Influences of Ce on the Formation of Chlorophyll-Protein Complexes in Chloroplasts of Cucumber Leaves

It has been proven that the Ce content of cucumber (Cucumis sativus L. ) leaves was enhanced with the increase of CeC13 concentration in Hoagland solution. The Chl a/b ratio of cucumber leaves in the control was the same as that in the treated plant, both being 3.67. However, under lower light intensity, the Chl a/b ratio in leaves of the contral was 2.72 whereas that of the treated leaves was 2.86. It showed that only under lower light intensity Ce could decrease the contents of chlorophyll b in leaves. The authors also evidenced that Ce was able to accelerate the formation of chlorophyll-protein complexes of PS Ⅰ and 110 kD polypeptide and decrease the light harvesting complex protein and 27 kD polypeptide.     

Removal of magnesium by Mg-dechelatase is a major step in the chlorophyll-degrading pathway in Ginkgo biloba in the process of autumnal tints

Autumnal tints are one of the most manifest and fascinating natural phenomena, but the mechanism of chlorophyll (Chl)-breakdown in deciduous trees has not been fully elucidated. In this study, we analyzed the composition of Chl-related compounds and determined the activities of initial Chl-degrading enzymes in Ginkgo leaves at various stages in the process of autumnal coloring. Only pheophytin a (Pheo a, Mg-free Chl a) was detected in yellow leaves by HPLC analysis, and the activity of Mg-dechelatase in yellow leaves was found to be higher than in green leaves. These findings showed that the removal of magnesium from Chl a occurred in advance of dephytylation in the Ginkgo.     

Molecular cloning and function analysis of the stay green gene in rice

Chloroplasts undergo drastic morphological and physiological changes during senescence with a visible symptom of chlorophyll (Chl) degradation. A stay green mutant was identified and then isolated from the japonica rice (Oryza sativa) cv. Huazhiwu by gamma-ray irradiation. The stay green mutant was characterized by Chl retention, stable Chl-protein complexes, and stable thylakoid membrane structures, but lost its photosynthetic competence during senescence. The gene, designated Stay Green Rice (SGR), was cloned by a positional cloning strategy encoding an ancient protein containing a putative chloroplast transit peptide. SGR protein was found in both soluble and thylakoid membranes in rice. SGR, like the gene for pheophorbide a oxygenase (PaO), was constitutively expressed, but was upregulated by dark-induced senescence in rice leaves. Senescence-induced expression of SGR and PaO was enhanced by ABA, but inhibited by cytokinin. Overexpression of SGR reduced the number of lamellae in the grana thylakoids and reduced the Chl content of normally growing leaves. This indicates that upregulation of SGR increases Chl breakdown during senescence in rice. A small quantity of chlorophyllide a accumulated in sgr leaves, but this also accumulated in wild-type rice leaves during senescence. Some pheophorbide a was detected in sgr leaves in the dark. According to these observations, we propose that SGR may be involved in regulating or taking part in the activity of PaO, and then may influence Chl breakdown and degradation of pigment-protein complex.     

Photosynthetic activities in the petunia corolla

Pink Petuniahybrida (cv Hit Parade Rosa) corollas were found to contain photosynthetically active chloroplasts. The corolla chloroplasts were similar to those of green leaves in size and structure. The chlorophyll (Chl) content of Petunia corollas increased during early stages of flower development, reaching a maximum just before anthesis. Chloroplasts isolated from corollas at this stage, carried out photosystem I-dependent electron transport at rates which were two-thirds of those measured in chloroplasts from green leaves, but full chain electron transport at only one-quarter of the rate carried out by chloroplasts from green leaves. Both the light saturated rate and the quantum yield for electron transport were lower in corolla chloroplasts, which also required lower intensities for light saturation. Reduced efficiency of photosystem II photoreactions in the corolla was also indicated by the ratio between variable and constant components of Chl fluorescence, which was lower in corollas compared to green leaves. The induction time of Chl fluorescence was at least three times shorter in corollas compared to green leaves, indicating a smaller number of functional photosystem II centers (per Chl) in the corolla. It is suggested that corolla chloroplasts of Petunia might have a role in flower developmental processes.     

