水稻温敏型突变体叶片间断失绿的超微结构

在短时降温诱导下,水稻温敏型突变体１１０３ｓ（Ｏｒｙｚａ　ｓａｔｉｖａ　ｓｓｐ．ｉｎｄｉｃａ）植株间断失绿性状表达（临界温度２３．１℃）过程中,叶绿体含量的增减与叶色变化相符。电镜观察发现,性状表达时叶片间断失绿区叶绿体内部结构发生退化,呈现基粒垛叠片层数的异常减少,或基粒消失仅剩基粒残迹,有的甚至整个叶绿体为高电子密度的囊泡状结构。但在同一叶片的绿区,叶绿体仅表现基粒片层数减少、排列不规则,嗜锇小球聚集。在叶片失绿区的复绿过程中,叶绿体的这些变化又可逆转,内部结构重建,最后整个叶绿体结构基本恢复正常。水稻温敏突变体１１０３ｓ叶片间断失绿性状表达过程,实质上是一个由温度调控的叶绿体结构退化与修复的可逆过程。     

水稻温敏失绿突变体的Rubisco活化酶活性、蛋白质与氨基酸组分的变化

在水稻温敏失绿突变性状表达过程中,对其Rutbsico含量、Rubsico活化酶活性,全叶蛋白及游离氨基酸组分变化进行测定。结果表明：突变体的Rubisco结构和含量与野生型一样,保持相对稳定;而其Rubisco活化酶活性则随一个分子量为56．2kD(PI=4．5)的特异蛋白质的存在与消失发生明显改变。当突变性状表达时,分子量为56.2kD(PI=4．5)的特异蛋白消失,其Rubisco活化酶活性下降;当叶片失绿区域复绿时,56.2kD(PI=4．5)特异蛋白出现,则Rubisco活化酶活性上升。这一密切地相关关系表明,突变体的Rubisco活化酶活性变化在光合作用过程中,除与自身结构和含量有关外,还与叶片中这一特异蛋白的存在密切相关,它可能是Rubisco活化酶活性的调节蛋白。这种调节具体表现在氨基酸代谢上,是对上游氨基酸的阻遏调控,从而使叶绿体的结构物质合成受阻,最终导致类囊体膜的退化。     

水稻温敏失绿突变体的Rubisco活化酶活性、蛋白质与氨基酸组分的变化

在水稻温敏失绿突变性状表达过程中,对其Rubsico 含量、Rubsico 活化酶活性,全叶蛋白及游离氨基酸组分变化进行测定。结果表明:突变体的Rubisco 结构和含量与野生型一样,保持相对稳定;而其Rubisco 活化酶活性则随一个分子量为56.2kD(PI=4.5)的特异蛋白质的存在与消失发生明显改变。当突变性状表达时,分子量为56.2kD(PT=4.5)的特异蛋白消失,其Rubisco 活化酶活性下降;当叶片失绿区域复绿时,56.2kD(PI=4.5)特异蛋白出现,则Rubisco 活化酶活性上升。这一密切地相关关系表明,突变体的Rubisco 活化酶活性变化在光合作用过程中,除与自身结构和含量有关外,还与叶片中这一特异蛋白的存在密切相关,它可能是Rubisco 活化酶活性的调节蛋白。这种调节具体表现在氨基酸代谢上,是对上游氨基酸的阻遏调控,从而使叶绿体的结构物质合成受阻,最终导致类囊体膜的退化。     

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。     

Heat shock increases thermotolerance of photosynthetic electron transport and the content of chloroplast membranes and lipids in wheat leaves

Preliminary heating of 15-16-day-old wheat (Triticum aestivum L.) plants for 3 h at 37–38°C (heat shock, HS) increased the tolerance of photosynthetic electron transport (determined as the reduction of 2,6-dichlorophenol indophenol by isolated chloroplasts) toward heating of leaves at 42–48°C in high light (100 klx). At the same time, HS did not affect the activity of the xanthophyll cycle reactions in the 30–48°C temperature range. HS exposure induced an increase in the thylakoid length, the number of grana, and the average number of thylakoids per granum. The volume of the thylakoid system increased 1.4-fold. Such indices as the total content of chlorophylls (a + b), the chlorophyll a/b ratio, as well as the contents of individual carotenoids, chloroplast membrane proteins, and the soluble leaf proteins remained unchanged. The de novo photosynthetic membrane formation was accompanied by the 1.5-fold increase in major chloroplast lipids. It was concluded that, in mature wheat chloroplasts, HS induced the formation of thylakoids characterized by a changed molecular structure and by increased lipid/protein and lipid/chlorophyll ratios.     

