Reevaluation of the Effect of Calcium Ions on Auxin-induced Elongation

The mechanism by which calcium ions inhibit cell elongation has been reinvestigated. Growth-inhibiting levels of calcium, when applied to isolated walls (in vitro treatment) do not decrease cell wall extensibility as measured by the Instron technique. Thus, the hypothesis that calcium inhibits growth by forming wall-stiffening calcium bridges must be abandoned. Treatment of living auxin-treated sections with calcium (in vivo treatment) does cause a decrease in the subsequently measured wall extensibility, but this decline appears to be simply a consequence of the growth inhibition rather than its cause. Growth-inhibiting levels of calcium do not appreciably reduce the rate of auxin-enhanced H(+) excretion. Pretreatment with calcium does not reduce the capacity of walls to undergo acid-activated wall loosening in the absence of calcium. High concentrations of CaCl(2) (0.02 m) cause an initial elastic shrinkage of Avena sections comparable to that caused by the same osmolarity of mannitol, but the subsequent growth inhibition is too great to be explained by an osmotic inhibition. Calcium ions do inhibit H(+)-induced extension of frozen-thawed sections under tension. The growth-inhibitory effects of calcium, then, may be ascribed to a direct inhibition exerted by calcium ions on the H(+)-induced wall-loosening process.     

Properties and Possible Physiological Significance of Cell Wall Calcium Binding in Etiolated Pea Epicotyls

Baydoun, E. A-H. and Brett, C. T. 1988. Properties and possiblephysiological significance of cell wall calcium binding in etiolatedpea epicotyls.—J. exp. Bot. 39: 199–208. The binding of ⁴⁵Ca²⁺ ions to cell walls prepared from pea epicotylswas examined in young and old parts of the epicotyl, and wasfound to be considerably greater, on a carbohydrate basis, inthe older, non-growing cells. A similar comparison between light-and dark-grown stems showed greater binding in the dark-grownstems. The polygalacturonase-insensitive component of the bindingcontained at least three types of binding with different affinities,and had an apparent pK of 4.3. The specificity of the bindingfor calcium ions was examined and a considerable degree of specificitywas observed. The specificity of inhibition by calcium of epicotylelongation was similar to the specificity of calcium binding.A specific calcium chelator, EGTA, when present at a concentrationof above 10 mol m^–3, promoted the extension of matureregions of the epicotyl, while inhibiting extension of youngertissue. Key words: Cell wall, calcium, pea epicotyl     

Stimulation of Endomitotic DNA Synthesis and Cell Elongation by Gibberellic Acid in Epicotyls Grown from Gamma-irradiated Pea Seeds

Large doses of γ-irradiation, given to air-dried pea seeds, inhibit the endomitotic DNA synthesis in pea epicotyls during germination in darkness. The cortex cells of the etiolated epicotyls reach only the 4 C DNA level, whereas cortex cells of unirradiated seeds reach the 8 C DNA level. Epicotyl elongation and cell elongation are also reduced.     

On the Nature and Origin of the Calcium Asymmetry Arising during Gravitropic Response in Etiolated Pea Epicotyls

