Brassinolide, a growth-promoting steroidal lactone

Brassinolide (BR), a naturally-occurring-steroidal lactone from rape ( Brassica napus L.) pollen, was compared with auxin for activity in a number of bioassay systems. Responses similar to IAA were elicited by BR in bioassays based upon bean hypocotyl hook opening, elongation of maize mesocotyl, pea epicotyl and azuki bean epicotyl sections, and fresh weight increase in Jerusalem artichoke (2,4-D used) and pea epicotyl sections. The azuki bean and dwarf pea epicotyl bioassays were much more responsive to BR than IAA (at 10 μ M ). Responses approximately two-fold greater in magnitude were elicited by IAA in the maize mesocotyl, bean hypocotyl hook and Jerusalem artichoke bioassays. Little or no response was elicited by BR (0.01 to μ M ) in the cress root or decapitated pea-lateral bud bioassays. A powerful synergism between BR and IAA was observed in the azuki bean, pea epicotyl and bean hypocotyl hook bioassays. Although, as previously reported, other steroidal substances are active in some of the bioassay systems tested, none compared with BR in magnitude and diversity of elicited responses.     

Stabilization of Cortical Microtubules in Maize Mesocotyl Cells by Gibberellin A3

Effects of GA₃ on the stability of cortical microtubules (MTs)were studied in mesocotyl cells of etiolated maize seedlings.Propyzamide, an MT-disrupting agent specific for plant tubulin,disrupted cortical MTs in cells in the upper regions of mesocotylsof GA₃-untreated seedlings and caused swelling of the cells.GA₃ prevented propyzamide from disrupting MTs and from causingsuch swelling. Chilling of mesocotyls at 4°C for 60 minbrought about the disruption of cortical MTs in cells in theupper regions of mesocotyls of GA₃-untreated seedlings, butnot in cells in corresponding regions of GA₃-treated seedlings,suggesting that treatment with GA₃ increased the stability ofcortical MTs in maize mesocotyl cells. Cortical MTs in protoplastsisolated from mesocotyls of GA₃-untreated seedlings failed towithstand chilling at 0°C for 90 min, while those in protoplastsisolated from mesocotyls of GA₃-treated seedlings withstoodchilling successfully. It appears that the cell wall is notinvolved in the stabilization of cortical MTs by GA₃ in maizemesocotyl cells. (Received July 6, 1993; Accepted November 29, 1993)     

Effect of Calcium Chloride Concentration on Growth and Sporulation of Saprolegnia terrestris Cookson ex Seymour

As CaCl₂ concentration in the growth medium was reduced from27.4 to 0.107 µM the range of oospore number per oogoniumin Saprolegnia terrestris increased from 1 to 8 to 1 to 25 ormore, oospore size decreased and the precision of correlationbetween oospore number per oogonium and oogonial diameter decreased,but colony d. wt was not affected. The threshold CaCl₂concentrationrequired to give the lower range of oospore number per oogoniumwas independent of the concentration of other mineral nutrientsin the growth medium. The number of oogonia formed per unitarea of colony was affected by the concentration both of CaCl₂and of other mineral nutrients. Oospores formed in 0.107 µMCaCl₂ medium were often multinucleate but otherwise of normalappearance. The oospore abortion rate was increased from 3.09per cent at 27.4 µM CaCl₂ to 8.2 per cent at 0.107 µMCaCl₂. calcium, oogonia, oospores, Saprolegnia terrestris Cookson ex Seymour     

Light or ethylene treatments induce transverse cell enlargement in etiolated maize mesocotyls

Dark-grown maize seedlings (hybrid WF-9 × 38-11) exposed for 1 or more hours to white light and then returned to darkness developed mesocotyls with enlarged apical diameters. This swelling response was an all-or-none response, and the fraction of the seedling population that showed the response depended on seedling age at irradiation. Irradiation of the coleoptile alone was nearly as effective in causing this response as was irradiation of the nodal region of the epicotyl, but irradiation of the mesocotyl base was ineffective. Removal of the coleoptile prior to irradiation did not prevent the formation of the light-induced swelling. Exogenously applied C₂H₄ (10 microliters per liter) for 24 hours in dark also induced swelling of the mesocotyl. The swelling induced in the intact seedlings was localized in the apical mesocotyl tissues with either light or C₂H₄ treatment, and maximal response to both treatments occurred with 3- to 4-day-old seedlings. Swelling of the mesocotyl was the result of transverse cell enlargement, not increase of cell numbers. The evidence suggests that light and C₂H₄ induce mesocotyl swelling in intact maize shoots by a common mechanism.     

