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Enhanced Sensitivity to Ethylene in Nitrogen- or Phosphate-Starved Roots of Zea mays L. during Aerenchyma Formation.

Adventitious roots of maize (Zea mays L. cv TX 5855), grown in a well-oxygenated nutrient solution, were induced to form cortical gas spaces (aerenchyma) by temporarily omitting nitrate and ammonium (-N), or phosphate (-P), from the solution. Previously this response was shown (MC Drew, CJ He, PW Morgan [1989] Plant Physiology 91: 266-271) to be associated with a slower rate of ethylene biosynthesis, contrasting with the induction of aerenchyma by hypoxia during which ethylene production is strongly stimulated. In the present paper, we show that aerenchyma formation induced by nutrient starvation was blocked, under noninjurious conditions, by addition of low concentrations of Ag(+), an inhibitor of ethylene action, or of aminoethoxyvinyl glycine, an inhibitor of ethylene biosynthesis. When extending roots were exposed to low concentrations of ethylene in air sparged through the nutrient solution, N or P starvation enhanced the sensitivity to exogenous ethylene at concentrations as low as 0.05 microliters ethylene per liter air, promoting a more rapid and extensive formation of aerenchyma than in unstarved roots. We conclude that temporary deprivation of N or P enhances the sensitivity of ethylene-responsive cells of the root cortex, leading to cell lysis and aerenchyma.

Journal Article↗

Calcium and boron for in vitro rooting of Nothofagus nervosa.

Several mineral compounds influence adventitious rooting, and those containing boron and calcium, play an important role. They are associated to the enzyme regulation, cell division, cellular metabolism (induction stage) and roots enlargement (expression stage). The aims of this work were to study the variation of the endogenous calcium and boron concentrations along the rooting, and determine their optimal concentrations in the rooting basal medium of Nothofagus nervosa. Significant variation in endogenous calcium concentrations was detected along the rooting phase, strongly correlated with rhizogenesis phenomenons and linked to total peroxidase variation as cited elsewhere. Calcium and boron concentrations in the basal medium significantly affected the rooting. Treatment containing 60 mg.L-1 Ca and 1 mg.L-1 B produced root formation in fewer days than other combinations. The variation of these nutrient concentrations in the medium is a useful tool to improve microcutting rooting. Roots are obtained earlier and in a shorter period of time, finally resulting in more homogeneous microplants with a better developed rooting system.

Boron↗

Cortical development in roots of the aquatic plant Pontederia cordata (Pontederiaceae).

Adventitious roots of marsh-grown Pontederia cordata were examined to determine cortical development and structure. The innermost layer of the ground meristem forms the endodermis and aerenchymatous cortex. The outermost layer of the early ground meristem undergoes a precise pattern of oblique and periclinal cell divisions to produce a single or double layer of prohypodermis with an anchor cell for each radial file of aerenchyma cells. At maturity, endodermal cell walls are modified only by narrow Casparian bands. The central regions of the ground meristem become proaerenchyma and exhibit asymmetric cell division and expansion. They produce an aerenchymatous zone with barrel-shaped large cells and irregularly shaped small cells traversing the aerenchyma horizontally along radii; some crystalliferous cells with raphides are present in the aerenchyma. The walls of the hypodermis are modified early by polyphenols. The outermost layer of the hypodermis later matures into an exodermis with Casparian bands that are impermeable to berberine, an apoplastic tracer dye. The nonexodermal layer(s) of the hypodermis has suberin-modified walls. Radial files of aerenchyma are usually connected by narrow protuberances near their midpoints, the aerenchyma lacunae having been produced by expansion of cells along walls lining intercellular spaces. We are terming this type of aerenchyma development, which is neither schizogenous nor lysigenous, "differential expansion."

Journal Article↗

Biochemical character of the purified OsRAA1, a novel rice protein with GTP-binding activity, and its expression pattern in Oryza sativa.

OsRAA1, as previously reported, is a novel conserved protein in plants and plays an important role in rice root development. Overexpression of OsRAA1 results in reduced growth of primary roots and an increased number of adventitious roots. The biochemical functions and expression patterns of OsRAA1, however, remain poorly understood. To obtain purified OsRAA1 for biochemical analysis, the coding region was amplified by RT-PCR and expressed as a fusion protein with glutathione S-transferase in Escherichia coli. The antibodies to OsRAA1 were prepared by a synthetic 15-residue peptide (YYEDPSLFQFHKRGS) cross-linked with bovine serum albumin. Results of isotope labeling experiments suggested that OsRAA1 had binding activities with [alpha-32P]-GTP. The immunoprecipitation data showed that OsRAA1 had tissue-specific expression in roots and spikes rather than young shoots and leaves, which was consistent with its transcriptional expression. Our results indicate that OsRAA1 GTP-binding activity may contribute to the regulation mechanism of root development mediated by OsRAA1.

