Effect of Benzyladenine on DNA, RNA, Protein, and Chlorophyll Contents in Intact Bean Leaves: Differential Responses to Benzyladenine According to Leaf Age

Benzyladenine (BA) was applied to intact bean (Phaseolus vulgaris) leaves at different stages of their growth. Changes in the amounts of cellular constituents resulting from the different treatments were followed and compared. RNA, protein, and chlorophyll contents, dry weight, fresh weight, and leaf area per single leaf continued increasing when leaves were treated with BA from an early stage, whereas in untreated leaves all these values levelled off or declined with advancing age. Besides these changes, BA treatment induced an increase in the DNA content. Changes in RNA content was more remarkable in response to application or deprival of BA treatment than the corresponding ones in protein and chlorophyll contents. The pattern of response to BA varied greatly according to the age at which the leaf received the treatment. As leaves aged, they lost the ability to increase their area and fresh weight in response to BA. However, continuous treatment with BA from an early stage kept the leaves young and able to respond.     

Effect of benzyladenine on RNA and protein synthesis in intact bean leaves at various stages of ageing

Primary leaves of intact bean plants ( Phaseolus vulgaris L.) were treated with benzyladenine (BA) at different stages of ageing, BA promoted the synthesis of RNA, and soluble and insoluble proteins. The effects of BA stimulation differed depending on the age at which the leaf received the hormone treatment. In leaves attached to the plant, BA appeared to stimulate the rate of synthesis more than the rate of decomposition of RNA and protein, resulting in a net increase in RNA and protein. Both chloroplast and cytoplasmic ribosomes were still observed in intact yellowish green leaves. Polysomes in the cytoplasm increased remarkably when BA treatment was begun at late stages.     

Regulation of the carotenoid content and chloroplast development by levulinic acid

Chloroplast development and chlorophyll biosynthesis are co-regulated. To understand the mechanism of regulation of chloroplast biogenesis by chlorophyll, development of the photosynthetic apparatus was monitored during greening of etiolated barley leaf discs in the presence of levulinic acid, an inhibitor of chlorophyll biosynthesis. Although not a direct inhibitor of carotenoid biosynthesis, treatment by levulinic acid resulted in a linear reduction in both chlorophyll and carotenoid contents. Chlorophyll biosynthesis appeared to control that of carotenes. In the presence of levulinic acid, photosystem II (PSII) activity decreased while photosystem I (PSI) activity increased when expressed on a chlorophyll basis. However, the activities of both photosystem I and II decreased when expressed on a per plastid basis. As expected, in the presence of low amounts of chlorophyll, the light-harvesting chlorophyll-protein complex II (LHCPII) was not visible in Coomassie-stained gels in 20 m M levulinic acidtreated tissues, but was detected as a faint band by immunoblotting. This small amount of the LHCPII induced significant amounts of grana stacking, which was monitored as an increase in the ratio of variable to maximum fluorescence. When levulinic acid was washed from the leaf discs and the latter allowed to green in its absence, the chlorophyll and carotenoid contents and the photosynthetic activities approached the control values. Levulinic acid could be used to arrest the light-induced chloroplast development at a desired phase of greening and removed by washing the leaves to restore the developmental process without any apparent toxic effect. Results demonstrate that biosynthesis of carotenes is regulated by that of chlorophylls and extremely low amounts of the LHCPII can induce grana stacking.     

Effect of Benzyladenine on Diurnal Changes in DNA Content per Chloroplast and Chloroplast Replication in Intact Bean Leaves

The effect of benzyladenine (BA) on the diurnal changes in DNAand Chl contents per chloroplast and chloroplast replicationin primary leaves of bean plants (Phaseolus vulgaris L.) grownunder a 16 h light/8 h dark cycle was studied. Experiments weremade on primary leaves in the early expansion phase, where celldivision had been completed but chloroplasts were replicating.In untreated controls, chloroplast number, Chl content and freshweight per leaf showed daily periodic changes. Chl content perchloroplast increased in the light period every day, and freshweight per leaf increased most rapidly in the early dark period.Chloroplast number per leaf increased rapidly in the early darkperiod on day 9, though the increase began a little earlierand was less sharp on days 8 and 10. During these periods, DNAcontent per chloroplast was decreasing due to chloroplast divisionas chloroplast DNA (ctDNA) per leaf remained unchanged throughoutthe experimental period. BA induced increases in Chi contentper chloroplast, ctDNA content and fresh weight per leaf within6 h of its application, regardless of whether it was appliedat or 10 h after the beginning of the light period. Applicationof BA at 10 h in the light period shifted the start of chloroplastreplication by 6 h compared to that in untreated controls. However,when BA was applied at the beginning of illumination, the startof chloroplast replication showed the same relative change intime as above. 5-Fluorodeoxyuridine (5-FdU) promptly preventedBA-induced increase in Chl content and chloroplast number perleaf as well as ctDNA content per leaf.     

