Effects of desiccation on the 77°K fluorescence properties of the liverwort Porella navicularis and the isolated lichen green alga Trebouxia pyriformis

The uptake of the auxin type herbicide 2,4-D into rice seedlings ( Oryza sativa L. cv. Dunghan Shali) and its effects on the K⁺, NH⁺₄ and NO₃ ion uptake and the K⁺ content were investigated at different pH values. A short incubation of the roots in 0.01 m M 2,4-D caused a marked ion uptake inhibition only at low pH. The non-auxin type herbicide benthiocarb did not produce such an inhibitory effect. Lowering of the pH in the external medium led to an increased 2,4-D uptake by the roots. These results can be explained by the increased H⁺ permeability of the membranes, allowing a more rapid entrance of 2,4-D into the root cells, thereby inhibiting the active ion uptake. Rice roots not subjected to 2,4-D treatment responded to H⁺ stress with an increased anomalous K⁺ uptake and a decreased K⁺ content. With reference to the effects of pH changes on the ion and 2,4-D uptake, possible transport mechanism of NH⁺₄ and 2,4-D are briefly discussed.     

Induction of anthocyanin synthesis in relation to embryogenesis in a carrot suspension culture: Correlation of metabolic differentiation with morphological differentiation

A system in which anthocyanin synthesis could be induced under a defined condition, was established in a carrot suspension culture. A cell suspension culture of carrot ( Daucus carota L. cv. Kurodagosun) was subcultured for more than a year in a medium containing 5 × 10⁻⁷ M 2,4-dichlorophenoxyacetic acid (2,4-D). At every subculture the cultures were sieved through nylon screens and the cells and cell clusters collected in the size range of 31–81 μm were transferred to a fresh medium. When the cells were transferred to a medium without auxin, synthesis of anthocyanin was induced. Zeatin promoted anthocyanin synthesis in a medium lacking auxin, with maximum yields of anthocyanin obtained at 10⁻⁷ to 10⁻⁸ M zeatin, 2,4-D at higher concentrations than 10⁻⁷ M inhibited anthocyanin synthesis completely. The sieved cells were fractionated by Ficoll density gradient centrifugation. Somatic embryos were formed in the fraction of higher density (>14% of Ficoll) in a medium containing 10⁻⁷ M zeatin but lacking auxin, while synthesis of anthocyanin was hardly observed. On the other hand, cells in the fraction of lower density (<12% of Ficoll) synthesized anthocyanin in the same medium, but formed few embryos. Forty to fifty percent of the total cells in this lighter cell fraction synthesized anthocyanin at a maximum. The similarity between anthocyanin synthesis and embryogenesis was observed in the time course as well as in the effects of growth regulators. The correlation between metabolic and morphological differentiation is discussed.     

Effect of 2,4-dichlorophenoxyacetic acid on polysomal profiles and polyadenylated RNA in excised tuber tissue of Jerusalem artichoke (Helianthus tuberosus)

Dormant tuber tissue of Jerusalem artichoke ( Helianthus tuberosus L.) can be stimulated by wounding to initiate RNA and protein synthesis. No DNA synthesis or cell divisions occur unless an auxin is provided. Changes in polysomal profiles and levels of Poly(A)⁺-RNA in response to wounding and auxin treatment were studied. Polysomes were isolated at various times after excision and incubation of tissue in the presence or absence of 10⁻⁵ M 2,4-dichlorophenoxyacetic acid. Polysomal profiles were studied by sucrose density gradient centrifugation. Dormant tissue contained ribosomes mainly in monosome form. Within 4 h of excision, a significant increase in the polysomal fraction was observed both in control and auxin-treated tissue. Increases in polysomes continued during the next 20 h. Poly(A)⁺-RNA was isolated from total polysomal RNA by oligo(dT)-cellulose column chromatography. There was a large increase in the amount of poly(A)⁺-RNA within 4 h of excision. During the first 43 h of incubation, levels of total polysomal RNA as well as poly(A)⁺-RNA in tissue treated with 2,4-dichlorophenoxyacetic acid were significantly higher than those in controls.     

