Features of Grana Organization in Pea Chloroplasts

Two fractions of membrane fragments—the pellets precipitated at 1300 and 20000 g (fractions G1.3 and G20, respectively)—were isolated from pea (Pisum sativum L.) chloroplasts after solubilization with digitonin. These fragments assigned to grana displayed the following differences: (1) in spectra of low-temperature fluorescence, the ratio of short-wave and long-wave band intensities, as well as integrated intensity of the whole spectrum, were higher for G1.3 than for G20 fraction; (2) in excitation spectra of long-wave fluorescence, the ratio of peaks at 650 and 680 nm and integrated intensity of the spectrum were higher for G1.3 than for G20 fraction; and (3) the shapes of fluorescence excitation spectra differed for G1.3 and G20. These results indicate that the two fractions examined differed in proportion of photosystem I and photosystem II complexes, as well as in organization of these complexes. The size of light-harvesting antenna was larger in PSI complexes of G1.3 fraction, owing, in particular, to a higher content of chlorophyll a/b-protein complexes in this fraction. After repeated digitonin fragmentation of G1.3 and G20 preparations, more than 80% of G1.3 fraction was decomposed into lighter fragments, whereas G20 fraction was resistant to fragmentation (it lost about 10% of its material). Analysis of the data suggests the presence of two structurally different types of thylakoids in grana. The yield of G20 fraction (about 20%) is comparable to the ratio between the number of intergranal thylakoids, connected to granum in pea chloroplasts, and the total number of thylakoids in this granum. Based on these data, we assume that G20 fraction represent the fragments of intergranal thylakoids that extend into the granum.__________Translated from Fiziologiya Rastenii, Vol. 52, No. 4, 2005, pp. 499–506.Original Russian Text Copyright © 2005 by Kochubei, Shevchenko, Bondarenko.     

Pigments and Gas Exchange Characteristics in Leaves of Chlorophyll Mutants of Pisum sativum

Quantitative and qualitative characteristics of pigment composition and gas exchange were studied in chlorophyll mutants of pea, Pisum sativum L.: chlorotica 2004 and 2014. The mutant 2004 had light-green color, whereas the mutant 2014 has yellow-green leaves and stems; they contained about 80 and 50% of chlorophyll, respectively, compared to the initial line. cv. Torsdag. Leaves of the mutant 2004 had significantly lower carotene content and accumulated more lutein and violaxanthin. In the mutant 2014, the contents of chlorophyll and all carotenoids were reduced almost proportionally. The quantum efficiency of photosynthesis was by 29–30% lower in the mutants, and it was 1.5–2 times higher in F₁ hybrids, as compared to control plants. Our data allow us to conclude that the impairment of photosynthesis in the mutant 2014 is caused by the changed mesostructure of leaves, whereas in the mutant 2004, it may be caused by an impairment of photosystem reaction centers.     

Fine Chloroplast Structure in Cucumber and Pea Leaves Developed under Red Light

Photosynthetic activity, the content of various photosynthetic pigments, and the chloroplast ultrastructure were examined in the leaves of cucumber (Cucumis sativus L.) and pea (Pisum sativum L.) plants of different ages grown under red light (600–700 nm, 100 W/m²). In pea leaves tolerant to red-light irradiation, chloroplast ultrastructure did not essentially change. In the first true leaves of cucumber plants susceptible to red-light irradiation, we observed a considerable increase in the number and size of plastoglobules, the appearance of chloroplasts lacking grana or containing only infrequent grana, and stromal thylakoids. In the upper leaves of 22-day-old cucumber plants, the chloroplast structure was essentially similar to that of the control chloroplasts in white light, and we therefore suppose that these plants have acclimated to red light.     

Chloroplast ultrastructure changes in Pisum sativum associated with supplementary ultraviolet (UV-B) radiation

Pea plants (Pisum sativum L. cv. Greenfeast) were grown and exposed to supplementary UV-B radiation from day 17 after planting under growth cabinet conditions. The effects of this exposure on the ultrastructure of chloroplasts and the total soluble sugar and starch concentrations were estimated. Supplementary UV-B radiation was shown to damage the structure of chloroplasts, as manifested by dilation of thylakoid membranes, a progressive disruption of the thylakoid structure and disintegration of the double membrane envelope surrounding the chloroplast, accompanied by the accumulation of large starch grains. Diurnal changes observed in starch concentration suggest that the higher concentration of starch in supplementary UV-B-treated leaves is due to its immobilization, rather than to any increase in starch synthesis: soluble sugars accumulated and remained at a higher level and then later declined.     

