Stimulation by Ethylene of Chlorophyll Biosynthesis in Dark-grown Cucumber Cotyledons

Five-day-old etiolated cucumber (Cucumis sativus L. var. Alpha Green) cotyledons produced more chlorophyll over a 4-hour illumination period after a prolonged exposure (12 to 72 hours) in the dark to ethylene concentrations ranging from 0.1 to 10 μl/l. Intact seedlings and excised cotyledons responded in the same way to this treatment. This effect does not involve a shortening of the lag phase of chlorophyll accumulation. Exposure of cotyledons to ethylene during the illumination period did not produce the same stimulatory effect on chlorophyll synthesis and, under certain conditions, chlorophyll synthesis was slightly inhibited by exposure to ethylene in the light.     

Effects of Compounds that Influence Calcium Absorption and of Calcium-Calmodulin Inhibitors on Zeatin Induced Growth and Chlorophyll Synthesis in Excised Cucumber Cotyledons

Previous results indicate that cytokinin action requires formationof Ca-calmodulin. We evaluated that hypothesis for cytokinin(zeatin) induced growth and chlorophyll formation in excisedcucumber (Cucumis sativus L.) cotyledons using several compoundsexpected to increase or decrease cytosolic levels of Ca²⁺ andthereby perturb formation of Ca-calmodulin. We also studiedeffects of eight drugs known to inhibit action of Ca-calmodulinin animals. Most of the results indicate no special requirementof Ca-calmodulin for cytokinin action in these organs. (Received November 24, 1988; Accepted April 24, 1989)     

低温光照下与黄瓜子叶叶绿素降低有关的酶促反应

黄瓜子叶在低温光照下处理时,随着时间的延长,Chl含量和CAT活性显著下降,POD和AsA-POD在开始24h活性亦有下降趋势,随后POD活性显著增加,而AsA-POD活性仅稍有提高。Chl含量的减少和CAT活性降低之间呈正相关,与POD活性的增加呈负相关。低温、光照处理的黄瓜子叶内Chl的降低可被亚胺环己酮(CHM)延缓,被AsA、Vit E、没食子酸丙酯(PG)和DCMU抑制,但被H_2O_2,乙醇酸钠和甲基紫精(MV)加速。     

Evidence that Ethylene is not Involved in Red-light-induced Stimulation of Chlorophyll Formation in Etiolated Cucumber Cotyledons

The involvement of ethylene in red-light-induced stimulationof chlorophyll (Chl) formation was studied because one of thered-light effects on Chl formation (the lateappearing effect)interacts with the ethylene effect in 3-day-old excised etiolatedcotyledons of cucumber (Cucumis sativus L. cv. Aonagajibai).Ethylene production by etiolated cotyledons of intact seedlingsin the dark is enhanced by a red-light pulse, but the effectdoes not occur in excised cotyledons. Application of ethylenein the dark to 3-day-old intact seedlings has little effecton Chl formation in the cotyledon during subsequent continuousillumination, although ethylene pretreatment of 5-day-old seedlingssignificantly stimulates Chl formation. Removal of endogenousethylene by mercuric perchlorate [Hg(ClO₄)₂] does not specificallysuppress the red-light action on Chl formation in both attachedand excised cotyledons. Inhibition of ethylene synthesis byaminoethoxyvinylglycine does not affect the red-light effecton Chl formation in excised cotyledons. These facts indicatethat ethylene does not operate as a mediator of red light instimulating Chi formation in either attached or excised cotyledons. (Received December 13, 1981; Accepted March 30, 1981)     

The Influence of Atrazine, Benzyladenine, and the Embryonic Axis on Chlorophyll Synthesis in Cucumber and Squash Cotyledons

Excision of the embryonic axis prior to 3 1/2 days of germination in the dark followed by 8-h of light decreased the total chlorophyll content of cucumber cotyledons but not squash cotyledons. Benzyladenine stimulated the accumulation of chlorophyll in the cotyledons of intact embryos and excised cotyledons in both cucumber and squash. Gibberellic acid had no effect. Atrazine inhibited chlorophyll formation in excised squash cotyledons. Benzyladenine also increased the carotenoid and xanthophyll content in the cotyledons from intact squash seedlings. The results suggest that pigment synthesis in cotyledons may be controlled by a number of substances produced in the embryonic axis and that cytokinin-like benzyladenine can simulate the action of one of them.     