Physiological, structural, and immunological characterization of leaf and chloroplast development in cotton

Summary Many of the studies of chloroplast ontogeny in higher plants have utilized suboptimal conditions of light and growth to assess development. In this study, we utilized structural, immunological, and physiological techniques to examine the development of the chloroplast in fieldgrown cotton (Gossypium hirsutum cv. MD 51 ne). Our youngest leaf sample developmentally was completely folded upon itself and about 0.5 cm in length; leaves of this same plastochron were followed for three weeks to the fully expanded leaf. The chloroplasts at the earliest stage monitored had almost all of the lamellae in small, relatively electron-opaque grana, with relatively few thylakoids which were not appressed on at least one surface. During the development of the thylakoids, the membranes increase in complexity, with considerable stroma lamellae development and an increase in the number of thylakoids per granum. Besides the increase in complexity, both the size and numbers of the chloroplast increase during the development of the leaf. Developmental changes in six thylakoid proteins, five stromal proteins, and one peroxisomal protein were monitored by quantitative immunocytochemistry. Even at the earliest stages of development, the plastids are equipped with the proteins required to carry out both light and dark reactions of photosynthesis. Several of the proteins follow three phases of accumulation: a relatively high density at early stages, a linear increase to keep step with chloroplast growth, and a final accumulation in the mature chloroplast. Photosystem-II(PS II)-related proteins are present at their highest densities early in development, with an accumulation of other parts of the photosynthetic apparatus at a latter stage. The early accumulation of PS-II-related proteins correlates with the much lower ratio of chlorophylla tob in the younger leaves and with the changes in fluorescence transients. These data indicate that some of the conclusions on chloroplast development based upon studies of intercalary meristems of monocots or the greening of etiolated plants may not be adequate to explain development of chloroplasts in leaves from apical meristems grown under natural conditions.Abbreviations CF1 chloroplast coupling factor 1 - chl chlorophyll - DAP days after planting - LHC light-harvesting chlorophyll-a/b-binding protein - OEC oxygen-evolving complex of photosystem II - PBS phosphate-buffered saline - PS photosystem - RuBisCo ribulose bisphosphate carboxylase/oxygenase     

Chlorophyll,carotenoid, and lipid content in Triticum sativum L. plastid envelopes,prolamellar bodies,stroma lamellae,and grana

The pigment and lipid content, expressed on a protein basis, is compared in wheat etioplast and chloroplast membrane fractions. Chloroplast envelopes contain less carotenoid and 1/3 more lipid than etioplast envelopes. The minute amount of chlorophyll and carotenoid found in chloroplast envelopes could be due to thylakoid contamination. Prolamellar bodies and grana have nearly the same amount of total lipid and total carotenoid per mg of protein although their respective compositions differ. On a protein basis, the lipid, chlorophyll, and carotenoid contents are lower (2.3, 10, and 20 times, respectively) in stroma lamellae than in grana membranes, but the latter contains a higher proportion of -carotene, chlorophyll a, and sulfolipid.This research represents partial fulfillment of the thesis Doctorat d'Etat ès Sciences requirements of the author     

金叶连翘叶片色素含量和解剖结构研究

金叶连翘不同冠层的成熟叶片呈现为不同颜色。以朝鲜连翘深绿色叶为对照,观察金叶连翘冠层上、中、下位叶色,测定其叶片大小和叶绿素a、叶绿素b、总叶绿素及类胡萝卜素含量,同时观察分析叶片横切面解剖结构,旨在阐明叶片色素含量和解剖结构对叶色的影响。研究表明:上层黄色、中层黄绿色、下层浅绿色,黄、黄绿、浅绿色叶总叶绿素含量分别是对照组的0.51%、4.44%和66.47%,均极显著低于对照(P <0.01),但黄绿叶的叶绿素a/b比值显著升高(P <0.05),黄、黄绿叶的总叶绿素/类胡萝卜素比值极显著降低(P <0.01)。黄、黄绿叶的叶绿体发育停滞于单片层时期,类囊体分化程度低,浅绿叶类囊基粒片层肿胀;黄叶细胞器降解,栅栏组织细胞形状难以辨别,黄绿叶上表皮细胞凸起。金叶连翘属于总叶绿素及叶绿素b合成减少型突变体,表现为叶绿素严重缺失,类胡萝卜素相对含量升高;其叶绿体发育停滞,类囊体结构异常,是金叶连翘叶片呈现不同颜色的主要因素,与其叶片解剖显微结构无关。     