The Effect of Root Hypoxia and Iron Deficiency on the Photosynthesis, Biochemical Composition, and Structure of Pea Chloroplasts

The combined effect of root hypoxia and iron deficiency on biochemical composition, photosynthetic indices, and structure of pea (Pisum sativum L.) chloroplasts were investigated. Both factors suppressed chlorophyll accumulation and leaf photosynthetic activity, causing chlorosis. It was shown, that iron deficiency reduced more severe the light-harvesting complexes of photosystems (PS), and root hypoxia, the reaction center complexes of the photosystem I (PSI) and photosystem II (PSII). The combined action of both factors was stronger than the effect of each factor. However, even in yellow and almost white leaves, chloroplasts contained small amounts of all pigment–protein complexes and maintained weak photosynthetic activity, although their structure was poorly developed and comprised only vesicles and small thylakoids capable to form contacts and small grana. The conclusion is that the mechanisms of root hypoxia and iron deficiency destructive action are different and these factors differently and independently influenced leaf chloroplasts.     

菲白竹组培苗白化、绿化突变体的超微结构及15个叶绿体编码基因的表达

以菲白竹（Pleioblastus fortunei）组培苗绿化突变体及经60Coγ射线辐射获得的白化突变体为实验材料,从生长特性、超微结构及15个叶绿体编码基因的拷贝数及转录水平方面进行了研究。结果显示,白化突变体具有分化不定芽和不定根的能力,其叶肉细胞中叶绿体缺失或结构不完整,基因拷贝数与野生型无明显差异,部分基因转录水平低于野生型,psbA基因的转录水平高于野生型。绿化突变体叶色一致且能稳定遗传,与野生型绿色组织细胞的超微结构相比,其叶绿体基粒片层数量少,结构松散,基因拷贝数低或相当于野生型,多数叶绿体编码基因转录水平较野生型呈下调趋势,atpI基因转录水平高于野生型。psaA基因在两种突变体中均不表达,仅在野生型中有微弱表达。     

UV-B对设施桃叶片光合功能及叶绿体超微结构的影响

对温室栽培的油桃中油5号(Prunus persica var. nectarina cv.‘Zhongyou5’)适量补充UV-B,分析其对桃叶片光合功能及叶绿体超微结构的影响。结果表明,UV-B处理下各色素含量均有不同程度的增加,其中叶绿素b的含量和净光合速率(P_n)提升幅度较大。相较于未补充UV-B的桃树(对照), UV-B处理的F_v/F_m无显著变化, F_v’/F_m’比值、光化学猝灭系数(qP)、非光化学猝灭系数(qN)以及PSII实际光化学量子效率(Φ_PSII)均有显著或极显著升高。透射电镜结果显示,UV-B处理下叶绿体基质片层空隙小,堆叠紧密,叶绿体外膜边缘清晰。可见,温室内适量补充UV-B可快速改善叶片叶绿体的超微结构,提升叶绿素分子捕获光能及向PSII传递的能力,增大PSII反应中心的开放程度,提高实际光能转化效率和PSII电子传递量子效率,提高叶片的光合功能。该研究为设施果树光合性能改善和UV-B合理利用提供了理论依据。     

The Sym31Gene Responsible for Bacteroid Differentiation Is Involved in Nitrate-Dependent Nodule Formation in Pea Plants

The effects of exogenous nitrate on the number of developing nodules and their leghemoglobin content in the original pea (Pisum sativumL.) line and its symbiotic mutants were studied. Mutation in the Sym31gene conferred the tolerance to nitrate in the corresponding pea line and manifested itself as the number of nodules independent of the nitrate concentration. Thus, the Sym31gene was identified as the only known symbiotic gene involved in both the differentiation of symbiotic compartments and the nitrate-dependent process of nodule formation. The presence of leghemoglobin in double mutants (sym13, sym31) indicates the possibility of the complementary contribution of these genes in the control of leghemoglobin synthesis.     