Chlorophyll b to chlorophyll a conversion precedes chlorophyll degradation in Hordeum vulgare L.

This study reveals by in vivo deuterium labeling that in higher plants chlorophyll (Chl) b is converted to Chl a before degradation. For this purpose, de-greening of excised green primary leaves of barley (Hordeum vulgare) was induced by permanent darkness in the presence of heavy water (80 atom % (2)H). The resulting Chl a catabolite in the plant extract was subjected to chemical degradation by chromic acid. 3-(2-Hydroxyethyl)-4-methyl-maleimide, the key fragment that originates from the Chl catabolite, was isolated. High resolution (1)H-, (2)H-NMR and mass spectroscopy unequivocally demonstrates that a fraction of this maleimide fragment consists of a mono-deuterated methyl group. These results suggest that Chl b is converted into Chl a before degradation. Quantification proves that the initial ratio of Chl a:Chl b in the green plant is preserved to about 60-70% in the catabolite composition isolated from yellowing leaves. The incorporation of only one deuterium atom indicates the involvement of two distinguishable redox enzymes during the conversion.     

Characterization of a zebra mutant of rice with increased susceptibility to light stress

The rice zebra mutant TCM248 is a single recessive mutant. This mutant develops transverse-striped leaves with green and white sectors under alternate light/dark growth conditions. Mutants that were grown under a higher light intensity during the light period showed a more intense striped phenotype. The white tissues contained abnormal chloroplasts with few internal membrane structures, while the green tissues in the mutants contained normal chloroplasts. The white tissue contained only trace amounts of Chls and carotenoids, and mRNA accumulation of nuclear genes encoding chloroplast proteins (rbcS, cab) was strongly suppressed compared to that in the wild type plants. A series of growth condition shift experiments demonstrated that the mutant displayed the striped phenotype only if it was exposed to the alternate light/dark growth conditions during a limited stage of early leaf development. These data suggest that the zebra gene is involved in the acquisition of photoprotective capacity of the plants and that this gene functions at an early stage of chloroplast differentiation.     

Effect of BA upon the Amount of Pchlide and NPR protein and the Activity of NPR in Cucumber Cotyledons

When cucumber seedlings (Cucumis sativus L. cv. Aonagajibai) were treated with aqueous solution of BA (10–5mol/1 and 10–8mol/l) by spraying application, the amount of NPR protein was increased and the NPR activity was promoted. The contents of protochlorophyl, lide (Pchlide) and NADPH as the substrates of the enzyme reaction, were also increased remarkably. 2. The amount of Pchlide and NPR protein decreased rapidly when treated with red light. BA plays the role in the protection against the damage of enzyme activity by light and then decreasing the destruction of NPR protein. BA increased the content of Pchlide and NADPH, so the NPR activity was maintained at a higher level although in light condition. 3. Because that BA increased the content of NPR protein and the reaction substrate such as NADPH and Pchlide, the NPR activity was promoted. So, the rate of chlorophyll biosynthesis was rapid, and the content of chlorophyll increased. The leaf turns to dark green.     

Structure-function correlation during the etioplast-chloroplast transition in cucumber cotyledonary leaves

We studied the development of chloroplasts from etioplasts in the cotyledonary leaves of 4-d-old dark-grown cucumber (Cucumis sativus) seedlings after irradiation (20 μmol m^-2 s^-1). Upon irradiation, the triggering of chlorophyll (Chl) synthesis and accumulation showed a relatively short lag phase. The irradiation of etiolated seedlings initiated the synthesis of apoproteins of pigment-protein complexes. While Chl-protein 2 (CP2) was detected at 6 h after irradiation, CP1 only after 29 h. The appearance and accumulation of some of the apoproteins were monitored by Western-blotting. LHC2 apoprotein was detected after a 6 h-irradiation. The amounts of D1 protein of photosystem (PS) 2 and PsaA/B protein of PS1 were quantitated by ELISA. Further, the thylakoid membrane function during this time period in terms of PS1- and PS2-mediated electron transfer activity and intersystem electron pool size were analyzed. While PS1 activity was detected after 4 h, PS2-mediated O₂ evolution was detected only after a 17 h-irradiation. F_v/F_m value of Chl a fluorescence measurements indicated that the photochemical efficiency of these leaves reached its maximum after 29 h of irradiation. The intersystem pool size of cotyledonary leaves was equivalent to that of the control cotyledonary leaves grown for 25 h under continuous irradiation. Thus etioplasts develop into fully functional chloroplasts after approximately 25 h when 4 d-dark grown cucumber seedlings are continuously moderately irradiated. The development of photosynthetic electron transport chain seems to be limited in time at the level of PS2, possibly at the donor side. This revised version was published online in September 2006 with corrections to the Cover Date.     

