old inoculated plants No IAA oxidase or peroxidase activities were associated with the bacterial cells. Native polyacrylamide gel electrophoresis demonstrated that washes of P. putida-inoculated roots contained two zones of peroxidase activity. Only the more anodic bands were detected in washes from noninoculated roots. Ion exchange and molecular sizing gel chromatography of washes from P. putida-colonized roots separated two fractions of peroxidase activity. One fraction corresponded to the anodic bands detected in washes of P. putida inoculated and in noninoculated roots. A second fraction corresponded to the less anodic zone of peroxidase, which was characteristic of P. putida-inoculated plants. This peroxidase had a higher IAA oxidase to peroxidase ratio than the more anodic, common enzyme. The changes in root surface peroxidases caused by colonization by a saprophytic bacterium are discussed with reference to plant-pathogen interactions.     

IAA Oxidase preparations from fresh and aged Ipomoea batatas tuber discs

IAA oxidase preparations from fresh sweet potato tuber discs oxidized IAA only in the presence of added phenolic cofactors, and the pH optimum for enzyme activity depended on the cofactor used. Ageing of tuber discs, either by aeration in distilled water or by incubation on moist filter paper, resulted in increased peroxidase and phenol-stimulated IAA oxidase activities, as well as the development of IAA oxidase activity in the absence of added cofactors. High phenolase activity of fresh tuber discs decreased considerably with ageing. Phenol-stimulated IAA oxidase activity reached maximal levels before IAA oxidase activity in the absence of added cofactors. Enzyme preparations from aged tuber discs had double pH optima, similar to those previously described for sweet potato root IAA oxidase preparations. IAA in the concentration range 10^?4 to 10^?2 M inhibited the increase in peroxidase and IAA oxidase activities with ageing. DCP-stimulated IAA oxidase activities in preparations from both fresh and aged sweet potato tuber discs were inhibited by manganous ion.     

The Role of Auxins in Root-Knot Nematode-Induced Growth on Excised Tobacco Stem Segments

Root-knot nematodes, Meloidogyne incognita, induced lumps of callus tissue on the cambial surfaces of peeled tobacco stem segments cultured in vitro. Except for a layer 1 to 3 cells thick, callus was limited to the basal ends of control segments. Indole-3-acetic acid (IAA) applied in agar blocks to the centers of stem segments, when it had any effect on the cambial surface, induced streaks of callus extending from the blocks toward the basal ends of the segments. IAA in agar blocks also increased callus growth at the basal ends of the segments, increased the growth of pith on the undersides of the segments, promoted root initiation, but inhibited bud initiation. Nematodes produced none of these effects, nor did they change the type of organs induced by various concentrations of IAA in the medium. Callus tissue did grow on the cambial surface of stem segments surrounding agar blocks containing 2,3,5-triiodobenzoic acid, an inhibitor of polar auxin transport. Paraffin sections showed that the nematodes were confined to the callus tissue on the cambial surfaces of the segments. Except for occasional syncytia and areas of cell division, nematode-induced callus was composed of thin-walled, irregularly shaped cells arising from the cambium. Differences between the responses of tobacco stem segments to root-knot nematodes and IAA-agar blocks indicate that auxins were not freed from the plant tissue nor secreted by the nematodes. Instead, it is suggested that nematodes enabled the tissue to retain and use endogenous auxins that otherwise would have been transported to the basal ends of the segments.     

生长素和模拟微重力效应对大白菜不定根形态发生的影响

在生长素诱导下,大白菜(Brassicacampestris．Spp．Pekinensis)的下胚轴切段显示了一定的发根能力,其中,0．4—1．0mg／LIAA显著地促进大白菜不定根的发生。在生长素诱导24h后,可借助显微切片观察到下胚轴切面明显的解剖学变化首先是中柱鞘内靠近韧皮部的薄壁细胞的细胞质与细胞核变浓,染色加深,部分细胞分裂;随后是分裂的细胞团增大,逐渐形成根原基并分化出根冠。当下胚轴切段培养5天后,大量不定根穿破皮层,达到肉眼可见的程度。同一外植体中不定根的发育是不同步的,下胚轴不同部位的切段具有不同的发根能力;当下胚轴切段在培养基上反插时,提高外源IAA可修饰根发生的极性,提高蔗糖浓度能增强IAA的修饰作用;在模拟微重力效应条件下,不定根发生的极性没有明显变化,但是,增加了外植体对IAA诱导发根的敏感性。本结果为进一步研究不定根发生的分子机制建立了试验系统。     

Developmental changes in the gibberellin-induced growth response in stem segments of light-grown pea genotypes