Activation of Chloroplast Fructose-1, 6-Bisphosphatase of Pisum sativum by Mole Mass Change

Separation of extracts, obtained from isolated intact P. sativum chloroplasts, by fast protein liquid chromatography (FPLC) on superose 6, reveals a 1,400 kDa-FBPase II form at pH 6.0 and a 380 kDa form at pH 7.5. Addition of F1,6P₂, Mg⁺⁺ and ATP cause dissociation of the large form into the smaller one, which leads to an approximate 4-fold increase in activity. Reversibility of the mole mass change could be shown for the influence of pH and of fructose-1, 6-bisphosphate on purified enzyme samples, separated from crude leaf extracts. Compared to thelarge enzyme form, the small form has higher activity and is specific for the substrate fructose-1, 6-bisphosphate, while the large form is not. Activation of FBPase II in the light and inactivation in the dark is discussed on the basis of different oligomeric forms of the enzyme caused by changes in the concentration of intermediates and effectors in the chloroplast stroma. The conclusion is drawn that oligomerization of key enzymes might provide an effective mechanism for enzyme activation/inactivation in vivo.     

Characteristics of Photosystem I Complexes in the End Membranes of Pea Granal Thylakoids

Two fractions of the light fragments enriched in the photosystem I (PSI) complexes were obtained from pea (Pisum sativum L.) thylakoids by digitonin treatment and subsequent differential centrifugation. The ratio of chlorophyll a to chlorophyll b, chlorophyll/P700 spectra of low-temperature fluorescence, and excitation spectra of long-wave fluorescence were measured. These characteristics were shown to be different due to variation in the size and composition of the light-harvesting antenna of PSI complexes present in the particles obtained. The larger antenna size of one of the fractions was related to the incorporation of the pool of light-harvesting complex II (LHCII). A comparison with the data available allowed us to identify these particles as fragments of intergranal thylakoids and end membranes of granal thylakoids. The suggestion that an increase in the PSI light-harvesting antenna in intergranal thylakoids is related to the attachment of phosphorylated LHCII is discussed.     

Red-light-induced Changes in the Distribution of Xanthoxin in Pea Seedlings

The distribution of xanthoxin (Xan), was determined in light-grown, 20-d-old pea (Pisum sativum L. cv. Progress No. 9) seedlings. The cis,trans-xanthoxin (c,t-Xan) and the trans,trans-xanthoxin (t,t-Xan) were more abundant in the young leaves and terminal bud; their concentrations in leaves were 2 - 3 times those in internodes of the same nodes. After the onset of red-light-irradiation, the concentration of both Xan isomers in 7-d-old dark-grown pea seedlings increased after a 12-h lag time. The increased level of Xan was greatest in the terminal bud and decreased to lower parts of the seedlings. The ratio of c,t-Xan to t,t-Xan concentration in the seedlings was about 2:3.     

Organization of the Marginal Regions of Pea Granal Thylakoids

Thylakoids of pea chloroplasts isolated from plants grown during various time intervals from June to August were subjected to fragmentation. Using a modified procedure, a fraction of larger particles was separated from those previously considered as fragments of intergranal thylakoids. The particles of the fraction isolated were identified as fragments of marginal regions of granal thylakoids (margins). The relative yield of these fragments depended on the time interval of plant growth. Two types of low-temperature fluorescence spectra corresponding to a high and low yield of the fraction were detected. The characteristics of the first one were a high fluorescence intensity in the short-wave region and the presence of bands with maxima at 687 and 696 nm emitted by photosystem II (PSII). The ratio of PSII to PSI complexes (PSII/PSI) in the fractions characterized by a low and high yield varied from 1 to 5. The analysis of excitation spectra of long-wave fluorescence of PSI showed that PSI complexes in the margin fragments obtained at a low fraction yield were depleted in chlorophyll forms with a 682-nm absorption maximum and enriched in those with a 668-nm maximum. Since an increase in the yield of the margin-fragment fraction is due to an increased unstacking of granal thylakoids, the differences in the characteristics of fragments obtained with a low and a high yield reflect the changes in the composition of granal thylakoids in the direction from the margin to the centrum, that is, a decrease in the relative content of PSI complexes and alterations in the composition and size of its light-harvesting antenna. The consistency between the data obtained and the present view concerning the different functions of PSI located in different thylakoid regions is discussed.     