Adaptation of corn roots to exogenously applied auxin

The ability of intact primary roots of corn ( Zea mays L. Bear Hybrid WF 9 × 38) to adapt to growth-inhibitory concentrations of auxin was studied using a highly sensitive position sensor transducer to measure growth. The timing, concentration dependence and temperature dependence of adaptation were studied as well as the time course of loss of adaptation upon removal of auxin. The rate of root elongation is inhibited 80% within 40 min after application of 10⁻⁷ M IAA. Within 90 min growth rate begins to recover. For concentrations of IAA equal to or greater than 10⁻⁷ M , recovery of growth rate (adaptation) is incomplete. Corn roots show a similar pattern of adaptation to the synthetic auxins NAA and 2,4-D. The Q₁₀ for adaptation is high (3.2) and comparable to that for root growth (3.3). Upon removal of exogenous IAA, loss of adaptation occurs with full sensitivity to the hormone regained within 20 min.
Based on the auxin specificity and the Q₁₀ for adaptation it is concluded that adaptation occurs neither by a change in the auxin degradation capacity of the root nor by a diffusional redistribution of applied auxin. It is suggested that adaptation involves metabolic processes, perhaps a metabolically dependent alteration of the number or affinity of auxin binding sites.     

Herbicidal activity of 4-chloroindoleacetic acid and other auxins on pea, barley and mustard

The natural auxins, 4-chloroindoleacetic acid and its methyl ester have strong herbicidal effects on pea, Pisum sativum , a plant in which they occur naturally. The standard herbicide, 2,4-dichlorophenoxyacetic acid (2,4-D) is only 5 times more effective than 4-chloroindoleacetic acid. The I₅₀, the dose inhibiting yield by 50%, for 4-chloroindoleacetic acid and its methyl ester is 0.5 kg ha⁻¹ or 15 mg kg⁻¹ fresh weight, close to the concentration of 4-chloroindoleacetic acid methyl ester in maturing pea seeds. Naphthaleneacetic acid and indoleacetic acid are also inhibitory, but at much higher concentrations. In its inhibiting effect on white mustard, Sinapis alba , 4-chloroindoleacetic acid approximates that of 2,4-D, whereas in barley, Hordeum vulgare , it is a stronger herbicide than 2,4-D. All auxins tested killed white mustard at low doses, but none killed barley. Both 4-chloroindoleacetic acid and 2,4-D killed pea. The chloroindole auxins of pea may be the hypothetic death hormones or senescence factors that are secreted from the developing seeds into the parent plant which is strongly inhibited or killed and from which the nutrients are mobilized and translocated to the seeds. The action mechanism of auxin type herbicides may be to simulate the action of endogenous herbicides.     

2,4-Dichlorophenoxyacetic acid inhibition of potassium transport in barley seedlings

Growth, potassium uptake and translocation as well as transpiration rates were measured in intact low-salt barley seedlings ( Hordeum vulgare L. cv. Union) in the presence of different 2,4-D concentrations at pH 6.5. Growth was only affected at 10^-3 M .
Above 10^-7 M 2,4-D both uptake by the roots and transport to the shoots were inhibited. The inhibition at 10^-5 M remained constant for at least 24 h. Furthermore inhibition of uptake was measurable within 1 h. Excised roots and roots of intact plants showed the same uptake pattern.
It is suggested that the observed effects were caused by 2,4-D-induced changes in uptake and translocation systems in the roots. Pre-treatment with 10^-5 M 2,4-D had no effect upon subsequent potassium uptake. Transpiration was reduced within 1 h in 10^-4 or 10^-3 M 2,4-D, probably due to changes in water transport or root permeability.     

Tonoplast H+ pumps are activated during callus formation of tuber tissues of Jerusalem artichoke (Helianthus tuberosus)

Changes in tonoplast H⁺-ATPase (EC 3.6.1.3) and H⁺–PPase (EC 3.6.1.1) activities were examined during the early period of callus formation in tuber tissues of Jerusalem artichoke ( Helianthus tuberosus L.). In callus-forming tissues cultured on a medium containing 2,4-D, the ATP-dependent H⁺-translocation activity of tonoplast vesicles increased 3-fold after a 2-day lag phase, while the ATP-hydrolytic activity and amount of tonoplast H⁺-ATPase protein were relatively constant after the lag phase. In the control tissue disks cultured on a medium free of 2,4-D, large declines in ATP-hydrolytic and ATP-dependent H⁺-translocation activities were observed. By contrast, the PP-dependent H⁺-translocation activity of tonoplast vesicles increased about 8-fold during the first 3 days of culture without any lag phase, and regardless of the presence of 2,4-D in the culture medium. However, the PP-hydrolytic activity and amount of H⁺-PPase protein did not change during the culture period, independently of callus formation. Transfer of the control tissue disks to the 2,4-D-containing medium, however, resulted in a further rapid stimulation of PP-dependent H⁺-translocation as well as an activation of ATP-dependent H⁺-translocation. These results suggest that both tonoplast H⁺ pumps are involved in callus formation of tuber tissues of Jerusalem artichoke.     