日光温室光温因子对黄瓜叶绿体超微结构及其功能的影响

在日光温室内,研究了光温因子对黄瓜叶绿体超微结构及其功能的影响．结果表明,因季节之间光、温条件不同,日光温室黄瓜叶片显微结构和叶绿体超微结构有一定差异,1月份光照弱叶肉细胞较大,而5月份光照强叶绿体数较多．在该试验条件下,未发现叶片光合速率与叶绿体超微结构之间有直接或密切的相关性．在各生长季节其光合速率均为第4叶>初展叶>基部叶,与叶龄及各叶位的受光量有关．如果将不同叶位叶放在相同的光照下,则差异明显减少．黄瓜叶片的叶肉细胞、叶绿体和淀粉粒的大小以及叶绿体数、基粒数、基粒厚度、基粒片层数都随叶位的下降而呈增加趋势。不同品种、同品种不同生长时期的叶片显微结构和叶绿体超微结构及其功能也有一定的差异．限制日光温室冬季黄瓜光合作用的主要因素是光照弱、有效光照时数少,而在晴天温度的限制作用相对较小。阴天因光照弱而导致的室内低温则是限制黄瓜生长的关键因素．     

水杨酸和茉莉酸甲酯对丹参幼苗叶片显微结构、光合及非结构糖积累的影响

研究了水杨酸(SA)和茉莉酸甲酯(MeJA)处理对丹参(Salvia miltiorrhiza Bunge)幼苗叶片显微结构、叶片光合能力及幼苗中非结构糖积累的影响.结果显示:SA处理增加了丹参幼苗叶片气孔密度;叶肉细胞排列紧密、体积减小,叶肉细胞内叶绿体数目减少,但叶绿体体积增大,叶绿体基粒片层结构的数目增加;叶片中叶绿素a、b含量、叶气孔导度、蒸腾速率以及净光合速率均增加;同时,幼苗根中和叶片中酸性转化酶活性降低,幼苗地上部分蔗糖含量及可溶性糖总量显著高于对照.MeJA处理减少了叶片气孔密度,气孔发育畸形;叶肉细胞间隙增大,栅栏细胞层数减少,叶肉细胞内叶绿体数目减少,叶绿体体积减小,叶绿体基粒片层结构被破坏;叶片中叶绿素a及类胡萝卜素含量、叶片的净光合速率低于对照,叶气孔导度、蒸腾速率增强;同时,幼苗根中及叶中酸性转化酶活性增加,幼苗根中蔗糖含量及可溶性糖总量显著低于对照.可见,SA处理能促进植物叶片显微结构发育,增强叶片光合能力,抑制蔗糖降解并促进蔗糖积累;而MeJA处理则破坏了植物叶片显微结构,降低了叶片光合能力,促进了蔗糖降解并减少蔗糖积累.     

不同叶色水稻叶绿体密度及基粒结构的计算机图象分析

水稻叶绿体计算机图象分析表明,随着叶片色级的提高,叶绿体表面积密度、体积密度以及两者的比值都相应增加。深色稻叶基粒堆直径与高度、类囊体垛叠数与类囊体厚度、叶绿素与类胡萝卜素含量、气孔导度与净光合率均大于浅色叶片。深色叶片基粒堆密集,有些基粒类囊体出现沿叶绿体长轴方向排列整齐现象;浅色叶片基粒堆稀疏,其中较大的基粒类囊体与长轴呈倾斜排列。     

Benzyladenine-induced increase in DNA content per chloroplast in intact bean leaves

Benzyladenine (BA) treatment was found to induce chloroplast DNA (ctDNA) synthesis after it had stopped in primary leaves of light-grown intact bean plants (Phaseolus vulgaris L.). The leaves were treated with BA from 7 days after sowing. Chloroplasts were isolated and the ctDNA content per chloroplast was determined. Chloroplast division occurred until 13 days after sowing in untreated leaves. BA stimulated the division keeping the level of ctDNA content per chloroplast the same as that in the untreated controls. After the division period, the ctDNA content per chloroplast increased in BA-treated leaves, but not in controls. Consequently, ctDNA per leaf (or per cell) increased immediately after the beginning of BA treatment, but remained constant in the control leaves.     

