一个新的水稻叶绿素缺失黄叶突变体的特征及基因分子定位

从粳稻"嘉花1号"60Coγ射线辐照的后代中筛选到一个叶绿素缺失黄叶突变体(yl11),与野生型"嘉花1号"相比该突变体表现为全生育期植株叶片呈黄色,叶绿素含量以及净光合速率明显下降,叶绿体发育不完善,并且伴随着株高等主要农艺性状的变化。遗传分析表明,该突变性状受一对隐性核基因(yl11)控制。该突变体与籼稻"培矮64S"杂交生产的F2、F3群体中的分离出突变体型920个单株作为定位群体,利用SSR和InDel分子标记将yl11基因定位在水稻第11染色体长臂上的MM2199和ID21039分子标记之间,其物理距离约为110kb,目前该区域内没有发现与水稻叶绿素合成/叶绿体发育相关已知功能基因。研究结果为今后对该基因的克隆和功能分析奠定了基础。     

一个新的水稻叶绿素缺失黄叶突变体的特征及基因分子定位

从粳稻“嘉花1号”⁶⁰Coγ射线辐照的后代中筛选到一个叶绿素缺失黄叶突变体(yl11), 与野生型“嘉花1号”相比该突变体表现为全生育期植株叶片呈黄色, 叶绿素含量以及净光合速率明显下降, 叶绿体发育不完善, 并且伴随着株高等主要农艺性状的变化。遗传分析表明, 该突变性状受一对隐性核基因(yl11)控制。该突变体与籼稻“培矮64S”杂交生产的F₂、F₃群体中的分离出突变体型920个单株作为定位群体, 利用SSR和InDel分子标记将yl11基因定位在水稻第11染色体长臂上的MM2199和ID21039分子标记之间, 其物理距离约为110 kb, 目前该区域内没有发现与水稻叶绿素合成/叶绿体发育相关已知功能基因。研究结果为今后对该基因的克隆和功能分析奠定了基础。     

Characterization of a rice chlorophyll-deficient mutant using the T-DNA gene-trap system

We have previously generated a large pool of T-DNA insertional lines in rice. In this study, we screened those T-DNA pools for rice mutants that had defective chlorophylls. Among the 1,995 lines examined in the T2 generation, 189 showed a chlorophyll-deficient phenotype that segregated as a single recessive locus. Among the mutants, 10 lines were beta-glucuronidase (GUS)-positive in the leaves. Line 9-07117 has a T-DNA insertion into the gene that is highly homologous to XANTHA-F in barley and CHLH in ARABIDOPSIS: This OsCHLH gene encodes the largest subunit of the rice Mg-chelatase, a key enzyme in the chlorophyll branch of the tetrapyrrole biosynthetic pathway. In the T2 and T3 generations, the chlorina mutant phenotypes are co-segregated with the T-DNA. We have identified two additional chlorina mutants that have a Tos17 insertion in the OsCHLH gene. Those phenotypes were co-segregated with Tos17 in the progeny. GUS assays and RNA blot analysis showed that expression of the OsCHLH gene is light inducible, while TEM analysis revealed that the thylakoid membrane of the mutant chloroplasts is underdeveloped. The chlorophyll content was very low in the OschlH mutants. This is the first report that T-DNA insertional mutagenesis can be used for functional analysis of rice genes.     

Leaf anatomy and chloroplast ultrastructure of Mn-deficient orange plants

Leaf samples of Mn-deficient and Mn-sufficient (control) ‘Navelate’ orange plants grown in a greenhouse were taken to investigate the effects of Mn deficiency in leaf structure and chloroplast ultrastructure. Total leaf chlorophyll concentration was significantly lower in Mn-deficient plants than in control ones. Entire lamina thickness was not altered due to Mn deficiency. However, Mn deficiency resulted in disorganization of mesophyll cells, mainly of palisade parenchyma cells. The number of mesophyll chloroplasts per cellular area and their length were both affected negatively. The membranous system of chloroplasts was also disorganized. The percentages of starch grains and plastoglobuli per chloroplast of Mn-deficient leaves were significantly greater than those of control leaves.     