Influence of exogenously supplied potassium and sodium salts on the abscisic acid-induced proline accumulation in barley leaf segments

A stimulation of the abscisic acid (ABA)-induced increase in proline was observed in leaf segments of barley ( Hordeum vulgare L. cv. Georgie) if K⁺ or Na⁺ were supplied in the external medium as salts of monovalent anions such as NO₃, Br, Cr and I, but not when sulphate or phosphate were used. To a lesser extent, the effect was evident also with RbCl, but it did not occur when chlorides of Li⁺. Cs⁺, NH₄⁺, Mg:⁺ and Ca²⁺ were used. Both KC1 and NaCl in the concentration range 2–100 m M influence the ABA-dependent proline accumulation to the same extent; the increase induced was about 100% at 10 m M , and reached a maximum between 60 and 100 m M. The effect is not due to the osmotic activity of the salts and does not seem to depend on changes in K⁺ and Na⁺ levels within the leaf tissue, but it is somehow linked to their external concentration. The existence of a specific interaction between ABA and K⁺ or Na⁺, possibly at the cell membrane level, is proposed.     

Effect of auxins on spermidine uptake into carrot protoplasts

The effect of an auxin, indole-3-acetic-acid (IAA), on spermidine uptake into protoplasts of carrot ( Daucus carota L. cv. Ingrid) was studied. In the presence of 1 m M Ca²⁺, IAA (10⁻⁷ to 10⁻⁴ M ) enchances [¹⁴C]-spermidine uptake into carrot protoplasts, while no stimulation occurs in the absence of Ca²⁺. The time course of the uptake with and without IAA is very rapid and reaches saturation within 1 to 2 min. Preincubation of protoplasts with IAA inhibits the spermidine uptake. La³⁺, known not to penetrate the plasmalemma, exerts the same effect as Ca²⁺, but gives lower uptake values than Ca²⁺. The application of vanadate, an ATPase inhibitor, strongly inhibits IAA-stimulated spermidine uptake, suggesting that an energy-dependent mechanism may be involved in this transport. Neither spermidine nor Ca²⁺ alone stimulate IAA uptake. The synthetic auxin 2,4-dichlorophenoxyacetic acid, yields the same results as IAA with regard to time course of spermidine uptake with and without preincubation while, unlike IAA, no significant effect was observed on the Ca²⁺ -induced increase of spermidine uptake.     

Auxin-stimulated ethylene production in fern gametophytes and sporophytes

The auxins indole-3-acetic acid (IAA) and 2,4-dichlorophenoxyacetic acid (2,4-D) stimulated ethylene production from gametophytes of the fern Pteridium aquilinum (L.) Kuhn. var. latiusculum (Desv) underw. ex Heller and sporophytes of the ferns Matteuccia struthiopteris (L.)Todaro and Polystichum munitum (Kaulf.) Presl. Treatment with Co²⁺ or l -α -(2-aminoethoxyvinyl)-glycine (AVG) eleminated or significantly reduced the stimulatory effects of IAA. Treatment with 1-aminocyclopropane-1-carboxylic acid (ACC) resulted in significantly greater rates of ethylene production from all tissues tested. Based on their response to auxin, ACC, AVG and Co²⁺, the ethylene biosynthetic pathway in these three lower vascular plants appears similar to that existing in angiosperms.     