Correlated Changes in the Activity,Amount of Protein,and Abundance of Transcript of NADPH:Protochlorophyllide Oxidoreductase and Chlorophyll Accumulation during Greening of Cucumber Cotyledons

Changes in the activity and abundance of NADPH:protochlorophyllide oxidoreductase (NPR) and the abundance of mRNA encoding it were examined during the greening of 5-d-old etiolated cucumber cotyledons under continuous illumination. To measure NPR activity in the extracts from fully greened tissues, we have developed an improved method of assay. Upon exposure of etiolated cotyledons to light, NPR activity decreased rapidly within the first 2 h of exposure. Thereafter, enzymatic activity increased transiently, reaching a submaximum level at 12 h, and decreased slowly. The level of immunodetectable NPR protein followed the same pattern of changes during 96 h of greening as observed for NPR activity. The NPR mRNA in etiolated cotyledons disappeared quickly in the 1st h of irradiation. However, the level of mRNA increased thereafter to reach 3-fold or more of the dark level at 12 h and then decreased. The changes in the activity, protein level, and mRNA level after the first rapid decreases corresponded chronologically and nearly paralleled the increase in the rate of chlorophyll accumulation. These findings suggest that the greening of cucumber cotyledons is regulated basically by the level of NPR protein without activation or repression of enzymatic activity and that NPR mRNA increased by light maintains the level of enzyme protein necessary for greening.     

竹柏2种颜色叶片的光合特性研究

为了解竹柏(Podocarpus nagi)的光合特性,以3 a生全绿叶和花叶竹柏为材料,测定其光合色素含量和气体交换参数。结果表明,全绿叶竹柏叶片的叶绿素a、叶绿素b、类胡萝卜素、叶绿素a+b、叶绿素a/b和叶绿素a+b/类胡萝卜素均显著高于花叶竹柏;全绿叶竹柏叶片的初始量子效率、最大光合速率和暗呼吸速率均显著高于花叶,而光饱和点和光补偿点均显著低于花叶;全绿叶竹柏叶片的初始羧化效率、光合速率、CO2饱和点和光呼吸速率均高于花叶,而CO2补偿点低于花叶。2种颜色叶片的气孔导度、蒸腾速率和水分利用效率均随着光合有效辐射的增大而增大,且均表现为全绿叶花叶,而胞间CO2浓度则相反,表现为花叶全绿叶。因此,全绿叶竹柏利用弱光的能力强于花叶竹柏,而花叶竹柏利用强光的能力更强,在园林绿化配置中,可根据2种颜色叶片的光合特性合理配置。     

Effect of Different Light Quality on Photosynthetic Characteristics of Cucumber Leaves

Cucumber (Cucumis sativus L. cultivar "Changchun Mici") seedlings were cultured in Hoagland solution under irradiation with different light spectra (8 h per day) for 20 days. The red light (λmax 658 nm, λ1/2 25 nm), blue light (λmax 450 nm, λ1/2 43 nm) and white fluorescent light possessed the same fluent rate (20 μmol· m-2·s-1 ). The experimental results showed that chlorophyll content of the leaves grown under white light was 7 % and 22.4% higher than those in red and blue light, respectively. Compared with white and blue light, red light induced a lower Chl a/b ratio and a higher level of Chl b in the cucumber leaves. Measurements of the low temperature (77 K) fluorescence emission spectra and kinetics of Chl a fluorescence induction of the leaves proved that the leaves grown under red light expressed the highest PSⅡ and the lowest PSⅠactivities while the leaves under blue light had the lowest PSⅡand the highest PSⅠ activities. The O2 evolution rate of red light-grown leaves was 44.9% higher than that of the white light-grown leaves, while blue light effect was similar to that of white in respect of O2 evolution. It is concluded that light quality is an important factor in regulating the development and activities of PSⅡ and PSⅡand the O2 evolution of photosynthesis in cucumber leaves.     