遮荫对南美蟛蜞菊、蟛蜞菊及其自然杂交种叶片显微结构及叶绿体超微结构的影响

利用JSM-6360LV型扫描电镜和JEM-1010型透射电镜,观察了南美蟛蜞菊、蟛蜞菊及其自然杂交种新近成熟和老熟叶片的解剖结构及叶绿体超微结构。结果表明:遮荫后该杂交种与其亲本新近成熟叶片均表现为上下表皮气孔密度、叶片总厚度及上下表皮厚度、栅栏组织、海绵组织厚度减小,叶绿体肿胀变形,基粒片层垛叠程度增加,淀粉粒增多变大;遮荫后杂交种老熟叶片总厚度及上表皮、栅栏组织、海绵组织厚度增加,入侵种的下表皮厚度及本地种的上表皮厚度增加,叶绿体超微结构在遮荫后均出现严重损伤,基粒片层类囊体结构边缘溶解等。说明三种蟛蜞菊属物种及各物种不同叶龄叶片对弱光条件的响应存在差异;杂交种叶片显微及超微结构在不同光照下的变化介于亲本之间,对遮荫环境能较好适应。     

A Vertical Gradient of the Chloroplast Abundance among Leaves of Chenopodium album

The abundances of chloroplasts in leaves on the main stems ofChenopodium album at different height levels were investigatedin relation to the photosynthetic capacity and light environmentof the leaves. (1) The number of chloroplasts per mesophyllcell decreased with descending position of leaves, except foryoung developing leaves at the top of plants that had smallerchloroplast numbers per cell than matured leaves beneath them.Contents of chlorophyll and ribulose-1,5-bisphosphate carboxylase/oxygenaseper leaf area that were highest in the topmost young leavesand decreased with decreasing height level indicate that thereis a vertical gradient of chloroplast abundance per leaf areadecreasing from the top of the leaf canopy with depth. (2) Light-saturatingrate of photosynthetic oxygen evolution per leaf area of maturedleaves decreased more steeply with decreasing leaf positionthan the chloroplast number per cell. Gradients of chlorophylland the enzyme protein contents were also steeper than thatof the chloroplast number. Loss of photosynthesis in lower leavesis, therefore, ascribed partly to loss of whole chloroplastsand partly to reduced photosynthetic capacities of the remainingchloroplasts. (3) The chloroplast number per cell in newly expandedsecond leaves was comparable to those in leaves that have developedat later stages of the plant growth but decreased graduallyduring leaf senescence both in the dark and light. The formationof the vertical gradient of chloroplast abundance is, therefore,ascribed to loss of whole chloroplasts during senescence ofleaves. (4) Irradiance a leaf receives decreased sharply fromthe top of the canopy with depth. The physiological or ecophysiologicalsignificance of the vertical distribution of chloroplasts amongleaves was discussed taking light environments of leaves intoconsideration. (Received July 31, 1995; Accepted October 20, 1995)     

Some factors affecting the chloroplast replication in the mossPlagiomnium trichomanes

Summary Some factors affecting the chloroplast replication were studied using the leaf cells of the mossPlagiomnium trichomanes. There was a significant positive correlation between chloroplast number per cell and cell volume in leaves of any developmental stage. However, when the detached leaves were cultured on nutrient agar, it was observed that the chloroplast replication occurred without cell enlargement regardless of the developmental stage of leaves. This implies that cell enlargement is not an essential factor for the chloroplast replication, but one of the environmental factors affecting it. Light is essential for the chloroplast replication which response to the light intensity. In the dark, there was little increase in chloroplast number per cell. With a light intensity of 50 lux, the increase rate of chloroplast number per cell was about half of that with 3,000 lux. Day length also affected significantly the chloroplast replication.     

THE EFFECT OF LIGHT INTENSITY AND TEMPERATURE ON PLANT GROWTH AND CHLOROPLAST ULTRASTRUCTURE IN SOYBEAN

Soybean plants grown in controlled environment cabinets under light intensities of 220 w/m² or 90 w/m² (400–700 nm) and day to night temperatures of 27.5–22.5 C or 20.0–12.5 C in all combinations, exhibited differences in growth rate, leaf anatomy, chloroplast ultrastructure, and leaf starch, chlorophyll, and chloroplast lipid contents. Leaves grown under the lower light intensity at both temperatures had palisade mesophyll chloroplasts containing well-formed grana. The corresponding leaves developed under the higher light intensity had very rudimentary grana. Chloroplasts from high temperature and high light had grana consisting of two or three appressed thylakoids, while grana from the low temperature were confined to occasional thylakoid overlap. Spongy mesophyll chloroplasts were less sensitive to growth conditions. Transfer experiments showed that the ultrastructure of chloroplasts from mature leaves could be modified by changing the conditions, though the effect was less marked than when the leaf was growing.     