Gibberellin Control of Microtubule Arrangement in the Mesocotyl Epidermal Cells of the d5 Mutant of Zea mays L.

The mesocotyl length of dark-grown maize seedlings is much shorterin the dwarf (d₅) than in the normal. The elongating zone ofd₅ mesocotyl is restricted to 1 mm below the coleoptilar node,while that of the normal extends to almost 4 mm. When the 4mm portion below the node was marked at 1 mm intervals and designatedas A, B, C and D from the top, gibberellin (GA₃) particularlystimulated elongation of the A–B region of d₅ and alsoC to some extent. Electronmicroscopic observation of microtubulesin the epidermal cells of these regions of d₅ showed that microtubuleswere longitudinally and/or randomly oriented in all other regionsexcept the region immediately below the node. Following 100µM GA₃ treatment for 21 h, the microtubule arrangementbecame transverse to the cell axis in A and B. Even in C a significantnumber of transversely oriented microtubules were observed.On the other hand, in normal seedlings microtubules were orientedtransversely to the cell axis in all four regions regardlessof GA₃ treatment. Results indicate that the dwarf habit of d₅seedlings can be ascribed to their possession of a small populationof elongating cells, the microtubules of which are predominantlyoriented transversely to the cell axis as a result of insufficientsupply of endogenous gibberellin. ¹Present address: Department of Cell Biology, National Institutefor Basic Biology, Myodaiji, Okazaki 444, Japan. (Received December 17, 1985; Accepted March 10, 1986)     

Basic nuclear proteins in the Oömycete fungus Achlya bisexualis

Summary A comparison was made of the basic proteins extracted from the chromatin and nuclei of Achlya bisexualis, Blastocladiella emersonii, and Pisum sativum. Extraction of purified chromatin and nuclei of the Chytridiomycete, B. emersonii followed by gel electrophoresis produced no detectable protein bands. Extractions of purified nuclei and chromatin by either mineral acid or CaCl₂ from the Oömycete A. bisexualis resulted in several protein bands following separation by disc gel electrophoresis. Monitoring the nucleic acids during the nuclear isolation procedure as well as comparing electropherograms of basic nuclear proteins with basic ribosomal proteins suggests no significant contamination of the nuclear preparations with ribosomes. Carboxymethyl cellulose chromatography of the A. bisexualis nuclear basic proteins resolved three distinct fractions. Gel electrophoresis of the CM cellulose fractions indicated heterogeneity of each fraction. Amino acid analysis of the CM fractions showed that they were all lysine-rich and meet the basisity requirements of histones.     

Effect of auxin on mesocotyl elongation of dark-grown maize under different seeding depths

In order to elucidate the physiological mechanism of maize mesocotyl elongation induced by auxin under different seeding depths, seeds of five maize inbred lines, including 3681-4 line tolerant to deep seeding, were treated with IAA and triiodobenzoic acid (TIBA) under seeding depths of 20 or 2 cm. Under deep seeding conditions, maize mesocotyls grew by 1.5–2.0 times faster than under shallow seeding conditions. IAA (10⁻⁶ to 10⁻⁴ M) applied to roots stimulated mesocotyl elongation only of 3681-4 line and only under deep seeding conditions. TIBA (10⁻⁵ and 10⁻⁴ M) applied to roots inhibited mesocotyl elongation in all lines, but only 3681-4 was sensitive to 10⁻⁶ M TIBA. IAA promoted only cell elongation, and TIBA inhibited both cell elongation and cell division. After IAA and TIBA treatments, endogenous IAA content changed in parallel with the mesocotyl growth rate under different seeding depths. Furthermore, ABP1 gene expression changed in parallel with the mesocotyl growth rate under deep seeding conditions. Therefore, deep seeding tolerance of 3681-4 line was achieved due to auxin-regulated rapid mesocotyl elongation.     