Effects of Cytokinin and Light on Polyamines during the Greening Response of Cucumber Cotyledons

The cucumber cotyledon greening test was used as a model systemto study possible relationships between cytokinins and polyaminesduring the greening process. When cytokinin was applied to dark-growncotyledons, large increases in chlorophyll and putrescine levelswere observed. However, inhibition of putrescine biosynthesiswith D-arginine and difluoromethylarginine did not affect chlorophyllproduction, and applied polyamines proved inhibitory to greening.Addition of 50 mM K^$ to the cytokinin treatments increased chlorophyllsynthesis but caused a marked reduction in putrescine levels.These results indicate that the large increase in putrescinecontent that derives from cytokinin treatment of the cotyledonsis not essential for the cytokinin-induced greening response. ¹Present address: Crop Science Department, University of Guelph,Guelph, Ontario, Canada, NIG 2W1. ²Present address: Agrogen Biotechnologies Inc., 520W. 6th Ave.,Vancouver, B.C., Canada, V5Z 4H5. (Received June 29, 1987; Accepted October 9, 1987)     

A Comparative Study of Chlorophyll Production in Cotyledons

Chlorophyll production was studied in four species with cotyledonsdiffering widely in photosynthetic potential. When cotyledons were detached from dark-grown seedlings at intervalsduring germination and exposed to light for 24–48 h, maximumchlorophyll production was found after 8 days of dark-germinationin french bean, runner bean, and lupin, and after 5 days inmustard. Chlorophyll formation was reduced when the cotyledonswere detached earlier or later than this time, and during thefirst 1–2 days of germination the cotyledons producedvery little chlorophyll. With later times of excision the abilityto accumulate chlorophyll increased, rapidly in lupin and mustardand more slowly in the beans. A comparison of chlorophyll accumulation by attached cotyledonsand those excised during the first 24 h of germination showeda fourfold enhancement of chlorophyll content in detached french-beancotyledons and a fourfold reduction in chlorophyll content indetached mustard cotyledons, whilst chlorophyll accumulationwas similar in both the attached and detached cotyledons oflupin. The results are discussed in relation to the possible promotiveand inhibitory influence of the seedling axis on greening ofcotyledons differing in their potential for expansion and inthe amount of reserve material stored.     

The Effects of Cytokinins on Unfolding of Cotyledons and Opening of Hypocotylar Hooks of Cucumber

Benzyladenine (BA), zeatin and kinetin caused the unfolding of cucumber (Cucumis sativus L. Var. Jing-ye No. 4) cotyledons. But the other plant hormones e.g. gibberellic acid (GA3), indoleacetic acid(IAA), ethylene (ethrel), abscisic acid (ABA) and other related compound e.g. KCl, adenine were not so. This response was caused by even lower concentration (0.01 ppm). Therefore, it could be used as a bioassay for cytokinins. KCl affaeted synergistically on cytokinin-induced unfolding. Basally applied BA (10 ppm) stimulated the opening of the intact hypocotylar hook. The light affacted synergistically on BA-induced opening. However, basally applied BA prevented the opening of decapitated hooks, especially in the light. The cotyledons were found to play a part of effects in maintaining the hook.     

Inter-organ Control of Greening in Etiolated Cucumber Cotyledons

Inter-organ control of greening in etiolated cucumber (Cucumis sativus L. cv. Aonagajibae) cotyledons was investigated. Four- or six-day-old excised or intact etiolated cucumber cotyledons were illuminated under aerobic conditions. Excised cotyledons without hypocotyl hooks produced chlorophyll without a prolonged lag phase and the rate of chlorophyll formation was not depressed if they were illuminated immediately after excision. If the excised cotyledons were incubated in the dark before illumination, chlorophyll accumulation at the end of 6 h of continuous illumination was remarkably lowered as the dark period lengthened, especially in 6-day-old cotyledons. The rapid loss of chlorophyll-forming capacity of excised cotyledons during dark preincubation suggests a stimulatory effect of hypocotyls on the greening in cotyledons. The treatment of excised cotyledons with bleeding sap in the dark for 18 h resulted in the promotion of chlorophyll formation during subsequent continuous illumination. Partial fractionation of bleeding sap with organic solvents and paper chromatography indicates that the active substances showed the same behavior as cytokinins. These facts add weight to the hypothesis that cytokinins from roots flow into cotyledons and stimulate greening.     