Seven day old etiolated pea epicotyls were loaded symmetrically with ³H-indole 3-acetic acid (IAA) or ⁴⁵Ca²⁺, then subjected to 1.5 hours of 1g gravistimulation. Epidermal peels taken from top and bottom surfaces after 90 minutes showed an increase in IAA on the lower side and of Ca²⁺ on the upper side. Inhibitors of IAA movement (TIBA, 9-hydroxyfluorene carboxylic acid) block the development of both IAA and Ca²⁺ asymmetries, but substances known to interfere with normal Ca²⁺ transport (nitrendipine, nisoldipine, Bay K 8644, A 23187) do not significantly alter either IAA or Ca²⁺ asymmetries. These substances, however, are active in modifying both Ca²⁺ uptake and efflux through oat and pea leaf protoplast membranes. We conclude that the ⁴⁵Ca²⁺ fed to pea epicotyls occurs largely in the cell wall, and that auxin movement is primary and Ca²⁺ movement secondary in gravitropism. We hypothesize that apoplastic Ca²⁺ changes during graviresponse because it is displaced by H⁺ secreted through auxin-induced proton release. This proposed mechanism is supported by localized pH experiments, in which filter paper soaked in various buffers was applied to one side of a carborundum-abraded epicotyls. Buffer at pH 3 increases calcium loss from the side to which it is applied, whereas pH 7 buffer decreases it. Moreover, 10 micromolar IAA and 1 micromolar fusicoccin, which promote H⁺ efflux, increase Ca²⁺ release from pea epicotyl segments, whereas cycloheximide, which inhibits H⁺ efflux, has the reverse effect. We suggest that Ca²⁺ does not redistribute actively during gravitropism: the asymmetry arises because of its release from the wall adjacent to the region of high IAA concentration, proton secretion, and growth. Thus, the asymmetric distribution of Ca²⁺ appears to be a consequence of growth stimulation, not a critical step in the early phase of the graviresponse.     

Comparison of Auxin-induced and Acid-induced Elongation in Soybean Hypocotyl

Acid-induced growth was compared to auxin-induced growth. After a transient pH 4-induced increase in the elongation rate was completed, auxin could still induce an enhanced rate of elongation in soybean (Glycine max) hypocotyl segments. This auxin response occurred both when the medium was changed to pH 6 before auxin addition, and when the auxin was added directly to the pH 4 medium. This postacid response to auxin was persistent, and quite unlike a postacid response to acid, which was again shortlived. One mm N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid (pH 7) inhibited the first response to auxin (the first response to auxin being similar to the acid response), but not the second response. This did not appear to be simply a hydrogen ion neutralizing effect, however, since a 50-fold increase in buffer concentration at pH 6 did not inhibit the first response. Decrease in the pH of the external medium, previously shown to occur with excised soybean hypocotyl segments, was not affected by auxin. Furthermore, this pH drop, during which the cells appear to be adjusting their external pH to about 5.4, did not result in an increased rate of elongation. Addition of auxin after the equilibrium pH had been attained did not alter the pH, but it did increase the rate of elongation, eliciting a normal auxin response. It was concluded that hydrogen ions do not mediate in long term auxin-induced elongation in soybean hypocotyl.     

高钙逆境对拟南芥根部基因表达的影响

土壤钙含量过高会对植物正常生长发育形成严重的威胁.这是中国西南喀斯特区域农业生产和环境保护的重要限制因素.了解植物对高钙胁迫的响应机制,可以促进喀斯特地区可持续农业发展和生态环境保护.为揭示植物对高钙胁迫相应的分子机理,本研究充分利用拟南芥(Arabidpsis thaliana)丰富的生物信息资源,系统比较和分析了对照和高钙胁迫处理情况下拟南芥根部的基因表达情况.40 mmol/L CaCl2处理明显抑制了拟南芥根部生长.转录组分析表明,高钙处理下,拟南芥根部乙烯、脱落酸和茉莉酸途径表达上调,其中,茉莉酸合成、响应和代谢途径相关基因表达均上调,说明这3个激素相关途径,对植物应对高钙胁迫具有重要的作用,尤其是茉莉酸相关途径.     

Inhibitory Effect of Calmodulin Antagonists and Calcium Antagonists on Fertilization-Induced Cyanide-Insensitive Respiration in Sea Urchin Eggs

During initial several minutes after fertilization, sea urchin eggs exhibited high rate of respiration which was only slightly inhibited by cyanide. This cyanide-insensitive respiration was inhibited by calcium antagonists, diltiazem and verapamil, and calmodulin antagonists, N-(6-aminohexyl)-5-chloro-1-naphthalensulfonamide hydrochloride (W-7), N-(6-aminohexyl)-1-naphthalenesulfonamide hydrochloride (W-5) and chlorpromazine, which were added within 1 min after insemination. The inhibitory effect of W-7 on cyanide-insensitive respiration was higher than that of W-5. Cyanide-sensitive respiration of fertilized eggs observed after this initial period was not inhibited by these compounds. Ca²⁺ influx in eggs just after fertilization was inhibited by calcium antagonists but was rather enhanced by calmodulin antagonists. Fertilization-induced stimulation of cyanide-insensitive respiration probably results from calmodulin-dependent reactions which are activated by Ca²⁺ influx.     