Animals↗

In vitro regeneration of roots of Phyla nodiflora and Leptadenia reticulata, and comparison of roots from cultured and natural plants for secondary metabolites.

Adventitious roots, generated using leaf explants of P. nodiflora, and meristem explants of L. reticulata, were cultured on Murashige and Skoog (MS) medium supplemented with napthylacetic acid (2 microM) and indole butyric acid (3 microM) respectively. After 30 days, subculturing of roots in liquid MS medium with napthylacetic acid (1.5 microM) for P. nodiflora and indole butyric acid (3 microM) for L. reticulata afforded considerable increase in root mass. HPTLC profiles and microscopic examination of transverse sections of in vitro and naturally grown roots provided information on secondary metabolite accumulation vis-à-vis developmental stages of the root.

Animals↗

Effects of arbuscular mycorrhizal colonization and phosphorus application on nuclear ploidy in Allium porrum plants.

Arbuscular mycorrhizal (AM) colonization can strongly affect the plant cell nucleus, causing displacement from the periphery to the center of the cell, hypertrophy and polyploidization. The hypertrophy response has been shown in a variety of AM plants whilst polyploidization has been reported only in Lycopersicon esculentum, a multiploid species with a small genome. In order to determine whether polyploidization is a general plant response to AM colonization, analyses were performed on Allium porrum, a plant with a large genome, which is much less subject to polyploidization than L. esculentum. The ploidy status of leaves, complete root systems and four zones of the adventitious roots was investigated in relation to phosphorus content, AM colonization and root differentiation in A. porrum plants grown under two different regimes of phosphate nutrition in order to distinguish direct effects of the fungus from those of improved nutrition. Results showed the presence of two nuclear populations (2C and 4C) in all treatments and samples. Linear regression analyses suggested a general negative correlation between phosphorus content and the proportion of 2C nuclei. The percentage of 2C nuclei (and consequently that of 4C nuclei), was also influenced by AM colonization, differentiation and ageing of the root cells, which resulted in earlier occurrence, in time and space, of polyploid nuclei.

Cell Nucleus↗

Expressional analysis of an EREBP transcription factor gene OsEBP-89 in rice.

OsEBP-89 gene encodes an ethylene responsive element binding protein (EREBP) transcription factor from rice (Oryza sativa). Northern blot analysis revealed that OsEBP-89 was expressed in root, stem, seeds, flowers and leaves of rice. Histochemical assay showed that GUS expressed mainly in phloem of vascular tissues of the root and stem transition region (RST), basal part of sheath roots, stem node and basal part of adventitious roots, also in endosperm of seeds in transgenic rice harboring OsEBP-89/GUS construct (pNSG). A sequence of region from C279 to C97 was found to play an important role for OsEBP-89 genes expression though promoter deletion assay. The possible function of OsEBP-89 gene was discussed.

Gene Expression Profiling↗

Amyloplast distribution in hairy roots induced by infection with Agrobacterium rhizogenes.

To elucidate the rapid and plagiotropic growth of hairy root induced by A. rhizogenes, a root apex was investigated with respect to it's amyloplast deposition, activity of alpha-amylase and glucose content. The amyloplasts distributed in the hairy roots were fewer than those of the adventitious root. Since auxin availability is enhanced in hairy roots, it could affect the statolith degradation by elevating alpha-amylase activity so that the energy requirement for rapid growth could be fulfilled as represented of glucose content. Consequently, it is suggested the overall decrease of starch grains could result in the lack of gravi-response in hairy roots.

Daucus carota↗

An easy and versatile embedding method for transverse sections.

In several research areas, transverse sections are indispensable for studying structural aspects of specimens. However, the oriented embedding of small cylindrical samples can become problematic, especially when transverse sections at right angles to the main axis of the object are desired. Here, we describe an easy and low-cost technique for oriented embedding of small (psi < 500 micro m) as well as of larger specimens (psi > 500 micro m). The usefulness of the technique is demonstrated for roots and stamens of Arabidopsis thaliana and for adventitious roots of Asplenium demerkense, as examples of small and larger cylindrical samples, respectively. Furthermore, several types of resin (glycol methacrylate, epoxy and acrylic resins) were successfully tested, showing the applicability of the technique for light and electron microscopy and for immunolocalizations. In conclusion, the principle of the technique can be extended to several resins and a wide variety of specimen types, such as stems, leaves and textile fibres. The originality of the technique lies in its simplicity combined with its high efficiency to produce well-oriented transverse sections.

Arabidopsis↗

Effect of La(III) on the growth and aging of root of loquat plantlet in vitro.