Potassium channels in barley: cloning, functional characterization and expression analyses in relation to leaf growth and development

It is not known how the uptake and retention of the key osmolyte K⁺ in cells are mediated in growing leaf tissue. In the present study on the growing leaf 3 of barley, we have cloned the full-length coding sequence of three genes which encode putative K⁺ channels ( HvAKT1 , HvAKT2 , HvKCO1 / HvTPK1 ), and of one gene which encodes a putative K⁺ transporter ( HvHAK4 ). The functionality of the gene products of HvAKT1 and HvAKT2 was tested through expression in Xenopus laevis oocytes. Both are inward-rectifying K⁺ channels which are inhibited by Cs⁺. Function of HvAKT1 in oocytes requires co-expression of a calcineurin-interacting protein kinase ( At CIPK23) and a calcineurin B-like protein (AtCBL9) from Arabidopsis , showing cross-species complementation of function. In planta , HvAKT1 is expressed primarily in roots, but is also expressed in leaf tissue. HvAKT2 is expressed particularly in leaf tissue, and HvHAK4 is expressed particularly in growing leaf tissue. Within leaves, HvAKT1 and HvAKT2 are expressed predominantly in mesophyll. Expression of genes changes little in response to low external K⁺ or salinity, despite major changes in K⁺ concentrations and osmolality of cells. Possible contributions of HvAKT1 , HvAKT2 , HvKCO1 and HvHAK4 to regulation of K⁺ relations of growing barley leaf cells are discussed.     

葡萄风信子叶片及花器官组织培养研究

以4种葡萄风信子Muscari armeniacum、Muscari aucheri ‘Dark Eyes’、Muscari aucheri ‘Ocean Magic’、Muscari azureum的叶片以及M.aucheri ‘Dark Eyes’的花器官为外植体,进行诱导愈伤组织分化的研究,建立了葡萄风信子的组织培养体系。研究结果表明：葡萄风信子抽梗期叶片最适升汞灭菌时间为7 min;筛选出四种品种的叶片最佳诱导愈伤组织培养基均为：MS+2,4-D 0.5 mg·L^-1+TDZ 0.1 mg·L^-1;同一叶片不同部位的愈伤组织诱导率由高到低依次是叶片基部、叶片中部、叶片尖端;M.aucheri ‘Dark Eyes’品种的花器官出愈率由高到低依次是花梗、花瓣、花药;筛选出M.aucheri ‘Dark Eyes’花药的最佳诱导培养基：MS+2,4-D 0.1 mg·L^-1+6-BA 0.5 mg·L^-1。     

Oxygen influences benzyladenine and 2,4-dichlorophenoxyacetic acid levels in cultured embryogenic tissue of Norway spruce

It is known that reducing the partial pressure of O₂ influences the induction of somatic embryogenesis. We tested the hypothesis that O₂ causes changes in the endogenous levels of exogenously supplied benzyladenine (BA) or 2,4-dichlorophenoxyacetic acid (2,4-D). Embryogenic tissue of Picea abies was incubated under reduced (2.5, 5 kPa) and ambient (21 kPa) levels of O₂ for 1, 3, 7 and 11 days and the endogenous concentrations of BA and 2,4-D were measured. For all treatments the concentration of BA in the tissue increased until the third day. At day 3, the ratio of BA in the tissue relative to the initial concentration in the medium, was 3.9, 2.8 and 1.9 for tissue incubated under 2.5, 5 and 21 kPa O₂, respectively. The BA concentration then declined gradually. Uptake of 2,4-D was inhibited at low O₂ levels. However, 2,4-D gradually accumulated in tissue grown under hypoxia, so that high levels were reached by day 11. These shifts in the BA and 2,4-D levels also caused a transient increase in the BA to 2,4-D ratio in tissue incubated under hypoxia. Although relevant for the previously reported effects of oxygen on induction of embryogenic tissue, it is unlikely that oxygen-induced alterations in BA and 2,4-D levels alone suffice to explain these findings.     