Effect of {alpha}-Tomatine on the Permeability of Plant Storage Tissues

The effect of the steroidal glycoalkaloid -tomatine on effluxof cell material from disks of beetroot, potato, apple, andcarrot has been investigated. A concentration of 1 mM tomatinecaused large increases in efflux from all tissues but 10 µMhad no effect. Leakage from potato and apple, but not from beetrootand carrot, was stimulated by 100 µM. tomatine, althoughlosses were smaller. Alkaloid-induced efflux of cell materialwas reduced considerably by addition of 1 mM CaCl₂ and to alesser extent by 1 mM MgCl₂. Cholesterol and its acetate didnot reduce leakage from beetroot but higher concentrations ofthe former caused small reductions in loss of material frompotato. The significance of these findings in relation to theaction of other membrane-active compounds is discussed.     

Salt Stress Induced Nuclear and DNA Degradation in Meristematic Cells of Barley Roots

Root growth of barley (Hordeum vulgare L., cv. Akashinriki)was inhibited by 200 raM NaCl, when 1 mM CaCl₂ was present inthe hydroponic culture solution. Increasing the CaCl₂ up to10 mM partially prevented this inhibition. However, inhibitionalso occurred with 100 mM NaCl in the presence of 0.1 mM CaCl₂.The nuclei of meristematic cells in roots in which growth hadbeen inhibited by salt stress were studied after staining withDAPI (4',6-diamino-2-phenylindol). Nuclear deformation of thecells occurred with 12 h of salt stress with 500 mM NaCl, andwas followed by degradation. The nuclear degradation was alsoobserved when the roots were exposed to more than 300 mM NaClfor 24 h. Biochemical analysis revealed that nuclear degradationwas accompanied by apoptosis-like DNA fragmentation. The intracellularmechanisms of nuclear degradation in cells after salt stressare discussed. ¹Emertius professor, Okayama University.     

Chromatin fractionation of normal and virus-transformed cells in culture

A simple technique for the obtaining of purified chromatin fractions from mammalian cells in culture is described. The procedure involves the isolation of clean nuclei in 0.30 M sucrose, 1.5 mM MgCl₂, 0.2 mM CaCl₂, 0.01 M Tris HCl pH 7.2, followed by sonication and differential centrifugation to collect specific chromatin fractions. Heterochromatin of SV-40 and Rous sarcoma virus transformed 3T3 cells showed a 6- to 7-fold enrichment in satellite DNA while a 2- to 3-fold enrichment in repetitive DNA was obtained in established and RSV transformed cells of Microtus agrestis. This method will facilitate the search for the site of integration of oncogenic viruses in the chromatin of mammalian cells.     

The Maize Glutamine Synthetase GS1- Gene is Preferentially Expressed in Kernel Pedicels and is Developmentally-Regulated

The pedicel region of Zea mays kernels contains a unique formof maize glutamine synthetase (GS), GS_pl. RNA blot analysisusing GS gene-specific probes revealed that the expression ofthe GS.₂ gene was specific to the pedicel and that it increasedin the kernels during development. This pattern of the maizeGS.₂ gene expression is consistent with the tissue specificityof the GS_pl protein and suggests that it encodes the GS_pl isoformof maize GS. (Received September 16, 1997; Accepted January 19, 1998)     

Synthesis of Isozymes of Superoxide Dismutase in Maize Leaves in Response to O3, SO2 and Elevated O2