低温与光对黄瓜幼苗子叶光合电子传递活性的影响

25～30℃和30 μmol m~(-2)s~(-1)光下培养的黄瓜幼苗,在黑暗下经 1～7℃处理24h或5℃处理24～72h,光合电子传递活性受不同程度的抑制;其抑制部位主要在PSⅡ氧化侧;随温度的降低和时间的延长,抑制部位可发展至PSⅡ及之后的电子递体上,但尚未影响PSⅠ的活性。160μmol m~(-2)s~(-1)的光强加重低温对电子传递活性的抑制,光强越高,则加重的程度越高;抑制部位从PSⅡ氧化侧发展至PSⅡ反应中心以及PSⅠ。     

Effect of Benzyladenine Treatment Duration on delta-Aminolevulinic Acid Accumulation in the Dark, Chlorophyll Lag Phase Abolition, and Long-Term Chlorophyll Production in Excised Cotyledons of Dark-Grown Cucumber Seedlings

Cotyledons excised from dark-grown cucumber (Cucumis sativus L. cv. Aonagajibai) seedlings were incubated in the dark with the cytokinin benzyladenine for different time periods. Then, various greening parameters were examined, including protochlorophyll(ide) to chlorophyll(ide) photoconversion and δ-aminolevulinic acid accumulations in the dark, both triggered by a 5-minute red-light pulse.     

激动素在黄瓜子叶器官分化中的作用

采用在培养初期分阶段供应激动素 (KT)的方法 ,研究了其对黄瓜子叶器官分化的影响 ,并对全程供应和不供应KT下子叶中内源多胺和内源激素的变化进行了动态测定。结果表明 :1)供应KT使不定根分化率下降 ,不影响营养芽的分化 ,而对于花芽分化则是必需的。2 )供应KT使多胺中腐胺 (Put)含量变化最为显著 ,在培养第 2、 6d出现峰值 ,在第 4d呈低谷 ,对照则在第 4d出现峰值。 3)供应KT使培养 2d的子叶中吲哚乙酸 (IAA)和脱落酸(ABA)含量显著降低 ,4d时IAA含量显著升高。据上结果 ,就这些生理变化和花芽分化之间是否存在相关进行了充分讨论。     

The Effects of Tentoxin on Chlorophyll Synthesis and Plastid Structure in Cucumber and Cabbage

To determine if chlorosis caused by tentoxin, a toxin produced by Alternaria tenuis Nees., is due to interference with chlorophyll synthesis directly or to disruption of normal chloroplast development, the effects of the toxin on these processes in cucumber (Cucumis sativus L.) and cabbage (Brassica oleracea L., var. capitata) were studied. Cucumber cotyledons are highly sensitive to the toxin but exhibited no interference with the conversion of protochlorophyll(ide) to chlorophyll(ide) or with the general time course pattern of chlorophyll synthesis, although there was a 90% reduction in chlorophyll concentration. In cabbage, which shows no chlorosis in the presence of the toxin, there was a slight stimulation of chlorophyll synthesis in the presence of the toxin. Electron microscopy revealed that in cucumber, toxin treatment interferes with development of prolamellar bodies and lamellae, and results in deformed plastids. No such effects were noted in toxin-treated cabbage tissues. Plastids in toxin-treated cotyledons of both cucumber and cabbage contained more starch than plastids in nontreated tissues. It was concluded that tentoxin acts through disruption of normal plastid development, rather than through direct interference with chlorophyll synthesis.     

The Yellowing of Attached and Detached Cucumber Cotyledons

Measurements of various constituents and the activities of variousenzymes have been made in attached and detached cotyledons ofCucumis sativus. The complex changes observed suggest that thereis a reproducible pattern. Comparison of results for attachedand detached material shows that, although both become yellow,the other biochemical changes associated with yellowing arenot the same.     