Effect of Calcium on the Structure and Template Activity of Pea Chromatin

The absorption spectrum of native pea chromatin solubilizedunder minimal shearing conditions changed with increasing Ca²⁺concentration; the ratio of maximum to minimum absorption decreasedand the maximum absorption peak shifted to a longer wavelength.The concentration of Ca²⁺ to cause half complete sedimentationof chromatin was much lower for the solubilized native chromatin(more condensed and larger in size) than for the sonicated chromatin(less condensed and smaller in size). Solubilized native chromatinshowed a two-step melting profile in the absence of Ca²⁺. In the presence of Ca²⁺ the two T_ms disappeared and a new higherT_m appeared. Template activity of solubilized native chromatinincreased 3-fold upon dispersion and fragmentation by sonication.Addition of a small amount of ethylene glycol-bis (ß-aminoethylether)-N, N'-tetraacetic acid (EGTA) promoted the template activityof solubilized native chromatin, but not that of sonicated ordenatured DNA. The effect of EGTA was reversed by Ca²⁺. Thechromatin reconstituted in the presence of EGTA showed a lowerT_m than the chromatin reconstituted in the presence of Ca²⁺.The relationship between chromatin structure and its templateactivity is discussed in relation to Ca²⁺. (Received August 12, 1985; Accepted December 7, 1985)     

Hydroxyproline Formation and Its Relation to Auxin-induced Cell Elongation in the Avena Coleoptile

A study has been made of the effects on hydroxyproline formation of 4 factors that influence the rate of cell elongation in the Avena coleoptile; auxin, sugars, an external osmoticum, and actinomycin D. Hydroxyproline formation is increased by a combination of auxin and sucrose, but is affected to a much lesser extent by either factor alone. Its formation is inhibited by an external osmoticum but is scarcely affected by actinomycin D. The lack of correlation between the amount of hydroxyproline synthesis and the growth rate suggests that hydroxyproline formation is not involved in the actual process of wall loosening. It is suggested, instead, that if the wall is to retain its capacity for rapid extension, those hemicelluloses which are incorporated into it by intussusception rather than by apposition must be attached to a hydroxyproline-protein.     

The Association of Ascorbate and Ascorbate Oxidase in the Apoplast with IAA-Enhanced Elongation of Epicotyls from Vigna angularis

The level of (ascorbic acid (AA) plus dehydroascorbic acid (DHA))and the ratio of the level of AA to that of AA plus DHA in intercellularwashing fluid (IWF) of epicotyl segments from Vigna angularisdecreased from 2.8±0.3 to 1.2±0.5nmol (g fr wt)^–1and from 0.23±0.03 to 0.13±0.01, respectively,after incubation of the segments without IAA for 20 h at 27°C.However, these values changed to 5.3±1.7 nmol (g fr wt)^–1and 0.07±0.05 after incubation with 0.1 mM IAA. The activityof cell wall-bound ascorbate oxidase increased by about 20%and 70% after incubation without IAA and with IAA, respectively.However, the activity of cell wall-bound peroxidase was notaffected by incubation with or without IAA. The activities ofascorbate oxidase and peroxidase in IWF decreased by about 85and 75% after incubation without IAA. IAA did not affect thesedecreases to any great extent. A lignin-like compound was formedduring the incubation of epicotyl segments in the absence ofIAA. Formation of this compound was inhibited by IAA. The resultssuggest that one of the causes of the enhancement of elongationgrowth by IAA is the inhibition of peroxidase-dependent lignificationas a result of increases in levels of AA and DHA and in ascorbateoxidase activity. (Received August 16, 1993; Accepted December 6, 1993)     

Effects of Extreme Acceleration on the Germination, Growth and Cell Wall Composition of Pea Epicotyls