Dismantling of Arabidopsis thaliana mesophyll cell chloroplasts during natural leaf senescence

One of the earliest events in the process of leaf senescence is dismantling of chloroplasts. Mesophyll cell chloroplasts from rosette leaves were studied in Arabidopsis thaliana undergoing natural senescence. The number of chloroplasts decreased by only 17% in fully yellow leaves, and chloroplasts were found to undergo progressive photosynthetic and ultrastructural changes as senescence proceeded. In ultrastructural studies, an intact tonoplast could not be visualized, thus, a 35S-GFP::δ-TIP line with a GFP-labeled tonoplast was used to demonstrate that chloroplasts remain outside of the tonoplast even at late stages of senescence. Chloroplast DNA was measured by real-time PCR at four different chloroplast loci, and a fourfold decrease in chloroplast DNA per chloroplast was noted in yellow senescent leaves when compared to green leaves from plants of the same age. Although chloroplast DNA did decrease, the chloroplast/nuclear gene copy ratio was still 31:1 in yellow leaves. Interestingly, mRNA levels for the four loci differed: psbA and ndhB mRNAs remained abundant late into senescence, while rpoC1 and rbcL mRNAs decreased in parallel to chloroplast DNA. Together, these data demonstrate that, during senescence, chloroplasts remain outside of the vacuole as distinct organelles while the thylakoid membranes are dismantled internally. As thylakoids were dismantled, Rubisco large subunit, Lhcb1, and chloroplast DNA levels declined, but variable levels of mRNA persisted.     

表达阳离子抗菌肽的转基因棉花表型异常

用基因枪轰击茎尖法将人工合成的阳离子抗菌肽CEMA基因导入棉花中,获得3株转基因棉花,它们均出现嵌合体表型。在表型不正常的枝条上,叶片扭曲畸形,出现灰绿色与绿色条纹相间的花叶,花器发育异常、无花粉、人工授粉不能结实。花叶区的叶绿素含量降低约20％,叶肉栅栏组织细胞局部缺损,维管束及其管状分子数目减少。叶片灰绿色区叶肉细胞的次生壁明显增厚,局部区域吞噬现象明显,叶绿体内基粒片层数目较少,嗜锇小体明显较多,类囊体腔不明显。PCR分析表明,仅在不正常枝条的花叶中检测到外源CEMA基因。     

建兰水晶艺叶片的超微结构和转录组学分析

水晶艺建兰(Cymbidium ensifolium)因其叶片上呈现出白色透明状,犹如水晶而得名,观赏价值高,但是其形成机理不明确。该研究以建兰‘铁骨水晶’为试验材料,通过对水晶叶片和绿色叶片进行显微结构和超微结构观察,并结合转录组测序等方法,探索建兰水晶艺叶片形成的原因。结果表明:(1)建兰‘铁骨水晶’水晶叶片比绿色叶片薄,叶肉细胞数量减少,形状不规则,且叶绿体含量少;水晶叶片的表皮气孔数量较绿色叶片显著减少;水晶叶片的叶肉细胞中叶绿体结构发育不良,叶绿体双膜和类囊体膜模糊,细胞中存在着大量的嗜锇颗粒。(2)转录组数据分析显示,水晶叶片中与光合作用-天线蛋白、光合作用等代谢途径相关的基因表达量显著下降,而与色素合成代谢途径相关的基因表达量上升。研究推测,建兰水晶艺叶形成的原因可能是由于与光合作用相关的基因表达量降低,导致叶绿体发育不良,叶绿素合成受阻,从而形成白色透明状叶片。     

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.     

Characteristics of Photosynthate Partitioning during Chloroplast Development in Avena Leaves

The first leaves (40 millimeters long) of 4-day-old light-grown Avena sativa L. cv Victory I seedlings contained a complete age sequence of cells from the base to the tip, and within these tissues all stages of chloroplast development could be observed. Although chloroplasts underwent progressive development, a marked increase in number of thylakoids per granum, in chloroplast volume, and in chlorophyll content occurred in the region between 20 and 30 millimeters from the base. Photosynthetic CO₂ fixation (per unit chlorophyll) increased markedly during chloroplast development and closely followed structural changes in chloroplasts. It was also found that the partitioning of photosynthates differed greatly in the segment from 30 to 40 millimeters (at the tip of the leaf) compared with the segment nearer to the leaf base, although both total ¹⁴CO₂ fixation and chlorophyll content per segment did not change significantly along the length of the leaves. As the thylakoid system reached full maturation, partitioning of photosynthates into sucrose increased but partitioning decreased into starch, lipids, and phosphorylated intermediates.     