建兰叶艺品种光合色素含量及叶绿素荧光特性分析

以建兰栽培品种‘八宝奇珍’Cymbidium ensifolium ‘Ba Bao Qi Zhen’为材料,对其正常绿色叶片及黄化变异叶片的光合色素含量、叶绿素荧光参数进行比较。结果表明,‘八宝奇珍’黄化变异叶片的叶绿素总量、叶绿素a 和叶绿素b 含量均显著低于正常绿色叶片,且随着黄化面积的增大呈现递减趋势;黄化变异叶片的初始荧光量(Fo)、最大荧光产量(Fm)、Kautsky 诱导效应最大荧光(Fp)、稳态光适应光化学淬灭系数(qP)以及光适应稳态荧光产量(Ft_Lss)均显著低于正常绿色叶片;PSⅡ原初光能转化效率(Fv/Fm)与正常绿色叶片无明显差异;稳态非光化学荧光淬灭系数(NPQ)高于正常绿色叶片。     

Enhanced antioxidant protection at the early stages of leaf expansion in ginkgo under natural environmental conditions

Photosynthetic pigments, gas exchange, chlorophyll (Chl) a fluorescence kinetics, antioxidant enzymes and chloroplast ultrastructure were investigated in ginkgo (Ginkgo biloba L.) leaves from emergence to full size. Under natural conditions, the net photosynthetic rate (P_N), contents of Chl a, Chl b and total soluble proteins and fresh and dry leaf mass gradually increased during leaf expansion. The maximum photochemical efficiency of photosystem (PS) 2 (variable to maximum fluorescence ratio, F_v/F_m) was considerably higher at the early stages of leaf development than in fully expanded leaves. During daily course, only reversible decrease in F_v/F_m was distinguished at various stages, implying that no photo-damage occurred. Absorption flux per cross section (CS) and trapped energy flux per CS were significantly lower in newly expanding leaves compared with fully expanded ones, however, dissipated energy flux per CS was only slightly lower in expanding leaves. The ratio of carotenoids (Car)/Chl decreased gradually during leaf expansion due to increasing Chl content. Moreover, activities of the antioxidant enzymes, such as superoxide dismutase, ascorbate peroxidase, catalase and peroxidase, increased at the early stages of leaf expansion. The appearance of osmiophilic granules in fully expanded leaves further proves that photo-protection is significantly strengthened at the early stages of leaf expansion.     

叶角、光呼吸和热耗散协同作用减轻大豆幼叶光抑制

研究了大豆叶片逐步展开过程中的色素组成、气体交换、荧光动力学以及叶片角度等特性。随着叶片展开程度的增加 ,叶绿素含量和叶绿素 a/ b比值增加 ;光合速率 (Pn)也增加 ,揭示叶片展开过程中光合机构是逐步完善的。自然状态下 ,不同展开程度的叶片均未发生明显的光抑制 ;但将叶片平展并暴露在 12 0 0μmol/ (m2 · s)光下时幼叶发生严重的光抑制 ,伴随叶面积的增加光抑制程度减轻。强光下 ,尽管幼叶光呼吸 (Pr)的测定值较低 ,但幼叶光呼吸与总光合之比 (Pr/ Pm)较高。将叶片平展置于强光下时 ,幼叶的实际光化学效率 (ΦPSII)明显下调 ,非光化学猝灭 (NPQ)大幅增加 ;幼叶叶黄素库较大 ,光下积累较多的脱环氧化组分 ,揭示幼叶依赖叶黄素循环的热耗散增强。自然条件下测量叶片角度 ,观察到在叶片展开过程中叶柄夹角逐渐增加 ;日动态过程中幼叶的悬挂角随光强增加而明显减小 ,完全展开叶的悬挂角变化幅度很小。叶片角度的变化使实际照射到幼叶叶表的光强减少。推测较强的光呼吸、依赖叶黄素循环的热耗散以及较大的叶角变化可能是自然状态下幼叶未发生严重光抑制的原因     