Pea (Pisum sativum) seedlings were germinated and grown in darknessfor 5 d whilst being subjected to continuous accelerations ofbetween 1 ? g and 1054 ? g. Increased acceleration retardedthe elongation of both epicotyls and roots, and inhibited lateralroot growth. Changes in the cell wall carbohydrate and lignincomponents were also recognized. Nevertheless, increasing accelerationfrom 1 ? g to 1054 ? g had no effect on germination and peaswere able to germinate and grow successfully at up to approximately10 000 ? g. Key words: Acceleration, cell walls, Pisum     

The Movement of Calcium in Germinating Pea Seeds

Pea seeds contain less calcium than phosphorus, potassium ormagnesium; more than half of this calcium is located in thetesta. Peas at either end of a pod have more calcium than thosein the middle. When pea seeds are allowed to germinate in water,less than 30 per cent of the cotyledonary calcium moved to thegrowing axis during the first 15 days of germination, whereas70–90 per cent of magnesium, potassium and phosphate wasexported. Various attempts to increase the amount of calciumexported were not successful. When radioactive calcium was appliedto the cotyledons, essentially no movement to the axis was observedunder conditions where extensive movement of radioactive phosphateoccurred.     

Effect of Some Calcium Antagonists on Muscarinic Receptor-Mediated Cyclic GMP Formation

Several calcium antagonists were screened for their effect on muscarinic acetylcholine receptor-mediated cyclic GMP formation in murine neuroblastoma cells (clone N1E-115). Mn2+, Ni2+, and verapamil rapidly antagonized the response noncompetitively, with the following order of potency: verapamil greater than Mn2+ greater than Ni2+. The effects of Mn2+ and Ni2+, but not those of verapamil, were largely reversed by increasing extracellular calcium concentration. Additional effects of these agents included displacement of [3H]quinuclidinyl benzilate binding by verapamil and elevation of cyclic GMP levels by Mn2+ and Ni2+ in the absence of agonists. These results are in support of the hypothesis that receptor-mediated cyclic GMP formation by these cells is dependent upon entry of calcium into the cell and demonstrate the complexity of the effects of calcium antagonists.     

Changes in Nucleic Acids in Phytochrome-dependent Elongation of the Alaska Pea Epicotyl

Red light, which produces the physiologically active form of phytochrome (Pfr), inhibited epicotyl elongation in intact dark-grown Alaska pea seedlings. This red light response was detectable 3 hours after the light treatment and became pronounced after 5 hours. The growth inhibition was completely reversed by far red light applied immediately after the red or by pretreatment of the seedlings with the plant hormone gibberellin A₃.     

Actinomycin D Inhibits the GA3-Induced Elongation of Azuki Bean Epicotyls and the Reorientation of Cortical Microtubules

Actinomycin D inhibited the elongation of epicotyl segmentsfrom azuki bean seedlings that was induced by simultaneous treatmentwith IAA and GA₃. The drug also inhibited the elongation ofthe segments that was caused by IAA alone when it was appliedtogether with IAA. When the segments were pretreated with GA₃and then incubated with IAA, GA₃ promoted the elongation causedby IAA and brought about a predominance of transverse corticalmicrotubules (MTs) in the epidermal cells of the segments. Thechange in the arrangement of MTs caused by pretreatment withGA₃ was evident 1 h after the start of subsequent incubationwith IAA when the effect of pretreatment with GA₃ on the elongationhad not yet become apparent. Pretreatment with GA₃ did not causeany change in the arrangement of MTs when GA₃-pretreated segmentswere not incubated subsequently with IAA. Although actinomycinD applied before treatment with IAA did not inhibit the IAA-inducedelongation, the drug diminished the promotion of the elongationcaused by pretreatment with GA₃ and prevented GA₃ from bringingabout a predominance of transverse MTs when the drug was appliedduring the pretreatment with GA₃. GA₃-induced synthesis of mRNA seems to be involved in the promotionby GA₃ of IAA-induced elongation and in the GA₃-induced rearrangementof cortical MTs. (Received June 15, 1993; Accepted August 16, 1993)