The State of Photosystem I Complexes in the Central Regions of Pea Granal Thylakoids

Grana-core and grana-margin fragments were obtained from pea (Pisum sativum L.) thylakoids, and both fractions contained photosystem I (PSI) complexes. The yield of these fractions exhibited variations for the plants grown during various periods of the summer season. Low-temperature fluorescence spectra, excitation spectra of long-wave fluorescence, and P700 kinetic characteristics were recorded for these fractions. PSI complexes in central granal regions were associated with PSII and the light-harvesting complexes of PSII, which followed from the excitation spectra of long-wave fluorescence and the kinetic characteristics of P700 light oxidation and dark reduction. The characteristics of the margin regions were changed depending on the fraction yield. If the yield was low, marginal fragments contained mainly PSI complexes. When the yield increased, PSI associates with PSII appeared. A spatial distribution and state of PSI complexes in granal thylakoids are discussed as related to the size and composition of the light-harvesting antenna.     

烟草NtFtsZ2-1基因参与叶绿体分裂和扩大的调节

叶绿体虽然是植物细胞内一种极其重要的细胞器,但其分裂的分子机制尚不很清楚。已经证明FtsZ蛋白作为真核细胞分裂装置的一个关键成分,参与叶绿体的分裂过程。烟草的FtsZ基因属于2个不同的家族,在对NtFtsZ1家族成员研究的基础上,用正义和反义表达技术研究了NtFtsZ2家族成员NtFtsZ2-1基因在转基因烟草中的功能。显微分析结果表明NtFtsZ2-1基因的表达水平异常增强或减弱都会严重干扰叶绿体的正常分裂过程,导致叶绿体在形态和数目上的异常（体积明显增大,数目显著减少）,而单个叶肉细胞中叶绿体的总表面积在正反义转基因烟草和野生型烟草之间保持了相对稳定,没有发生明显的变化。同时还证明NtFtsZ2-1基因表达的变化对叶绿素含量和叶绿体的光合作用能力没有直接的影响。据此我们认为NtFtsZ2-1基因参与叶绿体的分裂和体积的扩大,其表达水平的波动会改变植物中叶绿体的数目和大小,而且在叶绿体的数目与体积之间可能存在一种补偿机制,保证叶绿体能最大限度地吸收光能,从而使光合作用得以正常进行。     

菲白竹组培苗白化、绿化突变体的超微结构及15个叶绿体编码基因的表达

以菲白竹(Pleioblastus fortunei)组培苗绿化突变体及经⁶⁰Co γ射线辐射获得的白化突变体为实验材料, 从生长特性、超微结构及15个叶绿体编码基因的拷贝数及转录水平方面进行了研究。结果显示, 白化突变体具有分化不定芽和不定根的能力, 其叶肉细胞中叶绿体缺失或结构不完整, 基因拷贝数与野生型无明显差异, 部分基因转录水平低于野生型, psbA基因的转录水平高于野生型。绿化突变体叶色一致且能稳定遗传, 与野生型绿色组织细胞的超微结构相比, 其叶绿体基粒片层数量少, 结构松散, 基因拷贝数低或相当于野生型, 多数叶绿体编码基因转录水平较野生型呈下调趋势, atpI基因转录水平高于野生型。psaA基因在两种突变体中均不表达, 仅在野生型中有微弱表达。     