A chlorophyll-deficient rice mutant with impaired chlorophyllide esterification in chlorophyll biosynthesis

Chlorophyll (Chl) synthase catalyzes esterification of chlorophyllide to complete the last step of Chl biosynthesis. Although the Chl synthases and the corresponding genes from various organisms have been well characterized, Chl synthase mutants have not yet been reported in higher plants. In this study, a rice (Oryza Sativa) Chl-deficient mutant, yellow-green leaf1 (ygl1), was isolated, which showed yellow-green leaves in young plants with decreased Chl synthesis, increased level of tetrapyrrole intermediates, and delayed chloroplast development. Genetic analysis demonstrated that the phenotype of ygl1 was caused by a recessive mutation in a nuclear gene. The ygl1 locus was mapped to chromosome 5 and isolated by map-based cloning. Sequence analysis revealed that it encodes the Chl synthase and its identity was verified by transgenic complementation. A missense mutation was found in a highly conserved residue of YGL1 in the ygl1 mutant, resulting in reduction of the enzymatic activity. YGL1 is constitutively expressed in all tissues, and its expression is not significantly affected in the ygl1 mutant. Interestingly, the mRNA expression of the cab1R gene encoding the Chl a/b-binding protein was severely suppressed in the ygl1 mutant. Moreover, the expression of some nuclear genes associated with Chl biosynthesis or chloroplast development was also affected in ygl1 seedlings. These results indicate that the expression of nuclear genes encoding various chloroplast proteins might be feedback regulated by the level of Chl or Chl precursors.     

Electron transport and chloroplast ultrastructure of a chlorophyll deficient mutant of wheat

A non-lethal chlorophyll deficient mutation was induced by use of the chemical mutagen ethyl methanesulfonate. Chloroplasts from the control and mutant plants were found to be very similar ultrastructurally. Thylakoid membrane volume was only slightly greater in plastids from the control as compared with plastids from the mutant. The chlorophyll content of the mutant was reduced by over 60%. This decrease in chlorophyll was not accompanied by a similar decrease in electron transport. Uncoupled electron transport rate based on a unit chlorophyll basis was nearly twice as great for mutant chloroplasts as for control plastids. However, electron transport rate based on a unit membrane volume was similar in mutant and control plants. At high irradiance the relative quantum requirement of the control and mutant was similar when expressed on membrane volume.     

Abnormal regulation of photosynthetic electron transport in a chloroplast ycf9 inactivation mutant

The ycf9 (orf62) gene of the plastid genome encodes a 6.6-kDa protein (ORF62) of thylakoid membranes. To elucidate the role of the ORF62 protein, the coding region of the gene was disrupted with an aadA cassette, yielding mutant plants that were nearly (more than 95%) homoplasmic for ycf9 inactivation. The ycf9 mutant had no altered phenotype under standard growth conditions, but its growth rate was severely reduced under suboptimal irradiances. On the other hand, it was less susceptible to photodamage than the wild type. ycf9 inactivation resulted in a clear reduction in protein amounts of CP26, the NAD(P)H dehydrogenase complex, and the plastid terminal oxidase. Furthermore, depletion of ORF62 led to a faster flow of electrons to photosystem I without a change in the maximum electron transfer capacity of photosystem II. Despite the reduction of CP26 in the mutant thylakoids, no differences in PSII oxygen evolution rates were evident even at low light intensities. On the other hand, the ycf9 mutant presented deficiencies in the capacity for PSII-independent electron transport (ferredoxin-dependent cyclic electron transport and NAD(P)H dehydrogenase-mediated plastoquinone reduction). Altogether, it is shown that depletion of ORF62 leads to anomalies in the photosynthetic electron transfer chain and in the regulation of electron partitioning among the different routes of electron transport.     

Light metabolism and chloroplast structure in chlorophyll-deficient tobacco mutants

In tobacco mutants which contain 1/8 to 1/30 of the normal chlorophyll content per leaf area the content of yellow pigments (carotenoids) is also diminished but less in proportion to the chlorophyll content. The pale yellow-green mutant grows and matures provided that light intensity and temperature make up for the chlorophyll deficiency. In most green plants and algae light saturation of photosynthesis is reached between 5000 and 12,000 ergs/sec.cm2. The mutants continue to give higher photosynthetic rates until the incident intensity reaches 50,000 ergs/sec.cm2. While often unable to compensate their respiration at intensities at which the normal green plant approaches saturation, the pale yellow-green leaves are able to provide the mutant plant with two to three times the absolute amount of carbon dioxide assimilation per hour and leaf area at 50,000 ergs/sec.cm2 and 20 degrees to 25 degrees C. These observations are valid for red light lambda > 600 m mu. In blue light lambda < 575 m mu (below saturation levels) the mutants separate into two classes, one in which absorption by some carotenoid enhances the photosynthetic rate and the other in which the absorbing pigments are inactive and therefore depress the rate strongly. The unusual kinetics of photosynthesis in these chlorophyll-deficient tobacco mutants is reflected in the structure of their chloroplasts which we found to be of a kind thus far not described for healthy, normally growing, higher plants.     