Matters, G. L. and Scandalios, J. G. 1987. Synthesis of isozymesof superoxide dismutase in maize leaves in response to O₃ SO₂and elevated O₂.—J. exp. Bot 38: 842–852. The activities of the enzymes superoxide dismutase (SOD) andcatalase were determined in maize leaves treated with O₃or SO₂for8 h, or with elevated levels of oxygen for up to 96 h. NeitherO₃nor SO₂significantly increased the levels of superoxide dismutaseor catalase activity. However, after 72 h in an atmosphere containing90% oxygen, superoxide dismutase activity was increased, butnot the activities of catalase, ascorbate pcroxidase, and malatedehydrogenase. Immunological analysis showed that amounts ofthe cytosolic superoxide dismutase isozymes, SOD-2 and SOD-4,were increased by the elevated oxygen but not the chroloplast(SOD-1) or mitochondrial (SOD-3) isozymes. Immunoprecipitationof translation products of leaf polysomes indicated that thehigher levels of SOD-2 and SOD-4 were due to increased amountsof polysome-bound mRNA coding for these proteins. The specificresponse of SOD-2 and SOD-4 to 90% oxygen treatments contrastswith the increase in all SOD isozymes in maize leaves treatedwith the herbicide paraquat. Key words: Air pollutants, maize, oxidative stress, oxygen, superoxide dismutase     

Effects of incubation of nuclei from a long-day duckweed, Lemna gibba G3, under different ionic conditions on the activity of RNA synthesis

The effects of medium compositions used for incubation of nucleiisolated from a duckweed, Lemna gibba G3, on maintenance ofRNA synthetic activity were examined. The activity in the isolatednuclei decreased rapidly when they were incubated at 30^?C. Partof this decrease was regulated by the ionic conditions of theincubation medium. The optimum concentration of MgCl₂ for themaintenance of the activity was 30 mM. MgCl₂ could be replacedby CaCl₂. Potassium ion concentration above 100 mM caused adrastic decrease in activity. Magnesium ion had two differentfunctions: stabilization of the binding of RNA polymerase Iand of stimulating factors to nuclei. The patterns of SephacrylS-200 column chromatography of the stimulating factors presentin the nuclei during the day phase in a diurnal cycle were differentfrom those of the night phase. This pattern also changed accordingto whether the nuclei were extracted with a low (3 min) Mg mediumor an optimum (30 mM) Mg medium. We will discuss the role ofmagnesiumion in the occurrence of the diurnal rhythm of RNAsynthesis in the duckweed. (Received September 26, 1978; )     

Characterization of Exudation from Excised Roots of Onion, Allium cepa: II. CHEMICAL COMPOSITION OF EXUDATE

The exudate of excised onion roots bathed in simple KCl/CaCl₂solutions contains a variety of inorganic cations and anionstogether with quantities of organic molecules. This complexityof composition contrasts with the simplicity of the exudatesof maize roots under identical conditions and presumably reflectstransfer of these materials from the onion bulb to the rootsduring growth.     

Effects of CaCl2-washing on EPR Signals of Cytochrome b559 in Photosystem II Preparation from Spinach Chloroplasts

Washing of PS II preparation by 1 M CaCl₂ inactivates oxygenevolution without loss of bound manganese [Ono and Inoue (1983)FEBS Lett. 164: 255]. Most of the high-potential Cyt b₅₅₀, whichamounts to about a half of the total Cyt b₅₅₉ in untreated preparation,was converted to its low-potential form by CaCl₂-washing. Theeffect was similar to that of Tris-washing. The peak positionof the g_s band of the EPR spectrum of the CaCl₂-washed preparation(g=2.95) was the same as that of the low potential form of untreatedpreparation but was slightly different from that of the Tris-washedor heat-treated preparation (g=2.98). ¹ Present address: Department of Biology, Faculty of Science,Tokyo Metropolitan University, Fukazawa 2-1-1, Setagaya, Tokyo158, Japan. (Received November 14, 1984; Accepted January 30, 1985)     

Inducing salt tolerance in maize (Zea mays L.) through seed priming with chloride salts: Growth and ion transport at early growth stages