The effects of GA on stem elongation were studied using segments from one tall and three dwarf light-grown pea genotypes varying in endogenous hormone content. Stem segments were cut at two distinct ages: when the fourth internode was at about 6–13% of full expansion (early-expansion) or at 18–25% of full expansion (mid-expansion). Light microscopy and flow cytometry were used to demonstrate that GA does not induce cell division in excised pea stem segments. The growth studied here was strictly elongation. Measurement of final segment length after 48 hours and high resolution measurement of growth kinetics over 20 hours using an angular position transducer were done on segments treated with hormone solutions. Our data indicate that the action of GA on stem elongation can be classified into two distinct modes. The first, apparent in early-expansion stem segments, shows distinct growth kinetics and is independent of the endogenous IAA concentration of the segments. Quantitation of IAA by GC/MS in early-expansion segments of wild type pea incubated with gibberellin shows that an increase in IAA concentration is part of the GA response in such segments. The second mode of GA action is evinced in mid-expansion segments. Whereas there is no short term (<20 h) response to GA alone (as determined by growth kinetics), there is a long term (48 h) response whose magnitude decreases across the genotypes with decreasing endogenous hormone content. Growth responses indicate that in mid-expansion segments exogenous GA acts by enhancing IAA action but appears to be unable to augment endogenous IAA content. Contradictory reports of the response of excised stem segments to GA can be reconciled when tissue genotype and developmental stage are considered.     

生长素和模拟微重力效应对大白菜不定根形态发生的影响

Under the induction of indole-3-acetic acid (IAA), adventitious roots were differentiated on hypocotyl segments derived from seedlings of Chinese cabbage (Brassica campestris spp. pekinensis). IAA at concentration of 0.4-1.0 mg/L in solid MS medium incited many adventitious roots on hypocotyl segments. The earliest anatomic changes were observed on cut surface of hypocotyl segments under optical microscope 24 hours after IAA treatment: cytoplasmic and nuclear density became higher in a few of parenchytmatous cells adjacent to phloem in tissue of pericycle, followed by cell divisions. Lately, the dividing cells expanded and developed into root primordium from which root cap was differentiated. After five days, most roots protruded through hypocotyl cortex and appeared just below the cut surface. The rooting capacity of the segments derived from three regions of each hypocotyl was different. High level of IAA modified the polarity of root formation on segment inserted upside down and sucrose increased the function of IAA. Additionally, microgravity did not significantly change the rooting polarity under the condition of stimulated microgravity, but it increased the competence of explants to IAA treatment. The results presented here provided an experimental system for further investigation of molecular events associated with adventitious root initiation.     

Effect on Root Growth of Endogenous and Applied IAA and ABA: A Critical Reexamination

Applications of indole-3yl-acetic acid (IAA) and abscisic acid (ABA) were done on two-day-old intact maize (cv LG 11) roots. The effect of the treatment on the root growth depends on their initial elongation rate. The slow growing roots were all inhibited by exogenous IAA and ABA at any concentrations used whereas for the fast growing roots their elongation was promoted by these two hormones at low concentrations. Quantitative analyses of endogenous IAA and ABA were performed using the gas chromatography-mass spectrometry technique. Detection and quantification of endogenous IAA and ABA were done on the zone of the root implicated in elongation. These techniques were achieved by electron impact on the IAA-Me-heptafluorobutyryl derivative and by negative ion chemical ionization with NH₃ on the ABA-Me ester derivative. A negative correlation between the growth and the endogenous content of these two hormones was obtained. ABA presented a larger range of endogenous level than IAA on the whole population of roots tested. When using applied IAA and ABA at different concentrations the same differentiating effect on the growth was observed. This allowed us to conclude that for identical concentrations, IAA has a more powerful effect on root elongation than ABA. Present results are discussed in relation to previous data related to the role of IAA and ABA in the growth and gravireaction of maize roots.     

Distribution of Indole-3-Acetic Acid in the Apoplast and Symplast of Squash (Cucurbita maxima) Hypocotyls

The concentration of endogenous IAA was higher in an apoplasticsolution (2.3xl0^–7M) than in a symplastic solution (0.5x 10^–7 M) obtained from segments of etiolated squash (Cucurbitamaxima Duch.) hypocotyls. Exogenously applied IAA (10^–5M) increased the level of IAA in both the apoplastic and thesymplastic solution. The concentration of IAA in the apoplasticsolution increased to 75% of the concentration of exogenousIAA in 4 h, but that in the symplastic solution increased onlyto 23% of the concentration of exogenous IAA. These resultsdemonstrate that the concentration of endogenous IAA is higherin the apoplast than in the symplast of squash hypocotyls, andthey suggest that IAA exerts its physiological effects, at leastto some extent, from the outside of cells. (Received September 20, 1996; Accepted January 10, 1997)     

Indole-3-acetic Acid Levels and the Role of Indole-3-acetic Acid Oxidase in the Normal Root and Club-root of Cabbage

The auxin content of club-root (Plasmodiophora brassicae Wor.) is 50–100 times higher than that of normal cabbage root. The importance of this difference in the disease development is discussed. Both normal root and club-root of cabbage contain allosteric IAA oxidase and IAA oxidase with ordinary kinetic properties. In normal cabbage root the allosteric one is associated with cell fractions sedimenting at 20,000 × g and 105,000 × g, in club-root it remains in the supernatant after 105,000 × g centrifugation. IAA oxidase with conventional kinetic properties is present in both these tissues in the cell fraction sedimenting at 10,000 × g, which contains mainly cell wall fragments. It is concluded that IAA oxidase is not primarily involved in regulation of the endogenous IAA level.