Effects of low nocturnal temperature on photosynthetic characteristics and chloroplast ultrastructure of winter rapeseed

We investigated the effects of low nocturnal temperature on photosynthetic apparatus of winter rapeseed (Brassica campestris L.). An artificial climate chamber was used to simulate the effects of low nocturnal temperature on seedling and stomatal morphology, chloroplast ultrastructure, photosynthetic parameters, and dry matter distribution and accumulation in two winter rapeseed cultivars, Longyou-7 (ultra coldresistant) and Tianyou-2 (weak cold resistance). Compared with those at diurnal/nocturnal temperatures of 20°/10°C (control), rapeseed seedlings at 20°/5°C had increased leaf chlorophyll content, deepened green leaf color, decreased stomatal conductance (G_s), intercellular CO₂ concentration (C_i), and photosynthetic rate (P_n), and improved root/shoot ratio; the majority of stomata remained open in Longyou-7 while those in Tianyou-2 were mostly closed or semi-closed. At diurnal/nocturnal temperatures of 20°/–5°C, rapeseed seedlings had decreased leaf chlorophyll content with increased C_i but decreased G_s and P_n; Tianyou-2 exhibited ruptured chloroplast membrane, dissolved grana, broken stroma lamella, and decreased root/shoot ratio, whereas Longyou-7 had chloroplasts retaining partial structure of grana with a small amount of starch granules in guard cells. Low nocturnal temperature damaged the photosynthetic membrane of chloroplasts and reduced P_n in the leaves of winter rapeseed influencing photosynthetic processes in this crop. The reduction of P_n was mainly related to stomatal limitation at diurnal/nocturnal temperatures of 20°/5°C and non-stomatal limitation at diurnal/nocturnal temperatures of 20°/–5°C.     

Comparative analysis of ultrastructure,antioxidant enzyme activities,and photosynthetic performance in rice mutant 812HS prone to photooxidation

Under optimal conditions, most of the light energy is used to drive electron transport. However, when the light energy exceeds the capacity of photosynthesis, the overall photosynthetic efficiency drops down. The present study investigated the effects of high light on rice photooxidation-prone mutant 812HS, characterized by a mutation of leaf photooxidation 1 gene, and its wild type 812S under field conditions. Our results showed no significant difference between 812HS and 812S before exposure to high sunlight. However, during exposure to high light, shoot tips of 812HS turned yellow and their chlorophyll (Chl) content decreased. Transmission electron microscopy showed that photooxidation resulted in significant damage of chloroplast ultrastructure. It was confirmed also by inhibited photophosphorylation and reduced ATP content. The decreased coupling factor of ATP, Ca²⁺-ATPase and Mg²⁺-ATPase activities also verified these results. Further, significantly enhanced activities of antioxidative enzymes were observed during photooxidation. Malondialdehyde, hydrogen peroxide, and the superoxide generation rates also increased. Chl a fluorescence analysis found that the performance index and maximum quantum yield of PSII declined on August 4, 20 days after high-light treatment. Net photosynthetic rate also decreased and substomatal CO₂ concentration increased in 812HS at the same time. In conclusion, our findings indicated that excessive energy triggered the production of toxic reactive oxygen species and promoted lipid peroxidation in 812HS plants, causing severe damage to cell membranes, degradation of photosynthetic pigments and proteins, and ultimately inhibition of photosynthesis.     

弱光对不结球白菜光合特性与叶绿体超微结构的影响

研究了不结球白菜耐弱光品种“矮王”和不耐弱光品种“绿优”经弱光处理后,光合特性和叶绿体超微结构的变化.结果表明:遮荫处理后,“矮王”的总叶绿素和叶绿素b含量大幅度上升;2个品种的净光合速率（P_n）、光补偿点（LCP）、光饱和点（(LSP）、表观量子效率（AQY）、羧化效率（CE）均呈下降趋势;除光补偿点外,“矮王”的下降幅度均小于“绿优”.叶绿素荧光参数分析表明,在弱光条件下,“绿优”的光系统Ⅱ（PSII）受到伤害,F_v/F_m、ETR、Φ_PSⅡ和q_P的下降幅度均大于耐弱光品种“矮王”.电镜超微观察结果显示:“绿优”的叶绿体内部出现裂缝和孔洞,多数基质片层断裂,基粒片层解体,线粒体出现内含物减少、发生空洞的现象;而“矮王”的叶绿体结构良好,线粒体发育正常.表明弱光处理可以影响植物的光合机构,但对不同品种的影响不同.     