短期夜间低温后番茄光合作用及叶绿体超微结构的恢复效应

为明确短期夜间低温后番茄光合作用在常温下的恢复效应,研究了番茄幼苗经夜间15 ℃（对照）、12 ℃、9 ℃和6 ℃处理7 d后植株光合作用和叶绿体超微结构的变化.结果表明：短期夜间低温处理有利于恢复期番茄叶片的光合作用;12 ℃和9 ℃处理叶片叶绿素含量均可恢复至对照水平,6 ℃处理叶绿素含量则不可恢复;夜间低温处理并未严重抑制叶片气孔的形成,相反刺激了恢复期叶片气孔的形成与发育;6 ℃处理后叶绿体基粒片层排列较松散,片层数减少.恢复7 d时,夜间处理温度越低,叶绿体中淀粉粒越小、数量越少,基粒片层排列越整齐致密.另外,短期夜间低温处理促进了番茄植株叶片在恢复期的光合产物运输.     

Ultrastructure of Mesophyll Cells and Pigment Content in Senescing Leaves of Maize and Barley

Leaf senescence is a genetically regulated stage in the plant life cycle leading to death. Ultrastructural analysis of a particular region of the leaf and even of a particular mesophyll cell can give a clear picture of the time development of the process. In this study we found relations between changes in mesophyll cell ultrastructure and pigment concentration in every region of the leaf during leaf senescence in maize and barley. Our observations demonstrated that each mesophyll cell undergoes a similar senescence sequence of events: a) chromatin condensation, b) degradation of thylakoid membranes and an increase in the number of plastoglobules, c) damage to internal mitochondrial membrane and chloroplast destruction. Degradation of chloroplast structure is not fully correlated with changes in photosynthetic pigment content; chlorophyll and carotenoid content remained at a rather high level in the final stage of chloroplast destruction. We also compared the dynamics of leaf senescence between maize and barley. We showed that changes to the mesophyll cells do not occur at the same time in different parts of the leaf. The senescence damage begins at the base and moves to the top of the leaf. The dynamics of mesophyll cell senescence is different in leaves of both analyzed plant species; in the initial stages, the process was faster in barley whereas in the later stages the process occurred more quickly in maize. At the final stage, the oldest barley mesophyll cells were more damaged than maize cells of the same age.     

Protein reutilisation in corms of Colchicum autumnale

Colchicum autumnale L. is a monocotyledonous geophyte with hysteranthous leaves, i.e. flowering and leaf growth occur in different time periods. Because after the starch, the second prominent storage compound of corm is represented by proteins, we were interested in nitrogen remobilisation during the annual life cycle of C. autumnale. In this context the content of soluble and insoluble proteins were measured in parallel with determination of some exo-and endopeptidase activities. Our results indicate that the continual proteolysis occurs in both mother and new daughter corms during the whole life-cycle of the plant. L-Ala-aminopeptidase and trypsin-like endopeptidase were the most active peptidases in both mother and daughter corms. As the protein level of mother corm did not change significantly during the development of the future above-ground part under the soil surface (the first, autumnal developmental stage), the developmental profile of nitrate reductase activity was estimated followed by evaluation of total nitrogen and amino acid contents. Significant activity of root nitrate reductase was detected in the roots only in the second, vernal stage. Our results showed that the stored proteins constituted a relevant nitrogen source partly required by the growth processes of the late autumnal stage, but mainly by the intensive growth of leaves and reproductive structures during the second, photosynthetically active stage of the life-cycle.     

Effect of Salicylic Acid on Leaf Anatomy and Chloroplast Ultrastructure of Barley Plants

Light and electron microscopy were used to relate histological and ultrastructural differences of barley leaves treated with different concentrations of salicylic acid (SA, 100 µM-1 mM). Light microscopy revealed that the thickness of all leaf tissue components decreased in SA-treated plants. The effect was most pronounced on the width of the adaxial epidermis and on the size of the bulliform cells. The chloroplast ultrastructure was also affected by SA treatment. Swelling of grana thylakoids in various degrees, coagulation of the stroma, and increase in chloroplast volume were observed. 1 mM SA caused a vast destruction of the whole plastid structure.