The study was conducted to determine whether salt tolerance could be induced in maize at germination stage by soaking of seeds for 8 h in distilled water or in 200 meq·L⁻¹ of NaCl, KCl, CaCl₂·2H₂O. Both primed and un-primed seeds were subjected for 14 days to 0, 100 or 200 mol·m⁻³ NaCl under controlled conditions. Although all priming agents were effective in alleviating adverse effects of salt stress on maize at germination stage, CaCl₂·2H₂O proved to be more effective since the seeds primed with this salt had significantly higher final germination, rate of germination and fresh and dry weights of plumules and radicles than those treated with other salts or distilled water. Concentration of Na⁺, K⁺ and Ca²⁺ increased significantly in all parts of germinating seeds of maize seeds primed with NaCl, KCl, or CaCl₂·2H₂O, respectively. In addition, seeds primed with CaCl₂·2H₂O were the highest in Cl⁻ accumulation in all parts of the germinating seeds, followed by seeds treated with NaCl and KCl. Most of the Ca²⁺ was retained in seeds and mesocotyl, because of which, transport of this ion to plumules and radicles was low.     

Auxin and Ethylene Control of Growth in Seedlings of Zea mays L. and Avena sativa L

The effects of applied ethylene on the growth of coleoptilesand mesocotyls of etiolated monocot seedlings (oat and maize)have been compared with those on the epicotyl of a dicot seedling(the etiolated pea). Significant inhibition of elongation by ethylene (10 µll^–1for 24 h) was found in intact seedlings of all three species,but lateral expansion growth was observed only in the pea internodeand oat mesocotyl tissue. The sensitivity of the growth of seedlingparts to ethylene is in the decreasing order pea internode,oat coleoptile and oat mesocotyl, with maize exhibiting theleast growth response. Although excised segments of mesocotyland coleoptile or pea internode all exhibit enhanced elongationgrowth in IAA solutions (10^–6–2 ? 10^–5 moll^–1), no consistent effects were found in ethylene. Ethyleneproduction in segments was significantly enhanced by applicationof auxin (IAA, 10^–5 mol l^–6 or less) in all tissuesexcept those of the eat mesocotyl. Segments of maize show a slow rate of metabolism of applied[2-¹⁴C]IAA (30 per cent converted to other metabolites within9 h) and a high capacity for polar auxin transport. Ethylene(10 µl l^–1 for 24 h) has little effect on eitherof these processes. The oat has a smaller capacity for polartransport than maize and the rate ef metabolism of auxin isas fast as in the pea (90 per cent metabolized in 6 h). Althoughethylene pretreatment does not change the rate of auxin metabolismin oat, there is a marked reduction in auxin transport. It is proposed that the insensitivity of maize seedlings toethylene is related to the supply and persistence of auxin whichcould protect the seedling against the effects of applied orendogenously produced ethylene. Although the mesocotyl of oatis sensitive to applied ethylene it may be in part protectedagainst ethylene in vivo by the absence of an auxin-enhancedethylene production system. The results are discussed in relationto a model for the auxin and ethylene control of cell growthin the pea.     

Some Basic Psychophysics of Calcium Salt Solutions

Detection thresholds and the taste qualities of suprathresholdconcentrations of calcium salt solutions were assessed. Averagetaste detection thresholds for calcium chloride (CaCl₂), lactate(CaLa), hydroxide, phosphate and gluconate ranged between 8and 50 mM, with no reliable differences among the various salts.Between-subject variability ranged over four orders of magnitudeand reliability coefficients for repeated detection thresholdtests of CaCl₂ averaged r = 0.52. In an odor detection test,subjects could reliably discriminate 100 but not 1 mM CaCl₂and CaLa from water. The taste of suprathreshold concentrations(1–100 mM) of CaCl₂ and CaLa was considered unpleasant.At 1 mM, CaCl₂ solution was rated as 35% bitter, 32% sour, 29%sweet and 4% salty. At higher concentrations the sweet componentdiminished and the salty component increased, so that 100 mMCaCl₂ was rated as 44% bitter, 20% sour, 1% sweet and 35% salty.CaLa solutions were considered to be significantly less bitterand marginally more sour than equimolar CaCl₂ solutions. Thus,the taste of calcium varied with both the form and concentrationof salt tested, but included both sour and bitter components.Saltiness was identified only in high (