Responses of photosynthetic properties and chloroplast ultrastructure of Bryum argenteum from a desert biological soil crust to elevated ultraviolet‐B radiation

Our understanding of plant responses to enhanced ultraviolet‐B (UV‐B) radiation has improved over recent decades. However, research on cryptogams is scarce and it remains controversial whether UV‐B radiation causes changes in physiology related to photosynthesis. To investigate the effects of supplementary UV‐B radiation on photosynthesis and chloroplast ultrastructure in Bryum argenteum Hedw., specimens were cultured for 10 days under four UV‐B treatments (2.75, 3.08, 3.25 and 3.41 W m^–2), simulating depletion of 0% (control), 6%, 9% and 12% of stratospheric ozone at the latitude of Shapotou, a temperate desert area of northwest China. Analyses showed malondialdehyde content significantly increased, whereas chlorophyll (Chl) fluorescence parameters and Chl contents decreased with increased UV‐B intensity. These results corresponded with changes in thylakoid protein complexes and chloroplast ultrastructure. Overall, enhanced UV‐B radiation leads to significant decreases in photosynthetic function and serious destruction of the chloroplast ultrastructure of B. argenteum. The degree of negative influences increased with the intensity of UV‐B radiation. These results may not only provide a potential mechanism for supplemental UV‐B effects on photosynthesis of moss crust, but also establish a theoretical basis for further studies of adaptation and response mechanisms of desert ecosystems under future ozone depletion.     

Leaf senescence in a non-yellowing mutant of Festuca pratensis: Photosynthesis and photosynthetic electron transport

The photosynthetic capacity of detached leaves of a non-yellowing mutant of Festuca pratensis Huds. declined during senescence at a similar rate to that in a normal cultivar. Respiratory oxygen uptake in the dark continued at similar rates in both genotypes during several days of senescence. In chloroplasts isolated from leaves at intervals after excision, the rate of photosystem I (PS I)-mediated methyl viologen reduction using reduced N,N,N,N-tetramethyl-p-phenylene diamine as electron donor also declined in both genotypes, possibly due to loss of integrity of the photosynthetic apparatus in the cytochrome f-plastocyanin region. There was a similar fall in PS II electron transport using water as electron donor and measured at the rate of reduction of 2,6-dichlorophenolindophenol. Partial restoration of this activity by the addition of diphenyl carbazide was evidence for lability of the oxygen-evolving complex during senescence. An accentuated difference between mutant and normal material in this case indicated that the mutant retains a greater number of functional PS II centres. Changes in the light-saturation characteristics of the two photosystems have been discussed in relation to the organization of the photosynthetic membranes during senescence.Abbreviations and symbols DCMU 3-(3,4-dichlorophenyl)-1,1-dimethylurea - DCPIP 2,6-dichlorophenolindophenol - DMSO dimethyl sulphoxide - DPC diphenyl carbazide - MV methyl viologen - PS I, PS II photosystem I, II - TMPD N,N,N,N-tetramethyl-p-phenylene diamine     

High photosynthetic efficiency of a rice (Oryza sativa L.) xantha mutant

Comparative analysis revealed that a xantha rice mutant (cv. Huangyu B) had higher ratios of chlorophyll (Chl) a/b and carotenoids/Chl, and higher photosynthetic efficiency than its wild type parent (cv. II32 B). Unexpectedly, the mutant had higher net photosynthetic rate (P _N) than II32 B. This might have resulted from its lower non-photochemical quenching (q_N) but higher maximal photochemical efficiency (F_V/F_M), higher excitation energy capture efficiency of photosystem 2 (PS2) reaction centres (F_V′/F_M′), higher photochemical quenching (q_P), higher effective PS2 quantum yield (Φ_PS2), and higher non-cyclic electron transport rate (ETR). This is the first report of a chlorophyll mutant that has higher photosynthetic efficiency and main Chl fluorescence parameters than its wild type. This mutant could become a unique material both for the basic research on photosynthesis and for the development of high yielding rice cultivars.     

等渗NaCl、干旱胁迫对番茄幼苗光合特性及叶绿体超微结构的影响

为分析等渗盐分和水分胁迫对番茄叶片光合功能的影响,选用耐盐性不同的4种基因型番茄(小果型的辽园红玛瑙、野生醋栗番茄、大果型的金田粉冠、超402)进行等渗的140 mmol·L^-1NaCl和15%PEG6000模拟盐分和水分胁迫.结果表明: 在光合特性方面,处理12 d后,两种胁迫导致4种不同基因型番茄的叶绿素含量(叶绿素a、叶绿素b、叶绿素总量)、光合速率、气孔导度、胞间CO₂浓度、蒸腾效率降低,气孔限制值升高.两种胁迫导致的光合下降由气孔性因素所致.等渗的盐分胁迫对番茄的光合系统损伤大于水分胁迫,这是因为盐分胁迫除了渗透胁迫还会导致离子伤害.4种不同基因型番茄中,耐盐型的辽园红玛瑙具有高光合特性,金田粉冠光合效率最差.在叶绿体超微结构方面,两种胁迫会造成番茄叶片的气孔密度增加,气孔张开率降低,叶绿体长度增加,宽度变小,长宽比增大,叶绿体内基粒数减少,嗜锇颗粒数增多.盐分胁迫下两种番茄叶绿体结构的影响大于水分胁迫,耐盐种醋栗番茄气孔变化小于盐敏感品种金田粉冠.     