NCS对离体萝卜子叶转绿期间叶绿素荧光参数及类囊体膜蛋白复合物的影响

用PAM 2000型便携式荧光测定系统测定了不同浓度N arc iclas ine(NCS)处理的离体萝卜子叶的荧光动力学参数:初始荧光(F0)、光化学淬灭(qP)、光系统Ⅱ最大光化学效率(Fv/Fm)、光合电子传递速率(ETR).结果表明,NCS对各种叶绿素荧光参数均显示出明显的抑制作用,而且NCS的抑制作用随其浓度的增加而逐渐增强;利用蓝绿温和胶电泳技术分析萝卜子叶类囊体膜蛋白复合物发现,转绿期间叶绿体中光系统Ⅱ(PSⅡ)和光系统捕光色素蛋白复合体的含量增加均被NCS抑制.表明从水仙鳞茎分泌物中分离提取的生物活性物质NCS能够明显抑制离体萝卜子叶的光下转绿,而且NCS对离体萝卜子叶光下转绿的抑制是多位点的.     

Effects of Ammonium and Potassium Ions on Some Physiological and Biochemical Processes of Excised Cucumber Cotyledons

Incubation of excised cucumber cotyledons (Cucumis sativus L.) with NH₄Cl solutions exceeding 0.001 M inhibited their greening, fresh weight increases, and incorporation of ¹⁴C-leucine into insoluble N compounds. The absorption of ¹⁴C-leucine during incubation and retention of moisture by the excised cotyledons after incubation were greatly diminished by the NH₄Cl treatments. Treatment with KCl solutions of the same concentrations as the NH₄Cl stimulated the greening, fresh weight increases, and the absorption and incorporation of ¹⁴C-leucine. Desiccation of cotyledons stored at 5°C for 10 days was inhibited by dilute KCl solutions. The toxicity of NH₄Cl was alleviated by KCl solutions at low concentrations.     

Effects of Acifluorfen on Endogenous Antioxidants and Protective Enzymes in Cucumber (Cucumis sativus L.) Cotyledons

The herbicide acifluorfen (2-chloro-4-(trifluoromethyl)phenoxy-2-nitrobenzoate) causes strong photooxidative destruction of pigments and lipids in sensitive plant species. Antioxidants and oxygen radical scavengers slow the bleaching action of the herbicide. The effect of acifluorfen on glutathione and ascorbate levels in cucumber (Cucumis sativus L.) cotyledon discs was investigated to assess the relationship between herbicide activity and endogenous antioxidants. Acifluorfen decreased the levels of glutathione and ascorbate over 50% in discs exposed to less than 1.5 hours of white light (450 microeinsteins per square meter per second). Coincident increases in dehydroascorbate and glutathione disulfide were not observed. Acifluorfen also caused the rapid depletion of ascorbate in far-red light grown plants which were photosynthetically incompetent.

Glutathione reductase, dehydroascorbate reductase, superoxide dismutase, ascorbate oxidase, ascorbate free radical reductase, peroxidase, and catalase activities rapidly decreased in acifluorfen-treated tissue exposed to white light. None of the enzymes were inhibited in vitro by the herbicide. Acifluorfen causes irreversible photooxidative destruction of plant tissue, in part, by depleting endogenous antioxidants and inhibiting the activities of protective enzymes.

     

Inhibition of Pigment Biosynthesis in Cucumber Cotyledons by Isoxaflutole

Isoxaflutole [5-cyclopropyl-4-(2-methylsulphonyl-4-trifluromethylbenzoyl)isoxazole] is a new preemergence herbicide for broad-spectrum weed control in maize. The effect of isoxaflutole on chlorophyll (Chl) and carotenoid (Car) biosynthesis was investigated in cucumber (Cucumis sativus L.) cotyledons. Etiolated tissue was incubated with 5 mM isoxaflutole for 24 h and irradiated (60 mol m^-2 s^-1). The irradiation for 3 h did not reduce Chl a, Chl b, and Car contents, but after a 28-h irradiation the contents of Chl a and Car decreased by 35 and 15 %, respectively, and the content of Chl b increased by 24 %. Increasing the concentration of isoxaflutole beyond 5 mM resulted in reduction of Chl a (71 %), Chl b (20 %), and Car (31 %) contents. Similarly, increase in irradiance from 60 to 180 mol m^-2 s^-1 resulted in larger reduction of Chl and Car contents. Exogenously supplied 5-aminolevulinic acid did not reverse the isoxaflutole-inhibited Chl synthesis, whereas an exogenously supplied homogentisic acid lactone reversed the inhibition of pigment synthesis due to isoxaflutole.