Pathogenesis-related protein b1' in plants and in cell suspension cultures of Nicotiana glutinosa Nicotiana debneyi and an amphidiploid cross (N. glutinosa×N. debneyi)

The expression of PR-protein b₁' in plants and cell suspension cultures of Nicotiana glutinosa L., Nicotiana debneyi Domin, and an amphidiploid cross of these two species, a hybrid, has been investigated. An enzyme linked immunosorbent assay has been employed to determine the concentration of PR-protein b₁' in extracts. The PR-Protein b₁' was constitutively produced in intact plants of the hybrid (around 25 μg g⁻¹ leaf tissue), while only trace amounts of the protein (< 50 ng g⁻¹ leaf tissue) were found in plants of the two parents. In suspension culture, the concentrations of PR-protein b₁' were 8, 0.4 and less than 0.1 mg l⁻¹ medium for the hybrid. N. debneyi and N. glutinosa , respectively. Only trace amounts of the protein were found in extracts from cells. Seven days after infection by tobacco mosaic virus (TMV) the concentration of PR-protein b₁' in leaves of N. glutinosa was 22.5 μg g⁻¹ leaf tissue. In N. debneyi and the hybrid a relatively limited induction of PR-protein b₁' by TMV was observed. The influence of various phenoxyacetic acids on the expression of PR-protein b₁' in the 3 cell cultures has been investigated. Cultures of N. glutinosa responded to treatments with 2,4-D and 2,4,5-T while cultures of N. debneyi and the hybrid were essentially unaffected. In the former case a concentration of 5–10 mg l⁻¹ 2,4,5-T was optimal and cells were most responsive to the treatment 4 days after subcultivation. The concentration of PR-protein b₁' in elicited cell cultures of N. glutinosa was 2 to 4 mg l⁻¹ medium.     

Cation fluxes in the saps of Sinapis alba during the floral transition

Plants of Sinapis alba L. were induced to flower by either a single long day or a single displaced short day. The levels of three cations. Ca²⁺, Mg²⁺ and K⁺, were measured by atomic absorption spectrophotometry in exudates from roots, leaves and apical stem tips. The export of all three cations out of the root system (root exudate) was increased in induced as compared to non-induced plants. No changes were observed in cation export out of the mature leaves (leaf exudate). The supply of cations to the apical bud (apical exudate) did not originate from the phloem and, so, should mainly be of apoplastic origin. Only the supply of Ca²⁺ to the apical bud was increased, not the supply of Mg²⁺ or K⁺. The increase in Ca²⁺ supply was transient and occurred at about the same time as a conspicuous stimulation of cell division, previously detected in the apical bud.     

Role of Ca2+ in Differentiation Mediated by Nerve Growth Factor and Dibutyryl Cyclic AMP in PC12 Cells

Abstract: Nerve growth factor (NGF) and dibutyryl cyclic AMP (dbcAMP) have synergistic effects on the neurite outgrowth of rat pheochromocytoma PC12 cells. The sites of interaction between NGF and dbcAMP have been studied extensively; however, the role of Ca²⁺ in differentiation induced by the two agents remains unclear. To understand whether intracellular Ca²⁺ is involved in the differentiation induced by the two agents, PC12 cells were treated with NGF, dbcAMP, or NGF plus dbcAMP for 2 days, and then effects on neurite outgrowth, ATP-induced Ca²⁺ influx, and Ca²⁺ mobilization from intracellular Ca²⁺ pools were examined. NGF or dbcAMP alone enhanced neurite outgrowth and Ca²⁺ accumulation by nonmitochondrial Ca²⁺ pools or the thapsigargin (TG)-sensitive Ca²⁺ pool. The dbcAMP acted synergistically with NGF to increase neurite outgrowth and to enlarge the TG-sensitive Ca²⁺ pool. The synergistic effect occurred within the first hour of treatment with dbcAMP plus NGF. On the other hand, dbcAMP abolished NGF's ability to enhance ATP-induced influx of extracellular Ca²⁺. Therefore, NGF and dbcAMP induced different effects on Ca²⁺ signaling pathways through two different but interacting pathways. In PC12 cells pretreated with TG to deplete the TG-sensitive Ca²⁺ pool, the dbcAMP- or dbcAMP plus NGF-mediated neurite outgrowth was significantly inhibited, whereas NGF-mediated neurite outgrowth was not affected by TG pretreatment. Our results suggest that the intracellular nonmitochondrial Ca²⁺ pools were changed in the differentiation process and were necessary for the synergistic effect of NGF and dbcAMP.     