外源亚精胺对NaCl胁迫下毕氏海蓬子光合参数和叶绿体超微结构的影响

在0、100、300、500和700 mmol·L-1NaCl胁迫条件下比较了喷施0.1mmol·L-1亚精胺(Spd)对毕氏海蓬子(Salicomia bigelovii Torr.)幼苗叶绿素含量、净光合速率、气孔导度、胞间CO2浓度和叶绿体超微结构的影响.结果表明:毕氏海蓬子的叶绿素含量、净光合速率和气孔导度均呈低浓度条件下(0、100和300 mmol·L-1NaCl)升高、高浓度条件下(500和700 mmol·L-1NaCl)降低的趋势,在300 mmol·L-1 NaCl胁迫条件下达到最高值:胞间CO2浓度则呈低浓度NaCl胁迫条件下降低、500 mmol·L-1NaCl条件下升高、700 mmol·L-1NaCl条件下略降低的趋势;在0～500 mmol·L-1NaCl胁迫条件下叶绿素a/b值变化不明显,但在700 mmol·L-1NaCl条件下急剧降低.在低浓度NaCl胁迫条件下,叶绿体整体膨胀,类囊体片层结构松散,但叶绿体和类囊体结构仍保持完整;而经500和700mmol·L-1NaCl处理后,叶绿体超微结构被严重破坏,叶绿体膜结构破裂、类囊体结构松散呈放射状、有些叶绿体完全解体.而在相应的NaCl胁迫条件下喷施0.1 mmol·L-1Spd,毕氏海蓬子的叶绿素含量、净光合速率、气孔导度和胞间CO2浓度虽然也呈现出相同的变化趋势,但其数值均显著高于对照(未喷施Spd);且叶绿体超微结构的损伤程度也轻于对照.研究结果说明:喷施外源Spd能够减缓NaCl胁迫对毕氏海蓬子的伤害作用.     

The effects of reduced amounts of lipid unsaturation on chloroplast ultrastructure and photosynthesis in a mutant of Arabidopsis

A mutant of Arabidopsis thaliana with reduced content of C_18:3 and C_16:3 fatty acids in membrane lipids exhibited a 45% reduction in the cross-sectional area of chloroplasts and had a decrease of similar magnitude in the amount of chloroplast lamellar membranes. The reduction in chloroplast size was partially compensated by a 45% increase in the number of chloroplasts per cell in the mutant. When expressed on a chlorophyll basis the rates of CO₂-fixation and photosynthetic electron transport were not affected by these changes. Fluorescence polarization measurements indicated that the fluidity of the thylakoid membranes was not significantly altered by the mutation. Similarly, on the basis of temperature-induced fluorescence yield enhancement measurements, there was no significant effect on the thermal stability of chlorophyll-protein complexes in the mutant. These observations suggest that the high content of trienoic fatty acids in chloroplast lipids may be an important factor regulating organelle biogenesis but is not required to support normal levels of the photosynthetic activities associated with the thylakoid membranes.     

UV-B辐射增强对芦苇光合生理及叶绿体超微结构的影响

在UV-B辐照增强条件下,研究不同辐照梯度对芦苇光合特性、光合色素含量及叶绿体超微结构的影响. 结果表明: 与自然光照相比,UV-B辐照增强显著降低芦苇叶片净光合速率、气孔导度、蒸腾速率,且随辐照强度增大,降低程度加剧,胞间CO₂浓度升高,光合效率显著降低;与自然光照相比,UV-B辐照增强显著降低芦苇叶片光合色素含量(包括叶绿素a、叶绿素b、总叶绿素和类胡萝卜素),且随辐照强度增大,降低程度加剧;UV-B辐射增强条件下,叶绿体超微结构遭到破坏,表现为叶绿体结构变形,类囊体片层排列稀疏紊乱、膨胀甚至模糊不清,并且UV-B辐射强度越大,损伤越大,高强度UV-B辐射对叶绿体超微结构的影响大于低强度辐射.