The effect of lanthanum on the adventitious root growth, activities of superoxide dismutase (SOD), catalase (CAT), and peroxidase (POD), O2* production rate, MDA content, and the relative plasma membrane permeability in roots of loquat test-tube plantlet were studied. Addition of lanthanum chloride (0.5 micromol/L) to the culture medium significantly increased the length of roots, increased the dry weight, enhanced the activities of SOD, CAT, and POD, decreased O2* production rate, malondialdehyde (MDA) contents, and the relative plasma membrane permeability. The relatively stable membrane structure of cell could defer the root aging of the plantlet in vitro. There are important theory meaning and practical value in applying LaCl3 in the rooting medium to raise the rate of rooting and transplant for wood plant.

Aging↗

Rooting cofactor activity of plant phytoalexins.

The pterocarpinoid phytoalexins, glyceollin, pisatin, and phaseollin, stimulated adventitious root formation in a mung bean rooting bioassay only in the presence of indoleacetic acid (rooting cofactor activity). Relatively low (5 to 50 micrograms per milliliter) concentrations of the phytoalexins were effective. The phytoalexins also increased the numbers of root primordia formed, therefore suggesting that they interacted with an early process in root formation.

Journal Article↗

Mycorrhiza-induced differential response to a yellows disease in tomato.

The protective effects induced by arbuscular mycorrhizal (AM) fungi against a phytoplasma of the Stolbur group have been investigated in tomato by morphometry and flow cytometry. Symptoms induced by the phytoplasma were less severe when the plants also harboured AM fungi. Morphological parameters such as shoot and root fresh weight, shoot height, internode length, leaf number and adventitious root diameter were closer to those of healthy plants when arbuscular mycorrhiza were present. Reduced nuclear senescence was observed in AM plants infected with phytoplasmas; the percentages of nuclear populations with different ploidy levels were intermediate between AM and phytoplasma-infected plants. The mechanisms underlying these interactions are discussed and a direct action of the AM fungus is hypothesized.

Flow Cytometry↗

Peroxidases, lignin and anatomy during in vitro and ex vitro rooting of gardenia (Gardenia jasminoides Ellis) microshoots.

In vitro and ex vitro rooting of gardenia (Gardenia jasminoides Ellis) microshoots with or without indolic-3-butyric acid (IBA) was studied in order to improve acclimatization of microplants after root formation and transplantation. Peroxidase (POD) activity and isoforms, lignin content and anatomical observations were evaluated in the course of the three interdependent phases (induction, initiation and expression) of microshoot rooting. Microshoots treated or not treated with IBA achieved high rooting percentages both in vitro and ex vitro. At the end of the 2-week acclimatization period, the percentage of surviving microplants ranged from 80% to 100%, for in vitro and ex vitro rooted microshoots, respectively. Microshoots rooted in vitro and ex vitro showed a relationship between rooting and POD activity but in a different time course. It appeared that root formation occurred after the microshoots had reached and passed a peak of maximum enzyme activity. In all treatments, electrophoretic analysis (native PAGE) of PODs revealed the appearance of one anionic and three cationic POD isoforms (C(1), C(3) and C(4)). An additional cationic POD isoform (C(2)) appeared only in the ex vitro rooting. The lignin content was similar in microshoots rooted both in vitro and ex vitro. The sequential anatomical changes during the rooting process were similar in both in vitro and ex vitro rooting treatments. In the case of in vitro rooting, pith cells had vacuoles entirely filled with a dark substance, while in the case of ex vitro rooting, pith cells contained many amyloplasts. The origin of the adventitious roots, in both rooting conditions, was located in the cambial ring. Roots with organized tissue systems emerged from the microshoot stem 10-14 days after the root induction treatments; on day 10 for rooting in vitro, while a 4-day delay was noted in microshoots rooted ex vitro.

Acclimatization↗

Signals controlling root suckering and adventitious shoot formation in aspen (Populus tremuloides).

We determined the effects of removal of leaves, stem axillary buds, or the entire shoot on root suckering (adventitious shoot formation by roots) and basal stem sprouts in 3- and 4-year-old potted seedlings of aspen (Populus tremuloides Michx.). The greatest number of root suckers (67.9 +/- 8.5 per plant) emerged after excision of the entire shoot. Defoliated and debudded stems were the major source of inhibitory agents for root suckering, although axillary buds and developing new leaves also exerted a significant inhibitory effect. Removal of mature leaves had only a minor effect on root suckering. Removal of a continuous band of bark (girdling) at the base of the stem consistently stimulated growth of adventitious shoots from the stem below the girdle and occasionally promoted root suckering. Exogenous application of indole-3-acetic acid to excised stumps inhibited root suckering and basal stem sprouting. Naphthylphthalamic acid (NPA), an auxin polar transport inhibitor, had no effect on root suckering or stem sprouting when it was applied to the bark of the basal stem. However, NPA significantly increased root suckering when it was applied to the exposed surface of xylem after girdling. These results suggest that polar transport of auxin in the xylem parenchyma is an important inhibitor of root suckering. On decapitated stems, vacuum extraction of xylem sap from the root system lowered the frequency of root suckering compared with decapitation alone, indicating that substance(s) originating in the root system also play a significant role in controlling root suckering.