Protein kinase inhibitor K-252a and fusicoccin induce similar initial changes in ion transport of parsley suspension cells

The protein kinase inhibitor K-252a induces a rapid, transient decrease of extracellular pH and [K⁺], and a concomitant increase in extracellular [Ca²⁺] in suspensions of cultured parsley cells. These effects are subsequently reversed. As with K-252a, fusicoccin also induces similar changes in pH and extracellular [Ca²⁺], but reversion does not occur. Acidification by HCI also leads to an increase in external [Ca²⁺], suggesting that the changes in extracellular [Ca²⁺] are mainly due to a pH-dependent displacement of Ca²⁺ ionically bound to the cell wall. The artificial acidification by HCI is rapidly followed by cell-mediated alkalinization, a process associated with K² release and rebinding of Ca²⁺. Any change in external pH or [K⁺] induced by K-252a, fusicoccin, or HCI is followed by an uptake of ⁴⁵Ca²⁺ into cellular pools. The results show that K-252a may be a valuable tool for studying the complex regulation of ion transport which may involve changes in the phosphorylation of unknown proteins.     

Effects of chloride and sodium on gas exchange parameters and water relations of Citrus plants

Gas exchange parameters, water relations and Na⁺/Cl^- content were measured on leaves of one-year-old sweet orange ( Citrus sinensis [L.] Osbeck cv. Hamlin) seedlings grown at increasing levels of salinity. Different salts (NaCl, KCl and NaNO₃) were used to separate the effects of Cl⁻ and Na⁺ on the investigated parameters. The chloride salts reduced plant dry weight and increased defoliation. Accumulation of Cl⁻ in the leaf tissue caused a sharp reduction in photosynthesis and stomatal conductance. By contrast, these parameters were not affected by leaf Na⁺ concentrations of up to 478 m M in the tissue water. Leaf water potentials reached values near −1.8 MPa at high NaCl and KCl supplies. This reduction was offset by a decrease in the osmotic potential so that turgor was maintained at or above control values. The changes in osmotic potential were closely correlated with changes in leaf proline concentrations. Addition of Ca²⁺ (as calcium acetate) increased growth and halved defoliation of salt stressed plants. Furthermore, calcium acetate decreased the concentration of Cl⁻ and Na⁺ in the leaves, and increased photosynthesis and stomatal conductance. Calcium acetate also counteracted the reductions in leaf water and osmotic potentials induced by salinity. In addition, calcium acetate inhibited the accumulation of proline in the leaves which affected the reduction in osmotic potential. These results indicate that adverse effects of salinity in Citrus leaves are caused by accumulation of chloride.     

Acute effects of the sodium salt of 2,4-D on the early developmental stages of bleak, Alburnm alburnus

The acute effects of a herbicide based on 2,4-D on embryos and larvae of bleak, Alburnus alburnus L. exposed to concentrations of 25–3200 mg 1⁻¹ for 12–48 hours were investigated. The 2,4-D delayed or stopped the development of embryos in their early stages, caused behavioural changes and morphological alterations. The L.C.₅₀ values for embryos varied between 12–9 and 15·9–4mg 1⁻¹ and for larvae from 51·6 to 111·2mg 1⁻¹ according to the time of exposure. The level of 2,4-D in Lake Balaton, or similar shallow lakes, without substantial injury of the littoral fish fauna should not surpass 0·5–1·0 mg 1⁻¹.     

Role of sodium in the ABA‐mediated long‐term growth response of bean to salt stress

Long‐term salt effects on plant growth have often been related to direct ion toxicity due to the accumulation of high ion concentrations in plant tissue. This work examines the relative importance of endogenous ABA, as well as Na⁺ and Cl⁻ toxicity, in the inhibition of leaf growth and photosynthesis, in bean plants grown at 1, 25, 50 and 75 m M NaCl until the fruit‐bearing stage. All salt‐treated plants showed very high leaf Cl⁻ concentrations, with little difference between plants exposed to 50 or 75 m M NaCl. The 25 and 50 mM salt‐treated plants were able to successfully exclude Na⁺ from their leaves, and only suffered an initial decline in the rate of leaf growth. Plants exposed to 75 m M NaCl showed an increase in Na⁺ leaf concentrations with an accompanying decrease in growth and photosynthesis as salt exposure progressed. A high correlation was found between leaf Na⁺ and leaf growth. Leaf ABA significantly increased with salt supply, and was highly correlated with both leaf Na⁺ and leaf growth. Our results suggest that in bean plants under long‐term salt stress, leaf ABA may participate in the regulation of leaf growth, and leaf Na⁺ would be at least partly responsible for increased ABA levels.