Regeneration of protochlorophyllide in green and greening leaves of plants with varying proportions of protochlorophyllide forms in darkness

During illumination of dark-grown plants protochlorophyllide (Pchlide) is continuously transformed to chlorophyllide (Chlide). Different dark-grown plants, maize ( Zea mays cv. Sundance), wheat ( Triticum aestivum cv. Kosack), pea ( Pisum sativum cv. Kelwedon wonder), the lip1 mutant of pea, and the aurea mutant of tomato ( Solanum lycopersicum ), have various ratios of spectral Pchlide forms in darkness. When the plants were illuminated and then returned to darkness Pchlide re-accumulated. The proportions of different Pchlide forms within the pool of re-accumulated Pchlide were followed by low temperature fluorescence emission and excitation spectra in green and greening leaves. After 1 h of illumination the spectral characteristics of regenerated Pchlide forms mirrored those of Pchlide in dark-grown plants and were thus species dependent. After a prolonged illumination period (24 h) as well as in fully green leaves energy transfer to chlorophyll (Chl) masked the presence of long-wavelength Pchlide in the fluorescence emission spectra. However, excitation spectra showed Pchlide absorption around 650 nm and its flash-induced disappearance confirmed its nature of phototransformable Pchlide. In fact the excitation spectra showed that the proportions of different Pchlide forms in green leaves highly resembled the proportions of Pchlide forms in dark-grown leaves and were specific for the plant variety. Thus Chl formation in both dark-grown and light-grown leaves can occur in a similar way through the main photoactive long-wavelength form of Pchlide.     

Role of chlorophyllase in chlorophyll homeostasis and post-harvest breakdown in Piper betle L. leaf

Piper betle L., a dioecious shade-loving perennial climber is one of the important Pan-Asiatic plants. More than hundred landraces having marked variation in leaf chlorophyll (Chl) content are in cultivation in India. In this study, role of chlorophyllase (Chlase) in Chl homeostasis and post-harvest breakdown was investigated in two contrasting P. betle landraces Kapoori Vellaikodi (KV) with light green and Khasi Shillong (KS) with dark green leaves. The two landraces showed negative correlation between Chl content and Chlase activity in fresh as well as stored leaves. Accumulation of chlorophyllide a (Chlid a) was correlated with the level of Chlase activity, which was higher in KV than KS. The overall response of abscisic acid (ABA) and benzylaminopurine (BAP) was similar in KV and KS, however, the time-course was different. ABA-induced Chl loss was accompanied by rise in Chlase activity in KV and KS and the delay in Chl loss by BAP was accompanied by reduction in Chlase activity. While there were significant differences in Chlase activity in KV and KS, only minor differences were observed in the enzyme properties like pH and temperature optima, Km and Vmax. No landrace-related differences were observed on the effect of metal ions and functional group reagents/amino acid effectors on Chlase activity. These results showed that despite significant differences in Chl content and Chlase activity between landraces KV and KS, the properties of Chlase were similar. The findings show that in P. betle Chlase is involved in Chl homeostasis and also in Chl degradation during post-harvest storage and responds to hormonal regulations. These findings might be useful in predicting the stability of Chl during post-harvest storage and also the shelf-life in other P. betle landraces.     

Structural mechanism and photoprotective function of water-soluble chlorophyll-binding protein

A water-soluble chlorophyll-binding protein (WSCP) is the single known instance of a putative chlorophyll (Chl) carrier in green plants. Recently the photoprotective function of WSCP has been demonstrated by EPR measurements; the light-induced singlet-oxygen formation of Chl in the WSCP tetramer is about four times lower than that of unbound Chl. This paper describes the crystal structure of the WSCP-Chl complex purified from leaves of Lepidium virginicum (Virginia pepperweed) to clarify the mechanism of its photoprotective function. The WSCP-Chl complex is a homotetramer comprising four protein chains of 180 amino acids and four Chl molecules. At the center of the complex one hydrophobic cavity is formed in which all of the four Chl molecules are tightly packed and isolated from bulk solvent. With reference to the novel Chl-binding mode, we propose that the photoprotection mechanism may be based on the inhibition of physical contact between the Chl molecules and molecular oxygen.