Three-dimensional organization of higher-plant chloroplast thylakoid membranes revealed by electron tomography

The light-harvesting and energy-transducing functions of the chloroplast are performed within an intricate lamellar system of membranes, called thylakoid membranes, which are differentiated into granum and stroma lamellar domains. Using dual-axis electron microscope tomography, we determined the three-dimensional organization of the chloroplast thylakoid membranes within cryo-immobilized, freeze-substituted lettuce (Lactuca sativa) leaves. We found that the grana are built of repeating units that consist of paired layers formed by bifurcations of stroma lamellar sheets, which fuse within the granum body. These units are rotated relative to each other around the axis of the granum cylinder. One of the layers that makes up the pair bends upwards at its edge and fuses with the layer above it, whereas the other layer bends in the opposite direction and merges with the layer below. As a result, each unit in the granum is directly connected to its neighbors as well as to the surrounding stroma lamellae. This highly connected morphology has important consequences for the formation and function of the thylakoid membranes as well as for their stacking/unstacking response to variations in light conditions.     

Changes in the biochemical composition, structure, and function of pea leaf chloroplasts in iron deficiency and root anoxia

A combined effect of iron deficiency and root anoxia on the biochemical composition, function, and structure of pea leaf chloroplasts was studied. It was found that the chlorosis of apical leaves in response to iron deficiency was determined by the reduction of light-harvesting complexes I and II. Under root anoxia, complexes of the reaction centers of photosystems I and II degraded first. Weak activity was preserved even in yellow and white leaves under the effect of both factors. The ultrastructure of leaf chloroplasts gradually degraded. Initially, intergranal thylakoid sites were reduced, and the longitudinal orientation of grana was disturbed. However, yellow and white leaves still retained small thylakoids and grana. It is concluded that the degrading effects of iron deficiency and root anoxia on the complex composition and leaf chloroplast structure and function are additive because of their autonomous mechanisms.     

Changes in the Biochemical Composition, Structure, and Function of Pea Leaf Chloroplasts in Iron Deficiency and Root Anoxia

A combined effect of iron deficiency and root anoxia on the biochemical composition, function, and structure of pea leaf chloroplasts was studied. It was found that the chlorosis of apical leaves in response to iron deficiency was determined by the reduction of light-harvesting complexes I and II. Under root anoxia, complexes of the reaction centers of photosystems I and II degraded first. Weak activity was preserved even in yellow and white leaves under the effect of both factors. The ultrastructure of leaf chloroplasts gradually degraded. Initially, intergranal thylakoid sites were reduced, and the longitudinal orientation of grana was disturbed. However, yellow and white leaves still retained small thylakoids and grana. It is concluded that the degrading effects of iron deficiency and root anoxia on the complex composition and leaf chloroplast structure and function are additive because of their autonomous mechanisms.     

Transformation of plastids in the leaves of Acer negundo L. var. odessanum (H. Rothe).

The leaves of Acer negundo L. var. odessanum (H. Rothe), if permanently exposed to strong sunlight, do not green, but remain yellow and finally become bleached. In yellow leaves the plastids contain single thylakoids and no grana. In plastids of bleached leaves, however, only vesicles are present. The concentration of chlorophylls and photosynthetic activity are much lower in those leaves than in the green ones. If the illumination is reduced (e.g. by shading) both the yellow and the bleached leaves become greenish, and even fully green after a few days at a sufficiently low light intensity. The plastids of yellow-green leaves contain small grana. In dark green leaves the thylakoid system of the chloroplasts is normally developed forming true grana, regardless of whether the leaves were originally green, or became green by shading the yellow or bleached ones. Their pigment concentration and photosynthetic activity are also normal. If green leaves are exposed to sunlight they do not yellow or bleach. During a 3-week period the structure of the thylakoid system did not perceivably change, with the exception that large plastoglobules formed in the stroma.     

Morphometric Analysis of Chloroplasts of Cotton Leaf and Fruiting Organs

We examined morphological and ultrastructural differences in chloroplasts of cotton leaves and the fruiting organs, bract, and capsule wall to advance our understanding of their commonly observed differences in photosynthetic efficiency. Chloroplasts from leaves were large (7.1 μm long in cross section), lens shaped with a well developed membrane system differentiated into grana and stroma lamellae that occupied the large cross-sectional area (12.3 μm²) of the organelle. A few small plastoglobuli and starch grains were scattered in the stroma region. The bract chloroplasts were correlative of leaf chloroplasts in size (6.8 μm in length) and shape with the exception that the bract chloroplasts exhibited greater thylakoid number per granum (15.8) than the leaf chloroplasts (10.5). In contrast to leaf and bract, the capsule wall chloroplasts were smaller in size (4.3 μm) and cross sectional area (6.8 μm²) than either the leaf or bract. The most intriguing feature of the capsule wall chloroplasts was its domination by large starch granules (5.3 μm²) in the stroma which filled the whole chloroplast coercing the membrane system to move towards the periphery of the organelle. Grana number and thylakoids per granum were lowest in the capsule wall chloroplasts. This revised version was published online in July 2006 with corrections to the Cover Date.     