Indoleacetic Acids↗

Dependency of Iron Reduction on Development of a Unique Root Morphology in Ficus benjamina L.

The activity of the Fe(3+) reductase of excised adventitious roots of Ficus benjamina L., grown in hydroponic culture without iron, was determined by a colorometric assay simplified by the use of a microplate reader. Reductase activity remained the same from pH 4.5 to 6.5 and decreased sharply above pH 6.5. Acetate buffer inhibited reduction. During early stages of root growth, excised roots did not exhibit Fe(3+) reductase activity. After several weeks and extensive root system development, Fe(3+) reduction still was not detectable in primary roots, but intermediate and high rates of reduction occurred in lateral and newly formed root clusters, respectively. Clustered roots only developed on plants grown at 0 or very low (<1 micromolar) iron. Microscopic examination revealed the root cluster to be composed of up to 30 lateral roots, usually less than 1 millimeter in diameter and 1 centimeter in length, that were completely covered with root hairs.

Journal Article↗

Effects of flooding on leaf dynamics and other seedling responses in flood-tolerant Alnus japonica and flood-intolerant Betula platyphylla var. japonica.

Two-year-old seedlings of Alnus japonica Steud. and Betula platyphylla var. japonica Hara were flooded from mid-June to early November, to study the effects of flooding on seedling survival and growth, morphological changes in stems and roots, leaf emergence, leaf fall, and leaf longevity. In A. japonica, growth was not affected by flooding, except for a slight decrease in height growth, but some morphological changes of stems and roots were observed, i.e., stem base hypertrophy, hypertrophied lenticels, formation of adventitious roots and development of new roots. In B. platyphylla var. japonica, growth was severely reduced by flooding and all seedlings died by the 20th week of flooding, without showing any adaptive morphological changes in stems or roots. Flooding induced rapid depression of leaf emergence, promoted leaf abscission, and reduced leaf longevity in B. platyphylla var. japonica. In contrast, in A. japonica, basal leaf senescence was delayed in flooded seedlings, thereby extending leaf longevity compared with unflooded seedlings.

Journal Article↗

Synthesis and biological activities of 4-chloroindole-3-acetic acid and its esters.

4-Chloroindole-3-acetic acid (4-Cl-IAA) and its esters were synthesized from 2-chloro-6-nitrotoluene as the starting material. The biological activities of 4-CI-IAA and its esters were determined by four bioassays. Except for the tert-butyl ester, 4-Cl-IAA and its esters had stronger elongation activity toward Avena coleoptiles than had indole-3-acetic acid. The biological activities of the methyl, ethyl and allyl esters were as strong as the activity of the free acid. All the esters, except for the tert-butyl, inhibited Chinese cabbage hypocotyl growth more than the free acid did, and all the esters induced severe swelling and formation of numerous lateral roots in black gram seedlings even at a low concentration. Furthermore, adventitious root formation was strongly promoted in Serissa japonica cuttings by all the esters. The root formation-promoting activities of the ethyl and allyl esters were about three times the value for indole-3-butyric acid which is used to promote and accelerate root formation in plant cuttings.

Avena↗

Expression, tissue distribution and subcellular localization of dehydrin TAS14 in salt-stressed tomato plants.

We previously isolated and characterized TAS14, and mRNA that is induced in tomato upon osmotic stress or abscisic acid (ABA) treatment and that shares expression and sequence characteristics with other dehydrin genes in different species. Affinity-purified antibodies against TAS14 protein were used to study the expression of TAS14 protein, both in seedlings and mature plants, its tissue distribution and its subcellular localization. TAS14 protein was not detected in 4-day-old seedlings but accumulated after ABA, NaCl or mannitol treatments. In NaCl-treated seedlings, some protein was detectable after 6 h of treatment and reached maximal levels between 24 and 48 h. Concentrations ranging from 5 to 12.5 g/l NaCl induced the protein to similar levels. In salt-stressed mature plants, TAS14 was expressed abundantly and continuously in aerial parts, but only slightly and transiently in roots. Immunocytochemical analysis of salt-treated plants showed TAS14 accumulated in adventitious root primordia and associated to the provascular and vascular tissues in stems and leaves. Immunogold electron microscopy localized TAS14 protein both in the cytosol and in the nucleus, associated to the nucleolus and euchromatin. Since TAS14 is a phosphoprotein in vivo, the classes of protein kinases potentially responsible for its in vivo phosphorylation were tested in in vitro phosphorylation assays. TAS14 protein was phosphorylated in vitro by both casein kinase II and cAMP-dependent protein kinase.

Abscisic Acid↗