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。     

盐胁迫对转AtNHX1基因杨树光合特性与叶绿体超微结构的影响

以欧美107杨(Populus×euramericana ‘Neva',Wt)和转拟南芥液泡膜Na~+/H~+逆向转运蛋白基因AtNHX1的欧美107杨新品系(Tr) 幼苗为材料,研究了高低度盐胁迫对两品系幼苗光合色素含量、光合参数和叶绿体超微结构的影响,以阐明转AtNHX1基因杨树的耐盐性与其光合作用及叶绿体结构之间的关系.结果表明:(1)盐处理后,两品系叶片叶绿素含量、类胡萝卜素含量、净光合速率、蒸腾速率和气孔导度均下降,且高盐度处理下降幅度更大;同等盐度处理下,Tr品系叶片叶绿素含量、净光合速率和气孔导度的下降幅度显著低于Wt品系,且在高盐度处理间差异更大;两品系杨树叶片P_n下降的原因在低盐处理时以气孔限制为主,而在高盐下则是气孔限制和非气孔限制共同作用的结果.(2)盐胁迫对T_r 品系叶片叶绿体超微结构的影响较轻,其在高盐下仍保持了较好的内部结构;盐胁迫Wt品系叶绿体则缩皱成球形,内部结构趋向简单,以至解体,脂质球显著增多.可见,盐胁迫导致杨树叶绿体结构破坏而引起叶绿体色素含量下降,最终降低其光合作用效率;同等盐度胁迫下,转AtNHX1基因品系叶片保持了较完整的叶绿体超微结构、更高的叶绿素含量,能维持较好的光合状态,从而表现出较高的耐盐能力.     

The Involvement of Endogenous Phenolic Compounds in the Response of Pea Seedling Roots to Inoculation with Rhizobium leguminosarum at Various Temperatures

Endogenous phenolic compounds (PC) affecting Rhizobium leguminosarum bv. viceae propagation were isolated from the roots of etiolated pea (Pisum sativum L.) seedlings before and within one or two day after inoculation. It was established that, during the first day after inoculation, PC-induced stimulation of bacterial growth in roots was replaced by its inhibition, which was somewhat more pronounced at 8°C. The ratio between PC fractions was also changed during the first day after inoculation, especially strongly at low temperature; and this was evidently the cause for Rhizobium growth inhibition in root cells.     

毛竹茎秆叶绿体超微结构及其发射荧光光谱特征

为了探讨毛竹（Phyllostachys pubescens）茎秆的光合特性,以1龄和3龄毛竹为材料,观察了茎秆和叶中叶绿体的超微结构,测定了光合色素含量以及发射荧光光谱。结果表明：茎秆中叶绿体发育完整,其类囊体垛叠程度高于叶,并含有淀粉粒。茎秆中叶绿素总含量、类胡萝卜素及Chla／b含量显著低于叶（FI〈O．05）。茎秆发射荧光光谱在735nm处没有明显的主峰,1龄和3龄毛竹茎秆光系统lI与光系统I的半峰宽比值分别比叶降低了7．0％和11．3％（P〈0．05）,峰高比值比叶分别增加了6．5％和18．3％（P〈0．05）。四阶导数光谱在650—800nm波长范围内出现了6个极大值,代表LHCII、CP43、CP47、RCI和ILHCI的发射荧光峰以及PSI和PSII的发射荧光副振峰：其中,茎秆中RCI和LHCI特征发射荧光峰与叶相比有不同程度的红移。表明毛竹茎秆叶绿体通过提高Chlb的相对含量和增加类囊体垛叠以及降低LHCI含量,来适应毛竹茎秆以红光为主的光环境。进而协调激发能在2个光系统间的分配。     

271份豌豆种质资源农艺性状遗传多样性分析

以来自五大洲57个国家和地区的271份豌豆资源为材料,利用主成分分析和聚类分析的方法,评价其2个质量性状和7个数量性状的遗传变异水平。结果表明,参试资源的农艺性状具有丰富的遗传多样性。其中遗传多样性指数最高的是始荚节位(2.0590),其次是主茎节数(2.0421);性状变异系数最大的是小区产量(64.874%),其次是百粒重(61.870%)。采用SPSS 22.0软件对参试资源7个数量性状的主成分分析结果表明,前3个主成分因子累计贡献率达66.021%,Ward法聚类将参试的271份豌豆资源划分为4大类群,其中第Ⅱ类群属于高秆、大粒、高产种质,具有很高的丰产潜力,为我国杂交育种亲本选择提供了原材料。对取材数大于等于5的来自于四大洲17个国家的参试资源进一步分析发现,不同地理区域资源间遗传变异显著,其中印度的遗传多样性指数最高(1.7533),巴基斯坦的最低(0.9685),波兰的变异系数和遗传多样性都较高。综合分析国外种质资源的农艺性状,有助于我国豌豆育种项目亲本选配。