花粒期光照对夏玉米光合特性和叶绿体超微结构的影响

在大田条件下,以夏玉米品种‘登海605’为试验材料,研究花粒期不同光照强度(正常光照、开花至收获期遮阴和开花至收获期增光)对夏玉米叶片光合、荧光性能和叶绿体超微结构的影响.结果表明:与对照相比,花粒期遮阴影响叶绿体排布及内部结构发育,基粒个数和基粒片层数均有不同程度减少,叶片的净光合速率、蒸腾速率、气孔导度、叶绿素含量下降,PSⅡ反应中心的实际光化学效率和最大光化学效率降低,非光化学淬灭系数数值增加,导致产量降低;增光后叶绿体结构良好,基粒片层排列紧致、清晰且数量增加,PSⅡ反应中心的实际光化学效率增加,净光合速率、蒸腾速率、气孔导度、叶绿素含量上升,叶片光合性能增强,产量增加.即花粒期遮阴破坏了夏玉米叶片叶绿体超微结构,降低了叶片光合能力,产量下降;花粒期增光增加了叶肉细胞中叶绿体的基粒和基粒片层,导致基粒片层排列紧密有序,有利于增加作物产量潜力.     

Comparative ultrastructural properties of pallisade tissue and spongy tissue chloroplasts from normal and mutant leaves of Pisum sativum L.*

Abstract. The ultrastructure of chloroplasts from palisade and spongy tissue was studied in order to analyse the adaptation of chloroplasts to the light gradient within the bifacial leaves of pea. Chloroplasts of two nuclear gene mutants of Pisum sativum (chlorotica-29 and chlorophyll b-less 130A), grown under normal light conditions, were compared with the wild type (WT) garden-pea cv. ‘Dippes Gelbe Viktoria’. The differentiation of the thylakoid membrane system of plastids from normal pea leaves exhibited nearly the same degree of grana formation in palisade and in spongy tissue. Using morphometrical measurements, only a slight increase in grana stacking capacity was found in chloroplasts of spongy tissue. In contrast, chloroplasts of mutant leaves differed in grana development in palisade and spongy tissue, respectively. Their thylakoid systems appeared to be disorganized and not developed as much as in chloroplasts from normal pea leaves. Grana contained fewer lamellae per granum, the number of grana per chloroplast section was reduced and the length of appressed thylakoid regions was decreased. Nevertheless, chloroplasts of the mutants were always differentiated into grana and stroma thylakoids. The structural changes observed and the reduction of the total chlorophyll content correlated with alterations in the polypeptide composition of thylakoid membrane preparations from mutant chloroplasts. In sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE), polypeptide bands with a relative molecular mass of 27 and 26 kilodalton (kD) were markedly reduced in mutant chloroplasts. These two polypeptides represented the major apoproteins of the light harvesting chlorophyll a/b complex from photosystem II (LHC-II) as inferred from a comparison with the electrophoretic mobility of polypeptides isolated from the LHC-II.     

烟草叶片衰老期过程中的蛋白质组学分析

大田烟叶生产过程中因打顶打叉的处理,改变了烟叶正常的衰老模式。为研究这一特殊的衰老机制,我们自旺长期开始,对‘云烟87’不同发育阶段烟株的中部叶片,进行形态观测、生理生化分析及蛋白质组学检测。结果显示：随着烟叶的逐渐成熟和衰老,烟草的叶色逐渐变黄,叶片逐渐变短、变窄,厚度减少;解剖结构清晰看到栅栏组织和海绵组织从最初的整齐排列到逐渐排列紊乱,组织细胞间轮廓不明显,细胞间隙明显增大;亚显微观测表明,淀粉粒在叶绿体中逐渐积累,类囊体片层结构被挤散,叶绿体膜被撑破。生理与生化分析表明衰老过程伴随着光合作用速率下降,光合色素降解加速,呼吸代谢的增加,这可能与衰老叶片中叶绿体逐渐崩塌和细胞膜透性增加相一致。iTRAQ标记方法共检测到不同发育阶段432个差异表达蛋白质,其中注释到308个与多种生命过程相关。蛋白差异富集分析表明,烟草叶片衰老过程中与光合作用等合成代谢相关蛋白多下调表达,而逆境反应及呼吸作用等分解代谢相关蛋白多上调表达。